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1

Idiotypic Study of a Bispecific Thyroglobulin and Thyroperoxidase Monoclonal Antibody  

Microsoft Academic Search

We have previously established that thyroperoxidase (TPO), a major thyroid antigen involved in autoimmune thyroid diseases, interacts with an idiotype present on human and mouse antibodies directed to thryoglobulin (TG), another thyroid autoantigen. In order to characterize the TPO-reactive idio-type, we selected a TG monoclonal antibody (mAb J7 B49.15) which bound to TPO and cross-reacted with human bispecific TG and

Maggy Villa; Josée-Martine Durand-Gorde; Pierre Carayon; Jean Ruf

1996-01-01

2

Production of a bi-specific monoclonal antibody recognizing mouse kappa light chains and horseradish peroxidase  

Microsoft Academic Search

The production of a bi-specific monoclonal antibody that simultaneously recognizes mouse kappa light chains and horseradish peroxidase (HRP) for use as a general developing reagent in a wide variety of immunobased techniques is described. This antibody, named McC10, was produced by the fusion of an aminopterin-sensitive interspecies hybridoma which secretes rat monoclonal antibodies against HRP (RAP2·Ag) and splenocytes from a

R. L. Kenigsberg; A. C. Cuello

1990-01-01

3

Gene Transfer by Retrovirus-Derived Shuttle Vectors in the Generation of Murine Bispecific Monoclonal Antibodies  

Microsoft Academic Search

The present study reports on the use of gene transfer by retrovirus-derived shuttle vectors in the generation of hybrid hybridomas secreting bispecific monoclonal antibodies. neo- and dhfr^*- genes were infected into distinct murine hybridomas, thus conferring a dominant resistance trait to geneticin (G418) and to methotrexate. The vectors employed were replication-deficient and dependent on complementation by a helper virus provided

L. B. Demonte; P. Nistico; R. Tecce; P. Dellabona; M. Momo; A. Anichini; M. Mariani; P. G. Natali; F. Malavasi

1990-01-01

4

Bispecific Single Domain Antibodies  

Microsoft Academic Search

\\u000a Monoclonal antibodies are now widely recognized as therapeutic molecules and more than 25 molecules have been approved in\\u000a the United States and other countries. Despite these successes, the clinical activity of these molecules is still far from\\u000a optimal and new solutions have to be found, especially in the field of cancer therapy. The potential of bispecific antibodies,\\u000a capable of simultaneously

Patrick Chames; Daniel Baty

5

Antitumor immunity: easy as 1, 2, 3 with monoclonal bispecific trifunctional antibodies?  

PubMed

Monoclonal antibodies occupy an increasing niche in the arsenal available to treat cancer. Several developments have rendered this the fastest growing sector in the pharmaceutical industry. Traditionally, antibodies were developed to block key signaling molecules implicated in tumor progression. However, antibodies also recruit additional immune effector mechanisms against tumors, a property that may be exploited for clinical benefit. Bispecific antibodies represent one such strategy in which elements derived from two monoclonal antibodies are incorporated into a single molecular species. Commonly, the bispecific approach is used to achieve simultaneous cross-linking of CD3 and a tumor antigen such as epithelial cell adhesion molecule (EpCAM), thereby recruiting T-cell activation to the tumor cell surface. A further sophistication involves the engineering of trifunctional derivatives such as the clinically approved agent, catumaxomab. Catumaxomab has antigen-binding arms that engage CD3 and EpCAM and a constant domain that recruits Fc receptor-bearing cells, notably monocytes, dendritic cells, and natural killer cells. Owing to this triangular binding capability, catumaxomab can activate both innate and adaptive immune effector mechanisms in addition to promoting immunologic memory. Recent data indicate that this agent can also promote immunogenic cell death, particularly when used in combination with selected chemotherapeutic agents such as oxaliplatin. PMID:24014596

Maher, John; Adami, Antonella A

2013-09-15

6

Human neutrophil interactions of a bispecific monoclonal antibody targeting tumor and human Fc??RIII  

Microsoft Academic Search

2B1 is a bispecific murine monoclonal antibody (bsmAb) targeting the c-erbB-2 and CD16 (Fc?RIII) antigens. c-erbB-2 is over-expressed\\u000a by a variety of adenocarcinomas, and CD16, the low-affinity Fc? receptor for aggregated immunoglobulins, is expressed by polymorphonuclear\\u000a leukocytes (PMN), natural killer (NK) cells and differentiated mononuclear phagocytes. 2B1 potentiates the in vitro lysis\\u000a of c-erbB-2 over-expressing tumors by NK cells and

L. M. Weiner; R. Katherine Alpaugh; Anne R. Amoroso; Gregory P. Adams; David B. Ring; Malcolm W. Barth

1996-01-01

7

Alternative Scaffolds as Bispecific Antibody Mimetics  

Microsoft Academic Search

\\u000a The use of non-immunoglobulin-based protein scaffolds for engineering of specific recognition was first described some 15\\u000a years ago and has matured as a discipline in parallel with the rapidly expanding monoclonal antibody field. As bispecific\\u000a antibodies and antibody fragments have come into focus lately, the corresponding development of bispecific alternative scaffolds\\u000a is also emerging. Here, the concept of alternative scaffold

John Löfblom; Fredrik Y. Frejd

8

Bispecific Antibodies: Molecules That Enable Novel Therapeutic Strategies  

Microsoft Academic Search

Bispecific antibodies are unique in the sense that they can bind simultaneously two different antigens. This property enables the development of therapeutic strategies that are not possible with conventional monoclonal antibodies. The large panel of imaginative bispecific antibody formats that has been developed reflects the strong interest for these molecules. Although in many cases the manufacturing of clinical grade material

Nicolas Fischer; Olivier Léger

2007-01-01

9

Bispecific antibodies for cancer therapy: the light at the end of the Patrick Chames1  

E-print Network

1 Bispecific antibodies for cancer therapy: the light at the end of the tunnel? Patrick Chames1, bispecific, cancer, therapy, clinical trials Abbreviation mAb: monoclonal antibodies, bsAbs: bispecificTE: bispecific T cell engager, MHC: major histocompatibility, DC: dendritic cells, NK: natural killer cells, TEM

Paris-Sud XI, Université de

10

A revival of bispecific antibodies  

Microsoft Academic Search

Bispecific antibodies usually do not occur in nature but are constructed by recombinant DNA or cell-fusion technologies. Most are designed to recruit cytotoxic effector cells of the immune system effectively against pathogenic target cells. This complex task explains why, after more than 15 years of extensive research, many different formats of bispecific antibodies have been developed but only a few

Peter Kufer; Ralf Lutterbüse; Patrick A. Baeuerle

2004-01-01

11

Chemical generation of bispecific antibodies  

PubMed Central

Bispecific antibodies (BsAbs) are regarded as promising therapeutic agents due to their ability to simultaneously bind two different antigens. Several bispecific modalities have been developed, but their utility is limited due to problems with stability and manufacturing complexity. Here we report a versatile technology, based on a scaffold antibody and pharmacophore peptide heterodimers, that enables rapid generation and chemical optimization of bispecific antibodies, which are termed bispecific CovX-Bodies. Two different peptides are joined together using a branched azetidinone linker and fused to the scaffold antibody under mild conditions in a site-specific manner. Whereas the pharmacophores are responsible for functional activities, the antibody scaffold imparts long half-life and Ig-like distribution. The pharmacophores can be chemically optimized or replaced with other pharmacophores to generate optimized or unique bispecific antibodies. As a prototype, we developed a bispecific antibody that binds both vascular endothelial growth factor (VEGF) and angiopoietin-2 (Ang2) simultaneously, inhibits their function, shows efficacy in tumor xenograft studies, and greatly augments the antitumor effects of standard chemotherapy. This unique antiangiogenic bispecific antibody is in phase-1 clinical trials. PMID:21149738

Doppalapudi, Venkata R.; Huang, Jie; Liu, Dingguo; Jin, Ping; Liu, Bin; Li, Lingna; Desharnais, Joel; Hagen, Crystal; Levin, Nancy J.; Shields, Michael J.; Parish, Michelle; Murphy, Robert E.; Del Rosario, Joselyn; Oates, Bryan D.; Lai, Jing-Yu; Matin, Marla J.; Ainekulu, Zemeda; Bhat, Abhijit; Bradshaw, Curt W.; Woodnutt, Gary; Lerner, Richard A.; Lappe, Rodney W.

2010-01-01

12

Bispecific Antibodies and Gene Therapy  

Microsoft Academic Search

\\u000a Gene therapy is the transfer of therapeutic genes, via gene transfer vectors, into patients for therapeutic purposes. Different\\u000a gene therapy strategies are being pursued, including long-term gene correction of monogenetic diseases, eradication of tumor\\u000a cells in cancer patients, or genetic vaccination for infectious diseases. Bispecific antibodies and gene therapy are connected\\u000a in two ways. First, bispecific antibodies are tools of

Dirk M. Nettelbeck

13

Treatment of Hodgkin's disease with bispecific antibodies  

Microsoft Academic Search

Summary Bispecific monoclonal antibodies (Bi-MAbs) with dual spe- cificity for tumor-associated antigens (TAA) and a triggering molecule of an immunologic effector cell, respectively, open the possibility to specifically target to and activate cytotoxic effector cells (macrophages, T-cells, NK cells) at the rumor site. Using appropriately designed Bi-MAbs and unstimulat- ed human NK cells and T-cells, respectively, we were able to

F. Hartmann; C. Renner; W. Jung; U. Sahin; M. Pfreundschuh

1996-01-01

14

Induction of multiple anti-c-erbB-2 specificities accompanies a classical idiotypic cascade following 2B1 bispecific monoclonal antibody treatment  

Microsoft Academic Search

The bispecific monoclonal antibody (bsmAb) 2B1, targeting the extracellular domain of c-erbB-2, the protein product of the\\u000a HER-2\\/neu proto-oncogene, and Fc?RIII (CD16), expressed by human natural killer cells, neutrophils and differentiated monocytes, mediates\\u000a the specific cytotoxic activity of these effector cells to tumor cells. A group of 24 patients with c-erbB-2-overexpressing\\u000a tumors were treated with intravenously administered 2B1 in a

J. I. Clark; R. Katherine Alpaugh; Margaret von Mehren; Josephine Schultz; Julie R. Gralow; Martin A. Cheever; David B. Ring; L. M. Weiner

1997-01-01

15

Catumaxomab: a bispecific trifunctional antibody.  

PubMed

The trifunctional bispecific monoclonal antibody catumaxomab has two binding specificities directed at epithelial cell adhesion molecule (EpCAM) and the T-cell antigen CD3. With its Fc-fragment, catumaxomab additionally binds accessory cells such as dendritic cells, macrophages and natural killer cells. The trifunctional approach thus leads to unrestricted but specific killing of epithelial tumor cells by major histocompatibility complex without the need for preactivation or external costimulation. The tumor-associated antigen EpCAM is strongly expressed in carcinomas of various origins including colon, rectum, ovarian, gastric, esophagus, lung, pancreas, breast and head and neck. Expression of EpCAM is often associated with an unfavorable prognosis in patients with breast cancer. Catumaxomab has been approved in Europe for the intraperitoneal treatment of malignant ascites in patients with EpCAM-positive epithelial tumors when standard therapy is not available or is no longer feasible. Basic preclinical and clinical findings with different routes of catumaxomab administration in various indications are summarized and discussed in this review. PMID:19927225

Sebastian, M; Kuemmel, A; Schmidt, M; Schmittel, A

2009-08-01

16

Bispecific antibodies for cancer therapy  

PubMed Central

With 23 approvals in the US and other countries and four approvals outside US, antibodies are now widely recognized as therapeutic molecules. The therapeutic and commercial successes met by rituximab, trastuzumab, cetuximab and other mAbs have inspired antibody engineers to improve the efficacy of these molecules. Consequently, a new wave of antibodies with engineered Fc leading to much higher effector functions such as antibody-dependent cell-mediated cytotoxicity or complement-dependent cytotoxicity is being evaluated in the clinic, and several approvals are expected soon. In addition, research on a different class of antibody therapeutics, bispecific antibodies, has recently led to outstanding clinical results, and the first approval of the bispecific antibody catumaxomab, a T cell retargeting agent that was approved in the European Union in April 2009. This review describes the most recent advances and clinical study results in the field of bispecific antibodies, a new class of molecules that might outshine conventional mAbs as cancer immunotherapeutics in a near future. PMID:20073127

Baty, Daniel

2009-01-01

17

Bispecific Antibodies from Hybrid Hybridoma  

Microsoft Academic Search

\\u000a Hybrid hybridomas (also termed quadromas or tetradomas) are man-made cell lines that secrete bispecific antibodies (bsAb)\\u000a with two different specificities being able to crosslink two distinct molecules. Such antibodies do not occur in nature and\\u000a have been originally developed to improve immunohistochemical staining procedures and immunoassays (Milstein and Cuello 1983;\\u000a Suresh et al. 1986). Interestingly, the fusion of two immunoglobulin-producing

Gerhard Moldenhauer

18

Recombinant Bispecific Antibodies for Cancer Therapy  

Microsoft Academic Search

\\u000a Bispecific antibodies are molecules capable of simultaneously binding to two different antigens. While initially bispecific\\u000a antibodies have been developed mainly for cellular cancer immunotherapy through retargeting of effector cells to tumor cells,\\u000a recent developments include also dual targeting strategies and the retargeting of effector molecules, e.g. in radioimmunotherapy.\\u000a In addition to various applications, a plethora of bispecific antibody formats have

Dafne Müller; Roland E. Kontermann

19

Time resolved native ion-mobility mass spectrometry to monitor dynamics of IgG4 Fab arm exchange and "bispecific" monoclonal antibody formation.  

PubMed

Monoclonal antibodies (mAbs) and derivatives such as antibody-drug conjugates (ADC) and bispecific antibodies (bsAb), are the fastest growing class of human therapeutics. Most of the therapeutic antibodies currently on the market and in clinical trials are chimeric, humanized, and human immunoglobulin G1 (IgG1). An increasing number of IgG2s and IgG4s that have distinct structural and functional properties are also investigated to develop products that lack or have diminished antibody effector functions compared to IgG1. Importantly, wild type IgG4 has been shown to form half molecules (one heavy chain and one light chain) that lack interheavy chain disulfide bonds and form intrachain disulfide bonds. Moreover, IgG4 undergoes a process of Fab-arm exchange (FAE) in which the heavy chains of antibodies of different specificities can dissociate and recombine in bispecific antibodies both in vitro and in vivo. Here, native mass spectrometry (MS) and time-resolved traveling wave ion mobility MS (TWIM-MS) were used for the first time for online monitoring of FAE and bsAb formation using Hz6F4-2v3 and natalizumab, two humanized IgG4s which bind to human Junctional Adhesion Molecule-A (JAM-A) and alpha4 integrin, respectively. In addition, native MS analysis of bsAb/JAM-A immune complexes revealed that bsAb can bind up to two antigen molecules, confirming that the Hz6F4 family preferentially binds dimeric JAM-A. Our results illustrate how IM-MS can rapidly assess bsAb structural heterogeneity and be easily implemented into MS workflows for bsAb production follow up and bsAb/antigen complex characterization. Altogether, these results provide new MS-based methodologies for in-depth FAE and bsAb formation monitoring. Native MS and IM-MS will play an increasing role in next generation biopharmaceutical product characterization like bsAbs, antibody mixtures, and antibody-drug conjugates (ADC) as well as for biosimilar and biobetter antibodies. PMID:24007193

Debaene, François; Wagner-Rousset, Elsa; Colas, Olivier; Ayoub, Daniel; Corvaďa, Nathalie; Van Dorsselaer, Alain; Beck, Alain; Cianférani, Sarah

2013-10-15

20

Bispecific antibody and its clinical applications in cancer  

Microsoft Academic Search

Bispecific antibody (BsAb) usually consists of two different antigen-binding arms, by which it is capable of simultaneously\\u000a binding to target cells and effector cells, and can directly mediate the killing of target cells by retargeting and activating\\u000a effector cells. The development of BsAb research goes through three main stages: chemical crosslinking of murine-derived monoclonal\\u000a antibody, hybrid hybridomas and engineered BsAb.

Yuanfu Xu; Chunzheng Yang; Zhenping Zhu

2001-01-01

21

Production of Native Bispecific Antibodies in Rabbits  

Microsoft Academic Search

BackgroundA natural bispecific antibody, which can be produced by exchanging Fab arms of two IgG4 molecules, was first described in allergic patients receiving therapeutic injections with two distinct allergens. However, no information has been published on the production of natural bispecific antibody in animals. Even more important, establishment of an animal model is a useful approach to investigate and characterize

Wei Wang; Ruihuan Xu; Jinming Li; Vladimir Brusic

2010-01-01

22

A Novel Redox Method for Rapid Production of Functional Bi-Specific Antibodies For Use in Early Pilot Studies  

PubMed Central

We demonstrate here a rapid alternative method for the production of functional bi-specific antibodies using the mild reducing agent 2-mercaptoethanesulfonic acid sodium salt (MESNA). Following reduction of a mixture of two monoclonal antibodies with MESNA to break inter heavy chain bonds, this solution is dialysed under oxidising conditions and antibodies are allowed to reform. During this reaction a mixture of antibodies is formed, including parental antibodies and bi-specific antibody. Bi-specific antibodies are purified over two sequential affinity columns. Following purification, bi-specificity of antibodies is determined in enzyme-linked immunosorbent assays and by flow cytometry. Using this redox method we have been successful in producing hybrid and same-species bi-specific antibodies in a time frame of 6–10 working days, making this production method a time saving alternative to the time-consuming traditional heterohybridoma technology for the production of bi-specific antibodies for use in early pilot studies. The use of both rat and mouse IgG antibodies forming a rat/mouse bi-specific antibody as well as producing a pure mouse bi-specific antibody and a pure rat bi-specific antibody demonstrates the flexibility of this production method. PMID:21811628

Carlring, Jennifer; De Leenheer, Evy; Heath, Andrew William

2011-01-01

23

Generation of Bispecific Antibodies by Chemical Conjugation  

Microsoft Academic Search

\\u000a Bispecifc antibodies (bsAbs) are emerging as a promising new class of biotherapeutics. Although Ig domain fusion by DNA engineering\\u000a is the prevalent methodology for producing bsAbs, a significant number of studies are being performed with chemically crosslinked\\u000a Abs. By using different starting material and various conjugation strategies, bispecific fragments, full-length Abs, or combinations\\u000a thereof can be generated. Two types or

Diego Ellerman; Justin M. Scheer

24

Induction of adaptive Anti-HER2/neu immune responses in a Phase 1B/2 trial of 2B1 bispecific murine monoclonal antibody in metastatic breast cancer (E3194): a trial coordinated by the Eastern Cooperative Oncology Group.  

PubMed

2B1 is a bispecific murine monoclonal antibody that binds to the extracellular domains of HER2/neu and FcgammaRIII. 2B1 efficiently promotes the lysis of tumor cells overexpressing HER2/neu by natural killer cells and mononuclear phagocytes that express the FcgammaRIII A isoform. Here, we report the results of E3194, a phase 1B/2 trial conducted by the Eastern Cooperative Oncology Group that employed 2B1 therapy in 20 women with metastatic breast cancer. The median age was 51 years. All but 1 patient had received prior chemotherapy. After the first dose, 3 of the initial 8 patients experienced dose-limiting toxicities that required dose-reduction. The nature of these dose-limiting toxicities resulted in a reduced dose from 2.5 mg/m/d to 1 mg/m/d in the remaining 12 patients. Objective antitumor responses were not seen. However, 2B1 therapy induced adaptive immune responses to both intracellular and extracellular domains of HER2/neu. Even though 2B1 antibody therapy did not show activity in metastatic breast cancer at the current administered doses, the ability of this antibody to induce detectable immune responses against an important tumor antigen has implications for understanding the mechanisms by which antibodies that mediate antibody-directed cellular cytotoxicity may exert their clinical antitumor effects. PMID:17457220

Borghaei, Hossein; Alpaugh, R Katherine; Bernardo, Patricia; Palazzo, Irma E; Dutcher, Janice P; Venkatraj, Usha; Wood, William C; Goldstein, Lori; Weiner, Louis M

2007-01-01

25

FcalphaRI (CD89) as a novel trigger molecule for bispecific antibody therapy.  

PubMed

Promising results from clinical trials with unconjugated antibodies stimulated renewed interest in immune effector mechanisms of monoclonal antibodies (MoAbs). We investigated the potential of IgA as antibody isotype for cell- or complement-mediated tumor cell lysis and assessed the potential of its myeloid Fc receptor, FcalphaRI (CD89), as trigger molecule for bispecific antibody (BsAb)-mediated immunotherapy. Comparing hapten-directed antibodies of human IgA2 with IgG1 or IgG3 isotypes, we found all three to mediate effective killing of sensitized tumor target cells in whole blood assays. Analysis of effector mechanisms showed IgG-mediated lysis to be predominantly complement-dependent, whereas IgA-dependent killing was primarily effector cell-mediated. A comparison of effector cell populations in antibody-dependent cell-mediated cytotoxicity (ADCC) showed neutrophils to be most important for IgA-dependent tumor cell killing, involving FcalphaRI as shown with Fc receptor blocking antibodies. Reverse ADCC experiments against target cells sensitized with Fc receptor antibodies, or assays with FcalphaRI-directed bispecific antibodies confirmed FcalphaRI as effective trigger molecule in polymorphonuclear neutrophil (PMN)-mediated lysis. During granulocyte colony-stimulating factor (G-CSF ) therapy, (FcalphaRI x HER-2/neu) bispecific antibodies induced enhanced killing of HER-2/neu positive SK-BR-3 breast cancer cells in whole blood assays. This enhanced cytotoxicity was paralleled by increased PMN counts, which lead to higher effector to target cell ratios in G-CSF-primed blood. Furthermore, bispecific antibodies, directed to FcalphaRI and Candida albicans, enhanced neutrophils' phagocytosis of fungi. In summary, these results identify IgA as an effective antibody isotype for immunotherapy, working primarily via FcalphaRI on neutrophils. They suggest FcalphaRI-directed bispecific antibodies and G-CSF to be an attractive combination for malignant or infectious diseases. PMID:9373259

Valerius, T; Stockmeyer, B; van Spriel, A B; Graziano, R F; van den Herik-Oudijk, I E; Repp, R; Deo, Y M; Lund, J; Kalden, J R; Gramatzki, M; van de Winkel, J G

1997-12-01

26

Immune recruitment by bispecific antibodies for the treatment of Hodgkin disease  

Microsoft Academic Search

For the treatment of Hodgkin lymphoma, bispecific monoclonal antibodies (bi-mAbs) were established which recognize the Hodgkin-associated CD30 antigen with one arm and the CD3 or CD28 antigen on T lymphocytes or the CD16 antigen on natural killer (NK) cells with the second arm. The NK cell-activating !-CD16\\/CD30 antibody was able to retarget human NK cells toward CD30+ target cells and

Ligia da Costa; Christoph Renner; Frank Hartmann; Michael Pfreundschuh

2000-01-01

27

Synthesis of Bispecific Antibodies with Genetically Encoded Unnatural Amino Acids  

PubMed Central

Bispecific antibodies were constructed using genetically encoded unnatural amino acids with orthogonal chemical reactivity. A two-step process afforded homogeneous products in excellent yield. Using this approach, we synthesized an anti-HER2/anti-CD3 bispecific antibody, which efficiently crosslinked HER2+ cells and CD3+ cells. In vitro effector-cell mediated cytotoxicity was observed at picomolar concentrations. PMID:22642368

Kim, Chan Hyuk; Axup, Jun Y.; Dubrovska, Anna; Kazane, Stephanie A.; Hutchins, Benjamin A.; Wold, Erik D.; Smider, Vaughn V.; Schultz, Peter G.

2013-01-01

28

Therapeutic Recombinant Monoclonal Antibodies  

ERIC Educational Resources Information Center

During the last two decades, the rapid growth of biotechnology-derived techniques has led to a myriad of therapeutic recombinant monoclonal antibodies with significant clinical benefits. Recombinant monoclonal antibodies can be obtained from a number of natural sources such as animal cell cultures using recombinant DNA engineering. In contrast to…

Bakhtiar, Ray

2012-01-01

29

Bispecific antibodies with natural architecture produced by co-culture of bacteria expressing two distinct half-antibodies.  

PubMed

By enabling the simultaneous engagement of two distinct targets, bispecific antibodies broaden the potential utility of antibody-based therapies. However, bispecific-antibody design and production remain challenging, owing to the need to incorporate two distinct heavy and light chain pairs while maintaining natural nonimmunogenic antibody architecture. Here we present a bispecific-antibody production strategy that relies on co-culture of two bacterial strains, each expressing a half-antibody. Using this approach, we produce 28 unique bispecific antibodies. A bispecific antibody against the receptor tyrosine kinases MET and EGFR binds both targets monovalently, inhibits their signaling, and suppresses MET and EGFR-driven cell and tumor growth. Our strategy allows rapid generation of bispecific antibodies from any two existing antibodies and yields milligram to gram quantities of bispecific antibodies sufficient for a wide range of discovery and preclinical applications. PMID:23831709

Spiess, Christoph; Merchant, Mark; Huang, Arthur; Zheng, Zhong; Yang, Nai-Ying; Peng, Jing; Ellerman, Diego; Shatz, Whitney; Reilly, Dorothea; Yansura, Daniel G; Scheer, Justin M

2013-08-01

30

Identification of Natural Bispecific Antibodies against Cyclic Citrullinated Peptide and Immunoglobulin G in Rheumatoid Arthritis  

Microsoft Academic Search

BackgroundPrevious studies indicate that natural bispecific antibodies can be readily produced in vivo when the body is simultaneously stimulated with 2 distinct antigens. Patients with rheumatoid arthritis (RA) usually exhibit persistent immune responses to various autoantigens, raising the possibility that natural bispecific antibodies against 2 distinct autoantigens might exist.Methodology\\/Principal FindingsWe identified the presence of natural bispecific antibodies against cyclic citrullinated

Wei Wang; Jinming Li; Niels Câmara

2011-01-01

31

Interleukin12 increases bispecific-antibody-mediated natural killer cell cytotoxicity against human tumors  

Microsoft Academic Search

The combination of CD16\\/CD30 bispecific monoclonal antibodies (bi-mAb) and unstimulated human resting natural killer (NK)\\u000a cells can cure about 50% of mice with severe combined immunodeficiency (SCID) bearing subcutaneously growing established Hodgkin’s\\u000a lymphoma. As interleukin-2 (IL-2) and IL-12 have been shown to increase NK cell activity, we tested the capacity of these\\u000a cytokines to increase bi-mAb-mediated NK cell cytotoxicity against

Ugur Sahin; Sylvia Kraft-Bauer; Sascha Ohnesorge; M. Pfreundschuh; Christoph Renner

1996-01-01

32

Quantification of cell surface proteins with bispecific antibodies  

PubMed Central

Flow cytometry is an established method for fast and accurate quantitation of cellular protein levels and requires fluorescently labeled antibodies as well as calibration standards. A critical step for quantitation remains the production of suitable detection antibodies with a precisely defined ratio of antigen-binding sites to fluorophores. Problems often arise as a consequence of inefficient and unspecific labeling which can influence antibody properties. In addition, the number of incorporated fluorophores necessitates a special normalization step for quantitation. To address these problems, we constructed different mono- and bivalent bispecific antibodies with binding site(s) for the cell surface antigens, cMET, EGFR1/HER1, ErbB2/HER2 or ErbB3/HER3 and with an additional digoxigenin-binding single-chain Fv fusion. The fluorophore Cy5 was covalently coupled to digoxigenin and quantitatively bound by the bispecific antibody. A panel of tumor cell lines was assessed under different culture conditions for absolute receptor expression levels of the indicated antigens and the data were set in relation to mRNA, gene count and immunoblot data. We could reproducibly quantify these receptors, omit the otherwise required normalization step and demonstrate the superiority of a 1 + 1 bispecific antibody. The same antibodies were also used to quantify the number of proteins in intracellular vesicles in confocal microscopy. The antibodies can be stored like regular antibodies and can be coupled with different digoxigenin-labeled fluorophores which makes them excellent tools for FACS and imaging-based experiments. PMID:23960142

Panke, C.; Weininger, D.; Haas, A.; Schelter, F.; Schlothauer, T.; Bader, S.; Sircar, R.; Josel, H.P.; Baer, U.; Burtscher, H.; Mundigl, O.; Grote, M.; Brinkmann, U.; Sustmann, C.

2013-01-01

33

Immunotherapy of Human Tumors with T-Cell-activating Bispecific Antibodies: Stimulation of Cytotoxic Pathways in Vivo1  

Microsoft Academic Search

Bispecific monoclonal antibodies (Bi-mAbs) specific for a tumor-asso- ciated antigen and the CD3 or CD28 antigen on T lymphocytes represent one of the most successful experimental strategies for the immunotherapy of cancer. We report that the in vivo administration of both a-CD3\\/CD30 and a-CD28\\/CD30 Bi-mAbs results in the specific activation of xenotrans- planted, resting human T cells infiltrating the CD30-positive

Stefan Bauer; Christoph Renner; Jan-Peter Juwana; Gerhard Held; Sascha Ohnesorge; Klaus Gerlach; Michael Pfreundschuh

1999-01-01

34

Recent advances in the generation of bispecific antibodies for tumor immunotherapy.  

