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Sample records for a549 xenograft model

  1. Irradiation-Dependent Effects on Tumor Perfusion and Endogenous and Exogenous Hypoxia Markers in an A549 Xenograft Model

    SciTech Connect

    Fokas, Emmanouil; Haenze, Joerg; Kamlah, Florentine; Eul, Bastian G.; Lang, Nico; Keil, Boris; Heverhagen, Johannes T.; Engenhart-Cabillic, Rita; An Hanxiang; Rose, Frank

    2010-08-01

    Purpose: Hypoxia is a major determinant of tumor radiosensitivity, and microenvironmental changes in response to ionizing radiation (IR) are often heterogenous. We analyzed IR-dependent changes in hypoxia and perfusion in A549 human lung adenocarcinoma xenografts. Materials and Methods: Immunohistological analysis of two exogenously added chemical hypoxic markers, pimonidazole and CCI-103F, and of the endogenous marker Glut-1 was performed time dependently after IR. Tumor vessels and apoptosis were analyzed using CD31 and caspase-3 antibodies. Dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) and fluorescent beads (Hoechst 33342) were used to monitor vascular perfusion. Results: CCI-103F signals measuring the fraction of hypoxic areas after IR were significantly decreased by approximately 50% when compared with pimonidazole signals, representing the fraction of hypoxic areas from the same tumors before IR. Interestingly, Glut-1 signals were significantly decreased at early time point (6.5 h) after IR returning to the initial levels at 30.5 h. Vascular density showed no difference between irradiated and control groups, whereas apoptosis was significantly induced at 10.5 h post-IR. DCE-MRI indicated increased perfusion 1 h post-IR. Conclusions: The discrepancy between the hypoxic fractions of CCI-103F and Glut-1 forces us to consider the possibility that both markers reflect different metabolic alterations of tumor microenvironment. The reliability of endogenous markers such as Glut-1 to measure reoxygenation in irradiated tumors needs further consideration. Monitoring tumor microvascular response to IR by DCE-MRI and measuring tumor volume alterations should be encouraged.

  2. Tumor growth affects the metabonomic phenotypes of multiple mouse non-involved organs in an A549 lung cancer xenograft model

    PubMed Central

    Xu, Shan; Tian, Yuan; Hu, Yili; Zhang, Nijia; Hu, Sheng; Song, Dandan; Wu, Zhengshun; Wang, Yulan; Cui, Yanfang; Tang, Huiru

    2016-01-01

    The effects of tumorigenesis and tumor growth on the non-involved organs remain poorly understood although many research efforts have already been made for understanding the metabolic phenotypes of various tumors. To better the situation, we systematically analyzed the metabolic phenotypes of multiple non-involved mouse organ tissues (heart, liver, spleen, lung and kidney) in an A549 lung cancer xenograft model at two different tumor-growth stages using the NMR-based metabonomics approaches. We found that tumor growth caused significant metabonomic changes in multiple non-involved organ tissues involving numerous metabolic pathways, including glycolysis, TCA cycle and metabolisms of amino acids, fatty acids, choline and nucleic acids. Amongst these, the common effects are enhanced glycolysis and nucleoside/nucleotide metabolisms. These findings provided essential biochemistry information about the effects of tumor growth on the non-involved organs. PMID:27329570

  3. PKM2 Thr454 phosphorylation increases its nuclear translocation and promotes xenograft tumor growth in A549 human lung cancer cells.

    PubMed

    Yu, Zhenhai; Huang, Liangqian; Qiao, Pengyun; Jiang, Aifang; Wang, Li; Yang, Tingting; Tang, Shengjian; Zhang, Wei; Ren, Chune

    2016-05-13

    Pyruvate kinase M2 (PKM2) is a key enzyme of glycolysis which is highly expressed in many tumor cells, and plays an important role in the Warburg effect. In previous study, we found PIM2 phosphorylates PKM2 at Thr454 residue (Yu, etl 2013). However, the functions of PKM2 Thr454 modification in cancer cells still remain unclear. Here we find PKM2 translocates into the nucleus after Thr454 phosphorylation. Replacement of wild type PKM2 with a mutant (T454A) enhances mitochondrial respiration, decreases pentose phosphate pathway, and enhances chemosensitivity in A549 cells. In addition, the mutant (T454A) PKM2 reduces xenograft tumor growth in nude mice. These findings demonstrate that PKM2 T454 phosphorylation is a potential therapeutic target in lung cancer. PMID:27045080

  4. Inhibition of Tumor Growth and Angiogenesis by a Lysophosphatidic Acid Antagonist in a Engineered Three-dimensional Lung Cancer Xenograft Model

    PubMed Central

    Xu, Xiaoyu; Prestwich, Glenn D

    2009-01-01

    BACKGROUND We developed an engineered three-dimensional (3-D) tumor xenograft model of non-small cell lung cancer (NSCLC) in nude mice, and used this model to evaluate a dual-activity inhibitor of lysophosphatidic acid (LPA) biosynthesis and receptor activation. METHODS First, BrP-LPA, a pan-antagonist for four LPA receptors and inhibitor of the lyosphospholipase D activity of autotaxin, was examined for inhibition of cell migration and cell invasion by human NSCLC A549 cells. Second, A549 cells were encapsulated in 3-D in three semi-synthetic ECMs based on chemically-modified glycosaminoglycans, and injected subcutaneously in nude mice. Tumor volume and vascularity were deteremined as a function of sECM composition. Third, engineered NSCLC xenografts were formed from A549 cells in either Extracel-HP or Matrigel, and mice were treated with four intraperitoneal injections of 3 mg/kg of BrP-LPA. RESULTS First, BrP-LPA inhibited cell migration and invasiveness of A549 cells in vitro. Second, tumor growth and microvessel formation for 3-D encapsulated A549 cells in vivo in nude mice increased in the order: buffer only < Extracel < Extracel-HP < Extracel-HP containing growth factors plus laminin. Third, tumor volumes increased rapidly in both Matrigel and Extracel-HP encapsulated A549 cells, and tumor growth was markedly inhibited by BrP-LPA treatment. Finally, tumor vascularization was dramatically reduced in the A549 tumors treated with BrP-LPA. CONCLUSIONS Engineered A549 lung tumors can be created by 3-D encapsulation in an ECM substitute with user controlled composition. The engineered tumors regress and lose vascularity in response to a dual activity inhibitor of the LPA signaling pathway. PMID:20143443

  5. Vitamin D Analogs Potentiate the Antitumor Effect of Imatinib Mesylate in a Human A549 Lung Tumor Model

    PubMed Central

    Maj, Ewa; Filip-Psurska, Beata; Świtalska, Marta; Kutner, Andrzej; Wietrzyk, Joanna

    2015-01-01

    In previous papers, we presented data on studies on the anticancer activity of the vitamin D3 analogs, named PRI-2191 and PRI-2205, in different cancer models. In this study, we showed the improved antiproliferative activity of a combination of imatinib mesylate (Gleevec, GV) and cytostatic agents in in vitro studies, when used with a third compound, namely PRI-2191, in an A549 human lung cancer model. Furthermore, we analyzed the influence of both PRI-2191, as well as PRI-2205 on the anticancer activity of GV in mice bearing A549 tumors. The route of PRI-2191 analog administration showed a significant impact on the outcome of GV treatment: subcutaneous injection was more efficient and less toxic than oral gavage. Moreover, both vitamin D compounds increased the anticancer activity of GV; however, they might also potentiate some adverse effects. We also evaluated in tumor tissue the expression of VEGF, PDGF-BB, vitamin D receptor, CYP27B1, CYP24, p53 and Bcl-2, as well as PDGF receptors: α and β. We observed the upregulation of p53 expression and the downregulation of Bcl-2, as well as VEGF in A549 tumors as a result of the tested treatment. However, vitamin D analogs did not significantly influence the expression of these proteins. PMID:26580599

  6. Vitamin D Analogs Potentiate the Antitumor Effect of Imatinib Mesylate in a Human A549 Lung Tumor Model.

    PubMed

    Maj, Ewa; Filip-Psurska, Beata; Świtalska, Marta; Kutner, Andrzej; Wietrzyk, Joanna

    2015-11-13

    In previous papers, we presented data on studies on the anticancer activity of the vitamin D₃ analogs, named PRI-2191 and PRI-2205, in different cancer models. In this study, we showed the improved antiproliferative activity of a combination of imatinib mesylate (Gleevec, GV) and cytostatic agents in in vitro studies, when used with a third compound, namely PRI-2191, in an A549 human lung cancer model. Furthermore, we analyzed the influence of both PRI-2191, as well as PRI-2205 on the anticancer activity of GV in mice bearing A549 tumors. The route of PRI-2191 analog administration showed a significant impact on the outcome of GV treatment: subcutaneous injection was more efficient and less toxic than oral gavage. Moreover, both vitamin D compounds increased the anticancer activity of GV; however, they might also potentiate some adverse effects. We also evaluated in tumor tissue the expression of VEGF, PDGF-BB, vitamin D receptor, CYP27B1, CYP24, p53 and Bcl-2, as well as PDGF receptors: α and β. We observed the upregulation of p53 expression and the downregulation of Bcl-2, as well as VEGF in A549 tumors as a result of the tested treatment. However, vitamin D analogs did not significantly influence the expression of these proteins.

  7. Vitamin D Analogs Potentiate the Antitumor Effect of Imatinib Mesylate in a Human A549 Lung Tumor Model.

    PubMed

    Maj, Ewa; Filip-Psurska, Beata; Świtalska, Marta; Kutner, Andrzej; Wietrzyk, Joanna

    2015-01-01

    In previous papers, we presented data on studies on the anticancer activity of the vitamin D₃ analogs, named PRI-2191 and PRI-2205, in different cancer models. In this study, we showed the improved antiproliferative activity of a combination of imatinib mesylate (Gleevec, GV) and cytostatic agents in in vitro studies, when used with a third compound, namely PRI-2191, in an A549 human lung cancer model. Furthermore, we analyzed the influence of both PRI-2191, as well as PRI-2205 on the anticancer activity of GV in mice bearing A549 tumors. The route of PRI-2191 analog administration showed a significant impact on the outcome of GV treatment: subcutaneous injection was more efficient and less toxic than oral gavage. Moreover, both vitamin D compounds increased the anticancer activity of GV; however, they might also potentiate some adverse effects. We also evaluated in tumor tissue the expression of VEGF, PDGF-BB, vitamin D receptor, CYP27B1, CYP24, p53 and Bcl-2, as well as PDGF receptors: α and β. We observed the upregulation of p53 expression and the downregulation of Bcl-2, as well as VEGF in A549 tumors as a result of the tested treatment. However, vitamin D analogs did not significantly influence the expression of these proteins. PMID:26580599

  8. Modeling Leukemogenesis in the Zebrafish Using Genetic and Xenograft Models.

    PubMed

    Rajan, Vinothkumar; Dellaire, Graham; Berman, Jason N

    2016-01-01

    The zebrafish is a widely accepted model to study leukemia. The major advantage of studying leukemogenesis in zebrafish is attributed to its short life cycle and superior imaging capacity. This chapter highlights using transgenic- and xenograft-based models in zebrafish to study a specific leukemogenic mutation and analyze therapeutic responses in vivo. PMID:27464808

  9. A human fetal prostate xenograft model of developmental estrogenization

    PubMed Central

    Saffarini, Camelia M.; McDonnell-Clark, Elizabeth V.; Amin, Ali; Boekelheide, Kim

    2015-01-01

    Prostate cancer is a common disease in older men. Rodent models have demonstrated that an early and later-life exposure to estrogen can lead to cancerous lesions, and implicated hormonal dysregulation as an avenue for developing future prostate neoplasia. This study utilizes a human fetal prostate xenograft model to study the role of estrogen in the progression of human disease. Histopathological lesions were assessed in 7, 30, 90, 200, and 400-day human prostate xenografts. Gene expression for cell cycle, tumor suppressors, and apoptosis-related genes (i.e. CDKN1A, CASP9, ESR2, PTEN, and TP53) were performed for 200-day estrogen-treated xenografts. Glandular hyperplasia was observed in xenografts given both an initial and secondary exposure to estradiol in both 200 and 400-day xenografts. Persistent estrogenic effects were verified using immunohistochemical markers for cytokeratin 10, p63, and estrogen receptor-α. This model provides data on the histopathological state of the human prostate following estrogenic treatment, which can be utilized in understanding the complicated pathology associated with prostatic disease and early- and later-life estrogenic exposures. PMID:25633637

  10. Genomic profiling of patient-derived colon cancer xenograft models.

    PubMed

    Lee, Won-Suk; Kim, Hye-Youn; Seok, Jae Yeon; Jang, Ho Hee; Park, Yeon Ho; Kim, So-Young; Shin, Dong Bok; Hong, Suntaek

    2014-12-01

    Recent evidence suggests that patient derived xenograft (PDX) models can maintain certain pathological and molecular features of the original disease. However, these characterizations are limited to immunohistochemistry or by tissue microarray analysis. We conducted a high-throughput sequencing of primary colon tumor and PDX has not been reported yet. Fresh primary colon cancer tissues that originate from surgery were implanted into the subcutaneous space of 6- to 8-week-old female BALB/c nu/nu or NOD/SCID mice and serially passaged in vivo. Ion AmpliSeq Cancer Hotspot Panel v2 (Ion Torrent) was used to detect frequent somatic mutations and similarity of molecular characteristics between the 10 patient tumors and matched PDX. Histologic and immunohistochemical analyses revealed a high degree of pathologic similarity including histologic architecture and expression of CEA, CK7, and CD20 between the patient and xenograft tumors. In 80% cases, all of the somatic mutations detected in primary tumor were concordantly detected in PDX models. However, 2 PDX models showed gained mutations such as PIK3CA or FBWX7 mutation. Ten patient-derived advanced colon cancer xenograft models were established. These models maintained the key characteristic features of the original tumors, suggesting useful tool for preclinical personalized medicine platform.

  11. Prioritizing therapeutic targets using patient-derived xenograft models.

    PubMed

    Lodhia, K A; Hadley, A M; Haluska, P; Scott, C L

    2015-04-01

    Effective systemic treatment of cancer relies on the delivery of agents with optimal therapeutic potential. The molecular age of medicine has provided genomic tools that can identify a large number of potential therapeutic targets in individual patients, heralding the promise of personalized treatment. However, determining which potential targets actually drive tumor growth and should be prioritized for therapy is challenging. Indeed, reliable molecular matches of target and therapeutic agent have been stringently validated in the clinic for only a small number of targets. Patient-derived xenografts (PDXs) are tumor models developed in immunocompromised mice using tumor procured directly from the patient. As patient surrogates, PDX models represent a powerful tool for addressing individualized therapy. Challenges include humanizing the immune system of PDX models and ensuring high quality molecular annotation, in order to maximize insights for the clinic. Importantly, PDX can be sampled repeatedly and in parallel, to reveal clonal evolution, which may predict mechanisms of drug resistance and inform therapeutic strategy design.

  12. Patient-derived xenograft models in gynecologic malignancies.

    PubMed

    Scott, Clare L; Mackay, Helen J; Haluska, Paul

    2014-01-01

    In the era of targeted therapies, patients with gynecologic malignancies have not yet been major beneficiaries of this new class of agents. This may reflect the fact that the main tumor types-ovarian, uterine, and cervical--are a highly heterogeneous group of cancers with variable response to standard chemotherapies and the lack of models in which to study the diversity of these cancers. Cancer-derived cell lines fail to adequately recapitulate molecular hallmarks of specific cancer subsets and complex microenvironments, which may be critical for sensitivity to targeted therapies. Patient-derived xenografts (PDX) generated from fresh human tumor without prior in vitro culture, combined with whole genome expression, gene copy number, and sequencing analyses, could dramatically aid the development of novel therapies for gynecologic malignancies. Gynecologic tumors can be engrafted in immunodeficient mice with a high rate of success and within a reasonable time frame. The resulting PDX accurately recapitulates the patient's tumor with respect to histologic, molecular, and in vivo treatment response characteristics. Orthotopic PDX develop complications relevant to the clinic, such as ascites and bowel obstruction, providing opportunities to understand the biology of these clinical problems. Thus, PDX have great promise for improved understanding of gynecologic malignancies, serve as better models for designing novel therapies and clinical trials, and could underpin individualized, directed therapy for patients from whom such models have been established.

  13. Sp1 inhibition-mediated upregulation of VEGF 165 b induced by rh-endostatin enhances antiangiogenic and anticancer effect of rh-endostatin in A549.

    PubMed

    Li, Zhen-yu; Zhu, Fang; Hu, Jian-li; Peng, Gang; Chen, Jing; Zhang, Sheng; Chen, Xu; Zhang, Rui-guang; Chen, Ling-juan; Liu, Pian; Luo, Ming; Sun, Zhi-hua; Ren, Jing-hua; Huang, Li-li; Wu, Gang

    2011-08-01

    Recombinant human endostatin (rh-endostatin), a potential antiangiogenic agent, is used in non-small cell lung carcinoma treatment and represses vascular endothelial cell growth factor (VEGF) levels in tumor cell. However, precise affection of rh-endostatin on the proangiogenic VEGF isoforms (VEGF(165)) or antiangiogenic VEGF isoforms (VEGF(165)b) is not clear. We therefore tested the hypothesis that rh-endostatin could alter expression of these isoforms to regulate tumor growth. A549 cells were exposed to rh-endostatin, and the expression of VEGF(165) and VEGF(165)b was detected. The role of SP1 as a regulator of isoform expression was investigated. We then examined the anticancer and antiangiogenic efficacy of rh-endostatin in combination with exogenous VEGF(165)b against A549 cells, EA.HY 926 cells and xenograft model of human lung cancer. rh-Endostatin reduced VEGF(165) and induced VEGF(165)b as well as inhibited SP1 in A549 cells. SP1 inhibitor (betulinic acid) also developed those changes. VEGF(165)b-rh-endostatin combination was highly synergistic and inhibited growth, survival, and migration of A549 cells, VEGF-mediated VEGFR2 phosphorylation in EA.HY 926 cells, and tumor growth in xenograft model of human lung cancer. rh-Endostatin downregulates proangiogenic vascular endothelial growth factor A (VEGFA) isoform and upregulates antiangiogenic VEGFA isoform, possibly through inhibition of SP1. Furthermore, VEGF(165)b sensitizes A549 to rh-endostatin treatment and enhances the anticancer effect of rh-endostatin.

  14. Modeling of Chronic Myeloid Leukemia: An Overview of In Vivo Murine and Human Xenograft Models.

    PubMed

    Sontakke, Pallavi; Jaques, Jenny; Vellenga, Edo; Schuringa, Jan Jacob

    2016-01-01

    Over the past years, a wide variety of in vivo mouse models have been generated in order to unravel the molecular pathology of Chronic Myeloid Leukemia (CML) and to develop and improve therapeutic approaches. These models range from (conditional) transgenic models, knock-in models, and murine bone marrow retroviral transduction models followed by transplantation. With the advancement of immunodeficient xenograft models, it has become possible to use human stem/progenitor cells for in vivo studies as well as cells directly derived from CML patients. These models not only mimic CML but also have been instrumental in uncovering various fundamental mechanisms of CML disease progression and tyrosine kinase inhibitor (TKI) resistance. With the availability of iPSC technology, it has become feasible to derive, maintain, and expand CML subclones that are at least genetically identical to those in patients. The following review provides an overview of all murine as well as human xenograft models for CML established till date. PMID:27642303

  15. Modeling of Chronic Myeloid Leukemia: An Overview of In Vivo Murine and Human Xenograft Models

    PubMed Central

    Vellenga, Edo

    2016-01-01

    Over the past years, a wide variety of in vivo mouse models have been generated in order to unravel the molecular pathology of Chronic Myeloid Leukemia (CML) and to develop and improve therapeutic approaches. These models range from (conditional) transgenic models, knock-in models, and murine bone marrow retroviral transduction models followed by transplantation. With the advancement of immunodeficient xenograft models, it has become possible to use human stem/progenitor cells for in vivo studies as well as cells directly derived from CML patients. These models not only mimic CML but also have been instrumental in uncovering various fundamental mechanisms of CML disease progression and tyrosine kinase inhibitor (TKI) resistance. With the availability of iPSC technology, it has become feasible to derive, maintain, and expand CML subclones that are at least genetically identical to those in patients. The following review provides an overview of all murine as well as human xenograft models for CML established till date.

  16. Modeling of Chronic Myeloid Leukemia: An Overview of In Vivo Murine and Human Xenograft Models

    PubMed Central

    Vellenga, Edo

    2016-01-01

    Over the past years, a wide variety of in vivo mouse models have been generated in order to unravel the molecular pathology of Chronic Myeloid Leukemia (CML) and to develop and improve therapeutic approaches. These models range from (conditional) transgenic models, knock-in models, and murine bone marrow retroviral transduction models followed by transplantation. With the advancement of immunodeficient xenograft models, it has become possible to use human stem/progenitor cells for in vivo studies as well as cells directly derived from CML patients. These models not only mimic CML but also have been instrumental in uncovering various fundamental mechanisms of CML disease progression and tyrosine kinase inhibitor (TKI) resistance. With the availability of iPSC technology, it has become feasible to derive, maintain, and expand CML subclones that are at least genetically identical to those in patients. The following review provides an overview of all murine as well as human xenograft models for CML established till date. PMID:27642303

  17. Genetically Engineered Cancer Models, But Not Xenografts, Faithfully Predict Anticancer Drug Exposure in Melanoma Tumors

    PubMed Central

    Combest, Austin J.; Roberts, Patrick J.; Dillon, Patrick M.; Sandison, Katie; Hanna, Suzan K.; Ross, Charlene; Habibi, Sohrab; Zamboni, Beth; Müller, Markus; Brunner, Martin; Sharpless, Norman E.

    2012-01-01

    Background. Rodent studies are a vital step in the development of novel anticancer therapeutics and are used in pharmacokinetic (PK), toxicology, and efficacy studies. Traditionally, anticancer drug development has relied on xenograft implantation of human cancer cell lines in immunocompromised mice for efficacy screening of a candidate compound. The usefulness of xenograft models for efficacy testing, however, has been questioned, whereas genetically engineered mouse models (GEMMs) and orthotopic syngeneic transplants (OSTs) may offer some advantages for efficacy assessment. A critical factor influencing the predictability of rodent tumor models is drug PKs, but a comprehensive comparison of plasma and tumor PK parameters among xenograft models, OSTs, GEMMs, and human patients has not been performed. Methods. In this work, we evaluated the plasma and tumor dispositions of an antimelanoma agent, carboplatin, in patients with cutaneous melanoma compared with four different murine melanoma models (one GEMM, one human cell line xenograft, and two OSTs). Results. Using microdialysis to sample carboplatin tumor disposition, we found that OSTs and xenografts were poor predictors of drug exposure in human tumors, whereas the GEMM model exhibited PK parameters similar to those seen in human tumors. Conclusions. The tumor PKs of carboplatin in a GEMM of melanoma more closely resembles the tumor disposition in patients with melanoma than transplanted tumor models. GEMMs show promise in becoming an improved prediction model for intratumoral PKs and response in patients with solid tumors. PMID:22993143

  18. Establishment of Patient-Derived Xenograft (PDX) Models of Human Breast Cancer.

    PubMed

    Zhang, Xiaomei; Lewis, Michael T

    2013-03-01

    Patient-derived xenograft (PDX) models of human breast cancer are proving useful for preclinical evaluation of experimental therapeutics. However, until recently, generation of PDX models reflecting the full spectrum of human breast cancers has been an elusive goal. We recently developed a method for establishing serially transplantable, phenotypically stable, human breast cancer xenograft models in immunocompromised mice with comparatively high efficiency (overall ∼25%). These xenografts represent the major clinically defined subtypes of breast cancer [e.g. estrogen receptor positive (ER+), HER2 positive (HER2+), and "triple negative" (TN) breast cancers]. This method, and methods being developed in other laboratories, may soon allow for conducting "animal clinical trials" once sufficient numbers of clinically relevant models are generated. Curr. Protoc. Mouse Biol. 3:21-29 © 2013 by John Wiley & Sons, Inc.

  19. Establishment and characterization of a canine xenograft model of inflammatory mammary carcinoma.

    PubMed

    Camacho, L; Peña, L; González Gil, A; Cáceres, S; Díez, L; Illera, J C

    2013-12-01

    Canine inflammatory mammary cancer (IMC) and human inflammatory breast cancer (IBC) are the most aggressive form of mammary/breast cancer. Both species naturally develop it, sharing epidemiological, clinical and histological characteristics. Thus, IMC has been suggested as a model to study the human disease. We have developed the first IMC xenograft model in SCID mice. Xenografts reproduced the histological features from the primary tumor, were highly aggressive and showed dermal tumor emboli, distinctive hallmarks of IMC/IBC. This model was hormone receptors positive and HER2 negative. Our findings showed that estrogens and androgens are locally produced in tissues. Factors related to tumor vascularization showed positive expression and xenografts with the highest expression of all analyzed vascular factors had the highest rate of tumor proliferation. The role of steroid hormones and the angio/lymphangiogenic properties found in this model, provide additional knowledge for future interventions in the diagnosis, treatment and prevention of the disease.

  20. Adenovirus-Mediated Expression of the p14 Fusion-Associated Small Transmembrane Protein Promotes Cancer Cell Fusion and Apoptosis In Vitro but Does Not Provide Therapeutic Efficacy in a Xenograft Mouse Model of Cancer

    PubMed Central

    Wong, Carmen M.; Poulin, Kathy L.; Tong, Grace; Christou, Carin; Kennedy, Michael A.; Falls, Theresa; Bell, John C.; Parks, Robin J.

    2016-01-01

    Adenoviruses (Ads) are used in numerous preclinical and clinical studies for delivery of anti-cancer therapeutic genes. Unfortunately, Ad has a poor ability to distribute throughout a tumor mass after intratumoral injection, and infects cells primarily within the immediate area of the injection tract. Thus, Ad-encoded transgene expression is typically limited to only a small percentage of cells within the tumor. One method to increase the proportion of the tumor impacted by Ad is through expression of fusogenic proteins. Infection of a single cell with an Ad vector encoding a fusogenic protein should lead to syncytium formation with adjacent cells, effectively spreading the effect of Ad and Ad-encoded therapeutic transgenes to a greater percentage of the tumor mass. Moreover, syncytium formation can be cytotoxic, suggesting that such proteins may be effective sole therapeutics. We show that an early region 1 (E1)-deleted Ad expressing reptilian reovirus p14 fusion-associated small transmembrane (FAST) protein caused extensive cell fusion in the replication-permissive 293 cell line and at high multiplicity of infection in non-permissive human lung adenocarcinoma A549 cells in vitro. FAST protein expression in the A549 cancer cell line led to a loss of cellular metabolic activity and membrane integrity, which correlated with induction of apoptosis. However, in an A549 xenograft CD-1 nude mouse cancer model, Ad-mediated FAST gene delivery did not induce detectable cell fusion, reduce tumor burden nor enhance mouse survival compared to controls. Taken together, our results show that, although AdFAST can enhance cancer cell killing in vitro, it is not effective as a sole therapeutic in the A549 tumor model in vivo. PMID:26986751

  1. Adenovirus-Mediated Expression of the p14 Fusion-Associated Small Transmembrane Protein Promotes Cancer Cell Fusion and Apoptosis In Vitro but Does Not Provide Therapeutic Efficacy in a Xenograft Mouse Model of Cancer.

    PubMed

    Wong, Carmen M; Poulin, Kathy L; Tong, Grace; Christou, Carin; Kennedy, Michael A; Falls, Theresa; Bell, John C; Parks, Robin J

    2016-01-01

    Adenoviruses (Ads) are used in numerous preclinical and clinical studies for delivery of anti-cancer therapeutic genes. Unfortunately, Ad has a poor ability to distribute throughout a tumor mass after intratumoral injection, and infects cells primarily within the immediate area of the injection tract. Thus, Ad-encoded transgene expression is typically limited to only a small percentage of cells within the tumor. One method to increase the proportion of the tumor impacted by Ad is through expression of fusogenic proteins. Infection of a single cell with an Ad vector encoding a fusogenic protein should lead to syncytium formation with adjacent cells, effectively spreading the effect of Ad and Ad-encoded therapeutic transgenes to a greater percentage of the tumor mass. Moreover, syncytium formation can be cytotoxic, suggesting that such proteins may be effective sole therapeutics. We show that an early region 1 (E1)-deleted Ad expressing reptilian reovirus p14 fusion-associated small transmembrane (FAST) protein caused extensive cell fusion in the replication-permissive 293 cell line and at high multiplicity of infection in non-permissive human lung adenocarcinoma A549 cells in vitro. FAST protein expression in the A549 cancer cell line led to a loss of cellular metabolic activity and membrane integrity, which correlated with induction of apoptosis. However, in an A549 xenograft CD-1 nude mouse cancer model, Ad-mediated FAST gene delivery did not induce detectable cell fusion, reduce tumor burden nor enhance mouse survival compared to controls. Taken together, our results show that, although AdFAST can enhance cancer cell killing in vitro, it is not effective as a sole therapeutic in the A549 tumor model in vivo. PMID:26986751

  2. Anti-tumor effect of bevacizumab on a xenograft model of feline mammary carcinoma

    PubMed Central

    MICHISHITA, Masaki; OHTSUKA, Aya; NAKAHIRA, Rei; TAJIMA, Tsuyoshi; NAKAGAWA, Takayuki; SASAKI, Nobuo; ARAI, Toshiro; TAKAHASHI, Kimimasa

    2015-01-01

    Feline mammary carcinomas are characterized by rapid progression and metastases. Vascular endothelial growth factor (VEGF) is a key regulator of tumor angiogenesis, proliferation and metastasis. The present study aimed to investigate the effects of a single drug therapy of bevacizumab on a xenograft model of feline mammary carcinoma expressing VEGF protein. Bevacizumab treatment suppressed tumor growth by inhibiting angiogenesis and enhancing apoptosis; however, it did not affect the tumor proliferation index. Thus, bevacizumab had anti-tumor effects on a xenograft model, and this may be useful for the treatment of feline mammary carcinoma. PMID:26616000

  3. Combined Vascular Endothelial Growth Factor Receptor and Epidermal Growth Factor Receptor (EGFR) Blockade Inhibits Tumor Growth in Xenograft Models of EGFR Inhibitor Resistance

    PubMed Central

    Naumov, George N.; Nilsson, Monique B.; Cascone, Tina; Briggs, Alexandra; Straume, Oddbjorn; Akslen, Lars A.; Lifshits, Eugene; Byers, Lauren Averett; Xu, Li; Wu, Hua-kang; Jänne, Pasi; Kobayashi, Susumu; Halmos, Balazs; Tenen, Daniel; Tang, Xi M.; Engelman, Jeffrey; Yeap, Beow; Folkman, Judah; Johnson, Bruce E.; Heymach, John V.

    2010-01-01

    Purpose The epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKI) gefitinib and erlotinib benefit some non–small cell lung cancer (NSCLC) patients, but most do not respond (primary resistance) and those who initially respond eventually progress (acquired resistance). EGFR TKI resistance is not completely understood and has been associated with certain EGFR and K-RAS mutations and MET amplification. Experimental Design We hypothesized that dual inhibition of the vascular endothelial growth factor (VEGF) and EGFR pathways may overcome primary and acquired resistance. We investigated the VEGF receptor/EGFR TKI vandetanib, and the combination of bevacizumab and erlotinib in vivo using xenograft models of EGFR TKI sensitivity, primary resistance, and three models of acquired resistance, including models with mutated K-RAS and secondary EGFR T790M mutation. Results Vandetanib, gefitinib, and erlotinib had similar profiles of in vitro activity and caused sustained tumor regressions in vivo in the sensitive HCC827 model. In all four resistant models, vandetanib and bevacizumab/erlotinib were significantly more effective than erlotinib or gefitinib alone. Erlotinib resistance was associated with a rise in both host and tumor-derived VEGF but not EGFR secondary mutations in the KRAS mutant-bearing A549 xenografts. Dual inhibition reduced tumor endothelial proliferation compared with VEGF or EGFR blockade alone, suggesting that the enhanced activity of dual inhibition is due at least in part to antiendothelial effects. Conclusion These studies suggest that erlotinib resistance may be associated with a rise in both tumor cell and host stromal VEGF and that combined blockade of the VEGFR and EGFR pathways can abrogate primary or acquired resistance to EGFR TKIs. This approach merits further evaluation in NSCLC patients. PMID:19447865

  4. Human skeletal muscle xenograft as a new preclinical model for muscle disorders

    PubMed Central

    Zhang, Yuanfan; King, Oliver D.; Rahimov, Fedik; Jones, Takako I.; Ward, Christopher W.; Kerr, Jaclyn P.; Liu, Naili; Emerson, Charles P.; Kunkel, Louis M.; Partridge, Terence A.; Wagner, Kathryn R.

    2014-01-01

    Development of novel therapeutics requires good animal models of disease. Disorders for which good animal models do not exist have very few drugs in development or clinical trial. Even where there are accepted, albeit imperfect models, the leap from promising preclinical drug results to positive clinical trials commonly fails, including in disorders of skeletal muscle. The main alternative model for early drug development, tissue culture, lacks both the architecture and, usually, the metabolic fidelity of the normal tissue in vivo. Herein, we demonstrate the feasibility and validity of human to mouse xenografts as a preclinical model of myopathy. Human skeletal muscle biopsies transplanted into the anterior tibial compartment of the hindlimbs of NOD-Rag1null IL2rγnull immunodeficient host mice regenerate new vascularized and innervated myofibers from human myogenic precursor cells. The grafts exhibit contractile and calcium release behavior, characteristic of functional muscle tissue. The validity of the human graft as a model of facioscapulohumeral muscular dystrophy is demonstrated in disease biomarker studies, showing that gene expression profiles of xenografts mirror those of the fresh donor biopsies. These findings illustrate the value of a new experimental model of muscle disease, the human muscle xenograft in mice, as a feasible and valid preclinical tool to better investigate the pathogenesis of human genetic myopathies and to more accurately predict their response to novel therapeutics. PMID:24452336

  5. Biological Analysis of Human CML Stem Cells; Xenograft Model of Chronic Phase Human Chronic Myeloid Leukemia.

    PubMed

    Abraham, Sheela A

    2016-01-01

    Xenograft mouse models have been instrumental in expanding our knowledge of hematopoiesis and can provide a functional description of stem cells that possess engrafting potential. Here we describe methodology outlining one way of analyzing human malignant cells that are able to engraft immune compromised mice. Using models such as these will allow researchers to gain valuable insight into the primitive leukemic subtypes that evade current therapy regimes and are critical to understand, in order to eradicate malignancy. PMID:27581148

  6. Mouse Xenograft Model for Mesothelioma | NCI Technology Transfer Center | TTC

    Cancer.gov

    The National Cancer Institute is seeking parties interested in collaborative research to co-develop, evaluate, or commercialize a new mouse model for monoclonal antibodies and immunoconjugates that target malignant mesotheliomas. Applications of the technology include models for screening compounds as potential therapeutics for mesothelioma and for studying the pathology of mesothelioma.

  7. Establishment of an Orthotopic Xenograft Mice Model of Retinoblastoma Suitable for Preclinical Testing.

    PubMed

    Cassoux, Nathalie; Thuleau, Aurélie; Assayag, Franck; Aerts, Isabelle; Decaudin, Didier

    2015-04-01

    Retinoblastoma is a rare cancer that occurs during childhood. The goal of current and future therapeutic strategies is to conserve the eye and visual function without using external beam radiotherapy, which is known to increase the risk of secondary cancers in genetically predisposed patients. Multimodality therapy (usually intravenous but also intra-arterial and intravitreal chemotherapy, transpupillary thermotherapy, cryotherapy, or brachytherapy) has recently improved the eye salvage rate in retinoblastoma and has led to a decreased need for external beam radiotherapy. However, the treatment of advanced intraocular retinoblastoma remains a real challenge, especially in cases of vitreous and subretinal seeding. There is a need for alternative and less toxic therapies as well as for better ways to administer the drugs. Animal models are an integral part of preclinical research in the field of oncology. This paper describes the different xenograft rodent models published in the literature so far. We will also describe a new orthotopic xenografted retinoblastoma model in immunodeficient mice, which is suitable for preclinical assays. The xenograft model was established from tumor tissue obtained directly from surgical samples and closely mimics human retinoblastoma.

  8. Establishment of an Orthotopic Xenograft Mice Model of Retinoblastoma Suitable for Preclinical Testing.

    PubMed

    Cassoux, Nathalie; Thuleau, Aurélie; Assayag, Franck; Aerts, Isabelle; Decaudin, Didier

    2015-04-01

    Retinoblastoma is a rare cancer that occurs during childhood. The goal of current and future therapeutic strategies is to conserve the eye and visual function without using external beam radiotherapy, which is known to increase the risk of secondary cancers in genetically predisposed patients. Multimodality therapy (usually intravenous but also intra-arterial and intravitreal chemotherapy, transpupillary thermotherapy, cryotherapy, or brachytherapy) has recently improved the eye salvage rate in retinoblastoma and has led to a decreased need for external beam radiotherapy. However, the treatment of advanced intraocular retinoblastoma remains a real challenge, especially in cases of vitreous and subretinal seeding. There is a need for alternative and less toxic therapies as well as for better ways to administer the drugs. Animal models are an integral part of preclinical research in the field of oncology. This paper describes the different xenograft rodent models published in the literature so far. We will also describe a new orthotopic xenografted retinoblastoma model in immunodeficient mice, which is suitable for preclinical assays. The xenograft model was established from tumor tissue obtained directly from surgical samples and closely mimics human retinoblastoma. PMID:27171982

  9. Establishment of an Orthotopic Xenograft Mice Model of Retinoblastoma Suitable for Preclinical Testing

    PubMed Central

    Cassoux, Nathalie; Thuleau, Aurélie; Assayag, Franck; Aerts, Isabelle; Decaudin, Didier

    2015-01-01

    Retinoblastoma is a rare cancer that occurs during childhood. The goal of current and future therapeutic strategies is to conserve the eye and visual function without using external beam radiotherapy, which is known to increase the risk of secondary cancers in genetically predisposed patients. Multimodality therapy (usually intravenous but also intra-arterial and intravitreal chemotherapy, transpupillary thermotherapy, cryotherapy, or brachytherapy) has recently improved the eye salvage rate in retinoblastoma and has led to a decreased need for external beam radiotherapy. However, the treatment of advanced intraocular retinoblastoma remains a real challenge, especially in cases of vitreous and subretinal seeding. There is a need for alternative and less toxic therapies as well as for better ways to administer the drugs. Animal models are an integral part of preclinical research in the field of oncology. This paper describes the different xenograft rodent models published in the literature so far. We will also describe a new orthotopic xenografted retinoblastoma model in immunodeficient mice, which is suitable for preclinical assays. The xenograft model was established from tumor tissue obtained directly from surgical samples and closely mimics human retinoblastoma. PMID:27171982

  10. Transplantation of Tissue-Engineered Cartilage in an Animal Model (Xenograft and Autograft): Construct Validation.

    PubMed

    Nemoto, Hitoshi; Watson, Deborah; Masuda, Koichi

    2015-01-01

    Tissue engineering holds great promise for cartilage repair with minimal donor-site morbidity. The in vivo maturation of a tissue-engineered construct can be tested in the subcutaneous tissues of the same species for autografts or of immunocompromised animals for allografts or xenografts. This section describes detailed protocols for the surgical transplantation of a tissue-engineered construct into an animal model to assess construct validity.

  11. Suppression of tumor growth in lung cancer xenograft model mice by poly(sorbitol-co-PEI)-mediated delivery of osteopontin siRNA.

    PubMed

    Cho, Won-Young; Hong, Seong-Ho; Singh, Bijay; Islam, Mohammad Ariful; Lee, Somin; Lee, Ah Young; Gankhuyag, Nomundelger; Kim, Ji-Eun; Yu, Kyeong-Nam; Kim, Kwang-Ho; Park, Young-Chan; Cho, Chong-Su; Cho, Myung-Haing

    2015-08-01

    Small interfering RNA (siRNA)-mediated gene silencing represents a promising strategy for treating diseases such as cancer; however, specific gene silencing requires an effective delivery system to overcome the instability and low transfection efficiency of siRNAs. To address this issue, a polysorbitol-based transporter (PSOT) was prepared by low molecular weight branched polyethylenimine (bPEI) crosslinked with sorbitol diacrylate (SDA). Osteopontin (OPN) gene, which is highly associated with non-small cell lung cancer (NSCLC) was targeted by siRNA therapy using siRNA targeting OPN (siOPN). Characterization study confirmed that PSOT formed compact complexes with siOPN and protected siOPN against degradation by RNase. PSOT/siOPN complexes demonstrated low cytotoxicity and enhanced transfection efficiency in vitro, suggesting that this carrier may be suitable for gene silencing. In the A549 and H460 lung cancer cell lines, PSOT/siOPN complexes demonstrated significant silencing efficiency at both RNA and protein levels. To study in vivo tumor growth suppression, two lung cancer cell-xenograft mouse models were prepared and PSOT/siOPN complexes were delivered into the mice through intravenous injection. The siOPN-treated groups demonstrated significantly reduced OPN expression at both the RNA and protein levels as well as suppression of tumor volume and weight. Taken together, siOPN delivery using PSOT may present an effective and novel therapeutic system for lung cancer treatment.

  12. Patient-Derived Tumor Xenograft Models of Breast Cancer.

    PubMed

    Suarez, Christopher D; Littlepage, Laurie E

    2016-01-01

    The need for model systems that more accurately predict patient outcome has led to a renewed interest and a rapid development of orthotopic transplantation models designed to grow, expand, and study patient-derived human breast tumor tissue in mice. After implanting a human breast tumor piece into a mouse mammary fat pad and allowing the tumor to grow in vivo, the tumor tissue can be either harvested and immediately implanted into mice or can be stored as tissue pieces in liquid nitrogen for surgical implantation at a later time. Here, we describe the process of surgically implanting patient-derived breast tumor tissue into the mammary gland of nonobese diabetic-severe combined immunodeficiency (NOD-SCID) mice and harvesting tumor tissue for long-term storage in liquid nitrogen.

  13. Primary esophageal and gastro-esophageal junction cancer xenograft models: clinicopathological features and engraftment.

    PubMed

    Dodbiba, Lorin; Teichman, Jennifer; Fleet, Andrew; Thai, Henry; Sun, Bin; Panchal, Devang; Patel, Devalben; Tse, Alvina; Chen, Zhuo; Faluyi, Olusola O; Renouf, Daniel J; Girgis, Hala; Bandarchi, Bizhan; Schwock, Joerg; Xu, Wei; Bristow, Robert G; Tsao, Ming-Sound; Darling, Gail E; Ailles, Laurie E; El-Zimaity, Hala; Liu, Geoffrey

    2013-04-01

    There are very few xenograft models available for the study of esophageal (E) and gastro-esophageal junction (GEJ) cancer. Using a NOD/SCID model, we implanted 90 primary E and GEJ tumors resected from patients and six endoscopic biopsy specimens. Of 69 resected tumors with histologically confirmed viable adenocarcinoma or squamous cell carcinoma, 22 (32%) was engrafted. One of 11 tumors, considered to have had a complete pathological response to neo-adjuvant chemo-radiation, also engrafted. Of the 23 patients whose tumors were engrafted, 65% were male; 30% were early stage while 70% were late stage; 22% received neo-adjuvant chemo-radiation; 61% were GEJ cancers. Engraftment occurred in 18/54 (33%) adenocarcinomas and 5/16 (31%) squamous cell carcinomas. Small endoscopic biopsy tissue had a 50% (3/6) engraftment rate. Of the factors analyzed, pretreatment with chemo-radiation and well/moderate differentiation showed significantly lower correlation with engraftment (P<0.05). In the subset of patients who did not receive neo-adjuvant chemo-radiation, 18/41 (44%) engrafted compared with those with pretreatment where 5/29 (17%, P=0.02) engrafted. Primary xenograft lines may be continued through 4-12 passages. Xenografts maintained similar histology and morphological characteristics with only minor variations even after multiple passaging in most instances.

  14. Berberine inhibits human tongue squamous carcinoma cancer tumor growth in a murine xenograft model.

    PubMed

    Ho, Yung-Tsuan; Yang, Jai-Sing; Lu, Chi-Cheng; Chiang, Jo-Hua; Li, Tsai-Chung; Lin, Jen-Jyh; Lai, Kuang-Chi; Liao, Ching-Lung; Lin, Jaung-Geng; Chung, Jing-Gung

    2009-09-01

    Our primary studies showed that berberine induced apoptosis in human tongue cancer SCC-4 cells in vitro. But there is no report to show berberine inhibited SCC-4 cancer cells in vivo on a murine xenograft animal model. SCC-4 tumor cells were implanted into mice and groups of mice were treated with vehicle, berberine (10mg/kg of body weight) and doxorubicin (4mg/kg of body weight). The tested agents were injected once per four days intraperitoneally (i.p.), with treatment starting 4 weeks prior to cells inoculation. Treatment with 4mg/kg of doxorubicin or with 10mg/kg of berberine resulted in a reduction in tumor incidence. Tumor size in xenograft mice treated with 10mg/kg berberine was significantly smaller than that in the control group. Our findings indicated that berbeirne inhibits tumor growth in a xenograft animal model. Therefore, berberine may represent a tongue cancer preventive agent and can be used in clinic. PMID:19303753

  15. Patient Derived Xenograft Models: An Emerging Platform for Translational Cancer Research

    PubMed Central

    Hidalgo, Manuel; Amant, Frederic; Biankin, Andrew V.; Budinská, Eva; Byrne, Annette T.; Caldas, Carlos; Clarke, Robert B.; de Jong, Steven; Jonkers, Jos; Mælandsmo, Gunhild Mari; Roman-Roman, Sergio; Seoane, Joan; Trusolino, Livio; Villanueva, Alberto

    2014-01-01

    Recently, there has been increasing interest in the development and characterization of patient derived tumor xenograft (PDX) models for cancer research. PDX models mostly retain the principal histological and genetic characteristics of their donor tumor and remain stable across passages. These models have been shown to be predictive of clinical outcomes and are being used for preclinical drug evaluation, biomarker identification, biological studies, and personalized medicine strategies. This paper summarizes the current state of the art in this field including methodological issues, available collections, practical applications, challenges and shortcoming, and future directions, and introduces a European consortium of PDX models. PMID:25185190

  16. The anti-Fn14 antibody BIIB036 inhibits tumor growth in xenografts and patient derived primary tumor models and enhances efficacy of chemotherapeutic agents in multiple xenograft models.

    PubMed

    Michaelson, Jennifer S; Kelly, Rebecca; Yang, Lu; Zhang, Xiamei; Wortham, Kathleen; Joseph, Ingrid B J K

    2012-07-01

    Agonistic antibodies targeting Fn14, the receptor for TWEAK, have demonstrated anti-tumor activity in xenograft models. Herein, we further explore the therapeutic potential of the humanized anti-Fn14 agonistic antibody, BIIB036, as a single agent and in combination with standard of care cancer therapeutics. Pharmacokinetic studies of BIIB036 in tumor-bearing mice revealed a half-life of approximately three days suggesting twice a week dosing would be necessary to maintain efficacy. However, in multiple xenograft models, BIIB036 treatment resulted in extended tumor growth inhibition up to 40-50 d following cessation of dosing, suggesting that frequent administration of BIIB036 may not be necessary to maintain prolonged anti-tumor activity. Subsequent xenograft studies revealed that maximal efficacy was achieved with BIIB036 dosing once every two weeks, by either intraperitoneal or subcutaneous administration. Xenograft tumors that were initially treated with BIBI036 and then re-grew up to 1000 mm³ following cessation of the first cycle of treatment remained sensitive to a second cycle of treatment. BIIB036 was also evaluated in patient derived primary colon tumor models, where efficacy compared favorably with a standard of care agent. Lastly, BIIB036 enhanced the efficacy of several standard of care chemotherapeutics, including paclitaxel in MDA-MBA-231 breast tumor xenografts, paclitaxel or carboplatin in HOP62 non-small cell lung xenografts, and 5-FU in NCI-N87 gastric xenografts, with no overlapping toxicities. These studies thus establish BIIB036 as a promising therapeutic agent with durable anti-tumor activity in human xenografts as well as patient derived primary tumor models, and enhanced activity and tolerability in combination with standard of care chemotherapeutics. Taken together, the data presented herein suggest that BIIB036 warrants evaluation in the clinic.

  17. Patient-derived xenograft models of squamous cell carcinoma of the uterine cervix.

    PubMed

    Rofstad, Einar K; Simonsen, Trude G; Huang, Ruixia; Andersen, Lise Mari K; Galappathi, Kanthi; Ellingsen, Christine; Wegner, Catherine S; Hauge, Anette; Gaustad, Jon-Vidar

    2016-04-10

    Patient-derived xenograft (PDX) models of cancer are considered to reflect the biology and treatment response of human tumors to a larger extent than xenograft models initiated from established cell lines. The characterization of a panel of four novel PDX models of cervical carcinoma of the uterine cervix is described in this communication. The outcome of treatment differed substantially among the donor patients, and the PDX models were found to mirror the histology, aggressiveness, and metastatic propensity of the donor patients' tumors. Two of the models (BK-12 and LA-19) were highly metastatic, one model (ED-15) was poorly metastatic, and one model (HL-16) was non-metastatic. The primary tumors of the two highly metastatic models showed high density of intratumoral lymphatics, whereas the other two models did not develop intratumoral lymphatics. The potential of the models to metastasize to lymph nodes was associated with high expression of both angiogenesis-related genes and cancer stem cell-related genes. The models may be highly valuable for studying mechanisms linking lymph node metastasis to lymphangiogenesis, hemangiogenesis, and the presence of cancer stem cells. PMID:26828134

  18. Patient-derived xenograft models of squamous cell carcinoma of the uterine cervix.

    PubMed

    Rofstad, Einar K; Simonsen, Trude G; Huang, Ruixia; Andersen, Lise Mari K; Galappathi, Kanthi; Ellingsen, Christine; Wegner, Catherine S; Hauge, Anette; Gaustad, Jon-Vidar

    2016-04-10

    Patient-derived xenograft (PDX) models of cancer are considered to reflect the biology and treatment response of human tumors to a larger extent than xenograft models initiated from established cell lines. The characterization of a panel of four novel PDX models of cervical carcinoma of the uterine cervix is described in this communication. The outcome of treatment differed substantially among the donor patients, and the PDX models were found to mirror the histology, aggressiveness, and metastatic propensity of the donor patients' tumors. Two of the models (BK-12 and LA-19) were highly metastatic, one model (ED-15) was poorly metastatic, and one model (HL-16) was non-metastatic. The primary tumors of the two highly metastatic models showed high density of intratumoral lymphatics, whereas the other two models did not develop intratumoral lymphatics. The potential of the models to metastasize to lymph nodes was associated with high expression of both angiogenesis-related genes and cancer stem cell-related genes. The models may be highly valuable for studying mechanisms linking lymph node metastasis to lymphangiogenesis, hemangiogenesis, and the presence of cancer stem cells.

  19. Ophiopogonin B induces apoptosis, mitotic catastrophe and autophagy in A549 cells.

    PubMed

    Chen, Meijuan; Guo, Yuanyuan; Zhao, Ruolin; Wang, Xiaoxia; Jiang, Miao; Fu, Haian; Zhang, Xu

    2016-07-01

    Ophiopogonin B (OP-B), a saponin compound isolated from Radix Ophiopogon japonicus, was verified to inhibit cell proliferation in numerous non-small cell lung cancer (NSCLC) cells in our previous study. However, the precise mechanisms of action have remained unclear. In the present study, we mainly investigated the effects of OP-B on adenocarcinoma A549 cells to further elaborate the underlying mechanisms of OP-B in different NSCLC cell lines. Detection by high content screening (HCS) and TUNEL assay verified that OP-B induced apoptosis in this cell line, while detection of Caspase-3, Bcl-2 and Bax showed that OP-B induced cell death was caspase and mitochondrial independent. Further experiments showed that OP-B induced cell cycle arrest in the S and G2/M phases by inhibiting the expression of Myt1 and phosphorylation of Histone H3 (Ser10), which resulted in mitotic catastrophe in the cells. Transmission electron microscopy (TEM) observation of cell micro-morphology combined with detection of Atgs by western blot analysis showed that OP-B induced autophagy in this cell line. Autophagy inhibition by the lysosome inhibitor CQ or Beclin1-siRNA knockdown both attenuated cell viability, demonstrated that autophagy also being the vital reason resulted in cell death. More importantly, the xenograft model using A549 cells provided further evidence of the inhibition of OP-B on tumor proliferation. Immunohistochemistry detection of LC3 and Tunel assay both verified that high dose of OP-B (75 mg/kg) induced autophagy and apoptosis in vivo, and western blot detection of p-Histone H3 (Ser10), Survivin and XIAP further indicated the molecular mechanism of OP-B in vivo. As our findings revealed, multiple types of cell death overlapped in OP-B treated A549 cells, it displayed multitarget characteristics of the compounds extracted from the Chinese herbal, which may be used as candidate anticancer medicine in clinic.

  20. Generation of Prostate Cancer Patient Derived Xenograft Models from Circulating Tumor Cells.

    PubMed

    Williams, Estrelania S; Rodriguez-Bravo, Veronica; Rodriquez-Bravo, Veronica; Chippada-Venkata, Uma; De Ia Iglesia-Vicente, Janis; Gong, Yixuan; Galsky, Matthew; Oh, William; Cordon-Cardo, Carlos; Domingo-Domenech, Josep

    2015-10-20

    Patient derived xenograft (PDX) models are gaining popularity in cancer research and are used for preclinical drug evaluation, biomarker identification, biologic studies, and personalized medicine strategies. Circulating tumor cells (CTC) play a critical role in tumor metastasis and have been isolated from patients with several tumor types. Recently, CTCs have been used to generate PDX experimental models of breast and prostate cancer. This manuscript details the method for the generation of prostate cancer PDX models from CTCs developed by our group. Advantages of this method over conventional PDX models include independence from surgical sample collection and generating experimental models at various disease stages. Density gradient centrifugation followed by red blood cell lysis and flow cytometry depletion of CD45 positive mononuclear cells is used to enrich CTCs from peripheral blood samples collected from patients with metastatic disease. The CTCs are then injected into immunocompromised mice; subsequently generated xenografts can be used for functional studies or harvested for molecular characterization. The primary limitation of this method is the negative selection method used for CTC enrichment. Despite this limitation, the generation of PDX models from CTCs provides a novel experimental model to be applied to prostate cancer research.

  1. The chick chorioallantoic membrane as an in vivo xenograft model for Burkitt lymphoma

    PubMed Central

    2014-01-01

    Background Burkitt lymphoma (BL) is an aggressive malignancy that arises from B-cells and belongs to the group of Non-Hodgkin lymphomas (NHL). Due to the lack of appropriate in vivo models NHL research is mainly performed in vitro. Here, we studied the use of the chick chorioallantoic membrane (CAM) for the generation of human BL xenograft tumors, which we compared with known characteristics of the human disease. Methods In order to generate experimental BL tumors, we inoculated human BL2B95 and BL2-GFP cells on the CAM. BL2B95 xenograft-tumors were grown for seven days and subsequently analyzed with transmission electron and immunofluorescence microscopy, as well as histological staining approaches. BL2-GFP cells were studied at regular intervals up to seven days, and their metastatic behavior was visualized with intravital immunofluorescence techniques. Results Xenografted BL2B95 cells formed solid tumors in the CAM model with a Ki67-index greater than 90%, preservation of typical tumor markers (CD10, CD19, CD20), a ‘starry sky’ morphology, production of agyrophilic fibers in the stroma, formation of blood and lymphatic vessels and lymphogenic dissemination of BL2B95 to distant sites. We identified macrophages, lymphocytes and heterophilic granulocytes (chick homolog of neutrophils) as the most abundant immune cells in the experimental tumors. BL2-GFP cells could be traced in real-time during their distribution in the CAM, and the first signs for their dissemination were visible after 2-3 days. Conclusions We show that xenografted BL2B95 cells generate tumors in the CAM with a high degree of cellular, molecular and proliferative concord with the human disease, supporting the application of the CAM model for NHL research with a focus on tumor-stroma interactions. Additionally we report that BL2-GFP cells, grafted on the CAM of ex ovo cultured chick embryos, provide a powerful tool to study lymphogenic dissemination in real-time. PMID:24884418

  2. Development and rescue of human familial hypercholesterolaemia in a xenograft mouse model

    PubMed Central

    Bissig-Choisat, Beatrice; Wang, Lili; Legras, Xavier; Saha, Pradip K.; Chen, Leon; Bell, Peter; Pankowicz, Francis P.; Hill, Matthew C.; Barzi, Mercedes; Leyton, Claudia Kettlun; Leung, Hon-Chiu Eastwood; Kruse, Robert L.; Himes, Ryan W.; Goss, John A.; Wilson, James M.; Chan, Lawrence; Lagor, William R.; Bissig, Karl-Dimiter

    2015-01-01

    Diseases of lipid metabolism are a major cause of human morbidity, but no animal model entirely recapitulates human lipoprotein metabolism. Here we develop a xenograft mouse model using hepatocytes from a patient with familial hypercholesterolaemia caused by loss-of-function mutations in the low-density lipoprotein receptor (LDLR). Like familial hypercholesterolaemia patients, our familial hypercholesterolaemia liver chimeric mice develop hypercholesterolaemia and a 'humanized‘ serum profile, including expression of the emerging drug targets cholesteryl ester transfer protein and apolipoprotein (a), for which no genes exist in mice. We go on to replace the missing LDLR in familial hypercholesterolaemia liver chimeric mice using an adeno-associated virus 9-based gene therapy and restore normal lipoprotein profiles after administration of a single dose. Our study marks the first time a human metabolic disease is induced in an experimental animal model by human hepatocyte transplantation and treated by gene therapy. Such xenograft platforms offer the ability to validate human experimental therapies and may foster their rapid translation into the clinic. PMID:26081744

  3. Pathologic Correlates of Primary Central Nervous System Lymphoma Defined in an Orthotopic Xenograft Model

    PubMed Central

    Kadoch, Cigall; Dinca, Eduard B.; Voicu, Ramona; Chen, Lingjing; Nguyen, Diana; Parikh, Seema; Karrim, Juliana; Shuman, Marc A.; Lowell, Clifford A.; Treseler, Patrick A.; James, C. David; Rubenstein, James L.

    2014-01-01

    Purpose The prospect for advances in the treatment of patients with primary central nervous system lymphoma (PCNSL) is likely dependent on the systematic evaluation of its pathobiology. Animal models of PCNSL are needed to facilitate the analysis of its molecular pathogenesis and for the efficient evaluation of novel therapeutics. Experimental Design We characterized the molecular pathology of CNS lymphoma tumors generated by the intracerebral implantation of Raji B lymphoma cells in athymic mice. Lymphoma cells were modified for bioluminescence imaging to facilitate monitoring of tumor growth and response to therapy. In parallel, we identified molecular features of lymphoma xenograft histopathology that are evident in human PCNSL specimens. Results Intracerebral Raji tumors were determined to faithfully reflect the molecular pathogenesis of PCNSL, including the predominant immunophenotypic state of differentiation of lymphoma cells and their reactive microenvironment. We show the expression of interleukin-4 by Raji and other B lymphoma cell lines in vitro and by Raji tumors in vivo and provide evidence for a role of this cytokine in the M2 polarization of lymphoma macrophages both in the murine model and in diagnostic specimens of human PCNSL. Conclusion Intracerebral implantation of Raji cells results in a reproducible and invasive xenograft model, which recapitulates the histopathology and molecular features of PCNSL, and is suitable for preclinical testing of novel agents. We also show for the first time the feasibility and accuracy of tumor bioluminescence in the monitoring of a highly infiltrative brain tumor. PMID:19276270

  4. Molecular characterization of patient-derived human pancreatic tumor xenograft models for preclinical and translational development of cancer therapeutics.

    PubMed

    Mattie, Mike; Christensen, Ashley; Chang, Mi Sook; Yeh, William; Said, Suzanne; Shostak, Yuriy; Capo, Linnette; Verlinsky, Alla; An, Zili; Joseph, Ingrid; Zhang, Yi; Kumar-Ganesan, Sathish; Morrison, Karen; Stover, David; Challita-Eid, Pia

    2013-10-01

    Preclinical evaluation of novel cancer agents requires models that accurately reflect the biology and molecular characteristics of human tumors. Molecular profiles of eight pancreatic ductal adenocarcinoma patient tumors were compared to corresponding passages of xenografts obtained by grafting tumor fragments into immunocompromised mice. Molecular characterization was performed by copy number analysis, gene expression and microRNA microarrays, mutation analysis, short tandem repeat (STR) profiling, and immunohistochemistry. Xenografts were found to be highly representative of their respective tumors, with a high degree of genetic stability observed by STR profiling and mutation analysis. Copy number variation (CNV) profiles of early and late xenograft passages were similar, with recurrent losses on chromosomes 1p, 3p, 4q, 6, 8p, 9, 10, 11q, 12p, 15q, 17, 18, 20p, and 21 and gains on 1q, 5p, 8q, 11q, 12q, 13q, 19q, and 20q. Pearson correlations of gene expression profiles of tumors and xenograft passages were above 0.88 for all models. Gene expression patterns between early and late passage xenografts were highly stable for each individual model. Changes observed in xenograft passages largely corresponded to human stromal compartment genes and inflammatory processes. While some differences exist between the primary tumors and corresponding xenografts, the molecular profiles remain stable after extensive passaging. Evidence for stability in molecular characteristics after several rounds of passaging lends confidence to clinical relevance and allows for expansion of models to generate the requisite number of animals required for cohorts used in drug screening and development studies.

  5. Developmental Exposure to Estrogen Alters Differentiation and Epigenetic Programming in a Human Fetal Prostate Xenograft Model

    PubMed Central

    Saffarini, Camelia M.; McDonnell-Clark, Elizabeth V.; Amin, Ali; Huse, Susan M.; Boekelheide, Kim

    2015-01-01

    Prostate cancer is the most frequent non-cutaneous malignancy in men. There is strong evidence in rodents that neonatal estrogen exposure plays a role in the development of this disease. However, there is little information regarding the effects of estrogen in human fetal prostate tissue. This study explored early life estrogen exposure, with and without a secondary estrogen and testosterone treatment in a human fetal prostate xenograft model. Histopathological lesions, proliferation, and serum hormone levels were evaluated at 7, 30, 90, and 200-day time-points after xenografting. The expression of 40 key genes involved in prostatic glandular and stromal growth, cell-cycle progression, apoptosis, hormone receptors and tumor suppressors was evaluated using a custom PCR array. Epigenome-wide analysis of DNA methylation was performed on whole tissue, and laser capture-microdissection (LCM) isolated epithelial and stromal compartments of 200-day prostate xenografts. Combined initial plus secondary estrogenic exposures had the most severe tissue changes as revealed by the presence of hyperplastic glands at day 200. Gene expression changes corresponded with the cellular events in the KEGG prostate cancer pathway, indicating that initial plus secondary exposure to estrogen altered the PI3K-Akt signaling pathway, ultimately resulting in apoptosis inhibition and an increase in cell cycle progression. DNA methylation revealed that differentially methylated CpG sites significantly predominate in the stromal compartment as a result of estrogen-treatment, thereby providing new targets for future investigation. By using human fetal prostate tissue and eliminating the need for species extrapolation, this study provides novel insights into the gene expression and epigenetic effects related to prostate carcinogenesis following early life estrogen exposure. PMID:25799167

  6. The Zebrafish Xenograft Platform: Evolution of a Novel Cancer Model and Preclinical Screening Tool.

    PubMed

    Wertman, Jaime; Veinotte, Chansey J; Dellaire, Graham; Berman, Jason N

    2016-01-01

    Animal xenografts of human cancers represent a key preclinical tool in the field of cancer research. While mouse xenografts have long been the gold standard, investigators have begun to use zebrafish (Danio rerio) xenotransplantation as a relatively rapid, robust and cost-effective in vivo model of human cancers. There are several important methodological considerations in the design of an informative and efficient zebrafish xenotransplantation experiment. Various transgenic fish strains have been created that facilitate microscopic observation, ranging from the completely transparent casper fish to the Tg(fli1:eGFP) fish that expresses fluorescent GFP protein in its vascular tissue. While human cancer cell lines have been used extensively in zebrafish xenotransplantation studies, several reports have also used primary patient samples as the donor material. The zebrafish is ideally suited for transplanting primary patient material by virtue of the relatively low number of cells required for each embryo (between 50 and 300 cells), the absence of an adaptive immune system in the early zebrafish embryo, and the short experimental timeframe (5-7 days). Following xenotransplantation into the fish, cells can be tracked using in vivo or ex vivo measures of cell proliferation and migration, facilitated by fluorescence or human-specific protein expression. Importantly, assays have been developed that allow for the reliable detection of in vivo human cancer cell growth or inhibition following administration of drugs of interest. The zebrafish xenotransplantation model is a unique and effective tool for the study of cancer cell biology.

  7. Modulating testicular mass in xenografting: a model to explore testis development and endocrine function.

    PubMed

    Schlatt, Stefan; Gassei, Kathrin; Westernströer, Birgit; Ehmcke, Jens

    2010-08-01

    The hypothalamic-pituitary-gonadal (HPG) axis is involved in both the regulation of growth of the developing testis and in controlling spermatogenic and steroidogenic activity in the adult testis. Here, we develop a novel testicular xenografting model to examine to which degree testicular growth and function are controlled by intra- and extratesticular factors. Two or eight halves of neonatal Djungarian hamster testes were implanted into intact, hemicastrated, or castrated nude mouse recipients, and the development of the grafts under reduced or increased competition of testicular tissue was monitored and analyzed. We hypothesized that the outgrowth of the testicular grafts is influenced by the total amount of testicular tissue present in a host and that less testicular tissue in a host would result in more extended outgrowth of the grafts. Our results reveal that the hypothesis is wrong, because implanted hamster testis tissue irrespectively of the grafting condition grows to a similar size revealing an intrinsic mechanism for testicular growth. In contrast, similar size of seminal vesicle as bio-indicator of androgen levels in all hosts revealed that the steroidogenic activity is independent from the mass of testicular tissue and that steroid levels are extrinsically regulated by the recipient's HPG axis. We propose that the model of testicular xenografting provides highly valuable options to explore testicular growth and endocrine regulation of the HPG axis.

  8. The Zebrafish Xenograft Platform: Evolution of a Novel Cancer Model and Preclinical Screening Tool.

    PubMed

    Wertman, Jaime; Veinotte, Chansey J; Dellaire, Graham; Berman, Jason N

    2016-01-01

    Animal xenografts of human cancers represent a key preclinical tool in the field of cancer research. While mouse xenografts have long been the gold standard, investigators have begun to use zebrafish (Danio rerio) xenotransplantation as a relatively rapid, robust and cost-effective in vivo model of human cancers. There are several important methodological considerations in the design of an informative and efficient zebrafish xenotransplantation experiment. Various transgenic fish strains have been created that facilitate microscopic observation, ranging from the completely transparent casper fish to the Tg(fli1:eGFP) fish that expresses fluorescent GFP protein in its vascular tissue. While human cancer cell lines have been used extensively in zebrafish xenotransplantation studies, several reports have also used primary patient samples as the donor material. The zebrafish is ideally suited for transplanting primary patient material by virtue of the relatively low number of cells required for each embryo (between 50 and 300 cells), the absence of an adaptive immune system in the early zebrafish embryo, and the short experimental timeframe (5-7 days). Following xenotransplantation into the fish, cells can be tracked using in vivo or ex vivo measures of cell proliferation and migration, facilitated by fluorescence or human-specific protein expression. Importantly, assays have been developed that allow for the reliable detection of in vivo human cancer cell growth or inhibition following administration of drugs of interest. The zebrafish xenotransplantation model is a unique and effective tool for the study of cancer cell biology. PMID:27165359

  9. Small-sample inference for incomplete longitudinal data with truncation and censoring in tumor xenograft models.

    PubMed

    Tan, Ming; Fang, Hong-Bin; Tian, Guo-Liang; Houghton, Peter J

    2002-09-01

    In cancer drug development, demonstrating activity in xenograft models, where mice are grafted with human cancer cells, is an important step in bringing a promising compound to humans. A key outcome variable is the tumor volume measured in a given period of time for groups of mice given different doses of a single or combination anticancer regimen. However, a mouse may die before the end of a study or may be sacrificed when its tumor volume quadruples, and its tumor may be suppressed for some time and then grow back. Thus, incomplete repeated measurements arise. The incompleteness or missingness is also caused by drastic tumor shrinkage (<0.01 cm3) or random truncation. Because of the small sample sizes in these models, asymptotic inferences are usually not appropriate. We propose two parametric test procedures based on the EM algorithm and the Bayesian method to compare treatment effects among different groups while accounting for informative censoring. A real xenograft study on a new antitumor agent, temozolomide, combined with irinotecan is analyzed using the proposed methods.

  10. Hypoxia is not required for human endometrial breakdown or repair in a xenograft model of menstruation.

    PubMed

    Coudyzer, Pauline; Lemoine, Pascale; Jordan, Bénédicte F; Gallez, Bernard; Galant, Christine; Nisolle, Michelle; Courtoy, Pierre J; Henriet, Patrick; Marbaix, Etienne

    2013-09-01

    Menstrual endometrial breakdown induced by estradiol and progesterone withdrawal is regularly attributed to vasospasm of spiral arteries causing ischemia and hypoxia. We investigated whether hypoxia actually occurred in an in vivo model of menstruation. Three complementary approaches were used to look for signs of hypoxia in fragments of human functionalis xenografted to ovariectomized immunodeficient mice bearing pellets-releasing estradiol and progesterone, and then deprived of ovarian steroids. Hormone withdrawal 21 d after grafting induced menstrual breakdown and MMP expression within 4 d. Local partial oxygen pressure (pO2) was measured by electron paramagnetic resonance using implanted lithium phtalocyanine crystals. In mice with hormone maintenance until sacrifice, pO2 was low one week after grafting (14.8±3.4 mmHg) but increased twofold from the second week when tissue was largely revascularized. After 3 wk, pO2 was not modified by hormone withdrawal but was slightly increased on hormone reimpregnation 4 d after removal (34.7±6.1 mmHg) by comparison with hormone maintenance (27.1±8.6 mmHg). These results were confirmed using fluorescence quenching-based OxyLite measurements. In a further search for signs of hypoxia, we did not find significant HIF1-α immunostaining, nor pimonidazole adducts after hormone withdrawal. We conclude that hypoxia is not needed to trigger menstrual-like tissue breakdown or repair in human endometrial xenograft.

  11. Prolonged exposure to acetaminophen reduces testosterone production by the human fetal testis in a xenograft model.

    PubMed

    van den Driesche, Sander; Macdonald, Joni; Anderson, Richard A; Johnston, Zoe C; Chetty, Tarini; Smith, Lee B; McKinnell, Chris; Dean, Afshan; Homer, Natalie Z; Jorgensen, Anne; Camacho-Moll, Maria E; Sharpe, Richard M; Mitchell, Rod T

    2015-05-20

    Most common male reproductive disorders are linked to lower testosterone exposure in fetal life, although the factors responsible for suppressing fetal testosterone remain largely unknown. Protracted use of acetaminophen during pregnancy is associated with increased risk of cryptorchidism in sons, but effects on fetal testosterone production have not been demonstrated. We used a validated xenograft model to expose human fetal testes to clinically relevant doses and regimens of acetaminophen. Exposure to a therapeutic dose of acetaminophen for 7 days significantly reduced plasma testosterone (45% reduction; P = 0.025) and seminal vesicle weight (a biomarker of androgen exposure; 18% reduction; P = 0.005) in castrate host mice bearing human fetal testis xenografts, whereas acetaminophen exposure for just 1 day did not alter either parameter. Plasma acetaminophen concentrations (at 1 hour after the final dose) in exposed host mice were substantially below those reported in humans after a therapeutic oral dose. Subsequent in utero exposure studies in rats indicated that the acetaminophen-induced reduction in testosterone likely results from reduced expression of key steroidogenic enzymes (Cyp11a1, Cyp17a1). Our results suggest that protracted use of acetaminophen (1 week) may suppress fetal testosterone production, which could have adverse consequences. Further studies are required to establish the dose-response and treatment-duration relationships to delineate the maximum dose and treatment period without this adverse effect.

  12. Development of a Patient-Derived Xenograft Model Using Brain Tumor Stem Cell Systems to Study Cancer.

    PubMed

    Chokshi, Chirayu; Dhillon, Manvir; McFarlane, Nicole; Venugopal, Chitra; Singh, Sheila K

    2016-01-01

    Patient-derived xenograft (PDX) models provide an excellent platform to understand cancer initiation and development in vivo. In the context of brain tumor initiating cells (BTICs), PDX models allow for characterization of tumor formation, growth, and recurrence, in a clinically relevant in vivo system. Here, we detail procedures to harvest, culture, characterize, and orthotopically inject human BTICs derived from patient samples.

  13. Genomic characterization of a large panel of patient-derived hepatocellular carcinoma xenograft tumor models for preclinical development.

    PubMed

    Gu, Qingyang; Zhang, Bin; Sun, Hongye; Xu, Qiang; Tan, Yexiong; Wang, Guan; Luo, Qin; Xu, Weiguo; Yang, Shuqun; Li, Jian; Fu, Jing; Chen, Lei; Yuan, Shengxian; Liang, Guibai; Ji, Qunsheng; Chen, Shu-Hui; Chan, Chi-Chung; Zhou, Weiping; Xu, Xiaowei; Wang, Hongyang; Fang, Douglas D

    2015-08-21

    Lack of clinically relevant tumor models dramatically hampers development of effective therapies for hepatocellular carcinoma (HCC). Establishment of patient-derived xenograft (PDX) models that faithfully recapitulate the genetic and phenotypic features of HCC becomes important. In this study, we first established a cohort of 65 stable PDX models of HCC from corresponding Chinese patients. Then we showed that the histology and gene expression patterns of PDX models were highly consistent between xenografts and case-matched original tumors. Genetic alterations, including mutations and DNA copy number alterations (CNAs), of the xenografts correlated well with the published data of HCC patient specimens. Furthermore, differential responses to sorafenib, the standard-of-care agent, in randomly chosen xenografts were unveiled. Finally, in the models expressing high levels of FGFR1 gene according to the genomic data, FGFR1 inhibitor lenvatinib showed greater efficacy than sorafenib. Taken together, our data indicate that PDX models resemble histopathological and genomic characteristics of clinical HCC tumors, as well as recapitulate the differential responses of HCC patients to the standard-of-care treatment. Overall, this large collection of PDX models becomes a clinically relevant platform for drug screening, biomarker discovery and translational research in preclinical setting.

  14. Genomic characterization of a large panel of patient-derived hepatocellular carcinoma xenograft tumor models for preclinical development

    PubMed Central

    Sun, Hongye; Xu, Qiang; Tan, Yexiong; Wang, Guan; Luo, Qin; Xu, Weiguo; Yang, Shuqun; Li, Jian; Fu, Jing; Chen, Lei; Yuan, Shengxian; Liang, Guibai; Ji, Qunsheng; Chen, Shu-Hui; Chan, Chi-Chung; Zhou, Weiping; Xu, Xiaowei; Wang, Hongyang; Fang, Douglas D.

    2015-01-01

    Lack of clinically relevant tumor models dramatically hampers development of effective therapies for hepatocellular carcinoma (HCC). Establishment of patient-derived xenograft (PDX) models that faithfully recapitulate the genetic and phenotypic features of HCC becomes important. In this study, we first established a cohort of 65 stable PDX models of HCC from corresponding Chinese patients. Then we showed that the histology and gene expression patterns of PDX models were highly consistent between xenografts and case-matched original tumors. Genetic alterations, including mutations and DNA copy number alterations (CNAs), of the xenografts correlated well with the published data of HCC patient specimens. Furthermore, differential responses to sorafenib, the standard-of-care agent, in randomly chosen xenografts were unveiled. Finally, in the models expressing high levels of FGFR1 gene according to the genomic data, FGFR1 inhibitor lenvatinib showed greater efficacy than sorafenib. Taken together, our data indicate that PDX models resemble histopathological and genomic characteristics of clinical HCC tumors, as well as recapitulate the differential responses of HCC patients to the standard-of-care treatment. Overall, this large collection of PDX models becomes a clinically relevant platform for drug screening, biomarker discovery and translational research in preclinical setting. PMID:26062443

  15. Efficacy of protracted temozolomide dosing is limited in MGMT unmethylated GBM xenograft models

    PubMed Central

    Cen, Ling; Carlson, Brett L.; Pokorny, Jenny L.; Mladek, Ann C.; Grogan, Patrick T.; Schroeder, Mark A.; Decker, Paul A.; Anderson, S. Keith; Giannini, Caterina; Wu, Wenting; Ballman, Karla V.; Kitange, Gaspar J.; Sarkaria, Jann N.

    2013-01-01

    Background Temozolomide (TMZ) is important chemotherapy for glioblastoma multiforme (GBM), but the optimal dosing schedule is unclear. Methods The efficacies of different clinically relevant dosing regimens were compared in a panel of 7 primary GBM xenografts in an intracranial therapy evaluation model. Results Protracted TMZ therapy (TMZ daily M–F, 3 wk every 4) provided superior survival to a placebo-treated group in 1 of 4 O6-DNA methylguanine-methyltransferase (MGMT) promoter hypermethylated lines (GBM12) and none of the 3 MGMT unmethylated lines, while standard therapy (TMZ daily M–F, 1 wk every 4) provided superior survival to the placebo-treated group in 2 of 3 MGMT unmethylated lines (GBM14 and GBM43) and none of the methylated lines. In comparing GBM12, GBM14, and GBM43 intracranial specimens, both GBM14 and GBM43 mice treated with protracted TMZ had a significant elevation in MGMT levels compared with placebo. Similarly, high MGMT was found in a second model of acquired TMZ resistance in GBM14 flank xenografts, and resistance was reversed in vitro by treatment with the MGMT inhibitor O6-benzylguanine, demonstrating a mechanistic link between MGMT overexpression and TMZ resistance in this line. Additionally, in an analysis of gene expression data, comparison of parental and TMZ-resistant GBM14 demonstrated enrichment of functional ontologies for cell cycle control within the S, G2, and M phases of the cell cycle and DNA damage checkpoints. Conclusions Across the 7 tumor models studied, there was no consistent difference between protracted and standard TMZ regimens. The efficacy of protracted TMZ regimens may be limited in a subset of MGMT unmethylated tumors by induction of MGMT expression. PMID:23479134

  16. Human polyethylene granuloma tissues inhibit bone healing in a novel xenograft animal model.

    PubMed

    Esposito, Christina I; Oliver, Rema A; Campbell, Patricia A; Yu, Yan; Walter, William L; Walter, William K; Walsh, William R

    2014-06-01

    During revision of a conventional polyethylene joint replacement, surgeons usually remove the source of osteolysis (polyethylene) but cannot always remove all of the polyethylene granuloma tissues. We developed a human/rat xenograft model to investigate the effects of polyethylene granuloma tissues on bone healing. Human osteoarthritic and periprosthetic tissues collected during primary and revision hip arthroplasty surgeries were transplanted into the distal femora of athymic nude rats. After 3 weeks in vivo, there was a significant difference in the bone volume fraction (Vf ) between empty, primary, and revision defects (p = 0.02), with a lower Vf in defects with revision granuloma tissues compared to defects with primary osteoarthritic tissues. Polyethylene granuloma tissues in trabecular bone defects inhibited bone healing. Therefore, debridement around a metal-on-polyethylene hip replacement may shorten the time it takes to achieve secondary stability around a revision hip replacement.

  17. Treatment of malignant effusion by oncolytic virotherapy in an experimental subcutaneous xenograft model of lung cancer

    PubMed Central

    2013-01-01

    Background Malignant pleural effusion (MPE) is associated with advanced stages of lung cancer and is mainly dependent on invasion of the pleura and expression of vascular endothelial growth factor (VEGF) by cancer cells. As MPE indicates an incurable disease with limited palliative treatment options and poor outcome, there is an urgent need for new and efficient treatment options. Methods In this study, we used subcutaneously generated PC14PE6 lung adenocarcinoma xenografts in athymic mice that developed subcutaneous malignant effusions (ME) which mimic pleural effusions of the orthotopic model. Using this approach monitoring of therapeutic intervention was facilitated by direct observation of subcutaneous ME formation without the need of sacrificing mice or special imaging equipment as in case of MPE. Further, we tested oncolytic virotherapy using Vaccinia virus as a novel treatment modality against ME in this subcutaneous PC14PE6 xenograft model of advanced lung adenocarcinoma. Results We demonstrated significant therapeutic efficacy of Vaccinia virus treatment of both advanced lung adenocarcinoma and tumor-associated ME. We attribute the efficacy to the virus-mediated reduction of tumor cell-derived VEGF levels in tumors, decreased invasion of tumor cells into the peritumoral tissue, and to viral infection of the blood vessel-invading tumor cells. Moreover, we showed that the use of oncolytic Vaccinia virus encoding for a single-chain antibody (scAb) against VEGF (GLAF-1) significantly enhanced mono-therapy of oncolytic treatment. Conclusions Here, we demonstrate for the first time that oncolytic virotherapy using tumor-specific Vaccinia virus represents a novel and promising treatment modality for therapy of ME associated with advanced lung cancer. PMID:23635329

  18. CysLT(1)R antagonists inhibit tumor growth in a xenograft model of colon cancer.

    PubMed

    Savari, Sayeh; Liu, Minghui; Zhang, Yuan; Sime, Wondossen; Sjölander, Anita

    2013-01-01

    The expression of the inflammatory G-protein coupled receptor CysLT1R has been shown to be upregulated in colon cancer patients and associated with poor prognosis. The present study investigated the correlation between CysLT1R and colon cancer development in vivo using CysLT1R antagonists (ZM198,615 or Montelukast) and the nude mouse xenograft model. Two drug administration regimens were established. The first regimen was established to investigate the importance of CysLT1R in tumor initiation. Nude mice were inoculated with 50 µM CysLT1R antagonist-pretreated HCT-116 colon cancer cells and received continued treatment (5 mg/kg/day, intraperitoneally). The second regimen aimed to address the role of CysLT1R in tumor progression. Nude mice were inoculated with non-pretreated HCT-116 cells and did not receive CysLT1R antagonist treatment until recordable tumor appearance. Both regimens resulted in significantly reduced tumor size, attributed to changes in proliferation and apoptosis as determined by reduced Ki-67 levels and increased levels of p21(WAF/Cip1) (P<0.01), cleaved caspase 3, and the caspase-cleaved product of cytokeratin 18. Decreased levels of VEGF (P<0.01) and reduced vessel size (P<0.05) were also observed, the latter only in the ZM198,615-pretreatment group. Furthermore, we performed a series of in vitro studies using the colon cancer cell line HCT-116 and CysLT1R antagonists. In addition to significant reductions in cell proliferation, adhesion and colony formation, we observed induction of cell cycle arrest and apoptosis in a dose-dependent manner. The ability of Montelukast to inhibit growth of human colon cancer xenograft was further validated by using two additional colon cancer cell lines, SW-480 and HT-29. Our results demonstrate that CysLT1R antagonists inhibit growth of colon cancer xenografts primarily by reducing proliferation and inducing apoptosis of the tumor cells.

  19. Utility of a human-mouse xenograft model and in vivo near-infrared fluorescent imaging for studying wound healing.

    PubMed

    Shanmugam, Victoria K; Tassi, Elena; Schmidt, Marcel O; McNish, Sean; Baker, Stephen; Attinger, Christopher; Wang, Hong; Shara, Nawar; Wellstein, Anton

    2015-12-01

    To study the complex cellular interactions involved in wound healing, it is essential to have an animal model that adequately mimics the human wound microenvironment. Currently available murine models are limited because wound contraction introduces bias into wound surface area measurements. The purpose of this study was to demonstrate utility of a human-mouse xenograft model for studying human wound healing. Normal human skin was harvested from elective abdominoplasty surgery, xenografted onto athymic nude (nu/nu) mice, and allowed to engraft for 3 months. The graft was then wounded using a 2-mm punch biopsy. Wounds were harvested on sequential days to allow tissue-based markers of wound healing to be followed sequentially. On the day of wound harvest, mice were injected with XenoLight RediJect cyclooxygenase-2 (COX-2) probe and imaged according to package instructions. Immunohistochemistry confirms that this human-mouse xenograft model is effective for studying human wound healing in vivo. Additionally, in vivo fluorescent imaging for inducible COX-2 demonstrated upregulation from baseline to day 4 (P = 0·03) with return to baseline levels by day 10, paralleling the reepithelialisation of the wound. This human-mouse xenograft model, combined with in vivo fluorescent imaging provides a useful mechanism for studying molecular pathways of human wound healing.

  20. Patient-Derived Gastric Carcinoma Xenograft Mouse Models Faithfully Represent Human Tumor Molecular Diversity.

    PubMed

    Zhang, Tianwei; Zhang, Lin; Fan, Shuqiong; Zhang, Meizhuo; Fu, Haihua; Liu, Yuanjie; Yin, Xiaolu; Chen, Hao; Xie, Liang; Zhang, Jingchuan; Gavine, Paul R; Gu, Yi; Ni, Xingzhi; Su, Xinying

    2015-01-01

    Patient-derived cancer xenografts (PDCX) generally represent more reliable models of human disease in which to evaluate a potential drugs preclinical efficacy. However to date, only a few patient-derived gastric cancer xenograft (PDGCX) models have been reported. In this study, we aimed to establish additional PDGCX models and to evaluate whether these models accurately reflected the histological and genetic diversities of the corresponding patient tumors. By engrafting fresh patient gastric cancer (GC) tissues into immune-compromised mice (SCID and/or nude mice), thirty two PDGCX models were established. Histological features were assessed by a qualified pathologist based on H&E staining. Genomic comparison was performed for several biomarkers including ERBB1, ERBB2, ERBB3, FGFR2, MET and PTEN. These biomarkers were profiled to assess gene copy number by fluorescent in situ hybridization (FISH) and/or protein expression by immunohistochemistry (IHC). All 32 PDGCX models retained the histological features of the corresponding human tumors. Furthermore, among the 32 models, 78% (25/32) highly expressed ERBB1 (EGFR), 22% (7/32) were ERBB2 (HER2) positive, 78% (25/32) showed ERBB3 (HER3) high expression, 66% (21/32) lost PTEN expression, 3% (1/32) harbored FGFR2 amplification, 41% (13/32) were positive for MET expression and 16% (5/32) were MET gene amplified. Between the PDGCX models and their parental tumors, a high degree of similarity was observed for FGFR2 and MET gene amplification, and also for ERBB2 status (agreement rate = 94~100%; kappa value = 0.81~1). Protein expression of PTEN and MET also showed moderate agreement (agreement rate = 78%; kappa value = 0.46~0.56), while ERBB1 and ERBB3 expression showed slight agreement (agreement rate = 59~75%; kappa value = 0.18~0.19). ERBB2 positivity, FGFR2 or MET gene amplification was all maintained until passage 12 in mice. The stability of the molecular profiles observed across subsequent passages within the

  1. New mouse xenograft model modulated by tumor-associated fibroblasts for human multi-drug resistance in cancer

    PubMed Central

    MA, YAN; LIN, ZHIQIANG; FALLON, JOHN K.; ZHAO, QIANG; LIU, DAN; WANG, YONGJUN; LIU, FENG

    2015-01-01

    We developed an MDR tumor model that is modulated by tumor-associated fibroblasts. Studies on proliferation of tumor cell lines including paclitaxel-sensitive and resistant cell lines were performed. The expressions of P-gp and α-smooth muscle actin (α-SMA) antigen were evaluated by immunohistochemistry and western blot analysis. Quantitative P-gp analyses of different cell lines were accomplished by nanoUPLC-MS/MS. Tumor cell colony formation assay and established xenograft model was used to investigate the relationship between P-gp expression, fibroblast levels and tumorigenesis. The mouse xenograft model was developed after co-inoculation with MDR tumor cells and NIH/3T3 fibroblast cells. There was no correlation between tumorigenesis in vivo and the growth rate of cells in vitro. The proliferation among different cell lines had no significant differences, but the P-gp expression and tumor growth in the xenograft model were fairly different. P-gp determination and α-SMA immunofluorescence staining clarified the relationship between P-gp expression, fibroblast levels and tumorigenesis. It was more difficult for tumor cells with higher P-gp levels to recruit fibroblasts in vivo, resulting in lower tumorigenesis due to the lack of structural and chemical support during tumor progression. In the established paclitaxel-resistant mouse xenograft model, no obvious antitumor effect was observed after Taxol treatment, but a significant decrease in tumor size for the group treated with gemcitabine sensitive to the model. The results show that the added fibroblasts do not disturb the applicability of the model in MDR. Therefore, this mouse xenograft MDR model could serve as an effective tool for MDR research. PMID:26352907

  2. The Anti-Proliferative Effect of Boron Neutron Capture Therapy in a Prostate Cancer Xenograft Model

    PubMed Central

    Yoshikawa, Yuki; Takai, Tomoaki; Ibuki, Naokazu; Hirano, Hajime; Nomi, Hayahito; Kawabata, Shinji; Kiyama, Satoshi; Miyatake, Shin-Ichi; Kuroiwa, Toshihiko; Suzuki, Minoru; Kirihata, Mitsunori; Azuma, Haruhito

    2015-01-01

    Purpose Boron neutron capture therapy (BNCT) is a selective radiation treatment for tumors that preferentially accumulate drugs carrying the stable boron isotope, 10B. BNCT has been evaluated clinically as an alternative to conventional radiation therapy for the treatment of brain tumors, and more recently, recurrent advanced head and neck cancer. Here we investigated the effect of BNCT on prostate cancer (PCa) using an in vivo mouse xenograft model that we have developed. Materials and Methods Mice bearing the xenotransplanted androgen-independent human PCa cell line, PC3, were divided into four groups: Group 1: untreated controls; Group 2: Boronophenylalanine (BPA); Group 3: neutron; Group 4: BPA-mediated BNCT. We compared xenograft growth among these groups, and the body weight and any motility disturbance were recorded. Immunohistochemical (IHC) studies of the proliferation marker, Ki-67, and TUNEL staining were performed 9 weeks after treatment. Results The in vivo studies demonstrated that BPA-mediated BNCT significantly delayed tumor growth in comparison with the other groups, without any severe adverse events. There was a significant difference in the rate of freedom from gait abnormalities between the BPA-mediated BNCT group and the other groups. The IHC studies revealed that BNCT treatment significantly reduced the number of Ki-67-positive cells in comparison with the controls (mean±SD 6.9±1.5 vs 12.7±4.0, p<0.05), while there was no difference in the number of apoptotic cells, suggesting that BPA-mediated BNCT reduced PCa progression without affecting apoptosis at 9 weeks post-treatment. Conclusions This study has provided the first preclinical proof-of-principle data to indicate that BPA-mediated BNCT reduces the in vivo growth of PCa. Although further studies will be necessary, BNCT might be a novel potential treatment for PCa. PMID:26325195

  3. Growth hormone receptor antagonism suppresses tumour regrowth after radiotherapy in an endometrial cancer xenograft model.

