Natural Killer Cell Immunotherapy Targeting Cancer Stem Cells

PubMed Central

Luna, Jesus I; Grossenbacher, Steven K.; Murphy, William J; Canter, Robert J

2017-01-01

Introduction Standard cytoreductive cancer therapy, such as chemotherapy and radiotherapy, are frequently resisted by a small portion of cancer cells with “stem-cell” like properties including quiescence and repopulation. Immunotherapy represents a breakthrough modality for improving oncologic outcomes in cancer patients. Since the success of immunotherapy is not contingent on target cell proliferation, it may also be uniquely suited to address the problem of resistance and repopulation exerted by cancer stem cells (CSCs). Areas covered Natural killer (NK) cells have long been known for their ability to reject allogeneic hematopoietic stem cells, and there are increasing data demonstrating that NK cells can selectively identify and lyse CSCs. In this report, we review the current knowledge of CSCs and NK cells and highlight recent studies that support the concept that NK cells are capable of targeting CSC in solid tumors, especially in the context of combination therapy simultaneously targeting non-CSCs and CSCs. Expert Opinion Unlike cytotoxic cancer treatments, NK cells are able to target and eliminate quiescent/non-proliferating cells such as CSCs, and these enigmatic cells are an important source of relapse and metastasis. NK targeting of CSCs represents a novel and potentially high impact method to capitalize on the intrinsic therapeutic potential of NK cells. PMID:27960589

Orchestration of transplantation tolerance by regulatory dendritic cell therapy or in-situ targeting of dendritic cells.

PubMed

Morelli, Adrian E; Thomson, Angus W

2014-08-01

Extensive research in murine transplant models over the past two decades has convincingly demonstrated the ability of regulatory dendritic cells (DCregs) to promote long-term allograft survival. We review important considerations regarding the source of therapeutic DCregs (donor or recipient) and their mode of action, in-situ targeting of DCregs, and optimal therapeutic regimens to promote DCreg function. Recent studies have defined protocols and mechanisms whereby ex-vivo-generated DCregs of donor or recipient origin subvert allogeneic T-cell responses and promote long-term organ transplant survival. Particular interest has focused on how donor antigen is acquired, processed and presented by autologous dendritic cells, on the stability of DCregs, and on in-situ targeting of dendritic cells to promote their tolerogenic function. New evidence of the therapeutic efficacy of DCregs in a clinically relevant nonhuman primate organ transplant model and production of clinical grade DCregs support early evaluation of DCreg therapy in human graft recipients. We discuss strategies currently used to promote dendritic cell tolerogenicity, including DCreg therapy and in-situ targeting of dendritic cells, with a view to improved understanding of underlying mechanisms and identification of the most promising strategies for therapeutic application.

ESAT-6 Targeting to DEC205+ Antigen Presenting Cells Induces Specific-T Cell Responses against ESAT-6 and Reduces Pulmonary Infection with Virulent Mycobacterium tuberculosis.

PubMed

Silva-Sánchez, Aarón; Meza-Pérez, Selene; Flores-Langarica, Adriana; Donis-Maturano, Luis; Estrada-García, Iris; Calderón-Amador, Juana; Hernández-Pando, Rogelio; Idoyaga, Juliana; Steinman, Ralph M; Flores-Romo, Leopoldo

2015-01-01

Airways infection with Mycobacterium tuberculosis (Mtb) is contained mostly by T cell responses, however, Mtb has developed evasion mechanisms which affect antigen presenting cell (APC) maturation/recruitment delaying the onset of Ag-specific T cell responses. Hypothetically, bypassing the natural infection routes by delivering antigens directly to APCs may overcome the pathogen's naturally evolved evasion mechanisms, thus facilitating the induction of protective immune responses. We generated a murine monoclonal fusion antibody (α-DEC-ESAT) to deliver Early Secretory Antigen Target (ESAT)-6 directly to DEC205+ APCs and to assess its in vivo effects on protection associated responses (IFN-γ production, in vivo CTL killing, and pulmonary mycobacterial load). Treatment with α-DEC-ESAT alone induced ESAT-6-specific IFN-γ producing CD4+ T cells and prime-boost immunization prior to Mtb infection resulted in early influx (d14 post-infection) and increased IFN-γ+ production by specific T cells in the lungs, compared to scarce IFN-γ production in control mice. In vivo CTL killing was quantified in relevant tissues upon transferring target cells loaded with mycobacterial antigens. During infection, α-DEC-ESAT-treated mice showed increased target cell killing in the lungs, where histology revealed cellular infiltrate and considerably reduced bacterial burden. Targeting the mycobacterial antigen ESAT-6 to DEC205+ APCs before infection expands specific T cell clones responsible for early T cell responses (IFN-γ production and CTL activity) and substantially reduces lung bacterial burden. Delivering mycobacterial antigens directly to APCs provides a unique approach to study in vivo the role of APCs and specific T cell responses to assess their potential anti-mycobacterial functions.

ESAT-6 Targeting to DEC205+ Antigen Presenting Cells Induces Specific-T Cell Responses against ESAT-6 and Reduces Pulmonary Infection with Virulent Mycobacterium tuberculosis

PubMed Central

Silva-Sánchez, Aarón; Meza-Pérez, Selene; Flores-Langarica, Adriana; Donis-Maturano, Luis; Estrada-García, Iris; Calderón-Amador, Juana; Hernández-Pando, Rogelio; Idoyaga, Juliana; Flores-Romo, Leopoldo

2015-01-01

Airways infection with Mycobacterium tuberculosis (Mtb) is contained mostly by T cell responses, however, Mtb has developed evasion mechanisms which affect antigen presenting cell (APC) maturation/recruitment delaying the onset of Ag-specific T cell responses. Hypothetically, bypassing the natural infection routes by delivering antigens directly to APCs may overcome the pathogen’s naturally evolved evasion mechanisms, thus facilitating the induction of protective immune responses. We generated a murine monoclonal fusion antibody (α-DEC-ESAT) to deliver Early Secretory Antigen Target (ESAT)-6 directly to DEC205+ APCs and to assess its in vivo effects on protection associated responses (IFN-γ production, in vivo CTL killing, and pulmonary mycobacterial load). Treatment with α-DEC-ESAT alone induced ESAT-6-specific IFN-γ producing CD4+ T cells and prime-boost immunization prior to Mtb infection resulted in early influx (d14 post-infection) and increased IFN-γ+ production by specific T cells in the lungs, compared to scarce IFN-γ production in control mice. In vivo CTL killing was quantified in relevant tissues upon transferring target cells loaded with mycobacterial antigens. During infection, α-DEC-ESAT-treated mice showed increased target cell killing in the lungs, where histology revealed cellular infiltrate and considerably reduced bacterial burden. Targeting the mycobacterial antigen ESAT-6 to DEC205+ APCs before infection expands specific T cell clones responsible for early T cell responses (IFN-γ production and CTL activity) and substantially reduces lung bacterial burden. Delivering mycobacterial antigens directly to APCs provides a unique approach to study in vivo the role of APCs and specific T cell responses to assess their potential anti-mycobacterial functions. PMID:25915045

Cooperative tumour cell membrane targeted phototherapy

NASA Astrophysics Data System (ADS)

Kim, Heegon; Lee, Junsung; Oh, Chanhee; Park, Ji-Ho

2017-06-01

The targeted delivery of therapeutics using antibodies or nanomaterials has improved the precision and safety of cancer therapy. However, the paucity and heterogeneity of identified molecular targets within tumours have resulted in poor and uneven distribution of targeted agents, thus compromising treatment outcomes. Here, we construct a cooperative targeting system in which synthetic and biological nanocomponents participate together in the tumour cell membrane-selective localization of synthetic receptor-lipid conjugates (SR-lipids) to amplify the subsequent targeting of therapeutics. The SR-lipids are first delivered selectively to tumour cell membranes in the perivascular region using fusogenic liposomes. By hitchhiking with extracellular vesicles secreted by the cells, the SR-lipids are transferred to neighbouring cells and further spread throughout the tumour tissues where the molecular targets are limited. We show that this tumour cell membrane-targeted delivery of SR-lipids leads to uniform distribution and enhanced phototherapeutic efficacy of the targeted photosensitizer.

Ablation of proliferating neural stem cells during early life is sufficient to reduce adult hippocampal neurogenesis.

PubMed

Youssef, Mary; Krish, Varsha S; Kirshenbaum, Greer S; Atsak, Piray; Lass, Tamara J; Lieberman, Sophie R; Leonardo, E David; Dranovsky, Alex

2018-05-09

Environmental exposures during early life, but not during adolescence or adulthood, lead to persistent reductions in neurogenesis in the adult hippocampal dentate gyrus (DG). The mechanisms by which early life exposures lead to long-term deficits in neurogenesis remain unclear. Here, we investigated whether targeted ablation of dividing neural stem cells during early life is sufficient to produce long-term decreases in DG neurogenesis. Having previously found that the stem cell lineage is resistant to long-term effects of transient ablation of dividing stem cells during adolescence or adulthood (Kirshenbaum et al., 2014), we used a similar pharmacogenetic approach to target dividing neural stem cells for elimination during early life periods sensitive to environmental insults. We then assessed the Nestin stem cell lineage in adulthood. We found that the adult neural stem cell reservoir was depleted following ablation during the first postnatal week, when stem cells were highly proliferative, but not during the third postnatal week, when stem cells were more quiescent. Remarkably, ablating proliferating stem cells during either the first or third postnatal week led to reduced adult neurogenesis out of proportion to the changes in the stem cell pool, indicating a disruption of the stem cell function or niche following stem cell ablation in early life. These results highlight the first three postnatal weeks as a series of sensitive periods during which elimination of dividing stem cells leads to lasting alterations in adult DG neurogenesis and stem cell function. These findings contribute to our understanding of the relationship between DG development and adult neurogenesis, as well as suggest a possible mechanism by which early life experiences may lead to lasting deficits in adult hippocampal neurogenesis. This article is protected by copyright. All rights reserved. © 2018 Wiley Periodicals, Inc.

A gene expression profile indicative of early stage HER2 targeted therapy response.

PubMed

O'Neill, Fiona; Madden, Stephen F; Clynes, Martin; Crown, John; Doolan, Padraig; Aherne, Sinéad T; O'Connor, Robert

2013-07-01

Efficacious application of HER2-targetting agents requires the identification of novel predictive biomarkers. Lapatinib, afatinib and neratinib are tyrosine kinase inhibitors (TKIs) of HER2 and EGFR growth factor receptors. A panel of breast cancer cell lines was treated with these agents, trastuzumab, gefitinib and cytotoxic therapies and the expression pattern of a specific panel of genes using RT-PCR was investigated as a potential marker of early drug response to HER2-targeting therapies. Treatment of HER2 TKI-sensitive SKBR3 and BT474 cell lines with lapatinib, afatinib and neratinib induced an increase in the expression of RB1CC1, ERBB3, FOXO3a and NR3C1. The response directly correlated with the degree of sensitivity. This expression pattern switched from up-regulated to down-regulated in the HER2 expressing, HER2-TKI insensitive cell line MDAMB453. Expression of the CCND1 gene demonstrated an inversely proportional response to drug exposure. A similar expression pattern was observed following the treatment with both neratinib and afatinib. These patterns were retained following exposure to traztuzumab and lapatinib plus capecitabine. In contrast, gefitinib, dasatinib and epirubicin treatment resulted in a completely different expression pattern change. In these HER2-expressing cell line models, lapatinib, neratinib, afatinib and trastuzumab treatment generated a characteristic and specific gene expression response, proportionate to the sensitivity of the cell lines to the HER2 inhibitor.Characterisation of the induced changes in expression levels of these genes may therefore give a valuable, very early predictor of the likely extent and specificity of tumour HER2 inhibitor response in patients, potentially guiding more specific use of these agents.

Applying cognitive training to target executive functions during early development.

PubMed

Wass, Sam V

2015-01-01

Developmental psychopathology is increasingly recognizing the importance of distinguishing causal processes (i.e., the mechanisms that cause a disease) from developmental outcomes (i.e., the symptoms of the disorder as it is eventually diagnosed). Targeting causal processes early in disordered development may be more effective than waiting until outcomes are established and then trying to reverse the pathogenic process. In this review, I evaluate evidence suggesting that neural and behavioral plasticity may be greatest at very early stages of development. I also describe correlational evidence suggesting that, across a number of conditions, early emerging individual differences in attentional control and working memory may play a role in mediating later-developing differences in academic and other forms of learning. I review the currently small number of studies that applied direct and indirect cognitive training targeted at young individuals and discuss methodological challenges associated with targeting this age group. I also discuss a number of ways in which early, targeted cognitive training may be used to help us understand the developmental mechanisms subserving typical and atypical cognitive development.

Applying cognitive training to target executive functions during early development

PubMed Central

Wass, Sam V.

2015-01-01

Developmental psychopathology is increasingly recognizing the importance of distinguishing causal processes (i.e., the mechanisms that cause a disease) from developmental outcomes (i.e., the symptoms of the disorder as it is eventually diagnosed). Targeting causal processes early in disordered development may be more effective than waiting until outcomes are established and then trying to reverse the pathogenic process. In this review, I evaluate evidence suggesting that neural and behavioral plasticity may be greatest at very early stages of development. I also describe correlational evidence suggesting that, across a number of conditions, early emerging individual differences in attentional control and working memory may play a role in mediating later-developing differences in academic and other forms of learning. I review the currently small number of studies that applied direct and indirect cognitive training targeted at young individuals and discuss methodological challenges associated with targeting this age group. I also discuss a number of ways in which early, targeted cognitive training may be used to help us understand the developmental mechanisms subserving typical and atypical cognitive development. PMID:24511910

Early Hematopoietic Zinc Finger Protein Prevents Tumor Cell Recognition by Natural Killer Cells1

PubMed Central

La Rocca, Rosanna; Fulciniti, Mariateresa; Lakshmikanth, Tadepally; Mesuraca, Maria; Ali, Talib Hassan; Mazzei, Valerio; Amodio, Nicola; Catalano, Lucio; Rotoli, Bruno; Ouerfelli, Ouathek; Grieco, Michele; Gulletta, Elio; Bond, Heather M.; Morrone, Giovanni; Ferrone, Soldano; Carbone, Ennio

2009-01-01

Early hematopoietic zinc finger/zinc finger protein 521 (EHZF/ZNF521) is a novel zinc finger protein expressed in hematopoietic stem and progenitor cells and is down-regulated during their differentiation. Its transcript is also abundant in some hematopoietic malignancies. Analysis of the changes in the antigenic profile of cells transfected with EHZF cDNA revealed up-regulation of HLA class I cell surface expression. This phenotypic change was associated with an increased level of HLA class I H chain, in absence of detectable changes in the expression of other Ag-processing machinery components. Enhanced resistance of target cells to NK cell-mediated cytotoxicity was induced by enforced expression of EHZF in the cervical carcinoma cell line HeLa and in the B lymphoblastoid cell line IM9. Preincubation of transfected cells with HLA class I Ag-specific mAb restored target cell susceptibility to NK cell-mediated lysis, indicating a specific role for HLA class I Ag up-regulation in the NK resistance induced by EHZF. A potential clinical significance of these findings is further suggested by the inverse correlation between EHZF and MHC class I expression levels, and autologous NK susceptibility of freshly explanted multiple myeloma cells. PMID:19342626

The c-Myb target gene neuromedin U functions as a novel cofactor during the early stages of erythropoiesis

PubMed Central

Gambone, Julia E.; Dusaban, Stephanie S.; Loperena, Roxana; Nakata, Yuji

2011-01-01

The requirement of c-Myb during erythropoiesis spurred an interest in identifying c-Myb target genes that are important for erythroid development. Here, we determined that the neuropeptide neuromedin U (NmU) is a c-Myb target gene. Silencing NmU, c-myb, or NmU's cognate receptor NMUR1 expression in human CD34+ cells impaired burst-forming unit-erythroid (BFU-E) and colony-forming unit-erythroid (CFU-E) formation compared with control. Exogenous addition of NmU peptide to NmU or c-myb siRNA-treated CD34+ cells rescued BFU-E and yielded a greater number of CFU-E than observed with control. No rescue of BFU-E and CFU-E growth was observed when NmU peptide was exogenously added to NMUR1 siRNA-treated cells compared with NMUR1 siRNA-treated cells cultured without NmU peptide. In K562 and CD34+ cells, NmU activated protein kinase C-βII, a factor associated with hematopoietic differentiation-proliferation. CD34+ cells cultured under erythroid-inducing conditions, with NmU peptide and erythropoietin added at day 6, revealed an increase in endogenous NmU and c-myb gene expression at day 8 and a 16% expansion of early erythroblasts at day 10 compared to cultures without NmU peptide. Combined, these data strongly support that the c-Myb target gene NmU functions as a novel cofactor for erythropoiesis and expands early erythroblasts. PMID:21378276

Nanoparticles target early-stage breast cancer metastasis in vivo

NASA Astrophysics Data System (ADS)

Goldman, Evgeniya; Zinger, Assaf; da Silva, Dana; Yaari, Zvi; Kajal, Ashima; Vardi-Oknin, Dikla; Goldfeder, Mor; Schroeder, Josh E.; Shainsky-Roitman, Janna; Hershkovitz, Dov; Schroeder, Avi

2017-10-01

Despite advances in cancer therapy, treating cancer after it has metastasized remains an unmet clinical challenge. In this study we demonstrate that 100 nm liposomes target triple-negative murine breast-cancer metastases post intravenous administration. Metastatic breast cancer was induced in BALB/c mice either experimentally, by a tail vein injection of 4T1 cells, or spontaneously, after implanting a primary tumor xenograft. To track their biodistribution in vivo the liposomes were labeled with multi-modal diagnostic agents, including indocyanine green and rhodamine for whole-animal fluorescent imaging, gadolinium for magnetic resonance imaging (MRI), and europium for a quantitative biodistribution analysis. The accumulation of liposomes in the metastases peaked at 24 h post the intravenous administration, similar to the time they peaked in the primary tumor. The efficiency of liposomal targeting to the metastatic tissue exceeded that of a non-liposomal agent by 4.5-fold. Liposomes were detected at very early stages in the metastatic progression, including metastatic lesions smaller than 2 mm in diameter. Surprisingly, while nanoparticles target breast cancer metastasis, they may also be found in elevated levels in the pre-metastatic niche, several days before metastases are visualized by MRI or histologically in the tissue. This study highlights the promise of diagnostic and therapeutic nanoparticles for treating metastatic cancer, possibly even for preventing the onset of the metastatic dissemination by targeting the pre-metastatic niche.

Optoacoustic imaging of gold nanoparticles targeted to breast cancer cells

NASA Astrophysics Data System (ADS)

Eghtedari, Mohammad; Motamedi, Massoud; Popov, Vsevolod L.; Kotov, Nicholas A.; Oraevsky, Alexander A.

2004-07-01

Optoacoustic Tomography (OAT) is a rapidly growing technology that enables noninvasive deep imaging of biological tissues based on their light absorption. In OAT, the interaction of a pulsed laser with tissue increases the temperature of the absorbing components in a confined volume of tissue. Rapid perturbation of the temperature (<1°C) deep within tissue produces weak acoustic waves that easily travel to the surface of the tissue with minor attenuation. Abnormal angiogenesis in a malignant tumor, that increases its blood content, makes a native contrast for optoacoustic imaging; however, the application of OAT for early detection of malignant tumors requires the enhancement of optoacoustic signals originated from tumor by using an exogenous contrast agent. Due to their strong absorption, we have used gold nanoparticles (NP) as a contrast agent. 40nm spherical gold nanoparticles were attached to monoclonal antibody to target cell surface of breast cancer cells. The targeted cancer cells were implanted at depth of 5-6cm within a gelatinous object that optically resembles human breast. Experimental sensitivity measurements along with theoretical analysis showed that our optoacoustic imaging system is capable of detecting a phantom breast tumor with the volume of 0.15ml, which is composed of 25 million NP-targeted cancer cells, at a depth of 5 centimeters in vitro.

Stage-specific control of early B cell development by the transcription factor Ikaros

PubMed Central

Gültekin, Sinan; Dakic, Aleksandar; Axelsson, Elin; Minnich, Martina; Ebert, Anja; Werner, Barbara; Roth, Mareike; Cimmino, Luisa; Dickins, Ross A.; Zuber, Johannes; Jaritz, Markus; Busslinger, Meinrad

2018-01-01

Ikaros is an essential regulator of lymphopoiesis. Here, we studied the B-cell-specific function of Ikaros by conditional Ikzf1 inactivation in pro-B cells. B-cell development was arrested at an aberrant ‘pro-B’ cell stage characterized by increased cell adhesion and loss of pre-B cell receptor signaling. Ikaros was found to activate genes coding for pre-BCR signal transducers and to repress genes involved in the downregulation of pre-BCR signaling and upregulation of the integrin signaling pathway. Unexpectedly, derepression of Aiolos expression could not compensate for the loss of Ikaros in pro-B cells. Ikaros induced or suppressed active chromatin at regulatory elements of activated or repressed target genes. Notably, Ikaros binding and target gene expression was dynamically regulated at distinct stages of early B-lymphopoiesis. PMID:24509509

A gene expression profile indicative of early stage HER2 targeted therapy response

PubMed Central

2013-01-01

Background Efficacious application of HER2-targetting agents requires the identification of novel predictive biomarkers. Lapatinib, afatinib and neratinib are tyrosine kinase inhibitors (TKIs) of HER2 and EGFR growth factor receptors. A panel of breast cancer cell lines was treated with these agents, trastuzumab, gefitinib and cytotoxic therapies and the expression pattern of a specific panel of genes using RT-PCR was investigated as a potential marker of early drug response to HER2-targeting therapies. Results Treatment of HER2 TKI-sensitive SKBR3 and BT474 cell lines with lapatinib, afatinib and neratinib induced an increase in the expression of RB1CC1, ERBB3, FOXO3a and NR3C1. The response directly correlated with the degree of sensitivity. This expression pattern switched from up-regulated to down-regulated in the HER2 expressing, HER2-TKI insensitive cell line MDAMB453. Expression of the CCND1 gene demonstrated an inversely proportional response to drug exposure. A similar expression pattern was observed following the treatment with both neratinib and afatinib. These patterns were retained following exposure to traztuzumab and lapatinib plus capecitabine. In contrast, gefitinib, dasatinib and epirubicin treatment resulted in a completely different expression pattern change. Conclusions In these HER2-expressing cell line models, lapatinib, neratinib, afatinib and trastuzumab treatment generated a characteristic and specific gene expression response, proportionate to the sensitivity of the cell lines to the HER2 inhibitor. Characterisation of the induced changes in expression levels of these genes may therefore give a valuable, very early predictor of the likely extent and specificity of tumour HER2 inhibitor response in patients, potentially guiding more specific use of these agents. PMID:23816254

Targeting Vaccine-Induced Extrafollicular Pathway of B Cell Differentiation Improves Rabies Postexposure Prophylaxis

PubMed Central

Haley, Shannon L.; Tzvetkov, Evgeni P.; Meuwissen, Samantha; Plummer, Joseph R.

2017-01-01

ABSTRACT Vaccine-induced B cells differentiate along two pathways. The follicular pathway gives rise to germinal centers (GCs) that can take weeks to fully develop. The extrafollicular pathway gives rise to short-lived plasma cells (PCs) that can rapidly secrete protective antibodies within days of vaccination. Rabies virus (RABV) postexposure prophylaxis (PEP) requires rapid vaccine-induced humoral immunity for protection. Therefore, we hypothesized that targeting extrafollicular B cell responses for activation would improve the speed and magnitude of RABV PEP. To test this hypothesis, we constructed, recovered, and characterized a recombinant RABV-based vaccine expressing murine B cell activating factor (BAFF) (rRABV-mBAFF). BAFF is an ideal molecule to improve early pathways of B cell activation, as it links innate and adaptive immunity, promoting potent B cell responses. Indeed, rRABV-mBAFF induced a faster, higher antibody response in mice and enhanced survivorship in PEP settings compared to rRABV. Interestingly, rRABV-mBAFF and rRABV induced equivalent numbers of GC B cells, suggesting that rRABV-mBAFF augmented the extrafollicular B cell pathway. To confirm that rRABV-mBAFF modulated the extrafollicular pathway, we used a signaling lymphocytic activation molecule (SLAM)-associated protein (SAP)-deficient mouse model. In response to antigen, SAP-deficient mice form extrafollicular B cell responses but do not generate GCs. rRABV-mBAFF induced similar anti-RABV antibody responses in SAP-deficient and wild-type mice, demonstrating that BAFF modulated immunity through the extrafollicular and not the GC B cell pathway. Collectively, strategies that manipulate pathways of B cell activation may facilitate the development of a single-dose RABV vaccine that replaces current complicated and costly RABV PEP. IMPORTANCE Effective RABV PEP is currently resource- and cost-prohibitive in regions of the world where RABV is most prevalent. In order to diminish the

Dosimetric evaluation of radionuclides for VCAM-1-targeted radionuclide therapy of early brain metastases.

PubMed

Falzone, Nadia; Ackerman, Nicole L; Rosales, Liset de la Fuente; Bernal, Mario A; Liu, Xiaoxuan; Peeters, Sarah Gja; Soto, Manuel Sarmiento; Corroyer-Dulmont, Aurélien; Bernaudin, Myriam; Grimoin, Elisa; Touzani, Omar; Sibson, Nicola R; Vallis, Katherine A

2018-01-01

Brain metastases develop frequently in patients with breast cancer, and present a pressing therapeutic challenge. Expression of vascular cell adhesion molecule 1 (VCAM-1) is upregulated on brain endothelial cells during the early stages of metastasis and provides a target for the detection and treatment of early brain metastases. The aim of this study was to use a model of early brain metastasis to evaluate the efficacy of α-emitting radionuclides, 149 Tb, 211 At, 212 Pb, 213 Bi and 225 Ac; β-emitting radionuclides, 90 Y, 161 Tb and 177 Lu; and Auger electron (AE)-emitters 67 Ga, 89 Zr, 111 In and 124 I, for targeted radionuclide therapy (TRT). Histologic sections and two photon microscopy of mouse brain parenchyma were used to inform a cylindrical vessel geometry using the Geant4 general purpose Monte Carlo (MC) toolkit with the Geant4-DNA low energy physics models. Energy deposition was evaluated as a radial function and the resulting phase spaces were superimposed on a DNA model to estimate double-strand break (DSB) yields for representative β- and α-emitters, 177 Lu and 212 Pb. Relative biological effectiveness (RBE) values were determined by only evaluating DNA damage due to physical interactions. 177 Lu produced 2.69 ± 0.08 DSB per GbpGy, without significant variation from the lumen of the vessel to a radius of 100 µm. The DSB yield of 212 Pb included two local maxima produced by the 6.1 MeV and 8.8 MeV α-emissions from decay products, 212 Bi and 212 Po, with yields of 7.64 ± 0.12 and 9.15 ± 0.24 per GbpGy, respectively. Given its higher DSB yield 212 Pb may be more effective for short range targeting of early micrometastatic lesions than 177 Lu. MC simulation of a model of early brain metastases provides invaluable insight into the potential efficacy of α-, β- and AE-emitting radionuclides for TRT. 212 Pb, which has the attributes of a theranostic radionuclide since it can be used for SPECT imaging, showed a favorable dose profile and RBE.

Targeting B Cells and Plasma Cells in Autoimmune Diseases

PubMed Central

Hofmann, Katharina; Clauder, Ann-Katrin; Manz, Rudolf Armin

2018-01-01

Success with B cell depletion using rituximab has proven the concept that B lineage cells represent a valid target for the treatment of autoimmune diseases, and has promoted the development of other B cell targeting agents. Present data confirm that B cell depletion is beneficial in various autoimmune disorders and also show that it can worsen the disease course in some patients. These findings suggest that B lineage cells not only produce pathogenic autoantibodies, but also significantly contribute to the regulation of inflammation. In this review, we will discuss the multiple pro- and anti-inflammatory roles of B lineage cells play in autoimmune diseases, in the context of recent findings using B lineage targeting therapies. PMID:29740441

Target or barrier? The cell wall of early- and later-diverging plants vs cadmium toxicity: differences in the response mechanisms

PubMed Central

Parrotta, Luigi; Guerriero, Gea; Sergeant, Kjell; Cai, Giampiero; Hausman, Jean-Francois

2015-01-01

Increasing industrialization and urbanization result in emission of pollutants in the environment including toxic heavy metals, as cadmium and lead. Among the different heavy metals contaminating the environment, cadmium raises great concern, as it is ecotoxic and as such can heavily impact ecosystems. The cell wall is the first structure of plant cells to come in contact with heavy metals. Its composition, characterized by proteins, polysaccharides and in some instances lignin and other phenolic compounds, confers the ability to bind non-covalently and/or covalently heavy metals via functional groups. A strong body of evidence in the literature has shown the role of the cell wall in heavy metal response: it sequesters heavy metals, but at the same time its synthesis and composition can be severely affected. The present review analyzes the dual property of plant cell walls, i.e., barrier and target of heavy metals, by taking Cd toxicity as example. Following a summary of the known physiological and biochemical responses of plants to Cd, the review compares the wall-related mechanisms in early- and later-diverging land plants, by considering the diversity in cell wall composition. By doing so, common as well as unique response mechanisms to metal/cadmium toxicity are identified among plant phyla and discussed. After discussing the role of hyperaccumulators’ cell walls as a particular case, the review concludes by considering important aspects for plant engineering. PMID:25814996

Amiodarone affects Ebola virus binding and entry into target cells.

PubMed

Salata, Cristiano; Munegato, Denis; Martelli, Francesco; Parolin, Cristina; Calistri, Arianna; Baritussio, Aldo; Palù, Giorgio

2018-03-02

Ebola Virus Disease is one of the most lethal transmissible infections characterized by a high fatality rate. Several research studies have aimed to identify effective antiviral agents. Amiodarone, a drug used for the treatment of arrhythmias, has been shown to inhibit filovirus infection in vitro by acting at the early step of the viral replication cycle. Here we demonstrate that amiodarone reduces virus binding to target cells and slows down the progression of the viral particles along the endocytic pathway. Overall our data support the notion that amiodarone interferes with Ebola virus infection by affecting cellular pathways/targets involved in the viral entry process.

Pharmacologic suppression of target cell recognition by engineered T cells expressing chimeric T-cell receptors.

PubMed

Alvarez-Vallina, L; Yañez, R; Blanco, B; Gil, M; Russell, S J

2000-04-01

Adoptive therapy with autologous T cells expressing chimeric T-cell receptors (chTCRs) is of potential interest for the treatment of malignancy. To limit possible T-cell-mediated damage to normal tissues that weakly express the targeted tumor antigen (Ag), we have tested a strategy for the suppression of target cell recognition by engineered T cells. Jurkat T cells were transduced with an anti-hapten chTCR tinder the control of a tetracycline-suppressible promoter and were shown to respond to Ag-positive (hapten-coated) but not to Ag-negative target cells. The engineered T cells were then reacted with hapten-coated target cells at different effector to target cell ratios before and after exposure to tetracycline. When the engineered T cells were treated with tetracycline, expression of the chTCR was greatly decreased and recognition of the hapten-coated target cells was completely suppressed. Tetracycline-mediated suppression of target cell recognition by engineered T cells may be a useful strategy to limit the toxicity of the approach to cancer gene therapy.

Immune Cell Targets of Infection at the Tick-Skin Interface during Powassan Virus Transmission.

PubMed

Hermance, Meghan E; Santos, Rodrigo I; Kelly, Brent C; Valbuena, Gustavo; Thangamani, Saravanan

2016-01-01

Powassan virus (POWV) is a tick-borne flavivirus that can result in a severe neuroinvasive disease with 50% of survivors displaying long-term neurological sequelae. Human POWV cases have been documented in Canada, the United States, and Russia. Although the number of reported POWV human cases has increased in the past fifteen years, POWV remains one of the less studied human pathogenic flaviviruses. Ixodes ticks are the vectors for POWV, and the virus is transmitted to a host's skin very early during the tick feeding process. Central to the successful transmission of a tick-borne pathogen are complex interactions between the host immune response and early tick-mediated immunomodulation, all of which initially occur at the skin interface. In our prior work, we examined the cutaneous immune gene expression during the early stages of POWV-infected Ixodes scapularis feeding. The present study serves to further investigate the skin interface by identifying early cell targets of infection at the POWV-infected tick feeding site. An in vivo infection model consisting of POWV-infected ticks feeding on mice for short durations was used in this study. Skin biopsies from the tick feeding sites were harvested at various early time points, enabling us to examine the skin histopathology and detect POWV viral antigen in immune cells present at the tick feeding site. The histopathology from the present study demonstrates that neutrophil and mononuclear cell infiltrates are recruited earlier to the feeding site of a POWV-infected tick versus an uninfected tick. This is the first report demonstrating that macrophages and fibroblasts contain POWV antigens, which suggests that they are early cellular targets of infection at the tick feeding site. These data provide key insights towards defining the complex interactions between the host immune response and early tick-mediated immunomodulation.

Immune Cell Targets of Infection at the Tick-Skin Interface during Powassan Virus Transmission

PubMed Central

Hermance, Meghan E.; Santos, Rodrigo I.; Kelly, Brent C.; Valbuena, Gustavo; Thangamani, Saravanan

2016-01-01

Powassan virus (POWV) is a tick-borne flavivirus that can result in a severe neuroinvasive disease with 50% of survivors displaying long-term neurological sequelae. Human POWV cases have been documented in Canada, the United States, and Russia. Although the number of reported POWV human cases has increased in the past fifteen years, POWV remains one of the less studied human pathogenic flaviviruses. Ixodes ticks are the vectors for POWV, and the virus is transmitted to a host’s skin very early during the tick feeding process. Central to the successful transmission of a tick-borne pathogen are complex interactions between the host immune response and early tick-mediated immunomodulation, all of which initially occur at the skin interface. In our prior work, we examined the cutaneous immune gene expression during the early stages of POWV-infected Ixodes scapularis feeding. The present study serves to further investigate the skin interface by identifying early cell targets of infection at the POWV-infected tick feeding site. An in vivo infection model consisting of POWV-infected ticks feeding on mice for short durations was used in this study. Skin biopsies from the tick feeding sites were harvested at various early time points, enabling us to examine the skin histopathology and detect POWV viral antigen in immune cells present at the tick feeding site. The histopathology from the present study demonstrates that neutrophil and mononuclear cell infiltrates are recruited earlier to the feeding site of a POWV-infected tick versus an uninfected tick. This is the first report demonstrating that macrophages and fibroblasts contain POWV antigens, which suggests that they are early cellular targets of infection at the tick feeding site. These data provide key insights towards defining the complex interactions between the host immune response and early tick-mediated immunomodulation. PMID:27203436

A targeted neuroglial reporter line generated by homologous recombination in human embryonic stem cells.

PubMed

Xue, Haipeng; Wu, Sen; Papadeas, Sophia T; Spusta, Steve; Swistowska, Anna Maria; MacArthur, Chad C; Mattson, Mark P; Maragakis, Nicholas J; Capecchi, Mario R; Rao, Mahendra S; Zeng, Xianmin; Liu, Ying

2009-08-01

In this study, we targeted Olig2, a basic helix-loop-helix transcription factor that plays an important role in motoneuron and oligodendrocyte development, in human embryonic stem cell (hESC) line BG01 by homologous recombination. One allele of Olig2 locus was replaced by a green fluorescent protein (GFP) cassette with a targeting efficiency of 5.7%. Targeted clone R-Olig2 (like the other clones) retained pluripotency, typical hESC morphology, and a normal parental karyotype 46,XY. Most importantly, GFP expression recapitulated endogenous Olig2 expression when R-Olig2 was induced by sonic hedgehog and retinoic acid, and GFP-positive cells could be purified by fluorescence-activated cell sorting. Consistent with previous reports on rodents, early GFP-expressing cells appeared biased to a neuronal fate, whereas late GFP-expressing cells appeared biased to an oligodendrocytic fate. This was corroborated by myoblast coculture, transplantation into the rat spinal cords, and whole genome expression profiling. The present work reports an hESC reporter line generated by homologous recombination targeting a neural lineage-specific gene, which can be differentiated and sorted to obtain pure neural progenitor populations.

Construction of ultrasonic nanobubbles carrying CAIX polypeptides to target carcinoma cells derived from various organs.

PubMed

Zhu, Lianhua; Guo, Yanli; Wang, Luofu; Fan, Xiaozhou; Xiong, Xingyu; Fang, Kejing; Xu, Dan

2017-09-29

Ultrasound molecular imaging is a novel diagnostic approach for tumors, whose key link is the construction of targeted ultrasound contrast agents. However, available targeted ultrasound contrast agents for molecular imaging of tumors are only achieving imaging in blood pool or one type tumor. No targeted ultrasound contrast agents have realized targeted ultrasound molecular imaging of tumor parenchymal cells in a variety of solid tumors so far. Carbonic anhydrase IX (CAIX) is highly expressed on cell membranes of various malignant solid tumors, so it's a good target for ultrasound molecular imaging. Here, targeted nanobubbles carrying CAIX polypeptides for targeted binding to a variety of malignant tumors were constructed, and targeted binding ability and ultrasound imaging effect in different types of tumors were evaluated. The mean diameter of lipid targeted nanobubbles was (503.7 ± 78.47) nm, and the polypeptides evenly distributed on the surfaces of targeted nanobubbles, which possessed the advantages of homogenous particle size, high stability, and good safety. Targeted nanobubbles could gather around CAIX-positive cells (786-O and Hela cells), while they cannot gather around CAIX-negative cells (BxPC-3 cells) in vitro, and the affinity of targeted nanobubbles to CAIX-positive cells were significantly higher than that to CAIX-negative cells (P < 0.05). Peak intensity and duration time of targeted nanobubbles and blank nanobubbles were different in CAIX-positive transplanted tumor tissues in vivo (P < 0.05). Moreover, targeted nanobubbles in CAIX-positive transplanted tumor tissues produced higher peak intensity and longer duration time than those in CAIX-negative transplanted tumor tissues (P < 0.05). Finally, immunofluorescence not only confirmed targeted nanobubbles could pass through blood vessels to enter in tumor tissue spaces, but also clarified imaging differences of targeted nanobubbles in different types of transplanted tumor tissues

Vitamin C, a Multi-Tasking Molecule, Finds a Molecular Target in Killing Cancer Cells.

PubMed

Li, Robert

2016-03-01

Early work in the 1970s by Linus Pauling, a twice-honored Nobel laureate, led to his proposal of using high-dose vitamin C to treat cancer patients. Over the past several decades, a number of studies in animal models as well as several small-scale clinical studies have provided substantial support of Linus Pauling's early proposal. Production of reactive oxygen species (ROS) via oxidation of vitamin C appears to be a major underlying event, leading to the selective killing of cancer cells. However, it remains unclear how vitamin C selectively kills cancer cells while sparing normal cells and what the molecular targets of high-dose vitamin C are. In a recent article published in Science (2015 December 11; 350(6266):1391-6. doi: 10.1126/science.aaa5004), Yun et al. reported that vitamin C selectively kills KRAS and BRAF mutant colorectal cancer cells by targeting glyceraldehyde 3-phosphate dehydrogenase (GAPDH) through an ROS-dependent mechanism. This work by Yun et al. along with other findings advances our current understanding of the molecular basis of high-dose vitamin C-mediated cancer cell killing, which will likely give an impetus to the continued research efforts aiming to further decipher the novel biochemistry of vitamin C and its unique role in cancer therapy.

Human immune cell targeting of protein nanoparticles - caveospheres

NASA Astrophysics Data System (ADS)

Glass, Joshua J.; Yuen, Daniel; Rae, James; Johnston, Angus P. R.; Parton, Robert G.; Kent, Stephen J.; de Rose, Robert

2016-04-01

Nanotechnology has the power to transform vaccine and drug delivery through protection of payloads from both metabolism and off-target effects, while facilitating specific delivery of cargo to immune cells. However, evaluation of immune cell nanoparticle targeting is conventionally restricted to monocultured cell line models. We generated human caveolin-1 nanoparticles, termed caveospheres, which were efficiently functionalized with monoclonal antibodies. Using this platform, we investigated CD4+ T cell and CD20+ B cell targeting within physiological mixtures of primary human blood immune cells using flow cytometry, imaging flow cytometry and confocal microscopy. Antibody-functionalization enhanced caveosphere binding to targeted immune cells (6.6 to 43.9-fold) within mixed populations and in the presence of protein-containing fluids. Moreover, targeting caveospheres to CCR5 enabled caveosphere internalization by non-phagocytic CD4+ T cells--an important therapeutic target for HIV treatment. This efficient and flexible system of immune cell-targeted caveosphere nanoparticles holds promise for the development of advanced immunotherapeutics and vaccines.

The expression of human natural killer cell receptors in early life.

PubMed

Sundström, Y; Nilsson, C; Lilja, G; Kärre, K; Troye-Blomberg, M; Berg, L

2007-01-01

Natural killer (NK) cells play an important role in tumour immunosurveillance and the early defence against viral infections. Recognition of altered cells (i.e. infected- or tumour-cells) is achieved through a multiple receptor recognition strategy which gives the NK cells inhibitory or activating signals depending on the ligands present on the target cell. NK cells originate from the bone marrow where they develop and proliferate. However, further maturation processes and homeostasis of NK cells in peripheral blood are not well understood. To determine the proportions of cells and the expression of NK cell receptors, mononuclear cells from children at three time points during early childhood were compared, i.e. cord blood (CB), 2 and 5 years of age. The proportion of NK cells was high in CB, but the interferon-gamma (IFN-gamma) production low compared to later in life. In contrast, the proportion of T cells was low in CB. This may indicate a deviation of the regulatory function of NK cells in CB compared to later in life, implying an importance of innate immunity in early life before the adaptive immune system matures. Additionally, we found that the proportion of LIR-1(+) NK cells increased with increasing age while CD94(+)NKG2C(-) (NKG2A(+)) NK cells and the level of expression of NKG2D, NKp30 and NKp46 decreased with age. These age related changes in NK cell populations defined by the expression of activating and inhibitory receptors may be the result of pathogen exposure and/or a continuation of the maturation process that begins in the bone marrow.

Targeting Vaccine-Induced Extrafollicular Pathway of B Cell Differentiation Improves Rabies Postexposure Prophylaxis.

PubMed

Haley, Shannon L; Tzvetkov, Evgeni P; Meuwissen, Samantha; Plummer, Joseph R; McGettigan, James P

2017-04-15

Vaccine-induced B cells differentiate along two pathways. The follicular pathway gives rise to germinal centers (GCs) that can take weeks to fully develop. The extrafollicular pathway gives rise to short-lived plasma cells (PCs) that can rapidly secrete protective antibodies within days of vaccination. Rabies virus (RABV) postexposure prophylaxis (PEP) requires rapid vaccine-induced humoral immunity for protection. Therefore, we hypothesized that targeting extrafollicular B cell responses for activation would improve the speed and magnitude of RABV PEP. To test this hypothesis, we constructed, recovered, and characterized a recombinant RABV-based vaccine expressing murine B cell activating factor (BAFF) (rRABV-mBAFF). BAFF is an ideal molecule to improve early pathways of B cell activation, as it links innate and adaptive immunity, promoting potent B cell responses. Indeed, rRABV-mBAFF induced a faster, higher antibody response in mice and enhanced survivorship in PEP settings compared to rRABV. Interestingly, rRABV-mBAFF and rRABV induced equivalent numbers of GC B cells, suggesting that rRABV-mBAFF augmented the extrafollicular B cell pathway. To confirm that rRABV-mBAFF modulated the extrafollicular pathway, we used a signaling lymphocytic activation molecule (SLAM)-associated protein (SAP)-deficient mouse model. In response to antigen, SAP-deficient mice form extrafollicular B cell responses but do not generate GCs. rRABV-mBAFF induced similar anti-RABV antibody responses in SAP-deficient and wild-type mice, demonstrating that BAFF modulated immunity through the extrafollicular and not the GC B cell pathway. Collectively, strategies that manipulate pathways of B cell activation may facilitate the development of a single-dose RABV vaccine that replaces current complicated and costly RABV PEP. IMPORTANCE Effective RABV PEP is currently resource- and cost-prohibitive in regions of the world where RABV is most prevalent. In order to diminish the requirements for

Smart Plasmonic Glucose Nanosensors as Generic Theranostic Agents for Targeting-Free Cancer Cell Screening and Killing.

PubMed

Chen, Limei; Li, Haijuan; He, Haili; Wu, Haoxi; Jin, Yongdong

2015-07-07

Fast and accurate identification of cancer cells from healthy normal cells in a simple, generic way is very crucial for early cancer detection and treatment. Although functional nanoparticles, like fluorescent quantum dots and plasmonic Au nanoparticles (NPs), have been successfully applied for cancer cell imaging and photothermal therapy, they suffer from the main drawback of needing time-consuming targeting preparation for specific cancer cell detection and selective ablation. The lack of a generic and effective method therefore limits their potential high-throughput cancer cell preliminary screening and theranostic applications. We report herein a generic in vitro method for fast, targeting-free (avoiding time-consuming preparations of targeting moiety for specific cancer cells) visual screening and selective killing of cancer cells from normal cells, by using glucose-responsive/-sensitive glucose oxidase-modified Ag/Au nanoshells (Ag/Au-GOx NSs) as a smart plasmonic theranostic agent. The method is generic to some extent since it is based on the distinct localized surface plasmon resonance (LSPR) responses (and colors) of the smart nanoprobe with cancer cells (typically have a higher glucose uptake level) and normal cells.

Prediction of intracellular exposure bridges the gap between target- and cell-based drug discovery

PubMed Central

Gordon, Laurie J.; Wayne, Gareth J.; Almqvist, Helena; Axelsson, Hanna; Seashore-Ludlow, Brinton; Treyer, Andrea; Lundbäck, Thomas; West, Andy; Hann, Michael M.; Artursson, Per

2017-01-01

Inadequate target exposure is a major cause of high attrition in drug discovery. Here, we show that a label-free method for quantifying the intracellular bioavailability (Fic) of drug molecules predicts drug access to intracellular targets and hence, pharmacological effect. We determined Fic in multiple cellular assays and cell types representing different targets from a number of therapeutic areas, including cancer, inflammation, and dementia. Both cytosolic targets and targets localized in subcellular compartments were investigated. Fic gives insights on membrane-permeable compounds in terms of cellular potency and intracellular target engagement, compared with biochemical potency measurements alone. Knowledge of the amount of drug that is locally available to bind intracellular targets provides a powerful tool for compound selection in early drug discovery. PMID:28701380

Membrane nanotubes facilitate long-distance interactions between natural killer cells and target cells

PubMed Central

Chauveau, Anne; Aucher, Anne; Eissmann, Philipp; Vivier, Eric; Davis, Daniel M.

2010-01-01

Membrane nanotubes are membranous tethers that physically link cell bodies over long distances. Here, we present evidence that nanotubes allow human natural killer (NK) cells to interact functionally with target cells over long distances. Nanotubes were formed when NK cells contacted target cells and moved apart. The frequency of nanotube formation was dependent on the number of receptor/ligand interactions and increased on NK cell activation. Most importantly, NK cell nanotubes contained a submicron scale junction where proteins accumulated, including DAP10, the signaling adaptor that associates with the activating receptor NKG2D, and MHC class I chain-related protein A (MICA), a cognate ligand for NKG2D, as occurs at close intercellular synapses between NK cells and target cells. Quantitative live-cell fluorescence imaging suggested that MICA accumulated at small nanotube synapses in sufficient numbers to trigger cell activation. In addition, tyrosine-phosphorylated proteins and Vav-1 accumulated at such junctions. Functionally, nanotubes could aid the lysis of distant target cells either directly or by moving target cells along the nanotube path into close contact for lysis via a conventional immune synapse. Target cells moving along the nanotube path were commonly polarized such that their uropods faced the direction of movement. This is the opposite polarization than for normal cell migration, implying that nanotubes can specifically drive target cell movement. Finally, target cells that remained connected to an NK cell by a nanotube were frequently lysed, whereas removing the nanotube using a micromanipulator reduced lysis of these target cells. PMID:20212116

Src Kinase: A Novel Target of Early-Stage ER-Negative Breast Cancer

DTIC Science & Technology

2012-03-01

patients with early stage ErbB2-overexpressing biopsies and ER- atypia . 13 REFERENCES: 1. Jordan VC. Tamoxifen for breast cancer prevention. Proc Soc...Summary01-03-2012 Src Kinase: A Novel Target of Early-Stage ER-Negative Breast Cancer Shalini Jain University of Texas M.D. Anderson Cancer Center Houston...SUBTITLE “Src Kinase: A Novel Target of Early-Stage ER-Negative Breast Cancer” 5a. CONTRACT NUMBER W81XWH-11-1-0004 5b. GRANT NUMBER

Cell-specific targeting by heterobivalent ligands.

PubMed

Josan, Jatinder S; Handl, Heather L; Sankaranarayanan, Rajesh; Xu, Liping; Lynch, Ronald M; Vagner, Josef; Mash, Eugene A; Hruby, Victor J; Gillies, Robert J

2011-07-20

Current cancer therapies exploit either differential metabolism or targeting to specific individual gene products that are overexpressed in aberrant cells. The work described herein proposes an alternative approach--to specifically target combinations of cell-surface receptors using heteromultivalent ligands ("receptor combination approach"). As a proof-of-concept that functionally unrelated receptors can be noncovalently cross-linked with high avidity and specificity, a series of heterobivalent ligands (htBVLs) were constructed from analogues of the melanocortin peptide ligand ([Nle(4), dPhe(7)]-α-MSH) and the cholecystokinin peptide ligand (CCK-8). Binding of these ligands to cells expressing the human Melanocortin-4 receptor and the Cholecystokinin-2 receptor was analyzed. The MSH(7) and CCK(6) were tethered with linkers of varying rigidity and length, constructed from natural and/or synthetic building blocks. Modeling data suggest that a linker length of 20-50 Å is needed to simultaneously bind these two different G-protein coupled receptors (GPCRs). These ligands exhibited up to 24-fold enhancement in binding affinity to cells that expressed both (bivalent binding), compared to cells with only one (monovalent binding) of the cognate receptors. The htBVLs had up to 50-fold higher affinity than that of a monomeric CCK ligand, i.e., Ac-CCK(6)-NH(2). Cell-surface targeting of these two cell types with labeled heteromultivalent ligand demonstrated high avidity and specificity, thereby validating the receptor combination approach. This ability to noncovalently cross-link heterologous receptors and target individual cells using a receptor combination approach opens up new possibilities for specific cell targeting in vivo for therapy or imaging.

Cell-Specific Targeting by Heterobivalent Ligands

PubMed Central

Josan, Jatinder S.; Handl, Heather L.; Sankaranarayanan, Rajesh; Xu, Liping; Lynch, Ronald M.; Vagner, Josef; Mash, Eugene A.; Hruby, Victor J.; Gillies, Robert J.

2012-01-01

Current cancer therapies exploit either differential metabolism or targeting to specific individual gene products that are overexpressed in aberrant cells. The work described herein proposes an alternative approach—to specifically target combinations of cell-surface receptors using heteromultivalent ligands (“receptor combination approach”). As a proof-of-concept that functionally unrelated receptors can be noncovalently cross-linked with high avidity and specificity, a series of heterobivalent ligands (htBVLs) were constructed from analogues of the melanocortin peptide ligand ([Nle4, DPhe7]-α-MSH) and the cholecystokinin peptide ligand (CCK-8). Binding of these ligands to cells expressing the human Melanocortin-4 receptor and the Cholecystokinin-2 receptor was analyzed. The MSH(7) and CCK(6) were tethered with linkers of varying rigidity and length, constructed from natural and/or synthetic building blocks. Modeling data suggest that a linker length of 20–50 Å is needed to simultaneously bind these two different G-protein coupled receptors (GPCRs). These ligands exhibited up to 24-fold enhancement in binding affinity to cells that expressed both (bivalent binding), compared to cells with only one (monovalent binding) of the cognate receptors. The htBVLs had up to 50-fold higher affinity than that of a monomeric CCK ligand, i.e., Ac-CCK(6)-NH2. Cell-surface targeting of these two cell types with labeled heteromultivalent ligand demonstrated high avidity and specificity, thereby validating the receptor combination approach. This ability to noncovalently cross-link heterologous receptors and target individual cells using a receptor combination approach opens up new possibilities for specific cell targeting in vivo for therapy or imaging. PMID:21639139

Enterocolitis induced by autoimmune targeting of enteric glial cells: A possible mechanism in Crohn's disease?

NASA Astrophysics Data System (ADS)

Cornet, Anne; Savidge, Tor C.; Cabarrocas, Julie; Deng, Wen-Lin; Colombel, Jean-Frederic; Lassmann, Hans; Desreumaux, Pierre; Liblau, Roland S.

2001-11-01

Early pathological manifestations of Crohn's disease (CD) include vascular disruption, T cell infiltration of nerve plexi, neuronal degeneration, and induction of T helper 1 cytokine responses. This study demonstrates that disruption of the enteric glial cell network in CD patients represents another early pathological feature that may be modeled after CD8+ T cell-mediated autoimmune targeting of enteric glia in double transgenic mice. Mice expressing a viral neoself antigen in astrocytes and enteric glia were crossed with specific T cell receptor transgenic mice, resulting in apoptotic depletion of enteric glia to levels comparable in CD patients. Intestinal and mesenteric T cell infiltration, vasculitis, T helper 1 cytokine production, and fulminant bowel inflammation were characteristic hallmarks of disease progression. Immune-mediated damage to enteric glia therefore may participate in the initiation and/or the progression of human inflammatory bowel disease.

Tracking targeted bimodal nanovaccines: immune responses and routing in cells, tissue, and whole organism.