PubMed

Bispecific antibodies are currently in clinical and preclinical development for the treatment of various cancers. Designed to direct and enhance the body's immune response to specific tumors, bispecific antibodies consist of a targeting domain (typically an antibody fragment that binds to a tumor antigen) linked to a triggering arm that is specific for a cell-surface molecule capable of mediating a phagocytic or lytic response by macrophages, natural killer cells, T-cells, or other effector cells. Recent animal studies have confirmed the therapeutic potential of bispecific antibodies; however, results from clinical trials so far have been less promising and have revealed clear limitations of these molecules, such as immunogenicity and severe side effects caused by mass release of inflammatory cytokines. A second generation of bispecific molecules, bispecific single-chain antibodies and hetero-oligomeric-engineered antibodies, has now been produced by using DNA recombinant technology; these may theoretically improve tumor targeting and minimize side effects, eventually replacing the full-length bispecific antibodies. Over the next few years, several recombinant bispecific antibodies are expected to enter clinical trials and ultimately emerge as new pharmaceutical products. This review briefly summarizes major trends in the development of bispecific antibodies for tumor therapy, and describes a rationale for the generation of novel recombinant molecules. PMID:15603258

Kipriyanov, Sergey M; Le Gall, Fabrice

2004-03-01

35

Bispecific small molecule–antibody conjugate targeting prostate cancer  

PubMed Central

Bispecific antibodies, which simultaneously target CD3 on T cells and tumor-associated antigens to recruit cytotoxic T cells to cancer cells, are a promising new approach to the treatment of hormone-refractory prostate cancer. Here we report a site-specific, semisynthetic method for the production of bispecific antibody-like therapeutics in which a derivative of the prostate-specific membrane antigen-binding small molecule DUPA was selectively conjugated to a mutant ?CD3 Fab containing the unnatural amino acid, p-acetylphenylalanine, at a defined site. Homogeneous conjugates were generated in excellent yields and had good solubility. The efficacy of the conjugate was optimized by modifying the linker structure, relative binding orientation, and stoichiometry of the ligand. The optimized conjugate showed potent and selective in vitro activity (EC50 ?100 pM), good serum half-life, and potent in vivo activity in prophylactic and treatment xenograft mouse models. This semisynthetic approach is likely to be applicable to the generation of additional bispecific agents using drug-like ligands selective for other cell-surface receptors. PMID:24127589

Kim, Chan Hyuk; Axup, Jun Y.; Lawson, Brian R.; Yun, Hwayoung; Tardif, Virginie; Choi, Sei Hyun; Zhou, Quan; Dubrovska, Anna; Biroc, Sandra L.; Marsden, Robin; Pinstaff, Jason; Smider, Vaughn V.; Schultz, Peter G.

2013-01-01

36

Bispecific antibodies retarget murine T cell cytotoxicity against syngeneic breast cancer in vitro and in vivo  

Microsoft Academic Search

Bispecific antibodies with specificity for CD3 and a tumor antigen can redirect cytolytic T cells to kill tumor targets, regardless of their natural specificity. To assess the clinical potential of bispecific antibodies for treatment of human cancers we have, in the present study, adapted a totally syngeneic mouse model to the targeting of mouse T cells against mouse tumors in

M. Belen Moreno; Julie A. Titus; Michael S. Cole; J. Yun Tso; Nhat Le; Chang H. Paik; Tibor Bakács; Charles M. Zacharchuk; David M. Segal; John R. Wunderlich

1995-01-01

37

MAbs . Author manuscript Bispecific antibodies for cancer therapy: the light at the end of the tunnel?  

E-print Network

. Unfortunately, none of them are able to cure cancer as single agent. Several clinical outcomes and animalMAbs . Author manuscript Page /1 10 Bispecific antibodies for cancer therapy: the light at the end conventional mAbs as cancer immunotherapeutics in a near future. MESH Keywords Animals ; Antibodies, Bispecific

Paris-Sud XI, Université de

38

Bispecific Antibody Pretargeting for Improving Cancer Imaging and Therapy  

SciTech Connect

The main objective of this project was to evaluate pretargeting systems that use a bispecific antibody (bsMAb) to improve the detection and treatment of cancer. A bsMAb has specificity to a tumor antigen, which is used to bind the tumor, while the other specificity is to a peptide that can be radiolabeled. Pretargeting is the process by which the unlabeled bsMAb is given first, and after a sufficient time (1-2 days) is given for it to localize in the tumor and clear from the blood, a small molecular weight radiolabeled peptide is given. According to a dynamic imaging study using a 99mTc-labeled peptide, the radiolabeled peptide localizes in the tumor in less than 1 hour, with > 80% of it clearing from the blood and body within this same time. Tumor/nontumor targeting ratios that are nearly 50 times better than that with a directly radiolabeled Fab fragment have been observed (Sharkey et al., ''Signal amplification in molecular imaging by a multivalent bispecific nanobody'' submitted). The bsMAbs used in this project have been composed of 3 antibodies that will target antigens found in colorectal and pancreatic cancers (CEA, CSAp, and MUC1). For the ''peptide binding moiety'' of the bsMAb, we initially examined an antibody directed to DOTA, but subsequently focused on another antibody directed against a novel compound, HSG (histamine-succinyl-glycine).

Sharkey, Robert M.

2005-02-04

39

Camel Single-domain Antibodies as Modular Building Units in Bispecific and Bivalent Antibody Constructs  

Microsoft Academic Search

Single-domain antibodies against various antigens are isolated from the unique heavy-chain antibodies of immunized camels and llamas. These minimal sized binders are very robust and bind the antigen with high affinity in a monomeric state. We evaluated the feasibil- ity to produce soluble, functional bispecific and bivalent antibodies in Escherichia coli with camel single-domain antibody fragments as building blocks. Two

Katja Els Conrath; Mark Lauwereys; Lode Wyns; Serge Muyldermans

2001-01-01

40

Ipilimumab augments antitumor activity of bispecific antibody-armed T cells  

PubMed Central

Background Ipilimumab is an antagonistic monoclonal antibody against cytotoxic T-lymphocyte antigen-4 (CTLA-4) that enhances antitumor immunity by inhibiting immunosuppressive activity of regulatory T cells (Treg). In this study, we investigated whether inhibiting Treg activity with ipilimumab during ex vivo T cell expansion could augment anti-CD3-driven T cell proliferation and enhance bispecific antibody (BiAb)-redirected antitumor cytotoxicity of activated T cells (ATC). Methods PBMC from healthy individuals were stimulated with anti-CD3 monoclonal antibody with or without ipilimumab and expanded for 10-14 days. ATC were harvested and armed with anti-CD3 x anti-EGFR BiAb (EGFRBi) or anti-CD3 x anti-CD20 BiAb (CD20Bi) to test for redirected cytotoxicity against COLO356/FG pancreatic cancer cell line or Burkitt’s lymphoma cell line (Daudi). Results In PBMC from healthy individuals, the addition of ipilimumab at the initiation of culture significantly enhanced T cell proliferation (p?=?0.0029). ATC grown in the presence of ipilimumab showed significantly increased mean tumor-specific cytotoxicity at effector:target (E:T) ratio of 25:1 directed at COLO356/FG and Daudi by 37.71% (p?bispecific antibodies. PMID:25008236

2014-01-01

41

Treatment of refractory Hodgkin’s disease with an anti-CD16\\/CD30 bispecific antibody  

Microsoft Academic Search

A group of 15 patients with refractory Hodgkin?s disease were treated in a phase I\\/II trial with the natural-killer (NK)-cell-activating\\u000a bispecific monoclonal antibody HRS-3\\/A9, which is directed against the Fc? receptor III (CD16 antigen) and the Hodgkin’s associated\\u000a CD30 antigen. The antibody was given four times every 3?–?4 days, starting with 1 mg\\/m2. The treatment was well tolerated and the

Cristoph Renner; Frank Hartmann; Michael Pfreundschuh

1997-01-01

42

Biodistribution studies with tumor-targeting bispecific antibodies reveal selective accumulation at the tumor site  

PubMed Central

Bispecific antibodies are proteins that bind two different antigens and may retarget immune cells with a binding moiety specific for a leukocyte marker. A binding event in blood could in principle prevent antibody extravasation and accumulation at the site of disease. In this study, we produced and characterized two tetravalent bispecific antibodies that bind with high affinity to the alternatively-spliced EDB domain of fibronectin, a tumor-associated antigen. The bispecific antibodies simultaneously engaged the cognate antigens (murine T cell co-receptor CD3 and hen egg lysozyme) and selectively accumulated on murine tumors in vivo. The results, which were in agreement with predictions based on pharmacokinetic modeling and antibody binding characteristics, confirmed that bispecific antibodies can reach abluminal targets without being blocked by peripheral blood leukocytes. PMID:23032949

List, Thomas; Neri, Dario

2012-01-01

43

Bispecific Antibodies: Developments and Current Perspectives  

Microsoft Academic Search

\\u000a All naturally occurring antibodies are multifunctional molecules, combining antigen-binding activity and Fc-mediated effector\\u000a functions within a single molecule (Schroeder and Cavacini 2010). Antigen binding can lead to direct neutralization of the\\u000a antigen, i.e., being antagonistic, but can also have agonistic activities, e.g., through activation of receptors (Fig. 1.1).\\u000a The Fc region is capable of mediating further effector functions, which include antibody-dependent

Roland E. Kontermann

44

A series of anti-CEA/anti-DOTA bispecific antibody formats evaluated for pre-targeting: comparison of tumor uptake and blood clearance  

PubMed Central

A series of anti-tumor/anti-chelate bispecific antibody formats were developed for pre-targeted radioimmunotherapy. Based on the anti-carcinoembryonic antigen humanized hT84.66-M5A monoclonal antibody and the anti-DOTA C8.2.5 scFv antibody fragment, this cognate series of bispecific antibodies were radioiodinated to determine their tumor targeting, biodistribution and pharmacokinetic properties in a mouse xenograft tumor model. The in vivo biodistribution studies showed that all the bispecific antibodies exhibited specific high tumor uptake but the tumor targeting was approximately one-half of the parental anti-CEA mAb due to faster blood clearance. Serum stability and FcRn studies showed no apparent reason for the faster blood clearance. A dual radiolabel biodistribution study revealed that the 111In-DOTA bispecific antibody had increased liver and spleen uptake, not seen for the 125I-version due to metabolism and release of the radioiodine from the cells. These data suggest increased clearance of the antibody fusion formats by the mononuclear phagocyte system. Importantly, a pre-targeted study showed specific tumor uptake of 177Lu-DOTA and a tumor : blood ratio of 199 : 1. This pre-targeted radiotherapeutic and substantial reduction in the radioactive exposure to the bone marrow should enhance the therapeutic potential of RIT. PMID:23175797

Yazaki, Paul J.; Lee, Brian; Channappa, Divya; Cheung, Chia-Wei; Crow, Desiree; Chea, Junie; Poku, Erasmus; Li, Lin; Andersen, Jan Terje; Sandlie, Inger; Orcutt, Kelly Davis; Wittrup, K.Dane; Shively, John E.; Raubitschek, Andrew; Colcher, David

2013-01-01

45

Development of a human IgG4 bispecific antibody for dual targeting of interleukin-4 (IL-4) and interleukin-13 (IL-13) cytokines.  

PubMed

Human bispecific antibodies have great potential for the treatment of human diseases. Although human IgG1 bispecific antibodies have been generated, few attempts have been reported in the scientific literature that extend bispecific antibodies to other human antibody isotypes. In this paper, we report our work expanding the knobs-into-holes bispecific antibody technology to the human IgG4 isotype. We apply this approach to generate a bispecific antibody that targets IL-4 and IL-13, two cytokines that play roles in type 2 inflammation. We show that IgG4 bispecific antibodies can be generated in large quantities with equivalent efficiency and quality and have comparable pharmacokinetic properties and lung partitioning, compared with the IgG1 isotype. This work broadens the range of published therapeutic bispecific antibodies with natural surface architecture and provides additional options for the generation of bispecific antibodies with differing effector functions through the use of different antibody isotypes. PMID:23880771

Spiess, Christoph; Bevers, Jack; Jackman, Janet; Chiang, Nancy; Nakamura, Gerald; Dillon, Michael; Liu, Hongbin; Molina, Patricia; Elliott, J Michael; Shatz, Whitney; Scheer, Justin M; Giese, Glen; Persson, Josefine; Zhang, Yin; Dennis, Mark S; Giulianotti, James; Gupta, Prateek; Reilly, Dorothea; Palma, Enzo; Wang, Jianyong; Stefanich, Eric; Scheerens, Heleen; Fuh, Germaine; Wu, Lawren C

2013-09-13

46

Targeting Human T-Lymphocytes with Bispecific Antibodies to React against Human Ovarian Carcinoma Cells Growing in nu\\/nu Mice  

Microsoft Academic Search

In the present study we tested whether human T-cells from normal donors can be targeted against human ovarian carcinoma cells and block i.p. growth of an established tumor in immunodeficientmice. For targeting we used chemically cross-linked bispecific monoclonal antibodies (mAbs) reacting with CD3 on the T-cells and with cell-surface antigens selectively expressed by tumor cells. The tumor model consisted of

Maria A. Garrido; Maria J. Valdayo; David F. Winkler; Julie A. Titus; Toby T. Hecht; Pilar Perez; David M. Segal; John R. Wunderlich

47

A Universal Pretargeting System for Cancer Detection and Therapy Using Bispecific Antibody1  

Microsoft Academic Search

Multistep targeting systems represent highly selective alternatives to targeting systems using directly radiolabeled antibodies for diagnostic and therapeutic applications. A flexible bispecific antibody (bsMAb) multi- step, pretargeting system that potentially can be developed for use with a variety of different imaging or therapeutic agents is described herein. The flexibility of this system is based on use of an antibody directed

Robert M. Sharkey; William J. McBride; Habibe Karacay; Ken Chang; Gary L. Griffiths; Hans J. Hansen; David M. Goldenberg

48

Monoclonal antibody therapy of cancer  

Microsoft Academic Search

The most significant recent advances in the application of monoclonal antibodies (mAbs) to oncology have been the introduction and approval of bevacizumab (Avastin), an anti–vascular endothelial growth factor antibody, and of cetuximab (Erbitux), an anti–epidermal growth factor antibody. In combination with standard chemotherapy regimens, bevacizumab significantly prolongs the survival of patients with metastatic cancers of the colorectum, breast and lung.

Gregory P Adams; Louis M Weiner

2005-01-01

49

Uses of monoclonal antibody 8H9  

DOEpatents

This invention provides a composition comprising an effective amount of monoclonal antibody 8H9 or a derivative thereof and a suitable carrier. This invention provides a pharmaceutical composition comprising an effective amount of monoclonal antibody 8H9 or a derivative thereof and a pharmaceutically acceptable carrier. This invention also provides an antibody other than the monoclonal antibody 8H9 comprising the complementary determining regions of monoclonal antibody 8H9 or a derivative thereof, capable of binding to the same antigen as the monoclonal antibody 8H9. This invention provides a substance capable of competitively inhibiting the binding of monoclonal antibody 8H9. This invention also provides an isolated scFv of monoclonal antibody 8H9 or a derivative thereof. This invention also provides the 8H9 antigen. This invention also provides different uses of the monoclonal antibody 8H9 or its derivative.

Cheung, Nai-Kong V.

2013-04-09

50

Progress in overcoming the chain association issue in bispecific heterodimeric IgG antibodies.  

PubMed

The development of bispecific antibodies has attracted substantial interest, and many different formats have been described. Those specifically containing an Fc part are mostly tetravalent, such as stabilized IgG-scFv fusions or dual-variable domain (DVD) IgGs. However, although they exhibit IgG-like properties and technical developability, these formats differ in size and geometry from classical IgG antibodies. Thus, considerable efforts focus on bispecific heterodimeric IgG antibodies that more closely mimic natural IgG molecules. The inherent chain association problem encountered when producing bispecific heterodimeric IgG antibodies can be overcome by several methods. While technologies like knobs-into-holes (KiH) combined with a common light chain or the CrossMab technology enforce the correct chain association, other approaches, e.g., the dual-acting Fab (DAF) IgGs, do not rely on a heterodimeric Fc part. This review discusses the state of the art in bispecific heterodimeric IgG antibodies, with an emphasis on recent progress. PMID:22925968

Klein, Christian; Sustmann, Claudio; Thomas, Markus; Stubenrauch, Kay; Croasdale, Rebecca; Schanzer, Jürgen; Brinkmann, Ulrich; Kettenberger, Hubert; Regula, Jörg T; Schaefer, Wolfgang

2012-01-01

51

Progress in overcoming the chain association issue in bispecific heterodimeric IgG antibodies  

PubMed Central

The development of bispecific antibodies has attracted substantial interest, and many different formats have been described. Those specifically containing an Fc part are mostly tetravalent, such as stabilized IgG-scFv fusions or dual-variable domain (DVD) IgGs. However, although they exhibit IgG-like properties and technical developability, these formats differ in size and geometry from classical IgG antibodies. Thus, considerable efforts focus on bispecific heterodimeric IgG antibodies that more closely mimic natural IgG molecules. The inherent chain association problem encountered when producing bispecific heterodimeric IgG antibodies can be overcome by several methods. While technologies like knobs-into-holes (KiH) combined with a common light chain or the CrossMab technology enforce the correct chain association, other approaches, e.g., the dual-acting Fab (DAF) IgGs, do not rely on a heterodimeric Fc part. This review discusses the state of the art in bispecific heterodimeric IgG antibodies, with an emphasis on recent progress. PMID:22925968

Klein, Christian; Sustmann, Claudio; Thomas, Markus; Stubenrauch, Kay; Croasdale, Rebecca; Schanzer, Jürgen; Brinkmann, Ulrich; Kettenberger, Hubert; Regula, Jörg T.; Schaefer, Wolfgang

2012-01-01

52

Therapeutic assessment of SEED: a new engineered antibody platform designed to generate mono- and bispecific antibodies.  

PubMed

The strand-exchange engineered domain (SEED) platform was designed to generate asymmetric and bispecific antibody-like molecules, a capability that expands therapeutic applications of natural antibodies. This new protein engineered platform is based on exchanging structurally related sequences of immunoglobulin within the conserved CH3 domains. Alternating sequences from human IgA and IgG in the SEED CH3 domains generate two asymmetric but complementary domains, designated AG and GA. The SEED design allows efficient generation of AG/GA heterodimers, while disfavoring homodimerization of AG and GA SEED CH3 domains. Using a clinically validated antibody (C225), we tested whether Fab derivatives constructed on the SEED platform retain desirable therapeutic antibody features such as in vitro and in vivo stability, favorable pharmacokinetics, ligand binding and effector functions including antibody-dependent cell-mediated cytotoxicity and complement-dependent cytotoxicity. In addition, we tested SEED with combinations of binder domains (scFv, VHH, Fab). Mono- and bivalent Fab-SEED fusions retain full binding affinity, have excellent biochemical and biophysical stability, and retain desirable antibody-like characteristics conferred by Fc domains. Furthermore, SEED is compatible with different combinations of Fab, scFv and VHH domains. Our assessment shows that the new SEED platform expands therapeutic applications of natural antibodies by generating heterodimeric Fc-analog proteins. PMID:21498564

Muda, Marco; Gross, Alec W; Dawson, Jessica P; He, Chaomei; Kurosawa, Emmi; Schweickhardt, Rene; Dugas, Melanie; Soloviev, Maria; Bernhardt, Anna; Fischer, David; Wesolowski, John S; Kelton, Christie; Neuteboom, Berend; Hock, Bjoern

2011-05-01

53

A novel bispecific antibody format enables simultaneous bivalent and monovalent co-engagement of distinct target antigens.  

PubMed

Bispecific antibodies based on full-length antibody structures are more optimal than fragment-based formats because they benefit from the favorable properties of the Fc region. However, the homodimeric nature of Fc effectively imposes bivalent binding on all current full-length bispecific antibodies, an attribute that can result in nonspecific activation of cross-linked receptors. We engineered a novel bispecific format, referred to as mAb-Fv, that utilizes a heterodimeric Fc region to enable monovalent co-engagement of a second target antigen in a full-length context. mAb-Fv constructs co-targeting CD16 and CD3 were expressed and purified as heterodimeric species, bound selectively to their co-target antigens, and mediated potent cytotoxic activity by NK cells and T cells, respectively. The capacity to co-engage distinct target antigens simultaneously with different valencies is an improved feature for bispecific antibodies with promising therapeutic implications. PMID:22123055

Moore, Gregory L; Bautista, Cristina; Pong, Erik; Nguyen, Duc-Hanh T; Jacinto, Jonathan; Eivazi, Araz; Muchhal, Umesh S; Karki, Sher; Chu, Seung Y; Lazar, Greg A

2011-01-01

54

A novel bispecific antibody format enables simultaneous bivalent and monovalent co-engagement of distinct target antigens  

PubMed Central

Bispecific antibodies based on full-length antibody structures are more optimal than fragment-based formats because they benefit from the favorable properties of the Fc region. However, the homodimeric nature of Fc effectively imposes bivalent binding on all current full-length bispecific antibodies, an attribute that can result in nonspecific activation of cross-linked receptors. We engineered a novel bispecific format, referred to as mAb-Fv, that utilizes a heterodimeric Fc region to enable monovalent co-engagement of a second target antigen in a full-length context. mAb-Fv constructs co-targeting CD16 and CD3 were expressed and purified as heterodimeric species, bound selectively to their co-target antigens and mediated potent cytotoxic activity by NK cells and T cells, respectively. The capacity to co-engage distinct target antigens simultaneously with different valencies is an improved feature for bispecific antibodies with promising therapeutic implications. PMID:22123055

Bautista, Cristina; Pong, Erik; Nguyen, Duc-Hanh T; Jacinto, Jonathan; Eivazi, Araz; Muchhal, Umesh S; Karki, Sher; Chu, Seung Y; Lazar, Greg A

2011-01-01

55

Immunoglobulin domain crossover as a generic approach for the production of bispecific IgG antibodies.  

PubMed

We describe a generic approach to assemble correctly two heavy and two light chains, derived from two existing antibodies, to form human bivalent bispecific IgG antibodies without use of artificial linkers. Based on the knobs-into-holes technology that enables heterodimerization of the heavy chains, correct association of the light chains and their cognate heavy chains is achieved by exchange of heavy-chain and light-chain domains within the antigen binding fragment (Fab) of one half of the bispecific antibody. This "crossover" retains the antigen-binding affinity but makes the two arms so different that light-chain mispairing can no longer occur. Applying the three possible "CrossMab" formats, we generated bispecific antibodies against angiopoietin-2 (Ang-2) and vascular endothelial growth factor A (VEGF-A) and show that they can be produced by standard techniques, exhibit stabilities comparable to natural antibodies, and bind both targets simultaneously with unaltered affinity. Because of its superior side-product profile, the CrossMab(CH1-CL) was selected for in vivo profiling and showed potent antiangiogenic and antitumoral activity. PMID:21690412

Schaefer, Wolfgang; Regula, Jörg T; Bähner, Monika; Schanzer, Jürgen; Croasdale, Rebecca; Dürr, Harald; Gassner, Christian; Georges, Guy; Kettenberger, Hubert; Imhof-Jung, Sabine; Schwaiger, Manfred; Stubenrauch, Kay G; Sustmann, Claudio; Thomas, Markus; Scheuer, Werner; Klein, Christian

2011-07-01

56

Initiation of humoral and cellular immune responses in patients with refractory Hodgkin's disease by treatment with an anti-CD16\\/CD30 bispecific antibody  

Microsoft Academic Search

Fifteen patients with refractory Hodgkin's disease were treated in a dose-escalation trial with the bispecific monoclonal\\u000a antibody (bi-mAb) HRS-3\\/A9, which is directed against the Fc? receptor III (CD16 antigen) and the Hodgkin's-associated CD30\\u000a antigen. Treatment consisted of four cycles of four bi-mAb infusions given over 1?h every 3–4 days at different dose levels\\u000a ranging from 1?mg\\/m2 to 64?mg\\/m2. Measurable serum

Christoph Renner; Frank Hartmann; Wolfram Jung; Christina Deisting; Marieta Juwana; Michael Pfreundschuh

2000-01-01

57

Monoclonal antibody therapeutics with up to five specificities  

PubMed Central

The recognition that few human diseases are thoroughly addressed by mono-specific, monoclonal antibodies (mAbs) continues to drive the development of antibody therapeutics with additional specificities and enhanced activity. Historically, efforts to engineer additional antigen recognition into molecules have relied predominantly on the reformatting of immunoglobulin domains. In this report we describe a series of fully functional mAbs to which additional specificities have been imparted through the recombinant fusion of relatively short polypeptides sequences. The sequences are selected for binding to a particular target from combinatorial libraries that express linear, disulfide-constrained, or domain-based structures. The potential for fusion of peptides to the N- and C- termini of both the heavy and light chains affords the bivalent expression of up to four different peptides. The resulting molecules, called zybodies, can gain up to four additional specificities, while retaining the original functionality and specificity of the scaffold antibody. We explore the use of two clinically significant oncology antibodies, trastuzumab and cetuximab, as zybody scaffolds and demonstrate functional enhancements in each case. The affect of fusion position on both peptide and scaffold function is explored, and penta-specific zybodies are demonstrated to simultaneously engage five targets (ErbB2, EGFR, IGF-1R, Ang2 and integrin ?v?3). Bispecific, trastuzumab-based zybodies targeting ErbB2 and Ang2 are shown to exhibit superior efficacy to trastuzumab in an angiogenesis-dependent xenograft tumor model. A cetuximab-based bispecific zybody that targeting EGFR and ErbB3 simultaneously disrupted multiple intracellular signaling pathways; inhibited tumor cell proliferation; and showed efficacy superior to that of cetuximab in a xenograft tumor model. PMID:23575268

LaFleur, David W.; Abramyan, Donara; Kanakaraj, Palanisamy; Smith, Rodger G.; Shah, Rutul R.; Wang, Geping; Yao, Xiao-Tao; Kankanala, Spandana; Boyd, Ernie; Zaritskaya, Liubov; Nam, Viktoriya; Puffer, Bridget A.; Buasen, Pete; Kaithamana, Shashi; Burnette, Andrew F.; Krishnamurthy, Rajesh; Patel, Dimki; Roschke, Viktor V.; Kiener, Peter A.; Hilbert, David M.; Barbas III, Carlos F.

2013-01-01

58

Uses of monoclonal antibody 8H9  

DOEpatents

This invention provides a composition comprising an effective amount of monoclonal antibody 8H9 or a derivative thereof and a suitable carrier. This invention provides a pharmaceutical composition comprising an effective amount of monoclonal antibody 8H9 or a derivative thereof and a pharmaceutically acceptable carrier. This invention also provides an antibody other than the monoclonal antibody 8H9 comprising the complementary determining regions of monoclonal antibody 8H9 or a derivative thereof, capable of binding to the same antigen as the monoclonal antibody 8H9. This invention provides a substance capable of competitively inhibiting the binding of monoclonal antibody 8H9. This invention also provides an isolated scFv of monoclonal antibody 8H9 or a derivative thereof. This invention also provides the 8H9 antigen. This invention also provides a method of inhibiting the growth of tumor cells comprising contacting said tumor cells with an appropriate amount of monoclonal antibody 8H9 or a derivative thereof.