    PubMed

    Evans, Angharad; Jamieson, Stephen M F; Liu, Dong-Xu; Wilson, William R; Perry, Jo K

    2016-08-28

    Human GH expression is associated with poor survival outcomes for endometrial cancer patients, enhanced oncogenicity of endometrial cancer cells and reduced sensitivity to ionising radiation in vitro, suggesting that GH is a potential target for anticancer therapy. However, whether GH receptor inhibition sensitises to radiotherapy in vivo has not been tested. In the current study, we evaluated whether the GH receptor antagonist, pegvisomant (Pfizer), sensitises to radiotherapy in vivo in an endometrial tumour xenograft model. Subcutaneous administration of pegvisomant (20 or 100 mg/kg/day, s.c.) reduced serum IGF1 levels by 23% and 68%, respectively, compared to vehicle treated controls. RL95-2 xenografts grown in immunodeficient NIH-III mice were treated with vehicle or pegvisomant (100 mg/kg/day), with or without fractionated gamma radiation (10 × 2.5 Gy over 5 days). When combined with radiation, pegvisomant significantly increased the median time tumours took to reach 3× the pre-radiation treatment volume (49 days versus 72 days; p = 0.001). Immunohistochemistry studies demonstrated that 100 mg/kg pegvisomant every second day was sufficient to abrogate MAP Kinase signalling throughout the tumour. In addition, treatment with pegvisomant increased hypoxic regions in irradiated tumours, as determined by immunohistochemical detection of pimonidazole adducts, and decreased the area of CD31 labelling in unirradiated tumours, suggesting an anti-vascular effect. Pegvisomant did not affect intratumoral staining for HIF1α, VEGF-A, CD11b, or phospho-EGFR. Our results suggest that blockade of the human GH receptor may improve the response of GH and/or IGF1-responsive endometrial tumours to radiation.

  4. Survivin Antisense Oligonucleotides Effectively Radiosensitize Colorectal Cancer Cells in Both Tissue Culture and Murine Xenograft Models

    SciTech Connect

    Roedel, Franz; Capalbo, Gianni; Weiss, Christian; Roedel, Claus

    2008-05-01

    Purpose: Survivin shows a radiation resistance factor in colorectal cancer. In the present study, we determined whether survivin messenger RNA levels in patients with rectal cancer predict tumor response after neoadjuvant radiochemotherapy and whether inhibition of survivin by the use of antisense oligonucleotides (ASOs) enhances radiation responses. Methods and Materials: SW480 colorectal carcinoma cells were transfected with survivin ASO (LY2181308) and irradiated with doses ranging from 0-8 Gy. Survivin expression, cell-cycle distribution, {gamma}H2AX fluorescence, and induction of apoptosis were monitored by means of immunoblotting, flow cytometry, and caspase 3/7 activity. Clonogenic survival was determined by using a colony-forming assay. An SW480 xenograft model was used to investigate the effect of survivin attenuation and irradiation on tumor growth. Furthermore, survivin messenger RNA levels were studied in patient biopsy specimens by using Affymetrix microarray analysis. Results: In the translational study of 20 patients with rectal cancer, increased survivin levels were associated with significantly greater risk of local tumor recurrence (p = 0.009). Treatment of SW480 cells with survivin ASOs and irradiation resulted in an increased percentage of apoptotic cells, caspase 3/7 activity, fraction of cells in the G{sub 2}/M phase, and H2AX phosphorylation. Clonogenic survival decreased compared with control-treated cells. Furthermore, treatment of SW480 xenografts with survivin ASOs and irradiation resulted in a significant delay in tumor growth. Conclusion: Survivin appears to be a molecular biomarker in patients with rectal cancer. Furthermore, in vitro and in vivo data suggest a potential role of survivin as a molecular target to improve treatment response to radiotherapy in patients with rectal cancer.

  5. Cetuximab intensifies the ADCC activity of adoptive NK cells in a nude mouse colorectal cancer xenograft model

    PubMed Central

    Chen, Shanshan; Li, Xuechun; Chen, Rongming; Yin, Mingang; Zheng, Qiuhong

    2016-01-01

    Natural killer (NK) cells, discovered ~40 years ago, are believed to be the most effective cytotoxic lymphocytes to counteract cancer; however, adoptive NK cell therapy in vivo has encountered certain limitations, including a lack of specificity. The drug cetuximab can mediate antibody dependent cell mediated cytotoxicity (ADCC) activity through NK cells in vivo, and has been approved for the first-line treatment of epidermal growth factor receptor (EGFR)-positive metastatic colorectal cancer (CRC). However, the ADCC activity of adoptive NK cells, induced by cetuximab in a nude mouse CRC xenograft model, has not been previously reported. The aim of the present study was to explore the ADCC activity of cetuximab combined with adoptive NK cells in CRC xenograft models with various EGFR expressions. The nude mouse xenograft models were established by subcutaneously injecting LOVO or SW620 cells. The mice were then randomly divided into 6 groups: Phosphate-buffered saline, cetuximab, human immunoglobulin G (hIgG), NK cells, hIgG plus NK cells and cetuximab plus NK cells. The ADCC antitumor activity was evaluated in these CRC models. The results indicated that the cetuximab plus NK cells group showed the greatest tumor inhibition effect compared with the NK cells group in LOVO xenograft tumor models with positive EGFR expression. However, the combination of cetuximab and NK cells did not show a stronger tumor inhibitory effect against the SW620 xenograft tumor models compared with the efficiency of NK cells. In conclusion, cetuximab could intensify the ADCC antitumor activity of adoptive NK cells towards CRC with an increased EGFR expression. The combination of cetuximab and NK cells may be a potential immunotherapy for metastatic CRC patients with positive EGFR expression.

  6. Cetuximab intensifies the ADCC activity of adoptive NK cells in a nude mouse colorectal cancer xenograft model

    PubMed Central

    Chen, Shanshan; Li, Xuechun; Chen, Rongming; Yin, Mingang; Zheng, Qiuhong

    2016-01-01

    Natural killer (NK) cells, discovered ~40 years ago, are believed to be the most effective cytotoxic lymphocytes to counteract cancer; however, adoptive NK cell therapy in vivo has encountered certain limitations, including a lack of specificity. The drug cetuximab can mediate antibody dependent cell mediated cytotoxicity (ADCC) activity through NK cells in vivo, and has been approved for the first-line treatment of epidermal growth factor receptor (EGFR)-positive metastatic colorectal cancer (CRC). However, the ADCC activity of adoptive NK cells, induced by cetuximab in a nude mouse CRC xenograft model, has not been previously reported. The aim of the present study was to explore the ADCC activity of cetuximab combined with adoptive NK cells in CRC xenograft models with various EGFR expressions. The nude mouse xenograft models were established by subcutaneously injecting LOVO or SW620 cells. The mice were then randomly divided into 6 groups: Phosphate-buffered saline, cetuximab, human immunoglobulin G (hIgG), NK cells, hIgG plus NK cells and cetuximab plus NK cells. The ADCC antitumor activity was evaluated in these CRC models. The results indicated that the cetuximab plus NK cells group showed the greatest tumor inhibition effect compared with the NK cells group in LOVO xenograft tumor models with positive EGFR expression. However, the combination of cetuximab and NK cells did not show a stronger tumor inhibitory effect against the SW620 xenograft tumor models compared with the efficiency of NK cells. In conclusion, cetuximab could intensify the ADCC antitumor activity of adoptive NK cells towards CRC with an increased EGFR expression. The combination of cetuximab and NK cells may be a potential immunotherapy for metastatic CRC patients with positive EGFR expression. PMID:27602116

  7. Establishment and characterisation of patient-derived xenografts as paraclinical models for gastric cancer

    PubMed Central

    Choi, Yoon Young; Lee, Jae Eun; Kim, Hyunki; Sim, Moon Hee; Kim, Ka-Kyung; Lee, Gunho; Kim, Hyoung-Il; An, Ji Yeong; Hyung, Woo Jin; Kim, Choong-Bai; Noh, Sung Hoon; Kim, Sangwoo; Cheong, Jae-Ho

    2016-01-01

    The patient-derived xenograft (PDX) model is emerging as a promising translational platform to duplicate the characteristics of tumours. However, few studies have reported detailed histological and genomic analyses for model fidelity and for factors affecting successful model establishment of gastric cancer. Here, we generated PDX tumours surgically-derived from 62 gastric cancer patients. Fifteen PDX models were successfully established (24.2%, 15/62) and passaged to maintain tumours in immune-compromised mice. Diffuse type and low tumour cell percentage were negatively correlated with success rates (p = 0.005 and p = 0.025, respectively), while reducing ex vivo and overall procedure times were positively correlated with success rates (p = 0.003 and p = 0.01, respectively). The histology and genetic characteristics of PDX tumour models were stable over subsequent passages. Lymphoma transformation occurred in five cases (33.3%, 5/15), and all were in the NOG mouse, with none in the nude mouse. Together, the present study identified Lauren classification, tumour cell percentages, and ex vivo times along with overall procedure times, as key determinants for successful PDX engraftment. Furthermore, genetic and histological characteristics were highly consistent between primary and PDX tumours, which provide realistic paraclinical models, enabling personalised development of treatment options for gastric cancer. PMID:26926953

  8. Establishment and characterisation of patient-derived xenografts as paraclinical models for gastric cancer.

    PubMed

    Choi, Yoon Young; Lee, Jae Eun; Kim, Hyunki; Sim, Moon Hee; Kim, Ka-Kyung; Lee, Gunho; Kim, Hyoung-Il; An, Ji Yeong; Hyung, Woo Jin; Kim, Choong-Bai; Noh, Sung Hoon; Kim, Sangwoo; Cheong, Jae-Ho

    2016-03-01

    The patient-derived xenograft (PDX) model is emerging as a promising translational platform to duplicate the characteristics of tumours. However, few studies have reported detailed histological and genomic analyses for model fidelity and for factors affecting successful model establishment of gastric cancer. Here, we generated PDX tumours surgically-derived from 62 gastric cancer patients. Fifteen PDX models were successfully established (24.2%, 15/62) and passaged to maintain tumours in immune-compromised mice. Diffuse type and low tumour cell percentage were negatively correlated with success rates (p = 0.005 and p = 0.025, respectively), while reducing ex vivo and overall procedure times were positively correlated with success rates (p = 0.003 and p = 0.01, respectively). The histology and genetic characteristics of PDX tumour models were stable over subsequent passages. Lymphoma transformation occurred in five cases (33.3%, 5/15), and all were in the NOG mouse, with none in the nude mouse. Together, the present study identified Lauren classification, tumour cell percentages, and ex vivo times along with overall procedure times, as key determinants for successful PDX engraftment. Furthermore, genetic and histological characteristics were highly consistent between primary and PDX tumours, which provide realistic paraclinical models, enabling personalised development of treatment options for gastric cancer.

  9. Patient-derived xenograft models for pancreatic adenocarcinoma demonstrate retention of tumor morphology through incorporation of murine stromal elements.

    PubMed

    Delitto, Daniel; Pham, Kien; Vlada, Adrian C; Sarosi, George A; Thomas, Ryan M; Behrns, Kevin E; Liu, Chen; Hughes, Steven J; Wallet, Shannon M; Trevino, Jose G

    2015-05-01

    Direct implantation of viable surgical specimens provides a representative preclinical platform in pancreatic adenocarcinoma. Patient-derived xenografts consistently demonstrate retained tumor morphology and genetic stability. However, the evolution of the tumor microenvironment over time remains poorly characterized in these models. This work specifically addresses the recruitment and incorporation of murine stromal elements into expanding patient-derived pancreatic adenocarcinoma xenografts, establishing the integration of murine cells into networks of invading cancer cells. In addition, we provide methods and observations in the establishment and maintenance of a patient-derived pancreatic adenocarcinoma xenograft model. A total of 25 histologically confirmed pancreatic adenocarcinoma specimens were implanted subcutaneously into nonobese diabetic severe combined immunodeficiency mice. Patient demographics, staging, pathological analysis, and outcomes were analyzed. After successful engraftment of tumors, histological and immunofluorescence analyses were performed on explanted tumors. Pancreatic adenocarcinoma specimens were successfully engrafted in 15 (60%) of 25 attempts. Successful engraftment does not appear to correlate with clinicopathologic factors or patient survival. Tumor morphology is conserved through multiple passages, and tumors retain metastatic potential. Interestingly, despite morphological similarity between passages, human stromal elements do not appear to expand with invading cancer cells. Rather, desmoplastic murine stroma dominates the xenograft microenvironment after the initial implantation. Recruitment of stromal elements in this manner to support and maintain tumor growth represents a novel avenue for investigation into tumor-stromal interactions.

  10. Development of Patient Derived Xenograft Models of Overt Spontaneous Breast Cancer Metastasis: A Cautionary Note

    PubMed Central

    Paez-Ribes, Marta; Man, Shan; Xu, Ping; Kerbel, Robert S.

    2016-01-01

    Several approaches are being evaluated to improve the historically limited value of studying transplanted primary tumors derived by injection of cells from established cell lines for predicting subsequent cancer therapy outcomes in patients and clinical trials. These approaches include use of genetically engineered mouse models (GEMMs) of spontaneous tumors, or patient tumor tissue derived xenografts (PDXs). Almost all such therapy studies utilizing such models involve treatment of established primary tumors. An alternative approach we have developed involves transplanted human tumor xenografts derived from established cell lines to treat mice with overt visceral metastases after primary tumor resection. The rationale is to mimic the more challenging circumstance of treating patients with late stage metastatic disease. These metastatic models entail prior in vivo selection of heritable, phenotypically stable variants with increased aggressiveness for spontaneous metastasis; they were derived by orthotopic injection of tumor cells followed by primary tumor resection and serial selection of distant spontaneous metastases, from which variant cell lines having a more aggressive heritable metastatic phenotype were established. We attempted to adopt this strategy for breast cancer PDXs. We studied five breast cancer PDXs, with the emphasis on two, called HCI-001 and HCI-002, both derived from triple negative breast cancer patients. However significant technical obstacles were encountered. These include the inherent slow growth rates of PDXs, the rarity of overt spontaneous metastases (detected in only 3 of 144 mice), very high rates of tumor regrowths at the primary tumor resection site, the failure of the few human PDX metastases isolated to manifest a more aggressive metastatic phenotype upon re-transplantation into new hosts, and the formation of metastases which were derived from de novo mouse thymomas arising in aged SCID mice that we used for the experiments. We

  11. Application of a Patient Derived Xenograft Model for Predicative Study of Uterine Fibroid Disease.

    PubMed

    Fritsch, Martin; Schmidt, Nicole; Gröticke, Ina; Frisk, Anna-Lena; Keator, Christopher S; Koch, Markus; Slayden, Ov D

    2015-01-01

    Human uterine fibroids, benign tumors derived from the smooth muscle layers of the uterus, impose a major health burden to up to 50% of premenopausal women in their daily life. To improve our understanding of this disease, we developed and characterized a patient-derived xenograft model by subcutaneous transplantation of pieces of human uterine fibroid tissue into three different strains of severe combined immunodeficient mice. Engrafted uterine fibroid tissue preserved the classical morphology with interwoven bundles of smooth muscle cells and an abundant deposition of collagenous matrix, similar to uterine fibroids in situ. The grafts expressed both estrogen receptor 1 and progesterone receptor. Additionally, both receptors were up-regulated by estrogen treatment. Growth of the fibroid grafts was dependent on 17β-estradiol and progesterone supplementation at levels similar to women with the disease and was studied for up to 60 days at maximum. Co-treatment with the antiprogestin mifepristone reduced graft growth (four independent donors, p<0.0001 two-sided t-test), as did treatment with the mTOR inhibitor rapamycin (three independent donors, p<0.0001 two-sided t-test). This in vivo animal model preserves the main histological and functional characteristics of human uterine fibroids, is amenable to intervention by pharmacological treatment, and can thus serve as an adequate model for the development of novel therapies. PMID:26588841

  12. Evaluating patient-derived colorectal cancer xenografts as preclinical models by comparison with patient clinical data.

    PubMed

    Nunes, Manoel; Vrignaud, Patricia; Vacher, Sophie; Richon, Sophie; Lièvre, Astrid; Cacheux, Wulfran; Weiswald, Louis-Bastien; Massonnet, Gerald; Chateau-Joubert, Sophie; Nicolas, André; Dib, Colette; Zhang, Weidong; Watters, James; Bergstrom, Donald; Roman-Roman, Sergio; Bièche, Ivan; Dangles-Marie, Virginie

    2015-04-15

    Development of targeted therapeutics required translationally relevant preclinical models with well-characterized cancer genome alterations. Here, by studying 52 colorectal patient-derived tumor xenografts (PDX), we examined key molecular alterations of the IGF2-PI3K and ERBB-RAS pathways and response to cetuximab. PDX molecular data were compared with that published for patient colorectal tumors in The Cancer Genome Atlas. We demonstrated a significant pattern of mutual exclusivity of genomic abnormalities in the IGF2-PI3K and ERBB-RAS pathways. The genomic anomaly frequencies observed in microsatellite stable PDX reproduce those detected in nonhypermutated patient tumors. We found frequent IGF2 upregulation (16%), which was mutually exclusive with IRS2, PIK3CA, PTEN, and INPP4B alterations, supporting IGF2 as a potential drug target. In addition to maintaining the genomic and histologic diversity, correct preclinical models need to reproduce drug response observed in patients. Responses of PDXs to cetuximab recapitulate also clinical data in patients, with partial or complete response in 15% (8 of 52) of PDXs and response strictly restricted to KRAS wild-type models. The response rate reaches 53% (8 of 15) when KRAS, BRAF, and NRAS mutations are concomitantly excluded, proving a functional cross-validation of predictive biomarkers obtained retrospectively in patients. Collectively, these results show that, because of their clinical relevance, colorectal PDXs are appropriate tools to identify both new targets, like IGF2, and predictive biomarkers of response/resistance to targeted therapies.

  13. Application of a Patient Derived Xenograft Model for Predicative Study of Uterine Fibroid Disease.

    PubMed

    Fritsch, Martin; Schmidt, Nicole; Gröticke, Ina; Frisk, Anna-Lena; Keator, Christopher S; Koch, Markus; Slayden, Ov D

    2015-01-01

    Human uterine fibroids, benign tumors derived from the smooth muscle layers of the uterus, impose a major health burden to up to 50% of premenopausal women in their daily life. To improve our understanding of this disease, we developed and characterized a patient-derived xenograft model by subcutaneous transplantation of pieces of human uterine fibroid tissue into three different strains of severe combined immunodeficient mice. Engrafted uterine fibroid tissue preserved the classical morphology with interwoven bundles of smooth muscle cells and an abundant deposition of collagenous matrix, similar to uterine fibroids in situ. The grafts expressed both estrogen receptor 1 and progesterone receptor. Additionally, both receptors were up-regulated by estrogen treatment. Growth of the fibroid grafts was dependent on 17β-estradiol and progesterone supplementation at levels similar to women with the disease and was studied for up to 60 days at maximum. Co-treatment with the antiprogestin mifepristone reduced graft growth (four independent donors, p<0.0001 two-sided t-test), as did treatment with the mTOR inhibitor rapamycin (three independent donors, p<0.0001 two-sided t-test). This in vivo animal model preserves the main histological and functional characteristics of human uterine fibroids, is amenable to intervention by pharmacological treatment, and can thus serve as an adequate model for the development of novel therapies.

  14. Application of a Patient Derived Xenograft Model for Predicative Study of Uterine Fibroid Disease

    PubMed Central

    Gröticke, Ina; Frisk, Anna-Lena; Keator, Christopher S.; Koch, Markus; Slayden, Ov D.

    2015-01-01

    Human uterine fibroids, benign tumors derived from the smooth muscle layers of the uterus, impose a major health burden to up to 50% of premenopausal women in their daily life. To improve our understanding of this disease, we developed and characterized a patient-derived xenograft model by subcutaneous transplantation of pieces of human uterine fibroid tissue into three different strains of severe combined immunodeficient mice. Engrafted uterine fibroid tissue preserved the classical morphology with interwoven bundles of smooth muscle cells and an abundant deposition of collagenous matrix, similar to uterine fibroids in situ. The grafts expressed both estrogen receptor 1 and progesterone receptor. Additionally, both receptors were up-regulated by estrogen treatment. Growth of the fibroid grafts was dependent on 17β-estradiol and progesterone supplementation at levels similar to women with the disease and was studied for up to 60 days at maximum. Co-treatment with the antiprogestin mifepristone reduced graft growth (four independent donors, p<0.0001 two-sided t-test), as did treatment with the mTOR inhibitor rapamycin (three independent donors, p<0.0001 two-sided t-test). This in vivo animal model preserves the main histological and functional characteristics of human uterine fibroids, is amenable to intervention by pharmacological treatment, and can thus serve as an adequate model for the development of novel therapies. PMID:26588841

  15. Patient-derived xenograft models to improve targeted therapy in epithelial ovarian cancer treatment.

    PubMed

    Scott, Clare L; Becker, Marc A; Haluska, Paul; Samimi, Goli

    2013-12-04

    Despite increasing evidence that precision therapy targeted to the molecular drivers of a cancer has the potential to improve clinical outcomes, high-grade epithelial ovarian cancer (OC) patients are currently treated without consideration of molecular phenotype, and predictive biomarkers that could better inform treatment remain unknown. Delivery of precision therapy requires improved integration of laboratory-based models and cutting-edge clinical research, with pre-clinical models predicting patient subsets that will benefit from a particular targeted therapeutic. Patient-derived xenografts (PDXs) are renewable tumor models engrafted in mice, generated from fresh human tumors without prior in vitro exposure. PDX models allow an invaluable assessment of tumor evolution and adaptive response to therapy. PDX models have been applied to pre-clinical drug testing and biomarker identification in a number of cancers including ovarian, pancreatic, breast, and prostate cancers. These models have been shown to be biologically stable and accurately reflect the patient tumor with regards to histopathology, gene expression, genetic mutations, and therapeutic response. However, pre-clinical analyses of molecularly annotated PDX models derived from high-grade serous ovarian cancer (HG-SOC) remain limited. In vivo response to conventional and/or targeted therapeutics has only been described for very small numbers of individual HG-SOC PDX in conjunction with sparse molecular annotation and patient outcome data. Recently, two consecutive panels of epithelial OC PDX correlate in vivo platinum response with molecular aberrations and source patient clinical outcomes. These studies underpin the value of PDX models to better direct chemotherapy and predict response to targeted therapy. Tumor heterogeneity, before and following treatment, as well as the importance of multiple molecular aberrations per individual tumor underscore some of the important issues addressed in PDX models.

  16. Safety and efficacy of quadrapeutics versus chemoradiation in head and neck carcinoma xenograft model.

    PubMed

    Lukianova-Hleb, Ekaterina Y; Kim, Yoo-Shin; Aryasomayajula, Bhawani; Boulikas, Teni; Phan, Jack; Hung, Mien-Chie; Torchilin, Vladimir P; O'Neill, Brian E; Lapotko, Dmitri O

    2015-01-01

    Chemoradiation is the strongest anti-tumor therapy but in resistant unresectable cancers it often lacks safety and efficacy. We compared our recently developed cell-level combination approach, quadrapeutics, to chemoradiation therapy to establish pre-clinical data for its biodistribution, safety and efficacy in head and neck squamous cell carcinoma (HNSCC), as a clinically challenging aggressive and resistant cancer. In vitro and in vivo models of four carcinomas were treated with standard chemoradiation and quadrapeutics using identical drug and radiation doses. We applied liposomal cisplatin or doxorubicin, colloidal gold, near-infrared laser pulses and radiation, all at low safe doses. The final evaluation used a xenograft model of HNSCC. Quadrapeutics enhanced standard chemoradiation in vitro by reducing head and neck cancer cell proliferation by 1000-fold, inhibiting tumor growth in vivo by 34-fold and improving animal survival by 5-fold, and reducing the side effects to a negligible level. In quadrapeutics, we observed an "inversion" of the drug efficacy of two standard drugs: doxorubicin, a low efficacy drug for the cancers studied, was two times more efficient than cisplatin, the first choice drug in clinic for HNSCC. The radical therapeutic gain of quadrapeutics resulted from the intracellular synergy of the four components employed which we administered in a specific sequence, while the reduction in the toxicity was due to the low doses of all four components. The biodistribution, safety and efficacy data for quadrapeutics in HNSCC ensure its high translational potential and justify the possibility of clinical trials. PMID:26885444

  17. Residual Disease in a Novel Xenograft Model of RUNX1-Mutated, Cytogenetically Normal Acute Myeloid Leukemia

    PubMed Central

    Sivagnanalingam, Umayal; Balys, Marlene; Eberhardt, Allison; Wang, Nancy; Myers, Jason R.; Ashton, John M.; Becker, Michael W.; Calvi, Laura M.; Mendler, Jason H.

    2015-01-01

    Cytogenetically normal acute myeloid leukemia (CN-AML) patients harboring RUNX1 mutations have a dismal prognosis with anthracycline/cytarabine-based chemotherapy. We aimed to develop an in vivo model of RUNX1-mutated, CN-AML in which the nature of residual disease in this molecular disease subset could be explored. We utilized a well-characterized patient-derived, RUNX1-mutated CN-AML line (CG-SH). Tail vein injection of CG-SH into NOD scid gamma mice led to leukemic engraftment in the bone marrow, spleen, and peripheral blood within 6 weeks. Treatment of leukemic mice with anthracycline/cytarabine-based chemotherapy resulted in clearance of disease from the spleen and peripheral blood, but persistence of disease in the bone marrow as assessed by flow cytometry and secondary transplantation. Whole exome sequencing of CG-SH revealed mutations in ASXL1, CEBPA, GATA2, and SETBP1, not previously reported. We conclude that CG-SH xenografts are a robust, reproducible in vivo model of CN-AML in which to explore mechanisms of chemotherapy resistance and novel therapeutic approaches. PMID:26177509

  18. Tryptophan PET Imaging of the Kynurenine Pathway in Patient-Derived Xenograft Models of Glioblastoma

    PubMed Central

    Guastella, Anthony R.; Michelhaugh, Sharon K.; Klinger, Neil V.; Kupsky, William J.; Polin, Lisa A.; Muzik, Otto; Juhász, Csaba; Mittal, Sandeep

    2016-01-01

    Increasing evidence demonstrates the immunosuppressive kynurenine pathway’s (KP) role in the pathophysiology of human gliomas. To study the KP in vivo, we used the noninvasive molecular imaging tracer α-[11C]-methyl-l-tryptophan (AMT). The AMT-positron emission tomography (PET) has shown high uptake in high-grade gliomas and predicted survival in patients with recurrent glioblastoma (GBM). We generated patient-derived xenograft (PDX) models from dissociated cells, or tumor fragments, from 5 patients with GBM. Mice bearing subcutaneous tumors were imaged with AMT-PET, and tumors were analyzed to detect the KP enzymes indoleamine 2,3-dioxygenase (IDO) 1, IDO2, tryptophan 2,3-dioxygenase, kynureninase, and kynurenine 3-monooxygenase. Overall, PET imaging showed robust tumoral AMT uptake in PDX mice with prolonged tracer accumulation over 60 minutes, consistent with AMT trapping seen in humans. Immunostained tumor tissues demonstrated positive detection of multiple KP enzymes. Furthermore, intracranial implantation of GBM cells was performed with imaging at both 9 and 14 days postimplant, with a marked increase in AMT uptake at 14 days and a corresponding high level of tissue immunostaining for KP enzymes. These results indicate that our PDX mouse models recapitulate human GBM, including aberrant tryptophan metabolism, and offer an in vivo system for development of targeted therapeutics for patients with GBM. PMID:27151136

  19. Characterization of novel genomic alterations and therapeutic approaches using acute megakaryoblastic leukemia xenograft models

    PubMed Central

    Thiollier, Clarisse; Lopez, Cécile K.; Gerby, Bastien; Ignacimouttou, Cathy; Poglio, Sandrine; Duffourd, Yannis; Guégan, Justine; Rivera-Munoz, Paola; Bluteau, Olivier; Mabialah, Vinciane; Diop, M’Boyba; Wen, Qiang; Petit, Arnaud; Bauchet, Anne-Laure; Reinhardt, Dirk; Bornhauser, Beat; Gautheret, Daniel; Lecluse, Yann; Landman-Parker, Judith; Radford, Isabelle; Vainchenker, William; Dastugue, Nicole; de Botton, Stéphane; Dessen, Philippe; Bourquin, Jean-Pierre; Crispino, John D.; Ballerini, Paola; Bernard, Olivier A.; Pflumio, Françoise

    2012-01-01

    Acute megakaryoblastic leukemia (AMKL) is a heterogeneous disease generally associated with poor prognosis. Gene expression profiles indicate the existence of distinct molecular subgroups, and several genetic alterations have been characterized in the past years, including the t(1;22)(p13;q13) and the trisomy 21 associated with GATA1 mutations. However, the majority of patients do not present with known mutations, and the limited access to primary patient leukemic cells impedes the efficient development of novel therapeutic strategies. In this study, using a xenotransplantation approach, we have modeled human pediatric AMKL in immunodeficient mice. Analysis of high-throughput RNA sequencing identified recurrent fusion genes defining new molecular subgroups. One subgroup of patients presented with MLL or NUP98 fusion genes leading to up-regulation of the HOX A cluster genes. A novel CBFA2T3-GLIS2 fusion gene resulting from a cryptic inversion of chromosome 16 was identified in another subgroup of 31% of non–Down syndrome AMKL and strongly associated with a gene expression signature of Hedgehog pathway activation. These molecular data provide useful markers for the diagnosis and follow up of patients. Finally, we show that AMKL xenograft models constitute a relevant in vivo preclinical screening platform to validate the efficacy of novel therapies such as Aurora A kinase inhibitors. PMID:23045605

  20. Characterization of novel genomic alterations and therapeutic approaches using acute megakaryoblastic leukemia xenograft models.

    PubMed

    Thiollier, Clarisse; Lopez, Cécile K; Gerby, Bastien; Ignacimouttou, Cathy; Poglio, Sandrine; Duffourd, Yannis; Guégan, Justine; Rivera-Munoz, Paola; Bluteau, Olivier; Mabialah, Vinciane; Diop, M'boyba; Wen, Qiang; Petit, Arnaud; Bauchet, Anne-Laure; Reinhardt, Dirk; Bornhauser, Beat; Gautheret, Daniel; Lecluse, Yann; Landman-Parker, Judith; Radford, Isabelle; Vainchenker, William; Dastugue, Nicole; de Botton, Stéphane; Dessen, Philippe; Bourquin, Jean-Pierre; Crispino, John D; Ballerini, Paola; Bernard, Olivier A; Pflumio, Françoise; Mercher, Thomas

    2012-10-22

    Acute megakaryoblastic leukemia (AMKL) is a heterogeneous disease generally associated with poor prognosis. Gene expression profiles indicate the existence of distinct molecular subgroups, and several genetic alterations have been characterized in the past years, including the t(1;22)(p13;q13) and the trisomy 21 associated with GATA1 mutations. However, the majority of patients do not present with known mutations, and the limited access to primary patient leukemic cells impedes the efficient development of novel therapeutic strategies. In this study, using a xenotransplantation approach, we have modeled human pediatric AMKL in immunodeficient mice. Analysis of high-throughput RNA sequencing identified recurrent fusion genes defining new molecular subgroups. One subgroup of patients presented with MLL or NUP98 fusion genes leading to up-regulation of the HOX A cluster genes. A novel CBFA2T3-GLIS2 fusion gene resulting from a cryptic inversion of chromosome 16 was identified in another subgroup of 31% of non-Down syndrome AMKL and strongly associated with a gene expression signature of Hedgehog pathway activation. These molecular data provide useful markers for the diagnosis and follow up of patients. Finally, we show that AMKL xenograft models constitute a relevant in vivo preclinical screening platform to validate the efficacy of novel therapies such as Aurora A kinase inhibitors. PMID:23045605

  1. Imaging Tumor Variation in Response to Photodynamic Therapy in Pancreatic Cancer Xenograft Models

    SciTech Connect

    Samkoe, Kimberley S.; Chen, Alina; Rizvi, Imran; O'Hara, Julia A.; Hoopes, P. Jack; Pereira, Stephen P.; Hasan, Tayyaba; Pogue, Brian W.

    2010-01-15

    Purpose: A treatment monitoring study investigated the differential effects of orthotopic pancreatic cancer models in response to interstitial photodynamic therapy (PDT), and the validity of using magnetic resonance imaging as a surrogate measure of response was assessed. Methods and Materials: Different orthotopic pancreatic cancer xenograft models (AsPC-1 and Panc-1) were used to represent the range of pathophysiology observed in human beings. Identical dose escalation studies (10, 20, and 40J/cm) using interstitial verteporfin PDT were performed, and magnetic resonance imaging with T2-weighted and T1-weighted contrast were used to monitor the total tumor volume and the vascular perfusion volume, respectively. Results: There was a significant amount of necrosis in the slower-growing Panc-1 tumor using high light dose, although complete necrosis was not observed. Lower doses were required for the same level of tumor kill in the faster-growing AsPC-1 cell line. Conclusions: The tumor growth rate and vascular pattern of the tumor affect the optimal PDT treatment regimen, with faster-growing tumors being relatively easier to treat. This highlights the fact that therapy in human beings shows a heterogeneous range of outcomes, and suggests a need for careful individualized treatment outcomes assessment in clinical work.

  2. Tryptophan PET Imaging of the Kynurenine Pathway in Patient-Derived Xenograft Models of Glioblastoma.

    PubMed

    Guastella, Anthony R; Michelhaugh, Sharon K; Klinger, Neil V; Kupsky, William J; Polin, Lisa A; Muzik, Otto; Juhász, Csaba; Mittal, Sandeep

    2016-01-01

    Increasing evidence demonstrates the immunosuppressive kynurenine pathway's (KP) role in the pathophysiology of human gliomas. To study the KP in vivo, we used the noninvasive molecular imaging tracer α-[(11)C]-methyl-l-tryptophan (AMT). The AMT-positron emission tomography (PET) has shown high uptake in high-grade gliomas and predicted survival in patients with recurrent glioblastoma (GBM). We generated patient-derived xenograft (PDX) models from dissociated cells, or tumor fragments, from 5 patients with GBM. Mice bearing subcutaneous tumors were imaged with AMT-PET, and tumors were analyzed to detect the KP enzymes indoleamine 2,3-dioxygenase (IDO) 1, IDO2, tryptophan 2,3-dioxygenase, kynureninase, and kynurenine 3-monooxygenase. Overall, PET imaging showed robust tumoral AMT uptake in PDX mice with prolonged tracer accumulation over 60 minutes, consistent with AMT trapping seen in humans. Immunostained tumor tissues demonstrated positive detection of multiple KP enzymes. Furthermore, intracranial implantation of GBM cells was performed with imaging at both 9 and 14 days postimplant, with a marked increase in AMT uptake at 14 days and a corresponding high level of tissue immunostaining for KP enzymes. These results indicate that our PDX mouse models recapitulate human GBM, including aberrant tryptophan metabolism, and offer an in vivo system for development of targeted therapeutics for patients with GBM.

  3. Safety and efficacy of quadrapeutics versus chemoradiation in head and neck carcinoma xenograft model

    PubMed Central

    Lukianova-Hleb, Ekaterina Y; Kim, Yoo-Shin; Aryasomayajula, Bhawani; Boulikas, Teni; Phan, Jack; Hung, Mien-Chie; Torchilin, Vladimir P; O’Neill, Brian E; Lapotko, Dmitri O

    2015-01-01

    Chemoradiation is the strongest anti-tumor therapy but in resistant unresectable cancers it often lacks safety and efficacy. We compared our recently developed cell-level combination approach, quadrapeutics, to chemoradiation therapy to establish pre-clinical data for its biodistribution, safety and efficacy in head and neck squamous cell carcinoma (HNSCC), as a clinically challenging aggressive and resistant cancer. In vitro and in vivo models of four carcinomas were treated with standard chemoradiation and quadrapeutics using identical drug and radiation doses. We applied liposomal cisplatin or doxorubicin, colloidal gold, near-infrared laser pulses and radiation, all at low safe doses. The final evaluation used a xenograft model of HNSCC. Quadrapeutics enhanced standard chemoradiation in vitro by reducing head and neck cancer cell proliferation by 1000-fold, inhibiting tumor growth in vivo by 34-fold and improving animal survival by 5-fold, and reducing the side effects to a negligible level. In quadrapeutics, we observed an “inversion” of the drug efficacy of two standard drugs: doxorubicin, a low efficacy drug for the cancers studied, was two times more efficient than cisplatin, the first choice drug in clinic for HNSCC. The radical therapeutic gain of quadrapeutics resulted from the intracellular synergy of the four components employed which we administered in a specific sequence, while the reduction in the toxicity was due to the low doses of all four components. The biodistribution, safety and efficacy data for quadrapeutics in HNSCC ensure its high translational potential and justify the possibility of clinical trials. PMID:26885444

  4. Differential antitumor activity of aflibercept and bevacizumab in patient-derived xenograft models of colorectal cancer.

    PubMed

    Chiron, Marielle; Bagley, Rebecca G; Pollard, Jack; Mankoo, Parminder K; Henry, Christophe; Vincent, Loïc; Geslin, Catherine; Baltes, Nina; Bergstrom, Donald A

    2014-06-01

    The recombinant fusion protein aflibercept (ziv-aflibercept in the United States) binds VEGF-A, VEGF-B, and placental growth factor (PlGF). The monoclonal antibody bevacizumab binds VEGF-A. Recent studies hypothesized that dual targeting of VEGF/PlGF is more beneficial than targeting either ligand. We compared activity of aflibercept versus bevacizumab in 48 patient-derived xenograft (PDX) colorectal cancer models. Nude mice engrafted subcutaneously with PDX colorectal cancer tumors received biweekly aflibercept, bevacizumab, or vehicle injections. Differential activity between aflibercept and bevacizumab, determined by mouse (m), human (h), VEGF-A, and PlGF levels in untreated tumors, was measured. Aflibercept induced complete tumor stasis in 31 of 48 models and bevacizumab in 2 of 48. Based on statistical analysis, aflibercept was more active than bevacizumab in 39 of 48 models; in 9 of 39 of these models, bevacizumab was considered inactive. In 9 of 48 remaining models, aflibercept and bevacizumab had similar activity. Tumor levels of hVEGF-A (range 776-56,039 pg/mg total protein) were ∼16- to 1,777-fold greater than mVEGF-A (range 8-159 pg/mg total protein). Tumor levels of mPlGF (range 104-1,837 pg/mg total protein) were higher than hPlGF (range 0-543 pg/mg total protein) in 47 of 48 models. Tumor cells were the major source of VEGF; PlGF was primarily produced by tumor stroma. Because tumor levels of hVEGF-A were far greater than mVEGF-A, bevacizumab's inability to bind mVEGF-A is unlikely to explain higher and more consistent aflibercept activity. Neutralizing PlGF and VEGFR-1 activation may be a factor and should be investigated in future studies. In these colorectal cancer PDX models, aflibercept demonstrated greater antitumor activity than bevacizumab.

  5. Establishment and characterization of esophageal squamous cell carcinoma patient-derived xenograft mouse models for preclinical drug discovery.

    PubMed

    Zhang, Jingchuan; Jiang, Dongxian; Li, Xiaojing; Lv, Jing; Xie, Liang; Zheng, Li; Gavine, Paul R; Hu, Qin; Shi, Yuan; Tan, Lijie; Ge, Di; Xu, Songtao; Li, Leon; Zhu, Lifang; Hou, Yingyong; Wang, Qun

    2014-08-01

    The purpose of this study was to establish and characterize patient-derived esophageal squamous cell carcinoma xenograft (PDECX) mice for utilization in antitumor drug discovery. A total of 96 esophageal squamous cell carcinoma (ESCC) tissues from Chinese patients were transplanted subcutaneously into immunodeficient mice. Histology, EGFR, K-ras, B-raf, and PIK3CA mutations, and HER2 gene amplifications were analyzed in both patient tumors and mouse xenograft tissues using immunohistochemistry, mutant-enriched liquid chip sequencing and fluorescence in situ hybridization assays, respectively. Furthermore, in vivo efficacy studies using five PDECX mice harboring a variety of genetic aberrations were performed using the chemotherapy agents 5-fluorouracil (5-FU) and cisplatin. Thirty-seven PDECX mouse models were successfully established in immunodeficient mice. Pathological analysis revealed similar histological architecture and degrees of differentiation between patient ESCC and xenografted tumors. No mutations were identified in EGFR, K-ras, and B-raf genes in either xenograft models or patient ESCC tissues. In contrast, PIK3CA gene mutations were detected in 12.5% (12/96) ESCC patients and 18.9% (7/37) PDECX models. Interestingly, patient ESCC tissues exhibiting HER2 overexpression or gene amplification were unable to survive in immunodeficient mice. Further analysis showed that PDECX models carrying HER2 2+ expression had no response to 5-FU/cisplatin, compared with HER2-negative models. In conclusion, a panel of PDECX mouse models, which include PIK3CA mutant and HER2-positive models, was established and characterized thus mimicking the current clinical genetic setting of esophageal carcinoma. The sensitivity of HER2-negative ESCC models to chemotherapy supports stratification approaches in the treatment of esophageal carcinoma patients and warrants further investigation of the impact of PI3KCA on treatment response.

  6. Patient-derived xenograft mouse models of pseudomyxoma peritonei recapitulate the human inflammatory tumor microenvironment.

    PubMed

    Kuracha, Murali R; Thomas, Peter; Loggie, Brian W; Govindarajan, Venkatesh

    2016-04-01

    Pseudomyxoma peritonei (PMP) is a neoplastic syndrome characterized by peritoneal tumor implants with copious mucinous ascites. The standard of care for PMP patients is aggressive cytoreductive surgery performed in conjunction with heated intraperitoneal chemotherapy. Not all patients are candidates for these procedures and a majority of the patients will have recurrent disease. In addition to secreted mucin, inflammation and fibrosis are central to PMP pathogenesis but the molecular processes that regulate tumor-stromal interactions within the peritoneal tumor microenvironment remain largely unknown. This knowledge is critical not only to elucidate PMP pathobiology but also to identify novel targets for therapy. Here, we report the generation of patient-derived xenograft (PDX) mouse models for PMP and assess the ability of these models to replicate the inflammatory peritoneal microenvironment of human PMP patients. PDX mouse models of low- and high-grade PMP were generated and were of a similar histopathology as human PMP. Cytokines previously shown to be elevated in human PMP were also elevated in PDX ascites. Significant differences in IL-6 and IL-8/KC/MIP2 were seen between human and PDX ascites. Interestingly, these cytokines were mostly secreted by mouse-derived, tumor-associated stromal cells rather than by human-derived PMP tumor cells. Our data suggest that the PMP PDX mouse models are especially suited to the study of tumor-stromal interactions that regulate the peritoneal inflammatory environment in PMP as the tumor and stromal cells in these mouse models are of human and murine origins, respectively. These mouse models are therefore, likely to be useful in vivo surrogates for testing and developing novel therapeutic treatment interventions for PMP.

  7. Comprehensive analysis of leukocytes, vascularization and matrix metalloproteinases in human menstrual xenograft model.

    PubMed

    Guo, Yong; He, Bin; Xu, Xiangbo; Wang, Jiedong

    2011-02-17

    In our previous study, menstrual-like changes in mouse were provoked through the pharmacologic withdrawal of progesterone with mifepristone following induction of decidualization. However, mouse is not a natural menstruation animal, and the menstruation model using external stimuli may not truly reflect the occurrence and development of the human menstrual process. Therefore, we established a model of menstruation based on human endometrial xenotransplantation. In this model, human endometrial tissues were transplanted subcutaneously into SCID mice that were ovarectomized and supplemented with estrogen and progestogen by silastic implants with a scheme imitating the endocrinological milieu of human menstrual cycle. Morphology, hormone levels, and expression of vimentin and cytokeratin markers were evaluated to confirm the menstrual-like changes in this model. With 28 days of hormone treatment, transplanted human endometrium survived and underwent proliferation, differentiation and disintegration, similar to human endometrium in vivo. Human CD45+ cells showed a peak of increase 28 days post-transplantation. Three days after progesterone withdrawal, mouse CD45+ cells increased rapidly in number and were significantly greater than human CD45+ cell counts. Mouse CD31+ blood vascular-like structures were detected in both transplanted and host tissues. After progesterone withdrawal, the expression levels of matrix metalloproteinases (MMP) 1, 2, and 9 were increased. In summary, we successfully established a human endometrial xenotransplantation model in SCID mice, based on the results of menstrual-like changes in which MMP-1, 2 and 9 are involved. We showed that leukocytes are originated from in situ proliferation in human xenografts and involved in the occurrence of menstruation. This model will help to further understand the occurrence, growth, and differentiation of the endometrium and the underlying mechanisms of menstruation.

  8. Xenograft Models of Primary Acute Myeloid Leukemia for the Development of Imaging Strategies and Evaluation of Novel Targeted Therapies.

    PubMed

    Gelebart, Pascal; Popa, Mihaela; McCormack, Emmet

    2016-01-01

    Despite the tremendous progress made in the comprehension of acute myeloid leukemia (AML) over the last 30 years most patients die from their disease. Our understanding of AML has relied on an intensive in-vitro research approach, based on AML cell lines as well as primary AML patient cells. However, experimental insight into the early events of AML leukemogenesis before they become clinically observable is not possible in humans. Thus, preclinical animal models have served the purpose to extend our knowledge of the disease as well as to develop innovative therapeutic strategies. Today, xenograft models using patient-derived neoplastic/leukemia cells represent the strategy of choice for preclinical studies of AML. These models exhibit several key advantages over AML cell lines. In fact, patient-derived cells, in contrast to AML cell lines, encompass the entire complexity of AML disease and can therefore provide more trustworthy results on the efficacy outcome of novel therapies. One other important aspect in the development of xenograft models of AML is the possibility to use imaging techniques to monitor in-vivo the progression of the disease. Imaging techniques also authorize the evaluation of the efficacy of an experimental treatment on tumor growth. This review will focus on the description of xenograft models of AML and will provide researchers and clinicians an overview of how these models have been used for the development of new therapeutic options and new imaging approaches to study AML in-vivo.

  9. Fine-tuning patient-derived xenograft models for precision medicine approaches in leukemia.

    PubMed

    Francis, Olivia L; Milford, Terry-Ann M; Beldiman, Cornelia; Payne, Kimberly J

    2016-03-01

    Many leukemias are characterized by well-known mutations that drive oncogenesis. Mice engineered with these mutations provide a foundation for understanding leukemogenesis and identifying therapies. However, data from whole genome studies provide evidence that malignancies are characterized by multiple genetic alterations that vary between patients, as well as inherited genetic variation that can also contribute to oncogenesis. Improved outcomes will require precision medicine approaches-targeted therapies tailored to malignancies in each patient. Preclinical models that reflect the range of mutations and the genetic background present in patient populations are required to develop and test the combinations of therapies that will be used to provide precision medicine therapeutic strategies. Patient-derived xenografts (PDX) produced by transplanting leukemia cells from patients into immune deficient mice provide preclinical models where disease mechanisms and therapeutic efficacy can be studied in vivo in context of the genetic variability present in patient tumors. PDX models are possible because many elements in the bone marrow microenvironment show cross-species activity between mice and humans. However, several cytokines likely to impact leukemia cells are species-specific with limited activity on transplanted human leukemia cells. In this review we discuss the importance of PDX models for developing precision medicine approaches to leukemia treatment. We illustrate how PDX models can be optimized to overcome a lack of cross-species cytokine activity by reviewing a recent strategy developed for use with a high-risk form of B-cell acute lymphoblastic leukemia (B-ALL) that is characterized by overexpression of CRLF2, a receptor component for the cytokine, TSLP.

  10. Systematic repurposing screening in xenograft models identifies approved drugs with novel anti-cancer activity.

    PubMed

    Roix, Jeffrey J; Harrison, S D; Rainbolt, Elizabeth A; Meshaw, Kathryn R; McMurry, Avery S; Cheung, Peter; Saha, Saurabh

    2014-01-01

    Approved drugs target approximately 400 different mechanisms of action, of which as few as 60 are currently used as anti-cancer therapies. Given that on average it takes 10-15 years for a new cancer therapeutic to be approved, and the recent success of drug repurposing for agents such as thalidomide, we hypothesized that effective, safe cancer treatments may be found by testing approved drugs in new therapeutic settings. Here, we report in-vivo testing of a broad compound collection in cancer xenograft models. Using 182 compounds that target 125 unique target mechanisms, we identified 3 drugs that displayed reproducible activity in combination with the chemotherapeutic temozolomide. Candidate drugs appear effective at dose equivalents that exceed current prescription levels, suggesting that additional pre-clinical efforts will be needed before these drugs can be tested for efficacy in clinical trials. In total, we suggest drug repurposing is a relatively resource-intensive method that can identify approved medicines with a narrow margin of anti-cancer activity.

  11. Monitoring breast tumor progression by photoacoustic measurements: a xenograft mice model study

    NASA Astrophysics Data System (ADS)

    Priya, Mallika; Satish Rao, Bola Sadashiva; Chandra, Subhash; Datta, Anirbit; Nayak, Subramanya G.; Mahato, Krishna Kishore

    2015-10-01

    The current study reports the photoacoustic spectroscopy-based assessment of breast tumor progression in a nude mice xenograft model. The tumor was induced through subcutaneous injection of MCF-7 cells in female nude mice and was monitored for 20 days until the tumor volume reached 1000 mm3. The tumor tissues were extracted at three different time points (days 10, 15, and 20) after tumor inoculation and subjected to photoacoustic spectral recordings in time domain ex vivo at 281 nm pulsed laser excitations. The spectra were converted into the frequency domain using the fast Fourier transformed tools of MATLAB® algorithms and further utilized to extract seven statistical features (mean, median, area under the curve, variance and standard deviation, skewness and kurtosis) from each time point sample to assess the tumor growth with wavelet principal component analysis based logistic regression analysis performed on the data. The prediction accuracies of the analysis for day 10 versus day 15, day 15 versus day 20, and day 10 versus day 20 were found to be 92.31, 87.5, and 95.2%, respectively. Also, receiver operator characteristics area under the curve analysis for day 10 versus day 15, day 15 versus day 20, and day 10 versus day 20 were found to be 0.95, 0.85, and 0.93, respectively. The ability of photoacoustic measurements in the objective assessment of tumor progression has been clearly demonstrated, indicating its clinical potential.

  12. Reversing Cancer Multidrug Resistance in Xenograft Models via Orchestrating Multiple Actions of Functional Mesoporous Silica Nanoparticles.

    PubMed

    Yang, Debin; Wang, Tingfang; Su, Zhigui; Xue, Lingjing; Mo, Ran; Zhang, Can

    2016-08-31

    A multistimuli responsive drug delivery system (DDS) based on sulfhydryl and amino-cofunctionalized mesoporous silica nanoparticles (SH/NH2-MSNs) has been developed, in which the multifunctional hyaluronic acid (HA) derivatives were grafted onto the SH/NH2-MSNs by disulfide bonds for targeting delivery, controlling drug release and reversing multidrug resistance (MDR). The doxorubicin (Dox) loaded multifunctional HA derivatives modified mesoporous silica nanoparticles (Dox/HHS-MSNs) were enzyme and redox sensitive, which could respond to the intracellular stimuli of hyaluronidase (HAase) and glutathione (GSH) successively and prevent drug leakage before reaching the tumor tissues. The cellular uptake experiments showed that Dox/HHS-MSNs were vulnerable to be endocytosed into the Dox-resistant human breast adenocarcinoma (MCF-7/ADR) cells, efficiently realized the endolysosomal escape and remained in the cytoplasm. Because of orchestrating multiple actions above including active targeting, endolysosomal escape and efficient multilevel drug release, Dox/HHS-MSNs could induce the strongest apoptosis and cytotoxicity of MCF-7/ADR cells. Furthermore, a series of in vivo studies on MCF-7/ADR tumor-bearing xenograft mouse models demonstrated that Dox/HHS-MSNs possessed the enhanced tumor-targeting capacity and the best therapeutic efficacy to reverse cancer MDR. PMID:27420116

  13. Mitochondrially targeted wild-type p53 induces apoptosis in a solid human tumor xenograft model

    PubMed Central

    Palacios, Gustavo; Crawford, Howard C.; Vaseva, Angelina; Moll, Ute M.

    2013-01-01

    Classic but also novel roles of p53 are becoming increasingly well characterized. We previously showed that ex vivo retroviral transfer of mitochondrially targeted wild type p53 (mitop53) in the Eμ-myc mouse lymphoma model efficiently induces tumor cell killing in vivo. In an effort to further explore the therapeutic potential of mitop53 for its pro-apoptotic effect in solid tumors, we generated replication-deficient recombinant human Adenovirus type 5 vectors. We show here that adenoviral delivery of mitop53 by intratumoral injection into HCT116 human colon carcinoma xenograft tumors in nude mice is surprisingly effective, resulting in tumor cell death of comparable potency to conventional p53. These apoptotic effects in vivo were confirmed by Ad5-mitop53 mediated cell death of HCT116 cells in culture. Together, these data provide encouragement to further explore the potential for novel mitop53 proteins in cancer therapy to execute the shortest known circuitry of p53 death signaling. PMID:18719383

  14. Application of a patient-derived xenograft model in cytolytic viral activation therapy for nasopharyngeal carcinoma.

    PubMed

    Hsu, Cheng-Lung; Kuo, Yung-Chia; Huang, Yenlin; Huang, Yin-Cheng; Lui, Kar-Wai; Chang, Kai-Ping; Lin, Tung-Liang; Fan, Hsien-Chi; Lin, An-Chi; Hsieh, Chia-Hsun; Lee, Li-Yu; Wang, Hung-Ming; Li, Hsin-Pai; Chang, Yu-Sun

    2015-10-13

    Nasopharyngeal carcinoma (NPC) is an Epstein Barr virus (EBV)-related malignancy in which the tumor microenvironment plays a pivotal role in tumor progression. Here, we developed two patient-derived xenograft (PDX) mouse lines from engrafted NPC metastatic tumors. Positive staining for EBV-encoded small RNAs confirmed that these tumors harbored EBV, and gene expression profile analyses further showed that the PDX was highly similar to the primary parent tumor. In vivo drug screening using the PDX system demonstrated that gemcitabine had the best antitumor effect among the tested drugs. The donor of this PDX also showed excellent responsiveness to gemcitabine treatment. The combination of gemcitabine and valproic acid exerted synergistic antitumor effects. Further addition of ganciclovir to this two-drug combination regimen enhanced cytolytic viral activation, yielding the best antitumor response among tested regimens. Treatment with this three-drug combination regimen decreased plasma EBV-DNA load, tumor viral concentration, and the number of viable tumor cells to a greater extent than the two-drug gemcitabine and valproic acid combination. These results highlight the value of PDX models in the development of EBV-targeted strategies to treat NPC.

  15. Patient-derived xenograft models of breast cancer and their predictive power.

    PubMed

    Whittle, James R; Lewis, Michael T; Lindeman, Geoffrey J; Visvader, Jane E

    2015-02-10

    Despite advances in the treatment of patients with early and metastatic breast cancer, mortality remains high due to intrinsic or acquired resistance to therapy. Increased understanding of the genomic landscape through massively parallel sequencing has revealed somatic mutations common to specific subtypes of breast cancer, provided new prognostic and predictive markers, and highlighted potential therapeutic targets. Evaluating new targets using established cell lines is limited by the inexact correlation between responsiveness observed in cell lines versus that elicited in the patient. Patient-derived xenografts (PDXs) generated from fresh tumor specimens recapitulate the diversity of breast cancer and reflect histopathology, tumor behavior, and the metastatic properties of the original tumor. The high degree of genomic preservation evident across primary tumors and their matching PDXs over serial passaging validate them as important preclinical tools. Indeed, there is accumulating evidence that PDXs can recapitulate treatment responses of the parental tumor. The finding that tumor engraftment is an independent and poor prognostic indicator of patient outcome represents the first step towards personalized medicine. Here we review the utility of breast cancer PDX models to study the clonal evolution of tumors and to evaluate novel therapies and drug resistance.

  16. In vitro cytotoxicity of gold nanorods in A549 cells.

    PubMed

    Tang, Ying; Shen, Yafeng; Huang, Libin; Lv, Gaojian; Lei, Changhai; Fan, Xiaoyan; Lin, Fangxing; Zhang, Yuxia; Wu, Lihui; Yang, Yongji

    2015-03-01

    Gold nanoparticles, which have unique physicochemical characteristics, are being used for an increasingly wide range of applications in biomedical research. In this study, gold nanorods (width of 25 nm, length of 52 nm) were found to be internalized by A549 cells and were primarily localized in the lysosomes and membranous vesicles. The integrity of the membranes of A549 cells exposed to gold nanorods for 4h was damaged, as indicated by laser scanning confocal microscopy (LSCM). Increased lactate dehydrogenase (LDH) leakage and decreased cell viability further indicated the concentration-dependent cytotoxicity of the gold nanorods to the A549 cells. Reactive oxygen species (ROS) production was induced in the A549 cells by the gold nanorods, and this effect was positively correlated with the concentration of the gold nanorods. The results of this study indicated that exposure to gold nanorods caused dose-dependent cytotoxicity in A549 cells and that oxidative stress may be the main factor causing cytotoxicity.

  17. Spheroid culture of LuCaP 136 patient-derived xenograft enables versatile preclinical models of prostate cancer.

    PubMed

    Valta, Maija P; Zhao, Hongjuan; Saar, Matthias; Tuomela, Johanna; Nolley, Rosalie; Linxweiler, Johannes; Sandholm, Jouko; Lehtimäki, Jaakko; Härkönen, Pirkko; Coleman, Ilsa; Nelson, Peter S; Corey, Eva; Peehl, Donna M

    2016-04-01

    LuCaP serially transplantable patient-derived xenografts (PDXs) are valuable preclinical models of locally advanced or metastatic prostate cancer. Using spheroid culture methodology, we recently established cell lines from several LuCaP PDXs. Here, we characterized in depth the features of xenografts derived from LuCaP 136 spheroid cultures and found faithful retention of the phenotype of the original PDX. In vitro culture enabled luciferase transfection into LuCaP 136 spheroids, facilitating in vivo imaging. We showed that LuCaP 136 spheroids formed intratibial, orthotopic, and subcutaneous tumors when re-introduced into mice. Intratibial tumors responded to castration and were highly osteosclerotic. LuCaP 136 is a realistic in vitro-in vivo preclinical model of a subtype of bone metastatic prostate cancer.

  18. Strigolactone analogs act as new anti-cancer agents in inhibition of breast cancer in xenograft model

    PubMed Central

    Mayzlish-Gati, Einav; Laufer, Dana; Grivas, Christopher F; Shaknof, Julia; Sananes, Amiram; Bier, Ariel; Ben-Harosh, Shani; Belausov, Eduard; Johnson, Michael D; Artuso, Emma; Levi, Oshrat; Genin, Ola; Prandi, Cristina; Khalaila, Isam; Pines, Mark; Yarden, Ronit I; Kapulnik, Yoram; Koltai, Hinanit

    2015-01-01

    Strigolactones (SLs) are a novel class of plant hormones. Previously, we found that analogs of SLs induce growth arrest and apoptosis in breast cancer cell lines. These compounds also inhibited the growth of breast cancer stem cell enriched-mammospheres with increased potency. Furthermore, strigolactone analogs inhibited growth and survival of colon, lung, prostate, melanoma, osteosarcoma and leukemia cancer cell lines. To further examine the anti-cancer activity of SLs in vivo, we have examined their effects on growth and viability of MDA-MB-231 tumor xenografts model either alone or in combination with paclitaxel. We show that strigolactone act as new anti-cancer agents in inhibition of breast cancer in xenograft model. In addition we show that SLs affect the integrity of the microtubule network and therefore may inhibit the migratory phenotype of the highly invasive breast cancer cell lines that were examined. PMID:26192476

  19. Design, synthesis, and validation of Axl-targeted monoclonal antibody probe for microPET imaging in human lung cancer xenograft.

    PubMed

    Liu, Shuanglong; Li, Dan; Guo, Jiacong; Canale, Nicolette; Li, Xiuqing; Liu, Ren; Krasnoperov, Valery; Gill, Parkash S; Conti, Peter S; Shan, Hong; Li, Zibo

    2014-11-01

    Accumulating experimental evidence indicates that overexpression of the oncogenic receptor tyrosine kinase, Axl, plays a key role in the tumorigenesis and metastasis of various types of cancer. The objective of this study is to design a novel imaging probe based on the monoclonal antibody, h173, for microPET imaging of Axl expression in human lung cancer. A bifunctional chelator, DOTA, was conjugated to h173, followed by radiolabeling with (64)Cu. The binding of DOTA-h173 to the Axl receptor was first evaluated by a cell uptake assay and flow cytometry analysis using human lung cancer cell lines. The probe (64)Cu-DOTA-h173 was further evaluated by microPET imaging, and ex vivo histology studies in the Axl-positive A549 tumors. In vitro cellular study showed that Axl probe, (64)Cu-DOTA-h173, was highly immuno-reactive with A549 cells. Western blot analysis confirmed that Axl is highly expressed in the A549 cell line. For microPET imaging, the A549 xenografts demonstrated a significantly higher (64)Cu-DOTA-h173 uptake compared to the NCI-H249 xenograft (a negative control model). Furthermore, (64)Cu-DOTA-h173 uptake in A549 is significantly higher than that of (64)Cu-DOTA-hIgG. Immuno-fluorescence staining was consistent with the in vivo micro-PET imaging results. In conclusion, (64)Cu-DOTA-h173 could be potentially used as a probe for noninvasive imaging of Axl expression, which could collect important information regarding tumor response to Axl-targeted therapeutic interventions.

  20. Combination of Vandetanib, Radiotherapy, and Irinotecan in the LoVo Human Colorectal Cancer Xenograft Model

    SciTech Connect

    Wachsberger, Phyllis; Burd, Randy; Ryan, Anderson; Daskalakis, Constantine; Dicker, Adam P.

    2009-11-01

    Purpose: The tumor growth kinetics of the human LoVo colorectal xenograft model was assessed in response to vandetanib, an orally available receptor tyrosine kinase inhibitor, radiotherapy (RT), or irinotecan (CPT-11), as single therapies and in combination. Methods and Materials: LoVo cells were injected subcutaneously into the right hind limb (5x10{sup 6} cells in 100muL phosphate-buffered saline) of athymic NCR NUM mice and tumors were grown to a volume of 200-300 mm{sup 3} before treatment. Vandetanib was administered at 50 mg/kg daily orally for 14 days starting on Day 1. RT was given as three fractions (3x3 Gy) on Days 1, 2, and 3. CPT-11 was given at 15 mg/kg intraperitoneally on Days 1 and 3. Tumor volumes were measured on a daily basis and calculated by measuring tumor diameters with digital calipers in two orthogonal dimensions. Results: All three single treatments (vandetanib, CPT-11, and radiation) significantly slowed LoVo colorectal tumor growth. Vandetanib significantly increased the antitumor effects of CPT-11 and radiation when given in combination with either of these treatments. These treatment combinations resulted in a slow tumor growth rate during the 2 weeks of vandetanib administration. The triple combination of vandetanib, CPT-11, and radiation produced the most marked improvement in response as observed by measurable shrinkage of tumors during the first week of treatment. Conclusions: The tumor growth delay kinetics observed in this study of the LoVo colorectal model suggest concurrent and sustained post-sequencing of vandetanib with cytotoxic therapy may be beneficial in tumors of this type.

  1. Novel CHOP activator LGH00168 induces necroptosis in A549 human lung cancer cells via ROS-mediated ER stress and NF-κB inhibition

    PubMed Central

    Ma, Yi-ming; Peng, Yan-min; Zhu, Qiong-hua; Gao, An-hui; Chao, Bo; He, Qiao-jun; Li, Jia; Hu, You-hong; Zhou, Yu-bo

    2016-01-01

    Aim: C/EBP homologous protein (CHOP) is a transcription factor that is activated at multiple levels during ER stress and plays an important role in ER stress-induced apoptosis. In this study we identified a novel CHOP activator, and further investigated its potential to be a therapeutic agent for human lung cancer. Methods: HEK293-CHOP-luc reporter cells were used in high-throughput screening (HTS) to identify CHOP activators. The cytotoxicity against cancer cells in vitro was measured with MTT assay. The anticancer effects were further examined in A549 human non-small cell lung cancer xenograft mice. The mechanisms underlying CHOP activation were analyzed using luciferase assays, and the anticancer mechanisms were elucidated in A549 cells. Results: From chemical libraries of 50 000 compounds, LGH00168 was identified as a CHOP activator, which showed cytotoxic activities against a panel of 9 cancer cell lines with an average IC50 value of 3.26 μmol/L. Moreover, administration of LGH00168 significantly suppressed tumor growth in A549 xenograft bearing mice. LGH00168 activated CHOP promoter via AARE1 and AP1 elements, increased DR5 expression, decreased Bcl-2 expression, and inhibited the NF-κB pathway. Treatment of A549 cells with LGH00168 (10 μmol/L) did not induce apoptosis, but lead to RIP1-dependent necroptosis, accompanied by cell swelling, plasma membrane rupture, lysosomal membrane permeabilization, MMP collapse and caspase 8 inhibition. Furthermore, LGH00168 (10 and 20 μmol/L) dose-dependently induced mito-ROS production in A549 cells, which was reversed by the ROS scavenger N-acetyl-L-cysteine (NAC, 10 mmol/L). Moreover, NAC significantly diminished LGH00168-induced CHOP activation, NF-κB inhibition and necroptosis in A549 cells. Conclusion: LGH00168 is a CHOP activator that inhibits A549 cell growth in vitro and lung tumor growth in vivo. PMID:27264312

  2. Pulsed high-intensity focused ultrasound therapy enhances targeted delivery of cetuximab to colon cancer xenograft model in mice.

    PubMed

    Park, Min Jung; Kim, Young-Sun; Yang, Jehoon; Sun, Woo Chul; Park, Hajan; Chae, Sun Young; Namgung, Mi-Sun; Choi, Kyu-Sil

    2013-02-01

    Our aim was to evaluate whether pulsed high-intensity focused ultrasound (HIFU) therapy enhances the effect of an epidermal growth factor receptor-targeted chemotherapeutic drug, cetuximab, in treating human colon cancer xenografts in a mouse model. Balb/c nude mice with subcutaneous xenografts of HT-29 cells were randomly categorized into control (n = 9), pulsed HIFU alone (n = 10), cetuximab monotherapy (n = 8) or combined pulsed HIFU and cetuximab therapy (n = 9) group. Cetuximab, pulsed HIFU therapy, or both were administered three times per week starting from day 8 after tumor cell injection. Based on tumor growth curves up to 34 days, the combination therapy group showed more suppressed tumor growth than all other groups (p < 0.05). The final relative tumor volumes were 5.4 ± 2.1, 5.2 ± 1.3, 4.8 ± 1.8, and 3.1 ± 0.9 for control, pulsed HIFU alone, cetuximab monotherapy, and combination therapy groups, respectively. In conclusion, pulsed HIFU therapy appears to enhance the anti-tumor effect of epidermal growth factor receptor-targeted cetuximab on human colon cancer xenograft models in mice. PMID:23219035

  3. Biodistribution of charged F(ab')2 photoimmunoconjugates in a xenograft model of ovarian cancer.

    PubMed

    Duska, L R; Hamblin, M R; Bamberg, M P; Hasan, T

    1997-01-01

    The effect of charge modification of photoimmunoconjugates (PICs) on their biodistribution in a xenograft model of ovarian cancer was investigated. Chlorin(e6)c(e6) was attached site specifically to the F(ab')2 fragment of the murine monoclonal antibody OC125, directed against human ovarian cancer cells, via poly-1-lysine linkers carrying cationic or anionic charges. Preservation of immunoreactivity was checked by enzyme-linked immunosorbent assay (ELISA). PICs were radiolabelled with 125I and compared with non-specific rabbit IgG PICs after intraperitoneal (i.p.) injection into nude mice. Samples were taken from normal organs and tumour at 3 h and 24 h. Tumour to normal 125I ratios showed that the cationic OC125F(ab')2 PIC had the highest tumour selectivity. Ratios for c(e6) were uniformly higher than for 125I, indicating that c(e6) became separated from 125I. OC125F(ab')2 gave highest tissue values of 125I, followed by cationic OC125F(ab')2 PIC; other species were much lower. The amounts of c(e6) delivered per gram of tumour were much higher for cationic OC125F(ab')2 PIC than for other species. The results indicate that cationic charge stimulates the endocytosis and lysosomal degradation of the OC125F(ab')2-pl-c(e6) that has bound to the i.p. tumour. Positively charged PICs may have applications in the i.p. photoimmunotherapy of minimal residual ovarian cancer. PMID:9062404

  4. Preclinical evaluation of transcriptional targeting strategy for human hepatocellular carcinoma in an orthotopic xenograft mouse model.

    PubMed

    Sia, Kian Chuan; Huynh, Hung; Chung, Alexander Yaw Fui; Ooi, London Lucien Peng Jin; Lim, Kiat Hon; Hui, Kam Man; Lam, Paula Yeng Po

    2013-08-01

    Gene regulation of many key cell-cycle players in S-, G(2) phase, and mitosis results from transcriptional repression in their respective promoter regions during the G(0) and G(1) phases of cell cycle. Within these promoter regions are phylogenetically conserved sequences known as the cell-cycle-dependent element (CDE) and cell-cycle genes homology regions (CHR) sites. Thus, we hypothesize that transcriptional regulation of cell-cycle regulation via the CDE/CHR region together with liver-specific apolipoprotein E (apoE)-hAAT promoter could bring about a selective transgene expression in proliferating human hepatocellular carcinoma. We show that the newly generated vector AH-6CC-L2C could mediate hepatocyte-targeted luciferase gene expression in tumor cells and freshly isolated short-term hepatocellular carcinoma cultures from patient biopsy. In contrast, normal murine and human hepatocytes infected with AH-6CC-L2C expressed minimal or low luciferase activities. In the presence of prodrug 5-fluorocytosine (5-FC), AH-6CC-L2C effectively suppressed the growth of orthotopic hepatocellular carcinoma patient-derived xenograft mouse model via the expression of yeast cytosine deaminase (yCD) that converts 5-FC to anticancer metabolite 5-fluoruracil. More importantly, we show that combination treatment of AH-6CC-L2C with an EZH2 inhibitor, DZNep, that targets EpCAM-positive hepatocellular carcinoma, can bring about a greater therapeutic efficacy compared with a single treatment of virus or inhibitor. Our study showed that targeting proliferating human hepatocellular carcinoma cells through the transcriptional control of therapeutic gene could represent a feasible approach against hepatocellular carcinoma.

  5. Endogenous retrovirus induces leukemia in a xenograft mouse model for primary myelofibrosis

    PubMed Central

    Triviai, Ioanna; Ziegler, Marion; Bergholz, Ulla; Oler, Andrew J.; Stübig, Thomas; Prassolov, Vladimir; Fehse, Boris; Kozak, Christine A.; Kröger, Nicolaus; Stocking, Carol

    2014-01-01

    The compound immunodeficiencies in nonobese diabetic (NOD) inbred mice homozygous for the Prkdcscid and Il2rgnull alleles (NSG mice) permit engraftment of a wide-range of primary human cells, enabling sophisticated modeling of human disease. In studies designed to define neoplastic stem cells of primary myelofibrosis (PMF), a myeloproliferative neoplasm characterized by profound disruption of the hematopoietic microenvironment, we observed a high frequency of acute myeloid leukemia (AML) in NSG mice. AML was of mouse origin, confined to PMF-xenografted mice, and contained multiple clonal integrations of ecotropic murine leukemia virus (E-MuLV). Significantly, MuLV replication was not only observed in diseased mice, but also in nontreated NSG controls. Furthermore, in addition to the single ecotropic endogenous retrovirus (eERV) located on chromosome 11 (Emv30) in the NOD genome, multiple de novo germ-line eERV integrations were observed in mice from each of four independent NSG mouse colonies. Analysis confirmed that E-MuLV originated from the Emv30 provirus and that recombination events were not necessary for virus replication or AML induction. Pathogenicity is thus likely attributable to PMF-mediated paracrine stimulation of mouse myeloid cells, which serve as targets for retroviral infection and transformation, as evidenced by integration into the Evi1 locus, a hotspot for retroviral-induced myeloid leukemia. This study thus corroborates a role of paracrine stimulation in PMF disease progression, underlines the importance of target cell type and numbers in MuLV-induced disease, and mandates awareness of replicating MuLV in NOD immunodeficient mice, which can significantly influence experimental results and their interpretation. PMID:24912157

  6. Endogenous retrovirus induces leukemia in a xenograft mouse model for primary myelofibrosis.

    PubMed

    Triviai, Ioanna; Ziegler, Marion; Bergholz, Ulla; Oler, Andrew J; Stübig, Thomas; Prassolov, Vladimir; Fehse, Boris; Kozak, Christine A; Kröger, Nicolaus; Stocking, Carol

    2014-06-10

    The compound immunodeficiencies in nonobese diabetic (NOD) inbred mice homozygous for the Prkdc(scid) and Il2rg(null) alleles (NSG mice) permit engraftment of a wide-range of primary human cells, enabling sophisticated modeling of human disease. In studies designed to define neoplastic stem cells of primary myelofibrosis (PMF), a myeloproliferative neoplasm characterized by profound disruption of the hematopoietic microenvironment, we observed a high frequency of acute myeloid leukemia (AML) in NSG mice. AML was of mouse origin, confined to PMF-xenografted mice, and contained multiple clonal integrations of ecotropic murine leukemia virus (E-MuLV). Significantly, MuLV replication was not only observed in diseased mice, but also in nontreated NSG controls. Furthermore, in addition to the single ecotropic endogenous retrovirus (eERV) located on chromosome 11 (Emv30) in the NOD genome, multiple de novo germ-line eERV integrations were observed in mice from each of four independent NSG mouse colonies. Analysis confirmed that E-MuLV originated from the Emv30 provirus and that recombination events were not necessary for virus replication or AML induction. Pathogenicity is thus likely attributable to PMF-mediated paracrine stimulation of mouse myeloid cells, which serve as targets for retroviral infection and transformation, as evidenced by integration into the Evi1 locus, a hotspot for retroviral-induced myeloid leukemia. This study thus corroborates a role of paracrine stimulation in PMF disease progression, underlines the importance of target cell type and numbers in MuLV-induced disease, and mandates awareness of replicating MuLV in NOD immunodeficient mice, which can significantly influence experimental results and their interpretation.

  7. Generation of Pediatric Leukemia Xenograft Models in NSG-B2m Mice: Comparison with NOD/SCID Mice.

    PubMed

    Gopalakrishnapillai, Anilkumar; Kolb, E Anders; Dhanan, Priyanka; Bojja, Aruna Sri; Mason, Robert W; Corao, Diana; Barwe, Sonali P

    2016-01-01

    Generation of orthotopic xenograft mouse models of leukemia is important to understand the mechanisms of leukemogenesis, cancer progression, its cross talk with the bone marrow microenvironment, and for preclinical evaluation of drugs. In these models, following intravenous injection, leukemic cells home to the bone marrow and proliferate there before infiltrating other organs, such as spleen, liver, and the central nervous system. Moreover, such models have been shown to accurately recapitulate the human disease and correlate with patient response to therapy and prognosis. Thus, various immune-deficient mice strains have been used with or without recipient preconditioning to increase engraftment efficiency. Mice homozygous for the severe combined immune deficiency (SCID) mutation and with non-obese diabetic background (NOD/SCID) have been used in the majority of leukemia xenograft studies. Later, NOD/SCID mice deficient for interleukin 2 receptor gamma chain (IL2Rγ) gene called NSG mice became the model of choice for leukemia xenografts. However, engraftment of leukemia cells without irradiation preconditioning still remained a challenge. In this study, we used NSG mice with null alleles for major histocompatibility complex class I beta2-microglobulin (β2m) called NSG-B2m. This is a first report describing the 100% engraftment efficiency of pediatric leukemia cell lines and primary samples in NSG-B2m mice in the absence of host preconditioning by sublethal irradiation. We also show direct comparison of the engraftment efficiency and growth rate of pediatric acute leukemia cells in NSG-B2m and NOD/SCID mice, which showed 80-90% engraftment efficiency. Secondary and tertiary xenografts in NSG-B2m mice generated by injection of cells isolated from the spleens of leukemia-bearing mice also behaved similar to the primary patient sample. We have successfully engrafted 25 acute lymphoblastic leukemia (ALL) and 5 acute myeloid leukemia (AML) patient samples with

  8. Generation of Pediatric Leukemia Xenograft Models in NSG-B2m Mice: Comparison with NOD/SCID Mice.

    PubMed

    Gopalakrishnapillai, Anilkumar; Kolb, E Anders; Dhanan, Priyanka; Bojja, Aruna Sri; Mason, Robert W; Corao, Diana; Barwe, Sonali P

    2016-01-01

    Generation of orthotopic xenograft mouse models of leukemia is important to understand the mechanisms of leukemogenesis, cancer progression, its cross talk with the bone marrow microenvironment, and for preclinical evaluation of drugs. In these models, following intravenous injection, leukemic cells home to the bone marrow and proliferate there before infiltrating other organs, such as spleen, liver, and the central nervous system. Moreover, such models have been shown to accurately recapitulate the human disease and correlate with patient response to therapy and prognosis. Thus, various immune-deficient mice strains have been used with or without recipient preconditioning to increase engraftment efficiency. Mice homozygous for the severe combined immune deficiency (SCID) mutation and with non-obese diabetic background (NOD/SCID) have been used in the majority of leukemia xenograft studies. Later, NOD/SCID mice deficient for interleukin 2 receptor gamma chain (IL2Rγ) gene called NSG mice became the model of choice for leukemia xenografts. However, engraftment of leukemia cells without irradiation preconditioning still remained a challenge. In this study, we used NSG mice with null alleles for major histocompatibility complex class I beta2-microglobulin (β2m) called NSG-B2m. This is a first report describing the 100% engraftment efficiency of pediatric leukemia cell lines and primary samples in NSG-B2m mice in the absence of host preconditioning by sublethal irradiation. We also show direct comparison of the engraftment efficiency and growth rate of pediatric acute leukemia cells in NSG-B2m and NOD/SCID mice, which showed 80-90% engraftment efficiency. Secondary and tertiary xenografts in NSG-B2m mice generated by injection of cells isolated from the spleens of leukemia-bearing mice also behaved similar to the primary patient sample. We have successfully engrafted 25 acute lymphoblastic leukemia (ALL) and 5 acute myeloid leukemia (AML) patient samples with

  9. Generation of Pediatric Leukemia Xenograft Models in NSG-B2m Mice: Comparison with NOD/SCID Mice

    PubMed Central

    Gopalakrishnapillai, Anilkumar; Kolb, E. Anders; Dhanan, Priyanka; Bojja, Aruna Sri; Mason, Robert W.; Corao, Diana; Barwe, Sonali P.

    2016-01-01

    Generation of orthotopic xenograft mouse models of leukemia is important to understand the mechanisms of leukemogenesis, cancer progression, its cross talk with the bone marrow microenvironment, and for preclinical evaluation of drugs. In these models, following intravenous injection, leukemic cells home to the bone marrow and proliferate there before infiltrating other organs, such as spleen, liver, and the central nervous system. Moreover, such models have been shown to accurately recapitulate the human disease and correlate with patient response to therapy and prognosis. Thus, various immune-deficient mice strains have been used with or without recipient preconditioning to increase engraftment efficiency. Mice homozygous for the severe combined immune deficiency (SCID) mutation and with non-obese diabetic background (NOD/SCID) have been used in the majority of leukemia xenograft studies. Later, NOD/SCID mice deficient for interleukin 2 receptor gamma chain (IL2Rγ) gene called NSG mice became the model of choice for leukemia xenografts. However, engraftment of leukemia cells without irradiation preconditioning still remained a challenge. In this study, we used NSG mice with null alleles for major histocompatibility complex class I beta2-microglobulin (β2m) called NSG-B2m. This is a first report describing the 100% engraftment efficiency of pediatric leukemia cell lines and primary samples in NSG-B2m mice in the absence of host preconditioning by sublethal irradiation. We also show direct comparison of the engraftment efficiency and growth rate of pediatric acute leukemia cells in NSG-B2m and NOD/SCID mice, which showed 80–90% engraftment efficiency. Secondary and tertiary xenografts in NSG-B2m mice generated by injection of cells isolated from the spleens of leukemia-bearing mice also behaved similar to the primary patient sample. We have successfully engrafted 25 acute lymphoblastic leukemia (ALL) and 5 acute myeloid leukemia (AML) patient samples with

  10. Knock down of the dual functional protein apurinic /apyrimidinic endonuclease 1 enhances the killing effect of hematoporphrphyrin derivative-mediated photodynamic therapy on non-small cell lung cancer cells in vitro and in a xenograft model.

    PubMed

    Yang, Zhen-Zhou; Li, Meng-Xia; Zhang, Yun-Song; Xiang, De-Bing; Dai, Nan; Zeng, Lin-Li; Li, Zeng-Peng; Wang, Ge; Wang, Dong

    2010-01-01

    Photodynamic therapy (PDT) is considered to be effective treatment for many cancers including lung cancer, head and neck cancers, and prostate cancer. It uses the combination of nontoxic photosensitizers and harmless visible light to generate reactive oxygen species and kill cells. However, DNA repair and reactive oxygen species-induced signaling pathway activation play crucial roles in cellular response to PDT and may also result in therapeutic limitation of PDT. To improve the cancer therapeutic efficacy of PDT, we targeted apurinic/apyrimidinic endonuclease (APE1), which is essential for both DNA repair and redox regulation of gene transcription, as a potential candidate for PDT combined gene therapy. In our study, an adenovirus-mediated APE1 silencing strategy was introduced to test its therapeutic enhancement for the non-small cell lung cancer cell line A549 both in vitro and in vivo after hematoporphrphyrin derivative (HpD)-mediated PDT. The adenovirus vector Ad5/F35-shAPE1 was validated to significantly suppress the protein expression of APE1 in cultured A549 cell and in its xenograft of nude mice. Ad5/F35-shAPE1 effectively inhibited APE1 protein upregulation induced by PDT and resulted in an increase in A549 cell killing by photoirradiation compared with the hematoporphrphyrin derivative-PDT alone group. Ad5/F35-shAPE1 suppressed the DNA repair capacity for single-strand breaks and abolished the activation of some stress-related transcription factors such as hypoxia-induced factor (HIF)-1 that consequently lead to increased cell apoptosis after PDT. Additionally, knock down of APE1 enhanced the tumor suppression efficacy of PDT on the A549 xenograft. Our study indicated that APE1-targeted gene therapy combined with PDT is a promising strategy for enhancement of the efficacy of PDT in treatment of non-small cell lung cancer.

  11. Xenograft models for undifferentiated pleomorphic sarcoma not otherwise specified are essential for preclinical testing of therapeutic agents

    PubMed Central

    Becker, Marc; Graf, Claudine; Tonak, Marcus; Radsak, Markus P.; Bopp, Tobias; Bals, Robert; Bohle, Rainer M.; Theobald, Matthias; Rommens, Pol-Maria; Proschek, Dirk; Wehler, Thomas C.

    2016-01-01

    Undifferentiated pleomorphic sarcoma not otherwise specified belongs to the heterogeneous group of soft tissue tumors. It is preferentially located in the upper and lower extremities of the body, and surgical resection remains the only curative treatment. Preclinical animal models are crucial to improve the development of novel chemotherapeutic agents for the treatment of undifferentiated pleomorphic sarcoma. However, this approach has been hampered by the lack of reproducible animal models. The present study established two xenograft animal models generated from stable non-clonal cell cultures, and investigated the difference in chemotherapeutic effects on tumor growth between undifferentiated pleomorphic sarcoma in vivo and in vitro. The cell cultures were generated from freshly isolated tumor tissues of two patients with undifferentiated pleomorphic sarcoma. For the in vivo analysis, these cells were injected subcutaneously into immunodeficient mice. The mice were monitored for tumor appearance and treated with the most common or innovative chemotherapeutic agents available to date. Furthermore, the same drugs were administered to in vitro cell cultures. The most effective tumor growth inhibition in vitro was observed with doxorubicin and the histone deacetylase inhibitor suberoylanilide hydroxamic acid (SAHA), also known as vorinostat. In the in vivo xenograft mouse model, the combination of doxorubicin and the tyrosine kinase inhibitor pazopanib induced a significant tumor reduction. By contrast, treatment with vorinostat did not reduce the tumor growth. Taken together, the results obtained from drug testing in vitro differed significantly from the in vivo results. Therefore, the novel and reproducible xenograft animal model established in the present study demonstrated that in vivo models are required to test potential chemotherapeutic agents for the treatment of undifferentiated pleomorphic sarcoma prior to clinical use, since animal models are more similar

  12. Comparative efficacy of 177Lu and 90Y for Anti-CD20 Pretargeted Radioimmunotherapy in Murine Lymphoma Xenograft Models

    DOE PAGES

    Frost, Sofia H. L.; Frayo, Shani L.; Miller, Brian W.; Orozco, Johnnie J.; Booth, Garrett C.; Hylarides, Mark D.; Lin, Yukang; Green, Damian J.; Gopal, Ajay K.; Pagel, John M.; et al

    2015-03-18

    approaches in these human lymphoma xenograft models.« less

  13. Comparative Efficacy of 177Lu and 90Y for Anti-CD20 Pretargeted Radioimmunotherapy in Murine Lymphoma Xenograft Models

    PubMed Central

    Frost, Sofia H. L.; Frayo, Shani L.; Miller, Brian W.; Orozco, Johnnie J.; Booth, Garrett C.; Hylarides, Mark D.; Lin, Yukang; Green, Damian J.; Gopal, Ajay K.; Pagel, John M.; Bäck, Tom A.; Fisher, Darrell R.; Press, Oliver W.