PubMed

Cruz, Luis J; Tacken, Paul J; Zeelenberg, Ingrid S; Srinivas, Mangala; Bonetto, Fernando; Weigelin, Bettina; Eich, Christina; de Vries, I Jolanda; Figdor, Carl G

2014-12-01

Dendritic cells (DCs) are the most potent antigen-presenting cells (APCs), involved in the induction of immunity and currently exploited for antitumor immunotherapies. An optimized noninvasive imaging modality capable of determining and quantifying DC-targeted nanoparticle (NP) trajectories could provide valuable information regarding therapeutic vaccine outcome. Here, targeted poly(d,l-lactide-co-glycolide) nanoparticles (PLGA NPs) recognizing DC receptors were equipped with superparamagnetic iron oxide particles (SPIO) or gold nanoparticles with fluorescently labeled antigen. The fluorescent label allowed for rapid analysis and quantification of DC-specific uptake of targeted PLGA NPs in comparison to uptake by other cells. Transmission electron microscopy (TEM) showed that a fraction of the encapsulated antigen reached the lysosomal compartment of DCs, where SPIO and gold were already partially released. However, part of the PLGA NPs localized within the cytoplasm, as confirmed by confocal microscopy. DCs targeted with NPs carrying SPIO or fluorescent antigen were detected within lymph nodes as early as 1 h after injection by magnetic resonance imaging (MRI). Despite the fact that targeting did not markedly affect PLGA NP biodistribution on organism and tissue level, it increased delivery of NPs to DCs residing in peripheral lymph nodes and resulted in enhanced T cell proliferation. In conclusion, two imaging agents within a single carrier allows tracking of targeted PLGA NPs at the subcellular, cellular, and organismal levels, thereby facilitating the rational design of in vivo targeted vaccination strategies.

Photochemical Targeting Of Phagocytic Trabecular Meshwork Cells Using Chlorin E6 Coupled Microspheres

NASA Astrophysics Data System (ADS)

Latina, M. A.; Kobsa, P. H.; Rakestraw, S. L.; Crean, E. A.; Hasan, T.; Yarmush, M. L.

1989-03-01

We have investigated a novel and efficient delivery system utilizing photosensitizer-coupled-latex microspheres to photochemically target and kill phagocytic trabecular meshwork (TM) cells. TM cells are the most actively phagocytic cells within the anterior chamber of the eye and are located within an optically accessible discrete band. This delivery system, along with the property of cell photocytosis, will achieve double selectivity by combining preferential localization of the photosensitizer to the target cells with spatial localization of illumination on the target cells. All experiments were performed with preconfluent bovine TM cells, 3rd to 4th passage, plated in 15 mm wells. Chlorin e6 monoethylene diamine monoamide was conjugated to the surface of 1.0 Am MX Duke Scientific fluorescent latex microspheres. Spectroscopic analysis revealed an average of 1.3 x 10 -17 moles of chlorin e6 per microsphere. TM cells were incubated for 18 hours with 5 x 10 7 microspheres/ml in MEM with 10% FCS, washed with MEM, and irradiated through fresh media using an argon-pumped dye laser emitting .2 W at 660 nm. A dose-survival study indicated that energy doses of 10 J/cm2 or greater resulted in greater than 95% cell death as determined by ethidium bromide exclusion. Cell death could be demonstrated as early as 4 hours post-irradiation. TM cells incubated with a solution of chlorin e6 at a concentration equal to that conjugated to the microspheres showed no cell death. Unirradiated controls also showed no cell death.

Targeting Cell Polarity Machinery to Exhaust Breast Cancer Stem Cells

DTIC Science & Technology

2016-10-01

AWARD NUMBER: W81XWH-15-1-0644 TITLE: Targeting Cell Polarity Machinery to Exhaust Breast Cancer Stem Cells PRINCIPAL INVESTIGATOR: Chun-Ju...U.S. Army Medical Research and Materiel Command Fort Detrick, Maryland 21702-5012 DISTRIBUTION STATEMENT: Approved for Public Release...Targeting Cell Polarity Machinery to Exhaust Breast Cancer Stem Cells 5a. CONTRACT NUMBER 5b. GRANT NUMBER W81XWH-15-1-0644 5c. PROGRAM ELEMENT

Targeted deletion of p97 (VCP/CDC48) in mouse results in early embryonic lethality.

PubMed

Müller, J M M; Deinhardt, K; Rosewell, I; Warren, G; Shima, D T

2007-03-09

The highly conserved AAA ATPase p97 (VCP/CDC48) has well-established roles in cell cycle progression, proteasome degradation and membrane dynamics. Gene disruption in Saccromyces cerevisiae, Drosophila melanogaster and Trypanosoma brucei demonstrated that p97 is essential in unicellular and multicellular organisms. To explore the requirement for p97 in mammalian cell function and embryogenesis, we disrupted the p97 locus by gene targeting. Heterozygous p97+/- mice were indistinguishable from their wild-type littermates, whereas homozygous mutants did not survive to birth and died at a peri-implantation stage. These results show that p97 is an essential gene for early mouse development.

A hypothesis of target cell formation in sickle cell disease.

PubMed

Wong, P

2016-08-01

A fraction of erythrocytes appear as target cells in stained blood smears in sickle cell disease, due to a inheritance of the hemoglobin variant Hb S, polymerizing upon deoxygenation. These cells appear in a three dimension as thin cups. A process of their formation in this disease is proposed based on a band 3-based mechanism of the erythrocyte shape control, able to explain the erythrocyte echinocytosis by glucose depletion. It indicates that their formation is due to a stomatocytogenic slow outward transport of the dibasic form of endogenous Pi with an H(+) by band 3, promoted by the decrease of the Donnan ratio, which decreases cell pH and volume, attributed by a decrease of cell KCl concentration by the higher efflux of K(+)Cl(-) cotransport and Ca(2+) activation of the Gardos channel. Its implications are briefly discussed with respect to target cells per se, target cell formation in other hemoglobinopathies, acquired and inherited disorders of the lipid metabolism and dehydrated hereditary stomatocytosis as well as a stomatocyte presence in a double heterozygote of Hb S and Hb C and of an involvement of the process of target cell formation in acanthocytosis in acquired and inherited disorders. Copyright © 2016 Elsevier Ltd. All rights reserved.

Evolving phage vectors for cell targeted gene delivery.

PubMed

Larocca, David; Burg, Michael A; Jensen-Pergakes, Kristen; Ravey, Edward Prenn; Gonzalez, Ana Maria; Baird, Andrew

2002-03-01

We adapted filamentous phage vectors for targeted gene delivery to mammalian cells by inserting a mammalian reporter gene expression cassette (GFP) into the vector backbone and fusing the pIII coat protein to a cell targeting ligand (i.e. FGF2, EGF). Like transfection with animal viral vectors, targeted phage gene delivery is concentration, time, and ligand dependent. Importantly, targeted phage particles are specific for the appropriate target cell surface receptor. Phage have distinct advantages over existing gene therapy vectors because they are simple, economical to produce at high titer, have no intrinsic tropism for mammalian cells, and are relatively simple to genetically modify and evolve. Initially transduction by targeted phage particles was low resulting in foreign gene expression in 1-2% of transfected cells. We increased transduction efficiency by modifying both the transfection protocol and vector design. For example, we stabilized the display of the targeting ligand to create multivalent phagemid-based vectors with transduction efficiencies of up to 45% in certain cell lines when combined with genotoxic treatment. Taken together, these studies establish that the efficiency of phage-mediated gene transfer can be significantly improved through genetic modification. We are currently evolving phage vectors with enhanced cell targeting, increased stability, reduced immunogenicity and other properties suitable for gene therapy.

Targeted antibody-mediated depletion of murine CD19 CAR T cells permanently reverses B cell aplasia

PubMed Central

Paszkiewicz, Paulina J.; Fräßle, Simon P.; Srivastava, Shivani; Sommermeyer, Daniel; Hudecek, Michael; Sadelain, Michel; Liu, Lingfeng; Jensen, Michael C.; Riddell, Stanley R.; Busch, Dirk H.

2016-01-01

The adoptive transfer of T cells that have been genetically modified to express a CD19-specific chimeric antigen receptor (CAR) is effective for treating human B cell malignancies. However, the persistence of functional CD19 CAR T cells causes sustained depletion of endogenous CD19+ B cells and hypogammaglobulinemia. Thus, there is a need for a mechanism to ablate transferred T cells after tumor eradication is complete to allow recovery of normal B cells. Previously, we developed a truncated version of the epidermal growth factor receptor (EGFRt) that is coexpressed with the CAR on the T cell surface. Here, we show that targeting EGFRt with the IgG1 monoclonal antibody cetuximab eliminates CD19 CAR T cells both early and late after adoptive transfer in mice, resulting in complete and permanent recovery of normal functional B cells, without tumor relapse. EGFRt can be incorporated into many clinical applications to regulate the survival of gene-engineered cells. These results support the concept that EGFRt represents a promising approach to improve safety of cell-based therapies. PMID:27760047

Targeted antibody-mediated depletion of murine CD19 CAR T cells permanently reverses B cell aplasia.

PubMed

Paszkiewicz, Paulina J; Fräßle, Simon P; Srivastava, Shivani; Sommermeyer, Daniel; Hudecek, Michael; Drexler, Ingo; Sadelain, Michel; Liu, Lingfeng; Jensen, Michael C; Riddell, Stanley R; Busch, Dirk H

2016-11-01

The adoptive transfer of T cells that have been genetically modified to express a CD19-specific chimeric antigen receptor (CAR) is effective for treating human B cell malignancies. However, the persistence of functional CD19 CAR T cells causes sustained depletion of endogenous CD19+ B cells and hypogammaglobulinemia. Thus, there is a need for a mechanism to ablate transferred T cells after tumor eradication is complete to allow recovery of normal B cells. Previously, we developed a truncated version of the epidermal growth factor receptor (EGFRt) that is coexpressed with the CAR on the T cell surface. Here, we show that targeting EGFRt with the IgG1 monoclonal antibody cetuximab eliminates CD19 CAR T cells both early and late after adoptive transfer in mice, resulting in complete and permanent recovery of normal functional B cells, without tumor relapse. EGFRt can be incorporated into many clinical applications to regulate the survival of gene-engineered cells. These results support the concept that EGFRt represents a promising approach to improve safety of cell-based therapies.

Highly sensitive and specific colorimetric detection of cancer cells via dual-aptamer target binding strategy.

PubMed

Wang, Kun; Fan, Daoqing; Liu, Yaqing; Wang, Erkang

2015-11-15

Simple, rapid, sensitive and specific detection of cancer cells is of great importance for early and accurate cancer diagnostics and therapy. By coupling nanotechnology and dual-aptamer target binding strategies, we developed a colorimetric assay for visually detecting cancer cells with high sensitivity and specificity. The nanotechnology including high catalytic activity of PtAuNP and magnetic separation & concentration plays a vital role on the signal amplification and improvement of detection sensitivity. The color change caused by small amount of target cancer cells (10 cells/mL) can be clearly distinguished by naked eyes. The dual-aptamer target binding strategy guarantees the detection specificity that large amount of non-cancer cells and different cancer cells (10(4) cells/mL) cannot cause obvious color change. A detection limit as low as 10 cells/mL with detection linear range from 10 to 10(5) cells/mL was reached according to the experimental detections in phosphate buffer solution as well as serum sample. The developed enzyme-free and cost effective colorimetric assay is simple and no need of instrument while still provides excellent sensitivity, specificity and repeatability, having potential application on point-of-care cancer diagnosis. Copyright © 2015 Elsevier B.V. All rights reserved.

Statistical Modeling of Single Target Cell Encapsulation

PubMed Central

Moon, SangJun; Ceyhan, Elvan; Gurkan, Umut Atakan; Demirci, Utkan

2011-01-01

High throughput drop-on-demand systems for separation and encapsulation of individual target cells from heterogeneous mixtures of multiple cell types is an emerging method in biotechnology that has broad applications in tissue engineering and regenerative medicine, genomics, and cryobiology. However, cell encapsulation in droplets is a random process that is hard to control. Statistical models can provide an understanding of the underlying processes and estimation of the relevant parameters, and enable reliable and repeatable control over the encapsulation of cells in droplets during the isolation process with high confidence level. We have modeled and experimentally verified a microdroplet-based cell encapsulation process for various combinations of cell loading and target cell concentrations. Here, we explain theoretically and validate experimentally a model to isolate and pattern single target cells from heterogeneous mixtures without using complex peripheral systems. PMID:21814548

Lower early postnatal oxygen saturation target and risk of ductus arteriosus closure failure.

PubMed

Inomata, Kei; Taniguchi, Shinji; Yonemoto, Hiroki; Inoue, Takeshi; Kawase, Akihiko; Kondo, Yuichi

2016-11-01

Early postnatal hyperoxia is a major risk factor for retinopathy of prematurity (ROP) in extremely premature infants. To reduce the occurrence of ROP, we adopted a lower early postnatal oxygen saturation (SpO 2 ) target range (85-92%) from April 2011. Lower SpO 2 target range, however, may lead to hypoxemia and an increase in the risk of ductus arteriosus (DA) closure failure. The aim of this study was therefore to determine whether a lower SpO 2 target range, during the early postnatal stage, increases the risk of DA closure failure. Infants born at <28 weeks' gestation were enrolled in this study. Oxygen saturation target range during the first postnatal 72 h was 84-100% in study period 1 and 85-92% in period 2. Eighty-two infants were included in period 1, and 61 were included in period 2. The lower oxygen saturation target range increased the occurrence of hypoxemia during the first postnatal 72 h. Prevalence of DA closure failure in period 2 (21%) was significantly higher than that in period 1 (1%). On multivariate logistic regression analysis, the lower oxygen saturation target range was an independent risk factor for DA closure failure. Lower early postnatal oxygen saturation target range increases the risk of DA closure failure. © 2016 Japan Pediatric Society.

Cysts mark the early stage of metastatic tumor development in non-small cell lung cancer

PubMed Central

Thakur, Chitra; Rapp, Ulf R.; Rudel, Thomas

2018-01-01

Identifying metastatic tumor growth at an early stage has been one of the biggest challenges in the treatment of lung cancer. By genetic lineage tracing approach in a conditional model of Non-Small Cell Lung Cancer (NSCLC) in mice, we demonstrate that cystic lesions represent an early stage of metastatic invasion. We generated a mouse model for NSCLC which incorporated a heritable DsRed fluorescent tag driven by the ubiquitous CAG promoter in the alveolar type II cells of the lung. We found early cystic lesions in a secondary organ (liver) that lacked the expression of bona fide lung makers namely Scgb1a1 and surfactant protein C Sftpc and were DsRed positive hence identifying lung as their source of origin. This demonstrates the significant potential of alveolar type II cells in orchestrating the process of metastasis, rendering it as one of the target cell types of the lung of therapeutic importance in human NSCLC. PMID:29464089

Oxidant-induced DNA damage of target cells.

PubMed Central

Schraufstätter, I; Hyslop, P A; Jackson, J H; Cochrane, C G

1988-01-01

In this study we examined the leukocytic oxidant species that induce oxidant damage of DNA in whole cells. H2O2 added extracellularly in micromolar concentrations (10-100 microM) induced DNA strand breaks in various target cells. The sensitivity of a specific target cell was inversely correlated to its catalase content and the rate of removal of H2O2 by the target cell. Oxidant species produced by xanthine oxidase/purine or phorbol myristate acetate-stimulated monocytes induced DNA breakage of target cells in proportion to the amount of H2O2 generated. These DNA strand breaks were prevented by extracellular catalase, but not by superoxide dismutase. Cytotoxic doses of HOCl, added to target cells, did not induce DNA strand breakage, and myeloperoxidase added extracellularly in the presence of an H2O2-generating system, prevented the formation of DNA strand breaks in proportion to its H2O2 degrading capacity. The studies also indicated that H2O2 formed hydroxyl radical (.OH) intracellularly, which appeared to be the most likely free radical responsible for DNA damage: .OH was detected in cells exposed to H2O2; the DNA base, deoxyguanosine, was hydroxylated in cells exposed to H2O2; and intracellular iron was essential for induction of DNA strand breaks. PMID:2843565

The B-cell identity factor Pax5 regulates distinct transcriptional programmes in early and late B lymphopoiesis

PubMed Central

Revilla-i-Domingo, Roger; Bilic, Ivan; Vilagos, Bojan; Tagoh, Hiromi; Ebert, Anja; Tamir, Ido M; Smeenk, Leonie; Trupke, Johanna; Sommer, Andreas; Jaritz, Markus; Busslinger, Meinrad

2012-01-01

Pax5 controls the identity and development of B cells by repressing lineage-inappropriate genes and activating B-cell-specific genes. Here, we used genome-wide approaches to identify Pax5 target genes in pro-B and mature B cells. In these cell types, Pax5 bound to 40% of the cis-regulatory elements defined by mapping DNase I hypersensitive (DHS) sites, transcription start sites and histone modifications. Although Pax5 bound to 8000 target genes, it regulated only 4% of them in pro-B and mature B cells by inducing enhancers at activated genes and eliminating DHS sites at repressed genes. Pax5-regulated genes in pro-B cells account for 23% of all expression changes occurring between common lymphoid progenitors and committed pro-B cells, which identifies Pax5 as an important regulator of this developmental transition. Regulated Pax5 target genes minimally overlap in pro-B and mature B cells, which reflects massive expression changes between these cell types. Hence, Pax5 controls B-cell identity and function by regulating distinct target genes in early and late B lymphopoiesis. PMID:22669466

Intradermal injection of an anti-Langerin-HIVGag fusion vaccine targets epidermal Langerhans cells in nonhuman primates and can be tracked in vivo.

PubMed

Salabert, Nina; Todorova, Biliana; Martinon, Frédéric; Boisgard, Raphaël; Zurawski, Gerard; Zurawski, Sandra; Dereuddre-Bosquet, Nathalie; Cosma, Antonio; Kortulewski, Thierry; Banchereau, Jacques; Levy, Yves; Le Grand, Roger; Chapon, Catherine

2016-03-01

The development of new immunization strategies requires a better understanding of early molecular and cellular events occurring at the site of injection. The skin is particularly rich in immune cells and represents an attractive site for vaccine administration. Here, we specifically targeted vaccine antigens to epidermal Langerhans cells (LCs) using a fusion protein composed of HIV antigens and a monoclonal antibody targeting Langerin. We developed a fluorescence imaging approach to visualize, in vivo, the vaccine-targeted cells. Studies were performed in nonhuman primates (NHPs) because of their relevance as a model to assess human vaccines. We directly demonstrated that in NHPs, intradermally injected anti-Langerin-HIVGag specifically targets epidermal LCs and induces rapid changes in the LC network, including LC activation and migration out of the epidermis. Vaccine targeting of LCs significantly improved anti-HIV immune response without requirement of an adjuvant. Although the co-injection of the TLR-7/8 synthetic ligand, R-848 (resiquimod), with the vaccine, did not enhance significantly the antibody response, it stimulated recruitment of HLA-DR+ inflammatory cells to the site of immunization. This study allowed us to characterize the dynamics of early local events following the injection of a vaccine-targeted epidermal LCs and R-848. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Visualization of early influenza A virus trafficking in human dendritic cells using STED microscopy.

PubMed

Baharom, Faezzah; Thomas, Oliver S; Lepzien, Rico; Mellman, Ira; Chalouni, Cécile; Smed-Sörensen, Anna

2017-01-01

Influenza A viruses (IAV) primarily target respiratory epithelial cells, but can also replicate in immune cells, including human dendritic cells (DCs). Super-resolution microscopy provides a novel method of visualizing viral trafficking by overcoming the resolution limit imposed by conventional light microscopy, without the laborious sample preparation of electron microscopy. Using three-color Stimulated Emission Depletion (STED) microscopy, we visualized input IAV nucleoprotein (NP), early and late endosomal compartments (EEA1 and LAMP1 respectively), and HLA-DR (DC membrane/cytosol) by immunofluorescence in human DCs. Surface bound IAV were internalized within 5 min of infection. The association of virus particles with early endosomes peaked at 5 min when 50% of NP+ signals were also EEA1+. Peak association with late endosomes occurred at 15 min when 60% of NP+ signals were LAMP1+. At 30 min of infection, the majority of NP signals were in the nucleus. Our findings illustrate that early IAV trafficking in human DCs proceeds via the classical endocytic pathway.

Transferrin-mediated rapid targeting, isolation, and detection of circulating tumor cells by multifunctional magneto-dendritic nanosystem.

PubMed

Banerjee, Shashwat S; Jalota-Badhwar, Archana; Satavalekar, Sneha D; Bhansali, Sujit G; Aher, Naval D; Mascarenhas, Russel R; Paul, Debjani; Sharma, Somesh; Khandare, Jayant J

2013-06-01

A multicomponent magneto-dendritic nanosystem (MDNS) is designed for rapid tumor cell targeting, isolation, and high-resolution imaging by a facile bioconjugation approach. The highly efficient and rapid-acting MDNS provides a convenient platform for simultaneous isolation and high-resolution imaging of tumor cells, potentially leading towards an early diagnosis of cancer. Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

[Clinical Advanced in Early-stage ALK-positive Non-small Cell Lung Cancer Patients].

PubMed

Gao, Qiongqiong; Jiang, Xiangli; Huang, Chun

2017-02-20

Lung cancer is the leading cause of cancer death in China. Non-small cell lung cancer (NSCLC) accounts for 85% of lung cancer cases, with the majority of the cases diagnosed at the advanced stage. Molecular targeted therapy is becoming the focus attention for advanced NSCLC. Echinoderm microtubule-associated protein-like 4 gene and the anaplastic lymphoma kinase gene (EML4-ALK) is among the most common molecular targets of NSCLC; its specific small-molecule tyrosine kinase inhibitors (TKIs) are approved for use in advanced NSCLC cases of ALK-positive. However, the influence of EML4-ALK fusion gene on the outcome of early-stage NSCLC cases and the necessity of application of TKIs for early-stage ALK-positive NSCLC patients are still uncertain. In this paper, we summarized the progression of testing methods for ALK-positive NSCLC patients as well as clinicopathological implication, outcome, and necessity of application of TKIs for early-stage ALK-positive NSCLC patients.

MOLECULARLY TARGETED THERAPIES IN NON-SMALL CELL LUNG CANCER ANNUAL UPDATE 2014

PubMed Central

Morgensztern, Daniel; Campo, Meghan J.; Dahlberg, Suzanne E.; Doebele, Robert C.; Garon, Edward; Gerber, David E.; Goldberg, Sarah B.; Hammerman, Peter S.; Heist, Rebecca; Hensing, Thomas; Horn, Leora; Ramalingam, Suresh S.; Rudin, Charles M.; Salgia, Ravi; Sequist, Lecia; Shaw, Alice T.; Simon, George R.; Somaiah, Neeta; Spigel, David R.; Wrangle, John; Johnson, David; Herbst, Roy S.; Bunn, Paul; Govindan, Ramaswamy

2015-01-01

There have been significant advances in the understanding of the biology and treatment of non-small cell lung cancer (NSCLC) over the past few years. A number of molecularly targeted agents are in the clinic or in development for patients with advanced NSCLC (Table 1). We are beginning to understand the mechanisms of acquired resistance following exposure to tyrosine kinase inhibitors in patients with oncogene addicted NSCLC. The advent of next generation sequencing has enabled to study comprehensively genomic alterations in lung cancer. Finally, early results from immune checkpoint inhibitors are very encouraging. This review summarizes recent advances in the area of cancer genomics, targeted therapies and immunotherapy. PMID:25535693

Designing oral vaccines targeting intestinal dendritic cells.

PubMed

Devriendt, Bert; De Geest, Bruno G; Cox, Eric

2011-04-01

Most pathogens colonize and invade the host at mucosal surfaces, such as the lung and the intestine. To combat intestinal pathogens the induction of local adaptive immune responses is required, which is mainly achieved through oral vaccination. However, most vaccines are ineffective when given orally owing to the hostile environment in the gastrointestinal tract. The encapsulation of antigens in biodegradable microparticulate delivery systems enhances their immunogenicity; however, the uptake of these delivery systems by intestinal immune cells is rather poor. Surface decoration of the particulates with targeting ligands could increase the uptake and mediate the selective targeting of the vaccine to intestinal antigen-presenting cells, including dendritic cells. In this review, current knowledge on dendritic cell subsets is discussed, along with progress in the development of selective antigen targeting to these cells, in addition to focusing on data obtained in mice and, where possible, the pig, as a non-rodent animal model for humans. Moreover, the potential use and benefits of Fcγ receptor-mediated targeting of antigen delivery systems are highlighted. In conclusion, dendritic cell targeting ligands grafted on antigen carrier systems should preferably bind to a conserved endocytotic receptor, facilitating the design of a multispecies vaccine platform, which could elicit robust protective immune responses against enteric pathogens.

Efficient priming of CD4 T cells by Langerin-expressing dendritic cells targeted with porcine epidemic diarrhea virus spike protein domains in pigs.

PubMed

Subramaniam, Sakthivel; Cao, Dianjun; Tian, Debin; Cao, Qian M; Overend, Christopher; Yugo, Danielle M; Matzinger, Shannon R; Rogers, Adam J; Heffron, C Lynn; Catanzaro, Nicholas; Kenney, Scott P; Opriessnig, Tanja; Huang, Yao-Wei; Labarque, Geoffrey; Wu, Stephen Q; Meng, Xiang-Jin

2017-01-02

Porcine epidemic diarrhea virus (PEDV) first emerged in the United States in 2013 causing high mortality and morbidity in neonatal piglets with immense economic losses to the swine industry. PEDV is an alpha-coronavirus replicating primarily in porcine intestinal cells. PEDV vaccines are available in Asia and Europe, and conditionally-licensed vaccines recently became available in the United States but the efficacies of these vaccines in eliminating PEDV from swine populations are questionable. In this study, the immunogenicity of a subunit vaccine based on the spike protein of PEDV, which was directly targeted to porcine dendritic cells (DCs) expressing Langerin, was assessed. The PEDV S antigen was delivered to the dendritic cells through a single-chain antibody specific to Langerin and the targeted cells were stimulated with cholera toxin adjuvant. This approach, known as "dendritic cell targeting," greatly improved PEDV S antigen-specific T cell interferon-γ responses in the CD4 pos CD8 pos T cell compartment in pigs as early as 7days upon transdermal administration. When the vaccine protein was targeted to Langerin pos DCs systemically through intramuscular vaccination, it induced higher serum IgG and IgA responses in pigs, though these responses require a booster dose, and the magnitude of T cell responses were lower as compared to transdermal vaccination. We conclude that PEDV spike protein domains targeting Langerin-expressing dendritic cells significantly increased CD4 T cell immune responses in pigs. The results indicate that the immunogenicity of protein subunit vaccines can be greatly enhanced by direct targeting of the vaccine antigens to desirable dendritic cell subsets in pigs. Copyright © 2016 Elsevier B.V. All rights reserved.

Shared target antigens on cancer cells and tissue stem cells: go or no-go for CAR T cells?

PubMed

Hombach, Andreas A; Abken, Hinrich

2017-02-01

Adoptive therapy with chimeric antigen receptor (CAR) T cells redirected towards CD19 produces remissions of B cell malignancies, however, it also eradicates healthy B cells sharing the target antigen. Such 'on-target off-tumor' toxicity raises serious safety concerns when the target antigen is also expressed by tissue stem cells, with the risk of lasting tissue destruction. Areas covered: We discuss CAR T cell targeting of activation antigens versus lineage associated antigens on the basis of recent experimental and animal data and the literature in the field. Expert commentary: Targeting an activation associated antigen which is transiently expressed by stem cells seems to be safe, like CAR T cells targeting CD30 spare CD30 + hematopoietic stem and progenitor cells while eliminating CD30 + lymphoma cells, whereas targeting lineage associated antigens which increase in expression during cell maturation, like folate receptor-β and CD123, is of risk to destruct tissue stem cells.

Targeting tumor cell motility to prevent metastasis

PubMed Central

Palmer, Trenis D.; Ashby, William J.; Lewis, John D.; Zijlstra, Andries

2011-01-01

Mortality and morbidity in patients with solid tumors invariably results from the disruption of normal biological function caused by disseminating tumor cells. Tumor cell migration is under intense investigation as the underlying cause of cancer metastasis. The need for tumor cell motility in the progression of metastasis has been established experimentally and is supported empirically by basic and clinical research implicating a large collection of migration-related genes. However, there are few clinical interventions designed to specifically target the motility of tumor cells and adjuvant therapy to specifically prevent cancer cell dissemination is severely limited. In an attempt to define motility targets suitable for treating metastasis, we have parsed the molecular determinants of tumor cell motility into five underlying principles including cell autonomous ability, soluble communication, cell-cell adhesion, cell-matrix adhesion, and integrating these determinants of migration on molecular scaffolds. The current challenge is to implement meaningful and sustainable inhibition of metastasis by developing clinically viable disruption of molecular targets that control these fundamental capabilities. PMID:21664937

VEGF is a Promising Therapeutic Target for the Treatment of Clear Cell Carcinoma of the Ovary

PubMed Central

Mabuchi, Seiji; Kawase, Chiaki; Altomare, Deborah A.; Morishige, Kenichirou; Hayashi, Masami; Sawada, Kenjiro; Ito, Kimihiko; Terai, Yoshito; Nishio, Yukihiro; Klein-Szanto, Andres J.; Burger, Robert A.; Ohmichi, Masahide; Testa, Joseph R.; Kimura, Tadashi

2010-01-01

This study examined the role of VEGF as a therapeutic target in clear cell carcinoma (CCC) of the ovary, which has been regarded as a chemoresistant histological subtype. Immunohistochemical analysis using tissue microarrays of 98 primary ovarian cancers revealed that VEGF was strongly expressed both in early stage and advanced stage CCC of the ovary. In early stage CCCs, patients who had tumors with high levels of VEGF had significantly shorter survival than those with low levels of VEGF. In vitro experiments revealed that VEGF expression was significantly higher in cisplatin-refractory human clear cell carcinoma cells (RMG1-CR and KOC7C-CR), compared to the respective parental cells (RMG1 and KOC7C) in the presence of cisplatin. In vivo treatment with bevacizumab markedly inhibited the growth of both parental CCC cells-derived (RMG1 and KOC7C) and cisplatin-refractory CCC cells-derived (RMG1-CR and KOC7C-CR) tumors as a result of inhibition of tumor angiogenesis. The results of the current study indicate that VEGF is frequently expressed and can be a promising therapeutic target in the management of CCC. Bevacizumab may be efficacious not only as a first-line treatment but also as a second-line treatment of recurrent disease in patients previously treated with cisplatin. PMID:20663925

Photoacoustic imaging with an acoustic lens detects prostate cancer cells labeled with PSMA-targeting near-infrared dye-conjugates

NASA Astrophysics Data System (ADS)

Dogra, Vikram; Chinni, Bhargava; Singh, Shalini; Schmitthenner, Hans; Rao, Navalgund; Krolewski, John J.; Nastiuk, Kent L.

2016-06-01

There is an urgent need for sensitive and specific tools to accurately image early stage, organ-confined human prostate cancers to facilitate active surveillance and reduce unnecessary treatment. Recently, we developed an acoustic lens that enhances the sensitivity of photoacoustic imaging. Here, we report the use of this device in conjunction with two molecular imaging agents that specifically target the prostate-specific membrane antigen (PSMA) expressed on the tumor cell surface of most prostate cancers. We demonstrate successful imaging of phantoms containing cancer cells labeled with either of two different PSMA-targeting agents, the ribonucleic acid aptamer A10-3.2 and a urea-based peptidomimetic inhibitor, each linked to the near-infrared dye IRDye800CW. By specifically targeting cells with these agents linked to a dye chosen for optimal signal, we are able to discriminate prostate cancer cells that express PSMA.

Single-cell multi-omics sequencing of mouse early embryos and embryonic stem cells.

PubMed

Guo, Fan; Li, Lin; Li, Jingyun; Wu, Xinglong; Hu, Boqiang; Zhu, Ping; Wen, Lu; Tang, Fuchou

2017-08-01

Single-cell epigenome sequencing techniques have recently been developed. However, the combination of different layers of epigenome sequencing in an individual cell has not yet been achieved. Here, we developed a single-cell multi-omics sequencing technology (single-cell COOL-seq) that can analyze the chromatin state/nucleosome positioning, DNA methylation, copy number variation and ploidy simultaneously from the same individual mammalian cell. We used this method to analyze the reprogramming of the chromatin state and DNA methylation in mouse preimplantation embryos. We found that within < 12 h of fertilization, each individual cell undergoes global genome demethylation together with the rapid and global reprogramming of both maternal and paternal genomes to a highly opened chromatin state. This was followed by decreased openness after the late zygote stage. Furthermore, from the late zygote to the 4-cell stage, the residual DNA methylation is preferentially preserved on intergenic regions of the paternal alleles and intragenic regions of maternal alleles in each individual blastomere. However, chromatin accessibility is similar between paternal and maternal alleles in each individual cell from the late zygote to the blastocyst stage. The binding motifs of several pluripotency regulators are enriched at distal nucleosome depleted regions from as early as the 2-cell stage. This indicates that the cis-regulatory elements of such target genes have been primed to an open state from the 2-cell stage onward, long before pluripotency is eventually established in the ICM of the blastocyst. Genes may be classified into homogeneously open, homogeneously closed and divergent states based on the chromatin accessibility of their promoter regions among individual cells. This can be traced to step-wise transitions during preimplantation development. Our study offers the first single-cell and parental allele-specific analysis of the genome-scale chromatin state and DNA

Single-cell multi-omics sequencing of mouse early embryos and embryonic stem cells

PubMed Central

Guo, Fan; Li, Lin; Li, Jingyun; Wu, Xinglong; Hu, Boqiang; Zhu, Ping; Wen, Lu; Tang, Fuchou

2017-01-01

Single-cell epigenome sequencing techniques have recently been developed. However, the combination of different layers of epigenome sequencing in an individual cell has not yet been achieved. Here, we developed a single-cell multi-omics sequencing technology (single-cell COOL-seq) that can analyze the chromatin state/nucleosome positioning, DNA methylation, copy number variation and ploidy simultaneously from the same individual mammalian cell. We used this method to analyze the reprogramming of the chromatin state and DNA methylation in mouse preimplantation embryos. We found that within < 12 h of fertilization, each individual cell undergoes global genome demethylation together with the rapid and global reprogramming of both maternal and paternal genomes to a highly opened chromatin state. This was followed by decreased openness after the late zygote stage. Furthermore, from the late zygote to the 4-cell stage, the residual DNA methylation is preferentially preserved on intergenic regions of the paternal alleles and intragenic regions of maternal alleles in each individual blastomere. However, chromatin accessibility is similar between paternal and maternal alleles in each individual cell from the late zygote to the blastocyst stage. The binding motifs of several pluripotency regulators are enriched at distal nucleosome depleted regions from as early as the 2-cell stage. This indicates that the cis-regulatory elements of such target genes have been primed to an open state from the 2-cell stage onward, long before pluripotency is eventually established in the ICM of the blastocyst. Genes may be classified into homogeneously open, homogeneously closed and divergent states based on the chromatin accessibility of their promoter regions among individual cells. This can be traced to step-wise transitions during preimplantation development. Our study offers the first single-cell and parental allele-specific analysis of the genome-scale chromatin state and DNA

Gold nano-popcorn attached SWCNT hybrid nanomaterial for targeted diagnosis and photothermal therapy of human breast cancer cells.

PubMed

Beqa, Lule; Fan, Zhen; Singh, Anant Kumar; Senapati, Dulal; Ray, Paresh Chandra

2011-09-01

Breast cancer presents greatest challenge in health care in today's world. The key to ultimately successful treatment of breast cancer disease is an early and accurate diagnosis. Current breast cancer treatments are often associated with severe side effects. Driven by the need, we report the design of novel hybrid nanomaterial using gold nano popcorn-attached single wall carbon nanotube for targeted diagnosis and selective photothermal treatment. Targeted SK-BR-3 human breast cancer cell sensing have been performed in 10 cancer cells/mL level, using surface enhanced Raman scattering of single walls carbon nanotube's D and G bands. Our data show that S6 aptamer attached hybrid nanomaterial based SERS assay is highly sensitive to targeted human breast cancer SK-BR-3 cell line and it will be able to distinguish it from other non targeted MDA-MB breast cancer cell line and HaCaT normal skin cell line. Our results also show that 10 min of photothermal therapy treatment by 1.5 W/cm(2) power, 785 nm laser is enough to kill cancer cells very effectively using S6 aptamer attached hybrid nanomaterials. Possible mechanisms for targeted sensing and operating principle for highly efficient photothermal therapy have been discussed. Our experimental results reported here open up a new possibility for using aptamers modified hybrid nanomaterial for reliable diagnosis and targeted therapy of cancer cell lines quickly.

Cytotoxic T cells use mechanical force to potentiate target cell killing

PubMed Central

Basu, Roshni; Whitlock, Benjamin M.; Husson, Julien; Le Floc’h, Audrey; Jin, Weiyang; Oyler-Yaniv, Alon; Dotiwala, Farokh; Giannone, Gregory; Hivroz, Claire; Biais, Nicolas; Lieberman, Judy; Kam, Lance C.; Huse, Morgan

2016-01-01

SUMMARY The immunological synapse formed between a cytotoxic T lymphocyte (CTL) and an infected or transformed target cell is a physically active structure capable of exerting mechanical force. Here, we investigated whether synaptic forces promote the destruction of target cells. CTLs kill by secreting toxic proteases and the pore forming protein perforin into the synapse. Biophysical experiments revealed a striking correlation between the magnitude of force exertion across the synapse and the speed of perforin pore formation on the target cell, implying that force potentiates cytotoxicity by enhancing perforin activity. Consistent with this interpretation, we found that increasing target cell tension augmented pore formation by perforin and killing by CTLs. Our data also indicate that CTLs coordinate perforin release and force exertion in space and time. These results reveal an unappreciated physical dimension to lymphocyte function and demonstrate that cells use mechanical forces to control the activity of outgoing chemical signals. PMID:26924577

Cytotoxic T Cells Use Mechanical Force to Potentiate Target Cell Killing.

PubMed

Basu, Roshni; Whitlock, Benjamin M; Husson, Julien; Le Floc'h, Audrey; Jin, Weiyang; Oyler-Yaniv, Alon; Dotiwala, Farokh; Giannone, Gregory; Hivroz, Claire; Biais, Nicolas; Lieberman, Judy; Kam, Lance C; Huse, Morgan

2016-03-24

The immunological synapse formed between a cytotoxic T lymphocyte (CTL) and an infected or transformed target cell is a physically active structure capable of exerting mechanical force. Here, we investigated whether synaptic forces promote the destruction of target cells. CTLs kill by secreting toxic proteases and the pore forming protein perforin into the synapse. Biophysical experiments revealed a striking correlation between the magnitude of force exertion across the synapse and the speed of perforin pore formation on the target cell, implying that force potentiates cytotoxicity by enhancing perforin activity. Consistent with this interpretation, we found that increasing target cell tension augmented pore formation by perforin and killing by CTLs. Our data also indicate that CTLs coordinate perforin release and force exertion in space and time. These results reveal an unappreciated physical dimension to lymphocyte function and demonstrate that cells use mechanical forces to control the activity of outgoing chemical signals. Copyright © 2016 Elsevier Inc. All rights reserved.

Cell-to-Cell Transmission Can Overcome Multiple Donor and Target Cell Barriers Imposed on Cell-Free HIV

PubMed Central

Ilinskaya, Anna; Dorjbal, Batsukh; Truong, Rosaline; Derse, David; Uchil, Pradeep D.; Heidecker, Gisela; Mothes, Walther

2013-01-01

Virus transmission can occur either by a cell-free mode through the extracellular space or by cell-to-cell transmission involving direct cell-to-cell contact. The factors that determine whether a virus spreads by either pathway are poorly understood. Here, we assessed the relative contribution of cell-free and cell-to-cell transmission to the spreading of the human immunodeficiency virus (HIV). We demonstrate that HIV can spread by a cell-free pathway if all the steps of the viral replication cycle are efficiently supported in highly permissive cells. However, when the cell-free path was systematically hindered at various steps, HIV transmission became contact-dependent. Cell-to-cell transmission overcame barriers introduced in the donor cell at the level of gene expression and surface retention by the restriction factor tetherin. Moreover, neutralizing antibodies that efficiently inhibit cell-free HIV were less effective against cell-to-cell transmitted virus. HIV cell-to-cell transmission also efficiently infected target T cells that were relatively poorly susceptible to cell-free HIV. Importantly, we demonstrate that the donor and target cell types influence critically the extent by which cell-to-cell transmission can overcome each barrier. Mechanistically, cell-to-cell transmission promoted HIV spread to more cells and infected target cells with a higher proviral content than observed for cell-free virus. Our data demonstrate that the frequently observed contact-dependent spread of HIV is the result of specific features in donor and target cell types, thus offering an explanation for conflicting reports on the extent of cell-to-cell transmission of HIV. PMID:23308151

(−)-Epigallocatechin Gallate Targets Notch to Attenuate the Inflammatory Response in the Immediate Early Stage in Human Macrophages

PubMed Central

Wang, Tengfei; Xiang, Zemin; Wang, Ya; Li, Xi; Fang, Chongye; Song, Shuang; Li, Chunlei; Yu, Haishuang; Wang, Han; Yan, Liang; Hao, Shumei; Wang, Xuanjun; Sheng, Jun

2017-01-01

Inflammation plays important roles at different stages of diabetes mellitus, tumorigenesis, and cardiovascular diseases. (−)-Epigallocatechin gallate (EGCG) can attenuate inflammatory responses effectively. However, the immediate early mechanism of EGCG in inflammation remains unclear. Here, we showed that EGCG attenuated the inflammatory response in the immediate early stage of EGCG treatment by shutting off Notch signaling and that the effect did not involve the 67-kDa laminin receptor, the common receptor for EGCG. EGCG eliminated mature Notch from the cell membrane and the nuclear Notch intercellular domain, the active form of Notch, within 2 min by rapid degradation via the proteasome pathway. Transcription of the Notch target gene was downregulated simultaneously. Knockdown of Notch 1/2 expression by RNA interference impaired the downregulation of the inflammatory response elicited by EGCG. Further study showed that EGCG inhibited lipopolysaccharide-induced inflammation and turned off Notch signaling in human primary macrophages. Taken together, our results show that EGCG targets Notch to regulate the inflammatory response in the immediate early stage. PMID:28443100

Single-Cell Droplet Microfluidic Screening for Antibodies Specifically Binding to Target Cells.

PubMed

Shembekar, Nachiket; Hu, Hongxing; Eustace, David; Merten, Christoph A

2018-02-20

Monoclonal antibodies are a main player in modern drug discovery. Many antibody screening formats exist, each with specific advantages and limitations. Nonetheless, it remains challenging to screen antibodies for the binding of cell-surface receptors (the most important class of all drug targets) or for the binding to target cells rather than purified proteins. Here, we present a high-throughput droplet microfluidics approach employing dual-color normalized fluorescence readout to detect antibody binding. This enables us to obtain quantitative data on target cell recognition, using as little as 33 fg of IgG per assay. Starting with an excess of hybridoma cells releasing unspecific antibodies, individual clones secreting specific binders (of target cells co-encapsulated into droplets) could be enriched 220-fold after sorting 80,000 clones in a single experiment. This opens the way for therapeutic antibody discovery, especially since the single-cell approach is in principle also applicable to primary human plasma cells. Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.

Modeling activity and target-dependent developmental cell death of mouse retinal ganglion cells ex vivo.

PubMed

Voyatzis, Sylvie; Muzerelle, Aude; Gaspar, Patricia; Nicol, Xavier

2012-01-01

Programmed cell death is widespread during the development of the central nervous system and serves multiple purposes including the establishment of neural connections. In the mouse retina a substantial reduction of retinal ganglion cells (RGCs) occurs during the first postnatal week, coinciding with the formation of retinotopic maps in the superior colliculus (SC). We previously established a retino-collicular culture preparation which recapitulates the progressive topographic ordering of RGC projections during early post-natal life. Here, we questioned whether this model could also be suitable to examine the mechanisms underlying developmental cell death of RGCs. Brn3a was used as a marker of the RGCs. A developmental decline in the number of Brn3a-immunolabelled neurons was found in the retinal explant with a timing that paralleled that observed in vivo. In contrast, the density of photoreceptors or of starburst amacrine cells increased, mimicking the evolution of these cell populations in vivo. Blockade of neural activity with tetrodotoxin increased the number of surviving Brn3a-labelled neurons in the retinal explant, as did the increase in target availability when one retinal explant was confronted with 2 or 4 collicular slices. Thus, this ex vivo model reproduces the developmental reduction of RGCs and recapitulates its regulation by neural activity and target availability. It therefore offers a simple way to analyze developmental cell death in this classic system. Using this model, we show that ephrin-A signaling does not participate to the regulation of the Brn3a population size in the retina, indicating that eprhin-A-mediated elimination of exuberant projections does not involve developmental cell death.

Identification of type IV collagen exposure as a molecular imaging target for early detection of thoracic aortic dissection

PubMed Central

Xu, Ke; Xu, Chen; Zhang, Yanzhenzi; Qi, Feiran; Yu, Bingran; Li, Ping; Jia, Lixin; Li, Yulin; Xu, Fu-jian; Du, Jie

2018-01-01

Thoracic aortic dissection (TAD) is an aggressive and life-threatening vascular disease and there is no effective means of early diagnosis of dissection. Type IV collagen (Col-IV) is a major component of the sub-endothelial basement membrane, which is initially exposed followed by endothelial injury as early-stage event of TAD. So, we want to build a noninvasive diagnostic method to detect early dissection by identifying the exposed Col-IV via MRI. Methods: Col-IV-targeted magnetic resonance/ fluorescence dual probe (Col-IV-DOTA-Gd-rhodamine B; CDR) was synthesized by amide reaction and coordination reaction. Flow cytometry analysis was used to evaluate the cell viability of SMC treated with CDR and fluorescence assays were used to assess the Col-IV targeting ability of CDR in vitro. We then examined the sensitivity and specificity of CDR at different stages of TAD via MRI and bioluminescence imaging in vivo. Results: The localization of Col-IV (under the intima) was observed by histology images. CDR bound specifically to Col-IV-expressing vascular smooth muscle cells and BAPN-induced dissected aorta. The CDR signal was co-detected by magnetic resonance imaging (MRI) and bioluminescence imaging as early as 2 weeks after BAPN administration (pre-dissection stage). The ability to detect rupture of dissected aorta was indicated by a strong normalized signal enhancement (NSE) in vivo. Moreover, NSE was negatively correlated with the time of dissection rupture after BAPN administration (r2 = 0.8482). Conclusion: As confirmed by in vivo studies, the CDR can identify the exposed Col-IV in degenerated aorta to monitor the progress of aortic dissection from the early stage to the rupture via MRI. Thus, CDR-enhanced MRI proposes a potential method for dissection screening, and for monitoring disease progression and therapeutic response. PMID:29290819

Orchestration of transplantation tolerance by regulatory dendritic cell therapy or in situ targeting of dendritic cells

PubMed Central

Morelli, Adrian E.; Thomson, Angus W.

2014-01-01

Purpose of review Extensive research in murine transplant models over the past two decades has convincingly demonstrated the ability of regulatory dendritic cells (DCreg) to promote long-term allograft survival. We review important considerations regarding the source of therapeutic DCreg (donor or recipient) and their mode of action, in situ targeting of DCreg, and optimal therapeutic regimens to promote DCreg function. Recent findings Recent studies have defined protocols and mechanisms whereby ex vivo-generated DCreg of donor or recipient origin subvert allogeneic T cell responses and promote long-term organ transplant survival. Particular interest has focused on how donor antigen (Ag) is acquired, processed and presented by autologous DCs, on the stability of DCreg, and on in situ targeting of DC to promote their tolerogenic function. New evidence of the therapeutic efficacy of DCreg in a clinically-relevant non-human primate organ transplant model and production of clinical grade DCreg support early evaluation of DCreg therapy in human graft recipients. Summary We discuss strategies currently used to promote DC tolerogenicity, including DCreg therapy and in situ targeting of DC, with a view to improved understanding of underlying mechanisms and identification of the most promising strategies for therapeutic application. PMID:24926700

Controversies in targeted therapy of adult T cell leukemia/lymphoma: ON target or OFF target effects?

PubMed

Nasr, Rihab; El Hajj, Hiba; Kfoury, Youmna; de Thé, Hugues; Hermine, Olivier; Bazarbachi, Ali

2011-06-01

Adult T cell leukemia/lymphoma (ATL) represents an ideal model for targeted therapy because of intrinsic chemo-resistance of ATL cells and the presence of two well identified targets: the HTLV-I retrovirus and the viral oncoprotein Tax. The combination of zidovudine (AZT) and interferon-alpha (IFN) has a dramatic impact on survival of ATL patients. Although the mechanism of action remains unclear, arguments in favor or against a direct antiviral effect will be discussed. Yet, most patients relapse and alternative therapies are mandatory. IFN and arsenic trioxide induce Tax proteolysis, synergize to induce apoptosis in ATL cells and cure Tax-driven ATL in mice through specific targeting of leukemia initiating cell activity. These results provide a biological basis for the clinical success of arsenic/IFN/AZT therapy in ATL patients and suggest that both extinction of viral replication (AZT) and Tax degradation (arsenic/IFN) are needed to cure ATL.

Antibody-targeted interleukin 2 stimulates T-cell killing of autologous tumor cells.

PubMed Central

Gillies, S D; Reilly, E B; Lo, K M; Reisfeld, R A

1992-01-01

A genetically engineered fusion protein consisting of a chimeric anti-ganglioside GD2 antibody (ch14.18) and interleukin 2 (IL2) was tested for its ability to enhance the killing of autologous GD2-expressing melanoma target cells by a tumor-infiltrating lymphocyte line (660 TIL). The fusion of IL2 to the carboxyl terminus of the immunoglobulin heavy chain did not reduce IL2 activity as measured in a standard proliferation assay using either mouse or human T-cell lines. Antigen-binding activity was greater than that of the native chimeric antibody. The ability of resting 660 TIL cells to kill their autologous GD2-positive target cells was enhanced if the target cells were first coated with the fusion protein. This stimulation of killing was greater than that of uncoated cells in the presence of equivalent or higher concentrations of free IL2. Such antibody-cytokine fusion proteins may prove useful in targeting the biological effect of IL2 and other cytokines to tumor cells and in this way stimulate their immune destruction. Images PMID:1741398

Reproductive Toxicity of T Cells in Early Life: Abnormal Immune Development and Postnatal Diseases.

PubMed

Liu, Han-Xiao; Jiang, Aifang; Chen, Ting; Qu, Wen; Yan, Hui-Yi; Ping, Jie

2017-01-01

Immunity is a balanced status with adequate biological defenses to recognize and fight "non-self", as well as adequate tolerance to recognize "self". To maintain this immune homeostasis, a well-organized T cell immune network is required, which in part depends on the well-controlled development of alternative effector T cells, with different cytokine repertoires. Recent researches have pointed that developing fetal T cells network is a remarkably sensitive toxicological target for adverse factors in early life. Epidemiological and experimental studies showed an inseparable relationship between T cell developmental toxicity and immune diseases in adults. Considering that the inflammatory and immune disorders have become a growing health problem worldwide, increasing attention is now being paid to the T cell developmental toxicity. We propose that adverse factors may have programming effects on the crucial functions of immune system during early life which is critical for fetal T cell development and the establishment of the distinct T cell repertoires balance. The permanently disturbed intrathymic or peripheral T cell development may in turn lead to the immune disorders in later life. In this manuscript, we reviewed how adverse factors affected T cell development in early-life with the consequence of the immune dysfunction and immune diseases, and further elucidate the mechanisms. These mechanisms will be helpful in prevention and treatment of the increased prevalence of immune diseases by interfering those pathways. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Semiconductor nanocrystal-aptamer bioconjugate probes for specific prostate carcinoma cell targeting

NASA Astrophysics Data System (ADS)

Shieh, Felice; Lavery, Laura; Chu, Chitai T.; Richards-Kortum, Rebecca; Ellington, Andrew D.; Korgel, Brian A.

2005-04-01

Cancer of the prostate affects approximately 1 in 11 men. Current early screening for prostate cancer utilizes digital rectal examinations to detect anomalies in the prostate gland and blood test screenings for upregulated levels of prostate specific antigen (PSA). Many of these tests are invasive and can often be inconclusive as PSA levels may be heightened due to benign factors. Prostate specific membrane antigen (PSMA), a well-characterized integral membrane protein, is expressed in virtually all prostate cancers and often correlates with cancer aggressiveness. Therefore, it may be used as an indicator of cancer growth and metastases. PSMA-specific antibodies have been identified and conjugated to fluorescent markers for cancer cell targeting; however, both the antibodies and markers possess significant limitations in their pharmaceutical and diagnostic value. Here we report the use of semiconductor nanocrystals bioconjugated to PSMA-specific aptamer recognition molecules for prostate carcinoma cell targeting. The nanocrystal/aptamer bioconjugates are small biocompatible probes with the potential for color-tunability for multicolor imaging. Ongoing in vitro and in vivo research seeks to introduce these nanoparticle bioconjugates into medical diagnostics.