Cheung, Nai-Kong V

2013-08-06

59

Signal amplification in molecular imaging by pretargeting a multivalent, bispecific antibody  

Microsoft Academic Search

Here we describe molecular imaging of cancer using signal amplification of a radiotracer in situ by pretargeting a multivalent, bispecific antibody to carcinoembryonic antigen (CEA), which subsequently also captures a radioactive hapten-peptide. Human colon cancer xenografts as small as ?0.15 g were disclosed in nude mice within 1 h of giving the radiotracer, with tumor\\/blood ratios increased by ?40-fold (?10:1

Robert M Sharkey; Thomas M Cardillo; Edmund A Rossi; Chien-Hsing Chang; Habibe Karacay; William J McBride; Hans J Hansen; Ivan D Horak; David M Goldenberg

2005-01-01

60

Use of trifunctional bispecific antibodies to prevent graft versus host disease induced by allogeneic lymphocytes  

Microsoft Academic Search

A trifunctional bispecific antibody (BiLu) directed against murine CD3 and human epithelial-cell adhesion molecule (Ep- CAM) was tested for its ability to improve cell-mediated adoptive immunotherapy in a murine model of B16 melanoma cells transfected with human EpCAM. Intraperi- toneal inoculation of naive C57BL\\/6 (C57) splenocytes induced lethal graft versus host disease (GVHD) in 85% to 97% of sublethally irradiated

Shoshana Morecki; Horst Lindhofer; Elena Yacovlev; Yael Gelfand; Shimon Slavin

2006-01-01

61

Monoclonal antibodies to the human mammary gland  

Microsoft Academic Search

Mouse monoclonal antibodies have been raised to the human milk fat globule membrane. The distribution of the antigens detected by four of the antibodies has been examined in formalin-fixed, paraffin-embedded human tissues by light microscopic immunocytochemistry. The four antibodies stain lactating breast and normal resting breast. Two exclusively stain the luminal membranes of breast epithelial cells. A third antibody stains

C. S. Foster; P. A. W. Edwards; E. A. Dinsdale; A. M. Neville

1982-01-01

62

LC-MS characterization and purity assessment of a prototype bispecific antibody  

PubMed Central

Bispecific IgG asymmetric (heterodimeric) antibodies offer enhanced therapeutic efficacy, but present unique challenges for drug development. These challenges are related to the proper assembly of heavy and light chains. Impurities such as symmetric (homodimeric) antibodies can arise with improper assembly. A new method to assess heterodimer purity of such bispecific antibody products is needed because traditional separation-based purity assays are unable to separate or quantify homodimer impurities. This paper presents a liquid chromatography-mass spectrometry (LC-MS)-based method for evaluating heterodimeric purity of a prototype asymmetric antibody containing two different heavy chains and two identical light chains. The heterodimer and independently expressed homodimeric standards were characterized by two complementary LC-MS techniques: Intact protein mass measurement of deglycosylated antibody and peptide map analyses. Intact protein mass analysis was used to check molecular integrity and composition. LC-MSE peptide mapping of Lys-C digests was used to verify protein sequences and characterize post-translational modifications, including C-terminal truncation species. Guided by the characterization results, a heterodimer purity assay was demonstrated by intact protein mass analysis of pure deglycosylated heterodimer spiked with each deglycosylated homodimeric standard. The assay was capable of detecting low levels (2%) of spiked homodimers in conjunction with co-eluting half antibodies and multiple mass species present in the homodimer standards and providing relative purity differences between samples. Detection of minor homodimer and half-antibody C-terminal truncation species at levels as low as 0.6% demonstrates the sensitivity of the method. This method is suitable for purity assessment of heterodimer samples during process and purification development of bispecific antibodies, e.g., clone selection. PMID:23884083

Woods, R. Jeremy; Xie, Michael Hongwei; Von Kreudenstein, Thomas Spreter; Ng, Gordon Y.; Dixit, Surjit B.

2013-01-01

63

Monoclonal antibodies that detect live salmonellae.  

PubMed Central

Nine immunoglobulin G and nine immunoglobulin M murine monoclonal antibody-producing hybridomas reactive with live Salmonella bacteria were obtained from several fusions of immune spleen cells and Sp2/0 myeloma cells. The antibodies were selected by the magnetic immunoluminescence assay. The monoclonal antibodies were reactive with serogroups A, B, C1, C2, D, E, and K and Salmonella choleraesuis subsp. diarizonae. Each monoclonal antibody proved to be reactive with a distinct serotype. Clinical isolates belonging to these Salmonella serogroups could be detected. Reactivity with non-Salmonella bacteria proved to be minor. Images PMID:1476430

Torensma, R; Visser, M J; Aarsman, C J; Poppelier, M J; van Beurden, R; Fluit, A C; Verhoef, J

1992-01-01

64

Improved monoclonal antibodies to halodeoxyuridine  

DOEpatents

The development, method of production, characterization and methods of use of two hybridomas, CIdU-1 (ATCC Accession No. HB-8321) and CIdU-2 (ATCC Accession No. HB-8320), are described. These secrete IgG/sub 1/(K) immunoglobulins that react with halodeoxyuridine (HdU or halodU) such as bromo, chloro, fluoro and iodo deoxyuridine (BrdU, CldU, FdU and IdU), whether these are free in solution or incorporated into single stranded DNA in whole cells. The antibodies do not react with naturally occurring free nucleic acids or with deoxyribonucleic acid (DNA) or ribonucleic acid (RNA) polymers. These antibodies are suitable for use in enzyme immunoassays for free CldU, FdU, IdU and BrdU and for detecting cells with these nucleotides incorporated into them. The monoclonal antibodies are useful in the detection of the sensitivity of tumor cells to specific chemotherapeutic agents, in the measurement of the rate of cellular DNA synthesis, in the measurement of the rate of proliferation of normal and malignant cells and in the detection of HPRT deficiency in cells. 1 tab.

Vanderlaan, M.; Dolbeare, F.A.; Gray, J.W.; Thomas, C.B.

1983-10-18

65

Monoclonal Antibodies for the Treatment of Cancer  

PubMed Central

Over the past decade, the clinical utility of monoclonal antibodies has been realized and antibodies are now a mainstay for the treatment of cancer. Antibodies have the unique capacity to target and kill tumor cells while simultaneously activating immune effectors to kill tumor cells through the complement cascade or antibody-dependent cellular cytotoxicity (ADCC). This multifaceted mechanism of action combined with target specificity underlies the capacity of antibodies to elicit anti-tumor responses while minimizing the frequency and magnitude of adverse events. This review will focus on mechanisms of action, clinical applications and putative mechanisms of resistance to monoclonal antibody therapy in the context of cancer. PMID:22245472

Shuptrine, Casey; Surana, Rishi; Weiner, Louis M.

2012-01-01

66

Original article Monoclonal antibodies against goldfish  

E-print Network

immunoglobulin and antibody levels by ELISA in carp (Cyprinus carpio) AK Siwicki C Vergnet J Charlemagne M Dunier decreased until day 28. monoclonal antibody / ELISA / ELISPOT / antibody-secreting cells / Cyprinus carpio suite. anticorps monoc/ona//EL/S/Cyprinus carpio

Paris-Sud XI, Université de

67

Anti-CD22/CD20 Bispecific Antibody with Enhanced Trogocytosis for Treatment of Lupus  

PubMed Central

The humanized anti-CD22 antibody, epratuzumab, has demonstrated therapeutic activity in clinical trials of lymphoma, leukemia and autoimmune diseases, treating currently over 1500 cases of non-Hodgkin lymphoma, acute lymphoblastic leukemias, Waldenström’s macroglobulinemia, Sjögren’s syndrome, and systemic lupus erythematosus. Because epratuzumab reduces on average only 35% of circulating B cells in patients, and has minimal antibody-dependent cellular cytotoxicity and negligible complement-dependent cytotoxicity when evaluated in vitro, its therapeutic activity may not result completely from B-cell depletion. We reported recently that epratuzumab mediates Fc/FcR-dependent membrane transfer from B cells to effector cells via trogocytosis, resulting in a substantial reduction of multiple BCR modulators, including CD22, CD19, CD21, and CD79b, as well as key cell adhesion molecules, including CD44, CD62L, and ?7 integrin, on the surface of B cells in peripheral blood mononuclear cells obtained from normal donors or SLE patients. Rituximab has clinical activity in lupus, but failed to achieve primary endpoints in a Phase III trial. This is the first study of trogocytosis mediated by bispecific antibodies targeting neighboring cell-surface proteins, CD22, CD20, and CD19, as demonstrated by flow cytometry and immunofluorescence microscopy. We show that, compared to epratuzumab, a bispecific hexavalent antibody comprising epratuzumab and veltuzumab (humanized anti-CD20 mAb) exhibits enhanced trogocytosis resulting in major reductions in B-cell surface levels of CD19, CD20, CD21, CD22, CD79b, CD44, CD62L and ?7-integrin, and with considerably less immunocompromising B-cell depletion that would result with anti-CD20 mAbs such as veltuzumab or rituximab, given either alone or in combination with epratuzumab. A CD22/CD19 bispecific hexavalent antibody, which exhibited enhanced trogocytosis of some antigens and minimal B-cell depletion, may also be therapeutically useful. The bispecific antibody is a candidate for improved treatment of lupus and other autoimmune diseases, offering advantages over administration of the two parental antibodies in combination. PMID:24841238

Rossi, Edmund A.; Chang, Chien-Hsing; Goldenberg, David M.

2014-01-01

68

Application of 300× enhanced fluorescence on a plasmonic chip modified with a bispecific antibody to a sensitive immunosensor.  

PubMed

The grating substrate covered with a metal layer, a plasmonic chip, and a bispecific antibody can play a key role in the sensitive detection of a marker protein with an immunosensor, because of the provision of an enhanced fluorescence signal and the preparation of a sensor surface densely modified with capture antibody, respectively. In this study, one of the tumor markers, a soluble epidermal growth factor receptor (sEGFR), was selected as the target to be detected. The ZnO- and silver-coated plasmonic chip with precise regularity and the appropriate duty ratio in the periodic structure further enhanced the fluorescence intensity. As for sensor surface modification with capture antibody, a bispecific antibody (anti-sEGFR and anti-ZnO antibody), the concentrated bispecific antibody solution was found to nonlinearly form a surface densely immobilized with antibody, because the binding process of a bispecific antibody to the ZnO surface can be a competitive process with adsorption of phosphate. As a result, the interface on the plasmonic chip provided a 300× enhanced fluorescence signal compared with that on a ZnO-coated glass slide, and therefore sEGFR was found to be quantitatively detected in a wide concentration range from 10 nM to 700 fM on our plasmonic surface. PMID:23945148

Tawa, Keiko; Umetsu, Mitsuo; Nakazawa, Hikaru; Hattori, Takamitsu; Kumagai, Izumi

2013-09-11

69

Cost modeling for monoclonal antibody manufacturing  

E-print Network

The Novartis BioPharmOps division is responsible for manufacturing large molecule products, including monoclonal antibodies, for late stage clinical trials and commercial sales. The BioPharmOps site in Huningue, France is ...

Simpson, Christina M. (Christina Margaret)

2011-01-01

70

Monoclonal Antibody That Defines Human Myoepithelium  

NASA Astrophysics Data System (ADS)

We have isolated a mouse monoclonal antibody that, upon immunohistochemical localization in frozen sections, displays specificity for human myoepithelial cells in the resting mammary gland, sweat glands, and salivary glands. Furthermore, this antibody was strongly and homogeneously reactive with frozen sections of 3 of 60 breast carcinoma specimens. Using immunolocalization techniques in conjunction with polyacrylamide gel electrophoresis, we have determined that the reactivity of this monoclonal antibody is directed toward a 51,000-dalton keratin polypeptide. The potential uses of this antibody in the prognosis of human mammary carcinoma and in understanding the role of the myoepithelium in development and differentiation are discussed.

Dairkee, Shahnaz Hashmi; Blayney, Carlene; Smith, Helene S.; Hackett, Adeline J.

1985-11-01

71

Preparation of astatine-labeled monoclonal antibodies  

SciTech Connect

In the cationic state astatine forms a stable complex with diethylenetriaminepentaacetic acid. Thanks to this complex, astatine can be bound to monoclonal antibodies of the RYa{sub 1} type. The most favorable conditions for preparing astatine-labeled antibodies are established. The chromatographic analysis and electromigration experiments showed that astatine is firmly linked to a biomolecule in vitro and it did not escape from labeled monoclonal antibodies even under treatment with such highly effective astatine-complexing agent as thiourea. The immune activity of astatine-labeled antibodies did not change even after 20 h.

Milesz, S.; Norseev, Yu.V.; Szucs, Z. [Joint Inst. for Nuclear Research, Dubna (Russian Federation)]|[Central Inst. of Physical Research, Budapest (Hungary)

1995-07-01

72

[Therapeutic monoclonal antibodies in oncology].  

PubMed

Advances in bioengineering have lead to the possibility to conduct large scale production of monoclonal antibodies (MoAB) and to reduce progressively the murine component from 30% (chimeric MoAB) to 5% (humanized MoAB) to 0% (human MoAB). Three types of extracellular components are targeted in solid tumours: (1) Growth factors with transmembrane tyrosine kinase receptors either of tumour cells (IGF1) or endothelial cells (Bevacizumab). Bevacizumab has activity additive to that of chemotherapy in advanced colorectal, non squamous lung, ovarian, metastatic breast cancers and glioblastomas; (2) Extracellular domain of those transmembrane receptors: EGFR in colorectal cancer if no activating of KRAS with cetuximab and panitumumab, head and neck carcinomas with radiotherapy, and probably squamous lung cancers. Anti-ERBB2 MoAb are now a constitutive part of therapy of ERBB2 positive breast cancers at any stage; (3) Differentiation cluster regulating relationship ot tumour and stromal cells and in particular immunologic effectors. This is the case of anti-CTLA4 MoAB ipilimumab which activates and amplifies immunological cytotoxic response against melanoma with improved survival. These activities are achieved to the expense of class, target related toxicity conditioned by expression of the target on normal cells and or mechanism of action (immunological toxicity with ipilimumab). Of note a synergistic or additive activity with valid treatment regimens of targeted cancers and now targeted small molecules. PMID:22863361

Bouzid, K; Bedairia, N; Marty, M

2012-08-01

73

Heterodimeric bispecific antibody-derivatives against CD19 and CD16 induce effective antibody-dependent cellular cytotoxicity against B-lymphoid tumor cells  

Microsoft Academic Search

Bispecific scFv antibody-derivatives (bsscFvs) recruiting natural killer (NK) cells for the lysis of malignant cells have therapeutic potential. However, a bsscFv specific for the B-lymphoid tumor antigen CD19 and the trigger molecule CD16 on NK cells had similar affinities for both antigens (42 and 58nM, respectively) and was not optimal for cytotoxicity. Therefore, a bispecific tribody (bsTb) was constructed with

Christian Kellner; Joerg Bruenke; Heike Horner; Joerg Schubert; Michael Schwenkert; Kristin Mentz; Karin Barbin; Christoph Stein; Matthias Peipp; Bernhard Stockmeyer; Georg H. Fey

2011-01-01

74

Synthesis and Evaluation of an Anti-MLC1 × Anti-CD90 Bispecific Antibody for Targeting and Retaining Bone-Marrow Derived Multipotent Stromal Cells in Infarcted Myocardium  

PubMed Central

A key issue regarding the use of stem cells in cardiovascular regenerative medicine is their retention in target tissues. Here, we have generated and assessed a bispecific antibody heterodimer designed to improve the retention of bone marrow–derived multipotent stromal cells (BMMSC) in cardiac tissue damaged by myocardial infarction. The heterodimer comprises an anti-human CD90 monoclonal antibody (mAb) (clone 5E10) and an anti-myosin light chain 1 (MLC1) mAb (clone MLM508) covalently cross-linked by a bis-aryl hydrazone. We modified the anti-CD90 antibody with a pegylated-4-formylbenzamide moiety to a molar substitution ratio (MSR) of 2.6 and the anti-MLC1 antibody with a 6-hydrazinonicotinamide moiety to a MSR of 0.9. The covalent modifications had no significant deleterious effect on mAb epitope binding. Furthermore, the binding of anti-CD90 antibody to BMMSCs did not prevent their differentiation into adipo-, chondro-, or osteogenic lineages. Modified antibodies were combined under mild conditions (RT, pH 6, 1 h) in the presence of a catalyst (aniline) to allow for rapid generation of the covalent bis-aryl hydrazone, which was monitored at A354. We evaluated epitope immunoreactivity for each mAb in the construct. Flow cytometry demonstrated binding of the bispecific construct to BMMSCs that was competed by free anti-CD90 mAb, verifying that modification and cross-linking were not detrimental to the anti-CD90 complementarity-determining region. Similarly, ELISA-based assays demonstrated bispecific antibody binding to plastic-immobilized recombinant MLC1. Excess anti-MLC1 mAb competed for bispecific antibody binding. Finally, the anti-CD90 × anti-MLC1 bispecific antibody construct induced BMMSC adhesion to plastic-immobilized MLC1 that was resistant to shear stress, as measured in parallel-plate flow chamber assays. We used mAbs that bind both human antigens and the respective pig homologues. Thus, the anti-CD90 × anti-MLC1 bispecific antibody may be used in large animal studies of acute myocardial infarction and may provide a starting point for clinical studies. PMID:21749133

Gundlach, C. William; Caivano, Amy; Cabreira-Hansen, Maria da Graca; Gahremanpour, Amir; Brown, Wells S.; Zheng, Yi; McIntyre, Bradley W.; Willerson, James T.; Dixon, Richard A.F.; Perin, Emerson C.; Woodside, Darren G.

2011-01-01

75

Single-domain antibody-based and linker-free bispecific antibodies targeting Fc?RIII induce potent antitumor activity without recruiting regulatory T cells.  

PubMed

Antibody-dependent cell-mediated cytotoxicity, one of the most prominent modes of action of antitumor antibodies, suffers from important limitations due to the need for optimal interactions with Fc? receptors. In this work, we report the design of a new bispecific antibody format, compact and linker-free, based on the use of llama single-domain antibodies that are capable of circumventing most of these limitations. This bispecific antibody format was created by fusing single-domain antibodies directed against the carcinoembryonic antigen and the activating Fc?RIIIa receptor to human C? and CH1 immunoglobulin G1 domains, acting as a natural dimerization motif. In vitro and in vivo characterization of these Fab-like bispecific molecules revealed favorable features for further development as a therapeutic molecule. They are easy to produce in Escherichia coli, very stable, and elicit potent lysis of tumor cells by human natural killer cells at picomolar concentrations. Unlike conventional antibodies, they do not engage inhibitory Fc?RIIb receptor, do not compete with serum immunoglobulins G for receptor binding, and their cytotoxic activity is independent of Fc glycosylation and Fc?RIIIa polymorphism. As opposed to anti-CD3 bispecific antitumor antibodies, they do not engage regulatory T cells as these latter cells do not express Fc?RIII. Studies in nonobese diabetic/severe combined immunodeficient gamma mice xenografted with carcinoembryonic antigen-positive tumor cells showed that Fab-like bispecific molecules in the presence of human peripheral blood mononuclear cells significantly slow down tumor growth. This new compact, linker-free bispecific antibody format offers a promising approach for optimizing antibody-based therapies. PMID:23757164

Rozan, Caroline; Cornillon, Amélie; Pétiard, Corinne; Chartier, Martine; Behar, Ghislaine; Boix, Charlotte; Kerfelec, Brigitte; Robert, Bruno; Pčlegrin, André; Chames, Patrick; Teillaud, Jean-Luc; Baty, Daniel

2013-08-01

76

Multimodal Cancer Therapy Involving Oncolytic Newcastle Disease Virus, Autologous Immune Cells, and Bi-Specific Antibodies  

PubMed Central

This paper focuses on oncolytic Newcastle disease virus (NDV). This paper summarizes (i) the peculiarities of this virus as an anti-cancer and immune stimulatory agent and (ii) the approaches to further harness this virus as a vector to combat cancer. Special emphasis is given on combining virus therapy with cell therapy and on improving tumor targeting. The review will include some of the authors work on NDV, bi-specific antibodies, and cell therapy as building blocks for a new perspective of multimodal cancer therapy. The broad anti-tumor immune reactivation includes innate and adaptive, tumor antigen (TA) specific and TA independent activities PMID:25309868

Schirrmacher, Volker; Fournier, Philippe

2014-01-01

77

Glycosylation pattern of humanized IgG-like bispecific antibody produced by recombinant CHO cells  

Microsoft Academic Search

The glycosylation pattern of a humanized anti-EGFR×anti-CD3 bispecific single-chain diabody with an Fc portion (hEx3-scDb-Fc)\\u000a produced by recombinant Chinese hamster ovary cells was evaluated and compared with those of a recombinant humanized anti-IL-8\\u000a antibody (IgG1) and human serum IgG. N-Linked oligosaccharide structures were estimated by two-dimensional high-performance liquid chromatography and matrix-assisted\\u000a laser desorption\\/ionization time-of-flight mass spectrometry. No sialylation was observed

Wook-Dong Kim; Miwako Tokunaga; Hiroyuki Ozaki; Takuya Ishibashi; Kohsuke Honda; Hiroyuki Kajiura; Kazuhito Fujiyama; Ryutaro Asano; Izumi Kumagai; Takeshi Omasa; Hisao Ohtake

2010-01-01

78

Optimization of multivalent bispecific antibodies and immunocytokines with improved in vivo properties.  

PubMed

Multifunctional antibody-based biologics, such as bispecific antibodies and immunocytokines, can be difficult to produce with sufficient yield and stability, and often exhibit inferior pharmacokinetics. Dock-and-Lock (DNL) is a modular method that combines recombinant engineering with site-specific conjugation, allowing the construction of various complex, yet defined, biostructures with multivalency and multispecificity. The technology platform exploits the natural interaction between two interactive human protein binding domains that are modified to provide covalent fusion. We explored the potential application of a new class of IgG-based DNL modules with an anchor domain fused at the C-terminal end of the kappa light chain (C(k)), instead of the C-terminal end of the Fc. Two C(k)-derived prototypes, an anti-CD22/CD20 bispecific hexavalent antibody, comprising epratuzumab (anti-CD22) and four Fabs of veltuzumab (anti-CD20), and a CD20-targeting immunocytokine, comprising veltuzumab and four molecules of interferon-?2b, were compared to their Fc-derived counterparts. The Ck-based conjugates exhibited superior Fc-effector functions in vitro, as well as improved pharmacokinetics, stability, and anti-lymphoma activity in vivo. These results favor the selection of DNL conjugates with the C(k)-design for future clinical development. PMID:23116517

Rossi, Edmund A; Chang, Chien-Hsing; Cardillo, Thomas M; Goldenberg, David M

2013-01-16

79

Use of trifunctional bispecific antibodies to prevent graft versus host disease induced by allogeneic lymphocytes.  

PubMed

A trifunctional bispecific antibody (BiLu) directed against murine CD3 and human epithelial-cell adhesion molecule (EpCAM) was tested for its ability to improve cell-mediated adoptive immunotherapy in a murine model of B16 melanoma cells transfected with human EpCAM. Intraperitoneal inoculation of naive C57BL/6 (C57) splenocytes induced lethal graft versus host disease (GVHD) in 85% to 97% of sublethally irradiated (BALB/c x C57BL/6) F1 (F1) hosts inoculated intraperitoneally with a sublethal or lethal dose of melanoma cells. BiLu antibodies given intraperitoneally concomitantly with alloreactive C57 cells effectively prevented GVHD-related and tumor-related death in 16 of 25 F1 mice inoculated with a sublethal tumor-cell dose and in 10 of 20 mice inoculated with a lethal tumor-cell dose over a follow-up period of more than 200 days. BiLu treatment also efficiently prevented severe GVHD, which was induced by high doses of BALB/c-derived splenocytes. Trifunctional bispecific antibodies (TbsAbs) capable of cross-linking T lymphocytes, natural killer, and other FcgammaR-positive effector cells, via their Fc region, to the tumor cells may be applied together with adoptive allogeneic-cell therapy to maximize antitumor responses while acting on GVHD in patients with minimal residual disease. PMID:16234351

Morecki, Shoshana; Lindhofer, Horst; Yacovlev, Elena; Gelfand, Yael; Slavin, Shimon

2006-02-15

80

Phase I clinical trial of the bispecific antibody MDX-H210 (anti-Fc?RI × anti-HER-2\\/neu) in combination with Filgrastim (G-CSF) for treatment of advanced breast cancer  

Microsoft Academic Search

A phase I study of the bispecific antibody MDX-H210 in combination with granulocyte colony-stimulating factor (G-CSF) was performed in stage IV breast carcinoma patients, overexpressing HER-2\\/neu. MDX-H210, constructed by crosslinking antigen binding fragments (F(ab?) fragments) of monoclonal antibody (mAb) H22 to Fc gamma receptor I (Fc?RI), and mAb 520C9 to HER-2\\/neu, respectively, mediates the lysis of tumour cells in vitro,

R Repp; H H van Ojik; T Valerius; G Groenewegen; G Wieland; C Oetzel; B Stockmeyer; W Becker; M Eisenhut; H Steininger; Y M Deo; G H Blijham; J R Kalden; J G J van de Winkel; M Gramatzki

2003-01-01

81

Production of Monoclonal Antibody against Human Nestin  

PubMed Central

We have employed a peptide-based antibody generation protocol for producing antibody against human nestin. Using a 12-mer synthetic peptide from repetitive region of human nestin protein devoid of any N- or O-glyco-sylation sequences, we generated a mouse monoclonal antibody capable of recognizing human, mouse, bovine, and rat nestin. A wide variety of nestin proteins ranging from 140–250 kDa was detected by this antibody. This antibody is highly specific and functional in applications such as ELISA, flow cytometry, immunocytochemistry, and Western blot assays. PMID:23407796

Hadavi, Reza; Zarnani, Amir Hassan; Ahmadvand, Negah; Mahmoudi, Ahmad Reza; Bayat, Ali Ahmad; Mahmoudian, Jafar; Sadeghi, Mohammad-Reza; Soltanghoraee, Haleh; Akhondi, Mohammad Mehdi; Tarahomi, Majid; Jeddi-Tehrani, Mahmood; Rabbani, Hodjattallah

2010-01-01

82

An Alternative Chemical Redox Method for the Production of Bispecific Antibodies: Implication in Rapid Detection of Food Borne Pathogens  

PubMed Central

Bi-functional antibodies with the ability to bind two unrelated epitopes have remarkable potential in diagnostic and bio-sensing applications. In the present study, bispecific antibodies that recognize human red blood cell (RBC) and the food borne pathogen Listeria monocytogenes (L. monocytogenes) were engineered. The procedure involves initial reduction of a mixture of anti-RBC and anti-Listeria antibodies followed by gradual re-oxidation of the reduced disulphides. This facilitates association of the separated antibody chains and formation of hybrid immunoglobulins with affinity for the L. monocytogenes and human RBC. The bispecific antibodies caused the agglutination of the RBCs only in the presence of L. monocytogenes cells. The agglutination process necessitated the specific presence of L. monocytogenes and the red colored clumps formed were readily visible with naked eyes. The RBC agglutination assay described here provides a remarkably simple approach for the rapid and highly specific screening of various pathogens in their biological niches. PMID:24637674

Owais, Mohammad; Kazmi, Shadab; Tufail, Saba; Zubair, Swaleha

2014-01-01

83

Heterodimeric bispecific single-chain variable-fragment antibodies against EpCAM and CD16 induce effective antibody-dependent cellular cytotoxicity against human carcinoma cells.  

PubMed

A heterodimeric bispecific biological recombinant drug was synthesized by splicing DNA fragments from two fully humanized single-chain variable-fragment (scFV) antibody fragments forming a novel drug simultaneously recognizing the CD16 natural killer (NK) cell marker and the cancer marker epithelial cell adhesion molecule (EpCAM). The drug precipitously enhanced the killing of human carcinomas of the prostate, breast, colon, head, and neck even at very low effector:target ratios. The drug EpCAM16 rendered even nonactivated NK cell-proficient killers and activated them to kill via degranulation and cytokine production. Studies show that bispecific antibodies can be used to induce proficient killing of the carcinoma targets that ordinarily are resistant to NK-mediated killing. Apparently, the innate immune system can be effectively recruited to kill cancer cells using the bispecific antibody platform and EpCAM targeting. PMID:23611188

Vallera, Daniel A; Zhang, Bin; Gleason, Michelle K; Oh, Seunguk; Weiner, Louis M; Kaufman, Dan S; McCullar, Valarie; Miller, Jeffrey S; Verneris, Michael R

2013-05-01

84

Heterodimeric Bispecific Single-Chain Variable-Fragment Antibodies Against EpCAM and CD16 Induce Effective Antibody-Dependent Cellular Cytotoxicity Against Human Carcinoma Cells  

PubMed Central

Abstract A heterodimeric bispecific biological recombinant drug was synthesized by splicing DNA fragments from two fully humanized single-chain variable-fragment (scFV) antibody fragments forming a novel drug simultaneously recognizing the CD16 natural killer (NK) cell marker and the cancer marker epithelial cell adhesion molecule (EpCAM). The drug precipitously enhanced the killing of human carcinomas of the prostate, breast, colon, head, and neck even at very low effector:target ratios. The drug EpCAM16 rendered even nonactivated NK cell-proficient killers and activated them to kill via degranulation and cytokine production. Studies show that bispecific antibodies can be used to induce proficient killing of the carcinoma targets that ordinarily are resistant to NK-mediated killing. Apparently, the innate immune system can be effectively recruited to kill cancer cells using the bispecific antibody platform and EpCAM targeting. PMID:23611188

Zhang, Bin; Gleason, Michelle K.; Oh, Seunguk; Weiner, Louis M.; Kaufman, Dan S.; McCullar, Valarie; Miller, Jeffrey S.; Verneris, Michael R.

2013-01-01

85

Monoclonal antibody expression in mammalian cells.  

PubMed

In the past two decades, the production levels for monoclonal antibodies in mammalian expression systems have improved dramatically. Single cell productivity for monoclonal antibodies has increased 20-50 fold due to the improvements in expression hosts, expression vectors, cell culture media, and production processes. However, most of these improvements are proprietary to large pharmaceutical/biotech companies and involve large steel-tank bioreactors. Therefore, these processes are difficult for small companies and academic labs to reproduce. Transient expression in mammalian cells has recently been used very widely for monoclonal antibody expression. Cell line and expression vector engineering increased expression levels to several hundred milligrams per liter. The availability of highly effective transfection reagents and disposable bioreactors make the transient expression process an efficient and cost-effective way to make recombinant antibodies in large quantity. Here, we describe the protocols for small- to mid-scale transient expression of monoclonal antibodies in shake-flasks and for large-scale production in WAVE bioreactors. PMID:22907362

Zhang, Richard Yi; Shen, Wenyan David

2012-01-01

86

Therapeutic potential of monovalent monoclonal antibodies  

NASA Astrophysics Data System (ADS)

One therapeutic use for monoclonal antibody technology1 is the elimination of categories of unwanted cells by virtue of their distinct cell surface antigens. The efficiency of cell destruction by complement lysis or opsonization depends on a number of factors such as antibody specificity and isotype as well as certain properties of the target antigen2. In some instances cells can escape destruction by redistributing and eventually losing the antigen-antibody complexes from their surface3,4. This process, known as antigenic modulation, generally depends on bivalent antibody binding. Starting from the observation that rabbit antisera can be made more effective at killing tumour cells if they are first rendered univalent by limited proteolysis, we have now prepared a number of monovalent rat monoclonal antibodies to human cell-surface antigens. We find that these antibodies are no longer able to bring about modulation of their target antigens and have an enhanced facility for lysis with human complement. These special properties should greatly increase the therapeutic potential of monoclonal antibodies.