    2015-01-01

    these human lymphoma xenograft models. PMID:25785845

  14. Novel Dedifferentiated Liposarcoma Xenograft Models Reveal PTEN Down-Regulation as a Malignant Signature and Response to PI3K Pathway Inhibition

    PubMed Central

    Smith, Kathleen B.; Tran, Linh M.; Tam, Brenna M.; Shurell, Elizabeth M.; Li, Yunfeng; Braas, Daniel; Tap, William D.; Christofk, Heather R.; Dry, Sarah M.; Eilber, Fritz C.; Wu, Hong

    2014-01-01

    Liposarcoma is a type of soft tissue sarcoma that exhibits poor survival and a high recurrence rate. Treatment is generally limited to surgery and radiation, which emphasizes the need for better understanding of this disease. Because very few in vivo and in vitro models can reproducibly recapitulate the human disease, we generated several xenograft models from surgically resected human dedifferentiated liposarcoma. All xenografts recapitulated morphological and gene expression characteristics of the patient tumors after continuous in vivo passages. Importantly, xenograftability was directly correlated with disease-specific survival of liposarcoma patients. Thus, the ability for the tumor of a patient to engraft may help identify those patients who will benefit from more aggressive treatment regimens. Gene expression analyses highlighted the association between xenograftability and a unique gene expression signature, including down-regulated PTEN tumor-suppressor gene expression and a progenitor-like phenotype. When treated with the PI3K/AKT/mTOR pathway inhibitor rapamycin alone or in combination with the multikinase inhibitor sorafenib, all xenografts responded with increased lipid content and a more differentiated gene expression profile. These human xenograft models may facilitate liposarcoma research and accelerate the generation of readily translatable preclinical data that could ultimately influence patient care. PMID:23416162

  15. Intratumoral injection of taxol in vivo suppresses A549 tumor showing cytoplasmic vacuolization.

    PubMed

    Wang, Chaoyang; Chen, Tongsheng

    2012-04-01

    Based on our recent in vitro studies, this report was designed to explore the mechanism by which high concentration of taxol (70 µM) induced paraptosis-like cell death in human lung carcinoma (A549) cells, and to evaluate the therapeutic efficacy of taxol using A549 tumor-bearing mice in vivo. Exposure of cells to taxol induced time-dependent cytotoxicity and cytoplasmic vacuolization without the involvement of Bax, Bak, Mcl-1, Bcl-XL, and caspase-3. Although taxol treatment induced activating transcription factor 6 (ATF6) cleavage indicative of endoplasmic reticulum (ER) stress, silencing ATF6 by shATF6 did not prevent taxol-induced both cytotoxcity and cytoplasmic vacuolization, suggesting that taxol-induced cytoplasmic vacuolization and cell death were not due to ER stress. Moreover, taxol-treated cells did not show DNA fragmentation and loss of mitochondrial membrane potential, the typical characteristics of apoptosis. In addition, taxol-induced cytoplasmic vacuolization did not show the cellular lysis, the characteristics of oncosis, and positive of β-galactosidase, the characteristic of senescence, indicating that taxol induced paraptosis-like cell death is neither oncosis nor senescence. Moreover, our in vivo data showed that intratumoral injection of taxol (50 mg/kg) in A549 tumor xenograft mice on day 1 and day 19 potently suppressed tumor growth showing significant ER vacuolization without toxicity. In conclusion, high concentration of taxol exhibits a significant anticancer activity by inducing paraptosis-like cell death in vitro and in vivo, without significant toxicity, suggesting a promising therapeutic strategy for apoptosis-resistance cancer by inducing ER vacuolization.

  16. Appropriateness of Using Patient-Derived Xenograft Models for Pharmacologic Evaluation of Novel Therapies for Esophageal/Gastro-Esophageal Junction Cancers

    PubMed Central

    Dodbiba, Lorin; Teichman, Jennifer; Fleet, Andrew; Thai, Henry; Starmans, Maud H. W.; Navab, Roya; Chen, Zhuo; Girgis, Hala; Eng, Lawson; Espin-Garcia, Osvaldo; Shen, Xiaowei; Bandarchi, Bizhan; Schwock, Joerg; Tsao, Ming-Sound; El-Zimaity, Hala; Der, Sandy D.; Xu, Wei; Bristow, Robert G.; Darling, Gail E.; Boutros, Paul C.

    2015-01-01

    The high morbidity and mortality of patients with esophageal (E) and gastro-esophageal junction (GEJ) cancers, warrants new pre-clinical models for drug testing. The utility of primary tumor xenografts (PTXGs) as pre-clinical models was assessed. Clinicopathological, immunohistochemical markers (p53, p16, Ki-67, Her-2/neu and EGFR), and global mRNA abundance profiles were evaluated to determine selection biases of samples implanted or engrafted, compared with the underlying population. Nine primary E/GEJ adenocarcinoma xenograft lines were further characterized for the spectrum and stability of gene/protein expression over passages. Seven primary esophageal adenocarcinoma xenograft lines were treated with individual or combination chemotherapy. Tumors that were implanted (n=55) in NOD/SCID mice had features suggestive of more aggressive biology than tumors that were never implanted (n=32). Of those implanted, 21/55 engrafted; engraftment was associated with poorly differentiated tumors (p=0.04) and older patients (p=0.01). Expression of immunohistochemical markers were similar between patient sample and corresponding xenograft. mRNA differences observed between patient tumors and first passage xenografts were largely due to loss of human stroma in xenografts. mRNA patterns of early vs late passage xenografts and of small vs large tumors of the same passage were similar. Complete resistance was present in 2/7 xenografts while the remaining tumors showed varying degrees of sensitivity, that remained constant across passages. Because of their ability to recapitulate primary tumor characteristics during engraftment and across serial passaging, PTXGs can be useful clinical systems for assessment of drug sensitivity of human E/GEJ cancers. PMID:25826681

  17. Correlation between tumor engraftment in patient-derived xenograft models and clinical outcomes in colorectal cancer patients.

    PubMed

    Oh, Bo Young; Lee, Woo Yong; Jung, Sungwon; Hong, Hye Kyung; Nam, Do-Hyun; Park, Yoon Ah; Huh, Jung Wook; Yun, Seong Hyeon; Kim, Hee Cheol; Chun, Ho-Kyung; Cho, Yong Beom

    2015-06-30

    Despite numerous studies involving patient-derived xenograft (PDX) models, few studies have investigated the relationship between the ability of the tumor to engraft (tumorigenicity) and the clinical features of colorectal cancer (CRC). The aim of this study was to determine whether tumorigenicity correlates with clinical outcomes of CRC patients. We included 241 CRC patients who underwent radical surgery from 2010 to 2013. PDX models were established by implanting tumor fragments obtained from these patients into the subcutaneous layer of immunodeficient mice. Xenografts were successfully established from 62.2%. Successful engraftment was associated with advanced stage (p < 0.001) and moderate/poor differentiation (p = 0.029). Three-year disease-free survival (DFS) rates were lower for patients with tumorigenicity (p = 0.011). In stage III patients, tumorigenicity was an independent predictor of poor DFS (p = 0.034). In addition, mutation of TP53 was most frequently detected in stage III patients with tumorigenicity. Two models of stage IV disease without KRAS mutations showed high sensitivity to EGFR-targeted agents, while none of the models with KRAS mutations showed high sensitivity. In conclusion, PDX models may provide an effective preclinical tool for predicting cancer progression and could be used to further genomic and pharmacologic research on personalized treatments.

  18. Single-agent cytarabine is insufficient for the treatment of human mantle cell lymphoma in mouse xenograft model.

    PubMed

    Klanova, M; Soukup, T; Molinsky, J; Lateckova, L; Vockova, P; Alam, M; Zivny, J; Trneny, M; Klener, P

    2016-01-01

    Mantle cell lymphoma is an aggressive type of B-cell non-Hodgkin lymphoma with adverse prognosis. It was demonstrated that alternation of CHOP and DHAP chemotherapy improved outcome of mantle cell lymphoma patients. However, which components of DHAP, cisplatin, cytarabine, or both, were responsible for the improved outcome remained unclear. To answer this question, antitumor efficacies of equally toxic doses of cytarabine, cisplatin, and three different combinations were compared in vivo using mouse xenograft models of mantle cell lymphoma. We demonstrated that cisplatin, alone or with cytarabine, is significantly superior to single-agent cytarabine in both eliminating lymphoma cells and suppressing their proliferation rate. PMID:27468882

  19. Xenograft and genetically engineered mouse model systems of osteosarcoma and Ewing's sarcoma: tumor models for cancer drug discovery

    PubMed Central

    Sampson, Valerie B; Kamara, Davida F; Kolb, E Anders

    2014-01-01

    Introduction There are > 75 histological types of solid tumors that are classified into two major groups: bone and soft-tissue sarcomas. These diseases are more prevalent in children, and pediatric sarcomas tend to be highly aggressive and rapidly progressive. Sarcomas in adults may follow a more indolent course, but aggressive tumors are also common. Sarcomas that are metastatic at diagnosis, or recurrent following therapy, remain refractory to current treatment options with dismal overall survival rates. A major focus of clinical trials, for patients with sarcoma, is to identify novel and more effective therapeutic strategies targeted to genomic or proteomic aberrations specific to the malignant cells. Critical to the understanding of the potential for targeted therapies are models of disease that are representative of clinical disease and predictive of relevant clinical responses. Areas covered In this article, the authors discuss the use of mouse xenograft models and genetically engineered mice in cancer drug discovery. The authors provide a special focus on models for the two most common bone sarcomas: osteosarcoma (OS) and Ewing's sarcoma (ES). Expert opinion Predicting whether a new anticancer agent will have a positive therapeutic index in patients with OS and ES remains a challenge. The use of mouse sarcoma models for understanding the mechanisms involved in the response of tumors to new treatments is an important step in the process of drug discovery and the development of clinically relevant therapeutic strategies for these diseases. PMID:23844615

  20. Efficacy of an Fc-modified anti-CD123 antibody (CSL362) combined with chemotherapy in xenograft models of acute myelogenous leukemia in immunodeficient mice

    PubMed Central

    Lee, Erwin M.; Yee, Dean; Busfield, Samantha J.; McManus, Julie F.; Cummings, Nik; Vairo, Gino; Wei, Andrew; Ramshaw, Hayley S.; Powell, Jason A.; Lopez, Angel F.; Lewis, Ian D.; McCall, Martin N.; Lock, Richard B.

    2015-01-01

    The prognosis of older patients with acute myelogenous leukemia is generally poor. The interleukin-3 receptor α-chain (CD123) is highly expressed on the surface of acute leukemia cells compared with normal hematopoietic stem cells. CSL362 is a fully humanized, CD123-neutralizing monoclonal antibody containing a modified Fc structure, which enhances human natural killer cell antibody-dependent cell-mediated cytotoxicity. Six continuous acute myelogenous leukemia xenografts established from patient explants and characterized by cell and molecular criteria, produced progressively lethal disease 42-202 days after transplantation. CSL362 alone reduced engraftment of one of four and three of four acute myelogenous leukemia xenografts in the bone marrow and peripheral organs, respectively. A cytarabine and daunorubicin regimen was optimized using this model to identify potentially synergistic interactions with CSL362. Cytarabine/daunorubicin improved the survival of mice engrafted with four of four acute myelogenous leukemia xenografts by 31–41 days. Moreover, CSL362 extended the survival of cytarabine/daunorubicin-treated mice for two of two acute myelogenous leukemia xenografts, while augmentation of natural killer cell-deficient NSG mice with adoptively transferred human natural killer cells improved survival against a single xenograft. Interestingly, this enhanced CSL362 efficacy was lost in the absence of chemotherapy. This study shows that acute myelogenous leukemia xenografts provide a platform for the evaluation of new therapeutics, simulating complex in vivo interactions, and that the in vivo efficacy of CSL362 supports continued clinical development of this drug. PMID:26130514

  1. Application of Coiled Coil Peptides in Liposomal Anticancer Drug Delivery Using a Zebrafish Xenograft Model.

    PubMed

    Yang, Jian; Shimada, Yasuhito; Olsthoorn, René C L; Snaar-Jagalska, B Ewa; Spaink, Herman P; Kros, Alexander

    2016-08-23

    The complementary coiled coil forming peptides E4 [(EIAALEK)4] and K4 [(KIAALKE)4] are known to trigger liposomal membrane fusion when tethered to lipid vesicles in the form of lipopeptides. In this study, we examined whether these coiled coil forming peptides can be used for drug delivery applications. First, we prepared E4 peptide modified liposomes containing the far-red fluorescent dye TO-PRO-3 iodide (E4-Lipo-TP3) and confirmed that E4-liposomes could deliver TP3 into HeLa cells expressing K4 peptide on the membrane (HeLa-K) under cell culture conditions in a selective manner. Next, we prepared doxorubicin-containing E4-liposomes (E4-Lipo-DOX) and confirmed that E4-liposomes could also deliver DOX into HeLa-K cells. Moreover, E4-Lipo-DOX showed enhanced cytotoxicity toward HeLa-K cells compared to free doxorubicin. To prove the suitability of E4/K4 coiled coil formation for in vivo drug delivery, we injected E4-Lipo-TP3 or E4-Lipo-DOX into zebrafish xenografts of HeLa-K. As a result, E4-liposomes delivered TP3 to the implanted HeLa-K cells, and E4-Lipo-DOX could suppress cancer proliferation in the xenograft when compared to nontargeted conditions (i.e., zebrafish xenograft with free DOX injection). These data demonstrate that coiled coil formation enables drug selectivity and efficacy in vivo. It is envisaged that these findings are a step forward toward biorthogonal targeting systems as a tool for clinical drug delivery. PMID:27504667

  2. Development and analysis of patient-derived xenograft mouse models in intravascular large B-cell lymphoma.

    PubMed

    Shimada, K; Shimada, S; Sugimoto, K; Nakatochi, M; Suguro, M; Hirakawa, A; Hocking, T D; Takeuchi, I; Tokunaga, T; Takagi, Y; Sakamoto, A; Aoki, T; Naoe, T; Nakamura, S; Hayakawa, F; Seto, M; Tomita, A; Kiyoi, H

    2016-07-01

    Intravascular large B-cell lymphoma (IVLBCL) is a distinct disease entity with the peculiar characteristic that tumor cells proliferate within vessels. Despite recent advances in understanding the disease from clinical aspects, the underlying pathogenesis remains unknown. Here we demonstrate analyses of IVLBCL biology using four xenograft mouse models established from primary IVLBCL samples. In all four models, the main characteristic of IVLBCL tumor cell proliferation within vessels was retained. Time-lapse engraftment analyses revealed that the tumor cells initially engrafted and proliferated in the sinusoids and vessels in the liver and then engrafted and proliferated in multiple organs. Intriguingly, serial passage of tumor cells from the adrenal gland of a transplanted mouse developed from primary patient bone marrow cells into a second mouse showed that the tumor cells mainly distributed into the adrenal gland in the second mouse, implying the existence of clonal selection and/or evolution at engraftment of a specific organ. Gene expression profiling analyses demonstrated that the gene set associated with cell migration was enriched for normal peripheral blood B cells, indicating that inhibition of cell migration might be involved in IVLBCL pathogenesis. In conclusion, the mouse xenograft models described here are essential tools for uncovering IVLBCL biology.

  3. Intrahepatic Xenograft of Cutaneous T-Cell Lymphoma Cell Lines: A Useful Model for Rapid Biological and Therapeutic Evaluation.

    PubMed

    Andrique, Laetitia; Poglio, Sandrine; Prochazkova-Carlotti, Martina; Kadin, Marshall Edward; Giese, Alban; Idrissi, Yamina; Beylot-Barry, Marie; Merlio, Jean-Philippe; Chevret, Edith

    2016-07-01

    Cutaneous T-cell lymphomas (CTCLs) are a heterogeneous group of diseases primarily involving the skin that could have an aggressive course with circulating blood cells, especially in Sézary syndrome and transformed mycosis fungoides. So far, few CTCL cell lines have been adapted for in vivo experiments and their tumorigenicity has not been adequately assessed, hampering the use of a reproducible model for CTCL biological evaluation. In fact, both patient-derived xenografts and cell line xenografts at subcutaneous sites failed to provide a robust tool, because engraftment was dependent on mice strain and cell line subtype. Herein, we describe an original method of intrahepatic injection into adult NOD.Cg-Prkdc(scid)Il2rg(tm1Wjl)/SzJ mice liver of both aggressive (My-La, HUT78, HH, MAC2A, and MAC2B) and indolent (FE-PD and MAC1) CTCL cell lines. Six of the seven CTCL cell lines were grafted with a high rate of success (80%). Moreover, this model provided a quick (15 days) and robust assay for in vivo evaluation of CTCL cell lines tumorigenicity and therapeutic response in preclinical studies. Such a reproducible model can be therefore used for further functional studies and in vivo drug testing. PMID:27181405

  4. Pseudotyped AAV Vector-Mediated Gene Transfer in a Human Fetal Trachea Xenograft Model: Implications for In Utero Gene Therapy for Cystic Fibrosis

    PubMed Central

    Leung, Alice; Katz, Anna B.; Lim, Foong-Yen; Habli, Mounira; Jones, Helen N.; Wilson, James M.; Crombleholme, Timothy M.

    2012-01-01

    Background Lung disease including airway infection and inflammation currently causes the majority of morbidities and mortalities associated with cystic fibrosis (CF), making the airway epithelium and the submucosal glands (SMG) novel target cells for gene therapy in CF. These target cells are relatively inaccessible to postnatal gene transfer limiting the success of gene therapy. Our previous work in a human-fetal trachea xenograft model suggests the potential benefit for treating CF in utero. In this study, we aim to validate adeno-associated virus serotype 2 (AAV2) gene transfer in a human fetal trachea xenograft model and to compare transduction efficiencies of pseudotyping AAV2 vectors in fetal xenografts and postnatal xenograft controls. Methodology/Principal Findings Human fetal trachea or postnatal bronchus controls were xenografted onto immunocompromised SCID mice for a four-week engraftment period. After injection of AAV2/2, 2/1, 2/5, 2/7 or 2/8 with a LacZ reporter into both types of xenografts, we analyzed for transgene expression in the respiratory epithelium and SMGs. At 1 month, transduction by AAV2/2 and AAV2/8 in respiratory epithelium and SMG cells was significantly greater than that of AAV2/1, 2/5, and 2/7 in xenograft tracheas. Efficiency in SMG transduction was significantly greater in AAV2/8 than AAV2/2. At 3 months, AAV2/2 and AAV2/8 transgene expression was >99% of respiratory epithelium and SMG. At 1 month, transduction efficiency of AAV2/2 and AAV2/8 was significantly less in adult postnatal bronchial xenografts than in fetal tracheal xenografts. Conclusions/Significance Based on the effectiveness of AAV vectors in SMG transduction, our findings suggest the potential utility of pseudotyped AAV vectors for treatment of cystic fibrosis. The human fetal trachea xenograft model may serve as an effective tool for further development of fetal gene therapy strategies for the in utero treatment of cystic fibrosis. PMID:22937069

  5. The use of thermographic imaging to evaluate therapeutic response in human tumour xenograft models.

    PubMed

    Hussain, Nosheen; Connah, David; Ugail, Hassan; Cooper, Patricia A; Falconer, Robert A; Patterson, Laurence H; Shnyder, Steven D

    2016-01-01

    Non-invasive methods to monitor tumour growth are an important goal in cancer drug development. Thermographic imaging systems offer potential in this area, since a change in temperature is known to be induced due to changes within the tumour microenvironment. This study demonstrates that this imaging modality can be applied to a broad range of tumour xenografts and also, for the first time, the methodology's suitability to assess anti-cancer agent efficacy. Mice bearing subcutaneously implanted H460 lung cancer xenografts were treated with a novel vascular disrupting agent, ICT-2552, and the cytotoxin doxorubicin. The effects on tumour temperature were assessed using thermographic imaging over the first 6 hours post-administration and subsequently a further 7 days. For ICT-2552 a significant initial temperature drop was observed, whilst for both agents a significant temperature drop was seen compared to controls over the longer time period. Thus thermographic imaging can detect functional differences (manifesting as temperature reductions) in the tumour response to these anti-cancer agents compared to controls. Importantly, these effects can be detected in the first few hours following treatment and therefore the tumour is observable non-invasively. As discussed, this technique will have considerable 3Rs benefits in terms of reduction and refinement of animal use. PMID:27491535

  6. The use of thermographic imaging to evaluate therapeutic response in human tumour xenograft models

    PubMed Central

    Hussain, Nosheen; Connah, David; Ugail, Hassan; Cooper, Patricia A.; Falconer, Robert A.; Patterson, Laurence H.; Shnyder, Steven D.

    2016-01-01

    Non-invasive methods to monitor tumour growth are an important goal in cancer drug development. Thermographic imaging systems offer potential in this area, since a change in temperature is known to be induced due to changes within the tumour microenvironment. This study demonstrates that this imaging modality can be applied to a broad range of tumour xenografts and also, for the first time, the methodology’s suitability to assess anti-cancer agent efficacy. Mice bearing subcutaneously implanted H460 lung cancer xenografts were treated with a novel vascular disrupting agent, ICT-2552, and the cytotoxin doxorubicin. The effects on tumour temperature were assessed using thermographic imaging over the first 6 hours post-administration and subsequently a further 7 days. For ICT-2552 a significant initial temperature drop was observed, whilst for both agents a significant temperature drop was seen compared to controls over the longer time period. Thus thermographic imaging can detect functional differences (manifesting as temperature reductions) in the tumour response to these anti-cancer agents compared to controls. Importantly, these effects can be detected in the first few hours following treatment and therefore the tumour is observable non-invasively. As discussed, this technique will have considerable 3Rs benefits in terms of reduction and refinement of animal use. PMID:27491535

  7. RAD001 (everolimus) inhibits tumour growth in xenograft models of human hepatocellular carcinoma.

    PubMed

    Huynh, Hung; Chow, K H Pierce; Soo, Khee Chee; Toh, Han Chong; Choo, Su Pin; Foo, Kian Fong; Poon, Donald; Ngo, Van Chanh; Tran, Evelyn

    2009-07-01

    Hepatocellular carcinoma (HCC) is the fifth most common malignancy worldwide and highly resistant to available chemotherapies. Mammalian target of rapamycin (mTOR) functions to regulate protein translation, angiogenesis and cell cycle progression in many cancers including HCC. In the present study, subcutaneous patient-derived HCC xenografts were used to study the effects of an mTOR inhibitor, RAD001 (everolimus), on tumour growth, apoptosis and angiogenesis. We report that oral administration of RAD001 to mice bearing patient-derived HCC xenografts resulted in a dose-dependent inhibition of tumour growth. RAD001-induced growth suppression was associated with inactivation of downstream targets of mTOR, reduction in VEGF expression and microvessel density, inhibition of cell proliferation, up-regulation of p27(Kip1) and down-regulation of p21(Cip1/Waf1), Cdk-6, Cdk-2, Cdk-4, cdc-25C, cyclin B1 and c-Myc. Our data indicate that the mTOR pathway plays an important role in angiogenesis, cell cycle progression and proliferation of liver cancer cells. Our study provides a strong rationale for clinical investigation of mTOR inhibitor RAD001 in patients with HCC.

  8. Activin type IB receptor signaling in prostate cancer cells promotes lymph node metastasis in a xenograft model

    SciTech Connect

    Nomura, Masatoshi; Tanaka, Kimitaka; Wang, Lixiang; Goto, Yutaka; Mukasa, Chizu; Ashida, Kenji; Takayanagi, Ryoichi

    2013-01-04

    Highlights: Black-Right-Pointing-Pointer ActRIB signaling induces Snail and S100A4 expressions in prostate cancer cells. Black-Right-Pointing-Pointer The prostate cancer cell lines expressing an active form of ActRIB were established. Black-Right-Pointing-Pointer ActRIB signaling promotes EMT and lymph node metastasis in xenograft model. -- Abstract: Activin, a member of the transforming growth factor-{beta} family, has been known to be a growth and differentiating factor. Despite its pluripotent effects, the roles of activin signaling in prostate cancer pathogenesis are still unclear. In this study, we established several cell lines that express a constitutive active form of activin type IB receptor (ActRIBCA) in human prostate cancer cells, ALVA41 (ALVA-ActRIBCA). There was no apparent change in the proliferation of ALVA-ActRIBCA cells in vitro; however, their migratory ability was significantly enhanced. In a xenograft model, histological analysis revealed that the expression of Snail, a cell-adhesion-suppressing transcription factor, was dramatically increased in ALVA-ActRIBCA tumors, indicating epithelial mesenchymal transition (EMT). Finally, mice bearing ALVA-ActRIBCA cells developed multiple lymph node metastases. In this study, we demonstrated that ActRIBCA signaling can promote cell migration in prostate cancer cells via a network of signaling molecules that work together to trigger the process of EMT, and thereby aid in the aggressiveness and progression of prostate cancers.

  9. Therapeutic Effects of Microbubbles Added to Combined High-Intensity Focused Ultrasound and Chemotherapy in a Pancreatic Cancer Xenograft Model

    PubMed Central

    Yu, Mi Hye; Kim, Hae Ri; Kim, Bo Ram; Park, Eun-Joo; Kim, Hoe Suk; Han, Joon Koo; Choi, Byung Ihn

    2016-01-01

    Objective To investigate whether high-intensity focused ultrasound (HIFU) combined with microbubbles enhances the therapeutic effects of chemotherapy. Materials and Methods A pancreatic cancer xenograft model was established using BALB/c nude mice and luciferase-expressing human pancreatic cancer cells. Mice were randomly assigned to five groups according to treatment: control (n = 10), gemcitabine alone (GEM; n = 12), HIFU with microbubbles (HIFU + MB, n = 11), combined HIFU and gemcitabine (HIGEM; n = 12), and HIGEM + MB (n = 13). After three weekly treatments, apoptosis rates were evaluated using the terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling assay in two mice per group. Tumor volume and bioluminescence were monitored using high-resolution 3D ultrasound imaging and in vivo bioluminescence imaging for eight weeks in the remaining mice. Results The HIGEM + MB group showed significantly higher apoptosis rates than the other groups (p < 0.05) and exhibited the slowest tumor growth. From week 5, the tumor-volume-ratio relative to the baseline tumor volume was significantly lower in the HIGEM + MB group than in the control, GEM, and HIFU + MB groups (p < 0.05). Despite visible distinction, the HIGEM and HIGEM + MB groups showed no significant differences. Conclusion High-intensity focused ultrasound combined with microbubbles enhances the therapeutic effects of gemcitabine chemotherapy in a pancreatic cancer xenograft model. PMID:27587968

  10. Desferrioxamine enhances AIDS-associated Kaposi's sarcoma tumor development in a xenograft model.

    PubMed

    Simonart, Thierry; Boelaert, Johan R; Andrei, Graciela; van den Oord, Joost J; Degraef, Chantale; Hermans, Philippe; Noel, Jean-Christophe; Van Vooren, Jean-Paul; Heenen, Michel; De Clercq, Erik; Snoeck, Robert

    2002-07-10

    Iron is suspected to be involved in the induction and/or progression of various human tumors. More particularly, we have previously shown that iron may be involved in the pathogenesis of Kaposi's sarcoma (KS). We have also shown that the iron chelator desferrioxamine (DFO) has a potent anti-KS activity in vitro, suggesting that it may represent a potential therapeutic approach for the treatment of KS. The present study was designed to investigate the effect of DFO on the growth of human KS xenografts in immunodeficient mice. Unexpectedly, we found that mice treated with DFO (400 mg/kg, 3 times weekly) (n = 30) exhibited a marked enhancement of tumor growth compared with control mice (n = 33) (230 +/- 134 mm(2) versus 143 +/- 70 mm p < 0.01). No enhancement of tumor growth was seen in mice treated with iron-saturated DFO. At least 2 findings suggest that this paradoxic pro-KS activity occurred independently of mice iron stores. First, treatment with DFO had only a marginal effect on ferritin and hematocrit levels. Second, induction of effective iron depletion by an iron-poor diet (6.7 mg iron/kg diet) (n = 23) did not have a deleterious effect on the growth of the KS xenografts. The lesions obtained from the DFO-treated animals exhibited a significantly decreased apoptotic index (p < 0.05), indicating that some antiapoptotic mechanism induced by DFO may be operating in vivo to favour tumor growth. In conclusion, our data show that DFO has a stimulatory effect on KS growth in immunodeficient mice, suggesting that this drug is not indicated in patients with KS.

  11. Transferrin Receptor Targeted Lipopolyplexes for Delivery of Antisense Oligonucleotide G3139 in a Murine K562 Xenograft Model

    PubMed Central

    Zhang, Xulang; Koh, Chee Guan; Yu, Bo; Liu, Shujun; Piao, Longzhu; Marcucci, Guido; Lee, Robert J.; Lee, L. James

    2013-01-01

    Purpose Transferrin (Tf) conjugated lipopolyplexes (LPs) carrying G3139, an antisense oligonucleotide for Bcl-2, were synthesized and evaluated in Tf receptor positive K562 erythroleukemia cells and then in a murine K562 xenograft model. Materials and Methods Particle size and Zeta potentials of transferrin conjugated lipopolyplexs containing G3139 (Tf-LP-G3139) were measured by Dynamic Light Scattering and ZetaPALS. In vitro and in vivo sample’s Bcl-2 downregulation was analyzed using Western blot and tumor tissue samples also exhibited by immunohistochemistry method. For athymic mice bearing with K562 xenograft tumors, tumor growth inhibition and survival rate were investigated. Nanoparticle distribution in 3-D cell cluster was observed by Laser scan confocal microscopy. IL-12 production in the plasma was measured by ELISA kit. Results In vitro, Tf-LP-G3139 was more effective in inducing down regulation of Bcl-2 in K562 cells than non-targeted LP-G3139, free G3139 and mismatched control ODN-G4126 in the same formulation. In vivo Tf-LP-G3139 was less effective than free G3139 in Bcl-2 down regulation. 3-D cell cluster model diffusion results indeed indicated limited penetration of the LPs into the cell cluster. Finally, the therapeutic efficacies of Tf-LP-G3139 and free G3139 were determined in the K562 xenograft model. Tf-LP-G3139 showed slower plasma clearance, higher AUC, and greater accumulation in the tumor compared to free G3139. In addition, Tf-LP-G3139 was found to be more effective in tumor growth inhibition and prolonging mouse survival than free G3139. This was associated with increased spleen weight and IL-12 production in the plasma. Conclusion The role of the immune system in the therapeutic response obtained with the Tf-LPs is necessary and in vitro 3-D cell cluster model can be a potential tool to evaluate the nanoparticle distribution. PMID:19291371

  12. Whole Exome Sequencing of Rapid Autopsy Tumors and Xenograft Models Reveals Possible Driver Mutations Underlying Tumor Progression.

    PubMed

    Xie, Tao; Musteanu, Monica; Lopez-Casas, Pedro P; Shields, David J; Olson, Peter; Rejto, Paul A; Hidalgo, Manuel

    2015-01-01

    Pancreatic Ductal Adenocarcinoma (PDAC) is a highly lethal malignancy due to its propensity to invade and rapidly metastasize and remains very difficult to manage clinically. One major hindrance towards a better understanding of PDAC is the lack of molecular data sets and models representative of end stage disease. Moreover, it remains unclear how molecularly similar patient-derived xenograft (PDX) models are to the primary tumor from which they were derived. To identify potential molecular drivers in metastatic pancreatic cancer progression, we obtained matched primary tumor, metastases and normal (peripheral blood) samples under a rapid autopsy program and performed whole exome sequencing (WES) on tumor as well as normal samples. PDX models were also generated, sequenced and compared to tumors. Across the matched data sets generated for three patients, there were on average approximately 160 single-nucleotide mutations in each sample. The majority of mutations in each patient were shared among the primary and metastatic samples and, importantly, were largely retained in the xenograft models. Based on the mutation prevalence in the primary and metastatic sites, we proposed possible clonal evolution patterns marked by functional mutations affecting cancer genes such as KRAS, TP53 and SMAD4 that may play an important role in tumor initiation, progression and metastasis. These results add to our understanding of pancreatic tumor biology, and demonstrate that PDX models derived from advanced or end-stage likely closely approximate the genetics of the disease in the clinic and thus represent a biologically and clinically relevant pre-clinical platform that may enable the development of effective targeted therapies for PDAC.

  13. Celecoxib enhanced the cytotoxic effect of cisplatin in chemo-resistant gastric cancer xenograft mouse models through a cyclooxygenase-2-dependent manner.

    PubMed

    Xu, Hong-Bin; Shen, Fu-Ming; Lv, Qian-Zhou

    2016-04-01

    Our previous study suggested that co-administration of celecoxib increased chemo-sensitivity of multidrug-resistant human gastric cancer SGC-7901/DDP cells to cisplatin (DDP) in vitro. The present study was designed to investigate whether celecoxib had the similar activities in vivo. SGC-7901/DDP and SGC-7901 xenograft mouse models were established. At the end of the experiment, cisplatin treatment alone significantly inhibited tumor growth in SGC-7901 xenograft, as compared with that in SGC-7901/DDP xenograft, suggesting that it maintained cisplatin sensitivity. When cisplatin and celecoxib were co-administrated, their antitumor activities were augmented in SGC-7901/DDP xenograft. The levels of Ki67 and PCNA after combination therapy were significantly decreased in SGC-7901/DDP xenograft, as compared with those of cisplatin treatment alone. Moreover, examining the apoptotic index by TUNEL assay showed similar results. Further studies demonstrated the inhibitory effect of celecoxib on cyclooxygenase-2 and P-glycoprotein expression was the possible reason to increase sensitivity of SGC-7901/DDP cells to cisplatin in vivo. However, the ratio of thromboxane B2 and prostaglandin F1α was elevated after celecoxib treatment in mice. This has been proposed to increase the risk of thrombogenesis. Further studies are required to evaluate the efficacy and safety of celecoxib for reducing chemo-resistance in gastric cancer. PMID:26879869

  14. PEG-liposomal oxaliplatin potentialization of antitumor efficiency in a nude mouse tumor-xenograft model of colorectal carcinoma.

    PubMed

    Yang, Chuang; Liu, Hai-Zhong; Lu, Wei-Dong; Fu, Zhong-Xue

    2011-06-01

    The non-selectivity of chemotherapeutics between normal tissue and pathological sites poses a challenge for the treatment strategy for advanced colorectal carcinoma. To obtain sufficient antitumor activity, optimization of the therapeutic regimen is of great importance. We investigated PEG-liposomal oxaliplatin potentialization of antitumor efficiency in a nude mouse tumor-xenograft model of colorectal carcinoma. A tumor-bearing nude mouse model, intravenous injections of (Dio)-labeled PEG-liposomes via tail vein and fluorescence imaging with in vivo imaging system were employed. Mice were treated with free L-oHP, PEG-liposomal L-oHP via the tail vein, followed by analysis of the accumulation of L-oHP in tumor tissues by high-performance liquid chromatography (HPLC), observation of the tumor volume and the survival rate. Apoptosis and proliferation of tumors were detected by TUNEL assay and immunohistochemistry. The mRNA and protein levels of Bcl-2, Bax, caspase-3 (P17) and Ki-67 were determined by RT-PCR and Western blotting. Fluorescence imaging with in vivo imaging showed PEG-liposome targeting in tumor tissues. After intravenous injections of PEG-liposomal oxaliplatin, tumor tissue maximum accumulation of L-oHP was 9.37 ± 0.79 µg/g at 24 h; The tumor volume was significantly suppressed, and mice showed longer survival, compared with the free oxaliplatin group. Apoptosis increased, but proliferation decreased in tumor tissues. The mRNA expression of Bcl-2 and Ki-67 was down-regulated, while Bax and caspase-3 expression was up-regulated. Protein expression of Bcl-2 was down-regulated, while Bax and P17 expression was up-regulated. The results indicate that PEG-liposomal oxaliplatin can improve antitumor efficiency in a nude mouse tumor-xenograft model of colorectal carcinoma.

  15. Aminomethylphosphonic acid inhibits growth and metastasis of human prostate cancer in an orthotopic xenograft mouse model

    PubMed Central

    Parajuli, Keshab Raj; Zhang, Qiuyang; Liu, Sen; You, Zongbing

    2016-01-01

    Aminomethylphosphonic acid (AMPA) has been shown to inhibit prostate cancer cell growth in vitro. The purpose of the present study was to determine if AMPA could inhibit growth and metastasis of prostate cancer in vivo. Human prostate cancer PC-3-LacZ-luciferase cells were implanted into the ventral lateral lobes of the prostate in 39 athymic Nu/Nu nude male mice. Seven days later, mice were randomized into the control group (n = 14, treated intraperitoneally with phosphate buffered saline), low dose group (n = 10, treated intraperitoneally with AMPA at 400 mg/kg body weight/day), and high dose group (n = 15, treated intraperitoneally with AMPA at 800 mg/kg body weight/day). Tumor growth and metastasis were examined every 4-7 days by bioluminescence imaging of live mice. We found that AMPA treatment significantly inhibited growth and metastasis of orthotopic xenograft prostate tumors and prolonged the survival time of the mice. AMPA treatment decreased expression of BIRC2 and activated caspase 3, leading to increased apoptosis in the prostate tumors. AMPA treatment decreased expression of cyclin D1. AMPA treatment also reduced angiogenesis in the prostate tumors. Taken together, these results demonstrate that AMPA can inhibit prostate cancer growth and metastasis, suggesting that AMPA may be developed into a therapeutic agent for the treatment of prostate cancer. PMID:26840261

  16. A Novel Synthetic Smoothened Antagonist Transiently Inhibits Pancreatic Adenocarcinoma Xenografts in a Mouse Model

    PubMed Central

    Strand, Martin F.; Wilson, Steven R.; Dembinski, Jennifer L.; Holsworth, Daniel D.; Khvat, Alexander; Okun, Ilya; Petersen, Dirk; Krauss, Stefan

    2011-01-01

    Background Hedgehog (Hh) signaling is over-activated in several solid tumors where it plays a central role in cell growth, stroma recruitment and tumor progression. In the Hh signaling pathway, the Smoothened (SMO) receptor comprises a primary drug target with experimental small molecule SMO antagonists currently being evaluated in clinical trials. Principal Findings Using Shh-Light II (Shh-L2) and alkaline phosphatase (AP) based screening formats on a “focused diversity” library we identified a novel small molecule inhibitor of the Hh pathway, MS-0022 (2-bromo-N-(4-(8-methylimidazo[1,2-a]pyridin-2-yl)phenyl)benzamide). MS-0022 showed effective Hh signaling pathway inhibition at the level of SMO in the low nM range, and Hh pathway inhibition downstream of Suppressor of fused (SUFU) in the low µM range. MS-0022 reduced growth in the tumor cell lines PANC-1, SUIT-2, PC-3 and FEMX in vitro. MS-0022 treatment led to a transient delay of tumor growth that correlated with a reduction of stromal Gli1 levels in SUIT-2 xenografts in vivo. Significance We document the in vitro and in vivo efficacy and bioavailability of a novel small molecule SMO antagonist, MS-0022. Although MS-0022 primarily interferes with Hh signaling at the level of SMO, it also has a downstream inhibitory effect and leads to a stronger reduction of growth in several tumor cell lines when compared to related SMO antagonists. PMID:21698280

  17. Whole transcriptome profiling of patient-derived xenograft models as a tool to identify both tumor and stromal specific biomarkers.

    PubMed

    Bradford, James R; Wappett, Mark; Beran, Garry; Logie, Armelle; Delpuech, Oona; Brown, Henry; Boros, Joanna; Camp, Nicola J; McEwen, Robert; Mazzola, Anne Marie; D'Cruz, Celina; Barry, Simon T

    2016-04-12

    The tumor microenvironment is emerging as a key regulator of cancer growth and progression, however the exact mechanisms of interaction with the tumor are poorly understood. Whilst the majority of genomic profiling efforts thus far have focused on the tumor, here we investigate RNA-Seq as a hypothesis-free tool to generate independent tumor and stromal biomarkers, and explore tumor-stroma interactions by exploiting the human-murine compartment specificity of patient-derived xenografts (PDX).Across a pan-cancer cohort of 79 PDX models, we determine that mouse stroma can be separated into distinct clusters, each corresponding to a specific stromal cell type. This implies heterogeneous recruitment of mouse stroma to the xenograft independent of tumor type. We then generate cross-species expression networks to recapitulate a known association between tumor epithelial cells and fibroblast activation, and propose a potentially novel relationship between two hypoxia-associated genes, human MIF and mouse Ddx6. Assessment of disease subtype also reveals MMP12 as a putative stromal marker of triple-negative breast cancer. Finally, we establish that our ability to dissect recruited stroma from trans-differentiated tumor cells is crucial to identifying stem-like poor-prognosis signatures in the tumor compartment.In conclusion, RNA-Seq is a powerful, cost-effective solution to global analysis of human tumor and mouse stroma simultaneously, providing new insights into mouse stromal heterogeneity and compartment-specific disease markers that are otherwise overlooked by alternative technologies. The study represents the first comprehensive analysis of its kind across multiple PDX models, and supports adoption of the approach in pre-clinical drug efficacy studies, and compartment-specific biomarker discovery. PMID:26980748

  18. Whole transcriptome profiling of patient-derived xenograft models as a tool to identify both tumor and stromal specific biomarkers

    PubMed Central

    Bradford, James R.; Wappett, Mark; Beran, Garry; Logie, Armelle; Delpuech, Oona; Brown, Henry; Boros, Joanna; Camp, Nicola J.; McEwen, Robert; Mazzola, Anne Marie; D'Cruz, Celina; Barry, Simon T.

    2016-01-01

    The tumor microenvironment is emerging as a key regulator of cancer growth and progression, however the exact mechanisms of interaction with the tumor are poorly understood. Whilst the majority of genomic profiling efforts thus far have focused on the tumor, here we investigate RNA-Seq as a hypothesis-free tool to generate independent tumor and stromal biomarkers, and explore tumor-stroma interactions by exploiting the human-murine compartment specificity of patient-derived xenografts (PDX). Across a pan-cancer cohort of 79 PDX models, we determine that mouse stroma can be separated into distinct clusters, each corresponding to a specific stromal cell type. This implies heterogeneous recruitment of mouse stroma to the xenograft independent of tumor type. We then generate cross-species expression networks to recapitulate a known association between tumor epithelial cells and fibroblast activation, and propose a potentially novel relationship between two hypoxia-associated genes, human MIF and mouse Ddx6. Assessment of disease subtype also reveals MMP12 as a putative stromal marker of triple-negative breast cancer. Finally, we establish that our ability to dissect recruited stroma from trans-differentiated tumor cells is crucial to identifying stem-like poor-prognosis signatures in the tumor compartment. In conclusion, RNA-Seq is a powerful, cost-effective solution to global analysis of human tumor and mouse stroma simultaneously, providing new insights into mouse stromal heterogeneity and compartment-specific disease markers that are otherwise overlooked by alternative technologies. The study represents the first comprehensive analysis of its kind across multiple PDX models, and supports adoption of the approach in pre-clinical drug efficacy studies, and compartment-specific biomarker discovery. PMID:26980748

  19. Whole transcriptome profiling of patient-derived xenograft models as a tool to identify both tumor and stromal specific biomarkers.

    PubMed

    Bradford, James R; Wappett, Mark; Beran, Garry; Logie, Armelle; Delpuech, Oona; Brown, Henry; Boros, Joanna; Camp, Nicola J; McEwen, Robert; Mazzola, Anne Marie; D'Cruz, Celina; Barry, Simon T

    2016-04-12

    The tumor microenvironment is emerging as a key regulator of cancer growth and progression, however the exact mechanisms of interaction with the tumor are poorly understood. Whilst the majority of genomic profiling efforts thus far have focused on the tumor, here we investigate RNA-Seq as a hypothesis-free tool to generate independent tumor and stromal biomarkers, and explore tumor-stroma interactions by exploiting the human-murine compartment specificity of patient-derived xenografts (PDX).Across a pan-cancer cohort of 79 PDX models, we determine that mouse stroma can be separated into distinct clusters, each corresponding to a specific stromal cell type. This implies heterogeneous recruitment of mouse stroma to the xenograft independent of tumor type. We then generate cross-species expression networks to recapitulate a known association between tumor epithelial cells and fibroblast activation, and propose a potentially novel relationship between two hypoxia-associated genes, human MIF and mouse Ddx6. Assessment of disease subtype also reveals MMP12 as a putative stromal marker of triple-negative breast cancer. Finally, we establish that our ability to dissect recruited stroma from trans-differentiated tumor cells is crucial to identifying stem-like poor-prognosis signatures in the tumor compartment.In conclusion, RNA-Seq is a powerful, cost-effective solution to global analysis of human tumor and mouse stroma simultaneously, providing new insights into mouse stromal heterogeneity and compartment-specific disease markers that are otherwise overlooked by alternative technologies. The study represents the first comprehensive analysis of its kind across multiple PDX models, and supports adoption of the approach in pre-clinical drug efficacy studies, and compartment-specific biomarker discovery.

  20. Targeting Tumor Vasculature Endothelial Cells and Tumor Cells for Immunotherapy of Human Melanoma in a Mouse Xenograft Model

    NASA Astrophysics Data System (ADS)

    Hu, Zhiwei; Sun, Ying; Garen, Alan

    1999-07-01

    An immunotherapy treatment for cancer that targets both the tumor vasculature and tumor cells has shown promising results in a severe combined immunodeficient mouse xenograft model of human melanoma. The treatment involves systemic delivery of an immunoconjugate molecule composed of a tumor-targeting domain conjugated to the Fc effector domain of human IgG1. The effector domain induces a cytolytic immune response against the targeted cells by natural killer cells and complement. Two types of targeting domains were used. One targeting domain is a human single-chain Fv molecule that binds to a chondroitin sulfate proteoglycan expressed on the surface of most human melanoma cells. Another targeting domain is factor VII (fVII), a zymogen that binds with high specificity and affinity to the transmembrane receptor tissue factor (TF) to initiate the blood coagulation cascade. TF is expressed by endothelial cells lining the tumor vasculature but not the normal vasculature, and also by many types of tumor cells including melanoma. Because the binding of a fVII immunoconjugate to TF might cause disseminated intravascular coagulation, the active site of fVII was mutated to inhibit coagulation without affecting the affinity for TF. The immunoconjugates were encoded as secreted molecules in a replication-defective adenovirus vector, which was injected into the tail vein of severe combined immunodeficient mice. The results demonstrate that a mutated fVII immunoconjugate, administered separately or together with a single-chain Fv immunoconjugate that binds to the tumor cells, can inhibit the growth or cause regression of an established human tumor xenograft. This procedure could be effective in treating a broad spectrum of human solid tumors that express TF on vascular endothelial cells and tumor cells.

  1. Abrogation of STAT3 signaling cascade by zerumbone inhibits proliferation and induces apoptosis in renal cell carcinoma xenograft mouse model.

    PubMed

    Shanmugam, Muthu K; Rajendran, Peramaiyan; Li, Feng; Kim, Chulwon; Sikka, Sakshi; Siveen, Kodappully Sivaraman; Kumar, Alan Prem; Ahn, Kwang Seok; Sethi, Gautam

    2015-10-01

    Persistent activation of signal transducer and activator of transcription 3 (STAT3) is one of the characteristic features of renal cell carcinoma (RCC) and often linked to its deregulated proliferation, survival, and angiogenesis. In the present report, we investigated whether zerumbone, a sesquiterpene, exerts its anticancer effect through modulation of STAT3 activation pathway. The pharmacological effect of zerumbone on STAT3 activation, associated protein kinases and phosphatase, and apoptosis was investigated using both RCC cell lines and xenograft mouse model. We observed that zerumbone suppressed STAT3 activation in a dose- and time-dependent manner in RCC cells. The suppression was mediated through the inhibition of activation of upstream kinases c-Src, Janus-activated kinase 1, and Janus-activated kinase 2. Pervanadate treatment reversed zerumbone-induced downregulation of STAT3, suggesting the involvement of a tyrosine phosphatase. Indeed, we found that zerumbone induced the expression of tyrosine phosphatase SHP-1 that correlated with its ability to inhibit STAT3 activation. Interestingly, deletion of SHP-1 gene by siRNA abolished the ability of zerumbone to inhibit STAT3 activation. The inhibition of STAT3 activation by zerumbone also caused the suppression of the gene products involved in proliferation, survival, and angiogenesis. Finally, when administered i.p., zerumbone inhibited STAT3 activation in tumor tissues and the growth of human RCC xenograft tumors in athymic nu/nu mice without any side effects. Overall, our results suggest for the first time that zerumbone is a novel blocker of STAT3 signaling cascade and thus has an enormous potential for the treatment of RCC and other solid tumors.

  2. Nilotinib reverses ABCB1/P-glycoprotein-mediated multidrug resistance but increases cardiotoxicity of doxorubicin in a MDR xenograft model.

    PubMed

    Zhou, Zhi-Yong; Wan, Li-Li; Yang, Quan-Jun; Han, Yong-Long; Li, Dan; Lu, Jin; Guo, Cheng

    2016-09-30

    The BCR-Abl tyrosine kinase inhibitor (TKI), nilotinib, was developed to surmount resistance or intolerance to imatinib in patients with Philadelphia-positive chronic myelogenous leukemia. Recent studies have shown that nilotinib induces potent sensitization to anticancer agents by blocking the functions of ABCB1/P-glycoprotein (P-gp) in multidrug resistance (MDR). However, changes in P-gp expression or function affect the cardiac disposition and prolong the presence of both doxorubicin (DOX) and doxorubicinol (DOXol) in cardiac tissue, thus, enhancing the risk of cardiotoxicity. In this study, we used a MDR xenograft model to evaluate the antitumor activity, tissue distribution and cardiotoxicity of DOX when co-administered with nilotinib. This information will provide more insight into the pharmacological role of nilotinib in MDR reversal and the risk of DOX cardiotoxicity. Our results showed that nilotinib significantly enhanced DOX cytotoxicity and increased intracellular rhodamine 123 accumulation in MG63/DOX cells in vitro and strongly enhanced DOX inhibition of growth of P-gp-overexpressing MG63/DOX cell xenografts in nude mice. Additionally, nilotinib significantly increased DOX and DOXol accumulation in serum, heart, liver and tumor tissues. Importantly, nilotinib induced a disproportionate increase in DOXol in cardiac tissue. In the co-administration group, CBR1 and AKR1A1 protein levels were significantly increased in cardiac tissue, with more severe necrosis and vacuole formation. These results indicate that nilotinib reverses P-gp- mediated MDR by blocking the efflux function and potentiates DOX-induced cardiotoxicity. These findings represent a guide for the design of future clinical trials and studies of pharmacokinetic interactions and may be useful in guiding the use of nilotinib in combination therapy of cancer in clinical practice.

  3. Molecular Imaging of Bcr-Abl Phosphokinase in a Xenograft Model*

    PubMed Central

    Wu, Ji Yuan; Yang, David J.; Angelo, Laura S.; Kohanim, Saady; Kurzrock, Razelle

    2009-01-01

    The purpose of this study was to determine whether the Bcr-Abl tyrosine kinase can be assessed by gamma imaging using an 111Indium-labeled anti-phosphotyrosine antibody, and if the response to treatment with imatinib could be detected using this imaging technique. Anti-phosphotyrosine antibody (APT) was labeled with indium (111In) using ethylenedicysteine (EC) as a chelator. To determine if 111In-EC-APT could assess a non-receptor tyrosine kinase, xenografts of the human chronic myelogenous leukemia (CML) cell line K562 were used. Gamma scintigraphy of the tumor-bearing mice, before and after imatinib treatment, was obtained 1, 24, and 48 hours after they were given 111In-EC-APT (100 uCi/mouse, i.v.). 111In-EC-APT is preferentially taken up by Bcr-Abl-bearing tumor cells when compared to 111In-EC-BSA or 111In-EC-IgG1 controls, and comparable to the level of uptake of 111In-EC-Bcr-Abl. Imatinib treatment resulted in decreased expression of phosphorylated Bcr-Abl by Western blot analysis, which correlated with early (four days after starting imatinib) kinase down-regulation as assessed by imaging using 111In-EC-APT. The optimal time to imaging was 24 and 48 hours after injection of 111In-EC-APT. Although tumor regression was insignificant on day 4 after starting imatinib treatment, it was marked by day 14. 111In-EC-APT can assess intracellular phosphokinase activity, and down-regulation of phosphokinase activity predates tumor regression. This technique may therefore be useful in the clinic to detect the presence of phosphokinase activity, and for early prediction of response. PMID:19258427

  4. FL118, a novel camptothecin analogue, overcomes irinotecan and topotecan resistance in human tumor xenograft models

    PubMed Central

    Ling, Xiang; Liu, Xiaojun; Zhong, Kai; Smith, Nicholas; Prey, Joshua; Li, Fengzhi

    2015-01-01

    Irinotecan and topotecan are the only camptothecin analogues approved by the FDA for cancer treatment. However, inherent and/or acquired irinotecan and topotecan resistance is a challenging issue in clinical practice. In this report, we showed that FL118, a novel camptothecin analogue, effectively obliterated human xenograft tumors that acquire irinotecan and topotecan resistance. Consistent with this finding, Pharmacokinetics studies indicated that FL118 rapidly clears from circulation, while effectively accumulating in tumors with a long elimination half-life. Consistent with our previous studies on irinotecan, FL118 exhibited ≥25 fold more effectiveness than topotecan at inhibiting cancer cell growth and colony formation; we further showed that although topotecan can inhibit the expression of survivin, Mcl-1, XIAP or cIAP2, its effectiveness is about 10-100 fold weaker than FL118. Lastly, in contrast to both SN-38 (active metabolite of irinotecan) and topotecan are substrates of the efflux pump proteins P-gp/MDR1 and ABCG2/BCRP, FL118 is not a substrate of P-gp and ABCG2. Consistently, sildenafil, a multiple efflux pump inhibitor, sensitized SN-38 much more than these of the ABCG2-selective inhibitor KO143 in growth inhibition of SW620 and HCT-8 cells. In contrast, both inhibitors showed no effect on FL118 efficacy. Given that both P-gp and ABCG2 express in SW620 and HCT-8 cells and FL118 is not a substrate for P-gp and ABCG2, this suggests that FL118 appears to bypass multiple efflux pump protein-induced resistance, which may contribute to FL118 overcoming irinotecan and topotecan resistance in vivo. These new findings provide renewed perspectives for further development of FL118 for clinical applications. PMID:26692923

  5. Effect of melatonin on tumor growth and angiogenesis in xenograft model of breast cancer.

    PubMed

    Jardim-Perassi, Bruna Victorasso; Arbab, Ali S; Ferreira, Lívia Carvalho; Borin, Thaiz Ferraz; Varma, Nadimpalli R S; Iskander, A S M; Shankar, Adarsh; Ali, Meser M; de Campos Zuccari, Debora Aparecida Pires

    2014-01-01

    As neovascularization is essential for tumor growth and metastasis, controlling angiogenesis is a promising tactic in limiting cancer progression. Melatonin has been studied for their inhibitory properties on angiogenesis in cancer. We performed an in vivo study to evaluate the effects of melatonin treatment on angiogenesis in breast cancer. Cell viability was measured by MTT assay after melatonin treatment in triple-negative breast cancer cells (MDA-MB-231). After, cells were implanted in athymic nude mice and treated with melatonin or vehicle daily, administered intraperitoneally 1 hour before turning the room light off. Volume of the tumors was measured weekly with a digital caliper and at the end of treatments animals underwent single photon emission computed tomography (SPECT) with Technetium-99m tagged vascular endothelial growth factor (VEGF) C to detect in vivo angiogenesis. In addition, expression of pro-angiogenic/growth factors in the tumor extracts was evaluated by membrane antibody array and collected tumor tissues were analyzed with histochemical staining. Melatonin in vitro treatment (1 mM) decreased cell viability (p<0.05). The breast cancer xenografts nude mice treated with melatonin showed reduced tumor size and cell proliferation (Ki-67) compared to control animals after 21 days of treatment (p<0.05). Expression of VEGF receptor 2 decreased significantly in the treated animals compared to that of control when determined by immunohistochemistry (p<0.05) but the changes were not significant on SPECT (p>0.05) images. In addition, there was a decrease of micro-vessel density (Von Willebrand Factor) in melatonin treated mice (p<0.05). However, semiquantitative densitometry analysis of membrane array indicated increased expression of epidermal growth factor receptor and insulin-like growth factor 1 in treated tumors compared to vehicle treated tumors (p<0.05). In conclusion, melatonin treatment showed effectiveness in reducing tumor growth and cell

  6. In vitro and in vivo studies on the inhibitory effects of myocardial cell culture medium on growth of a human lung adenocarcinoma cell line, A549

    PubMed Central

    Zheng, Y.; Zhou, J.; Fu, S.Z.; Fan, J.; Wu, J.B.

    2016-01-01

    Background Although the heart is one of the body’s vital organs, with an abundant blood supply, metastasis to the heart is considered rare. In a previous study, we found that the myocardial microenvironment might contain a low molecular weight natural tumour suppressor. The present study was designed to investigate the inhibitory effect of cardiac myocyte–conditioned medium (cmcm) on the growth of A549 human lung adenocarcinoma cells in vitro and in vivo. Methods An mtt assay was used to detect the inhibition ratio with respect to A549 proliferation. Human lung adenocarcinoma cells (A549 cell strain) were transplanted subcutaneously into nude mice to produce tumours. The xenograft tumour growth in mice was observed after selected drug administration. Results After treatment with cmcm and cisplatin (Cis), A549 cell viability significantly declined (p < 0.001). The cell viability in the cmcm and Cis groups were 53.42% ± 3.45% and 58.45% ± 6.39% respectively. Growth of implanted tumour cells in vivo was significantly inhibited in the cmcm group, the group treated with recombinant human adenovirus–p53, and the Cis-treated group compared with a control group. The inhibition rates were 41.44% in the cmcm group, 41.34% in the p53 group, and 64.50% in the Cis group. Lung metastasis capacity was significantly reduced in the presence of cmcm (p < 0.05). Lung metastasis inhibition rates in mice were 56.52% in the cmcm group, 47.83% in the p53 group, and 82.61% in the Cis group. With cmcm, the lives of A549-tumour-bearing mice could be significantly prolonged without any effect on weight loss. Conclusions Use of cmcm has the effect of reducing A549 cell viability, tumour volume, and lung metastasis rate, while prolonging survival duration without severe toxicity. PMID:26966411

  7. TLE1 promotes EMT in A549 lung cancer cells through suppression of E-cadherin

    PubMed Central

    Yao, Xin; Ireland, Shubha Kale; Pham, Tri; Temple, Brandi; Chen, Renwei; Raj, Madhwa HG; Biliran, Hector

    2014-01-01

    The Groucho transcriptional corepressor TLE1 protein has recently been shown to be a putative lung specific oncogene, but its underlying oncogenic activity in lung cancer has not been fully elucidated. In this report, we investigated whether TLE1 regulates lung cancer aggressiveness using the human lung adenocarcinoma cell line A549 as a model system. Through a combination of genetic approaches, we found that TLE1 potentiates Epithelial-to-Mesenchymal Transition (EMT) in A549 cells in part through suppression of the tumor suppressor gene E-cadherin. Exogenous expression of TLE1 in A549 cells resulted in heightened EMT phenotypes (enhanced fibroblastoid morphology and increased cell migratory potential) and in molecular alterations characteristic of EMT (downregulation of the epithelial marker E-cadherin and upregulation of the mesenchymal marker Vimentin). Conversely, downregulation of endogenous TLE1 expression in these cells resulted in reversal of basal EMT characterized by a cuboidal-like epithelial cell phenotype, reduced cell motility, and upregulated E-cadherin expression. Mechanistic studies showed that TLE1 suppresses E-cadherin expression at the transcriptional level in part by recruiting Histone Deacetylase (HDAC) activity to the E-cadherin promoter. Consistently, the HDAC inhibitor TSA partially reversed the TLE1-induced E-cadherin downregulation and cell migration, suggesting a role for HDACs in TLE1-mediated transcriptional repression of E-cadherin and EMT function. These findings uncover a novel role of TLE1 in regulating EMT in A549 cells through its repressive effect on E-cadherin and provide a mechanism for TLE1 oncogenic activity in lung cancer. PMID:25446087

  8. Lipocortin 1 mediates dexamethasone-induced growth arrest of the A549 lung adenocarcinoma cell line.

    PubMed Central

    Croxtall, J D; Flower, R J

    1992-01-01

    The synthetic glucocorticoid dexamethasone (1 microM to 1 pM) strongly (maximum greater than 80%) inhibits proliferation of the A549 human lung adenocarcinoma line (EC50 greater than 1 nM) and leads to the appearance, or a further increase (approximately 3-fold) in the expression on the cell surface, of the calcium and phospholipid binding protein lipocortin (annexin) 1. Both these effects, which are shared by hydrocortisone (1 microM) but not by progesterone or aldosterone (1 microM), are inhibited by the antiglucocorticoids RU38486 and RU43044 (1 microM). The nonsteroidal antiinflammatory drugs indomethacin (1 microM) and naproxen (10 microM) and human recombinant lipocortin 1 (0.05-5.0 micrograms/ml) also produce growth arrest in this cell line. During proliferation A549 cells spontaneously release prostaglandin E2 [10-20 ng (28-57 pmol) per ml per 5-day period] into the growth medium. In concentrations that cause growth-arrest, dexamethasone, indomethacin, and lipocortin 1 abolish the generation of this eicosanoid by A549 cells. Prostaglandin E2 itself (0.01-1 pM) stimulates cell growth and partially reverses (approximately 50%) the inhibition of growth caused by dexamethasone and indomethacin. Addition of the neutralizing anti-lipocortin 1 monoclonal antibody 1A (5 micrograms/ml), but not the nonneutralizing anti-lipocortin monoclonal antibody 1B, substantially reversed (greater than 80%) the inhibitory activity of dexamethasone on both growth and prostaglandin E2 synthesis. The generation of prostaglandin E2 by A549 cells seems to be an important regulator of cell proliferation in vitro and the dexamethasone-induced suppression of proliferation in this model is attributable to eicosanoid inhibition caused by lipocortin 1. Images PMID:1533045

  9. Artesunate induces AIF-dependent apoptosis in A549 cells

    NASA Astrophysics Data System (ADS)

    Zhou, Chen-juan; Chen, Tong-Sheng

    2012-03-01

    Artesunate (ART), a semi-synthetic derivative of the sesquiterpene artemisinin extracted from the Chinese herb Artemisia annua, exerts a broad spectrum of clinical activity against human cancers. It has been shown that ART induces cancer cells death through apoptosis pathway. This study investigated whether ART treatment induced reactive oxygen species (ROS)-dependent cell death in the apoptosis fashion in human lung adenocarconoma A549 cell line and the proapoptotic protein apoptosis inducing factor (AIF) is involved in ART-induced apoptosis. Cells treated with ART exhibited typical apoptotic morphology as chromatin condensation, margination and shrunken nucleus. ART treatment also induced a loss of mitochondrial membrane potential and AIF release from mitochondria. Silencing AIF can remarkable attenuated ART-induced apoptosis. Collectively, ART induces apoptosis by caspase-independent intrinsic pathway in A549 cells.

  10. Phloretin induces apoptosis of non-small cell lung carcinoma A549 cells via JNK1/2 and p38 MAPK pathways.

    PubMed

    Min, Jie; Huang, Kenan; Tang, Hua; Ding, Xinyu; Qi, Chen; Qin, Xiong; Xu, Zhifei

    2015-12-01

    Phloretin (Ph) existing in apples, pears and various vegetables is known to have antitumor activities in several cancer cell lines. However, little is known about its effect on human lung cancer cells. The aim of the present study was to see whether Ph could induce apoptosis of non-small cell lung cancer (NSCLC) cells, and explore the possible underlying mechanism of action. We found that Ph markedly induced cell apoptosis of NSCLC cell line A549, and inhibited the migration of A549 cells in a dose-dependent manner. The expression level of BAX, cleaved caspase-3 and -9, and degraded form of PARP was increased and Bcl-2 was decreased after Ph treatment. In addition, the phosphorylation of P38 MAPK, ERK1/2 and JNK1/2 was increased in a dose‑dependent manner in parallel with Ph treatment. Inhibition of P38 MAPK and JNK1/2 by specific inhibitors significantly abolished the Ph-induced activation of the caspase-3 and -9. In vivo tumor-suppression assay further indicated that Ph (20 mg/kg) displayed a more significant inhibitory effect on A549 xenografts in tumor growth. All these findings indicate that Ph is able to inhibit NSCLC A549 cell growth by inducing apoptosis through P38 MAPK and JNK1/2 pathways, and therefore may prove to be an adjuvant to the treatment of NSCLC.

  11. Phloretin induces apoptosis of non-small cell lung carcinoma A549 cells via JNK1/2 and p38 MAPK pathways.

    PubMed

    Min, Jie; Huang, Kenan; Tang, Hua; Ding, Xinyu; Qi, Chen; Qin, Xiong; Xu, Zhifei

    2015-12-01

    Phloretin (Ph) existing in apples, pears and various vegetables is known to have antitumor activities in several cancer cell lines. However, little is known about its effect on human lung cancer cells. The aim of the present study was to see whether Ph could induce apoptosis of non-small cell lung cancer (NSCLC) cells, and explore the possible underlying mechanism of action. We found that Ph markedly induced cell apoptosis of NSCLC cell line A549, and inhibited the migration of A549 cells in a dose-dependent manner. The expression level of BAX, cleaved caspase-3 and -9, and degraded form of PARP was increased and Bcl-2 was decreased after Ph treatment. In addition, the phosphorylation of P38 MAPK, ERK1/2 and JNK1/2 was increased in a dose‑dependent manner in parallel with Ph treatment. Inhibition of P38 MAPK and JNK1/2 by specific inhibitors significantly abolished the Ph-induced activation of the caspase-3 and -9. In vivo tumor-suppression assay further indicated that Ph (20 mg/kg) displayed a more significant inhibitory effect on A549 xenografts in tumor growth. All these findings indicate that Ph is able to inhibit NSCLC A549 cell growth by inducing apoptosis through P38 MAPK and JNK1/2 pathways, and therefore may prove to be an adjuvant to the treatment of NSCLC. PMID:26503828

  12. Phloretin induces apoptosis of non-small cell lung carcinoma A549 cells via JNK1/2 and p38 MAPK pathways

    PubMed Central

    MIN, JIE; LI, XU; HUANG, KENAN; TANG, HUA; DING, XINYU; QI, CHEN; QIN, XIONG; XU, ZHIFEI

    2015-01-01

    Phloretin (Ph) existing in apples, pears and various vegetables is known to have antitumor activities in several cancer cell lines. However, little is known about its effect on human lung cancer cells. The aim of the present study was to see whether Ph could induce apoptosis of non-small cell lung cancer (NSCLC) cells, and explore the possible underlying mechanism of action. We found that Ph markedly induced cell apoptosis of NSCLC cell line A549, and inhibited the migration of A549 cells in a dose-dependent manner. The expression level of BAX, cleaved caspase-3 and -9, and degraded form of PARP was increased and Bcl-2 was decreased after Ph treatment. In addition, the phosphorylation of P38 MAPK, ERK1/2 and JNK1/2 was increased in a dose-dependent manner in parallel with Ph treatment. Inhibition of P38 MAPK and JNK1/2 by specific inhibitors significantly abolished the Ph-induced activation of the caspase-3 and -9. In vivo tumor-suppression assay further indicated that Ph (20 mg/kg) displayed a more significant inhibitory effect on A549 xenografts in tumor growth. All these findings indicate that Ph is able to inhibit NSCLC A549 cell growth by inducing apoptosis through P38 MAPK and JNK1/2 pathways, and therefore may prove to be an adjuvant to the treatment of NSCLC. PMID:26503828

  13. Patient-derived orthotopic xenografts: better mimic of metastasis than subcutaneous xenografts.

    PubMed

    Hoffman, Robert M

    2015-08-01

    The majority of human solid tumours do not metastasize when grown subcutaneously in immunocompromised mice; this includes patient-derived xenograft (PDX) models. However, orthotopic implantation of intact tumour tissue can lead to metastasis that mimics that seen in patients. These patient-derived orthotopic xenograft (PDOX) models have a long history and might better recapitulate human tumours than PDX models.

  14. Development of a Patient-Derived Xenograft (PDX) of Breast Cancer Bone Metastasis in a Zebrafish Model

    PubMed Central

    Mercatali, Laura; La Manna, Federico; Groenewoud, Arwin; Casadei, Roberto; Recine, Federica; Miserocchi, Giacomo; Pieri, Federica; Liverani, Chiara; Bongiovanni, Alberto; Spadazzi, Chiara; de Vita, Alessandro; van der Pluijm, Gabri; Giorgini, Andrea; Biagini, Roberto; Amadori, Dino; Ibrahim, Toni; Snaar-Jagalska, Ewa

    2016-01-01

    Bone metastasis is a complex process that needs to be better understood in order to help clinicians prevent and treat it. Xenografts using patient-derived material (PDX) rather than cancer cell lines are a novel approach that guarantees more clinically realistic results. A primary culture of bone metastasis derived from a 67-year-old patient with breast cancer was cultured and then injected into zebrafish (ZF) embryos to study its metastatic potential. In vivo behavior and results of gene expression analyses of the primary culture were compared with those of cancer cell lines with different metastatic potential (MCF7 and MDA-MB-231). The MCF7 cell line, which has the same hormonal receptor status as the bone metastasis primary culture, did not survive in the in vivo model. Conversely, MDA-MB-231 disseminated and colonized different parts of the ZF, including caudal hematopoietic tissues (CHT), revealing a migratory phenotype. Primary culture cells disseminated and in later stages extravasated from the vessels, engrafting into ZF tissues and reaching the CHT. Primary cell behavior reflected the clinical course of the patient’s medical history. Our results underline the potential for using PDX models in bone metastasis research and outline new methods for the clinical application of this in vivo model. PMID:27556456

  15. Development of a Patient-Derived Xenograft (PDX) of Breast Cancer Bone Metastasis in a Zebrafish Model.

    PubMed

    Mercatali, Laura; La Manna, Federico; Groenewoud, Arwin; Casadei, Roberto; Recine, Federica; Miserocchi, Giacomo; Pieri, Federica; Liverani, Chiara; Bongiovanni, Alberto; Spadazzi, Chiara; de Vita, Alessandro; van der Pluijm, Gabri; Giorgini, Andrea; Biagini, Roberto; Amadori, Dino; Ibrahim, Toni; Snaar-Jagalska, Ewa

    2016-08-22

    Bone metastasis is a complex process that needs to be better understood in order to help clinicians prevent and treat it. Xenografts using patient-derived material (PDX) rather than cancer cell lines are a novel approach that guarantees more clinically realistic results. A primary culture of bone metastasis derived from a 67-year-old patient with breast cancer was cultured and then injected into zebrafish (ZF) embryos to study its metastatic potential. In vivo behavior and results of gene expression analyses of the primary culture were compared with those of cancer cell lines with different metastatic potential (MCF7 and MDA-MB-231). The MCF7 cell line, which has the same hormonal receptor status as the bone metastasis primary culture, did not survive in the in vivo model. Conversely, MDA-MB-231 disseminated and colonized different parts of the ZF, including caudal hematopoietic tissues (CHT), revealing a migratory phenotype. Primary culture cells disseminated and in later stages extravasated from the vessels, engrafting into ZF tissues and reaching the CHT. Primary cell behavior reflected the clinical course of the patient's medical history. Our results underline the potential for using PDX models in bone metastasis research and outline new methods for the clinical application of this in vivo model.

  16. Diffuse large B-cell lymphoma patient-derived xenograft models capture the molecular and biological heterogeneity of the disease.

    PubMed

    Chapuy, Bjoern; Cheng, Hongwei; Watahiki, Akira; Ducar, Matthew D; Tan, Yuxiang; Chen, Linfeng; Roemer, Margaretha G M; Ouyang, Jing; Christie, Amanda L; Zhang, Liye; Gusenleitner, Daniel; Abo, Ryan P; Farinha, Pedro; von Bonin, Frederike; Thorner, Aaron R; Sun, Heather H; Gascoyne, Randy D; Pinkus, Geraldine S; van Hummelen, Paul; Wulf, Gerald G; Aster, Jon C; Weinstock, David M; Monti, Stefano; Rodig, Scott J; Wang, Yuzhuo; Shipp, Margaret A

    2016-05-01

    Diffuse large B-cell lymphoma (DLBCL) is a heterogeneous disease defined by transcriptional classifications, specific signaling and survival pathways, and multiple low-frequency genetic alterations. Preclinical model systems that capture the genetic and functional heterogeneity of DLBCL are urgently needed. Here, we generated and characterized a panel of large B-cell lymphoma (LBCL) patient-derived xenograft (PDX) models, including 8 that reflect the immunophenotypic, transcriptional, genetic, and functional heterogeneity of primary DLBCL and 1 that is a plasmablastic lymphoma. All LBCL PDX models were subjected to whole-transcriptome sequencing to classify cell of origin and consensus clustering classification (CCC) subtypes. Mutations and chromosomal rearrangements were evaluated by whole-exome sequencing with an extended bait set. Six of the 8 DLBCL models were activated B-cell (ABC)-type tumors that exhibited ABC-associated mutations such as MYD88, CD79B, CARD11, and PIM1. The remaining 2 DLBCL models were germinal B-cell type, with characteristic alterations of GNA13, CREBBP, and EZH2, and chromosomal translocations involving IgH and either BCL2 or MYC Only 25% of the DLBCL PDX models harbored inactivating TP53 mutations, whereas 75% exhibited copy number alterations of TP53 or its upstream modifier, CDKN2A, consistent with the reported incidence and type of p53 pathway alterations in primary DLBCL. By CCC criteria, 6 of 8 DLBCL PDX models were B-cell receptor (BCR)-type tumors that exhibited selective surface immunoglobulin expression and sensitivity to entospletinib, a recently developed spleen tyrosine kinase inhibitor. In summary, we have established and characterized faithful PDX models of DLBCL and demonstrated their usefulness in functional analyses of proximal BCR pathway inhibition.

  17. Metallofullerene-based Nanoplatform for Brain Tumor Brachytherapy and Longitudinal Imaging in a Murine Orthotopic Xenograft Model

    PubMed Central

    Shultz, Michael D.; Wilson, John D.; Fuller, Christine E.; Zhang, Jianyuan; Dorn, Harry C.

    2011-01-01

    Purpose: To demonstrate in an orthotopic xenograft brain tumor model that a functionalized metallofullerene (f-Gd3N@C80) can enable longitudinal tumor imaging and, when radiolabeled with lutetium 177 (177Lu) and tetraazacyclododecane tetraacetic acid (DOTA) (177Lu-DOTA-f-Gd3N@C80), provide an anchor to deliver effective brachytherapy. Materials and Methods: All experiments involving the use of mice were carried out in accordance with protocols approved by the institutional animal care and use committee. Human glioblastoma U87MG cells were implanted by using stereotactic procedures into the brains of 37 female athymic nude-Foxn1nu mice and allowed to develop into a tumor for 8 days. T1- and T2-weighted magnetic resonance (MR) imaging was performed in five mice. Biodistribution studies were performed in 12 mice at four time points over 7 days to evaluate gadolinium content. Survival studies involved 20 mice that received infusion of a nanoplatform by means of convection-enhanced delivery (CED) 8 days after tumor implantation. Mice in survival studies were divided into two groups: one comprised untreated mice that received f-Gd3N@C80 alone and the other comprised mice treated with brachytherapy that received 1.11 MBq of 177Lu-DOTA-f-Gd3N@C80. Survival data were evaluated by using Kaplan-Meier statistical methods. Results: MR imaging showed extended tumor retention (25.6% ± 1.2 of the infused dose at 52 days, confirmed with biodistribution studies) of the f-Gd3N@C80 nanoplatform, which enabled longitudinal imaging. Successful coupling of 177Lu to the f-Gd3N@C80 surface was achieved by using a bifunctional macrocyclic chelator. The extended tumor retention allowed for effective brachytherapy, as indicated by extended survival time (>2.5 times that of the untreated group) and histologic signs of radiation-induced tumor damage. Conclusion: The authors have developed a multimodal nanoplatform and have demonstrated longitudinal tumor imaging, prolonged intratumoral probe

  18. The isothiocyanate erucin abrogates telomerase in hepatocellular carcinoma cells in vitro and in an orthotopic xenograft tumour model of HCC.

    PubMed

    Herz, Corinna; Hertrampf, Anke; Zimmermann, Stefan; Stetter, Nadine; Wagner, Meike; Kleinhans, Claudia; Erlacher, Miriam; Schüler, Julia; Platz, Stefanie; Rohn, Sascha; Mersch-Sundermann, Volker; Lamy, Evelyn

    2014-12-01

    In contrast to cancer cells, most normal human cells have no or low telomerase levels which makes it an attractive target for anti-cancer drugs. The small molecule sulforaphane from broccoli is known for its cancer therapeutic potential in vitro and in vivo. In animals and humans it was found to be quickly metabolized into 4-methylthiobutyl isothiocyanate (MTBITC, erucin) which we recently identified as strong selective apoptosis inducer in hepatocellular carcinoma (HCC) cells. Here, we investigated the relevance of telomerase abrogation for cytotoxic efficacy of MTBITC against HCC. The drug was effective against telomerase, independent from TP53 and MTBITC also blocked telomerase in chemoresistant subpopulations. By using an orthotopic human liver cancer xenograft model, we give first evidence that MTBITC at 50 mg/KG b.w./d significantly decreased telomerase activity in vivo without affecting enzyme activity of adjacent normal tissue. Upon drug exposure, telomerase decrease was consistent with a dose-dependent switch to anti-survival, cell arrest and apoptosis in our in vitro HCC models. Blocking telomerase by the specific inhibitor TMPyP4 further sensitized cancer cells to MTBITC-mediated cytotoxicity. Overexpression of hTERT, but not enzyme activity deficient DNhTERT, protected against apoptosis; neither DNA damage nor cytostasis induction by MTBITC was prevented by hTERT overexpression. These findings imply that telomerase enzyme activity does not protect against MTBITC-induced DNA damage but impacts signalling processes upstream of apoptosis execution level.

  19. The isothiocyanate erucin abrogates telomerase in hepatocellular carcinoma cells in vitro and in an orthotopic xenograft tumour model of HCC

    PubMed Central

    Herz, Corinna; Hertrampf, Anke; Zimmermann, Stefan; Stetter, Nadine; Wagner, Meike; Kleinhans, Claudia; Erlacher, Miriam; Schüler, Julia; Platz, Stefanie; Rohn, Sascha; Mersch-Sundermann, Volker; Lamy, Evelyn

    2014-01-01

    In contrast to cancer cells, most normal human cells have no or low telomerase levels which makes it an attractive target for anti-cancer drugs. The small molecule sulforaphane from broccoli is known for its cancer therapeutic potential in vitro and in vivo. In animals and humans it was found to be quickly metabolized into 4-methylthiobutyl isothiocyanate (MTBITC, erucin) which we recently identified as strong selective apoptosis inducer in hepatocellular carcinoma (HCC) cells. Here, we investigated the relevance of telomerase abrogation for cytotoxic efficacy of MTBITC against HCC. The drug was effective against telomerase, independent from TP53 and MTBITC also blocked telomerase in chemoresistant subpopulations. By using an orthotopic human liver cancer xenograft model, we give first evidence that MTBITC at 50 mg/KG b.w./d significantly decreased telomerase activity in vivo without affecting enzyme activity of adjacent normal tissue. Upon drug exposure, telomerase decrease was consistent with a dose-dependent switch to anti-survival, cell arrest and apoptosis in our in vitro HCC models. Blocking telomerase by the specific inhibitor TMPyP4 further sensitized cancer cells to MTBITC-mediated cytotoxicity. Overexpression of hTERT, but not enzyme activity deficient DNhTERT, protected against apoptosis; neither DNA damage nor cytostasis induction by MTBITC was prevented by hTERT overexpression. These findings imply that telomerase enzyme activity does not protect against MTBITC-induced DNA damage but impacts signalling processes upstream of apoptosis execution level. PMID:25256442

  20. Optimizing lutetium 177-anti-carbonic anhydrase IX radioimmunotherapy in an intraperitoneal clear cell renal cell carcinoma xenograft model.

    PubMed

    Muselaers, Constantijn H J; Oosterwijk, Egbert; Bos, Desirée L; Oyen, Wim J G; Mulders, Peter F A; Boerman, Otto C

    2014-01-01

    A new approach in the treatment of clear cell renal carcinoma (ccRCC) is radioimmunotherapy (RIT) using anti-carbonic anhydrase IX (CAIX) antibody G250. To investigate the potential of RIT with lutetium 177 (177Lu)-labeled G250, we conducted a protein dose escalation study and subsequently an RIT study in mice with intraperitoneally growing ccRCC lesions. Mice with intraperitoneal xenografts were injected with 1, 3, 10, 30, or 100 μg of G250 labeled with 10 MBq indium 111 (111In) to determine the optimal protein dose. The optimal protein dose determined with imaging and biodistribution studies was used in a subsequent RIT experiment in three groups of 10 mice with intraperitoneal SK-RC-52 tumors. One group received 13 MBq 177Lu-DOTA-G250, a control group received 13 MBq nonspecific 177Lu-MOPC21, and the second control group was not treated and received 20 MBq 111In-DOTA-G250. The optimal G250 protein dose to target ccRCC in this model was 10 μg G250. Treatment with 13 MBq 177Lu-DOTA-G250 was well tolerated and resulted in significantly prolonged median survival (139 days) compared to controls (49-53 days, p  =  .015), indicating that RIT has potential in this metastatic ccRCC model.

  1. Antitumor activity of [Pt(O,O'-acac)(γ-acac)(DMS)] in mouse xenograft model of breast cancer.

    PubMed

    Muscella, A; Vetrugno, C; Migoni, D; Biagioni, F; Fanizzi, F P; Fornai, F; De Pascali, S A; Marsigliante, S

    2014-01-01

    The higher and selective cytotoxicity of [Pt(O,O'-acac)(γ-acac)(DMS)] toward cancer cell in both immortalized cell lines and in breast cancer cells in primary cultures, stimulated a pre-clinical study so as to evaluate its therapeutic potential in vivo. The efficacy of [Pt(O,O'-acac)(γ-acac)(DMS)] was assessed using a xenograft model of breast cancer developed by injection of MCF-7 cells in the flank of BALB/c nude mice. Treatment of solid tumor-bearing mice with [Pt(O,O'-acac)(γ-acac)(DMS)] induced up to 50% reduction of tumor mass compared with an average 10% inhibition recorded in cisplatin-treated animals. Thus, chemotherapy with [Pt(O,O'-acac)(γ-acac)(DMS)] was much more effective than cisplatin. We also demonstrated enhanced in vivo pharmacokinetics, biodistribution and tolerability of [Pt(O,O'-acac)(γ-acac)(DMS)] when compared with cisplatin administered in Wistar rats. Pharmacokinetics studies with [Pt(O,O'-acac)(γ-acac)(DMS)] revealed prolonged Pt persistence in systemic blood circulation and decreased nefrotoxicity and hepatotoxicity, major target sites of cisplatin toxicity. Overall, [Pt(O,O'-acac)(γ-acac)(DMS)] turned out to be extremely promising in terms of greater in vivo anticancer activity, reduced nephrotoxicity and acute toxicity compared with cisplatin. PMID:24457958

  2. Antitumor activity of [Pt(O,O'-acac)(γ-acac)(DMS)] in mouse xenograft model of breast cancer

    PubMed Central

    Muscella, A; Vetrugno, C; Migoni, D; Biagioni, F; Fanizzi, F P; Fornai, F; De Pascali, S A; Marsigliante, S

    2014-01-01

    The higher and selective cytotoxicity of [Pt(O,O′-acac)(γ-acac)(DMS)] toward cancer cell in both immortalized cell lines and in breast cancer cells in primary cultures, stimulated a pre-clinical study so as to evaluate its therapeutic potential in vivo. The efficacy of [Pt(O,O′-acac)(γ-acac)(DMS)] was assessed using a xenograft model of breast cancer developed by injection of MCF-7 cells in the flank of BALB/c nude mice. Treatment of solid tumor-bearing mice with [Pt(O,O′-acac)(γ-acac)(DMS)] induced up to 50% reduction of tumor mass compared with an average 10% inhibition recorded in cisplatin-treated animals. Thus, chemotherapy with [Pt(O,O′-acac)(γ-acac)(DMS)] was much more effective than cisplatin. We also demonstrated enhanced in vivo pharmacokinetics, biodistribution and tolerability of [Pt(O,O′-acac)(γ-acac)(DMS)] when compared with cisplatin administered in Wistar rats. Pharmacokinetics studies with [Pt(O,O′-acac)(γ-acac)(DMS)] revealed prolonged Pt persistence in systemic blood circulation and decreased nefrotoxicity and hepatotoxicity, major target sites of cisplatin toxicity. Overall, [Pt(O,O′-acac)(γ-acac)(DMS)] turned out to be extremely promising in terms of greater in vivo anticancer activity, reduced nephrotoxicity and acute toxicity compared with cisplatin. PMID:24457958

  3. The impact of bone morphogenetic protein 4 (BMP4) on breast cancer metastasis in a mouse xenograft model.

    PubMed

    Ampuja, M; Alarmo, E L; Owens, P; Havunen, R; Gorska, A E; Moses, H L; Kallioniemi, A

    2016-06-01

    Bone morphogenetic protein 4 (BMP4) is a key regulator of cell proliferation and differentiation. In breast cancer cells, BMP4 has been shown to reduce proliferation in vitro and interestingly, in some cases, also to induce migration and invasion. Here we investigated whether BMP4 influences breast cancer metastasis formation by using a xenograft mouse model. MDA-MB-231 breast cancer cells were injected intracardially into mice and metastasis formation was monitored using bioluminescence imaging. Mice treated with BMP4 developed metastases slightly earlier as compared to control animals but the overall number of metastases was similar in both groups (13 in the BMP4 group vs. 12 in controls). In BMP4-treated mice, bone metastases were more common (10 vs. 7) but adrenal gland metastases were less frequent (1 vs. 5) than in controls. Immunostaining revealed no differences in signaling activation, proliferation rate, blood vessel formation, EMT markers or the number of cancer-associated fibroblasts between the treatment groups. In conclusion, BMP4 caused a trend towards accelerated metastasis formation, especially in bone. More work is needed to uncover the long-term effects of BMP4 and the clinical relevance of these findings.

  4. The Addition of A Pregnenolone Pendant Group Enhances the Anticancer Properties of Titanocene Dichloride in a MCF-7 Xenograft Model

    PubMed Central

    Ramos, Gladiany; Loperena, Yaliz; Ortiz, Giovanni; Reyes, Fiorella; Szeto, Ada; Vera, Jose; Velez, Javier; Morales, Jessica; Morrero, Deborah; Castillo, Linnette; Dharmawardhane, Surangani; Melendez, Enrique; Washington, A. Valance

    2014-01-01

    Background/Aim Titanocene dichloride held great promise as a chemotherapeutic compound in preclinical studies. However, subsequent clinical trials revealed hepatoxicity and nephrotoxicity, which limited its use in clinical applications. Therefore, we used steroid pendant groups to improve the targeting of titanocene in the MCF-7 breast cancer cell line, and demonstrated a 10-fold lower effective dose compared to titanocene in in vitro assays. The aim of the present study was to test the efficacy of a titanocene functionalized with pregnenolone (Ti-Preg) in an in vivo breast cancer model. Materials and Methods Xenografts from the MCF7 breast cancer cell line were implanted into athymic nu/nu mice to evaluate the potential of Ti-Preg as an anti-breast cancer agent. Results Ti-Preg demonstrated a significant inhibition of MCF-7 tumor growth when compared to vehicle and to titanocene controls. Conclusion Our findings demonstrate the potential of steroid pendent groups for targeting chemotherapeutics to steroid hormone-dependent cancer. PMID:24692689

  5. Systemically administered AAV9-sTRAIL combats invasive glioblastoma in a patient-derived orthotopic xenograft model

    PubMed Central

    Crommentuijn, Matheus HW; Kantar, Rami; Noske, David P; Vandertop, W Peter; Badr, Christian E; Würdinger, Thomas; Maguire, Casey A; Tannous, Bakhos A

    2016-01-01

    Adeno-associated virus (AAV) vectors expressing tumoricidal genes injected directly into brain tumors have shown some promise, however, invasive tumor cells are relatively unaffected. Systemic injection of AAV9 vectors provides widespread delivery to the brain and potentially the tumor/microenvironment. Here we assessed AAV9 for potential glioblastoma therapy using two different promoters driving the expression of the secreted anti-cancer agent sTRAIL as a transgene model; the ubiquitously active chicken β-actin (CBA) promoter and the neuron-specific enolase (NSE) promoter to restrict expression in brain. Intravenous injection of AAV9 vectors encoding a bioluminescent reporter showed similar distribution patterns, although the NSE promoter yielded 100-fold lower expression in the abdomen (liver), with the brain-to-liver expression ratio remaining the same. The main cell types targeted by the CBA promoter were astrocytes, neurons and endothelial cells, while expression by NSE promoter mostly occurred in neurons. Intravenous administration of either AAV9-CBA-sTRAIL or AAV9-NSE-sTRAIL vectors to mice bearing intracranial patient-derived glioblastoma xenografts led to a slower tumor growth and significantly increased survival, with the CBA promoter having higher efficacy. To our knowledge, this is the first report showing the potential of systemic injection of AAV9 vector encoding a therapeutic gene for the treatment of brain tumors. PMID:27382645

  6. Selective In Vivo Targeting of Human Liver Tumors by Optimized AAV3 Vectors in a Murine Xenograft Model

    PubMed Central

    Wang, Yuan; Zhang, Yuanhui; Ejjigani, Anila; Yin, Zifei; Lu, Yuan; Wang, Lina; Wang, Meng; Li, Jun; Hu, Zhongbo; Aslanidi, George V.; Zhong, Li; Gao, Guangping

    2014-01-01

    Abstract Current challenges for recombinant adeno-associated virus (rAAV) vector–based cancer treatment include the low efficiency and the lack of specificity in vivo. rAAV serotype 3 (rAAV3) vectors have previously been shown to be ineffective in normal mouse tissues following systemic administration. In the present study, we report that rAAV3 vectors can efficiently target and transduce various human liver cancer cells in vivo. Elimination of specific surface-exposed serine and threonine residues on rAAV3 capsids results in further augmentation in the transduction efficiency of these vectors, without any change in the viral tropism and cellular receptor interactions. In addition, we have identified a potential chemotherapy drug, shikonin, as a multifunctional compound to inhibit liver tumor growth as well as to significantly enhance the efficacy of rAAV vector-based gene therapy in vivo. Furthermore, we also document that suppression of tumorigenesis in a human liver cancer xenograft model can be achieved through systemic administration of the optimized rAAV3 vectors carrying a therapeutic gene, and shikonin at a dose that does not lead to liver damage. Our research provides a novel means to achieve not only targeted delivery but also the potential for gene therapy of human liver cancer. PMID:25296041

  7. Selective in vivo targeting of human liver tumors by optimized AAV3 vectors in a murine xenograft model.

    PubMed

    Ling, Chen; Wang, Yuan; Zhang, Yuanhui; Ejjigani, Anila; Yin, Zifei; Lu, Yuan; Wang, Lina; Wang, Meng; Li, Jun; Hu, Zhongbo; Aslanidi, George V; Zhong, Li; Gao, Guangping; Srivastava, Arun; Ling, Changquan

    2014-12-01

    Current challenges for recombinant adeno-associated virus (rAAV) vector-based cancer treatment include the low efficiency and the lack of specificity in vivo. rAAV serotype 3 (rAAV3) vectors have previously been shown to be ineffective in normal mouse tissues following systemic administration. In the present study, we report that rAAV3 vectors can efficiently target and transduce various human liver cancer cells in vivo. Elimination of specific surface-exposed serine and threonine residues on rAAV3 capsids results in further augmentation in the transduction efficiency of these vectors, without any change in the viral tropism and cellular receptor interactions. In addition, we have identified a potential chemotherapy drug, shikonin, as a multifunctional compound to inhibit liver tumor growth as well as to significantly enhance the efficacy of rAAV vector-based gene therapy in vivo. Furthermore, we also document that suppression of tumorigenesis in a human liver cancer xenograft model can be achieved through systemic administration of the optimized rAAV3 vectors carrying a therapeutic gene, and shikonin at a dose that does not lead to liver damage. Our research provides a novel means to achieve not only targeted delivery but also the potential for gene therapy of human liver cancer.