Gold Nano Popcorn Attached SWCNT Hybrid Nanomaterial for Targeted Diagnosis and Photothermal Therapy of Human Breast Cancer Cells

PubMed Central

Beqa, Lule; Fan, Zhen; Singh, Anant Kumar; Senapati, Dulal; Ray, Paresh Chandra

2011-01-01

Breast cancer presents greatest challenge in health care in today’s world. The key to ultimately successful treatment of breast cancer disease is an early and accurate diagnosis. Current breast cancer treatments are often associated with severe side effects. Driven by the need, we report the design of novel hybrid nanomaterial using gold nano popcorn-attached single wall carbon nanotube for targeted diagnosis and selective photothermal treatment. Targeted SK-BR-3 human breast cancer cell sensing have been performed in 10 cancer cells/mL level, using surface enhanced Raman scattering of single walls carbon nanotube’s D and G bands. Our data show that S6 aptamer attached hybrid nanomaterial based SERS assay is highly sensitive to targeted human breast cancer SK-BR-3 cell line and it will be able to distinguish it from other non targeted MDA-MB breast cancer cell line and HaCaT normal skin cell line. Our results also show that 10 minutes of photothermal therapy treatment by 1.5 W/cm2 power, 785 nm laser is enough to kill cancer cells very effectively using S6 aptamer attached hybrid nanomaterials. Possible mechanisms for targeted sensing and operating principle for highly efficient photothermal therapy have been discussed. Our experimental results reported here open up a new possibility for using aptamers modified hybrid nanomaterial for reliable diagnosis and targeted therapy of cancer cell lines quickly. PMID:21842867

Triple DMARD treatment in early rheumatoid arthritis modulates synovial T cell activation and plasmablast/plasma cell differentiation pathways

PubMed Central

Wechalekar, Mihir D.; Guo, Yanxia; Yin, Xuefeng; Weedon, Helen; Proudman, Susanna M.; Smith, Malcolm D.; Nagpal, Sunil

2017-01-01

Objectives This study sought to investigate the genome-wide transcriptional effects of a combination of disease modifying anti-rheumatic drugs (tDMARD; methotrexate, sulfasalazine and hydroxychloroquine) in synovial tissues obtained from early rheumatoid arthritis (RA) patients. While combination DMARD strategies have been investigated for clinical efficacy, very little data exists on the potential molecular mechanism of action. We hypothesized that tDMARD would impact multiple biological pathways, but the specific pathways were unknown. Methods Paired synovial biopsy samples from early RA patients before and after 6 months of tDMARD therapy were collected by arthroscopy (n = 19). These biopsies as well as those from subjects with normal synovium (n = 28) were profiled by total RNA sequencing. Results Large differences in gene expression between RA and control biopsies (over 5000 genes) were identified. Despite clinical efficacy, the expression of a restricted set of less than 300 genes was reversed after 6 months of treatment. Many genes remained elevated, even in patients who achieved low disease activity. Interestingly, tDMARD downregulated genes included those involved in T cell activation and signaling and plasmablast/plasma cell differentiation and function. Conclusions We have identified transcriptomic signatures that characterize synovial tissue from RA patients with early disease. Analysis after 6 months of tDMARD treatment highlight consistent alterations in expression of genes related to T cell activation and plasmablast/plasma cell differentiation. These results provide novel insight into the biology of early RA and the mechanism of tDMARD action and may help identify novel drug targets to improve rates of treatment-induced disease remission. PMID:28863153

Triple DMARD treatment in early rheumatoid arthritis modulates synovial T cell activation and plasmablast/plasma cell differentiation pathways.

PubMed

Walsh, Alice M; Wechalekar, Mihir D; Guo, Yanxia; Yin, Xuefeng; Weedon, Helen; Proudman, Susanna M; Smith, Malcolm D; Nagpal, Sunil

2017-01-01

This study sought to investigate the genome-wide transcriptional effects of a combination of disease modifying anti-rheumatic drugs (tDMARD; methotrexate, sulfasalazine and hydroxychloroquine) in synovial tissues obtained from early rheumatoid arthritis (RA) patients. While combination DMARD strategies have been investigated for clinical efficacy, very little data exists on the potential molecular mechanism of action. We hypothesized that tDMARD would impact multiple biological pathways, but the specific pathways were unknown. Paired synovial biopsy samples from early RA patients before and after 6 months of tDMARD therapy were collected by arthroscopy (n = 19). These biopsies as well as those from subjects with normal synovium (n = 28) were profiled by total RNA sequencing. Large differences in gene expression between RA and control biopsies (over 5000 genes) were identified. Despite clinical efficacy, the expression of a restricted set of less than 300 genes was reversed after 6 months of treatment. Many genes remained elevated, even in patients who achieved low disease activity. Interestingly, tDMARD downregulated genes included those involved in T cell activation and signaling and plasmablast/plasma cell differentiation and function. We have identified transcriptomic signatures that characterize synovial tissue from RA patients with early disease. Analysis after 6 months of tDMARD treatment highlight consistent alterations in expression of genes related to T cell activation and plasmablast/plasma cell differentiation. These results provide novel insight into the biology of early RA and the mechanism of tDMARD action and may help identify novel drug targets to improve rates of treatment-induced disease remission.

Cell-targeted platinum nanoparticles and nanoparticle clusters.

PubMed

Papst, Stefanie; Brimble, Margaret A; Evans, Clive W; Verdon, Daniel J; Feisst, Vaughan; Dunbar, P Rod; Tilley, Richard D; Williams, David E

2015-06-21

Herein, we report the facile preparation of cell-targeted platinum nanoparticles (PtNPs), through the design of peptides that, as a single molecule added in small concentration during the synthesis, control the size of PtNP clusters during their growth, stabilise the PtNPs in aqueous suspension and enable the functionalisation of the PtNPs with a versatile range of cell-targeting ligands. Water-soluble PtNPs targeted respectively at blood group antigens and at integrin receptors are demonstrated.

Imaging of targeted lipid microbubbles to detect cancer cells using third harmonic generation microscopy

PubMed Central

Harpel, Kaitlin; Baker, Robert Dawson; Amirsolaimani, Babak; Mehravar, Soroush; Vagner, Josef; Matsunaga, Terry O.; Banerjee, Bhaskar; Kieu, Khanh

2016-01-01

The use of receptor-targeted lipid microbubbles imaged by ultrasound is an innovative method of detecting and localizing disease. However, since ultrasound requires a medium between the transducer and the object being imaged, it is impractical to apply to an exposed surface in a surgical setting where sterile fields need be maintained and ultrasound gel may cause the bubbles to collapse. Multiphoton microscopy (MPM) is an emerging tool for accurate, label-free imaging of tissues and cells with high resolution and contrast. We have recently determined a novel application of MPM to be used for detecting targeted microbubble adherence to the upregulated plectin-receptor on pancreatic tumor cells. Specifically, the third-harmonic generation response can be used to detect bound microbubbles to various cell types presenting MPM as an alternative and useful imaging method. This is an interesting technique that can potentially be translated as a diagnostic tool for the early detection of cancer and inflammatory disorders. PMID:27446711

Human induced pluripotent stem cells labeled with fluorescent magnetic nanoparticles for targeted imaging and hyperthermia therapy for gastric cancer.

PubMed

Li, Chao; Ruan, Jing; Yang, Meng; Pan, Fei; Gao, Guo; Qu, Su; Shen, You-Lan; Dang, Yong-Jun; Wang, Kan; Jin, Wei-Lin; Cui, Da-Xiang

2015-09-01

Human induced pluripotent stem (iPS) cells exhibit great potential for generating functional human cells for medical therapies. In this paper, we report for use of human iPS cells labeled with fluorescent magnetic nanoparticles (FMNPs) for targeted imaging and synergistic therapy of gastric cancer cells in vivo. Human iPS cells were prepared and cultured for 72 h. The culture medium was collected, and then was co-incubated with MGC803 cells. Cell viability was analyzed by the MTT method. FMNP-labeled human iPS cells were prepared and injected into gastric cancer-bearing nude mice. The mouse model was observed using a small-animal imaging system. The nude mice were irradiated under an external alternating magnetic field and evaluated using an infrared thermal mapping instrument. Tumor sizes were measured weekly. iPS cells and the collected culture medium inhibited the growth of MGC803 cells. FMNP-labeled human iPS cells targeted and imaged gastric cancer cells in vivo, as well as inhibited cancer growth in vivo through the external magnetic field. FMNP-labeled human iPS cells exhibit considerable potential in applications such as targeted dual-mode imaging and synergistic therapy for early gastric cancer.

MiR-615 inhibits cell proliferation, migration and invasion by targeting EGFR in human glioblastoma.

PubMed

Ji, Yanwei; Sun, Qingshan; Zhang, Jianbin; Hu, Haoran

2018-05-15

MiR-615 and epidermal growth factor receptor (EGFR) are associated with a number of disease processes and pathogenesis. However, little is known about the mechanisms of miR-615 and EGFR in human glioblastoma multiforme (GBM). Here, we found that down-regulation of miR-615 expression occurred in GBM tissues and cells, and was inversely correlated with overall survival, relapse-free survival, WHO grade as well as EGFR expression. We further determined that miR-615 functions as a tumor suppressor by inhibiting GBM cell proliferation, cell cycle, migration and invasion, and promoting cell apoptosis. In-vivo assay validated the inhibition effect of miR-615 on tumor growth and EGFR expression. Luciferase reporter assays demonstrated that miR-615 targeted the 3'-untranslated region (3'-UTR) of EGFR. Besides, over-expression of EGFR reversed the inhibition effects of miR-615, while silencing of EGFR aggravated these inhibition effects. In conclusions, we identified that miR-615 plays a tumor suppressor role in GBM cell proliferation, migration and invasion by targeting EGFR expression, and miR-615 may act as a novel biomarker for early diagnosis or therapeutic targets of GBM. Copyright © 2018 Elsevier Inc. All rights reserved.

Controversies in Targeted Therapy of Adult T Cell Leukemia/Lymphoma: ON Target or OFF Target Effects?

PubMed Central

Nasr, Rihab; Hajj, Hiba El; Kfoury, Youmna; de Thé, Hugues; Hermine, Olivier; Bazarbachi, Ali

2011-01-01

Adult T cell leukemia/lymphoma (ATL) represents an ideal model for targeted therapy because of intrinsic chemo-resistance of ATL cells and the presence of two well identified targets: the HTLV-I retrovirus and the viral oncoprotein Tax. The combination of zidovudine (AZT) and interferon-alpha (IFN) has a dramatic impact on survival of ATL patients. Although the mechanism of action remains unclear, arguments in favor or against a direct antiviral effect will be discussed. Yet, most patients relapse and alternative therapies are mandatory. IFN and arsenic trioxide induce Tax proteolysis, synergize to induce apoptosis in ATL cells and cure Tax-driven ATL in mice through specific targeting of leukemia initiating cell activity. These results provide a biological basis for the clinical success of arsenic/IFN/AZT therapy in ATL patients and suggest that both extinction of viral replication (AZT) and Tax degradation (arsenic/IFN) are needed to cure ATL. PMID:21994752

Triggering receptor expressed on myeloid cells 2 (TREM2): a potential therapeutic target for Alzheimer disease?

PubMed

Deming, Yuetiva; Li, Zeran; Benitez, Bruno A; Cruchaga, Carlos

2018-06-20

There are currently no effective therapeutics for Alzheimer disease (AD). Clinical trials targeting amyloid beta thus far have shown very little benefit and only in the earliest stages of disease. These limitations have driven research to identify alternative therapeutic targets, one of the most promising is the triggering receptor expressed on myeloid cells 2 (TREM2). Areas covered: Here, we review the literature to-date and discuss the potentials and pitfalls for targeting TREM2 as a potential therapeutic for AD. We focus on research in animal and cell models for AD and central nervous system injury models which may help in understanding the role of TREM2 in disease. Expert opinion: Studies suggest TREM2 plays a key role in AD pathology; however, results have been conflicting about whether TREM2 is beneficial or harmful. More research is necessary before designing TREM2-targeting therapies. Successful therapeutics will most likely be administered early in disease.

Early aggressive intraocular pressure lowering, target intraocular pressure, and a novel concept for glaucoma care.

PubMed

Singh, Kuldev; Shrivastava, Anurag

2008-11-01

Early diagnosis of glaucomatous optic nerve damage offers the potential for early treatment which may prevent vision loss from this neurodegenerative disease. Even in patients who do not begin early treatment, early diagnosis allows for better monitoring of disease at a stage where the stakes are lower relative to later stages of the disease. For these reasons, early diagnosis of glaucomatous optic nerve disease is desirable and opens the door for appropriately aggressive therapy. The target intraocular pressure (IOP) concept is widely employed by glaucoma practitioners. Although there have been no randomized clinical trials or other high-quality studies showing the benefits of setting a target IOP versus not using this approach, there is ample evidence that lowering IOP slows glaucoma progression and, in general, lower is better, regardless of disease stage and baseline IOP level. We propose an alternative approach to managing glaucoma without the use of the target IOP concept and suggest that this market IOP concept should be compared with the target IOP approach in an appropriately powered comparative clinical trial.

The SPOR Domain, a Widely Conserved Peptidoglycan Binding Domain That Targets Proteins to the Site of Cell Division.

PubMed

Yahashiri, Atsushi; Jorgenson, Matthew A; Weiss, David S

2017-07-15

Sporulation-related repeat (SPOR) domains are small peptidoglycan (PG) binding domains found in thousands of bacterial proteins. The name "SPOR domain" stems from the fact that several early examples came from proteins involved in sporulation, but SPOR domain proteins are quite diverse and contribute to a variety of processes that involve remodeling of the PG sacculus, especially with respect to cell division. SPOR domains target proteins to the division site by binding to regions of PG devoid of stem peptides ("denuded" glycans), which in turn are enriched in septal PG by the intense, localized activity of cell wall amidases involved in daughter cell separation. This targeting mechanism sets SPOR domain proteins apart from most other septal ring proteins, which localize via protein-protein interactions. In addition to SPOR domains, bacteria contain several other PG-binding domains that can exploit features of the cell wall to target proteins to specific subcellular sites. Copyright © 2017 American Society for Microbiology.

"Kill" the messenger: Targeting of cell-derived microparticles in lupus nephritis.

PubMed

Nielsen, Christoffer T; Rasmussen, Niclas S; Heegaard, Niels H H; Jacobsen, Søren

2016-07-01

Immune complex (IC) deposition in the glomerular basement membrane (GBM) is a key early pathogenic event in lupus nephritis (LN). The clarification of the mechanisms behind IC deposition will enable targeted therapy in the future. Circulating cell-derived microparticles (MPs) have been proposed as major sources of extracellular autoantigens and ICs and triggers of autoimmunity in LN. The overabundance of galectin-3-binding protein (G3BP) along with immunoglobulins and a few other proteins specifically distinguish circulating MPs in patients with systemic lupus erythematosus (SLE), and this is most pronounced in patients with active LN. G3BP co-localizes with deposited ICs in renal biopsies from LN patients supporting a significant presence of MPs in the IC deposits. G3BP binds strongly to glomerular basement membrane proteins and integrins. Accordingly, MP surface proteins, especially G3BP, may be essential for the deposition of ICs in kidneys and thus for the ensuing formation of MP-derived electron dense structures in the GBM, and immune activation in LN. This review focuses on the notion of targeting surface molecules on MPs as an entirely novel treatment strategy in LN. By targeting MPs, a double hit may be achieved by attenuating both the autoantigenic fueling of immune complexes and the triggering of the adaptive immune system. Thereby, early pathogenic events may be blocked in contrast to current treatment strategies that primarily target and modulate later events in the cellular and humoral immune response. Copyright © 2016 Elsevier B.V. All rights reserved.

Heterogeneity in Oct4 and Sox2 Targets Biases Cell Fate in 4-Cell Mouse Embryos.

PubMed

Goolam, Mubeen; Scialdone, Antonio; Graham, Sarah J L; Macaulay, Iain C; Jedrusik, Agnieszka; Hupalowska, Anna; Voet, Thierry; Marioni, John C; Zernicka-Goetz, Magdalena

2016-03-24

The major and essential objective of pre-implantation development is to establish embryonic and extra-embryonic cell fates. To address when and how this fundamental process is initiated in mammals, we characterize transcriptomes of all individual cells throughout mouse pre-implantation development. This identifies targets of master pluripotency regulators Oct4 and Sox2 as being highly heterogeneously expressed between blastomeres of the 4-cell embryo, with Sox21 showing one of the most heterogeneous expression profiles. Live-cell tracking demonstrates that cells with decreased Sox21 yield more extra-embryonic than pluripotent progeny. Consistently, decreasing Sox21 results in premature upregulation of the differentiation regulator Cdx2, suggesting that Sox21 helps safeguard pluripotency. Furthermore, Sox21 is elevated following increased expression of the histone H3R26-methylase CARM1 and is lowered following CARM1 inhibition, indicating the importance of epigenetic regulation. Therefore, our results indicate that heterogeneous gene expression, as early as the 4-cell stage, initiates cell-fate decisions by modulating the balance of pluripotency and differentiation. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

Low MITF/AXL ratio predicts early resistance to multiple targeted drugs in melanoma.

PubMed

Müller, Judith; Krijgsman, Oscar; Tsoi, Jennifer; Robert, Lidia; Hugo, Willy; Song, Chunying; Kong, Xiangju; Possik, Patricia A; Cornelissen-Steijger, Paulien D M; Geukes Foppen, Marnix H; Kemper, Kristel; Goding, Colin R; McDermott, Ultan; Blank, Christian; Haanen, John; Graeber, Thomas G; Ribas, Antoni; Lo, Roger S; Peeper, Daniel S

2014-12-15

Increased expression of the Microphthalmia-associated transcription factor (MITF) contributes to melanoma progression and resistance to BRAF pathway inhibition. Here we show that the lack of MITF is associated with more severe resistance to a range of inhibitors, while its presence is required for robust drug responses. Both in primary and acquired resistance, MITF levels inversely correlate with the expression of several activated receptor tyrosine kinases, most frequently AXL. The MITF-low/AXL-high/drug-resistance phenotype is common among mutant BRAF and NRAS melanoma cell lines. The dichotomous behaviour of MITF in drug response is corroborated in vemurafenib-resistant biopsies, including MITF-high and -low clones in a relapsed patient. Furthermore, drug cocktails containing AXL inhibitor enhance melanoma cell elimination by BRAF or ERK inhibition. Our results demonstrate that a low MITF/AXL ratio predicts early resistance to multiple targeted drugs, and warrant clinical validation of AXL inhibitors to combat resistance of BRAF and NRAS mutant MITF-low melanomas.

MicroRNA-137 Affects Proliferation and Migration of Placenta Trophoblast Cells in Preeclampsia by Targeting ERRα.

PubMed

Lu, Tan-Min; Lu, Wei; Zhao, Long-Jun

2016-06-06

To investigate the effects of microRNA-137 (miRNA-137) in proliferation and migration of placenta trophoblast cells of preeclampsia and the targeting gene of miRNA-137. A total of 134 cases of puerperants were divided into normal pregnancy (n = 50), mild preeclampsia (n = 38), and severe preeclampsia groups (n = 46). MiRNA-137, estrogen-related receptor α (ERRα), and wingless INT (WNT)11 messenger RNAs (mRNAs) were measured in placental tissue and trophoblast cells after transfection, and ERRα protein in placental tissues was detected by immunohistochemistry. The target genes of miRNA-137, trophoblast cell proliferation, migration, and invasion abilities were detected. Both ERRα and WNT11 proteins in the trophoblast cells were measured after transfection. Relative expressions of miRNA-137 were higher, and positive expression rates and relative expression levels of ERRα protein were lower in mild and severe preeclampsia and early- and late-onset preeclampsia than in normal pregnancy group (all P < .05). MiRNA-137 in the placental tissues was negatively correlated with ERRα protein (P < .05). Luciferase reporter gene assay analysis showed that ERRα was a direct target gene of miRNA-137. Absorbance values, relative scratch-covered areas, cell membrane permeable rate, ERRα, and WNT11 mRNA and protein relative expressions were significantly lower, while cells at G1/G0 phase were higher in miRNA-137 mimic group than those in the blank, negative control, and miRNA-137 inhibitor group. MiRNA-137 significantly reduced the proliferation and migration of placenta trophoblast cells of preeclampsia by targeting ERRα, which might be a potential target for gene therapy. © The Author(s) 2016.

SERS detection and targeted ablation of lymphoma cells using functionalized Ag nanoparticles

NASA Astrophysics Data System (ADS)

Yao, Qian; Cao, Fei; Feng, Chao; Zhao, Yan; Wang, Xiuhong

2016-03-01

Lymphoma is a heterogeneous group of malignancies of the lymphoid tissue, and is prevalent worldwide affecting both children and adults with a high mortality rate. There is in dire need of accurate and noninvasive approaches for early detection of the disease. Herein, we report a facile way to fabricate silver nanoparticle based nanoprobe by incorporating the corner-stone immunotherapeutic drug Rituxan for simultaneous detection and ablation of lymphoma cells in vitro. The fabricated nanoprobe can detect CD20 positive single lymphoma cell by surface enhanced Raman scattering technique with high specificity. The engineered nanoprobe retains the same antibody property as intact drug via Antibody-Dependent Cell-mediated Cytotoxicity (ADCC) analysis. The nanoprobe efficiently eradicates lymphoma cells in vitro. By integrating the advantages of sensitive SERS detection with targeted ablation capabilities of immunotherapeutic drug through site specificity, this nanoprobe can be applied as outstanding tools in living imaging, cancer diagnosis and treatment.

Bruton tyrosine kinase inhibitors: a promising novel targeted treatment for B cell lymphomas

PubMed Central

Aalipour, Amin; Advani, Ranjana H.

2015-01-01

Summary Constitutive or aberrant signalling of the B cell receptor signalling cascade has been implicated in the propagation and maintenance of a variety of B cell malignancies. Small molecule inhibitors of Bruton tyrosine kinase (BTK), a protein early in this cascade and specifically expressed in B cells, have emerged as a new class of targeted agents. There are several BTK inhibitors, including ONO-WG-307, LFM-A13, dasatinib, CC-292, and PCI-32765 (ibrutinib), in preclinical and/or clinical development of which ibrutinib is currently in phase III trials. Recent clinical data suggest significant activity of ibrutinib as a first in class oral inhibitor of BTK. This review provides an overview of ongoing clinical studies of BTK inhibitors. PMID:24111579

Cell-permeable nanobodies for targeted immunolabelling and antigen manipulation in living cells

NASA Astrophysics Data System (ADS)

Herce, Henry D.; Schumacher, Dominik; Schneider, Anselm F. L.; Ludwig, Anne K.; Mann, Florian A.; Fillies, Marion; Kasper, Marc-André; Reinke, Stefan; Krause, Eberhard; Leonhardt, Heinrich; Cardoso, M. Cristina; Hackenberger, Christian P. R.

2017-08-01

Functional antibody delivery in living cells would enable the labelling and manipulation of intracellular antigens, which constitutes a long-thought goal in cell biology and medicine. Here we present a modular strategy to create functional cell-permeable nanobodies capable of targeted labelling and manipulation of intracellular antigens in living cells. The cell-permeable nanobodies are formed by the site-specific attachment of intracellularly stable (or cleavable) cyclic arginine-rich cell-penetrating peptides to camelid-derived single-chain VHH antibody fragments. We used this strategy for the non-endocytic delivery of two recombinant nanobodies into living cells, which enabled the relocalization of the polymerase clamp PCNA (proliferating cell nuclear antigen) and tumour suppressor p53 to the nucleolus, and thereby allowed the detection of protein-protein interactions that involve these two proteins in living cells. Furthermore, cell-permeable nanobodies permitted the co-transport of therapeutically relevant proteins, such as Mecp2, into the cells. This technology constitutes a major step in the labelling, delivery and targeted manipulation of intracellular antigens. Ultimately, this approach opens the door towards immunostaining in living cells and the expansion of immunotherapies to intracellular antigen targets.

Targeting c-KIT (CD117) by dasatinib and radotinib promotes acute myeloid leukemia cell death.

PubMed

Heo, Sook-Kyoung; Noh, Eui-Kyu; Kim, Jeong Yi; Jeong, Yoo Kyung; Jo, Jae-Cheol; Choi, Yunsuk; Koh, SuJin; Baek, Jin Ho; Min, Young Joo; Kim, Hawk

2017-11-10

Dasatinib and radotinib are oral BCR-ABL tyrosine kinase inhibitors that were developed as drugs for the treatment of chronic myeloid leukemia. We report here that the c-KIT (CD117) targeting with dasatinib and radotinib promotes acute myeloid leukemia (AML) cell death, and c-KIT endocytosis is essential for triggering c-KIT-positive AML cell death by dasatinib and radotinib during the early stages. In addition, dasatinib and radotinib reduce heat shock protein 90β (HSP90β) expression and release Apaf-1 in c-KIT-positive AML cells. Finally, this activates a caspase-dependent apoptotic pathway in c-KIT-positive AML cells. Moreover, the inhibition of c-KIT endocytosis by dynamin inhibitor (DY) reversed cell viability and c-KIT expression by dasatinib and radotinib. HSP90β expression was recovered by DY in c-KIT-positive AML cells as well. Furthermore, the effect of radotinib on c-KIT and HSP90β showed the same pattern in a xenograft animal model using HEL92.1.7 cells. Therefore, dasatinib and radotinib promote AML cell death by targeting c-KIT. Taken together, these results indicate that dasatinib and radotinib treatment have a potential role in anti-leukemic therapy on c-KIT-positive AML cells.

Induction of viral interference by IPNV-carrier cells on target cells: A cell co-culture study.

PubMed

Parreño, Ricardo; Torres, Susana; Almagro, Lucía; Belló-Pérez, Melissa; Estepa, Amparo; Perez, Luis

2016-11-01

IPNV is a salmonid birnavirus that possesses the ability to establish asymptomatic persistent infections in a number of valuable fish species. The presence of IPNV may interfere with subsequent infection by other viruses. In the present study we show that an IPNV-carrier cell line (EPC IPNV ) can induce an antiviral state in fresh EPC by co-cultivating both cell types in three different ways: a "droplet" culture system, a plastic chamber setup, and a transmembrane (Transwell ® ) system. All three cell co-culture methods were proven useful to study donor/target cell interaction. Naïve EPC cells grown in contact with EPC IPNV cells develop resistance to VHSV superinfection. The transmembrane system seems best suited to examine gene expression in donor and target cells separately. Our findings point to the conclusion that one or more soluble factors produced by the IPNV carrier culture induce the innate immune response within the target cells. This antiviral response is associated to the up-regulation of interferon (ifn) and mx gene expression in target EPC cells. To our knowledge this is the first article describing co-culture systems to study the interplay between virus-carrier cells and naive cells in fish. Copyright © 2016 The Author(s). Published by Elsevier Ltd.. All rights reserved.

Targeting Premalignant Lesions - Implications for Early Breast Cancer Detection and Intervention

DTIC Science & Technology

2017-04-01

lesions. Peptide conjugated AgNP were injected intravenously in mice and mammary glands were isolated and analyzed for nanoparticle accumulation by silver ...Furthermore, these probes will be used to develop targeted therapeutic nanoparticles for early intervention in breast cancer. 2. KEYWORDS...cancer (Months 18-24) (To be done) Specific Aim 3: Target premalignant lesions utilizing peptide-conjugated nanoparticles to prevent/delay

Low Doses of Gamma-Irradiation Induce an Early Bystander Effect in Zebrafish Cells Which Is Sufficient to Radioprotect Cells

PubMed Central

Pereira, Sandrine; Malard, Véronique; Ravanat, Jean-Luc; Davin, Anne-Hélène; Armengaud, Jean; Foray, Nicolas; Adam-Guillermin, Christelle

2014-01-01

The term “bystander effect” is used to describe an effect in which cells that have not been exposed to radiation are affected by irradiated cells though various intracellular signaling mechanisms. In this study we analyzed the kinetics and mechanisms of bystander effect and radioadaptation in embryonic zebrafish cells (ZF4) exposed to chronic low dose of gamma rays. ZF4 cells were irradiated for 4 hours with total doses of gamma irradiation ranging from 0.01–0.1 Gy. In two experimental conditions, the transfer of irradiated cells or culture medium from irradiated cells results in the occurrence of DNA double strand breaks in non-irradiated cells (assessed by the number of γ-H2AX foci) that are repaired at 24 hours post-irradiation whatever the dose. At low total irradiation doses the bystander effect observed does not affect DNA repair mechanisms in targeted and bystander cells. An increase in global methylation of ZF4 cells was observed in irradiated cells and bystander cells compared to control cells. We observed that pre-irradiated cells which are then irradiated for a second time with the same doses contained significantly less γ-H2AX foci than in 24 h gamma-irradiated control cells. We also showed that bystander cells that have been in contact with the pre-irradiated cells and then irradiated alone present less γ-H2AX foci compared to the control cells. This radioadaptation effect is significantly more pronounced at the highest doses. To determine the factors involved in the early events of the bystander effect, we performed an extensive comparative proteomic study of the ZF4 secretomes upon irradiation. In the experimental conditions assayed here, we showed that the early events of bystander effect are probably not due to the secretion of specific proteins neither the oxidation of these secreted proteins. These results suggest that early bystander effect may be due probably to a combination of multiple factors. PMID:24667817

Selection of Phage Display Peptides Targeting Human Pluripotent Stem Cell-Derived Progenitor Cell Lines.

PubMed

Bignone, Paola A; Krupa, Rachel A; West, Michael D; Larocca, David

2016-01-01

The ability of human pluripotent stem cells (hPS) to both self-renew and differentiate into virtually any cell type makes them a promising source of cells for cell-based regenerative therapies. However, stem cell identity, purity, and scalability remain formidable challenges that need to be overcome for translation of pluripotent stem cell research into clinical applications. Directed differentiation from hPS cells is inefficient and residual contamination with pluripotent cells that have the potential to form tumors remains problematic. The derivation of scalable (self-renewing) embryonic progenitor stem cell lines offers a solution because they are well defined and clonally pure. Clonally pure progenitor stem cell lines also provide a means for identifying cell surface targeting reagents that are useful for identification, tracking, and repeated derivation of the corresponding progenitor stem cell types from additional hPS cell sources. Such stem cell targeting reagents can then be applied to the manufacture of genetically diverse banks of human embryonic progenitor cell lines for drug screening, disease modeling, and cell therapy. Here we present methods to identify human embryonic progenitor stem cell targeting peptides by selection of phage display libraries on clonal embryonic progenitor cell lines and demonstrate their use for targeting quantum dots (Qdots) for stem cell labeling.

Integrative analysis reveals CD38 as a therapeutic target for plasma cell-rich pre-disease and established rheumatoid arthritis and systemic lupus erythematosus.

PubMed

Cole, Suzanne; Walsh, Alice; Yin, Xuefeng; Wechalekar, Mihir D; Smith, Malcolm D; Proudman, Susanna M; Veale, Douglas J; Fearon, Ursula; Pitzalis, Costantino; Humby, Frances; Bombardieri, Michele; Axel, Amy; Adams, Homer; Chiu, Christopher; Sharp, Michael; Alvarez, John; Anderson, Ian; Madakamutil, Loui; Nagpal, Sunil; Guo, Yanxia

2018-05-02

Plasmablasts and plasma cells play a key role in many autoimmune diseases, such as rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE). This study was undertaken to evaluate the potential of targeting CD38 as a plasma cell/plasmablast depletion mechanism by daratumumab in the treatment of patients with RA and SLE. RNA-sequencing analysis of synovial biopsies from various stages of RA disease progression, flow cytometry analysis of peripheral blood mononuclear cells (PBMC) from patients with RA or SLE and healthy donors, immunohistochemistry assessment (IHC) of synovial biopsies from patients with early RA, and ex vivo immune cell depletion assays using daratumumab (an anti-CD38 monoclonal antibody) were used to assess CD38 as a therapeutic target. We demonstrated that the plasma cell/plasmablast-related genes CD38, XBP1, IRF4, PRDM1, IGJ and TNFSF13B are significantly up-regulated in synovial biopsies from patients with arthralgia, undifferentiated arthritis (UA), early RA and established RA as compared to healthy controls and control patients with osteoarthritis. In addition, the highest CD38 expression was observed on plasma cells and plasmablasts compared to natural killer (NK) cells, classical dendritic cells (DCs), plasmacytoid DCs (pDCs) and T cells, in blood from healthy controls and patients with SLE and RA. Furthermore, IHC showed CD38 staining in the same region as CD3 and CD138 staining in synovial tissue biopsies from patients with early RA. Most importantly, our data show for the first time that daratumumab effectively depletes plasma cells/plasmablasts in PBMC from patients with SLE and RA in a dose-dependent manner ex vivo. These results indicate that CD38 may be a potential target for RA disease interception and daratumumab should be evaluated clinically for the treatment of both RA and SLE.

IgG1-iS18 impedes the adhesive and invasive potential of early and late stage malignant melanoma cells.

PubMed

Munien, Carmelle; Rebelo, Thalia M; Ferreira, Eloise; Weiss, Stefan F T

2017-02-15

The 37kDa/67kDa laminin receptor (LRP/LR) is a non-integrin laminin receptor which is overexpressed in tumorigenic cells and supports progression of cancer via promoting metastasis, angiogenesis and telomerase activity and impediment of apoptosis. The present study investigates the role of LRP/LR on the metastatic potential of early (A375) and late (A375SM) stage malignant melanoma cells. Flow cytometry revealed that both early and late stage malignant melanoma cells display high levels of LRP/LR on their cell surface. Flow cytometry and western blot analysis showed that late stage malignant melanoma cells display significantly higher total and cell surface LRP/LR levels in comparison to early stage malignant melanoma cells and the poorly invasive breast cancer (MCF-7) control cell line. Targeting LRP/LR using the LRP/LR specific antibody IgG1-iS18 resulted in a significant reduction of the adhesive potential to laminin-1 and the invasive potential through the 'ECM-simulating' Matrigel™ of both early and late stage malignant melanoma cells. Furthermore, Pearson's correlation coefficient confirmed that increased LRP levels correlate with the increased invasive and adhesive potential in early and late stage melanoma cells. Thus, blocking LRP/LR using the IgG1-iS18 antibody may therefore be a promising therapeutic strategy for early and late stage malignant melanoma treatment. Copyright © 2017 Elsevier Inc. All rights reserved.

The Quest for Targets Executing MYC-Dependent Cell Transformation.

PubMed

Hartl, Markus

2016-01-01

MYC represents a transcription factor with oncogenic potential converting multiple cellular signals into a broad transcriptional response, thereby controlling the expression of numerous protein-coding and non-coding RNAs important for cell proliferation, metabolism, differentiation, and apoptosis. Constitutive activation of MYC leads to neoplastic cell transformation, and deregulated MYC alleles are frequently observed in many human cancer cell types. Multiple approaches have been performed to isolate genes differentially expressed in cells containing aberrantly activated MYC proteins leading to the identification of thousands of putative targets. Functional analyses of genes differentially expressed in MYC-transformed cells had revealed that so far more than 40 upregulated or downregulated MYC targets are actively involved in cell transformation or tumorigenesis. However, further systematic and selective approaches are required for determination of the known or yet unidentified targets responsible for processing the oncogenic MYC program. The search for critical targets in MYC-dependent tumor cells is exacerbated by the fact that during tumor development, cancer cells progressively evolve in a multistep process, thereby acquiring their characteristic features in an additive manner. Functional expression cloning, combinatorial gene expression, and appropriate in vivo tests could represent adequate tools for dissecting the complex scenario of MYC-specified cell transformation. In this context, the central goal is to identify a minimal set of targets that suffices to phenocopy oncogenic MYC. Recently developed genomic editing tools could be employed to confirm the requirement of crucial transformation-associated targets. Knowledge about essential MYC-regulated genes is beneficial to expedite the development of specific inhibitors to interfere with growth and viability of human tumor cells in which MYC is aberrantly activated. Approaches based on the principle of

The Quest for Targets Executing MYC-Dependent Cell Transformation

PubMed Central

Hartl, Markus

2016-01-01

MYC represents a transcription factor with oncogenic potential converting multiple cellular signals into a broad transcriptional response, thereby controlling the expression of numerous protein-coding and non-coding RNAs important for cell proliferation, metabolism, differentiation, and apoptosis. Constitutive activation of MYC leads to neoplastic cell transformation, and deregulated MYC alleles are frequently observed in many human cancer cell types. Multiple approaches have been performed to isolate genes differentially expressed in cells containing aberrantly activated MYC proteins leading to the identification of thousands of putative targets. Functional analyses of genes differentially expressed in MYC-transformed cells had revealed that so far more than 40 upregulated or downregulated MYC targets are actively involved in cell transformation or tumorigenesis. However, further systematic and selective approaches are required for determination of the known or yet unidentified targets responsible for processing the oncogenic MYC program. The search for critical targets in MYC-dependent tumor cells is exacerbated by the fact that during tumor development, cancer cells progressively evolve in a multistep process, thereby acquiring their characteristic features in an additive manner. Functional expression cloning, combinatorial gene expression, and appropriate in vivo tests could represent adequate tools for dissecting the complex scenario of MYC-specified cell transformation. In this context, the central goal is to identify a minimal set of targets that suffices to phenocopy oncogenic MYC. Recently developed genomic editing tools could be employed to confirm the requirement of crucial transformation-associated targets. Knowledge about essential MYC-regulated genes is beneficial to expedite the development of specific inhibitors to interfere with growth and viability of human tumor cells in which MYC is aberrantly activated. Approaches based on the principle of

Targeting human breast cancer cells by an oncolytic adenovirus using microRNA-targeting strategy.

PubMed

Shayestehpour, Mohammad; Moghim, Sharareh; Salimi, Vahid; Jalilvand, Somayeh; Yavarian, Jila; Romani, Bizhan; Mokhtari-Azad, Talat

2017-08-15

MicroRNA-targeting strategy is a promising approach that enables oncolytic viruses to replicate in tumor cells but not in normal cells. In this study, we targeted adenoviral replication toward breast cancer cells by inserting ten complementary binding sites for miR-145-5p downstream of E1A gene. In addition, we evaluated the effect of increasing miR-145 binding sites on inhibition of virus replication. Ad5-control and adenoviruses carrying five or ten copies of miR145-5p target sites (Ad5-5miR145T, Ad5-10miR145T) were generated and inoculated into MDA-MB-453, BT-20, MCF-7 breast cancer cell lines and human mammary epithelial cells (HMEpC). Titer of Ad5-10miR145T in HMEpC was significantly lower than Ad5-control titer. Difference between the titer of these two viruses at 12, 24, 36, and 48h after infection was 1.25, 2.96, 3.06, and 3.77 log TCID 50 . No significant difference was observed between the titer of both adenoviruses in MDA-MB-453, BT-20 and MCF-7 cells. The infectious titer of adenovirus containing 10 miR-145 binding sites in HMEpC cells at 24, 36, and 48h post-infection was 1.7, 2.08, and 4-fold, respectively, lower than the titer of adenovirus carrying 5 miR-145 targets. Our results suggest that miR-145-targeting strategy provides selectivity for adenovirus replication in breast cancer cells. Increasing the number of miRNA binding sites within the adenoviral genome confers more selectivity for viral replication in cancer cells. Copyright © 2017. Published by Elsevier B.V.

Cell cycle-tailored targeting of metastatic melanoma: Challenges and opportunities.

PubMed

Haass, Nikolas K; Gabrielli, Brian

2017-07-01

The advent of targeted therapies of metastatic melanoma, such as MAPK pathway inhibitors and immune checkpoint antagonists, has turned dermato-oncology from the "bad guy" to the "poster child" in oncology. Current targeted therapies are effective, although here is a clear need to develop combination therapies to delay the onset of resistance. Many antimelanoma drugs impact on the cell cycle but are also dependent on certain cell cycle phases resulting in cell cycle phase-specific drug insensitivity. Here, we raise the question: Have combination trials been abandoned prematurely as ineffective possibly only because drug scheduling was not optimized? Firstly, if both drugs of a combination hit targets in the same melanoma cell, cell cycle-mediated drug insensitivity should be taken into account when planning combination therapies, timing of dosing schedules and choice of drug therapies in solid tumors. Secondly, if the combination is designed to target different tumor cell subpopulations of a heterogeneous tumor, one drug effective in a particular subpopulation should not negatively impact on the other drug targeting another subpopulation. In addition to the role of cell cycle stage and progression on standard chemotherapeutics and targeted drugs, we discuss the utilization of cell cycle checkpoint control defects to enhance chemotherapeutic responses or as targets themselves. We propose that cell cycle-tailored targeting of metastatic melanoma could further improve therapy outcomes and that our real-time cell cycle imaging 3D melanoma spheroid model could be utilized as a tool to measure and design drug scheduling approaches. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Sonoporation of endothelial cells by vibrating targeted microbubbles.

PubMed

Kooiman, Klazina; Foppen-Harteveld, Miranda; van der Steen, Antonius F W; de Jong, Nico

2011-08-25

Molecular imaging using ultrasound makes use of targeted microbubbles. In this study we investigated whether these microbubbles could also be used to induce sonoporation in endothelial cells. Lipid-coated microbubbles were targeted to CD31 and insonified at 1 MHz at low peak negative acoustic pressures at six sequences of 10 cycle sine-wave bursts. Vibration of the targeted microbubbles was recorded with the Brandaris-128 high-speed camera (~13 million frames per second). In total, 31 cells were studied that all had one microbubble (1.2-4.2 micron in diameter) attached per cell. After insonification at 80 kPa, 30% of the cells (n=6) had taken up propidium iodide, while this was 20% (n=1) at 120 kPa and 83% (n=5) at 200 kPa. Irrespective of the peak negative acoustic pressure, uptake of propidium iodide was observed when the relative vibration amplitude of targeted microbubbles was greater than 0.5. No relationship was found between the position of the microbubble on the cell and induction of sonoporation. This study shows that targeted microbubbles can also be used to induce sonoporation, thus making it possible to combine molecular imaging and drug delivery. Copyright © 2011 Elsevier B.V. All rights reserved.

Enhanced cellular uptake of LHRH-conjugated PEG-coated magnetite nanoparticles for specific targeting of triple negative breast cancer cells.

PubMed

Hu, J; Obayemi, J D; Malatesta, K; Košmrlj, A; Soboyejo, W O

2018-07-01

Targeted therapy is an emerging technique in cancer detection and treatment. This paper presents the results of a combined experimental and theoretical study of the specific targeting and entry of luteinizing hormone releasing hormone (LHRH)-conjugated PEG-coated magnetite nanoparticles into triple negative breast cancer (TNBC) cells and normal breast cells. The conjugated nanoparticles structures, cellular uptake of PEG-coated magnetite nanoparticles (MNPs) and LHRH-conjugated PEG-coated magnetite nanoparticles (LHRH-MNPs) into breast cancer cells and normal breast cells were investigated using a combination of transmission electron microscope, optical and confocal fluorescence microscopy techniques. The results show that the presence of LHRH enhances the uptake of LHRH-MNPs into TNBC cells. Nanoparticle entry into breast cancer cells is also studied using a combination of thermodynamics and kinetics models. The trends in the predicted nanoparticle entry times (into TNBC cells) and the size ranges of the engulfed nanoparticles (within the TNBC cells) are shown to be consistent with experimental observations. The implications of the results are then discussed for the specific targeting of TNBCs with LHRH-conjugated PEG-coated magnetite nanoparticles for the early detection and treatment of TNBC. Copyright © 2018. Published by Elsevier B.V.

Targeted silver nanoparticles for ratiometric cell phenotyping

NASA Astrophysics Data System (ADS)

Willmore, Anne-Mari A.; Simón-Gracia, Lorena; Toome, Kadri; Paiste, Päärn; Kotamraju, Venkata Ramana; Mölder, Tarmo; Sugahara, Kazuki N.; Ruoslahti, Erkki; Braun, Gary B.; Teesalu, Tambet

2016-04-01

Affinity targeting is used to deliver nanoparticles to cells and tissues. For efficient targeting, it is critical to consider the expression and accessibility of the relevant receptors in the target cells. Here, we describe isotopically barcoded silver nanoparticles (AgNPs) as a tool for auditing affinity ligand receptors in cells. Tumor penetrating peptide RPARPAR (receptor: NRP-1) and tumor homing peptide GKRK (receptor: p32) were used as affinity ligands on the AgNPs. The binding and uptake of the peptide-functionalized AgNPs by cultured PPC-1 prostate cancer and M21 melanoma cells was dependent on the cell surface expression of the cognate peptide receptors. Barcoded peptide-functionalized AgNPs were synthesized from silver and palladium isotopes. The cells were incubated with a cocktail of the barcoded nanoparticles [RPARPAR (R), GKRK (K), and control], and cellular binding and internalization of each type of nanoparticle was assessed by inductively coupled plasma mass spectrometry. The results of isotopic analysis were in agreement with data obtained using optical methods. Using ratiometric measurements, we were able to classify the PPC-1 cell line as mainly NRP-1-positive, with 75 +/- 5% R-AgNP uptake, and the M21 cell line as only p32-positive, with 89 +/- 9% K-AgNP uptake. The isotopically barcoded multiplexed AgNPs are useful as an in vitro ratiometric phenotyping tool and have potential uses in functional evaluation of the expression of accessible homing peptide receptors in vivo.Affinity targeting is used to deliver nanoparticles to cells and tissues. For efficient targeting, it is critical to consider the expression and accessibility of the relevant receptors in the target cells. Here, we describe isotopically barcoded silver nanoparticles (AgNPs) as a tool for auditing affinity ligand receptors in cells. Tumor penetrating peptide RPARPAR (receptor: NRP-1) and tumor homing peptide GKRK (receptor: p32) were used as affinity ligands on the AgNPs. The

Dendritic Cells Program Non-Immunogenic Prostate-Specific T Cell Responses Beginning at Early Stages of Prostate Tumorigenesis

PubMed Central

Mihalyo, Marianne A.; Hagymasi, Adam T.; Slaiby, Aaron M.; Nevius, Erin E.; Adler, Adam J.

2010-01-01

BACKGROUND Prostate cancer promotes the development of T cell tolerance towards prostatic antigens, potentially limiting the efficacy of prostate cancer vaccines targeting these antigens. Here, we sought to determine the stage of disease progression when T cell tolerance develops, as well as the role of steady state dendritic cells (DC) and CD4+CD25+ T regulatory cells (Tregs) in programming tolerance. METHODS The response of naïve HA-specific CD4+ T cells were analyzed following adoptive transfer into Pro-HA × TRAMP transgenic mice harboring variably-staged HA-expressing prostate tumors on two genetic backgrounds that display different patterns and kinetics of tumorigenesis. The role of DC and Tregs in programming HA-specific CD4 cell responses were assessed via depletion. RESULTS HA-specific CD4 cells underwent non-immunogenic responses at all stages of tumorigenesis in both genetic backgrounds. These responses were completely dependent on DC, but not appreciably influenced by Tregs. CONCLUSIONS These results suggest that tolerogenicity is an early and general property of prostate tumors. PMID:17221844

A novel double-targeted nondrug delivery system for targeting cancer stem cells

PubMed Central

Qiao, Shupei; Zhao, Yufang; Geng, Shuai; Li, Yong; Hou, Xiaolu; Liu, Yi; Lin, Feng-Huei; Yao, Lifen; Tian, Weiming

2016-01-01

Instead of killing cancer stem cells (CSCs), the conventional chemotherapy used for cancer treatment promotes the enrichment of CSCs, which are responsible for tumor growth, metastasis, and recurrence. However, most therapeutic agents are only able to kill a small proportion of CSCs by targeting one or two cell surface markers or dysregulated CSC pathways, which are usually shared with normal stem cells (NSCs). In this study, we developed a novel nondrug delivery system for the dual targeting of CSCs by conjugating hyaluronic acid (HA) and grafting the doublecortin-like kinase 1 (DCLK1) monoclonal antibody to the surface of poly(ethylene glycol) (PEG)–poly(d,l-lactide-co-glycolide) (PLGA) nanoparticles (NPs), which can specifically target CD44 receptors and the DCLK1 surface marker – the latter was shown to possess the capacity to distinguish between CSCSs and NSCs. The size and morphology of these NPs were characterized by dynamic light scattering (DLS), transmission electron microscopy (TEM), and scanning electron microscopy (SEM). This was followed by studies of NP encapsulation efficiency and in vitro drug release properties. Then, the cytotoxicity of the NPs was tested via Cell Counting Kit-8 assay. Finally, the 4T1 CSCs were obtained from the alginate-based platform, which we developed as an in vitro tumor model. Tumor-bearing nude mice were used as in vivo models to systematically detect the ability of NPs to target CSCs. Our results showed that the DCLK1–HA–PEG–PLGA NPs exhibited a targeting effect toward CSCs both in vitro and in vivo. These findings have important implications for the rational design of drug delivery systems that target CSCs with high efficacy. PMID:27994463

Advanced cell therapies: targeting, tracking and actuation of cells with magnetic particles.

PubMed

Connell, John J; Patrick, P Stephen; Yu, Yichao; Lythgoe, Mark F; Kalber, Tammy L

2015-01-01

Regenerative medicine would greatly benefit from a new platform technology that enabled measurable, controllable and targeting of stem cells to a site of disease or injury in the body. Superparamagnetic iron-oxide nanoparticles offer attractive possibilities in biomedicine and can be incorporated into cells, affording a safe and reliable means of tagging. This review describes three current and emerging methods to enhance regenerative medicine using magnetic particles to guide therapeutic cells to a target organ; track the cells using MRI and assess their spatial localization with high precision and influence the behavior of the cell using magnetic actuation. This approach is complementary to the systemic injection of cell therapies, thus expanding the horizon of stem cell therapeutics.

Self-targeted salinomycin-loaded DSPE-PEG-methotrexate nanomicelles for targeting both head and neck squamous cell carcinoma cancer cells and cancer stem cells.

PubMed

Zhu, Minhui; Chen, Shicai; Hua, Libo; Zhang, Caiyun; Chen, Mengjie; Chen, Donghui; Dong, Yinmei; Zhang, Yingying; Li, Meng; Song, Xianmin; Chen, Huaiwen; Zheng, Hongliang

2017-02-01

To target both head and neck squamous cell carcinoma (HNSCC) cells and cancer stem cells (CSCs) by salinomycin-loaded DSPE-PEG-MTX (synthesized using DSPE-PEG2000-NH2 and methotrexate) nanomicelles (M-SAL-MTX). The characterization, antitumor activity and mechanism of M-SAL-MTX were evaluated. M-SAL-MTX showed enhanced inhibitory effect toward both HNSCC CSCs and non-CSCs compared with a single treatment of methotrexate and salinomycin. In nude mice-bearing HNSCC xenografts, M-SAL-MTX suppressed tumor growth more effectively than other controls including combination of methotrexate and salinomycin. Therefore, M-SAL-MTX may provide a strategy for treating HNSCC by targeting both HNSCC CSCs and HNSCC cells.

An innovative pre-targeting strategy for tumor cell specific imaging and therapy

NASA Astrophysics Data System (ADS)

Qin, Si-Yong; Peng, Meng-Yun; Rong, Lei; Jia, Hui-Zhen; Chen, Si; Cheng, Si-Xue; Feng, Jun; Zhang, Xian-Zheng

2015-08-01

A programmed pre-targeting system for tumor cell imaging and targeting therapy was established based on the ``biotin-avidin'' interaction. In this programmed functional system, transferrin-biotin can be actively captured by tumor cells with the overexpression of transferrin receptors, thus achieving the pre-targeting modality. Depending upon avidin-biotin recognition, the attachment of multivalent FITC-avidin to biotinylated tumor cells not only offered the rapid fluorescence labelling, but also endowed the pre-targeted cells with targeting sites for the specifically designed biotinylated peptide nano-drug. Owing to the successful pre-targeting, tumorous HepG2 and HeLa cells were effectively distinguished from the normal 3T3 cells via fluorescence imaging. In addition, the self-assembled peptide nano-drug resulted in enhanced cell apoptosis in the observed HepG2 cells. The tumor cell specific pre-targeting strategy is applicable for a variety of different imaging and therapeutic agents for tumor treatments.A programmed pre-targeting system for tumor cell imaging and targeting therapy was established based on the ``biotin-avidin'' interaction. In this programmed functional system, transferrin-biotin can be actively captured by tumor cells with the overexpression of transferrin receptors, thus achieving the pre-targeting modality. Depending upon avidin-biotin recognition, the attachment of multivalent FITC-avidin to biotinylated tumor cells not only offered the rapid fluorescence labelling, but also endowed the pre-targeted cells with targeting sites for the specifically designed biotinylated peptide nano-drug. Owing to the successful pre-targeting, tumorous HepG2 and HeLa cells were effectively distinguished from the normal 3T3 cells via fluorescence imaging. In addition, the self-assembled peptide nano-drug resulted in enhanced cell apoptosis in the observed HepG2 cells. The tumor cell specific pre-targeting strategy is applicable for a variety of different imaging

Non-targeted plasma metabolome of early and late lactation gilts

USDA-ARS?s Scientific Manuscript database

Female pigs nursing their first litter (first-parity gilts) have increased energy requirements not only to support their piglets, but they themselves are still maturing. Non-targeted plasma metabolomics were used to investigate the differences between (1) post-farrowing and weaning (early or late l...

Stromal cells in breast cancer as a potential therapeutic target

PubMed Central

Dykes, Samantha S.; Hughes, Veronica S.; Wiggins, Jennifer M.; Fasanya, Henrietta O.; Tanaka, Mai; Siemann, Dietmar

2018-01-01

Breast cancer in the United States is the second most commonly diagnosed cancer in women. About 1 in 8 women will develop invasive breast cancer over the course of her lifetime and breast cancer remains the second leading cause of cancer-related death. In pursuit of novel therapeutic strategies, researchers have examined the tumor microenvironment as a potential anti-cancer target. In addition to neoplastic cells, the tumor microenvironment is composed of several critical normal cell types, including fibroblasts, vascular and lymph endothelial cells, osteoclasts, adipocytes, and immune cells. These cells have important roles in healthy tissue stasis, which frequently are altered in tumors. Indeed, tumor-associated stromal cells often contribute to tumorigenesis, tumor progression, and metastasis. Consequently, these host cells may serve as a possible target in anti-tumor and anti-metastatic therapeutic strategies. Targeting the tumor associated host cells offers the benefit that such cells do not mutate and develop resistance in response to treatment, a major cause of failure in cancer therapeutics targeting neoplastic cells. This review discusses the role of host cells in the tumor microenvironment during tumorigenesis, progression, and metastasis, and provides an overview of recent developments in targeting these cell populations to enhance cancer therapy efficacy.

Mast cell proteases as pharmacological targets

PubMed Central

Caughey, George H.