Cobbold, S. P.; Waldmann, H.

1984-03-01

87

Improving biophysical properties of a bispecific antibody scaffold to aid developability: quality by molecular design.  

PubMed

While the concept of Quality-by-Design is addressed at the upstream and downstream process development stages, we questioned whether there are advantages to addressing the issues of biologics quality early in the design of the molecule based on fundamental biophysical characterization, and thereby reduce complexities in the product development stages. Although limited number of bispecific therapeutics are in clinic, these developments have been plagued with difficulty in producing materials of sufficient quality and quantity for both preclinical and clinical studies. The engineered heterodimeric Fc is an industry-wide favorite scaffold for the design of bispecific protein therapeutics because of its structural, and potentially pharmacokinetic, similarity to the natural antibody. Development of molecules based on this concept, however, is challenged by the presence of potential homodimer contamination and stability loss relative to the natural Fc. We engineered a heterodimeric Fc with high heterodimeric specificity that also retains natural Fc-like biophysical properties, and demonstrate here that use of engineered Fc domains that mirror the natural system translates into an efficient and robust upstream stable cell line selection process as a first step toward a more developable therapeutic. PMID:23924797

Von Kreudenstein, Thomas Spreter; Escobar-Carbrera, Eric; Lario, Paula I; D'Angelo, Igor; Brault, Karine; Kelly, John; Durocher, Yves; Baardsnes, Jason; Woods, R Jeremy; Xie, Michael Hongwei; Girod, Pierre-Alain; Suits, Michael D L; Boulanger, Martin J; Poon, David K Y; Ng, Gordon Y K; Dixit, Surjit B

2013-01-01

88

Improving biophysical properties of a bispecific antibody scaffold to aid developability  

PubMed Central

While the concept of Quality-by-Design is addressed at the upstream and downstream process development stages, we questioned whether there are advantages to addressing the issues of biologics quality early in the design of the molecule based on fundamental biophysical characterization, and thereby reduce complexities in the product development stages. Although limited number of bispecific therapeutics are in clinic, these developments have been plagued with difficulty in producing materials of sufficient quality and quantity for both preclinical and clinical studies. The engineered heterodimeric Fc is an industry-wide favorite scaffold for the design of bispecific protein therapeutics because of its structural, and potentially pharmacokinetic, similarity to the natural antibody. Development of molecules based on this concept, however, is challenged by the presence of potential homodimer contamination and stability loss relative to the natural Fc. We engineered a heterodimeric Fc with high heterodimeric specificity that also retains natural Fc-like biophysical properties, and demonstrate here that use of engineered Fc domains that mirror the natural system translates into an efficient and robust upstream stable cell line selection process as a first step toward a more developable therapeutic. PMID:23924797

Von Kreudenstein, Thomas Spreter; Escobar-Carbrera, Eric; Lario, Paula I.; D’Angelo, Igor; Brault, Karine; Kelly, John F; Durocher, Yves; Baardsnes, Jason; Woods, R. Jeremy; Xie, Michael Hongwei; Girod, Pierre-Alain; Suits, Michael D. L.; Boulanger, Martin J.; Poon, David K. Y.; Ng, Gordon Y.; Dixit, Surjit B.

2013-01-01

89

The use of bispecific antibodies in tumor cell and tumor vasculature directed immunotherapy  

Microsoft Academic Search

To overcome dose limiting toxicities and to increase efficacy of immunotherapy of cancer, a number of strategies are under development for selectively redirecting effector cells\\/molecules towards tumor cells. Many of these strategies exploit the specificity of tumor associated antigen recognition by monoclonal antibodies. Using either hybridoma fusion, chemical derivatization or molecular biology technology, antibodies with dual specificity can be constructed.

Grietje Molema; Bart Jan Kroesen; Wijnand Helfrich; Dirk K. F. Meijer

2000-01-01

90

Antibody dependent cellular phagocytosis (ADCP) and antibody dependent cellular cytotoxicity (ADCC) of breast cancer cells mediated by bispecific antibody, MDX-210  

Microsoft Academic Search

Background: MDX-210 is a bispecific antibody (BsAb) with specificity for both the proto-oncogene product of HER-2\\/neu (c-erbB-2) and Fc?RI (CD64). HER-2\\/neu is overexpressed in malignant tissue of approximately 30% of patients with breast cancer, and Fc?RI is expressed on human monocytes, macrophages, and IFN-? activated granulocytes. We investigated phagocytosis and cytolysis of cultured human breast cancer cells by human monocyte-derived

Michiaki Watanabe; Paul K. Wallace; Tibor Keler; Yashwant M. Deo; Charuwan Akewanlop; Daniel F. Hayes

1999-01-01

91

Magic Bullets and Monoclonals: An Antibody Tale  

NSDL National Science Digital Library

FASEB Breakthroughs in Bioscience article. This most recent article describes the century of fundamental immunology research that led to todayĂÂs cutting edge monoclonal antibody therapies, used to treat millions of patients for several types of cancer, autoimmune and inflammatory disorders, and infectious disease.

Margie Patlak (Federation of American Societies for Experimental Biology Office of Public Affairs)

2010-07-12

92

Structure and specificity of lamprey monoclonal antibodies  

E-print Network

Structure and specificity of lamprey monoclonal antibodies Brantley R. Herrin*§ , Matthew N. Alder, December 10, 2007 (sent for review November 26, 2007) Adaptive immunity in jawless vertebrates (lamprey in both lamprey and hagfish (1­3). The germline VLR genes are incomplete in that they have coding regions

Ronquist, Fredrik

93

Monoclonal antibodies against chicken interleukin-6  

Technology Transfer Automated Retrieval System (TEKTRAN)

Monoclonal antibodies (mAb) were produced against a recombinant (r) chicken interleukin-6 (IL-6). Eight mAbs that were produced were tested for isotype; ability to inhibit recombinant forms of chicken (ch), human (h) and murine (m) IL-6; and recognition of rchIL-6 by Western immunoblotting. The mA...

94

Potent inhibition of local and disseminated tumor growth in immunocompetent mouse models by a bispecific antibody construct specific for Murine CD3  

Microsoft Academic Search

Bispecific single-chain antibody constructs specific for human CD3 have been extensively studied for antitumor activity in\\u000a human xenograft models using severe combined immunodeficient mice supplemented with human T cells. High efficacy at low effector-to-target\\u000a ratios, independence of T cell costimuli and a potent activation of previously unstimulated polyclonal T cells were identified\\u000a as hallmarks of this class of bispecific antibodies.

Bernd Schlereth; Petra Kleindienst; Iduna Fichtner; Grit Lorenczewski; Klaus Brischwein; Sandra Lippold; Antonio da Silva; Mathias Locher; Roman Kischel; Ralf Lutterbüse; Peter Kufer; Patrick A. Baeuerle

2006-01-01

95

CD3×CD19 bispecific antibodies and CD28 costimulation for locoregional treatment of low-malignancy non-Hodgkin’s lymphoma  

Microsoft Academic Search

In advance of using bispecific antibodies for the treatment of B cell lymphoma in humans, we analysed CD3×CD19 bispecific\\u000a antibodies for their capacity to induce T cell activiation in cell suspensions from follicular lymphoma lymph nodes. Here,\\u000a we demostrate that the lack of costimulatory molecules, such as members of the B7 family, on the tumour cells resulted in\\u000a insufficient activation

Oliver Manzkem; Sandra Titzer; Hans Tesch; Volker Diehl; Heribert Bohlen

1997-01-01

96

Monoclonal antibodies against the aster yellows agent.  

PubMed

Hybridoma clones secreting specific monoclonal antibodies against the aster yellows agent, a mycoplasma-like organism, were produced by using partially purified salivary gland preparations from infected leafhopper vectors as the immunogen. After 3947 hybridomas from 20 independent fusions were screened for specific antibody against the aster yellows agent, two table clones were obtained. With these monoclonal antibodies the aster yellows agent in diseased lettuce, periwinkles, and inoculative insects was specifically identified by enzyme-linked immunosorbent assay. The aster yellows agent was serologically differentiated from the mycoplasma-like organisms associated with ash yellows, loofah witches'-broom, paulownia witches'-broom, sweet potato witches'-broom, peanut rosette, maize bushy stunt, and elm phloem necrosis. PMID:17757867

Lin, C P; An Chen, T

1985-03-01

97

Systemic administration of a bispecific antibody targeting EGFRvIII successfully treats intracerebral glioma  

PubMed Central

Bispecific antibodies (bscAbs), particularly those of the bispecific T-cell engager (BiTE) subclass, have been shown to effectively redirect T cells against cancer. Previous efforts to target antigens expressed in both tumors and normal tissues have produced significant toxicity, however. Moreover, like other large molecules, bscAbs may be restricted from entry into the “immunologically privileged” CNS. A tumor-specific mutation of the epidermal growth factor receptor, EGFRvIII, is a constitutively activated tyrosine kinase not found in normal tissues but frequently expressed in glioblastomas and many other neoplasms. Because it is localized solely to tumor tissue, EGFRvIII presents an ideal target for immunotherapy. Here we report the preclinical evaluation of an EGFRvIII-targeted BiTE, bscEGFRvIIIxCD3. Our results show that bscEGFRvIIIxCD3 activates T cells to mediate potent and antigen-specific lysis of EGFRvIII-expressing gliomas in vitro (P < 0.001) at exceedingly low concentrations (10 ng/mL) and effector-to-target ratios (2.5:1). Treatment with i.v. bscEGFRvIIIxCD3 yielded extended survival in mice with well-established intracerebral tumors (P < 0.05) and achieved durable complete cure at rates up to 75%. Antitumor efficacy was significantly abrogated on blockade of EGFRvIII binding, demonstrating the need for target antigen specificity both in vitro and in vivo. These results demonstrate that BiTEs can be used to elicit functional antitumor immunity in the CNS, and that peptide blockade of BiTE-mediated activity may greatly enhance the safety profile for antibody-redirected T-cell therapies. Finally, bscEGFRvIIIxCD3 represents a unique advancement in BiTE technology given its exquisite tumor specificity, which enables precise elimination of cancer without the risk of autoimmune toxicity. PMID:23248284

Choi, Bryan D.; Kuan, Chien-Tsun; Cai, Mingqing; Archer, Gary E.; Mitchell, Duane A.; Gedeon, Patrick C.; Sanchez-Perez, Luis; Pastan, Ira; Bigner, Darell D.; Sampson, John H.

2013-01-01

98

Serovar determination of Chlamydia trachomatis isolates by using type-specific monoclonal antibodies.  

PubMed Central

A panel of 15 monoclonal antibodies was prepared that could distinguish among the 15 serovars of Chlamydia trachomatis. Twelve of these antibodies were specific for a single serovar (A, B, C, D, E, F, G, H, I, K, L1, and L2) and three were specific for two serovars (B/Ba, C/J, and C/L3). Ten of the serovar-specific and two of the bispecific antibodies were shown by immunoblotting to recognize epitopes on the major outer membrane protein. These data provide evidence that such epitopes are closely correlated with and may be partly responsible for the antigenic variations detected by microimmunofluorescence that distinguish the currently recognized serovars. When used in a radioimmunoassay, these antibodies correctly identified the serovar of 17 strains that had been serotyped by the microimmunofluorescence test. In addition, we found that the chlamydial antigen derived from 1.0 cm2 of an infected HeLa cell monolayer was sufficient to allow serotyping with these antibodies. Thus, these monoclonal antibodies may provide a rapid and reliable alternative to mouse immunization and microimmunofluorescence for serotyping of clinical isolates. Images PMID:2422202

Newhall, W J; Terho, P; Wilde, C E; Batteiger, B E; Jones, R B

1986-01-01

99

Visualization of effective tumor targeting by CD8+ natural killer T cells redirected with bispecific antibody F(ab')(2)HER2xCD3.  

PubMed

HER2 is an attractive immunotherapeutic target for neoplastic disease because this cell surface molecule is overexpressed on a large fraction of malignant tumor cells. To directly assess therapeutic responses to targeted therapy by noninvasive in vivo imaging in small animals, human HER2-expressing ovarian carcinoma cells were genetically modified with a firefly luciferase gene, and light emission was used for visualization of tumor growth and response to therapy. This imaging approach was able to demonstrate in real-time tumor regression in a HER2 xenograft mouse model by adoptive transfer of in vitro induced and expanded cytotoxic CD8+ natural killer T (NKT) cells retargeted with a humanized bispecific antibody F(ab')(2)HER2xCD3. Immunotherapy with effector cells alone or a humanized monoclonal antibody anti-p185(HER2) (4D5-8) resulted in significant but slower reduction in tumor burden. Long-term survival of tumor xenografts correlated inversely with visible residual tumor burden. In vitro, F(ab')(2)HER2xCD3 substantially augmented cytotoxic activity of CD8+ NKT cells. By flow-sorting, CD8+ NKT cells coexpressing CD56 were found to have the highest redirected killing ability. Treatment with concanamycin A or EGTA abrogated CD8+ NKT cytotoxicity indicating that perforin is a major pathway of tumor cell lysis. In contrast, when CD8+ NKT cell were cross-linked with F(ab')(2)HER2xCD3 neither the immunosuppressants cyclosporine A and FK506, nor the increase of intracellular cyclic AMP by dibutyryl cyclic AMP were able to inhibit cytotoxicity demonstrating that signaling via the CD3 antigen changes the biological activity of non-MHC-restricted effector cells. These studies have demonstrated that CD8+ NKT cells can be successfully redirected to tumor cells using bispecific antibodies and offer a promising strategy for adoptive immunotherapy of neoplastic diseases. PMID:12384539

Scheffold, Christian; Kornacker, Martin; Scheffold, Yolanda C; Contag, Christopher H; Negrin, Robert S

2002-10-15

100

Enhanced killing of primary ovarian cancer by retargeting autologous cytokine-induced killer cells with bispecific antibodies: a preclinical study.  

PubMed

Cytokine-induced killer (CIK) cells are ex vivo activated and expanded CD8+ natural killer T cells that have been shown to have antitumor activity. This is the first study exploring cell killing of primary ovarian carcinoma cells with and without bispecific antibodies. Primary cancer cells and autologous CIK cells were collected from women with epithelial ovarian cancer. Bispecific antibodies against cancer antigen-125 (BSAbxCA125) and Her2 (BSAbxHer2) were developed using chemical heteroconjugation. On fluorescence-activated cell sorting analysis, the expansion of CIK cells resulted in a significant increase of CD3+CD8+ and CD3+CD56+ T cells. With enhancement by bispecific antibodies, the mean percent lysis in a 51Cr release assay of fresh ovarian cancer cells exposed to autologous CIK cells increased from 21.7 +/- 0.3% to 89.4 +/- 2.1% at an E:T ratio of 100:1 (P < 0.001). Anti-NKG2D antibodies attenuated the CIK activity by 56.8% on primary cells (P < 0.001). In a xenograft severe combined immunodeficient mouse model, real-time tumor regression and progression was visualized using a noninvasive in vivo bioluminescence imaging system. Four hours after CIK cell injection, we were able to visualize CD8+NKG2D+ CIK cells infiltrating Her2-expressing cancer cells on fluorescence microscopy. Mice that underwent adoptive transfer of CIK cells redirected with BSAbxCA125 and BSAbxHer2 had significant reduction in tumor burden (P < 0.001 and P < 0.001) and improvement in survival (P = 0.05 and P = 0.006) versus those treated with CIK cells alone. Bispecific antibodies significantly enhanced the cytotoxicity of CIK cells in primary ovarian cancer cells and in our in vivo mouse model. The mechanism of cytolysis seems to be mediated in part by the NKG2D receptor. PMID:16551871

Chan, John K; Hamilton, Chad A; Cheung, Michael K; Karimi, Mobin; Baker, Jeanette; Gall, Jonathan M; Schulz, Stephan; Thorne, Steve H; Teng, Nelson N; Contag, Christopher H; Lum, Lawrence G; Negrin, Robert S

2006-03-15

101

SPECT assay of radiolabeled monoclonal antibodies  

SciTech Connect

The accurate determination of the biodistribution of radiolabeled monoclonal antibodies (MoAbs) is important for calculation of dosimetry and evaluation of pharmacokinetic variables such as antibody dose and route of administration. The hypothesis of this application is that the biodistribution of radiolabeled monoclonal antibodies (MoAbs) can be quantitatively determined using single photon emission computed tomography (SPECT). The major thrusts during the third year include the continued development and evaluation of improved 3D SPECT acquisition and reconstruction approaches to improve quantitative imaging of radiolabeled monoclonal antibodies (MoAbs), and the implementation and evaluation of algorithms to register serial SPECT image data sets, or to register 3D SPECT images with 3D image data sets acquired from positron emission tomography (PEI) and magnetic resonance images (MRI). The research has involved the investigation of statistical models and iterative reconstruction algorithms that accurately account for the physical characteristics of the SPECT acquisition system. It is our belief that SPECT quantification can be improved by accurately modeling the physical processes such as attenuation, scatter, geometric collimator response, and other factors that affect the measured projection data.

Jaszczak, R.J.

1992-02-01

102

Preclinical studies with Fc(gamma)R bispecific antibodies and granulocyte colony-stimulating factor-primed neutrophils as effector cells against HER-2/neu overexpressing breast cancer.  

PubMed

Immunotherapies directed to the proto-oncogene product HER-2/neu, which is overexpressed on a subset of breast and other carcinomas, currently receive considerable attention. We have investigated cell-mediated effector mechanisms of HER-2/neu antibodies against breast cancer cell lines. Compared to unfractionated control blood, whole blood from patients during granulocyte colony-stimulating factor (G-CSF) treatment exhibits significantly enhanced lysis (P < 0.001) of SK-BR-3 cells in the presence of HER-2/neu antibody 520C9. The extent of tumor cell killing correlated positively (r = 0.74) to polymorphonuclear neutrophil (PMN) blood counts. Fractionation of whole blood into plasma, mononuclear cells, and PMNs showed major killing capacity to reside in the granulocyte fraction. PMNs were efficiently cytolytic with a panel of HER-2/neu antibodies and against various breast cancer cell lines. Experiments with blocking antibodies to Fc(gamma)R documented Fc(gamma)RII (CD32) as the major trigger molecule for monoclonal antibody 502C9-mediated cytotoxicity. Killing via 520C9 was significantly influenced by an allotypic polymorphism of Fc(gamma)RIIa, the CD32 molecule expressed on PMNs. In reverse antibody-dependent cell-mediated cytotoxicity experiments with a panel of HER-2/neu-directed bispecific antibodies, Fc(gamma)RIII (CD16) proved to be an efficient trigger molecule in blood from healthy volunteers. During G-CSF treatment, however, Fc(gamma)RI (CD64)-expressed on monocytes and G-CSF primed, but not on healthy donor PMNs-became the predominant cytotoxic trigger molecule. Thus, G-CSF application increased effector cell numbers for HER-2/neu-directed immunotherapy, and G-CSF primed PMNs proved particularly effective with a [HER-2/neu x Fc(gamma)RI] bispecific antibody. These findings support clinical trials with HER-2/neu-directed antibodies in combination with G-CSF in breast cancer patients overexpressing HER-2/neu. PMID:9044847

Stockmeyer, B; Valerius, T; Repp, R; Heijnen, I A; Bühring, H J; Deo, Y M; Kalden, J R; Gramatzki, M; van de Winkel, J G

1997-02-15

103

Anti-CD16/CD30 bispecific antibody treatment for Hodgkin's disease: role of infusion schedule and costimulation with cytokines.  

PubMed

The natural killer cell-activating anti-CD16/CD30 bispecific monoclonal antibody (BiMAb) had shown efficacy in a Phase I/II trial of refractory Hodgkin's disease (HD). To gain additional information on clinical efficacy and to investigate the effects of different application schedules and the concomitant application of cytokines, we performed a second randomized pilot trial using this BiMAb in patients with refractory HD. Patients received 4 x 25 mg HRS-3/A9 either as a continuous infusion for 4 days or as a 1-h infusion every other day. In case of an objective response, retreatment was attempted after 4 weeks; in case of stable disease (SD), a second course was given after prestimulation with interleukin 2 and followed by granulocyte macrophage colony-stimulating factor s.c. A total of 16 heavily pretreated patients received one to four BiMAb courses. Overall, we observed one complete remission and three partial remissions lasting 5-9 months (three of four of these responses occurred after continuous BiMAb infusion) and four cases of SD for 3 to >6 months. Interleukin 2 pretreatment before the second BiMAb course resulted in a significant increase of circulating natural killer cells in all five patients treated. This coincided with the conversion of two cases of SD into one complete remission and one partial remission. HRS-3/A9-related side effects consisted of mild fever in only six patients. In summary, this second trial confirmed the antitumor efficacy of this BiMAb against HD and the minor toxicity of this BiMAb. Coadministration of cytokines might contribute to an augmented antitumor activity, and additional clinical trials are warranted to optimize this novel treatment modality. PMID:11448899

Hartmann, F; Renner, C; Jung, W; da Costa, L; Tembrink, S; Held, G; Sek, A; König, J; Bauer, S; Kloft, M; Pfreundschuh, M

2001-07-01

104

Innovative Monoclonal Antibody Therapies in Multiple Sclerosis  

PubMed Central

The recent years have witnessed great efforts in establishing new therapeutic options for multiple sclerosis (MS), especially for relapsing–remitting disease courses. In particular, the application of monoclonal antibodies provide innovative approaches allowing for blocking or depleting specific molecular targets, which are of interest in the pathogenesis of MS. While natalizumab received approval by the US Food and Drug Administration and the European Medicines Agency in 2006 as the first monoclonal antibody in MS therapy, rituximab, alemtuzumab, and daclizumab were successfully tested for relapsing-remitting MS in small cohorts in the meantime. Here, we review the data available from these recent phase II trials and at the same time critically discuss possible pitfalls which may be relevant for clinical practice. The results of these studies may not only broaden our therapeutic options in the near future, but also provide new insights into disease pathogenesis. PMID:21180564

Kieseier, Bernd C.

2008-01-01

105

Recent developments in monoclonal antibody radiolabeling techniques  

SciTech Connect

Monoclonal antibodies (MAbs) have shown the potential to serve as selective carriers of radionuclides to specific in vivo antigens. Accordingly, there has been an intense surge of research activity in an effort to develop and evaluate MAb-based radiopharmaceuticals for tumor imaging (radioimmunoscintigraphy) and therapy (radioimmunotherapy), as well as for diagnosing nonmalignant diseases. A number of problems have recently been identified, related to the MAbs themselves and to radiolabeling techniques, that comprise both the selectivity and the specificity of the in vivo distribution of radiolabeled MAbs. This paper will address some of these issues and primarily discuss recent developments in the techniques for radiolabeling monoclonal antibodies that may help resolve problems related to the poor in vivo stability of the radiolabel and may thus produce improved biodistribution. Even though many issues are identical with therapeutic radionuclides, the discussion will focus mainly on radioimmunoscintigraphic labels. 78 refs., 6 tabs.

Srivastava, S.C.; Mease, R.C.

1989-01-01

106

T cell activation and retargeting using staphylococcal enterotoxin B and bispecific antibody: An effective in vivo antitumor strategy  

Microsoft Academic Search

The aim of this work was to test for cure and immunity in a micrometastatic tumor model using in vivo T cell activation with\\u000a staphylococcal enterotoxin B (SEB) and retargeting with antitumor×anti-CD3 F(ab?)2 bispecific antibodies (bsAb). All studies were performed in C3H\\/HeN mice using syngeneic tumor cell lines. For survival studies,\\u000a mice were injected intravenously on day 0 with CL62

Lewis E. Porterm; Heidi Nelson; I. Ethem Gecim; David C. Rice; Claude Thibault; Andrei I. Chapoval

1997-01-01

107

Bispecific Antibody-Mediated Immunotherapy of the BCL, Lymphoma: Increased Efficacy With Multiple Injections and CD28Induced Costimulation  

Microsoft Academic Search

The BCL, lymphoma in Balblc mice can be successfully treated with bispecific (antLCD3 x anti-idiotype) antibodies (BSABs). In these experiments, animals were injected intra- peritoneally (IP) with 5 x 10s tumor cells (day 0) and treated with one single intravenous (IV) injection of 5 pg BSAB (day 9). Because cross-linking of the CD3 complex is not in itself sufficient to

Christian Demanet; Jan Brissinck; Jan De Jonge; Kris Thielemans

1996-01-01

108

Targeted adeno-associated virus vector transduction of nonpermissive cells mediated by a bispecific F(ab'?)2 antibody  

Microsoft Academic Search

We have developed a system for the targeted delivery of adeno–associated virus (AAV) vectors. Targeting is achieved via a bispecific F(ab´)2 antibody that mediates a novel interaction between the AAV vector and a specific cell surface receptor expressed on human megakaryocytes. Targeted AAV vectors were able to transduce megakaryocyte cell lines, DAMI and MO7e, which were nonpermissive for normal AAV

Jurgen Kleinschmidt; Richard C. Boucher; R. Jude Samulski; Jeffrey S. Bartlett

1999-01-01

109

Monoclonal antibody targets, kills leukemia cells  

Cancer.gov

Researchers at the University of California, San Diego Moores Cancer Center have identified a humanized monoclonal antibody that targets and directly kills chronic lymphocytic leukemia (CLL) cells. The findings, published in the online Early Edition of the Proceedings of the National Academy of Sciences on March 25, 2013 represent a potential new therapy for treating at least some patients with CLL, the most common type of blood cancer in the United States.

110

Preclinical Safety Evaluation of Monoclonal Antibodies  

Microsoft Academic Search

Monoclonal antibodies (mAbs) are a well-established product class of biotechnology-derived pharmaceuticals for treating multiple\\u000a diseases. A growing number of mAbs are being tested in clinical trials worldwide. Many of the second generation mAbs entering\\u000a the clinic today are highly engineered, produced from recombinant cell lines, and present new safety challenges for regulators\\u000a and industry scientists responsible for their safety evaluation.

C. M. Lynch; I. S. Grewal

111

Mechanisms of G-CSF- or GM-CSF-stimulated tumor cell killing by Fc receptor-directed bispecific antibodies.  

PubMed

Studies with gene-modified mice have recently reinforced the importance of Fc receptor-mediated effector mechanisms for the therapeutic efficacy of rituxan and herceptin - two clinically approved antibodies for the treatment of tumor patients. We investigated Fc receptor-dependent tumor cell killing by mononuclear and granulocytic effector cells - comparing human IgG1 antibodies against CD20 or HER-2/neu with their respective FcgammaRI (CD64)-, FcgammaRIII (CD16)-, or FcalphaRI (CD89)-directed bispecific derivatives. With blood from healthy donors as effector source, human IgG1 and FcgammaRIII (CD16)-directed bispecific antibodies proved most effective in recruiting mononuclear effector cells, whereas tumor cell killing by granulocytes was most potently triggered by FcalphaRI-directed bispecific constructs. Granulocyte-mediated tumor cell lysis was significantly enhanced when blood from G-CSF- or GM-CSF-treated patients was investigated. Interestingly, however, both myeloid growth factors improved effector cell recruitment by different mechanisms, which were furthermore dependent on the tumor target antigen, and on the selected cytotoxic Fc receptor. PMID:11223072

Stockmeyer, B; Elsässer, D; Dechant, M; Repp, R; Gramatzki, M; Glennie, M J; van de Winkel, J G; Valerius, T

2001-02-01

112

Microencapsulation of therapeutic bispecific antibodies producing cells: immunotherapeutic organoids for cancer management.  

PubMed

Abstract Regardless of the important therapeutic advances developed over the last years for the management of cancer, the fact is that many patients still suffer from a tremendous reduction on their quality of life due to lack of complete selectivity of conventionally administered chemotherapeutic drugs. In the search of more efficacious tumor-targeted therapies, the use of bispecific antibodies (bsAbs) capable of simultaneous binding to tumor-associated antigens and to an activating receptor, such as CD3, has emerged as a promising approach. With the intention to complementing and improving this cancer immunotherapy, human HEK-293 cells have been genetically modified ex vivo to secrete a recombinant anti-CEA (carcinoembryonic antigen)?×?anti-CD3 bsAb. After encapsulation in alginate-poly-l-lysine microcapsules, bsAb-secreting HEK-293 cells were monitorized for several weeks. This system has proved to be feasible for the maintenance of cell growth and recombinant antibody production giving proof-of-concept of its use as immunotherapeutic organoids in cancer treatment. PMID:25338126

Saenz Del Burgo, Laura; Compte, Marta; Aceves, Mónica; Hernández, Rosa María; Sanz, Laura; Álvarez-Vallina, Luis; Pedraz, Jose Luis

2015-02-01

113

Monoclonal antibodies and method for detecting dioxins and dibenzofurans  

DOEpatents

Compositions of matter are described which include five monoclonal antibodies that react with dioxins and dibenzofurans, and the five hybridomas that produce these monoclonal antibodies. In addition, a method for the use of these antibodies in a sensitive immunoassay for dioxins and dibenzofurans is given, which permits detection of these pollutants in samples at concentrations in the range of a few parts per billion.