  8. The impact of bone morphogenetic protein 4 (BMP4) on breast cancer metastasis in a mouse xenograft model.

    PubMed

    Ampuja, M; Alarmo, E L; Owens, P; Havunen, R; Gorska, A E; Moses, H L; Kallioniemi, A

    2016-06-01

    Bone morphogenetic protein 4 (BMP4) is a key regulator of cell proliferation and differentiation. In breast cancer cells, BMP4 has been shown to reduce proliferation in vitro and interestingly, in some cases, also to induce migration and invasion. Here we investigated whether BMP4 influences breast cancer metastasis formation by using a xenograft mouse model. MDA-MB-231 breast cancer cells were injected intracardially into mice and metastasis formation was monitored using bioluminescence imaging. Mice treated with BMP4 developed metastases slightly earlier as compared to control animals but the overall number of metastases was similar in both groups (13 in the BMP4 group vs. 12 in controls). In BMP4-treated mice, bone metastases were more common (10 vs. 7) but adrenal gland metastases were less frequent (1 vs. 5) than in controls. Immunostaining revealed no differences in signaling activation, proliferation rate, blood vessel formation, EMT markers or the number of cancer-associated fibroblasts between the treatment groups. In conclusion, BMP4 caused a trend towards accelerated metastasis formation, especially in bone. More work is needed to uncover the long-term effects of BMP4 and the clinical relevance of these findings. PMID:26970275

  9. Carbon nanotube lipid drug approach for targeted delivery of a chemotherapy drug in a human breast cancer xenograft animal model.

    PubMed

    Shao, Wei; Paul, Arghya; Zhao, Bin; Lee, Crystal; Rodes, Laetitia; Prakash, Satya

    2013-12-01

    Carbon nanotube (CNT) possesses excellent properties as a drug carrier. To overcome the challenge of drug functionalization with CNT, we have developed a lipid-drug approach for efficient drug loading onto CNT, in which a long chain lipid molecule is conjugated to the drug molecule so that the lipid-drug can be loaded directly onto CNT through binding of the lipid 'tail' in the drug molecule to CNT surfaces via hydrophobic interactions. In a proof-of-concept study, drug paclitaxel (PTX) was conjugated with a non-toxic lipid molecule docosanol for functionalization with CNT. Folic acid was also conjugated to CNT for targeted drug delivery. High level of drug loading onto SWNT could be achieved by lipid-drug approach. Conjugation of FA to SWNT-lipid-PTX led to an increase in cell penetration capacity, and the targeted SWNT-lipid-PTX showed much improved drug efficacy in vitro in comparison to free drug Taxol and non-targeted SWNT-lipid-PTX at 48 h (78.5% vs. 31.6% and 59.1% in cytotoxicity respectively, p < 0.01). In vivo analysis using a human breast cancer xenograft mice model also confirmed the improved drug efficacy. The targeted SWNT-lipid-PTX was found non-toxic as evaluated by biochemical analysis using blood samples, and by histological analysis of major organs.

  10. Smad6 determines BMP-regulated invasive behaviour of breast cancer cells in a zebrafish xenograft model

    PubMed Central

    de Boeck, Miriam; Cui, Chao; Mulder, Aat A; Jost, Carolina R; Ikeno, Souichi; ten Dijke, Peter

    2016-01-01

    The transforming growth factor-β (TGF-β) family is known to play critical roles in cancer progression. While the dual role of TGF-β is well described, the function of bone morphogenetic proteins (BMPs) is unclear. In this study, we established the involvement of Smad6, a BMP-specific inhibitory Smad, in breast cancer cell invasion. We show that stable overexpression of Smad6 in breast cancer MCF10A M2 cells inhibits BMP signalling, thereby mitigating BMP6-induced suppression of mesenchymal marker expression. Using a zebrafish xenograft model, we demonstrate that overexpression of Smad6 potentiates invasion of MCF10A M2 cells and enhances the aggressiveness of breast cancer MDA-MB-231 cells in vivo, whereas a reversed phenotype is observed after Smad6 knockdown. Interestingly, BMP6 pre-treatment of MDA-MB-231 cells induced cluster formation at the invasive site in the zebrafish. BMP6 also stimulated cluster formation of MDA-MB-231 cells co-cultured on Human Microvascular Endothelial Cells (HMEC)-1 in vitro. Electron microscopy illustrated an induction of cell-cell contact by BMP6. The clinical relevance of our findings is highlighted by a correlation of high Smad6 expression with poor distant metastasis free survival in ER-negative cancer patients. Collectively, our data strongly indicates the involvement of Smad6 and BMP signalling in breast cancer cell invasion in vivo. PMID:27113436

  11. Prediction of drug distribution in subcutaneous xenografts of human tumor cell lines and healthy tissues in mouse: application of the tissue composition-based model to antineoplastic drugs.

    PubMed

    Poulin, Patrick; Chen, Yung-Hsiang; Ding, Xiao; Gould, Stephen E; Hop, Cornelis Eca; Messick, Kirsten; Oeh, Jason; Liederer, Bianca M

    2015-04-01

    Advanced tissue composition-based models can predict the tissue-plasma partition coefficient (Kp ) values of drugs under in vivo conditions on the basis of in vitro and physiological input data. These models, however, focus on healthy tissues and do not incorporate data from tumors. The objective of this study was to apply a tissue composition-based model to six marketed antineoplastic drugs (docetaxel, DOC; doxorubicin, DOX; gemcitabine, GEM; methotrexate, MTX; topotecan, TOP; and fluorouracil, 5-FU) to predict their Kp values in three human tumor xenografts (HCT-116, H2122, and PC3) as well as in healthy tissues (brain, muscle, lung, and liver) under steady-state in vivo conditions in female NCR nude mice. The mechanisms considered in the tissue/tumor composition-based model are the binding to lipids and to plasma proteins, but the transporter effect was also investigated. The method consisted of analyzing tissue composition, performing the pharmacokinetics studies in mice, and calculating the corresponding in vivo Kp values. Analyses of tumor composition indicated that the tumor xenografts contained no or low amounts of common transporters by contrast to lipids. The predicted Kp values were within twofold and threefold of the measured values in 77% and 93% of cases, respectively. However, predictions for brain for each drug, for liver for MTX, and for each tumor xenograft for GEM were disparate from the observed values, and, therefore, not well served by the model. Overall, this study is the first step toward the mechanism-based prediction of Kp values of small molecules in healthy and tumor tissues in mouse when no transporter and permeation limitation effect is evident. This approach will be useful in selecting compounds based on their abilities to penetrate human cancer xenografts with a physiologically based pharmacokinetic (PBPK) model, thereby increasing therapeutic index for chemotherapy in oncology study.

  12. Ovarian Tumor Attachment, Invasion, and Vascularization Reflect Unique Microenvironments in the Peritoneum: Insights from Xenograft and Mathematical Models

    PubMed Central

    Steinkamp, Mara P.; Winner, Kimberly Kanigel; Davies, Suzy; Muller, Carolyn; Zhang, Yong; Hoffman, Robert M.; Shirinifard, Abbas; Moses, Melanie; Jiang, Yi; Wilson, Bridget S.

    2013-01-01

    Ovarian cancer relapse is often characterized by metastatic spread throughout the peritoneal cavity with tumors attached to multiple organs. In this study, interaction of ovarian cancer cells with the peritoneal tumor microenvironment was evaluated in a xenograft model based on intraperitoneal injection of fluorescent SKOV3.ip1 ovarian cancer cells. Intra-vital microscopy of mixed GFP-red fluorescent protein (RFP) cell populations injected into the peritoneum demonstrated that cancer cells aggregate and attach as mixed spheroids, emphasizing the importance of homotypic adhesion in tumor formation. Electron microscopy provided high resolution structural information about local attachment sites. Experimental measurements from the mouse model were used to build a three-dimensional cellular Potts ovarian tumor model (OvTM) that examines ovarian cancer cell attachment, chemotaxis, growth, and vascularization. OvTM simulations provide insight into the relative influence of cancer cell–cell adhesion, oxygen availability, and local architecture on tumor growth and morphology. Notably, tumors on the mesentery, omentum, or spleen readily invade the “open” architecture, while tumors attached to the gut encounter barriers that restrict invasion and instead rapidly expand into the peritoneal space. Simulations suggest that rapid neovascularization of SKOV3.ip1 tumors is triggered by constitutive release of angiogenic factors in the absence of hypoxia. This research highlights the importance of cellular adhesion and tumor microenvironment in the seeding of secondary ovarian tumors on diverse organs within the peritoneal cavity. Results of the OvTM simulations indicate that invasion is strongly influenced by features underlying the mesothelial lining at different sites, but is also affected by local production of chemotactic factors. The integrated in vivo mouse model and computer simulations provide a unique platform for evaluating targeted therapies for ovarian cancer

  13. Biosynthesized Platinum Nanoparticles Inhibit the Proliferation of Human Lung-Cancer Cells in vitro and Delay the Growth of a Human Lung-Tumor Xenograft in vivo

    PubMed Central

    Yogesh, Bendale; Vineeta, Bendale; Rammesh, Natu; Saili, Paul

    2016-01-01

    Objectives: Lung cancer remains a deadly disease with unsatisfactory overall survival. Cisplatin, a standard platinum (Pt)-based chemotherapeutic agent, has the potential to inhibit the growth of lung cancer. Its use, however, is occasionally limited by severe organ toxicity. However, until now, no systematic study has been conducted to verify its efficacy with proper experimental support in vivo. Therefore, we examined whether biosynthesized Pt nanoparticles (NPs) inhibited human lung cancer in vitro and in vivo to validate their use in alternative and complementary medicine. Methods: We evaluated the in vitro and the in vivo anticancer efficiencies of biosynthesized Pt NPs in a subcutaneous xenograft model with A549 cells. Severe combined immune deficient mice (SCID) were divided into four groups: group 1 being the vehicle control group and groups 2, 3 and 4 being the experimental groups. Once the tumor volume had reached 70 ─ 75 mm3, the progression profile of the tumor growth kinetics and the body weights of the mice were measured every week for 6 weeks after oral administration of Pt NPs. Doses of Pt NPs of 500, 1,000 and 2,000 mg/kg of body weight were administered to the experimental groups and a dose of honey was administered to the vehicle control group. The efficacy was quantified by using the delay in tumor growth following the administration of Pt NPs of A549 human-lung-cancer xenografts growing in SCID mice. Results: The in vitro cytotoxicity evaluation indicated that Pt NPs, in a dose-dependent manner, inhibited the growth of A549 cells, and the in vivo evaluation showed that Pt NPs at the mid and high doses effectively inhibited and delayed the growth of lung cancer in SCID mice. Conclusion: These findings confirm the antitumor properties of biosynthesized Pt NPs and suggest that they may be a cost-effective alternative for the treatment of patients with lung cancer. PMID:27386144

  14. The HDACi Panobinostat Shows Growth Inhibition Both In Vitro and in a Bioluminescent Orthotopic Surgical Xenograft Model of Ovarian Cancer

    PubMed Central

    Helland, Øystein; Popa, Mihaela; Bischof, Katharina; Gjertsen, Bjørn Tore; McCormack, Emmet; Bjørge, Line

    2016-01-01

    Background In most epithelial ovarian carcinomas (EOC), epigenetic changes are evident, and overexpression of histone deacetylases (HDACs) represents an important manifestation. In this study, we wanted to evaluate the effects of the novel HDAC inhibitor (HDACi) panobinostat, both alone and in combination with carboplatin, on ovarian cancer cell lines and in a murine bioluminescent orthotopic surgical xenograft model for EOC. Methods The effects of panobinostat, both alone and in combination with carboplatin, on proliferation and apoptosis in ovarian cancer cell lines, were evaluated using colony and WST-1 assays, Hoechst staining and flow cytometry analysis. In addition, mechanisms were characterised by western blotting and phosphoflow analysis. Immuno-deficient mice were engrafted orthotopically with SKOV-3luc+ cells and serial bioluminescence imaging monitored the effects of treatment with panobinostat and/or carboplatin and/or surgery. Survival parameters were also measured. Results Panobinostat treatment reduced cell growth and diminished cell viability, as shown by the induced cell cycle arrest and apoptosis in vitro. We observed increased levels of cleaved PARP and caspase-3, downregulation of cdc2 protein kinase, acetylation of H2B and higher pH2AX expression. The combined administration of carboplatin and panobinostat synergistically increased the anti-tumour effects compared to panobinostat or carboplatin treatment alone. In our novel ovarian cancer model, the mice showed significantly higher rates of survival when treated with panobinostat, carboplatin or a combination of both, compared to the controls. Panobinostat was as efficient as carboplatin regarding prolongation of survival. No significant additional effect on survival was observed when surgery was combined with carboplatin/panobinostat treatment. Conclusions Panobinostat demonstrates effective in vitro growth inhibition in ovarian cancer cells. The efficacy of panobinostat and carboplatin was

  15. The BET bromodomain inhibitor JQ1 suppresses growth of pancreatic ductal adenocarcinoma in patient-derived xenograft models.

    PubMed

    Garcia, P L; Miller, A L; Kreitzburg, K M; Council, L N; Gamblin, T L; Christein, J D; Heslin, M J; Arnoletti, J P; Richardson, J H; Chen, D; Hanna, C A; Cramer, S L; Yang, E S; Qi, J; Bradner, J E; Yoon, K J

    2016-02-18

    The primary aim of this study was to evaluate the antitumor efficacy of the bromodomain inhibitor JQ1 in pancreatic ductal adenocarcinoma (PDAC) patient-derived xenograft (tumorgraft) models. A secondary aim of the study was to evaluate whether JQ1 decreases expression of the oncogene c-Myc in PDAC tumors, as has been reported for other tumor types. We used five PDAC tumorgraft models that retain specific characteristics of tumors of origin to evaluate the antitumor efficacy of JQ1. Tumor-bearing mice were treated with JQ1 (50 mg/kg daily for 21 or 28 days). Expression analyses were performed with tumors harvested from host mice after treatment with JQ1 or vehicle control. An nCounter PanCancer Pathways Panel (NanoString Technologies) of 230 cancer-related genes was used to identify gene products affected by JQ1. Quantitative RT-PCR, immunohistochemistry and immunoblots were carried out to confirm that changes in RNA expression reflected changes in protein expression. JQ1 inhibited the growth of all five tumorgraft models (P<0.05), each of which harbors a KRAS mutation; but induced no consistent change in expression of c-Myc protein. Expression profiling identified CDC25B, a regulator of cell cycle progression, as one of the three RNA species (TIMP3, LMO2 and CDC25B) downregulated by JQ1 (P<0.05). Inhibition of tumor progression was more closely related to decreased expression of nuclear CDC25B than to changes in c-Myc expression. JQ1 and other agents that inhibit the function of proteins with bromodomains merit further investigation for treating PDAC tumors. Work is ongoing in our laboratory to identify effective drug combinations that include JQ1.

  16. A Novel Bufalin Derivative Exhibited Stronger Apoptosis-Inducing Effect than Bufalin in A549 Lung Cancer Cells and Lower Acute Toxicity in Mice

    PubMed Central

    Liu, Miao; Feng, Li-Xing; Sun, Peng; Liu, Wang; Wu, Wan-Ying; Jiang, Bao-Hong; Yang, Min; Hu, Li-Hong; Guo, De-An; Liu, Xuan

    2016-01-01

    BF211 is a synthetic molecule derived from bufalin (BF). The apoptosis-inducing effect of BF211 was stronger than that of BF while the acute toxicity of BF211 was much lower than that of BF. BF211 exhibited promising concentration-dependent anti-cancer effects in nude mice inoculated with A549 cells in vivo. The growth of A549 tumor xenografts was almost totally blocked by treatment with BF211 at 6 mg/kg. Notably, BF and BF211 exhibited differences in their binding affinity and kinetics to recombinant proteins of the α subunits of Na+/K+-ATPase. Furthermore, there was a difference in the effects of BF or BF211 on inhibiting the activity of porcine cortex Na+/K+-ATPase and in their time-dependent effects on intracellular Ca2+ levels in A549 cells. The time-dependent effects of BF or BF211 on the activation of Src, which was mediated by the Na+/K+-ATPase signalosome, in A549 cells were also different. Both BF and BF211 could induce apoptosis-related cascades, such as activation of caspase-3 and the cleavage of PARP (poly ADP-ribose polymerase) in A549 cells, in a concentration-dependent manner; however, the effects of BF211 on apoptosis-related cascades was stronger than that of BF. The results of the present study supported the importance of binding to the Na+/K+-ATPase α subunits in the mechanism of cardiac steroids and also suggested the possibility of developing new cardiac steroids with a stronger anti-cancer activity and lower toxicity as new anti-cancer agents. PMID:27459387

  17. A multifunctional drug combination shows highly potent therapeutic efficacy against human cancer xenografts in athymic mice.

    PubMed

    Liu, Xiu-Jun; Zheng, Yan-Bo; Li, Yi; Wu, Shu-Ying; Zhen, Yong-Su

    2014-01-01

    The tumor microenvironment plays a crucial role during tumor development. Integrated combination of drugs that target tumor microenvironment is a promising approach to anticancer therapy. Here, we report a multifunctional combination of low-cytotoxic drugs composed of dipyridamole, bestatin and dexamethasone (DBDx) which mainly acts on the tumor microenvironment shows highly potent antitumor efficacy in vivo. In mouse hepatoma H22 model, the triple drug combination showed synergistic and highly potent antitumor efficacy. The combination indices of various combinations of the triple drugs were between 0.2 and 0.5. DBDx inhibited the growth of a panel of human tumor xenografts and showed no obvious systemic toxicity. At tolerated doses, DBDx suppressed the growth of human hepatocellular carcinoma BEL-7402, HepG2, and lung adenocarcinoma A549 xenografts by 94.5%, 93.7% and 96.9%, respectively. Clonogenic assay demonstrated that DBDx showed weak cytotoxicity. Western blot showed that Flk1 and Nos3 were down-regulated in the DBDx-treated group. Proteomic analysis showed that DBDx mainly affected the metabolic process and immune system process; in addition, the angiogenesis and VEGF signaling pathway were also affected. Conclusively, DBDx, a multifunctional drug combination of three low-cytotoxic drugs, shows synergistic and highly potent antitumor efficacy evidently mediated by the modulation of tumor microenvironment. Based on its low-cytotoxic attributes and its broad-spectrum antitumor therapeutic efficacy, this multifunctional combination might be useful in the treatment of cancers, especially those refractory to conventional chemotherapeutics.

  18. Mouse Models in Prostate Cancer Translational Research: From Xenograft to PDX

    PubMed Central

    del Vecchio, Vitale; Palma, Giuseppe; Barbieri, Antonio; Falco, Michela; Luciano, Antonio; De Biase, Davide; Perdonà, Sisto; Facchini, Gaetano; Arra, Claudio

    2016-01-01

    Despite the advancement of clinical and preclinical research on PCa, which resulted in the last five years in a decrement of disease incidence by 3-4%, it remains the most frequent cancer in men and the second for mortality rate. Based on this evidence we present a brief dissertation on numerous preclinical models, comparing their advantages and disadvantages; among this we report the PDX mouse models that show greater fidelity to the disease, in terms of histopathologic features of implanted tumor, gene and miRNA expression, and metastatic pattern, well describing all tumor progression stages; this characteristic encourages the translation of preclinical results. These models become particularly useful in meeting the need of new treatments identification that eradicate PCa bone metastases growing, clarifying pathway of angiogenesis, identifying castration-resistant stem-like cells, and studying the antiandrogen therapies. Also of considerable interest are the studies of 3D cell cultures derived from PDX, which have the ability to maintain PDX cell viability with continued native androgen receptor expression, also showing a differential sensitivity to drugs. 3D PDX PCa may represent a diagnostic platform for the rapid assessment of drugs and push personalized medicine. Today the development of preclinical models in vitro and in vivo is necessary in order to obtain increasingly reliable answers before reaching phase III of the drug discovery. PMID:27294148

  19. Mouse Models in Prostate Cancer Translational Research: From Xenograft to PDX.

    PubMed

    Rea, Domenica; Del Vecchio, Vitale; Palma, Giuseppe; Barbieri, Antonio; Falco, Michela; Luciano, Antonio; De Biase, Davide; Perdonà, Sisto; Facchini, Gaetano; Arra, Claudio

    2016-01-01

    Despite the advancement of clinical and preclinical research on PCa, which resulted in the last five years in a decrement of disease incidence by 3-4%, it remains the most frequent cancer in men and the second for mortality rate. Based on this evidence we present a brief dissertation on numerous preclinical models, comparing their advantages and disadvantages; among this we report the PDX mouse models that show greater fidelity to the disease, in terms of histopathologic features of implanted tumor, gene and miRNA expression, and metastatic pattern, well describing all tumor progression stages; this characteristic encourages the translation of preclinical results. These models become particularly useful in meeting the need of new treatments identification that eradicate PCa bone metastases growing, clarifying pathway of angiogenesis, identifying castration-resistant stem-like cells, and studying the antiandrogen therapies. Also of considerable interest are the studies of 3D cell cultures derived from PDX, which have the ability to maintain PDX cell viability with continued native androgen receptor expression, also showing a differential sensitivity to drugs. 3D PDX PCa may represent a diagnostic platform for the rapid assessment of drugs and push personalized medicine. Today the development of preclinical models in vitro and in vivo is necessary in order to obtain increasingly reliable answers before reaching phase III of the drug discovery.

  20. RAD1901: a novel, orally bioavailable selective estrogen receptor degrader that demonstrates antitumor activity in breast cancer xenograft models

    PubMed Central

    Shomali, Maysoun; Paquin, Dotty; Lyttle, C. Richard; Hattersley, Gary

    2015-01-01

    Agents that inhibit estrogen production, such as aromatase inhibitors or those that directly block estrogen receptor (ER) activity, such as selective estrogen receptor modulators and selective estrogen receptor degraders, are routinely used in the treatment of ER-positive breast cancers. However, although initial treatment with these agents is often successful, many women eventually relapse with drug-resistant breast cancers. To overcome some of the challenges associated with current endocrine therapies and to combat the development of resistance, there is a need for more durable and more effective ER-targeted therapies. Here we describe and characterize a novel, orally bioavailable small-molecule selective estrogen receptor degrader, RAD1901, and evaluate its therapeutic potential for the treatment of breast cancer. RAD1901 selectively binds to and degrades the ER and is a potent antagonist of ER-positive breast cancer cell proliferation. Importantly, RAD1901 produced a robust and profound inhibition of tumor growth in MCF-7 xenograft models. In an intracranial MCF-7 model, RAD1901-treated animals survived longer than those treated with either control or fulvestrant, suggesting the potential benefit of RAD1901 in the treatment of ER-positive breast cancer that has metastasized to the brain. Finally, RAD1901 preserved ovariectomy-induced bone loss and prevented the uterotropic effects of E2, suggesting that it may act selectively as an agonist in bone but as an antagonist in breast and uterine tissues. RAD1901 is currently under clinical study in postmenopausal women with ER-positive advanced breast cancer. PMID:26164151

  1. Human cytomegalovirus infection leads to elevated levels of transplant arteriosclerosis in a humanized mouse aortic xenograft model.

    PubMed

    Abele-Ohl, S; Leis, M; Wollin, M; Mahmoudian, S; Hoffmann, J; Müller, R; Heim, C; Spriewald, B M; Weyand, M; Stamminger, T; Ensminger, S M

    2012-07-01

    Recent findings emphasized an important role of human cytomegalovirus (HCMV) infection in the development of transplant arteriosclerosis. Therefore, the aim of this study was to develop a human peripheral blood lymphocyte (hu-PBL)/Rag-2(-/-) γc(-/-) mouse-xenograft-model to investigate both immunological as well as viral effector mechanisms in the progression of transplant arteriosclerosis. For this, sidebranches from the internal mammary artery were recovered during coronary artery bypass graft surgery, tissue-typed and infected with HCMV. Then, size-matched sidebranches were implanted into the infrarenal aorta of Rag-2(-/-) γc(-/-) mice. The animals were reconstituted with human peripheral blood mononuclear cells (PBMCs) 7 days after transplantation. HCMV-infection was confirmed by Taqman-PCR and immunofluorescence analyses. Arterial grafts were analyzed by histology on day 40 after transplantation. PBMC-reconstituted Rag-2(-/-) γc(-/-) animals showed splenic chimerism levels ranging from 1-16% human cells. After reconstitution, Rag-2(-/-) γc(-/-) mice developed human leukocyte infiltrates in their grafts and vascular lesions that were significantly elevated after infection. Cellular infiltration revealed significantly increased ICAM-1 and PDGF-R-β expression after HCMV-infection of the graft. Arterial grafts from unreconstituted Rag-2(-/-) γc(-/-) recipients showed no vascular lesions. These data demonstrate a causative relationship between HCMV-infection as an isolated risk factor and the development of transplant-arteriosclerosis in a humanized mouse arterial-transplant-model possibly by elevated ICAM-1 and PDGF-R-β expression.

  2. Targeted Hsp70 expression combined with CIK-activated immune reconstruction synergistically exerts antitumor efficacy in patient-derived hepatocellular carcinoma xenograft mouse models.

    PubMed

    Hu, Huanzhang; Qiu, Yinghe; Guo, Minggao; Huang, Yao; Fang, Lin; Peng, Zhangxiao; Ji, Weidan; Xu, Yang; Shen, Shuwen; Yan, Yan; Huang, Xuandong; Zheng, Junnian; Su, Changqing

    2015-01-20

    The patient-derived tumor xenograft (PDTX) models can reproduce a similar natural genetic background and similar biological behaviors to tumor cells in patients, which is conducive to the assessment of personalized cancer treatment. In this study, to verify the targeting and effectiveness of the therapeutic strategy using a Survivin promoter-regulated oncolytic adenovirus expressing Hsp70, the PDTX models of hepatocellular carcinoma (HCC) were established in nude mice and the cytokine-induced killer (CIK) cells were intravenously infused into mice to partially reconstruct the mouse immune function. The results demonstrated that, either the immune anti-tumor effect caused by CIK cell infusion or the oncolytic effect generated by oncolytic adenovirus replication was very limited. However, the synergistic tumor inhibitory effect was significantly enhanced after treatments with oncolytic adenovirus expressing Hsp70 combined with CIK cells. Oncolytic adenovirus mediated the specific expression of Hsp70 in cancer tissues allowed the CIK chemotaxis, and induce the infiltration of CD3+ T cells in tumor stroma, thereby exhibiting anti-tumor activity. The anti-tumor effect was more effective for the highly malignant tumor xenografts with highly Survivin expression. This strategy can synergistically activate multiple anti-tumor mechanisms and exert effective anti-tumor activities that have a significant inhibitory effect against the growth of HCC xenografts.

  3. Multicolor Fluorescent Intravital Live Microscopy (FILM) for Surgical Tumor Resection in a Mouse Xenograft Model

    PubMed Central

    Thurber, Greg M.; Figueiredo, Jose L.; Weissleder, Ralph

    2009-01-01

    Background Complete surgical resection of neoplasia remains one of the most efficient tumor therapies. However, malignant cell clusters are often left behind during surgery due to the inability to visualize and differentiate them against host tissue. Here we establish the feasibility of multicolor fluorescent intravital live microscopy (FILM) where multiple cellular and/or unique tissue compartments are stained simultaneously and imaged in real time. Methodology/Principal Findings Theoretical simulations of imaging probe localization were carried out for three agents with specificity for cancer cells, stromal host response, or vascular perfusion. This transport analysis gave insight into the probe pharmacokinetics and tissue distribution, facilitating the experimental design and allowing predictions to be made about the localization of the probes in other animal models and in the clinic. The imaging probes were administered systemically at optimal time points based on the simulations, and the multicolor FILM images obtained in vivo were then compared to conventional pathological sections. Our data show the feasibility of real time in vivo pathology at cellular resolution and molecular specificity with excellent agreement between intravital and traditional in vitro immunohistochemistry. Conclusions/Significance Multicolor FILM is an accurate method for identifying malignant tissue and cells in vivo. The imaging probes distributed in a manner similar to predictions based on transport principles, and these models can be used to design future probes and experiments. FILM can provide critical real time feedback and should be a useful tool for more effective and complete cancer resection. PMID:19956597

  4. The in ovo CAM-assay as a xenograft model for sarcoma.

    PubMed

    Sys, Gwen M L; Lapeire, Lore; Stevens, Nikita; Favoreel, Herman; Forsyth, Ramses; Bracke, Marc; De Wever, Olivier

    2013-01-01

    Sarcoma is a very rare disease that is heterogeneous in nature, all hampering the development of new therapies. Sarcoma patients are ideal candidates for personalized medicine after stratification, explaining the current interest in developing a reproducible and low-cost xenotransplant model for this disease. The chick chorioallantoic membrane is a natural immunodeficient host capable of sustaining grafted tissues and cells without species-specific restrictions. In addition, it is easily accessed, manipulated and imaged using optical and fluorescence stereomicroscopy. Histology further allows detailed analysis of heterotypic cellular interactions. This protocol describes in detail the in ovo grafting of the chorioallantoic membrane with fresh sarcoma-derived tumor tissues, their single cell suspensions, and permanent and transient fluorescently labeled established sarcoma cell lines (Saos-2 and SW1353). The chick survival rates are up to 75%. The model is used to study graft- (viability, Ki67 proliferation index, necrosis, infiltration) and host (fibroblast infiltration, vascular ingrowth) behavior. For localized grafting of single cell suspensions, ECM gel provides significant advantages over inert containment materials. The Ki67 proliferation index is related to the distance of the cells from the surface of the CAM and the duration of application on the CAM, the latter determining a time frame for the addition of therapeutic products. PMID:23892612

  5. Porphyrin lipid nanoparticles for enhanced photothermal therapy in a patient-derived orthotopic pancreas xenograft cancer model

    NASA Astrophysics Data System (ADS)

    MacLaughlin, Christina M.; Ding, Lili; Jin, Cheng; Cao, Pingjiang; Siddiqui, Iram; Hwang, David M.; Chen, Juan; Wilson, Brian C.; Zheng, Gang; Hedley, David W.

    2016-03-01

    Local disease control is a major problem in the treatment of pancreatic cancer, because curative-intent surgery is only possible in a minority of patients, and radiotherapy cannot be delivered in curative doses. Despite the promise of photothermal therapy (PTT) for ablation of pancreatic tumors, this approach remains under investigated. Using photothermal sensitizers in combination with laser light for PTT can result in more efficient conversion of light energy to heat, and confinement of thermal destruction to the tumor, thus sparing adjacent organs and vasculature. Porphyrins have been previously employed as photosensitizers for PDT and PTT, however their incorporation in to "porphysomes", lipid-based nanoparticles each containing ~80,000 porphyrins through conjugation of pyropheophorbide to phospholipids, carries two distinct advantages: 1) high-density porphyrin packing imparts the nanoparticles with enhanced photonic properties for imaging and phototherapy; 2) the enhanced permeability and retention effect may be exploited for optimal delivery of porphysomes to the tumor region thus high payload porphyrin delivery. The feasibility of porphysome-enhanced PTT for pancreatic cancer treatment was investigated using a patient-derived orthotopic pancreas xenograft tumor model. Uptake of porphysomes at the orthotopic tumor site was validated using ex vivo fluorescence imaging of intact organs of interest. The accumulation of porphysomes in orthotopic tumor microstructure was also confirmed by fluorescence imaging of excised tissue slices. PTT progress was monitored as changes in tumor surface temperature using IR optical imaging. Histological analyses were conducted to examine microstructure changes in tissue morphology, and the viability of remaining tumor tissues following exposure to heat. These studies may also provide insight as to the contribution of heat sink in application of thermal therapies to highly vascularized pancreatic tumors.

  6. Naltrindole Inhibits Human Multiple Myeloma Cell Proliferation In Vitro and in a Murine Xenograft Model In Vivo

    PubMed Central

    Mundra, Jyoti Joshi; Terskiy, Alexandra

    2012-01-01

    It has been demonstrated previously that immune cell activation and proliferation were sensitive to the effects of naltrindole, a nonpeptidic δ-opioid receptor-selective antagonist; therefore, we hypothesized that human multiple myeloma (MM) would be a valuable model for studying potential antineoplastic properties of naltrindole. [3H]naltrindole exhibited saturable, low-affinity binding to intact human MM cells; however, the pharmacological profile of the binding site differed considerably from the properties of δ-, κ-, and μ-opioid receptors, and opioid receptor mRNA was not detected in MM cells by reverse transcriptase-polymerase chain reaction. Naltrindole inhibited the proliferation of cultured human U266 MM cells in a time- and dose-dependent manner with an EC50 of 16 μM. The naltrindole-induced inhibition of U266 cell proliferation was not blocked by a 10-fold molar excess of naltrexone, a nonselective opioid antagonist. Additive inhibition of MM cell proliferation was observed when using a combination of naltrindole with the histone deacetylase inhibitor sodium valproate, the proteasome inhibitor bortezomib, the glucocorticoid receptor agonist dexamethasone, and the HMG CoA reductase inhibitor simvastatin. Treatment of U266 cells with naltrindole significantly decreased the level of the active, phosphorylated form of the kinases, extracellular signal-regulated kinase and Akt, which may be related to its antiproliferative activity. The antiproliferative activity of naltrindole toward MM cells was maintained in cocultures of MM and bone marrow-derived stromal cells, mimicking the bone marrow microenvironment. In vivo, naltrindole significantly decreased tumor cell volumes in human MM cell xenografts in severe combined immunodeficient mice. We hypothesize that naltrindole inhibits the proliferation of MM cells through a nonopioid receptor-dependent mechanism. PMID:22537770

  7. Combined magnetic resonance, fluorescence, and histology imaging strategy in a human breast tumor xenograft model

    PubMed Central

    Jiang, Lu; Greenwood, Tiffany R.; Amstalden van Hove, Erika R.; Chughtai, Kamila; Raman, Venu; Winnard, Paul T.; Heeren, Ron; Artemov, Dmitri; Glunde, Kristine

    2014-01-01

    Applications of molecular imaging in cancer and other diseases frequently require combining in vivo imaging modalities, such as magnetic resonance and optical imaging, with ex vivo optical, fluorescence, histology, and immunohistochemical (IHC) imaging, to investigate and relate molecular and biological processes to imaging parameters within the same region of interest. We have developed a multimodal image reconstruction and fusion framework that accurately combines in vivo magnetic resonance imaging (MRI) and magnetic resonance spectroscopic imaging (MRSI), ex vivo brightfield and fluorescence microscopic imaging, and ex vivo histology imaging. Ex vivo brightfield microscopic imaging was used as an intermediate modality to facilitate the ultimate link between ex vivo histology and in vivo MRI/MRSI. Tissue sectioning necessary for optical and histology imaging required generation of a three-dimensional (3D) reconstruction module for 2D ex vivo optical and histology imaging data. We developed an external fiducial marker based 3D reconstruction method, which was able to fuse optical brightfield and fluorescence with histology imaging data. Registration of 3D tumor shape was pursued to combine in vivo MRI/MRSI and ex vivo optical brightfield and fluorescence imaging data. This registration strategy was applied to in vivo MRI/MRSI, ex vivo optical brightfield/fluorescence, as well as histology imaging data sets obtained from human breast tumor models. 3D human breast tumor data sets were successfully reconstructed and fused with this platform. PMID:22945331

  8. Uptake of verteporfin by orthotopic xenograft pancreas models with different levels of aggression

    NASA Astrophysics Data System (ADS)

    O'Hara, Julia; Samkoe, Kimberley S.; Chen, Alina; Hoopes, P. Jack; Rizvi, Imran; Hasan, Tayyaba; Pogue, Brian W.

    2009-06-01

    Pancreatic cancer is an aggressive disease with a poor prognosis, usually treated with chemoradiation therapy. Interstitial photodynamic therapy is a potentially effective adjuvant treatment that is under development. In the current study, two orthotopic pancreatic cancer models (AsPC-1 and Panc-1), have been characterized with respect to growth rates, morphology and liposomal drug (Verteporfin) uptake and distribution in SCID mice. Fluorescence of Verteporfin was measured in liver and tumor in vivo using a PDT fluorescence dosimeter with measurements taken before and up to one hour after tail vein injection. Fluorescence reached a plateau by about 15 minutes and did not decrease over the first hour. At time points from 15 minutes to 24 hrs, the internal organs (kidney, spleen, pancreas, tumor, muscle, lung, liver, and skin were excised and scanned on a Typhoon imager. The ratio of fluorescence in tumor versus normal tissues was analyzed with image processing, calculated at each time point and compared to in vivo results. Tissue distribution of Verteporfin in relation to functional vasculature marked by DiOc7 was carried out on frozen sections. Final analysis will result in determination of the ideal time point to administer light to achieve maximum tumor destruction while preserving normal tissue.

  9. Effect of Citrus bergamia juice on human neuroblastoma cells in vitro and in metastatic xenograft models.

    PubMed

    Navarra, M; Ursino, M R; Ferlazzo, N; Russo, M; Schumacher, U; Valentiner, U

    2014-06-01

    Neuroblastoma is the most common extracranial pediatric solid tumor with poor prognosis in children with disseminated stage of disease. A number of studies show that molecules largely distributed in commonly consumed fruits and vegetables may have anti-tumor activity. In this study we evaluate the effect of Citrus bergamia (bergamot) juice (BJ) in vitro and in a spontaneous metastatic neuroblastoma SCID mouse model. Qualitative and quantitative characterizations of BJ flavonoid fractions were performed by RP-HPLC/PDA/MS. We show that BJ significantly affects SK-N-SH and LAN-1 cell proliferation in vitro, but fails to reduce primary tumor weight in vivo. Moreover, BJ reduced cell adhesiveness and invasion of LAN-1 and SK-N-SH cells in vitro and the number of pulmonary metastases under consideration of the number of tumor cells in the blood in mice inoculated with LAN-1 cells in vivo. These effects without any apparent sign of systemic toxicity confirm the potential clinical interest of BJ and lay the basis for further investigation in cancer.

  10. Supersonic Shear Wave Elastography of Response to Anti-cancer Therapy in a Xenograft Tumor Model.

    PubMed

    Chamming's, Foucauld; Le-Frère-Belda, Marie-Aude; Latorre-Ossa, Heldmuth; Fitoussi, Victor; Redheuil, Alban; Assayag, Franck; Pidial, Laetitia; Gennisson, Jean-Luc; Tanter, Mickael; Cuénod, Charles-André; Fournier, Laure S

    2016-04-01

    Our objective was to determine if supersonic shear wave elastography (SSWE) can detect changes in stiffness of a breast cancer model under therapy. A human invasive carcinoma was implanted in 22 mice. Eleven were treated with an anti-angiogenic therapy and 11 with glucose for 24 d. Tumor volume and stiffness were assessed during 2 wk before treatment and 0, 7, 12, 20 and 24 d after the start of therapy using SSWE. Pathology was assessed after 12 and 24 d of treatment. We found that response to therapy was associated with early softening of treated tumors only, resulting in a significant difference from non-treated tumors after 12 d of treatment (p = 0.03). On pathology, large areas of necrosis were observed at 12 d in treated tumors. Although treatment was still effective, treated tumors subsequently stiffened during a second phase of the treatment (days 12-24), with a small amount of necrosis observed on pathology on day 24. In conclusion, SSWE was able to measure changes in the stiffness of tumors in response to anti-cancer treatment. However, stiffness changes associated with good response to treatment may change over time, and increased stiffness may also reflect therapy efficacy. PMID:26746382

  11. Human brain tumor xenografts in nude mice as a chemotherapy model.

    PubMed

    Houchens, D P; Ovejera, A A; Riblet, S M; Slagel, D E

    1983-06-01

    Two human brain tumors which were previously established in nude mice were used to determine antitumor efficacy of various therapeutic agents. These tumors were a medulloblastoma (TE-671) and a glioma (U-251) with mass doubling times of 3.5 and 5.5 days respectively as subcutaneous implants in nude mice. Intracranial (i.c.) tumor challenge was accomplished by inoculating tissue culture-grown cells of either tumor into the right cerebral hemisphere to a depth of 3 mm. Median survival time (MST) in untreated mice with 10(5) i.c. injected TE-671 cells was approximately 30 days and 53 days in the U-251 tumor. With 2 X 10(5) U-251 tumor cells the MST was 27-31 days. Groups of mice which had been inoculated with tumor were treated with various doses and schedules of antineoplastic compounds by the i.p. route. The TE-671 tumor responded to AZQ treatment with an increase in life span (ILS) of 37% compared to untreated controls and an ILS of 30% with CCNU treatment. BCNU and PCNU were ineffective. With the U-251 tumor BCNU produced an ILS of greater than 60%, with 75% cures, greater than 112% ILS with PCNU and 49% ILS with CCNU. Neither tumor responded to procarbazine, PALA, dianhydrogalactitol, D-O-norleucine or dibromodulcitol. The U-251 tumor was treated on various schedules and doses with BCNU and found to respond well on late as well as early treatment. A new drug (rapamycin) being investigated by the NCI was found to be very effective against the U-251 tumor. This model system should prove valuable in assessing the effects of various chemotherapeutic modalities against brain tumors.

  12. Evaluation of the NOD/SCID xenograft model for glucocorticoid-regulated gene expression in childhood B-cell precursor acute lymphoblastic leukemia

    PubMed Central

    2011-01-01

    Background Glucocorticoids such as prednisolone and dexamethasone are critical drugs used in multi-agent chemotherapy protocols used to treat acute lymphoblastic leukemia (ALL), and response to glucocorticoids is highly predictive of outcome. The NOD/SCID xenograft mouse model of ALL is a clinically relevant model in which the mice develop a systemic leukemia which retains the fundamental biological characteristics of the original disease. Here we report a study evaluating the NOD/SCID xenograft mouse model to investigate glucocorticoid-induced gene expression. Cells from a glucocorticoid-sensitive xenograft derived from a child with B-cell precursor ALL were inoculated into NOD/SCID mice. When highly engrafted the mice were randomized into groups of 4 to receive dexamethasone 15 mg/kg by intraperitoneal injection or vehicle control. Leukemia cells were harvested from mice spleens at 0, 8, 24 or 48 hours thereafter, and gene expression analyzed on Illumina WG-6_V3 chips, comparing all groups to time 0 hours. Results The 8 hour dexamethasone-treated timepoint had the highest number of significantly differentially expressed genes, with fewer observed at the 24 and 48 hour timepoints, and with minimal changes seen across the time-matched controls. When compared to publicly available datasets of glucocorticoid-induced gene expression from an in vitro cell line study and from an in vivo study of patients with ALL, at the level of pathways, expression changes in the 8 hour xenograft samples showed a similar response to patients treated with glucocorticoids. Replicate analysis revealed that at the 8 hour timepoint, a dataset with high signal and differential expression, using data from 3 replicates instead of 4 resulted in excellent recovery scores of > 0.9. However at other timepoints with less signal very poor recovery scores were obtained with 3 replicates. Conclusions The NOD/SCID xenograft mouse model provides a reproducible experimental system in which to

  13. A novel xenograft model to study the role of TSLP-induced CRLF2 signals in normal and malignant human B lymphopoiesis.

    PubMed

    Francis, Olivia L; Milford, Terry-Ann M; Martinez, Shannalee R; Baez, Ineavely; Coats, Jacqueline S; Mayagoitia, Karina; Concepcion, Katherine R; Ginelli, Elizabeth; Beldiman, Cornelia; Benitez, Abigail; Weldon, Abby J; Arogyaswamy, Keshav; Shiraz, Parveen; Fisher, Ross; Morris, Christopher L; Zhang, Xiao-Bing; Filippov, Valeri; Van Handel, Ben; Ge, Zheng; Song, Chunhua; Dovat, Sinisa; Su, Ruijun Jeanna; Payne, Kimberly J

    2016-04-01

    Thymic stromal lymphopoietin (TSLP) stimulates in-vitro proliferation of human fetal B-cell precursors. However, its in-vivo role during normal human B lymphopoiesis is unknown. Genetic alterations that cause overexpression of its receptor component, cytokine receptor-like factor 2 (CRLF2), lead to high-risk B-cell acute lymphoblastic leukemia implicating this signaling pathway in leukemogenesis. We show that mouse thymic stromal lymphopoietin does not stimulate the downstream pathways (JAK/STAT5 and PI3K/AKT/mTOR) activated by the human cytokine in primary high-risk leukemia with overexpression of the receptor component. Thus, the utility of classic patient-derived xenografts for in-vivo studies of this pathway is limited. We engineered xenograft mice to produce human thymic stromal lymphopoietin (+T mice) by injection with stromal cells transduced to express the cytokine. Control (-T) mice were produced using stroma transduced with control vector. Normal levels of human thymic stromal lymphopoietin were achieved in sera of +T mice, but were undetectable in -T mice. Patient-derived xenografts generated from +T as compared to -T mice showed a 3-6-fold increase in normal human B-cell precursors that was maintained through later stages of B-cell development. Gene expression profiles in high-risk B-cell acute lymphoblastic leukemia expanded in +T mice indicate increased mTOR pathway activation and are more similar to the original patient sample than those from -T mice. +T/-T xenografts provide a novel pre-clinical model for understanding this pathway in B lymphopoiesis and identifying treatments for high-risk B-cell acute lymphoblastic leukemia with overexpression of cytokine-like factor receptor 2. PMID:26611474

  14. Gamma-amino butyric acid inhibits the nicotine-imposed stimulatory challenge in xenograft models of non-small cell lung carcinoma.

    PubMed

    Al-Wadei, H A N; Al-Wadei, M H; Ullah, M F; Schuller, H M

    2012-02-01

    Non-small cell lung carcinoma (NSCLC) is the leading type of lung cancer; smoking is a documented risk factor. Nicotinic acetylcholine receptor (nAChR)-mediated intracellular signaling in response to nicotine has recently been implicated in the growth regulation of NSCLC. In the current study nude mice carrying xenografts of the human lung NSCLC cell lines NCI-H322 or NCI-H441 were used as animal models. Nicotine administration and gamma aminobutyric acid (GABA) treatment lasted for 30 days. Catecholamines, cortisol, GABA, and cAMP were analyzed in blood and tumor tissues by immunoassays. Expression of nicotinic receptors and effector proteins in the xenografts was assessed by Western blotting. Our data indicate that nicotine stimulated the growth of NSCLC xenografts via modulation of nAChR upregulation and activation of cAMP signaling. The nicotine-treated group showed an enhanced level of stress neurotransmitters and second messenger cAMP in serum, blood cellular fraction, and xenograft tissues. Activation of critical proteins in the oncogenic pathway, including CREB, ERK, Akt, and Src, and upregulation of α-4 and α-7 subunits of nAChR provided mechanistic insight for the observed stimulatory effect of nicotine. Interestingly, GABA, being an antagonist to cAMP signaling, showed a promising intervention by reversing the stimulatory effect of nicotine on cancer growth and all signaling pathways. GABA has potential to lower the risk of NSCLC among smokers and could be used to enhance the clinical outcome of standard cancer intervention strategies.

  15. A novel xenograft model to study the role of TSLP-induced CRLF2 signals in normal and malignant human B lymphopoiesis

    PubMed Central

    Francis, Olivia L.; Milford, Terry-Ann M.; Martinez, Shannalee R.; Baez, Ineavely; Coats, Jacqueline S.; Mayagoitia, Karina; Concepcion, Katherine R.; Ginelli, Elizabeth; Beldiman, Cornelia; Benitez, Abigail; Weldon, Abby J.; Arogyaswamy, Keshav; Shiraz, Parveen; Fisher, Ross; Morris, Christopher L.; Zhang, Xiao-Bing; Filippov, Valeri; Van Handel, Ben; Ge, Zheng; Song, Chunhua; Dovat, Sinisa; Su, Ruijun Jeanna; Payne, Kimberly J.

    2016-01-01

    Thymic stromal lymphopoietin (TSLP) stimulates in vitro proliferation of human fetal B-cell precursors. However, its in vivo role during normal human B lymphopoiesis is unknown. Genetic alterations that cause overexpression of its receptor component, cytokine receptor-like factor 2 (CRLF2), lead to high-risk B-cell acute lymphoblastic leukemia implicating this signaling pathway in leukemogenesis. We show that mouse thymic stromal lymphopoietin does not stimulate the downstream pathways (JAK/STAT5 and PI3K/AKT/mTOR) activated by the human cytokine in primary high-risk leukemia with overexpression of the receptor component. Thus, the utility of classic patient-derived xenografts for in vivo studies of this pathway is limited. We engineered xenograft mice to produce human thymic stromal lymphopoietin (+T mice) by injection with stromal cells transduced to express the cytokine. Control (−T) mice were produced using stroma transduced with control vector. Normal levels of human thymic stromal lymphopoietin were achieved in sera of +T mice, but were undetectable in −T mice. Patient-derived xenografts generated from +T as compared to −T mice showed a 3–6-fold increase in normal human B-cell precursors that was maintained through later stages of B-cell development. Gene expression profiles in high-risk B-cell acute lymphoblastic leukemia expanded in +T mice indicate increased mTOR pathway activation and are more similar to the original patient sample than those from −T mice. +T/−T xenografts provide a novel pre-clinical model for understanding this pathway in B lymphopoiesis and identifying treatments for high-risk B-cell acute lymphoblastic leukemia with overexpression of cytokine-like factor receptor 2. PMID:26611474

  16. Rapamycin targeting mTOR and hedgehog signaling pathways blocks human rhabdomyosarcoma growth in xenograft murine model

    SciTech Connect

    Kaylani, Samer Z.; Xu, Jianmin; Srivastava, Ritesh K.; Kopelovich, Levy; Pressey, Joseph G.; Athar, Mohammad

    2013-06-14

    Graphical abstract: Intervention of poorly differentiated RMS by rapamycin: In poorly differentiated RMS, rapamycin blocks mTOR and Hh signaling pathways concomitantly. This leads to dampening in cell cycle regulation and induction of apoptosis. This study provides a rationale for the therapeutic intervention of poorly differentiated RMS by treating patients with rapamycin alone or in combination with other chemotherapeutic agents. -- Highlights: •Rapamycin abrogates RMS tumor growth by modulating proliferation and apoptosis. •Co-targeting mTOR/Hh pathways underlie the molecular basis of effectiveness. •Reduction in mTOR/Hh pathways diminish EMT leading to reduced invasiveness. -- Abstract: Rhabdomyosarcomas (RMS) represent the most common childhood soft-tissue sarcoma. Over the past few decades outcomes for low and intermediate risk RMS patients have slowly improved while patients with metastatic or relapsed RMS still face a grim prognosis. New chemotherapeutic agents or combinations of chemotherapies have largely failed to improve the outcome. Based on the identification of novel molecular targets, potential therapeutic approaches in RMS may offer a decreased reliance on conventional chemotherapy. Thus, identification of effective therapeutic agents that specifically target relevant pathways may be particularly beneficial for patients with metastatic and refractory RMS. The PI3K/AKT/mTOR pathway has been found to be a potentially attractive target in RMS therapy. In this study, we provide evidence that rapamycin (sirolimus) abrogates growth of RMS development in a RMS xenograft mouse model. As compared to a vehicle-treated control group, more than 95% inhibition in tumor growth was observed in mice receiving parenteral administration of rapamycin. The residual tumors in rapamycin-treated group showed significant reduction in the expression of biomarkers indicative of proliferation and tumor invasiveness. These tumors also showed enhanced apoptosis

  17. Preventing T cell rejection of pig xenografts.

    PubMed

    Higginbotham, Laura; Ford, Mandy L; Newell, Kenneth A; Adams, Andrew B

    2015-11-01

    Xenotransplantation is a potential solution to the limited supply of donor organs. While early barriers to xenograft acceptance, such as hyperacute rejection, are now largely avoided through genetic engineering, the next frontier in successful xenograft survival will require prevention of T cell-mediated rejection. Most successful immunosuppressive regimens in xenotransplantation utilize T cell depletion with antibody therapy. Additionally, the use of T cell costimulatory blockade - specifically blockade of the CD40-CD154 pathway - shows promise with several reports of long-term xenograft survival. Additional therapies, such as transgenic expression of T cell coinhibitory molecules or transfer of immunomodulatory cells to promote tolerance, may be necessary to achieve reliable long-term xenograft acceptance. Further studies in pre-clinical models are essential in order to optimize these regimens prior to trials in patients.

  18. Comparative efficacy of 177Lu and 90Y for Anti-CD20 Pretargeted Radioimmunotherapy in Murine Lymphoma Xenograft Models

    SciTech Connect

    Frost, Sofia H. L.; Frayo, Shani L.; Miller, Brian W.; Orozco, Johnnie J.; Booth, Garrett C.; Hylarides, Mark D.; Lin, Yukang; Green, Damian J.; Gopal, Ajay K.; Pagel, John M.; Bäck, Tom A.; Fisher, Darrell R.; Press, Oliver W.; Afrin, Farhat

    2015-03-18

    % cured with 177Lu-PRIT. Toxicities were comparable with both isotopes. Conclusion 90Y was therapeutically superior to 177Lu for streptavidin-biotin PRIT approaches in these human lymphoma xenograft models.

  19. Investigational drug MLN0128, a novel TORC1/2 inhibitor, demonstrates potent oral antitumor activity in human breast cancer xenograft models.

    PubMed

    Gökmen-Polar, Yesim; Liu, Yi; Toroni, Rachel A; Sanders, Kerry L; Mehta, Rutika; Badve, Sunil; Rommel, Christian; Sledge, George W

    2012-12-01

    Aberrant activation of the mammalian target of rapamycin (mTOR) signaling plays an important role in breast cancer progression and represents a potential therapeutic target for breast cancer. In this study, we report the impact of the investigational drug MLN0128, a potent and selective small molecule active-site TORC1/2 kinase inhibitor, on tumor growth and metastasis using human breast cancer xenograft models. We assessed in vitro antiproliferative activity of MLN0128 in a panel of breast cancer cell lines. We next evaluated the impact of MLN0128 on tumor growth, angiogenesis and metastasis using mammary fat pad xenograft models of a non-VEGF (ML20) and a VEGF-driven (MV165) MCF-7 sublines harboring PIK3CA mutations. MLN0128 potently inhibited cell proliferation in various breast cancer cell lines harboring PIK3CA (IC(50): 1.5-53 nM), PTEN (IC(50): 1-149 nM), KRAS, and/or BRAF mutations (IC(50): 13-162 nM), and in human endothelial cells (IC(50): 33-40 nM) in vitro. In vivo, MLN0128 decreased primary tumor growth significantly in both non-VEGF (ML20; p = 0.05) and VEGF-driven MCF-7 (MV165; p = 0.014) xenograft models. MLN0128 decreased the phosphorylation of Akt, S6, 4E-BP1, and NDRG1 in both models. In contrast, rapamycin increased Akt activity and failed to reduce the phosphorylation of 4E-BP1, PRAS40, and NDRG1. VEGF-induced lung metastasis in MV165 is inhibited by MLN0128 and rapamycin. In conclusion, MLN0128 inhibits TORC1/2-dependent signaling in preclinical models of breast cancer. MLN0128 appears to be superior in blocking mTORC1/2 signaling in contrast to rapamycin. Our findings support the clinical research of MLN0128 in patients with breast cancer and metastasis.

  20. Antitumor effect of Deoxypodophyllotoxin on human breast cancer xenograft transplanted in BALB/c nude mice model.

    PubMed

    Khaled, Meyada; Belaaloui, Ghania; Jiang, Zhen-Zhou; Zhu, Xiong; Zhang, Lu-Yong

    2016-10-01

    Recently, biologically active compounds isolated from plants used in herbal medicine have been the center of interest. Deoxypodophyllotoxin (DPT), structurally closely related to the lignan podophyllotoxin, was found to be a potent antitumor and antiproliferative agent, in several tumor cells, in vitro. However, DPT has not been used clinically yet because of the lack of in vivo studies. This study is the first report demonstrating the antitumor effect of DPT on MDA-MB-231 human breast cancer xenografts in nude mice. DPT, significantly, inhibited the growth of MDA-MB-231 xenograft in BALB/c nude mice. The T/C value (the value of the relative tumor volume of treatment group compared to the control group) of groups treated with 5, 10, and 20 mg/kg of intravenous DPT-HP-β-CD was 42.87%, 34.04% and 9.63%, respectively, suggesting the positive antitumor activity of DPT. In addition, the antitumor effect of DPT-HP-β-CD (20 mg/kg) in human breast cancer MDA-MB-231 xenograft was more effective than etoposide (VP-16) (20 mg/kg) and docetaxel (20 mg/kg). These findings suggest that this drug is a promising chemotherapy candidate against human breast carcinoma. PMID:27578026

  1. Evaluation of Trastuzumab Anti-Tumor Efficacy and its Correlation with HER-2 Status in Patient-Derived Gastric Adenocarcinoma Xenograft Models.

    PubMed

    Chen, Hao; Ye, Qingqing; Lv, Jing; Ye, Peng; Sun, Yun; Fan, Shuqiong; Su, Xinying; Gavine, Paul; Yin, Xiaolu

    2015-09-01

    The aim of the study was to investigate trastuzumab anti-tumor efficacy and its correlation with HER-2 status in primary xenograft models derived from Chinese patients with gastric adenocarcinoma. Patient-derived gastric adenocarcinoma xenograft (PDGAX) mouse models were firstly generated by implanting gastric adenocarcinoma tissues from patients into immune deficient mice. A high degree of histological and molecular similarity between the PDGAX mouse models and their corresponding patients' gastric adenocarcinoma tissues was shown by pathological observation, HER-2 expression, HER-2 gene copy number, and mutation detection. Based on Hoffmann's criteria in gastric cancer, three models (PDGAX001, PDGAX003 and PDGAX005) were defined as HER-2 positive with fluorescence in situ hybridization (FISH) amplification or immunohistochemistry (IHC) 2+/ 3+, while two models (PDGAX002, PDGAX004) were defined as HER-2 negative. Upon trastuzumab treatment, significant tumor regression (105 % TGI) was observed in model PDGAX005 (TP53 wt), while moderate sensitivity (26 % TGI) was observed in PDGAX003, and resistance was observed in PDGAX001, 002 and 004. A significant increase in HER-2 gene copy number was only observed in PDGAX005 (TP53 wt). Interestingly, trastuzumab showed no efficacy in PDGAX001 (HER2 IHC 3+ and FISH amplification, but with mutant TP53). Consistent with this finding, phosphor-HER2 modulation by trastuzumab was observed in model PDGAX005, but not in PDGAX001.

  2. Preservation of KIT genotype in a novel pair of patient-derived orthotopic xenograft mouse models of metastatic pediatric CNS germinoma.

    PubMed

    Lindsay, Holly; Huang, Yulun; Du, Yuchen; Braun, Frank K; Teo, Wan Yee; Kogiso, Mari; Qi, Lin; Zhang, Huiyuan; Zhao, Sibo; Mao, Hua; Lin, Frank; Baxter, Patricia; Su, Jack M; Terashima, Keita; Perlaky, Laszlo; Chintagumpala, Murali; Adesina, Adekunle; Lau, Ching C; Williams Parsons, D; Li, Xiao-Nan

    2016-05-01

    Metastatic intracranial germinoma is difficult to treat. Although the proto-oncogene KIT is recognized as one of the most frequent genetic abnormalities in CNS germinoma, the development of new target therapeutic agents for CNS germinoma is hampered by the lack of clinically-relevant animal models that replicate the mutated or over-expressed KIT. CNS germinoma tumor cells from five pediatric patients were directly implanted into the brains of Rag2/severe combined immune deficiency mice. Once established, the xenograft tumors were sub-transplanted in vivo in mouse brains. Characterization of xenograft tumors were performed through histologic and immunohistochemical staining, and KIT mutation analysed with quantitative pyro-sequencing. Expression of putative cancer stem cell markers (CD133, CD15, CD24, CD44, CD49f) was analyzed through flow cytometry. Two patient-derived orthotopic xenograft (PDOX) models (IC-6999GCT and IC-9302GCT) were established from metastatic germinoma and serially sub-transplanted five times in mouse brains. Similar to the original patient tumors, they both exhibited faint expression (+) of PLAP, no expression (-) of β-HCG and strong (+++) expression of KIT. KIT mutation (D816H), however, was only found in IC-9320GCT. This mutation was maintained during the five in vivo tumor passages with an increased mutant allele frequency compared to the patient tumor. Expression of putative cancer stem cell markers CD49f and CD15 was also detected in a small population of tumor cells in both models. This new pair of PDOX models replicated the key biological features of pediatric intracranial germinoma and should facilitate the biological and pre-clinical studies for metastatic intracranial germinomas.

  3. Establishment of a patient-derived orthotopic Xenograft (PDOX) model of HER-2-positive cervical cancer expressing the clinical metastatic pattern.

    PubMed

    Hiroshima, Yukihiko; Zhang, Yong; Zhang, Nan; Maawy, Ali; Mii, Sumiyuki; Yamamoto, Mako; Uehara, Fuminari; Miwa, Shinji; Yano, Shuya; Murakami, Takashi; Momiyama, Masashi; Chishima, Takashi; Tanaka, Kuniya; Ichikawa, Yasushi; Bouvet, Michael; Murata, Takuya; Endo, Itaru; Hoffman, Robert M

    2015-01-01

    Squamous cell carcinoma of the cervix, highly prevalent in the developing world, is often metastatic and treatment resistant with no standard treatment protocol. Our laboratory pioneered the patient-derived orthotopic xenograft (PDOX) nude mouse model with the technique of surgical orthotopic implantation (SOI). Unlike subcutaneous transplant patient-derived xenograft (PDX) models, PDOX models metastasize. Most importantly, the metastasis pattern correlates to the patient. In the present report, we describe the development of a PDOX model of HER-2-positive cervical cancer. Metastasis after SOI in nude mice included peritoneal dissemination, liver metastasis, lung metastasis as well as lymph node metastasis reflecting the metastatic pattern in the donor patient. Metastasis was detected in 4 of 6 nude mice with primary tumors. Primary tumors and metastases in the nude mice had histological structures similar to the original tumor and were stained by an anti-HER-2 antibody in the same pattern as the patient's cancer. The metastatic pattern, histology and HER-2 tumor expression of the patient were thus preserved in the PDOX model. In contrast, subcutaneous transplantation of the patient's cervical tumors resulted in primary growth but not metastasis.

  4. Efficacy of Tumor-Targeting Salmonella A1-R on a Melanoma Patient-Derived Orthotopic Xenograft (PDOX) Nude-Mouse Model

    PubMed Central

    Yamamoto, Mako; Zhao, Ming; Hiroshima, Yukihiko; Zhang, Yong; Shurell, Elizabeth; Eilber, Fritz C.; Bouvet, Michael; Noda, Makoto; Hoffman, Robert M.

    2016-01-01

    Tumor-targeting Salmonella enterica serovar Typhimurium A1-R (Salmonella A1-R) had strong efficacy on a melanoma patient-derived orthotopic xenograft (PDOX) nude-mouse model. GFP-expressing Salmonella A1-R highly and selectively colonized the PDOX melanoma and significantly suppressed tumor growth (p = 0.021). The combination of Salmonella A1-R and cisplatinum (CDDP), both at low-dose, also significantly suppressed the growth of the melanoma PDOX (P = 0.001). Salmonella A1-R has future clinical potential for combination chemotherapy with CDDP of melanoma, a highly-recalcitrant cancer. PMID:27500926

  5. Castration induces up-regulation of intratumoral androgen biosynthesis and androgen receptor expression in an orthotopic VCaP human prostate cancer xenograft model.

    PubMed

    Knuuttila, Matias; Yatkin, Emrah; Kallio, Jenny; Savolainen, Saija; Laajala, Teemu D; Aittokallio, Tero; Oksala, Riikka; Häkkinen, Merja; Keski-Rahkonen, Pekka; Auriola, Seppo; Poutanen, Matti; Mäkelä, Sari

    2014-08-01

    Androgens are key factors involved in the development and progression of prostate cancer (PCa), and PCa growth can be suppressed by androgen deprivation therapy. In a considerable proportion of men receiving androgen deprivation therapy, however, PCa progresses to castration-resistant PCa (CRPC), making the development of efficient therapies challenging. We used an orthotopic VCaP human PCa xenograft model to study cellular and molecular changes in tumors after androgen deprivation therapy (castration). Tumor growth was monitored through weekly serum prostate-specific antigen measurements, and mice with recurrent tumors after castration were randomized to treatment groups. Serum prostate-specific antigen concentrations showed significant correlation with tumor volume. Castration-resistant tumors retained concentrations of intratumoral androgen (androstenedione, testosterone, and 5α-dihydrotestosterone) at levels similar to tumors growing in intact hosts. Accordingly, castration induced up-regulation of enzymes involved in androgen synthesis (CYP17A1, AKR1C3, and HSD17B6), as well as expression of full-length androgen receptor (AR) and AR splice variants (AR-V1 and AR-V7). Furthermore, AR target gene expression was maintained in castration-resistant xenografts. The AR antagonists enzalutamide (MDV3100) and ARN-509 suppressed PSA production of castration-resistant tumors, confirming the androgen dependency of these tumors. Taken together, the findings demonstrate that our VCaP xenograft model exhibits the key characteristics of clinical CRPC and thus provides a valuable tool for identifying druggable targets and for testing therapeutic strategies targeting AR signaling in CRPC.

  6. Gene expression profiling upon (212) Pb-TCMC-trastuzumab treatment in the LS-174T i.p. xenograft model.

    PubMed

    Yong, Kwon J; Milenic, Diane E; Baidoo, Kwamena E; Kim, Young-Seung; Brechbiel, Martin W

    2013-10-01

    Recent studies have demonstrated that therapy with (212) Pb-TCMC-trastuzumab resulted in (1) induction of apoptosis, (2) G2/M arrest, and (3) blockage of double-strand DNA damage repair in LS-174T i.p. (intraperitoneal) xenografts. To further understand the molecular basis of the cell killing efficacy of (212) Pb-TCMC-trastuzumab, gene expression profiling was performed with LS-174T xenografts 24 h after exposure to (212) Pb-TCMC-trastuzumab. DNA damage response genes (84) were screened using a quantitative real-time polymerase chain reaction array (qRT-PCR array). Differentially regulated genes were identified following exposure to (212) Pb-TCMC-trastuzumab. These included genes involved in apoptosis (ABL, GADD45α, GADD45γ, PCBP4, and p73), cell cycle (ATM, DDIT3, GADD45α, GTSE1, MKK6, PCBP4, and SESN1), and damaged DNA binding (DDB) and repair (ATM and BTG2). The stressful growth arrest conditions provoked by (212) Pb-TCMC-trastuzumab were found to induce genes involved in apoptosis and cell cycle arrest in the G2/M phase. The expression of genes involved in DDB and single-strand DNA breaks was also enhanced by (212) Pb-TCMC-trastuzumab while no modulation of genes involved in double-strand break repair was apparent. Furthermore, the p73/GADD45 signaling pathway mediated by p38 kinase signaling may be involved in the cellular response, as evidenced by the enhanced expression of genes and proteins of this pathway. These results further support the previously described cell killing mechanism by (212) Pb-TCMC-trastuzumab in the same LS-174T i.p. xenograft. Insight into these mechanisms could lead to improved strategies for rational application of radioimmunotherapy using α-particle emitters.

  7. Exosomes: decreased sensitivity of lung cancer A549 cells to cisplatin.

    PubMed

    Xiao, Xia; Yu, Shaorong; Li, Shuchun; Wu, Jianzhong; Ma, Rong; Cao, Haixia; Zhu, Yanliang; Feng, Jifeng

    2014-01-01

    Exosomes are small extracellular membrane vesicles of endocytic origin released by many cells that could be found in most body fluids. The main functions of exosomes are cellular communication and cellular waste clean-up. This study was conducted to determine the involvement of exosomes in the regulation of sensitivity of the lung cancer cell line A549 to cisplatin (DDP). When DDP was added to A549 cells, exosomes secretion was strengthened. Addition of the secreted exosomes to other A549 cells increased the resistance of these A549 cells to DDP. Upon exposure of A549 to DDP, the expression levels of several miRNAs and mRNAs reportedly associated with DDP sensitivity changed significantly in exosomes; these changes may mediate the resistance of A549 cells to DDP. Exosomes released by A549 cells during DDP exposure decreased the sensitivity of other A549 cells to DDP, which may be mediated by miRNAs and mRNAs exchange by exosomes via cell-to-cell communication. Although the detailed mechanism of resistance remains unclear, we believed that inhibition of exosomes formation and release might present a novel strategy for lung cancer treatment in the future. PMID:24586853

  8. XactMice: humanizing mouse bone marrow enables microenvironment reconstitution in a patient-derived xenograft model of head and neck cancer.

    PubMed

    Morton, J J; Bird, G; Keysar, S B; Astling, D P; Lyons, T R; Anderson, R T; Glogowska, M J; Estes, P; Eagles, J R; Le, P N; Gan, G; McGettigan, B; Fernandez, P; Padilla-Just, N; Varella-Garcia, M; Song, J I; Bowles, D W; Schedin, P; Tan, A-C; Roop, D R; Wang, X-J; Refaeli, Y; Jimeno, A

    2016-01-21

    The limitations of cancer cell lines have led to the development of direct patient-derived xenograft models. However, the interplay between the implanted human cancer cells and recruited mouse stromal and immune cells alters the tumor microenvironment and limits the value of these models. To overcome these constraints, we have developed a technique to expand human hematopoietic stem and progenitor cells (HSPCs) and use them to reconstitute the radiation-depleted bone marrow of a NOD/SCID/IL2rg(-/-) (NSG) mouse on which a patient's tumor is then transplanted (XactMice). The human HSPCs produce immune cells that home into the tumor and help replicate its natural microenvironment. Despite previous passage on nude mice, the expression of epithelial, stromal and immune genes in XactMice tumors aligns more closely to that of the patient tumor than to those grown in non-humanized mice-an effect partially facilitated by human cytokines expressed by both the HSPC progeny and the tumor cells. The human immune and stromal cells produced in the XactMice can help recapitulate the microenvironment of an implanted xenograft, reverse the initial genetic drift seen after passage on non-humanized mice and provide a more accurate tumor model to guide patient treatment.

  9. Downregulation of peroxiredoxin-1 by β-elemene enhances the radiosensitivity of lung adenocarcinoma xenografts.

    PubMed

    Li, Guoquan; Xie, Bingbing; Li, Xiaolong; Chen, Yinghai; Xu, Yinghui; Xu-Welliver, Meng; Zou, Lijuan

    2015-03-01

    β-elemene, the active component of elemene (1-methyl-1-vinyl-2,4-diisopropenyl-cyclohexane), is a naturally occurring compound isolated from the traditional Chinese medicinal herb Curcuma wenyujin. Studies have confirmed that β-elemene enhances the radiosensitivity of lung cancer cell lines such as A549, by multiple pathways; however, their underlying mechanisms and pathways are yet to be elucidated. In the present study, two-dimensional differential in-gel electrophoresis and matrix-assisted laser desorption/ionization time-of-flight tandem mass spectrometry were used to profile the different proteins in A549 cell xenograft models of both treatment groups. The protein/mRNA expression was assessed by reverse transcription-polymerase chain reaction and western blotting techniques in tumor samples from all treatment groups. As a critical player in redox regulation of cancer cells, inhibition of peroxiredoxin-1 (Prx-1) may be an effective option for enhancing the tumor response to radiation. We further verified Prx-1 expression at the transcription and translation levels. β-elemene at a dose of 45 mg/kg had little effect on the Prx-1 protein expression, which was correlated with a moderate antitumor effect. However, a 45 mg/kg dose of β-elemene significantly inhibited the Prx-1 mRNA expression, thereby suggesting a possible influence on the transcriptional process, and radiation significantly increased the Prx-1 mRNA/protein expression compared to the control group (p<0.01). Notably, Prx-1 mRNA/protein expression was significantly lower in the β-elemene/radiation co-treatment group compared to the baseline levels in the control group (p<0.01). These results suggest that radiation-induced Prx-1 expression is directly or indirectly suppressed by β-elemene, thus suggesting a new pathway by which to reverse radioresistance.

  10. Vorinostat, an HDAC inhibitor attenuates epidermoid squamous cell carcinoma growth by dampening mTOR signaling pathway in a human xenograft murine model

    SciTech Connect

    Kurundkar, Deepali; Srivastava, Ritesh K.; Chaudhary, Sandeep C.; Ballestas, Mary E.; Kopelovich, Levy; Elmets, Craig A.; Athar, Mohammad

    2013-01-15

    Histone deacetylase (HDAC) inhibitors are potent anticancer agents and show efficacy against various human neoplasms. Vorinostat is a potent HDAC inhibitor and has shown potential to inhibit growth of human xenograft tumors. However, its effect on the growth of skin neoplasm remains undefined. In this study, we show that vorinostat (2 μM) reduced expression of HDAC1, 2, 3, and 7 in epidermoid carcinoma A431 cells. Consistently, it increased acetylation of histone H3 and p53. Vorinostat (100 mg/kg body weight, IP) treatment reduced human xenograft tumor growth in highly immunosuppressed nu/nu mice. Histologically, the vorinostat-treated tumor showed features of well-differentiation with large necrotic areas. Based on proliferating cell nuclear antigen (PCNA) staining and expression of cyclins D1, D2, E, and A, vorinostat seems to impair proliferation by down-regulating the expression of these proteins. However, it also induced apoptosis. The mechanism by which vorinostat blocks proliferation and makes tumor cells prone to apoptosis, involved inhibition of mTOR signaling which was accompanied by reduction in cell survival AKT and extracellular-signal regulated kinase (ERK) signaling pathways. Our data provide a novel mechanism-based therapeutic intervention for cutaneous squamous cell carcinoma (SCC). Vorinostat may be utilized to cure skin neoplasms in organ transplant recipient (OTR). These patients have high morbidity and surgical removal of these lesions which frequently develop in these patients, is difficult. -- Highlights: ► Vorinostat reduces SCC growth in a xenograft murine model. ► Vorinostat dampens proliferation and induces apoptosis in tumor cells. ► Diminution in mTOR, Akt and ERK signaling underlies inhibition in proliferation. ► Vorinostat by inhibiting HDACs inhibits epithelial–mesenchymal transition.

  11. Novel Effects of Simvastatin on Uterine Fibroids: In vitro and Patient-Derived Xenograft Mouse Model Study

    PubMed Central

    BORAHAY, Mostafa A.; VINCENT, Kathleen; MOTAMEDI, Massoud; SBRANA, Elena; KILIC, Gokhan S.; AL-HENDY, Ayman; BOEHNING, Darren

    2015-01-01

    Objective Uterine leiomyomas represent a common gynecologic problem with no satisfactory long-term medical treatment. The purpose of this study is to examine the effects of simvastatin on uterine leiomyoma, both in vitro and in vivo. Study Design This is a laboratory-based experimental study. For in vitro studies, we used human and rat leiomyoma cells. For in vivo studies, we used immunodeficient mice supplemented with estrogen/progesterone pellets xenografted with human leiomyoma tissue explant. Results For in vitro studies, cells were treated with different concentrations of simvastatin for 48 hours. Simvastatin induced dose-dependent apoptosis in leiomyoma cells as measured by a fluorometric caspase-3 activity assay, and inhibited proliferation as demonstrated by an MTT assay (both were significant at 5 and 10 μM). In addition, simvastatin decreased Akt signaling pathway phosphorylation as examined using Western blot analysis. For in vivo studies, animals were treated for 28 days with simvastatin (20 μg/ gm body weight/ day) vs vehicle control. The treatment inhibited tumor growth as measured weekly using calipers and/ or ultrasound (P<.01). Finally, simvastatin decreased expression of the proliferation marker Ki67 in xenograft tumor tissue as examined by immunohistochemistry (P=.02). Conclusion Simvastatin can be a promising treatment for uterine leiomyoma. Further studies, including pharmacokinetic and drug delivery studies, are required. PMID:25840272

  12. XH1--a new cervical carcinoma cell line and xenograft model of tumour invasion, 'metastasis' and regression.

    PubMed Central

    Han, X.; Lyle, R.; Eustace, D. L.; Jewers, R. J.; Parrington, J. M.; Das, A.; Chana, T.; Dagg, B.; Money, S.; Bates, T. D.

    1991-01-01

    A new cell line, XH1, has been derived from an invasive focally keratinising adenosquamous carcinoma of the cervix in a 32 year old patient. It has been maintained in long term monolayer culture for 26 months, and passaged over 100 times (much greater than 300 population doublings). It is aneuploid with a mean chromosome number of 78. Examination using two minisatellite hypervariable DNA probes has shown it to be different from other cell lines maintained in this laboratory and from HeLa. Two sublines, XH1a and XH1b, show marked differences in monolayer culture, growth in soft agar, and xenograft formation. XH1 and XH1a cells readily form subcutaneous xenografts, and lung colonies can be established by their intravenous injection. Subcutaneous injection of XH1b cells results in rapid cell growth for a few days after which the tumour undergoes degeneration and then regresses completely. The XH1 karyotype has many rearranged chromosomes. Parental XH1 cells and both sublines show integration of HPV16 into the genome. Images Figure 1 Figure 2 Figure 4 Figure 5 Figure 6 Figure 9 Figure 7 Figure 8 Figure 10 Figure 13 Figure 14 Figure 11 Figure 12 Figure 15 Figure 16 Figure 17 Figure 18 PMID:1911212

  13. A HLA-A2-restricted CTL epitope induces anti-tumor effects against human lung cancer in mouse xenograft model

    PubMed Central

    Chen, I-Hua; Liu, Hsin-Yu; Lin, Chen-Yuan; Chiang, I-Ping; Roffler, Steve; Chen, Hsin-Wei; Liu, Shih-Jen

    2016-01-01

    Cancer immunotherapy is attractive for antigen-specific T cell-mediated anti-tumor therapy, especially in induction of cytotoxic T lymphocytes. In this report, we evaluated human CTL epitope-induced anti-tumor effects in human lung cancer xenograft models. The tumor associated antigen L6 (TAL6) is highly expressed in human lung cancer cell lines and tumor specimens as compared to normal lung tissues. TAL6 derived peptides strongly inhibited tumor growth, cancer metastasis and prolonged survival time in HLA-A2 transgenic mice immunized with a formulation of T-helper (Th) peptide, synthetic CpG ODN, and adjuvant Montanide ISA-51 (ISA-51). Adoptive transfer of peptide-induced CTL cells from HLA-A2 transgenic mice into human tumor xenograft SCID mice significantly inhibited tumor growth. Furthermore, combination of CTL-peptide immunotherapy and gemcitabine additively improved the therapeutic effects. This pre-clinical evaluation model provides a useful platform to develop efficient immunotherapeutic drugs to treat lung cancer and demonstrates a promising strategy with benefit of antitumor immune responses worthy of further development in clinical trials. PMID:26621839

  14. Bufalin Inhibits HCT116 Colon Cancer Cells and Its Orthotopic Xenograft Tumor in Mice Model through Genes Related to Apoptotic and PTEN/AKT Pathways

    PubMed Central

    Wang, Jie; Chen, Chao; Wang, Shiying; Zhang, Yong; Yin, Peihao; Gao, Zhongxiang; Xu, Jie; Feng, Dianxu; Zuo, Qinsong; Zhao, Ronghua; Chen, Teng

    2015-01-01

    Aims. To investigate the anticolorectal cancer (CRC) effects of Bufalin, a bioactive polyhydroxysteroid from Venenum Bufonis, using HCT116 human CRC cell and an established orthotopic xenograft model in mice, and to explore the mechanisms of action. Material and Methods. Cultured HCT116 cells or BALB/c mice with orthotopic tumor were treated by Bufalin (positive control: 5-FU). Cell proliferation, apoptosis, and cycling were determined by MTT, Annexin V/PI staining, and flow cytometry, respectively. In mice, tumor inhibition rate and animal survival were calculated. The expressions of PTEN/phosphate-PTEN, AKT/phosphate-AKT, Bad, Bcl-xl, Bax, or Caspase-3 in cells and/or tumors were determined by Western blot or immunohistochemical staining. Results. Bufalin significantly inhibited cell proliferation and induced cell apoptosis and cycle arrest in a dose/time-dependent manner. In the animal model, Bufalin treatment resulted in significant inhibition of tumor growth and prolonged survival. In the Bufalin-treated cultured cells and/or xenograft tumors, the expressions of PTEN, Bad, Bax, and Caspase-3 were significantly increased, while p-AKT and Bcl-xL significantly decreased. Conclusions. Our results indicate that Bufalin inhibit cell proliferation and orthotopic tumor growth by inducing cell apoptosis through the intrinsic apoptotic pathway, which is of pivotal significance in the identification of an anticancer drug that may synergize with Bufalin. PMID:26770191

  15. A Novel Eg5 Inhibitor (LY2523355) Causes Mitotic Arrest and Apoptosis in Cancer Cells and Shows Potent Antitumor Activity in Xenograft Tumor Models.

    PubMed

    Ye, Xiang S; Fan, Li; Van Horn, Robert D; Nakai, Ryuichiro; Ohta, Yoshihisa; Akinaga, Shiro; Murakata, Chikara; Yamashita, Yoshinori; Yin, Tinggui; Credille, Kelly M; Donoho, Gregory P; Merzoug, Farhana F; Li, Heng; Aggarwal, Amit; Blanchard, Kerry; Westin, Eric H

    2015-11-01

    Intervention of cancer cell mitosis by antitubulin drugs is among the most effective cancer chemotherapies. However, antitubulin drugs have dose-limiting side effects due to important functions of microtubules in resting normal cells and are often rendered ineffective by rapid emergence of resistance. Antimitotic agents with different mechanisms of action and improved safety profiles are needed as new treatment options. Mitosis-specific kinesin Eg5 represents an attractive anticancer target for discovering such new antimitotic agents, because Eg5 is essential only in mitotic progression and has no roles in resting, nondividing cells. Here, we show that a novel selective Eg5 inhibitor, LY2523355, has broad target-mediated anticancer activity in vitro and in vivo. LY2523355 arrests cancer cells at mitosis and causes rapid cell death that requires sustained spindle-assembly checkpoint (SAC) activation with a required threshold concentration. In vivo efficacy of LY2523355 is highly dose/schedule-dependent, achieving complete remission in a number of xenograft tumor models, including patient-derived xenograft (PDX) tumor models. We further establish that histone-H3 phosphorylation of tumor and proliferating skin cells is a promising pharmacodynamic biomarker for in vivo anticancer activity of LY2523355. PMID:26304237

  16. A HLA-A2-restricted CTL epitope induces anti-tumor effects against human lung cancer in mouse xenograft model.

    PubMed

    Sher, Yuh-Pyng; Lin, Su-I; Chen, I-Hua; Liu, Hsin-Yu; Lin, Chen-Yuan; Chiang, I-Ping; Roffler, Steve; Chen, Hsin-Wei; Liu, Shih-Jen

    2016-01-01

    Cancer immunotherapy is attractive for antigen-specific T cell-mediated anti-tumor therapy, especially in induction of cytotoxic T lymphocytes. In this report, we evaluated human CTL epitope-induced anti-tumor effects in human lung cancer xenograft models. The tumor associated antigen L6 (TAL6) is highly expressed in human lung cancer cell lines and tumor specimens as compared to normal lung tissues. TAL6 derived peptides strongly inhibited tumor growth, cancer metastasis and prolonged survival time in HLA-A2 transgenic mice immunized with a formulation of T-helper (Th) peptide, synthetic CpG ODN, and adjuvant Montanide ISA-51 (ISA-51). Adoptive transfer of peptide-induced CTL cells from HLA-A2 transgenic mice into human tumor xenograft SCID mice significantly inhibited tumor growth. Furthermore, combination of CTL-peptide immunotherapy and gemcitabine additively improved the therapeutic effects. This pre-clinical evaluation model provides a useful platform to develop efficient immunotherapeutic drugs to treat lung cancer and demonstrates a promising strategy with benefit of antitumor immune responses worthy of further development in clinical trials. PMID:26621839

  17. Efficacy of JAK/STAT pathway inhibition in murine xenograft models of early T-cell precursor (ETP) acute lymphoblastic leukemia

    PubMed Central

    Maude, Shannon L.; Dolai, Sibasish; Delgado-Martin, Cristina; Vincent, Tiffaney; Robbins, Alissa; Selvanathan, Arthavan; Ryan, Theresa; Hall, Junior; Wood, Andrew C.; Tasian, Sarah K.; Hunger, Stephen P.; Loh, Mignon L.; Mullighan, Charles G.; Wood, Brent L.; Hermiston, Michelle L.; Grupp, Stephan A.; Lock, Richard B.

    2015-01-01

    Early T-cell precursor (ETP) acute lymphoblastic leukemia (ALL) is a recently described subtype of T-ALL characterized by a unique immunophenotype and genomic profile, as well as a high rate of induction failure. Frequent mutations in cytokine receptor and Janus kinase (JAK)/signal transducer and activator of transcription (STAT) signaling pathways led us to hypothesize that ETP-ALL is dependent on JAK/STAT signaling. Here we demonstrate aberrant activation of the JAK/STAT pathway in ETP-ALL blasts relative to non-ETP T-ALL. Moreover, ETP-ALL showed hyperactivation of STAT5 in response to interleukin-7, an effect that was abrogated by the JAK1/2 inhibitor ruxolitinib. In vivo, ruxolitinib displayed activity in 6 of 6 patient-derived murine xenograft models of ETP-ALL, with profound single-agent efficacy in 5 models. Ruxolitinib treatment decreased peripheral blast counts relative to pretreatment levels and compared with control (P < .01) in 5 of 6 ETP-ALL xenografts, with marked reduction in mean splenic blast counts (P < .01) in 6 of 6 samples. Surprisingly, both JAK/STAT pathway activation and ruxolitinib efficacy were independent of the presence of JAK/STAT pathway mutations, raising the possibility that the therapeutic potential of ruxolitinib in ETP-ALL extends beyond those cases with JAK mutations. These findings establish the preclinical in vivo efficacy of ruxolitinib in ETP-ALL, a biologically distinct subtype for which novel therapies are needed. PMID:25645356

  18. Increased COX-2 expression in epithelial and stromal cells of high mammographic density tissues and in a xenograft model of mammographic density.