2015-01-01

Mast cells are rich in proteases, which are the major proteins of intracellular granules and are released with histamine and heparin by activated cells. Most of these proteases are active in the granule as well outside of the mast cell when secreted, and can cleave targets near degranulating mast cells and in adjoining tissue compartments. Some proteases released from mast cells reach the bloodstream and may have far-reaching actions. In terms of relative amounts, the major mast cell proteases include the tryptases, chymases, cathepsin G, carboxypeptidase A3, dipeptidylpeptidase I/cathepsin C, and cathepsins L and S. Some mast cells also produce granzyme B, plasminogen activators, and matrix metalloproteinases. Tryptases and chymases are almost entirely mast cell-specific, whereas other proteases, such as cathepsins G, C, and L are expressed by a variety of inflammatory cells. Carboxypeptidase A3 expression is a property shared by basophils and mast cells. Other proteases, such as mastins, are largely basophil-specific, although human basophils are protease-deficient compared with their murine counterparts. The major classes of mast cell proteases have been targeted for development of therapeutic inhibitors. Also, a human β-tryptase has been proposed as a potential drug itself, to inactivate of snake venins. Diseases linked to mast cell proteases include allergic diseases, such as asthma, eczema, and anaphylaxis, but also include non-allergic diseases such inflammatory bowel disease, autoimmune arthritis, atherosclerosis, aortic aneurysms, hypertension, myocardial infarction, heart failure, pulmonary hypertension and scarring diseases of lungs and other organs. In some cases, studies performed in mouse models suggest protective or homeostatic roles for specific proteases (or groups of proteases) in infections by bacteria, worms and other parasites, and even in allergic inflammation. At the same time, a clearer picture has emerged of differences in the properties

Targeting Stromal Recruitment by Prostate Cancer Cells

DTIC Science & Technology

2006-03-01

Ensinger, C., Tumer , Z., Tommerup, N. et al.: Hedgehog signaling in small-cell lung cancer : frequent in vivo but a rare event in vitro. Lung Cancer , 52...W81XWH-04-1-0157 TITLE: Targeting Stromal Recruitment by Prostate Cancer Cells PRINCIPAL INVESTIGATOR: Jingxian Zhang, Ph.D...DATES COVERED (From - To) 15 Feb 2004 – 14 Feb 2006 4. TITLE AND SUBTITLE 5a. CONTRACT NUMBER Targeting Stromal Recruitment by Prostate Cancer

Magnetic stem cell targeting to the inner ear

NASA Astrophysics Data System (ADS)

Le, T. N.; Straatman, L.; Yanai, A.; Rahmanian, R.; Garnis, C.; Häfeli, U. O.; Poblete, T.; Westerberg, B. D.; Gregory-Evans, K.

2017-12-01

Severe sensorineural deafness is often accompanied by a loss of auditory neurons in addition to injury of the cochlear epithelium and hair cell loss. Cochlear implant function however depends on a healthy complement of neurons and their preservation is vital in achieving optimal results. We have developed a technique to target mesenchymal stem cells (MSCs) to a deafened rat cochlea. We then assessed the neuroprotective effect of systematically delivered MSCs on the survival and function of spiral ganglion neurons (SGNs). MSCs were labeled with superparamagnetic nanoparticles, injected via the systemic circulation, and targeted using a magnetized cochlea implant and external magnet. Neurotrophic factor concentrations, survival of SGNs, and auditory function were assessed at 1 week and 4 weeks after treatments and compared against multiple control groups. Significant numbers of magnetically targeted MSCs (>30 MSCs/section) were present in the cochlea with accompanied elevation of brain-derived neurotrophic factor and glial cell-derived neurotrophic factor levels (p < 0.001). In addition we saw improved survival of SGNs (approximately 80% survival at 4 weeks). Hearing threshold levels in magnetically targeted rats were found to be significantly better than those of control rats (p < 0.05). These results indicate that magnetic targeting of MSCs to the cochlea can be accomplished with a magnetized cochlear permalloy implant and an external magnet. The targeted stem cells release neurotrophic factors which results in improved SGN survival and hearing recovery. Combining magnetic cell-based therapy and cochlear implantation may improve cochlear implant function in treating deafness.

Surface-modified gold nanorods for specific cell targeting

NASA Astrophysics Data System (ADS)

Wang, Chan-Ung; Arai, Yoshie; Kim, Insun; Jang, Wonhee; Lee, Seonghyun; Hafner, Jason H.; Jeoung, Eunhee; Jung, Deokho; Kwon, Youngeun

2012-05-01

Gold nanoparticles (GNPs) have unique properties that make them highly attractive materials for developing functional reagents for various biomedical applications including photothermal therapy, targeted drug delivery, and molecular imaging. For in vivo applications, GNPs need to be prepared with very little or negligible cytotoxicitiy. Most GNPs are, however, prepared using growth-directing surfactants such as cetyl trimethylammonium bromide (CTAB), which are known to have considerable cytotoxicity. In this paper, we describe an approach to remove CTAB to a non-toxic concentration. We optimized the conditions for surface modification with methoxypolyethylene glycol thiol (mPEG), which replaced CTAB and formed a protective layer on the surface of gold nanorods (GNRs). The cytotoxicities of pristine and surface-modified GNRs were measured in primary human umbilical vein endothelial cells and human cell lines derived from hepatic carcinoma cells, embryonic kidney cells, and thyroid papillary carcinoma cells. Cytotoxicity assays revealed that treating cells with GNRs did not significantly affect cell viability except for thyroid papillary carcinoma cells. Thyroid cancer cells were more susceptible to residual CTAB, so CTAB had to be further removed by dialysis in order to use GNRs for thyroid cell targeting. PEGylated GNRs are further modified to present monoclonal antibodies that recognize a specific surface marker, Na-I symporter, for thyroid cells. Antibody-conjugated GNRs specifically targeted human thyroid cells in vitro.

Chimeric antigen receptor T cells targeting Fc μ receptor selectively eliminate CLL cells while sparing healthy B cells.

PubMed

Faitschuk, Elena; Hombach, Andreas A; Frenzel, Lukas P; Wendtner, Clemens-Martin; Abken, Hinrich

2016-09-29

Adoptive cell therapy of chronic lymphocytic leukemia (CLL) with chimeric antigen receptor (CAR)-modified T cells targeting CD19 induced lasting remission of this refractory disease in a number of patients. However, the treatment is associated with prolonged "on-target off-tumor" toxicities due to the targeted elimination of healthy B cells demanding more selectivity in targeting CLL cells. We identified the immunoglobulin M Fc receptor (FcμR), also known as the Fas apoptotic inhibitory molecule-3 or TOSO, as a target for a more selective treatment of CLL by CAR T cells. FcμR is highly and consistently expressed by CLL cells; only minor levels are detected on healthy B cells or other hematopoietic cells. T cells with a CAR specific for FcμR efficiently responded toward CLL cells, released a panel of proinflammatory cytokines and lytic factors, like soluble FasL and granzyme B, and eliminated the leukemic cells. In contrast to CD19 CAR T cells, anti-FcμR CAR T cells did not attack healthy B cells. T cells with anti-FcμR CAR delayed outgrowth of Mec-1-induced leukemia in a xenograft mouse model. T cells from CLL patients in various stages of the disease, modified by the anti-FcμR CAR, purged their autologous CLL cells in vitro without reducing the number of healthy B cells, which is the case with anti-CD19 CAR T cells. Compared with the currently used therapies, the data strongly imply a superior therapeutic index of anti-FcμR CAR T cells for the treatment of CLL. © 2016 by The American Society of Hematology.

Identifying the Target Cell in Primary Simian Immunodeficiency Virus (SIV) Infection: Highly Activated Memory CD4+ T Cells Are Rapidly Eliminated in Early SIV Infection In Vivo

PubMed Central

Veazey, Ronald S.; Tham, Irene C.; Mansfield, Keith G.; DeMaria, MaryAnn; Forand, Amy E.; Shvetz, Daniel E.; Chalifoux, Laura V.; Sehgal, Prabhat K.; Lackner, Andrew A.

2000-01-01

It has recently been shown that rapid and profound CD4+ T-cell depletion occurs almost exclusively within the intestinal tract of simian immunodeficiency virus (SIV)-infected macaques within days of infection. Here we demonstrate (by three- and four-color flow cytometry) that this depletion is specific to a definable subset of CD4+ T cells, namely, those having both a highly and/or acutely activated (CD69+ CD38+ HLA-DR+) and memory (CD45RA− Leu8−) phenotype. Moreover, we demonstrate that this subset of helper T cells is found primarily within the intestinal lamina propria. Viral tropism for this particular cell type (which has been previously suggested by various studies in vitro) could explain why profound CD4+ T-cell depletion occurs in the intestine and not in peripheral lymphoid tissues in early SIV infection. Furthermore, we demonstrate that an acute loss of this specific subset of activated memory CD4+ T cells may also be detected in peripheral blood and lymph nodes in early SIV infection. However, since this particular cell type is present in such small numbers in circulation, its loss does not significantly affect total CD4+ T cell counts. This finding suggests that SIV and, presumably, human immunodeficiency virus specifically infect, replicate in, and eliminate definable subsets of CD4+ T cells in vivo. PMID:10590091

Surface-modified CMOS IC electrochemical sensor array targeting single chromaffin cells for highly parallel amperometry measurements.

PubMed

Huang, Meng; Delacruz, Joannalyn B; Ruelas, John C; Rathore, Shailendra S; Lindau, Manfred

2018-01-01

Amperometry is a powerful method to record quantal release events from chromaffin cells and is widely used to assess how specific drugs modify quantal size, kinetics of release, and early fusion pore properties. Surface-modified CMOS-based electrochemical sensor arrays allow simultaneous recordings from multiple cells. A reliable, low-cost technique is presented here for efficient targeting of single cells specifically to the electrode sites. An SU-8 microwell structure is patterned on the chip surface to provide insulation for the circuitry as well as cell trapping at the electrode sites. A shifted electrode design is also incorporated to increase the flexibility of the dimension and shape of the microwells. The sensitivity of the electrodes is validated by a dopamine injection experiment. Microwells with dimensions slightly larger than the cells to be trapped ensure excellent single-cell targeting efficiency, increasing the reliability and efficiency for on-chip single-cell amperometry measurements. The surface-modified device was validated with parallel recordings of live chromaffin cells trapped in the microwells. Rapid amperometric spikes with no diffusional broadening were observed, indicating that the trapped and recorded cells were in very close contact with the electrodes. The live cell recording confirms in a single experiment that spike parameters vary significantly from cell to cell but the large number of cells recorded simultaneously provides the statistical significance.

Methods for Surgical Targeting of the STN in Early-Stage Parkinson’s Disease

PubMed Central

Camalier, Corrie R.; Konrad, Peter E.; Gill, Chandler E.; Kao, Chris; Remple, Michael R.; Nasr, Hana M.; Davis, Thomas L.; Hedera, Peter; Phibbs, Fenna T.; Molinari, Anna L.; Neimat, Joseph S.; Charles, David

2013-01-01

Patients with Parkinson’s disease (PD) experience progressive neurological decline, and future interventional therapies are thought to show most promise in early stages of the disease. There is much interest in therapies that target the subthalamic nucleus (STN) with surgical access. While locating STN in advanced disease patients (Hoehn–Yahr Stage III or IV) is well understood and routinely performed at many centers in the context of deep brain stimulation surgery, the ability to identify this nucleus in early-stage patients has not previously been explored in a sizeable cohort. We report surgical methods used to target the STN in 15 patients with early PD (Hoehn–Yahr Stage II), using a combination of image guided surgery, microelectrode recordings, and clinical responses to macrostimulation of the region surrounding the STN. Measures of electrophysiology (firing rates and root mean squared activity) have previously been found to be lower than in later-stage patients, however, the patterns of electrophysiology seen and dopamimetic macrostimulation effects are qualitatively similar to those seen in advanced stages. Our experience with surgical implantation of Parkinson’s patients with minimal motor symptoms suggest that it remains possible to accurately target the STN in early-stage PD using traditional methods. PMID:24678307

Myoepithelial cell-specific expression of stefin A as a suppressor of early breast cancer invasion.

PubMed

Duivenvoorden, Hendrika M; Rautela, Jai; Edgington-Mitchell, Laura E; Spurling, Alex; Greening, David W; Nowell, Cameron J; Molloy, Timothy J; Robbins, Elizabeth; Brockwell, Natasha K; Lee, Cheok Soon; Chen, Maoshan; Holliday, Anne; Selinger, Cristina I; Hu, Min; Britt, Kara L; Stroud, David A; Bogyo, Matthew; Möller, Andreas; Polyak, Kornelia; Sloane, Bonnie F; O'Toole, Sandra A; Parker, Belinda S

2017-12-01

Mammography screening has increased the detection of early pre-invasive breast cancers, termed ductal carcinoma in situ (DCIS), increasing the urgency of identifying molecular regulators of invasion as prognostic markers to predict local relapse. Using the MMTV-PyMT breast cancer model and pharmacological protease inhibitors, we reveal that cysteine cathepsins have important roles in early-stage tumorigenesis. To characterize the cell-specific roles of cathepsins in early invasion, we developed a DCIS-like model, incorporating an immortalized myoepithelial cell line (N1ME) that restrained tumor cell invasion in 3D culture. Using this model, we identified an important myoepithelial-specific function of the cysteine cathepsin inhibitor stefin A in suppressing invasion, whereby targeted stefin A loss in N1ME cells blocked myoepithelial-induced suppression of breast cancer cell invasion. Enhanced invasion observed in 3D cultures with N1ME stefin A-low cells was reliant on cathepsin B activation, as addition of the small molecule inhibitor CA-074 rescued the DCIS-like non-invasive phenotype. Importantly, we confirmed that stefin A was indeed abundant in myoepithelial cells in breast tissue. Use of a 138-patient cohort confirmed that myoepithelial stefin A (cystatin A) is abundant in normal breast ducts and low-grade DCIS but reduced in high-grade DCIS, supporting myoepithelial stefin A as a candidate marker of lower risk of invasive relapse. We have therefore identified myoepithelial cell stefin A as a suppressor of early tumor invasion and a candidate marker to distinguish patients who are at low risk of developing invasive breast cancer, and can therefore be spared further treatment. Copyright © 2017 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. Copyright © 2017 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Cell-type-specific, Aptamer-functionalized Agents for Targeted Disease Therapy

PubMed Central

Zhou, Jiehua; Rossi, John J.

2014-01-01

One hundred years ago, Dr. Paul Ehrlich popularized the “magic bullet” concept for cancer therapy in which an ideal therapeutic agent would only kill the specific tumor cells it targeted. Since then, “targeted therapy” that specifically targets the molecular defects responsible for a patient's condition has become a long-standing goal for treating human disease. However, safe and efficient drug delivery during the treatment of cancer and infectious disease remains a major challenge for clinical translation and the development of new therapies. The advent of SELEX technology has inspired many groundbreaking studies that successfully adapted cell-specific aptamers for targeted delivery of active drug substances in both in vitro and in vivo models. By covalently linking or physically functionalizing the cell-specific aptamers with therapeutic agents, such as siRNA, microRNA, chemotherapeutics or toxins, or delivery vehicles, such as organic or inorganic nanocarriers, the targeted cells and tissues can be specifically recognized and the therapeutic compounds internalized, thereby improving the local concentration of the drug and its therapeutic efficacy. Currently, many cell-type-specific aptamers have been developed that can target distinct diseases or tissues in a cell-type-specific manner. In this review, we discuss recent advances in the use of cell-specific aptamers for targeted disease therapy, as well as conjugation strategies and challenges. PMID:24936916

Trispecific antibodies for CD16A-directed NK cell engagement and dual-targeting of tumor cells.

PubMed

Gantke, Thorsten; Weichel, Michael; Herbrecht, Carmen; Reusch, Uwe; Ellwanger, Kristina; Fucek, Ivica; Eser, Markus; Müller, Thomas; Griep, Remko; Molkenthin, Vera; Zhukovsky, Eugene A; Treder, Martin

2017-09-01

Bispecific antibodies that redirect the lytic activity of cytotoxic immune effector cells, such as T- and NK cells, onto tumor cells have emerged as a highly attractive and clinically validated treatment modality for hematological malignancies. Advancement of this therapeutic concept into solid tumor indications, however, is hampered by the scarcity of targetable antigens that are surface-expressed on tumor cells but demonstrate only limited expression on healthy tissues. To overcome this limitation, the concept of dual-targeting, i.e. the simultaneous targeting of two tumor-expressed surface antigens with limited co-expression on non-malignant cells, with multispecific antibodies has been proposed to increase tumor selectivity of antibody-induced effector cell cytotoxicity. Here, a novel CD16A (FcγRIIIa)-directed trispecific, tetravalent antibody format, termed aTriFlex, is described, that is capable of redirecting NK cell cytotoxicity to two surface-expressed antigens. Using a BCMA/CD200-based in vitro model system, the potential use of aTriFlex antibodies for dual-targeting and selective induction of NK cell-mediated target cell lysis was investigated. Bivalent bispecific target cell binding was found to result in significant avidity gains and up to 17-fold increased in vitro potency. These data suggest trispecific aTriFlex antibodies may support dual-targeting strategies to redirect NK cell cytotoxicity with increased selectivity to enable targeting of solid tumor antigens. © The Author 2017. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Dendritic cell targeted vaccines: Recent progresses and challenges

PubMed Central

Chen, Pengfei; Liu, Xinsheng; Sun, Yuefeng; Zhou, Peng; Wang, Yonglu; Zhang, Yongguang

2016-01-01

ABSTRACT Dendritic cells (DCs) are known to be a set of morphology, structure and function of heterogeneous professional antigen presenting cells (APCs), as well as the strongest functional antigen presenting cells, which can absorb, process and present antigens. As the key regulators of innate and adaptive immune responses, DCs are at the center of the immune system and capable of interacting with both B cells and T cells, thereby manipulating the humoral and cellular immune responses. DCs provide an essential link between the innate and adaptive immunity, and the strong immune activation function of DCs and their properties of natural adjuvants, make them a valuable target for antigen delivery. Targeting antigens to DC-specific endocytic receptors in combination with the relevant antibodies or ligands along with immunostimulatory adjuvants has been recently recognized as a promising strategy for designing an effective vaccine that elicits a strong and durable T cell response against intracellular pathogens and cancer. This opinion article provides a brief summary of the rationales, superiorities and challenges of existing DC-targeting approaches. PMID:26513200

Effect of a Targeted Early Literacy Intervention for English Learners

ERIC Educational Resources Information Center

Arellano, Elizabeth Michelle

2013-01-01

This study examined the effectiveness of a targeted early literacy intervention among Spanish-speaking kindergarten English Learners (ELs). Using a Response to Intervention (RtI) framework, participants were screened in English to ensure a need for additional literacy support. Selected students were then screened in Spanish, and students with…

Superior Control of HIV-1 Replication by CD8+ T Cells Targeting Conserved Epitopes: Implications for HIV Vaccine Design

PubMed Central

Kunwar, Pratima; Hawkins, Natalie; Dinges, Warren L.; Liu, Yi; Gabriel, Erin E.; Swan, David A.; Stevens, Claire E.; Maenza, Janine; Collier, Ann C.; Mullins, James I.; Hertz, Tomer; Yu, Xuesong; Horton, Helen

2013-01-01

A successful HIV vaccine will likely induce both humoral and cell-mediated immunity, however, the enormous diversity of HIV has hampered the development of a vaccine that effectively elicits both arms of the adaptive immune response. To tackle the problem of viral diversity, T cell-based vaccine approaches have focused on two main strategies (i) increasing the breadth of vaccine-induced responses or (ii) increasing vaccine-induced responses targeting only conserved regions of the virus. The relative extent to which set-point viremia is impacted by epitope-conservation of CD8+ T cell responses elicited during early HIV-infection is unknown but has important implications for vaccine design. To address this question, we comprehensively mapped HIV-1 CD8+ T cell epitope-specificities in 23 ART-naïve individuals during early infection and computed their conservation score (CS) by three different methods (prevalence, entropy and conseq) on clade-B and group-M sequence alignments. The majority of CD8+ T cell responses were directed against variable epitopes (p<0.01). Interestingly, increasing breadth of CD8+ T cell responses specifically recognizing conserved epitopes was associated with lower set-point viremia (r = - 0.65, p = 0.009). Moreover, subjects possessing CD8+ T cells recognizing at least one conserved epitope had 1.4 log10 lower set-point viremia compared to those recognizing only variable epitopes (p = 0.021). The association between viral control and the breadth of conserved CD8+ T cell responses may be influenced by the method of CS definition and sequences used to determine conservation levels. Strikingly, targeting variable versus conserved epitopes was independent of HLA type (p = 0.215). The associations with viral control were independent of functional avidity of CD8+ T cell responses elicited during early infection. Taken together, these data suggest that the next-generation of T-cell based HIV-1 vaccines should focus on strategies that

Selective tumor cell targeting by the disaccharide moiety of bleomycin.

PubMed

Yu, Zhiqiang; Schmaltz, Ryan M; Bozeman, Trevor C; Paul, Rakesh; Rishel, Michael J; Tsosie, Krystal S; Hecht, Sidney M

2013-02-27

In a recent study, the well-documented tumor targeting properties of the antitumor agent bleomycin (BLM) were studied in cell culture using microbubbles that had been derivatized with multiple copies of BLM. It was shown that BLM selectively targeted MCF-7 human breast carcinoma cells but not the "normal" breast cell line MCF-10A. Furthermore, it was found that the BLM analogue deglycobleomycin, which lacks the disaccharide moiety of BLM, did not target either cell line, indicating that the BLM disaccharide moiety is necessary for tumor selectivity. Not resolved in the earlier study were the issues of whether the BLM disaccharide moiety alone is sufficient for tumor cell targeting and the possible cellular uptake of the disaccharide. In the present study, we conjugated BLM, deglycoBLM, and BLM disaccharide to the cyanine dye Cy5**. It was found that the BLM and BLM disaccharide conjugates, but not the deglycoBLM conjugate, bound selectively to MCF-7 cells and were internalized. The same was also true for the prostate cancer cell line DU-145 (but not for normal PZ-HPV-7 prostate cells) and for the pancreatic cancer cell line BxPC-3 (but not for normal SVR A221a pancreas cells). The targeting efficiency of the disaccharide was only slightly less than that of BLM in MCF-7 and DU-145 cells and comparable to that of BLM in BxPC-3 cells. These results establish that the BLM disaccharide is both necessary and sufficient for tumor cell targeting, a finding with obvious implications for the design of novel tumor imaging and therapeutic agents.

Protein Tyrosine Phosphatase PRL2 Mediates Notch and Kit Signals in Early T Cell Progenitors.

PubMed

Kobayashi, Michihiro; Nabinger, Sarah C; Bai, Yunpeng; Yoshimoto, Momoko; Gao, Rui; Chen, Sisi; Yao, Chonghua; Dong, Yuanshu; Zhang, Lujuan; Rodriguez, Sonia; Yashiro-Ohtani, Yumi; Pear, Warren S; Carlesso, Nadia; Yoder, Mervin C; Kapur, Reuben; Kaplan, Mark H; Daniel Lacorazza, Hugo; Zhang, Zhong-Yin; Liu, Yan

2017-04-01

The molecular pathways regulating lymphoid priming, fate, and development of multipotent bone marrow hematopoietic stem and progenitor cells (HSPCs) that continuously feed thymic progenitors remain largely unknown. While Notch signal is indispensable for T cell specification and differentiation, the downstream effectors are not well understood. PRL2, a protein tyrosine phosphatase that regulates hematopoietic stem cell proliferation and self-renewal, is highly expressed in murine thymocyte progenitors. Here we demonstrate that protein tyrosine phosphatase PRL2 and receptor tyrosine kinase c-Kit are critical downstream targets and effectors of the canonical Notch/RBPJ pathway in early T cell progenitors. While PRL2 deficiency resulted in moderate defects of thymopoiesis in the steady state, de novo generation of T cells from Prl2 null hematopoietic stem cells was significantly reduced following transplantation. Prl2 null HSPCs also showed impaired T cell differentiation in vitro. We found that Notch/RBPJ signaling upregulated PRL2 as well as c-Kit expression in T cell progenitors. Further, PRL2 sustains Notch-mediated c-Kit expression and enhances stem cell factor/c-Kit signaling in T cell progenitors, promoting effective DN1-DN2 transition. Thus, we have identified a critical role for PRL2 phosphatase in mediating Notch and c-Kit signals in early T cell progenitors. Stem Cells 2017;35:1053-1064. © 2016 AlphaMed Press.

A dendritic cell targeted vaccine induces long-term HIV-specific immunity within the gastrointestinal tract.

PubMed

Ruane, D; Do, Y; Brane, L; Garg, A; Bozzacco, L; Kraus, T; Caskey, M; Salazar, A; Trumpheller, C; Mehandru, S

2016-09-01

Despite significant therapeutic advances for HIV-1 infected individuals, a preventative HIV-1 vaccine remains elusive. Studies focusing on early transmission events, including the observation that there is a profound loss of gastrointestinal (GI) CD4(+) T cells during acute HIV-1 infection, highlight the importance of inducing HIV-specific immunity within the gut. Here we report on the generation of cellular and humoral immune responses in the intestines by a mucosally administered, dendritic cell (DC) targeted vaccine. Our results show that nasally delivered α-CD205-p24 vaccine in combination with polyICLC, induced polyfunctional immune responses within naso-pulmonary lymphoid sites that disseminated widely to systemic and mucosal (GI tract and the vaginal epithelium) sites. Qualitatively, while α-CD205-p24 prime-boost immunization generated CD4(+) T-cell responses, heterologous prime-boost immunization with α-CD205-p24 and NYVAC gag-p24 generated high levels of HIV-specific CD4(+) and CD8(+) T cells within the GI tract. Finally, DC-targeting enhanced the amplitude and longevity of vaccine-induced immune responses in the GI tract. This is the first report of a nasally delivered, DC-targeted vaccine to generate HIV-specific immune responses in the GI tract and will potentially inform the design of preventative approaches against HIV-1 and other mucosal infections.

An innovative pre-targeting strategy for tumor cell specific imaging and therapy.

PubMed

Qin, Si-Yong; Peng, Meng-Yun; Rong, Lei; Jia, Hui-Zhen; Chen, Si; Cheng, Si-Xue; Feng, Jun; Zhang, Xian-Zheng

2015-09-21

A programmed pre-targeting system for tumor cell imaging and targeting therapy was established based on the "biotin-avidin" interaction. In this programmed functional system, transferrin-biotin can be actively captured by tumor cells with the overexpression of transferrin receptors, thus achieving the pre-targeting modality. Depending upon avidin-biotin recognition, the attachment of multivalent FITC-avidin to biotinylated tumor cells not only offered the rapid fluorescence labelling, but also endowed the pre-targeted cells with targeting sites for the specifically designed biotinylated peptide nano-drug. Owing to the successful pre-targeting, tumorous HepG2 and HeLa cells were effectively distinguished from the normal 3T3 cells via fluorescence imaging. In addition, the self-assembled peptide nano-drug resulted in enhanced cell apoptosis in the observed HepG2 cells. The tumor cell specific pre-targeting strategy is applicable for a variety of different imaging and therapeutic agents for tumor treatments.

Strategies to target non-T-cell HIV reservoirs.

PubMed

Sacha, Jonah B; Ndhlovu, Lishomwa C

2016-07-01

A central question for the HIV cure field is to determine new ways to target clinically relevant, latently and actively replicating HIV-infected cells beyond resting memory CD4 T cells, particularly in anatomical areas of low drug penetrability. HIV eradication strategies being positioned for targeting HIV for extinction in the CD4 T-cell compartment may also show promise in non-CD4 T-cells reservoirs. Furthermore, several exciting novel therapeutic approaches specifically focused on HIV clearance from non-CD4 T-cell populations are being developed. Although reservoir validity in these non-CD4 T cells continues to remain debated, this review will highlight recent advances and make an argument as to their clinical relevancy as we progress towards an HIV cure.

Growth hormone is a cellular senescence target in pituitary and nonpituitary cells

PubMed Central

Chesnokova, Vera; Zhou, Cuiqi; Ben-Shlomo, Anat; Zonis, Svetlana; Tani, Yuji; Ren, Song-Guang; Melmed, Shlomo

2013-01-01

Premature proliferative arrest in benign or early-stage tumors induced by oncoproteins, chromosomal instability, or DNA damage is associated with p53/p21 activation, culminating in either senescence or apoptosis, depending on cell context. Growth hormone (GH) elicits direct peripheral metabolic actions as well as growth effects mediated by insulin-like growth factor 1 (IGF1). Locally produced peripheral tissue GH, in contrast to circulating pituitary-derived endocrine GH, has been proposed to be both proapoptotic and prooncogenic. Pituitary adenomas expressing and secreting GH are invariably benign and exhibit DNA damage and a senescent phenotype. We therefore tested effects of nutlin-induced p53-mediated senescence in rat and human pituitary cells. We show that DNA damage senescence induced by nutlin triggers the p53/p21 senescent pathway, with subsequent marked induction of intracellular pituitary GH in vitro. In contrast, GH is not induced in cells devoid of p53. Furthermore we show that p53 binds specific GH promoter motifs and enhances GH transcription and secretion in senescent pituitary adenoma cells and also in nonpituitary (human breast and colon) cells. In vivo, treatment with nutlin results in up-regulation of both p53 and GH in the pituitary gland, as well as increased GH expression in nonpituitary tissues (lung and liver). Intracrine GH acts in pituitary cells as an apoptosis switch for p53-mediated senescence, likely protecting the pituitary adenoma from progression to malignancy. Unlike in the pituitary, in nonpituitary cells GH exerts antiapoptotic properties. Thus, the results show that GH is a direct p53 transcriptional target and fulfills criteria as a p53 target gene. Induced GH is a readily measurable cell marker for p53-mediated cellular senescence. PMID:23940366

Potential targets for lung squamous cell carcinoma

Cancer.gov

Researchers have identified potential therapeutic targets in lung squamous cell carcinoma, the second most common form of lung cancer. The Cancer Genome Atlas (TCGA) Research Network study comprehensively characterized the lung squamous cell carcinoma gen

N-acetylgalactosamine-functionalized dendrimers as hepatic cancer cell-targeted carriers.

PubMed

Medina, Scott H; Tekumalla, Venkatesh; Chevliakov, Maxim V; Shewach, Donna S; Ensminger, William D; El-Sayed, Mohamed E H

2011-06-01

There is an urgent need for novel polymeric carriers that can selectively deliver a large dose of chemotherapeutic agents into hepatic cancer cells to achieve high therapeutic activity with minimal systemic side effects. PAMAM dendrimers are characterized by a unique branching architecture and a large number of chemical surface groups suitable for coupling of chemotherapeutic agents. In this article, we report the coupling of N-acetylgalactosamine (NAcGal) to generation 5 (G5) of poly(amidoamine) (PAMAM-NH₂) dendrimers via peptide and thiourea linkages to prepare NAcGal-targeted carriers used for targeted delivery of chemotherapeutic agents into hepatic cancer cells. We describe the uptake of NAcGal-targeted and non-targeted G5 dendrimers into hepatic cancer cells (HepG2) as a function of G5 concentration and incubation time. We examine the contribution of the asialoglycoprotein receptor (ASGPR) to the internalization of NAcGal-targeted dendrimers into hepatic cancer cells through a competitive inhibition assay. Our results show that uptake of NAcGal-targeted G5 dendrimers into hepatic cancer cells occurs via ASGPR-mediated endocytosis. Internalization of these targeted carriers increased with the increase in G5 concentration and incubation time following Michaelis-Menten kinetics characteristic of receptor-mediated endocytosis. These results collectively indicate that G5-NAcGal conjugates function as targeted carriers for selective delivery of chemotherapeutic agents into hepatic cancer cells. Copyright © 2011 Elsevier Ltd. All rights reserved.

Cell cycle proteins as promising targets in cancer therapy.

PubMed

Otto, Tobias; Sicinski, Piotr

2017-01-27

Cancer is characterized by uncontrolled tumour cell proliferation resulting from aberrant activity of various cell cycle proteins. Therefore, cell cycle regulators are considered attractive targets in cancer therapy. Intriguingly, animal models demonstrate that some of these proteins are not essential for proliferation of non-transformed cells and development of most tissues. By contrast, many cancers are uniquely dependent on these proteins and hence are selectively sensitive to their inhibition. After decades of research on the physiological functions of cell cycle proteins and their relevance for cancer, this knowledge recently translated into the first approved cancer therapeutic targeting of a direct regulator of the cell cycle. In this Review, we focus on proteins that directly regulate cell cycle progression (such as cyclin-dependent kinases (CDKs)), as well as checkpoint kinases, Aurora kinases and Polo-like kinases (PLKs). We discuss the role of cell cycle proteins in cancer, the rationale for targeting them in cancer treatment and results of clinical trials, as well as the future therapeutic potential of various cell cycle inhibitors.

A Cell-Based Approach to Early Pancreatic Cancer Detection

DTIC Science & Technology

2016-10-01

Award Number: W81XWH-15-1-0457 TITLE: A Cell -Based Approach to Early Pancreatic Cancer Detection PRINCIPAL INVESTIGATOR: Dr. Ben Stanger...SUBTITLE 5a. CONTRACT NUMBER A Cell -Based Approach to Early Pancreatic Cancer Detection 5b. GRANT NUMBER W81XWH-15-1-0457 5c. PROGRAM ELEMENT NUMBER 6...tumor cells from mouse blood by depleting the sample of white blood cells (WBCs). Furthermore, the RNA profile of these cells can be assessed by

Imatinib-loaded polyelectrolyte microcapsules for sustained targeting of BCR-ABL+ leukemia stem cells.

PubMed

Palamà, Ilaria E; Leporatti, Stefano; de Luca, Emanuela; Di Renzo, Nicola; Maffia, Michele; Gambacorti-Passerini, Carlo; Rinaldi, Ross; Gigli, Giuseppe; Cingolani, Roberto; Coluccia, Addolorata M L

2010-04-01

The lack of sensitivity of chronic myeloid leukemia (CML) stem cells to imatinib mesylate (IM) commonly leads to drug dose escalation or early disease relapses when therapy is stopped. Here, we report that packaging of IM into a biodegradable carrier based on polyelectrolyte microcapsules increases drug retention and antitumor activity in CML stem cells, also improving the ex vivo purging of malignant progenitors from patient autografts. Microparticles/capsules were obtained by layer-by-layer (LbL) self-assembly of oppositely charged polyelectrolyte multilayers on removable calcium carbonate (CaCO(3)) templates and loaded with or without IM. A leukemic cell line (KU812) and CD34(+) cells freshly isolated from healthy donors or CML patients were tested. Polyelectrolyte microcapsules (PMCs) with an average diameter of 3 microm, fluorescently labelled multilayers sensitive to the action of intracellular proteases and 95-99% encapsulation efficiency of IM, were prepared. Cell uptake efficiency of such biodegradable carriers was quantified in KU812, leukemic and normal CD34(+) stem cells (range: 70-85%), and empty PMCs did not impact cell viability. IM-loaded PMCs selectively targeted CML cells, by promoting apoptosis at doses that exert only cytostatic effects by IM alone. More importantly, residual CML cells from patient leukapheresis products were reduced or eliminated more efficiently by using IM-loaded PMCs compared with freely soluble IM, with a purging efficiency of several logs. No adverse effects on normal CD34(+) stem-cell survival and their clonogenic potential was noticed in long-term cultures of hematopoietic progenitors in vitro. This pilot study provides the proof-of-principle for the clinical application of biodegradable IM-loaded PMC as feasible, safe and effective ex vivo purging agents to target CML stem cells, in order to improve transplant outcome of resistant/relapsed patients or reduce IM dose escalation.

Folate-conjugated immunoglobulin targets melanoma tumor cells for NK cell effector functions

PubMed Central

Skinner, Cassandra C.; McMichael, Elizabeth L.; Jaime-Ramirez, Alena C.; Abrams, Zachary B.; Lee, Robert J.; Carson, William E.

2016-01-01

The folate receptor (FR) is over-expressed on the vascular side of cancerous cells including those of the breast, ovaries, testes, and cervix. We hypothesized that a folate-conjugated immunoglobulin (F-IgG) would bind to the FR that is over-expressed on melanoma tumor cells to target these cells for lysis by natural killer (NK) cells. Folate receptor expression was confirmed in the Mel-39 (human melanoma) cell line by flow cytometry and immunoblot analysis, using KB (human oral epithelial) and F01 (human melanoma) as a positive and negative control, respectively. FR-positive and negative cell lines were treated with F-IgG or control immunoglobulin G (C-IgG) in the presence or absence of cytokines in order to determine NK cell ability to lyse FR-positive cell lines. NK cell activation was significantly upregulated and lysis of Mel 39 tumor cells enhanced following treatment with F-IgG, as compared to C-IgG at all effector:target (E:T) ratios (p<0.01). This trend was further enhanced by NK cell stimulation with the activating cytokine interleukin-12 (IL-12). NK cell production of cytokines such as interferon-gamma (IFN-γ), macrophage inflammatory protein 1 alpha (MIP-1α), and regulated on activation normal T-cell expressed and secreted (RANTES) were also significantly increased in response to co-stimulation with IL-12 stimulation and F-IgG-coated Mel 39 target cells, as compared to controls (p<0.01). In contrast, F-IgG did not bind to the FR-negative cell line F01 and had no significant effect on NK cell lysis or cytokine production. This research indicates the potential use of F-IgG for its ability to induce an immune response from NK cells against FR-positive melanoma tumor cells which can be further enhanced by the addition of cytokines. PMID:27035691

Pregnancy at early age is associated with a reduction of progesterone-responsive cells and epithelial Wnt signaling in human breast tissue.

PubMed

Muenst, Simone; Mechera, Robert; Däster, Silvio; Piscuoglio, Salvatore; Ng, Charlotte K Y; Meier-Abt, Fabienne; Weber, Walter P; Soysal, Savas D

2017-04-04

Pregnancy at early age is the most significant modifiable factor which consistently decreases lifetime breast cancer risk. However, the underlying mechanisms haven't been conclusively identified. Studies in mice suggest a reduction in progesterone-receptor (PR) sensitive epithelial cells as well as a downregulation of the Wnt signaling pathway as being one of the main mechanisms for the protective effect of early pregnancy. The aim of our study was to validate these findings in humans. We collected benign breast tissue of 125 women who had been stratified according to age at first pregnancy and the occurrence of subsequent breast cancer, and performed immunohistochemistry for PR, Wnt4 and the Wnt-target Versican. The number of PR positive epithelial cells was significantly lower in the group of women with early pregnancy and no subsequent breast cancer compared to the group of nulliparous women with subsequent invasive breast cancer (p = 0.0135). In women with early pregnancy, expression of Versican and Wnt4 was significantly lower compared to nulliparous women (p = 0.0036 and p = 0.0241 respectively), and Versican expression was also significant lower compared to women with late pregnancy (p < 0.0001). Our results confirm prior observations in mice and suggest a role of downregulation of epithelial Wnt signaling in the protective effect of early pregnancy in humans. This results in a decreased proliferation of stem/progenitor cells; therefore, the Wnt signaling pathway may represent a potential target for breast cancer prevention in humans.

Curcumin: a promising agent targeting cancer stem cells.

PubMed

Zang, Shufei; Liu, Tao; Shi, Junping; Qiao, Liang

2014-01-01

Cancer stem cells are a subset of cells that are responsible for cancer initiation and relapse. They are generally resistant to the current anticancer agents. Successful anticancer therapy must consist of approaches that can target not only the differentiated cancer cells, but also cancer stem cells. Emerging evidence suggested that the dietary agent curcumin exerted its anti-cancer activities via targeting cancer stem cells of various origins such as those of colorectal cancer, pancreatic cancer, breast cancer, brain cancer, and head and neck cancer. In order to enhance the therapeutic potential of curcumin, this agent has been modified or used in combination with other agents in the experimental therapy for many cancers. In this mini-review, we discussed the effect of curcumin and its derivatives in eliminating cancer stem cells and the possible underlying mechanisms.

The mechanism of T-cell mediated cytotoxicity. VI. T-cell projections and their role in target cell killing.

PubMed Central

Sanderson, C J; Glauert, A M

1979-01-01

Electron micrographs of material fixed during the first 10 min of a T-cell cytotoxic system showed T-cell projections and T-cell burrowing into target cells. These observations were made possible by using a system with a very high rate of killing. The projections vary in shape and size, and can push deeply into the target cell, distorting organelles in their path, including the nucleus. The projections contain fine fibrillar material, to the exclusion of organelles. They push the target cell membrane in front of them to form pockets approximating to the shape of the projection. Areas of close contact occur between the projections and the target cell membrane, particularly at the leading edges. The likelihood that these projections develop as a result of contact with specific antigen, and are involved in the cytotoxic mechanism is discussed. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 Figure 9 Figure 10 Figure 11 Figure 12 Figure 13 Figure 14 Figure 15 Figure 16 PMID:311336

Targeting survival pathways in chronic myeloid leukaemia stem cells

PubMed Central

Sinclair, A; Latif, A L; Holyoake, T L

2013-01-01

Chronic myeloid leukaemia (CML) is a clonal myeloproliferative disorder characterized by the presence of a fusion oncogene BCR-ABL, which encodes a protein with constitutive TK activity. The implementation of tyrosine kinase inhibitors (TKIs) marked a major advance in CML therapy; however, there are problems with current treatment. For example, relapse occurs when these drugs are discontinued in the majority of patients who have achieved a complete molecular response on TKI and these agents are less effective in patients with mutations in the BCR-ABL kinase domain. Importantly, TKI can effectively target proliferating mature cells, but do not eradicate quiescent leukaemic stem cells (LSCs), therefore allowing disease persistence despite treatment. It is essential that alternative strategies are used to target the LSC population. BCR-ABL activation is responsible for the modulation of different signalling pathways, which allows the LSC fraction to evade cell death. Several pathways have been shown to be modulated by BCR-ABL, including PI3K/AKT/mTOR, JAK-STAT and autophagy signalling pathways. Targeting components of these survival pathways, alone or in combination with TKI, therefore represents an attractive potential therapeutic approach for targeting the LSC. However, many pathways are also active in normal stem cells. Therefore, potential targets must be validated to effectively eradicate CML stem cells while sparing normal counterparts. This review summarizes the main pathways modulated in CML stem cells, the recent developments and the use of novel drugs to target components in these pathways which may be used to target the LSC population. Linked Articles This article is part of a themed section on Emerging Therapeutic Aspects in Oncology. To view the other articles in this section visit http://dx.doi.org/10.1111/bph.2013.169.issue-8 PMID:23517124

Identification of a Novel Drug Lead That Inhibits HCV Infection and Cell-to-Cell Transmission by Targeting the HCV E2 Glycoprotein

DOE PAGES

Al Olaby, Reem R.; Cocquerel, Laurence; Zemla, Adam; ...

2014-10-30

We report that Hepatitis C Virus (HCV) infects 200 million individuals worldwide. Although several FDA approved drugs targeting the HCV serine protease and polymerase have shown promising results, there is a need for better drugs that are effective in treating a broader range of HCV genotypes and subtypes without being used in combination with interferon and/or ribavirin. Recently, two crystal structures of the core of the HCV E2 protein (E2c) have been determined, providing structural information that can now be used to target the E2 protein and develop drugs that disrupt the early stages of HCV infection by blocking E2’smore » interaction with different host factors. Using the E2c structure as a template, we have created a structural model of the E2 protein core (residues 421–645) that contains the three amino acid segments that are not present in either structure. Computational docking of a diverse library of 1,715 small molecules to this model led to the identification of a set of 34 ligands predicted to bind near conserved amino acid residues involved in the HCV E2: CD81 interaction. We used surface plasmon resonance detection to screen the ligand set for binding to recombinant E2 protein, and the best binders were subsequently tested to identify compounds that inhibit the infection of Huh-7 cells by HCV. One compound, 281816, blocked E2 binding to CD81 and inhibited HCV infection in a genotype-independent manner with IC50’s ranging from 2.2 µM to 4.6 µM. 281816 blocked the early and late steps of cell-free HCV entry and also abrogated the cell-to-cell transmission of HCV. Collectively the results obtained with this new structural model of E2c suggest the development of small molecule inhibitors such as 281816 that target E2 and disrupt its interaction with CD81 may provide a new paradigm for HCV treatment.« less

Identification of a Novel Drug Lead That Inhibits HCV Infection and Cell-to-Cell Transmission by Targeting the HCV E2 Glycoprotein

PubMed Central

Al Olaby, Reem R.; Cocquerel, Laurence; Zemla, Adam; Saas, Laure; Dubuisson, Jean; Vielmetter, Jost; Marcotrigiano, Joseph; Khan, Abdul Ghafoor; Catalan, Felipe Vences; Perryman, Alexander L.; Freundlich, Joel S.; Forli, Stefano; Levy, Shoshana; Balhorn, Rod; Azzazy, Hassan M.

2014-01-01

Hepatitis C Virus (HCV) infects 200 million individuals worldwide. Although several FDA approved drugs targeting the HCV serine protease and polymerase have shown promising results, there is a need for better drugs that are effective in treating a broader range of HCV genotypes and subtypes without being used in combination with interferon and/or ribavirin. Recently, two crystal structures of the core of the HCV E2 protein (E2c) have been determined, providing structural information that can now be used to target the E2 protein and develop drugs that disrupt the early stages of HCV infection by blocking E2’s interaction with different host factors. Using the E2c structure as a template, we have created a structural model of the E2 protein core (residues 421–645) that contains the three amino acid segments that are not present in either structure. Computational docking of a diverse library of 1,715 small molecules to this model led to the identification of a set of 34 ligands predicted to bind near conserved amino acid residues involved in the HCV E2: CD81 interaction. Surface plasmon resonance detection was used to screen the ligand set for binding to recombinant E2 protein, and the best binders were subsequently tested to identify compounds that inhibit the infection of Huh-7 cells by HCV. One compound, 281816, blocked E2 binding to CD81 and inhibited HCV infection in a genotype-independent manner with IC50’s ranging from 2.2 µM to 4.6 µM. 281816 blocked the early and late steps of cell-free HCV entry and also abrogated the cell-to-cell transmission of HCV. Collectively the results obtained with this new structural model of E2c suggest the development of small molecule inhibitors such as 281816 that target E2 and disrupt its interaction with CD81 may provide a new paradigm for HCV treatment. PMID:25357246

Identification of a novel drug lead that inhibits HCV infection and cell-to-cell transmission by targeting the HCV E2 glycoprotein.

PubMed

Al Olaby, Reem R; Cocquerel, Laurence; Zemla, Adam; Saas, Laure; Dubuisson, Jean; Vielmetter, Jost; Marcotrigiano, Joseph; Khan, Abdul Ghafoor; Vences Catalan, Felipe; Perryman, Alexander L; Freundlich, Joel S; Forli, Stefano; Levy, Shoshana; Balhorn, Rod; Azzazy, Hassan M

2014-01-01

Hepatitis C Virus (HCV) infects 200 million individuals worldwide. Although several FDA approved drugs targeting the HCV serine protease and polymerase have shown promising results, there is a need for better drugs that are effective in treating a broader range of HCV genotypes and subtypes without being used in combination with interferon and/or ribavirin. Recently, two crystal structures of the core of the HCV E2 protein (E2c) have been determined, providing structural information that can now be used to target the E2 protein and develop drugs that disrupt the early stages of HCV infection by blocking E2's interaction with different host factors. Using the E2c structure as a template, we have created a structural model of the E2 protein core (residues 421-645) that contains the three amino acid segments that are not present in either structure. Computational docking of a diverse library of 1,715 small molecules to this model led to the identification of a set of 34 ligands predicted to bind near conserved amino acid residues involved in the HCV E2: CD81 interaction. Surface plasmon resonance detection was used to screen the ligand set for binding to recombinant E2 protein, and the best binders were subsequently tested to identify compounds that inhibit the infection of Huh-7 cells by HCV. One compound, 281816, blocked E2 binding to CD81 and inhibited HCV infection in a genotype-independent manner with IC50's ranging from 2.2 µM to 4.6 µM. 281816 blocked the early and late steps of cell-free HCV entry and also abrogated the cell-to-cell transmission of HCV. Collectively the results obtained with this new structural model of E2c suggest the development of small molecule inhibitors such as 281816 that target E2 and disrupt its interaction with CD81 may provide a new paradigm for HCV treatment.

Identification of a Novel Drug Lead That Inhibits HCV Infection and Cell-to-Cell Transmission by Targeting the HCV E2 Glycoprotein

DOE Office of Scientific and Technical Information (OSTI.GOV)

Al Olaby, Reem R.; Cocquerel, Laurence; Zemla, Adam

We report that Hepatitis C Virus (HCV) infects 200 million individuals worldwide. Although several FDA approved drugs targeting the HCV serine protease and polymerase have shown promising results, there is a need for better drugs that are effective in treating a broader range of HCV genotypes and subtypes without being used in combination with interferon and/or ribavirin. Recently, two crystal structures of the core of the HCV E2 protein (E2c) have been determined, providing structural information that can now be used to target the E2 protein and develop drugs that disrupt the early stages of HCV infection by blocking E2’smore » interaction with different host factors. Using the E2c structure as a template, we have created a structural model of the E2 protein core (residues 421–645) that contains the three amino acid segments that are not present in either structure. Computational docking of a diverse library of 1,715 small molecules to this model led to the identification of a set of 34 ligands predicted to bind near conserved amino acid residues involved in the HCV E2: CD81 interaction. We used surface plasmon resonance detection to screen the ligand set for binding to recombinant E2 protein, and the best binders were subsequently tested to identify compounds that inhibit the infection of Huh-7 cells by HCV. One compound, 281816, blocked E2 binding to CD81 and inhibited HCV infection in a genotype-independent manner with IC50’s ranging from 2.2 µM to 4.6 µM. 281816 blocked the early and late steps of cell-free HCV entry and also abrogated the cell-to-cell transmission of HCV. Collectively the results obtained with this new structural model of E2c suggest the development of small molecule inhibitors such as 281816 that target E2 and disrupt its interaction with CD81 may provide a new paradigm for HCV treatment.« less

Breast cancer stem cells, EMT and therapeutic targets

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kotiyal, Srishti; Bhattacharya, Susinjan, E-mail: s.bhattacharya@jiit.ac.in

Highlights: • Therapeutic targeting or inhibition of the key molecules of signaling pathways can control growth of breast cancer stem cells (BCSCs). • Development of BCSCs also involves miRNA interactions. • Therapeutic achievement can be done by targeting identified targets in the BCSC pathways. - Abstract: A small heterogeneous population of breast cancer cells acts as seeds to induce new tumor growth. These seeds or breast cancer stem cells (BCSCs) exhibit great phenotypical plasticity which allows them to undergo “epithelial to mesenchymal transition” (EMT) at the site of primary tumor and a future reverse transition. Apart from metastasis they aremore » also responsible for maintaining the tumor and conferring it with drug and radiation resistance and a tendency for post-treatment relapse. Many of the signaling pathways involved in induction of EMT are involved in CSC generation and regulation. Here we are briefly reviewing the mechanism of TGF-β, Wnt, Notch, TNF-α, NF-κB, RTK signalling pathways which are involved in EMT as well as BCSCs maintenance. Therapeutic targeting or inhibition of the key/accessory players of these pathways could control growth of BCSCs and hence malignant cancer. Additionally several miRNAs are dysregulated in cancer stem cells indicating their roles as oncogenes or tumor suppressors. This review also lists the miRNA interactions identified in BCSCs and discusses on some newly identified targets in the BCSC regulatory pathways like SHIP2, nicastrin, Pin 1, IGF-1R, pro-inflammatory cytokines and syndecan which can be targeted for therapeutic achievements.« less

Neurotrophins Acting Via TRKB Receptors Activate the JAGGED1-NOTCH2 Cell-Cell Communication Pathway to Facilitate Early Ovarian Development

PubMed Central

Dorfman, Mauricio D.; Kerr, Bredford; Garcia-Rudaz, Cecilia; Paredes, Alfonso H.; Dissen, Gregory A.

2011-01-01

Tropomyosin-related kinase (TRK) receptor B (TRKB) mediates the supportive actions of neurotrophin 4/5 and brain-derived neurotrophic factor on early ovarian follicle development. Absence of TRKB receptors reduces granulosa cell (GC) proliferation and delays follicle growth. In the present study, we offer mechanistic insights into this phenomenon. DNA array and quantitative PCR analysis of ovaries from TrkB-null mice revealed that by the end of the first week of postnatal life, Jagged1, Hes1, and Hey2 mRNA abundance is reduced in the absence of TRKB receptors. Although Jagged1 encodes a NOTCH receptor ligand, Hes1 and Hey2 are downstream targets of the JAGGED1-NOTCH2 signaling system. Jagged1 is predominantly expressed in oocytes, and the abundance of JAGGED1 is decreased in TrkB−/− oocytes. Lack of TRKB receptors also resulted in reduced expression of c-Myc, a NOTCH target gene that promotes entry into the cell cycle, but did not alter the expression of genes encoding core regulators of cell-cycle progression. Selective restoration of JAGGED1 synthesis in oocytes of TrkB−/− ovaries via lentiviral-mediated transfer of the Jagged1 gene under the control of the growth differentiation factor 9 (Gdf9) promoter rescued c-Myc expression, GC proliferation, and follicle growth. These results suggest that neurotrophins acting via TRKB receptors facilitate early follicle growth by supporting a JAGGED1-NOTCH2 oocyte-to-GC communication pathway, which promotes GC proliferation via a c-MYC-dependent mechanism. PMID:22028443

Early and late HIV-1 membrane fusion events are impaired by sphinganine lipidated peptides that target the fusion site.

PubMed

Klug, Yoel A; Ashkenazi, Avraham; Viard, Mathias; Porat, Ziv; Blumenthal, Robert; Shai, Yechiel

2014-07-15

Lipid-conjugated peptides have advanced the understanding of membrane protein functions and the roles of lipids in the membrane milieu. These lipopeptides modulate various biological systems such as viral fusion. A single function has been suggested for the lipid, binding to the membrane and thus elevating the local concentration of the peptide at the target site. In the present paper, we challenged this argument by exploring in-depth the antiviral mechanism of lipopeptides, which comprise sphinganine, the lipid backbone of DHSM (dihydrosphingomyelin), and an HIV-1 envelope-derived peptide. Surprisingly, we discovered a partnership between the lipid and the peptide that impaired early membrane fusion events by reducing CD4 receptor lateral diffusion and HIV-1 fusion peptide-mediated lipid mixing. Moreover, only the joint function of sphinganine and its conjugate peptide disrupted HIV-1 fusion protein assembly and folding at the later fusion steps. Via imaging techniques we revealed for the first time the direct localization of these lipopeptides to the virus-cell and cell-cell contact sites. Overall, the findings of the present study may suggest lipid-protein interactions in various biological systems and may help uncover a role for elevated DHSM in HIV-1 and its target cell membranes.

Plasmonic nanobubbles for target cell-specific gene and drug delivery and multifunctional processing of heterogeneous cell systems

NASA Astrophysics Data System (ADS)

Lukianova-Hleb, Ekaterina Y.; Huye, Leslie E.; Brenner, Malcolm K.; Lapotko, Dmitri O.