Vanderlaan, Martin (San Ramon, CA); Stanker, Larry H. (Livermore, CA); Watkins, Bruce E. (Livermore, CA); Bailey, Nina R. (Berkley, CA)

1989-01-01

114

Monoclonal antibodies specific for sickle cell hemoglobin  

SciTech Connect

Two mouse hybridoma cell lines were isolated which produce monoclonal antibodies that bind hemoglobin S. The mice were immunized with peptide-protein conjugates to stimulate a response to the amino terminal peptide of the beta chain of hemoglobin S, where the single amino acid difference between A and S occurs. Immunocharacterization of the antibodies shows that they bind specifically to the immunogen peptide and to hemoglobin S. The specificity for S is high enough that one AS cell in a mixture with a million AA cells is labeled by antibody, and such cells can be analyzed by flow cytometry. Immunoblotting of electrophoretic gels allows definitive identification of hemoglobin S as compared with other hemoglobins with similar electrophoretic mobility. 12 references, 4 figures.

Jensen, R.H.; Vanderlaan, M.; Grabske, R.J.; Branscomb, E.W.; Bigbee, W.L.; Stanker, L.H.

1985-01-01

115

Leukocyte analysis using monoclonal antibodies in human glomerulonephritis  

Microsoft Academic Search

Leukocyte analysis using monoclonal antibodies in human glomerulonephritis. The leukocyte subpopulations were analyzed within both the glomeruli and the interstitium in renal biopsies from 145 patients with various forms of glomerulonephritis. Cells were identified by monoclonal antibodies to leukocyte cell–surface antigens and immunoper-oxidase labelling. Leukocytes, as defined by a monoclonal antibody to the leukocyte common antigen (PHM1), were present in

David H Hooke; David C Gee; Robert C Atkins

1987-01-01

116

Retargeting of human lymphocytes against human ovarian carcinoma cells by bispecific antibodies: from laboratory to clinic  

Microsoft Academic Search

Summary  We have selected a monoclonal antibody (MOv18) reactive with ovarian carcinoma, which exhibits a restricted tumor specificity,\\u000a a high affinity constant and which recognizes a 38-kDa glycoprotein homogeneously expressed on the cell surface of 90% of\\u000a human ovarian carcinomas. Localization studies with radiolabelled MOv18 showed that MOv18 could specifically reach ovarian\\u000a carcinoma cells growing in the peritoneal cavity of nu\\/nu

Delia Mezzanzanica; Silvana Canevari; Maria I. Colnaghi

1992-01-01

117

Monoclonal antibody to growth hormone receptors  

SciTech Connect

Using hybridoma technology, monoclonal antibodies to growth hormone receptors can be prepared in large quantities with only a few micrograms of purified antigen by in vitro immunization or by immunization of larger quantities of less pure material. The discussion centers on areas most pertinent to receptors such as receptor preparation, immunization procedure, fusion method, screening assay, and identification of the immunoglobulin class. The specificity of the antibody for growth hormone receptor was examined by testing the ability of the ascitic fluid to inhibit binding of (/sup 125/I) growth hormone to the prolactin receptors on rabbit mammary gland membranes and the inhibition of /sup 125/I-labelled rat growth hormone binding to rabbit liver membrane.

Simpson, J.S.A.; Friesen, H.G.

1985-01-01

118

Bispecific anti-CD3 x anti-HER2 antibody mediates T cell cytolytic activity to HER2-positive colorectal cancer in vitro and in vivo.  

PubMed

Targeting HER2 overexpressed breast cancer cells with anti?HER2 monoclonal antibodies inhibits tumor growth. Here we investigated whether HER2 can serve as a target for T cell-mediated immunotherapy of human colorectal carcinoma. Specific cytolytic activity of activated T cells (ATCs) armed with anti?CD3 x anti?HER2 bispecific antibody (HER2Bi-Ab) against HER2+ tumor cells was evaluated by bioluminescent signal generated by luciferase reporter on tumor cells in vitro and in vivo. In contrast to unarmed ATCs, increased cytotoxic activity of HER2Bi-armed ATCs against HER2+ tumor cells was observed. Moreover, HER2Bi-armed ATCs expressed higher level of activation marker CD69 and secreted significantly higher levels of IFN-? than the unarmed ATC counterpart. In addition, compared with anti?HER2 mAb (Herceptin®) or unarmed ATC, HER2Bi-armed ATCs showed significant suppression against colorectal carcinoma cells. In colorectal tumor cell xenograft mice, infusion of HER2Bi-armed ATCs successfully inhibited the growth of Colo205-luc cells. The HER2Bi-armed ATCs with anti-tumor effects may provide a promising immunotherapy for colorectal carcinoma in the future. PMID:25242665

Han, Huamin; Ma, Juan; Zhang, Keming; Li, Wei; Liu, Changzhen; Zhang, Yu; Zhang, Ganlin; Ma, Pan; Wang, Lei; Zhang, Ge; Tao, Hua; Gao, Bin

2014-12-01

119

The bi-specific CD3 x NCAM antibody: a model to preactivate T cells prior to tumour cell lysis.  

PubMed

To target the neural cell adhesion molecule (NCAM, CD56) on neuroblastoma by T cell-based immunotherapy we have generated a bi-specific CD3 x NCAM antibody (OE-1). This antibody can be used to redirect T cells to NCAM+ cells. Expectedly, the antibody binds specifically to NCAM+ neuroblastoma cells and CD3+ T cells. OE-1 induces T cell activation, expansion and effector function in peripheral blood mononuclear cell (PBMC)-derived CD4+ and CD8+ T cells. T cell activation was shown to depend on the presence of normal natural killer (NK) cells in the culture. Interestingly, while PBMC- derived T cells were activated by OE-1, NK cells were almost completely depleted, suggesting that T cells activated by OE-1 deleted the NK cells. Activated CD4+ and CD8+ T cells differentiate into a larger CCR7+ central memory and a smaller CCR7- effector memory cell population. Most importantly, preactivated T cells were highly cytotoxic for neuroblastoma cells. In eight of 11 experiments tumour-directed cytotoxicity was enhanced when NK cells were present during preactivation with OE-1. These data strongly support a bi-phasic therapeutic concept of primarily stimulating T cells with the bi-specific antibody in the presence of normal NCAM+ cells to induce T cell activation, migratory capacity and finally tumour cell lysis. PMID:14616785

Jensen, M; Ernestus, K; Kemshead, J; Klehr, M; Von Bergwelt-Baildon, M S; Schinköthe, T; Schultze, J L; Berthold, F

2003-11-01

120

DETECTION OF ROTAVIRUS IN HUMAN STOOLS BY USING MONOCLONAL ANTIBODY  

EPA Science Inventory

A monoclonal antibody, 3F7, which reacts with the common rotavirus antigen on the sixth viral gene product was prepared. It was used in a direct monoclonal antibody radioimmunoassay (RIA) as a diagnostic reagent for detection in 3.5 hours of rotavirus in human pediatric stool spe...

121

Human proteasomes analysed with monoclonal antibodies.  

PubMed Central

The proteasome or multicatalytic endopeptidase from eukaryotic cells consists of at least 14 subunits that fall into two families, alpha and beta. Subunit-specific monoclonal antibodies against ten different subunits of human proteasomes have been produced, together with an antibody that reacts with a motif (prosbox 1), common to alpha-type subunits. Four of the subunit-specific antibodies were able to precipitate proteasomes. The subunit composition of HeLa-cell proteasomes precipitated with these four different antibodies were identical, as judged from two-dimensional electrophoresis. One of the four antibodies was used to obtain proteasomes from cell lines (HeLa, Daudi, IMR90 and BSC-1) and human tissues (placenta, kidney, and liver). Electrophoretic analysis of these proteasomes, combined with peptide mapping of some subunits, suggests that they all contain 14 types of subunits as their major constituents. However, one subunit was present in two isoelectric isoforms in all cells examined. Two other subunits occurred in two or three isoelectric isoforms in placenta, liver and kidney, but not in the cell cultures. Extracts of human cells (HeLa, IMR90, Daudi and erythrocytes) were analysed by non-denaturing electrophoresis and immunoblotting. All of the 11 subunits detected by antibodies were present in a pair of ATP-stabilized protein complexes, presumed to be the 26 S proteinase, and in a doublet of complexes which migrated more slowly than purified proteasomes. Besides being present in proteasomes, one subunit was also found to occur in the free state in cell extracts. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 PMID:7826336

Hendil, K B; Kristensen, P; Uerkvitz, W

1995-01-01

122

The immunodiagnosis of lung cancer with monoclonal antibodies.  

PubMed

Lung cancer is responsible for much suffering and death worldwide. The only hope for cure is therapy applied in an early phase, and all methods of diagnosis should be aimed at this goal. This paper reviews the development of the use of monoclonal antibodies in the diagnosis of lung cancer. Relevant data since the publication of the technology of producing monoclonal antibodies in 1975 to the present are summarized. The authors evaluate the progress of the immunodiagnosis of lung cancer by monoclonal antibodies from pleural effusion, bone marrow, sputum, bronchial lavage, and bronchial brush (immunocytochemistry). They collect recent data on the immunohistochemistry of biopsy materials and of removed tissues. They evaluate radioimmuno-imaging (radioimmuno-scintigraphy) and immuno-PET as in vivo macroscopic diagnostic methods of lung cancer by monoclonal antibodies as well as the help monoclonal antibodies provide in radioimmuno-guided surgery or immunoimage-guided, focally ablative therapy of this disease. PMID:16127376

Egri, Gábor; Takáts, Alajos

2005-09-01

123

Production of a monoclonal antibody specific for seminomas and dysgerminomas.  

PubMed Central

A monoclonal antibody (M2A, IgG2a) was produced against a cultured human ovarian epithelial adenocarcinoma cell line, HEY. Monoclonal antibody M2A reacted with a glycoprotein of molecular weight 40,000 on the surface of HEY cells. The affinity constant of the monoclonal antibody M2A for HEY cells was 10(9) M-1, and the number of binding sites on HEY cells was 2 X 10(4) per cell. The monoclonal antibody produced positive immunoperoxidase staining of fetal (but not adult) testis and of seminomas and dysgerminomas but did not stain various normal adult tissues or other gonadal or extragonadal tumors. Monoclonal antibody M2A may be useful for confirming a histological diagnosis of seminoma and dysgerminoma. Images PMID:3523489

Bailey, D; Baumal, R; Law, J; Sheldon, K; Kannampuzha, P; Stratis, M; Kahn, H; Marks, A

1986-01-01

124

Characterization and utilization of monoclonal antibodies reactive to Yersinia pseudotuberculosis.  

PubMed

The 3 murine monoclonal antibodies, Yps1, Yps2 and Yps3 reactive to Y. pseudotuberculosis can be stabilized and all were found to be of IgG type. Monoclonal antibody, Yps1, recognized a glycoprotein antigen of the organism with reactivity at the 55-75 kDa region, while Yps2 and Yps3 recognized protein antigens of Y. pseudotuberculosis 65 kDa and 26-28 kDa molecular weight regions, respectively. The specificity of monoclonal antibodies was tested using dot ELISA and Western blotting with whole cell organisms or whole cell sonicated soluble antigens of different Yersinia species, Salmonella typhi, Klebsiella pnemoniae, Streptococcus abortus-equi and Escherichia coli. Monoclonal antibody, Yps1 exhibited cross-reactivity with soluble antigens and whole cell preparations of Y. pestis. Yps2 cross-reacted to soluble antigens of all the tested bacteria. Reactivity of monoclonal antibody, Yps3 was restricted to Y. pseudotuberculosis and Y. pestis with soluble antigen preparations. No reaction was observed with Yps2 and Yps3 to whole cell organism preparations from tested bacteria including Y. pseudotuberculosis. The co-agglutination reagent prepared by sensitizing staphylococcal cells with Yps1 monoclonal antibody produced a positive agglutination with all the 4 Y. pseudotuberculosis isolates and the 3 Y. pestis strains tested. Sandwich dot ELISA using monospecific antisera as a capture antibody and a monoclonal antibody, and Yps3 as a revealing antibody had a high level of specificity in detecting Y. pseudotuberculosis antigens. PMID:15115097

Jain, Reena; Tuteja, Urmil; Batra, Harsh Vardhan

2003-12-01

125

A Fc?RIII-engaging bispecific antibody expands the range of HER2-expressing breast tumors eligible to antibody therapy  

PubMed Central

Trastuzumab is established as treatment of HER2high metastatic breast cancers but many limitations impair its efficacy. Here, we report the design of a Fab-like bispecific antibody (HER2bsFab) that displays a moderate affinity for HER2 and a unique, specific and high affinity for Fc?RIII. In vitro characterization showed that ADCC was the major mechanism of action of HER2bsFab as no significant HER2-driven effect was observed. HER2bsFab mediated ADCC at picomolar concentration against HER2high, HER2low as well as trastuzumab-refractive cell lines. In vivo HER2bsFab potently inhibited HER2high tumor growth by recruitment of mouse Fc?RIII and IV-positive resident effector cells and more importantly, exhibited a net superiority over trastuzumab at inhibiting HER2low tumor growth. Moreover, Fc?RIIIA-engagement by HER2bsFab was independent of V/F158 polymorphism and induced a stronger NK cells activation in response to target cell recognition. Thus, taking advantage of its epitope specificity and affinity for HER2 and Fc?RIIIA, HER2bsFab exhibits potent anti-tumor activity against HER2low tumors while evading most of trastuzumab Fc-linked limitations thereby potentially enlarging the number of patients eligible for breast cancer immunotherapy. PMID:24979648

Turini, Marc; Chames, Patrick; Bruhns, Pierre

2014-01-01

126

Complement in monoclonal antibody therapy of cancer.  

PubMed

Monoclonal antibodies (mAb) have been used as targeted treatments against cancer for more than a decade, with mixed results. Research is needed to understand mAb mechanisms of action with the goal of improving the efficacy of currently used mAbs and guiding the design of novel mAbs. While some mAb-induced tumor cell killing is a result of direct effects on tumor cell signaling, mAb opsonization of tumor cells also triggers activation of immune responses due to complement activation and engagement of antibody receptors on immune effector cells. In fact, complement has been shown to play an important role in modulating the anti-tumor activity of many mAb through complement-dependent cytotoxicity, antibody-dependent cytotoxicity, and through indirect effects by modulating the tumor microenvironment. Complement activity can have both agonistic and antagonistic effects on these processes. How the balance of such effects impacts on the clinical efficacy of mAb therapy remains unclear. In this review, we discuss the mAbs currently approved for cancer treatment and examine how complement can impact their efficacy with a focus on how this information might be used to improve the clinical efficacy of mAb treatment. PMID:24906530

Rogers, Laura M; Veeramani, Suresh; Weiner, George J

2014-08-01

127

In situ production of therapeutic monoclonal antibodies.  

PubMed

The use of antibodies as a treatment for disease has it origins in experiments performed in the 1890s, and since these initial experiments, monoclonal antibodies (mAbs) have become one of the fastest growing therapeutic classes for the treatment of cancer, autoimmune disease, and infectious diseases. However, treatment with therapeutic mAbs often requires high doses given via long infusions or multiple injections, which, coupled with the prohibitively high cost associated with the production of clinical-grade proteins and the transient serum half-lives that necessitate multiple administrations to gain therapeutic benefits, makes large-scale treatment of patients, especially patients in the developing world, difficult. Due to their low-cost and rapid scalability, nucleic acid-based approaches to deliver antibody gene sequences for in situ mAb production have gained substantial traction. In this review, we discuss new approaches to produce therapeutic mAbs in situ to overcome the need for the passive infusion of purified protein. PMID:25578347

Suscovich, Todd J; Alter, Galit

2015-02-01

128

The birth pangs of monoclonal antibody therapeutics  

PubMed Central

This paper examines the development and termination of nebacumab (Centoxin®), a human IgM monoclonal antibody (mAb) drug frequently cited as one of the notable failures of the early biopharmaceutical industry. The non-approval of Centoxin in the United States in 1992 generated major concerns at the time about the future viability of any mAb therapeutics. For Centocor, the biotechnology company that developed Centoxin, the drug posed formidable challenges in terms of safety, clinical efficacy, patient selection, the overall economic costs of health care, as well as financial backing. Indeed, Centocor's development of the drug brought it to the brink of bankruptcy. This article shows how many of the experiences learned with Centoxin paved the way for the current successes in therapeutic mAb development. PMID:22531443

2012-01-01

129

Monitoring therapeutic monoclonal antibodies in brain tumor.  

PubMed

Bevacizumab induces normalization of abnormal blood vessels, making them less leaky. By binding to vascular endothelial growth factor, it indirectly attacks the vascular tumor mass. The optimal delivery of targeted therapies including monoclonal antibodies or anti-angiogenesis drugs to the target tissue highly depends on the blood-brain barrier permeability. It is therefore critical to investigate how drugs effectively reach the tumor. In situ investigation of drug distribution could provide a better understanding of pharmacological agent action and optimize chemotherapies for solid tumors. We developed an imaging method coupled to protein identification using matrix-assisted laser desorption/ionization mass spectrometry. This approach monitored bevacizumab distribution within the brain structures, and especially within the tumor, without any labeling. PMID:25484065

Ait-Belkacem, Rima; Berenguer, Caroline; Villard, Claude; Ouafik, L'Houcine; Figarella-Branger, Dominique; Beck, Alain; Chinot, Olivier; Lafitte, Daniel

2014-01-01

130

SPECT assay of radiolabeled monoclonal antibodies  

SciTech Connect

The long-term goal of this research project is to develop methods to improve the utility of single photon emission computed tomography (SPECI) to quantify the biodistribution of monoclonal antibodies (MoAbs) labeled with clinically relevant radionuclides ({sup 123}I, {sup 131}I, and {sup 111}In) and with another radionuclide,{sup 211}At, recently used in therapy. We describe here our progress in developing quantitative SPECT methodology for {sup 111}In and {sup 123}I. We have focused our recent research thrusts on the following aspects of SPECT: (1) The development of improved SPECT hardware, such as improved acquisition geometries. (2) The development of better reconstruction methods that provide accurate compensation for the physical factors that affect SPECT quantification. (3) The application of carefully designed simulations and experiments to validate our hardware and software approaches.

Jaszczak, R.J.

1992-02-01

131

Functional comparison of single-chain and two-chain anti-CD3-based bispecific antibodies in gene immunotherapy applications  

PubMed Central

Gene therapy to achieve in vivo secretion of recombinant anti-CD3 x anti-tumor bispecific antibodies in cancer patients is being explored as a strategy to counterbalance rapid renal elimination, thereby sustaining levels of bispecific antibodies in the therapeutic range. Here, we performed a comparative analysis between single- and two-chain configurations for anti-CD3 x anti-CEA (carcinoembryonic antigen) bispecific antibodies secreted by genetically-modified human cells. We demonstrate that tandem single-chain variable fragment (scFv) antibodies and two-chain diabodies are expressed as soluble secreted proteins with similar yields. However, we found significant differences in their biological functionality (i.e., antigen binding) and in their ability to induce non-specific T cell activation. Whereas single-chain tandem scFvs induced human T cell activation and proliferation in an antigen-independent manner, secreted two-chain diabodies exerted almost no proliferative stimulus when human T cells were cultured alone or in co-cultures with CEA negative cells. Thus, our data suggest that two-chain diabodies are preferable to single-chain tandem scFvs for immunotherapeutic strategies comprising in vivo secretion of bispecific antibodies aiming to recruit and activate anticancer specific lymphocytic effector T cells. PMID:25057445

Compte, Marta; Álvarez-Cienfuegos, Ana; Nuńez-Prado, Natalia; Sainz-Pastor, Noelia; Blanco-Toribio, Ana; Pescador, Nuria; Sanz, Laura; Álvarez-Vallina, Luis

2014-01-01

132

Monoclonal antibodies to plant plasma-membrane antigens  

Microsoft Academic Search

Murine monoclonal antibodies to membrane antigens were generated by immunization with a crude cellular membrane preparation from suspension-cultured cells of Nicotiana glutinosa L. From a panel of thirteen monoclonal antibodies, seven were found to be directed against antigens present on the plasma-membrane by immunofluorescence visualization of antibody binding to the surface of isolated protoplasts. The corresponding set of plasma-membrane antigen(s)

P. M. Norman; V. P. M. Wingate; M. S. Fitter; C. J. Lamb

1986-01-01

133

Specific activation of resting T cells against CA19-9+ tumor cells by an anti-CD3/CA19-9 bispecific antibody in combination with a costimulatory anti-CD28 antibody.  

PubMed

Specific activation of resting lymphocytes for tumor targeting can be achieved by bispecific monoclonal antibodies (bi-mAb) with specificity for tumor antigens and T-cell-activating antigens, respectively, in combination with a costimulatory anti-CD28 antibody. We describe the generation and function of a bi-mAb with specificity for CD3 and for the tumor antigen CA19-9. The bi-mAb OKT3/NSI19-9 was generated by somatic fusion of two hybridoma lines secreting antibodies against CA19-9 and CD3, respectively. A hybrid/hybridoma was established, and its bi-mAb was characterized. In combination with a costimulatory anti-CD28 mAb resting peripheral lymphocytes could be activated specifically with T-cell proliferation and secretion of high amounts of interferon-gamma. On specific T-cell activation, bi-mAb OKT3/NSI19-9 could also redirect the cytotoxic effects of these T cells toward CA19-9+ tumor cells in vitro. Our results indicate that specific activation of resting T cells with bi-mAb OKT3/NSI19-9 in combination with an anti-CD28 mAb can activate resting T cells specifically and leads to antigen-dependent bi-mAb-mediated cytotoxicity against CA19-9+ target cells. This approach may offer new perspectives for the specific immunotherapy of CA19-9+ tumors. PMID:9336739

Hombach, A; Tillmann, T; Jensen, M; Heuser, C; Sircar, R; Diehl, V; Kruis, W; Pohl, C

1997-09-01

134

Autoantibody potential of cancer therapeutic monoclonal antibodies.  

PubMed

We and others have reported that multiple autoantibodies are unmasked in human polyclonal antibody preparations after exposure to physiological oxidizing agents (hemin) or electromotive force. We now have asked if oxidation unmasks autoantibody reactivities in monoclonal antibodies (mAb). To do this, we have studied 9 FDA approved mAb used therapeutically, including 4 chimeric, 4 humanized and 1 chemically modified chimeric Fab that were exposed to the physiological oxidizing agent hemin at 36 degrees C for 20 hr. These mAb were studied for autoantibody activity to phospholipids and DNA before and after oxidation with hemin and found to develop autoantibody activities after oxidation, while retaining their original specificity as measured by mAb anti-glycophorin A binding of erythrocytes, CD 19 binding to B lymphocytes and anti-HLA-A29 binding to A29-positive lymphocytes. The finding that certain mAb have the potential to unmask autoantibody activities as a consequence of exposure to physiological redox reactions in vitro gives pause to our present understanding of the immunological basis of tolerance and concern for potential autoimmune side effects in patients receiving mAb for diagnosis or treatment. PMID:19904753

McIntyre, John A; Faulk, And W Page

2010-07-15

135

Clinical laboratory applications of monoclonal antibodies.  

PubMed Central

Monoclonal antibody (MAb) technology is well recognized as a significant development for producing specific serologic reagents to a wide variety of antigens in unlimited amounts. These reagents have provided the means for developing a number of highly specific and reproducible immunological assays for rapid and accurate diagnosis of an extensive list of diseases, including infectious diseases. The impact that MAbs have had in characterizing infectious disease pathogens, as well as their current and future applications for use in clinical microbiology laboratories, is reviewed. In addition, the advantages (and disadvantages) of the use of MAbs in a number of immunoassays, such as particle agglutination, radioimmunoassays, enzyme-linked immunosorbent assays, immunofluorescent-antibody assays, and immunohistology, are explored, including the use of these reagents in novel test system assays. Also, nucleic acid probe technology is compared with the use of MAbs from the perspective of their respective applications in the diagnosis of infectious disease agents. There is no question that hybridoma technology has the potential to alter significantly the methods currently used in most clinical microbiology laboratories. PMID:3058298

Payne, W J; Marshall, D L; Shockley, R K; Martin, W J

1988-01-01

136

Monoclonal antibody specific for a pigmentation associated antigen  

SciTech Connect

Monoclonal antibody TA99, which specifically binds to a pigmentation associated antigen present on melanoma cells is described. Additionally, the hybridoma cell line deposited with the ATCC under Accession Number HB 8704 from which the antibody is derived, as well as methods for using the antibody are described.

Thomson, T.M.; Mattes, M.J.; Old, L.J.; Lloyd, K.O

1989-01-17

137

T-cell activation and B-cell depletion in chimpanzees treated with a bispecific anti-CD19\\/anti-CD3 single-chain antibody construct  

Microsoft Academic Search

BscCD19xCD3 is a bispecific single-chain antibody construct with exceptional cytotoxic potency in vitro and in vivo. Here, we have investigated the biological activity of bscCD19xCD3 in chimpanzee, the only animal species identified in which bscCD19xCD3 showed bispecific binding, redirected B-cell lysis and cytokine production comparable to human cells. Pharmacokinetic analysis following 2-h intravenous infusion of 0.06, 0.1 or 0.12 ?g\\/kg of

Bernd Schlereth; Cornelia Quadt; Torsten Dreier; Peter Kufer; Grit Lorenczewski; Nadja Prang; Christian Brandl; Sandra Lippold; Kathy Cobb; Kathleen Brasky; Eugen Leo; Ralf Bargou; Krishna Murthy; Patrick A. Baeuerle

2006-01-01

138

Regulatory T cells are redirected to kill glioblastoma by an EGFRvIII-targeted bispecific antibody  

PubMed Central

Regulatory T cells (Tregs) play a central role in in tumor escape from immunosurveillance. We report that a bispecific T-cell engager (BiTE) targeting a mutated form of the epidermal growth factor receptor, i.e., EGFRvIII, potently redirects Tregs to kill glioblastoma through the granzyme-perforin pathway. PMID:24475376

Choi, Bryan D; Gedeon, Patrick C; Sanchez-Perez, Luis; Bigner, Darell D; Sampson, John H

2013-01-01

139

Anti-Mesothelin Monoclonal Antibodies for the Treatment of Cancer  

Cancer.gov

The National Cancer Institute, Laboratory of Molecular Biology seeks parties interested in collaborative research to further co-develop monoclonal antibodies for the treatment of mesothelin-expressing cancers.

140

Production and characterization of yeast killer toxin monoclonal antibodies.  

PubMed Central

Monoclonal antibodies were obtained after fusion of mouse myeloma cells with spleen cells isolated from mice primed with a crude extract of yeast killer toxin produced by a strain of Hansenula anomala. Hybridomas were selected by specific immunoassay reaction of their fluid with crude yeast killer toxin extract. Among the monoclonal antibodies, which were characterized by the Western blot technique, one (designated KT4) proved to have precipitating properties, thus permitting the neutralization of the killer activity of the toxin. Experiments in double immunodiffusion showed that monoclonal antibody KT4 produced homologous precipitin bands by reacting with either the crude toxin used as immunogen or a toxic extract of Hansenula mrakii. It is suggested that these monoclonal antibodies will be useful for the purification, characterization, and understanding of the bioactions of yeast killer toxins. Images PMID:2434524

Polonelli, L; Morace, G

1987-01-01

141

DEVELOPMENT OF MONOCLONAL ANTIBODIES AGAINST FATHEAD MINNOW (PIMEPHALES PROMELAS) VITELLOGENIN  

EPA Science Inventory

We have obtained a panel of monoclonal antibodies directed against fathead minnow vitellogenin (Vtg) for use in sensitive ELISAs to quantify the response of exposure in vivo to estrogen or estrogen mimics....

142

Murine monoclonal antibodies specific for virulent Treponema pallidum (Nichols).  