    PubMed

    Chew, G L; Huo, C W; Huang, D; Hill, P; Cawson, J; Frazer, H; Hopper, J L; Haviv, I; Henderson, M A; Britt, K; Thompson, E W

    2015-08-01

    Mammographic density (MD) adjusted for age and body mass index is one of the strongest known risk factors for breast cancer. Given the high attributable risk of MD for breast cancer, chemoprevention with a safe and available agent that reduces MD and breast cancer risk would be beneficial. Cox-2 has been implicated in MD-related breast cancer risk, and was increased in stromal cells in high MD tissues in one study. Our study assessed differential Cox-2 expression in epithelial and stromal cells in paired samples of high and low MD human breast tissue, and in a validated xenograft biochamber model of MD. We also examined the effects of endocrine treatment upon Cox-2 expression in high and low MD tissues in the MD xenograft model. Paired high and low MD human breast tissue samples were immunostained for Cox-2, then assessed for differential expression and staining intensity in epithelial and stromal cells. High and low MD human breast tissues were separately maintained in biochambers in mice treated with Tamoxifen, oestrogen or placebo implants, then assessed for percentage Cox-2 staining in epithelial and stromal cells. Percentage Cox-2 staining was greater for both epithelial (p = 0.01) and stromal cells (p < 0.0001) of high compared with low MD breast tissues. In high MD biochamber tissues, percentage Cox-2 staining was greater in stromal cells of oestrogen-treated versus placebo-treated tissues (p = 0.05).

  19. 2'-(2-bromohexadecanoyl)-paclitaxel conjugate nanoparticles for the treatment of non-small cell lung cancer in an orthotopic xenograft mouse model.

    PubMed

    Peng, Lei; Schorzman, Allison N; Ma, Ping; Madden, Andrew J; Zamboni, William C; Benhabbour, Soumya Rahima; Mumper, Russell J

    2014-01-01

    A nanoparticle (NP) formulation with 2'-(2-bromohexadecanoyl)-paclitaxel (Br-16-PX) conjugate was developed in these studies for the treatment of non-small cell lung cancer (NSCLC). The lipophilic paclitaxel conjugate Br-C16-PX was synthesized and incorporated into lipid NPs where the 16-carbon chain enhanced drug entrapment in the drug delivery system and improved in vivo pharmacokinetics. The electron-withdrawing bromine group was used to facilitate the conversion of Br-C16-PX to paclitaxel at the tumor site. The developed system was evaluated in luciferase-expressing A549 cells in vitro and in an orthotopic NSCLC mouse model. The results demonstrated that the Br-C16-PX NPs had a higher maximum tolerated dose (75 mg/kg) than Taxol (19 mg/kg) and provided significantly longer median survival (88 days versus 70 days, P<0.05) in the orthotopic NSCLC model. An improved pharmacokinetic profile was observed for the Br-C16-PX NPs at 75 mg/kg compared to Taxol at 19 mg/kg. The area under the concentration versus time curve (AUC)₀₋₉₆ h of Br-C16-PX from the NPs was 91.7-fold and 49.6-fold greater than Taxol in plasma and tumor-bearing lungs, respectively, which provided sustained drug exposure and higher antitumor efficacy in the NP-treated group.

  20. Evaluating the Anticancer Properties of Liposomal Copper in a Nude Mouse Xenograft Model of Human Prostate Cancer: Formulation, In Vitro, In Vivo, Histology and Tissue Distribution Studies

    PubMed Central

    Wang, Yan; Zeng, San; Lin, Tien-Min; Krugner-Higby, Lisa; Lyman, Doug; Steffen, Dana; Xiong, May P.

    2014-01-01

    Purpose Although copper (Cu) complexes have been investigated as anticancer agents, there has been no description of Cu itself as a cancer killing agent. A stealth liposomal Cu formulation (LpCu) was studied in vitro and in vivo. Methods LpCu was evaluated in prostate cancer origin PC-3 cells by a metabolic cytotoxicity assay, by monitoring reactive oxygen species (ROS), and by flow cytometry. LpCu efficacy was evaluated in vivo using intratumoral and intravenous injections into mice bearing PC-3 xenograft tumors. Toxicology was assessed by performing hematological and blood biochemistry assays, and tissue histology and Cu distribution was investigated by elemental analysis. Results LpCu and free Cu salts displayed similar levels of cell metabolic toxicity and ROS. Flow cytometry indicated that the mechanisms of cell death were both apoptosis and necrosis. Animals injected i.t. with 3.5 mg/kg or i.v. with 3.5 and 7.0 mg/kg LpCu exhibited significant tumor growth inhibition. Kidney and eye were the main organs affected by Cu-mediated toxicities, but spleen and liver were the major organs of Cu deposition. Conclusions LpCu was effective at reducing tumor burden in the xenograft prostate cancer model. There was histological evidence of Cu toxicity in kidneys and eyes of animals treated at the maximum tolerated dose of LpCu 7.0 mg/kg. PMID:24848339

  1. Pre-transplant antibody screening and anti-CD154 costimulation blockade promote long-term xenograft survival in a pig-to-primate kidney transplant model

    PubMed Central

    Higginbotham, Laura; Mathews, Dave; Breeden, Cynthia A.; Song, Mingqing; Farris, Alton Brad; Larsen, Christian P.; Ford, Mandy L.; Lutz, Andrew J.; Tector, Matthew; Newell, Kenneth A.; Tector, A. Joseph; Adams, Andrew B.

    2016-01-01

    Xenotransplantation has the potential to alleviate the organ shortage that prevents many patients with end-stage renal disease from enjoying the benefits of kidney transplantation. Despite significant advances in other models, pig-to-primate kidney xenotransplantation has met limited success. Preformed anti-pig antibodies are an important component of the xenogeneic immune response. To address this, we screened a cohort of 34 rhesus macaques for anti-pig antibody levels. We then selected animals with both low and high titers of anti-pig antibodies to proceed with kidney transplant from galactose-α1,3-galactose knockout/CD55 transgenic pig donors. All animals received T-cell depletion followed by maintenance therapy with costimulation blockade (either anti-CD154 mAb or belatacept), mycophenolate mofetil, and steroid. The animal with the high titer of anti-pig antibody rejected the kidney xenograft within the first week. Low-titer animals treated with anti-CD154 antibody, but not belatacept exhibited prolonged kidney xenograft survival (>133 and >126 vs. 14 and 21 days, respectively). Long-term surviving animals treated with the anti-CD154-based regimen continue to have normal kidney function and preserved renal architecture without evidence of rejection on biopsies sampled at day 100. This description of the longest reported survival of pig-to-non-human primate kidney xenotransplantation, now >125 days, provides promise for further study and potential clinical translation. PMID:25847130

  2. Metformin impairs Rho GTPase signaling to induce apoptosis in neuroblastoma cells and inhibits growth of tumors in the xenograft mouse model of neuroblastoma

    PubMed Central

    Kumar, Ambrish; Al-Sammarraie, Nadia; DiPette, Donald J.; Singh, Ugra S.

    2014-01-01

    Metformin has been shown to inhibit tumor growth in xenograft rodent models of adult cancers, and various human clinical trials are in progress. However, the precise molecular mechanisms of metformin action are largely unknown. In the present study we examined the anti-tumor activity of metformin against neuroblastoma, and determined the underlying signaling mechanisms. Using human neuroblastoma xenograft mice, we demonstrated that oral administration of metformin (100 and 250 mg/kg body weight) significantly inhibited the growth of tumors. The interference of metformin in spheroid formation further confirmed the anti-tumor activity of metformin. In tumors, the activation of Rac1 (GTP-Rac1) and Cdc42 (GTP-Cdc42) was increased while RhoA activation (GTP-RhoA) was decreased by metformin. It also induced phosphorylation of JNK and inhibited the phosphorylation of ERK1/2 without affecting p38 MAP Kinase. Infection of cells by adenoviruses expressing dominant negative Rac1 (Rac1-N17), Cdc42 (Cdc42-N17) or constitutively active RhoA (RhoA-V14), or incubation of cells with pharmacological inhibitors of Rac1 (NSC23766) or Cdc42 (ML141) significantly protected neuroblastoma cells from metformin-induced apoptosis. Additionally, inhibition of JNK activity along with Rac1 or Cdc42 attenuated cytotoxic effects of metformin. These studies demonstrated that metformin impairs Rho GTPases signaling to induce apoptosis via JNK pathway. PMID:25365944

  3. Metformin impairs Rho GTPase signaling to induce apoptosis in neuroblastoma cells and inhibits growth of tumors in the xenograft mouse model of neuroblastoma.

    PubMed

    Kumar, Ambrish; Al-Sammarraie, Nadia; DiPette, Donald J; Singh, Ugra S

    2014-11-30

    Metformin has been shown to inhibit tumor growth in xenograft rodent models of adult cancers, and various human clinical trials are in progress. However, the precise molecular mechanisms of metformin action are largely unknown. In the present study we examined the anti-tumor activity of metformin against neuroblastoma, and determined the underlying signaling mechanisms. Using human neuroblastoma xenograft mice, we demonstrated that oral administration of metformin (100 and 250 mg/kg body weight) significantly inhibited the growth of tumors. The interference of metformin in spheroid formation further confirmed the anti-tumor activity of metformin. In tumors, the activation of Rac1 (GTP-Rac1) and Cdc42 (GTP-Cdc42) was increased while RhoA activation (GTP-RhoA) was decreased by metformin. It also induced phosphorylation of JNK and inhibited the phosphorylation of ERK1/2 without affecting p38 MAP Kinase. Infection of cells by adenoviruses expressing dominant negative Rac1 (Rac1-N17), Cdc42 (Cdc42-N17) or constitutively active RhoA (RhoA-V14), or incubation of cells with pharmacological inhibitors of Rac1 (NSC23766) or Cdc42 (ML141) significantly protected neuroblastoma cells from metformin-induced apoptosis. Additionally, inhibition of JNK activity along with Rac1 or Cdc42 attenuated cytotoxic effects of metformin. These studies demonstrated that metformin impairs Rho GTPases signaling to induce apoptosis via JNK pathway.

  4. Sunitinib (SUTENT, SU11248) suppresses tumor growth and induces apoptosis in xenograft models of human hepatocellular carcinoma.

    PubMed

    Huynh, H; Ngo, V C; Choo, S P; Poon, D; Koong, H N; Thng, C H; Toh, H C; Zheng, L; Ong, L C; Jin, Y; Song, I C; Chang, A P C; Ong, H S; Chung, A Y F; Chow, P K H; Soo, K C

    2009-09-01

    Hepatocellular carcinoma (HCC) is the fifth most common and third deadliest primary neoplasm. Since HCC is a particularly vascular solid tumor, we determined the antitumor and antiangiogenic activities of sunitinib malate, a potent inhibitor of two receptors involved in angiogenesis - vascular endothelial growth factor receptor (VEGFR) and platelet-derived growth factor receptor (PDGFR). In the present study, we reported that treatment of HepG2 and SK-Hep-1 cells with sunitinib led to growth inhibition and apoptosis in a dose-dependent fashion. Sunitinib inhibited phosphorylation of VEGFR-2 at Tyr951 and PDGFR-beta at Tyr1021 both in vitro and in vivo. Sunitinib also suppressed tumor growth of five patient-derived xenografts. Sunitinib-induced tumor growth inhibition was associated with increased apoptosis, reduced microvessel density and inhibition of cell proliferation. This study provides a strong rationale for further clinical investigation of sunitinib in patients with hepatocellular carcinoma.

  5. Comparative Efficacy of 177Lu and 90Y for Anti-CD20 Pretargeted Radioimmunotherapy in Murine Lymphoma Xenograft Models

    SciTech Connect

    Frost, Sophia; Frayo, Shani; Miller, Brian W.; Orozco, Johnnie J.; Booth, Garrett C.; Hylarides, Mark; Lin, Yukang; Green, Damian J.; Gopal, Ajay K.; Pagel, John M.; Back, Tom; Fisher, Darrell R.; Press, Oliver W.

    2015-03-01

    Pretargeted radioimmunotherapy (PRIT) is a multi-step method of selectively delivering high doses of radiotherapy to tumor cells while minimizing exposure to surrounding tissues. Yttrium-90 (90Y) and lutetium-177 (177Lu) are two of the most promising beta-particle emitting radionuclides used for radioimmunotherapy, which despite having similar chemistries differ distinctly in terms of radiophysical features. These differences may have important consequences for the absorbed dose to tumors and normal organs. Whereas 90Y has been successfully applied in a number of preclinical and clinical radioimmunotherapy settings, there have been few published pretargeting studies with 177Lu. We therefore compared the therapeutic potential of targeting either 90Y or 177Lu to human B-cell lymphoma xenografts in mice.

  6. ERK1/2 activation modulates pyocyanin-induced toxicity in A549 respiratory epithelial cells.

    PubMed

    Forbes, Amanda; Davey, Andrew K; Perkins, Anthony V; Grant, Gary D; McFarland, Amelia J; McDermott, Catherine M; Anoopkumar-Dukie, Shailendra

    2014-02-01

    Pyocyanin (PCN), a virulence factor produced by Pseudomonas aeruginosa, has many damaging effects on mammalian cells. Several lines of evidence suggest that this damage is primarily mediated by its ability to generate oxidative stress. However mechanisms underlying PCN-induced oxidative injury remain unclear. Although oxidative stress and subsequent MAPK signaling has been shown to modulate cell death in other models, its role in PCN-induced cytotoxicity remains unknown. Therefore the aim of this study was to investigate the role of redox-sensitive MAPK in PCN-induced toxicity in A549 cells. Here we show that PCN (50μM) rapidly increased ERK1/2 phosphorylation after 5min. Pre-treatment of A549 cells with the MEK1/2 inhibitor U0126 (10μM) decreased PCN-induced ERK1/2 phosphorylation and protected cells against apoptosis and cell injury suggesting a role for ERK signalling. In contrast, JNK and p38 MAPK phosphorylation remained unchanged following exposure to PCN and pretreatment with either the JNK or p38 MAPK inhibitors (10μM SP600125 and 10μM SB203580, respectively) did not afford protection against PCN toxicity. This would suggest that PCN-induced cytotoxicity appears to occur independently of JNK and p38 MAPK signaling pathways. Finally, although we confirm that oxidative stress contributes to PCN-induced toxicity, our data suggest the contribution of oxidative stress is independent of ERK1/2 signaling. These findings may provide insight for novel targeted therapies to reduce PCN-mediated lung injury in patients with chronic P. aeruginosa respiratory infections.

  7. Pharmacokinetic-pharmacodynamic modeling of the antitumor effect of TM208 and EGFR-TKI resistance in human breast cancer xenograft mice

    PubMed Central

    Ji, Xi-wei; Ji, Shuang-min; Li, Run-tao; Wu, Ke-hua; Zhu, Xiao; Lu, Wei; Zhou, Tian-yan

    2016-01-01

    Aim: The novel anticancer compound TM208 is an EGFR tyrosine kinase inhibitor (EGFR-TKI). Since the development of resistance to EGFR-TKIs is a major challenge in their clinical usage, we investigated the profiles of resistance following continuous treatment with TM208 in human breast cancer xenograft mice, and identified the relationship between the tumor pEGFR levels and tumor growth inhibition. Methods: Female BALB/c nude mice were implanted with human breast cancer MCF-7 cells, and the xenograft mice received TM208 (50 or 150 mg·kg−1·d−1, ig) or vehicle for 18 d. The pharmacokinetics (PK) and pharmacodynamics (PD) of TM208 were evaluated. Results: The PK properties of TM208 were described by a one-compartment model with first-order absorption kinetics. Our study showed the inhibitory effects of TM208 on tumor pEGFR levels gradually reached a maximum effect, after which it became weaker over time, which was characterized by a combined tolerance/indirect response PD model with an estimated EC50 (55.9 μg/L), as well as three parameters ('a' of 27.2%, 'b' of 2730%, 'c' of 0.58 h−1) denoting the maximum, extent and rate of resistance, respectively. The relationship between the tumor pEGFR levels and tumor growth inhibition was characterized by a combined logistic tumor growth/transit compartment model with estimated parameters associated with tumor growth characteristics kng (0.282 day−1), drug potency kTM208 (0.0499 cm3/day) and the kinetics of tumor cell death k1 (0.141 day−1), which provided insight into drug mechanisms and behaviors. Conclusion: The proposed PK/PD model provides a better understanding of the pharmacological properties of TM208 in the treatment of breast cancer. Furthermore, simulation based on a tolerance model allows prediction of the occurrence of resistance. PMID:27133303

  8. In Silico cancer cell versus stroma cellularity index computed from species-specific human and mouse transcriptome of xenograft models: towards accurate stroma targeting therapy assessment

    PubMed Central

    2014-01-01

    Background The current state of the art for measuring stromal response to targeted therapy requires burdensome and rate limiting quantitative histology. Transcriptome measures are increasingly affordable and provide an opportunity for developing a stromal versus cancer ratio in xenograft models. In these models, human cancer cells are transplanted into mouse host tissues (stroma) and together coevolve into a tumour microenvironment. However, profiling the mouse or human component separately remains problematic. Indeed, laser capture microdissection is labour intensive. Moreover, gene expression using commercial microarrays introduces significant and underreported cross-species hybridization errors that are commonly overlooked by biologists. Method We developed a customized dual-species array, H&M array, and performed cross-species and species-specific hybridization measurements. We validated a new methodology for establishing the stroma vs cancer ratio using transcriptomic data. Results In the biological validation of the H&M array, cross-species hybridization of human and mouse probes was significantly reduced (4.5 and 9.4 fold reduction, respectively; p < 2x10-16 for both, Mann-Whitney test). We confirmed the capability of the H&M array to determine the stromal to cancer cells ratio based on the estimation of cellularity index of mouse/human mRNA content in vitro. This new metrics enable to investigate more efficiently the stroma-cancer cell interactions (e.g. cellularity) bypassing labour intensive requirement and biases of laser capture microdissection. Conclusion These results provide the initial evidence of improved and cost-efficient analytics for the investigation of cancer cell microenvironment, using species-specificity arrays specifically designed for xenografts models. PMID:25079962

  9. A novel orally available inhibitor of focal adhesion signaling increases survival in a xenograft model of diffuse large B-cell lymphoma with central nervous system involvement.

    PubMed

    Bosch, Rosa; Moreno, María José; Dieguez-Gonzalez, Rebeca; Céspedes, María Virtudes; Gallardo, Alberto; Trias, Manuel; Grañena, Albert; Sierra, Jorge; Casanova, Isolda; Mangues, Ramon

    2013-08-01

    Central nervous system dissemination is a relatively uncommon but almost always fatal complication in diffuse large B-cell lymphoma patients. Optimal therapy for central nervous involvement in this malignancy has not been established. In this paper, we aimed to evaluate the therapeutic effect of E7123, a celecoxib derivative that inhibits focal adhesion signaling, in a novel xenograft model of diffuse large B-cell lymphoma with central nervous system involvement. Cells obtained after disaggregation of HT subcutaneous tumors (HT-SC cells) were intravenously injected in NOD/SCID mice. These mice received oral vehicle or 75 mg/kg of E7123 daily until they were euthanized for weight loss or signs of sickness. The antitumor effect of E7123 was validated in an independent experiment using a bioluminescent mouse model. Intravenously injected HT-SC cells showed higher take rate and higher central nervous system tropism (associated with increased expression of β1-integrin and p130Cas proteins) than HT cells. The oral administration of E7123 significantly increased survival time in 2 independent experiments using mice injected with unmodified or bioluminescent HT-SC cells. We have developed a new xenograft model of diffuse large B-cell lymphoma with central nervous system involvement that can be used in the pre-clinical evaluation of new drugs for this malignancy. E7123 is a new, well-tolerated and orally available therapeutic agent that merits further investigation since it may improve current management of diffuse large B-cell lymphoma patients with central nervous system involvement.

  10. Scopoletin, an active principle of tree tobacco (Nicotiana glauca) inhibits human tumor vascularization in xenograft models and modulates ERK1, VEGF-A, and FGF-2 in computer model.

    PubMed

    Tabana, Yasser M; Hassan, Loiy Elsir A; Ahamed, Mohamed B Khadeer; Dahham, Saad S; Iqbal, Muhammad Adnan; Saeed, Mohammed A A; Khan, Md Shamsuddin S; Sandai, Doblin; Majid, Aman S Abdul; Oon, Chern Ein; Majid, Amin Malik S A

    2016-09-01

    We recently reported the antineovascularization effect of scopoletin on rat aorta and identified its potential anti-angiogenic activity. Scopoletin could be useful as a systemic chemotherapeutic agent against angiogenesis-dependent malignancies if its antitumorigenic activity is investigated and scientifically proven using a suitable human tumor xenograft model. In the present study, bioassay-guided (anti-angiogenesis) phytochemical investigation was conducted on Nicotiana glauca extract which led to the isolation of scopoletin. Further, anti-angiogenic activity of scopoletin was characterized using ex vivo, in vivo and in silico angiogenesis models. Finally, the antitumorigenic efficacy of scopoletin was studied in human colorectal tumor xenograft model using athymic nude mice. For the first time, an in vivo anticancer activity of scopoletin was reported and characterized using xenograft models. Scopoletin caused significant suppression of sprouting of microvessels in rat aortic explants with IC50 (median inhibitory concentration) 0.06μM. Scopoletin (100 and 200mg/kg) strongly inhibited (59.72 and 89.4%, respectively) vascularization in matrigel plugs implanted in nude mice. In the tumor xenograft model, scopoletin showed remarkable inhibition on tumor growth (34.2 and 94.7% at 100 and 200mg/kg, respectively). Tumor histology revealed drastic reduction of the extent of vascularization. Further, immunostaining of CD31 and NG2 receptors in the histological sections confirmed the antivascular effect of scopoletin in tumor vasculature. In computer modeling, scopoletin showed strong ligand affinity and binding energies toward the following angiogenic factors: protein kinase (ERK1), vascular endothelial growth factor A (VEGF-A), and fibroblast growth factor 2 (FGF-2). These results suggest that the antitumor activity of scopoletin may be due to its strong anti-angiogenic effect, which may be mediated by its effective inhibition of ERK1, VEGF-A, and FGF-2. PMID:27133199

  11. Protective effect of curcumin against formaldehyde-induced genotoxicity in A549 Cell Lines.

    PubMed

    Zhang, Ben-Yan; Shi, Yu-Qin; Chen, Xin; Dai, Juan; Jiang, Zhong-Fa; Li, Ning; Zhang, Zhi-Bing

    2013-12-01

    Formaldehyde is ubiquitous in the environment. It is known to be a genotoxic substance. We hypothesized that reactive oxygen species (ROS) and lipid peroxidation are involved in formaldehyde-induced genotoxicity in human lung cancer cell lines A549. To test this hypothesis, we investigated the effects of antioxidant on formaldehyde-induced genotoxicity in A549 Cell Lines. Formaldehyde exposure caused induction of DNA-protein cross-links (DPCs). Curcumin is an important antioxidant. Formaldehyde significantly increased malondialdehyde (MDA) levels, and decreased superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) activity. In addition, the activation of NF-κB and AP-1 were induced by formaldehyde treatment. Pretreatment with curcumin counteracted formaldehyde-induced oxidative stress, ameliorated DPCs and attenuated activation of NF-κB and AP-1 in A549 Cell Lines. These results, taken together, suggest that formaldehyde induced genotoxicity through its ROS and lipid peroxidase activity and caused DPCs effects in A549 cells.

  12. An Orally Bioavailable, Indole-3-glyoxylamide Based Series of Tubulin Polymerization Inhibitors Showing Tumor Growth Inhibition in a Mouse Xenograft Model of Head and Neck Cancer.

    PubMed

    Colley, Helen E; Muthana, Munitta; Danson, Sarah J; Jackson, Lucinda V; Brett, Matthew L; Harrison, Joanne; Coole, Sean F; Mason, Daniel P; Jennings, Luke R; Wong, Melanie; Tulasi, Vamshi; Norman, Dennis; Lockey, Peter M; Williams, Lynne; Dossetter, Alexander G; Griffen, Edward J; Thompson, Mark J

    2015-12-10

    A number of indole-3-glyoxylamides have previously been reported as tubulin polymerization inhibitors, although none has yet been successfully developed clinically. We report here a new series of related compounds, modified according to a strategy of reducing aromatic ring count and introducing a greater degree of saturation, which retain potent tubulin polymerization activity but with a distinct SAR from previously documented libraries. A subset of active compounds from the reported series is shown to interact with tubulin at the colchicine binding site, disrupt the cellular microtubule network, and exert a cytotoxic effect against multiple cancer cell lines. Two compounds demonstrated significant tumor growth inhibition in a mouse xenograft model of head and neck cancer, a type of the disease which often proves resistant to chemotherapy, supporting further development of the current series as potential new therapeutics.

  13. Murine xenograft model demonstrates significant radio-sensitising effect of liposomal doxorubicin in a combination therapy for Feline Injection Site Sarcoma.

    PubMed

    Petznek, H; Kleiter, M; Tichy, A; Fuchs-Baumgartinger, A; Hohenadl, C

    2014-10-01

    Therapy of cats suffering from feline injection site sarcomas (FISS) is still a challenging problem, as the recurrence rate after surgery is up to 70%. Four FISS-derived primary tumour cell lines and corresponding xenograft tumour mouse models were established to evaluate the efficacy of a concomitant chemo-/radiation therapy with doxorubicin. In vitro, strongly depending upon the timing of administration, pre-treatment with 0.25 µmol doxorubicin resulted in a significant enhancement of radiation-induced (3.5 Gy) tumour cell death. This result was confirmed in vivo, where only the combined chemo-/radiation therapy resulted in a significant reduction in tumour growth compared to the respective mono-therapies with either doxorubicin or radiation. These results support the use of the concomitant chemo-/radiation therapy for adjuvant treatment of FISS, particularly in advanced or recurrent disease where surgery alone is no longer feasible. PMID:25104320

  14. Trichomonas vaginalis induces cytopathic effect on human lung alveolar basal carcinoma epithelial cell line A549.

    PubMed

    Salvador-Membreve, Daile Meek C; Jacinto, Sonia D; Rivera, Windell L

    2014-12-01

    Trichomonas vaginalis, the causative agent of trichomoniasis is generally known to inhabit the genitourinary tract. However, several case reports with supporting molecular and immunological identifications have documented its occurrence in the respiratory tract of neonates and adults. In addition, the reports have documented that its occurrence is associated with respiratory failures. The medical significance or consequence of this association is unclear. Thus, to establish the possible outcome from the interaction of T. vaginalis with lung cells, the cytopathic effects of the parasites were evaluated using monolayer cultures of the human lung alveolar basal carcinoma epithelial cell line A549. The possible effect of association of T. vaginalis with A549 epithelial cells was analyzed using phase-contrast, scanning electron microscopy and fluorescence microscopy. MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide), crystal-violet and TUNEL (terminal deoxynucleotidyltransferase-mediated dUTP nick-end labelling) assays were conducted for cytotoxicity testing. The results demonstrate that T. vaginalis: (1) adheres to A549 epithelial cells, suggesting a density-dependent parasite-cell association; (2) adherence on A549 is through flagella, membrane and axostyle; (3) causes cell detachment and cytotoxicity (50-72.4%) to A549 and this effect is a function of parasite density; and (4) induces apoptosis in A549 about 20% after 6 h of incubation. These observations indicate that T. vaginalis causes cytopathic effects on A549 cell. To date, this is the first report showing a possible interaction of T. vaginalis with the lung cells using A549 monolayer cultures. Further studies are recommended to completely elucidate this association.

  15. Radiation-Induced Bystander Effects in A549 Cells Exposed to 6 MV X-rays.

    PubMed

    Yang, Shuning; Xu, Jing; Shao, Weixian; Geng, Chong; Li, Jia; Guo, Feng; Miao, Hui; Shen, Wenbin; Ye, Tao; Liu, Yazhou; Xu, Haiting; Zhang, Xuguang

    2015-07-01

    The aim of the study is to explore the bystander effects in A549 cells that have been exposed to 6MV X-ray. Control group, irradiated group, irradiated conditioned medium (ICM)-received group, and fresh medium group were designed in this study. A549 cells in the logarithmic growth phase were irradiated with 6MV X-ray at 0, 0.5, 1, 1.5, and 2. In ICM-received group, post-irradiation A549 cells were cultured for 3 h and were transferred into non-irradiated A549 cells for further cultivation. Clone forming test was applied to detect the survival fraction of cells. Annexin V-FITC/PI double-staining assay was used to detect the apoptosis of A549 cells 24, 48, 72, and 96 h after 2-Gy 6MV X-ray irradiation, and the curves of apoptosis were drawn. The changes in the cell cycles 4, 48, 72, and 96 h after 2-Gy 6MV X-ray irradiation were detected using PI staining flow cytometry. With the increase of irradiation dose, the survival fraction of A549 cells after the application of 0.5 Gy irradiation was decreasing continuously. In comparison to the control group, the apoptosis rate of the ICM-received group was increased in a time-dependent pattern, with the highest apoptosis rate observed at 72 h (p < 0.05). Cell count in G2/M stages was obviously increased compared with that of the control group (p < 0.05), with the highest count observed at 72 h, after which G2/M stage arrest was diminished. ICM can cause apparent A549 cell damage, indicating that 6MV X-ray irradiation can induce bystander effect on A549 cells, which reaches a peak at 72 h.

  16. Scanning Acoustic Microscopy-A Novel Noninvasive Method to Determine Tumor Interstitial Fluid Pressure in a Xenograft Tumor Model.

    PubMed

    Hofmann, Matthias; Pflanzer, Ralph; Habib, Anowarul; Shelke, Amit; Bereiter-Hahn, Jürgen; Bernd, August; Kaufmann, Roland; Sader, Robert; Kippenberger, Stefan

    2016-06-01

    Elevated tumor interstitial fluid pressure (TIFP) is a prominent feature of solid tumors and hampers the transmigration of therapeutic macromolecules, for example, large monoclonal antibodies, from tumor-supplying vessels into the tumor interstitium. TIFP values of up to 40 mm Hg have been measured in experimental solid tumors using two conventional invasive techniques: the wick-in-needle and the micropuncture technique. We propose a novel noninvasive method of determining TIFP via ultrasonic investigation with scanning acoustic microscopy at 30-MHz frequency. In our experimental setup, we observed for the impedance fluctuations in the outer tumor hull of A431-vulva carcinoma-derived tumor xenograft mice. The gain dependence of signal strength was quantified, and the relaxation of tissue was calibrated with simultaneous hydrostatic pressure measurements. Signal patterns from the acoustical images were translated into TIFP curves, and a putative saturation effect was found for tumor pressures larger than 3 mm Hg. This is the first noninvasive approach to determine TIFP values in tumors. This technique can provide a potentially promising noninvasive assessment of TIFP and, therefore, can be used to determine the TIFP before treatment approach as well to measure therapeutic efficacy highlighted by lowered TFP values. PMID:27267834

  17. Filamentous, mixed micelles of triblock copolymers enhance tumor localization of indocyanine green in a murine xenograft model

    PubMed Central

    Kim, Tae Hee; Mount, Christopher W; Dulken, Benjamin W; Ramos, Jenelyn; Fu, Caroline J; Khant, Htet A; Chiu, Wah; Gombotz, Wayne R; Pun, Suzie H

    2012-01-01

    Polymeric micelles formed by the self-assembly of amphiphilic block copolymers can be used to encapsulate hydrophobic drugs for tumor-delivery applications. Filamentous carriers with high aspect ratios offer potential advantages over spherical carriers, including prolonged circulation times. In this work, mixed micelles comprised of poly (ethylene oxide)-poly-[(R)-3-hydroxybutyrate]-poly (ethylene oxide) (PEO-PHB-PEO) and Pluronic F-127 (PF-127) were used to encapsulate a near-infrared fluorophore. The micelle formulations were assessed for tumor accumulation after tail vein injection to xenograft tumor-bearing mice by non-invasive optical imaging. The mixed micelle formulation that facilitated the highest tumor accumulation was shown by cryo-electron microscopy to be filamentous in structure compared to spherical structures of pure PF-127 micelles. In addition, increased dye loading efficiency and dye stability was attained in this mixed micelle formulation compared to pure PEO-PHB-PEO micelles. Therefore, the optimized PEO-PHB-PEO/PF-127 mixed micelle formulation offers advantages for cancer delivery over micelles formed from the individual copolymer components. PMID:22118658

  18. Filamentous, mixed micelles of triblock copolymers enhance tumor localization of indocyanine green in a murine xenograft model.

    PubMed

    Kim, Tae Hee; Mount, Christopher W; Dulken, Benjamin W; Ramos, Jenelyn; Fu, Caroline J; Khant, Htet A; Chiu, Wah; Gombotz, Wayne R; Pun, Suzie H

    2012-01-01

    Polymeric micelles formed by the self-assembly of amphiphilic block copolymers can be used to encapsulate hydrophobic drugs for tumor-delivery applications. Filamentous carriers with high aspect ratios offer potential advantages over spherical carriers, including prolonged circulation times. In this work, mixed micelles composed of poly(ethylene oxide)-poly[(R)-3-hydroxybutyrate]-poly(ethylene oxide) (PEO-PHB-PEO) and Pluronic F-127 (PF-127) were used to encapsulate a near-infrared fluorophore. The micelle formulations were assessed for tumor accumulation after tail vein injection to xenograft tumor-bearing mice by noninvasive optical imaging. The mixed micelle formulation that facilitated the highest tumor accumulation was shown by cryo-electron microscopy to be filamentous in structure compared to spherical structures of pure PF-127 micelles. In addition, increased dye loading efficiency and dye stability were attained in this mixed micelle formulation compared to pure PEO-PHB-PEO micelles. Therefore, the optimized PEO-PHB-PEO/PF-127 mixed micelle formulation offers advantages for cancer delivery over micelles formed from the individual copolymer components.

  19. Silencing of APE1 enhances sensitivity of human hepatocellular carcinoma cells to radiotherapy in vitro and in a xenograft model.

    PubMed

    Cun, Yanping; Dai, Nan; Xiong, Chengjie; Li, Mengxia; Sui, Jiangdong; Qian, Chengyuan; Li, Zheng; Wang, Dong

    2013-01-01

    Resistance to radiotherapy is a key limitation for the treatment of human hepatocellular carcinoma (HCC). To overcome this problem, we investigated the correlation between radioresistance and the human apurinic/apyrimidinic endonuclease (APE1), a bifunctional protein, which plays an important role in DNA repair and redox regulation activity of transcription factors. In the present study, we examined the radiosensitivity profiles of three human HCC cell lines, HepG2, Hep3B, and MHCC97L, using the adenoviral vector Ad5/F35-mediated APE1 siRNA (Ad5/F35-siAPE1). The p53 mutant cell lines MHCC97L showed radioresistance, compared with HepG2 and Hep3B cells. APE1 was strongly expressed in MHCC97L cells and was induced by irradiation in a dose-dependent manner, and Ad5/F35-siAPE1 effectively inhibited irradiation-induced APE1 and p53 expression. Moreover, silencing of APE1 significantly potentiated the growth inhibition and apoptosis induction by irradiation in all tested human HCC cell lines. In addition, Ad5/F35-siAPE1 significantly enhanced inhibition of tumor growth and potentiated cell apoptosis by irradiation both in HepG2 and MHCC97L xenografts. In conclusion, down regulation of APE1 could enhance sensitivity of human HCC cells to radiotherapy in vitro and in vivo.

  20. Anti-CD45 Pretargeted Radioimmunotherapy using Bismuth-213: High Rates of Complete Remission and Long-Term Survival in a Mouse Myeloid Leukemia Xenograft Model

    SciTech Connect

    Pagel, John M; Kenoyer, Aimee L; Back, Tom; Hamlin, Donald K; Wilbur, D Scott; Fisher, Darrell R; Park, Steven I; Frayo, Shani; Axtman, Amanda; Orgun, Nural; Orozoco, Johnnie; Shenoi, Jaideep; Lin, Yukang; Gopal, Ajay K; Green, Damian J; Appelbaum, Frederick R; Press, Oliver W

    2011-07-21

    Pretargeted radioimmunotherapy (PRIT) using an anti-CD45 antibody (Ab)-streptavidin (SA) conjugate and DOTA-biotin labeled with β-emitting radionuclides has been explored as a strategy to decrease relapse and toxicity. α-emitting radionuclides exhibit high cytotoxicity coupled with a short path-length, potentially increasing the therapeutic index and making them an attractive alternative to β-emitting radionuclides for patients with Acute Myeloid Leukemia (AML). Accordingly, we have used 213Bi in mice with human leukemia xenografts. Results demonstrated excellent localization of 213Bi-DOTA-biotin to tumors with minimal uptake into normal organs. After 10 minutes, 4.5 ± 1.1% of the injected dose of 213Bi was delivered per gram of tumor. α imaging demonstrated uniform radionuclide distribution within tumor tissue 45 minutes after 213Bi-DOTA-biotin injection. Radiation absorbed doses were similar to those observed using a β-emitting radionuclide (90Y) in the same model. We conducted therapy experiments in a xenograft model using a single-dose of 213Bi-DOTA-biotin given 24 hours after anti-CD45 Ab-SA conjugate. Among mice treated with anti-CD45 Ab-SA conjugate followed by 800 μCi of 213Bi- or 90Y-DOTA-biotin, 80% and 20%, respectively, survived leukemia-free for >100 days with minimal toxicity. These data suggest that anti-CD45 PRIT using an α-emitting radionuclide may be highly effective and minimally toxic for treatment of AML.

  1. Thymoquinone overcomes chemoresistance and enhances the anticancer effects of bortezomib through abrogation of NF-κB regulated gene products in multiple myeloma xenograft mouse model

    PubMed Central

    Siveen, Kodappully Sivaraman; Mustafa, Nurulhuda; Li, Feng; Kannaiyan, Radhamani; Ahn, Kwang Seok; Kumar, Alan Prem; Chng, Wee-Joo; Sethi1, Gautam

    2014-01-01

    Multiple myeloma (MM) is a B cell malignancy characterized by clonal proliferation of plasma cells in the bone marrow. With the advent of novel targeted agents, the median survival rate has increased to 5−7 years. However, majority of patients with myeloma suffer relapse or develop chemoresistance to existing therapeutic agents. Thus, there is a need to develop novel alternative therapies for the treatment of MM. Thus in the present study, we investigated whether thymoquinone (TQ), a bioactive constituent of black seed oil, could suppress the proliferation and induce chemosensitization in human myeloma cells and xenograft mouse model. Our results show that TQ inhibited the proliferation of MM cells irrespective of their sensitivity to doxorubicin, melphalan or bortezomib. Interestingly, TQ treatment also resulted in a significant inhibition in the proliferation of CD138+ cells isolated from MM patient samples in a concentration dependent manner. TQ also potentiated the apoptotic effects of bortezomib in various MM cell lines through the activation of caspase-3, resulting in the cleavage of PARP. TQ treatment also inhibited chemotaxis and invasion induced by CXCL12 in MM cells. Furthermore, in a xenograft mouse model, TQ potentiated the antitumor effects of bortezomib (p < 0.05, vehicle versus bortezomib + TQ; p < 0.05, bortezomib versus bortezomib + TQ), and this correlated with modulation of various markers for survival and angiogenesis, such as Ki-67, vascular endothelial growth factor (VEGF), Bcl-2 and p65 expression. Overall, our results demonstrate that TQ can enhance the anticancer activity of bortezomib in vitro and in vivo and may have a substantial potential in the treatment of MM. PMID:24504138

  2. Yeast-Derived β-Glucan Augments the Therapeutic Efficacy Mediated by Anti–Vascular Endothelial Growth Factor Monoclonal Antibody in Human Carcinoma Xenograft Models

    PubMed Central

    Salvador, Carolina; Li, Bing; Hansen, Richard; Cramer, Daniel E.; Kong, Maiying; Jun, Yan

    2008-01-01

    Purpose Bevacizumab is a recombinant IgG1humanized monoclonal antibody against vascular endothelial growth factor (VEGF). Its proposed mechanism of action is independent of immune effector functions. Many human carcinomas not only secrete VEGF but also express membrane-bound VEGF. In addition, VEGF receptors are expressed on tumor cells. It is hypothesized that bevacizumab could bind membrane-bound VEGF or VEGF-VEGF receptor complexes on tumors, thereby initiating potential immunologic consequences. We previously showed that yeast-derived β-glucan functions with antitumor antibodies that activate complement to recruit complement receptor 3– expressing leukocytes capable of mediating complement receptor 3– dependent cellular cytotoxicity of tumors opsonized with iC3b. In the current study, the therapeutic efficacy mediated by combining bevacizumab with yeast-derived β-glucan was studied in human carcinoma xenograft models. Experimental Design Human tumor cell lines were screened for membrane-bound VEGF expression both in vitro and in vivo. Complement activation mediated by bevacizumab was examined. Tumor cell lines positive or negative for membrane-bound VEGF expression were implanted in severe combined immunodeficient mice to establish xenograft models. Tumor-bearing mice were treated with different regimens. Tumor regression and long-term survival were recorded. Results Human ovarian carcinoma SKOV-3 cells expressed membrane-bound VEGF both in vitro and in vivo. Bevacizumab was bound to membrane-bound VEGF, activated complement, and synergized with β-glucan to elicit cellular cytotoxicity in vitro. In vivo study showed that β-glucan could significantly augment the therapeutic efficacy mediated by bevacizumab. Conclusions Yeast-derived β-glucan can synergize with anti-VEGF monoclonal antibody bevacizumab for the treatment of cancer with membrane-bound VEGF expression. PMID:18281559

  3. Restriction of dietary protein decreases mTORC1 in tumors and somatic tissues of a tumor-bearing mouse xenograft model

    PubMed Central

    Cummings, Nicole E.; Rastelli, Antonella L.; Gao, Feng; Cava, Edda; Bertozzi, Beatrice; Spelta, Francesco; Pili, Roberto

    2015-01-01

    Reduced dietary protein intake and intermittent fasting (IF) are both linked to healthy longevity in rodents, and are effective in inhibiting cancer growth. The molecular mechanisms underlying the beneficial effects of chronic protein restriction (PR) and IF are unclear, but may be mediated in part by a down-regulation of the IGF/mTOR pathway. In this study we compared the effects of PR and IF on tumor growth in a xenograft mouse model of breast cancer. We also investigated the effects of PR and IF on the mechanistic Target Of Rapamycin (mTOR) pathway, inhibition of which extends lifespan in model organisms including mice. The mTOR protein kinase is found in two distinct complexes, of which mTOR complex 1 (mTORC1) is responsive to acute treatment with amino acids in cell culture and in vivo. We found that both PR and IF inhibit tumor growth and mTORC1 phosphorylation in tumor xenografts. In somatic tissues, we found that PR, but not IF, selectively inhibits the activity of the amino acid sensitive mTORC1, while the activity of the second mTOR complex, mTORC2, was relatively unaffected by PR. In contrast, IF resulted in increased S6 phosphorylation in multiple metabolic tissues. Our work represents the first finding that PR may reduce mTORC1 activity in tumors and multiple somatic tissues, and suggest that PR may represent a highly translatable option for the treatment not only of cancer, but also other age-related diseases. PMID:26378060

  4. Restriction of dietary protein decreases mTORC1 in tumors and somatic tissues of a tumor-bearing mouse xenograft model.

    PubMed

    Lamming, Dudley W; Cummings, Nicole E; Rastelli, Antonella L; Gao, Feng; Cava, Edda; Bertozzi, Beatrice; Spelta, Francesco; Pili, Roberto; Fontana, Luigi

    2015-10-13

    Reduced dietary protein intake and intermittent fasting (IF) are both linked to healthy longevity in rodents, and are effective in inhibiting cancer growth. The molecular mechanisms underlying the beneficial effects of chronic protein restriction (PR) and IF are unclear, but may be mediated in part by a down-regulation of the IGF/mTOR pathway. In this study we compared the effects of PR and IF on tumor growth in a xenograft mouse model of breast cancer. We also investigated the effects of PR and IF on the mechanistic Target Of Rapamycin (mTOR) pathway, inhibition of which extends lifespan in model organisms including mice. The mTOR protein kinase is found in two distinct complexes, of which mTOR complex 1 (mTORC1) is responsive to acute treatment with amino acids in cell culture and in vivo. We found that both PR and IF inhibit tumor growth and mTORC1 phosphorylation in tumor xenografts. In somatic tissues, we found that PR, but not IF, selectively inhibits the activity of the amino acid sensitive mTORC1, while the activity of the second mTOR complex, mTORC2, was relatively unaffected by PR. In contrast, IF resulted in increased S6 phosphorylation in multiple metabolic tissues. Our work represents the first finding that PR may reduce mTORC1 activity in tumors and multiple somatic tissues, and suggest that PR may represent a highly translatable option for the treatment not only of cancer, but also other age-related diseases. PMID:26378060

  5. A Small Molecule Inhibitor of ETV1, YK-4-279, Prevents Prostate Cancer Growth and Metastasis in a Mouse Xenograft Model

    PubMed Central

    Rahim, Said; Minas, Tsion; Hong, Sung-Hyeok; Justvig, Sarah; Çelik, Haydar; Kont, Yasemin Saygideger; Han, Jenny; Kallarakal, Abraham T.; Kong, Yali; Rudek, Michelle A.; Brown, Milton L.; Kallakury, Bhaskar; Toretsky, Jeffrey A.; Üren, Aykut

    2014-01-01

    Background The erythroblastosis virus E26 transforming sequences (ETS) family of transcription factors consists of a highly conserved group of genes that play important roles in cellular proliferation, differentiation, migration and invasion. Chromosomal translocations fusing ETS factors to promoters of androgen responsive genes have been found in prostate cancers, including the most clinically aggressive forms. ERG and ETV1 are the most commonly translocated ETS proteins. Over-expression of these proteins in prostate cancer cells results in a more invasive phenotype. Inhibition of ETS activity by small molecule inhibitors may provide a novel method for the treatment of prostate cancer. Methods and Findings We recently demonstrated that the small molecule YK-4-279 inhibits biological activity of ETV1 in fusion-positive prostate cancer cells leading to decreased motility and invasion in-vitro. Here, we present data from an in-vivo mouse xenograft model. SCID-beige mice were subcutaneously implanted with fusion-positive LNCaP-luc-M6 and fusion-negative PC-3M-luc-C6 tumors. Animals were treated with YK-4-279, and its effects on primary tumor growth and lung metastasis were evaluated. YK-4-279 treatment resulted in decreased growth of the primary tumor only in LNCaP-luc-M6 cohort. When primary tumors were grown to comparable sizes, YK-4-279 inhibited tumor metastasis to the lungs. Expression of ETV1 target genes MMP7, FKBP10 and GLYATL2 were reduced in YK-4-279 treated animals. ETS fusion-negative PC-3M-luc-C6 xenografts were unresponsive to the compound. Furthermore, YK-4-279 is a chiral molecule that exists as a racemic mixture of R and S enantiomers. We established that (S)-YK-4-279 is the active enantiomer in prostate cancer cells. Conclusion Our results demonstrate that YK-4-279 is a potent inhibitor of ETV1 and inhibits both the primary tumor growth and metastasis of fusion positive prostate cancer xenografts. Therefore, YK-4-279 or similar compounds may be

  6. Genomic characterization of patient-derived xenograft models established from fine needle aspirate biopsies of a primary pancreatic ductal adenocarcinoma and from patient-matched metastatic sites

    PubMed Central

    de Abreu, Francine B.; Gardner, Timothy B.; Gordon, Stuart R.; Barth, Richard J.; Colacchio, Thomas A.; Wood, Matthew; Kacsoh, Balint Z.; Bouley, Stephanie J.; Cui, Jingxuan; Hamilton, Joanna; Choi, Jungbin A.; Lange, Joshua T.; Peterson, Jason D.; Padmanabhan, Vijayalakshmi; Tomlinson, Craig R.; Tsongalis, Gregory J.; Suriawinata, Arief A.; Smith, Kerrington D.

    2016-01-01

    N-of-1 trials target actionable mutations, yet such approaches do not test genomically-informed therapies in patient tumor models prior to patient treatment. To address this, we developed patient-derived xenograft (PDX) models from fine needle aspiration (FNA) biopsies (FNA-PDX) obtained from primary pancreatic ductal adenocarcinoma (PDAC) at the time of diagnosis. Here, we characterize PDX models established from one primary and two metastatic sites of one patient. We identified an activating KRAS G12R mutation among other mutations in these models. In explant cells derived from these PDX tumor models with a KRAS G12R mutation, treatment with inhibitors of CDKs (including CDK9) reduced phosphorylation of a marker of CDK9 activity (phospho-RNAPII CTD Ser2/5) and reduced viability/growth of explant cells derived from PDAC PDX models. Similarly, a CDK inhibitor reduced phospho-RNAPII CTD Ser2/5, increased apoptosis, and inhibited tumor growth in FNA-PDX and patient-matched metastatic-PDX models. In summary, PDX models can be constructed from FNA biopsies of PDAC which in turn can enable genomic characterization and identification of potential therapies. PMID:26934555

  7. Genomic characterization of patient-derived xenograft models established from fine needle aspirate biopsies of a primary pancreatic ductal adenocarcinoma and from patient-matched metastatic sites.

    PubMed

    Allaway, Robert J; Fischer, Dawn A; de Abreu, Francine B; Gardner, Timothy B; Gordon, Stuart R; Barth, Richard J; Colacchio, Thomas A; Wood, Matthew; Kacsoh, Balint Z; Bouley, Stephanie J; Cui, Jingxuan; Hamilton, Joanna; Choi, Jungbin A; Lange, Joshua T; Peterson, Jason D; Padmanabhan, Vijayalakshmi; Tomlinson, Craig R; Tsongalis, Gregory J; Suriawinata, Arief A; Greene, Casey S; Sanchez, Yolanda; Smith, Kerrington D

    2016-03-29

    N-of-1 trials target actionable mutations, yet such approaches do not test genomically-informed therapies in patient tumor models prior to patient treatment. To address this, we developed patient-derived xenograft (PDX) models from fine needle aspiration (FNA) biopsies (FNA-PDX) obtained from primary pancreatic ductal adenocarcinoma (PDAC) at the time of diagnosis. Here, we characterize PDX models established from one primary and two metastatic sites of one patient. We identified an activating KRAS G12R mutation among other mutations in these models. In explant cells derived from these PDX tumor models with a KRAS G12R mutation, treatment with inhibitors of CDKs (including CDK9) reduced phosphorylation of a marker of CDK9 activity (phospho-RNAPII CTD Ser2/5) and reduced viability/growth of explant cells derived from PDAC PDX models. Similarly, a CDK inhibitor reduced phospho-RNAPII CTD Ser2/5, increased apoptosis, and inhibited tumor growth in FNA-PDX and patient-matched metastatic-PDX models. In summary, PDX models can be constructed from FNA biopsies of PDAC which in turn can enable genomic characterization and identification of potential therapies.

  8. A novel synthetic compound exerts effective anti-tumour activity in vivo via the inhibition of tubulin polymerisation in A549 cells.

    PubMed

    Yan, Jun; Pang, Yanqing; Sheng, Jianfeng; Wang, Yali; Chen, Jie; Hu, Jinhui; Huang, Ling; Li, Xingshu

    2015-09-01

    Microtubules are critical elements that are involved in a wide range of cellular processes, and thus, they have become an attractive target for many anticancer drugs. A novel synthesised compound, 12P, was identified as new microtubule inhibitor. This compound inhibits tubulin polymerisation through binding to the colchicine-binding site of tubulin. 12P exhibits excellent anti-proliferative activities against a panel of human cancer cell lines, with IC₅₀ values range from 9 to 55nM. Interestingly, compound 12P also displayed equally potent cytotoxicity against several drug-resistant cell lines, and it showed high selectivity for active human umbilical vein endothelial cells (HUVECs). Further flow cytometric analysis showed that 12P induces G₂/M phase arrest and apoptosis in A549 cells. Cellular studies have revealed that the induction of apoptosis by 12P was associated with a collapse of mitochondrial membrane potential (MMP), accumulation of reactive oxygen species (ROS), alterations in the expression of some cell cycle-related proteins (e.g. Cyclin B1, Cdc25c, Cdc2) and some apoptosis-related proteins (e.g. Bax, Bad, Bcl-2, Bcl-xl). Importantly, 12P significantly reduced the growth of xenograft tumours of A549 cells in vivo (tumour inhibitory rate of 12P: 84.2%), without any loss of body weight. Taken together, these in vitro and in vivo results suggested that 12P may become a promising lead compound for the development of new anticancer drugs.

  9. Thymoquinone Inhibits Tumor Growth and Induces Apoptosis in a Breast Cancer Xenograft Mouse Model: The Role of p38 MAPK and ROS

    PubMed Central

    Woo, Chern Chiuh; Hsu, Annie; Kumar, Alan Prem; Sethi, Gautam; Tan, Kwong Huat Benny

    2013-01-01

    Due to narrow therapeutic window of cancer therapeutic agents and the development of resistance against these agents, there is a need to discover novel agents to treat breast cancer. The antitumor activities of thymoquinone (TQ), a compound isolated from Nigella sativa oil, were investigated in breast carcinoma in vitro and in vivo. Cell responses after TQ treatment were assessed by using different assays including MTT assay, annexin V-propidium iodide staining, Mitosox staining and Western blot. The antitumor effect was studied by breast tumor xenograft mouse model, and the tumor tissues were examined by histology and immunohistochemistry. The level of anti-oxidant enzymes/molecules in mouse liver tissues was measured by commercial kits. Here, we show that TQ induced p38 phosphorylation and ROS production in breast cancer cells. These inductions were found to be responsible for TQ’s anti-proliferative and pro-apoptotic effects. Moreover, TQ-induced ROS production regulated p38 phosphorylation but not vice versa. TQ treatment was found to suppress the tumor growth and this effect was further enhanced by combination with doxorubicin. TQ also inhibited the protein expression of anti-apoptotic genes, such as XIAP, survivin, Bcl-xL and Bcl-2, in breast cancer cells and breast tumor xenograft. Reduced Ki67 and increased TUNEL staining were observed in TQ-treated tumors. TQ was also found to increase the level of catalase, superoxide dismutase and glutathione in mouse liver tissues. Overall, our results demonstrated that the anti-proliferative and pro-apoptotic effects of TQ in breast cancer are mediated through p38 phosphorylation via ROS generation. PMID:24098377

  10. Nature of tumor control by permanently and transiently modified GD2 chimeric antigen receptor T cells in xenograft models of neuroblastoma.

    PubMed

    Singh, Nathan; Liu, Xiaojun; Hulitt, Jessica; Jiang, Shuguang; June, Carl H; Grupp, Stephan A; Barrett, David M; Zhao, Yangbing

    2014-11-01

    Chimeric antigen receptor (CAR) therapy has begun to demonstrate success as a novel treatment modality for hematologic malignancies. The success observed thus far has been with T cells permanently engineered to express chimeric receptors. T cells engineered using RNA electroporation represent an alternative with the potential for similar efficacy and greater safety when initially targeting novel antigens. Neuroblastoma is a common pediatric solid tumor with the potential to be targeted using immunotherapy. We performed xenograft studies in NSG mice in which we assessed the efficacy of both permanently modified and transiently modified CAR T cells directed against the neuroblastoma antigen GD2 in both local and disseminated disease models. Disease response was monitored by tumor volume measurement and histologic examination, as well as in vivo bioluminescence. RNA-modified GD2 CAR T cells mediated rapid tumor destruction when delivered locally. A single infusion of lentivirally modified GD2 CAR T cells resulted in long-term control of disseminated disease. Multiple infusions of RNA GD2 CAR T cells slowed the progression of disseminated disease and improved survival, but did not result in long-term disease control. Histologic examination revealed that the transiently modified cells were unable to significantly penetrate the tumor environment when delivered systemically, despite multiple infusions of CAR T cells. Thus, we demonstrate that RNA-modified GD2 CAR T cells can mediate effective antitumor responses in vivo, and permanently modified cells are able to control disseminated neuroblastoma in xenograft mice. Lack of long-term disease control by RNA-engineered cells resulted from an inability to penetrate the tumor microenvironment.

  11. Calculated and TLD-based absorbed dose estimates for I-131-labeled 3F8 monoclonal antibody in a human neuroblastoma xenograft nude mouse model.

    PubMed

    Ugur, O; Scott, A M; Kostakoglu, L; Hui, T E; Masterson, M E; Febo, R; Sgouros, G; Rosa, E; Mehta, B M; Fisher, D R

    1995-01-01

    Preclinical evaluation of the therapeutic potential of radiolabeled antibodies is commonly performed in a xenografted nude mouse model. To assess therapeutic efficacy it is important to estimate the absorbed dose to the tumor and normal tissues of the nude mouse. The current study was designed to accurately measure radiation does to human neuroblastoma xenografts and normal organs in nude mice treated with I-131-labeled 3F8 monoclonal antibody (MoAb) against disialoganglioside GD2 antigen. Absorbed dose estimates were obtained using two different approaches: (1) measurement with teflon-imbedded CaSO4:Dy mini-thermoluminescent dosimeters (TLDs) and (2) calculations using mouse S-factors. The calculated total dose to tumor one week after i.v. injection of the 50 microCi I-131-3F8 MoAb was 604 cGy. The corresponding decay corrected and not corrected TLD measurements were 109 +/- 9 and 48.7 +/- 3.4 cGy respectively. The calculated to TLD-derived dose ratios for tumor ranged from 6.1 at 24 h to 5.5 at 1 week. The light output fading rate was found to depend upon the tissue type within which the TLDs were implanted. The decay rate in tumor, muscle, subcutaneous tissue and in vitro, were 9.5, 5.0, 3.7 and 0.67% per day, respectively. We have demonstrated that the type of tissue in which the TLD was implanted strongly influenced the in vivo decay of light output. Even with decay correction, a significant discrepancy was observed between MIRD-based calculated and CaSO4:Dy mini-TLD measured absorbed doses. Batch dependence, pH of the tumor or other variables associated with TLDs which are not as yet well known may account for this discrepancy.

  12. Evaluation of 6-([18F] fluoroacetamido)-1-hexanoic-anilide (18F-FAHA) as imaging probe in tumor xenograft mice model

    NASA Astrophysics Data System (ADS)

    Li, Fiona; Cho, Sung Ju; Yu, Lihai; Hudson, Robert H. E.; Luyt, Leonard G.; Pin, Christopher L.; Kovacs, Michael S.; Koropatnick, James; Lee, Ting-Yim

    2016-03-01

    Alteration in genetic expression is as important as gene mutation in cancer development and proliferation. Epigenetic changes affect gene expression without altering the DNA sequence. Histone deacetylase (HDAC), an enzyme facilitating histone remodelling, can lead to silencing of tumor suppressor genes making HDAC inhibitors viable anticancer drugs against tumors with increased activity of the enzyme. In this study we evaluated 18F-fluroacetamido-1-hexanoicanilide (18F-FAHA), an artificial HDAC substrate, as imaging probe of HDAC activity of human tumor xenografts in immunocompromised host mice. Human breast and melanoma cell lines, MDA-MB-468 and MDA-MB-435 respectively, known to overexpress HDAC activity were xenografted into immunocompromised mice and HDAC activity was imaged using 18F-FAHA. The melanoma group was treated with saline, SAHA (suberoylanilide hydroxamic acid, an approved anticancer HDAC inhibitor) in DMSO, or DMSO as positive control. Tracer kinetic modelling and SUV were used to estimate HDAC activity from dynamic PET data. Both breast tumor and melanoma group showed great variability in binding rate constant (BRC) of 18F-FAHA suggesting highly variable inter- and intra-tumoral HDAC activity. For the SAHA treated melanoma group, HDAC activity, as monitored by BRC of 18F-FAHA, decreased more than the two (positive and negative) control groups but not tumor growth. Our preliminary study showed that noninvasive PET imaging with 18F-FAHA has the potential to identify patients for whom treatment with HDAC inhibitors are appropriate, to assess the effectiveness of that treatment as an early marker of target reduction, and also eliminate the need for invasive tissue biopsy to individualize treatment.

  13. 5α-reductase inhibition suppresses testosterone-induced initial regrowth of regressed xenograft prostate tumors in animal models.

    PubMed

    Masoodi, Khalid Z; Ramos Garcia, Raquel; Pascal, Laura E; Wang, Yujuan; Ma, Hei M; O'Malley, Katherine; Eisermann, Kurtis; Shevrin, Daniel H; Nguyen, Holly M; Vessella, Robert L; Nelson, Joel B; Parikh, Rahul A; Wang, Zhou

    2013-07-01

    Androgen deprivation therapy (ADT) is the standard treatment for patients with prostate-specific antigen progression after treatment for localized prostate cancer. An alternative to continuous ADT is intermittent ADT (IADT), which allows recovery of testosterone during off-cycles to stimulate regrowth and differentiation of the regressed prostate tumor. IADT offers patients a reduction in side effects associated with ADT, improved quality of life, and reduced cost with no difference in overall survival. Our previous studies showed that IADT coupled with 5α-reductase inhibitor (5ARI), which blocks testosterone conversion to DHT could prolong survival of animals bearing androgen-sensitive prostate tumors when off-cycle duration was fixed. To further investigate this clinically relevant observation, we measured the time course of testosterone-induced regrowth of regressed LuCaP35 and LNCaP xenograft tumors in the presence or absence of a 5ARI. 5α-Reductase inhibitors suppressed the initial regrowth of regressed prostate tumors. However, tumors resumed growth and were no longer responsive to 5α-reductase inhibition several days after testosterone replacement. This finding was substantiated by bromodeoxyuridine and Ki67 staining of LuCaP35 tumors, which showed inhibition of prostate tumor cell proliferation by 5ARI on day 2, but not day 14, after testosterone replacement. 5α-Reductase inhibitors also suppressed testosterone-stimulated proliferation of LNCaP cells precultured in androgen-free media, suggesting that blocking testosterone conversion to DHT can inhibit prostate tumor cell proliferation via an intracrine mechanism. These results suggest that short off-cycle coupled with 5α-reductase inhibition could maximize suppression of prostate tumor growth and, thus, improve potential survival benefit achieved in combination with IADT. PMID:23671262

  14. Anti-Tumoral Effects of Anti-Progestins in a Patient-Derived Breast Cancer Xenograft Model.

    PubMed

    Esber, Nathalie; Cherbonnier, Clément; Resche-Rigon, Michèle; Hamze, Abdallah; Alami, Mouad; Fagart, Jérôme; Loosfelt, Hugues; Lombès, Marc; Chabbert-Buffet, Nathalie

    2016-04-01

    Breast cancer is a hormone-dependent disease in which estrogen signaling targeting drugs fail in about 10 % due to resistance. Strong evidences highlighted the mitogen role of progesterone, its ligands, and the corresponding progesterone receptor (PR) isoforms in mammary carcinoma. Several PR antagonists have been synthesized; however, some of them are non-selective and led to side or toxic effects. Herein, we evaluated the anti-tumor activity of a commercially available PR modulator, ulipristal acetate (UPA), and a new selective and passive PR antagonist "APR19" in a novel preclinical approach based on patient-derived breast tumor (HBCx-34) xenografted in nude mice. As opposed to P4 that slightly reduces tumor volume, UPA and APR19 treatment for 42 days led to a significant 30 % reduction in tumor weight, accompanied by a significant 40 % retardation in tumor growth upon UPA exposure while a 1.5-fold increase in necrotic areas was observed in APR19-treated tumors. Interestingly, PR expression was upregulated by a 2.5-fold factor in UPA-treated tumors while APR19 significantly reduced expression of both PR and estrogen receptor α, indicating a potential distinct molecular mechanism among PR antagonists. Cell proliferation was clearly reduced in UPA group compared to vehicle conditions, as revealed by the significant reduction in Ki-67, Cyclin D1, and proliferating cell nuclear antigen (PCNA) expression. Likewise, an increase in activated, cleaved poly(ADP-ribose) polymerase (PARP) expression was also demonstrated upon UPA exposure. Collectively, our findings provide direct in vivo evidence for anti-progestin-mediated control of human breast cancer growth, given their anti-proliferative and pro-apoptotic activities, supporting a potential role in breast cancer therapy. PMID:26941094

  15. In vitro and in vivo antitumour activities of puerarin 6″-O-xyloside on human lung carcinoma A549 cell line via the induction of the mitochondria-mediated apoptosis pathway.

    PubMed

    Chen, Ti; Chen, Hui; Wang, Ying; Zhang, Jian

    2016-09-01

    Context Pueraria lobata (Leguminoseae) shows cytotoxic effects against cancer cells; however, its active components remain unclear. Objective This study investigated the antitumour activity of puerarin 6″-O-xyloside (POS) on the human lung carcinoma A549 cell line. Materials and methods The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was used to determine the cytotoxicity of POS (at 10, 20 and 40 μM) in vitro, and xenograft nude mice were established to evaluate the antitumour effect of POS (at 40 mg/kg/d) in vivo by 15 days intraperitoneal injection (ip). To explore its mechanism of action, flow cytometry was performed to determine the pro-apoptotic effect of POS (at 10, 20 and 40 μM). Subsequently, the expression of caspase-3, caspase-7, caspase-9, Bcl-2 and Bax in A549 cells were determined. Results POS showed significant cytotoxicity toward A549 cells (p < 0.05) by inducing apoptosis. Treatment with POS significantly upregulated the levels of caspase-3 (p < 0.01), caspase-7 (p < 0.01), caspase-9 (p < 0.01) and Bax (p < 0.01) in A549 cells, and Bcl-2 was downregulated (p < 0.01). Additionally, the in vivo animal study showed that POS significantly inhibited tumour growth in A549 cells (p < 0.01). Conclusion Our study demonstrated the POS has significant antitumour activities. The mechanisms are related to increased levels of caspase-3, caspase-7, caspase-9 and Bax, and reduced levels Bcl-2. PMID:26730946

  16. Patient-derived xenograft platform of OSCC: a renewable human bio-bank for preclinical cancer research and a new co-clinical model for treatment optimization.

    PubMed

    Sun, Shuyang; Zhang, Zhiyuan

    2016-03-01

    Advances in next-generation sequencing and bioinformatics have begun to reveal the complex genetic landscape in human cancer genomes, including oral squamous cell carcinoma (OSCC). Sophisticated preclinical models that fully represent intra- and inter-tumoral heterogeneity are required to understand the molecular diversity of cancer and achieve the goal of personalized therapies. Over the last decade, patient-derived xenograft (PDX) models generated from human tumor samples that can retain the histological and genetic features of their donor tumors have been shown to be the preferred preclinical tool in translational cancer research compared with other conventional preclinical models. Specifically, genetically well-defined PDX models can be applied to accelerate targeted antitumor drug development and biomarker discovery. Recently, we have successfully established and characterized an OSCC PDX panel as part of our tumor bio-bank for translational cancer research. In this paper, we discuss the establishment, characterization, and preclinical applications of the PDX models. In particular, we focus on the classification and applications of the PDX models based on validated annotations, including clinicopathological features, genomic profiles, and pharmacological testing information. We also explore the translational value of this well-annotated PDX panel in the development of co-clinical trials for patient stratification and treatment optimization in the near future. Although various limitations still exist, this preclinical approach should be further tested and improved.

  17. A 3D in vitro model of patient-derived prostate cancer xenograft for controlled interrogation of in vivo tumor-stromal interactions.

    PubMed

    Fong, Eliza L S; Wan, Xinhai; Yang, Jun; Morgado, Micaela; Mikos, Antonios G; Harrington, Daniel A; Navone, Nora M; Farach-Carson, Mary C

    2016-01-01

    Patient-derived xenograft (PDX) models better represent human cancer than traditional cell lines. However, the complex in vivo environment makes it challenging to employ PDX models to investigate tumor-stromal interactions, such as those that mediate prostate cancer (PCa) bone metastasis. Thus, we engineered a defined three-dimensional (3D) hydrogel system capable of supporting the co-culture of PCa PDX cells and osteoblastic cells to recapitulate the PCa-osteoblast unit within the bone metastatic microenvironment in vitro. Our 3D model not only maintained cell viability but also preserved the typical osteogenic phenotype of PCa PDX cells. Additionally, co-culture cellularity was maintained over that of either cell type cultured alone, suggesting that the PCa-osteoblast cross-talk supports PCa progression in bone, as is hypothesized to occur in patients with prostatic bone metastasis. Strikingly, osteoblastic cells co-cultured with PCa PDX tumoroids organized around the tumoroids, closely mimicking the architecture of PCa metastases in bone. Finally, tumor-stromal signaling mediated by the fibroblast growth factor axis tightly paralleled that in the in vivo counterpart. Together, these findings indicate that this 3D PCa PDX model recapitulates important pathological properties of PCa bone metastasis, and validate the use of this model for controlled and systematic interrogation of complex in vivo tumor-stromal interactions.

  18. Rapamycin enhances docetaxel-induced cytotoxicity in a androgen-independent prostate cancer xenograft model by survivin downregulation

    SciTech Connect

    Morikawa, Yasuyuki; Koike, Hidekazu; Sekine, Yoshitaka; Matsui, Hiroshi; Shibata, Yasuhiro; Ito, Kazuto; Suzuki, Kazuhiro

    2012-03-16

    Highlights: Black-Right-Pointing-Pointer Rapamycin (RPM) enhances the susceptibility of PC3 cells to docetaxel. Black-Right-Pointing-Pointer Low-dosage of docetaxel (DTX) did not reduce survivin expression levels in PC3 cells. Black-Right-Pointing-Pointer Combination treatment of RPM with DTX suppressed the expression of surviving. Black-Right-Pointing-Pointer SiRNA against survivin enhanced the susceptibility of PC3 cells to DTX. Black-Right-Pointing-Pointer RPM and DTX cotreatment inhibited PC3 cell growth and decreased surviving in vivo. -- Abstract: Background: Docetaxel is a first-line treatment choice in castration-resistant prostate cancer (CRPC). However, the management of CRPC remains an important challenge in oncology. There have been many reports on the effects of rapamycin, which is an inhibitor of the mammalian target of rapamycin (mTOR), in the treatment of carcinogenesis. We assessed the cytotoxic effects of the combination treatment of docetaxel and rapamycin in prostate cancer cells. Furthermore, we examined the relationship between these treatments and survivin, which is a member of the inhibitory apoptosis family. Methods: Prostate cancer cells were cultured and treated with docetaxel and rapamycin. The effects on proliferation were evaluated with the MTS assay. In addition, we evaluated the effect on proliferation of the combination treatment induced knockdown of survivin expression by small interfering RNA transfection and docetaxel. Protein expression levels were assayed using western blotting. PC3 cells and xenograft growth in nude mice were used to evaluate the in vivo efficacy of docetaxel and its combination with rapamycin. Results: In vitro and in vivo, the combination of rapamycin with docetaxel resulted in a greater inhibition of proliferation than treatment with rapamycin or docetaxel alone. In addition, in vitro and in vivo, rapamycin decreased basal surviving levels, and cotreatment with docetaxel further decreased these levels

  19. Longitudinal evaluation of the metabolic response of a tumor xenograft model to single fraction radiation therapy using magnetic resonance spectroscopy

    NASA Astrophysics Data System (ADS)

    Tessier, A. G.; Yahya, A.; Larocque, M. P.; Fallone, B. G.; Syme, A.

    2014-09-01

    Proton magnetic resonance spectroscopy (MRS) was used to evaluate the metabolic profile of human glioblastoma multiform brain tumors grown as xenografts in nude mice before, and at multiple time points after single fraction radiation therapy. Tumors were grown over the thigh in 16 mice in this study, of which 5 served as untreated controls and 11 had their tumors treated to 800 cGy with 200 kVp x-rays. Spectra were acquired within 24 h pre-treatment, and then at 3, 7 and 14 d post-treatment using a 9.4 T animal magnetic resonance (MR) system. For the untreated control tumors, spectra (1-2 per mouse) were acquired at different stages of tumor growth. Spectra were obtained with the PRESS pulse sequence using a 3  ×  3 × 3 mm3 voxel. Analysis was performed with the LCModel software platform. Six metabolites were profiled for this analysis: alanine (Ala), myo-inositol (Ins), taurine (Tau), creatine and phosphocreatine (Cr + PCr), glutamine and glutamate (Glu + Gln), and total choline (glycerophosphocholine + phosphocholine) (GPC + PCh). For the treated cohort, most metabolite/water concentration ratios were found to decrease in the short term at 3 and 7 d post-treatment, followed by an increase at 14 d post-treatment toward pre-treatment values. The lowest concentrations were observed at 7 d post-treatment, with magnitudes (relative to pre-treatment concentration ratios) of: 0.42  ±  24.6% (Ala), 0.43  ±  15.3% (Ins), 0.68  ±  27.9% (Tau), 0.52  ±  14.6% (GPC+PCh), 0.49  ±  21.0% (Cr + PCr) and 0.78  ±  24.5% (Glu + Gln). Control animals did not demonstrate any significant correlation between tumor volume and metabolite concentration, indicating that the observed kinetics were the result of the therapeutic intervention. We have demonstrated the feasibility of using MRS to follow multiple metabolic markers over time for the purpose of evaluating therapeutic response of tumors to radiation therapy. This study provides

  20. Long Term Culture of the A549 Cancer Cell Line Promotes Multilamellar Body Formation and Differentiation towards an Alveolar Type II Pneumocyte Phenotype

    PubMed Central

    Cooper, James Ross; Abdullatif, Muhammad Bilal; Burnett, Edward C.; Kempsell, Karen E.; Conforti, Franco; Tolley, Howard; Collins, Jane E.; Davies, Donna E.

    2016-01-01

    Pulmonary research requires models that represent the physiology of alveolar epithelium but concerns with reproducibility, consistency and the technical and ethical challenges of using primary or stem cells has resulted in widespread use of continuous cancer or other immortalized cell lines. The A549 ‘alveolar’ cell line has been available for over four decades but there is an inconsistent view as to its suitability as an appropriate model for primary alveolar type II (ATII) cells. Since most work with A549 cells involves short term culture of proliferating cells, we postulated that culture conditions that reduced proliferation of the cancer cells would promote a more differentiated ATII cell phenotype. We examined A549 cell growth in different media over long term culture and then used microarray analysis to investigate temporal regulation of pathways involved in cell cycle and ATII differentiation; we also made comparisons with gene expression in freshly isolated human ATII cells. Analyses indicated that long term culture in Ham’s F12 resulted in substantial modulation of cell cycle genes to result in a quiescent population of cells with significant up-regulation of autophagic, differentiation and lipidogenic pathways. There were also increased numbers of up- and down-regulated genes shared with primary cells suggesting adoption of ATII characteristics and multilamellar body (MLB) development. Subsequent Oil Red-O staining and Transmission Electron Microscopy confirmed MLB expression in the differentiated A549 cells. This work defines a set of conditions for promoting ATII differentiation characteristics in A549 cells that may be advantageous for studies with this cell line. PMID:27792742

  1. Study of gaseous benzene effects upon A549 lung epithelial cells using a novel exposure system.

    PubMed

    Mascelloni, Massimiliano; Delgado-Saborit, Juana Maria; Hodges, Nikolas J; Harrison, Roy M

    2015-08-19

    Volatile organic compounds (VOCs) are ubiquitous pollutants known to be present in both indoor and outdoor air arising from various sources. Indoor exposure has increasingly become a major cause of concern due to the effects that such pollutants can have on health. Benzene, along with toluene, is one of the main components of the VOC mixture and is a known carcinogen due to its genotoxic effects. The aim of this study was to test the feasibility of an in vitro model to study the short-term effects of exposure of lung cells to airborne benzene. We studied the effects of exposure on DNA and the production of reactive oxygen species (ROS) in A549 cells, exposed to various concentrations of benzene (0.03; 0.1; 0.3 ppm) in gaseous form using a custom designed cell exposure chamber. Results showed a concentration-dependent increase of DNA breaks and an increase of ROS production, confirming the feasibility of the experimental procedure and validating the model for further in vitro studies of exposure to other VOCs.

  2. Methyl methanesulfonate induces necroptosis in human lung adenoma A549 cells through the PIG-3-reactive oxygen species pathway.

    PubMed

    Jiang, Ying; Shan, Shigang; Chi, Linfeng; Zhang, Guanglin; Gao, Xiangjing; Li, Hongjuan; Zhu, Xinqiang; Yang, Jun

    2016-03-01

    Methyl methanesulfonate (MMS) is an alkylating agent that can induce cell death through apoptosis and necroptosis. The molecular mechanisms underlying MMS-induced apoptosis have been studied extensively; however, little is known about the mechanism for MMS-induced necroptosis. Therefore, we first established MMS-induced necroptosis model using human lung carcinoma A549 cells. It was found that, within a 24-h period, although MMS at concentrations of 50, 100, 200, 400, and 800 μM can induce DNA damage, only at higher concentrations (400 and 800 μM) MMS treatment lead to necroptosis in A549 cells, as it could be inhibited by the specific necroptotic inhibitor necrostatin-1, but not the specific apoptotic inhibitor carbobenzoxy-valyl-alanyl-aspartyl-[O-methyl]-fluoromethylketone (Z-VAD-fmk). MMS-induced necroptosis was further confirmed by the induction of the necroptosis biomarkers including the depletion of cellular NADH and ATP and leakage of LDH. This necroptotic cell death was also concurrent with the increased expression of p53, p53-induced gene 3 (PIG-3), high mobility group box-1 protein (HMGB1), and receptor interaction protein kinase (RIP) but not the apoptosis-associated caspase-3 and caspase-9 proteins. Elevated reactive oxygen species (ROS) level was also involved in this process as the specific ROS inhibitor (4-amino-2,4-pyrrolidine-dicarboxylic acid (APDC)) can inhibit the necroptotic cell death. Interestingly, knockdown of PIG-3 expression by small interfering RNA (siRNA) treatment can inhibit the generation of ROS. Taken together, these results suggest that MMS can induce necroptosis in A549 cells, probably through the PIG-3-ROS pathway.

  3. Radix Tetrastigma hemsleyani flavone inhibits proliferation, migration, and invasion of human lung carcinoma A549 cells

    PubMed Central

    Zhong, Liangrui; Zheng, Junxian; Sun, Qianqian; Wei, Kemin; Hu, Yijuan

    2016-01-01

    Radix Tetrastigma hemsleyani flavone (RTHF) is widely used as a traditional herb and has detoxification and anti-inflammatory effects. In this study, we investigated the potential effects of RTHF on the growth and metastasis of human lung adenocarcinoma A549 cells and evaluated its mechanisms. A549 cells were treated with RTHF at various concentrations for different periods. In vitro Cell Counting Kit-8 assay and colony formation methods showed that RTHF had dose- and time-dependent antiproliferation effects on A549 cells. A cell adhesion assay showed that RTHF decreased A549 cell adhesion in a dose-dependent manner. Cell invasion and migration were investigated using the Transwell assay and observed using an inverted microscope; the results showed that cell metastasis was significantly lower in the treatment group than that in the control group (P<0.01). Expression of metastasis-related matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) was detected by real-time polymerase chain reaction and Western blotting. The results showed that the expression of MMP-2, MMP-9, and TIMP-1 decreased, while that of TIMP-2 increased significantly in the RTHF group when compared with the results of the control group. These results show that RTHF exhibits antigrowth and antimetastasis activity in lung cancer A549 cells by decreasing the expression of MMP-2/-9 and TIMP-1 and increasing that of TIMP-2. PMID:26893573

  4. Combined Inhibition of Cyclin-Dependent Kinases (Dinaciclib) and AKT (MK-2206) Blocks Pancreatic Tumor Growth and Metastases in Patient-Derived Xenograft Models.

    PubMed

    Hu, Chaoxin; Dadon, Tikva; Chenna, Venugopal; Yabuuchi, Shinichi; Bannerji, Rajat; Booher, Robert; Strack, Peter; Azad, Nilofer; Nelkin, Barry D; Maitra, Anirban

    2015-07-01

    KRAS is activated by mutation in the vast majority of cases of pancreatic cancer; unfortunately, therapeutic attempts to inhibit KRAS directly have been unsuccessful. Our previous studies showed that inhibition of cyclin-dependent kinase 5 (CDK5) reduces pancreatic cancer growth and progression, through blockage of the centrally important RAL effector pathway, downstream of KRAS. In the current study, the therapeutic effects of combining the CDK inhibitor dinaciclib (SCH727965; MK-7965) with the pan-AKT inhibitor MK-2206 were evaluated using orthotopic and subcutaneous patient-derived human pancreatic cancer xenograft models. The combination of dinaciclib (20 mg/kg, i.p., three times a week) and MK-2206 (60 mg/kg, orally, three times a week) dramatically blocked tumor growth and metastasis in all eight pancreatic cancer models examined. Remarkably, several complete responses were induced by the combination treatment of dinaciclib and MK-2206. The striking results obtained in these models demonstrate that the combination of dinaciclib with the pan-AKT inhibitor MK-2206 is promising for therapeutic evaluation in pancreatic cancer, and strongly suggest that blocking RAL in combination with other effector pathways downstream from KRAS may provide increased efficacy in pancreatic cancer. Based on these data, an NCI-CTEP-approved multicenter phase I clinical trial for pancreatic cancer of the combination of dinaciclib and MK-2206 (NCT01783171) has now been opened. PMID:25931518

  5. Frankincense essential oil prepared from hydrodistillation of Boswellia sacra gum resins induces human pancreatic cancer cell death in cultures and in a xenograft murine model

    PubMed Central

    2012-01-01

    Background Regardless of the availability of therapeutic options, the overall 5-year survival for patients diagnosed with pancreatic cancer remains less than 5%. Gum resins from Boswellia species, also known as frankincense, have been used as a major ingredient in Ayurvedic and Chinese medicine to treat a variety of health-related conditions. Both frankincense chemical extracts and essential oil prepared from Boswellia species gum resins exhibit anti-neoplastic activity, and have been investigated as potential anti-cancer agents. The goals of this study are to identify optimal condition for preparing frankincense essential oil that possesses potent anti-tumor activity, and to evaluate the activity in both cultured human pancreatic cancer cells and a xenograft mouse cancer model. Methods Boswellia sacra gum resins were hydrodistilled at 78°C; and essential oil distillate fractions were collected at different durations (Fraction I at 0–2 h, Fraction II at 8–10 h, and Fraction III at 11–12 h). Hydrodistillation of the second half of gum resins was performed at 100°C; and distillate was collected at 11–12 h (Fraction IV). Chemical compositions were identified by gas chromatography–mass spectrometry (GC-MS); and total boswellic acids contents were quantified by high-performance liquid chromatography (HPLC). Frankincense essential oil-modulated pancreatic tumor cell viability and cytotoxicity were determined by colorimetric assays. Levels of apoptotic markers, signaling molecules, and cell cycle regulators expression were characterized by Western blot analysis. A heterotopic (subcutaneous) human pancreatic cancer xenograft nude mouse model was used to evaluate anti-tumor capability of Fraction IV frankincense essential oil in vivo. Frankincense essential oil-induced tumor cytostatic and cytotoxic activities in animals were assessed by immunohistochemistry. Results Longer duration and higher temperature hydrodistillation produced more abundant high molecular

  6. Human lung epithelial cell A549 proteome data after treatment with titanium dioxide and carbon black.

    PubMed

    Vuong, Ngoc Q; Goegan, Patrick; Mohottalage, Susantha; Breznan, Dalibor; Ariganello, Marianne; Williams, Andrew; Elisma, Fred; Karthikeyan, Subramanian; Vincent, Renaud; Kumarathasan, Premkumari

    2016-09-01

    Here, we have described the dataset relevant to the A549 cellular proteome changes after exposure to either titanium dioxide or carbon black particles as compared to the non-exposed controls, "Proteomic changes in human lung epithelial cells (A549) in response to carbon black and titanium dioxide exposures" (Vuong et al., 2016) [1]. Detailed methodologies on the separation of cellular proteins by 2D-GE and the subsequent mass spectrometry analyses using MALDI-TOF-TOF-MS are documented. Particle exposure-specific protein expression changes were measured via 2D-GE spot volume analysis. Protein identification was done by querying mass spectrometry data against SwissProt and RefSeq protein databases using Mascot search engine. Two-way ANOVA analysis data provided information on statistically significant A549 protein expression changes associated with particle exposures. PMID:27508218

  7. Human lung epithelial cell A549 proteome data after treatment with titanium dioxide and carbon black.

    PubMed

    Vuong, Ngoc Q; Goegan, Patrick; Mohottalage, Susantha; Breznan, Dalibor; Ariganello, Marianne; Williams, Andrew; Elisma, Fred; Karthikeyan, Subramanian; Vincent, Renaud; Kumarathasan, Premkumari

    2016-09-01

    Here, we have described the dataset relevant to the A549 cellular proteome changes after exposure to either titanium dioxide or carbon black particles as compared to the non-exposed controls, "Proteomic changes in human lung epithelial cells (A549) in response to carbon black and titanium dioxide exposures" (Vuong et al., 2016) [1]. Detailed methodologies on the separation of cellular proteins by 2D-GE and the subsequent mass spectrometry analyses using MALDI-TOF-TOF-MS are documented. Particle exposure-specific protein expression changes were measured via 2D-GE spot volume analysis. Protein identification was done by querying mass spectrometry data against SwissProt and RefSeq protein databases using Mascot search engine. Two-way ANOVA analysis data provided information on statistically significant A549 protein expression changes associated with particle exposures.

  8. Xylitol induces cell death in lung cancer A549 cells by autophagy.

    PubMed

    Park, Eunjoo; Park, Mi Hee; Na, Hee Sam; Chung, Jin

    2015-05-01

    Xylitol is a widely used anti-caries agent that has anti-inflammatory effects. We have evaluated the potential of xylitol in cancer treatment. It's effects on cell proliferation and cytotoxicity were measured by MTT assay and LDH assay. Cell morphology and autophagy were examined by immunostaining and immunoblotting. Xylitol inhibited cell proliferation in a dose-dependent manner in these cancer cells: A549, Caki, NCI-H23, HCT-15, HL-60, K562, and SK MEL-2. The IC50 of xylitol in human gingival fibroblast cells was higher than in cancer cells, indicating that it is more specific for cancer cells. Moreover, xylitol induced autophagy in A549 cells that was inhibited by 3-methyladenine, an autophagy inhibitor. These results indicate that xylitol has potential in therapy against lung cancer by inhibiting cell proliferation and inducing autophagy of A549 cells.

  9. Ursolic acid and oleanolic acid from Eriobotrya fragrans inhibited the viability of A549 cells.

    PubMed

    Yuan, Yuan; Gao, Yongshun; Song, Gang; Lin, Shunquan

    2015-02-01

    Loquat {Eriobotrya japonica (Lindl.)}, a kind of Chinese herb, has many efficacies such as anti-inflammatory, antimicrobial and curing chronic bronchitis. However, reports on the pharmacological action of wild loquat extract are limited. In this work, the A549 cell line was selected to study the inhibitory effect of ursolic acid and oleanolic acid (UA, OA) from the leaves of E. fragrans. Results showed that UA/OA inhibited A549 cell viability and induced apoptosis in a dose and time dependent manner. The cell fraction in the G0/G1 phase dramatically increased under treatment with UA/OA. Data showed that UA activated the expression of PARP. UA and OA down-regulated MMP-2 and Bcl-2; on the contrary, they up-regulated Bid. This work demonstrated that UA/OA extracted from wild loquat leaves can significantly inhibit the viability of A549 cells.

  10. Aptamer based electrochemical sensor for detection of human lung adenocarcinoma A549 cells

    NASA Astrophysics Data System (ADS)

    Sharma, Rachna; Varun Agrawal, Ved; Sharma, Pradeep; Varshney, R.; Sinha, R. K.; Malhotra, B. D.

    2012-04-01

    We report results of the studies relating to development of an aptamer-based electrochemical biosensor for detection of human lung adenocarcinoma A549 cells. The aminated 85-mer DNA aptamer probe specific for the A549 cells has been covalently immobilized onto silane self assembled monolayer (SAM) onto ITO surface using glutaraldehyde as the crosslinker. The results of cyclic voltammetry and differential pulse voltammetry studies reveal that the aptamer functionalized bioelectrode can specifically detect lung cancer cells in the concentration range of 103 to 107 cells/ml with detection limit of 103 cells/ml within 60 s. The specificity studies of the bioelectrode have been carried out with control KB cells. No significant change in response is observed for control KB cells as compared to that of the A549 target cells.