2014-03-01

Cell and gene cancer therapies require ex vivo cell processing of human grafts. Such processing requires at least three steps - cell enrichment, cell separation (destruction), and gene transfer - each of which requires the use of a separate technology. While these technologies may be satisfactory for research use, they are of limited usefulness in the clinical treatment setting because they have a low processing rate, as well as a low transfection and separation efficacy and specificity in heterogeneous human grafts. Most problematic, because current technologies are administered in multiple steps - rather than in a single, multifunctional, and simultaneous procedure - they lengthen treatment process and introduce an unnecessary level of complexity, labor, and resources into clinical treatment; all these limitations result in high losses of valuable cells. We report a universal, high-throughput, and multifunctional technology that simultaneously (1) inject free external cargo in target cells, (2) destroys unwanted cells, and (3) preserve valuable non-target cells in heterogeneous grafts. Each of these functions has single target cell specificity in heterogeneous cell system, processing rate > 45 mln cell/min, injection efficacy 90% under 96% viability of the injected cells, target cell destruction efficacy > 99%, viability of not-target cells >99% The developed technology employs novel cellular agents, called plasmonic nanobubbles (PNBs). PNBs are not particles, but transient, intracellular events, a vapor nanobubbles that expand and collapse in mere nanoseconds under optical excitation of gold nanoparticles with short picosecond laser pulses. PNBs of different, cell-specific, size (1) inject free external cargo with small PNBs, (2) Destroy other target cells mechanically with large PNBs and (3) Preserve non-target cells. The multi-functionality, precision, and high throughput of all-in-one PNB technology will tremendously impact cell and gene therapies and other

Targeted Sos1 deletion reveals its critical role in early T-cell development

PubMed Central

Kortum, Robert L.; Sommers, Connie L.; Alexander, Clayton P.; Pinski, John M.; Li, Wenmei; Grinberg, Alex; Lee, Jan; Love, Paul E.; Samelson, Lawrence E.

2011-01-01

Activation of the small G protein Ras is required for thymocyte differentiation. In thymocytes, Ras is activated by the Ras guanine exchange factors (RasGEFs) Sos1, Sos2, and RasGRP1. We report the development of a floxed allele of sos1 to assess the role of Sos1 during thymocyte development. Sos1 was required for pre–T-cell receptor (pre-TCR)– but not TCR-stimulated developmental signals. Sos1 deletion led to a partial block at the DN-to-DP transition. Sos1-deficient thymocytes showed reduced pre-TCR–stimulated proliferation, differentiation, and ERK phosphorylation. In contrast, TCR-stimulated positive selection, and negative selection under strong stimulatory conditions, remained intact in Sos1-deficient mice. Comparison of RasGEF expression at different developmental stages showed that relative to Sos2 and RasGRP1, Sos1 is most abundant in DN thymocytes, but least abundant in DP thymocytes. These data reveal that Sos1 is uniquely positioned to affect signal transduction early in thymocyte development. PMID:21746917

Early ultrasonographic evaluation of tumor thrombus level during sunitinib therapy for renal cell carcinoma.

PubMed

Sano, Futoshi; Fusayasu, Syusei; Otake, Shinji; Yamanaka, Hiroyuki; Tatenuma, Tomoyuki; Sakata, Ryoko; Makiyama, Kazuhide; Nakaigawa, Noboru; Yao, Masahiro; Kubota, Yoshinobu

2013-10-01

A 72-year-old man presenting with a 14-cm left renal mass, an inferior vena cava (IVC) tumor thrombus, and pulmonary metastases underwent renal mass biopsy that revealed clear cell renal cell carcinoma. Because of metastases and the extent of the tumor thrombus, sunitinib was administered, which resulted in a marked reduction in the tumor thrombus (from level III to level II after 11 weeks of treatment). Ultrasonography, preceding computed tomography, showed a slight shrinkage of the tumor thrombus level in the first 2 weeks. Therefore, ultrasound may be advantageous to monitor the IVC tumor thrombus level during the early phase of targeted therapy.

The Mechanism of Gene Targeting in Human Somatic Cells

PubMed Central

Kan, Yinan; Ruis, Brian; Lin, Sherry; Hendrickson, Eric A.

2014-01-01

Gene targeting in human somatic cells is of importance because it can be used to either delineate the loss-of-function phenotype of a gene or correct a mutated gene back to wild-type. Both of these outcomes require a form of DNA double-strand break (DSB) repair known as homologous recombination (HR). The mechanism of HR leading to gene targeting, however, is not well understood in human cells. Here, we demonstrate that a two-end, ends-out HR intermediate is valid for human gene targeting. Furthermore, the resolution step of this intermediate occurs via the classic DSB repair model of HR while synthesis-dependent strand annealing and Holliday Junction dissolution are, at best, minor pathways. Moreover, and in contrast to other systems, the positions of Holliday Junction resolution are evenly distributed along the homology arms of the targeting vector. Most unexpectedly, we demonstrate that when a meganuclease is used to introduce a chromosomal DSB to augment gene targeting, the mechanism of gene targeting is inverted to an ends-in process. Finally, we demonstrate that the anti-recombination activity of mismatch repair is a significant impediment to gene targeting. These observations significantly advance our understanding of HR and gene targeting in human cells. PMID:24699519

Targeted cellular ablation based on the morphology of malignant cells

NASA Astrophysics Data System (ADS)

Ivey, Jill W.; Latouche, Eduardo L.; Sano, Michael B.; Rossmeisl, John H.; Davalos, Rafael V.; Verbridge, Scott S.

2015-11-01

Treatment of glioblastoma multiforme (GBM) is especially challenging due to a shortage of methods to preferentially target diffuse infiltrative cells, and therapy-resistant glioma stem cell populations. Here we report a physical treatment method based on electrical disruption of cells, whose action depends strongly on cellular morphology. Interestingly, numerical modeling suggests that while outer lipid bilayer disruption induced by long pulses (~100 μs) is enhanced for larger cells, short pulses (~1 μs) preferentially result in high fields within the cell interior, which scale in magnitude with nucleus size. Because enlarged nuclei represent a reliable indicator of malignancy, this suggested a means of preferentially targeting malignant cells. While we demonstrate killing of both normal and malignant cells using pulsed electric fields (PEFs) to treat spontaneous canine GBM, we proposed that properly tuned PEFs might provide targeted ablation based on nuclear size. Using 3D hydrogel models of normal and malignant brain tissues, which permit high-resolution interrogation during treatment testing, we confirmed that PEFs could be tuned to preferentially kill cancerous cells. Finally, we estimated the nuclear envelope electric potential disruption needed for cell death from PEFs. Our results may be useful in safely targeting the therapy-resistant cell niches that cause recurrence of GBM tumors.

Targeted cellular ablation based on the morphology of malignant cells

PubMed Central

Ivey, Jill W.; Latouche, Eduardo L.; Sano, Michael B.; Rossmeisl, John H.; Davalos, Rafael V.; Verbridge, Scott S.

2015-01-01

Treatment of glioblastoma multiforme (GBM) is especially challenging due to a shortage of methods to preferentially target diffuse infiltrative cells, and therapy-resistant glioma stem cell populations. Here we report a physical treatment method based on electrical disruption of cells, whose action depends strongly on cellular morphology. Interestingly, numerical modeling suggests that while outer lipid bilayer disruption induced by long pulses (~100 μs) is enhanced for larger cells, short pulses (~1 μs) preferentially result in high fields within the cell interior, which scale in magnitude with nucleus size. Because enlarged nuclei represent a reliable indicator of malignancy, this suggested a means of preferentially targeting malignant cells. While we demonstrate killing of both normal and malignant cells using pulsed electric fields (PEFs) to treat spontaneous canine GBM, we proposed that properly tuned PEFs might provide targeted ablation based on nuclear size. Using 3D hydrogel models of normal and malignant brain tissues, which permit high-resolution interrogation during treatment testing, we confirmed that PEFs could be tuned to preferentially kill cancerous cells. Finally, we estimated the nuclear envelope electric potential disruption needed for cell death from PEFs. Our results may be useful in safely targeting the therapy-resistant cell niches that cause recurrence of GBM tumors. PMID:26596248

A missense mutation in myosin VIIA prevents aminoglycoside accumulation in early postnatal cochlear hair cells.

PubMed

Richardson, G P; Forge, A; Kros, C J; Marcotti, W; Becker, D; Williams, D S; Thorpe, J; Fleming, J; Brown, S D; Steel, K P

1999-11-28

Myosin VIIA is expressed by sensory hair cells in the inner ear and proximal tubule cells in the kidney, the two primary targets of aminoglycoside antibiotics. Using cochlear cultures prepared from early postnatal Myo7a6J mice with a missense mutation in the head region of the myosin VIIA molecule we show that this myosin is required for aminoglycoside accumulation in cochlear hair cells. Hair cells in homozygous mutant Myo7a6J cochlear cultures have disorganized hair bundles, but are otherwise morphologically normal and transduce. However, and in contrast to hair cells from heterozygous Myo7a6J cultures, the homozygous Myo7a6J hair cells do not accumulate [3H]gentamicin and do not exhibit an ototoxic response on exposure to aminoglycoside. Possible roles for myosin VIIA in the process of aminoglycoside accumulation are discussed.

The Osteogenic Niche Promotes Early-Stage Bone Colonization of Disseminated Breast Cancer Cells

PubMed Central

Wang, Hai; Yu, Cuijuan; Gao, Xia; Welte, Thomas; Muscarella, Aaron M.; Tian, Lin; Zhao, Hong; Zhao, Zhen; Du, Shiyu; Tao, Jianning; Lee, Brendan; Westbrook, Thomas F.; Wong, Stephen T. C.; Jin, Xin; Rosen, Jeffrey M.; Osborne, C. Kent; Zhang, Xiang H.-F.

2014-01-01

Summary Breast cancer bone micrometastases can remain asymptomatic for years before progressing into overt lesions. The biology of this process, including the microenvironment niche and supporting pathways, is unclear. We find that bone micrometastases predominantly reside in a niche that exhibits features of osteogenesis. Niche interactions are mediated by heterotypic adherens junctions (hAJs) involving cancer-derived E-cadherin and osteogenic N-cadherin, the disruption of which abolishes niche-conferred advantages. We further elucidate that hAJ activates the mTOR pathway in cancer cells, which drives the progression from single cells to micrometastases. Human datasets analyses support the roles of AJ and the mTOR pathway in bone colonization. Our study illuminates the initiation of bone colonization, and provides potential therapeutic targets to block progression toward osteolytic metastases. Significance In advanced stages, breast cancer bone metastases are driven by paracrine crosstalk among cancer cells, osteoblasts, and osteoclasts, which constitute a vicious osteolytic cycle. Current therapies targeting this process limit tumor progression, but do not improve patient survival. On the other hand, bone micrometastases may remain indolent for years before activating the vicious cycle, providing a therapeutic opportunity to prevent macrometastases. Here, we show that bone colonization is initiated in a microenvironment niche exhibiting active osteogenesis. Cancer and osteogenic cells form heterotypic adherens junctions, which enhance mTOR activity and drive early-stage bone colonization prior to osteolysis. These results reveal a strong connection between osteogenesis and micrometastasis and suggest potential therapeutic targets to prevent bone macrometastases. PMID:25600338

Pro-Tumoral Inflammatory Myeloid Cells as Emerging Therapeutic Targets.

PubMed

Szebeni, Gabor J; Vizler, Csaba; Nagy, Lajos I; Kitajka, Klara; Puskas, Laszlo G

2016-11-23

Since the observation of Virchow, it has long been known that the tumor microenvironment constitutes the soil for the infiltration of inflammatory cells and for the release of inflammatory mediators. Under certain circumstances, inflammation remains unresolved and promotes cancer development. Here, we review some of these indisputable experimental and clinical evidences of cancer related smouldering inflammation. The most common myeloid infiltrate in solid tumors is composed of myeloid-derived suppressor cells (MDSCs) and tumor-associated macrophages (TAMs). These cells promote tumor growth by several mechanisms, including their inherent immunosuppressive activity, promotion of neoangiogenesis, mediation of epithelial-mesenchymal transition and alteration of cellular metabolism. The pro-tumoral functions of TAMs and MDSCs are further enhanced by their cross-talk offering a myriad of potential anti-cancer therapeutic targets. We highlight these main pro-tumoral mechanisms of myeloid cells and give a general overview of their phenotypical and functional diversity, offering examples of possible therapeutic targets. Pharmacological targeting of inflammatory cells and molecular mediators may result in therapies improving patient condition and prognosis. Here, we review experimental and clinical findings on cancer-related inflammation with a major focus on creating an inventory of current small molecule-based therapeutic interventions targeting cancer-related inflammatory cells: TAMs and MDSCs.

Endocytosis of Cytotoxic Granules Is Essential for Multiple Killing of Target Cells by T Lymphocytes.

PubMed

Chang, Hsin-Fang; Bzeih, Hawraa; Schirra, Claudia; Chitirala, Praneeth; Halimani, Mahantappa; Cordat, Emmanuelle; Krause, Elmar; Rettig, Jens; Pattu, Varsha

2016-09-15

CTLs are serial killers that kill multiple target cells via exocytosis of cytotoxic granules (CGs). CG exocytosis is tightly regulated and has been investigated in great detail; however, whether CG proteins are endocytosed following exocytosis and contribute to serial killing remains unknown. By using primary CTLs derived from a knock-in mouse of the CG membrane protein Synaptobrevin2, we show that CGs are endocytosed in a clathrin- and dynamin-dependent manner. Following acidification, endocytosed CGs are recycled through early and late, but not recycling endosomes. CGs are refilled with granzyme B at the late endosome stage and polarize to subsequent synapses formed between the CTL and new target cells. Importantly, inhibiting CG endocytosis in CTLs results in a significant reduction of their cytotoxic activity. Thus, our data demonstrate that continuous endocytosis of CG membrane proteins is a prerequisite for efficient serial killing of CTLs and identify key events in this process. Copyright © 2016 by The American Association of Immunologists, Inc.

Prodrug strategy for cancer cell-specific targeting: A recent overview.

PubMed

Zhang, Xian; Li, Xiang; You, Qidong; Zhang, Xiaojin

2017-10-20

The increasing development of targeted cancer therapy provides extensive possibilities in clinical trials, and numerous strategies have been explored. The prodrug is one of the most promising strategies in targeted cancer therapy to improve the selectivity and efficacy of cytotoxic compounds. Compared with normal tissues, cancer cells are characterized by unique aberrant markers, thus inactive prodrugs targeting these markers are excellent therapeutics to release active drugs, killing cancer cells without damaging normal tissues. In this review, we explore an integrated view of potential prodrugs applied in targeted cancer therapy based on aberrant cancer specific markers and some examples are provided for inspiring new ideas of prodrug strategy for cancer cell-specific targeting. Copyright © 2017. Published by Elsevier Masson SAS.

In Situ Target Engagement Studies in Adherent Cells.

PubMed

Axelsson, Hanna; Almqvist, Helena; Otrocka, Magdalena; Vallin, Michaela; Lundqvist, Sara; Hansson, Pia; Karlsson, Ulla; Lundbäck, Thomas; Seashore-Ludlow, Brinton

2018-04-20

A prerequisite for successful drugs is effective binding of the desired target protein in the complex environment of a living system. Drug-target engagement has typically been difficult to monitor in physiologically relevant models, and with current methods, especially, while maintaining spatial information. One recent technique for quantifying drug-target engagement is the cellular thermal shift assay (CETSA), in which ligand-induced protein stabilization is measured after a heat challenge. Here, we describe a CETSA protocol in live A431 cells for p38α (MAPK14), where remaining soluble protein is detected in situ, using high-content imaging in 384-well, microtiter plates. We validate this assay concept using a number of known p38α inhibitors and further demonstrate the potential of this technology for chemical probe and drug discovery purposes by performing a small pilot screen for novel p38α binders. Importantly, this protocol creates a workflow that is amenable to adherent cells in their native state and yields spatially resolved target engagement information measurable at the single-cell level.

Fragments of Target Cells are Internalized into Retroviral Envelope Protein-Expressing Cells during Cell-Cell Fusion by Endocytosis

PubMed Central

Izumida, Mai; Kamiyama, Haruka; Suematsu, Takashi; Honda, Eri; Koizumi, Yosuke; Yasui, Kiyoshi; Hayashi, Hideki; Ariyoshi, Koya; Kubo, Yoshinao

2016-01-01

Retroviruses enter into host cells by fusion between viral and host cell membranes. Retroviral envelope glycoprotein (Env) induces the membrane fusion, and also mediates cell-cell fusion. There are two types of cell-cell fusions induced by the Env protein. Fusion-from-within is induced by fusion between viral fusogenic Env protein-expressing cells and susceptible cells, and virions induce fusion-from-without by fusion between adjacent cells. Although entry of ecotropic murine leukemia virus (E-MLV) requires host cell endocytosis, the involvement of endocytosis in cell fusion is unclear. By fluorescent microscopic analysis of the fusion-from-within, we found that fragments of target cells are internalized into Env-expressing cells. Treatment of the Env-expressing cells with an endocytosis inhibitor more significantly inhibited the cell fusion than that of the target cells, indicating that endocytosis in Env-expressing cells is required for the cell fusion. The endocytosis inhibitor also attenuated the fusion-from-without. Electron microscopic analysis suggested that the membrane fusion resulting in fusion-from-within initiates in endocytic membrane dents. This study shows that two types of the viral cell fusion both require endocytosis, and provides the cascade of fusion-from-within. PMID:26834711

An early history of T cell-mediated cytotoxicity.

PubMed

Golstein, Pierre; Griffiths, Gillian M

2018-04-16

After 60 years of intense fundamental research into T cell-mediated cytotoxicity, we have gained a detailed knowledge of the cells involved, specific recognition mechanisms and post-recognition perforin-granzyme-based and FAS-based molecular mechanisms. What could not be anticipated at the outset was how discovery of the mechanisms regulating the activation and function of cytotoxic T cells would lead to new developments in cancer immunotherapy. Given the profound recent interest in therapeutic manipulation of cytotoxic T cell responses, it is an opportune time to look back on the early history of the field. This Timeline describes how the early findings occurred and eventually led to current therapeutic applications.

The therapeutic potential of cell cycle targeting in multiple myeloma.

PubMed

Maes, Anke; Menu, Eline; Veirman, Kim De; Maes, Ken; Vand Erkerken, Karin; De Bruyne, Elke

2017-10-27

Proper cell cycle progression through the interphase and mitosis is regulated by coordinated activation of important cell cycle proteins (including cyclin-dependent kinases and mitotic kinases) and several checkpoint pathways. Aberrant activity of these cell cycle proteins and checkpoint pathways results in deregulation of cell cycle progression, which is one of the key hallmarks of cancer. Consequently, intensive research on targeting these cell cycle regulatory proteins identified several candidate small molecule inhibitors that are able to induce cell cycle arrest and even apoptosis in cancer cells. Importantly, several of these cell cycle regulatory proteins have also been proposed as therapeutic targets in the plasma cell malignancy multiple myeloma (MM). Despite the enormous progress in the treatment of MM the past 5 years, MM still remains most often incurable due to the development of drug resistance. Deregulated expression of the cyclins D is observed in virtually all myeloma patients, emphasizing the potential therapeutic interest of cyclin-dependent kinase inhibitors in MM. Furthermore, other targets have also been identified in MM, such as microtubules, kinesin motor proteins, aurora kinases, polo-like kinases and the anaphase promoting complex/cyclosome. This review will provide an overview of the cell cycle proteins and checkpoint pathways deregulated in MM and discuss the therapeutic potential of targeting proteins or protein complexes involved in cell cycle control in MM.

Cell Death Pathways in Mutant Rhodopsin Rat Models Identifies Genotype-Specific Targets Controlling Retinal Degeneration.

PubMed

Viringipurampeer, Ishaq A; Gregory-Evans, Cheryl Y; Metcalfe, Andrew L; Bashar, Emran; Moritz, Orson L; Gregory-Evans, Kevin

2018-06-18

Retinitis pigmentosa (RP) is a group of inherited neurological disorders characterized by rod photoreceptor cell death, followed by secondary cone cell death leading to progressive blindness. Currently, there are no viable treatment options for RP. Due to incomplete knowledge of the molecular signaling pathways associated with RP pathogenesis, designing therapeutic strategies remains a challenge. In particular, preventing secondary cone photoreceptor cell loss is a key goal in designing potential therapies. In this study, we identified the main drivers of rod cell death and secondary cone loss in the transgenic S334ter rhodopsin rat model, tested the efficacy of specific cell death inhibitors on retinal function, and compared the effect of combining drugs to target multiple pathways in the S334ter and P23H rhodopsin rat models. The primary driver of early rod cell death in the S334ter model was a caspase-dependent process, whereas cone cell death occurred though RIP3-dependent necroptosis. In comparison, rod cell death in the P23H model was via necroptotic signaling, whereas cone cell loss occurred through inflammasome activation. Combination therapy of four drugs worked better than the individual drugs in the P23H model but not in the S334ter model. These differences imply that treatment modalities need to be tailored for each genotype. Taken together, our data demonstrate that rationally designed genotype-specific drug combinations will be an important requisite to effectively target primary rod cell loss and more importantly secondary cone survival.

Cancer stem cell-targeted therapeutics and delivery strategies.

PubMed

Ahmad, Gulzar; Amiji, Mansoor M

2017-08-01

Cancer initiating or stem cells (CSCs) are a small population of cells in the tumor mass, which have been reported to be present in different types of cancers. CSCs usually reside within the tumor and are responsible for reoccurrence of cancer. The imprecise, inaccessible nature and increased efflux of conventional therapeutic drugs make these cells resistant to drugs. We discuss the specific markers for identification of these cells, role of CSCs in chemotherapy resistance and use of different therapeutic means to target them, including elucidation of specific cell markers, exploitation of different signaling pathways and use of nanotechnology. Area covered: This review covers cancer stem cell signaling which are used by these cells to maintain their quiescence, stemness and resistant phenotype, distinct cell surface markers, contribution of these cells in drug resistance, inevitability to cure cancer and use of nanotechnology to overcome this hurdle. Expert opinion: Cancer stem cells are the main culprit of our failure to cure cancer. In order to cure cancer along with other cells types in cancer, cancer stem cells need to be targeted in the tumor bed. Nanotechnology solutions can facilitate clinical translation of the therapeutics along with other emerging technologies to cure cancer.

Targeting sortilin in immune cells reduces proinflammatory cytokines and atherosclerosis

PubMed Central

Mortensen, Martin B.; Kjolby, Mads; Gunnersen, Stine; Larsen, Jakob V.; Palmfeldt, Johan; Falk, Erling; Nykjaer, Anders; Bentzon, Jacob F.

2014-01-01

Genome-wide association studies have identified a link between genetic variation at the human chromosomal locus 1p13.3 and coronary artery disease. The gene encoding sortilin (SORT1) has been implicated as the causative gene within the locus, as sortilin regulates hepatic lipoprotein metabolism. Here we demonstrated that sortilin also directly affects atherogenesis, independent of its regulatory role in lipoprotein metabolism. In a mouse model of atherosclerosis, deletion of Sort1 did not alter plasma cholesterol levels, but reduced the development of both early and late atherosclerotic lesions. We determined that sortilin is a high-affinity receptor for the proinflammatory cytokines IL-6 and IFN-γ. Moreover, macrophages and Th1 cells (both of which mediate atherosclerotic plaque formation) lacking sortilin had reduced secretion of IL-6 and IFN-γ, but not of other measured cytokines. Transfer of sortilin-deficient BM into irradiated atherosclerotic mice reduced atherosclerosis and systemic markers of inflammation. Together, these data demonstrate that sortilin influences cytokine secretion and that targeting sortilin in immune cells attenuates inflammation and reduces atherosclerosis. PMID:25401472

Variations of target volume definition and daily target volume localization in stereotactic body radiotherapy for early-stage non–small cell lung cancer patients under abdominal compression

DOE Office of Scientific and Technical Information (OSTI.GOV)

Han, Chunhui, E-mail: chan@coh.org; Sampath, Sagus; Schultheisss, Timothy E.

We aimed to compare gross tumor volumes (GTV) in 3-dimensional computed tomography (3DCT) simulation and daily cone beam CT (CBCT) with the internal target volume (ITV) in 4-dimensional CT (4DCT) simulation in stereotactic body radiotherapy (SBRT) treatment of patients with early-stage non–small cell lung cancer (NSCLC) under abdominal compression. We retrospectively selected 10 patients with NSCLC who received image-guided SBRT treatments under abdominal compression with daily CBCT imaging. GTVs were contoured as visible gross tumor on the planning 3DCT and daily CBCT, and ITVs were contoured using maximum intensity projection (MIP) images of the planning 4DCT. Daily CBCTs were registeredmore » with 3DCT and MIP images by matching of bony landmarks in the thoracic region to evaluate interfractional GTV position variations. Relative to MIP-based ITVs, the average 3DCT-based GTV volume was 66.3 ± 17.1% (range: 37.5% to 92.0%) (p < 0.01 in paired t-test), and the average CBCT-based GTV volume was 90.0 ± 6.7% (daily range: 75.7% to 107.1%) (p = 0.02). Based on bony anatomy matching, the center-of-mass coordinates for CBCT-based GTVs had maximum absolute shift of 2.4 mm (left-right), 7.0 mm (anterior-posterior [AP]), and 5.2 mm (superior-inferior [SI]) relative to the MIP-based ITV. CBCT-based GTVs had average overlapping ratio of 81.3 ± 11.2% (range: 45.1% to 98.9%) with the MIP-based ITV, and 57.7 ± 13.7% (range: 35.1% to 83.2%) with the 3DCT-based GTV. Even with abdominal compression, both 3DCT simulations and daily CBCT scans significantly underestimated the full range of tumor motion. In daily image-guided patient setup corrections, automatic bony anatomy-based image registration could lead to target misalignment. Soft tissue-based image registration should be performed for accurate treatment delivery.« less

Pros and Cons of Antigen-Presenting Cell Targeted Tumor Vaccines.

PubMed

Goyvaerts, Cleo; Breckpot, Karine

2015-01-01

In therapeutic antitumor vaccination, dendritic cells play the leading role since they decide if, how, when, and where a potent antitumor immune response will take place. Since the disentanglement of the complexity and merit of different antigen-presenting cell subtypes, antitumor immunotherapeutic research started to investigate the potential benefit of targeting these subtypes in situ. This review will discuss which antigen-presenting cell subtypes are at play and how they have been targeted and finally question the true meaning of targeting antitumor-based vaccines.

Purification-Free, Target-Selective Immobilization of a Protein from Cell Lysates.

PubMed

Cha, Jaehyun; Kwon, Inchan

2018-02-27

Protein immobilization has been widely used for laboratory experiments and industrial processes. Preparation of a recombinant protein for immobilization usually requires laborious and expensive purification steps. Here, a novel purification-free, target-selective immobilization technique of a protein from cell lysates is reported. Purification steps are skipped by immobilizing a target protein containing a clickable non-natural amino acid (p-azidophenylalanine) in cell lysates onto alkyne-functionalized solid supports via bioorthogonal azide-alkyne cycloaddition. In order to achieve a target protein-selective immobilization, p-azidophenylalanine was introduced into an exogenous target protein, but not into endogenous non-target proteins using host cells with amber codon-free genomic DNAs. Immobilization of superfolder fluorescent protein (sfGFP) from cell lysates is as efficient as that of the purified sfGFP. Using two fluorescent proteins (sfGFP and mCherry), the authors also demonstrated that the target proteins are immobilized with a minimal immobilization of non-target proteins (target-selective immobilization). © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

HIV-1 Nef sequesters MHC-I intracellularly by targeting early stages of endocytosis and recycling

PubMed Central

Dirk, Brennan S.; Pawlak, Emily N.; Johnson, Aaron L.; Van Nynatten, Logan R.; Jacob, Rajesh A.; Heit, Bryan; Dikeakos, Jimmy D.

2016-01-01

A defining characteristic of HIV-1 infection is the ability of the virus to persist within the host. Specifically, MHC-I downregulation by the HIV-1 accessory protein Nef is of critical importance in preventing infected cells from cytotoxic T-cell mediated killing. Nef downregulates MHC-I by modulating the host membrane trafficking machinery, resulting in the endocytosis and eventual sequestration of MHC-I within the cell. In the current report, we utilized the intracellular protein-protein interaction reporter system, bimolecular fluorescence complementation (BiFC), in combination with super-resolution microscopy, to track the Nef/MHC-I interaction and determine its subcellular localization in cells. We demonstrate that this interaction occurs upon Nef binding the MHC-I cytoplasmic tail early during endocytosis in a Rab5-positive endosome. Disruption of early endosome regulation inhibited Nef-dependent MHC-I downregulation, demonstrating that Nef hijacks the early endosome to sequester MHC-I within the cell. Furthermore, super-resolution imaging identified that the Nef:MHC-I BiFC complex transits through both early and late endosomes before ultimately residing at the trans-Golgi network. Together we demonstrate the importance of the early stages of the endocytic network in the removal of MHC-I from the cell surface and its re-localization within the cell, which allows HIV-1 to optimally evade host immune responses. PMID:27841315

Ion mediated targeting of cells with nanoparticles

NASA Astrophysics Data System (ADS)

Maheshwari, Vivek; Fu, Jinlong

2010-03-01

In eukaryotic cells, Ca^2+ ions are necessary for intracellular signaling, in activity of mitochondria and a variety of other cellular process that have been linked to cell apoptosis, proteins synthesis and cell-cycle regulation. Here we show that Ca^2+ ions, serving as the bio-compatible interface can be used to target Saccharomyces cerevisiae (SaC, baker's yeast), a model eukaryotic cell, with Au nanoparticles (10 nm). The Ca^2+ ions bind to the carboxylic acid groups in the citrate functionalized Au nanoparticles. This transforms the nanoparticles into micron long 1-D branched chain assemblies due to inter-particle dipole-dipole interaction and inter-particle bonding due to the divalent nature of the Ca^2+ ion. A similar transformation is observed with the use of divalent ions Mg^2+, Cd^2+ and Fe^2+. The 1-D assembly aids the interfacing of ion-nanoparticles on the cell by providing multiple contact points. Further monovalent ions such as Na^+ are also effective for the targeting of the cell with nanoparticles. However Na-Au nanoparticles are limited in their deposition as they exist in solution as single particles. The cells remain alive after the deposition process and their vitality is unaffected by the interfacing with ion-nanoparticles.

T-REX on-demand redox targeting in live cells.

PubMed

Parvez, Saba; Long, Marcus J C; Lin, Hong-Yu; Zhao, Yi; Haegele, Joseph A; Pham, Vanha N; Lee, Dustin K; Aye, Yimon

2016-12-01

This protocol describes targetable reactive electrophiles and oxidants (T-REX)-a live-cell-based tool designed to (i) interrogate the consequences of specific and time-resolved redox events, and (ii) screen for bona fide redox-sensor targets. A small-molecule toolset comprising photocaged precursors to specific reactive redox signals is constructed such that these inert precursors specifically and irreversibly tag any HaloTag-fused protein of interest (POI) in mammalian and Escherichia coli cells. Syntheses of the alkyne-functionalized endogenous reactive signal 4-hydroxynonenal (HNE(alkyne)) and the HaloTag-targetable photocaged precursor to HNE(alkyne) (also known as Ht-PreHNE or HtPHA) are described. Low-energy light prompts photo-uncaging (t 1/2 <1-2 min) and target-specific modification. The targeted modification of the POI enables precisely timed and spatially controlled redox events with no off-target modification. Two independent pathways are described, along with a simple setup to functionally validate known targets or discover novel sensors. T-REX sidesteps mixed responses caused by uncontrolled whole-cell swamping with reactive signals. Modification and downstream response can be analyzed by in-gel fluorescence, proteomics, qRT-PCR, immunofluorescence, fluorescence resonance energy transfer (FRET)-based and dual-luciferase reporters, or flow cytometry assays. T-REX targeting takes 4 h from initial probe treatment. Analysis of targeted redox responses takes an additional 4-24 h, depending on the nature of the pathway and the type of readouts used.

T-REX on-demand redox targeting in live cells

PubMed Central

Parvez, Saba; Long, Marcus J C; Lin, Hong-Yu; Zhao, Yi; Haegele, Joseph A; Pham, Vanha N; Lee, Dustin K; Aye, Yimon

2017-01-01

This protocol describes targetable reactive electrophiles and oxidants (T-REX)—a live-cell-based tool designed to (i) interrogate the consequences of specific and time-resolved redox events, and (ii) screen for bona fide redox-sensor targets. A small-molecule toolset comprising photocaged precursors to specific reactive redox signals is constructed such that these inert precursors specifically and irreversibly tag any HaloTag-fused protein of interest (POI) in mammalian and Escherichia coli cells. Syntheses of the alkyne-functionalized endogenous reactive signal 4-hydroxynonenal (HNE (alkyne)) and the HaloTag-targetable photocaged precursor to HNE (alkyne) (also known as Ht-PreHNE or HtPHA) are described. Low-energy light prompts photo-uncaging (t1/2 <1–2 min) and target-specific modification. The targeted modification of the POI enables precisely timed and spatially controlled redox events with no off-target modification. Two independent pathways are described, along with a simple setup to functionally validate known targets or discover novel sensors. T-REX sidesteps mixed responses caused by uncontrolled whole-cell swamping with reactive signals. Modification and downstream response can be analyzed by in-gel fluorescence, proteomics, qRT-PCR, immunofluorescence, fluorescence resonance energy transfer (FRET)-based and dual-luciferase reporters, or flow cytometry assays. T-REX targeting takes 4 h from initial probe treatment. Analysis of targeted redox responses takes an additional 4–24 h, depending on the nature of the pathway and the type of readouts used. PMID:27809314

Landscape phages and their fusion proteins targeted to breast cancer cells

PubMed Central

Fagbohun, Olusegun A.; Bedi, Deepa; Grabchenko, Natalia I.; Deinnocentes, Patricia A.; Bird, Richard C.; Petrenko, Valery A.

2012-01-01

Breast cancer is a leading cause of death among women in the USA. The efficacy of existing anticancer therapeutics can be improved by targeting them through conjugation with ligands binding to cellular receptors. Recently, we developed a novel drug targeting strategy based on the use of pre-selected cancer-specific ‘fusion pVIII proteins’ (fpVIII), as targeting ligands. To study the efficiency of this approach in animal models, we developed a panel of breast cancer cell-binding phages as a source of targeted fpVIIIs. Two landscape phage peptide libraries (8-mer f8/8 and 9-mer f8/9) were screened to isolate 132 phage variants that recognize breast carcinoma cells MCF-7 and ZR-75-1 and internalize into the cells. When tested for their interaction with the breast cancer cells in comparison with liver cancer cells HepG2, human mammary cells MCF-10A cells and serum, 16 of the phage probes selectively interacted with the breast cancer cells whereas 32 bound both breast and liver cancer cells. The most prominent cancer-specific phage DMPGTVLP, demonstrating sub-nanomolar Kd in interaction with target cells, was used for affinity chromatography of cellular membrane molecules to reveal its potential binding receptor. The isolated protein was identified by direct sequencing as cellular surface nucleolin. This conclusion was confirmed by inhibition of the phage–cell interaction with nucleolin antibodies. Other prominent phage binders VPTDTDYS, VEEGGYIAA, and DWRGDSMDS demonstrate consensus motifs common to previously identified cancer-specific peptides. Isolated phage proteins exhibit inherent binding specificity towards cancer cells, demonstrating the functional activity of the selected fused peptides. The selected phages, their peptide inserts and intact fusion proteins can serve as promising ligands for the development of targeted nanomedicines and their study in model mice with xenograft of human cells MCF-7 and ZR-75-1. PMID:22490956

Modular cell-internalizing aptamer nanostructure enables targeted delivery of large functional RNAs in cancer cell lines.

PubMed

Porciani, David; Cardwell, Leah N; Tawiah, Kwaku D; Alam, Khalid K; Lange, Margaret J; Daniels, Mark A; Burke, Donald H

2018-06-11

Large RNAs and ribonucleoprotein complexes have powerful therapeutic potential, but effective cell-targeted delivery tools are limited. Aptamers that internalize into target cells can deliver siRNAs (<15 kDa, 19-21 nt/strand). We demonstrate a modular nanostructure for cellular delivery of large, functional RNA payloads (50-80 kDa, 175-250 nt) by aptamers that recognize multiple human B cell cancer lines and transferrin receptor-expressing cells. Fluorogenic RNA reporter payloads enable accelerated testing of platform designs and rapid evaluation of assembly and internalization. Modularity is demonstrated by swapping in different targeting and payload aptamers. Both modules internalize into leukemic B cell lines and remained colocalized within endosomes. Fluorescence from internalized RNA persists for ≥2 h, suggesting a sizable window for aptamer payloads to exert influence upon targeted cells. This demonstration of aptamer-mediated, cell-internalizing delivery of large RNAs with retention of functional structure raises the possibility of manipulating endosomes and cells by delivering large aptamers and regulatory RNAs.

Dictyostelium cell death: early emergence and demise of highly polarized paddle cells.

PubMed

Levraud, Jean-Pierre; Adam, Myriam; Luciani, Marie-Françoise; de Chastellier, Chantal; Blanton, Richard L; Golstein, Pierre

2003-03-31

Cell death in the stalk of Dictyostelium discoideum, a prototypic vacuolar cell death, can be studied in vitro using cells differentiating as a monolayer. To identify early events, we examined potentially dying cells at a time when the classical signs of Dictyostelium cell death, such as heavy vacuolization and membrane lesions, were not yet apparent. We observed that most cells proceeded through a stereotyped series of differentiation stages, including the emergence of "paddle" cells showing high motility and strikingly marked subcellular compartmentalization with actin segregation. Paddle cell emergence and subsequent demise with paddle-to-round cell transition may be critical to the cell death process, as they were contemporary with irreversibility assessed through time-lapse videos and clonogenicity tests. Paddle cell demise was not related to formation of the cellulose shell because cells where the cellulose-synthase gene had been inactivated underwent death indistinguishable from that of parental cells. A major subcellular alteration at the paddle-to-round cell transition was the disappearance of F-actin. The Dictyostelium vacuolar cell death pathway thus does not require cellulose synthesis and includes early actin rearrangements (F-actin segregation, then depolymerization), contemporary with irreversibility, corresponding to the emergence and demise of highly polarized paddle cells.

Pseudomonas aeruginosa cells attached to a surface display a typical proteome early as 20 minutes of incubation

PubMed Central

Crouzet, Marc; Claverol, Stéphane; Lomenech, Anne-Marie; Le Sénéchal, Caroline; Costaglioli, Patricia; Barthe, Christophe; Garbay, Bertrand; Bonneu, Marc

2017-01-01

Biofilms are present in all environments and often result in negative effects due to properties of the biofilm lifestyle and especially antibiotics resistance. Biofilms are associated with chronic infections. Controlling bacterial attachment, the first step of biofilm formation, is crucial for fighting against biofilm and subsequently preventing the persistence of infection. Thus deciphering the underlying molecular mechanisms involved in attachment could allow discovering molecular targets from it would be possible to develop inhibitors against bacterial colonization and potentiate antibiotherapy. To identify the key components and pathways that aid the opportunistic pathogen Pseudomonas aeruginosa in attachment we performed for the first time a proteomic analysis as early as after 20 minutes of incubation using glass wool fibers as a surface. We compared the protein contents of the attached and unattached bacteria. Using mass spectrometry, 3043 proteins were identified. Our results showed that, as of 20 minutes of incubation, using stringent quantification criteria 616 proteins presented a modification of their abundance in the attached cells compared to their unattached counterparts. The attached cells presented an overall reduced gene expression and characteristics of slow-growing cells. The over-accumulation of outer membrane proteins, periplasmic folding proteins and O-antigen chain length regulators was also observed, indicating a profound modification of the cell envelope. Consistently the sigma factor AlgU required for cell envelope homeostasis was highly over-accumulated in attached cells. In addition our data suggested a role of alarmone (p)ppGpp and polyphosphate during the early attachment phase. Furthermore, almost 150 proteins of unknown function were differentially accumulated in the attached cells. Our proteomic analysis revealed the existence of distinctive biological features in attached cells as early as 20 minutes of incubation. Analysis of

The cancer cell adhesion resistome: mechanisms, targeting and translational approaches.

PubMed

Dickreuter, Ellen; Cordes, Nils

2017-06-27

Cell adhesion-mediated resistance limits the success of cancer therapies and is a great obstacle to overcome in the clinic. Since the 1990s, where it became clear that adhesion of tumor cells to the extracellular matrix is an important mediator of therapy resistance, a lot of work has been conducted to understand the fundamental underlying mechanisms and two paradigms were deduced: cell adhesion-mediated radioresistance (CAM-RR) and cell adhesion-mediated drug resistance (CAM-DR). Preclinical work has evidently demonstrated that targeting of integrins, adapter proteins and associated kinases comprising the cell adhesion resistome is a promising strategy to sensitize cancer cells to both radiotherapy and chemotherapy. Moreover, the cell adhesion resistome fundamentally contributes to adaptation mechanisms induced by radiochemotherapy as well as molecular drugs to secure a balanced homeostasis of cancer cells for survival and growth. Intriguingly, this phenomenon provides a basis for synthetic lethal targeted therapies simultaneously administered to standard radiochemotherapy. In this review, we summarize current knowledge about the cell adhesion resistome and highlight targeting strategies to override CAM-RR and CAM-DR.

Restricted T cell receptor repertoire in CLL-like monoclonal B cell lymphocytosis and early stage CLL.

PubMed

Blanco, Gonzalo; Vardi, Anna; Puiggros, Anna; Gómez-Llonín, Andrea; Muro, Manuel; Rodríguez-Rivera, María; Stalika, Evangelia; Abella, Eugenia; Gimeno, Eva; López-Sánchez, Manuela; Senín, Alicia; Calvo, Xavier; Abrisqueta, Pau; Bosch, Francesc; Ferrer, Ana; Stamatopoulos, Kostas; Espinet, Blanca

2018-01-01

Analysis of the T cell receptor (TR) repertoire of chronic lymphocytic leukemia-like monoclonal B cell lymphocytosis (CLL-like MBL) and early stage CLL is relevant for understanding the dynamic interaction of expanded B cell clones with bystander T cells. Here we profiled the T cell receptor β chain (TRB) repertoire of the CD4 + and CD8 + T cell fractions from 16 CLL-like MBL and 13 untreated, Binet stage A/Rai stage 0 CLL patients using subcloning analysis followed by Sanger sequencing. The T cell subpopulations of both MBL and early stage CLL harbored restricted TRB gene repertoire, with CD4 + T cell clonal expansions whose frequency followed the numerical increase of clonal B cells. Longitudinal analysis in MBL cases revealed clonal persistence, alluding to persistent antigen stimulation. In addition, the identification of shared clonotypes among different MBL/early stage CLL cases pointed towards selection of the T cell clones by common antigenic elements. T cell clonotypes previously described in viral infections and immune disorders were also detected. Altogether, our findings evidence that antigen-mediated TR restriction occurs early in clonal evolution leading to CLL and may further increase together with B cell clonal expansion, possibly suggesting that the T cell selecting antigens are tumor-related.

Application of stem cells in targeted therapy of breast cancer: a systematic review.

PubMed

Madjd, Zahra; Gheytanchi, Elmira; Erfani, Elham; Asadi-Lari, Mohsen

2013-01-01

The aim of this systematic review was to investigate whether stem cells could be effectively applied in targeted therapy of breast cancer. A systematic literature search was performed for original articles published from January 2007 until May 2012. Nine studies met the inclusion criteria for phase I or II clinical trials, of which three used stem cells as vehicles, two trials used autologous hematopoetic stem cells and in four trials cancer stem cells were targeted. Mesenchymal stem cells (MSCs) were applied as cellular vehicles to transfer therapeutic agents. Cell therapy with MSC can successfully target resistant cancers. Cancer stem cells were selectively targeted via a proteasome-dependent suicide gene leading to tumor regression. Wnt/β-catenin signaling pathway has been also evidenced to be an attractive CSC-target. This systematic review focused on two different concepts of stem cells and breast cancer marking a turning point in the trials that applied stem cells as cellular vehicles for targeted delivery therapy as well as CSC-targeted therapies. Applying stem cells as targeted therapy could be an effective therapeutic approach for treatment of breast cancer in the clinic and in therapeutic marketing; however this needs to be confirmed with further clinical investigations.

Targeting of non-dominant antigens as a vaccine strategy to broaden T-cell responses during chronic viral infection.

PubMed

Holst, Peter J; Jensen, Benjamin A H; Ragonnaud, Emeline; Thomsen, Allan R; Christensen, Jan P

2015-01-01

In this study, we compared adenoviral vaccine vectors with the capacity to induce equally potent immune responses against non-dominant and immunodominant epitopes of murine lymphocytic choriomeningitis virus (LCMV). Our results demonstrate that vaccination targeting non-dominant epitopes facilitates potent virus-induced T-cell responses against immunodominant epitopes during subsequent challenge with highly invasive virus. In contrast, when an immunodominant epitope was included in the vaccine, the T-cell response associated with viral challenge remained focussed on that epitope. Early after challenge with live virus, the CD8+ T cells specific for vaccine-encoded epitopes, displayed a phenotype typically associated with prolonged/persistent antigenic stimulation marked by high levels of KLRG-1, as compared to T cells reacting to epitopes not included in the vaccine. Notably, this association was lost over time in T cells specific for the dominant T cell epitopes, and these cells were fully capable of expanding in response to a new viral challenge. Overall, our data suggests a potential for broadening of the antiviral CD8+ T-cell response by selecting non-dominant antigens to be targeted by vaccination. In addition, our findings suggest that prior adenoviral vaccination is not likely to negatively impact the long-term and protective immune response induced and maintained by a vaccine-attenuated chronic viral infection.

Synchrotron FTIR microspectroscopy reveals early adipogenic differentiation of human mesenchymal stem cells at single-cell level

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liu, Zhixiao; University of Chinese Academy of Science, Beijing 100049; Tang, Yuzhao

Human mesenchymal stem cells (hMSCs) have been used as an ideal in vitro model to study human adipogenesis. However, little knowledge of the early stage differentiation greatly hinders our understanding on the mechanism of the adipogenesis processes. In this study, synchrotron radiation-based Fourier transform infrared (SR-FTIR) microspectroscopy was applied to track the global structural and compositional changes of lipids, proteins and nucleic acids inside individual hMSCs along the time course. The multivariate analysis of the SR-FTIR spectra distinguished the dynamic and significant changes of the lipids and nucleic acid at early differentiation stage. Importantly, changes of lipid structure during early daysmore » (Day 1–3) of differentiation might serve as a potential biomarker in identifying the state in early differentiation at single cell level. These results proved that SR-FTIR is a powerful tool to study the stem cell fate determination and early lipogenesis events. - Highlights: • Molecular events occur in the early adipogenic differentiation stage of hMSCs are studied by SR-FTIR. • SR-FTIR data suggest that lipids may play an important role in hMSCs determination. • As potential biomarkers, lipids peaks can identify the state of cell in early differentiation stage at single-cell level.« less

Concise Review: Emerging Drugs Targeting Epithelial Cancer Stem-Like Cells.

PubMed

Ahmed, Mehreen; Chaudhari, Kritika; Babaei-Jadidi, Roya; Dekker, Lodewijk V; Shams Nateri, Abdolrahman

2017-04-01

Increasing evidence suggests that cancer cell populations contain a small proportion of cells that display stem-like cell properties and which may be responsible for overall tumor maintenance. These cancer stem-like cells (CSCs) appear to have unique tumor-initiating ability and innate survival mechanisms that allow them to resist cancer therapies, consequently promoting relapses. Selective targeting of CSCs may provide therapeutic benefit and several recent reports have indicated this may be possible. In this article, we review drugs targeting CSCs, in selected epithelial cell-derived cancers. Stem Cells 2017;35:839-850. © 2017 AlphaMed Press.

MicroRNA-944 Affects Cell Growth by Targeting EPHA7 in Non-Small Cell Lung Cancer.

PubMed

Liu, Minxia; Zhou, Kecheng; Cao, Yi

2016-09-26

MicroRNAs (miRNAs) have critical roles in lung tumorigenesis and development. To determine aberrantly expressed miRNAs involved in non-small cell lung cancer (NSCLC) and investigate pathophysiological functions and mechanisms, we firstly carried out small RNA deep sequencing in NSCLC cell lines (EPLC-32M1, A549 and 801D) and a human immortalized cell line 16HBE, we then studied miRNA function by cell proliferation and apoptosis. cDNA microarray, luciferase reporter assay and miRNA transfection were used to investigate interaction between the miRNA and target gene. miR-944 was significantly down-regulated in NSCLC and had many putative targets. Moreover, the forced expression of miR-944 significantly inhibited the proliferation of NSCLC cells in vitro. By integrating mRNA expression data and miR-944-target prediction, we disclosed that EPHA7 was a potential target of miR-944, which was further verified by luciferase reporter assay and microRNA transfection. Our data indicated that miR-944 targets EPHA7 in NSCLC and regulates NSCLC cell proliferation, which may offer a new mechanism underlying the development and progression of NSCLC.

New Strategies in Engineering T-cell Receptor Gene-Modified T cells to More Effectively Target Malignancies.

PubMed

Schmitt, Thomas M; Stromnes, Ingunn M; Chapuis, Aude G; Greenberg, Philip D

2015-12-01

The immune system, T cells in particular, have the ability to target and destroy malignant cells. However, antitumor immune responses induced from the endogenous T-cell repertoire are often insufficient for the eradication of established tumors, as illustrated by the failure of cancer vaccination strategies or checkpoint blockade for most tumors. Genetic modification of T cells to express a defined T-cell receptor (TCR) can provide the means to rapidly generate large numbers of tumor-reactive T cells capable of targeting tumor cells in vivo. However, cell-intrinsic factors as well as immunosuppressive factors in the tumor microenvironment can limit the function of such gene-modified T cells. New strategies currently being developed are refining and enhancing this approach, resulting in cellular therapies that more effectively target tumors and that are less susceptible to tumor immune evasion. ©2015 American Association for Cancer Research.

Probing Xist RNA Structure in Cells Using Targeted Structure-Seq

PubMed Central

Rutenberg-Schoenberg, Michael; Simon, Matthew D.

2015-01-01

The long non-coding RNA (lncRNA) Xist is a master regulator of X-chromosome inactivation in mammalian cells. Models for how Xist and other lncRNAs function depend on thermodynamically stable secondary and higher-order structures that RNAs can form in the context of a cell. Probing accessible RNA bases can provide data to build models of RNA conformation that provide insight into RNA function, molecular evolution, and modularity. To study the structure of Xist in cells, we built upon recent advances in RNA secondary structure mapping and modeling to develop Targeted Structure-Seq, which combines chemical probing of RNA structure in cells with target-specific massively parallel sequencing. By enriching for signals from the RNA of interest, Targeted Structure-Seq achieves high coverage of the target RNA with relatively few sequencing reads, thus providing a targeted and scalable approach to analyze RNA conformation in cells. We use this approach to probe the full-length Xist lncRNA to develop new models for functional elements within Xist, including the repeat A element in the 5’-end of Xist. This analysis also identified new structural elements in Xist that are evolutionarily conserved, including a new element proximal to the C repeats that is important for Xist function. PMID:26646615

Cell targeting peptides as smart ligands for targeting of therapeutic or diagnostic agents: a systematic review.

PubMed

Mousavizadeh, Ali; Jabbari, Ali; Akrami, Mohammad; Bardania, Hassan

2017-10-01

Cell targeting peptides (CTP) are small peptides which have high affinity and specificity to a cell or tissue targets. They are typically identified by using phage display and chemical synthetic peptide library methods. CTPs have attracted considerable attention as a new class of ligands to delivery specifically therapeutic and diagnostic agents, because of the fact they have several advantages including easy synthesis, smaller physical sizes, lower immunogenicity and cytotoxicity and their simple and better conjugation to nano-carriers and therapeutic or diagnostic agents compared to conventional antibodies. In this systematic review, we will focus on the basic concepts concerning the use of cell-targeting peptides (CTPs), following the approaches of selecting them from peptide libraries. We discuss several developed strategies for cell-specific delivery of different cargos by CTPs, which are designed for drug delivery and diagnostic applications. Copyright © 2017 Elsevier B.V. All rights reserved.

Endothelial binding of beta toxin to small intestinal mucosal endothelial cells in early stages of experimentally induced Clostridium perfringens type C enteritis in pigs.

PubMed

Schumacher, V L; Martel, A; Pasmans, F; Van Immerseel, F; Posthaus, H

2013-07-01

Beta toxin (CPB) is known to be an essential virulence factor in the development of lesions of Clostridium perfringens type C enteritis in different animal species. Its target cells and exact mechanism of toxicity have not yet been clearly defined. Here, we evaluate the suitability of a neonatal piglet jejunal loop model to investigate early lesions of C. perfringens type C enteritis. Immunohistochemically, CPB was detected at microvascular endothelial cells in intestinal villi during early and advanced stages of lesions induced by C. perfringens type C. This was first associated with capillary dilatation and subsequently with widespread hemorrhage in affected intestinal segments. CPB was, however, not demonstrated on intestinal epithelial cells. This indicates a tropism of CPB toward endothelial cells and suggests that CPB-induced endothelial damage plays an important role in the early stages of C. perfringens type C enteritis in pigs.

Tumor-targeting domains for chimeric antigen receptor T cells.

PubMed

Bezverbnaya, Ksenia; Mathews, Ashish; Sidhu, Jesse; Helsen, Christopher W; Bramson, Jonathan L

2017-01-01

Immunotherapy with chimeric antigen receptor (CAR) T cells has been advancing steadily in clinical trials. Since the ability of engineered T cells to recognize intended tumor-associated targets is crucial for the therapeutic success, antigen-binding domains play an important role in shaping T-cell responses. Single-chain antibody and T-cell receptor fragments, natural ligands, repeat proteins, combinations of the above and universal tag-specific domains have all been used in the antigen-binding moiety of chimeric receptors. Here we outline the advantages and disadvantages of different domains, discuss the concepts of affinity and specificity, and highlight the recent progress of each targeting strategy.

Cytotoxic chemotherapy in the treatment of advanced renal cell carcinoma in the era of targeted therapy.