PubMed Central

Murine anti-Treponema pallidum (Nichols) lymphocyte hybridoma cell lines secreting monoclonal antibodies against a variety of treponemal antigens have been generated. Hybridomas isolated were of three major types: those that were directed specifically against T. pallidum antigens, those that were directed against treponemal group antigens (as evidenced by their cross-reactivity with T. phagedenis biotype Reiter antigens), and those that cross-reacted with both treponemal as well as rabbit host testicular tissue antigens. The majority (31 of 39 clones) of these anti-T. pallidum hybridomas, which produced monoclonal antibodies of mouse isotypes immunoglobulin G1 (IgG1), IgG2a, IgG2b, IgG3 or IgM, were directed specifically against T. pallidum and not other treponemal or rabbit antigens tested by radioimmunoassay. Four of these T. pallidum-specific hybridomas secreted monoclonal antibodies with greater binding affinity for "aged" rather than freshly isolated intact T. pallidum cells, suggesting a possible specificity for "unmasked" surface antigens of T. pallidum. Six anti-T. pallidum hybridomas produced complement-fixing monoclonal antibodies (IgG2a, IgG2b, or IgM) that were capable of immobilizing virulent treponemes in the T. pallidum immobilization (TPI) test; these may represent biologically active monoclonal antibodies against treponemal surface antigens. Three other hybridomas secreted monoclonal antibodies which bound to both T. pallidum and T. phagedenis biotype Reiter antigens, thus demonstrating a possible specificity for treponemal group antigens. Five hybridoma cell lines were also isolated which produced IgM monoclonal antibodies that cross-reacted with all treponemal and rabbit host testicular tissue antigens employed in the radioimmunoassays. This report describes the construction and characteristics of these hybridoma cell lines. The potential applications of the anti-T. pallidum monoclonal antibodies are discussed. PMID:7047388

Robertson, S M; Kettman, J R; Miller, J N; Norgard, M V

1982-01-01

143

Application of Quantitative Pharmacology in Development of Therapeutic Monoclonal Antibodies  

Microsoft Academic Search

The advancement of therapeutic monoclonal antibodies during various stages of the drug development process can be effectively\\u000a streamlined when appropriate translational strategies are applied. Design of successful translational strategies for development\\u000a of monoclonal antibodies should allow for understanding of the dose– and concentration–response relationships with respect\\u000a to both beneficial and toxic effects from early phases of drug development. Evaluation of

Mohammad Tabrizi; Cherryl Funelas; Hamza Suria

2010-01-01

144

Monoclonal antibodies in the treatment of immune thrombocytopenic purpura (ITP).  

PubMed

Immune thrombocytopenic purpura is characterized by antibody-mediated destruction of platelets and suboptimal platelet production. Initially the treatment of ITP includes corticosteroids, IgG-anti-D, and intravenous immunoglobulins. Splenectomy and monoclonal antibodies are usually considered for refractory and chronic ITP patients. There are new data suggesting that early administration of rituximab is important, and this antibody has been used as first-line therapy in adults. In this concise review the role of rituximab and other monoclonal antibodies is analyzed. These agents have the capability of sparing splenectomy and possibly curing the disease in some patients. PMID:22507772

Gómez-Almaguer, David

2012-04-01

145

Monoclonal antibody to dengue capsid protein  

PubMed Central

Dengue fever (DF) and dengue hemorrhagic fever/dengue shock syndrome (DHF/DSS) are considered the most important arthropod-borne viral diseases in terms of morbidity and mortality. The emergency and severity of dengue (Den) infections increase the necessity of an early, quick and effective dengue laboratory diagnostic. Viral isolation is considered a gold standard for diagnosis of dengue infection using monoclonal antibodies (mAbs) as a tool for determining serotype specificity. Alternatives have been used to improve sensitivity and time to dengue diagnosis. Based on the early expression of dengue C protein in the life cycle, we focused our study on the application of an anti-dengue 2 virus capsid protein mAb in dengue diagnosis. The kinetic expression of dengue-2 capsid in mosquito cells and its immuno-localization in experimentally infected suckling albin Swiss (OF-1) mice brain tissues was established. The results demonstrate the possible utility of this mAb in early dengue diagnosis versus traditional isolation. In addition, a preliminary study of an enzyme immunoassay method using 8H8 mAb for specific detection of dengue C protein antigen was performed, making possible recombinant C protein quantification. The results suggest that detection of dengue capsid protein could be useful in the diagnosis of early dengue infection. PMID:20061827

Vazquez, Y; Vazquez, S V; Capó; Torres, G; Caballero, Y; Sánchez, A; Limonta, D; Alvarez, M; Guzmán, MG

2009-01-01

146

Monoclonal antibodies against plant cell wall polysaccharides  

SciTech Connect

Monoclonal antibodies (McAbs) are useful tools to probe the structure of plant cell wall polysaccharides and to localize these polysaccharides in plant cells and tissues. Murine McAbs were generated against the pectic polysaccharide, rhamnogalacturonan I (RG-I), isolated from suspension-cultured sycamore cells. The McAbs that were obtained were grouped into three classes based upon their reactivities with a variety of plant polysaccharides and membrane glycoproteins. Eleven McAbs (Class I) recognize epitope(s) that appear to be immunodominant and are found in RG-I from sycamore and maize, citrus pectin, polygalacturonic acid, and membrane glycoproteins from suspension-cultured cells of sycamore, maize, tobacco, parsley, and soybean. A second group of five McAbs (Class II) recognize epitope(s) present in sycamore RG-I, but do not bind to any of the other polysaccharides or glycoproteins recognized by Class I. Lastly, one McAb (Class III) reacts with sycamore RG-I, sycamore and tamarind xyloglucan, and sycamore and rice glucuronoarabinoxylan, but does not bind to maize RG-I, polygalacturonic acid or the plant membrane glycoproteins recognized by Class I. McAbs in Classes II and III are likely to be useful in studies of the structure, biosynthesis and localization of plant cell wall polysaccharides.

Hahn, M.G.; Bucheli, E.; Darvill, A.; Albersheim, P. (Univ. of Georgia, Athens (USA))

1989-04-01

147

Comparative efficiencies of bispecific F(ab??) 2 and chimeric mouse\\/human IgG antibodies in recruiting cellular effectors for cytotoxicity via Fc? receptors  

Microsoft Academic Search

Summary The three forms of Fc? receptor carried by monocytes (Fc?RI, II) and natural killer (NK) cells (Fc?RIII) are all capable of mediating cell lysis. Here we compare the use of F(ab'?)2 bispecific antibodies, specifically targetting individual Fc?R, and chimeric IgG mouse\\/human antibodies which are capable of targetting all Fc?R, for their ability to mediate target cell destruction. The derivatives

John Greenman; Nancy Hogg; Suzanne Nikoletti; Christopher Slade; George Stevenson; Martin Glennie

1992-01-01

148

T-cell activation and cytokine production via a bispecific single-chain antibody fragment targeted to blood-stage malaria parasites  

Microsoft Academic Search

A novel bispecific single-chain antibody fragment (biscFv) has been constructed to address the possibility of a new ap- proach to malaria therapeutic drug devel- opment. The biscFv consists of 2 differ- ent single-chain antibody fragments linked by a flexible peptide linker (Gly4- Ser)3. Of the 2 scFv fragments, one is directed against a conserved epitope of the 19-kDa C-terminal fragment

Shigeto Yoshida; Tominari Kobayashi; Hiroyuki Matsuoka; Chisato Seki; William L. Gosnell; Sandra P. Chang; Akira Ishii

149

Human monoclonal antibodies against amyloid-beta from healthy adults  

Microsoft Academic Search

Two anti-amyloid-beta human antibody-producing cell lines were established from amyloid-beta (A?)-selected lymphocytes from peripheral blood of healthy adults. ELISA and Western blot analysis showed that the monoclonal antibodies bound with high affinity to the 43 amino acid-long amyloid-beta peptide. The antigen epitope of these antibodies encountered within amino acids 1–16 of the amyloid-beta peptide. The antibodies did not bind to

Valeria Geylis; Vitaly Kourilov; Zeev Meiner; Inger Nennesmo; Nenad Bogdanovic; Michael Steinitz

2005-01-01

150

Characterization of a monoclonal antibody to bovine xanthine oxidase.  

PubMed Central

The isolation of a hybridoma cell line, C-41, secreting monoclonal antibody to bovine xanthine oxidase (EC 1.2.3.2), is described. The specificity of this antibody was determined by solid-phase immunoassay, immunoblotting procedures, affinity chromatography, immunoelectrophoresis and precipitation techniques. The results are compared with those obtained in similar specificity studies on a previously described monoclonal antibody secreted by hybridoma cell line A-94 [Mather, Nace, Johnson & Goldsby (1980) Biochem. J. 188, 925-928]. This latter antibody appears to bind to xanthine oxidase only when the enzyme is immobilized on a solid support such as a plastic plate or nitrocellulose paper. Potential problems in the determination of the specificity of monoclonal antibodies, especially towards membrane proteins of unknown biological activity, are discussed. Images Fig. 2. Fig. 3. Fig. 6. Fig. 7. PMID:6378181

Kaetzel, C S; Mather, I H; Bruder, G; Madara, P J

1984-01-01

151

Stable IgG-like Bispecific Antibodies Directed toward the Type I Insulin-like Growth Factor Receptor Demonstrate Enhanced Ligand Blockade and Anti-tumor Activity  

PubMed Central

Bispecific antibodies (BsAbs) target multiple epitopes on the same molecular target or different targets. Although interest in BsAbs has persisted for decades, production of stable and active BsAbs has hindered their clinical evaluation. Here, we describe the production and characterization of tetravalent IgG-like BsAbs that combine the activities of allosteric and competitive inhibitors of the type-I insulin-like growth factor receptor (IGF-1R). The BsAbs, which were engineered for thermal stability, express well, demonstrate favorable biophysical properties, and recognize both epitopes on IGF-1R. Only one BsAb with a unique geometry, denoted BIIB4-5scFv, was capable of engaging all four of its binding arms simultaneously. All the BsAbs (especially BIIB4-5scFv) demonstrated enhanced ligand blocking over the single monoclonal antibodies (mAbs), particularly at high ligand concentrations. The pharmacokinetic profiles of two IgG-like BsAbs were tested in nude mice and shown to be comparable with that of the parental mAbs. The BsAbs, especially BIIB4-5scFv, demonstrated an improved ability to reduce the growth of multiple tumor cell lines and to inhibit ligand-induced IGF-1R signaling in tumor cells over the parental mAbs. BIIB4-5scFv also led to superior tumor growth inhibition over its parental mAbs in vivo. In summary, BsAbs that bridge multiple inhibitory mechanisms against a single target may generally represent a more effective strategy for intervention in oncology or other indications compared with traditional mAb therapy. PMID:21123183

Dong, Jianying; Sereno, Arlene; Snyder, William B.; Miller, Brian R.; Tamraz, Susan; Doern, Adam; Favis, Michael; Wu, Xiufeng; Tran, Hon; Langley, Emma; Joseph, Ingrid; Boccia, Antonio; Kelly, Rebecca; Wortham, Kathleen; Wang, Qin; Berquist, Lisa; Huang, Flora; Gao, Sharon X.; Zhang, Ying; Lugovskoy, Alexey; Martin, Shelly; Gouvis, Heather; Berkowitz, Steven; Chiang, Gisela; Reff, Mitchell; Glaser, Scott M.; Hariharan, Kandasamy; Demarest, Stephen J.

2011-01-01

152

Heterodimeric bispecific antibody-derivatives against CD19 and CD16 induce effective antibody-dependent cellular cytotoxicity against B-lymphoid tumor cells.  

PubMed

Bispecific scFv antibody-derivatives (bsscFvs) recruiting natural killer (NK) cells for the lysis of malignant cells have therapeutic potential. However, a bsscFv specific for the B-lymphoid tumor antigen CD19 and the trigger molecule CD16 on NK cells had similar affinities for both antigens (42 and 58nM, respectively) and was not optimal for cytotoxicity. Therefore, a bispecific tribody (bsTb) was constructed with two binding sites for CD19 and one for CD16. This bsTb contained a CD19-specific Fab fragment carrying a CD16-specific scFv fused to its light chain and a CD19-specific scFv fused to its heavy chain. The bsTb was compared with a bispecific bibody (bsBb) lacking the CD19-specific scFv. The bsTb had 3-fold greater avidity for CD19 than the bsBb (8 and 24nM, respectively), while both had equal affinity for CD16 (56nM). Both molecules mediated antibody-dependent cellular cytotoxicity (ADCC) of leukemia-derived SEM cells and primary cells from leukemia patients. The bsTb showed half-maximum effective concentrations (EC(50)) of 55pM and promoted equal lysis as the bsBb and the bsscFv at 6- and 12-fold lower concentrations, respectively. Among these three molecules the bsTb showed the most promising in vitro properties which are anticipated to be displayed also in vivo. PMID:21339041

Kellner, Christian; Bruenke, Joerg; Horner, Heike; Schubert, Joerg; Schwenkert, Michael; Mentz, Kristin; Barbin, Karin; Stein, Christoph; Peipp, Matthias; Stockmeyer, Bernhard; Fey, Georg H

2011-04-28

153

The Fc-region of a new class of intact bispecific antibody mediates activation of accessory cells and NK cells and induces direct phagocytosis of tumour cells  

PubMed Central

Bispecific antibodies (bsAb) are considered as promising tools for the elimination of disseminated tumour cells in a minimal residual disease situation. The bsAb-mediated recruitment of an immune effector cell in close vicinity of a tumour cell is thought to induce an antitumoural immune response. However, classical bispecific molecules activate only a single class of immune effector cell that may not yield optimal immune responses. We therefore constructed an intact bispecific antibody, BiUII (anti-CD3 × anti-EpCAM), that not only recognizes tumour cells and T lymphocytes with its two binding arms, but also binds and activates Fc?-receptor positive accessory cells through its Fc-region. We have demonstrated recently that activated accessory cells contribute to the bsAb-induced antitumoural activity. We now analyse this stimulation in more detail and demonstrate here the BiUll-induced upregulation of activation markers like CD83 and CD95 on accessory cells and the induction of neopterin and biopterin synthesis. Experiments with pure cell subpopulations revealed binding of BiUll to CD64+ accessory cells and CD16+ NK cells, but not to CD32+ B lymphocytes. We provide further evidence for the importance of the Fc-region in that this bispecific molecule stimulates Fc?-R-positive accessory cells to eliminate tumour cells in vitro by direct phagocytosis. © 2000 Cancer Research Campaign PMID:10901380

Zeidler, R; Mysliwietz, J; Csánady, M; Walz, A; Ziegler, I; Schmitt, B; Wollenberg, B; Lindhofer, H

2000-01-01

154

Monoclonal antibodies: versatile platforms for cancer immunotherapy  

Microsoft Academic Search

Antibodies are important therapeutic agents for cancer. Recently, it has become clear that antibodies possess several clinically relevant mechanisms of action. Many clinically useful antibodies can manipulate tumour-related signalling. In addition, antibodies exhibit various immunomodulatory properties and, by directly activating or inhibiting molecules of the immune system, antibodies can promote the induction of antitumour immune responses. These immunomodulatory properties can

Rishi Surana; Shangzi Wang; Louis M. Weiner

2010-01-01

155

Monoclonal antibody therapy in multiple sclerosis  

PubMed Central

Therapeutic approaches to multiple sclerosis (MS) are based on altering the functions of the immune system, either by using broad immunosuppressive drugs used for transplantation rejection and rheumatology, or by modulating them more discreetly with beta interferon and synthetic amino-acid copolymers. These strategies are only partially successful, have important safety and tolerability limitations, and have shown to be mostly effective in earlier stages of the disease, in which acute relapses dominate the clinical picture. For progressive phenotypes of MS there are currently no effective therapeutic options. As very specific and potent immunosuppressive agents, monoclonal antibodies (mAbs) may offer considerable advantages over other therapies for MS. During the last decade, anti-a4 integrin natalizumab became the first approved mAb for treatment of relapsing MS, after convincingly demonstrating clinically significant effects on two large Phase 3 trials. Moreover, the concept of disease remission was introduced for the first time to describe patients who show no signs of clinical or imaging markers of disease activity during therapy with natalizumab. Of the mAbs under development for MS, alemtuzumab and rituximab have also shown promising evidence of effectiveness and potentially expanded the therapeutic horizon to reversal of disease progression in early relapsing patients and progressive patients who previously had not been studied. However, the appearance of progressive multifocal leukoencephalopathy (PML) in natalizumab-treated MS patients, as well as in patients with lymphoma, lupus and rheumatoid arthritis, treated with rituximab and autoimmune-type complications in alemtuzumab-treated MS patients underlines the fact that extended efficacy comes with significant clinical risks. The challenge is then how best to utilize therapies that have evidently superior efficacy in a chronic disease of young adults to obtain the best benefit-risk ratio and how to monitor and prevent emergent safety concerns. PMID:21124072

2010-01-01

156

UTILIZATION OF MONOCLONAL ANTIBODIES FOR ANTIGENIC CHARACTERIZATION OF CORONAVIRUSES  

E-print Network

UTILIZATION OF MONOCLONAL ANTIBODIES FOR ANTIGENIC CHARACTERIZATION OF CORONAVIRUSES J.F. VAUTHEROT antibodies against Bovine Enteric Coronavirus (BECV strain G110) were obtained by fusion between SP2 isolate, Human Enteric Corona- virus (HECV), Bovine Respiratory Coronavirus (BRCV) and our BECV strains

Paris-Sud XI, Université de

157

MONOCLONAL ANTIBODY TO FENBENDAZOLE: UTILITY IN RESIDUE STUDIES  

Technology Transfer Automated Retrieval System (TEKTRAN)

A monoclonal antibody-based ELISA was developed for fenbendazole, a widely used benzimidazole anthelmintic, with approved uses in cattle and other food animals. The antibody was elicited using as hapten 2-succinamido-5(6)-phenylthiobenzimidazole, which was conjugated with bovine serum albumin to pro...

158

Monoclonal Antibody Therapy for Relapsed or Treatment-Resistant Neuroblastoma  

Cancer.gov

NCI is sponsoring two clinical trials of a monoclonal antibody called ch14.18, in combination with other drugs, to see if the antibody may be helpful for children or young adults (up to age 21) with relapsed or refractory high-risk neuroblastoma.

159

Specific Endocrine Tissue Marker Defined by a Monoclonal Antibody  

Microsoft Academic Search

One of two mouse monoclonal antibodies (LK2H10) produced by hybridoma technology against a human endocrine tumor (pheochromocytoma) demonstrated specific immunoreactivity for 69 normal and neoplastic endocrine cells and tissues known to contain secretory granules. This immunoreactivity was specific, since other normal tissues, tumors from endocrine cells without granules, and tumors from other nonendocrine tissues were negative when tested with antibody

Ricardo V. Lloyd; Barry S. Wilson

1983-01-01

160

A perspective of monoclonal antibodies: Past, present, and future  

SciTech Connect

In 1975, the development of the technique to produce monoclonal antibodies revolutionized the approach to cancer detection and therapy. Hundreds of monoclonal antibodies to the epitopes of tumor cells have been produced, providing more specific tools for probing the cellular elements of cancer. At the same time, these tools have disclosed greater complexity in the character of these cells and stimulated further investigation. Although there are antibodies to specific epitopes of neoplastic cells, this purity has not provided the improved detection and therapy of cancer first expected. Technical manipulations have provided limited improvement in results, but more sophisticated techniques, such as biologic response modifiers, may be required to attain clinical results that can be universally applied. The intense research in monoclonal antibodies and their application does offer promise that the goal of improved cancer detection and therapy will be forthcoming. 58 references.

DeLand, F.H. (Veterans Administration Medical Center, Syracuse, NY (USA))

1989-07-01

161

Serological classification of Neisseria gonorrhoeae with monoclonal antibodies.  

PubMed Central

Hybrid cells producing monoclonal antibodies against antigens of Neisseria gonorrhoeae were obtained by the polyethylene glycol-mediated fusion of mouse myeloma cells and lymphocytes from mice immunized with gonococcal protein I or outer membrane proteins. From four fusions, 16 phenotypically stable, independently cloned hybrid cell lines were selected for continued study. Each of the cell lines produced a characteristically different monoclonal antibody which reacted in immunoprecipitation assays with a unique antigenic determinant on protein I of the outer membrane complex of the bacteria. In antibody binding, immunofluorescence, and coagglutination assays these antibodies each reacted with a restricted group of N. gonorrhoeae strains. None of the monoclonal antibodies reacted with 17 other different species of Neisseria or with Branhamella catarrhalis. When tested on 34 N. gonorrhoeae reference serotyping strains, the monoclonal antibodies demonstrated serological relationships between the strains which paralleled those observed with conventional polyvalent antisera. These antibodies now provide standardized reagents for the rapid and precise serological characterization of many strains of N. gonorrhoeae. Images FIG. 1 FIG. 2 FIG. 3 FIG. 4 FIG. 5 PMID:6807844

Tam, M R; Buchanan, T M; Sandström, E G; Holmes, K K; Knapp, J S; Siadak, A W; Nowinski, R C

1982-01-01

162

Monoclonal Antibodies That Recognize Discrete Forms of Tubulin  

Microsoft Academic Search

Anti-tubulin antibodies secreted by plasmacytoma NSI-spleen cell hybrids were detected by an indirect binding assay. Different antibodies bound to different combinations of the tubulins as resolved by isoelectric focusing. Two monoclonal antibodies (TUB 2.1 and TUB 2.5) labeled only (i) the tubulin band on a polyacrylamide electropherogram and (ii) beta -tubulins as resolved by isoelectric focusing. The fraction that was

Illana Gozes; Colin J. Barnstable

1982-01-01

163

A general affinity method to purify peroxidase-tagged antibodies  

Microsoft Academic Search

Antibodies tagged with enzymes, e.g. horseradish peroxidase (HRPO) are used extensively in a broad range of immunoassay, immunohistochemical, and prodrug-based immunotherapeutic applications. These antibodies may be polyclonal, monoclonal, bispecific or genetically engineered in origin. Often, purification of the antibody is the single greatest obstacle to obtaining immunoprobes with high specific activity [Milstein and Cuello, Nature 305 (1983) 537]. We have

Donald R Husereau; Mavanur R Suresh

2001-01-01

164

Monoclonal antibody therapeutics with up to five specificities: functional enhancement through fusion of target-specific peptides.  

PubMed

The recognition that few human diseases are thoroughly addressed by mono-specific, monoclonal antibodies (mAbs) continues to drive the development of antibody therapeutics with additional specificities and enhanced activity. Historically, efforts to engineer additional antigen recognition into molecules have relied predominantly on the reformatting of immunoglobulin domains. In this report we describe a series of fully functional mAbs to which additional specificities have been imparted through the recombinant fusion of relatively short polypeptides sequences. The sequences are selected for binding to a particular target from combinatorial libraries that express linear, disulfide-constrained, or domain-based structures. The potential for fusion of peptides to the N- and C- termini of both the heavy and light chains affords the bivalent expression of up to four different peptides. The resulting molecules, called zybodies, can gain up to four additional specificities, while retaining the original functionality and specificity of the scaffold antibody. We explore the use of two clinically significant oncology antibodies, trastuzumab and cetuximab, as zybody scaffolds and demonstrate functional enhancements in each case. The affect of fusion position on both peptide and scaffold function is explored, and penta-specific zybodies are demonstrated to simultaneously engage five targets (ErbB2, EGFR, IGF-1R, Ang2 and integrin ?v?3). Bispecific, trastuzumab-based zybodies targeting ErbB2 and Ang2 are shown to exhibit superior efficacy to trastuzumab in an angiogenesis-dependent xenograft tumor model. A cetuximab-based bispecific zybody that targeting EGFR and ErbB3 simultaneously disrupted multiple intracellular signaling pathways; inhibited tumor cell proliferation; and showed efficacy superior to that of cetuximab in a xenograft tumor model. PMID:23575268

LaFleur, David W; Abramyan, Donara; Kanakaraj, Palanisamy; Smith, Rodger G; Shah, Rutul R; Wang, Geping; Yao, Xiao-Tao; Kankanala, Spandana; Boyd, Ernie; Zaritskaya, Liubov; Nam, Viktoriya; Puffer, Bridget A; Buasen, Pete; Kaithamana, Shashi; Burnette, Andrew F; Krishnamurthy, Rajesh; Patel, Dimki; Roschke, Viktor V; Kiener, Peter A; Hilbert, David M; Barbas, Carlos F

2013-01-01

165

Effective elimination of acute myeloid leukemic cells by recombinant bispecific antibody derivatives directed against CD33 and CD16.  

PubMed

Single-chain Fv triplebodies (sctb), consisting of a single polypeptide chain with 3 single-chain antibody variable fragments connected in tandem, were generated as antileukemic agents. A CD19-specific sctb of this format has previously been shown to be superior to a bispecific single-chain Fv antibody fragment (bsscFv) for the elimination of leukemic B-lineage cells, but corresponding targeted agents for the treatment of acute myeloid leukemia are still lacking. For this purpose, both a bsscFv and a sctb specific for CD33 and the trigger molecule CD16 (FcgammaRIII) were produced. The sctb displayed 3.5-fold greater avidity for CD33 than the bsscFv 33xds16, whereas both had close to equal affinity for CD16. In antibody-dependent cellular cytotoxicity (ADCC) reactions with human mononuclear cells as effectors, both the bsscFv 33xds16 and the sctb induced lysis of tumor cells with half maximum effective concentrations (EC50) in the low picomolar range. It is interesting to note that the sctb promoted equal lysis of human leukemia-derived cell lines at 10 to 200-fold lower concentrations than the bsscFv. Both molecules mediated ADCC of primary patient cells. In conclusion, both the bsscFv 33xds16 and the sctb 33xds16x33 eliminated acute myeloid leukemia cells in ADCC reactions, but the novel sctb format showed significantly greater specific activity. PMID:20551837

Singer, Heiko; Kellner, Christian; Lanig, Harald; Aigner, Michael; Stockmeyer, Bernhard; Oduncu, Fuat; Schwemmlein, Michael; Stein, Christoph; Mentz, Kristin; Mackensen, Andreas; Fey, Georg H

2010-01-01

166

Monoclonal hybridoma antibodies to human amyloid related protein SAA.  

PubMed Central

Problems concerning isolation and characterization of the amyloid related serum protein SAA in a pure form prompted us to make monoclonal antibodies to the protein. Protein SAA isolated by gel filtration under dissociating conditions was used for immunization of BALB/c mice, and spleen cells from a mouse producing high titred antiserum to SAA were fused with cells from the mouse plasmacytoma line P3U1. Antibody specificity to various preparations of protein SAA was tested using an indirect enzyme-linked immunosorbent assay. Monoclonal antibodies with specificity for SAA were obtained in addition to antibodies which reacted with both SAA and the related amyloid protein AA. Antibodies specific for one of the apoC proteins of the lipoprotein fraction were also produced showing that the SAA preparation used for immunization was contaminated with apoC proteins. PMID:7151332

Marhaug, G; Gaudernack, G; Bogen, B; Husby, G

1982-01-01

167

[Regulatory consequences for the use of monoclonal antibodies].  

PubMed

Köhler and Milstein published a method for the manufacture of mouse monoclonal antibodies of predefined specificity 1975 [1], a work rewarded with the Nobel Prize 1984. Since then, the field has developed rapidly with new production methods such as recombinant DNA technology, phage display and genetically engineered animals. Following the first clinical applications with a mouse monoclonal antibody, new classes as chimaeric, humanized and human monoclonal antibodies appeared, with the advantages of less adverse reactions and better efficacy. The development over more than 30 years resulted in more than 25 approved products on the market for various therapeutic applications, e.g. for the treatment of cancer, inflammatory diseases, heart disease and transplantation, and medicines for many more applications are currently under development. PMID:20035703

Lackner, Friedrich; Behr-Gross, Marie-Emmanuelle

2009-12-01

168

A bispecific single-chain antibody that mediates target cell-restricted, supra-agonistic CD28 stimulation and killing of lymphoma cells  

Microsoft Academic Search

We have previously reported that r28M, a recombinant bispecific single-chain antibody directed to a melanoma-associated proteoglycan (NG2) and the costimulatory CD28 molecule on T cells, induced T-cell activation, which resulted in tumor-cell killing. T-cell activation did not require a primary signal through the T-cell antigen receptor (TCR)\\/CD3 complex and depended on the presence of NG2-positive tumor cells. Here, we further

T Otz; L Große-Hovest; M Hofmann; H-G Rammensee; G Jung

2009-01-01

169

Production and characterization of monoclonal and polyclonal antibodies to forchlorfenuron.  

PubMed

The development of immunoassays for the detection of the plant growth regulator forchlorfenuron (CPPU) is described. To achieve that purpose, a set of CPPU derivatives has been obtained by the previous synthesis of the adequate p-aminophenyl alkanoic acid. Protein conjugates of these compounds have been used as immunogens to produce rabbit polyclonal antibodies and a collection of mouse monoclonal antibodies. Additionally, a battery of structural analogues of the target analyte has been synthesized and used for the characterization of antibody binding. This strategy has demonstrated that most antibodies followed Landsteiner's principle, although some monoclonal antibodies showing important deviations from this behavior have also been found. Finally, different assay formats have been developed with a variety of antibodies and conjugates, and a rapid procedure has been optimized for the indirect ELISA format using monoclonal and polyclonal antibodies. In the indirect competitive ELISA, assay IC50 values for CPPU below 0.5 nM were found with LODs as low as 0.013 nM. PMID:18989973

Suárez-Pantaleón, Celia; Mercader, Josep V; Agulló, Consuelo; Abad-Somovilla, Antonio; Abad-Fuentes, Antonio

2008-12-10

170

MONOCLONAL ANTIBODIES IDENTIFY CONSERVED EPITOPES ON THE POLYHEDRIN OF 'HELIOTHIS ZEA' NUCLEAR POLYHEDROSIS VIRUS  

EPA Science Inventory

Recent advances in monoclonal antibody techniques have provided an opportunity to simplify the procedures of serological identification of microorganisms. Because monoclonal antibodies are raised against individual antigenic determinants (epitopes), they can be used to screen wit...