  11. Inhibition of human tumor xenograft growth in nude mice by a conjugate of monoclonal antibody LA22 to epidermal growth factor receptor with anti-tumor antibiotics mitomycin C

    SciTech Connect

    Shao Wei; Zhao Shan; Liu Zhaofei; Zhang Jianzhong; Ma Shujun; Sato, J. Denry; Zhang Peng; Tong Mei; Han Jiping; Wang Yan; Bai Dongmei; Wang Fan . E-mail: wangfan@bjmu.edu.cn; Sun Le . E-mail: lsun@welsonpharma.com

    2006-10-20

    Anti-EGFR monoclonal antibodies LA22 and Erbitux bind to different epitopes of EGFR. The chemimmunoconjugates of MMC with LA22 or Erbitux were prepared, and in vitro cytotoxicity assays with A549 cells showed that LA22-MMC was much more potent than Erbitux or Erbitux-MMC. Viabilities of A549 cells treated with LA22-MMC, Erbitux or Erbitux-MMC were 35%, 94%, and 81%, respectively. Immunoscintigraphy of xenografts of human A431 and A549 cells in nude mice both showed that {sup 125}I-labeled-LA22-MMC enriched in tumor sites prominently. Most importantly, in vivo assays showed LA22-MMC was significantly more effective than free drug MMC in the treatment of subcutaneous xenografts of human A431 cells in nude mice (83% inhibition for LA22-MMC and 30% for MMC). We concluded that LA22-MMC could be a very potent drug for treatment of solid tumors.

  12. AZ17: a new bispecific drug targeting IL-6 and IL-23 with potential clinical use--improves psoriasis in a human xenograft transplantation model.

    PubMed

    Stenderup, Karin; Rosada, Cecilia; Shanebeck, Kurt; Brady, William; Van Brunt, Michael P; King, Gordon; Marelli, Marcello; Slagle, Paul; Xu, Hengyu; Nairn, Natalie W; Johnson, Jeffrey; Wang, Aijun A; Li, Gary; Thornton, Kenneth C; Dam, Tomas N; Grabstein, Kenneth H

    2015-10-01

    Targeting more than one molecule in multifactorial diseases involving several disease mediators may provide improved therapeutic efficacy. Psoriasis is a multifactorial disease in which interleukin (IL)-6 and IL-23 are important disease mediators because they facilitate development of Th17 cells; widely accepted to be associated with psoriasis. To meet the need for new therapeutics, we aimed to create a clinically relevant bispecific drug, by combining the inhibitory properties of anti-IL-6 and anti-IL-23 antibodies, exhibiting high affinity, high stability and the ability to be produced in high yield. The bispecific molecule AZ17 was created by combining high affinity binding domains originating from monoclonal antibodies targeting human IL-6 and IL-23. To allow for high and efficient production, AZ17 was assembled by site-specific bioconjugation from two individual single chain fragment variables that were synthesized separately in Escherichia coli. To improve stability and extend pharmacokinetics, a flexible poly-ethylene glycol molecule was used as linker. In preclinical psoriasis models, AZ17 reduced IL-23-induced ear inflammation and improved psoriasis in a xenograft transplantation model where psoriasis skin is transplanted onto immune-deficient mice. The data presented here suggest AZ17 to be a promising drug candidate in psoriasis and other inflammatory diseases associated with Th17 cell development.

  13. 3'-hydroxy-3,4,5,4'-tetramethoxystilbene, the metabolite of resveratrol analogue DMU-212, inhibits ovarian cancer cell growth in vitro and in a mice xenograft model.

    PubMed

    Piotrowska-Kempisty, Hanna; Ruciński, Marcin; Borys, Sylwia; Kucińska, Małgorzata; Kaczmarek, Mariusz; Zawierucha, Piotr; Wierzchowski, Marcin; Łażewski, Dawid; Murias, Marek; Jodynis-Liebert, Jadwiga

    2016-01-01

    In screening studies, the cytotoxic activity of four metabolites of resveratrol analogue 3,4,5,4'-tetramethoxystilbene (DMU-212) against A-2780 and SKOV-3 ovarian cancer cells was investigated. The most active metabolite, 3'-hydroxy-3,4,5,4'-tetramethoxystilbene (DMU-214), was chosen for further studies. The cytotoxicity of DMU-214 was shown to be higher than that of the parent compound, DMU-212, in both cell lines tested. Since DMU-212 was supposed to undergo metabolic activation through its conversion to DMU-214, an attempt was made to elucidate the mechanism of its anti-proliferative activity. We found that in SKOV-3 cells lacking p53, DMU-214 induced receptor-mediated apoptosis. In A-2780 cell line with expression of wild-type p53, DMU-214 modulated the expression pattern of p53-target genes driving intrinsic and extrinsic apoptosis pathways, as well as DNA repair and damage prevention. Regardless of the up-regulation of p48, p53R2, sestrins and Gaad45 genes involved in cancer cell DNA repair, we demonstrated the stronger anti-proliferative and pro-apoptotic effects of DMU-214 in A-2780 cells when compared to those in SKOV-3. Hence we verified DMU-214 activity in the xenograft model using SCID mice injected with A-2780 cells. The strong anti-proliferative activity of DMU-214 in the in vivo model allowed to suggest the tested compound as a potential therapeutic in ovarian cancer treatment. PMID:27585955

  14. A novel type of cellular senescence that can be enhanced in mouse models and human tumor xenografts to suppress prostate tumorigenesis

    PubMed Central

    Alimonti, Andrea; Nardella, Caterina; Chen, Zhenbang; Clohessy, John G.; Carracedo, Arkaitz; Trotman, Lloyd C.; Cheng, Ke; Varmeh, Shohreh; Kozma, Sara C.; Thomas, George; Rosivatz, Erika; Woscholski, Rudiger; Cognetti, Francesco; Scher, Howard I.; Pandolfi, Pier Paolo

    2010-01-01

    Irreversible cell growth arrest, a process termed cellular senescence, is emerging as an intrinsic tumor suppressive mechanism. Oncogene-induced senescence is thought to be invariably preceded by hyperproliferation, aberrant replication, and activation of a DNA damage checkpoint response (DDR), rendering therapeutic enhancement of this process unsuitable for cancer treatment. We previously demonstrated in a mouse model of prostate cancer that inactivation of the tumor suppressor phosphatase and tensin homolog deleted on chromosome 10 (Pten) elicits a senescence response that opposes tumorigenesis. Here, we show that Pten-loss–induced cellular senescence (PICS) represents a senescence response that is distinct from oncogene-induced senescence and can be targeted for cancer therapy. Using mouse embryonic fibroblasts, we determined that PICS occurs rapidly after Pten inactivation, in the absence of cellular proliferation and DDR. Further, we found that PICS is associated with enhanced p53 translation. Consistent with these data, we showed that in mice p53-stabilizing drugs potentiated PICS and its tumor suppressive potential. Importantly, we demonstrated that pharmacological inhibition of PTEN drives senescence and inhibits tumorigenesis in vivo in a human xenograft model of prostate cancer. Taken together, our data identify a type of cellular senescence that can be triggered in nonproliferating cells in the absence of DNA damage, which we believe will be useful for developing a “pro-senescence” approach for cancer prevention and therapy. PMID:20197621

  15. Induction of p53-independent growth inhibition in lung carcinoma cell A549 by gypenosides.

    PubMed

    Liu, Jung-Sen; Chiang, Tzu-Hsuan; Wang, Jinn-Shyan; Lin, Li-Ju; Chao, Wei-Chih; Inbaraj, Baskaran Stephen; Lu, Jyh-Feng; Chen, Bing-Huei

    2015-07-01

    The objectives of this study are to investigate antiproliferative effect and mechanisms of bioactive compounds from Gynostemma pentaphyllum (G. pentaphyllum) on lung carcinoma cell A549. Saponins, carotenoids and chlorophylls were extracted and fractionated by column chromatography, and were subjected to high-performance liquid chromatography-mass spectrometry analyses. The saponin fraction, which consisted mainly of gypenoside (Gyp) XXII and XXIII, rather than the carotenoid and chlorophyll ones, was effective in inhibiting A549 cell growth in a concentration- and a time-dependent manner as evaluated using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The estimated half maximal inhibitory concentration (IC50 ) of Gyp on A549 cells was 30.6 μg/ml. Gyp was further demonstrated to induce an apparent arrest of the A549 cell cycle at both the S phase and the G2/M phase, accompanied by a concentration- and a time-dependent increase in the proportions of both the early and late apoptotic cells. Furthermore, Gyp down-regulated cellular expression of cyclin A and B as well as BCL-2, while up-regulated the expression of BAX, DNA degradation factor 35 KD, poly [ADP-ribose] polymerase 1, p53, p21 and caspase-3. Nevertheless, both the treatment of a p53 inhibitor, pifithrin-α, and the small hairpin RNA-mediated p53 knockdown in the A549 cells did not alter the growth inhibition effect induced by Gyp. As a result, the cell cycle arrest and apoptosis of A549 cells induced by Gyp would most likely proceed through p53-independent pathway(s). PMID:25781909

  16. Impact of bevacizumab in combination with erlotinib on EGFR-mutated non-small cell lung cancer xenograft models with T790M mutation or MET amplification.

    PubMed

    Furugaki, Koh; Fukumura, Junko; Iwai, Toshiki; Yorozu, Keigo; Kurasawa, Mitsue; Yanagisawa, Mieko; Moriya, Yoichiro; Yamamoto, Kaname; Suda, Kenichi; Mizuuchi, Hiroshi; Mitsudomi, Tetsuya; Harada, Naoki

    2016-02-15

    Erlotinib (ERL), an epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor, shows notable efficacy against non-small cell lung cancer (NSCLC) harboring EGFR mutations. Bevacizumab (BEV), a humanized monoclonal antibody to vascular endothelial cell growth factor (VEGF), in combination with ERL (BEV+ERL) significantly extended progression-free survival in patients with EGFR-mutated NSCLC compared with ERL alone. However, the efficacy of BEV+ERL against EGFR-mutated NSCLC harboring T790M mutation or MET amplification, is unclear. Here, we examined the antitumor activity of BEV+ERL in four xenograft models of EGFR-mutated NSCLC (three harboring ERL resistance mutations). In the HCC827 models (exon 19 deletion: DEL), ERL significantly inhibited tumor growth by blocking EGFR signal transduction. Although there was no difference between ERL and BEV+ERL in maximum tumor growth inhibition, BEV+ERL significantly suppressed tumor regrowth during a drug-cessation period. In the HCC827-EPR model (DEL+T790M) and HCC827-vTR model (DEL+MET amplification), ERL reduced EGFR signal transduction and showed less pronounced but still significant tumor growth inhibition than in the HCC827 model. In these models, tumor growth inhibition was significantly stronger with BEV+ERL than with each single agent. In the NCI-H1975 model (L858R+T790M), ERL did not inhibit growth or EGFR signal transduction, and BEV+ERL did not inhibit growth more than BEV. BEV alone significantly decreased microvessel density in each tumor. In conclusion, addition of BEV to ERL did not enhance antitumor activity in primarily ERL-resistant tumors with T790M mutation; however, BEV+ERL enhanced antitumor activity in T790M mutation- or MET amplification-positive tumors as long as their growth remained significantly suppressed by ERL.

  17. Alkylator-Induced and Patient-Derived Xenograft Mouse Models of Therapy-Related Myeloid Neoplasms Model Clinical Disease and Suggest the Presence of Multiple Cell Subpopulations with Leukemia Stem Cell Activity.

    PubMed

    Jonas, Brian A; Johnson, Carl; Gratzinger, Dita; Majeti, Ravindra

    2016-01-01

    Acute myeloid leukemia (AML) is a heterogeneous group of aggressive bone marrow cancers arising from transformed hematopoietic stem and progenitor cells (HSPC). Therapy-related AML and MDS (t-AML/MDS) comprise a subset of AML cases occurring after exposure to alkylating chemotherapy and/or radiation and are associated with a very poor prognosis. Less is known about the pathogenesis and disease-initiating/leukemia stem cell (LSC) subpopulations of t-AML/MDS compared to their de novo counterparts. Here, we report the development of mouse models of t-AML/MDS. First, we modeled alkylator-induced t-AML/MDS by exposing wild type adult mice to N-ethyl-N-nitrosurea (ENU), resulting in several models of AML and MDS that have clinical and pathologic characteristics consistent with human t-AML/MDS including cytopenia, myelodysplasia, and shortened overall survival. These models were limited by their inability to transplant clinically aggressive disease. Second, we established three patient-derived xenograft models of human t-AML. These models led to rapidly fatal disease in recipient immunodeficient xenografted mice. LSC activity was identified in multiple HSPC subpopulations suggesting there is no canonical LSC immunophenotype in human t-AML. Overall, we report several new t-AML/MDS mouse models that could potentially be used to further define disease pathogenesis and test novel therapeutics. PMID:27428079

  18. Alkylator-Induced and Patient-Derived Xenograft Mouse Models of Therapy-Related Myeloid Neoplasms Model Clinical Disease and Suggest the Presence of Multiple Cell Subpopulations with Leukemia Stem Cell Activity

    PubMed Central

    Johnson, Carl; Gratzinger, Dita; Majeti, Ravindra

    2016-01-01

    Acute myeloid leukemia (AML) is a heterogeneous group of aggressive bone marrow cancers arising from transformed hematopoietic stem and progenitor cells (HSPC). Therapy-related AML and MDS (t-AML/MDS) comprise a subset of AML cases occurring after exposure to alkylating chemotherapy and/or radiation and are associated with a very poor prognosis. Less is known about the pathogenesis and disease-initiating/leukemia stem cell (LSC) subpopulations of t-AML/MDS compared to their de novo counterparts. Here, we report the development of mouse models of t-AML/MDS. First, we modeled alkylator-induced t-AML/MDS by exposing wild type adult mice to N-ethyl-N-nitrosurea (ENU), resulting in several models of AML and MDS that have clinical and pathologic characteristics consistent with human t-AML/MDS including cytopenia, myelodysplasia, and shortened overall survival. These models were limited by their inability to transplant clinically aggressive disease. Second, we established three patient-derived xenograft models of human t-AML. These models led to rapidly fatal disease in recipient immunodeficient xenografted mice. LSC activity was identified in multiple HSPC subpopulations suggesting there is no canonical LSC immunophenotype in human t-AML. Overall, we report several new t-AML/MDS mouse models that could potentially be used to further define disease pathogenesis and test novel therapeutics. PMID:27428079

  19. KRAS and PIK3CA mutation frequencies in patient-derived xenograft models of pancreatic and colorectal cancer are reflective of patient tumors and stable across passages.

    PubMed

    Tignanelli, Christopher J; Herrera Loeza, Silvia G; Yeh, Jen Jen

    2014-09-01

    One obstacle in the translation of advances in cancer research into the clinic is a deficiency of adequate preclinical models that recapitulate human disease. Patient-derived xenograft (PDX) models are established by engrafting patient tumor tissue into mice and are advantageous because they capture tumor heterogeneity. One concern with these models is that selective pressure could lead to mutational drift and thus be an inaccurate reflection of patient tumors. Therefore, we evaluated if mutational frequency in PDX models is reflective of patient populations and if crucial mutations are stable across passages. We examined KRAS and PIK3CA gene mutations from pancreatic ductal adenocarcinoma (PDAC) (n = 30) and colorectal cancer (CRC) (n = 37) PDXs for as many as eight passages. DNA was isolated from tumors and target sequences were amplified by polymerase chain reaction. KRAS codons 12/13 and PIK3CA codons 542/545/1047 were examined using pyrosequencing. Twenty-three of 30 (77%) PDAC PDXs had KRAS mutations and one of 30 (3%) had PIK3CA mutations. Fifteen of 37 (41%) CRC PDXs had KRAS mutations and three of 37 (8%) had PIK3CA mutations. Mutations were 100 per cent preserved across passages. We found that the frequency of KRAS (77%) and PIK3CA (3%) mutations in PDAC PDX was similar to frequencies in patient tumors (71 to 100% KRAS, 0 to 11% PIK3CA). Similarly, KRAS (41%) and PIK3CA (8%) mutations in CRC PDX closely paralleled patient tumors (35 to 51% KRAS, 12 to 21% PIK3CA). The accurate mirroring and stability of genetic changes in PDX models compared with patient tumors suggest that these models are good preclinical surrogates for patient tumors.

  20. Role of WNT1-inducible-signaling pathway protein 1 in etoposide resistance in lung adenocarcinoma A549 cells

    PubMed Central

    Xu, Yunhua; Lu, Shun

    2015-01-01

    Object: The aim of this study was to explore the role of WNT1-inducible-signaling Pathway Protein 1 (WISP-1) in etoposide resistance in lung adenocarcinoma A549 cells. Methods: WISP-1 overexpression A549 lung adenocarcinoma cell was established. After exposure to ultraviolet (UV) and etoposide, cell viability and apoptosis were evaluated. Moreover, western-blot was employed to examine the expression of apoptotic pathway proteins. In addition, a nude mice tumor model was established to examine the effect of WISP-1 overexpression in vivo and TUNEL staining was used to assess cell apoptosis of tumor tissue. Results: WISP-1 overexpression significantly increased cell viability and decreased cell apoptosis after treatment with UV and etoposide. Decreased expression of Bad and Bax and increased expression of Bcl-2 was found after etoposide treatment in WISP-1 overexpressed cells. A significantly increasing of tumor volume in WISP-1 overexpressed group was found and TUNEL staining revealed that decreased cell apoptosis in WISP-1 overexpressed group. Conclusion: Our results demonstrated that WISP-1 may have a facilitating role in etoposide resistance through increasing cell viability and decreasing cell apoptosis. PMID:26628978

  1. Generation and characterization of bioluminescent xenograft mouse models of MLL-related acute leukemias and in vivo evaluation of luciferase-targeting siRNA nanoparticles.

    PubMed

    Fazzina, Raffaella; Lombardini, Lorenza; Mezzanotte, Laura; Roda, Aldo; Hrelia, Patrizia; Pession, Andrea; Tonelli, Roberto

    2012-08-01

    Chromosomal translocations involving the MLL gene on 11q23 present frequent abnormalities in pediatric, adult and therapy-related acute leukemias, and are generally associated with aggressive disease and poor prognosis. Here, we report bioluminescent acute leukemia xenograft mouse models of the most frequent and aggressive MLL-related acute leukemias (infant and adult MLL-AF9, MLL-ENL, MLL-AF4). Four acute leukemia cell lines carrying MLL-related translocations were stably transduced with a firefly luciferase transgene and injected intravenously into NOD/SCID mice. Leukemia progression was monitored by in vivo bioluminescence imaging (BLI). All mice developed MLL-related acute leukemia. The four MLL-related acute leukemia models showed a different course of infant and adult MLL-AF9 acute myeloid leukemia, and a rapid aggressiveness of MLL-ENL acute lymphoblastic leukemia and MLL-AF4 acute biphenotypic leukemia. Tissue analysis and RT-PCR of bone marrow, spleen and liver from the mice confirmed the BL results. To validate BLI for the detection of a therapeutic response, systemic treatment with an anti-luciferase-targeting siRNA (siLuc) complexed with cationic nanoparticles was administered to mice with MLL-AF4 acute lymphoblastic leukemia. The BLI signal showed a reduction following treatment with siLuc compared to the control mice. These mouse models present MLL-related acute leukemia evolution similar to the human counterparts. Moreover, they are non-invasive, rapid and sensitive models, suitable for the in vivo study of MLL-related acute leukemias. Finally, BLI showed in vivo luminescence down modulation obtained by systemic treatment with luciferase-targeting siRNA nanoparticle complexes, confirming that these MLL-related leukemia mouse models are optimal for the evaluation and selection of delivery systems for siRNA and other new biotechnological pharmaceuticals.

  2. Selective small molecule Stat3 inhibitor reduces breast cancer tumor-initiating cells and improves recurrence free survival in a human-xenograft model.

    PubMed

    Dave, Bhuvanesh; Landis, Melissa D; Tweardy, David J; Chang, Jenny C; Dobrolecki, Lacey E; Wu, Meng-Fen; Zhang, Xiaomei; Westbrook, Thomas F; Hilsenbeck, Susan G; Liu, Dan; Lewis, Michael T

    2012-01-01

    Metastasis and disease relapse are hypothesized to result from tumor initiating cells (TICs). Previously, we have defined a CD44+/CD24-/low mammosphere-forming tumorigenic 493-gene signature in breast cancer. Stat3 was identified as a critical node in self-renewal based on an ongoing lentiviral shRNA screen being conducted in two breast cancer cell lines SUM159 and BT549. In corroborating work, targeting the SH2 domain of Stat3 with a novel small molecule decreased the percentage of cells expressing TIC markers (CD44+/CD24-/low and ALDH+) and mammosphere formation in p-Stat3 overexpressing human breast cancer xenografts in SCID-beige mice. Importantly, we observed a four-fold improvement in the 30-day recurrence-free survival relative to docetaxel alone with the addition of the Stat3 inhibitor in the chemoresistant tumor model. Thus, these findings provide a strong impetus for the development of selective Stat3 inhibitors in order to improve survival in patients with p-Stat3 overexpressing tumors. PMID:22879872

  3. Transforming growth factor-beta can suppress tumorigenesis through effects on the putative cancer stem or early progenitor cell and committed progeny in a breast cancer xenograft model.

    PubMed

    Tang, Binwu; Yoo, Naomi; Vu, Mary; Mamura, Mizuko; Nam, Jeong-Seok; Ooshima, Akira; Du, Zhijun; Desprez, Pierre-Yves; Anver, Miriam R; Michalowska, Aleksandra M; Shih, Joanna; Parks, W Tony; Wakefield, Lalage M

    2007-09-15

    The transforming growth factor-beta (TGF-beta) pathway has tumor-suppressor activity in many epithelial tissues. Because TGF-beta is a potent inhibitor of epithelial cell proliferation, it has been widely assumed that this property underlies the tumor-suppressor effect. Here, we have used a xenograft model of breast cancer to show that endogenous TGF-beta has the potential to suppress tumorigenesis through a novel mechanism, involving effects at two distinct levels in the hierarchy of cellular progeny that make up the epithelial component of the tumor. First, TGF-beta reduces the size of the putative cancer stem or early progenitor cell population, and second it promotes differentiation of a more committed, but highly proliferative, progenitor cell population to an intrinsically less proliferative state. We further show that reduced expression of the type II TGF-beta receptor correlates with loss of luminal differentiation in a clinical breast cancer cohort, suggesting that this mechanism may be clinically relevant. At a molecular level, the induction of differentiation by TGF-beta involves down-regulation of Id1, and forced overexpression of Id1 can promote tumorigenesis despite persistence of the antiproliferative effect of TGF-beta. These data suggest new roles for the TGF-beta pathway in regulating tumor cell dynamics that are independent of direct effects on proliferation.

  4. Tubeimoside-1 suppresses tumor angiogenesis by stimulation of proteasomal VEGFR2 and Tie2 degradation in a non-small cell lung cancer xenograft model

    PubMed Central

    Gu, Yuan; Körbel, Christina; Scheuer, Claudia; Nenicu, Anca; Menger, Michael D.; Laschke, Matthias W.

    2016-01-01

    Tubeimoside-1 (TBMS1) is a potent anti-tumor phytochemical. Its functional and molecular mode of action, however, remains elusive so far. Since angiogenesis is essential for tumor progression and metastasis, we herein investigated the anti-angiogenic effects of the compound. In a non-small cell lung cancer (NSCLC) xenograft model we found that treatment of CD1 nu/nu mice with TBMS1 (5mg/kg) significantly suppressed the growth and vascularization of NCI-H460 flank tumors. Moreover, TBMS1 dose-dependently reduced vascular sprouting in a rat aortic ring assay. In vitro, TBMS1 induced endothelial cell apoptosis without decreasing the viability of NSCLC tumor cells and inhibited the migration of endothelial cells by disturbing their actin filament organization. TBMS1 further stimulated the proteasomal degradation of vascular endothelial growth factor receptor-2 (VEGFR2) and Tie2 in endothelial cells, which down-regulated AKT/mTOR signaling. These findings indicate that TBMS1 represents a novel phytochemical for anti-angiogenic treatment of cancer and other angiogenesis-related diseases. PMID:26701724

  5. Tumor-targeting Salmonella typhimurium A1-R in combination with doxorubicin eradicate soft tissue sarcoma in a patient-derived orthotopic xenograft (PDOX) model

    PubMed Central

    Murakami, Takashi; DeLong, Jonathan; Eilber, Fritz C.; Zhao, Ming; Zhang, Yong; Zhang, Nan; Singh, Arun; Russell, Tara; Deng, Samantha; Reynoso, Jose; Quan, Cuong; Hiroshima, Yukihiko; Matsuyama, Ryusei; Chishima, Takashi; Tanaka, Kuniya; Bouvet, Michael; Chawla, Sant; Endo, Itaru; Hoffman, Robert M.

    2016-01-01

    A patient with high grade undifferentiated pleomorphic soft-tissue sarcoma from a striated muscle was grown orthotopically in the right biceps femoris muscle of mice to establish a patient-derived orthotopic xenograft (PDOX) model. Twenty PDOX mice were divided into 4 groups: G1, control without treatment; G2, Salmonella typhimurium (S. typhimurium)A1-R administered by intratumoral (i.t.) injection once a week for 4 weeks; G3, doxorubicin (DOX) administered by intraperitoneal (i.p.) injection once a week for 4 weeks; G4, S. typhimurium A1-R (i.t.) administered once a week for 2 weeks followed by i.p. doxorubicin once a week for 2 weeks. On day 25 from the initiation of treatment, tumor volume in G2, G3, and G4 was significantly lower than G1. Mice found without gross tumor included one mouse (20%) in G2; one mouse (20%) in G3; and 3 mice (60%) in G4. Body weight loss did not significantly differ between the 3 treated groups or from the untreated control. Histological examination revealed eradication of tumor only in G4 where mice were treated with S. typhimurium A1-R followed by DOX. Our present study indicates future clinical potential of combining S. typhimurium A1-R with chemotherapy such as DOX for soft tissue sarcoma patients. PMID:26859573

  6. Inhibition of p300 lysine acetyltransferase activity by luteolin reduces tumor growth in head and neck squamous cell carcinoma (HNSCC) xenograft mouse model

    PubMed Central

    Selvi, Ruthrotha B.; Swaminathan, Amrutha; Chatterjee, Snehajyoti; Shanmugam, Muthu K.; Li, Feng; Ramakrishnan, Gowsica B.; Siveen, Kodappully Sivaraman; Chinnathambi, Arunachalam; Zayed, M. Emam; Alharbi, Sulaiman Ali; Basha, Jeelan; Bhat, Akshay; Vasudevan, Madavan; Dharmarajan, Arunasalam; Sethi, Gautam; Kundu, Tapas K.

    2015-01-01

    Chromatin acetylation is attributed with distinct functional relevance with respect to gene expression in normal and diseased conditions thereby leading to a topical interest in the concept of epigenetic modulators and therapy. We report here the identification and characterization of the acetylation inhibitory potential of an important dietary flavonoid, luteolin. Luteolin was found to inhibit p300 acetyltransferase with competitive binding to the acetyl CoA binding site. Luteolin treatment in a xenografted tumor model of head and neck squamous cell carcinoma (HNSCC), led to a dramatic reduction in tumor growth within 4 weeks corresponding to a decrease in histone acetylation. Cells treated with luteolin exhibit cell cycle arrest and decreased cell migration. Luteolin treatment led to an alteration in gene expression and miRNA profile including up-regulation of p53 induced miR-195/215, let7C; potentially translating into a tumor suppressor function. It also led to down-regulation of oncomiRNAs such as miR-135a, thereby reflecting global changes in the microRNA network. Furthermore, a direct correlation between the inhibition of histone acetylation and gene expression was established using chromatin immunoprecipitation on promoters of differentially expressed genes. A network of dysregulated genes and miRNAs was mapped along with the gene ontology categories, and the effects of luteolin were observed to be potentially at multiple levels: at the level of gene expression, miRNA expression and miRNA processing. PMID:26517526

  7. Tumor-targeting Salmonella typhimurium A1-R in combination with doxorubicin eradicate soft tissue sarcoma in a patient-derived orthotopic xenograft (PDOX) model.

    PubMed

    Murakami, Takashi; DeLong, Jonathan; Eilber, Fritz C; Zhao, Ming; Zhang, Yong; Zhang, Nan; Singh, Arun; Russell, Tara; Deng, Samantha; Reynoso, Jose; Quan, Cuong; Hiroshima, Yukihiko; Matsuyama, Ryusei; Chishima, Takashi; Tanaka, Kuniya; Bouvet, Michael; Chawla, Sant; Endo, Itaru; Hoffman, Robert M

    2016-03-15

    A patient with high grade undifferentiated pleomorphic soft-tissue sarcoma from a striated muscle was grown orthotopically in the right biceps femoris muscle of mice to establish a patient-derived orthotopic xenograft (PDOX) model. Twenty PDOX mice were divided into 4 groups: G1, control without treatment; G2, Salmonella typhimurium (S. typhimurium)A1-R administered by intratumoral (i.t.) injection once a week for 4 weeks; G3, doxorubicin (DOX) administered by intraperitoneal (i.p.) injection once a week for 4 weeks; G4, S. typhimurium A1-R (i.t.) administered once a week for 2 weeks followed by i.p. doxorubicin once a week for 2 weeks. On day 25 from the initiation of treatment, tumor volume in G2, G3, and G4 was significantly lower than G1. Mice found without gross tumor included one mouse (20%) in G2; one mouse (20%) in G3; and 3 mice (60%) in G4. Body weight loss did not significantly differ between the 3 treated groups or from the untreated control. Histological examination revealed eradication of tumor only in G4 where mice were treated with S. typhimurium A1-R followed by DOX. Our present study indicates future clinical potential of combining S. typhimurium A1-R with chemotherapy such as DOX for soft tissue sarcoma patients.

  8. Formulation, Characterization, and Antitumor Properties of Trans- and Cis-Citral in the 4T1 Breast Cancer Xenograft Mouse Model

    PubMed Central

    Zeng, San; Kapur, Arvinder; Patankar, Manish S.; Xiong, May P.

    2015-01-01

    Purpose Citral is composed of a random mixture of two geometric stereoisomers geranial (trans-citral) and neral (cis-citral) yet few studies have directly compared their in vivo antitumor properties. A micelle formulation was therefore developed. Methods Geranial and neral were synthesized. Commercially-purchased citral, geranial, and neral were formulated in PEG-b-PCL (block sizes of 5000:10000, Mw/Mn 1.26) micelles. In vitro degradation, drug release, cytotoxicity, flow cytometry, and western blot studies were conducted. The antitumor properties of drug formulations (40 mg/kg and 80 mg/kg based on MTD studies) were evaluated on the 4T1 xenograft mouse model and tumor tissues were analyzed by western blot. Results Micelles encapsulated drugs with >50% LE at 5-40% drug to polymer (w/w), displayed sustained release (t1/2 of 8-9 hours), and improved drug stability at pH 5.0. The IC50 of drug formulations against 4T1 cells ranged from 1.4-9.9 μM. Western blot revealed that autophagy was the main cause of cytotoxicity. Geranial at 80 mg/kg was most effective at inhibiting tumor growth. Conclusions Geranial is significantly more potent than neral and citral at 80 mg/kg (p<0.001) and western blot of tumor tissues confirms that autophagy and not apoptosis is the major mechanism of tumor growth inhibition in p53-null 4T1 cells. PMID:25673043

  9. Assessing Metabolic Changes in Response to mTOR Inhibition in a Mantle Cell Lymphoma Xenograft Model Using AcidoCEST MRI

    PubMed Central

    Akhenblit, Paul J.; Hanke, Neale T.; Gill, Alexander; Persky, Daniel O.; Howison, Christine M.; Pagel, Mark D.; Baker, Amanda F.

    2016-01-01

    AcidoCEST magnetic resonance imaging (MRI) has previously been shown to measure tumor extracellular pH (pHe) with excellent accuracy and precision. This study investigated the ability of acidoCEST MRI to monitor changes in tumor pHe in response to therapy. To perform this study, we used the Granta 519 human mantle cell lymphoma cell line, which is an aggressive B-cell malignancy that demonstrates activation of the phosphatidylinositol-3-kinase/Akt/mammalian target of rapamycin (mTOR) pathway. We performed in vitro and in vivo studies using the Granta 519 cell line to investigate the efficacy and associated changes induced by the mTOR inhibitor, everolimus (RAD001). AcidoCEST MRI studies showed a statistically significant increase in tumor pHe of 0.10 pH unit within 1 day of initiating treatment, which foreshadowed a decrease in tumor growth of the Granta 519 xenograft model. AcidoCEST MRI then measured a decrease in tumor pHe 7 days after initiating treatment, which foreshadowed a return to normal tumor growth rate. Therefore, this study is a strong example that acidoCEST MRI can be used to measure tumor pHe that may serve as a marker for therapeutic efficacy of anticancer therapies. PMID:27140422

  10. Enhanced Replication of Hepatitis E Virus Strain 47832c in an A549-Derived Subclonal Cell Line

    PubMed Central

    Schemmerer, Mathias; Apelt, Silke; Trojnar, Eva; Ulrich, Rainer G.; Wenzel, Jürgen J.; Johne, Reimar

    2016-01-01

    Hepatitis E virus (HEV) is a human pathogen with increasing importance. The lack of efficient cell culture systems hampers systematic studies on its replication cycle, virus neutralization and inactivation. Here, several cell lines were inoculated with the HEV genotype 3c strain 47832c, previously isolated from a chronically infected transplant patient. At 14 days after inoculation the highest HEV genome copy numbers were found in A549 cells, followed by PLC/PRF/5 cells, whereas HepG2/C3A, Huh-7 Lunet BLR and MRC-5 cells only weakly supported virus replication. Inoculation of A549-derived subclone cell lines resulted in most cases in reduced HEV replication. However, the subclone A549/D3 was susceptible to lower virus concentrations and resulted in higher virus yields as compared to parental A549 cells. Transcriptome analysis indicated a downregulation of genes for carcinoembryonic antigen-related cell adhesion molecules (CEACAM) 5 and 6, and an upregulation of the syndecan 2 (SDC2) gene in A549/D3 cells compared to A549 cells. However, treatment of A549/D3 cells or A549 cells with CEACAM- or syndecan 2-specific antisera did not influence HEV replication. The results show that cells supporting more efficient HEV replication can be selected from the A549 cell line. The specific mechanisms responsible for the enhanced replication remain unknown. PMID:27690085

  11. Global Conservation of Protein Status between Cell Lines and Xenografts.

    PubMed

    Biau, Julian; Chautard, Emmanuel; Court, Frank; Pereira, Bruno; Verrelle, Pierre; Devun, Flavien; De Koning, Leanne; Dutreix, Marie

    2016-08-01

    Common preclinical models for testing anticancer treatment include cultured human tumor cell lines in monolayer, and xenografts derived from these cell lines in immunodeficient mice. Our goal was to determine how similar the xenografts are compared with their original cell line and to determine whether it is possible to predict the stability of a xenograft model beforehand. We studied a selection of 89 protein markers of interest in 14 human cell cultures and respective subcutaneous xenografts using the reverse-phase protein array technology. We specifically focused on proteins and posttranslational modifications involved in DNA repair, PI3K pathway, apoptosis, tyrosine kinase signaling, stress, cell cycle, MAPK/ERK signaling, SAPK/JNK signaling, NFκB signaling, and adhesion/cytoskeleton. Using hierarchical clustering, most cell culture-xenograft pairs cluster together, suggesting a global conservation of protein signature. Particularly, Akt, NFkB, EGFR, and Vimentin showed very stable protein expression and phosphorylation levels highlighting that 4 of 10 pathways were highly correlated whatever the model. Other proteins were heterogeneously conserved depending on the cell line. Finally, cell line models with low Akt pathway activation and low levels of Vimentin gave rise to more reliable xenograft models. These results may be useful for the extrapolation of cell culture experiments to in vivo models in novel targeted drug discovery. PMID:27567954

  12. Selenium pretreatment attenuates formaldehyde-induced genotoxicity in A549 cell lines.

    PubMed

    Shi, Yu-Qin; Chen, Xin; Dai, Juan; Jiang, Zhong-Fa; Li, Ning; Zhang, Ben-Yan; Zhang, Zhi-Bing

    2014-11-01

    Formaldehyde is a major industrial chemical and has been extensively used in the manufacture of synthetic resins and chemicals. Numerous studies indicate that formaldehyde can induce various genotoxic effects in vitro and in vivo. A recent study indicated that formaldehyde impaired antioxidant cellular defences and enhanced lipid peroxidation. Selenium is an important antioxidant. We hypothesized that reactive oxygen species (ROS) and lipid peroxidation are involved in formaldehyde-induced genotoxicity in human lung cancer cell line, A549 cell line. To test the hypothesis, we investigated the effects of selenium on formaldehyde-induced genotoxicity in A549 cell lines. The results indicated that exposure to formaldehyde showed the induction of DNA-protein cross-links (DPCs). Formaldehyde significantly increased the malondialdehyde levels and decreased the activities of superoxide dismutase and glutathione peroxidase. In addition, the activations of necrosis factor-κB (NF-κB) and activator protein 1 (AP-1) were induced by the formaldehyde treatment. The pretreatment with selenium counteracted the formaldehyde-induced oxidative stress, ameliorated DPCs and attenuated the activation of NF-κB and AP-1 in A549 cell lines. All the results suggested that the pretreatment with selenium attenuated the formaldehyde-induced genotoxicity through its ROS scavenging and anti-DPCs effects in A549 cell lines.

  13. Functional expression of nicotine influx transporter in A549 human alveolar epithelial cells.

    PubMed

    Tega, Yuma; Yuzurihara, Chihiro; Kubo, Yoshiyuki; Akanuma, Shin-ichi; Ehrhardt, Carsten; Hosoya, Ken-ichi

    2016-02-01

    Nicotine is a potent addictive alkaloid, and is rapidly absorbed through the alveoli of the lung. However, the transport mechanism of nicotine at the human alveolar epithelial barrier has not been investigated in great detail. In the present study, the transport mechanism of nicotine across alveolar epithelium was investigated in vitro using A549 cells, a human adenocarcinoma-derived cell line with an alveolar epithelial cell like phenotype. Nicotine uptake by A549 cells exhibited time-, temperature-, and concentration-dependence with a Km of 50.4 μM. These results suggest that a carrier-mediated transport process is involved in nicotine transport in human alveolar epithelial cells. Nicotine uptake by A549 cells was insensitive to change in extracellular pH. Moreover, nicotine uptake by A549 cells could be inhibited by organic cations such as verapamil and pyrilamine, but not typical substrates of organic cation transporters and β2-agonist. These results suggest that a novel, not yet molecularly identified, organic cation transporter plays a role in nicotine transport which is unlikely to interact with β2-agonist transport. This nicotine influx transporter in human alveolar epithelium might have implications for the rapid absorption of nicotine into the systemic circulation. PMID:26830082

  14. Effect of fucoidan from Turbinaria conoides on human lung adenocarcinoma epithelial (A549) cells.

    PubMed

    Alwarsamy, Madhavarani; Gooneratne, Ravi; Ravichandran, Ramanibai

    2016-11-01

    Fucoidan was purified from seaweed, Turbinaria conoides. Isolated fragments were characterized with NMR ((13)C, (1)H), Gas Chromatography-Mass Spectronomy (GC-MS) and HPLC analysis. The autohydrolysate of fucoidans consisted of sulfated fuco-oligosaccharides having the backbone of α-(1, 3)-linked fuco-pyranose derivatives and minor components of galactose, glucose, mannose and xylose sugars. Fucoidan induced a dose-dependent reduction in cell survival of lung cancer A549 cells by MTT assay (GI50, 75μg/mL). However, it was not cytotoxic to a non-tumorigenic human keratinocyte cell line of skin tissue (HaCaT) (GI50>1.0mg/mL). The apoptotic cells in fucoidan-treated A549 cells were visualized by laser confocal microscopy and cell cycle analysis showed induction of G0/G1 phase arrest of the cell progression cycle. Further, CFSE labeling and flow cytometry highlighted that fucoidan significantly (P<0.05) inhibited the proliferation rate of A549 cells by up to 2-fold compared with the control cells. It is concluded that fucoidan has the potential to act as an anti-proliferative agent on lung carcinoma (A549) cells.

  15. Oxidative stress and inflammatory response to printer toner particles in human epithelial A549 lung cells.

    PubMed

    Könczöl, Mathias; Weiß, Adilka; Gminski, Richard; Merfort, Irmgard; Mersch-Sundermann, Volker

    2013-02-01

    Reports on adverse health effects related to occupational exposure to toner powder are still inconclusive. Therefore, we have previously conducted an in vitro-study to characterize the genotoxic potential of three commercially available black printer toner powders in A549 lung cells. In these cell-based assays it was clearly demonstrated that the tested toner powders damage DNA and induce micronucleus (MN) formation. Here, we have studied the cytotoxic and proinflammatory potential of these three types of printer toner particles and the influence of ROS and NF-κB induction in order to unravel the underlying mechanisms. A549 cells were exposed to various concentrations of printer toner particle suspensions for 24 h. The toner particles were observed to exert significant cytotoxic effects in the WST-1 and neutral red (NR)-assays, although to a varying extent. Caspase 3/7 activity increased, while the mitochondrial membrane potential (MMP) was not affected. Particles of all three printer toner powders induced concentration-dependent formation of reactive oxygen species (ROS), as measured in the DCFH-DA assay. Furthermore, toner particle exposure enhanced interleukin-6 and interleukin-8 production, which is in agreement with activation of the transcription factor NF-κB in A549 cells shown by the electrophoretic mobility shift assay (EMSA). Therefore, it can be concluded that exposure of A549 lung cells to three selected printer toner powders caused oxidative stress through induction of ROS. Increased ROS formation may trigger genotoxic effects and activate proinflammatory pathways.

  16. Effect of fucoidan from Turbinaria conoides on human lung adenocarcinoma epithelial (A549) cells.

    PubMed

    Alwarsamy, Madhavarani; Gooneratne, Ravi; Ravichandran, Ramanibai

    2016-11-01

    Fucoidan was purified from seaweed, Turbinaria conoides. Isolated fragments were characterized with NMR ((13)C, (1)H), Gas Chromatography-Mass Spectronomy (GC-MS) and HPLC analysis. The autohydrolysate of fucoidans consisted of sulfated fuco-oligosaccharides having the backbone of α-(1, 3)-linked fuco-pyranose derivatives and minor components of galactose, glucose, mannose and xylose sugars. Fucoidan induced a dose-dependent reduction in cell survival of lung cancer A549 cells by MTT assay (GI50, 75μg/mL). However, it was not cytotoxic to a non-tumorigenic human keratinocyte cell line of skin tissue (HaCaT) (GI50>1.0mg/mL). The apoptotic cells in fucoidan-treated A549 cells were visualized by laser confocal microscopy and cell cycle analysis showed induction of G0/G1 phase arrest of the cell progression cycle. Further, CFSE labeling and flow cytometry highlighted that fucoidan significantly (P<0.05) inhibited the proliferation rate of A549 cells by up to 2-fold compared with the control cells. It is concluded that fucoidan has the potential to act as an anti-proliferative agent on lung carcinoma (A549) cells. PMID:27516266

  17. Identification of EP4 as a potential target for the treatment of castration-resistant prostate cancer using a novel xenograft model.

    PubMed

    Terada, Naoki; Shimizu, Yosuke; Kamba, Tomomi; Inoue, Takahiro; Maeno, Atsushi; Kobayashi, Takashi; Nakamura, Eijiro; Kamoto, Toshiyuki; Kanaji, Toshiya; Maruyama, Takayuki; Mikami, Yoshiki; Toda, Yoshinobu; Matsuoka, Toshiyuki; Okuno, Yasushi; Tsujimoto, Gozoh; Narumiya, Shuh; Ogawa, Osamu

    2010-02-15

    More effective therapeutic approaches for castration-resistant prostate cancer (CRPC) are urgently needed, thus reinforcing the need to understand how prostate tumors progress to castration resistance. We have established a novel mouse xenograft model of prostate cancer, KUCaP-2, which expresses the wild-type androgen receptor (AR) and which produces the prostate-specific antigen (PSA). In this model, tumors regress soon after castration, but then reproducibly restore their ability to proliferate after 1 to 2 months without AR mutation, mimicking the clinical behavior of CRPC. In the present study, we used this model to identify novel therapeutic targets for CRPC. Evaluating tumor tissues at various stages by gene expression profiling, we discovered that the prostaglandin E receptor EP4 subtype (EP4) was significantly upregulated during progression to castration resistance. Immunohistochemical results of human prostate cancer tissues confirmed that EP4 expression was higher in CRPC compared with hormone-naïve prostate cancer. Ectopic overexpression of EP4 in LNCaP cells (LNCaP-EP4 cells) drove proliferation and PSA production in the absence of androgen supplementation in vitro and in vivo. Androgen-independent proliferation of LNCaP-EP4 cells was suppressed when AR expression was attenuated by RNA interference. Treatment of LNCaP-EP4 cells with a specific EP4 antagonist, ONO-AE3-208, decreased intracellular cyclic AMP levels, suppressed PSA production in vitro, and inhibited castration-resistant growth of LNCaP-EP4 or KUCaP-2 tumors in vivo. Our findings reveal that EP4 overexpression, via AR activation, supports an important mechanism for castration-resistant progression of prostate cancer. Furthermore, they prompt further evaluation of EP4 antagonists as a novel therapeutic modality to treat CRPC.

  18. Anti-CCR7 therapy exerts a potent anti-tumor activity in a xenograft model of human mantle cell lymphoma

    PubMed Central

    2013-01-01

    Background The chemokine receptor CCR7 mediates lymphoid dissemination of many cancers, including lymphomas and epithelial carcinomas, thus representing an attractive therapeutic target. Previous results have highlighted the potential of the anti-CCR7 monoclonal antibodies to inhibit migration in transwell assays. The present study aimed to evaluate the in vivo therapeutic efficacy of an anti-CCR7 antibody in a xenografted human mantle cell lymphoma model. Methods NOD/SCID mice were either subcutaneously or intravenously inoculated with Granta-519 cells, a human cell line derived from a leukemic mantle cell lymphoma. The anti-CCR7 mAb treatment (3 × 200 μg) was started on day 2 or 7 to target lymphoma cells in either a peri-implantation or a post-implantation stage, respectively. Results The anti-CCR7 therapy significantly delayed the tumor appearance and also reduced the volumes of tumors in the subcutaneous model. Moreover, an increased number of apoptotic tumor cells was detected in mice treated with the anti-CCR7 mAb compared to the untreated animals. In addition, significantly reduced number of Granta-519 cells migrated from subcutaneous tumors to distant lymphoid organs, such as bone marrow and spleen in the anti-CCR7 treated mice. In the intravenous models, the anti-CCR7 mAb drastically increased survival of the mice. Accordingly, dissemination and infiltration of tumor cells in lymphoid and non-lymphoid organs, including lungs and central nervous system, was almost abrogated. Conclusions The anti-CCR7 mAb exerts a potent anti-tumor activity and might represent an interesting therapeutic alternative to conventional therapies. PMID:24305507

  19. Chloroquine inhibits cell growth and induces cell death in A549 lung cancer cells.

    PubMed

    Fan, Chuandong; Wang, Weiwei; Zhao, Baoxiang; Zhang, Shangli; Miao, Junying

    2006-05-01

    To investigate the effects of chloroquine diphosphate (CQ) on lung cancer cell growth, we treated A549 cells, a lung cancer cell line, with the drug at various concentrations (0.25-128 microM) for 24-72 h. The results showed that, at lower concentrations (from 0.25 to 32 microM), CQ inhibited the growth of A549 cells and, at the same time, it induced vacuolation with increased volume of acidic compartments (VAC). On the other hand, at higher concentrations (64-128 microM), CQ induced apoptosis at 24 h, while its effect of inducing vacuolation declined. The lactate dehydrogenase (LDH) assay showed that with the treatment of CQ 32-64 microM for 72 h or 128 microM for 48 h, CQ induced necrosis of A549 cells. To understand the possible mechanism by which CQ acts in A549 cells, we further incubated the cells with this drug at the concentrations of 32 or 128 microM in the presence of D609, a specific inhibitor of phosphatidylcholine-specific phospholipase C (PC-PLC). The results showed that D609 (50 microM) could inhibit the effects of CQ 32 microM on the viability and VAC, but it could not change the effects of CQ 128 microM on the same. Our data suggested that CQ inhibited A549 lung cancer cell growth at lower concentrations by increasing the volume of lysosomes and that PC-PLC might be involved in this process. The data also indicated that, at higher concentrations, CQ induced apoptosis and necrosis, but at this time its ability to increase the volume of lysosome gradually declined, and PC-PLC might not be implicated in the process. PMID:16413786

  20. Pharmacologic inhibition of MLK3 kinase activity blocks the in vitro migratory capacity of breast cancer cells but has no effect on breast cancer brain metastasis in a mouse xenograft model.

    PubMed

    Rhoo, Kun Hyoe; Granger, Megan; Sur, Joynita; Feng, Changyong; Gelbard, Harris A; Dewhurst, Stephen; Polesskaya, Oksana

    2014-01-01

    Brain metastasis of breast cancer is an important clinical problem, with few therapeutic options and a poor prognosis. Recent data have implicated mixed lineage kinase 3 (MLK3) in controlling the in vitro migratory capacity of breast cancer cells, as well as the metastasis of MDA-MB-231 breast cancer cells from the mammary fat pad to distant lymph nodes in a mouse xenograft model. We therefore set out to test whether MLK3 plays a role in brain metastasis of breast cancer cells. To address this question, we used a novel, brain penetrant, MLK3 inhibitor, URMC099. URMC099 efficiently inhibited the migration of breast cancer cells in an in vitro cell monolayer wounding assay, and an in vitro transwell migration assay, but had no effect on in vitro cell growth. We also tested the effect of URMC099 on tumor formation in a mouse xenograft model of breast cancer brain metastasis. This analysis showed that URMC099 had no effect on the either the frequency or size of breast cancer brain metastases. We conclude that pharmacologic inhibition of MLK3 by URMC099 can reduce the in vitro migratory capacity of breast cancer cells, but that it has no effect on either the frequency or size of breast cancer brain metastases, in a mouse xenograft model.

  1. Prognostic and functional importance of the engraftment-associated genes in the patient-derived xenograft models of triple-negative breast cancers.

    PubMed

    Moon, Hyeong-Gon; Oh, Keunhee; Lee, Jiwoo; Lee, Minju; Kim, Ju-Yeon; Yoo, Tae-Kyung; Seo, Myung Won; Park, Ae Kyung; Ryu, Han Suk; Jung, Eun-Jung; Kim, Namshin; Jeong, Seongmun; Han, Wonshik; Lee, Dong-Sup; Noh, Dong-Young

    2015-11-01

    We aimed to identify the factors affecting the successful tumor engraftment in breast cancer patient-derived xenograft (PDX) models. Further, we investigated the prognostic significance and the functional importance of the PDX engraftment-related genes in triple-negative breast cancers (TNBC). The clinico-pathologic features of 81 breast cancer patients whose tissues were used for PDX transplantation were analyzed to identify the factors affecting the PDX engraftment. A gene signature associated with the PDX engraftment was discovered and its clinical importance was tested in a publicly available dataset and in vitro assays. Nineteen out of 81 (23.4 %) transplanted tumors were successfully engrafted into the PDX models. The engraftment rate was highest in TNBC when compared to other subtypes (p = 0.001) and in recurrent or chemotherapy-resistant tumors compared to newly diagnosed primary tumors (p = 0.024). PDX tumors originated from the TNBC cases showed more rapid tumor growth in mice. Gene expression profiling showed that down-regulation of genes involved in the tumor-immune interaction was significantly associated with the successful PDX engraftment. The engraftment gene signature was associated with worse survival outcome when tested in publicly available mRNA datasets of TNBC cases. Among the engraftment-related genes, PHLDA2, TKT, and P4HA2 showed high expression in triple-negative breast cancer cell lines, and siRNA-based gene silencing resulted in reduced cell invasion and proliferation in vitro. Our results show that the PDX engraftment may reflect the aggressive phenotype in breast cancer. Genes associated with the PDX engraftment may provide a novel prognostic biomarker and therapeutic targets in TNBC.

  2. Prevention of EBV lymphoma development by oncolytic myxoma virus in a murine xenograft model of post-transplant lymphoproliferative disease

    SciTech Connect

    Kim, Manbok; Rahman, Masmudur M.; Cogle, Christopher R.

    2015-07-10

    Epstein–Barr virus (EBV) has been associated with a variety of epithelial and hematologic malignancies, including B-, T- and NK cell-lymphomas, Hodgkin's disease (HD), post-transplant lymphoproliferative diseases (LPDs), nasopharyngeal and gastric carcinomas, smooth muscle tumors, and HIV-associated lymphomas. Currently, treatment options for EBV-associated malignancies are limited. We have previously shown that myxoma virus specifically targets various human solid tumors and leukemia cells in a variety of animal models, while sparing normal human or murine tissues. Since transplant recipients of bone marrow or solid organs often develop EBV-associated post-transplant LPDs and lymphoma, myxoma virus may be of utility to prevent EBV-associated malignancies in immunocompromised transplant patients where treatment options are frequently limited. In this report, we demonstrate the safety and efficacy of myxoma virus purging as a prophylactic strategy for preventing post-transplant EBV-transformed human lymphomas, using a highly immunosuppressed mouse xenotransplantation model. This provides support for developing myxoma virus as a potential oncolytic therapy for preventing EBV-associated LPDs following transplantation of bone marrow or solid organ allografts. - Highlights: • Myxoma virus effectively infects and purges EBV lymphoma cells in vivo. • Oncolytic myxoma virus effectively eradicates oncogenic EBV tumorigenesis. • Ex vivo pre-treatment of myxoma virus can be effective as a preventive treatment modality for post-transplant lymphoproliferative diseases.

  3. Isolation of Pancreatic Cancer Cells from a Patient-Derived Xenograft Model Allows for Practical Expansion and Preserved Heterogeneity in Culture.

    PubMed

    Pham, Kien; Delitto, Daniel; Knowlton, Andrea E; Hartlage, Emily R; Madhavan, Ricky; Gonzalo, David H; Thomas, Ryan M; Behrns, Kevin E; George, Thomas J; Hughes, Steven J; Wallet, Shannon M; Liu, Chen; Trevino, Jose G

    2016-06-01

    Commercially available, highly passaged pancreatic cancer (PC) cell lines are of limited translational value. Attempts to overcome this limitation have primarily consisted of cancer cell isolation and culture directly from human PC specimens. However, these techniques are associated with exceedingly low success rates. Here, we demonstrate a highly reproducible culture of primary PC cell lines (PPCLs) from patient-derived xenografts, which preserve, in part, the intratumoral heterogeneity known to exist in PC. PPCL expansion from patient-derived xenografts was successful in 100% of attempts (5 of 5). Phenotypic analysis was evaluated with flow cytometry, immunofluorescence microscopy, and short tandem repeat profiling. Importantly, tumorigenicity of PPCLs expanded from patient-derived xenografts was assessed by subcutaneous injection into nonobese diabeteic.Cg-Prkdc(scid)Il2rg(tm1Wjl)/SzJ mice. Morphologically, subcutaneous injection of all PPCLs into mice yielded tumors with similar characteristics to the parent xenograft. PPCLs uniformly expressed class I human leukocyte antigen, epithelial cell adhesion molecule, and cytokeratin-19. Heterogeneity within each PPCL persisted in culture for the frequency of cells expressing the cancer stem cell markers CD44, CD133, and c-Met and the immunologic markers human leukocyte antigen class II and programmed death ligand 1. This work therefore presents a reliable method for the rapid expansion of primary human PC cells and, thereby, provides a platform for translational investigation and, importantly, potential personalized therapeutic approaches.

  4. Human Noxin is an anti-apoptotic protein in response to DNA damage of A549 non-small cell lung carcinoma.

    PubMed

    Won, Kyoung-Jae; Im, Joo-Young; Yun, Chae-Ok; Chung, Kyung-Sook; Kim, Young Joo; Lee, Jung-Sun; Jung, Young-Jin; Kim, Bo-Kyung; Song, Kyung Bin; Kim, Young-Ho; Chun, Ho-Kyung; Jung, Kyeong Eun; Kim, Moon-Hee; Won, Misun

    2014-06-01

    Human Noxin (hNoxin, C11Orf82), a homolog of mouse noxin, is highly expressed in colorectal and lung cancer tissues. hNoxin contains a DNA-binding C-domain in RPA1, which mediates DNA metabolic processes, such as DNA replication and DNA repair. Expression of hNoxin is associated with S phase in cancer cells and in normal cells. Expression of hNoxin was induced by ultraviolet (UV) irradiation. Knockdown of hNoxin caused growth inhibition of colorectal and lung cancer cells. The comet assay and western blot analysis revealed that hNoxin knockdown induced apoptosis through activation of p38 mitogen-activated protein kinase (MAPK)/p53 in non-small cell lung carcinoma A549 cells. Furthermore, simultaneous hNoxin knockdown and treatment with DNA-damaging agents, such as camptothecin (CPT) and UV irradiation, enhanced apoptosis, whereas Trichostatin A (TSA) did not. However, transient overexpression of hNoxin rescued cells from DNA damage-induced apoptosis but did not block apoptosis in the absence of DNA damage. These results suggest that hNoxin may be associated with inhibition of apoptosis in response to DNA damage. An adenovirus expressing a short hairpin RNA against hNoxin transcripts significantly suppressed the growth of A549 tumor xenografts, indicating that hNoxin knockdown has in vivo anti-tumor efficacy. Thus, hNoxin is a DNA damage-induced anti-apoptotic protein and potential therapeutic target in cancer.

  5. Establishment of a human multiple myeloma xenograft model in the chicken to study tumor growth, invasion and angiogenesis.

    PubMed

    Martowicz, Agnieszka; Kern, Johann; Gunsilius, Eberhard; Untergasser, Gerold

    2015-01-01

    Multiple myeloma (MM), a malignant plasma cell disease, remains incurable and novel drugs are required to improve the prognosis of patients. Due to the lack of the bone microenvironment and auto/paracrine growth factors human MM cells are difficult to cultivate. Therefore, there is an urgent need to establish proper in vitro and in vivo culture systems to study the action of novel therapeutics on human MM cells. Here we present a model to grow human multiple myeloma cells in a complex 3D environment in vitro and in vivo. MM cell lines OPM-2 and RPMI-8226 were transfected to express the transgene GFP and were cultivated in the presence of human mesenchymal cells and collagen type-I matrix as three-dimensional spheroids. In addition, spheroids were grafted on the chorioallantoic membrane (CAM) of chicken embryos and tumor growth was monitored by stereo fluorescence microscopy. Both models allow the study of novel therapeutic drugs in a complex 3D environment and the quantification of the tumor cell mass after homogenization of grafts in a transgene-specific GFP-ELISA. Moreover, angiogenic responses of the host and invasion of tumor cells into the subjacent host tissue can be monitored daily by a stereo microscope and analyzed by immunohistochemical staining against human tumor cells (Ki-67, CD138, Vimentin) or host mural cells covering blood vessels (desmin/ASMA). In conclusion, the onplant system allows studying MM cell growth and angiogenesis in a complex 3D environment and enables screening for novel therapeutic compounds targeting survival and proliferation of MM cells. PMID:25993267

  6. Inhibition of Cell Proliferation and Growth of Pancreatic Cancer by Silencing of Carbohydrate Sulfotransferase 15 In Vitro and in a Xenograft Model

    PubMed Central

    Shibazaki, Yuichiro; Yoneyama, Hiroyuki; Fujii, Masato; Hashiguchi, Taishi; Ito, Zensho; Kajihara, Mikio; Misawa, Takeyuki; Homma, Sadamu; Ohkusa, Toshifumi

    2015-01-01

    Chondroitin sulfate E (CS-E), a highly sulfated glycosaminoglycan, is known to promote tumor invasion and metastasis. Because the presence of CS-E is detected in both tumor and stromal cells in pancreatic ductal adenocarcinoma (PDAC), multistage involvement of CS-E in the development of PDAC has been considered. However, its involvement in the early stage of PDAC progression is still not fully understood. In this study, to clarify the direct role of CS-E in tumor, but not stromal, cells of PDAC, we focused on carbohydrate sulfotransferase 15 (CHST15), a specific enzyme that biosynthesizes CS-E, and investigated the effects of the CHST15 siRNA on tumor cell proliferation in vitro and growth in vivo. CHST15 mRNA is highly expressed in the human pancreatic cancer cell lines PANC-1, MIA PaCa-2, Capan-1 and Capan-2. CHST15 siRNA significantly inhibited the expression of CHST15 mRNA in these four cells in vitro. Silencing of the CHST15 gene in the cells was associated with significant reduction of proliferation and up-regulation of the cell cycle inhibitor-related gene p21CIP1/WAF1. In a subcutaneous xenograft tumor model of PANC-1 in nude mice, a single intratumoral injection of CHST15 siRNA almost completely suppressed tumor growth. Reduced CHST15 protein signals associated with tumor necrosis were observed with the treatment with CHST15 siRNA. These results provide evidence of the direct action of CHST15 on the proliferation of pancreatic tumor cells partly through the p21CIP1/WAF1 pathway. Thus, CHST15-CS-E axis-mediated tumor cell proliferation could be a novel therapeutic target in the early stage of PDAC progression. PMID:26642349

  7. Inhibition of Cell Proliferation and Growth of Pancreatic Cancer by Silencing of Carbohydrate Sulfotransferase 15 In Vitro and in a Xenograft Model.

    PubMed

    Takakura, Kazuki; Shibazaki, Yuichiro; Yoneyama, Hiroyuki; Fujii, Masato; Hashiguchi, Taishi; Ito, Zensho; Kajihara, Mikio; Misawa, Takeyuki; Homma, Sadamu; Ohkusa, Toshifumi; Koido, Shigeo

    2015-01-01

    Chondroitin sulfate E (CS-E), a highly sulfated glycosaminoglycan, is known to promote tumor invasion and metastasis. Because the presence of CS-E is detected in both tumor and stromal cells in pancreatic ductal adenocarcinoma (PDAC), multistage involvement of CS-E in the development of PDAC has been considered. However, its involvement in the early stage of PDAC progression is still not fully understood. In this study, to clarify the direct role of CS-E in tumor, but not stromal, cells of PDAC, we focused on carbohydrate sulfotransferase 15 (CHST15), a specific enzyme that biosynthesizes CS-E, and investigated the effects of the CHST15 siRNA on tumor cell proliferation in vitro and growth in vivo. CHST15 mRNA is highly expressed in the human pancreatic cancer cell lines PANC-1, MIA PaCa-2, Capan-1 and Capan-2. CHST15 siRNA significantly inhibited the expression of CHST15 mRNA in these four cells in vitro. Silencing of the CHST15 gene in the cells was associated with significant reduction of proliferation and up-regulation of the cell cycle inhibitor-related gene p21CIP1/WAF1. In a subcutaneous xenograft tumor model of PANC-1 in nude mice, a single intratumoral injection of CHST15 siRNA almost completely suppressed tumor growth. Reduced CHST15 protein signals associated with tumor necrosis were observed with the treatment with CHST15 siRNA. These results provide evidence of the direct action of CHST15 on the proliferation of pancreatic tumor cells partly through the p21CIP1/WAF1 pathway. Thus, CHST15-CS-E axis-mediated tumor cell proliferation could be a novel therapeutic target in the early stage of PDAC progression. PMID:26642349

  8. Antitumor activity and prolonged survival by carbon-nanotube-mediated therapeutic siRNA silencing in a human lung xenograft model.

    PubMed

    Podesta, Jennifer E; Al-Jamal, Khuloud T; Herrero, M Antonia; Tian, Bowen; Ali-Boucetta, Hanene; Hegde, Vikas; Bianco, Alberto; Prato, Maurizio; Kostarelos, Kostas

    2009-05-01

    Carbon nanotubes are novel nanomaterials that are thought to offer potential benefits to a variety of biomedical and clinical applications. In this study, the treatment of a human lung carcinoma model in vivo using siRNA sequences leading to cytotoxicity and cell death is carried out using either cationic liposomes (DOTAP:cholesterol) or amino-functionalized multi-walled carbon nanotubes (MWNT - NH(+)(3)). Validation for the most cytotoxic siRNA sequence using a panel of human carcinoma and murine cells reveals that the proprietary siTOX sequence is human specific and can lead to significant cytotoxic activities delivered both by liposome or MWNT - NH(+)(3) in vitro. A comparative study using both types of vector indicates that only MWNT - NH(+)(3):siRNA complexes administered intratumorally can elicit delayed tumor growth and increased survival of xenograft-bearing animals. siTOX delivery via the cationic MWNT - NH(+)(3) is biologically active in vivo by triggering an apoptotic cascade, leading to extensive necrosis of the human tumor mass. This suggests that carbon-nanotube-mediated delivery of siRNA by intratumoral administration leads to successful and statistically significant suppression of tumor volume, followed by a concomitant prolongation of survival of human lung tumor-bearing animals. The direct comparison between carbon nanotubes and liposomes demonstrates the potential advantages offered by carbon nanotubes for the intracellular delivery of therapeutic agents in vivo. The present work may act as the impetus for further studies to explore the therapeutic capacity of chemically functionalized carbon nanotubes to deliver siRNA directly into the cytoplasm of target cells and achieve effective therapeutic silencing in various disease indications where local delivery is feasible or desirable.

  9. Oral administration of naturally occurring chitosan-based nanoformulated green tea polyphenol EGCG effectively inhibits prostate cancer cell growth in a xenograft model

    PubMed Central

    Mukhtar, Hasan

    2014-01-01

    In preclinical animal models, several phytochemicals have shown excellent potential to be used as effective agents in preventing and treating many cancers. However, the limited bioavailability of active agents could be one reason for their restricted usefulness for human consumption. To overcome this limitation, we recently introduced the concept of nanochemoprevention by encapsulating useful bioactive food components for their slow and sustained release. Here, we report the synthesis, characterization and efficacy assessment of a nanotechnology-based oral formulation of chitosan nanoparticles encapsulating epigallocatechin-3-gallate (Chit-nanoEGCG) for the treatment of prostate cancer (PCa) in a preclinical setting. Chit-nanoEGCG with a size of <200nm diameter and encapsulating EGCG as determined by dynamic light scattering and transmission electron microscope showed slow release of EGCG in simulated gastric juice acidic pH and faster release in simulated intestinal fluid. The antitumor efficacy of Chit-nanoEGCG was assessed in subcutaneously implanted 22Rν1 tumor xenografts in athymic nude mice. Treatment with Chit-nanoEGCG resulted in significant inhibition of tumor growth and secreted prostate-specific antigen levels compared with EGCG and control groups. In tumor tissues of mice treated with Chit-nanoEGCG, compared with groups treated with EGCG and controls, there was significant (i) induction of poly (ADP-ribose) polymerases cleavage, (ii) increase in the protein expression of Bax with concomitant decrease in Bcl-2, (iii) activation of caspases and (iv) reduction in Ki-67 and proliferating cell nuclear antigen. Through this study, we propose a novel preventive and therapeutic modality for PCa using EGCG that addresses issues related to bioavailability. PMID:24072771

  10. A novel, selective inhibitor of fibroblast growth factor receptors that shows a potent broad spectrum of antitumor activity in several tumor xenograft models.

    PubMed

    Zhao, Genshi; Li, Wei-Ying; Chen, Daohong; Henry, James R; Li, Hong-Yu; Chen, Zhaogen; Zia-Ebrahimi, Mohammad; Bloem, Laura; Zhai, Yan; Huss, Karen; Peng, Sheng-Bin; McCann, Denis J

    2011-11-01

    The fibroblast growth factor receptors (FGFR) are tyrosine kinases that are present in many types of endothelial and tumor cells and play an important role in tumor cell growth, survival, and migration as well as in maintaining tumor angiogenesis. Overexpression of FGFRs or aberrant regulation of their activities has been implicated in many forms of human malignancies. Therefore, targeting FGFRs represents an attractive strategy for development of cancer treatment options by simultaneously inhibiting tumor cell growth, survival, and migration as well as tumor angiogenesis. Here, we describe a potent, selective, small-molecule FGFR inhibitor, (R)-(E)-2-(4-(2-(5-(1-(3,5-Dichloropyridin-4-yl)ethoxy)-1H-indazol-3yl)vinyl)-1H-pyrazol-1-yl)ethanol, designated as LY2874455. This molecule is active against all 4 FGFRs, with a similar potency in biochemical assays. It exhibits a potent activity against FGF/FGFR-mediated signaling in several cancer cell lines and shows an excellent broad spectrum of antitumor activity in several tumor xenograft models representing the major FGF/FGFR relevant tumor histologies including lung, gastric, and bladder cancers and multiple myeloma, and with a well-defined pharmacokinetic/pharmacodynamic relationship. LY2874455 also exhibits a 6- to 9-fold in vitro and in vivo selectivity on inhibition of FGF- over VEGF-mediated target signaling in mice. Furthermore, LY2874455 did not show VEGF receptor 2-mediated toxicities such as hypertension at efficacious doses. Currently, this molecule is being evaluated for its potential use in the clinic.

  11. Continuous administration of bevacizumab plus capecitabine, even after acquired resistance to bevacizumab, restored anti-angiogenic and antitumor effect in a human colorectal cancer xenograft model

    PubMed Central

    Iwai, Toshiki; Sugimoto, Masamichi; Harada, Suguru; Yorozu, Keigo; Kurasawa, Mitsue; Yamamoto, Kaname

    2016-01-01

    Vascular endothelial growth factor (VEGF)-neutralizing therapy with bevacizumab has become increasingly important for treating colorectal cancer. It was demonstrated that second-line chemotherapy together with bevacizumab after disease progression (PD) on first-line therapy including bevacizumab showed clinical benefits in metastatic colorectal and breast cancers (ML18147 trial, TANIA trial). One of the rationales for these trials was that the refractoriness to first-line therapy is caused by resistance to not so much bevacizumab as to the chemotherapeutic agents. Nevertheless, resistance to bevacizumab cannot be ruled out because VEGF-independent angiogenesis has been reported to be a mechanism of resistance to anti-VEGF therapy. In this study, we used a xenograft model with the human colon cancer HT-29 cells to investigate the mechanisms underlying the effect of continued administration of bevacizumab plus capecitabine even after resistance to bevacizumab was acquired. The combination of capecitabine plus bevacizumab exhibited significantly stronger antitumor and anti-angiogenic activities than did monotherapy with either agent. Capecitabine treatment significantly increased the intratumoral VEGF level compared with the control group; however, the combination with bevacizumab neutralized the VEGF. Among angiogenic factors other than VEGF, intratumoral galectin-3, which reportedly promotes angiogenesis both dependent on, and independently of VEGF, was significantly decreased in the capecitabine group and the combination group compared with the control group. In an in vitro experiment, 5-fluorouracil (5-FU), an active metabolite of capecitabine, inhibited galectin-3 production by HT-29 cells. These results suggested that capecitabine has a dual mode of action: namely, inhibition of tumor cell growth and inhibition of galectin-3 production by tumor cells. Thus, capecitabine and bevacizumab may work in a mutually complementary manner in tumor angiogenesis inhibition

  12. Dependence of Wilms tumor cells on signaling through insulin-like growth factor 1 in an orthotopic xenograft model targetable by specific receptor inhibition.

    PubMed

    Bielen, Aleksandra; Box, Gary; Perryman, Lara; Bjerke, Lynn; Popov, Sergey; Jamin, Yann; Jury, Alexa; Valenti, Melanie; Brandon, Alexis de Haven; Martins, Vanessa; Romanet, Vincent; Jeay, Sebastien; Raynaud, Florence I; Hofmann, Francesco; Robinson, Simon P; Eccles, Suzanne A; Jones, Chris

    2012-05-15

    We have previously demonstrated an increased DNA copy number and expression of IGF1R to be associated with poor outcome in Wilms tumors. We have now tested whether inhibiting this receptor may be a useful therapeutic strategy by using a panel of Wilms tumor cell lines. Both genetic and pharmacological targeting resulted in inhibition of downstream signaling through PI3 and MAP kinases, G(1) cell cycle arrest, and cell death, with drug efficacy dependent on the levels of phosphorylated IGF1R. These effects were further associated with specific gene expression signatures reflecting pathway inhibition, and conferred synergistic chemosensitisation to doxorubicin and topotecan. In the in vivo setting, s.c. xenografts of WiT49 cells resembled malignant rhabdoid tumors rather than Wilms tumors. Treatment with an IGF1R inhibitor (NVP-AEW541) showed no discernable antitumor activity and no downstream pathway inactivation. By contrast, Wilms tumor cells established orthotopically within the kidney were histologically accurate and exhibited significantly elevated insulin-like growth factor-mediated signaling, and growth was significantly reduced on treatment with NVP-AEW541 in parallel with signaling pathway ablation. As a result of the paracrine effects of enhanced IGF2 expression in Wilms tumor, this disease may be acutely dependent on signaling through the IGF1 receptor, and thus treatment strategies aimed at its inhibition may be useful in the clinic. Such efficacy may be missed if only standard ectopic models are considered as a result of an imperfect recapitulation of the specific tumor microenvironment.

  13. Zinc Ionophore (Clioquinol) Inhibition of Human ZIP1-Deficient Prostate Tumor Growth in the Mouse Ectopic Xenograft Model: A Zinc Approach for the Efficacious Treatment of Prostate Cancer

    PubMed Central

    Franklin, Renty B.; Zou, Jing; Zheng, Yao; Naslund, Michael J.; Costello, Leslie C.

    2016-01-01

    Prostate cancer remains the second leading cause of cancer deaths in males. This is mainly due to the absence of an available efficacious chemotherapy despite decades of research in pursuit of effective treatment approaches. A plausible target for the treatment is the established clinical relationship that the zinc levels in the malignant cells are markedly decreased compared to the normal epithelium in virtually all cases of prostate cancer, and at all stages malignancy. The decrease in zinc results from the downregulation of the functional zinc uptake transporter, ZIP1; which occurs during early development of prostate malignancy. This is an essential requirement for the development of malignancy to prevent the cytotoxic/tumor-suppressor effects of increased zinc on the premalignant and malignant cells. Thus prostate cancer is a ZIP1-deficient malignancy. This relationship provides the basis for a treatment regimen that will facilitate the uptake and accumulation of zinc into the premalignant and malignant cells. In this report we employed a zinc ionophore (clioquinol) approach in the treatment of mice with human ZIP1-deficient prostate tumors (ectopic xenograft model). Clioquinol treatment resulted in 85%inhibition of tumor growth due to the cytotoxic effects of zinc. Coupled with additional results from earlier studies, the compelling evidence provides a plausible approach for the effective treatment of human prostate cancer; including primary site malignancy, hormone-resistant cancer, and metastasis. Additionally, this approach might be effective in preventing the development of malignancy in individuals suspected of presenting with early development of malignancy. Clinical trials are now required in leading to the potential for an efficacious zinc-treatment approach, which is urgently needed for the treatment of prostate cancer. PMID:26878064

  14. Total lymphoid irradiation and discordant cardiac xenografts

    SciTech Connect

    Kaplan, E.; Dresdale, A.R.; Diehl, J.T.; Katzen, N.A.; Aronovitz, M.J.; Konstam, M.A.; Payne, D.D.; Cleveland, R.J. )

    1990-01-01

    Total lymphoid irradiation can prolong concordant cardiac xenografts. The effects of total lymphoid irradiation in a discordant xenograft model (guinea pig to rat) were studied with and without adjuvant pharmacologic immunosuppression. Inbred Lewis rats were randomly allocated to one of four groups. Group 1 (n = 6) served as a control group and rats received no immunosuppression. Group 2 (n = 5) received triple-drug therapy that consisted of intraperitoneal azathioprine (2 mg/kg), cyclosporine (20 mg/kg), and methylprednisolone (1 mg/kg) for 1 week before transplantation. Group 3 animals (n = 5) received 15 Gy of total lymphoid irradiation in 12 divided doses over a 3-week period. Group 4 (n = 6) received both triple-drug therapy and total lymphoid irradiation as described for groups 2 and 3. Complement-dependent cytotoxicity assay was performed to determine if a correlation between complement-dependent cytotoxicity and rejection-free interval existed. Rejection was defined as cessation of graft pulsation and was confirmed by histologic test results. Only groups 1 and 2 showed a difference in survival (group 1, 6.9 +/- 1.0 minutes; group 2, 14.2 +/- 2.7 minutes, p = 0.02). Although total lymphoid irradiation did decrease complement-dependent cytotoxicity, linear regression revealed no correlation between complement-dependent cytotoxicity and graft survival (coefficient of correlation, 0.30). Unlike concordant cardiac xenografts, total lymphoid irradiation with or without triple-drug therapy does not prolong graft survival.

  15. Blockade of the ERK pathway enhances the therapeutic efficacy of the histone deacetylase inhibitor MS-275 in human tumor xenograft models

    SciTech Connect

    Sakamoto, Toshiaki; Ozaki, Kei-ichi; Fujio, Kohsuke; Kajikawa, Shu-hei; Uesato, Shin-ichi; Watanabe, Kazushi; Tanimura, Susumu; Koji, Takehiko; Kohno, Michiaki

    2013-04-19

    Highlights: •Blockade of the ERK pathway enhances the anticancer efficacy of HDAC inhibitors. •MEK inhibitors sensitize human tumor xenografts to HDAC inhibitor cytotoxicity. •Such the enhanced efficacy is achieved by a transient blockade of the ERK pathway. •This drug combination provides a promising therapeutic strategy for cancer patients. -- Abstract: The ERK pathway is up-regulated in various human cancers and represents a prime target for mechanism-based approaches to cancer treatment. Specific blockade of the ERK pathway alone induces mostly cytostatic rather than pro-apoptotic effects, however, resulting in a limited therapeutic efficacy of the ERK kinase (MEK) inhibitors. We previously showed that MEK inhibitors markedly enhance the ability of histone deacetylase (HDAC) inhibitors to induce apoptosis in tumor cells with constitutive ERK pathway activation in vitro. To evaluate the therapeutic efficacy of such drug combinations, we administered the MEK inhibitor PD184352 or AZD6244 together with the HDAC inhibitor MS-275 in nude mice harboring HT-29 or H1650 xenografts. Co-administration of the MEK inhibitor markedly sensitized the human xenografts to MS-275 cytotoxicity. A dose of MS-275 that alone showed only moderate cytotoxicity thus suppressed the growth of tumor xenografts almost completely as well as induced a marked reduction in tumor cellularity when administered with PD184352 or AZD6244. The combination of the two types of inhibitor also induced marked oxidative stress, which appeared to result in DNA damage and massive cell death, specifically in the tumor xenografts. The enhanced therapeutic efficacy of the drug combination was achieved by a relatively transient blockade of the ERK pathway. Administration of both MEK and HDAC inhibitors represents a promising chemotherapeutic strategy with improved safety for cancer patients.

  16. In vivo sampling of Verteporfin uptake in pancreas cancer xenograft models: comparison of surface, oral, and interstitial measurements

    NASA Astrophysics Data System (ADS)

    Isabelle, Martin; O'Hara, Julia A.; Samkoe, Kimberley S.; Hoopes, P. Jack; Mosse, Sandy; Pereira, Stephen; Hasan, Tayyaba; Pogue, Brian W.

    2010-02-01

    Photodynamic therapy (PDT) mediated with Verteporfin is being investigated as a pancreatic cancer treatment in the cases for non-surgical candidates. Tissue response to PDT is based on a number of parameters including photosensitizer (PS) dose, light dose and time interval between light application and PS injection. In this study, PS uptake and distribution in animal leg muscle, oral cavity tissues, pancreas and tumor was measured in vivo using light-induced fluorescence spectroscopy (LIFS) via an Aurora Optics Inc. PDT fluorescence dosimeter. An orthotopic pancreatic cancer model (AsPC-1) was implanted in SCID mice and treated with the PS. Probe measurements were made using a surface probe and an interstitial needle probe before and up to one hour after intravenous tail vein injection of the PS. The study demonstrated that it is possible to correlate in-vivo LIFS measurements of the PS uptake in the pancreas with measurements taken from the oral cavity indicating that light dosimetry of PDT of the pancreas can be ascertained from the LIFS measurements in the oral cavity. These results emphasize the importance of light dosimetry in improving the therapeutic outcome of PDT through light dose adaptation to the relative in situ tissue PS concentration.

  17. A xenograft model of macrophage activation syndrome amenable to anti-CD33 and anti–IL-6R treatment

    PubMed Central

    Wunderlich, Mark; Devarajan, Mahima; Ravishankar, Navin; Sexton, Christina; Kumar, Ashish R.; Mizukawa, Benjamin; Mulloy, James C.

    2016-01-01

    Transgenic expression of key myelosupportive human cytokines in immune-deficient mice corrects for the lack of cross-species activities of stem cell factor (SCF), IL-3, and GM-CSF. When engrafted with human umbilical cord blood (UCB), these triple-transgenic mice produce BM and spleen grafts with much higher myeloid composition, relative to nontransgenic controls. Shortly after engraftment with UCB, these mice develop a severe, fatal macrophage activation syndrome (MAS) characterized by a progressive drop in rbc numbers, increased reticulocyte counts, decreased rbc half-life, progressive cytopenias, and evidence of chronic inflammation, including elevated human IL-6. The BM becomes strikingly hypocellular, and spleens are significantly enlarged with evidence of extramedullary hematopoiesis and activated macrophages engaged in hemophagocytosis. This manifestation of MAS does not respond to lymphocyte-suppressive therapies such as steroids, i.v. immunoglobulin, or antibody-mediated ablation of human B and T cells, demonstrating a lymphocyte-independent mechanism of action. In contrast, elimination of human myeloid cells using gemtuzumab ozogamicin (anti-CD33) completely reversed the disease. Additionally, the IL-6R antibody tocilizumab delayed progression and prolonged lifespan. This new model of MAS provides an opportunity for investigation of the mechanisms driving this disease and for the testing of directed therapies in a humanized mouse.

  18. Target therapy of multiple myeloma by PTX-NPs and ABCG2 antibody in a mouse xenograft model.

    PubMed

    Yang, Cuiping; Xiong, Fei; Dou, Jun; Xue, Jun; Zhan, Xi; Shi, Fangfang; Li, Miao; Wu, Songyan; Luo, Shouhua; Zhang, Tianzhu; Zhang, Yu; Ming, Ji; Gu, Ning

    2015-09-29

    Multiple myeloma (MM) remains to be an incurable disease. The purpose of this study was to evaluate the effect of ABCG2 monoclonal antibody (McAb) combined with paclitaxel (PTX) conjugated with Fe3O4 nanoparticles (NPs) on MM progressed from cancer stem cells (CSCs) in non-obese-diabetic/severe-combined-immunodeficiency (NOD/SCID) mouse model. Mice were injected with MM CSCs as marked by CD138-CD34- phenotypes through tail veins. The developed MM mice were examined by micro-computer tomography scanning, ultrasonography and enzyme-linked immunosorbent analysis. These mice were then intravenously treated with different combinations of NPs, PTX, McAb, PTX-NPs and melphalan/prednisone once a week for four weeks. The injected mice developed characteristic MM-associated syndromes, including lytic bone lesions, renal damages and proteinuria. All the treated mice showed decrease in bone lesions, renal damages and anemia but increase in apoptosis compared with the mice treated with NPs only. In particular, the treatment with ABCG2 McAb plus PTX-NPs induced the strongest therapeutic response and had an efficacy even better than that of melphalan/prednisone, a conventional regimen for MM patients. These data suggest that PTX-NPs with ABCG2 McAb can be developed into potential treatment regimens for patients with relapsed/refractory MM.

  19. Iron-Oxide-Based Nanovector for Tumor Targeted siRNA Delivery in an Orthotopic Hepatocellular Carcinoma Xenograft Mouse Model.

    PubMed

    Wang, Kui; Kievit, Forrest M; Sham, Jonathan G; Jeon, Mike; Stephen, Zachary R; Bakthavatsalam, Arvind; Park, James O; Zhang, Miqin

    2016-01-27

    Hepatocellular carcinoma (HCC) is one of the deadliest cancers worldwide. Small interfering RNA (siRNA) holds promise as a new class of therapeutics for HCC, as it can achieve sequence-specific gene knockdown with low cytotoxicity. However, the main challenge in the clinical application of siRNA lies in the lack of effective delivery approaches that need to be highly specific and thus incur low or no systemic toxicity. Here, a nonviral nanoparticle-based gene carrier is presented that can specifically deliver siRNA to HCC. The nanovector (NP-siRNA-GPC3 Ab) is made of an iron oxide core coated with chitosan-polyethylene glycol (PEG) grafted polyethyleneimine copolymer, which is further functionalized with siRNA and conjugated with a monoclonal antibody (Ab) against human glypican-3 (GPC3) receptor highly expressed in HCC. A rat RH7777 HCC cell line that coexpresses human GPC3 and firefly luciferase (Luc) is established to evaluate the nanovector. The nanoparticle-mediated delivery of siRNA against Luc effectively suppresses Luc expression in vitro without notable cytotoxicity. Significantly, NP-siLuc-GPC3 Ab administered intravenously in an orthotopic model of HCC is able to specifically bound to tumor and induce remarkable inhibition of Luc expression. The findings demonstrate the potential of using this nanovector for targeted delivery of therapeutic siRNA to HCC.

  20. Sunitinib Combined with Angiotensin-2 Type-1 Receptor Antagonists Induces More Necrosis: A Murine Xenograft Model of Renal Cell Carcinoma

    PubMed Central

    Verhoest, Grégory; Dolley-Hitze, Thibault; Jouan, Florence; Belaud-Rotureau, Marc-Antoine; Oger, Emmanuel; Bensalah, Karim; Arlot-Bonnemains, Yannick; Collet, Nicolas; Rioux-Leclercq, Nathalie; Vigneau, Cécile

    2014-01-01

    Background. Angiotensin-2 type-1 receptor antagonists not are only antihypertensive drugs but also can inhibit VEGF production. We hypothesised that adding telmisartan to sunitinib could potentiate the antiangiogenic effects. Material and Methods. 786-O cell lines were injected in nude mice. After tumor development, mice were divided into 4 groups: the first was the control group (DMSO), the second group was treated with sunitinib alone, the third group was treated with telmisartan alone, and the fourth group was treated with the combination. Drugs were orally administered every day for four weeks. Animals were sacrificed after treatment. Blood and tumor tissues were collected for analysis by immunohistochemistry, Western Blot, and ELISA methods. Results. All animals developed a ccRCC and ten in each group were treated. Using a kinetic model, tumors tended to grow slower in the combination group compared to others (P = 0.06). Compared to sunitinib alone, the addition of telmisartan significantly increased tissue necrosis (P = 0.038). Central microvascular density decreased (P = 0.0038) as well as circulating VEGF (P = 0.003). There was no significant variation in proliferation or apoptosis markers. Conclusion. The combination of sunitinib and telmisartan revealed an enhancement of the blockage of the VEGF pathway on renal tumor resulting in a decrease in neoangiogenesis and an increase in necrosis. PMID:24967411

  1. A xenograft model of macrophage activation syndrome amenable to anti-CD33 and anti–IL-6R treatment

    PubMed Central

    Wunderlich, Mark; Devarajan, Mahima; Ravishankar, Navin; Sexton, Christina; Kumar, Ashish R.; Mizukawa, Benjamin; Mulloy, James C.

    2016-01-01

    Transgenic expression of key myelosupportive human cytokines in immune-deficient mice corrects for the lack of cross-species activities of stem cell factor (SCF), IL-3, and GM-CSF. When engrafted with human umbilical cord blood (UCB), these triple-transgenic mice produce BM and spleen grafts with much higher myeloid composition, relative to nontransgenic controls. Shortly after engraftment with UCB, these mice develop a severe, fatal macrophage activation syndrome (MAS) characterized by a progressive drop in rbc numbers, increased reticulocyte counts, decreased rbc half-life, progressive cytopenias, and evidence of chronic inflammation, including elevated human IL-6. The BM becomes strikingly hypocellular, and spleens are significantly enlarged with evidence of extramedullary hematopoiesis and activated macrophages engaged in hemophagocytosis. This manifestation of MAS does not respond to lymphocyte-suppressive therapies such as steroids, i.v. immunoglobulin, or antibody-mediated ablation of human B and T cells, demonstrating a lymphocyte-independent mechanism of action. In contrast, elimination of human myeloid cells using gemtuzumab ozogamicin (anti-CD33) completely reversed the disease. Additionally, the IL-6R antibody tocilizumab delayed progression and prolonged lifespan. This new model of MAS provides an opportunity for investigation of the mechanisms driving this disease and for the testing of directed therapies in a humanized mouse. PMID:27699249

  2. Evaluating dynamic contrast-enhanced and photoacoustic CT to assess intra-tumor heterogeneity in xenograft mouse models

    NASA Astrophysics Data System (ADS)

    Stantz, Keith M.; Liu, Bo; Cao, Minsong; Reinecke, Dan; Dzemidzic, Mario; Liang, Yun; Kruger, Robert

    2006-03-01

    Purpose: To evaluate photoacoustic CT spectroscopy (PCT-S) and dynamic contrast-enhanced CT (DCE-CT) ability to measure parameters - oxygen saturation and vascular physiology - associated with the intra-tumor oxygenation status. Material and Methods: Breast (VEGF165 enhance MCF-7) and ovarian (SKOV3x) cancer cells were implanted into the fat pads and flanks of immune deficient mice and allowed to grow to a diameter of 8-15 mm. CT was used to determine physiological parameters by acquiring a sequence of scans over a 10 minute period after an i.v. injection of a radio-opaque contrast agent (Isovue). These time-dependent contrast-enhanced curves were fit to a two-compartmental model determining tumor perfusion, fractional plasma volume, permeability-surface area produce, and fractional interstitial volume on a voxel-by-voxel basis. After which, the tumors were imaged using photoacoustic CT (Optosonics, Inc., Indianapolis, IN 46202). The near infrared spectra (700-910 nm) within the vasculature was fit to linear combination of measured oxy- and deoxy-hemoglobin blood samples to obtain oxygen saturation levels (SaO II). Results: The PCT-S scanner was first calibrated using different samples of oxygenated blood, from which a statistical error ranging from 2.5-6.5% was measured and a plot of the hemoglobin dissociation curve was consistent with empirical formula. In vivo determination of tumor vasculature SaO II levels were measurably tracked, and spatially correlated to the periphery of the tumor. Tumor depend variations in SaO II - 0.32 (ovarian) and 0.60 (breast) - and in vascular physiology - perfusion, 1.03 and 0.063 mL/min/mL, and fractional plasma volume, 0.20 and 0.07 - were observed. Conclusion: Combined, PCT-S and CED-CT has the potential to measure intra-tumor levels of tumor oxygen saturation and vascular physiology, key parameters associated with hypoxia.