PubMed

Diamond, E; Molina, A M; Carbonaro, M; Akhtar, N H; Giannakakou, P; Tagawa, S T; Nanus, D M

2015-12-01

Renal cell carcinoma (RCC) is a heterogeneous disease with regards to histology, progression, and response to treatment. Cytotoxic chemotherapy has been extensively studied in metastatic RCC (mRCC). Responses in most studies are modest and the mechanisms of resistance remain poorly understood. Targeted therapies have significantly improved outcomes in mRCC; however, most patients eventually relapse and die of their disease. Early clinical data suggest that combinations of chemotherapy and targeted agents are clinically active and are well tolerated. We reviewed the available literature for published clinical trials incorporating traditional chemotherapeutic agents in the treatment of mRCC. These papers were identified through a Medline search and were included if they employed at least one chemotherapeutic agent in the treatment of mRCC. The literature was also reviewed for information regarding mechanisms of chemotherapy resistance. The data regarding the use of cytotoxic chemotherapy in mRCC consist of small, non-randomized phase I and II studies. The major response proportions with single agent chemotherapies are low but combination regimens either with other cytotoxic agents, cytokines, or targeted agents have demonstrated moderate activity. Disparate trial designs and lack of head to head clinical trials make it difficult to compare the efficacy of chemotherapy with that of immunotherapy or targeted agents. Chemotherapy is particularly useful in patients with collecting duct histology and predominantly sarcomatoid differentiation. Chemotherapy resistance may be mediated by overexpression of p-glycoprotein efflux pumps and the dysregulation of the microtubule-hypoxia inducible factor signaling axis. The role of cytotoxic chemotherapy in the treatment for clear cell RCC remains poorly defined. Cytotoxic chemotherapy is considered a standard of care in patients with mRCC with predominantly sarcomatoid differentiation and collecting duct RCC variants (Motzer et al

Optical cell monitoring system for underwater targets

NASA Astrophysics Data System (ADS)

Moon, SangJun; Manzur, Fahim; Manzur, Tariq; Demirci, Utkan

2008-10-01

We demonstrate a cell based detection system that could be used for monitoring an underwater target volume and environment using a microfluidic chip and charge-coupled-device (CCD). This technique allows us to capture specific cells and enumerate these cells on a large area on a microchip. The microfluidic chip and a lens-less imaging platform were then merged to monitor cell populations and morphologies as a system that may find use in distributed sensor networks. The chip, featuring surface chemistry and automatic cell imaging, was fabricated from a cover glass slide, double sided adhesive film and a transparent Polymethlymetacrylate (PMMA) slab. The optically clear chip allows detecting cells with a CCD sensor. These chips were fabricated with a laser cutter without the use of photolithography. We utilized CD4+ cells that are captured on the floor of a microfluidic chip due to the ability to address specific target cells using antibody-antigen binding. Captured CD4+ cells were imaged with a fluorescence microscope to verify the chip specificity and efficiency. We achieved 70.2 +/- 6.5% capturing efficiency and 88.8 +/- 5.4% specificity for CD4+ T lymphocytes (n = 9 devices). Bright field images of the captured cells in the 24 mm × 4 mm × 50 μm microfluidic chip were obtained with the CCD sensor in one second. We achieved an inexpensive system that rapidly captures cells and images them using a lens-less CCD system. This microfluidic device can be modified for use in single cell detection utilizing a cheap light-emitting diode (LED) chip instead of a wide range CCD system.

Biomarker evaluation of face transplant rejection: association of donor T cells with target cell injury.

PubMed

Lian, Christine Guo; Bueno, Ericka M; Granter, Scott R; Laga, Alvaro C; Saavedra, Arturo P; Lin, William M; Susa, Joseph S; Zhan, Qian; Chandraker, Anil K; Tullius, Stefan G; Pomahac, Bohdan; Murphy, George F

2014-06-01

This series of 113 sequential biopsies of full facial transplants provides findings of potential translational significance as well as biological insights that could prompt reexamination of conventional paradigms of effector pathways in skin allograft rejection. Serial biopsies before, during, and after rejection episodes were evaluated for clinicopathological assessment that in selected cases included specific biomarkers for donor-versus-recipient T cells. Histologic evidence of rejection included lymphocyte-associated injury to epidermal rete ridges, follicular infundibula, and dermal microvessels. Surprisingly, during active rejection, immune cells spatially associated with target cell injury consisted abundantly or predominantly of lymphocytes of donor origin with an immunophenotype typical of the resident memory T-cell subset. Current dogma assumes that skin allograft rejection is mediated by recipient T cells that attack epidermal targets, and the association of donor T cells with sites of target cell injury raises questions regarding the potential complexity of immune cell interactions in the rejection process. A more histopathologically refined and immune-based biomarker approach to assessment of rejection of facial transplants is now indicated.

Efficient Generation of Somatic Cell Nuclear Transfer-Competent Porcine Cells with Mutated Alleles at Multiple Target Loci by Using CRISPR/Cas9 Combined with Targeted Toxin-Based Selection System.

PubMed

Sato, Masahiro; Miyoshi, Kazuchika; Nakamura, Shingo; Ohtsuka, Masato; Sakurai, Takayuki; Watanabe, Satoshi; Kawaguchi, Hiroaki; Tanimoto, Akihide

2017-12-04

The recent advancement in genome editing such a CRISPR/Cas9 system has enabled isolation of cells with knocked multiple alleles through a one-step transfection. Somatic cell nuclear transfer (SCNT) has been frequently employed as one of the efficient tools for the production of genetically modified (GM) animals. To use GM cells as SCNT donor, efficient isolation of transfectants with mutations at multiple target loci is often required. The methods for the isolation of such GM cells largely rely on the use of drug selection-based approach using selectable genes; however, it is often difficult to isolate cells with mutations at multiple target loci. In this study, we used a novel approach for the efficient isolation of porcine cells with at least two target loci mutations by one-step introduction of CRISPR/Cas9-related components. A single guide (sg) RNA targeted to GGTA1 gene, involved in the synthesis of cell-surface α-Gal epitope (known as xenogenic antigen), is always a prerequisite. When the transfected cells were reacted with toxin-labeled BS-I-B₄ isolectin for 2 h at 37 C to eliminate α-Gal epitope-expressing cells, the surviving clones lacked α-Gal epitope expression and were highly expected to exhibit induced mutations at another target loci. Analysis of these α-Gal epitope-negative surviving cells demonstrated a 100% occurrence of genome editing at target loci. SCNT using these cells as donors resulted in the production of cloned blastocysts with the genotype similar to that of the donor cells used. Thus, this novel system will be useful for SCNT-mediated acquisition of GM cloned piglets, in which multiple target loci may be mutated.

Curcumin suppresses proliferation of colon cancer cells by targeting CDK2.

PubMed

Lim, Tae-Gyu; Lee, Sung-Young; Huang, Zunnan; Lim, Do Young; Chen, Hanyong; Jung, Sung Keun; Bode, Ann M; Lee, Ki Won; Dong, Zigang

2014-04-01

Curcumin, the yellow pigment of turmeric found in Southeast Indian food, is one of the most popular phytochemicals for cancer prevention. Numerous reports have demonstrated modulation of multiple cellular signaling pathways by curcumin and its molecular targets in various cancer cell lines. To identify a new molecular target of curcumin, we used shape screening and reverse docking to screen the Protein Data Bank against curcumin. Cyclin-dependent kinase 2 (CDK2), a major cell-cycle protein, was identified as a potential molecular target of curcumin. Indeed, in vitro and ex vivo kinase assay data revealed a dramatic suppressive effect of curcumin on CDK2 kinase activity. Furthermore, curcumin induced G1 cell-cycle arrest, which is regulated by CDK2 in HCT116 cells. Although the expression levels of CDK2 and its regulatory subunit, cyclin E, were not changed, the phosphorylation of retinoblastoma (Rb), a well-known CDK2 substrate, was reduced by curcumin. Because curcumin induced cell-cycle arrest, we investigated the antiproliferative effect of curcumin on HCT116 colon cancer cells. In this experiment, curcumin suppressed HCT116 cell proliferation effectively. To determine whether CDK2 is a direct target of curcumin, CDK2 expression was knocked down in HCT116 cells. As expected, HCT116 sh-CDK2 cells exhibited G1 arrest and reduced proliferation. Because of the low levels of CDK2 in HCT116 sh-CDK2 cells, the effects of curcumin on G1 arrest and cell proliferation were not substantially relative to HCT116 sh-control cells. From these results, we identified CDK2 as a direct target of curcumin in colon cancer cells.

Curcumin suppresses proliferation of colon cancer cells by targeting CDK2

PubMed Central

Lim, Tae-Gyu; Lee, Sung-Young; Huang, Zunnan; Lim, Do Young; Chen, Hanyong; Jung, Sung Keun; Bode, Ann M.; Lee, Ki Won; Dong, Zigang

2014-01-01

Curcumin, the yellow pigment of turmeric found in Southeast Indian food, is one of the most popular phytochemicals for cancer prevention. Numerous reports have demonstrated modulation of multiple cellular signaling pathways by curcumin and its molecular targets in various cancer cell lines. To identify a new molecular target of curcumin, we used shape screening and reverse docking to screen the protein data bank against curcumin. Cyclin dependent kinase 2 (CDK2), a major cell cycle protein, was identified as a potential molecular target of curcumin. Indeed, in vitro and ex vivo kinase assay data revealed a dramatic suppressive effect of curcumin on CDK2 kinase activity. Furthermore, curcumin induced G1 cell cycle arrest, which is regulated by CDK2 in HCT116 cells. Although the expression levels of CDK2 and its regulatory subunit, cyclin E, were not changed, the phosphorylation of Rb, a well-known CDK2 substrate, was reduced by curcumin. Because curcumin induced cell cycle arrest, we investigated the anti-proliferative effect of curcumin on HCT116 colon cancer cells. In this experiment, curcumin suppressed HCT116 cell proliferation effectively. To determine if CDK2 is a direct target of curcumin, CDK2 expression was knocked down in HCT116 cells. As expected, HCT116 sh-CDK2 cells exhibited G1 arrest and reduced proliferation. Because of the low levels of CDK2 in HCT116 sh-CDK2 cells, the effects of curcumin on G1 arrest and cell proliferation were not substantial relative to HCT116 sh-control cells. From these results, we identified CDK2 as a direct target of curcumin in colon cancer cells. PMID:24550143

Enhancing Oral Vaccine Potency by Targeting Intestinal M Cells

PubMed Central

Azizi, Ali; Kumar, Ashok; Diaz-Mitoma, Francisco; Mestecky, Jiri

2010-01-01

The immune system in the gastrointestinal tract plays a crucial role in the control of infection, as it constitutes the first line of defense against mucosal pathogens. The attractive features of oral immunization have led to the exploration of a variety of oral delivery systems. However, none of these oral delivery systems have been applied to existing commercial vaccines. To overcome this, a new generation of oral vaccine delivery systems that target antigens to gut-associated lymphoid tissue is required. One promising approach is to exploit the potential of microfold (M) cells by mimicking the entry of pathogens into these cells. Targeting specific receptors on the apical surface of M cells might enhance the entry of antigens, initiating the immune response and consequently leading to protection against mucosal pathogens. In this article, we briefly review the challenges associated with current oral vaccine delivery systems and discuss strategies that might potentially target mouse and human intestinal M cells. PMID:21085599

Microcinematographic and electron microscopic analysis of target cell lysis induced by cytotoxic T lymphocytes.

PubMed Central

Matter, A

1979-01-01

A study was carried out to determine the sequence of events of T-cell mediated target cell lysis in microcinematography and electron microscopy. Highly efficient cytotoxic T lymphocytes (CTL) were generated in vivo and in vitro using preimmunized spleen cells and purification procedures. Such CTL were highly specific. This specificity correlated well with the number of adhesions formed between CTL and targets and this criterion was used to study killer-target cell interaction. Microcinematography showed that target cell lysis at the single cell level, despite time variations, could be clearly separated into three phases: (a) a recognition phase, visible by random crawling of CTL over the target cell surface until firm contact was established; (b) a post-recognition phase, during which firm contact between CTL and target was maintained without gross modification of either cell; (c) a phase of target cell disintegration, mainly characterized by vigorous blebbing of the cell membrane resulting in a motionless carcass of the target cell but not in its total dissolution. Only later this carcass decayed and formed a necrotic ghost. Electron microscopic observations were put into sequence according to microcinematography. Post-recognition phase was characterized by a tight apposition of the membranes of CTL and target cell. No gap junctions could be observed. During target cell disintegration, profound cytoplasmic and nuclear changes occurred simultaneous with surface blebbing. Most noticeable were extensive internal vacuolization, mitochondrial swelling, nuclear pycnosis and dissolution of the nucleolus. These observations suggested that target cell lysis does not start with a surface phenomenon similar to complement lysis, but a process involving practically the whole cell simultaneously. It is conceivable, therefore, that the signal from the CTL is transmitted across the target cell, and that the switch to sudden cell death is manipulated deep inside the cell. Images

Colon-targeted delivery of live bacterial cell biotherapeutics including microencapsulated live bacterial cells

PubMed Central

Prakash, Satya; Malgorzata Urbanska, Aleksandra

2008-01-01

There has been an ample interest in delivery of therapeutic molecules using live cells. Oral delivery has been stipulated as best way to deliver live cells to humans for therapy. Colon, in particular, is a part of gastrointestinal (GI) tract that has been proposed to be an oral targeted site. The main objective of these oral therapy procedures is to deliver live cells not only to treat diseases like colorectal cancer, inflammatory bowel disease, and other GI tract diseases like intestinal obstruction and gastritis, but also to deliver therapeutic molecules for overall therapy in various diseases such as renal failure, coronary heart disease, hypertension, and others. This review provides a comprehensive summary of recent advancement in colon targeted live bacterial cell biotherapeutics. Current status of bacterial cell therapy, principles of artificial cells and its potentials in oral delivery of live bacterial cell biotherapeutics for clinical applications as well as biotherapeutic future perspectives are also discussed in our review. PMID:19707368

Meningeal mast cells affect early T cell central nervous system infiltration and blood-brain barrier integrity through TNF: a role for neutrophil recruitment?

PubMed

Sayed, Blayne A; Christy, Alison L; Walker, Margaret E; Brown, Melissa A

2010-06-15

Mast cells contribute to the pathogenesis of experimental autoimmune encephalomyelitis, a rodent model of the human demyelinating disease multiple sclerosis. Yet their site and mode of action is unknown. In both diseases, myelin-specific T cells are initially activated in peripheral lymphoid organs. However, for disease to occur, these cells must enter the immunologically privileged CNS through a breach in the relatively impermeable blood-brain barrier. In this study, we demonstrate that a dense population of resident mast cells in the meninges, structures surrounding the brain and spinal cord, regulate basal CNS barrier function, facilitating initial T cell CNS entry. Through the expression of TNF, mast cells recruit an early wave of neutrophils to the CNS. We propose that neutrophils in turn promote the blood-brain barrier breach and together with T cells lead to further inflammatory cell influx and myelin damage. These findings provide specific targets for intervention in multiple sclerosis as well as other immune-mediated CNS diseases.

Targeting Notch signalling pathway of cancer stem cells.

PubMed

Venkatesh, Vandana; Nataraj, Raghu; Thangaraj, Gopenath S; Karthikeyan, Murugesan; Gnanasekaran, Ashok; Kaginelli, Shanmukhappa B; Kuppanna, Gobianand; Kallappa, Chandrashekrappa Gowdru; Basalingappa, Kanthesh M

2018-01-01

Cancer stem cells (CSCs) have been defined as cells within tumor that possess the capacity to self-renew and to cause the heterogeneous lineages of cancer cells that comprise the tumor. CSCs have been increasingly identified in blood cancer, prostate, ovarian, lung, melanoma, pancreatic, colon, brain and many more malignancies. CSCs have slow growth rate and are resistant to chemotherapy and radiotherapy that lead to the failure of traditional current therapy. Eradicating the CSCs and recurrence, is promising aspect for the cure of cancer. The CSCs like any other stem cells activate the signal transduction pathways that involve the development and tissue homeostasis, which include Notch signaling pathway. The new treatment targets these pathway that control stem-cell replication, survival and differentiation that are under development. Notch inhibitors either single or in combination with chemotherapy drugs have been developed to treat cancer and its recurrence. This approach of targeting signaling pathway of CSCs represents a promising future direction for the therapeutic strategy to cure cancer.

Tuberin and PRAS40 are anti-apoptotic gatekeepers during early human amniotic fluid stem-cell differentiation.

PubMed

Fuchs, Christiane; Rosner, Margit; Dolznig, Helmut; Mikula, Mario; Kramer, Nina; Hengstschläger, Markus

2012-03-01

Embryoid bodies (EBs) are three-dimensional multicellular aggregates allowing the in vitro investigation of stem-cell differentiation processes mimicking early embryogenesis. Human amniotic fluid stem (AFS) cells harbor high proliferation potential, do not raise the ethical issues of embryonic stem cells, have a lower risk for tumor development, do not need exogenic induction of pluripotency and are chromosomal stable. Starting from a single human AFS cell, EBs can be formed accompanied by the differentiation into cells of all three embryonic germ layers. Here, we report that siRNA-mediated knockdown of the endogenous tuberous sclerosis complex-2 (TSC2) gene product tuberin or of proline-rich Akt substrate of 40 kDa (PRAS40), the two major negative regulators of mammalian target of rapamycin (mTOR), leads to massive apoptotic cell death during EB development of human AFS cells without affecting the endodermal, mesodermal and ectodermal cell differentiation spectrum. Co-knockdown of endogenous mTOR demonstrated these effects to be mTOR-dependent. Our findings prove this enzyme cascade to be an essential anti-apoptotic gatekeeper of stem-cell differentiation during EB formation. These data allow new insights into the regulation of early stem-cell maintenance and differentiation and identify a new role of the tumor suppressor tuberin and the oncogenic protein PRAS40 with the relevance for a more detailed understanding of the pathogenesis of diseases associated with altered activities of these gene products.

Adaptive early detection ML/PDA estimator for LO targets with EO sensors

NASA Astrophysics Data System (ADS)

Chummun, Muhammad R.; Kirubarajan, Thiagalingam; Bar-Shalom, Yaakov

2000-07-01

The batch Maximum Likelihood Estimator, combined with Probabilistic Data (ML-PDA), has been shown to be effective in acquiring low observable (LO) - low SNR - non-maneuvering targets in the presence of heavy clutter. The use of signal strength or amplitude information (AI) in the ML-PDA estimator with AI in a sliding-window fashion, to detect high- speed targets in heavy clutter using electro-optical (EO) sensors. The initial time and the length of the sliding-window are adjusted adaptively according to the information content of the received measurements. A track validation scheme via hypothesis testing is developed to confirm the estimated track, that is, the presence of a target, in each window. The sliding-window ML-PDA approach, together with track validation, enables early detection by rejecting noninformative scans, target reacquisition in case of temporary target disappearance and the handling of targets with speeds evolving over time. The proposed algorithm is shown to detect the target, which is hidden in as many as 600 false alarms per scan, 10 frames earlier than the Multiple Hypothesis Tracking (MHT) algorithm.

MicroRNA-93 inhibits tumor growth and early relapse of human colorectal cancer by affecting genes involved in the cell cycle.

PubMed

Yang, I-Ping; Tsai, Hsiang-Lin; Hou, Ming-Feng; Chen, Ku-Chung; Tsai, Pei-Chien; Huang, Szu-Wei; Chou, Wen-Wen; Wang, Jaw-Yuan; Juo, Suh-Hang Hank

2012-08-01

Colorectal cancer (CRC) is associated with high recurrence and mortality. Because deregulation of microRNAs is associated with CRC development and recurrence, the expression levels of microRNAs can be a simple and reliable biomarker to detect postoperative early relapse, thereby helping physicians to treat high-risk patients more efficiently. We used microRNA arrays and observed that microRNA-93 had substantially different expression levels in early (recurrence within 12 months after surgery) and non-early relapse CRC patients. The replication study, which included 35 early relapse and 42 non-early relapse subjects, further confirmed overexpression of microRNA-93 in non-early relapse samples. The in vitro and in vivo effects of microRNA-93 were investigated by examining cell proliferation, migration and invasion, as well as cell cycles, target-gene expression and xenograft in null mice. Cellular studies showed that the overexpression of microRNA-93 inhibited colon cancer cell proliferation and migration but not invasion. The cell cycle studies also revealed that microRNA-93 caused an accumulation of the G2 population. However, microRNA-93 could not induce cell apoptosis or necrosis. Functional studies showed that microRNA-93 could suppress CCNB1 protein expression leading to cell cycle arrest in the G2 phase. Moreover, microRNA-93 repressed expression of ERBB2, p21 and VEGF, all of which are involved in cell proliferation. MicroRNA-93 also suppressed tumor growth in null mice. This study showed that microRNA-93 can inhibit tumorigenesis and reduce the recurrence of CRC; these findings may have potential clinical applications for predicting the recurrence of CRC.

A new prospect in cancer therapy: targeting cancer stem cells to eradicate cancer.

PubMed

Chen, Li-Sha; Wang, An-Xin; Dong, Bing; Pu, Ke-Feng; Yuan, Li-Hua; Zhu, Yi-Min

2012-12-01

According to the cancer stem cell theory, cancers can be initiated by cancer stem cells. This makes cancer stem cells prime targets for therapeutic intervention. Eradicating cancer stem cells by efficient targeting agents may have the potential to cure cancer. In this review, we summarize recent breakthroughs that have improved our understanding of cancer stem cells, and we discuss the therapeutic strategy of targeting cancer stem cells, a promising future direction for cancer stem cell research.

Glioblastoma Stem Cells as a New Therapeutic Target for Glioblastoma.

PubMed

Kalkan, Rasime

2015-01-01

Primary and secondary glioblastomas (GBMs) are two distinct diseases. The genetic and epigenetic background of these tumors is highly variable. The treatment procedure for these tumors is often unsuccessful because of the cellular heterogeneity and intrinsic ability of the tumor cells to invade healthy tissues. The fatal outcome of these tumors promotes researchers to find out new markers associated with the prognosis and treatment planning. In this communication, the role of glioblastoma stem cells in tumor progression and the malignant behavior of GBMs are summarized with attention to the signaling pathways and molecular regulators that are involved in maintaining the glioblastoma stem cell phenotype. A better understanding of these stem cell-like cells is necessary for designing new effective treatments and developing novel molecular strategies to target glioblastoma stem cells. We discuss hypoxia as a new therapeutic target for GBM. We focus on the inhibition of signaling pathways, which are associated with the hypoxia-mediated maintenance of glioblastoma stem cells, and the knockdown of hypoxia-inducible factors, which could be identified as attractive molecular target approaches for GBM therapeutics.

Monoclonal antibodies targeting non-small cell lung cancer stem-like cells by multipotent cancer stem cell monoclonal antibody library.

PubMed

Cao, Kaiyue; Pan, Yunzhi; Yu, Long; Shu, Xiong; Yang, Jing; Sun, Linxin; Sun, Lichao; Yang, Zhihua; Ran, Yuliang

2017-02-01

Cancer stem cells (CSCs) are a rare subset of cancer cells that play a significant role in cancer initiation, spreading, and recurrence. In this study, a subpopulation of lung cancer stem-like cells (LCSLCs) was identified from non-small cell lung carcinoma cell lines, SPCA-1 and A549, using serum-free suspension sphere-forming culture method. A monoclonal antibody library was constructed using immunized BLAB/c mice with the multipotent CSC cell line T3A-A3. Flow cytometry analysis showed that 33 mAbs targeted antigens can be enriched in sphere cells compared with the parental cells of SPCA-1 and A549 cell lines. Then, we performed functional antibody screening including sphere-forming inhibiting and invasion inhibiting assay. The results showed that two antibodies, 12C7 and 9B8, notably suppressed the self-renewal and invasion of LCSLCs. Fluorescence-activated cell sorting (FACs) found that the positive cells recognized by mAbs, 12C7 or 9B8, displayed features of LCSLCs. Interestingly, we found that these two antibodies recognized different subsets of cells and their combination effect was superior to the individual effect both in vitro and in vivo. Tissue microarrays were applied to detect the expression of the antigens targeted by these two antibodies. The positive expression of 12C7 and 9B8 targeted antigen was 84.4 and 82.5%, respectively, which was significantly higher than that in the non-tumor lung tissues. In conclusion, we screened two potential therapeutic antibodies that target different subsets of LCSLCs.

Intestinal epithelial cells as producers but not targets of chronic TNF suffice to cause murine Crohn-like pathology

PubMed Central

Roulis, Manolis; Armaka, Maria; Manoloukos, Menelaos; Apostolaki, Maria; Kollias, George

2011-01-01

TNF plays a crucial role in the pathogenesis of Crohn disease. Dysregulated TNF production in mice that bear the genetic deletion of the TNF AU-rich regulatory elements (ARE) (TnfΔARE/+ mice) results in TNF receptor I (TNFRI)-dependent spontaneous Crohn-like pathology. Current concepts consider intestinal epithelial cell (IEC) responses to TNF to be critical for intestinal pathology, but the potential contribution of IEC-derived TNF in disease pathogenesis has not been addressed. In this study we examined whether IEC are sufficient as cellular targets or sources of TNF in the development of intestinal pathology. Using IEC-specific reactivation of a hypomorphic TnfΔAREneo allele in mice, we show that selective chronic overproduction of TNF by IEC suffices to cause full development of Crohn-like pathology. Epithelial TNF overexpression leads to early activation of the underlying intestinal myofibroblast, a cell type previously identified as a sufficient target of TNF for disease development in the TnfΔARE model. By contrast, restricted TNFRI expression on IEC although sufficient to confer IEC apoptosis after acute exogenous TNF administration, fails to induce pathology following chronic specific targeting of IEC by endogenous TNF in TnfΔARE/+ mice. Our results argue against IEC being early and sufficient responders to chronic TNF-mediated pathogenic signals and suggest that proinflammatory aberrations leading to chronic TNF production by IEC may initiate pathology in Crohn disease. PMID:21402942

HLA-targeted flow cytometric sorting of blood cells allows separation of pure and viable microchimeric cell populations.

PubMed

Drabbels, Jos J M; van de Keur, Carin; Kemps, Berit M; Mulder, Arend; Scherjon, Sicco A; Claas, Frans H J; Eikmans, Michael

2011-11-10

Microchimerism is defined by the presence of low levels of nonhost cells in a person. We developed a reliable method for separating viable microchimeric cells from the host environment. For flow cytometric cell sorting, HLA antigens were targeted with human monoclonal HLA antibodies (mAbs). Optimal separation of microchimeric cells (present at a proportion as low as 0.01% in artificial mixtures) was obtained with 2 different HLA mAbs, one targeting the chimeric cells and the other the background cells. To verify purity of separated cell populations, flow-sorted fractions of 1000 cells were processed for DNA analysis by HLA-allele-specific and Y-chromosome-directed real-time quantitative PCR assays. After sorting, PCR signals of chimeric DNA markers in the positive fractions were significantly enhanced compared with those in the presort samples, and they were similar to those in 100% chimeric control samples. Next, we demonstrate applicability of HLA-targeted FACS sorting after pregnancy by separating chimeric maternal cells from child umbilical cord mononuclear cells. Targeting allelic differences with anti-HLA mAbs with FACS sorting allows maximal enrichment of viable microchimeric cells from a background cell population. The current methodology enables reliable microchimeric cell detection and separation in clinical specimens.

Design and development of a multifunctional nano carrier system for imaging, drug delivery, and cell targeting in cancer research

NASA Astrophysics Data System (ADS)

Cho, Hoon-Sung

There has been an increasing need in the last decade for early diagnosis and treatment of cancer prior to the tumor mass becoming evident as anatomical anomaly. A major challenge in cancer diagnosis is to distinguish cancer cells from the surrounding, normal tissue. For early cancer diagnosis and treatment, a nano carrier system was designed and developed with key components uniquely structured according to biomedical and clinical requirements: targeting, drug storage capabilities, fluorescent emissions near the infrared range for in vivo imaging, and magnetic hyperthermia. For in vivo imaging, quantum dots with emissions near infrared range (˜800 nm) were conjugated onto the surface of carbon nanotubes and nanospheres consisting of a spherical polystyrene matrix (˜100 nm) and high fraction of superparamagnetic Fe3O4 nanoparticles (˜10 nm) embedded. The QDs on these nano carriers exhibited intense visible emissions using fluorescent spectroscopy and successfully facilitated in vivo soft tissue imaging in mice. For drug storage, the chemotherapeutic agent, paclitaxel (PTX) was loaded onto the surfaces of these nano-carriers by using a layer of biodegradable poly(lactic-co-glycolic acid) (PLGA). A cell-based cytotoxicity assay was employed to verify successful loading of pharmacologically active drug, PTX. Cell viability of human, metastatic PC3mm2 prostate cancer cells was assessed in the presence and absence of various nano-carrier populations using the MTT assay. For hyperthermia, Fe3O 4 nanoparticles were conjugated onto the surfaces of carbon nanotubes (CNT) and embedded into the nanospheres. Magnetization measurements showed nearly reversible hysteresis curves from the Fe3O4-conjugated CNTs and the magnetic nanospheres (MNS). Application of an alternating electromagnetic field effectively induced heating the solution of the Fe3O 4-conjugated CNTs and the magnetic nanospheres (MNS) into temperature ranges (up to 55ºC) suitable for therapeutic hyperthermia

Zn(II)-curc targets p53 in thyroid cancer cells.

PubMed

Garufi, Alessia; D'Orazi, Valerio; Crispini, Alessandra; D'Orazi, Gabriella

2015-10-01

TP53 mutation is a common event in many cancers, including thyroid carcinoma. Defective p53 activity promotes cancer resistance to therapies and a more malignant phenotype, acquiring oncogenic functions. Rescuing the function of mutant p53 (mutp53) protein is an attractive anticancer therapeutic strategy. Zn(II)-curc is a novel small molecule that has been shown to target mutp53 protein in several cancer cells, but its effect in thyroid cancer cells remains unclear. Here, we investigated whether Zn(II)-curc could affect p53 in thyroid cancer cells with both p53 mutation (R273H) and wild-type p53. Zn(II)-curc induced mutp53H273 downregulation and reactivation of wild-type functions, such as binding to canonical target promoters and target gene transactivation. This latter effect was similar to that induced by PRIMA-1. In addition, Zn(II)-curc triggered p53 target gene expression in wild-type p53-carrying cells. In combination treatments, Zn(II)-curc enhanced the antitumor activity of chemotherapeutic drugs, in both mutant and wild-type-carrying cancer cells. Taken together, our data indicate that Zn(II)-curc promotes the reactivation of p53 in thyroid cancer cells, providing in vitro evidence for a potential therapeutic approach in thyroid cancers.

Targeted gene panel sequencing in children with very early onset inflammatory bowel disease--evaluation and prospective analysis.

PubMed

Kammermeier, Jochen; Drury, Suzanne; James, Chela T; Dziubak, Robert; Ocaka, Louise; Elawad, Mamoun; Beales, Philip; Lench, Nicholas; Uhlig, Holm H; Bacchelli, Chiara; Shah, Neil

2014-11-01

Multiple monogenetic conditions with partially overlapping phenotypes can present with inflammatory bowel disease (IBD)-like intestinal inflammation. With novel genotype-specific therapies emerging, establishing a molecular diagnosis is becoming increasingly important. We have introduced targeted next-generation sequencing (NGS) technology as a prospective screening tool in children with very early onset IBD (VEOIBD). We evaluated the coverage of 40 VEOIBD genes in two separate cohorts undergoing targeted gene panel sequencing (TGPS) (n=25) and whole exome sequencing (WES) (n=20). TGPS revealed causative mutations in four genes (IL10RA, EPCAM, TTC37 and SKIV2L) discovered unexpected phenotypes and directly influenced clinical decision making by supporting as well as avoiding haematopoietic stem cell transplantation. TGPS resulted in significantly higher median coverage when compared with WES, fewer coverage deficiencies and improved variant detection across established VEOIBD genes. Excluding or confirming known VEOIBD genotypes should be considered early in the disease course in all cases of therapy-refractory VEOIBD, as it can have a direct impact on patient management. To combine both described NGS technologies would compensate for the limitations of WES for disease-specific application while offering the opportunity for novel gene discovery in the research setting. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.

ErbB-targeted CAR T-cell immunotherapy of cancer.

PubMed

Whilding, Lynsey M; Maher, John

2015-01-01

Chimeric antigen receptor (CAR) based immunotherapy has been under development for the last 25 years and is now a promising new treatment modality in the field of cancer immunotherapy. The approach involves genetically engineering T cells to target malignant cells through expression of a bespoke fusion receptor that couples an HLA-independent antigen recognition domain to one or more intracellular T-cell activating modules. Multiple clinical trials are now underway in several centers to investigate CAR T-cell immunotherapy of diverse hematologic and solid tumor types. The most successful results have been achieved in the treatment of patients with B-cell malignancies, in whom several complete and durable responses have been achieved. This review focuses on the preclinical and clinical development of CAR T-cell immunotherapy of solid cancers, targeted against members of the ErbB family.

New target genes of MITF-induced microRNA-211 contribute to melanoma cell invasion.

PubMed

Margue, Christiane; Philippidou, Demetra; Reinsbach, Susanne E; Schmitt, Martina; Behrmann, Iris; Kreis, Stephanie

2013-01-01

The non-coding microRNAs (miRNA) have tissue- and disease-specific expression patterns. They down-regulate target mRNAs, which likely impacts on most fundamental cellular processes. Differential expression patterns of miRNAs are currently being exploited for identification of biomarkers for early disease diagnosis, prediction of progression for melanoma and other cancers and as promising drug targets, since they can easily be inhibited or replaced in a given cellular context. Before successfully manipulating miRNAs in clinical settings, their precise expression levels, endogenous functions and thus their target genes have to be determined. MiR-211, a melanocyte lineage-specific small non-coding miRNA, is located in an intron of TRPM1, a target gene of the microphtalmia-associated transcription factor (MITF). By transcriptionally up-regulating TRPM1, MITF, which is critical for both melanocyte differentiation and survival and for melanoma progression, indirectly drives the expression of miR-211. Expression of this miRNA is often reduced in melanoma samples. Here, we investigated functional roles of miR-211 by identifying and studying new target genes. We show that MITF-correlated miR-211 expression levels are mostly but not always reduced in a panel of 11 melanoma cell lines and in primary and metastatic melanoma compared to normal melanocytes and nevi, respectively. MiR-211 itself only marginally impacted on cell invasion and migration, while perturbation of some new miR-211 target genes, such as AP1S2, SOX11, IGFBP5, and SERINC3 significantly increased invasion. These results and the variable expression levels of miR-211 raise serious doubts on the value of miR-211 as a melanoma tumor-suppressing miRNA and/or as a biomarker for melanoma.

GEM-loaded magnetic albumin nanospheres modified with cetuximab for simultaneous targeting, magnetic resonance imaging, and double-targeted thermochemotherapy of pancreatic cancer cells.

PubMed

Wang, Ling; An, Yanli; Yuan, Chenyan; Zhang, Hao; Liang, Chen; Ding, Fengan; Gao, Qi; Zhang, Dongsheng

2015-01-01

Targeted delivery is a promising strategy to improve the diagnostic imaging and therapeutic effect of cancers. In this paper, novel cetuximab (C225)-conjugated, gemcitabine (GEM)-containing magnetic albumin nanospheres (C225-GEM/MANs) were fabricated and applied as a theranostic nanocarrier to conduct simultaneous targeting, magnetic resonance imaging (MRI), and double-targeted thermochemotherapy against pancreatic cancer cells. Fe3O4 nanoparticles (NPs) and GEM co-loaded albumin nanospheres (GEM/MANs) were prepared, and then C225 was further conjugated to synthesize C225-GEM/MANs. Their morphology, mean particle size, GEM encapsulation ratio, specific cell-binding ability, and thermal dynamic profiles were characterized. The effects of discriminating different EGFR-expressing pancreatic cancer cells (AsPC-1 and MIA PaCa-2) and monitoring cellular targeting effects were assessed by targeted MRI. Lastly, the antitumor efficiency of double/C225/magnetic-targeted and nontargeted thermochemotherapy was compared with chemotherapy alone using 3-(4, 5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) and flow cytometry (FCM) assay. When treated with targeted nanospheres, AsPC-1 cells showed a significantly less intense MRI T2 signal than MIA PaCa-2 cells, while both cells had similar signal strength when incubated with nontargeted nanospheres. T2 signal intensity was significantly lower when magnetic and C225 targeting were combined, rather than used alone. The inhibitory and apoptotic rates of each thermochemotherapy group were significantly higher than those of the chemotherapy-alone groups. Additionally, both MTT and FCM analysis verified that double-targeted thermochemotherapy had the highest targeted killing efficiency among all groups. The C225-GEM/MANs can distinguish various EGFR-expressing live pancreatic cancer cells, monitor diverse cellular targeting effects using targeted MRI imaging, and efficiently mediate double-targeted thermochemotherapy

A Combined Omics Approach to Generate the Surface Atlas of Human Naive CD4+ T Cells during Early T-Cell Receptor Activation*

PubMed Central

Graessel, Anke; Hauck, Stefanie M.; von Toerne, Christine; Kloppmann, Edda; Goldberg, Tatyana; Koppensteiner, Herwig; Schindler, Michael; Knapp, Bettina; Krause, Linda; Dietz, Katharina; Schmidt-Weber, Carsten B.; Suttner, Kathrin

2015-01-01

Naive CD4+ T cells are the common precursors of multiple effector and memory T-cell subsets and possess a high plasticity in terms of differentiation potential. This stem-cell-like character is important for cell therapies aiming at regeneration of specific immunity. Cell surface proteins are crucial for recognition and response to signals mediated by other cells or environmental changes. Knowledge of cell surface proteins of human naive CD4+ T cells and their changes during the early phase of T-cell activation is urgently needed for a guided differentiation of naive T cells and may support the selection of pluripotent cells for cell therapy. Periodate oxidation and aniline-catalyzed oxime ligation technology was applied with subsequent quantitative liquid chromatography-tandem MS to generate a data set describing the surface proteome of primary human naive CD4+ T cells and to monitor dynamic changes during the early phase of activation. This led to the identification of 173 N-glycosylated surface proteins. To independently confirm the proteomic data set and to analyze the cell surface by an alternative technique a systematic phenotypic expression analysis of surface antigens via flow cytometry was performed. This screening expanded the previous data set, resulting in 229 surface proteins, which were expressed on naive unstimulated and activated CD4+ T cells. Furthermore, we generated a surface expression atlas based on transcriptome data, experimental annotation, and predicted subcellular localization, and correlated the proteomics result with this transcriptional data set. This extensive surface atlas provides an overall naive CD4+ T cell surface resource and will enable future studies aiming at a deeper understanding of mechanisms of T-cell biology allowing the identification of novel immune targets usable for the development of therapeutic treatments. PMID:25991687

Phenotypic high-throughput screening elucidates target pathway in breast cancer stem cell-like cells.

PubMed

Carmody, Leigh C; Germain, Andrew R; VerPlank, Lynn; Nag, Partha P; Muñoz, Benito; Perez, Jose R; Palmer, Michelle A J

2012-10-01

Cancer stem cells (CSCs) are resistant to standard cancer treatments and are likely responsible for cancer recurrence, but few therapies target this subpopulation. Due to the difficulty in propagating CSCs outside of the tumor environment, previous work identified CSC-like cells by inducing human breast epithelial cells into an epithelial-to-mesenchymal transdifferentiated state (HMLE_sh_ECad). A phenotypic screen was conducted against HMLE_sh_ECad with 300 718 compounds from the Molecular Libraries Small Molecule Repository to identify selective inhibitors of CSC growth. The screen yielded 2244 hits that were evaluated for toxicity and selectivity toward an isogenic control cell line. An acyl hydrazone scaffold emerged as a potent and selective scaffold targeting HMLE_sh_ECad. Fifty-three analogues were acquired and tested; compounds ranged in potency from 790 nM to inactive against HMLE_sh_ECad. Of the analogues, ML239 was best-in-class with an IC(50)= 1.18 µM against HMLE_sh_ECad, demonstrated a >23-fold selectivity over the control line, and was toxic to another CSC-like line, HMLE_shTwist, and a breast carcinoma cell line, MDA-MB-231. Gene expression studies conducted with ML239-treated cells showed altered gene expression in the NF-κB pathway in the HMLE_sh_ECad line but not in the isogenic control line. Future studies will be directed toward the identification of ML239 target(s).

Visualization of Endoplasmic Reticulum and Mitochondria in Aurantiochytrium limacinum by the Expression of EGFP with Cell Organelle-Specific Targeting/Retaining Signals.

PubMed

Okino, Nozomu; Wakisaka, Hiroyoshi; Ishibashi, Yohei; Ito, Makoto

2018-04-01

Thraustochytrids are single cell marine eukaryotes that produce large amounts of polyunsaturated fatty acids such as docosahexaenoic acid. In the present study, we report the visualization of endoplasmic reticulum (ER) and mitochondria in a type strain of the thraustochytrid, Aurantiochytrium limacinum ATCC MYA-1381, using the enhanced green fluorescent protein (EGFP) with specific targeting/retaining signals. We expressed the egfp gene with ER targeting/retaining signals from A. limacinum calreticulin or BiP/GRP78 in the thraustochytrid, resulting in the distribution of EGFP signals at the perinuclear region and near lipid droplets. ER-Tracker™ Red, an authentic fluorescent probe for the visualization of ER in mammalian cells, also stained the same region. We observed small lipid droplets generated from the visualized ER in the early growth phase of cell culture. Expression of the egfp gene with the mitochondria targeting signal from A. limacinum cytochrome c oxidase resulted in the localization of EGFP near the plasma membrane. The distribution of EGFP signals coincided with that of MitoTracker® Red CMXRos, which is used to visualize mitochondria in eukaryotes. The ER and mitochondria of A. limacinum were visualized for the first time by EGFP with thraustochytrid cell organelle-specific targeting/retaining signals. These results will contribute to classification of the intracellular localization of proteins expressed in ER and mitochondria as well as analyses of these cell organelles in thraustochytrids.

Detecting drug-target binding in cells using fluorescence-activated cell sorting coupled with mass spectrometry analysis.

PubMed

Wilson, Kris; Webster, Scott P; Iredale, John P; Zheng, Xiaozhong; Homer, Natalie Z; Pham, Nhan T; Auer, Manfred; Mole, Damian J

2017-12-15

The assessment of drug-target engagement for determining the efficacy of a compound inside cells remains challenging, particularly for difficult target proteins. Existing techniques are more suited to soluble protein targets. Difficult target proteins include those with challenging in vitro solubility, stability or purification properties that preclude target isolation. Here, we report a novel technique that measures intracellular compound-target complex formation, as well as cellular permeability, specificity and cytotoxicity-the toxicity-affinity-permeability-selectivity (TAPS) technique. The TAPS assay is exemplified here using human kynurenine 3-monooxygenase (KMO), a challenging intracellular membrane protein target of significant current interest. TAPS confirmed target binding of known KMO inhibitors inside cells. We conclude that the TAPS assay can be used to facilitate intracellular hit validation on most, if not all intracellular drug targets.

Detecting drug-target binding in cells using fluorescence-activated cell sorting coupled with mass spectrometry analysis

NASA Astrophysics Data System (ADS)

Wilson, Kris; Webster, Scott P.; Iredale, John P.; Zheng, Xiaozhong; Homer, Natalie Z.; Pham, Nhan T.; Auer, Manfred; Mole, Damian J.

2018-01-01

The assessment of drug-target engagement for determining the efficacy of a compound inside cells remains challenging, particularly for difficult target proteins. Existing techniques are more suited to soluble protein targets. Difficult target proteins include those with challenging in vitro solubility, stability or purification properties that preclude target isolation. Here, we report a novel technique that measures intracellular compound-target complex formation, as well as cellular permeability, specificity and cytotoxicity-the toxicity-affinity-permeability-selectivity (TAPS) technique. The TAPS assay is exemplified here using human kynurenine 3-monooxygenase (KMO), a challenging intracellular membrane protein target of significant current interest. TAPS confirmed target binding of known KMO inhibitors inside cells. We conclude that the TAPS assay can be used to facilitate intracellular hit validation on most, if not all intracellular drug targets.

Targeting myeloid-derived suppressor cells for cancer immunotherapy.

PubMed

Liu, Yijun; Wei, Guowei; Cheng, Wesley A; Dong, Zhenyuan; Sun, Han; Lee, Vincent Y; Cha, Soung-Chul; Smith, D Lynne; Kwak, Larry W; Qin, Hong

2018-05-31

Myeloid-derived suppressor cells (MDSCs) are a heterogeneous population of immature myeloid cells with an immune suppressive phenotype. They represent a critical component of the immune suppressive niche described in cancer, where they support immune escape and tumor progression through direct effects on both the innate and adaptive immune responses, largely by contributing to maintenance of a high oxidative stress environment. The number of MDSCs positively correlates with protumoral activity, and often diminishes the effectiveness of immunotherapies, which is particularly problematic with the emergence of personalized medicine. Approaches targeting MDSCs showed promising results in preclinical studies and are under active investigation in clinical trials in combination with various immune checkpoint inhibitors. In this review, we discuss MDSC targets and therapeutic approaches targeting MDSC that have the aim of enhancing the existing tumor therapies.

Many si/shRNAs can kill cancer cells by targeting multiple survival genes through an off-target mechanism

PubMed Central

van Dongen, Stijn; Haluck-Kangas, Ashley; Sarshad, Aishe A; Bartom, Elizabeth T; Kim, Kwang-Youn A; Scholtens, Denise M; Hafner, Markus; Zhao, Jonathan C; Murmann, Andrea E

2017-01-01

Over 80% of multiple-tested siRNAs and shRNAs targeting CD95 or CD95 ligand (CD95L) induce a form of cell death characterized by simultaneous activation of multiple cell death pathways preferentially killing transformed and cancer stem cells. We now show these si/shRNAs kill cancer cells through canonical RNAi by targeting the 3’UTR of critical survival genes in a unique form of off-target effect we call DISE (death induced by survival gene elimination). Drosha and Dicer-deficient cells, devoid of most miRNAs, are hypersensitive to DISE, suggesting cellular miRNAs protect cells from this form of cell death. By testing 4666 shRNAs derived from the CD95 and CD95L mRNA sequences and an unrelated control gene, Venus, we have identified many toxic sequences - most of them located in the open reading frame of CD95L. We propose that specific toxic RNAi-active sequences present in the genome can kill cancer cells. PMID:29063830

The NRF2-KEAP1 Pathway Is an Early Responsive Gene Network in Arsenic Exposed Lymphoblastoid Cells

PubMed Central

Córdova, Emilio J.; Martínez-Hernández, Angélica; Uribe-Figueroa, Laura; Centeno, Federico; Morales-Marín, Mirna; Koneru, Harsha; Coleman, Matthew A.; Orozco, Lorena

2014-01-01

Inorganic arsenic (iAs), a major environmental contaminant, has risen as an important health problem worldwide. More detailed identification of the molecular mechanisms associated with iAs exposure would help to establish better strategies for prevention and treatment. Although chronic iAs exposures have been previously studied there is little to no information regarding the early events of exposure to iAs. To better characterize the early mechanisms of iAs exposure we conducted gene expression studies using sublethal doses of iAs at two different time-points. The major transcripts differentially regulated at 2 hrs of iAs exposure included antioxidants, detoxificants and chaperones. Moreover, after 12 hrs of exposure many of the down-regulated genes were associated with DNA replication and S phase cell cycle progression. Interestingly, the most affected biological pathway by both 2 or 12 hrs of iAs exposure were the Nrf2-Keap1 pathway, represented by the highly up-regulated HMOX1 transcript, which is transcriptionally regulated by the transcription factor Nrf2. Additional Nrf2 targets included SQSTM1 and ABCB6, which were not previously associated with acute iAs exposure. Signalling pathways such as interferon, B cell receptor and AhR route were also responsive to acute iAs exposure. Since HMOX1 expression increased early (20 min) and was responsive to low iAs concentrations (0.1 µM), this gene could be a suitable early biomarker for iAs exposure. In addition, the novel Nrf2 targets SQSTM1 and ABCB6 could play an important and previously unrecognized role in cellular protection against iAs. PMID:24516582

Targeted delivery of cisplatin to prostate cancer cells by aptamer functionalized Pt(IV) prodrug-PLGA-PEG nanoparticles.

PubMed

Dhar, Shanta; Gu, Frank X; Langer, Robert; Farokhzad, Omid C; Lippard, Stephen J

2008-11-11

Cisplatin is used to treat a variety of tumors, but dose limiting toxicities or intrinsic and acquired resistance limit its application in many types of cancer including prostate. We report a unique strategy to deliver cisplatin to prostate cancer cells by constructing Pt(IV)-encapsulated prostate-specific membrane antigen (PSMA) targeted nanoparticles (NPs) of poly(D,L-lactic-co-glycolic acid) (PLGA)-poly(ethylene glycol) (PEG)-functionalized controlled release polymers. By using PLGA-b-PEG nanoparticles with PSMA targeting aptamers (Apt) on the surface as a vehicle for the platinum(IV) compound c,t,c-[Pt(NH(3))(2)(O(2)CCH(2)CH(2)CH(2)CH(2)CH(3))(2)Cl(2)] (1), a lethal dose of cisplatin was delivered specifically to prostate cancer cells. PSMA aptamer targeted delivery of Pt(IV) cargos to PSMA(+) LNCaP prostate cancer cells by endocytosis of the nanoparticle vehicles was demonstrated using fluorescence microscopy by colocalization of green fluorescent labeled cholesterol-encapsulated NPs and early endosome marker EEA-1. The choice of linear hexyl chains in 1 was the result of a systematic study to optimize encapsulation and controlled release from the polymer without compromising either feature. Release of cisplatin from the polymeric nanoparticles after reduction of 1 and formation of cisplatin 1,2-intrastrand d(GpG) cross-links on nuclear DNA was confirmed by using a monoclonal antibody for the adduct. A comparison between the cytotoxic activities of Pt(IV)-encapsulated PLGA-b-PEG NPs with the PSMA aptamer on the surface (Pt-NP-Apt), cisplatin, and the nontargeted Pt(IV)-encapsulated NPs (Pt-NP) against human prostate PSMA-overexpressing LNCaP and PSMA(-) PC3 cancer cells revealed significant differences. The effectiveness of PSMA targeted Pt-NP-Apt nanoparticles against the PSMA(+) LNCaP cells is approximately an order of magnitude greater than that of free cisplatin.

Variations of target volume definition and daily target volume localization in stereotactic body radiotherapy for early-stage non-small cell lung cancer patients under abdominal compression.

PubMed

Han, Chunhui; Sampath, Sagus; Schultheisss, Timothy E; Wong, Jeffrey Y C

2017-01-01

We aimed to compare gross tumor volumes (GTV) in 3-dimensional computed tomography (3DCT) simulation and daily cone beam CT (CBCT) with the internal target volume (ITV) in 4-dimensional CT (4DCT) simulation in stereotactic body radiotherapy (SBRT) treatment of patients with early-stage non-small cell lung cancer (NSCLC) under abdominal compression. We retrospectively selected 10 patients with NSCLC who received image-guided SBRT treatments under abdominal compression with daily CBCT imaging. GTVs were contoured as visible gross tumor on the planning 3DCT and daily CBCT, and ITVs were contoured using maximum intensity projection (MIP) images of the planning 4DCT. Daily CBCTs were registered with 3DCT and MIP images by matching of bony landmarks in the thoracic region to evaluate interfractional GTV position variations. Relative to MIP-based ITVs, the average 3DCT-based GTV volume was 66.3 ± 17.1% (range: 37.5% to 92.0%) (p < 0.01 in paired t-test), and the average CBCT-based GTV volume was 90.0 ± 6.7% (daily range: 75.7% to 107.1%) (p = 0.02). Based on bony anatomy matching, the center-of-mass coordinates for CBCT-based GTVs had maximum absolute shift of 2.4 mm (left-right), 7.0 mm (anterior-posterior [AP]), and 5.2 mm (superior-inferior [SI]) relative to the MIP-based ITV. CBCT-based GTVs had average overlapping ratio of 81.3 ± 11.2% (range: 45.1% to 98.9%) with the MIP-based ITV, and 57.7 ± 13.7% (range: 35.1% to 83.2%) with the 3DCT-based GTV. Even with abdominal compression, both 3DCT simulations and daily CBCT scans significantly underestimated the full range of tumor motion. In daily image-guided patient setup corrections, automatic bony anatomy-based image registration could lead to target misalignment. Soft tissue-based image registration should be performed for accurate treatment delivery. Copyright © 2017 American Association of Medical Dosimetrists. Published by Elsevier Inc. All rights reserved.

EGFR-targeted granzyme B expressed in NK cells enhances natural cytotoxicity and mediates specific killing of tumor cells.

PubMed

Oberoi, Pranav; Jabulowsky, Robert A; Bähr-Mahmud, Hayat; Wels, Winfried S

2013-01-01

Natural killer (NK) cells are highly specialized effectors of the innate immune system that hold promise for adoptive cancer immunotherapy. Their cell killing activity is primarily mediated by the pro-apoptotic serine protease granzyme B (GrB), which enters targets cells with the help of the pore-forming protein perforin. We investigated expression of a chimeric GrB fusion protein in NK cells as a means to augment their antitumoral activity. For selective targeting to tumor cells, we fused the epidermal growth factor receptor (EGFR) peptide ligand transforming growth factor α (TGFα) to human pre-pro-GrB. Established human NKL natural killer cells transduced with a lentiviral vector expressed this GrB-TGFα (GrB-T) molecule in amounts comparable to endogenous wildtype GrB. Activation of the genetically modified NK cells by cognate target cells resulted in the release of GrB-T together with endogenous granzymes and perforin, which augmented the effector cells' natural cytotoxicity against NK-sensitive tumor cells. Likewise, GrB-T was released into the extracellular space upon induction of degranulation with PMA and ionomycin. Secreted GrB-T fusion protein displayed specific binding to EGFR-overexpressing tumor cells, enzymatic activity, and selective target cell killing in the presence of an endosomolytic activity. Our data demonstrate that ectopic expression of a targeted GrB fusion protein in NK cells is feasible and can enhance antitumoral activity of the effector cells.

Adoptive therapy with CAR redirected T cells: the challenges in targeting solid tumors.

PubMed

Abken, Hinrich

2015-01-01

Recent spectacular success in the adoptive cell therapy of leukemia and lymphoma with chimeric antigen receptor (CAR)-modified T cells raised the expectations that this therapy may be efficacious in a wide range of cancer entities. The expectations are based on the predefined specificity of CAR T cells by an antibody-derived binding domain that acts independently of the natural T-cell receptor, recognizes targets independently of presentation by the major histocompatibility complex and allows targeting toward virtually any cell surface antigen. We here discuss that targeting CAR T cells toward solid tumors faces certain circumstances critical for the therapeutic success. Targeting tumor stroma and taking advantage of TRUCK cells, in other words, CAR T cells with inducible release of a transgenic payload, are some strategies envisaged to overcome current limitations in the near future.