171

Therapeutic efficacy of potent neutralizing HIV-1-specific monoclonal antibodies in SHIV-infected rhesus monkeys  

E-print Network

Human immunodeficiency virus type 1 (HIV-1)-specific monoclonal antibodies with extraordinary potency and breadth have recently been described. In humanized mice, combinations of monoclonal antibodies have been shown to ...

Barouch, Dan H.

172

A recombinant bispecific single-chain Fv antibody against HLA class II and FcgammaRIII (CD16) triggers effective lysis of lymphoma cells.  

PubMed

Bispecific antibodies offer the possibility of improving effector-cell recruitment for antibody therapy. For this purpose, a recombinant bispecific single-chain Fv antibody (bsscFv), directed against FcgammaRIII (CD16) and human leucocyte antigen (HLA) class II, was constructed and tested in functional assays. RNA from the hybridomas 3G8 and F3.3, reacting with CD16 and HLA class II, respectively, was used to generate phage display libraries. From these libraries, reactive phages were isolated and the bsscFv was constructed by connecting both single-chain Fv components through a 20 amino acid flexible linker. After expression in SF21 insect cells and chromatographic purification, the bsscFv bound specifically and simultaneously to both antigens. The affinities of the anti-CD16 and the anti-HLA class II scFv components of the bsscFv were 8.6 x 10(-8) mol/l and 13.7 x 10(-8) mol/l, respectively, which was approximately sevenfold lower than the F(ab) fragments of the parental antibodies. In antibody-dependent cellular cytotoxicity experiments with human mononuclear cells as effectors, the bsscFv-mediated specific lysis of both HLA class II-positive, malignant human B-lymphoid cell lines and primary cells from patients with chronic B-cell lymphocytic leukaemia. Optimal lysis was obtained at bsscFv concentrations of approximately 400 ng/ml, similar to the concentration required for maximum lysis by the corresponding chemically linked bispecific antibody. Thus, this recombinant bsscFv-antibody is an efficient molecule for effector-cell mediated lysis of malignant human B-lymphoid cells. PMID:15059139

Bruenke, Joerg; Fischer, Barbara; Barbin, Karin; Schreiter, Katja; Wachter, Yvonne; Mahr, Kerstin; Titgemeyer, Fritz; Niederweis, Michael; Peipp, Matthias; Zunino, Susan J; Repp, Roland; Valerius, Thomas; Fey, Georg H

2004-04-01

173

A novel CD19-directed recombinant bispecific antibody derivative with enhanced immune effector functions for human leukemic cells.  

PubMed

A novel bispecific antibody-derived recombinant protein targeting leukemias and lymphomas was designed, a single-chain Fv triple body (sctb) consisting of 1 polypeptide chain with 3 scFvs connected in tandem. The distal scFvs were specific for the tumor antigen CD19, and the central scFv for the trigger molecule CD16 (FcgammaRIII) on natural killer (NK) cells and macrophages. We had previously built a disulphide stabilized (ds) bsscFv [19 x 16] with monovalent binding for CD19 from ds components. The sctb ds[19 x 16 x 19] also used ds components and displayed 3-fold greater avidity for CD19 than the bsscFv (KD = 13 vs. 42 nM), whereas both had equal affinity for CD16 (KD = 58 nM). Plasma half-lives in mice were 4 and 2 hours for the sctb and the bsscFv, respectively. In antibody-dependent cellular cytotoxicity reactions with human mononuclear cells as effectors, the sctb promoted equal lysis of leukemic cell lines and primary cells from leukemia and lymphoma patients at 10-fold to 40-fold lower concentrations than the bsscFv. This new format may also be applicable to a variety of other tumor antigens and effector molecules. With half-maximum effective concentrations (EC50) in the low picomolar range, the sctb ds[19 x 16 x 19] is an attractive candidate for further preclinical evaluation. PMID:18833000

Kellner, Christian; Bruenke, Joerg; Stieglmaier, Julia; Schwemmlein, Michael; Schwenkert, Michael; Singer, Heiko; Mentz, Kristin; Peipp, Matthias; Lang, Peter; Oduncu, Fuat; Stockmeyer, Bernhard; Fey, Georg H

2008-01-01

174

Mechanisms of monoclonal antibody stabilization and release from silk biomaterials  

PubMed Central

The availability of stabilization and sustained delivery systems for antibody therapeutics remains a major clinical challenge, despite the growing development of antibodies for a wide range of therapeutic applications due to their specificity and efficacy. A mechanistic understanding of protein-matrix interactions is critical for the development of such systems and is currently lacking as a mode to guide the field. We report mechanistic insight to address this need by using well-defined matrices based on silk gels, in combination with a monoclonal antibody. Variables including antibody loading, matrix density, charge interactions, hydrophobicity and water access were assessed to clarify mechanisms involved in the release of antibody from the biomaterial matrix. The results indicate that antibody release is primarily governed by hydrophobic interactions and hydration resistance, which are controlled by silk matrix chemistry, peptide domain distribution and protein density. Secondary ionic repulsions are also critical in antibody stabilization and release. Matrix modification by free methionine incorporation was found to be an effective strategy for mitigating encapsulation induced antibody oxidation. Additionally, these studies highlight a characterization approach to improve the understanding and development of other protein sustained delivery systems, with broad applicability to the rapidly developing monoclonal antibody field. PMID:23859659

Guziewicz, Nicholas A.; Massetti, Andrew J.; Perez-Ramirez, Bernardo J.; Kaplan, David L.

2013-01-01

175

Cooperative Immunoassays: Ultrasensitive Assays with Mixed Monoclonal Antibodies  

NASA Astrophysics Data System (ADS)

Mixtures of certain monoclonal antibodies appear to bind human chorionic gonadotropin in a ``cooperative'' fashion because they form circular complexes with the hormone. Experiments illustrate how this property might be exploited to develop very sensitive immunoassays for human chorionic gonadotropin or any other antigen. Since the assays are not based on competitive inhibition between radiolabeled and unlabeled antigen, they are much more sensitive than a traditional radioimmunoassay in which either one of the same antibodies is used alone.

Ehrlich, Paul H.; Moyle, William R.

1983-07-01

176

The safety and side effects of monoclonal antibodies  

Microsoft Academic Search

Monoclonal antibodies (mAbs) are now established as targeted therapies for malignancies, transplant rejection, autoimmune and infectious diseases, as well as a range of new indications. However, administration of mAbs carries the risk of immune reactions such as acute anaphylaxis, serum sickness and the generation of antibodies. In addition, there are numerous adverse effects of mAbs that are related to their

Harald Kropshofer; Thomas Singer; Jane A. Mitchell; Andrew J. T. George; Trevor T. Hansel

2010-01-01

177

Production of monoclonal antibodies in COS and CHO cells  

Microsoft Academic Search

Recent advances in the generation of genetically engineered monoclonal antibodies have enhanced the importance of COS cells as expression systems for rapidly producing sufficient quantities of these proteins for preliminary biochemical and biophysical analysis. In order to meet the demand for clinical supplies, a gradual increase has occurred in the usage of dihydrofolate reductase negative (DHFR-) Chinese hamster ovary (CHO)

John J Trill; Allan R Shatzman; Ganguly Subinay

1995-01-01

178

Development of biodegradable nanocarriers loaded with a monoclonal antibody.  

PubMed

Treatments utilizing monoclonal antibody therapeutics against intracellular protein-protein interactions in cancer cells have been hampered by several factors, including poor intracellular uptake and rapid lysosomal degradation. Our current work examines the feasibility of encapsulating monoclonal antibodies within poly(lactic-co-glycolic acid) (PLGA) nanoparticles using a water/oil/water double emulsion solvent evaporation technique. This method can be used to prepare protective polymeric nanoparticles for transporting functional antibodies to the cytoplasmic compartment of cancer cells. Nanoparticles were formulated and then characterized using a number of physical and biological parameters. The average nanoparticle size ranged from 221 to 252 nm with a low polydispersity index. Encapsulation efficiency of 16%-22% and antibody loading of 0.3%-1.12% were observed. The antibody molecules were released from the nanoparticles in a sustained manner and upon release maintained functionality. Our studies achieved successful formulation of antibody loaded polymeric nanoparticles, thus indicating that a PLGA-based antibody nanoformulation is a promising intracellular delivery vehicle for a large number of new intracellular antibody targets in cancer cells. PMID:25690029

Gdowski, Andrew; Ranjan, Amalendu; Mukerjee, Anindita; Vishwanatha, Jamboor

2015-01-01

179

Purification of human monoclonal antibodies and their fragments.  

PubMed

This chapter summarizes the most common chromatographic mAb and mAb fragment purification methods, starting by elucidating the relevant properties of the compounds and introducing the various chromatography modes that are available and useful for this application. A focus is put on the capture step affinity and ion exchange chromatography. Aspects of scalability play an important role in judging the suitability of the methods. The chapter introduces also analytical chromatographic methods that can be utilized for quantification and purity control of the product. In the case of mAbs, for most purposes the purity obtained using an affinity capture step is sufficient. Polishing steps are required if material of particularly high purity needs to be generated. For mAb fragments, affinity chromatography is not yet fully established, and the capture step potentially may not provide material of high purity. Therefore, the available polishing techniques are touched upon briefly. In the case of mAb isoform and bispecific antibody purification, countercurrent chromatography techniques have been proven to be very useful and a part of this chapter has been dedicated to them, paying tribute to the rising interest in these antibody formats in research and industry. PMID:24037849

Müller-Späth, Thomas; Morbidelli, Massimo

2014-01-01

180

Monoclonal Antibody Cross-Reactions between Drosophila and Human Brain  

NASA Astrophysics Data System (ADS)

A panel of 146 monoclonal antibodies (MAbs), obtained with Drosophila melanogaster tissue as primary immunogen, was tested for cross-reactivity with the human central nervous system. Sites examined included spinal cord, cerebellum, hippocampus, and optic nerve. Nonnervous tissues tested were liver and lymph node. Approximately half of the antibodies reacted with one or more sites in the human central nervous system, identifying regional, cell class, and subcellular antigens. Some recognized neuronal, glial, or axonal subsets. Immunoblot analysis revealed that some antibodies reacted with similar antigen patterns in both species.

Miller, Carol A.; Benzer, Seymour

1983-12-01

181

Production and characterization of monoclonal antibodies against fumitremorgin B.  

PubMed

This paper reports the preparation and identification of two monoclonal antibodies against FTB, and the establishment of an indirect competitive ELISA methods for FTB determination in buckwheat, rice, and corn. Two of the hybridoma cell lines (1C9 and 2D10), which could produce specific antibodies against fumitremorgin B(FTB), were selected and developed. The affinity Kaff constants of the monoclonal antibodies with the coating antigen, FTBS-IgG, were found to be 6 x 10(8) M-1 and 9.8 x 10 M-1, respectively. The isotypes of the monoclonal antibodies are of two isotypes, IgG1 and IgM, respectively. The antibody titers were found around 1 x 10(6) and 1.5 x 10(6). The standard curves showed that as little as 5 pg of FTB in 50 mL could be detected, and the linear range of standard curve was from 10 pg to 1000 pg of standard FTB. There were no cross-reaction for McAbs in the assay system with some mycotoxins tested. The mean recovery rate from buckwheat spiked with 10-60 ng/g of FTB was 78-88.7%. PMID:10095931

Liu, J; Meng, Z H

1998-12-01

182

Combined inhibition of TNF? and IL-17 as therapeutic opportunity for treatment in rheumatoid arthritis: Development and characterization of a novel bispecific antibody.  

PubMed

Objective: As single cytokine inhibition of e.g. TNF? or IL-6 produces clinically meaningful responses in only about half of RA patients, this study is designed to investigate whether combined inhibition of TNF? and IL-17 has additive/synergistic effects in suppression of mesenchymal cell activation in vitro and inflammation and tissue destruction in arthritis in vivo. Methods: Cultures of human fibroblast-like synoviocytes (FLS) were stimulated with TNF?, IL-17 or a combination of both. Single/combined neutralizing antibodies against TNF? and IL-17 were used to interrogate in vitro cytokine responses and in vivo development of arthritis, bone and cartilage destruction in TNF?-transgenic mice. Bi-specific anti-TNF?/IL-17 antibodies were designed and tested for their potential to block cytokine responses in human FLS. Results: TNF? and IL-17 had additive/synergistic effects in promoting IL-6, -8 and G-CSF as well as MMP production in FLS. Bi-specific anti-TNF?/IL-17 antibodies showed superior efficacy to block cytokine and chemokine responses in vitro. Furthermore, dual versus single inhibition of both cytokines using neutralizing antibodies was more effective in inhibiting the development of inflammation, bone and cartilage destruction in arthritic mice. Conclusion: Combined blockade of TNF? and IL-17 is more effective in inhibiting cytokine, chemokine and matrix enzyme responses from human mesenchymal cells and in blocking tissue destruction associated with arthritis and also showed positive impact on rebalance of bone homeostasis. Bi-specific anti-TNF?/IL-17 antibodies may have superior efficacy in the treatment of arthritis and may overcome the limited therapeutic responses of single cytokine neutralization. © 2014 American College of Rheumatology. PMID:25303306

Fischer, Jens A A; Hueber, Axel J; Wilson, Stacy; Galm, Margarete; Baum, Wolfgang; Kitson, Chris; Auer, Johannes; Lorenz, Stefan; Moelleken, Jörg; Bader, Martin; Tissot, Alain C; Tan, Seng-Lai; Seeber, Stefan; Schett, Georg

2014-10-01

183

Sperm-immobilizing monoclonal antibody to human seminal plasma antigens.  

PubMed Central

Rat spleen cells immunized to human azoospermic semen (a mixture of seminal plasma components) and mouse myeloma cells (P3/X63 Ag8U1; P3U1) (Marguilies et al., 1976) were successfully fused with polyethylene glycol (PEG 1500) and 19 of 89 fused cell cultures were found to produce sperm-immobilizing antibody. The cells that produced antibody indicating the highest sperm-immobilizing activity were distributed into wells for further recloning and 10 clones producing sperm-immobilizing antibody were established. The clone (1C4) producing the highest antibody titre was found to produce a large amount of IgG in culture supernatants and to contain a mixture of rat and mouse chromosomes. It was proved by immunodiffusion test that the monoclonal antibody was produced to the human seminal plasma antigen No. 7 which is common to human milk protein. Using this hybridoma which produced a large amount of monoclonal sperm-immobilizing antibody, a new method could be developed for purifying human seminal plasma antigen by immunoaffinity chromatography with bound antibody from the hybridoma. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 PMID:6783353

Shigeta, M; Watanabe, T; Maruyama, S; Koyama, K; Isojima, S

1980-01-01

184

Rescue of impaired NK cell activity in hodgkin lymphoma with bispecific antibodies in vitro and in patients.  

PubMed

Natural killer (NK) cells represent a key component of the innate immune system against cancer. Nevertheless, malignant diseases arise in immunocompetent individuals despite tumor immunosurveillance. Hodgkin lymphoma (HL) is characterized by CD30(+) tumor cells and a massive infiltration of immune effector cells in affected lymph nodes. The latter obviously fail to eliminate the malignant cell population. Here, we tested for functional NK cell defects in HL and suggest an improvement of NK function by therapeutic means. We demonstrate that peripheral NK cells (pNK) from patients with HL fail to eliminate HL cell lines in ex vivo killing assays. Impaired NK cell function correlated with elevated serum levels of soluble ligands for NK cell receptors NKp30 (BAG6/BAT3) and NKG2D (MICA), factors known to constrict NK cell function. In vitro, NK cell cytotoxicity could be restored by an NKG2D/NKp30-independent bispecific antibody construct (CD30xCD16A). It artificially links the tumor receptor CD30 with the cytotoxicity NK cell receptor CD16A. Moreover, we observed that NK cells from patients treated with this construct were generally activated and displayed a restored cytotoxicity against HL target cells. These data suggest that reversible suppression of NK cell activity contributes to immune evasion in HL and can be antagonized therapeutically. PMID:23459515

Reiners, Katrin S; Kessler, Jörg; Sauer, Maike; Rothe, Achim; Hansen, Hinrich P; Reusch, Uwe; Hucke, Christian; Köhl, Ulrike; Dürkop, Horst; Engert, Andreas; von Strandmann, Elke Pogge

2013-04-01

185

Simulations of site-specific target-mediated pharmacokinetic models for guiding the development of bispecific antibodies.  

PubMed

Bispecific antibodies (BAbs) are novel constructs that are under development and show promise as new therapeutic modalities for cancer and autoimmune disorders. The aim of this study is to develop a semi-mechanistic modeling approach to elucidate the disposition of BAbs in plasma and possible sites of action in humans. Here we present two case studies that showcase the use of modeling to guide BAb development. In case one, a BAb is directed against a soluble and a membrane-bound ligand for treating systemic lupus erythematosus, and in case two, a BAb targets two soluble ligands as a potential treatment for ulcerative colitis and asthma. Model simulations revealed important differences between plasma and tissues, when evaluated for drug disposition and target suppression. Target concentrations at tissue sites and type (soluble vs membrane-bound), tissue-site binding, and binding affinity are all major determinants of BAb disposition and subsequently target suppression. For the presented case studies, higher doses and/or frequent dosing regimens are required to achieve 80 % target suppression in site specific tissue (the more relevant matrix) as compared to plasma. Site-specific target-mediated models may serve to guide the selection of first-in-human doses for new BAbs. PMID:25559227

Chudasama, Vaishali L; Zutshi, Anup; Singh, Pratap; Abraham, Anson K; Mager, Donald E; Harrold, John M

2015-02-01

186

Adsorption of monoclonal antibodies to glass microparticles.  

PubMed

Microparticulate glass represents a potential contamination to protein formulations that may occur as a result of processing conditions or glass types. The effect of added microparticulate glass to formulations of three humanized antibodies was tested. Under the three formulation conditions tested, all three antibodies adsorbed irreversibly at near monolayer surface coverages to the glass microparticles. Analysis of the secondary structure of the adsorbed antibodies by infrared spectroscopy reveal only minor perturbations as a result of adsorption. Likewise, front-face fluorescence quenching measurements reflected minimal tertiary structural changes upon adsorption. In contrast to the minimal effects on protein structure, adsorption of protein to suspensions of glass microparticles induced significant colloidal destabilization and flocculation of the suspension. PMID:20575075

Hoehne, Matthew; Samuel, Fauna; Dong, Aichun; Wurth, Christine; Mahler, Hanns-Christian; Carpenter, John F; Randolph, Theodore W

2011-01-01

187

Monoclonal hybridoma anti-cardiolipin antibodies from SLE mice.  

PubMed Central

To determine whether the anti-cardiolipin antibodies are identical with the lupus anticoagulant and other antibodies to phospholipids and DNA, we prepared monoclonal hybridoma autoantibodies to cardiolipin from SLE-prone MRL/lpr mice and characterized their specificity. Using a somatic cell hybridization technique, we established three hybridoma clones which produce antibodies to cardiolipin (CAL-1: IgG2b, k, CAL-2: IgM, k and CAL-3: IgM, k). These hybridoma antibodies preferentially reacted with cardiolipin and phosphatidylserine, weakly reacted with phosphatidylinositol, but not with other phospholipids such as phosphatidylcholine, sphingomyelin, phosphatidylethanolamine and VDRL antigen. Two hybridoma anti-cardiolipin antibodies bound to ssDNA and were found to act as the lupus anticoagulant when mixing activated partial thromboplastin time with cephalin. These autoantibodies may prove to be good tools for elucidating mechanisms of thrombosis, thrombocytopenia, fetal loss and other related manifestations found in patients with systemic lupus erythematosus. PMID:3146452

Ichikawa, K; Shimada, K; Nawata, Y; Ishii, T; Tomioka, H; Yoshida, S; Koike, T

1988-01-01

188

Bispecific antibodies targeting tumor-associated antigens and neutralizing complement regulators increase the efficacy of antibody-based immunotherapy in mice.  

PubMed

The efficacy of antibody-based immunotherapy is due to the activation of apoptosis, the engagement of antibody-dependent cellular cytotoxicity and complement-dependent cytotoxicity (CDC). We developed a novel strategy to enhance CDC using bispecific antibodies (bsAbs) that neutralize the C-regulators CD55 and CD59 to enhance C-mediated functions. Two bsAbs (MB20/55 and MB20/59) were designed to recognize CD20 on one side. The other side neutralizes CD55 or CD59. Analysis of CDC revealed that bsAbs could kill 4-25 times more cells than anti-CD20 recombinant antibody in cell lines or cells isolated from patients with chronic lymphocytic leukemia. The pharmacokinetics of the bsAbs was evaluated in a human-SCID model of Burkitt lymphoma. The distribution profile of bsAbs mimics the data obtained by studying the pharmacokinetics of anti-CD20 antibodies, showing a peak in the tumor mass 3-4 days after injection. The treatment with bsAbs completely prevented the development of human/SCID lymphoma. The tumor growth was blocked by the activation of the C cascade and by the recruitment of macrophages, polymorphonuclear and natural killer cells. This strategy can easily be applied to the other anti-tumor C-fixing antibodies currently used in the clinic or tested in preclinical studies using the same vector with the appropriate modifications.Leukemia advance online publication, 4 July 2014; doi:10.1038/leu.2014.185. PMID:24903480

Macor, P; Secco, E; Mezzaroba, N; Zorzet, S; Durigutto, P; Gaiotto, T; De Maso, L; Biffi, S; Garrovo, C; Capolla, S; Tripodo, C; Gattei, V; Marzari, R; Tedesco, F; Sblattero, D

2014-06-01

189

Monoclonal antibodies to novel myeloid antigens reveal human neutrophil heterogeneity.  

PubMed Central

Three cytotoxic murine monoclonal antibodies that recognize myeloid-specific antigens have been produced by immunization with normal human neutrophils or myeloblasts from a patient with acute myelomonocytic leukemia. Two of these, PMN 6 and PMN 29, are specific for neutrophils; the third monoclonal antibody, AML-2-23, is reactive with the majority of normal monocytes as well as a subpopulation of mature neutrophils. Although neutrophils from all individuals tested expressed these antigens, cytofluorographic analysis revealed that the percentage of cells bearing the PMN 6 and AML-2-23 antigens varied among individuals. Significant additional heterogeneity in the density of each antigen among antigen-bearing cells was also observed. All three antibodies efficiently mediated complement-dependent cytotoxicity of acute myelocytic leukemia cells yet were unreactive with lymphocytic leukemia cells. Neutrophil cytotoxicity was mediated by PMN 6 and PMN 29 but not by AML-2-23. On the other hand, AML-2-23, but not PMN 6 or PMN 29, was cytotoxic for normal monocytes and macrophages. These monoclonal antibodies may be of value in the study of normal neutrophil function and differentiation and may have clinical utility in diagnosis and therapy of myeloid leukemia. PMID:6752945

Ball, E D; Graziano, R F; Shen, L; Fanger, M W

1982-01-01

190

Affinity of monoclonal antibodies for Globo-series glycans.  

PubMed

Globo-series glycans are human cell-surface carbohydrates that include stem-cell marker SSEA-4 and cancer-cell antigen Globo H. These two hexasaccharides differ only in their terminal saccharide: N-acetylneuraminic acid in SSEA-4 and L-fucose in Globo H. Herein, we evaluated the affinity of the monoclonal antibodies ?-SSEA-4 and ?-GH for the glycans SSEA-4 and Globo H. Using fluorescence polarization, we find that the two monoclonal antibodies have affinity for their cognate glycan in the low nanomolar range, and have negligible affinity for the non-cognate glycan. Using surface plasmon resonance, we find that each cognate affinity is ?20-fold greater if the glycan is immobilized on a surface rather than free in solution. We conclude that the terminal saccharide plays a dominant role in the ability of monoclonal antibodies to recognize these Globo-series glycans and that the extraordinary specificity of these antibodies supports their use for identifying and sorting stem-cells (?-SSEA-4) and as an agent in cancer immunotherapy (?-GH). PMID:25163606

Eller, Chelcie H; Yang, Guangbin; Ouerfelli, Ouathek; Raines, Ronald T

2014-10-01

191

Affinity of monoclonal antibodies for Globo-series glycans  

PubMed Central

Globo-series glycans are human cell-surface carbohydrates that include stem-cell marker SSEA-4 and cancer-cell antigen Globo H. These two hexasaccharides differ only in their terminal saccharide: N-acetylneuraminic acid in SSEA-4 and l-fucose in Globo H. Herein, we evaluated the affinity of the monoclonal antibodies ?-SSEA-4 and ?-GH for the glycans SSEA-4 and Globo H. Using fluorescence polarization, we find that the two monoclonal antibodies have affinity for their cognate glycanin the low nanomolar range, and have negligible affinity for the non-cognate glycan. Using surface plasmon resonance, we find that each cognate affinity is ~20-fold greater if the glycanis immobilized on a surface rather than free in solution. We conclude that the terminal saccharide plays a dominant role in the ability of monoclonal antibodies to recognize these Globo-series glycans and that the extraordinary specificity of these antibodies supports their use for identifying and sorting stem-cells (?-SSEA-4) and as an agent in cancer immunotherapy (?-GH). PMID:25163606

Eller, Chelcie H.; Yang, Guangbin; Ouerfelli, Ouathek; Raines, Ronald T.

2014-01-01

192

Stimuli-responsive magnetic nanoparticles for monoclonal antibody purification.  

PubMed

Monoclonal antibodies (mAbs) are important therapeutic proteins. One of the challenges facing large-scale production of monoclonal antibodies is the capacity bottleneck in downstream processing, which can be circumvented by using magnetic stimuli-responsive polymer nanoparticles. In this work, stimuli-responsive magnetic particles composed of a magnetic poly(methyl methacrylate) core with a poly(N-isopropylacrylamide-co-acrylic acid) (P(NIPAM-co-AA)) shell cross-linked with N, N'-methylenebisacrylamide were prepared by miniemulsion polymerization. The particles were shown to have an average hydrodynamic diameter of 317 nm at 18°C, which decreased to 277 nm at 41°C due to the collapse of the thermo-responsive shell. The particles were superparamagnetic in behavior and exhibited a saturation magnetization of 12.6 emu/g. Subsequently, we evaluated the potential of these negatively charged stimuli-responsive magnetic particles in the purification of a monoclonal antibody from a diafiltered CHO cell culture supernatant by cation exchange. The adsorption of antibodies onto P(NIPAM-co-AA)-coated nanoparticles was highly selective and allowed for the recovery of approximately 94% of the mAb. Different elution strategies were employed providing highly pure mAb fractions with host cell protein (HCP) removal greater than 98%. By exploring the stimuli-responsive properties of the particles, shorter magnetic separation times were possible without significant differences in product yield and purity. PMID:23420794

Borlido, Luís; Moura, Leila; Azevedo, Ana M; Roque, Ana C A; Aires-Barros, Maria R; Farinha, José Paulo S

2013-06-01

193

Monoclonal antibodies that target pathological assemblies of Abeta.  

PubMed

Amyloid beta (Abeta) immunotherapy for Alzheimer's disease has shown initial success in mouse models of Alzheimer's disease and in human patients. However, because of meningoencephalitis in clinical trials of active vaccination, approaches using therapeutic antibodies may be preferred. As a novel antigen to generate monoclonal antibodies, the current study has used Abeta oligomers (amyloid beta-derived diffusible ligands, ADDLs), pathological assemblies known to accumulate in Alzheimer's disease brain. Clones were selected for the ability to discriminate Alzheimer's disease from control brains in extracts and tissue sections. These antibodies recognized Abeta oligomers and fibrils but not the physiologically prevalent Abeta monomer. Discrimination derived from an epitope found in assemblies of Abeta1-28 and ADDLs but not in other sequences, including Abeta1-40. Immunoneutralization experiments showed that toxicity and attachment of ADDLs to synapses in culture could be prevented. ADDL-induced reactive oxygen species (ROS) generation was also inhibited, establishing this response to be oligomer-dependent. Inhibition occurred whether ADDLs were prepared in vitro or obtained from Alzheimer's disease brain. As conformationally sensitive monoclonal antibodies that selectively immunoneutralize binding and function of pathological Abeta assemblies, these antibodies provide tools by which pathological Abeta assemblies from Alzheimer's disease brain might be isolated and evaluated, as well as offering a valuable prototype for new antibodies useful for Alzheimer's disease therapeutics. PMID:17116235

Lambert, Mary P; Velasco, Pauline T; Chang, Lei; Viola, Kirsten L; Fernandez, Sara; Lacor, Pascale N; Khuon, Daliya; Gong, Yuesong; Bigio, Eileen H; Shaw, Pamela; De Felice, Fernanda G; Krafft, Grant A; Klein, William L

2007-01-01

194

A trivalent anti-erbB2/anti-CD16 bispecific antibody retargeting NK cells against human breast cancer cells.  