  3. Endostatin enhances antitumor effect of tumor antigen-pulsed dendritic cell therapy in mouse xenograft model of lung carcinoma

    PubMed Central

    Liang, Jing; Liu, Xiaolin; Xie, Qi; Chen, Guoling; Li, Xingyu; Jia, Yanrui; Yin, Beibei; Qu, Xun; Li, Yan

    2016-01-01

    Objective To investigate the antitumor effect of endostatin combined with tumor antigen-pulsed dendritic cell (DC)-T cell therapy on lung cancer. Methods Transplanted Lewis lung cancer (LLC) models of C57BL/6 mice were established by subcutaneous injection of LLC cells in left extremity axillary. Tumor antigen-pulsed DC-T cells from spleen cells and bone of mice were cultured in vitro. Tumor-bearing mice were randomly divided into three groups, including DC-T+endostatin group, DC-T group, and phosphate-buffered saline (PBS) control group. Microvessel density (MVD) of tumor tissue in tumor-bearing mice was determined by immunohistochemistry (IHC). The expressions of vascular endothelial growth factor (VEGF) and hypoxia-inducible factor-1α (HIF-1α) were determined by Western blotting and IHC staining. The proportions of CD8+ T cells, mature dendritic cells (mDC), tumor-associated macrophages [TAM (M1/M2)], and myeloid-derived suppressor cells (MDSC) in suspended cells of tumor tissue were determined by flow cytometry. The expressions of interleukin (IL)-6, IL-10, IL-17, transforming growth factor-β (TGF-β) and interferon-γ (IFN-γ) in suspended cells of tumor tissue were detected by enzyme-linked immune sorbent assay (ELISA). Results DC-T cells combined with endostatin remarkably suppressed tumor growth. MVD of mice in DC-T+endostatin group was significantly lower than that of the control group and DC-T monotherapy group. The expressions of VEGF, IL-6 and IL-17 in tumors were markedly decreased, but IFN-γ and HIF-1α increased after treating with DC-T cells combined with endostatin, compared to control group and DC-T group. In the DC-T+endostatin group, the proportions of MDSC and TAM (M2 type) were significantly decreased, mDC and TAM (M1 type) were up-regulated, and CD8+ T cells were recruited to infiltrate tumors, in contrast to PBS control and DC-T monotherapy. DC-T cells combined with endostatin potently reduced the expressions of IL-6, IL-10, TGF-β and

  4. Apoptosis-Inducing Activity of Marine Sponge Haliclona sp. Extracts Collected from Kosrae in Nonsmall Cell Lung Cancer A549 Cells.

    PubMed

    Bae, Woori; Lim, Hyun Kyung; Kim, Kyoung Mee; Cho, Hyosun; Lee, Sun Yi; Jeong, Choon-Sik; Lee, Hyi-Seung; Jung, Joohee

    2015-01-01

    Although various anticancer drugs have been developed for the treatment of nonsmall cell lung cancer, chemotherapeutic efficacy is still limited. Natural products such as phytochemicals have been screened as novel alternative materials, but alternative funds such as marine bioresources remain largely untapped. Of these resources, marine sponges have undergone the most scrutiny for their biological activities, including antiinflammatory, antiviral, and anticancer properties. However, the biological mechanisms of the activities of these marine sponges are still unclear. We investigated the anticancer activity of marine sponges collected from Kosrae in Micronesia and examined their mechanisms of action using nonsmall cell lung cancer A549 cells as a model system. Of 20 specimens, the Haliclona sp. (KO1304-328) showed both dose- and time-dependent cytotoxicity. Further, methanol extracts of Haliclona sp. significantly inhibited cell proliferation and cell viability. A549 cells treated with Haliclona sp. demonstrated induced expression of c-Jun N-terminal kinase (JNK), p53, p21, caspase-8, and caspase-3. The percentage of apoptotic cells significantly increased in A549 cultures treated with Haliclona sp. These results indicate that Haliclona sp. induces apoptosis via the JNK-p53 pathway and caspase-8, suggesting that this marine sponge is a good resource for the development of drugs for treatment of nonsmall cell lung cancer. PMID:26236382

  5. Deactivation of A549 cancer cells in vitro by a dielectric barrier discharge plasma needle

    SciTech Connect

    Huang Jun; Chen Wei; Li Hui; Wang Xingquan; Lv Guohua; Wang Pengye; Khohsa, M. Latif; Guo Ming; Feng Kecheng; Yang Size

    2011-03-01

    An inactivation mechanism study on A549 cancer cells by means of a dielectric barrier discharge plasma needle is presented. The neutral red uptake assay provides a quantitative estimation of cell viability after plasma treatment. Experimental results show that the efficiency of argon plasma for the inactivation process is very dependent on power and treatment time. A 27 W power and 120 s treatment time along with 900 standard cubic centimeter per minute Ar flow and a nozzle-to-sample separation of 3 mm are the best parameters of the process. According to the argon emission spectra of the plasma jet and the optical microscope images of the A549 cells after plasma treatment, it is concluded that the reactive species (for example, OH and O) in the argon plasma play a major role in the cell deactivation.

  6. Upregulation of AQP3 and AQP5 induced by dexamethasone and ambroxol in A549 cells.

    PubMed

    Ben, Yong; Chen, Jie; Zhu, Rong; Gao, Lei; Bai, Chunxue

    2008-04-30

    Aquaporins (AQPs) are membrane channel proteins that play roles in the regulation of water permeability in many tissues. AQP1 and AQP5 expressed in lung provide the principal route for osmotically driven water transport. In the airways, AQP3 and AQP4 facilitate water transport. Dexamethasone and ambroxol are often used to treat patients with pulmonary diseases accompanied by airway hypersecretion. The role of AQPs in these effective treatments has not been addressed. In this study, we analyzed the expression of AQPs in a human airway epithelial cell line (A549 cells) and showed that AQP3 and 5, but not AQP1 and 4, were expressed in A549 cells. Both dexamethasone and ambroxol stimulated the expression of AQP3 and 5 at the mRNA and protein levels. The data suggest potential roles of AQP3 and 5 in the regulation of airway hypersecretion, perhaps ultimately providing a target for treating such diseases.

  7. G4-Tetra DNA Duplex Induce Lung Cancer Cell Apoptosis in A549 Cells

    NASA Astrophysics Data System (ADS)

    Xu, Xiaobo; Zhao, YiZhuo; Lu, Hu; Fu, Cuiping; Li, Xiao; Jiang, Liyan; Li, Shanqun

    2016-10-01

    The specific DNA is typically impermeable to the plasma membrane due to its natural characters, but DNA tetra structures (DTNs) can be readily uptake by cells in the absence of transfection agents, providing a new strategy to deliver DNA drugs. In this research, the delivery efficiency of tetrahedral DNA nanostructures was measured on adenocarcinomic human alveolar basal epithelial (A549) cells via delivering AS1411 (G4). The DNA tetra-AS1411 complex was rapidly and abundantly uptake by A549 cells, and the induced apoptosis was enhanced. Furthermore, biodistribution in mouse proved the rapid clearance from non-targeted organs in vivo. This study improved the understanding of potential function in DNA-based drug delivery and proved that DTNs-AS1411 could be potentially useful for the treatment of lung cancer.

  8. Brusatol Enhances the Radiosensitivity of A549 Cells by Promoting ROS Production and Enhancing DNA Damage

    PubMed Central

    Sun, Xiaohui; Wang, Qin; Wang, Yan; Du, Liqing; Xu, Chang; Liu, Qiang

    2016-01-01

    NF-E2-related factor 2 (Nrf2) has been identified as a master regulatory factor in the protection of cells from oxidative and electrophilic stress. However, overexpression of Nrf2 in lung cancer may cause chemoresistance, as well as radioresistance. In this study, we examined the relationship between radioresistance and Nrf2 protein levels in H1299, A549, and H460 cells, and finally chose the A549 cell line to continue with due to its strong radioresistance and high Nrf2 protein levels. We found that the Nrf2 inhibitor, brusatol, could prevent the increase and accumulation of Nrf2 after exposure to irradiation. Additionally, following treatment with 80 nM brusatol, A549 cells became sensitive to irradiation, suffering severe DNA damage. Combination treatment with brusatol and ionizing radiation (IR) can distinctly increase the level of reactive oxygen species in A549 cells, causing a 1.8-fold increase compared with the control, and a 1.4-fold increase compared with IR alone. In fact, in the treatment with both brusatol and IR, lung cancer cell proliferation is halted, gradually leading to cell death. Because Nrf2 is closely linked to DNA damage repair, inhibiting the function of Nrf2, as in brusatol treatment, may increase the DNA damage caused by radiotherapy or chemotherapy, possibly enhancing the efficacy of chemotherapeutic drugs. Our study is the first to demonstrate brusatol’s ability to enhance the responsiveness of lung cancer cells to irradiation, and its potential application as a natural sensitizer in radiotherapy. PMID:27347930

  9. Sinomenine inhibits A549 human lung cancer cell invasion by mediating the STAT3 signaling pathway

    PubMed Central

    Jiang, Shulong; Gao, Yebo; Hou, Wei; Liu, Rui; Qi, Xin; Xu, Xia; Li, Jie; Bao, Yanju; Zheng, Honggang; Hua, Baojin

    2016-01-01

    Increasing evidence suggests that the failure of lung cancer treatment may occur as a result of tumor invasion and metastasis. Signal transducer and activator of transcription 3 (STAT3), an epithelial-mesenchymal transition-inducing transcription factor, is a key signaling molecule involved in the proliferation, apoptosis, invasion and metastasis of tumor cells. Sinomenine is an alkaloid compound with an antineoplastic potential against a variety of cancer cells. The aim of the present study was to assess the antitumor mechanisms of sinomenine in the A549 human lung cancer cell line. The results demonstrated that sinomenine manifested dose-dependent cytotoxicity and induced apoptosis in A549 cells. The protein expression of Janus kinase 2, STAT3, phosphorylated-STAT3, Snail, N-cadherin and vimentin decreased in sinomenine-treated cells, while E-cadherin protein expression increased. The regulation of STAT3, N-cadherin and E-cadherin by sinomenine was further confirmed by reverse transcription-quantitative polymerase chain reaction and immunofluorescent staining. It was demonstrated that sinomenine exerts inhibitory effects on A549 human lung cancer cell invasion, possibly through the inhibition of STAT3 signaling. These results provide a novel insight into the role of sinomenine in the treatment of non-small cell lung cancer. PMID:27446441

  10. Wnt/{beta}-catenin signaling regulates cancer stem cells in lung cancer A549 cells

    SciTech Connect

    Teng, Ying; Wang, Xiuwen; Wang, Yawei; Ma, Daoxin

    2010-02-12

    Wnt/{beta}-catenin signaling plays an important role not only in cancer, but also in cancer stem cells. In this study, we found that {beta}-catenin and OCT-4 was highly expressed in cisplatin (DDP) selected A549 cells. Stimulating A549 cells with lithium chloride (LiCl) resulted in accumulation of {beta}-catenin and up-regulation of a typical Wnt target gene cyclin D1. This stimulation also significantly enhanced proliferation, clone formation, migration and drug resistance abilities in A549 cells. Moreover, the up-regulation of OCT-4, a stem cell marker, was observed through real-time PCR and Western blotting. In a reverse approach, we inhibited Wnt signaling by knocking down the expression of {beta}-catenin using RNA interference technology. This inhibition resulted in down-regulation of the Wnt target gene cyclin D1 as well as the proliferation, clone formation, migration and drug resistance abilities. Meanwhile, the expression of OCT-4 was reduced after the inhibition of Wnt/{beta}-catenin signaling. Taken together, our study provides strong evidence that canonical Wnt signaling plays an important role in lung cancer stem cell properties, and it also regulates OCT-4, a lung cancer stem cell marker.

  11. Selective killing effect of oxytetracycline, propafenone and metamizole on A549 or Hela cells

    PubMed Central

    Feng, Guihua

    2013-01-01

    Objective To determine the selective killing effect of oxytetracycline, propafenone and metamizole on A549 or Hela cells. Methods Proliferation assay, lactate dehydrogenase (LDH) assay, apoptosis detecting, flow cytometry and western blot were performed. Results It was found that treatment with propafenone at the concentration of 0.014 g/L or higher for 48 h could induce apoptosis in Hela cells greatly, while it was not observed in oxytetracycline and metamizole at the concentration of 0.20 g/L for 48 h. Oxytetracycline, propafenone and metamizole all displayed evident inhibitory effects on the proliferation of A549 cells. The results of LDH assay demonstrated that the drugs at the test range of concentration did not cause necrosis in the cells. Propafenone could elevate the protein level of P53 effectively (P<0.01). Conclusions Oxytetracycline, propafenone and metamizol (dipyrone) all displayed evident inhibitory effects on the proliferation of A549 cells. Propafenone also displayed evident inhibitory effects on the proliferation of Hela cells. PMID:24385693

  12. Safrole oxide induces apoptosis in A549 human lung cancer cells.

    PubMed

    Du, Aiying; Zhang, Shangli; Miao, Junying; Zhao, Baoxiang

    2004-09-01

    3,4-(Methylenedioxy)-1-(2',3'-epoxypropyl)-benzene (safrole oxide) was synthesized in the authors' laboratory. To investigate the effects of safrole oxide on the growth and apoptosis of A549 human lung cancer cells, the authors treated the cells with safrole oxide, 112.36 to 449.44 micromol/L, for 24 to 48 hours. The results showed that the drug led A549 cells to apoptosis and blocked cell cycle completely at G1 phase and partly at G(2)-M phase. To further study the correlated mechanism, the authors examined P53 and H-Ras protein expressions by using immunofluorescence assay. They found that the expression of P53 was dramatically up-regulated but the expression of H-Ras was hardly affected by safrole oxide, 224.72 micromol/L, within 24 hours. Taken together, these results revealed that safrole oxide could induce apoptosis of A549 cells and suggested that safrole oxide might perform its function by blocking cells completely at G1 phase and partly at G(2)-M phase, and also by up-regulating the expression of P53 protein. These findings would raise exciting possibilities for cancer therapy in future.

  13. PVM/MA-shelled selol nanocapsules promote cell cycle arrest in A549 lung adenocarcinoma cells

    PubMed Central

    2014-01-01

    Background Selol is an oily mixture of selenitetriacylglycerides that was obtained as a semi-synthetic compound containing selenite. Selol is effective against cancerous cells and less toxic to normal cells compared with inorganic forms of selenite. However, Selol’s hydrophobicity hinders its administration in vivo. Therefore, the present study aimed to produce a formulation of Selol nanocapsules (SPN) and to test its effectiveness against pulmonary adenocarcinoma cells (A549). Results Nanocapsules were produced through an interfacial nanoprecipitation method. The polymer shell was composed of poly(methyl vinyl ether-co-maleic anhydride) (PVM/MA) copolymer. The obtained nanocapsules were monodisperse and stable. Both free Selol (S) and SPN reduced the viability of A549 cells, whereas S induced a greater reduction in non-tumor cell viability than SPN. The suppressor effect of SPN was primarily associated to the G2/M arrest of the cell cycle, as was corroborated by the down-regulations of the CCNB1 and CDC25C genes. Apoptosis and necrosis were induced by Selol in a discrete percentage of A549 cells. SPN also increased the production of reactive oxygen species, leading to oxidative cellular damage and to the overexpression of the GPX1, CYP1A1, BAX and BCL2 genes. Conclusions This study presents a stable formulation of PVM/MA-shelled Selol nanocapsules and provides the first demonstration that Selol promotes G2/M arrest in cancerous cells. PMID:25149827

  14. MicroRNA-1228(*) inhibit apoptosis in A549 cells exposed to fine particulate matter.

    PubMed

    Li, Xiaobo; Ding, Zhen; Zhang, Chengcheng; Zhang, Xin; Meng, Qingtao; Wu, Shenshen; Wang, Shizhi; Yin, Lihong; Pu, Yuepu; Chen, Rui

    2016-05-01

    Studies have reported associations between fine particulate matter (PM2.5) and respiratory disorders; however, the underlying mechanism is not completely clear owing to the complex components of PM2.5. microRNAs (miRNAs) demonstrate tremendous regulation to target genes, which are sensitive to exogenous stimulation, and facilitate the integrative understood of biological responses. Here, significantly modulated miRNA were profiled by miRNA microarray, coupled with bioinformatic analysis; the potential biological function of modulated miRNA were predicted and subsequently validated by cell-based assays. Downregulation of miR-1228-5p (miR-1228(*)) expression in human A549 cells were associated with PM2.5-induced cellular apoptosis through a mitochondria-dependent pathway. Further, overexpression of miR-1228(*) rescued the cellular damages induced by PM2.5. Thus, our results demonstrate that PM2.5-induced A549 apoptosis is initiated by mitochondrial dysfunction and miR-1228(*) could protect A549 cells against apoptosis. The involved pathways and target genes might be used for future mechanistic studies.

  15. Dielectric barrier discharge plasma in Ar/O{sub 2} promoting apoptosis behavior in A549 cancer cells

    SciTech Connect

    Huang Jun; Li Hui; Chen Wei; Lv Guohua; Wang Xingquan; Zhang Guoping; Wang Pengye; Ostrikov, Kostya; Yang Size

    2011-12-19

    The Ar/O{sub 2} plasma needle in the induction of A549 cancer cells apoptosis process is studied by means of real-time observation. The entire process of programmed cell death is observed. The typical morphological changes of A549 apoptosis are detected by 4', 6-diamidino-2-phenylindole staining, for example, chromatin condensation and nuclear fragmentation. Cell viability is determined and quantified by neutral red uptake assay, and the survival rate of A549 from Ar/O{sub 2} plasmas is presented. Further spectral analysis indicates the reactive species, including O and OH play crucial roles in the cell inactivation.

  16. The soluble EP2 receptor FuEP2/Ex2 suppresses endometrial cancer cell growth in an orthotopic xenograft model in nude mice.

    PubMed

    Takahashi, Tetsuyuki; Ogawa, Hirohisa; Izumi, Keisuke; Uehara, Hisanori

    2011-07-01

    Endometrial cancer is one of the most common gynecologic malignancies and many factors influence in its growth and development. As in many other types of cancer, prostaglandin E(2) (PGE(2)) is thought to be an accelerator of cell proliferation and endometrial cancer progression. In this study, we examined the effect of FuEP2/Ex2, a soluble decoy receptor for PGE(2) on growth of endometrial cancer cells. A stable transfectant expressing FuEP2/Ex2 was established from human endometrial cancer Ishikawa cells (Ish-FuEP2/Ex2). Ish-FuEP2/Ex2 cells expressed FuEP2/Ex2 mRNA and protein. Expression levels of E-prostanoid receptor 1 (EP1), EP2, EP3, EP4, and F-prostanoid receptor (FP) were almost the same in Ish-FuEP2/Ex2 and vector control cells. Growth rates of Ish-FuEP2/Ex2 under normal culture conditions were also similar to vector control cells, although PGE(2)-induced growth stimulation was completely inhibited in Ish-FuEP2/Ex2 or by Ish-FuEP2/Ex2 culture medium. Moreover, phosphorylation of extracellular signal-regulated kinase (ERK) and induction of cyclooxygenase-2 (COX-2), vascular endothelial growth factor (VEGF), cyclin D1, and c-fos mRNA by PGE(2) were not observed in Ish-FuEP2/Ex2 and Ish-FuEP2/Ex2 culture medium-treated vector control cells, although they were found when treated with prostaglandin F(2α). An orthotopic xenograft model in athymic nude mice revealed that Ish-FuEP2/Ex2-injected mice had significantly decreased mean tumor area. The proportion of Ki-67-positive cells in the tumor lesion was also significantly lower in Ish-FuEP2/Ex2-injected mice. These findings suggest that an EP-targeting strategy using FuEP2/Ex2 may be of use in the treatment of endometrial cancer.

  17. Anti-CCR4 monoclonal antibody enhances antitumor immunity by modulating tumor-infiltrating Tregs in an ovarian cancer xenograft humanized mouse model

    PubMed Central

    Chang, De-Kuan; Peterson, Eric; Sun, Jiusong; Goudie, Calum; Drapkin, Ronny I.; Liu, Joyce F.; Matulonis, Ursula; Zhu, Quan; Marasco, Wayne A.

    2016-01-01

    ABSTRACT Recent studies have demonstrated that regulatory T cells (Tregs) are recruited to tumor sites where they can suppress antitumor immunity. The chemokine receptor CCR4 is expressed at high levels on functional CD4+CD25+FoxP3+ Tregs and production of the CCR4 ligand CCL22 by tumor cells and tumor-associated macrophages is associated with Treg recruitment to the tumor site. Here, we tested IgG1 and IgG4 isotypes of human anti-CCR4 mAb2-3 for their in vitro activity and in vivo capacity in a NSG mouse model bearing CCL22-secreting ovarian cancer (OvCA) xenograft to modulate Tregs and restore antitumor activity. Both mAb2-3 isotypes blocked in vitro chemoattraction of Tregs to CCL22-secreting OvCA cells. However, they differed in their in vivo mode of action with IgG1 causing Treg depletion and IgG4 blocking migration to the tumors. Primary T cells that were primed with OvCA-pulsed dendritic cells (DCs) demonstrated INFγ secretion that could be enhanced through Treg depletion by mAb2-3. Humanized mice reconstructed with allogeneic tumor-primed T cells (TP-T) were used to evaluate the restoration of OvCA immunity by depletion or blockade of Tregs with mAb2-3. We observed that IgG1 was more potent than IgG4 in inhibiting tumor growth. Mechanism studies demonstrated that mAb2-3 treatment lead to inhibition of IL-2 binding to its receptor. Further studies showed that mAb2-3 induced CD25 shedding (sCD25) from Tregs which lead to a decrease in IL-2-dependent survival. Together, the results demonstrate that mAb2-3 is an agonist antibody that can restore anti-OvCA immunity through modulation of Treg activity. PMID:27141347

  18. Analysis of the Lipidome of Xenografts Using MALDI-IMS and UHPLC-ESI-QTOF

    NASA Astrophysics Data System (ADS)

    Fernández, Roberto; Lage, Sergio; Abad-García, Beatriz; Barceló-Coblijn, Gwendolyn; Terés, Silvia; López, Daniel H.; Guardiola-Serrano, Francisca; Martín, M. Laura; Escribá, Pablo V.; Fernández, José A.

    2014-07-01

    Human tumor xenografts in immunodeficient mice are a very popular model to study the development of cancer and to test new drug candidates. Among the parameters analyzed are the variations in the lipid composition, as they are good indicators of changes in the cellular metabolism. Here, we present a study on the distribution of lipids in xenografts of NCI-H1975 human lung cancer cells, using MALDI imaging mass spectrometry and UHPLC-ESI-QTOF. The identification of lipids directly from the tissue by MALDI was aided by the comparison with identification using ESI ionization in lipid extracts from the same xenografts. Lipids belonging to PCs, PIs, SMs, DAG, TAG, PS, PA, and PG classes were identified and their distribution over the xenograft was determined. Three areas were identified in the xenograft, corresponding to cells in different metabolic stages and to a layer of adipose tissue that covers the xenograft.

  19. Physiologically based pharmacokinetic and pharmacodynamic modeling of an antagonist (SM-406/AT-406) of multiple inhibitor of apoptosis proteins (IAPs) in a mouse xenograft model of human breast cancer.

    PubMed

    Zhang, Tao; Li, Yanyan; Zou, Peng; Yu, Jing-yu; McEachern, Donna; Wang, Shaomeng; Sun, Duxin

    2013-09-01

    The inhibitors of apoptosis proteins (IAPs) are a class of key apoptosis regulators overexpressed or dysregulated in cancer. SM-406/AT-406 is a potent and selective small molecule mimetic of Smac that antagonizes the inhibitor of apoptosis proteins (IAPs). A physiologically based pharmacokinetic and pharmacodynamic (PBPK-PD) model was developed to predict the tissue concentration-time profiles of SM-406, the related onco-protein levels in tumor, and the tumor growth inhibition in a mouse model bearing human breast cancer xenograft. In the whole body physiologically based pharmacokinetic (PBPK) model for pharmacokinetics characterization, a well stirred (perfusion rate-limited) model was used to describe SM-406 pharmacokinetics in the lung, heart, kidney, intestine, liver and spleen, and a diffusion rate-limited (permeability limited) model was used for tumor. Pharmacodynamic (PD) models were developed to correlate the SM-406 concentration in tumor to the cIAP1 degradation, pro-caspase 8 decrease, CL-PARP accumulation and tumor growth inhibition. The PBPK-PD model well described the experimental pharmacokinetic data, the pharmacodynamic biomarker responses and tumor growth. This model may be helpful to predict tumor and plasma SM-406 concentrations in the clinic.

  20. Burkholderia pseudomallei Biofilm Promotes Adhesion, Internalization and Stimulates Proinflammatory Cytokines in Human Epithelial A549 Cells.

    PubMed

    Kunyanee, Chanikarn; Kamjumphol, Watcharaporn; Taweechaisupapong, Suwimol; Kanthawong, Sakawrat; Wongwajana, Suwin; Wongratanacheewin, Surasak; Hahnvajanawong, Chariya; Chareonsudjai, Sorujsiri

    2016-01-01

    Burkholderia pseudomallei is a Gram-negative bacterium that causes melioidosis. Inhalational exposure leading to pulmonary melioidosis is the most common clinical manifestation with significant mortality. However, the role of B. pseudomallei biofilm phenotype during bacterial-host interaction remains unclear. We hypothesize that biofilm phenotype may play a role in such interactions. In this study, B. pseudomallei H777 (biofilm wild type), B. pseudomallei M10 (biofilm mutant) and B. pseudomallei C17 (biofilm-complemented) strains were used to assess the contribution of biofilm to adhesion to human lung epithelial cells (A549), intracellular interactions, apoptosis/necrosis and impact on proinflammatory responses. Confocal laser scanning microscopy demonstrated that B. pseudomallei H777 and C17 produced biofilm, whereas M10 did not. To determine the role of biofilm in host interaction, we assessed the ability of each of the three strains to interact with the A549 cells at MOI 10. Strain H777 exhibited higher levels of attachment and invasion compared to strain M10 (p < 0.05). In addition, the biofilm-complemented strain, C17 exhibited restored bacterial invasion ability. Flow cytometry combined with a double-staining assay using annexin V and propidium iodide revealed significantly higher numbers of early apoptotic and late apoptotic A549 cells when these were infected with strain H777 (1.52%) and C17 (1.43%) compared to strain M10 (0.85%) (p < 0.05). Strains H777 and C17 were able to stimulate significant secretion of IL-6 and IL-8 compared with the biofilm mutant (p < 0.05). Together, these findings demonstrated the role of biofilm-associated phenotypes of B. pseudomallei in cellular pathogenesis of human lung epithelial cells with respect to initial attachment and invasion, apoptosis and proinflammatory responses. PMID:27529172

  1. Burkholderia pseudomallei Biofilm Promotes Adhesion, Internalization and Stimulates Proinflammatory Cytokines in Human Epithelial A549 Cells

    PubMed Central

    Kunyanee, Chanikarn; Kamjumphol, Watcharaporn; Taweechaisupapong, Suwimol; Kanthawong, Sakawrat; Wongwajana, Suwin; Wongratanacheewin, Surasak; Hahnvajanawong, Chariya

    2016-01-01

    Burkholderia pseudomallei is a Gram-negative bacterium that causes melioidosis. Inhalational exposure leading to pulmonary melioidosis is the most common clinical manifestation with significant mortality. However, the role of B. pseudomallei biofilm phenotype during bacterial-host interaction remains unclear. We hypothesize that biofilm phenotype may play a role in such interactions. In this study, B. pseudomallei H777 (biofilm wild type), B. pseudomallei M10 (biofilm mutant) and B. pseudomallei C17 (biofilm-complemented) strains were used to assess the contribution of biofilm to adhesion to human lung epithelial cells (A549), intracellular interactions, apoptosis/necrosis and impact on proinflammatory responses. Confocal laser scanning microscopy demonstrated that B. pseudomallei H777 and C17 produced biofilm, whereas M10 did not. To determine the role of biofilm in host interaction, we assessed the ability of each of the three strains to interact with the A549 cells at MOI 10. Strain H777 exhibited higher levels of attachment and invasion compared to strain M10 (p < 0.05). In addition, the biofilm-complemented strain, C17 exhibited restored bacterial invasion ability. Flow cytometry combined with a double-staining assay using annexin V and propidium iodide revealed significantly higher numbers of early apoptotic and late apoptotic A549 cells when these were infected with strain H777 (1.52%) and C17 (1.43%) compared to strain M10 (0.85%) (p < 0.05). Strains H777 and C17 were able to stimulate significant secretion of IL-6 and IL-8 compared with the biofilm mutant (p < 0.05). Together, these findings demonstrated the role of biofilm-associated phenotypes of B. pseudomallei in cellular pathogenesis of human lung epithelial cells with respect to initial attachment and invasion, apoptosis and proinflammatory responses. PMID:27529172

  2. Silencing BMP-2 expression inhibits A549 and H460 cell proliferation and migration

    PubMed Central

    2014-01-01

    Abstract Background Bone morphogenetic protein 2 (BMP-2) is a member of the TGF-β superfamily that is closely correlated with many malignancies, particularly lung cancer. However, the effects of silenced BMP-2 on lung cancer cell proliferation and migration are not clear. Methods Using quantitative real-time RT-PCR, BMP-2 mRNA expression was detected in 61 non-small cell lung cancer (NSCLC) samples. Survival curves were generated using follow-up data. Relationships between clinical or pathological characteristics and prognosis were analyzed. Cell viability assays and transwell migration assays were used to evaluate the effects of BMP-2 silencing on cell proliferation and migration of A549 and H460 cells. Results BMP-2 mRNA expression was higher in NSCLC tissues compared to matched adjacent normal tissues (P < 0.01). High BMP-2 expression levels were significantly associated with the occurrence of lymph node metastases and tumor stage (P < 0.05). There were significant differences in survival curves between groups with metastatic lymph nodes and non-metastatic lymph nodes, as well as between groups with low BMP-2 expression and groups with high BMP-2 expression. In addition, we observed decreased proliferation and migration rates of the NSCLC-derived cell lines A549 and H460 that were transfected with siBMP-2 (P < 0.05). Conclusion BMP-2 mRNA is overexpressed in NSCLC samples and is a risk factor for survival in patients with NSCLC. BMP-2 silencing can significantly inhibit A549 and H460 cell proliferation and migration. Virtual slides The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/4263254471298866 PMID:24946687

  3. Use of a Conformational Switching Aptamer for Rapid and Specific Ex Vivo Identification of Central Nervous System Lymphoma in a Xenograft Model

    PubMed Central

    Eschbacher, Jennifer; Nichols, Joshua; Mooney, Michael A.; Joy, Anna; Spetzler, Robert F.; Feuerstein, Burt G.; Preul, Mark C.; Anderson, Trent; Yan, Hao; Nakaji, Peter

    2015-01-01

    Improved tools for providing specific intraoperative diagnoses could improve patient care. In neurosurgery, intraoperatively differentiating non-operative lesions such as CNS B-cell lymphoma from operative lesions can be challenging, often necessitating immunohistochemical (IHC) procedures which require up to 24-48 hours. Here, we evaluate the feasibility of generating rapid ex vivo specific labeling using a novel lymphoma-specific fluorescent switchable aptamer. Our B-cell lymphoma-specific switchable aptamer produced only low-level fluorescence in its unbound conformation and generated an 8-fold increase in fluorescence once bound to its target on CD20-positive lymphoma cells. The aptamer demonstrated strong binding to B-cell lymphoma cells within 15 minutes of incubation as observed by flow cytometry. We applied the switchable aptamer to ex vivo xenograft tissue harboring B-cell lymphoma and astrocytoma, and within one hour specific visual identification of lymphoma was routinely possible. In this proof-of-concept study in human cell culture and orthotopic xenografts, we conclude that a fluorescent switchable aptamer can provide rapid and specific labeling of B-cell lymphoma, and that developing aptamer-based labeling approaches could simplify tissue staining and drastically reduce time to histopathological diagnoses compared with IHC-based methods. We propose that switchable aptamers could enhance expeditious, accurate intraoperative decision-making. PMID:25876071

  4. Artesunate suppresses tumor growth and induces apoptosis through the modulation of multiple oncogenic cascades in a chronic myeloid leukemia xenograft mouse model

    PubMed Central

    Kim, Chulwon; Lee, Jong Hyun; Kim, Sung-Hoon; Sethi, Gautam; Ahn, Kwang Seok

    2015-01-01

    Artesunate (ART), a semi-synthetic derivative of artemisinin, is one of the most commonly used anti-malarial drugs. Also, ART possesses anticancer potential albeit through incompletely understood molecular mechanism(s). Here, the effect of ART on various protein kinases, associated gene products, cellular response, and apoptosis was investigated. The in vivo effect of ART on the growth of human CML xenograft tumors in athymic nu/nu mice was also examined. In our preliminary experiments, we first observed that phosphorylation of p38, ERK, CREB, Chk-2, STAT5, and RSK proteins were suppressed upon ART exposure. Interestingly, ART induced the expression of SOCS-1 protein and depletion of SOCS-1 using siRNA abrogated the STAT5 inhibitory effect of the drug. Also various dephosphorylations caused by ART led to the suppression of various survival gene products and induced apoptosis through caspase-3 activation. Moreover, ART also substantially potentiated the apoptosis induced by chemotherapeutic agents. Finally, when administered intraperitoneally, ART inhibited p38, ERK, STAT5, and CREB activation in tumor tissues and the growth of human CML xenograft tumors in mice without exhibiting any significant adverse effects. Overall, our results suggest that ART exerts its anti-proliferative and pro-apoptotic effects through suppression of multiple signaling cascades in CML both in vitro and in vivo. PMID:25738364

  5. Use of a conformational switching aptamer for rapid and specific ex vivo identification of central nervous system lymphoma in a xenograft model.

    PubMed

    Georges, Joseph F; Liu, Xiaowei; Eschbacher, Jennifer; Nichols, Joshua; Mooney, Michael A; Joy, Anna; Spetzler, Robert F; Feuerstein, Burt G; Preul, Mark C; Anderson, Trent; Yan, Hao; Nakaji, Peter

    2015-01-01

    Improved tools for providing specific intraoperative diagnoses could improve patient care. In neurosurgery, intraoperatively differentiating non-operative lesions such as CNS B-cell lymphoma from operative lesions can be challenging, often necessitating immunohistochemical (IHC) procedures which require up to 24-48 hours. Here, we evaluate the feasibility of generating rapid ex vivo specific labeling using a novel lymphoma-specific fluorescent switchable aptamer. Our B-cell lymphoma-specific switchable aptamer produced only low-level fluorescence in its unbound conformation and generated an 8-fold increase in fluorescence once bound to its target on CD20-positive lymphoma cells. The aptamer demonstrated strong binding to B-cell lymphoma cells within 15 minutes of incubation as observed by flow cytometry. We applied the switchable aptamer to ex vivo xenograft tissue harboring B-cell lymphoma and astrocytoma, and within one hour specific visual identification of lymphoma was routinely possible. In this proof-of-concept study in human cell culture and orthotopic xenografts, we conclude that a fluorescent switchable aptamer can provide rapid and specific labeling of B-cell lymphoma, and that developing aptamer-based labeling approaches could simplify tissue staining and drastically reduce time to histopathological diagnoses compared with IHC-based methods. We propose that switchable aptamers could enhance expeditious, accurate intraoperative decision-making. PMID:25876071

  6. Identification of Biomarkers of Necrosis in Xenografts Using Imaging Mass Spectrometry

    NASA Astrophysics Data System (ADS)

    Fernández, Roberto; Garate, Jone; Lage, Sergio; Terés, Silvia; Higuera, Mónica; Bestard-Escalas, Joan; López, Daniel H.; Guardiola-Serrano, Francisca; Escribá, Pablo V.; Barceló-Coblijn, Gwendolyn; Fernández, José A.

    2016-02-01

    Xenografts are commonly used to test the effect of new drugs on human cancer. However, because of their heterogeneity, analysis of the results is often controversial. Part of the problem originates in the existence of tumor cells at different metabolic stages: from metastatic to necrotic cells, as it happens in real tumors. Imaging mass spectrometry is an excellent solution for the analysis of the results as it yields detailed information not only on the composition of the tissue but also on the distribution of the biomolecules within the tissue. Here, we use imaging mass spectrometry to determine the distribution of phosphatidylcholine (PC), phosphatidylethanolamine (PE), and their plasmanyl- and plasmenylether derivatives (PC-P/O and PE-P/O) in xenografts of five different tumor cell lines: A-549, NCI-H1975, BX-PC3, HT29, and U-87 MG. The results demonstrate that the necrotic areas showed a higher abundance of Na+ adducts and of PC-P/O species, whereas a large abundance of PE-P/O species was found in all the xenografts. Thus, the PC/PC-ether and Na+/K+ ratios may highlight the necrotic areas while an increase on the number of PE-ether species may be pointing to the existence of viable tumor tissues. Furthermore, the existence of important changes in the concentration of Na+ and K+ adducts between different tissues has to be taken into account while interpreting the imaging mass spectrometry results.

  7. Sensitivity of A-549 human lung cancer cells to nanoporous zinc oxide conjugated with Photofrin.

    PubMed

    Fakhar-e-Alam, Muhammad; Ali, Syed Muhammad Usman; Ibupoto, Zafar Hussain; Kimleang, Khun; Atif, M; Kashif, Muhammad; Loong, Foo Kai; Hashim, Uda; Willander, Magnus

    2012-05-01

    In the present study, we demonstrated the use of nanoporous zinc oxide (ZnO NPs) in photodynamic therapy. The ZnO NPs structure possesses a high surface to volume ratio due to its porosity and ZnO NPs can be used as an efficient photosensitizer carrier system. We were able to grow ZnO NPs on the tip of borosilicate glass capillaries (0.5 μm diameter) and conjugated this with Photofrin for efficient intracellular drug delivery. The ZnO NPs on the capillary tip could be excited intracellularly with 240 nm UV light, and the resultant 625 nm red light emitted in the presence of Photofrin activated a chemical reaction that produced reactive oxygen species (ROS). The procedure was tested in A-549 cells and led to cell death within a few minutes. The morphological changes in necrosed cells were examined by microscopy. The viability of control and treated A-549 cells with the optimum dose of UV/visible light was assessed using the MTT assay, and ROS were detected using a fluorescence microscopy procedure.

  8. Exendin-4 promotes extracellular-superoxide dismutase expression in A549 cells through DNA demethylation

    PubMed Central

    Yasuda, Hiroyuki; Mizukami, Koji; Hayashi, Mutsuna; Kamiya, Tetsuro; Hara, Hirokazu; Adachi, Tetsuo

    2016-01-01

    Exendin-4 is an agonist of the glucagon-like peptide 1 receptor (GLP-1R) and is used in the treatment of type 2 diabetes. Since human GLP-1R has been identified in various cells besides pancreatic cells, exendin-4 is expected to exert extrapancreatic actions. It has also been suggested to affect gene expression through epigenetic regulation, such as DNA methylation and/or histone modifications. Furthermore, the expression of extracellular-superoxide dismutase (EC-SOD), a major SOD isozyme that is crucially involved in redox homeostasis, is regulated by epigenetic factors. In the present study, we demonstrated that exendin-4 induced the demethylation of DNA in A549 cells, which, in turn, affected the expression of EC-SOD. Our results showed that the treatment with exendin-4 up-regulated the expression of EC-SOD through GLP-1R and demethylated some methyl-CpG sites (methylated cytosine at 5'-CG-3') in the EC-SOD gene. Moreover, the treatment with exendin-4 inactivated DNA methyltransferases (DNMTs), but did not change their expression levels. In conclusion, the results of the present study demonstrated for the first time that exendin-4 regulated the expression of EC-SOD by reducing the activity of DNMTs and demethylation of DNA within the EC-SOD promoter region in A549 cells. PMID:26798195

  9. COPD promotes migration of A549 lung cancer cells: the role of chemokine CCL21

    PubMed Central

    Kuźnar-Kamińska, Barbara; Mikuła-Pietrasik, Justyna; Sosińska, Patrycja; Książek, Krzysztof; Batura-Gabryel, Halina

    2016-01-01

    Patients with COPD develop lung cancer more frequently than healthy smokers. At the same time, molecular mediators promoting various aspects of cancer cell progression are still elusive. In this report, we examined whether COPD can be coupled with increased migration of non-small-cell lung cancer cells A549 and, if so, whether this effect may be related to altered production and activity of chemokines CCL21, CXCL5, and CXCL12. The study showed that the migration of A549 cells through the polycarbonate membrane and basement membrane extract toward a chemotactic gradient elicited by serum from patients with COPD was markedly higher as compared with serum from healthy donors. The concentration of CCL21 and CXCL12, but not CXCL5, in serum from patients with COPD was also increased. Experiments in which CCL21- and CXCL12-dependent signaling was blocked revealed that increased migration of the cancer cells upon treatment with serum from patients with COPD was mediated exclusively by CCL21. Collectively, our results indicate that COPD may contribute to the progression of lung cancer via CCL21-dependent intensification of cancer cell migration. PMID:27307721

  10. COPD promotes migration of A549 lung cancer cells: the role of chemokine CCL21.

    PubMed

    Kuźnar-Kamińska, Barbara; Mikuła-Pietrasik, Justyna; Sosińska, Patrycja; Książek, Krzysztof; Batura-Gabryel, Halina

    2016-01-01

    Patients with COPD develop lung cancer more frequently than healthy smokers. At the same time, molecular mediators promoting various aspects of cancer cell progression are still elusive. In this report, we examined whether COPD can be coupled with increased migration of non-small-cell lung cancer cells A549 and, if so, whether this effect may be related to altered production and activity of chemokines CCL21, CXCL5, and CXCL12. The study showed that the migration of A549 cells through the polycarbonate membrane and basement membrane extract toward a chemotactic gradient elicited by serum from patients with COPD was markedly higher as compared with serum from healthy donors. The concentration of CCL21 and CXCL12, but not CXCL5, in serum from patients with COPD was also increased. Experiments in which CCL21- and CXCL12-dependent signaling was blocked revealed that increased migration of the cancer cells upon treatment with serum from patients with COPD was mediated exclusively by CCL21. Collectively, our results indicate that COPD may contribute to the progression of lung cancer via CCL21-dependent intensification of cancer cell migration. PMID:27307721

  11. Alpha(v)beta5 integrins mediates Pseudomonas fluorescens interaction with A549 cells.

    PubMed

    Buommino, Elisabetta; Di Domenico, Marina; Paoletti, Iole; Fusco, Alessandra; De Gregorio, Vincenza; Cozza, Valentina; Rizzo, Antonietta; Tufano, Maria Antonietta; Donnarumma, Giovanna

    2014-01-01

    Interaction of pathogenic bacteria with human cells is usually an essential step in the infection process. The bacterial invasion is stimulated by microbial binding to mammalian extracellular matrix proteins such as vitronectin, fibronectin or integrins. We have recently shown that some strains isolated from a clinical environment are able to grow at/or above 37°C. In particular, we demonstrated that P. fluorescens AF181 binds specifically to the surface of A549 human respiratory epithelial cells and that adhesiveness modulates the inflammatory response. In this study, the involvement of Alpha(v)Beta5 integrins and its respective natural ligand vitronectin (VN) in P. fluorescens AF181 adherence and invasion was examined. The host cell cytoskeleton and cellular tyrosine kinases seem to be solicited during the P. fluorescens-respiratory cell interaction; consequently, cytochalasin D and genistein decreased the bacterial adherence and internalization. Gene silencing of α(v), β5 integrins and vitronectin reduced P. fluorescens adherence and internalization to A549 cells. Taken together, these findings suggest that Alpha(v)Beta5 integrins and their natural ligand VN are involved in P. fluorescens adherence and invasion in human epithelial cells.

  12. Middle infrared radiation induces G2/M cell cycle arrest in A549 lung cancer cells.

    PubMed

    Chang, Hsin-Yi; Shih, Meng-Her; Huang, Hsuan-Cheng; Tsai, Shang-Ru; Juan, Hsueh-Fen; Lee, Si-Chen

    2013-01-01

    There were studies investigating the effects of broadband infrared radiation (IR) on cancer cell, while the influences of middle-infrared radiation (MIR) are still unknown. In this study, a MIR emitter with emission wavelength band in the 3-5 µm region was developed to irradiate A549 lung adenocarcinoma cells. It was found that MIR exposure inhibited cell proliferation and induced morphological changes by altering the cellular distribution of cytoskeletal components. Using quantitative PCR, we found that MIR promoted the expression levels of ATM (ataxia telangiectasia mutated), ATR (ataxia-telangiectasia and Rad3-related and Rad3-related), TP53 (tumor protein p53), p21 (CDKN1A, cyclin-dependent kinase inhibitor 1A) and GADD45 (growth arrest and DNA-damage inducible), but decreased the expression levels of cyclin B coding genes, CCNB1 and CCNB2, as well as CDK1 (Cyclin-dependent kinase 1). The reduction of protein expression levels of CDC25C, cyclin B1 and the phosphorylation of CDK1 at Thr-161 altogether suggest G(2)/M arrest occurred in A549 cells by MIR. DNA repair foci formation of DNA double-strand breaks (DSB) marker γ-H2AX and sensor 53BP1 was induced by MIR treatment, it implies the MIR induced G(2)/M cell cycle arrest resulted from DSB. This study illustrates a potential role for the use of MIR in lung cancer therapy by initiating DSB and blocking cell cycle progression.

  13. Measuring Attachment and Internalization of Influenza A Virus in A549 Cells by Flow Cytometry.

    PubMed

    Pohl, Marie O; Stertz, Silke

    2015-01-01

    Attachment to target cells followed by internalization are the very first steps of the life cycle of influenza A virus (IAV). We provide here a detailed protocol for measuring relative changes in the amount of viral particles that attach to A549 cells, a human lung epithelial cell line, as well as in the amount of particles that are internalized into the cell. We use biotinylated virus which can be easily detected following staining with Cy3-labeled streptavidin (STV-Cy3). We describe the growth, purification and biotinylation of A/WSN/33, a widely used IAV laboratory strain. Cold-bound biotinylated IAV particles on A549 cells are stained with STV-Cy3 and measured using flow cytometry. To investigate uptake of viral particles, cold-bound virus is allowed to internalize at 37 °C. In order to differentiate between external and internalized viral particles, a blocking step is applied: Free binding spots on the biotin of attached virus on the cell surface are bound by unlabeled streptavidin (STV). Subsequent cell permeabilization and staining with STV-Cy3 then enables detection of internalized viral particles. We present a calculation to determine the relative amount of internalized virus. This assay is suitable to measure effects of drug-treatments or other manipulations on attachment or internalization of IAV. PMID:26575457

  14. High-Throughput Quantitative Proteomic Analysis of Dengue Virus Type 2 Infected A549 Cells

    PubMed Central

    Chiu, Han-Chen; Hannemann, Holger; Heesom, Kate J.; Matthews, David A.; Davidson, Andrew D.

    2014-01-01

    Disease caused by dengue virus is a global health concern with up to 390 million individuals infected annually worldwide. There are no vaccines or antiviral compounds available to either prevent or treat dengue disease which may be fatal. To increase our understanding of the interaction of dengue virus with the host cell, we analyzed changes in the proteome of human A549 cells in response to dengue virus type 2 infection using stable isotope labelling in cell culture (SILAC) in combination with high-throughput mass spectrometry (MS). Mock and infected A549 cells were fractionated into nuclear and cytoplasmic extracts before analysis to identify proteins that redistribute between cellular compartments during infection and reduce the complexity of the analysis. We identified and quantified 3098 and 2115 proteins in the cytoplasmic and nuclear fractions respectively. Proteins that showed a significant alteration in amount during infection were examined using gene enrichment, pathway and network analysis tools. The analyses revealed that dengue virus infection modulated the amounts of proteins involved in the interferon and unfolded protein responses, lipid metabolism and the cell cycle. The SILAC-MS results were validated for a select number of proteins over a time course of infection by Western blotting and immunofluorescence microscopy. Our study demonstrates for the first time the power of SILAC-MS for identifying and quantifying novel changes in cellular protein amounts in response to dengue virus infection. PMID:24671231

  15. Telomere shortening and cell senescence induced by perylene derivatives in A549 human lung cancer cells.

    PubMed

    Taka, Thanachai; Huang, Liming; Wongnoppavich, Ariyaphong; Tam-Chang, Suk-Wah; Lee, T Randall; Tuntiwechapikul, Wirote

    2013-02-15

    Cancer cells evade replicative senescence by re-expressing telomerase, which maintains telomere length and hence chromosomal integrity. Telomerase inhibition would lead cancer cells to senesce and therefore prevent cancer cells from growing indefinitely. G-quadruplex ligands can attenuate telomerase activity by inducing G-quadruplex formation at the 3'-overhang of telomere and at the human telomerase reverse transcriptase (hTERT) promoter; the former prevents telomerase from accessing the telomere, and the latter acts as a transcriptional silencer. The present investigation found that perylene derivatives PM2 and PIPER induced G-quadruplex formation from both telomeric DNA and the hTERT promoter region in vitro. Further, TRAP assay showed that these compounds inhibited telomerase in a dose-dependent manner. When A549 human lung cancer cells were treated with these compounds, hTERT expression was down-regulated. Moreover, the crude protein extract from these treated cells exhibited less telomerase activity. In the long-term treatment of A549 lung cancer cells with sub-cytotoxic dose of these perylenes, telomere shortening, reduction of cell proliferation and tumorigenicity, and cell senescence were observed. The results of this study indicate that perylene derivatives warrant further consideration as effective agents for cancer therapy.

  16. COPD promotes migration of A549 lung cancer cells: the role of chemokine CCL21.

    PubMed

    Kuźnar-Kamińska, Barbara; Mikuła-Pietrasik, Justyna; Sosińska, Patrycja; Książek, Krzysztof; Batura-Gabryel, Halina

    2016-01-01

    Patients with COPD develop lung cancer more frequently than healthy smokers. At the same time, molecular mediators promoting various aspects of cancer cell progression are still elusive. In this report, we examined whether COPD can be coupled with increased migration of non-small-cell lung cancer cells A549 and, if so, whether this effect may be related to altered production and activity of chemokines CCL21, CXCL5, and CXCL12. The study showed that the migration of A549 cells through the polycarbonate membrane and basement membrane extract toward a chemotactic gradient elicited by serum from patients with COPD was markedly higher as compared with serum from healthy donors. The concentration of CCL21 and CXCL12, but not CXCL5, in serum from patients with COPD was also increased. Experiments in which CCL21- and CXCL12-dependent signaling was blocked revealed that increased migration of the cancer cells upon treatment with serum from patients with COPD was mediated exclusively by CCL21. Collectively, our results indicate that COPD may contribute to the progression of lung cancer via CCL21-dependent intensification of cancer cell migration.

  17. Apoptosis inducing ability of silver decorated highly reduced graphene oxide nanocomposites in A549 lung cancer.

    PubMed

    Khan, Merajuddin; Khan, Mujeeb; Al-Marri, Abdulhadi H; Al-Warthan, Abdulrahman; Alkhathlan, Hamad Z; Siddiqui, Mohammed Rafiq H; Nayak, Vadithe Lakshma; Kamal, Ahmed; Adil, Syed F

    2016-01-01

    Recently, graphene and graphene-based materials have been increasingly used for various biological applications due to their extraordinary physicochemical properties. Here, we demonstrate the anticancer properties and apoptosis-inducing ability of silver doped highly reduced graphene oxide nanocomposites synthesized by employing green approach. These nano composites (PGE-HRG-Ag) were synthesized by using Pulicaria glutinosa extract (PGE) as a reducing agent and were evaluated for their anticancer properties against various human cancer cell lines with tamoxifen as the reference drug. A correlation between the amount of Ag nanoparticles on the surface of highly reduced graphene oxide (HRG) and the anticancer activity of nanocomposite was observed, wherein an increase in the concentration of Ag nanoparticles on the surface of HRG led to the enhanced anticancer activity of the nanocomposite. The nanocomposite PGE-HRG-Ag-2 exhibited more potent cytotoxicity than standard drug in A549 cells, a human lung cancer cell line. A detailed investigation was undertaken and Fluorescence activated cell sorting (FACS) analysis demonstrated that the nanocomposite PGE-HRG-Ag-2 showed G0/G1 phase cell cycle arrest and induced apoptosis in A549 cells. Studies such as, measurement of mitochondrial membrane potential, generation of reactive oxygen species (ROS) and Annexin V-FITC staining assay suggested that this compound induced apoptosis in human lung cancer cells. PMID:27022256

  18. Apoptosis inducing ability of silver decorated highly reduced graphene oxide nanocomposites in A549 lung cancer

    PubMed Central

    Khan, Merajuddin; Khan, Mujeeb; Al-Marri, Abdulhadi H; Al-Warthan, Abdulrahman; Alkhathlan, Hamad Z; Siddiqui, Mohammed Rafiq H; Nayak, Vadithe Lakshma; Kamal, Ahmed; Adil, Syed F

    2016-01-01

    Recently, graphene and graphene-based materials have been increasingly used for various biological applications due to their extraordinary physicochemical properties. Here, we demonstrate the anticancer properties and apoptosis-inducing ability of silver doped highly reduced graphene oxide nanocomposites synthesized by employing green approach. These nano composites (PGE-HRG-Ag) were synthesized by using Pulicaria glutinosa extract (PGE) as a reducing agent and were evaluated for their anticancer properties against various human cancer cell lines with tamoxifen as the reference drug. A correlation between the amount of Ag nanoparticles on the surface of highly reduced graphene oxide (HRG) and the anticancer activity of nanocomposite was observed, wherein an increase in the concentration of Ag nanoparticles on the surface of HRG led to the enhanced anticancer activity of the nanocomposite. The nanocomposite PGE-HRG-Ag-2 exhibited more potent cytotoxicity than standard drug in A549 cells, a human lung cancer cell line. A detailed investigation was undertaken and Fluorescence activated cell sorting (FACS) analysis demonstrated that the nanocomposite PGE-HRG-Ag-2 showed G0/G1 phase cell cycle arrest and induced apoptosis in A549 cells. Studies such as, measurement of mitochondrial membrane potential, generation of reactive oxygen species (ROS) and Annexin V-FITC staining assay suggested that this compound induced apoptosis in human lung cancer cells. PMID:27022256

  19. αTAT1 downregulation induces mitotic catastrophe in HeLa and A549 cells.

    PubMed

    Chien, J-Y; Tsen, S-D; Chien, C-C; Liu, H-W; Tung, C-Y; Lin, C-H

    2016-01-01

    α-Tubulin acetyltransferase 1 (αTAT1) controls reversible acetylation on Lys40 of α-tubulin and modulates multiple cellular functions. αTAT1 depletion induced morphological defects of touch receptor neurons in Caenorhabditis elegans and impaired cell adhesion and contact inhibition in mouse embryonic fibroblasts, however, no morphological or proliferation defects in human RPE-hTERT cells were found after αTAT1-specific siRNA treatment. Here, we compared the effect of three αTAT1-specific shRNAs on proliferation and morphology in two human cell lines, HeLa and A549. The more efficient two shRNAs induced mitotic catastrophe in both cell lines and the most efficient one also decreased F-actin and focal adhesions. Further analysis revealed that αTAT1 downregulation increased γ-H2AX, but not other DNA damage markers p-CHK1 and p-CHK2, along with marginal change in microtubule outgrowth speed and inter-kinetochore distance. Overexpression of αTAT1 could not precisely mimic the distribution and concentration of endogenous acetylated α-tubulin (Ac-Tu), although no overt phenotype change was observed, meanwhile, this could not completely prevent αTAT1 downregulation-induced deficiencies. We therefore conclude that efficient αTAT1 downregulation could impair actin architecture and induce mitotic catastrophe in HeLa and A549 cells through mechanisms partly independent of Ac-Tu. PMID:27551500

  20. Apoptosis inducing ability of silver decorated highly reduced graphene oxide nanocomposites in A549 lung cancer.

    PubMed

    Khan, Merajuddin; Khan, Mujeeb; Al-Marri, Abdulhadi H; Al-Warthan, Abdulrahman; Alkhathlan, Hamad Z; Siddiqui, Mohammed Rafiq H; Nayak, Vadithe Lakshma; Kamal, Ahmed; Adil, Syed F

    2016-01-01

    Recently, graphene and graphene-based materials have been increasingly used for various biological applications due to their extraordinary physicochemical properties. Here, we demonstrate the anticancer properties and apoptosis-inducing ability of silver doped highly reduced graphene oxide nanocomposites synthesized by employing green approach. These nano composites (PGE-HRG-Ag) were synthesized by using Pulicaria glutinosa extract (PGE) as a reducing agent and were evaluated for their anticancer properties against various human cancer cell lines with tamoxifen as the reference drug. A correlation between the amount of Ag nanoparticles on the surface of highly reduced graphene oxide (HRG) and the anticancer activity of nanocomposite was observed, wherein an increase in the concentration of Ag nanoparticles on the surface of HRG led to the enhanced anticancer activity of the nanocomposite. The nanocomposite PGE-HRG-Ag-2 exhibited more potent cytotoxicity than standard drug in A549 cells, a human lung cancer cell line. A detailed investigation was undertaken and Fluorescence activated cell sorting (FACS) analysis demonstrated that the nanocomposite PGE-HRG-Ag-2 showed G0/G1 phase cell cycle arrest and induced apoptosis in A549 cells. Studies such as, measurement of mitochondrial membrane potential, generation of reactive oxygen species (ROS) and Annexin V-FITC staining assay suggested that this compound induced apoptosis in human lung cancer cells.

  1. Iron stimulates plasma-activated medium-induced A549 cell injury

    PubMed Central

    Adachi, Tetsuo; Nonomura, Saho; Horiba, Minori; Hirayama, Tasuku; Kamiya, Tetsuro; Nagasawa, Hideko; Hara, Hirokazu

    2016-01-01

    Non-thermal atmospheric pressure plasma is applicable to living cells and has emerged as a novel technology for cancer therapy. Plasma has recently been shown to affect cells not only by direct irradiation, but also by indirect treatments with previously prepared plasma-activated medium (PAM). Iron is an indispensable element but is also potentially toxic because it generates the hydroxyl radical (•OH) in the presence of hydrogen peroxide (H2O2) via the Fenton reaction. The aim of the present study was to demonstrate the contribution of iron to PAM-induced A549 adenocarcinoma cell apoptosis. We detected the generation of •OH and elevation of intracellular ferrous ions in PAM-treated cells and found that they were inhibited by iron chelator. The elevations observed in ferrous ions may have been due to their release from the intracellular iron store, ferritin. Hydroxyl radical-induced DNA injury was followed by the activation of poly(ADP-ribose) polymerase-1, depletion of NAD+ and ATP, and elevations in intracellular Ca2+. The sensitivities of normal cells such as smooth muscle cells and keratinocytes to PAM were less than that of A549 cells. These results demonstrated that H2O2 in PAM and/or •OH generated in the presence of iron ions disturbed the mitochondrial-nuclear network in cancer cells. PMID:26865334

  2. Tumor-specific targeting by Bavituximab, a phosphatidylserine-targeting monoclonal antibody with vascular targeting and immune modulating properties, in lung cancer xenografts.

    PubMed

    Gerber, David E; Hao, Guiyang; Watkins, Linda; Stafford, Jason H; Anderson, Jon; Holbein, Blair; Öz, Orhan K; Mathews, Dana; Thorpe, Philip E; Hassan, Gedaa; Kumar, Amit; Brekken, Rolf A; Sun, Xiankai

    2015-01-01

    Bavituximab is a chimeric monoclonal antibody with immune modulating and tumor-associated vascular disrupting properties demonstrated in models of non-small cell lung cancer (NSCLC). The molecular target of Bavituximab, phosphatidylserine (PS), is exposed on the outer leaflet of the membrane bi-layer of malignant vascular endothelial cells and tumor cells to a greater extent than on normal tissues. We evaluated the tumor-targeting properties of Bavituximab for imaging of NSCLC xenografts when radiolabeled with (111)In through conjugation with a bifunctional chelating agent, 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA). In vitro binding of (111)In-DOTA-Bavituximab to PS was determined by enzyme-linked immunosorbent assay (ELISA). Biodistribution of (111)In-DOTA-Bavituximab was conducted in normal rats, which provided data for dosimetry calculation. Single-photon emission computed tomography/computed tomography (SPECT/CT) imaging was performed in athymic nude rats bearing A549 NSCLC xenografts. At the molar conjugation ratio of 0.54 DOTA per Bavituximab, the PS binding affinity of (111)In-DOTA-Bavituximab was comparable to that of unmodified Bavituximab. Based on the quantitative SPECT/CT imaging data analysis, (111)In-DOTA-Bavituximab demonstrated tumor-specific uptake as measured by the tumor-tomuscle ratio, which peaked at 5.2 at 72 hr post-injection. In contrast, the control antibody only presented a contrast of 1.2 at the same time point.These findings may underlie the diagnostic efficacy and relative low rates of systemic vascular and immune-related toxicities of this immunoconjugate. Future applications of (111)In-DOTA-bavituximab may include prediction of efficacy, indication of tumor immunologic status, or characterization of radiographic findings. PMID:26550540

  3. Genistein inhibits A549 human lung cancer cell proliferation via miR-27a and MET signaling

    PubMed Central

    Yang, Yang; Zang, Aimin; Jia, Youchao; Shang, Yanhong; Zhang, Zhuoqi; Ge, Kun; Zhang, Jinchao; Fan, Wufang; Wang, Bei

    2016-01-01

    Genistein is a soybean isoflavone; in its aglycone it has various biological activities. Animal experiments, clinical studies and epidemiological investigations suggest that genistein has preventative and curative functions for a number of diseases, particularly in cancer. The present study explored the potential anti-cancer effect of genistein by observing its role in inhibiting A549 human lung cancer cell proliferation and investigating the possible mechanism. A549 cells were exposed to various concentrations of genistein (0, 10, 25, 50, 100 and 200 µM; dissolved in physiological saline) for 1, 2 and 3 days. Subsequently, the viability of A549 cells was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, cell apoptosis was examined using a flow cytometer, caspase 3/9 activity was measured using commercial kits, reverse transcription quantitative polymerase chain reaction was used to analyze the miR-27a expression and western blotting was used to investigate MET protein expression. The results suggested a significant inhibition of A549 cell growth following treatment with genistein in a time- and dose-dependent manner. The current study also indicated that treatment with genistein significantly induces cell apoptosis and promotes caspase-3/9 activation of A549 cells in a dose-dependent manner. Further functional assays revealed that the anti-cancer effect of genistein activated microRNA-27a (miR-27a) expression levels and reduced MET protein expression in A549 cells. In conclusion, the present study demonstrates that genistein inhibits A549 human lung cancer cell proliferation. Furthermore, this study reports, for the first time, a correlation between the anti-cancer effect of genistein and miR-27a-mediated MET signaling.

  4. Genistein inhibits A549 human lung cancer cell proliferation via miR-27a and MET signaling

    PubMed Central

    Yang, Yang; Zang, Aimin; Jia, Youchao; Shang, Yanhong; Zhang, Zhuoqi; Ge, Kun; Zhang, Jinchao; Fan, Wufang; Wang, Bei

    2016-01-01

    Genistein is a soybean isoflavone; in its aglycone it has various biological activities. Animal experiments, clinical studies and epidemiological investigations suggest that genistein has preventative and curative functions for a number of diseases, particularly in cancer. The present study explored the potential anti-cancer effect of genistein by observing its role in inhibiting A549 human lung cancer cell proliferation and investigating the possible mechanism. A549 cells were exposed to various concentrations of genistein (0, 10, 25, 50, 100 and 200 µM; dissolved in physiological saline) for 1, 2 and 3 days. Subsequently, the viability of A549 cells was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, cell apoptosis was examined using a flow cytometer, caspase 3/9 activity was measured using commercial kits, reverse transcription quantitative polymerase chain reaction was used to analyze the miR-27a expression and western blotting was used to investigate MET protein expression. The results suggested a significant inhibition of A549 cell growth following treatment with genistein in a time- and dose-dependent manner. The current study also indicated that treatment with genistein significantly induces cell apoptosis and promotes caspase-3/9 activation of A549 cells in a dose-dependent manner. Further functional assays revealed that the anti-cancer effect of genistein activated microRNA-27a (miR-27a) expression levels and reduced MET protein expression in A549 cells. In conclusion, the present study demonstrates that genistein inhibits A549 human lung cancer cell proliferation. Furthermore, this study reports, for the first time, a correlation between the anti-cancer effect of genistein and miR-27a-mediated MET signaling. PMID:27602162

  5. Epigenetic induction of epithelial to mesenchymal transition by LCN2 mediates metastasis and tumorigenesis, which is abrogated by NF-κB inhibitor BRM270 in a xenograft model of lung adenocarcinoma.

    PubMed

    Mongre, Raj Kumar; Sodhi, Simrinder Singh; Sharma, Neelesh; Ghosh, Mrinmoy; Kim, Jeong Hyun; Kim, Nameun; Park, Yang Ho; Shin, Young Gyu; Kim, Sung Jin; Jiao, Zhang Jiao; Huynh, Do Luong; Jeong, Dong Kee

    2016-01-01

    Tumor initiating cancer stem-like cells (TICSCs) have recently become the object of intensive study. Human-Lipocalin-2 (hLCN2) acts as a biomarker for cancers. The aim of the present study was to explore new insights regarding the potential role of LCN2 in inducing epithelial to mesenchymal transition (EMT) by transfecting LCN2 into CD133+-A549-TICSCs and its cross-talk with the NF-κB signaling pathway in adenocarcinoma of the lung. Furthermore, EMT was confirmed by transcriptomic analysis, immunoblotting and immunocyto/histochemical analyses. Tumorigenesis and metastasis were confirmed by molecular therapeutics tracer 2DG infrared optical probe in BALB/cSIc-nude mice. It was observed that the CD133+-expressing-LCN2-A549 TICSCs population increased in adenocarcinoma of the lung compared to the normal lung tissue. The expressions of genes involved in stemness, adhesion, motility and drug efflux was higher in these cells than in their non-LCN2 expressing counterparts. The present study revealed that elevated expression of LCN2 significantly induced metastasis via EMT. Overexpression of LCN2 significantly increased stemness and tumor metastasis by modulating NF-κB cellular signaling. BRM270, a novel inhibitor of NF-κB plays a significant role in the EMT reversal. BRM270, a naturaceutical induces cell shrinkage, karyorrhexis and programmed cell death (PCD) which were observed by Hoechst 33342 staining while flow cytometry analysis showed significant (P<0.05) decrease in cell population from G0-G1 phases. Also, 2DG guided in vivo model revealed that BRRM270 significantly (P<0.0003) reduced tumor metastasis and increased percent survival in real-time with complete resection. An elaborate study on the novel concept with respect to linking of naturaceutics as selective and potential anticancer agent that eliminates the elevated LCN2 induced EMT and tumor dissemination through cooperation with the NF-κB signaling as the baseline data for the planning of new

  6. Targeting Tumor Cells with Anti-CD44 Antibody Triggers Macrophage-Mediated Immune Modulatory Effects in a Cancer Xenograft Model

    PubMed Central

    Maisel, Daniela; Birzele, Fabian; Voss, Edgar; Nopora, Adam; Bader, Sabine; Friess, Thomas; Goller, Bernhard; Laifenfeld, Daphna; Weigand, Stefan; Runza, Valeria

    2016-01-01

    CD44, a transmembrane receptor reported to be involved in various cellular functions, is overexpressed in several cancer types and supposed to be involved in the initiation, progression and prognosis of these cancers. Since the sequence of events following the blockage of the CD44-HA interaction has not yet been studied in detail, we profiled xenograft tumors by RNA Sequencing to elucidate the mode of action of the anti-CD44 antibody RG7356. Analysis of tumor and host gene-expression profiles led us to the hypothesis that treatment with RG7356 antibody leads to an activation of the immune system. Using cytokine measurements we further show that this activation involves the secretion of chemo-attractants necessary for the recruitment of immune cells (i.e. macrophages) to the tumor site. We finally provide evidence for antibody-dependent cellular phagocytosis (ADCP) of the malignant cells by macrophages. PMID:27463372

  7. Rho GTPases RhoA and Rac1 mediate effects of dietary folate on metastatic potential of A549 cancer cells through the control of cofilin phosphorylation.

    PubMed

    Oleinik, Natalia V; Helke, Kristi L; Kistner-Griffin, Emily; Krupenko, Natalia I; Krupenko, Sergey A

    2014-09-19

    Folate, an important nutrient in the human diet, has been implicated in cancer, but its role in metastasis is not established. We have shown previously that the withdrawal of medium folate leads to the inhibition of migration and invasion of A549 lung carcinoma cells. Here we have demonstrated that medium folate regulates the function of Rho GTPases by enabling their carboxyl methylation and translocation to plasma membrane. Conversely, the lack of folate leads to the retention of these proteins in endoplasmic reticulum. Folate also promoted the switch from inactive (GDP-bound) to active (GTP-bound) GTPases, resulting in the activation of downstream kinases p21-activated kinase and LIM kinase and phosphorylation of the actin-depolymerizing factor cofilin. We have further demonstrated that in A549 cells two GTPases, RhoA and Rac1, but not Cdc42, are immediate sensors of folate status: the siRNA silencing of RhoA or Rac1 blocked effects of folate on cofilin phosphorylation and cellular migration and invasion. The finding that folate modulates metastatic potential of cancer cells was confirmed in an animal model of lung cancer using tail vein injection of A549 cells in SCID mice. A folate-rich diet enhanced lung colonization and distant metastasis to lymph nodes and decreased overall survival (35 versus 63 days for mice on a folate-restricted diet). High folate also promoted epithelial-mesenchymal transition in cancer cells and experimental mouse tumors. Our study provides experimental evidence for a mechanism of metastasis promotion by dietary folate and highlights the interaction between nutrients and metastasis-related signaling.

  8. Resveratrol reduces IL-6 and VEGF secretion from co-cultured A549 lung cancer cells and adipose-derived mesenchymal stem cells.

    PubMed

    Sahin, Erhan; Baycu, Cengiz; Koparal, Ayse Tansu; Burukoglu Donmez, Dilek; Bektur, Ezgi

    2016-06-01

    Stem cell therapies are important treatment methodologies used in many areas of experimental or clinical medicine. In recent studies of cancer models, Mesenchymal stem cells (MSCs) suppressed the growth of cancer cells. However, also in some studies, stem cell treatments have been shown to induce cancer formation, increase tumor volume, induce the formation of new vessels, and lead to cancer invasion. The presence of MSC-secreted cytokines and their effects on cancer cells limits the reliability of MSC-based treatments. Resveratrol (trans-3,5,4'-trihydroxystilbene), an antioxidant found in red wine, has been shown to have therapeutic effects against several cancers. The aim of this study was to co-culture MSCs with A549 cancer cells to suppress the release of cancer-promoting cytokines from MSCs and to increase the applicability and reliability of stem cell therapies with resveratrol. MTT (3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) and neutral red cell viability assays were used to find safety dose of resveratrol. The MSCs secreted the cytokines IL-6 and VEGF, and the effect of resveratrol on these cytokines was analyzed by ELISA and western blot analysis of conditioned medium. One μM of resveratrol was found to be the safety dose for the A549 cancer cells and MSCs. We observed the highest release of IL-6 and VEGF from the co-cultured A549 cells and MSCs, and resveratrol was found to significantly decrease the release of these cytokines. Our study suggests that resveratrol exerts a positive effect on the release of cytokines. The safety dose of resveratrol can be administered together with stem cells during stem cell treatment.

  9. The synergistic antitumor effects of all-trans retinoic acid and C-phycocyanin on the lung cancer A549 cells in vitro and in vivo.

    PubMed

    Li, Bing; Gao, Mei-Hua; Chu, Xian-Ming; Teng, Lei; Lv, Cong-Yi; Yang, Peng; Yin, Qi-Feng

    2015-02-15

    The anticancer effects and mechanism of all-trans retinoic acid (ATRA), C-phycocyanin (C-PC) or ATRA+C-PC on the growth of A549 cells were studied in in vitro and in vivo experiments. The effects of C-PC and ATRA on the growth of A549 cells were determined. The expression of CDK-4 and caspase-3, and the cellular apoptosis levels were detected. The tumor model was established by subcutaneous injection of A549 cells to the left axilla of the NU/NU mice. The weights of tumor and the spleen were tested. The viabilities of T-cells and spleen cells, TNF levels, the expression of Bcl-2 protein and Cyclin D1 gene were examined. Results showed both C-PC and ATRA could inhibit the growth of tumor cells in vivo and in vitro. ATRA+C-PC cooperatively showed a higher antitumor activity. The dosage of ATRA was reduced when it was administered with C-PC together, and the toxicity was reduced as well. ATRA+C-PC could decrease CDK-4 but increase caspase-3 protein expression level and induce cell apoptosis. ATRA alone could lower the activities of T lymphocytes and spleen weights, but the combination with C-PC could effectively promote viability of T cells and spleen. C-PC+ATRA could up-regulate TNF, and down-regulate Bcl-2 and Cyclin D1 gene. The combination might inhibit tumor growth by inhibiting the progress of cell cycle, inducing cell apoptosis and enhancing the body immunity. PMID:25617793

  10. MR diffusion-weighted imaging-based subcutaneous tumour volumetry in a xenografted nude mouse model using 3D Slicer: an accurate and repeatable method

    PubMed Central

    Ma, Zelan; Chen, Xin; Huang, Yanqi; He, Lan; Liang, Cuishan; Liang, Changhong; Liu, Zaiyi

    2015-01-01

    Accurate and repeatable measurement of the gross tumour volume(GTV) of subcutaneous xenografts is crucial in the evaluation of anti-tumour therapy. Formula and image-based manual segmentation methods are commonly used for GTV measurement but are hindered by low accuracy and reproducibility. 3D Slicer is open-source software that provides semiautomatic segmentation for GTV measurements. In our study, subcutaneous GTVs from nude mouse xenografts were measured by semiautomatic segmentation with 3D Slicer based on morphological magnetic resonance imaging(mMRI) or diffusion-weighted imaging(DWI)(b = 0,20,800 s/mm2) . These GTVs were then compared with those obtained via the formula and image-based manual segmentation methods with ITK software using the true tumour volume as the standard reference. The effects of tumour size and shape on GTVs measurements were also investigated. Our results showed that, when compared with the true tumour volume, segmentation for DWI(P = 0.060–0.671) resulted in better accuracy than that mMRI(P < 0.001) and the formula method(P < 0.001). Furthermore, semiautomatic segmentation for DWI(intraclass correlation coefficient, ICC = 0.9999) resulted in higher reliability than manual segmentation(ICC = 0.9996–0.9998). Tumour size and shape had no effects on GTV measurement across all methods. Therefore, DWI-based semiautomatic segmentation, which is accurate and reproducible and also provides biological information, is the optimal GTV measurement method in the assessment of anti-tumour treatments. PMID:26489359

  11. 4-Nitroquinoline-1-oxide effects human lung adenocarcinoma A549 cells by regulating the expression of POLD4

    PubMed Central

    HUANG, QIN-MIAO; ZENG, YI-MING; ZHANG, HUA-PING; LV, LIANG-CHAO; YANG, DONG-YONG; LIN, HUI-HUANG

    2016-01-01

    The aim of the present study was to explore the expression of POLD4 in human lung adenocarcinoma A549 cells under 4-nitroquinoline-1-oxide (4NQO) stimulation to investigate the role of POLD4 in smoking-induced lung cancer. The lung cancer A549 cell line was treated with 4NQO, with or without MG132 (an inhibitor of proteasome activity), and subsequently the POLD4 level was determined by western blot analysis. Secondly, the cell sensitivity to 4NQO and Taxol was determined when the POLD4 expression level was downregulated by siRNA. The POLD4 protein levels in the A549 cells decreased following treatment with 4NQO; however, MG132 could reverse this phenotype. Downregulation of the POLD4 expression by siRNA enhanced A549 cell sensitivity to 4NQO, but not to Taxol. In conclusion, 4NQO affects human lung adenocarcinoma A549 cells by regulating the expression of POLD4. PMID:26998273

  12. In vitro anticancer activity of fucoidan from Turbinaria conoides against A549 cell lines.

    PubMed

    Marudhupandi, Thangapandi; Ajith Kumar, Thipramalai Thankappan; Lakshmanasenthil, Shanmugaasokan; Suja, Gunasekaran; Vinothkumar, Thirumalairaj

    2015-01-01

    The present study was conducted to evaluate the anticancer activity of fucoidan isolated from brown seaweed Turbinaria conoides. Extracted fucoidan contained 53 ± 0.69% of fucose and 38 ± 0.42% of sulphate, respectively. Functional groups and structural characteristics of the fucoidan were analyzed by FT-IR and NMR. In vitro anticancer effect was studied on A549 cell line. Fucoidan inhibited the growth of cancer cells in a dose-dependent manner and potent anticancer activities were 24.9-73.5% in the concentrations of 31.25-500 μg/ml. The CTC50 value against the cancer cell was found to be 45 μg/ml and the CTC50 value of normal Vero cell line is 325 μg/ml. This study suggests that the fucoidan from T. conoides could be significantly improved if the active component is further purified and tested for further investigation in various cancer cell lines.

  13. In Vitro Evaluation of 3-Arylcoumarin Derivatives in A549 Cell Line

    PubMed Central

    MUSA, MUSILIYU A.; JOSEPH, MOISE Y.; LATINWO, LEKAN M.; BADISA, VEERA; COOPERWOOD, JOHN S.

    2016-01-01

    Coumarins are naturally-occurring compounds with diverse and interesting biological activities. In the present study, we evaluated the in vitro cytotoxic effect of 8-(acetyloxy)-3-[4-(acetyloxy)phenyl]-2-oxo-2H-chromen-7-yl acetate (6); 8-(acetyloxy)-3-(4-methanesulfonyl phenyl)-2-oxo-2H-chromen-7-yl acetate (7); 4-(2-oxo-2H-chromen-3-yl)phenyl acetate (8); 3-(4-methanesulfonylphenyl)-2H-chromen-2-one (9); 4-(4-methyl-2-oxo-2H-chromen-3-yl)phenyl acetate (10); 3-(4-methanesulfonylphenyl)-4-methyl-2H-chromen-2-one (11); 8-(acetyloxy)-3-[4-(acetyloxy)phenyl]-4-methyl-2-oxo-2H-chromen-7-yl acetate (12); and 5-(acetyloxy)-3-[4-(acetyloxy) phenyl]-2-oxo-2H-chromen-7-yl acetate (13) in human lung (A549) cancer and normal lung (MRC-9) cell lines at different concentrations for 48 h using crystal violet dye binding assay. The cytotoxic effect of these coumarin derivatives were compared to the standard drug, docetaxel. Furthermore, the effect of the most active compound on the cell-cycle using propidium iodide, mitochondrial membrane potential (MMP) using tetramethyl rhodamine methyl ester (rhodamine-123) and reactive oxygen species (ROS) production using 2′,7′-dichlorofluorescin diacetate (PCFDA) were also evaluated. Results Compound 7 had the greatest cytotoxic effect (cytotoxic concentration, CC50=24 μM) and selectivity (MRC-9; CC50 >100 μM; inactive) in the A549 cell line, and caused cells to arrest in the S phase of the cell cycle, loss of MMP and increased ROS production in a concentration-dependent manner. Conclusion These findings suggest that compound 7 could serve as a new lead for the development of novel synthetic compounds with enhanced anticancer activity. PMID:25667442

  14. Effects of Fe3O4 Magnetic Nanoparticles on A549 Cells

    PubMed Central

    Watanabe, Masatoshi; Yoneda, Misao; Morohashi, Ayaka; Hori, Yasuki; Okamoto, Daiki; Sato, Akiko; Kurioka, Daisuke; Nittami, Tadashi; Hirokawa, Yoshifumi; Shiraishi, Taizo; Kawai, Kazuaki; Kasai, Hiroshi; Totsuka, Yukari

    2013-01-01

    Fe3O4 magnetic nanoparticles (MgNPs-Fe3O4) are widely used in medical applications, including magnetic resonance imaging, drug delivery, and in hyperthermia. However, the same properties that aid their utility in the clinic may potentially induce toxicity. Therefore, the purpose of this study was to investigate the cytotoxicity and genotoxicity of MgNPs-Fe3O4 in A549 human lung epithelial cells. MgNPs-Fe3O4 caused cell membrane damage, as assessed by the release of lactate dehydrogenase (LDH), only at a high concentration (100 μg/mL); a lower concentration (10 μg/mL) increased the production of reactive oxygen species, increased oxidative damage to DNA, and decreased the level of reduced glutathione. MgNPs-Fe3O4 caused a dose-dependent increase in the CD44+ fraction of A549 cells. MgNPs-Fe3O4 induced the expression of heme oxygenase-1 at a concentration of 1 μg/mL, and in a dose-dependent manner. Despite these effects, MgNPs-Fe3O4 had minimal effect on cell viability and elicited only a small increase in the number of cells undergoing apoptosis. Together, these data suggest that MgNPs-Fe3O4 exert little or no cytotoxicity until a high exposure level (100 μg/mL) is reached. This dissociation between elevated indices of cell damage and a small effect on cell viability warrants further study. PMID:23892599

  15. Effect of soluble nickel on cellular energy metabolism in A549 cells.

    PubMed

    Chen, Haobin; Costa, Max

    2006-10-01

    Iron is an essential nutrient to most organisms, and is actively involved in oxygen delivery, electron transport, DNA synthesis, and many other biochemical reactions important for cell survival. We previously reported that nickel (Ni) ion exposure decreases cellular iron level and converts cytosolic aconitase (c-aconitase) to iron-regulatory protein-1 in A549 cells (Chen H, Davidson T, Singleton S, Garrick MD, Costa M. Toxicol Appl Pharmacol 206:275-287, 2005). Here, we further investigated the effect of Ni ion exposure on the activity of mitochondrial iron-sulfur (Fe-S) enzymes and cellular energy metabolism. We found that acute Ni ion treatment up to 1 mM exhibits minimal toxicity in A549 cells. Ni ion treatment decreases the activity of several Fe-S enzymes related to cellular energy metabolism, including mitochondrial aconitase (m-aconitase), succinate dehydrogenase (SDH), and NADH:ubiquinone oxidoreductase (complex I). Low doses of Ni ion for 4 weeks resulted in an increased cellular glycolysis and NADH to NAD+ (NADH/NAD+) ratio, although glycolysis was inhibited at higher levels. Collectively, our results show that Ni ions decrease the activity of cellular iron (Fe)-containing enzymes, inhibit oxidative phosphorylation (OxPhos), and increase cellular glycolytic activity. Since increased glycolysis is one of the fundamental alterations of energy metabolism in cancer cells (the Warburg effect), the inhibition of Fe-S enzymes and subsequent changes in cellular energy metabolism caused by Ni ions may play an important role in Ni carcinogenesis. PMID:17018869

  16. Okadaic acid inhibits cell multiplication and induces apoptosis in a549 cells, a human lung adenocarcinoma cell line

    PubMed Central

    Wang, Renjun; Lv, Lili; Zhao, Yunfeng; Yang, Nana

    2014-01-01

    This essay aims to research the effect of okadaic acid (OA) on A549 cell multiplication, and cell apoptosis induced by OA was observed by cell morphology. MTT assay, trypan blue exclusion test (TBET), Giemsa staining method and acridine orange (AO) fluorescence staining assay were applied. The results of cell survival evaluated by TBET and colorimetric assay with 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) showed: The number of A549 cells was decreased in a dose-dependent manner. Cytomorphology observation of okadaic acid-treated cells showed that cells became shrinkage and turned round, some cells floated in the nutrient medium with nucleus agglutination broken, resulting in apoptotic bodies. Above-mentioned results indicated that OA exerted significantly inhibitory effect on A549 cell multiplication due to the apoptosis induced by OA. PMID:25232383

  17. Human Immunodeficiency Virus Type 1 Infection of Neural Xenografts

    NASA Astrophysics Data System (ADS)

    Cvetkovich, Therese A.; Lazar, Eliot; Blumberg, Benjamin M.; Saito, Yoshihiro; Eskin, Thomas A.; Reichman, Richard; Baram, David A.; del Cerro, Coca; Gendelman, Howard E.; del Cerro, Manuel; Epstein, Leon G.

    1992-06-01

    Human immunodeficiency virus type 1 (HIV-1) infection is highly specific for its human host. To study HIV-1 infection of the human nervous system, we have established a small animal model in which second-trimester (11 to 17.5 weeks) human fetal brain or neural retina is transplanted to the anterior chamber of the eye of immunosuppressed adult rats. The human xenografts vascularized, formed a blood-brain barrier, and differentiated, forming neurons and glia. The xenografts were infected with cell-free HIV-1 or with HIV-1-infected human monocytes. Analysis by polymerase chain reaction revealed HIV-1 sequences in DNA from xenograft tissue exposed to HIV-1 virions, and in situ hybridization demonstrated HIV-1 mRNA localized in macrophages and multinucleated giant cells. Pathological damage was observed only in neural xenografts containing HIV-1-infected human monocytes, supporting the hypothesis that these cells mediate neurotoxicity. This small animal model allows the study of direct and indirect effects of HIV-1 infection on developing human fetal neural tissues, and it should prove useful in evaluating antiviral therapies, which must ultimately target HIV-1 infection of the brain.