Gunslinger Effect and Müller-Lyer Illusion: Examining Early Visual Information Processing for Late Limb-Target Control.

PubMed

Roberts, James W; Lyons, James; Garcia, Daniel B L; Burgess, Raquel; Elliott, Digby

2017-07-01

The multiple process model contends that there are two forms of online control for manual aiming: impulse regulation and limb-target control. This study examined the impact of visual information processing for limb-target control. We amalgamated the Gunslinger protocol (i.e., faster movements following a reaction to an external trigger compared with the spontaneous initiation of movement) and Müller-Lyer target configurations into the same aiming protocol. The results showed the Gunslinger effect was isolated at the early portions of the movement (peak acceleration and peak velocity). Reacted aims reached a longer displacement at peak deceleration, but no differences for movement termination. The target configurations manifested terminal biases consistent with the illusion. We suggest the visual information processing demands imposed by reacted aims can be adapted by integrating early feedforward information for limb-target control.

Surface engineered antifouling optomagnetic SPIONs for bimodal targeted imaging of pancreatic cancer cells

PubMed Central

Wang, Xiaohui; Xing, Xiaohong; Zhang, Bingbo; Liu, Fengjun; Cheng, Yingsheng; Shi, Donglu

2014-01-01

Targeted imaging contrast agents for early pancreatic ductal adenocarcinoma diagnosis was developed using superparamagnetic iron oxide nanoparticles (SPIONs). For phase transfer of SPIONs, the hydrophobic SPIONs are first treated with tetrafluoroborate and then capped by bovine serum albumin (BSA) via ligand exchange. It was experimentally found that nitrosyl tetrafluoroborate pretreatment and proper structures of molecules are essential to the effective surface functionalization of SPIONs. Nonspecific binding was found to be significantly reduced by BSA surface functionalized hydrophobic SPIONs (BSA·SPIONs). The BSA·SPIONs were monodispersed with an average size of approximately 18.0 nm and stable in a wide pH range and various ionic strengths even after 7 days of storage. The longitudinal and transverse proton relaxation rate (r1, r2) values of the BSA·SPIONs were determined to be 11.6 and 154.2 s−1 per mM of Fe3+ respectively. The r2/r1 ratio of 13.3 ensured its application as the T2-weighted magnetic resonance imaging contrast agents. When conjugated with near-infrared fluorescent dye and monoclonal antibody, the dyeBSA·SPION-monoclonal antibody bioconjugates showed excellent targeting capability with minimal nonspecific binding in the bimodal imaging of pancreatic cancer cells. The experimental approach is facile, environmentally benign, and straightforward, which presents great promise in early cancer diagnosis. PMID:24741308

Personalized targeted therapy for esophageal squamous cell carcinoma

PubMed Central

Kang, Xiaozheng; Chen, Keneng; Li, Yicheng; Li, Jianying; D'Amico, Thomas A; Chen, Xiaoxin

2015-01-01

Esophageal squamous cell carcinoma continues to heavily burden clinicians worldwide. Researchers have discovered the genomic landscape of esophageal squamous cell carcinoma, which holds promise for an era of personalized oncology care. One of the most pressing problems facing this issue is to improve the understanding of the newly available genomic data, and identify the driver-gene mutations, pathways, and networks. The emergence of a legion of novel targeted agents has generated much hope and hype regarding more potent treatment regimens, but the accuracy of drug selection is still arguable. Other problems, such as cancer heterogeneity, drug resistance, exceptional responders, and side effects, have to be surmounted. Evolving topics in personalized oncology, such as interpretation of genomics data, issues in targeted therapy, research approaches for targeted therapy, and future perspectives, will be discussed in this editorial. PMID:26167067

Rabies virus co-localizes with early (Rab5) and late (Rab7) endosomal proteins in neuronal and SH-SY5Y cells.

PubMed

Ahmad, Waqas; Li, Yingying; Guo, Yidi; Wang, Xinyu; Duan, Ming; Guan, Zhenhong; Liu, Zengshan; Zhang, Maolin

2017-06-01

Rabies virus (RABV) is a highly neurotropic virus that follows clathrin-mediated endocytosis and pH-dependent pathway for trafficking and invasion into endothelial cells. Early (Rab5, EEA1) and late (Rab7, LAMP1) endosomal proteins play critical roles in endosomal sorting, maturity and targeting various molecular cargoes, but their precise functions in the early stage of RABV neuronal infection remain elusive. In this study, the relationship between enigmatic entry of RABV with these endosomal proteins into neuronal and SH-SY5Y cells was investigated. Immunofluorescence, TCID 50 titers, electron microscopy and western blotting were carried out to determine the molecular interaction of the nucleoprotein (N) of RABV with early or late endosomal proteins in these cell lines. The expression of N was also determined by down-regulating Rab5 and Rab7 in both cell lines through RNA interference. The results were indicative that N proficiently colocalized with Rab5/EEA1 and Rab7/LAMP1 in both cell lines at 24 and 48 h post-infection, while N titers significantly decreased in early infection of RABV. Down-regulation of Rab5 and Rab7 did not inhibit N expression, but it prevented productive infection via blocking the normal trafficking of RABV in a low pH environment. Ultrathin sections of cells studied by electron microscope also verified the close association of RABV with Rab5 and Rab7 in neurons. From the data it was concluded that primary entry of RABV strongly correlates with the kinetics of Rab-proteins present on early and late vesicles, which provides helpful clues to explain the early events of RABV in nerve cells.

Systematic Identification of Combinatorial Drivers and Targets in Cancer Cell Lines

PubMed Central

Tabchy, Adel; Eltonsy, Nevine; Housman, David E.; Mills, Gordon B.

2013-01-01

There is an urgent need to elicit and validate highly efficacious targets for combinatorial intervention from large scale ongoing molecular characterization efforts of tumors. We established an in silico bioinformatic platform in concert with a high throughput screening platform evaluating 37 novel targeted agents in 669 extensively characterized cancer cell lines reflecting the genomic and tissue-type diversity of human cancers, to systematically identify combinatorial biomarkers of response and co-actionable targets in cancer. Genomic biomarkers discovered in a 141 cell line training set were validated in an independent 359 cell line test set. We identified co-occurring and mutually exclusive genomic events that represent potential drivers and combinatorial targets in cancer. We demonstrate multiple cooperating genomic events that predict sensitivity to drug intervention independent of tumor lineage. The coupling of scalable in silico and biologic high throughput cancer cell line platforms for the identification of co-events in cancer delivers rational combinatorial targets for synthetic lethal approaches with a high potential to pre-empt the emergence of resistance. PMID:23577104

Systematic identification of combinatorial drivers and targets in cancer cell lines.

PubMed

Tabchy, Adel; Eltonsy, Nevine; Housman, David E; Mills, Gordon B

2013-01-01

There is an urgent need to elicit and validate highly efficacious targets for combinatorial intervention from large scale ongoing molecular characterization efforts of tumors. We established an in silico bioinformatic platform in concert with a high throughput screening platform evaluating 37 novel targeted agents in 669 extensively characterized cancer cell lines reflecting the genomic and tissue-type diversity of human cancers, to systematically identify combinatorial biomarkers of response and co-actionable targets in cancer. Genomic biomarkers discovered in a 141 cell line training set were validated in an independent 359 cell line test set. We identified co-occurring and mutually exclusive genomic events that represent potential drivers and combinatorial targets in cancer. We demonstrate multiple cooperating genomic events that predict sensitivity to drug intervention independent of tumor lineage. The coupling of scalable in silico and biologic high throughput cancer cell line platforms for the identification of co-events in cancer delivers rational combinatorial targets for synthetic lethal approaches with a high potential to pre-empt the emergence of resistance.

Microchimeric cells in systemic lupus erythematosus: targets or innocent bystanders?

PubMed

Stevens, A M

2006-01-01

During pregnancy maternal and fetal cells commute back and forth leading to fetal microchimerism in the mother and maternal microchimerism in the child that can persist for years after the birth. Chimeric fetal and maternal cells can be hematopoietic or can differentiate into somatic cells in multiple organs, potentially acting as targets for 'autoimmunity' and so have been implicated in the pathogenesis of autoimmune diseases that resemble graft-versus-host disease after stem cell transplantation. Fetal cells have been found in women with systemic lupus erythematosus, both in the blood and a target organ, the kidney, suggesting that they may be involved in pathogenesis. Future studies will address how the host immune system normally tolerates maternal and fetal cells or how the balance may change during autoimmunity.

Targeted therapy for localized non-small-cell lung cancer: a review

PubMed Central

Paleiron, Nicolas; Bylicki, Olivier; André, Michel; Rivière, Emilie; Grassin, Frederic; Robinet, Gilles; Chouaïd, Christos

2016-01-01

Targeted therapies have markedly improved the management of patients with advanced non-small-cell lung cancer (NSCLC), but their efficacy in localized NSCLC is less well established. The aim of this review is to analyze trials of targeted therapies in localized NSCLC. In patients with wild-type EGFR, tyrosine kinase inhibitors have shown no efficacy in Phase III trials. Few data are available for EGFR-mutated localized NSCLC, as routine biological profiling is not recommended. Available studies are small, often retrospectives, and/or conducted in a single-center making it difficult to draw firm conclusions. Ongoing prospective Phase III trials are comparing adjuvant tyrosine kinase inhibitor administration versus adjuvant chemotherapy. By analogy with the indication of bevacizumab in advanced NSCLC, use of antiangiogenic agents in the perioperative setting is currently restricted to nonsquamous NSCLC. Several trials of adjuvant or neoadjuvant bevacizumab are planned or ongoing, but for the moment there is no evidence of efficacy. Data on perioperative use of biomarkers in early-stage NSCLC come mainly from small, retrospective, uncontrolled studies. Assessment of customized adjuvant or neoadjuvant therapy in localized NSCLC (with or without oncogenic driver mutations) is a major challenge. PMID:27462164

Free Extracellular miRNA Functionally Targets Cells by Transfecting Exosomes from Their Companion Cells.

PubMed

Bryniarski, Krzysztof; Ptak, Wlodzimierz; Martin, Emilia; Nazimek, Katarzyna; Szczepanik, Marian; Sanak, Marek; Askenase, Philip W

2015-01-01

Lymph node and spleen cells of mice doubly immunized by epicutaneous and intravenous hapten application produce a suppressive component that inhibits the action of the effector T cells that mediate contact sensitivity reactions. We recently re-investigated this phenomenon in an immunological system. CD8+ T lymphocyte-derived exosomes transferred suppressive miR-150 to the effector T cells antigen-specifically due to exosome surface coat of antibody light chains made by B1a lymphocytes. Extracellular RNA (exRNA) is protected from plasma RNases by carriage in exosomes or by chaperones. Exosome transfer of functional RNA to target cells is well described, whereas the mechanism of transfer of exRNA free of exosomes remains unclear. In the current study we describe extracellular miR-150, extracted from exosomes, yet still able to mediate antigen-specific suppression. We have determined that this was due to miR-150 association with antibody-coated exosomes produced by B1a cell companions of the effector T cells, which resulted in antigen-specific suppression of their function. Thus functional cell targeting by free exRNA can proceed by transfecting companion cell exosomes that then transfer RNA cargo to the acceptor cells. This contrasts with the classical view on release of RNA-containing exosomes from the multivesicular bodies for subsequent intercellular targeting. This new alternate pathway for transfer of exRNA between cells has distinct biological and immunological significance, and since most human blood exRNA is not in exosomes may be relevant to evaluation and treatment of diseases.

Free Extracellular miRNA Functionally Targets Cells by Transfecting Exosomes from Their Companion Cells

PubMed Central

Bryniarski, Krzysztof; Ptak, Wlodzimierz; Martin, Emilia; Nazimek, Katarzyna; Szczepanik, Marian; Sanak, Marek; Askenase, Philip W.

2015-01-01

Lymph node and spleen cells of mice doubly immunized by epicutaneous and intravenous hapten application produce a suppressive component that inhibits the action of the effector T cells that mediate contact sensitivity reactions. We recently re-investigated this phenomenon in an immunological system. CD8+ T lymphocyte-derived exosomes transferred suppressive miR-150 to the effector T cells antigen-specifically due to exosome surface coat of antibody light chains made by B1a lymphocytes. Extracellular RNA (exRNA) is protected from plasma RNases by carriage in exosomes or by chaperones. Exosome transfer of functional RNA to target cells is well described, whereas the mechanism of transfer of exRNA free of exosomes remains unclear. In the current study we describe extracellular miR-150, extracted from exosomes, yet still able to mediate antigen-specific suppression. We have determined that this was due to miR-150 association with antibody-coated exosomes produced by B1a cell companions of the effector T cells, which resulted in antigen-specific suppression of their function. Thus functional cell targeting by free exRNA can proceed by transfecting companion cell exosomes that then transfer RNA cargo to the acceptor cells. This contrasts with the classical view on release of RNA-containing exosomes from the multivesicular bodies for subsequent intercellular targeting. This new alternate pathway for transfer of exRNA between cells has distinct biological and immunological significance, and since most human blood exRNA is not in exosomes may be relevant to evaluation and treatment of diseases. PMID:25923429

Large-scale generation of human iPSC-derived neural stem cells/early neural progenitor cells and their neuronal differentiation.

PubMed

D'Aiuto, Leonardo; Zhi, Yun; Kumar Das, Dhanjit; Wilcox, Madeleine R; Johnson, Jon W; McClain, Lora; MacDonald, Matthew L; Di Maio, Roberto; Schurdak, Mark E; Piazza, Paolo; Viggiano, Luigi; Sweet, Robert; Kinchington, Paul R; Bhattacharjee, Ayantika G; Yolken, Robert; Nimgaonka, Vishwajit L; Nimgaonkar, Vishwajit L

2014-01-01

Induced pluripotent stem cell (iPSC)-based technologies offer an unprecedented opportunity to perform high-throughput screening of novel drugs for neurological and neurodegenerative diseases. Such screenings require a robust and scalable method for generating large numbers of mature, differentiated neuronal cells. Currently available methods based on differentiation of embryoid bodies (EBs) or directed differentiation of adherent culture systems are either expensive or are not scalable. We developed a protocol for large-scale generation of neuronal stem cells (NSCs)/early neural progenitor cells (eNPCs) and their differentiation into neurons. Our scalable protocol allows robust and cost-effective generation of NSCs/eNPCs from iPSCs. Following culture in neurobasal medium supplemented with B27 and BDNF, NSCs/eNPCs differentiate predominantly into vesicular glutamate transporter 1 (VGLUT1) positive neurons. Targeted mass spectrometry analysis demonstrates that iPSC-derived neurons express ligand-gated channels and other synaptic proteins and whole-cell patch-clamp experiments indicate that these channels are functional. The robust and cost-effective differentiation protocol described here for large-scale generation of NSCs/eNPCs and their differentiation into neurons paves the way for automated high-throughput screening of drugs for neurological and neurodegenerative diseases.

Adoptive immunotherapy for B-cell malignancies with autologous chimeric antigen receptor modified tumor targeted T cells.

PubMed

Park, Jae H; Brentjens, Renier J

2010-04-01

Chemotherapy-resistant B-cell hematologic malignancies may be cured with allogeneic hematopoietic stem cell transplantation (HSCT), demonstrating the potential susceptibility of these tumors to donor T-cell mediated immune responses. However, high rates of transplant-related morbidity and mortality limit this approach. For this reason, there is an urgent need for less-toxic forms of immune-based cellular therapy to treat these malignancies. Adoptive transfer of autologous T cells genetically modified to express chimeric antigen receptors (CARs) targeted to specific tumor-associated antigens represents an attractive means of overcoming the limitations of conventional HSCT. To this end, investigators have generated CARs targeted to various antigens expressed by B-cell malignancies, optimized the design of these CARs to enhance receptor mediated T cell signaling, and demonstrated significant anti-tumor efficacy of the resulting CAR modified T cells both in vitro and in vivo mouse tumor models. These encouraging preclinical data have justified the translation of this approach to the clinical setting with currently 12 open clinical trials and one completed clinical trial treating various B-cell malignancies utilizing CAR modified T cells targeted to either the CD19 or CD20 B-cell specific antigens.

Vesicle-associated membrane protein 7 (VAMP-7) is essential for target cell killing in a natural killer cell line

DOE Office of Scientific and Technical Information (OSTI.GOV)

Marcet-Palacios, Marcelo; Odemuyiwa, Solomon O.; Coughlin, Jason J.

2008-02-15

Natural killer cells recognize and induce apoptosis in foreign, transformed or virus-infected cells through the release of perforin and granzymes from secretory lysosomes. Clinically, NK-cell mediated killing is a major limitation to successful allo- and xenotransplantation. The molecular mechanisms that regulate the fusion of granzyme B-containing secretory lysosomes to the plasma membrane in activated NK cells, prior to target cell killing, are not fully understood. Using the NK cell line YT-Indy as a model, we have investigated the expression of SNAP REceptors (SNAREs), both target (t-) and vesicular (v-) SNAREs, and their function in granzyme B-mediated target cell killing. Ourmore » data showed that YT-Indy cells express VAMP-7 and SNAP-23, but not VAMP-2. VAMP-7 was associated with granzyme B-containing lysosomal granules. Using VAMP-7 small interfering RNA (siRNA), we successfully knocked down the expression of VAMP-7 protein in YT-Indy to less than 10% of untreated cells in 24 h. VAMP7-deficient YT-Indy cells activated via co-culture with Jurkat cells released <1 ng/mL of granzyme B, compared to 1.5-2.5 {mu}g/mL from controls. Using Jurkat cells as targets, we showed a 7-fold reduction in NK cell-mediated killing by VAMP-7 deficient YT-Indy cells. Our results show that VAMP-7 is a crucial component of granzyme B release and target cell killing in the NK cell line YT-Indy. Thus, targeting VAMP-7 expression specifically with siRNA, following transplantation, may be a viable strategy for preventing NK cell-mediated transplant rejection, in vivo.« less

Tumor-targeting delivery of herb-based drugs with cell-penetrating/tumor-targeting peptide-modified nanocarriers

PubMed Central

Kebebe, Dereje; Liu, Yuanyuan; Wu, Yumei; Vilakhamxay, Maikhone; Liu, Zhidong; Li, Jiawei

2018-01-01

Cancer has become one of the leading causes of mortality globally. The major challenges of conventional cancer therapy are the failure of most chemotherapeutic agents to accumulate selectively in tumor cells and their severe systemic side effects. In the past three decades, a number of drug delivery approaches have been discovered to overwhelm the obstacles. Among these, nanocarriers have gained much attention for their excellent and efficient drug delivery systems to improve specific tissue/organ/cell targeting. In order to enhance targeting efficiency further and reduce limitations of nanocarriers, nanoparticle surfaces are functionalized with different ligands. Several kinds of ligand-modified nanomedicines have been reported. Cell-penetrating peptides (CPPs) are promising ligands, attracting the attention of researchers due to their efficiency to transport bioactive molecules intracellularly. However, their lack of specificity and in vivo degradation led to the development of newer types of CPP. Currently, activable CPP and tumor-targeting peptide (TTP)-modified nanocarriers have shown dramatically superior cellular specific uptake, cytotoxicity, and tumor growth inhibition. In this review, we discuss recent advances in tumor-targeting strategies using CPPs and their limitations in tumor delivery systems. Special emphasis is given to activable CPPs and TTPs. Finally, we address the application of CPPs and/or TTPs in the delivery of plant-derived chemotherapeutic agents. PMID:29563797

Deep magnetic capture of magnetically loaded cells for spatially targeted therapeutics.

PubMed

Huang, Zheyong; Pei, Ning; Wang, Yanyan; Xie, Xinxing; Sun, Aijun; Shen, Li; Zhang, Shuning; Liu, Xuebo; Zou, Yunzeng; Qian, Juying; Ge, Junbo

2010-03-01

Magnetic targeting has recently demonstrated potential in promoting magnetically loaded cell delivery to target lesion, but its application is limited by magnetic attenuation. For deep magnetic capture of cells for spatial targeting therapeutics, we designed a magnetic pole, in which the magnetic field density can be focused at a distance from the pole. As flowing through a tube served as a model of blood vessels, the magnetically loaded mesenchymal stem cells (MagMSCs) were highly enriched at the site distance from the magnetic pole. The cell capture efficiency was positively influenced by the magnetic flux density, and inversely influenced by the flow velocity, and well-fitted with the deductive value by theoretical considerations. It appeared to us that the spatially-focused property of the magnetic apparatus promises a new deep targeting strategy to promote homing and engraftment for cellular therapy. Copyright (c) 2009 Elsevier Ltd. All rights reserved.

MPN estimation of qPCR target sequence recoveries from whole cell calibrator samples.

PubMed

Sivaganesan, Mano; Siefring, Shawn; Varma, Manju; Haugland, Richard A

2011-12-01

DNA extracts from enumerated target organism cells (calibrator samples) have been used for estimating Enterococcus cell equivalent densities in surface waters by a comparative cycle threshold (Ct) qPCR analysis method. To compare surface water Enterococcus density estimates from different studies by this approach, either a consistent source of calibrator cells must be used or the estimates must account for any differences in target sequence recoveries from different sources of calibrator cells. In this report we describe two methods for estimating target sequence recoveries from whole cell calibrator samples based on qPCR analyses of their serially diluted DNA extracts and most probable number (MPN) calculation. The first method employed a traditional MPN calculation approach. The second method employed a Bayesian hierarchical statistical modeling approach and a Monte Carlo Markov Chain (MCMC) simulation method to account for the uncertainty in these estimates associated with different individual samples of the cell preparations, different dilutions of the DNA extracts and different qPCR analytical runs. The two methods were applied to estimate mean target sequence recoveries per cell from two different lots of a commercially available source of enumerated Enterococcus cell preparations. The mean target sequence recovery estimates (and standard errors) per cell from Lot A and B cell preparations by the Bayesian method were 22.73 (3.4) and 11.76 (2.4), respectively, when the data were adjusted for potential false positive results. Means were similar for the traditional MPN approach which cannot comparably assess uncertainty in the estimates. Cell numbers and estimates of recoverable target sequences in calibrator samples prepared from the two cell sources were also used to estimate cell equivalent and target sequence quantities recovered from surface water samples in a comparative Ct method. Our results illustrate the utility of the Bayesian method in accounting for

Activated ovarian endothelial cells promote early follicular development and survival.

PubMed

Kedem, Alon; Aelion-Brauer, Anate; Guo, Peipei; Wen, Duancheng; Ding, Bi-Sen; Lis, Raphael; Cheng, Du; Sandler, Vladislav M; Rafii, Shahin; Rosenwaks, Zev

2017-09-19

New data suggests that endothelial cells (ECs) elaborate essential "angiocrine factors". The aim of this study is to investigate the role of activated ovarian endothelial cells in early in-vitro follicular development. Mouse ovarian ECs were isolated using magnetic cell sorting or by FACS and cultured in serum free media. After a constitutive activation of the Akt pathway was initiated, early follicles (50-150 um) were mechanically isolated from 8-day-old mice and co-cultured with these activated ovarian endothelial cells (AOEC) (n = 32), gel (n = 24) or within matrigel (n = 27) in serum free media for 14 days. Follicular growth, survival and function were assessed. After 6 passages, flow cytometry showed 93% of cells grown in serum-free culture were VE-cadherin positive, CD-31 positive and CD 45 negative, matching the known EC profile. Beginning on day 4 of culture, we observed significantly higher follicular and oocyte growth rates in follicles co-cultured with AOECs compared with follicles on gel or matrigel. After 14 days of culture, 73% of primary follicles and 83% of secondary follicles co-cultured with AOEC survived, whereas the majority of follicles cultured on gel or matrigel underwent atresia. This is the first report of successful isolation and culture of ovarian ECs. We suggest that co-culture with activated ovarian ECs promotes early follicular development and survival. This model is a novel platform for the in vitro maturation of early follicles and for the future exploration of endothelial-follicular communication. In vitro development of early follicles necessitates a complex interplay of growth factors and signals required for development. Endothelial cells (ECs) may elaborate essential "angiocrine factors" involved in organ regeneration. We demonstrate that co-culture with ovarian ECs enables culture of primary and early secondary mouse ovarian follicles.

Liver cell-targeted delivery of therapeutic molecules.

PubMed

Kang, Jeong-Hun; Toita, Riki; Murata, Masaharu

2016-01-01

The liver is the largest internal organ in mammals and is involved in metabolism, detoxification, synthesis of proteins and lipids, secretion of cytokines and growth factors and immune/inflammatory responses. Hepatitis, alcoholic or non-alcoholic liver disease, hepatocellular carcinoma, hepatic veno-occlusive disease, and liver fibrosis and cirrhosis are the most common liver diseases. Safe and efficient delivery of therapeutic molecules (drugs, genes or proteins) into the liver is very important to increase the clinical efficacy of these molecules and to reduce their side effects in other organs. Several liver cell-targeted delivery systems have been developed and tested in vivo or ex vivo/in vitro. In this review, we discuss the literature concerning liver cell-targeted delivery systems, with a particular emphasis on the results of in vivo studies.

Metabolic and structural integrity of magnetic nanoparticle-loaded primary endothelial cells for targeted cell therapy.

PubMed

Orynbayeva, Zulfiya; Sensenig, Richard; Polyak, Boris

2015-05-01

To successfully translate magnetically mediated cell targeting from bench to bedside, there is a need to systematically assess the potential adverse effects of magnetic nanoparticles (MNPs) interacting with 'therapeutic' cells. Here, we examined in detail the effects of internalized polymeric MNPs on primary rat endothelial cells' structural intactness, metabolic integrity and proliferation potential. The intactness of cytoskeleton and organelles was studied by fluorescent confocal microscopy, flow cytometry and high-resolution respirometry. MNP-loaded primary endothelial cells preserve intact cytoskeleton and organelles, maintain normal rate of proliferation, calcium signaling and mitochondria energy metabolism. This study provides supportive evidence that MNPs at doses necessary for targeting did not induce significant adverse effects on structural integrity and functionality of primary endothelial cells - potential cell therapy vectors.

HPV16 early gene E5 specifically reduces miRNA-196a in cervical cancer cells

PubMed Central

Liu, Chanzhen; Lin, Jianfei; Li, Lianqin; Zhang, Yonggang; Chen, Weiling; Cao, Zeyi; Zuo, Huancong; Chen, Chunling; Kee, Kehkooi

2015-01-01

High-risk human papillomavirus (HPV) type 16, which is responsible for greater than 50% of cervical cancer cases, is the most prevalent and lethal HPV type. However, the molecular mechanisms of cervical carcinogenesis remain elusive, particularly the early steps of HPV infection that may transform normal cervical epithelium into a pre-neoplastic state. Here, we report that a group of microRNAs (microRNAs) were aberrantly decreased in HPV16-positive normal cervical tissues, and these groups of microRNAs are further reduced in cervical carcinoma. Among these miRNAs, miR196a expression is the most reduced in HPV16-infected tissues. Interestingly, miR196a expression is low in HPV16-positive cervical cancer cell lines but high in HPV16-negative cervical cancer cell lines. Furthermore, we found that only HPV16 early gene E5 specifically down-regulated miRNA196a in the cervical cancer cell lines. In addition, HoxB8, a known miR196a target gene, is up-regulated in the HPV16 cervical carcinoma cell line but not in HPV18 cervical cancer cell lines. Various doses of miR196a affected cervical cancer cell proliferation and apoptosis. Altogether, these results suggested that HPV16 E5 specifically down-regulates miR196a upon infection of the human cervix and initiates the transformation of normal cervix cells to cervical carcinoma. PMID:25563170

A 20-Amino Acid Module of Protein Kinase Cϵ Involved in Translocation and Selective Targeting at Cell-Cell Contacts*

PubMed Central

Diouf, Barthélémy; Collazos, Alejandra; Labesse, Gilles; Macari, Françoise; Choquet, Armelle; Clair, Philippe; Gauthier-Rouvière, Cécile; Guérineau, Nathalie C.; Jay, Philippe; Hollande, Frédéric; Joubert, Dominique

2009-01-01

In the pituitary gland, activated protein kinase C (PKC) isoforms accumulate either selectively at the cell-cell contact (α and ϵ) or at the entire plasma membrane (β1 and δ). The molecular mechanisms underlying these various subcellular locations are not known. Here, we demonstrate the existence within PKCϵ of a cell-cell contact targeting sequence (3CTS) that, upon stimulation, is capable of targeting PKCδ, chimerin-α1, and the PKCϵ C1 domain to the cell-cell contact. We show that this selective targeting of PKCϵ is lost upon overexpression of 3CTS fused to a (R-Ahx-R)4 (where Ahx is 6-aminohexanoic acid) vectorization peptide, reflecting a dominant-negative effect of the overexpressed 3CTS on targeting selectivity. 3CTS contains a putative amphipathic α-helix, a 14-3-3-binding site, and the Glu-374 amino acid, involved in targeting selectivity. We show that the integrity of the α-helix is important for translocation but that 14-3-3 is not involved in targeting selectivity. However, PKCϵ translocation is increased when PKCϵ/14-3-3 interaction is abolished, suggesting that phorbol 12-myristate 13-acetate activation may initiate two sets of PKCϵ functions, those depending on 14-3-3 and those depending on translocation to cell-cell contacts. Thus, 3CTS is involved in the modulation of translocation via its 14-3-3-binding site, in cytoplasmic desequestration via the α-helix, and in selective PKCϵ targeting at the cell-cell contact via Glu-374. PMID:19429675

E-selectin liposomal and nanotube-targeted delivery of doxorubicin to circulating tumor cells

PubMed Central

Mitchell, Michael J.; Chen, Christina S.; Ponmudi, Varun; Hughes, Andrew D.; King, Michael R.

2012-01-01

The presence of circulating tumor cells (CTCs) is believed to lead to the formation of secondary tumors via an adhesion cascade involving interaction between adhesion receptors of endothelial cells and ligands on CTCs. Many CTCs express sialylated carbohydrate ligands on their surfaces that adhere to selectin protein found on inflamed endothelial cells. We have investigated the feasibility of using immobilized selectin proteins as a targeting mechanism for CTCs under flow. Herein, targeted liposomal doxorubicin (L-DXR) was functionalized with recombinant human E-selectin (ES) and polyethylene glycol (PEG) to target and kill cancer cells under shear flow, both when immobilized along a microtube device or sheared in a cone-and-plate viscometer in a dilute suspension. Healthy circulating cells such as red blood cells were not targeted by this mechanism and were left to freely circulate, and minimal leukocyte death was observed. Halloysite nanotube (HNT)-coated microtube devices immobilized with nanoscale liposomes significantly enhanced the targeting, capture, and killing of cancer cells. This work demonstrates that E-selectin functionalized L-DXR, sheared in suspension or immobilized onto microtube devices, provides a novel approach to selectively target and deliver chemotherapeutics to CTCs in the bloodstream. PMID:22421423

Concise Review: Cell Surface N-Linked Glycoproteins as Potential Stem Cell Markers and Drug Targets.

PubMed

Boheler, Kenneth R; Gundry, Rebekah L

2017-01-01

Stem cells and their derivatives hold great promise to advance regenerative medicine. Critical to the progression of this field is the identification and utilization of antibody-accessible cell-surface proteins for immunophenotyping and cell sorting-techniques essential for assessment and isolation of defined cell populations with known functional and therapeutic properties. Beyond their utility for cell identification and selection, cell-surface proteins are also major targets for pharmacological intervention. Although comprehensive cell-surface protein maps are highly valuable, they have been difficult to define until recently. In this review, we discuss the application of a contemporary targeted chemoproteomic-based technique for defining the cell-surface proteomes of stem and progenitor cells. In applying this approach to pluripotent stem cells (PSCs), these studies have improved the biological understanding of these cells, led to the enhanced use and development of antibodies suitable for immunophenotyping and sorting, and contributed to the repurposing of existing drugs without the need for high-throughput screening. The utility of this latter approach was first demonstrated with human PSCs (hPSCs) through the identification of small molecules that are selectively toxic to hPSCs and have the potential for eliminating confounding and tumorigenic cells in hPSC-derived progeny destined for research and transplantation. Overall, the cutting-edge technologies reviewed here will accelerate the development of novel cell-surface protein targets for immunophenotyping, new reagents to improve the isolation of therapeutically qualified cells, and pharmacological studies to advance the treatment of intractable diseases amenable to cell-replacement therapies. Stem Cells Translational Medicine 2017;6:131-138. © 2016 The Authors Stem Cells Translational Medicine published by Wiley Periodicals, Inc. on behalf of AlphaMed Press.

Intracellular CXCR4+ cell targeting with T22-empowered protein-only nanoparticles

PubMed Central

Unzueta, Ugutz; Céspedes, María Virtudes; Ferrer-Miralles, Neus; Casanova, Isolda; Cedano, Juan; Corchero, José Luis; Domingo-Espín, Joan; Villaverde, Antonio; Mangues, Ramón; Vázquez, Esther

2012-01-01

Background Cell-targeting peptides or proteins are appealing tools in nanomedicine and innovative medicines because they increase the local drug concentration and reduce potential side effects. CXC chemokine receptor 4 (CXCR4) is a cell surface marker associated with several severe human pathologies, including colorectal cancer, for which intracellular targeting agents are currently missing. Results Four different peptides that bind CXCR4 were tested for their ability to internalize a green fluorescent protein-based reporter nanoparticle into CXCR4+ cells. Among them, only the 18 mer peptide T22, an engineered segment derivative of polyphemusin II from the horseshoe crab, efficiently penetrated target cells via a rapid, receptor-specific endosomal route. This resulted in accumulation of the reporter nanoparticle in a fully fluorescent and stable form in the perinuclear region of the target cells, without toxicity either in cell culture or in an in vivo model of metastatic colorectal cancer. Conclusion Given the urgent demand for targeting agents in the research, diagnosis, and treatment of CXCR4-linked diseases, including colorectal cancer and human immunodeficiency virus infection, T22 appears to be a promising tag for the intracellular delivery of protein drugs, nanoparticles, and imaging agents. PMID:22923991

Aptamer-Mediated Up-conversion Core/MOF Shell Nanocomposites for Targeted Drug Delivery and Cell Imaging

PubMed Central

Deng, Kerong; Hou, Zhiyao; Li, Xuejiao; Li, Chunxia; Zhang, Yuanxin; Deng, Xiaoran; Cheng, Ziyong; Lin, Jun

2015-01-01

Multifunctional nanocarriers for targeted bioimaging and drug delivery have attracted much attention in early diagnosis and therapy of cancer. In this work, we develop a novel aptamer-guided nanocarrier based on the mesoporous metal-organic framework (MOF) shell and up-conversion luminescent NaYF4:Yb3+/Er3+ nanoparticles (UCNPs) core for the first time to achieve these goals. These UCNPs, chosen as optical labels in biological assays and medical imaging, could emit strong green emission under 980 nm laser. The MOF structure based on iron (III) carboxylate materials [MIL-100 (Fe)] possesses high porosity and non-toxicity, which is of great value as nanocarriers for drug storage/delivery. As a unique nanoplatform, the hybrid inorganic-organic drug delivery vehicles show great promising for simultaneous targeted labeling and therapy of cancer cells. PMID:25597762

miR-1271 promotes non-small-cell lung cancer cell proliferation and invasion via targeting HOXA5

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang, Yongfang; Xu, Lianhong; Jiang, Lixin, E-mail: jianglx66766@163.com

2015-03-13

MicroRNAs (miRNAs) are short, non-coding RNAs (∼22 nt) that play important roles in the pathogenesis of human diseases by negatively regulating numerous target genes at posttranscriptional level. However, the role of microRNAs in lung cancer, particularly non-small-cell lung cancer (NSCLC), has remained elusive. In this study, two microRNAs, miR-1271 and miR-628, and their predicted target genes were identified differentially expressed in NSCLC by analyzing the miRNA and mRNA expression data from NSCLC tissues and their matching normal controls. miR-1271 and its target gene HOXA5 were selected for further investigation. CCK-8 proliferation assay showed that the cell proliferation was promoted by miR-1271more » in NSCLC cells, while miR-1271 inhibitor could significantly inhibited the proliferation of NSCLC cells. Interestingly, migration and invasion assay indicated that overexpression of miR-1271 could significantly promoted the migration and invasion of NSCLC cells, whereas miR-1271 inhibitor could inhibited both cell migration and invasion of NSCLC cells. Western blot showed that miR-1271 suppressed the protein level of HOXA5, and luciferase assays confirmed that miR-1271 directly bound to the 3'untranslated region of HOXA5. This study indicated indicate that miR-1271 regulates NSCLC cell proliferation and invasion, via the down-regulation of HOXA5. Thus, miR-1271 may represent a potential therapeutic target for NSCLC intervention. - Highlights: • Overexpression of miR-1271 promoted proliferation and invasion of NSCLC cells. • miR-1271 inhibitor inhibited the proliferation and invasion of NSCLC cells. • miR-1271 targets 3′ UTR of HOXA5 in NSCLC cells. • miR-1271 negatively regulates HOXA5 in NSCLC cells.« less

Elevated Ratio of Th17 Cell-Derived Th1 Cells (CD161(+)Th1 Cells) to CD161(+)Th17 Cells in Peripheral Blood of Early-Onset Rheumatoid Arthritis Patients.

PubMed

Kotake, Shigeru; Nanke, Yuki; Yago, Toru; Kawamoto, Manabu; Kobashigawa, Tsuyoshi; Yamanaka, Hisashi

2016-01-01

Rheumatoid arthritis (RA) is a chronic inflammatory disease characterized by the destruction of articular cartilage and bone with elevated levels of proinflammatory cytokines. It has been reported that IL-17 and Th17 cells play important roles in the pathogenesis of RA. Recently, plasticity in helper T cells has been demonstrated; Th17 cells can convert to Th1 cells. It remains to be elucidated whether this conversion occurs in the early phase of RA. Here, we tried to identify Th17 cells, Th1 cells, and Th17 cell-derived Th1 cells (CD161(+)Th1 cells) in the peripheral blood of early-onset RA patients. We also evaluated the effect of methotrexate on the ratio of Th17 cells in early-onset RA patients. The ratio of Th17 cell-derived Th1 cells to CD161(+)Th17 cells was elevated in the peripheral blood of early-onset RA patients. In addition, MTX reduced the ratio of Th17 cells but not Th1 cells. These findings suggest that IL-17 and Th17 play important roles in the early phase of RA; thus, anti-IL-17 antibodies should be administered to patients with RA in the early phase.

Elevated Ratio of Th17 Cell-Derived Th1 Cells (CD161+Th1 Cells) to CD161+Th17 Cells in Peripheral Blood of Early-Onset Rheumatoid Arthritis Patients

PubMed Central

Kotake, Shigeru; Nanke, Yuki; Yago, Toru; Kawamoto, Manabu; Kobashigawa, Tsuyoshi; Yamanaka, Hisashi

2016-01-01

Rheumatoid arthritis (RA) is a chronic inflammatory disease characterized by the destruction of articular cartilage and bone with elevated levels of proinflammatory cytokines. It has been reported that IL-17 and Th17 cells play important roles in the pathogenesis of RA. Recently, plasticity in helper T cells has been demonstrated; Th17 cells can convert to Th1 cells. It remains to be elucidated whether this conversion occurs in the early phase of RA. Here, we tried to identify Th17 cells, Th1 cells, and Th17 cell-derived Th1 cells (CD161+Th1 cells) in the peripheral blood of early-onset RA patients. We also evaluated the effect of methotrexate on the ratio of Th17 cells in early-onset RA patients. The ratio of Th17 cell-derived Th1 cells to CD161+Th17 cells was elevated in the peripheral blood of early-onset RA patients. In addition, MTX reduced the ratio of Th17 cells but not Th1 cells. These findings suggest that IL-17 and Th17 play important roles in the early phase of RA; thus, anti-IL-17 antibodies should be administered to patients with RA in the early phase. PMID:27123445

JAK2V617F expression in mice amplifies early hematopoietic cells and gives them a competitive advantage that is hampered by IFNα.

PubMed

Hasan, Salma; Lacout, Catherine; Marty, Caroline; Cuingnet, Marie; Solary, Eric; Vainchenker, William; Villeval, Jean-Luc

2013-08-22

The acquired gain-of-function V617F mutation in the Janus Kinase 2 (JAK2(V617F)) is the main mutation involved in BCR/ABL-negative myeloproliferative neoplasms (MPNs), but its effect on hematopoietic stem cells as a driver of disease emergence has been questioned. Therefore, we reinvestigated the role of endogenous expression of JAK2(V617F) on early steps of hematopoiesis as well as the effect of interferon-α (IFNα), which may target the JAK2(V617F) clone in humans by using knock-in mice with conditional expression of JAK2(V617F) in hematopoietic cells. These mice develop a MPN mimicking polycythemia vera with large amplification of myeloid mature and precursor cells, displaying erythroid endogenous growth and progressing to myelofibrosis. Interestingly, early hematopoietic compartments [Lin-, LSK, and SLAM (LSK/CD48-/CD150+)] increased with the age. Competitive repopulation assays demonstrated disease appearance and progressive overgrowth of myeloid, Lin-, LSK, and SLAM cells, but not lymphocytes, from a low number of engrafted JAK2(V617F) SLAM cells. Finally, IFNα treatment prevented disease development by specifically inhibiting JAK2(V617F) cells at an early stage of differentiation and eradicating disease-initiating cells. This study shows that JAK2(V617F) in mice amplifies not only late but also early hematopoietic cells, giving them a proliferative advantage through high cell cycling and low apoptosis that may sustain MPN emergence but is lost upon IFNα treatment.

A perspective on B-cell-targeting therapy for SLE.

PubMed

Looney, R John; Anolik, Jennifer; Sanz, Inaki

2010-02-01

In recent years, large controlled trials have tested several new agents for systemic lupus erythematosus (SLE). Unfortunately, none of these trials has met its primary outcome. This does not mean progress has not been made. In fact, a great deal has been learned about doing clinical trials in lupus and about the biological and clinical effects of the drugs being tested. Many of these drugs were designed to target B cells directly, e.g., rituximab, belimumab, epratuzumab, and transmembrane activator and calcium modulator and cyclophilin ligand interactor-immunoglobulin (TACI-Ig). The enthusiasm for targeting B cells derives from substantial evidence showing the critical role of B cells in murine models of SLE, as well promising results from multiple open trials with rituximab, a chimeric anti-CD20 monoclonal antibody that specifically depletes B cells (Martin and Chan in Immunity 20(5):517-527, 2004; Sobel et al. in J Exp Med 173:1441-1449, 1991; Silverman and Weisman in Arthritis Rheum 48:1484-1492, 2003; Silverman in Arthritis Rheum 52(4):1342, 2005; Shlomchik et al. in Nat Rev Immunol 1:147-153, 2001; Looney et al. in Arthritis Rheum 50:2580-2589, 2004; Lu et al. in Arthritis Rheum 61(4):482-487, 2009; Saito et al. in Lupus 12(10):798-800, 2003; van Vollenhoven et al. in Scand J Rheumatol 33(6):423-427, 2004; Sfikakis et al. Arthritis Rheum 52(2):501-513, 2005). Why have the controlled trials of B-cell-targeting therapies failed to demonstrate efficacy? Were there flaws in design or execution of these trials? Or, were promising animal studies and open trials misleading, as so often happens? This perspective discusses the current state of B-cell-targeting therapies for human lupus and the future development of these therapies.

VEGFR2-targeted fusion antibody improved NK cell-mediated immunosurveillance against K562 cells.

PubMed

Ren, Xueyan; Xie, Wei; Wang, Youfu; Xu, Menghuai; Liu, Fang; Tang, Mingying; Li, Chenchen; Wang, Min; Zhang, Juan

2016-08-01

MHC class I polypeptide-related sequence A (MICA), which is normally expressed on cancer cells, activates NK cells via NK group 2-member D pathway. However, some cancer cells escape NK-mediated immune surveillance by shedding membrane MICA causing immune suppression. To address this issue, we designed an antibody-MICA fusion targeting tumor-specific antigen (vascular endothelial growth factor receptor 2, VEGFR2) based on our patented antibody (mAb04) against VEGFR2. In vitro results demonstrate that the fusion antibody retains both the antineoplastic and the immunomodulatory activity of mAb04. Further, we revealed that it enhanced NK-mediated immunosurveillance against K562 cells through increasing degranulation and cytokine production of NK cells. The overall data suggest our new fusion protein provides a promising approach for cancer-targeted immunotherapy and has prospects for potential application of chronic myeloid leukemia.

Natural killer cells and regulatory T cells in early pregnancy loss

PubMed Central

SHARMA, SURENDRA

2015-01-01

Survival of the allogeneic embryo in the uterus depends on the maintenance of immune tolerance at the maternal-fetal interface. The pregnant uterus is replete with activated maternal immune cells. How this immune tolerance is acquired and maintained has been a topic of intense investigation. The key immune cells that predominantly populate the pregnant uterus are natural killer (NK) cells. In normal pregnancy, these cells are not killers, but rather provide a microenvironment that is pregnancy compatible and supports healthy placentation. In placental mammals, an array of highly orchestrated immune elements to support successful pregnancy outcome has been incorporated. This includes active cooperation between maternal immune cells, particularly NK cells, and trophoblast cells. This intricate process is required for placentation, immune regulation and to remodel the blood supply to the fetus. During the past decade, various types of maternal immune cells have been thought to be involved in cross-talk with trophoblasts and in programming immune tolerance. RegulatoryT cells (Tregs) have attracted a great deal of attention in promoting implantation and immune tolerance beyond implantation. However, what has not been fully addressed is how this immune-trophoblast axis breaks down during adverse pregnancy outcomes, particularly early pregnancy loss, and in response to unscheduled inflammation. Intense research efforts have begun to shed light on the roles of NK cells and Tregs in early pregnancy loss, although much remains to be unraveled in order to fully characterize the mechanisms underlying their detrimental activity. An increased understanding of host-environment interactions that lead to the cytotoxic phenotype of these otherwise pregnancy compatible maternal immune cells is important for prediction, prevention and treatment of pregnancy maladies, particularly recurrent pregnancy loss. In this review, we discuss relevant information from experimental and human

Catechol polymers for pH-responsive, targeted drug delivery to cancer cells.

PubMed

Su, Jing; Chen, Feng; Cryns, Vincent L; Messersmith, Phillip B

2011-08-10

A novel cell-targeting, pH-sensitive polymeric carrier was employed in this study for delivery of the anticancer drug bortezomib (BTZ) to cancer cells. Our strategy is based on facile conjugation of BTZ to catechol-containing polymeric carriers that are designed to be taken up selectively by cancer cells through cell surface receptor-mediated mechanisms. The polymer used as a building block in this study was poly(ethylene glycol), which was chosen for its ability to reduce nonspecific interactions with proteins and cells. The catechol moiety was exploited for its ability to bind and release borate-containing therapeutics such as BTZ in a pH-dependent manner. In acidic environments, such as in cancer tissue or the subcellular endosome, BTZ dissociates from the polymer-bound catechol groups to liberate the free drug, which inhibits proteasome function. A cancer-cell-targeting ligand, biotin, was presented on the polymer carriers to facilitate targeted entry of drug-loaded polymer carriers into cancer cells. Our study demonstrated that the cancer-targeting drug-polymer conjugates dramatically enhanced cellular uptake, proteasome inhibition, and cytotoxicity toward breast carcinoma cells in comparison with nontargeting drug-polymer conjugates. The pH-sensitive catechol-boronate binding mechanism provides a chemoselective approach for controlling the release of BTZ in targeted cancer cells, establishing a concept that may be applied in the future toward other boronic acid-containing therapeutics to treat a broad range of diseases. © 2011 American Chemical Society

Molecular Recognition of Human Liver Cancer Cells Using DNA Aptamers Generated via Cell-SELEX.

PubMed

Xu, Jiehua; Teng, I-Ting; Zhang, Liqin; Delgado, Stefanie; Champanhac, Carole; Cansiz, Sena; Wu, Cuichen; Shan, Hong; Tan, Weihong

2015-01-01

Most clinical cases of liver cancer cannot be diagnosed until they have evolved to an advanced stage, thus resulting in high mortality. It is well recognized that the implementation of early detection methods and the development of targeted therapies for liver cancer are essential to reducing the high mortality rates associated with this disease. To achieve these goals, molecular probes capable of recognizing liver cancer cell-specific targets are needed. Here we describe a panel of aptamers able to distinguish hepatocarcinoma from normal liver cells. The aptamers, which were selected by cell-based SELEX (Systematic Evolution of Ligands by Exponential Enrichment), have Kd values in the range of 64-349 nM toward the target human hepatoma cell HepG2, and also recognize ovarian cancer cells and lung adenocarcinoma. The proteinase treatment experiment indicated that all aptamers could recognize target HepG2 cells through surface proteins. This outcome suggested that these aptamers could be used as potential probes for further research in cancer studies, such as developing early detection assays, targeted therapies, and imaging agents, as well as for the investigation of common membrane proteins in these distinguishable cancers.

Targeting stromal glutamine synthetase in tumors disrupts tumor microenvironment-regulated cancer cell growth

USDA-ARS?s Scientific Manuscript database

Reactive stromal cells are an integral part of tumor microenvironment (TME) and interact with cancer cells to regulate their growth. Although targeting stromal cells could be a viable therapy to regulate the communication between TME and cancer cells, identification of stromal targets that make canc...

Targeting HIV Reservoir in Infected CD4 T Cells by Dual-Affinity Re-targeting Molecules (DARTs) that Bind HIV Envelope and Recruit Cytotoxic T Cells

PubMed Central

Sloan, Derek D.; Lam, Chia-Ying Kao; Irrinki, Alivelu; Liu, Liqin; Tsai, Angela; Pace, Craig S.; Kaur, Jasmine; Murry, Jeffrey P.; Balakrishnan, Mini; Moore, Paul A.; Johnson, Syd; Nordstrom, Jeffrey L.; Cihlar, Tomas; Koenig, Scott

2015-01-01

HIV reservoirs and production of viral antigens are not eliminated in chronically infected participants treated with combination antiretroviral therapy (cART). Novel therapeutic strategies aiming at viral reservoir elimination are needed to address chronic immune dysfunction and non-AIDS morbidities that exist despite effective cART. The HIV envelope protein (Env) is emerging as a highly specific viral target for therapeutic elimination of the persistent HIV-infected reservoirs via antibody-mediated cell killing. Dual-Affinity Re-Targeting (DART) molecules exhibit a distinct mechanism of action via binding the cell surface target antigen and simultaneously engaging CD3 on cytotoxic T lymphocytes (CTLs). We designed and evaluated Env-specific DARTs (HIVxCD3 DARTs) derived from known antibodies recognizing diverse Env epitopes with or without broadly neutralizing activity. HIVxCD3 DARTs derived from PGT121, PGT145, A32, and 7B2, but not VRC01 or 10E8 antibodies, mediated potent CTL-dependent killing of quiescent primary CD4 T cells infected with diverse HIV isolates. Similar killing activity was also observed with DARTs structurally modified for in vivo half-life extension. In an ex vivo model using cells isolated from HIV-infected participants on cART, combinations of the most potent HIVxCD3 DARTs reduced HIV expression both in quiescent and activated peripheral blood mononuclear cell cultures isolated from HIV-infected participants on suppressive cART. Importantly, HIVxCD3 DARTs did not induce cell-to-cell virus spread in resting or activated CD4 T cell cultures. Collectively, these results provide support for further development of HIVxCD3 DARTs as a promising therapeutic strategy for targeting HIV reservoirs. PMID:26539983

DNA breaks early in replication in B cell cancers

Cancer.gov

Research by scientists at the NCI has identified a new class of DNA sites in cells that break early in the replication process. They found that these break sites correlate with damage often seen in B cell cancers, such as diffuse large B cell lymphoma.

Circulating and disseminated tumor cells: diagnostic tools and therapeutic targets in motion

PubMed Central

Lin, Peter P.; Gires, Olivier

2017-01-01

Enumeration of circulating tumor cells (CTCs) in peripheral blood with the gold standard CellSearchTM has proven prognostic value for tumor recurrence and progression of metastatic disease. Therefore, the further molecular characterization of isolated CTCs might have clinical relevance as liquid biopsy for therapeutic decision-making and to monitor disease progression. The direct analysis of systemic cancer appears particularly important in view of the known disparity in expression of therapeutic targets as well as epithelial-to-mesenchymal transition (EMT)-based heterogeneity between primary and systemic tumor cells, which all substantially complicate monitoring and therapeutic targeting at present. Since CTCs are the potential precursor cells of metastasis, their in-depth molecular profiling should also provide a useful resource for target discovery. The present review will discuss the use of systemically spread cancer cells as liquid biopsy and focus on potential target antigens. PMID:27683128

Targeted delivery of celastrol to mesangial cells is effective against mesangioproliferative glomerulonephritis.

PubMed

Guo, Ling; Luo, Shi; Du, Zhengwu; Zhou, Meiling; Li, Peiwen; Fu, Yao; Sun, Xun; Huang, Yuan; Zhang, Zhirong

2017-10-12

Mesangial cells-mediated glomerulonephritis is a frequent cause of end-stage renal disease. Here, we show that celastrol is effective in treating both reversible and irreversible mesangioproliferative glomerulonephritis in rat models, but find that its off-target distributions cause severe systemic toxicity. We thus target celastrol to mesangial cells using albumin nanoparticles. Celastrol-albumin nanoparticles crosses fenestrated endothelium and accumulates in mesangial cells, alleviating proteinuria, inflammation, glomerular hypercellularity, and excessive extracellular matrix deposition in rat anti-Thy1.1 nephritis models. Celastrol-albumin nanoparticles presents lower drug accumulation than free celastrol in off-target organs and tissues, thereby minimizing celastrol-related systemic toxicity. Celastrol-albumin nanoparticles thus represents a promising treatment option for mesangioproliferative glomerulonephritis and similar glomerular diseases.Mesangial cell-mediated glomerulonephritis is a frequent cause of kidney disease. Here the authors show that celastrol loaded in albumin nanoparticles efficiently targets mesangial cells, and is effective in rat models.