  18. Human immunodeficiency virus type 1 infection of neural xenografts.

    PubMed Central

    Cvetkovich, T A; Lazar, E; Blumberg, B M; Saito, Y; Eskin, T A; Reichman, R; Baram, D A; del Cerro, C; Gendelman, H E; del Cerro, M

    1992-01-01

    Human immunodeficiency virus type 1 (HIV-1) infection is highly specific for its human host. To study HIV-1 infection of the human nervous system, we have established a small animal model in which second-trimester (11 to 17.5 weeks) human fetal brain or neural retina is transplanted to the anterior chamber of the eye of immunosuppressed adult rats. The human xenografts vascularized, formed a blood-brain barrier, and differentiated, forming neurons and glia. The xenografts were infected with cell-free HIV-1 or with HIV-1-infected human monocytes. Analysis by polymerase chain reaction revealed HIV-1 sequences in DNA from xenograft tissue exposed to HIV-1 virions, and in situ hybridization demonstrated HIV-1 mRNA localized in macrophages and multinucleated giant cells. Pathological damage was observed only in neural xenografts containing HIV-1-infected human monocytes, supporting the hypothesis that these cells mediate neurotoxicity. This small animal model allows the study of direct and indirect effects of HIV-1 infection on developing human fetal neural tissues, and it should prove useful in evaluating antiviral therapies, which must ultimately target HIV-1 infection of the brain. Images PMID:1594627

  19. α-Mangostin: A Dietary Antioxidant Derived from the Pericarp of Garcinia mangostana L. Inhibits Pancreatic Tumor Growth in Xenograft Mouse Model

    PubMed Central

    Mustafa, Ala; Fischer, Joseph W.; Singh, Ashok; Zhong, Weixiong; Shekhani, Mohammed Ozair; Meske, Louise; Havighurst, Thomas; Kim, KyungMann; Verma, Ajit Kumar

    2014-01-01

    Abstract Aims: Pancreatic cancer (PC) is the most aggressive malignant disease, ranking as the fourth most leading cause of cancer-related death among men and women in the United States. In this study, we provide evidence of chemotherapeutic effects of α-mangostin, a dietary antioxidant isolated from the pericarp of Garcinia mangostana L. against human PC. Results: The chemotherapeutic effect of α-mangostin was determined using four human PC cells (PL-45, PANC1, BxPC3, and ASPC1). α-Mangostin resulted in a significant inhibition of PC cells viability without having any effects on normal human pancreatic duct epithelial cells. α-Mangostin showed a dose-dependent increase of apoptosis in PC cells. Also, α-mangostin inhibited the expression levels of pNF-κB/p65Ser552, pStat3Ser727, and pStat3Tyr705. α-Mangostin inhibited DNA binding activity of nuclear factor kappa B (NF-κB) and signal transducer and activator 3 (Stat3). α-Mangostin inhibited the expression levels of matrix metallopeptidase 9 (MMP9), cyclin D1, and gp130; however, increased expression of tissue inhibitor of metalloproteinase 1 (TIMP1) was observed in PC cells. In addition, i.p. administration of α-mangostin (6 mg/kg body weight, 5 days a week) resulted in a significant inhibition of both primary (PL-45) and secondary (ASPC1) human PC cell-derived orthotopic and ectopic xenograft tumors in athymic nude mice. No sign of toxicity was observed in any of the mice administered with α-mangostin. α-Mangostin treatment inhibited the biomarkers of cell proliferation (Ki-67 and proliferating cell nuclear antigen [PCNA]) in the xenograft tumor tissues. Innovation: We present, for the first time, that dietary antioxidant α-mangostin inhibits the growth of PC cells in vitro and in vivo. Conclusion: These results suggest the potential therapeutic efficacy of α-mangostin against human PC. Antioxid. Redox Signal. 21, 682–699. PMID:24295217

  20. Feroniellin A-induced autophagy causes apoptosis in multidrug-resistant human A549 lung cancer cells.

    PubMed

    Kaewpiboon, Chutima; Surapinit, Serm; Malilas, Waraporn; Moon, Jeong; Phuwapraisirisan, Preecha; Tip-Pyang, Santi; Johnston, Randal N; Koh, Sang Seok; Assavalapsakul, Wanchai; Chung, Young-Hwa

    2014-04-01

    During the screening of natural chemicals that can reverse multidrug resistance in human A549 lung cancer cells resistant to etoposide (A549RT-eto), we discovered that Feroniellin A (FERO), a novel furanocoumarin, shows toxicity toward A549RT-eto cells in a dose- and time-dependent manner. FERO reduced the expression of NF-κB, leading to downregulation of P-glycoprotein (P-gp), encoded by MDR1, which eventually sensitized A549RT-eto cells to apoptosis. FERO specifically diminished transcription and promoter activity of MDR1 but did not inhibit the expression of other multidrug resistance genes MRP2 and BCRP. Moreover, co-administration of FERO with Bay11-7802, an inhibitor of NF-κB, accelerated apoptosis of A549RT-eto cells through decreased expression of P-gp, indicating that NF-κB is involved in multidrug resistance. Conversely, addition of Z-VAD, a pan-caspase inhibitor, blocked FERO-induced apoptosis in A549RT-eto cells but did not block downregulation of P-gp, indicating that a decrease in P-gp expression is necessary but not sufficient for FERO-induced apoptosis. Interestingly, we found that FERO also induces autophagy, which is characterized by the conversion of LC3 I to LC3 II, induction of GFP-LC3 puncta, enhanced expression of Beclin-1 and ATG5, and inactivation of mTOR. Furthermore, suppression of Beclin-1 by siRNA reduced FERO-induced apoptosis in A549RT-eto cells and activation of autophagy by rapamycin accelerated FERO-induced apoptosis, suggesting that autophagy plays an active role in FERO-induced apoptosis. Herein, we report that FERO reverses multidrug resistance in A549RT-eto cells and exerts its cytotoxic effect by induction of both autophagy and apoptosis, which suggests that FERO can be a useful anticancer drug for multidrug-resistant lung cancer. PMID:24535083

  1. Synthesis and cytotoxicity evaluation of 4-amino-4-dehydroxylarctigenin derivatives in glucose-starved A549 tumor cells.

    PubMed

    Lei, Min; Gan, Xianwen; Zhao, Kun; Yu, Qiang; Hu, Lihong

    2015-02-01

    The natural product arctigenin (ATG) demonstrated preferential cytotoxicity to cancer cells under glucose starvation. A series of 4-amino-4-dehydroxylarctigenin derivatives based on lead compound ATG were designed and synthesized by bioisosteric modifications. Their cytotoxicities were evaluated in glucose-starved A549 tumor cells and the results indicated that the 4-amino-4-dehydroxylarctigenin showed more potent cytotoxicity than arctigenin, and the further substituent group on 4-amino would result in the cytotoxicities decreased significantly. 4-Substituted-arctigenin could selectively target on glucose-starved A549 tumor cells which provide an alternative strategy for anticancer drug development with minimal normal tissue toxicity.

  2. Synthesis and cytotoxicity evaluation of 4-amino-4-dehydroxylarctigenin derivatives in glucose-starved A549 tumor cells.

    PubMed

    Lei, Min; Gan, Xianwen; Zhao, Kun; Yu, Qiang; Hu, Lihong

    2015-02-01

    The natural product arctigenin (ATG) demonstrated preferential cytotoxicity to cancer cells under glucose starvation. A series of 4-amino-4-dehydroxylarctigenin derivatives based on lead compound ATG were designed and synthesized by bioisosteric modifications. Their cytotoxicities were evaluated in glucose-starved A549 tumor cells and the results indicated that the 4-amino-4-dehydroxylarctigenin showed more potent cytotoxicity than arctigenin, and the further substituent group on 4-amino would result in the cytotoxicities decreased significantly. 4-Substituted-arctigenin could selectively target on glucose-starved A549 tumor cells which provide an alternative strategy for anticancer drug development with minimal normal tissue toxicity. PMID:25571795

  3. Feroniellin A-induced autophagy causes apoptosis in multidrug-resistant human A549 lung cancer cells.

    PubMed

    Kaewpiboon, Chutima; Surapinit, Serm; Malilas, Waraporn; Moon, Jeong; Phuwapraisirisan, Preecha; Tip-Pyang, Santi; Johnston, Randal N; Koh, Sang Seok; Assavalapsakul, Wanchai; Chung, Young-Hwa

    2014-04-01

    During the screening of natural chemicals that can reverse multidrug resistance in human A549 lung cancer cells resistant to etoposide (A549RT-eto), we discovered that Feroniellin A (FERO), a novel furanocoumarin, shows toxicity toward A549RT-eto cells in a dose- and time-dependent manner. FERO reduced the expression of NF-κB, leading to downregulation of P-glycoprotein (P-gp), encoded by MDR1, which eventually sensitized A549RT-eto cells to apoptosis. FERO specifically diminished transcription and promoter activity of MDR1 but did not inhibit the expression of other multidrug resistance genes MRP2 and BCRP. Moreover, co-administration of FERO with Bay11-7802, an inhibitor of NF-κB, accelerated apoptosis of A549RT-eto cells through decreased expression of P-gp, indicating that NF-κB is involved in multidrug resistance. Conversely, addition of Z-VAD, a pan-caspase inhibitor, blocked FERO-induced apoptosis in A549RT-eto cells but did not block downregulation of P-gp, indicating that a decrease in P-gp expression is necessary but not sufficient for FERO-induced apoptosis. Interestingly, we found that FERO also induces autophagy, which is characterized by the conversion of LC3 I to LC3 II, induction of GFP-LC3 puncta, enhanced expression of Beclin-1 and ATG5, and inactivation of mTOR. Furthermore, suppression of Beclin-1 by siRNA reduced FERO-induced apoptosis in A549RT-eto cells and activation of autophagy by rapamycin accelerated FERO-induced apoptosis, suggesting that autophagy plays an active role in FERO-induced apoptosis. Herein, we report that FERO reverses multidrug resistance in A549RT-eto cells and exerts its cytotoxic effect by induction of both autophagy and apoptosis, which suggests that FERO can be a useful anticancer drug for multidrug-resistant lung cancer.

  4. Direct and in vitro observation of growth hormone receptor molecules in A549 human lung epithelial cells by nanodiamond labeling

    NASA Astrophysics Data System (ADS)

    Cheng, C.-Y.; Perevedentseva, E.; Tu, J.-S.; Chung, P.-H.; Cheng, C.-L.; Liu, K.-K.; Chao, J.-I.; Chen, P.-H.; Chang, C.-C.

    2007-04-01

    This letter presents direct observation of growth hormone receptor in one single cancer cell using nanodiamond-growth hormone complex as a specific probe. The interaction of surface growth hormone receptor of A549 human lung epithelial cells with growth hormone was observed using nanodiamond's unique spectroscopic signal via confocal Raman mapping. The growth hormone molecules were covalent conjugated to 100nm diameter carboxylated nanodiamonds, which can be recognized specifically by the growth hormone receptors of A549 cell. The Raman spectroscopic signal of diamond provides direct and in vitro observation of growth hormone receptors in physiology condition in a single cell level.

  5. Identification of a novel pyrazolo[3,4-d]pyrimidine able to inhibit cell proliferation of a human osteogenic sarcoma in vitro and in a xenograft model in mice.

    PubMed

    Manetti, Fabrizio; Santucci, Annalisa; Locatelli, Giada A; Maga, Giovanni; Spreafico, Adriano; Serchi, Tommaso; Orlandini, Maurizio; Bernardini, Giulia; Caradonna, Nicola P; Spallarossa, Andrea; Brullo, Chiara; Schenone, Silvia; Bruno, Olga; Ranise, Angelo; Bondavalli, Francesco; Hoffmann, Oskar; Bologna, Mauro; Angelucci, Adriano; Botta, Maurizio

    2007-11-15

    New pyrazolo[3,4-d]pyrimidines were synthesized and found to inhibit Src phosphorylation in a cell-free assay. Some of them significantly reduced the growth of human osteogenic sarcoma (SaOS-2) cells. The best compound, in terms of inhibitory properties toward both Src and SaOS-2 cells, was further investigated and found to reduce bone resorption when used to treat mouse osteoclasts, without interfering with normal osteoblast growth. Moreover, its metabolic stability prompted its study on a human SaOS-2 xenograft tumor model in nude mice, where the compound reduced significantly both the volume and weight of the tumor. These experimental findings make the new compound an interesting hit in the field of bone-related diseases. PMID:17929792

  6. Intracellular dynamics and fate of polystyrene nanoparticles in A549 Lung epithelial cells monitored by image (cross-) correlation spectroscopy and single particle tracking.

    PubMed

    Deville, Sarah; Penjweini, Rozhin; Smisdom, Nick; Notelaers, Kristof; Nelissen, Inge; Hooyberghs, Jef; Ameloot, Marcel

    2015-10-01

    Novel insights in nanoparticle (NP) uptake routes of cells, their intracellular trafficking and subcellular targeting can be obtained through the investigation of their temporal and spatial behavior. In this work, we present the application of image (cross-) correlation spectroscopy (IC(C)S) and single particle tracking (SPT) to monitor the intracellular dynamics of polystyrene (PS) NPs in the human lung carcinoma A549 cell line. The ensemble kinetic behavior of NPs inside the cell was characterized by temporal and spatiotemporal image correlation spectroscopy (TICS and STICS). Moreover, a more direct interpretation of the diffusion and flow detected in the NP motion was obtained by SPT by monitoring individual NPs. Both techniques demonstrate that the PS NP transport in A549 cells is mainly dependent on microtubule-assisted transport. By applying spatiotemporal image cross-correlation spectroscopy (STICCS), the correlated motions of NPs with the early endosomes, late endosomes and lysosomes are identified. PS NPs were equally distributed among the endolysosomal compartment during the time interval of the experiments. The cotransport of the NPs with the lysosomes is significantly larger compared to the other cell organelles. In the present study we show that the complementarity of ICS-based techniques and SPT enables a consistent elaborate model of the complex behavior of NPs inside biological systems.

  7. Detection of DNA adducts of benzo[a]pyrene using immunoelectrophoresis with laser-induced fluorescence. Analysis of A549 cells.

    PubMed

    Tan, W G; Carnelley, T J; Murphy, P; Wang, H; Lee, J; Barker, S; Weinfeld, M; Le, X C

    2001-07-27

    Detection of benzo[a]pyrene diol epoxide (BPDE)-damaged DNA in a human lung carcinoma cell line (A549) has been performed using free zone affinity capillary electrophoresis with laser-induced fluorescence (LIF). Using BPDE as a model carcinogenic compound, the speed, sensitivity and specificity of this technique was demonstrated. Under free zone conditions, an antibody bound adduct was baseline-resolved from an unbound adduct in less than 2 min. The efficiencies of separation were in excess of 6 x 10(5) and 1 x 10(6) plates per meter for the antibody-bound and unbound adducts, respectively. Separation using a low ionic strength buffer permitted the use of a high electric field (830 V/cm) without the loss of resolving power. Using LIF detection, a concentration detection limit of roughly 3 x 10(-10) M was achieved for a 90-mer oligonuleotide containing a single BDPE. The use of formamide in the incubation buffer to enhance denaturing of DNA did not affect the stability of the complex between the antibody and the adducts. Using a fluorescently labeled BPDE-modified DNA adduct probe, a competitive assay was established to determine the levels of BPDE-DNA adducts in A549 cells.

  8. Preclinical evaluation of the anti-tumor effects of the natural isoflavone genistein in two xenograft mouse models monitored by [18F]FDG, [18F]FLT, and [64Cu]NODAGA-cetuximab small animal PET

    PubMed Central

    Honndorf, Valerie S.; Wiehr, Stefan; Rolle, Anna-Maria; Schmitt, Julia; Kreft, Luisa; Quintanilla-Martinez, Letitia; Kohlhofer, Ursula; Reischl, Gerald; Maurer, Andreas; Boldt, Karsten; Schwarz, Michael; Schmidt, Holger; Pichler, Bernd J.

    2016-01-01

    The natural phytoestrogen genistein is known as protein kinase inhibitor and tumor suppressor in various types of cancers. We studied its antitumor effect in two different xenograft models using positron emission tomography (PET) in vivo combined with ex vivo histology and nuclear magnetic resonance (NMR) metabolic fingerprinting. Procedures A431 and Colo205 tumor-bearing mice were treated with vehicle or genistein (500 mg/kg/d) over a period of 12 days. Imaging was performed with 2-deoxy-2-[18F]fluoro-D-glucose ([18F]FDG) and 3′-deoxy-3′-[18F]fluorothymidine ([18F] FLT). In a second study A431 tumor-bearing mice were treated with vehicle, genistein (500 mg/kg/d), cetuximab (1mg/3d) or a combination of the compounds and imaged using [18F]FDG, [18F]FLT and [64Cu]NODAGA-cetuximab. Data were compared to histology and principal components analysis (PCA) of NMR fingerprinting data. Results Genistein reduced tumor growth significantly in both xenografts. [18F] FLT uptake was consistent in both models and corresponded to histological findings and also PCA whereas [18F]FDG and [64Cu]NODAGA-cetuximab were not suitable for therapy monitoring. Conclusions As mono-therapy the natural isoflavone genistein has a powerful therapeutic effect in vivo on A431 and Colo205 tumors. [18F]FLT has superior consistency compared to the other tested tracers in therapy monitoring, while the treatment effect could be shown on the molecular level by histology and metabolic fingerprinting. PMID:27070087

  9. Therapeutic Silencing of Bcl-2 by Systemically Administered siRNA Nanotherapeutics Inhibits Tumor Growth by Autophagy and Apoptosis and Enhances the Efficacy of Chemotherapy in Orthotopic Xenograft Models of ER (−) and ER (+) Breast Cancer

    PubMed Central

    Tekedereli, Ibrahim; Alpay, S Neslihan; Akar, Ugur; Yuca, Erkan; Ayugo-Rodriguez, Cristian; Han, He-Dong; Sood, Anil K; Lopez-Berestein, Gabriel; Ozpolat, Bulent

    2013-01-01

    Bcl-2 is overexpressed in about a half of human cancers and 50–70% of breast cancer patients, thereby conferring resistance to conventional therapies and making it an excellent therapeutic target. Small interfering RNA (siRNA) offers novel and powerful tools for specific gene silencing and molecularly targeted therapy. Here, we show that therapeutic silencing of Bcl-2 by systemically administered nanoliposomal (NL)-Bcl-2 siRNA (0.15 mg siRNA/kg, intravenous) twice a week leads to significant antitumor activity and suppression of growth in both estrogen receptor-negative (ER(−)) MDA-MB-231 and ER-positive (+) MCF7 breast tumors in orthotopic xenograft models (P < 0.05). A single intravenous injection of NL-Bcl-2-siRNA provided robust and persistent silencing of the target gene expression in xenograft tumors. NL-Bcl-2-siRNA treatment significantly increased the efficacy of chemotherapy when combined with doxorubicin in both MDA-MB-231 and MCF-7 animal models (P < 0.05). NL-Bcl-2-siRNA treatment-induced apoptosis and autophagic cell death, and inhibited cyclin D1, HIF1α and Src/Fak signaling in tumors. In conclusion, our data provide the first evidence that in vivo therapeutic targeting Bcl-2 by systemically administered nanoliposomal-siRNA significantly inhibits growth of both ER(−) and ER(+) breast tumors and enhances the efficacy of chemotherapy, suggesting that therapeutic silencing of Bcl-2 by siRNA is a viable approach in breast cancers. PMID:24022053

  10. The feasibility of 18F-AlF-NOTA-PRGD2 PET/CT for monitoring early response of Endostar antiangiogenic therapy in human nasopharyngeal carcinoma xenograft model compared with 18F-FDG

    PubMed Central

    Liang, Sheng; Zhang, Caiyuan; Cheng, Weiwei; Hai, Wangxi; Yin, Bing; Wang, Dengbin

    2016-01-01

    Purpose Radiolabeled arginine-glycine-aspartic acid (RGD) peptides have been developed for PET imaging of integrin avβ3 in the tumor vasculature, leading to great potential for noninvasively evaluating tumor angiogenesis and monitoring antiangiogenic treatment. The aim of this study was to investigate a novel one-step labeled integrin-targeted tracer, 18F-AlF-NOTA-PRGD2, for PET/CT for detecting tumor angiogenesis and monitoring the early therapeutic efficacy of antiangiogenic agent Endostar in human nasopharyngeal carcinoma (NPC) xenograft model. Experimental design and results Mice bearing NPC underwent 18F-AlF-NOTA-PRGD2 PET/CT at baseline and after 2, 4, 7, and 14 days of consecutive treatment with Endostar or PBS, compared with 18F-FDG PET/CT. Tumors were harvested at all imaging time points for histopathological analysis with H & E and microvessel density (MVD) and integrin avβ3 immunostaining. The maximum percent injected dose per gram of body weight (%ID/gmax) tumor uptake of 18F-AlF-NOTA-PRGD2 PET/CT was significantly lower than that in the control group starting from day 2 (p < 0.01), much earlier and more accurately than that of 18F-FDG PET/CT. Moreover, a moderate linear correlation was observed between tumor MVD and the corresponding tumor uptake of 18F-AlF-NOTA-PRGD2 PET/CT (r = 0.853, p < 0.01). Conclusions 18F-AlF-NOTA-PRGD2 PET/CT can be used for in vivo angiogenesis imaging and monitoring early response to Endostar antiangiogenic treatment in NPC xenograft model, favoring its potential clinical translation. PMID:27029065

  11. Evaluation of Protein Profiles From Treated Xenograft Tumor Models Identifies an Antibody Panel for Formalin-fixed and Paraffin-embedded (FFPE) Tissue Analysis by Reverse Phase Protein Arrays (RPPA)*

    PubMed Central

    Bader, Sabine; Zajac, Magdalena; Friess, Thomas; Ruge, Elisabeth; Rieder, Natascha; Gierke, Berthold; Heubach, Yvonne; Thomas, Marlene; Pawlak, Michael

    2015-01-01

    Reverse phase protein arrays (RPPA) are an established tool for measuring the expression and activation status of multiple proteins in parallel using only very small amounts of tissue. Several studies have demonstrated the value of this technique for signaling pathway analysis using proteins extracted from fresh frozen (FF) tissue in line with validated antibodies for this tissue type; however, formalin fixation and paraffin embedding (FFPE) is the standard method for tissue preservation in the clinical setting. Hence, we performed RPPA to measure profiles for a set of 300 protein markers using matched FF and FFPE tissue specimens to identify which markers performed similarly using the RPPA technique in fixed and unfixed tissues. Protein lysates were prepared from matched FF and FFPE tissue specimens of individual tumors taken from three different xenograft models of human cancer. Materials from both untreated mice and mice treated with either anti-HER3 or bispecific anti-IGF-1R/EGFR monoclonal antibodies were analyzed. Correlations between signals from FF and FFPE tissue samples were investigated. Overall, 60 markers were identified that produced comparable profiles between FF and FFPE tissues, demonstrating significant correlation between the two sample types. The top 25 markers also showed significance after correction for multiple testing. The panel of markers covered several clinically relevant tumor signaling pathways and both phosphorylated and nonphosphorylated proteins were represented. Biologically relevant changes in marker expression were noted when RPPA profiles from treated and untreated xenografts were compared. These data demonstrate that, using appropriately selected antibodies, RPPA analysis from FFPE tissue is well feasible and generates biologically meaningful information. The identified panel of markers that generate similar profiles in matched fixed and unfixed tissue samples may be clinically useful for pharmacodynamic studies of drug effect

  12. King cobra (Ophiophagus hannah) venom L-amino acid oxidase induces apoptosis in PC-3 cells and suppresses PC-3 solid tumor growth in a tumor xenograft mouse model.

    PubMed

    Lee, Mui Li; Fung, Shin Yee; Chung, Ivy; Pailoor, Jayalakshmi; Cheah, Swee Hung; Tan, Nget Hong

    2014-01-01

    King cobra (Ophiophagus hannah) venom L-amino acid oxidase (OH-LAAO), a heat stable enzyme, has been shown to exhibit very potent anti-proliferative activity against human breast and lung tumorigenic cells but not in their non-tumorigenic counterparts. We further examine its in vitro and in vivo anti-tumor activity in a human prostate adenocarcinoma (PC-3) model. OH-LAAO demonstrated potent cytotoxicity against PC-3 cells with IC50 of 0.05 µg/mL after 72 h incubation in vitro. It induced apoptosis as evidenced with an increase in caspase-3/7 cleavages and an increase in annexin V-stained cells. To examine its in vivo anti-tumor activity, we treated PC-3 tumor xenograft implanted subcutaneously in immunodeficient NU/NU (nude) mice with 1 µg/g OH-LAAO given intraperitoneally (i.p.). After 8 weeks of treatment, OH-LAAO treated PC-3 tumors were markedly inhibited, when compared to the control group (P <0.05). TUNEL staining analysis on the tumor sections showed a significantly increase of apoptotic cells in the LAAO-treated animals. Histological examinations of the vital organs in these two groups showed no significant differences with normal tissues, indicating no obvious tissue damage. The treatment also did not cause any significant changes on the body weight of the mice during the duration of the study. These observations suggest that OH-LAAO cytotoxic effects may be specific to tumor xenografts and less to normal organs. Given its potent anti-tumor activities shown in vitro as well as in vivo, the king cobra venom LAAO can potentially be developed to treat prostate cancer and other solid tumors. PMID:24782648

  13. King cobra (Ophiophagus hannah) venom L-amino acid oxidase induces apoptosis in PC-3 cells and suppresses PC-3 solid tumor growth in a tumor xenograft mouse model.

    PubMed

    Lee, Mui Li; Fung, Shin Yee; Chung, Ivy; Pailoor, Jayalakshmi; Cheah, Swee Hung; Tan, Nget Hong

    2014-01-01

    King cobra (Ophiophagus hannah) venom L-amino acid oxidase (OH-LAAO), a heat stable enzyme, has been shown to exhibit very potent anti-proliferative activity against human breast and lung tumorigenic cells but not in their non-tumorigenic counterparts. We further examine its in vitro and in vivo anti-tumor activity in a human prostate adenocarcinoma (PC-3) model. OH-LAAO demonstrated potent cytotoxicity against PC-3 cells with IC50 of 0.05 µg/mL after 72 h incubation in vitro. It induced apoptosis as evidenced with an increase in caspase-3/7 cleavages and an increase in annexin V-stained cells. To examine its in vivo anti-tumor activity, we treated PC-3 tumor xenograft implanted subcutaneously in immunodeficient NU/NU (nude) mice with 1 µg/g OH-LAAO given intraperitoneally (i.p.). After 8 weeks of treatment, OH-LAAO treated PC-3 tumors were markedly inhibited, when compared to the control group (P <0.05). TUNEL staining analysis on the tumor sections showed a significantly increase of apoptotic cells in the LAAO-treated animals. Histological examinations of the vital organs in these two groups showed no significant differences with normal tissues, indicating no obvious tissue damage. The treatment also did not cause any significant changes on the body weight of the mice during the duration of the study. These observations suggest that OH-LAAO cytotoxic effects may be specific to tumor xenografts and less to normal organs. Given its potent anti-tumor activities shown in vitro as well as in vivo, the king cobra venom LAAO can potentially be developed to treat prostate cancer and other solid tumors.

  14. Evaluation of Protein Profiles From Treated Xenograft Tumor Models Identifies an Antibody Panel for Formalin-fixed and Paraffin-embedded (FFPE) Tissue Analysis by Reverse Phase Protein Arrays (RPPA).

    PubMed

    Bader, Sabine; Zajac, Magdalena; Friess, Thomas; Ruge, Elisabeth; Rieder, Natascha; Gierke, Berthold; Heubach, Yvonne; Thomas, Marlene; Pawlak, Michael

    2015-10-01

    Reverse phase protein arrays (RPPA) are an established tool for measuring the expression and activation status of multiple proteins in parallel using only very small amounts of tissue. Several studies have demonstrated the value of this technique for signaling pathway analysis using proteins extracted from fresh frozen (FF) tissue in line with validated antibodies for this tissue type; however, formalin fixation and paraffin embedding (FFPE) is the standard method for tissue preservation in the clinical setting. Hence, we performed RPPA to measure profiles for a set of 300 protein markers using matched FF and FFPE tissue specimens to identify which markers performed similarly using the RPPA technique in fixed and unfixed tissues. Protein lysates were prepared from matched FF and FFPE tissue specimens of individual tumors taken from three different xenograft models of human cancer. Materials from both untreated mice and mice treated with either anti-HER3 or bispecific anti-IGF-1R/EGFR monoclonal antibodies were analyzed. Correlations between signals from FF and FFPE tissue samples were investigated. Overall, 60 markers were identified that produced comparable profiles between FF and FFPE tissues, demonstrating significant correlation between the two sample types. The top 25 markers also showed significance after correction for multiple testing. The panel of markers covered several clinically relevant tumor signaling pathways and both phosphorylated and nonphosphorylated proteins were represented. Biologically relevant changes in marker expression were noted when RPPA profiles from treated and untreated xenografts were compared. These data demonstrate that, using appropriately selected antibodies, RPPA analysis from FFPE tissue is well feasible and generates biologically meaningful information. The identified panel of markers that generate similar profiles in matched fixed and unfixed tissue samples may be clinically useful for pharmacodynamic studies of drug effect

  15. Reduced 64Cu Uptake and Tumor Growth Inhibition by Knockdown of Human Copper Transporter 1 in Xenograft Mouse Model of Prostate Cancer

    PubMed Central

    Cai, Huawei; Wu, Jiu-sheng; Muzik, Otto; Hsieh, Jer-Tsong; Lee, Robert J.; Peng, Fangyu

    2015-01-01

    Copper is an element required for cell proliferation and angiogenesis. Human prostate cancer xenografts with increased 64Cu radioactivity were visualized previously by PET using 64CuCl2 as a radiotracer (64CuCl2 PET). This study aimed to determine whether the increased tumor 64Cu radioactivity was due to increased cellular uptake of 64Cu mediated by human copper transporter 1 (hCtr1) or simply due to nonspecific binding of ionic 64CuCl2 to tumor tissue. In addition, the functional role of hCtr1 in proliferation of prostate cancer cells and tumor growth was also assessed. Methods A lentiviral vector encoding short-hairpin RNA specific for hCtr1 (Lenti-hCtr1-shRNA) was constructed for RNA interference–mediated knockdown of hCtr1 expression in prostate cancer cells. The degree of hCtr1 knockdown was determined by Western blot, and the effect of hCtr1 knockdown on copper uptake and proliferation were examined in vitro by cellular 64Cu uptake and cell proliferation assays. The effects of hCtr1 knockdown on tumor uptake of 64Cu were determined by PET quantification and tissue radioactivity assay. The effects of hCtr1 knockdown on tumor growth were assessed by PET/CT and tumor size measurement with a caliper. Results RNA interference–mediated knockdown of hCtr1 was associated with the reduced cellular uptake of 64Cu and the suppression of prostate cancer cell proliferation in vitro. At 24 h after intravenous injection of the tracer 64CuCl2, the 64Cu uptake by the tumors with knockdown of hCtr1 (4.02 ± 0.31 percentage injected dose per gram [%ID/g] in Lenti-hCtr1-shRNA-PC-3 and 2.30 ± 0.59 %ID/g in Lenti-hCtr1-shRNA-DU-145) was significantly lower than the 64Cu uptake by the control tumors without knockdown of hCtr1 (7.21 ± 1.48 %ID/g in Lenti-SCR-shRNA-PC-3 and 5.57 ± 1.20 % ID/g in Lenti-SCR-shRNA-DU-145, P < 0.001) by PET quantification. Moreover, the volumes of prostate cancer xenograft tumors with knockdown of hCtr1 (179 ± 111 mm3 for Lenti-hCtr1-shRNA-PC-3

  16. Winter fine particulate matter from Milan induces morphological and functional alterations in human pulmonary epithelial cells (A549).

    PubMed

    Gualtieri, Maurizio; Mantecca, Paride; Corvaja, Viviana; Longhin, Eleonora; Perrone, Maria Grazia; Bolzacchini, Ezio; Camatini, Marina

    2009-07-10

    Samples of PM(2.5) were gravimetrically collected during the winter 2005/2006 in the urban area of Milan (North Italy). Samples were chemically characterized and the particles were detached from filters to determine their cytotoxic effects on the A549 cell line. Based on the potential toxicological relevance of its components, Milan winter PM(2.5) contained high concentrations of pro-oxidant transition metals and PAHs, while re-suspended particles showed a relatively high frequency of dimensional classes ranging from 40 nm to 300 nm. A549 cells exposed to particle suspensions showed a concentration-dependent decrease in viability, starting from 10 microg/cm(2). Phagocytosis of particles by A549 cells and particle aggregates were morphologically characterized and seemed to depend on both particle concentration and exposure time, with the majority of particles being engulfed in membrane-bound vacuoles after 24h of exposure. The ability of ultrafine particles to penetrate and spread throughout the cells was also verified. Cell membrane lysis and mitochondrial ultrastructural disruption appeared to be the main modifications induced by PM(2.5) on A549 cells. Concomitantly to the adverse effects observed in terms of cell mortality and ultrastructural lesions, a significant intracellular production of reactive oxygen species (ROS) was observed, suggesting that the cytotoxicity, exerted by the winter PM(2.5) in Milan, derived also from its oxidative potential, probably associated with particle-adsorbed metals and PAHs. PMID:19433270

  17. The synergistic effect of resveratrol in combination with cisplatin on apoptosis via modulating autophagy in A549 cells.

    PubMed

    Hu, Song; Li, Xiaolin; Xu, Rongrong; Ye, Lingyun; Kong, Hui; Zeng, Xiaoning; Wang, Hong; Xie, Weiping

    2016-06-01

    Several studies have shown that combination treatment with natural products and chemotherapy agents can improve the sensitivity and cytotoxicity of chemotherapy agents. Resveratrol, a natural product, has many biological effects including antitumor and antiviral activities, as well as vascular protective effect. The aim of this study is to investigate the synergistic anticancer effect of resveratrol in combination with cisplatin and the potential anticancer mechanisms involved in A549 cells. The results obtained from Cell Counting Kit-8 and isobolographic analysis demonstrated that combination of resveratrol and cisplatin resulted in synergistic cytotoxic effects in A549 cells. Results from Hoechst staining, flow cytometry and western blot analysis suggested that resveratrol enhanced cisplatin-mediated apoptosis. Meanwhile, the changes of LC3-II and P62 levels and formation of autophagosome suggested that resveratrol in combination with cisplatin triggered autophagy. More importantly, inhibiting autophagy by 3-methyladenine markedly attenuated the apoptosis caused by combination of resveratrol and cisplatin in A549 cells. Taken together, our study provides the first evidence that resveratrol combined with cisplatin synergistically induce apoptosis via modulating autophagic cell death in A549 cells. These findings also help us to understand the role of natural products in combination with chemotherapy agents in lung cancer.

  18. Induction of release and up-regulated gene expression of interleukin (IL)-8 in A549 cells by serine proteinases

    PubMed Central

    Wang, Haiyan; Zheng, Yanshan; He, Shaoheng

    2006-01-01

    Background Hypersecretion of cytokines and serine proteinases has been observed in asthma. Since protease-activated receptors (PARs) are receptors of several serine proteinases and airway epithelial cells are a major source of cytokines, the influence of serine proteinases and PARs on interleukin (IL)-8 secretion and gene expression in cultured A549 cells was examined. Results A549 cells express all four PARs at both protein and mRNA levels as assessed by flow cytometry, immunofluorescence microscopy and reverse transcription polymerase chain reaction (PCR). Thrombin, tryptase, elastase and trypsin induce a up to 8, 4.3, 4.4 and 5.1 fold increase in IL-8 release from A549 cells, respectively following 16 h incubation period. The thrombin, elastase and trypsin induced secretion of IL-8 can be abolished by their specific inhibitors. Agonist peptides of PAR-1, PAR-2 and PAR-4 stimulate up to 15.6, 6.6 and 3.5 fold increase in IL-8 secretion, respectively. Real time PCR shows that IL-8 mRNA is up-regulated by the serine proteinases tested and by agonist peptides of PAR-1 and PAR-2. Conclusion The proteinases, possibly through activation of PARs can stimulate IL-8 release from A549 cells, suggesting that they are likely to contribute to IL-8 related airway inflammatory disorders in man. PMID:16696869

  19. Apoptosis of human lung adenocarcinoma A549 cells induced by prodigiosin analogue obtained from an entomopathogenic bacterium Serratia marcescens.

    PubMed

    Zhou, Wei; Jin, Zhi-Xiong; Wan, Yong-Ji

    2010-12-01

    An entomopathogenic bacterial strain SCQ1 was isolated from silkworm (Bombyx mori) and identified as Serratia marcescens via 16S rRNA gene analysis. This strain produces a red pigment that causes acute septicemia of silkworm. The red pigment of strain SCQ1 was identified as prodigiosin analogue (PGA) with various reported biological activities. In this study, we found that low concentration of PGA showed significant anticancer activity in human lung adenocarcinoma A549 cells, but has little effect in human bone marrow stem cells, in vitro. By exposure to different concentrations of PGA for 24 h, morphological changes and the MTT assay showed that A549 cell line was very sensitive to PGA, with IC(50) value about 2.2 mg/L. Early stage of apoptosis was detected by flow cytometry while A549 cells were treated with PGA for 4 and 12 h, respectively. The proportion of dead cells was increased with treatment time or the concentrations of PGA, but it was inversely proportional to that of apoptotic cells. These results indicate that PGA obtained from strain SCQ1 induces apoptosis in A549 cells, but the molecular mechanisms of cell death are complicated, and the S. marcescens strain SCQ1 may serve as a source of the anticancer compound, PGA.

  20. Antimetastatic Effects of Phyllanthus on Human Lung (A549) and Breast (MCF-7) Cancer Cell Lines

    PubMed Central

    Lee, Sau Har; Jaganath, Indu Bala; Wang, Seok Mui; Sekaran, Shamala Devi

    2011-01-01

    Background Current chemotherapeutic drugs kill cancer cells mainly by inducing apoptosis. However, they become ineffective once cancer cell has the ability to metastasize, hence the poor prognosis and high mortality rate. Therefore, the purpose of this study was to evaluate the antimetastatic potential of Phyllanthus (P. niruri, P. urinaria, P. watsonii, and P. amarus) on lung and breast carcinoma cells. Methodology/Principal Findings Cytotoxicity of Phyllanthus plant extracts were first screened using the MTS reduction assay. They were shown to inhibit MCF-7 (breast carcinoma) and A549 (lung carcinoma) cells growth with IC50 values ranging from 50–180 µg/ml and 65–470 µg/ml for methanolic and aqueous extracts respectively. In comparison, they have lower toxicity on normal cells with the cell viability percentage remaining above 50% when treated up to 1000 µg/ml for both extracts. After determining the non-toxic effective dose, several antimetastasis assays were carried out and Phyllanthus extracts were shown to effectively reduce invasion, migration, and adhesion of both MCF-7 and A549 cells in a dose-dependent manner, at concentrations ranging from 20–200 µg/ml for methanolic extracts and 50–500 µg/ml for aqueous extracts. This was followed by an evaluation of the possible modes of cell death that occurred along with the antimetastatic activity. Phyllanthus was shown to be capable of inducing apoptosis in conjunction with its antimetastastic action, with more than three fold increase of caspases-3 and -7, the presence of DNA-fragmentation and TUNEL-positive cells. The ability of Phyllanthus to exert antimetastatic activities is mostly associated to the presence of polyphenol compounds in its extracts. Conclusions/Significance The presence of polyphenol compounds in the Phyllanthus plant is critically important in the inhibition of the invasion, migration, and adhesion of cancer cells, along with the involvement of apoptosis induction. Hence

  1. Anticancer activity of pyrithione zinc in oral cancer cells identified in small molecule screens and xenograft model: Implications for oral cancer therapy.

    PubMed

    Srivastava, Gunjan; Matta, Ajay; Fu, Guodong; Somasundaram, Raj Thani; Datti, Alessandro; Walfish, Paul G; Ralhan, Ranju

    2015-10-01

    Oral squamous cell carcinoma (OSCC) patients diagnosed in late stages have limited chemotherapeutic options, underscoring the great need for development of new anticancer agents for more effective disease management. We aimed to identify novel anticancer agents for OSCC using quantitative high throughput assays for screening six chemical libraries consisting of 5170 small molecule inhibitors. In depth characterization resulted in identification of pyrithione zinc (PYZ) as the most effective cytotoxic agent inhibiting cell proliferation and inducing apoptosis in OSCC cells in vitro. Further, treatment with PYZ reduced colony forming, migration and invasion potential of oral cancer cells in a dose-dependent manner. PYZ treatment also led to altered expression of several key components of the major signaling pathways including PI3K/AKT/mTOR and WNT/β-catenin in OSCC cells. In addition, treatment with PYZ also reduced expression of 14-3-3ζ, 14-3-3σ, cyclin D1, c-Myc and pyruvate kinase M2 (PKM2), proteins identified in our earlier studies to be involved in development and progression of OSCCs. Importantly, PYZ treatment significantly reduced tumor xenograft volume in immunocompromised NOD/SCID/Crl mice without causing apparent toxicity to normal tissues. Taken together, we demonstrate in vitro and in vivo efficacy of PYZ in OSCC. In conclusion, we identified PYZ in HTS assays and demonstrated in vitro and in vivo pre-clinical efficacy of PYZ as a novel anticancer therapeutic candidate in OSCC. PMID:26115765

  2. Tunicamycin potentiates cisplatin anticancer efficacy through the DPAGT1/Akt/ABCG2 pathway in mouse Xenograft models of human hepatocellular carcinoma.

    PubMed

    Hou, Helei; Sun, Hefen; Lu, Ping; Ge, Chao; Zhang, Lixing; Li, Hong; Zhao, Fangyu; Tian, Hua; Zhang, Lin; Chen, Taoyang; Yao, Ming; Li, Jinjun

    2013-12-01

    Hepatocellular carcinoma is highly chemoresistant, and ATP-binding cassette subfamily G member 2 (ABCG2) is thought to play a critical role in this drug resistance. The present study aims to develop effective therapeutic strategies to decrease ABCG2 expression level and to surmount drug resistance in hepatocellular carcinoma chemotherapy. First, we verified a positive correlation between the ABCG2 protein level and the drug resistance of hepatocellular carcinoma cell lines. ABCG2 was preferentially expressed in highly chemoresistant hepatocellular carcinoma cancer stem cells (CSC) enriched with CD133. In addition, ABCG2 was N-linked glycosylated in hepatocellular carcinoma cells, and this modification was involved in sustaining its protein stability. The N-linked glycosylation (NLG) inhibitor tunicamycin dramatically reduced ABCG2 expression, altered its subcellular localization, and reversed its drug efflux effect in multiple hepatocellular carcinoma cell lines. Furthermore, tunicamycin reduced the expression levels of several CSC markers and suppressed the tumorigenicity of CD133(+) CSCs. Tunicamycin combined with cisplatin (CDDP) inhibited proliferating cell nuclear antigen (PCNA) expression and increased the cleavage of PARP; this effect was partially rescued by the overexpression of ABCG2 or Akt-myr. The combination therapy more effectively suppressed tumor growth in xenograft mice than did single-agent therapy with either drug. Finally, the CDDP treatment combined with UDP-GlcNAc-dolichol-phosphate N-acetylglucosamine-1 phosphate transferase (DPAGT1) knockdown recapitulated the effect observed when CDDP was used in combination with tunicamycin. In summary, our results suggest that tunicamycin may reverse the drug resistance and improve the efficacy of combination treatments for hepatocellular carcinomas by targeting the DPAGT1/Akt/ABCG2 pathway.

  3. Capsaicin-induced genotoxic stress does not promote apoptosis in A549 human lung and DU145 prostate cancer cells.

    PubMed

    Lewinska, Anna; Jarosz, Paulina; Czech, Joanna; Rzeszutek, Iwona; Bielak-Zmijewska, Anna; Grabowska, Wioleta; Wnuk, Maciej

    2015-02-01

    Capsaicin is the major pungent component of the hot chili peppers of the genus Capsicum, which are consumed worldwide as a food additive. More recently, the selective action of capsaicin against cancer cells has been reported. Capsaicin was found to induce apoptosis and inhibit proliferation of a wide range of cancer cells in vitro, whereas being inactive against normal cells. As data on capsaicin-induced genotoxicity are limited and the effects of capsaicin against human lung A549 and DU145 prostate cancer cells were not explored in detail, we were interested in determining whether capsaicin-associated genotoxicity may also provoke A549 and DU145 cell death. Capsaicin-induced decrease in metabolic activity and cell proliferation, and changes in the cell cycle were limited to high concentrations used (≥ 100 μM), whereas, at lower concentrations, capsaicin stimulated both DNA double strand breaks and micronuclei production. Capsaicin was unable to provoke apoptotic cell death when used up to 250 μM concentrations. Capsaicin induced oxidative stress, but was ineffective in provoking the dissipation of the mitochondrial inner transmembrane potential. A different magnitude of p53 binding protein 1 (53BP1) recruitment contributed to diverse capsaicin-induced genotoxic effects in DU145 and A549 cells. Capsaicin was also found to be a DNA hypermethylating agent in A549 cells. In summary, we have shown that genotoxic effects of capsaicin may contribute to limited susceptibility of DU145 and A549 cancer cells to apoptosis in vitro, which may question the usefulness of capsaicin-based anticancer therapy, at least in a case of lung and prostate cancer. PMID:25813723

  4. MicroRNA-490-3p inhibits proliferation of A549 lung cancer cells by targeting CCND1

    SciTech Connect

    Gu, Haihua; Yang, Tao; Fu, Shaozi; Chen, Xiaofan; Guo, Lei; Ni, Yiming

    2014-01-31

    Highlights: • We examined the level of miR-490-3p in A549 lung cancer cells compared with normal bronchial epithelial cell line. • We are the first to show the function of miR-490-3p in A549 lung cancer cells. • We demonstrate CCND1 may be one of the targets of miR-490-3p. - Abstract: MicroRNAs (miRNAs) are small non-coding RNAs that negatively regulate the translation of messenger RNAs by binding their 3′-untranslated region (3′UTR). In this study, we found that miR-490-3p is significantly down-regulated in A549 lung cancer cells compared with the normal bronchial epithelial cell line. To better characterize the role of miR-490-3p in A549 cells, we performed a gain-of-function analysis by transfecting the A549 cells with chemically synthesized miR-490-3P mimics. Overexpression of miR-490-3P evidently inhibits cell proliferation via G1-phase arrest. We also found that forced expression of miR-490-3P decreased both mRNA and protein levels of CCND1, which plays a key role in G1/S phase transition. In addition, the dual-luciferase reporter assays indicated that miR-490-3P directly targets CCND1 through binding its 3′UTR. These findings indicated miR-490-3P could be a potential suppressor of cellular proliferation.

  5. Inhibition of lung cancer cells A549 and H460 by curcuminoid extracts and nanoemulsions prepared from Curcuma longa Linnaeus.

    PubMed

    Chang, Hong-Bin; Chen, Bing-Huei

    2015-01-01

    The objectives of this study were to explore the inhibition mechanism of lung cancer cells A549 and H460 by curcuminoid extracts and nanoemulsions prepared from Curcuma longa Linnaeus. In addition, human bronchus epithelial cell line BEAS-2B (normal cell) was selected for comparison. A high-performance liquid chromatography (HPLC) method was developed to separate and quantify the various curcuminoids in C. longa extract, including curcumin (1,714.5 μg/mL), demethoxycurcumin (1,147.4 μg/mL), and bisdemethoxycurcumin (190.2 μg/mL). A high-stability nanoemulsion composed of Tween 80, water, and curcuminoid extract was prepared, with mean particle size being 12.6 nm. The cell cycle was retarded at G2/M for both the curcuminoid extract and nanoemulsion treatments; however, the inhibition pathway may be different. H460 cells were more susceptible to apoptosis than A549 cells for both curcuminoid extract and nanoemulsion treatments. Growth of BEAS-2B remained unaffected for both the curcuminoid extract and nanoemulsion treatments, with a concentration range from 1 to 4 μg/mL. Also, the activities of caspase-3, caspase-8, and caspase-9 followed a dose-dependent increase for both A549 and H460 cells for both the treatments, accompanied by a dose-dependent increase in cytochrome C expression and a dose-dependent decrease in CDK1 expression. Interestingly, a dose-dependent increase in cyclin B expression was shown for A549 cells for both the treatments, while a reversed trend was found for H460 cells. Both mitochondria and death receptor pathways may be responsible for apoptosis of both A549 and H460 cells.

  6. Cabazitaxel-induced autophagy via the PI3K/Akt/mTOR pathway contributes to A549 cell death.

    PubMed

    Huo, Ruichao; Wang, Lili; Liu, Peijuan; Zhao, Yong; Zhang, Caiqin; Bai, Bing; Liu, Xueying; Shi, Changhong; Wei, Sanhua; Zhang, Hai

    2016-10-01

    Cabazitaxel has been used to treat castration-resistant prostate cancer since its approval by the US Food and Drug Administration in 2010. However, whether cabazitaxel may inhibit the proliferation of other tissue‑derived cancer cells, and its underlying mechanism, remains unknown. In the present study, the A549 lung adenocarcinoma cancer cell line was exposed to cabazitaxel, in order to investigate its cytotoxic effect and determine the underlying mechanism. The results demonstrated that cabazitaxel was able to induce autophagy in A549 cells, as evidenced by the formation of autophagosomes, upregulated LC3‑II expression and increased LC3 puncta. Cabazitaxel‑induced autophagy had a cytotoxic effect on A549 cells, as evidenced by the induction of cell death and cell cycle arrest at G2/M phase, which was independent of the apoptotic pathway. Furthermore, transfection with Beclin1 small interfering RNA and treatment with the autophagy inhibitor 3‑methyladenine protected cells from cabazitaxel‑induced cell death, thus confirming that cabazitaxel‑induced autophagy contributed to A549 cell death. In addition, cabazitaxel targeted the phosphoinositide 3‑kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) pathway to induce autophagy, as indicated by reduced phosphorylation of Akt and mTOR. In conclusion, the present study demonstrated that cabazitaxel exerts a cytotoxic effect on A549 cells by acting on the PI3K/Akt/mTOR pathway to promote autophagic cell death. This result supports the potential use of cabazitaxel as a chemotherapeutic agent for the treatment of lung cancer.

  7. Inhibition of lung cancer cells A549 and H460 by curcuminoid extracts and nanoemulsions prepared from Curcuma longa Linnaeus

    PubMed Central

    Chang, Hong-Bin; Chen, Bing-Huei

    2015-01-01

    The objectives of this study were to explore the inhibition mechanism of lung cancer cells A549 and H460 by curcuminoid extracts and nanoemulsions prepared from Curcuma longa Linnaeus. In addition, human bronchus epithelial cell line BEAS-2B (normal cell) was selected for comparison. A high-performance liquid chromatography (HPLC) method was developed to separate and quantify the various curcuminoids in C. longa extract, including curcumin (1,714.5 μg/mL), demethoxycurcumin (1,147.4 μg/mL), and bisdemethoxycurcumin (190.2 μg/mL). A high-stability nanoemulsion composed of Tween 80, water, and curcuminoid extract was prepared, with mean particle size being 12.6 nm. The cell cycle was retarded at G2/M for both the curcuminoid extract and nanoemulsion treatments; however, the inhibition pathway may be different. H460 cells were more susceptible to apoptosis than A549 cells for both curcuminoid extract and nanoemulsion treatments. Growth of BEAS-2B remained unaffected for both the curcuminoid extract and nanoemulsion treatments, with a concentration range from 1 to 4 μg/mL. Also, the activities of caspase-3, caspase-8, and caspase-9 followed a dose-dependent increase for both A549 and H460 cells for both the treatments, accompanied by a dose-dependent increase in cytochrome C expression and a dose-dependent decrease in CDK1 expression. Interestingly, a dose-dependent increase in cyclin B expression was shown for A549 cells for both the treatments, while a reversed trend was found for H460 cells. Both mitochondria and death receptor pathways may be responsible for apoptosis of both A549 and H460 cells. PMID:26345201

  8. Capsaicin-induced genotoxic stress does not promote apoptosis in A549 human lung and DU145 prostate cancer cells.

    PubMed

    Lewinska, Anna; Jarosz, Paulina; Czech, Joanna; Rzeszutek, Iwona; Bielak-Zmijewska, Anna; Grabowska, Wioleta; Wnuk, Maciej

    2015-02-01

    Capsaicin is the major pungent component of the hot chili peppers of the genus Capsicum, which are consumed worldwide as a food additive. More recently, the selective action of capsaicin against cancer cells has been reported. Capsaicin was found to induce apoptosis and inhibit proliferation of a wide range of cancer cells in vitro, whereas being inactive against normal cells. As data on capsaicin-induced genotoxicity are limited and the effects of capsaicin against human lung A549 and DU145 prostate cancer cells were not explored in detail, we were interested in determining whether capsaicin-associated genotoxicity may also provoke A549 and DU145 cell death. Capsaicin-induced decrease in metabolic activity and cell proliferation, and changes in the cell cycle were limited to high concentrations used (≥ 100 μM), whereas, at lower concentrations, capsaicin stimulated both DNA double strand breaks and micronuclei production. Capsaicin was unable to provoke apoptotic cell death when used up to 250 μM concentrations. Capsaicin induced oxidative stress, but was ineffective in provoking the dissipation of the mitochondrial inner transmembrane potential. A different magnitude of p53 binding protein 1 (53BP1) recruitment contributed to diverse capsaicin-induced genotoxic effects in DU145 and A549 cells. Capsaicin was also found to be a DNA hypermethylating agent in A549 cells. In summary, we have shown that genotoxic effects of capsaicin may contribute to limited susceptibility of DU145 and A549 cancer cells to apoptosis in vitro, which may question the usefulness of capsaicin-based anticancer therapy, at least in a case of lung and prostate cancer.

  9. Cabazitaxel-induced autophagy via the PI3K/Akt/mTOR pathway contributes to A549 cell death

    PubMed Central

    Huo, Ruichao; Wang, Lili; Liu, Peijuan; Zhao, Yong; Zhang, Caiqin; Bai, Bing; Liu, Xueying; Shi, Changhong; Wei, Sanhua; Zhang, Hai

    2016-01-01

    Cabazitaxel has been used to treat castration-resistant prostate cancer since its approval by the US Food and Drug Administration in 2010. However, whether cabazitaxel may inhibit the proliferation of other tissue-derived cancer cells, and its underlying mechanism, remains unknown. In the present study, the A549 lung adenocarcinoma cancer cell line was exposed to cabazitaxel, in order to investigate its cytotoxic effect and determine the underlying mechanism. The results demonstrated that cabazitaxel was able to induce autophagy in A549 cells, as evidenced by the formation of autophagosomes, upregulated LC3-II expression and increased LC3 puncta. Cabazitaxel-induced autophagy had a cytotoxic effect on A549 cells, as evidenced by the induction of cell death and cell cycle arrest at G2/M phase, which was independent of the apoptotic pathway. Furthermore, transfection with Beclin1 small interfering RNA and treatment with the autophagy inhibitor 3-methyladenine protected cells from cabazitaxel-induced cell death, thus confirming that cabazitaxel-induced autophagy contributed to A549 cell death. In addition, cabazitaxel targeted the phosphoinositide 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) pathway to induce autophagy, as indicated by reduced phosphorylation of Akt and mTOR. In conclusion, the present study demonstrated that cabazitaxel exerts a cytotoxic effect on A549 cells by acting on the PI3K/Akt/mTOR pathway to promote autophagic cell death. This result supports the potential use of cabazitaxel as a chemotherapeutic agent for the treatment of lung cancer. PMID:27572899

  10. Nanoparticles of Selaginella doederleinii leaf extract inhibit human lung cancer cells A549

    NASA Astrophysics Data System (ADS)

    Syaefudin; Juniarti, A.; Rosiyana, L.; Setyani, A.; Khodijah, S.

    2016-01-01

    The aim of the present study is to evaluate cytotoxicity effect of nanoparticles of Selaginella doederleinii (S. doederleinii) leaves extract. S. doederleinii was extracted by maceration method using 70%(v/v) ethanol as solvent. Phytochemical content was analyzed qualitatively by using Harborne and Thin Layer Chromatography (TLC) methods. Nanoparticle extract was prepared by ionic gelation using chitosan as encapsulant agent. Anticancer activity was performed by using 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay. The results showed that S. doederleinii contains of flavonoids. Nanoparticle of S. doederleinii leaves extract greatly inhibited A549 cells growth (cancer cells), with IC50 of 3% or 1020 μg/ml. These nanoparticles extract also inhibited the growth of Chang cells (normal cells), with IC50 of 4% or 1442 μg/ml. The effective concentration of nanoparticles extract which inhibits cancer cells without harming the normal cells is 0.5% or 167 μg/ml. Further studies are needed to obtain the concentration of nanoparticles extract which can selectively suppress cancer cells.

  11. Cytotoxic Effect of a Novel Synthesized Carbazole Compound on A549 Lung Cancer Cell Line

    PubMed Central

    Molatlhegi, Refilwe P.; Phulukdaree, Alisa; Anand, Krishnan; Gengan, Robert M.; Tiloke, Charlette; Chuturgoon, Anil A.

    2015-01-01

    Increased death rates due to lung cancer have necessitated the search for potential novel anticancer compounds such as carbazole derivatives. Carbazoles are aromatic heterocyclic compounds with anticancer, antibacterial and anti-inflammatory activity. The study investigated the ability of the novel carbazole compound (Z)-4-[9-ethyl-9aH-carbazol-3-yl) amino] pent-3-en-2-one (ECAP) to induce cytotoxicity of lung cancer cells and its mechanism of action. ECAP was synthesized as a yellow powder with melting point of 240-247 °C. The 3-(4,5-dimethythiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT), lipid peroxidation and comet assays were used to assess the cytotoxic effect of the compound on A549 lung cancer cells. Protein expression was determined using western blots, apoptosis was measured by luminometry (caspase-3/7, -8 and -9) assay and flow cytometry was used to measure phosphatidylserine (PS) externalisation. ECAP induced a p53 mediated apoptosis of lung cancer cells due to a significant reduction in the expression of antioxidant defence proteins (Nrf2 and SOD), Hsp70 (p < 0.02) and Bcl-2 (p < 0.0006), thereby up-regulating reactive oxygen species (ROS) production. This resulted in DNA damage (p < 0.0001), up-regulation of Bax expression and caspase activity and induction of apoptosis in lung cancer cells. The results show the anticancer potential of ECAP on lung cancer. PMID:26134408

  12. Taxol-induced paraptosis-like A549 cell death is not senescence

    NASA Astrophysics Data System (ADS)

    Wang, Chao-yang; Chen, Tong-Sheng

    2011-03-01

    Our previous studies have shown that taxol, a potent anticancer agent, induces caspase-independent cell death and cytoplasmic vacuolization in human lung cancer cells. However, the mechanisms of taxol-induced cytoplasmic vacuolization are poorly understood. Cytoplasmic vacuolization have been reported to be a characteristic of cell senescence. Here, we employed confocal fluorescence microscopy imaging to study the reversibility of taxol-induced cytoplasmic vacuolization and whether taxol triggers senescence in A549 cells. We found that taxol-induced cytoplasmic vacuolization at 6 or 9 h after treatment with taxol did not decrease but increase at 24 h or 72 h after refreshing the culture medium without taxol, indicating taxol-induced cytoplasmic vacuolization is irreversible. We used SA-β-Gal (senescence-associated β-galactosidase) to assess whether taxol-induced cell death in cytoplasmic vacuolization fashion is senescence, and found that hydrogen peroxide (H2O2)-treated, but not taxol-treated cells is significantly stained by the SA-β-Gal, a senescence testing kit, indicating that the form of taxol-induced cell death is not senescence.

  13. Raw and thermally treated cement asbestos exerts different cytotoxicity effects on A549 cells in vitro.

    PubMed

    Pugnaloni, Armanda; Lucarini, Guendalina; Rubini, Corrado; Smorlesi, Arianna; Tomasetti, Marco; Strafella, Elisabetta; Armeni, Tatiana; Gualtieri, Alessandro F

    2015-01-01

    Raw cement asbestos (RCA) undergoes a complete solid state transformation when heated at high temperatures. The secondary raw material produced, high temperatures-cement asbestos (HT-CA) is composed of newly-formed crystals in place of the asbestos fibers present in RCA. Our previous study showed that HT-CA exerts lower cytotoxic cell damage compared to RCA. Nevertheless further investigations are needed to deepen our understanding of pathogenic pathways involving oxidative and nitrative damage. Our aim is to deepen the understanding of the biological effects on A549 cells of these materials regarding DNA damage related proteins (p53, its isoform p73 and TRAIL) and nitric oxide (NO) production during inducible nitric oxide synthase (iNOS)-mediated inflammation. Increments of p53/p73 expression, iNOS positive cells and NO concentrations were found with RCA, compared to HT-CA and controls mainly at 48 h. Interestingly, ferrous iron causing reactive oxygen species (ROS)-mediated DNA damage was found in RCA as a contaminant. HT-CA thermal treatment induces a global recrystallization with iron in a crystal form poorly released in media. HT-CA slightly interferes with genome expression and exerts lower inflammatory potential compared to RCA on biological systems. It could represent a safe approach for storing or recycling asbestos and an environmentally friendly alternative to asbestos waste.

  14. Biosynthesis of gold nanoparticles and related cytotoxicity evaluation using A549 cells.

    PubMed

    Sathishkumar, M; Pavagadhi, S; Mahadevan, A; Balasubramanian, R

    2015-04-01

    Biosynthesis of gold nanoparticles (AuNPs) has become an attractive area of research as it is environmentally benign. The toxicity of AuNPs synthesized by chemical routes has been widely studied. However, little is known about the toxicity associated with the biological synthesis of AuNPs. The present study was carried out to synthesize AuNPs using star anise (Illicium verum; a commercially available spice in abundance)and evaluate its toxicity using human epithelial lung cells (A549) in comparison with AuNPs synthesized by the traditional chemical methods (using sodium citrate and sodium borohydride). Apart from cell viability, markers of oxidative stress (reduced glutathione) and cell death (caspases) were also evaluated to understand the mechanisms of toxicity. Cell viability was observed to be 65.7 percent and 72.3 percent in cells exposed to chemically synthesized AuNPs at the highest dose (200nM) as compared to 80.2 percent for biologically synthesized AuNPs. Protective coating/capping of AuNPs by various polyphenolic compounds present in star anise extract appears to be a major contributor to lower toxicity observed in biologically synthesized AuNPs.

  15. Therapeutic effect against human xenograft tumors in nude mice by the third generation microtubule stabilizing epothilones.

    PubMed

    Chou, Ting-Chao; Zhang, Xiuguo; Zhong, Zi-Yang; Li, Yong; Feng, Li; Eng, Sara; Myles, David R; Johnson, Robert; Wu, Nian; Yin, Ye Ingrid; Wilson, Rebecca M; Danishefsky, Samuel J

    2008-09-01

    The epothilones represent a promising class of natural product-based antitumor drug candidates. Although these compounds operate through a microtubule stabilization mechanism similar to that of taxol, the epothilones offer a major potential therapeutic advantage in that they retain their activity against multidrug-resistant cell lines. We have been systematically synthesizing and evaluating synthetic epothilone congeners that are not accessible through modification of the natural product itself. We report herein the results of biological investigations directed at two epothilone congeners: iso-fludelone and iso-dehydelone. Iso-fludelone, in particular, exhibits a number of properties that render it an excellent candidate for preclinical development, including biological stability, excellent solubility in water, and remarkable potency relative to other epothilones. In nude mouse xenograft settings, iso-fludelone was able to achieve therapeutic cures against a number of human cancer cell lines, including mammarian-MX-1, ovarian-SK-OV-3, and the fast-growing, refractory, subcutaneous neuroblastoma-SK-NAS. Strong therapeutic effect was observed against drug-resistant lung-A549/taxol and mammary-MCF-7/Adr xenografts. In addition, iso-fludelone was shown to exhibit a significant therapeutic effect against an intracranially implanted SK-NAS tumor. PMID:18755900

  16. Improving In Vivo High-Resolution CT Imaging of the Tumour Vasculature in Xenograft Mouse Models through Reduction of Motion and Bone-Streak Artefacts

    PubMed Central

    Kersemans, Veerle; Kannan, Pavitra; Beech, John S.; Bates, Russell; Irving, Benjamin; Gilchrist, Stuart; Allen, Philip D.; Thompson, James; Kinchesh, Paul; Casteleyn, Christophe; Schnabel, Julia; Partridge, Mike; Muschel, Ruth J.; Smart, Sean C.

    2015-01-01

    Introduction Preclinical in vivo CT is commonly used to visualise vessels at a macroscopic scale. However, it is prone to many artefacts which can degrade the quality of CT images significantly. Although some artefacts can be partially corrected for during image processing, they are best avoided during acquisition. Here, a novel imaging cradle and tumour holder was designed to maximise CT resolution. This approach was used to improve preclinical in vivo imaging of the tumour vasculature. Procedures A custom built cradle containing a tumour holder was developed and fix-mounted to the CT system gantry to avoid artefacts arising from scanner vibrations and out-of-field sample positioning. The tumour holder separated the tumour from bones along the axis of rotation of the CT scanner to avoid bone-streaking. It also kept the tumour stationary and insensitive to respiratory motion. System performance was evaluated in terms of tumour immobilisation and reduction of motion and bone artefacts. Pre- and post-contrast CT followed by sequential DCE-MRI of the tumour vasculature in xenograft transplanted mice was performed to confirm vessel patency and demonstrate the multimodal capacity of the new cradle. Vessel characteristics such as diameter, and branching were quantified. Results Image artefacts originating from bones and out-of-field sample positioning were avoided whilst those resulting from motions were reduced significantly, thereby maximising the resolution that can be achieved with CT imaging in vivo. Tumour vessels ≥ 77 μm could be resolved and blood flow to the tumour remained functional. The diameter of each tumour vessel was determined and plotted as histograms and vessel branching maps were created. Multimodal imaging using this cradle assembly was preserved and demonstrated. Conclusions The presented imaging workflow minimised image artefacts arising from scanner induced vibrations, respiratory motion and radiopaque structures and enabled in vivo CT imaging

  17. Three-dimensional quantitative structure-activity relationship study on anti-cancer activity of 3,4-dihydroquinazoline derivatives against human lung cancer A549 cells

    NASA Astrophysics Data System (ADS)

    Cho, Sehyeon; Choi, Min Ji; Kim, Minju; Lee, Sunhoe; Lee, Jinsung; Lee, Seok Joon; Cho, Haelim; Lee, Kyung-Tae; Lee, Jae Yeol

    2015-03-01

    A series of 3,4-dihydroquinazoline derivatives with anti-cancer activities against human lung cancer A549 cells were subjected to three-dimensional quantitative structure-activity relationship (3D-QSAR) studies using the comparative molecular similarity indices analysis (CoMSIA) approaches. The most potent compound, 1 was used to align the molecules. As a result, the best prediction was obtained with CoMSIA combined the steric, electrostatic, hydrophobic, hydrogen bond donor, and hydrogen bond acceptor fields (q2 = 0.720, r2 = 0.897). This model was validated by an external test set of 6 compounds giving satisfactory predictive r2 value of 0.923 as well as the scrambling stability test. This model would guide the design of potent 3,4-dihydroquinazoline derivatives as anti-cancer agent for the treatment of human lung cancer.

  18. Derivation of sperm from xenografted testis cells and tissues of the peccary (Tayassu tajacu).

    PubMed

    Campos-Junior, Paulo Henrique Almeida; Costa, Guilherme Mattos Jardim; Avelar, Gleide Fernandes; Lacerda, Samyra Maria Santos Nassif; da Costa, Nathália Nogueira; Ohashi, Otávio Mitio; Miranda, Moysés dos Santos; Barcelos, Lucíola Silva; Jorge, Erika Cristina; Guimarães, Diva Anelie; de França, Luiz Renato

    2014-03-01

    Because the collared peccary (Tayassu tajacu) has a peculiar Leydig cell cytoarchitecture, this species represents a unique mammalian model for investigating testis function. Taking advantage of the well-established and very useful testis xenograft technique, in the present study, testis tissue and testis cell suspensions from immature collared peccaries (n=4; 3 months old) were xenografted in SCID mice (n=48) and evaluated at 2, 4, 6, and 8 months after grafting. Complete spermatogenesis was observed at 6 and 8 months after testis tissue xenografting. However, probably due to de novo testis morphogenesis and low androgen secretion, functionally evaluated by the seminal vesicle weight, a delay in spermatogenesis progression was observed in the testis cell suspension xenografts, with the production of fertile sperm only at 8 months after grafting. Importantly, demonstrating that the peculiar testicular cytoarchitecture of the collared peccary is intrinsically programmed, the unique Leydig cell arrangement observed in this species was re-established after de novo testis morphogenesis. The sperm collected from the xenografts resulted in diploid embryos that expressed the paternally imprinted gene NNAT after ICSI. The present study is the first to demonstrate complete spermatogenesis with the production of fertile sperm from testis cell suspension xenografts in a wild mammalian species. Therefore, due to its unique testicular cytoarchitecture, xenograft techniques, particularly testis cell suspensions, may represent a new and very promising approach to evaluate testis morphogenesis and to investigate spermatogonial stem cell physiology and niche in the collared peccary.

  19. Erlotinib Inhibits Growth of a Patient-Derived Chordoma Xenograft

    PubMed Central

    Siu, I-Mei; Ruzevick, Jacob; Zhao, Qi; Connis, Nick; Jiao, Yuchen; Bettegowda, Chetan; Xia, Xuewei; Burger, Peter C.; Hann, Christine L.; Gallia, Gary L.

    2013-01-01

    Chordomas are rare primary bone tumors that occur along the neuraxis. Primary treatment is surgery, often followed by radiotherapy. Treatment options for patients with recurrence are limited and, notably, there are no FDA approved therapeutic agents. Development of therapeutic options has been limited by the paucity of preclinical model systems. We have established and previously reported the initial characterization of the first patient-derived chordoma xenograft model. In this study, we further characterize this model and demonstrate that it continues to resemble the original patient tumor histologically and immunohistochemically, maintains nuclear expression of brachyury, and is highly concordant with the original patient tumor by whole genome genotyping. Pathway analysis of this xenograft demonstrates activation of epidermal growth factor receptor (EGFR). In vitro studies demonstrate that two small molecule inhibitors of EGFR, erlotinib and gefitinib, inhibit proliferation of the chordoma cell line U-CH 1. We further demonstrate that erlotinib significantly inhibits chordoma growth in vivo. Evaluation of tumors post-treatment reveals that erlotinib reduces phosphorylation of EGFR. This is the first demonstration of antitumor activity in a patient-derived chordoma xenograft model and these findings support further evaluation of EGFR inhibitors in this disease. PMID:24260133

  20. Erlotinib inhibits growth of a patient-derived chordoma xenograft.

    PubMed

    Siu, I-Mei; Ruzevick, Jacob; Zhao, Qi; Connis, Nick; Jiao, Yuchen; Bettegowda, Chetan; Xia, Xuewei; Burger, Peter C; Hann, Christine L; Gallia, Gary L

    2013-01-01

    Chordomas are rare primary bone tumors that occur along the neuraxis. Primary treatment is surgery, often followed by radiotherapy. Treatment options for patients with recurrence are limited and, notably, there are no FDA approved therapeutic agents. Development of therapeutic options has been limited by the paucity of preclinical model systems. We have established and previously reported the initial characterization of the first patient-derived chordoma xenograft model. In this study, we further characterize this model and demonstrate that it continues to resemble the original patient tumor histologically and immunohistochemically, maintains nuclear expression of brachyury, and is highly concordant with the original patient tumor by whole genome genotyping. Pathway analysis of this xenograft demonstrates activation of epidermal growth factor receptor (EGFR). In vitro studies demonstrate that two small molecule inhibitors of EGFR, erlotinib and gefitinib, inhibit proliferation of the chordoma cell line U-CH 1. We further demonstrate that erlotinib significantly inhibits chordoma growth in vivo. Evaluation of tumors post-treatment reveals that erlotinib reduces phosphorylation of EGFR. This is the first demonstration of antitumor activity in a patient-derived chordoma xenograft model and these findings support further evaluation of EGFR inhibitors in this disease.

  1. Comparison of 18F-FES, 18F-FDG, and 18F-FMISO PET Imaging Probes for Early Prediction and Monitoring of Response to Endocrine Therapy in a Mouse Xenograft Model of ER-Positive Breast Cancer

    PubMed Central

    Yang, ZhongYi; Zhang, JianPing; Zhang, YongPing; Luo, JianMin; Zhang, YingJian

    2016-01-01

    Background There is an increasing need to characterize biological processes for early prediction and monitoring of response to endocrine therapy in breast cancer using multiple positron emission tomography (PET) imaging probes. However, use of more than two PET tracers in a single clinical trial is quite challenging. In this study we carried out a longitudinal investigation of 18F-FES, 18F-FDG, and 18F-FMISO PET imaging probes for early prediction and monitoring of response to endocrine therapy in a mouse xenograft model of estrogen receptor (ER)-positive breast cancer. Method ER+ human breast cancer ZR-75-1 models were established in female mice that were then randomly assigned to a treatment (fulvestrant, 5.0 mg/week for 21 days) or vehicle group. Micro-PET/CT imaging with 18F-FES, 18F-FDG, and 18F-FMISO was performed on days 0, 3, 14, and 21 after treatment. The uptake value (percentage injected dose per gram, %ID/g) for each probe in tumor (T) tissue and contralateral muscle (M) was measured for quantitative analysis and T/M calculation. Tumor volume was measured to record tumor growth at each time point. Tumor tissues were sampled for immunohistochemical staining of ER expression. Correlations for tumor volume and ERα levels with uptake data for the probe were tested. Results Uptake data for 18F-FES in ZR-75-1 tumor tissues corresponded well with tumor response to endocrine therapy, but not for 18F-FDG and 18F-FMISO, according to longitudinal micro-PET/CT imaging and quantitative correlation analysis. There was a significant positive correlation between 18F-FES uptake and ER levels (%ID/gmax r2 = 0.76, P< 0.05; T/M r2 = 0.82, P<0.05). Notably, 18F-FES uptake on day 3 was significantly correlated with the day 21/baseline tumor volume ratio (%ID/gmax r2 = 0.74, P < 0.05; T/M r2 = 0.78, P < 0.05). Conclusions Comparison of 18F-FES, 18F-FDG, and 18F-FMISO probes revealed that 18F-FES PET/CT molecular imaging can provide a precise early prediction of tumor

  2. Effects of Per2 overexpression on growth inhibition and metastasis, and on MTA1, nm23-H1 and the autophagy-associated PI3K/PKB signaling pathway in nude mice xenograft models of ovarian cancer.

    PubMed

    Wang, Zhaoxia; Li, Li; Wang, Yang

    2016-06-01

    The aim of the present study was to evaluate the association between Period2 (Per2) and the occurrence and development of ovarian cancer, in addition to evaluating the effect of this gene on the growth and metastasis of ovarian cancer in nude mice xenograft models. The detection of Per2 by reverse transcription‑quantitative polymerase chain reaction (RT‑qPCR) and western blotting methods at various stages of ovarian cancer in tumor tissue samples was conducted. Nude mice xenograft models of ovarian cancer were constructed using an ovarian cancer cell line and, using a gene transfection technique, exogenous infusion of the recombinant gene, Per2, was performed. To assess for the successful and stable expression of Per2 in the tumor tissue, levels of Per2 expression in the nude mice xenograft models were detected by RT‑qPCR. During the experimental period, the tumor volumes were measured every three days. Two weeks following treatment cessation, the nude mice were sacrificed and the tumor weight and volume were measured. Furthermore, detection of the changes in expression levels of metastasis‑associated gene 1 (MTA‑1) and tumor metastasis suppressor gene, non‑metastasis protein 23‑H1 (nm23‑H1), and the expression change of autophagy‑associated signal transduction pathway, phosphatidylinositol 3‑kinase (PI3K)/protein kinase B (PKB) kinase were analyzed. The findings demonstrated that with ovarian cancer stage development, the expression of Per2 gradually reduced or ceased. In addition, exogenous Per2 was successfully and stably expressed in nude mice tumor tissue samples. Furthermore, in the Per2 overexpression group, MTA‑1 protein expression was significantly reduced when compared with the phosphate‑buffered saline (PBS) control and empty plasmid groups, while nm23‑H1 protein expression was significantly higher when compared with those two groups. The expression levels of PI3K and PKB kinase, which are marker proteins of the autophagy

  3. Novel C-17-heteroaryl steroidal CYP17 inhibitors/antiandrogens: synthesis, in vitro biological activity, pharmacokinetics, and antitumor activity in the LAPC4 human prostate cancer xenograft model.

    PubMed

    Handratta, Venkatesh D; Vasaitis, Tadas S; Njar, Vincent C O; Gediya, Lalji K; Kataria, Ritesh; Chopra, Pankaj; Newman, Donnell; Farquhar, Rena; Guo, Zhiyong; Qiu, Yun; Brodie, Angela M H

    2005-04-21

    tentatively identified as 17-(1H-benzimidazol-1-yl)androsta-3-one. When tested in vivo, 5 proved to be very effective at inhibiting the growth of androgen-dependent LAPC4 human prostate tumor xenograft, while 6 was ineffective. Compound 5 (50 mg/kg/twice daily) resulted in a 93.8% reduction (P = 0.00065) in the mean final tumor volume compared with controls, and it was also significantly more effective than castration. To our knowledge, this is the first example of an antihormonal agent (an inhibitor of androgen synthesis (CYP17 inhibitor)/antiandrogen) that is significantly more effective than castration in suppression of androgen-dependent prostate tumor growth. In view of these impressive anticancer properties, compound 5 is a strong candidate for development for the treatment of human prostate cancer.

  4. Suitable parameter choice on quantitative morphology of A549 cell in epithelial–mesenchymal transition

    PubMed Central

    Ren, Zhou-Xin; Yu, Hai-Bin; Li, Jian-Sheng; Shen, Jun-Ling; Du, Wen-Sen

    2015-01-01

    Evaluation of morphological changes in cells is an integral part of study on epithelial to mesenchymal transition (EMT), however, only a few papers reported the changes in quantitative parameters and no article compared different parameters for demanding better parameters. In the study, the purpose was to investigate suitable parameters for quantitative evaluation of EMT morphological changes. A549 human lung adenocarcinoma cell line was selected for the study. Some cells were stimulated by transforming growth factor-β1 (TGF-β1) for EMT, and other cells were as control without TGF-β1 stimulation. Subsequently, cells were placed in phase contrast microscope and three arbitrary fields were captured and saved with a personal computer. Using the tools of Photoshop software, some cells in an image were selected, segmented out and exchanged into unique hue, and other part in the image was shifted into another unique hue. The cells were calculated with 29 morphological parameters by Image Pro Plus software. A parameter between cells with or without TGF-β1 stimulation was compared statistically and nine parameters were significantly different between them. Receiver operating characteristic curve (ROC curve) of a parameter was described with SPSS software and F-test was used to compare two areas under the curves (AUCs) in Excel. Among them, roundness and radius ratio were the most AUCs and were significant higher than the other parameters. The results provided a new method with quantitative assessment of cell morphology during EMT, and found out two parameters, roundness and radius ratio, as suitable for quantification. PMID:26182364

  5. Imaging and characterization of stretch-induced ATP release from alveolar A549 cells

    PubMed Central

    Grygorczyk, Ryszard; Furuya, Kishio; Sokabe, Masahiro

    2013-01-01

    Mechano-transduction at cellular and tissue levels often involves ATP release and activation of the purinergic signalling cascade. In the lungs, stretch is an important physical stimulus but its impact on ATP release, the underlying release mechanisms and transduction pathways are poorly understood. Here, we investigated the effect of unidirectional stretch on ATP release from human alveolar A549 cells by real-time luciferin–luciferase bioluminescence imaging coupled with simultaneous infrared imaging, to monitor the extent of cell stretch and to identify ATP releasing cells. In subconfluent (<90%) cell cultures, single 1 s stretch (10–40%)-induced transient ATP release from a small fraction (≤1.5%) of cells that grew in number dose-dependently with increasing extent of stretch. ATP concentration in the proximity (≤150 μm) of releasing cells often exceeded 10 μm, sufficient for autocrine/paracrine purinoreceptor stimulation of neighbouring cells. ATP release responses were insensitive to the putative ATP channel blockers carbenoxolone and 5-nitro-2-(3-phenylpropyl-amino) benzoic acid, but were inhibited by N-ethylmaleimide and bafilomycin. In confluent cell cultures, the maximal fraction of responding cells dropped to <0.2%, but was enhanced several-fold in the wound/scratch area after it was repopulated by new cells during the healing process. Fluo8 fluorescence experiments revealed two types of stretch-induced intracellular Ca2+ responses, rapid sustained Ca2+ elevations in a limited number of cells and delayed secondary responses in neighbouring cells, seen as Ca2+ waves whose propagation was consistent with extracellular diffusion of released ATP. Our experiments revealed that a single >10% stretch was sufficient to initiate intercellular purinergic signalling in alveolar cells, which may contribute to the regulation of surfactant secretion and wound healing. PMID:23247110

  6. Imaging and characterization of stretch-induced ATP release from alveolar A549 cells.

    PubMed

    Grygorczyk, Ryszard; Furuya, Kishio; Sokabe, Masahiro

    2013-03-01

    Abstract  Mechano-transduction at cellular and tissue levels often involves ATP release and activation of the purinergic signalling cascade. In the lungs, stretch is an important physical stimulus but its impact on ATP release, the underlying release mechanisms and transduction pathways are poorly understood. Here, we investigated the effect of unidirectional stretch on ATP release from human alveolar A549 cells by real-time luciferin-luciferase bioluminescence imaging coupled with simultaneous infrared imaging, to monitor the extent of cell stretch and to identify ATP releasing cells. In subconfluent (<90%) cell cultures, single 1 s stretch (10-40%)-induced transient ATP release from a small fraction (1.5%) of cells that grew in number dose-dependently with increasing extent of stretch. ATP concentration in the proximity (150 μm) of releasing cells often exceeded 10 μm, sufficient for autocrine/paracrine purinoreceptor stimulation of neighbouring cells. ATP release responses were insensitive to the putative ATP channel blockers carbenoxolone and 5-nitro-2-(3-phenylpropyl-amino) benzoic acid, but were inhibited by N-ethylmaleimide and bafilomycin. In confluent cell cultures, the maximal fraction of responding cells dropped to <0.2%, but was enhanced several-fold in the wound/scratch area after it was repopulated by new cells during the healing process. Fluo8 fluorescence experiments revealed two types of stretch-induced intracellular Ca(2+) responses, rapid sustained Ca(2+) elevations in a limited number of cells and delayed secondary responses in neighbouring cells, seen as Ca(2+) waves whose propagation was consistent with extracellular diffusion of released ATP. Our experiments revealed that a single >10% stretch was sufficient to initiate intercellular purinergic signalling in alveolar cells, which may contribute to the regulation of surfactant secretion and wound healing.

  7. Hyperoxia enhances VEGF release from A549 cells via post-transcriptional processes

    PubMed Central

    Shenberger, Jeffrey S.; Zhang, Lianqin; Powell, Richard J.; Barchowsky, Aaron

    2007-01-01

    Exposure of animals to hyperoxia decreases lung VEGF mRNA expression concomitant with an acute increase in VEGF protein within the epithelial lining fluid (ELF). The VEGF concentration in ELF is in excess of that found in the plasma, leading to the hypothesis that hyperoxia stimulates the release of VEGF protein from stores within the extracellular matrix. To test this hypothesis in a cell culture system, we exposed A549 cells to 95% O2 for 48 hrs followed by recovery in room air (RA) for 24 hrs. We found that Ox increased VEGF protein 2- to 3-fold within the medium at 48 hrs of exposure and during recovery. Heparin clearing revealed the medium to contain a 50:50 mixture of the heparin-binding (VEGF165) and heparin-non-binding (VEGF121) proteins and Ox to increase both proteins equally. Transcriptional activation of VEGF appears unlikely to explain the increase in VEGF protein as full-length and splice variant VEGF mRNA expression were unchanged by hyperoxia. Analysis of cell-associated VEGF proteins found that Ox increased the expression of VEGF121 and VEGF165 proteins. Blocking binding sites with exogenous heparin enhanced VEGF protein in the medium from RA grown cells while heparinase digestion of bound VEGF revealed a greater reserve of VEGF protein in RA cells. Collectively these findings indicate that hyperoxia enhances the expression of VEGF121/165 proteins and facilitates the release of VEGF165 from cell-associated stores. Increases in VEGF in ELF may represent an adaptive response fostering cell survival and type II cell proliferation in O2-induced lung injury. PMID:17664148

  8. Erlotinib Pretreatment Improves Photodynamic Therapy of Non-Small Cell Lung Carcinoma Xenografts via Multiple Mechanisms.

    PubMed

    Gallagher-Colombo, Shannon M; Miller, Joann; Cengel, Keith A; Putt, Mary E; Vinogradov, Sergei A; Busch, Theresa M

    2015-08-01

    Aberrant expression of the epidermal growth factor receptor (EGFR) is a common characteristic of many cancers, including non-small cell lung carcinoma (NSCLC), head and neck squamous cell carcinoma, and ovarian cancer. Although EGFR is currently a favorite molecular target for the treatment of these cancers, inhibition of the receptor with small-molecule inhibitors (i.e., erlotinib) or monoclonal antibodies (i.e., cetuximab) does not provide long-term therapeutic benefit as standalone treatment. Interestingly, we have found that addition of erlotinib to photodynamic therapy (PDT) can improve treatment response in typically erlotinib-resistant NSCLC tumor xenografts. Ninety-day complete response rates of 63% are achieved when erlotinib is administered in three doses before PDT of H460 human tumor xenografts, compared with 16% after PDT-alone. Similar benefit is found when erlotinib is added to PDT of A549 NCSLC xenografts. Improved response is accompanied by increased vascular shutdown, and erlotinib increases the in vitro cytotoxicity of PDT to endothelial cells. Tumor uptake of the photosensitizer (benzoporphyrin derivative monoacid ring A; BPD) is increased by the in vivo administration of erlotinib; nevertheless, this elevation of BPD levels only partially accounts for the benefit of erlotinib to PDT. Thus, pretreatment with erlotinib augments multiple mechanisms of PDT effect that collectively lead to large improvements in therapeutic efficacy. These data demonstrate that short-duration administration of erlotinib before PDT can greatly improve the responsiveness of even erlotinib-resistant tumors to treatment. Results will inform clinical investigation of EGFR-targeting therapeutics in conjunction with PDT.

  9. SchA-p85-FAK complex dictates isoform-specific activation of Akt2 and subsequent PCBP1-mediated post-transcriptional regulation of TGFβ-mediated epithelial to mesenchymal transition in human lung cancer cell line A549.

    PubMed

    Xue, Xinying; Wang, Xin; Liu, Yuxia; Teng, Guigen; Wang, Yong; Zang, Xuefeng; Wang, Kaifei; Zhang, Jinghui; Xu, Yali; Wang, Jianxin; Pan, Lei

    2014-08-01

    A post-transcriptional pathway by which TGF-β modulates expression of specific proteins, Disabled-2 (Dab2) and Interleukin-like EMT Inducer (ILEI), inherent to epithelial to mesenchymal transition (EMT) in murine epithelial cells through Akt2-mediated phosphorylation of poly r(C) binding protein (PCBP1), has been previously elucidated. The aims of the current study were to determine if the same mechanism is operative in the non-small cell lung cancer (NSCLC) cell line, A549, and to delineate the underlying mechanism. Steady-state transcript and protein expression levels of Dab2 and ILEI were examined in A549 cells treated with TGF-β for up to 48 h. Induction of translational de-repression in this model was quantified by polysomal fractionation followed by qRT-PCR. The underlying mechanism of isoform-specific activation of Akt2 was elucidated through a combination of co-immunoprecipitation studies. TGF-β induced EMT in A549 cells concomitant with translational upregulation of Dab2 and ILEI proteins through isoform-specific activation of Akt2 followed by phosphorylation of PCBP1 at serine-43. Our experiments further elucidated that the adaptor protein SchA is phosphorylated at tyrosine residues following TGF-β treatment, which initiated a signaling cascade resulting in the sequential recruitment of p85 subunit of PI3K and focal adhesion kinase (FAK). The SchA-FAK-p85 complex subsequently selectively recruited and activated Akt2, not Akt1. Inhibition of the p85 subunit through phosphorylated 1257 peptide completely attenuated EMT in these cells. We have defined the underlying mechanism responsible for isoform-specific recruitment and activation of Akt2, not Akt1, during TGF-β-mediated EMT in A549 cells. Inhibition of the formation of this complex thus represents an important and novel therapeutic target in metastatic lung carcinoma. PMID:24819169

  10. Inhibitory effect of ursolic acid and oleanolic acid from Eriobotrya fragrans on A549 cell viability in vivo.

    PubMed

    Gao, Y S; Yuan, Y; Song, G; Lin, S Q

    2016-01-01

    Loquat [Eriobotrya japonica (Lindl.)] is a traditional Chinese medicine, which has been used as an anti-inflammatory and for curing chronic bronchitis among other potential applications. Extracted ursolic acid (UA) and oleanolic acid (OA) from wild loquat were previously found capable of suppressing the proliferation of A549 cells in vitro. In the current study, nude mice were used to determine the inhibitory effect of UA and OA on tumor formation in vivo. The results demonstrate that UA and OA reduced the proliferation of A549 cells in nude mice, and increased the expression of Bid while decreasing the protein levels of MMP-2, Ki-67, and CD34. In this study, we identified potential antitumor activity in a wild loquat extract containing UA and OA, which demonstrates that traditional Chinese medicine may have a role in treating certain types of cancer. PMID:27323036

  11. Inhibitory effect of ursolic acid and oleanolic acid from Eriobotrya fragrans on A549 cell viability in vivo.

    PubMed

    Gao, Y S; Yuan, Y; Song, G; Lin, S Q

    2016-05-13

    Loquat [Eriobotrya japonica (Lindl.)] is a traditional Chinese medicine, which has been used as an anti-inflammatory and for curing chronic bronchitis among other potential applications. Extracted ursolic acid (UA) and oleanolic acid (OA) from wild loquat were previously found capable of suppressing the proliferation of A549 cells in vitro. In the current study, nude mice were used to determine the inhibitory effect of UA and OA on tumor formation in vivo. The results demonstrate that UA and OA reduced the proliferation of A549 cells in nude mice, and increased the expression of Bid while decreasing the protein levels of MMP-2, Ki-67, and CD34. In this study, we identified potential antitumor activity in a wild loquat extract containing UA and OA, which demonstrates that traditional Chinese medicine may have a role in treating certain types of cancer.

  12. Inhibition of signaling between human CXCR4 and zebrafish ligands by the small molecule IT1t impairs the formation of triple-negative breast cancer early metastases in a zebrafish xenograft model