Identification and validation nucleolin as a target of curcumol in nasopharyngeal carcinoma cells.

PubMed

Wang, Juan; Wu, Jiacai; Li, Xumei; Liu, Haowei; Qin, Jianli; Bai, Zhun; Chi, Bixia; Chen, Xu

2018-06-30

Identification of the specific protein target(s) of a drug is a critical step in unraveling its mechanisms of action (MOA) in many natural products. Curcumol, isolated from well known Chinese medicinal plant Curcuma zedoary, has been shown to possess multiple biological activities. It can inhibit nasopharyngeal carcinoma (NPC) proliferation and induce apoptosis, but its target protein(s) in NPC cells remains unclear. In this study, we employed a mass spectrometry-based chemical proteomics approach reveal the possible protein targets of curcumol in NPC cells. Cellular thermal shift assay (CETSA), molecular docking and cell-based assay was used to validate the binding interactions. Chemical proteomics capturing uncovered that NCL is a target of curcumol in NPC cells, Molecular docking showed that curcumol bound to NCL with an -7.8 kcal/mol binding free energy. Cell function analysis found that curcumol's treatment leads to a degradation of NCL in NPC cells, and it showed slight effects on NP69 cells. In conclusion, our results providing evidences that NCL is a target protein of curcumol. We revealed that the anti-cancer effects of curcumol in NPC cells are mediated, at least in part, by NCL inhibition. Many natural products showed high bioactivity, while their mechanisms of action (MOA) are very poor or completely missed. Understanding the MOA of natural drugs can thoroughly exploit their therapeutic potential and minimize their adverse side effects. Identification of the specific protein target(s) of a drug is a critical step in unraveling its MOA. Compound-centric chemical proteomics is a classic chemical proteomics approach which integrates chemical synthesis with cell biology and mass spectrometry (MS) to identify protein targets of natural products determine the drug mechanism of action, describe its toxicity, and figure out the possible cause of off-target. It is an affinity-based chemical proteomics method to identify small molecule-protein interactions

Viral Capsid DNA Aptamer Conjugates as Multivalent Cell Targeting Vehicles

PubMed Central

Tong, Gary J.; Hsiao, Sonny C.; Carrico, Zachary M.; Francis, Matthew B.

2009-01-01

Nucleic acid aptamers offer significant potential as convenient and evolvable targeting groups for drug delivery. To attach them to the surface of a genome-free viral capsid carrier, an efficient oxidative coupling strategy has been developed. The method involves the periodate-mediated reaction of phenylene diamine substituted oligonucleotides with aniline groups installed on the outer surface of the capsid shells. Up to 60 DNA strands can be attached to each viral capsid with no apparent loss of base-pairing capabilities or protein stability. The ability of the capsids to bind specific cellular targets was demonstrated through the attachment of a 41-nucleotide sequence that targets a tyrosine kinase receptor on Jurkat T cells. After the installation of a fluorescent dye on the capsid interior, capsids bearing the cell-targeting sequence showed significant levels of binding to the cells relative to control samples. Colocalization experiments using confocal microscopy indicated that the capsids were endocytosed and trafficked to lysosomes for degradation. These observations suggest that aptamer-labeled capsids could be used for the targeted drug delivery of acid-labile prodrugs that would be preferentially released upon lysosomal acidification. PMID:19603808

Interferon gamma peptidomimetic targeted to hepatic stellate cells ameliorates acute and chronic liver fibrosis in vivo.

PubMed

Bansal, Ruchi; Prakash, Jai; De Ruiter, Marieke; Poelstra, Klaas

2014-04-10

Hepatic stellate cells play a crucial role in the pathogenesis of hepatic fibrosis. Thus, pharmacological inhibition of pro-fibrotic activities of these cells might lead to an effective therapy for this disease. Among the potent anti-fibrotics, interferon gamma (IFNγ), a proinflammatory cytokine, is highly efficacious but it failed in clinical trials due to the poor efficacy and multiple adverse effects attributed to the ubiquitous IFNγ receptor (IFNγR) expression. To resolve these drawbacks, we chemically synthesized a chimeric molecule containing (a) IFNγ signaling peptide (IFNγ peptidomimetic, mimγ) that retains the agonistic activities of IFNγ but lacks an extracellular receptor recognition sequence for IFNγR; coupled via heterobifunctional PEG linker to (b) bicyclic platelet derived growth factor beta receptor (PDGFβR)-binding peptide (BiPPB) to induce internalization into the stellate cells that express PDGFβR. The synthesized targeted IFNγ peptidomimetic (mimγ-BiPPB) was extensively investigated for its anti-fibrotic and adverse effects in acute and chronic CCl4-induced liver fibrosis models in mice. Treatment with mimγ-BiPPB, after the onset of disease, markedly inhibited both early and established hepatic fibrosis as reflected by a reduced intrahepatic α-SMA, desmin and collagen-I mRNA expression and protein levels. While untargeted mimγ and BiPPB had no effect, and native IFNγ only induced a moderate reduction. Additionally, no off-target effects, e.g. systemic inflammation, were found with mimγ-BiPPB, which were substantially observed in mice treated with native IFNγ. The present study highlights the beneficial effects of a novel BiPPB mediated cell-specific targeting of IFNγ peptidomimetic to the disease-inducing cells and therefore represents a highly potential therapeutic approach to treat fibrotic diseases. Copyright © 2014 Elsevier B.V. All rights reserved.

Targeting inflammation in pancreatic cancer: Clinical translation

PubMed Central

Steele, Colin William; Kaur Gill, Nina Angharad; Jamieson, Nigel Balfour; Carter, Christopher Ross

2016-01-01

Preclinical modelling studies are beginning to aid development of therapies targeted against key regulators of pancreatic cancer progression. Pancreatic cancer is an aggressive, stromally-rich tumor, from which few people survive. Within the tumor microenvironment cellular and extracellular components exist, shielding tumor cells from immune cell clearance, and chemotherapy, enhancing progression of the disease. The cellular component of this microenvironment consists mainly of stellate cells and inflammatory cells. New findings suggest that manipulation of the cellular component of the tumor microenvironment is possible to promote immune cell killing of tumor cells. Here we explore possible immunogenic therapeutic strategies. Additionally extracellular stromal elements play a key role in protecting tumor cells from chemotherapies targeted at the pancreas. We describe the experimental findings and the pitfalls associated with translation of stromally targeted therapies to clinical trial. Finally, we discuss the key inflammatory signal transducers activated subsequent to driver mutations in oncogenic Kras in pancreatic cancer. We present the preclinical findings that have led to successful early trials of STAT3 inhibitors in pancreatic adenocarcinoma. PMID:27096033

miRNA-1297 induces cell proliferation by targeting phosphatase and tensin homolog in testicular germ cell tumor cells.

PubMed

Yang, Nian-Qin; Zhang, Jian; Tang, Qun-Ye; Guo, Jian-Ming; Wang, Guo-Min

2014-01-01

To investigate the role of miR-1297 and the tumor suppressor gene PTEN in cell proliferation of testicular germ cell tumors (TGCT). MTT assays were used to test the effect of miR-1297 on proliferation of the NCCIT testicular germ cell tumor cell line. In NCCIT cells, the expression of PTEN was assessed by Western blotting further. In order to confirm target association between miR-1297 and 3'-UTR of PTEN, a luciferase reporter activity assay was employed. Moreover, roles of PTEN in proliferation of NCCIT cells were evaluated by transfection of PTEN siRNA. Proliferation of NCCIT cells was promoted by miR-1297 in a concentration-dependent manner. In addition, miR-1297 could bind to the 3'-UTR of PTEN based on luciferase reporter activity assay, and reduced expression of PTEN at protein level was found. Proliferation of NCCIT cells was significantly enhanced after knockdown of PTEN by siRNA. miR-1297 as a potential oncogene could induce cell proliferation by targeting PTEN in NCCIT cells.

Target cell specific antibody-based photosensitizers for photodynamic therapy

NASA Astrophysics Data System (ADS)

Rosenblum, Lauren T.; Mitsunaga, Makoto; Kakareka, John W.; Morgan, Nicole Y.; Pohida, Thomas J.; Choyke, Peter L.; Kobayashi, Hisataka

2011-03-01

In photodynamic therapy (PDT), localized monochromatic light is used to activate targeted photosensitizers (PS) to induce cellular damage through the generation of cytotoxic species such as singlet oxygen. While first-generation PS passively targeted malignancies, a variety of targeting mechanisms have since been studied, including specifically activatable agents. Antibody internalization has previously been employed as a fluorescence activation system and could potentially enable similar activation of PS. TAMRA, Rhodamine-B and Rhodamine-6G were conjugated to trastuzumab (brand name Herceptin), a humanized monoclonal antibody with specificity for the human epidermal growth factor receptor 2 (HER2), to create quenched PS (Tra-TAM, Tra-RhoB, and Tra-Rho6G). Specific PDT with Tra-TAM and Tra-Rho6G, which formed covalently bound H-dimers, was demonstrated in HER2+ cells: Minimal cell death (<6%) was observed in all treatments of the HER2- cell line (BALB/3T3) and in treatments the HER2+ cell line (3T3/HER2) with light or trastuzumab only. There was significant light-induced cell death in HER2 expressing cells using Tra-TAM (3% dead without light, 20% at 50 J/cm2, 46% at 100 J/cm2) and Tra-Rho6G (5% dead without light, 22% at 50 J/cm2, 46% at 100 J/cm2). No efficacy was observed in treatment with Tra-RhoB, which was also non-specifically taken up by BALB/3T3 cells and which had weaker PS-antibody interactions (as demonstrated by visualization of protein and fluorescence on SDS-PAGE).

Delineating CD4 dependency of HIV-1: Adaptation to infect low level CD4 expressing target cells widens cellular tropism but severely impacts on envelope functionality

PubMed Central

Beauparlant, David; Rusert, Peter; Magnus, Carsten; Weber, Jacqueline; Uhr, Therese; Clapham, Paul R.; Metzner, Karin J.

2017-01-01

A hallmark of HIV-1 infection is the continuously declining number of the virus’ predominant target cells, activated CD4+ T cells. With diminishing CD4+ T cell levels, the capacity to utilize alternate cell types and receptors, including cells that express low CD4 receptor levels such as macrophages, thus becomes crucial. To explore evolutionary paths that allow HIV-1 to acquire a wider host cell range by infecting cells with lower CD4 levels, we dissected the evolution of the envelope-CD4 interaction under in vitro culture conditions that mimicked the decline of CD4high target cells, using a prototypic subtype B, R5-tropic strain. Adaptation to CD4low targets proved to severely alter envelope functions including trimer opening as indicated by a higher affinity to CD4 and loss in shielding against neutralizing antibodies. We observed a strikingly decreased infectivity on CD4high target cells, but sustained infectivity on CD4low targets, including macrophages. Intriguingly, the adaptation to CD4low targets altered the kinetic of the entry process, leading to rapid CD4 engagement and an extended transition time between CD4 and CCR5 binding during entry. This phenotype was also observed for certain central nervous system (CNS) derived macrophage-tropic viruses, highlighting that the functional perturbation we defined upon in vitro adaptation to CD4low targets occurs in vivo. Collectively, our findings suggest that CD4low adapted envelopes may exhibit severe deficiencies in entry fitness and shielding early in their evolution. Considering this, adaptation to CD4low targets may preferentially occur in a sheltered and immune-privileged environment such as the CNS to allow fitness restoring compensatory mutations to occur. PMID:28264054

TargetLink, a new method for identifying the endogenous target set of a specific microRNA in intact living cells.

PubMed

Xu, Yan; Chen, Yan; Li, Daliang; Liu, Qing; Xuan, Zhenyu; Li, Wen-Hong

2017-02-01

MicroRNAs are small non-coding RNAs acting as posttranscriptional repressors of gene expression. Identifying mRNA targets of a given miRNA remains an outstanding challenge in the field. We have developed a new experimental approach, TargetLink, that applied locked nucleic acid (LNA) as the affinity probe to enrich target genes of a specific microRNA in intact cells. TargetLink also consists a rigorous and systematic data analysis pipeline to identify target genes by comparing LNA-enriched sequences between experimental and control samples. Using miR-21 as a test microRNA, we identified 12 target genes of miR-21 in a human colorectal cancer cell by this approach. The majority of the identified targets interacted with miR-21 via imperfect seed pairing. Target validation confirmed that miR-21 repressed the expression of the identified targets. The cellular abundance of the identified miR-21 target transcripts varied over a wide range, with some targets expressed at a rather low level, confirming that both abundant and rare transcripts are susceptible to regulation by microRNAs, and that TargetLink is an efficient approach for identifying the target set of a specific microRNA in intact cells. C20orf111, one of the novel targets identified by TargetLink, was found to reside in the nuclear speckle and to be reliably repressed by miR-21 through the interaction at its coding sequence.

Cas9-mediated targeting of viral RNA in eukaryotic cells.

PubMed

Price, Aryn A; Sampson, Timothy R; Ratner, Hannah K; Grakoui, Arash; Weiss, David S

2015-05-12

Clustered, regularly interspaced, short palindromic repeats-CRISPR associated (CRISPR-Cas) systems are prokaryotic RNA-directed endonuclease machineries that act as an adaptive immune system against foreign genetic elements. Using small CRISPR RNAs that provide specificity, Cas proteins recognize and degrade nucleic acids. Our previous work demonstrated that the Cas9 endonuclease from Francisella novicida (FnCas9) is capable of targeting endogenous bacterial RNA. Here, we show that FnCas9 can be directed by an engineered RNA-targeting guide RNA to target and inhibit a human +ssRNA virus, hepatitis C virus, within eukaryotic cells. This work reveals a versatile and portable RNA-targeting system that can effectively function in eukaryotic cells and be programmed as an antiviral defense.

Cas9-mediated targeting of viral RNA in eukaryotic cells

PubMed Central

Price, Aryn A.; Sampson, Timothy R.; Ratner, Hannah K.; Grakoui, Arash; Weiss, David S.

2015-01-01

Clustered, regularly interspaced, short palindromic repeats–CRISPR associated (CRISPR-Cas) systems are prokaryotic RNA-directed endonuclease machineries that act as an adaptive immune system against foreign genetic elements. Using small CRISPR RNAs that provide specificity, Cas proteins recognize and degrade nucleic acids. Our previous work demonstrated that the Cas9 endonuclease from Francisella novicida (FnCas9) is capable of targeting endogenous bacterial RNA. Here, we show that FnCas9 can be directed by an engineered RNA-targeting guide RNA to target and inhibit a human +ssRNA virus, hepatitis C virus, within eukaryotic cells. This work reveals a versatile and portable RNA-targeting system that can effectively function in eukaryotic cells and be programmed as an antiviral defense. PMID:25918406

Pharmacological targets of breast cancer stem cells: a review.

PubMed

Pindiprolu, Sai Kiran S S; Krishnamurthy, Praveen T; Chintamaneni, Pavan Kumar

2018-05-01

Breast cancers contain small population of tumor-initiating cells called breast cancer stem cells (BCSCs), which are spared even after chemotherapy. Recently, BCSCs are implicated to be a cause of metastasis, tumor relapse, and therapy resistance in breast cancer. BCSCs have unique molecular mechanisms, which can be targeted to eliminate them. These include surface biomarkers, proteins involved in self-renewal pathways, drug efflux transporters, apoptotic/antiapoptotic proteins, autophagy, metabolism, and microenvironment regulation. The complex molecular mechanisms behind the survival of BCSCs and pharmacological targets for elimination of BCSCs are described in this review.

Cancer Nanotechnology: Recent Trends and Developments in Strategies for Targeting Cancer Cells to Improve Cancer Imaging and Treatment.

PubMed

Xu, Jingyao; Zhou, Xiaoling; Li, Yifei; Tian, Yudan

2017-01-01

Nanotechnology is a multidisciplinary field, which have the potential to cover applications in many subjects such as biology, chemistry and physics. The combined efforts of these subjects can lead to the successful engineering of nanodevices and nanovectors for targeted delivery and sensing/detection of cancer cells/tissues. The modulation of nanomaterials at surface and bulk level further adds value to this technology and develop strategies for early detection of precancerous and malignant cells from biological fluids. Furthermore, the novel nanotechnology-based imaging modalities have the prospects to offer non-invasive cancer imaging and treatment response study in real-time. This review covers the advantages of nanotechnology, which have been exploited for effective and targeted delivery of anti-cancer agents. Moreover, the initiatives taken by National Cancer Laboratory, USA to improve the clinical success of nanomedicines and nanovectors have also been comprehensively summarized. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

[Study on the hepatocytic cell targetability of liposomes].

PubMed

Hou, Xin-pu; Wang, Li; Wang, Xiang-tao; Li, Sha

2003-02-01

To target for hepatocytic cell, liposomes was modified by special ligand. Sterically stabilized liposomes (SSL) was conjugated with asialofeticin (AF), the ligand of asialoglycoprotein receptor (ASGP-R) of hepatocyte. ASGP-R-BLM is the ASGP-R reconstructed on bilayer lipid membrane (BLM). The recognition reaction between AF-SSL and ASGP-R-BLM can be monitored by the varieties of membrane electrical parameters. The targetability of AF-SSL mediated to hepatocyte was detected by radioisotopic labeled in vitro and in vivo. The therapeutic effect of antihepatocarcinoma was observed also. The lifetime of ASGP-R-BLM decreased with the added amount of AF-SSL. It was demonstrated that there was recognition reaction between AF-SSL and ASGP-R-BLM. The combination of AF-SSL with hepatocyte was significantly higher than that of SSL without AF-modified in vitro and in vivo. The survival time of rat for AF-SSL carriered ADM (adriamycin) group was much longer and the toxicities on heart, kidney and lung were lower than those SSL carried ADM group. It is possible to actively target the cell with specific receptor by ligand modified liposomes. The result prvide scientific basis of hepatocyte targeted liposomes.

Selective in vivo metabolic cell-labeling-mediated cancer targeting

PubMed Central

Wang, Hua; Wang, Ruibo; Cai, Kaimin; He, Hua; Liu, Yang; Yen, Jonathan; Wang, Zhiyu; Xu, Ming; Sun, Yiwen; Zhou, Xin; Yin, Qian; Tang, Li; Dobrucki, Iwona T; Dobrucki, Lawrence W; Chaney, Eric J; Boppart, Stephen A; Fan, Timothy M; Lezmi, Stéphane; Chen, Xuesi; Yin, Lichen; Cheng, Jianjun

2017-01-01

Distinguishing cancer cells from normal cells through surface receptors is vital for cancer diagnosis and targeted therapy. Metabolic glycoengineering of unnatural sugars provides a powerful tool to manually introduce chemical receptors onto the cell surface; however, cancer-selective labeling still remains a great challenge. Herein we report the design of sugars that can selectively label cancer cells both in vitro and in vivo. Specifically, we inhibit the cell-labeling activity of tetraacetyl-N-azidoacetylmannosamine (Ac4ManAz) by converting its anomeric acetyl group to a caged ether bond that can be selectively cleaved by cancer-overexpressed enzymes and thus enables the overexpression of azido groups on the surface of cancer cells. Histone deacetylase and cathepsin L-responsive acetylated azidomannosamine, one such enzymatically activatable Ac4ManAz analog developed, mediated cancer-selective labeling in vivo, which enhanced tumor accumulation of a dibenzocyclooctyne–doxorubicin conjugate via click chemistry and enabled targeted therapy against LS174T colon cancer, MDA-MB-231 triple-negative breast cancer and 4T1 metastatic breast cancer in mice. PMID:28192414

Essential Role of Chromatin Remodeling Protein Bptf in Early Mouse Embryos and Embryonic Stem Cells

PubMed Central

Landry, Joseph; Sharov, Alexei A.; Piao, Yulan; Sharova, Lioudmila V.; Xiao, Hua; Southon, Eileen; Matta, Jennifer; Tessarollo, Lino; Zhang, Ying E.; Ko, Minoru S. H.; Kuehn, Michael R.; Yamaguchi, Terry P.; Wu, Carl

2008-01-01

We have characterized the biological functions of the chromatin remodeling protein Bptf (Bromodomain PHD-finger Transcription Factor), the largest subunit of NURF (Nucleosome Remodeling Factor) in a mammal. Bptf mutants manifest growth defects at the post-implantation stage and are reabsorbed by E8.5. Histological analyses of lineage markers show that Bptf−/− embryos implant but fail to establish a functional distal visceral endoderm. Microarray analysis at early stages of differentiation has identified Bptf-dependent gene targets including homeobox transcriptions factors and genes essential for the development of ectoderm, mesoderm, and both definitive and visceral endoderm. Differentiation of Bptf−/− embryonic stem cell lines into embryoid bodies revealed its requirement for development of mesoderm, endoderm, and ectoderm tissue lineages, and uncovered many genes whose activation or repression are Bptf-dependent. We also provide functional and physical links between the Bptf-containing NURF complex and the Smad transcription factors. These results suggest that Bptf may co-regulate some gene targets of this pathway, which is essential for establishment of the visceral endoderm. We conclude that Bptf likely regulates genes and signaling pathways essential for the development of key tissues of the early mouse embryo. PMID:18974875

Comprehensive evaluation of Streptococcus sanguinis cell wall-anchored proteins in early infective endocarditis.

PubMed

Turner, Lauren Senty; Kanamoto, Taisei; Unoki, Takeshi; Munro, Cindy L; Wu, Hui; Kitten, Todd

2009-11-01

Streptococcus sanguinis is a member of the viridans group of streptococci and a leading cause of the life-threatening endovascular disease infective endocarditis. Initial contact with the cardiac infection site is likely mediated by S. sanguinis surface proteins. In an attempt to identify the proteins required for this crucial step in pathogenesis, we searched for surface-exposed, cell wall-anchored proteins encoded by S. sanguinis and then used a targeted signature-tagged mutagenesis (STM) approach to evaluate their contributions to virulence. Thirty-three predicted cell wall-anchored proteins were identified-a number much larger than those found in related species. The requirement of each cell wall-anchored protein for infective endocarditis was assessed in the rabbit model. It was found that no single cell wall-anchored protein was essential for the development of early infective endocarditis. STM screening was also employed for the evaluation of three predicted sortase transpeptidase enzymes, which mediate the cell surface presentation of cell wall-anchored proteins. The sortase A mutant exhibited a modest (approximately 2-fold) reduction in competitiveness, while the other two sortase mutants were indistinguishable from the parental strain. The combined results suggest that while cell wall-anchored proteins may play a role in S. sanguinis infective endocarditis, strategies designed to interfere with individual cell wall-anchored proteins or sortases would not be effective for disease prevention.

Novel therapeutic Strategies for Targeting Liver Cancer Stem Cells

PubMed Central

Oishi, Naoki; Wang, Xin Wei

2011-01-01

The cancer stem cell (CSC) hypothesis was first proposed over 40 years ago. Advances in CSC isolation were first achieved in hematological malignancies, with the first CSC demonstrated in acute myeloid leukemia. However, using similar strategies and technologies, and taking advantage of available surface markers, CSCs have been more recently demonstrated in a growing range of epithelial and other solid organ malignancies, suggesting that the majority of malignancies are dependent on such a compartment. Primary liver cancer consists predominantly of hepatocellular carcinoma (HCC) and intrahepatic cholangiocarcinoma (ICC). It is believed that hepatic progenitor cells (HPCs) could be the origin of some HCCs and ICCs. Furthermore, stem cell activators such as Wnt/β-catenin, TGF-β, Notch and Hedgehog signaling pathways also expedite tumorigenesis, and these pathways could serve as molecular targets to assist in designing cancer prevention strategies. Recent studies indicate that additional factors such as EpCAM, Lin28 or miR-181 may also contribute to HCC progression by targeting HCC CSCs. Various therapeutic drugs that directly modulate CSCs have been examined in vivo and in vitro. However, CSCs clearly have a complex pathogenesis, with a considerable crosstalk and redundancy in signaling pathways, and hence targeting single molecules or pathways may have a limited benefit for treatment. Many of the key signaling molecules are shared by both CSCs and normal stem cells, which add further challenges for designing molecularly targeted strategies specific to CSCs but sparing normal stem cells to avoid side effects. In addition to the direct control of CSCs, many other factors that are needed for the maintenance of CSCs, such as angiogenesis, vasculogenesis, invasion and migration, hypoxia, immune evasion, multiple drug resistance, and radioresistance, should be taken into consideration when designing therapeutic strategies for HCC. Here we provide a brief review of

Cancer active targeting by nanoparticles: a comprehensive review of literature

PubMed Central

Bazak, Remon; Houri, Mohamad; Achy, Samar El; Kamel, Serag

2016-01-01

Purpose Cancer is one of the leading causes of death, and thus, the scientific community has but great efforts to improve cancer management. Among the major challenges in cancer management is development of agents that can be used for early diagnosis and effective therapy. Conventional cancer management frequently lacks accurate tools for detection of early tumors and has an associated risk of serious side effects of chemotherapeutics. The need to optimize therapeutic ratio as the difference with which a treatment affects cancer cells versus healthy tissues lead to idea that it is needful to have a treatment that could act a the “magic bullet”—recognize cancer cells only. Nanoparticle platforms offer a variety of potentially efficient solutions for development of targeted agents that can be exploited for cancer diagnosis and treatment. There are two ways by which targeting of nanoparticles can be achieved, namely passive and active targeting. Passive targeting allows for the efficient localization of nanoparticles within the tumor microenvironment. Active targeting facilitates the active uptake of nanoparticles by the tumor cells themselves. Methods Relevant English electronic databases and scientifically published original articles and reviews were systematically searched for the purpose of this review. Results In this report, we present a comprehensive review of literatures focusing on the active targeting of nanoparticles to cancer cells, including antibody and antibody fragment-based targeting, antigen-based targeting, aptamer-based targeting, as well as ligand-based targeting. Conclusion To date, the optimum targeting strategy has not yet been announced, each has its own advantages and disadvantages even though a number of them have found their way for clinical application. Perhaps, a combination of strategies can be employed to improve the precision of drug delivery, paving the way for a more effective personalized therapy. PMID:25005786

Inhibitors targeting on cell wall biosynthesis pathway of MRSA.

PubMed

Hao, Haihong; Cheng, Guyue; Dai, Menghong; Wu, Qinghua; Yuan, Zonghui

2012-11-01

Methicillin resistant Staphylococcus aureus (MRSA), widely known as a type of new superbug, has aroused world-wide concern. Cell wall biosynthesis pathway is an old but good target for the development of antibacterial agents. Peptidoglycan and wall teichoic acids (WTAs) biosynthesis are two main processes of the cell wall biosynthesis pathway (CWBP). Other than penicillin-binding proteins (PBPs), some key factors (Mur enzymes, lipid I or II precursor, etc.) in CWBP are becoming attractive molecule targets for the discovery of anti-MRSA compounds. A number of new compounds, with higher affinity for PBPs or with inhibitory activity on such molecule targets in CWBP of MRSA, have been in the pipeline recently. This review concludes recent research achievements and provides a complete picture of CWBP of MRSA, including the peptidoglycan and wall teichoic acids synthesis pathway. The potential inhibitors targeting on CWBP are subsequently presented to improve development of novel therapeutic strategies for MRSA.

Targeting RAS-driven human cancer cells with antibodies to upregulated and essential cell-surface proteins.

PubMed

Martinko, Alexander J; Truillet, Charles; Julien, Olivier; Diaz, Juan E; Horlbeck, Max A; Whiteley, Gordon; Blonder, Josip; Weissman, Jonathan S; Bandyopadhyay, Sourav; Evans, Michael J; Wells, James A

2018-01-23

While there have been tremendous efforts to target oncogenic RAS signaling from inside the cell, little effort has focused on the cell-surface. Here, we used quantitative surface proteomics to reveal a signature of proteins that are upregulated on cells transformed with KRAS G12V , and driven by MAPK pathway signaling. We next generated a toolkit of recombinant antibodies to seven of these RAS-induced proteins. We found that five of these proteins are broadly distributed on cancer cell lines harboring RAS mutations. In parallel, a cell-surface CRISPRi screen identified integrin and Wnt signaling proteins as critical to RAS-transformed cells. We show that antibodies targeting CDCP1, a protein common to our proteomics and CRISPRi datasets, can be leveraged to deliver cytotoxic and immunotherapeutic payloads to RAS-transformed cancer cells and report for RAS signaling status in vivo. Taken together, this work presents a technological platform for attacking RAS from outside the cell. © 2018, Martinko et al.

Bypassing Protein Corona Issue on Active Targeting: Zwitterionic Coatings Dictate Specific Interactions of Targeting Moieties and Cell Receptors.

PubMed

Safavi-Sohi, Reihaneh; Maghari, Shokoofeh; Raoufi, Mohammad; Jalali, Seyed Amir; Hajipour, Mohammad J; Ghassempour, Alireza; Mahmoudi, Morteza

2016-09-07

Surface functionalization strategies for targeting nanoparticles (NP) to specific organs, cells, or organelles, is the foundation for new applications of nanomedicine to drug delivery and biomedical imaging. Interaction of NPs with biological media leads to the formation of a biomolecular layer at the surface of NPs so-called as "protein corona". This corona layer can shield active molecules at the surface of NPs and cause mistargeting or unintended scavenging by the liver, kidney, or spleen. To overcome this corona issue, we have designed biotin-cysteine conjugated silica NPs (biotin was employed as a targeting molecule and cysteine was used as a zwitterionic ligand) to inhibit corona-induced mistargeting and thus significantly enhance the active targeting capability of NPs in complex biological media. To probe the targeting yield of our engineered NPs, we employed both modified silicon wafer substrates with streptavidin (i.e., biotin receptor) to simulate a target and a cell-based model platform using tumor cell lines that overexpress biotin receptors. In both cases, after incubation with human plasma (thus forming a protein corona), cellular uptake/substrate attachment of the targeted NPs with zwitterionic coatings were significantly higher than the same NPs without zwitterionic coating. Our results demonstrated that NPs with a zwitterionic surface can considerably facilitate targeting yield of NPs and provide a promising new type of nanocarriers in biological applications.

Early transduction produces highly functional chimeric antigen receptor-modified virus-specific T-cells with central memory markers: a Production Assistant for Cell Therapy (PACT) translational application.

PubMed

Sun, Jiali; Huye, Leslie E; Lapteva, Natalia; Mamonkin, Maksim; Hiregange, Manasa; Ballard, Brandon; Dakhova, Olga; Raghavan, Darshana; Durett, April G; Perna, Serena K; Omer, Bilal; Rollins, Lisa A; Leen, Ann M; Vera, Juan F; Dotti, Gianpietro; Gee, Adrian P; Brenner, Malcolm K; Myers, Douglas G; Rooney, Cliona M

2015-01-01

multivirus-specific T-cells that allowed stable expression of high levels of a tumor directed CAR. Since a proportion of early-transduced CAR-VSTs had a central memory phenotype, they should expand and persist in vivo, simultaneously protecting against infection and targeting residual malignancy. This manufacturing strategy is currently under clinical investigation in patients receiving allogeneic HSCT for relapsed neuroblastoma and B-cell malignancies (NCT01460901 using a GD2.CAR and NCT00840853 using a CD19.CAR).

Instrumentation of Molecular Imaging on Site-Specific Targeting Fluorescent Peptide for Early Detection of Breast Cancer

NASA Astrophysics Data System (ADS)

Yu, Ping; Ma, Lixin

2012-02-01

In this work we developed two biomedical imaging techniques for early detection of breast cancer. Both image modalities provide molecular imaging capability to probe site-specific targeting dyes. The first technique, heterodyne CCD fluorescence mediated tomography, is a non-invasive biomedical imaging that uses fluorescent photons from the targeted dye on the tumor cells inside human breast tissue. The technique detects a large volume of tissue (20 cm) with a moderate resolution (1 mm) and provides the high sensitivity. The second technique, dual-band spectral-domain optical coherence tomography, is a high-resolution tissue imaging modality. It uses a low coherence interferometer to detect coherent photons hidden in the incoherent background. Due to the coherence detection, a high resolution (20 microns) is possible. We have finished prototype imaging systems for the development of both image modalities and performed imaging experiments on tumor tissues. The spectroscopic/tomographic images show contrasts of dense tumor tissues and tumor necrotic regions. In order to correlate the findings from our results, a diffusion-weighted magnetic resonance imaging (MRI) of the tumors was performed using a small animal 7-Telsa MRI and demonstrated excellent agreement.

A Cell-targeted Photodynamic Nanomedicine Strategy for Head & Neck Cancers

PubMed Central

Master, Alyssa; Malamas, Anthony; Solanki, Rachna; Clausen, Dana M.; Eiseman, Julie L.; Gupta, Anirban Sen

2013-01-01

Photodynamic Therapy (PDT) holds great promise for the treatment of head and neck (H&N) carcinomas where repeated loco-regional therapy often becomes necessary due to the highly aggressive and recurrent nature of the cancers. While interstitial light delivery technologies are being refined for PDT of H&N and other cancers, a parallel clinically relevant research area is the formulation of photosensitizers in nanovehicles that allow systemic administration yet preferential enhanced uptake in the tumor. This approach can render dual-selectivity of PDT, by harnessing both the drug and the light delivery within the tumor. To this end, we report on a cell-targeted nanomedicine approach for the photosensitizer silicon phthalocyanine-4 (Pc 4), by packaging it within polymeric micelles that are surface-decorated with GE11-peptides to promote enhanced cell-selective binding and receptor-mediated internalization in EGFR-overexpressing H&N cancer cells. Using fluorescence spectroscopy and confocal microscopy, we demonstrate in vitro that the EGFR-targeted Pc 4-nanoformulation undergoes faster and higher uptake in EGFR-overexpressing H&N SCC-15 cells. We further demonstrate that this enhanced Pc 4 uptake results in significant cell-killing and drastically reduced post-PDT clonogenicity. Building on this in vitro data, we demonstrate that the EGFR-targeted Pc 4-nanoformulation results in significant intra-tumoral drug uptake and subsequent enhanced PDT response, in vivo, in SCC-15 xenografts in mice. Altogether our results show significant promise towards a cell-targeted photodynamic nanomedicine for effective treatment of H&N carcinomas. PMID:23531079

Innovative T Cell-Targeted Therapy for Ovarian Cancer

DTIC Science & Technology

2012-10-01

from co-culture with EL4 -ROR1neg and EL4 -ROR1+ tumor targets. Ovarian cancer cell lines (A2780, EFO21, EFO27, IGROV1, OC314, and UPN251) were...profiled for ROR1 expression in normoxia (20% O2) and hypoxia (1% O2). Four-hour CRA was used to evaluate cytotoxicity against the OvCa and EL4 tumor...loaded aAPC for negative controls. EL4 is a murine T cell lymphoma cell line used to test specificity of CAR+ T cells with limited allo-reactivity

Antigen sensitivity of CD22-specific chimeric T cell receptors is modulated by target epitope distance from the cell membrane

PubMed Central

James, Scott E.; Greenberg, Philip D.; Jensen, Michael C.; Lin, Yukang; Wang, Jinjuan; Till, Brian G.; Raubitschek, Andrew A.; Forman, Stephen J.; Press, Oliver W.

2008-01-01

We have targeted CD22 as a novel tumor-associated antigen for recognition by human CTL genetically modified to express chimeric T cell receptors (cTCR) recognizing this surface molecule. CD22-specifc cTCR targeting different epitopes of the CD22 molecule promoted efficient lysis of target cells expressing high levels of CD22 with a maximum lytic potential that appeared to decrease as the distance of the target epitope from the target cell membrane increased. Targeting membrane-distal CD22 epitopes with cTCR+ CTL revealed defects in both degranulation and lytic granule targeting. CD22-specific cTCR+ CTL exhibited lower levels of maximum lysis and lower antigen sensitivity than CTL targeting CD20, which has a shorter extracellular domain than CD22. This diminished sensitivity was not a result of reduced avidity of antigen engagement, but instead reflected weaker signaling per triggered cTCR molecule when targeting membrane-distal epitopes of CD22. Both of these parameters were restored by targeting a ligand expressing the same epitope but constructed as a truncated CD22 molecule to approximate the length of a TCR:pMHC complex. The reduced sensitivity of CD22-specific cTCR+ CTL for antigen-induced triggering of effector functions has potential therapeutic applications, as such cells selectively lysed B cell lymphoma lines expressing high levels of CD22 but demonstrated minimal activity against autologous normal B cells, which express lower levels of CD22. Thus, our results demonstrate that cTCR signal strength – and consequently antigen sensitivity – can be modulated by differential choice of target epitopes with respect to distance from the cell membrane, allowing discrimination between targets with disparate antigen density. PMID:18453625

Specific elimination of CD133+ tumor cells with targeted oncolytic measles virus.

PubMed

Bach, Patricia; Abel, Tobias; Hoffmann, Christopher; Gal, Zoltan; Braun, Gundula; Voelker, Iris; Ball, Claudia R; Johnston, Ian C D; Lauer, Ulrich M; Herold-Mende, Christel; Mühlebach, Michael D; Glimm, Hanno; Buchholz, Christian J

2013-01-15

Tumor-initiating cells (TIC) are critical yet evasive targets for the development of more effective antitumoral strategies. The cell surface marker CD133 is frequently used to identify TICs of various tumor entities, including hepatocellular cancer and glioblastoma. Here, we describe oncolytic measles viruses (MV) retargeted to CD133. The viruses, termed MV-141.7 and MV-AC133, infected and selectively lysed CD133(+) tumor cells. Both viruses exerted strong antitumoral effects on human hepatocellular carcinoma growing subcutaneously or multifocally in the peritoneal cavity of nonobese diabetic/severe combined immunodeficient (NOD/SCID) mice. Notably, the CD133-targeted viruses were more effective in prolonging survival than the parental MV-NSe, which is currently assessed as oncolytic agent in clinical trials. Interestingly, target receptor overexpression or increased spreading kinetics through tumor cells were excluded as being causative for the enhanced oncolytic activity of CD133-targeted viruses. MV-141.7 was also effective in mouse models of orthotopic glioma tumor spheres and primary colon cancer. Our results indicate that CD133-targeted measles viruses selectively eliminate CD133(+) cells from tumor tissue, offering a key tool for research in tumor biology and cancer therapy.

Antibody-drug conjugate targeting CD46 eliminates multiple myeloma cells.

PubMed

Sherbenou, Daniel W; Aftab, Blake T; Su, Yang; Behrens, Christopher R; Wiita, Arun; Logan, Aaron C; Acosta-Alvear, Diego; Hann, Byron C; Walter, Peter; Shuman, Marc A; Wu, Xiaobo; Atkinson, John P; Wolf, Jeffrey L; Martin, Thomas G; Liu, Bin

2016-12-01

Multiple myeloma is incurable by standard approaches because of inevitable relapse and development of treatment resistance in all patients. In our prior work, we identified a panel of macropinocytosing human monoclonal antibodies against CD46, a negative regulator of the innate immune system, and constructed antibody-drug conjugates (ADCs). In this report, we show that an anti-CD46 ADC (CD46-ADC) potently inhibited proliferation in myeloma cell lines with little effect on normal cells. CD46-ADC also potently eliminated myeloma growth in orthometastatic xenograft models. In primary myeloma cells derived from bone marrow aspirates, CD46-ADC induced apoptosis and cell death, but did not affect the viability of nontumor mononuclear cells. It is of clinical interest that the CD46 gene resides on chromosome 1q, which undergoes genomic amplification in the majority of relapsed myeloma patients. We found that the cell surface expression level of CD46 was markedly higher in patient myeloma cells with 1q gain than in those with normal 1q copy number. Thus, genomic amplification of CD46 may serve as a surrogate for target amplification that could allow patient stratification for tailored CD46-targeted therapy. Overall, these findings indicate that CD46 is a promising target for antibody-based treatment of multiple myeloma, especially in patients with gain of chromosome 1q.

Inhibition of early T cell cytokine production by arsenic trioxide occurs independently of Nrf2.

PubMed

VanDenBerg, Kelly R; Freeborn, Robert A; Liu, Sheng; Kennedy, Rebekah C; Zagorski, Joseph W; Rockwell, Cheryl E

2017-01-01

Nuclear factor erythroid 2-related factor 2 (Nrf2) is a stress-activated transcription factor that induces a variety of cytoprotective genes. Nrf2 also mediates immunosuppressive effects in multiple inflammatory models. Upon activation, Nrf2 dissociates from its repressor protein, Keap1, and translocates to the nucleus where it induces Nrf2 target genes. The Nrf2-Keap1 interaction is disrupted by the environmental toxicant and chemotherapeutic agent arsenic trioxide (ATO). The purpose of the present study was to determine the effects of ATO on early events of T cell activation and the role of Nrf2 in those effects. The Nrf2 target genes Hmox-1, Nqo-1, and Gclc were all upregulated by ATO (1-2 μM) in splenocytes derived from wild-type, but not Nrf2-null, mice, suggesting that Nrf2 is activated by ATO in splenocytes. ATO also inhibited IFNγ, IL-2, and GM-CSF mRNA and protein production in wild-type splenocytes activated with the T cell activator, anti-CD3/anti-CD28. However, ATO also decreased production of these cytokines in activated splenocytes from Nrf2-null mice, suggesting the inhibition is independent of Nrf2. Interestingly, ATO inhibited TNFα protein secretion, but not mRNA expression, in activated splenocytes suggesting the inhibition is due to post-transcriptional modification. In addition, c-Fos DNA binding was significantly diminished by ATO in wild-type and Nrf2-null splenocytes activated with anti-CD3/anti-CD28, consistent with the observed inhibition of cytokine production by ATO. Collectively, this study suggests that although ATO activates Nrf2 in splenocytes, inhibition of early T cell cytokine production by ATO occurs independently of Nrf2 and may instead be due to impaired AP-1 DNA binding.

Engineering of Systematic Elimination of a Targeted Chromosome in Human Cells.

PubMed

Sato, Hiroshi; Kato, Hiroki; Yamaza, Haruyoshi; Masuda, Keiji; Nguyen, Huong Thi Nguyen; Pham, Thanh Thi Mai; Han, Xu; Hirofuji, Yuta; Nonaka, Kazuaki

2017-01-01

Embryonic trisomy leads to abortion or congenital genetic disorders in humans. The most common autosomal chromosome abnormalities are trisomy of chromosomes 13, 18, and 21. Although alteration of gene dosage is thought to contribute to disorders caused by extra copies of chromosomes, genes associated with specific disease phenotypes remain unclear. To generate a normal cell from a trisomic cell as a means of etiological analysis or candidate therapy for trisomy syndromes, we developed a system to eliminate a targeted chromosome from human cells. Chromosome 21 was targeted by integration of a DNA cassette in HeLa cells that harbored three copies of chromosome 21. The DNA cassette included two inverted loxP sites and a herpes simplex virus thymidine kinase (HSV-tk) gene. This system causes missegregation of chromosome 21 after expression of Cre recombinase and subsequently enables the selection of cells lacking the chromosome by culturing in a medium that includes ganciclovir (GCV). Cells harboring only two copies of chromosome 21 were efficiently induced by transfection of a Cre expression vector, indicating that this approach is useful for eliminating a targeted chromosome.

Therapeutic targeting of the p53 pathway in cancer stem cells

PubMed Central

Prabhu, Varun V.; Allen, Joshua E.; Hong, Bo; Zhang, Shengliang; Cheng, Hairong; El-Deiry, Wafik S.

2013-01-01

Introduction Cancer stem cells are a high profile drug target for cancer therapeutics due to their indispensable role in cancer progression, maintenance, and therapeutic resistance. Restoring wild-type p53 function is an attractive new therapeutic approach for the treatment of cancer due to the well-described powerful tumor suppressor function of p53. As emerging evidence intimately links p53 and stem cell biology, this approach also provides an opportunity to target cancer stem cells. Areas covered Therapeutic approaches to restore the function of wild-type p53, cancer and normal stem cell biology in relation to p53, and the downstream effects of p53 on cancer stem cells. Expert opinion The restoration of wild-type p53 function by targeting p53 directly, its interacting proteins, or its family members holds promise as a new class of cancer therapies. This review examines the impact that such therapies may have on normal and cancer stem cells based on the current evidence linking p53 signaling with these populations. PMID:22998602

Eliminating Cancer Stem Cells by Targeting Embryonic Signaling Pathways.

PubMed

Oren, Ohad; Smith, B Douglas

2017-02-01

Dramatic advances have been made in the understanding of cancer over the past decade. Prime among those are better appreciation of the biology of cancer and the development of targeted therapies. Despite these improvements, however, most tumors remain refractory to anti-cancer medications and frequently recur. Cancer Stem Cells (CSCs), which in some cases express markers of pluripotency (e.g., Oct-4), share many of the molecular features of normal stem cells. These cells have been hypothesised to play a role in tumor resistance and relapse. They exhibit dependence on many primitive regulatory pathways and may be best viewed in the context of embryonic signaling pathways. In this article, we review important embryonic signaling cascades and their differential expression in CSCs. We also discuss these pathways as actionable targets for novel therapies in hopes that eliminating cancer stem cells will lead to an improvement in overall survival for patients.

Folate receptor‐targeted aminoglycoside‐derived polymers for transgene expression in cancer cells

PubMed Central

Godeshala, Sudhakar; Nitiyanandan, Rajeshwar; Thompson, Brian; Goklany, Sheba; Nielsen, David R.

2016-01-01

Abstract Targeted delivery of anticancer therapeutics can potentially overcome the limitations associated with current chemotherapeutic regimens. Folate receptors are overexpressed in several cancers, including ovarian, triple‐negative breast and bladder cancers, making them attractive for targeted delivery of nucleic acid therapeutics to these tumors. This work describes the synthesis, characterization and evaluation of folic acid‐conjugated, aminoglycoside‐derived polymers for targeted delivery of transgenes to breast and bladder cancer cell lines. Transgene expression was significantly higher with FA‐conjugated aminoglycoside‐derived polymers than with Lipofectamine, and these polymers demonstrated minimal cytotoxicty. Competitive inhibition using free folic acid significantly reduced transgene expression efficacy of folate‐targeted polymers, suggesting a role for folate receptor‐mediated uptake. High efficacy FA‐targeted polymers were employed to deliver a plasmid expressing the TRAIL protein, which induced death in cancer cells. These results indicate that FA‐conjugated aminoglycoside‐derived polymers are promising for targeted delivery of nucleic acids to cancer cells that overexpress folate receptors. PMID:29313013

Update on B-cell targeted therapies for systemic lupus erythematosus.

PubMed

Mok, Chi Chiu

2014-06-01

Systemic lupus erythematosus (SLE) is a systemic autoimmune disease characterized by flares and remission, leading to accrual of organ damage over time as a result of persistent tissue inflammation and treatment-related complications. Novel therapies aiming at better treatment response and fewer adverse effects are being tested in the pipeline. This review summarizes the B-cell abnormalities observed in patients with SLE, and updates recent data on the efficacy and safety of B-cell targeted therapies in the treatment of SLE. The pitfalls of clinical trial design and future directions of the development of SLE therapeutics are discussed. The variability of clinical response to treatment in SLE reflects the clinical and immunological heterogeneity of the disease. The treatment plan for patients with SLE should be individualized with the aim of eradicating disease activity, preventing flares and minimizing treatment-related complications. Despite the disappointment of recent clinical trials, B-cell remains the promising target of future SLE therapies. Results from ongoing clinical trials on B-cell targeted biological agents are eagerly awaited.

Role of alveolar epithelial early growth response-1 (Egr-1) in CD8+ T cell-mediated lung injury.

PubMed

Ramana, Chilakamarti V; Cheng, Guang-Shing; Kumar, Aseem; Kwon, Hyung-Joo; Enelow, Richard I

2009-12-01

Influenza infection of the distal airways results in severe lung injury, a considerable portion of which is immunopathologic and attributable to the host responses. We have used a mouse model to specifically investigate the role of antiviral CD8(+) T cells in this injury, and have found that the critical effector molecule is TNF-alpha expressed by the T cells upon antigen recognition. Interestingly, the immunopathology which ensues is characterized by significant accumulation of host inflammatory cells, recruited by chemokines expressed by the target alveolar epithelial cells. In this study we analyzed the mechanisms involved in the induction of epithelial chemokine expression triggered by antigen-specific CD8(+) T cell recognition, and demonstrate that the early growth response-1 (Egr-1) transcription factor is rapidly induced in epithelial cells, both in vitro and ex vivo, and that this is a critical regulator of a host of inflammatory chemokines. Genetic deficiency of Egr-1 significantly abrogates both the chemokine expression and the immunopathologic injury associated with T cell recognition, and it directly regulates transcriptional activity of a model CXC chemokine, MIP-2. We further demonstrate that Egr-1 induction is triggered by TNF-alpha-dependent ERK activation, and inhibition of this pathway ablates Egr-1 expression. These findings suggest that Egr-1 may represent an important target in mitigating the immunopathology of severe influenza infection.

Role of alveolar epithelial Early growth response-1 (Egr-1) in CD8+ T Cell mediated Lung Injury

PubMed Central

Ramana, Chilakamarti V.; Cheng, Guang-Shing; Kumar, Aseem; Kwon, Hyung- Joo; Enelow, Richard I.

2009-01-01

Influenza infection of the distal airways results in severe lung injury, a considerable portion of which is immunopathologic and attributable to the host responses. We have used a mouse model to specifically investigate the role of antiviral CD8+ T cells in this injury, and have found that the critical effector molecule is TNF-α expressed by the T cells upon antigen recognition. Interestingly, the immunopathology which ensues is characterized by significant accumulation of host inflammatory cells, recruited by chemokines expressed by the target alveolar epithelial cells. In this study we analyzed the mechanisms involved in the induction of epithelial chemokine expression triggered by antigen-specific CD8+ T cell recognition, and demonstrate that the Early growth response-1 (Egr-1) transcription factor is rapidly induced in epithelial cells, both in vitro and ex vivo, and that this is a critical regulator of a host of inflammatory chemokines. Genetic deficiency of Egr-1 significantly abrogates both the chemokine expression and the immunopathologic injury associated with T cell recognition, and it directly regulates transcriptional activity of a model CXC chemokine, MIP-2. We further demonstrate that Egr-1 induction is triggered by TNF-α– dependent ERK activation, and inhibition of this pathway ablates Egr-1 expression. These findings suggest that Egr-1 may represent an important target in mitigating the immunopathology of severe influenza infection. PMID:19786304

Proposal for National Targets in the Framework of the European Reduction Goal for Early School Leaving

ERIC Educational Resources Information Center

Lastra-Bravo, Xavier B.; Tolón-Becerra, Alfredo; Salinas-Andújar, José A.

2013-01-01

According to the European Commission's "Europe 2020" strategy, the early school leaving (ESL) rate in European Union (EU) Member States must be reduced to a maximum of 10 per cent by 2020. This paper proposes a nonlinear distribution method based on dynamic targets for reducing the percentage of early school leavers. The aim of this…

Analysis of early mesothelial cell responses to Staphylococcus epidermidis isolated from patients with peritoneal dialysis-associated peritonitis.

PubMed

McGuire, Amanda L; Mulroney, Kieran T; Carson, Christine F; Ram, Ramesh; Morahan, Grant; Chakera, Aron

2017-01-01

The major complication of peritoneal dialysis (PD) is the development of peritonitis, an infection within the abdominal cavity, primarily caused by bacteria. PD peritonitis is associated with significant morbidity, mortality and health care costs. Staphylococcus epidermidis is the most frequently isolated cause of PD-associated peritonitis. Mesothelial cells are integral to the host response to peritonitis, and subsequent clinical outcomes, yet the effects of infection on mesothelial cells are not well characterised. We systematically investigated the early mesothelial cell response to clinical and reference isolates of S. epidermidis using primary mesothelial cells and the mesothelial cell line Met-5A. Using an unbiased whole genome microarray, followed by a targeted panel of genes known to be involved in the human antibacterial response, we identified 38 differentially regulated genes (adj. p-value < 0.05) representing 35 canonical pathways after 1 hour exposure to S. epidermidis. The top 3 canonical pathways were TNFR2 signaling, IL-17A signaling, and TNFR1 signaling (adj. p-values of 0.0012, 0.0012 and 0.0019, respectively). Subsequent qPCR validation confirmed significant differences in gene expression in a number of genes not previously described in mesothelial cell responses to infection, with heterogeneity observed between clinical isolates of S. epidermidis, and between Met-5A and primary mesothelial cells. Heterogeneity between different S. epidermidis isolates suggests that specific virulence factors may play critical roles in influencing outcomes from peritonitis. This study provides new insights into early mesothelial cell responses to infection with S. epidermidis, and confirms the importance of validating findings in primary mesothelial cells.

Regional delivery of mesothelin-targeted CAR T cell therapy generates potent and long-lasting CD4-dependent tumor immunity

PubMed Central

Adusumilli, Prasad S.; Cherkassky, Leonid; Villena-Vargas, Jonathan; Colovos, Christos; Servais, Elliot; Plotkin, Jason; Jones, David R.; Sadelain, Michel

2015-01-01

Translating the recent success of chimeric antigen receptor (CAR) T cell therapy for hematological malignancies to solid tumors will necessitate overcoming several obstacles, including inefficient T cell tumor infiltration and insufficient functional persistence. Taking advantage of an orthotopic model that faithfully mimics human pleural malignancy, we evaluated two routes of administration of mesothelin-targeted T cells using the M28z CAR. We found that intra-pleurally administered CAR T cells vastly out-performed systemically infused T cells, requiring 30-fold fewer M28z T cells to induce long-term complete remissions. Following intrapleural T cell administration, prompt in vivo antigen-induced T cell activation allowed robust CAR T cell expansion and effector differentiation, resulting in enhanced anti-tumor efficacy and functional T cell persistence for 200 days. Regional T cell administration also promoted efficient elimination of extrathoracic tumor sites. This therapeutic efficacy was dependent on early CD4+ T cell activation associated with a higher intra-tumoral CD4/CD8 cell ratios and CD28-dependent CD4+ T cell-mediated cytotoxicity. In contrast, intravenously delivered CAR T cells, even when accumulated at equivalent numbers in the pleural tumor, did not achieve comparable activation, tumor eradication or persistence. The remarkable ability of intrapleurally administered T cells to circulate and persist supports the concept of delivering optimal CAR T cell therapy through “regional distribution centers.” Based on these results, we are opening a phase I clinical trial to evaluate the safety of intrapleural administration of mesothelin-targeted CAR T cells in patients with primary or secondary pleural malignancies. PMID:25378643