PubMed

Bispecific antibody (BsAb) can physically cross-link immune cells to tumor cells, circumventing the proper structures for tumor cell-immune cell interactions and activating the cellular cytotoxic mechanisms. The optimal BsAb should target tumor cells with high affinity, but activate trigger molecules on cytotoxic cells by monovalent binding of Fab fragments. In the present study, a trivalent anti-erbB2/anti-CD16 BsAb was produced. This BsAb possesses bivalent arms specifically binding to the extracellular domain of erbB2 and monovalent Fab fragment redirecting NK cells. The recombinant protein could be expressed and purified from Escherichia coli as native proteins without refolding. It was fully functional in bispecific binding to SKBR3 and NK cells. The molecular size of this trivalent BsAb protein is larger than diabody and smaller than whole antibody and expected to have advantages for both high penetration of small antibody fragments and the slow circulation clearance of whole antibody. This novel protein may be an attractive target for further improvement and evaluation. PMID:14592414

Xie, Zhigang; Shi, Ming; Feng, Jiannan; Yu, Ming; Sun, Yingxun; Shen, Beifen; Guo, Ning

2003-11-14

195

Freezing-induced perturbation of tertiary structure of monoclonal antibody  

E-print Network

and methods Model protein: Monoclonal antibody (IgG) 0.5mg/ml Buffers: 10 mM Potassium Phosphate, pH 3,8 10 mM Sodium Acetate, pH 4 Salt: 0, 150mM Potassium Chloride 1-anilino-8-naphthalene sulfonate (ANS): 75?M Instruments: Quanta... Temperature (C) -30 -20 -10 0 10 20 0.0 5.0e+5 1.0e+6 1.5e+6 2.0e+6 2.5e+6 Reference [1] Reichert, J.M., et al., Monoclonal antibody successes in the clinic. Nat Biotechnol, 2005. 23(9): p. 1073-8. [2] Gabellieri, E. and G.B. Strambini, Perturbation...

Liu, Lu; Kueltzo, L. A.; Jones, L. S.; Carpenter, J. F.

2006-10-25

196

Adjuvant therapy of ovarian cancer with radioactive monoclonal antibody.  

PubMed Central

Fifty-two patients with epithelial ovarian cancer were treated with yttrium-90-labelled monoclonal antibody HMFG1 administered intraperitoneally following conventional surgery and chemotherapy as part of an extended phase I-II trial. The treatment was well tolerated and the only significant toxicity observed was reversible myelosuppression as previously described. Following conventional surgery and chemotherapy, 21 out of the 52 patients had no evidence of residual disease and were regarded as receiving treatment in an adjuvant setting. To date, two of these patients have died of their disease (follow-up 3-62 months, median follow-up 35 months). This extended phase I-II study suggests that patients with advanced ovarian cancer who achieve a complete remission following conventional therapy may benefit from further treatment with intraperitoneal radioactive monoclonal antibody. PMID:8347497

Hird, V.; Maraveyas, A.; Snook, D.; Dhokia, B.; Soutter, W. P.; Meares, C.; Stewart, J. S.; Mason, P.; Lambert, H. E.; Epenetos, A. A.

1993-01-01

197

Monoclonal antibodies for the detection of trace chemicals  

SciTech Connect

Problems in analytical chemistry may limit monitoring for trace organic residues by traditional chromatographic methods. For example, the cost and analysis time per sample may preclude adequate sampling, making the development of alternative technologies desirable. Immunoassays are one such alternative, with the potential for cost reduction by automation and parallel sample processing. A particularly significant advance in the past decade has been the development of monoclonal antibodies, which offer greater selectivity and reproducibility than conventional antisera. Immunoassays can be developed that use simple, field-portable instrumentation, give rapid results, and have detection limits of less than a part-per-billion. This paper reviews the general technology for developing monoclonal antibodies to small organic molecules using the immunoassay of 2,3,7,8-tetrachlorodibenzodioxin (2,3,7,8-TCDD) as an example. 18 refs., 2 figs., 1 tab.

Vanderlaan, M.; Van Emon, J.; Watkins, B.; Stanker, L.

1986-08-15

198

Targeted ?-Particle Radiotherapy with 211At-labeled Monoclonal Antibodies  

PubMed Central

An attractive feature of targeted radionuclide therapy is the ability to select radionuclides and targeting vehicles with characteristics that are best suited for a particular clinical application. One combination that has been receiving increasing attention is the use of monoclonal antibodies specifically reactive to receptors and antigens that are expressed on tumor cells to selectively deliver the ?-particle emitting radiohalogen 211At to malignant cell populations. Promising results have been obtained in preclinical models with multiple 211At-labeled mAbs; however, translation of concept to the clinic has been slow. Impediments to this process include limited radionuclide availability, the need for suitable radiochemistry methods operant at high activity levels, and the lack of data concerning toxicity of ?-particle emitters in humans. Nonetheless, two clinical trials have been initiated to date with 211At-labeled monoclonal antibodies and others are planned for the near future. PMID:17921029

Zalutsky, Michael R.; Reardon, David A.; Pozzi, Oscar R.; Vaidyanathan, Ganesan; Bigner, Darell D.

2007-01-01

199

Specific activation of resting T cells against tumour cells by bispecific antibodies and CD28-mediated costimulation is accompanied by Th1 differentiation and recruitment of MHC-independent cytotoxicity.  

PubMed

Specific activation of resting lymphocytes for tumour targeting can be achieved by bispecific monoclonal antibodies (bi-MoAbs) with specificity for tumour antigens and T cell-activating antigens in combination with a costimulatory anti-CD28 antibody. In this study we focus on the immunomodulatory function of an anti-CD3/CA19-9 bi-MoAb in combination with a costimulatory anti-CD28 antibody which may result not only in antigen-specific, T cell-mediated tumour cell lysis but also in recruitment of other cellular effector functions. In combination with costimulatory anti-CD28 antibodies, resting peripheral lymphocytes could be activated specifically to secrete high amounts of Th1 cytokines (IL-2, interferon-gamma (IFN-gamma)) characterizing a cellular immune response. In contrast, no IL-4 and only low amounts of IL-10 could be detected. Furthermore, bi-MoAb-mediated CA19-9-specific activation of T cells was accompanied by recruitment of MHC- and CA19-9-independent cytotoxicity, as was determined by lysis of different CA19-9-cell lines. This MHC-independent cytotxicity was mediated at least in part by activated natural killer (NK) cells, as depletion of CD16+ NK cells resulted in substantial decrease of cytotoxicity against CA19-9- targets. Our results indicate that specific activation of resting T cells with CD3-associated bi-MoAbs in combination with an anti-CD28 antibody leads to a Th1 differentiation pathway and is accompanied by recruitment of MHC-independent lymphokine-activated killer (LAK) cell cytotoxicity which can possibly be directed against a heterogeneous tumour. PMID:9158110

Hombach, A; Tillmann, T; Jensen, M; Heuser, C; Sircar, R; Diehl, V; Kruis, W; Pohl, C

1997-05-01

200

Monoclonal Antibody Epitope Mapping Describes Tailspike -Helix Folding and Aggregation Intermediates*S  

E-print Network

Monoclonal Antibody Epitope Mapping Describes Tailspike -Helix Folding and Aggregation, nine -tailspike antibody binding epitopes were characterized by measuring the binding of these mono that the antibody epitopes are distributed throughout the tailspike struc- ture, with several clustered

Clark, Patricia L.

201

Strain?specific monoclonal antibodies to Penicillium bilaii  

Microsoft Academic Search

Monoclonal antibodies (IgM class) to Penicillium bilaii (isolate PB?50) were developed and their specificity was determined against various fungi using enzyme?linked immunosorbent assays. Cross?reactivity in the range 0–6.5% was obtained for Fusarium, Aspergillus, Paecilomyces and Trichoderma species while 1.5–9.5% was obtained for P. lapidosum, P. crustosum, P. hiramayae, P. spinolosum and P. implicatum. Strains of P. bilaii (ATCC 22348 and

J. Zawistowski; L. Gosek; J. E. Cunningham

1993-01-01

202

Serological Studies of Dianthoviruses Using Monoclonal and Polyclonal Antibodies  

Microsoft Academic Search

SUMMARY In comparative serological studies of dianthoviruses, reverse passive haemag- glutination inhibition (RPHI) tests distinguished between sweet clover necrotic mosaic virus (SCNMV) and each of a Swedish isolate of red clover necrotic mosaic virus (RCNMV-SW), the clover primary leaf necrosis strain of RCNMV (RCNMV-C) and carnation ringspot virus (CRSV). Twenty-one monoclonal antibodies prepared against SCNMV were highly specific and failed

C. Hiruki; A. L. N. Rao; Y. Furuya; GINA FIGUEIREDO

1984-01-01

203

Positron emission tomographic imaging of tumors using monoclonal antibodies  

SciTech Connect

This research project is developing methods for utilizing positron emission tomography (PET) to increase the clinical potential of radiolabeled monoclonal antibodies (MAbs). This report describes the development of methods for labeling MAbs and their fragments with positron-emitting halogen nuclides, fluorine-18 and iodine-124. These nulides were selected because of the widespread availability of F-18 and because of our extensive experience in the development of new protein radiohalogenation methods.

Zalutsky, M.R.

1992-08-01

204

Monoclonal antibodies to human brain acetylcholinesterase: properties and applications  

Microsoft Academic Search

1.Acetylcholinesterase (AChE) was purified 20,000-fold in a 43% yield from 90 g of human cerebellum by combined immunoaffinity and ligand affinity chromatography. The purified enzyme migrated as a 68,000-dalton band during polyacrylamide gel electrophoresis under denaturing and reducing conditions.2.Balb\\/c mice were immunized with multiple 10-µg injections of this material in order to raise monoclonal antibodies to human brain AChE. Three

Zoltan Rakonczay; Stephen Brimijoin

1988-01-01

205

Modulation of tumor immunity by therapeutic monoclonal antibodies  

Microsoft Academic Search

The surveillance of tumors by the immune system of cancer patients and its impact on disease progression and patient survival\\u000a have been largely documented over the last years. In parallel, the use of therapeutic monoclonal antibodies (mAbs) in oncology\\u000a has gained a widespread recognition as it has made it possible to increase patient survival and to ameliorate the quality\\u000a of

Riad Abčs; Jean-Luc Teillaud

2011-01-01

206

A photosensitizer delivered by bispecific antibody redirected T lymphocytes enhances cytotoxicity against EpCAM-expressing carcinoma cells upon light irradiation.  

PubMed

Recently conducted clinical trials have provided impressive evidence that chemotherapy resistant metastatic melanoma and several hematological malignancies can be cured using adoptive T cell therapy or T cell-recruiting bispecific antibodies. However, a significant fraction of patients did not benefit from these treatments. Here we have evaluated the feasibility of a novel combination therapy which aims to further enhance the killing potential of bispecific antibody-redirected T lymphocytes by using these cells as targeted delivery system for photosensitizing agents. For a first in vitro proof-of-concept study, ex vivo activated human donor T cells were loaded with a poly(styrene sulfonate) (PSS)-complex of the model photosensitizer 5,10,15,20-tetrakis(3-hydroxyphenyl)porphyrin (mTHPP). In the absence of light and when loading with the water-soluble PSS/mTHPP-complex occurred at a tolerable concentration, viability and cytotoxic function of loaded T lymphocytes were not impaired. When "drug-enhanced" T cells were co-cultivated with EpCAM-expressing human carcinoma cells, mTHPP was transferred to target cells. Notably, in the presence of a bispecific antibody, which cross-links effector and target cells thereby inducing the cytolytic activity of cytotoxic T lymphocytes, significantly more photosensitizer was transferred. Consequently, upon irradiation of co-cultures, redirected drug-loaded T cells were more effective in killing A549 lung and SKOV-3 ovarian carcinoma cells than retargeted unloaded T lymphocytes. Particularly, the additive approach using redirected unloaded T cells in combination with appropriate amounts of separately applied PSS/mTHPP was less efficient as well. Thus, by loading T lymphocytes with a stimulus-sensitive anti-cancer drug, we were able to enhance the cytotoxic capacity of carrier cells. Photosensitizer boosted T cells could open new perspectives for adoptive T cell therapy as well as targeted photodynamic therapy. PMID:25449805

Blaudszun, André-René; Moldenhauer, Gerhard; Schneider, Marc; Philippi, Anja

2015-01-10

207

Detection of ERK activation by a novel monoclonal antibody.  

PubMed

The mitogen-activated protein kinase, ERK is activated by a dual phosphorylation on threonine and tyrosine residues. Using a synthetic diphospho peptide, we have generated a monoclonal antibody directed to the active ERK. The antibody specifically identified the active doubly phosphorylated, but not the inactive mono- or non- phosphorylated forms of ERKs. A direct correlation was observed between ERK activity and the intensity in Western blot of mitogen-activated protein kinases from several species. The antibody was proven suitable for immunofluorescence staining, revealing a transient reactivity with ERKs that were translocated to the nucleus upon stimulation. In conclusion, the antibody can serve as a useful tool in the study of ERK signaling in a wide variety of organisms. PMID:9188779

Yung, Y; Dolginov, Y; Yao, Z; Rubinfeld, H; Michael, D; Hanoch, T; Roubini, E; Lando, Z; Zharhary, D; Seger, R

1997-05-26

208

Monoclonal antibodies in the treatment of pancreatic cancer  

PubMed Central

Human pancreatic cancer is a malignant disease with almost equal incidence and mortality. Effective diagnostic and therapeutic strategies are still urgently needed to improve its survival rate. With advances in structural and functional genomics, recent work has focused on targeted molecular therapy using monoclonal antibodies. This review summarizes the target molecules on the tumor cell surface and normal tissue stroma, which are related to pancreatic cancer oncogenesis, tumor growth or resistance to chemotherapy, as well as molecules involved in regulating inflammation and host immunoresponses. Targeted molecules include cell-surface receptors, such as the EGF receptor, HER2, death receptor 5 and IGF-1 receptor. Effects of monoclonal antibodies against these target molecules alone or in combination with chemotherapy, small-molecule signal transduction inhibitors, or radiation therapy are also discussed. Also discussed are the use of toxin or radioisotope conjugates, and information relating to the use of these targeting agents in pancreatic cancer clinical trials. Although targeted molecular therapy with monoclonal antibodies has made some progress in pancreatic cancer treatment, especially in preclinical studies, its clinical application to improve the survival rate of pancreatic cancer patients requires further investigation. PMID:20046965

Huang, Zhi-Qiang; Buchsbaum, Donald J

2009-01-01

209

Human tumor antigens identified with monoclonal antibodies  

SciTech Connect

MoAbLc1 (IgM) and MoAbLc2 (IgG/sub 2a/) were produced against human lung carcinoma cell line (ChaGo). Lc1 recognizes a approx. = 330-kd/approx. = 310-kd glycoprotein complexes, and Lc2 recognizes a approx. = 60-kd/approx. = 47-kd protein complex. With a panel of cell lines of different tissue origin, Lc1 showed a more restricted reactivity to ChaGo; it cross-reacted with another lung carcinoma cell line (SK-Lc-2) and two breast carcinoma cell lines, but failed to react with cell lines of fetal lung, of colon, esophageal, prostate, stomach, and ovarian carcinomas, of B and T lymphoblastoid cells, neuroblastomas, glioblastoma, astrocytoma, and human peripheral blood lymphocytes. New and improved methods were developed for the production of indium-111-labeled MoAbs for tumor imaging. To facilitate the application of bicyclic anhydride diethylenetriaminepentaacetic acid (BADTPA) to In-111 labeling of antibodies, we have modified the original method by using C-14-labeled BADTPA, which allows precise quantitation of DTPA molecules incorporated. A new heterobifunctional reagent, 2,6-dioxo-N-(carboxyl)morpholine (DCM) was synthesized for chelating In-111 to MoAbs, and demonstrated higher retention of immunoreactivity of the labeled antibody.

AlSedairy, S.T.

1987-01-01

210

Cloned transgenic farm animals produce a bispecific antibody for T cell-mediated tumor cell killing  

Microsoft Academic Search

Complex recombinant antibody fragments for modulation of immune function such as tumor cell destruction have emerged at a rapid pace and diverse anticancer strategies are being developed to benefit patients. Despite improvements in molecule design and expression systems, the quantity and stability, e.g., of single-chain antibodies produced in cell culture, is often insufficient for treatment of human disease, and the

Ludger Grosse-Hovest; Sigrid Müller; Rosa Minoia; Eckhard Wolf; Valeri Zakhartchenko; Hendrik Wenigerkind; Caroline Lassnig; Urban Besenfelder; Mathias Müller; Simon D. Lytton; Gundram Jung; Gottfried Brem

2004-01-01

211

Varicella-zoster viral glycoproteins analyzed with monoclonal antibodies.  

PubMed Central

Monoclonal antibodies to varicella-zoster virus were used to study viral glycoproteins by immunoprecipitation and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Based on the viral glycoproteins immunoprecipitated, the five monoclonal antibodies fell into three groups. Two antibodies, 4B7 and 8G9 (group 1), immunoprecipitated a single glycoprotein of molecular weight (MW) 118,000 (118K glycoprotein) and had high neutralizing activity in the absence of complement. One antibody, 3C7 (group 2), which lacked neutralizing activity, immunoprecipitated two glycoproteins of MWs 120,000 and 118,000 and a glycoprotein giving a diffuse band in the region of 64,000 to 65,000. Pulse-chase experiments and experiments with monensin as an inhibitor of glycosylation suggested that the 120K polypeptide was derived by glycosylation of the 118K polypeptide and that a 43K antigen was processed into the 64 to 65K glycoprotein. Two antibodies, 3G8 and 4E6 (group 3), both had neutralizing activity only in the presence of complement, and both immunoprecipitated at least five polypeptides, with MWs ranging from 50,000 to 90,000. Antibody 3G8 was isotype immunoglobulin G2b (IgG2b), and its immunoprecipitating activity was stronger than that of 4E6, which was isotype IgG1. Pulse-chase experiments with antibody 3G8 showed that lower-MW glycopeptides chased into three polypeptides of MWs 90,000, 80,000, and 60,000 by 24 h. Immunoprecipitation experiments with antibody 3G8 on infected cells treated with glycosylation inhibitors 2-deoxyglucose, monensin, and tunicamycin, suggested that a prominent, early-appearing 70K polypeptide may have been processed into the glycoproteins of higher MWs and that the 60K polypeptide may have been derived by glycosylation of polypeptides of lower MWs. Images PMID:6090710

Forghani, B; Dupuis, K W; Schmidt, N J

1984-01-01

212

The effect of dexamethasone on polyclonal T cell activation and redirected target cell lysis as induced by a CD19\\/CD3-bispecific single-chain antibody construct  

Microsoft Academic Search

BiTE molecules comprise a new class of bispecific single-chain antibodies redirecting previously unstimulated CD8+ and CD4+ T cells for the elimination of target cells. One example is MT103 (MEDI-538; bscCD19xCD3), a CD19-specific BiTE that can\\u000a induce lysis of normal and malignant B cells at low picomolar concentrations, which is accompanied by T cell activation. Here,\\u000a we explored in cell culture

Christian Brandl; Cornelia Haas; Sandrine d’Argouges; Tanja Fisch; Peter Kufer; Klaus Brischwein; Nadja Prang; Ralf Bargou; JoAnn Suzich; Patrick A. Baeuerle; Robert Hofmeister

2007-01-01

213

Production, characterization, and applications of a murine monoclonal antibody to dog erythrocyte antigen 1.1.  

PubMed

A murine IgM monoclonal antibody, which recognizes dog erythrocyte antigen (DEA) 1.1, has been produced. The antibody correctly identified canine RBC possessing DEA 1.1 in a panel of RBC typed by an independent laboratory. Reactivity of the monoclonal antibody was compared with canine anti-DEA 1.1 antiserum with 163 RBC samples from 145 dogs. Results of agglutination tests with the 2 reagents were in agreement for all samples. A card agglutination test that uses the monoclonal antibody with blood is described. A monoclonal antibody-based test should facilitate blood typing for DEA 1.1 in clinical practice. PMID:1289333

Andrews, G A; Chavey, P S; Smith, J E

1992-11-15

214

Delivery of the ribosome-inactivating protein, gelonin, to lymphoma cells via CD22 and CD38 using bispecific antibodies.  

PubMed Central

It is well established that bispecific antibodies (BsAbs) can be used effectively in targeting the ribosome-inactivating protein (RIP), saporin, against neoplastic B cells. We have now extended this delivery system for use with gelonin. By measuring antigen-binding characteristics and epitope mapping a panel of anti-gelonin MAbs using the IAsys resonant mirror bisensor, we were able to rapidly select the most suitable for making BaAbs. The Fab' fragments from these MAbs were chemically conjugated with Fab' from either anti-CD22 or anti-CD38. Cytotoxicity assays showed that BsAbs were highly efficient at delivering gelonin to cultured Daudi cells and achieved levels of toxicity which correlated closely with the affinity of the BsAbs. Using pairs of anti-CD22 BsAbs we were able to generate bivalent BsAb-gelonin complexes which achieved IC50 values of 2 x 10(-11) M gelonin, a potency which is equivalent to that reached by saporin in this targeting system. However, because gelonin is 5-10 times less toxic than saporin, the therapeutic ratio for gelonin is superior, making it potentially a more useful agent for human treatment. Cytotoxicity assays and kinetic analysis showed that targeting gelonin via CD38 was 2-5 times less effective than delivery through CD22. However, with a pair of BsAbs designed to co-target gelonin via CD22 and CD38, the cytotoxicity achieved equalled that obtained with a pair of anti-CD22 BsAbs (IC50 = 1 x 10(-11) M). This important result suggests that the anti-CD38 helps bind the gelonin to the cell and is then 'dragged' or 'piggy-backed' into the cell by the anti-CD22 BsAb. The implication of these findings for cancer therapy is discussed. PMID:7734325

French, R. R.; Penney, C. A.; Browning, A. C.; Stirpe, F.; George, A. J.; Glennie, M. J.

1995-01-01

215

Recombinant human monoclonal antibodies to Ebola virus.  

PubMed

Human Fab (IgG1kappa) phage display libraries were constructed from bone marrow RNA from 2 donors who recovered from infection with Ebola (EBO) virus during the 1995 outbreak in Kikwit, Democratic Republic of the Congo. The libraries were initially panned against a radiation-inactivated EBO virus-infected Vero cell lysate, but only weak binders were identified. In contrast, panning against secreted EBO glycoprotein (SGP) resulted in Fabs showing very strong reactivity with SGP in ELISA. These Fabs also reacted with a virion membrane preparation. The Fabs were strongly positive in IFAs with cells infected with EBO (subtype Zaire) virus but negative with uninfected cells, with a characteristic punctate staining pattern in the cytoplasm. The Fabs showed weak or no reactivity with the virus cell lysate although donor serum did react. The Fabs are now being characterized in structural and functional terms. Major interest will focus on the ability of antibodies to neutralize EBO virus and, later, to protect animals against infection. PMID:9988189

Maruyama, T; Parren, P W; Sanchez, A; Rensink, I; Rodriguez, L L; Khan, A S; Peters, C J; Burton, D R

1999-02-01

216

Immunochemical characterization of anti-acetylcholinesterase inhibitory monoclonal antibodies.  

PubMed

Monoclonal antibodies (mAbs) were prepared against native or DFP-inhibited Torpedo californica acetylcholinesterase and native or DFP-, MEPQ-, and soman-inhibited fetal bovine serum acetylcholinesterase. The cross reactivity of these antibodies with acetylcholinesterases from various species and their ability to inhibit catalytic activity were determined. Eight antibodies were found to inhibit catalytic activity of either Torpedo or fetal bovine serum enzyme. In all cases the antibodies bound to the native form of the enzymes and in some cases even to the denatured form. None of the antibodies recognized human or horse serum butyrylcholinesterase. Sucrose density gradient centrifugation of enzyme-antibody complexes provided two types of profiles, one with multiple peaks, indicating numerous complexes between tetrameric forms of the enzyme, and the other with single peaks, demonstrating complex formation within the tetrameric form. Different antibodies appeared to interact with slightly different regions, but in all cases the binding encompassed the peripheral anionic site. Decrease in catalytic activity of the enzyme was most likely caused by conformational changes in the enzyme molecule resulting from interaction with these mAbs. PMID:7688272

Gentry, M K; Saxena, A; Ashani, Y; Doctor, B P

1993-06-01

217

Monkey-derived monoclonal antibodies against Plasmodium falciparum  

SciTech Connect

A system has been developed that allows efficient production of monkey monoclonal antibodies from owl monkeys. Splenocytes or peripheral blood lymphocytes from monkeys immune to the human malarial parasite, Plasmodium falciparum, were fused with P3X63 Ag8.653 mouse myelomas. The resulting hybridomas were screened by an indirect fluorescent antibody test for the production of monkey monoclonal antibodies (mAb) reactive with P. falciparum. Most of the mAb reacted with the P. falciparum merozoites and immunoprecipitated a parasite-derived glycoprotein having a relative molecular weight of 185,000. These mAb gave a minimum of five different immunoprecipitation patterns, thus demonstrating that a large number of polypeptides obtained when parasitized erythrocytes are solubilized share epitopes with this large glycoprotein. In addition, mAb were obtained that reacted with antigens associated with the infected erythrocyte membrane. One of these mAb bound a M/sub r/ 95,000 antigen. Radioimmunoprecipitation assays using /sup 125/T-antibodies were done.

Stanley, H.A.; Reese, R.T.

1985-09-01

218

Monoclonal anti-A antibody removal by synthetic A antigen immobilized on specific antibody filters.  

PubMed

Removal of blood group anti-A and anti-B antibodies can prevent hyperacute organ rejection in ABO-incompatible transplantation. We are developing an extracorporeal-specific antibody filter (SAF) as an immunoadsorption device for direct removal of ABO blood group antibodies from whole blood, without the need for plasma separation and plasma exchange. A hollow fiber-based small scale SAF (mini-SAF) device was fabricated and synthetic A antigen, Atrisaccharide (Atri) conjugated to activated polyacrylic acid, was immobilized on the fiber lumen surface. Monoclonal antibody anti-A IgM were specifically removed up to 70% of initial antibodies using mini-SAF device. The monoclonal anti-A capture experiments on mini-SAF indicated that antibody removal relative to the initial concentration is independent of inlet concentration in the beginning; however, as the surface starts saturating with bound antibodies, removal becomes dependent on inlet concentration. No significant effect of flow rate on removal rate was observed. The radial diffusion and axial convection-based mathematical model developed for unsteady state antibody removal was in good agreement with the experimental data and showed that the antibody removal rate can be maximized by increasing the antibody-binding capacity of the SAF. PMID:17705231

Gautam, Shalini; Korchagina, Elena Y; Bovin, Nicolai V; Federspiel, William J

2008-03-01

219

Seeing Better through a MIST: Evaluation of Monoclonal Recombinant Antibody Fragments on  

E-print Network

to the identification of monoclonal binders ob- tained from the selections. Here, we present a new approach to reduceSeeing Better through a MIST: Evaluation of Monoclonal Recombinant Antibody Fragments the requirements of antibody micro- arrays and to obtain a great variety of antibodies, new technologies

Konthur, Zoltán

220

Epitope mapping of a monoclonal antibody against human thrombin by H/D-exchange mass spectrometry  

E-print Network

Epitope mapping of a monoclonal antibody against human thrombin by H/D-exchange mass spectrometry February 18, 2002; ACCEPTED February 18, 2002) Abstract The epitope of a monoclonal antibody raised against. The antibody epitope was identified as the surface of thrombin that retained deuterium in the presence

Komives, Elizabeth A.

221

Using monoclonal antibodies to prevent mucosal transmission of epidemic infectious diseases.  

PubMed Central

Passive immunization with antibodies has been shown to prevent a wide variety of diseases. Recent advances in monoclonal antibody technology are enabling the development of new methods for passive immunization of mucosal surfaces. Human monoclonal antibodies, produced rapidly, inexpensively, and in large quantities, may help prevent respiratory, diarrheal, and sexually transmitted diseases on a public health scale. PMID:10081672

Zeitlin, L.; Cone, R. A.; Whaley, K. J.

1999-01-01

222

F(ab')2 fragment of a gp41 NHR-trimer-induced IgM monoclonal antibody neutralizes HIV-1 infection and blocks viral fusion by targeting the conserved gp41 pocket.  

PubMed

Using a recombinant protein N46FdFc that mimics the HIV-1 gp41 N-helix trimer to immunize mice, we identified the first IgM monoclonal antibody 18D3 that specifically bound to the conserved gp41 pocket. Its F(ab')2 fragment potently inhibited HIV-1 Env-mediated cell-cell fusion and neutralized infection by laboratory-adapted and primary HIV-1 isolates with different subtypes and tropism, including the T20-resistant variants. This F(ab')2 fragment can be used to develop a bispecific broad neutralizing monoclonal antibody or HIV-1 inactivator as a novel immunotherapeutic for treatment and prevention of HIV-1 infection. PMID:24141089

Lu, Lu; Wei, Meili; Chen, Yanxia; Xiong, Weiliang; Yu, Fei; Qi, Zhi; Jiang, Shibo; Pan, Chungen

2013-11-01

223

Distinct epitopes of Ik gene products identified by monoclonal antibodies.  

PubMed

Analysis of reactivity of monoclonal anti-Iak alloantibodies, obtained by fusion between NS 1 myeloma and spleen cells from alloimmune A. TH mice, permitted the identification of 9 distinct determinants of the Ik gene products. Competitive binding experiments indicated that 2 private epitopes (defined by H8-109.13 and H8-138.4 antibodies) of the I-Ak product could be separated, thereby apparently splitting the Ia.2 specificity. A public determinant of the I-Ak molecule (identified by H8-15.9 antibody) was found expressed not only on the I-A products of the H-2b, H-2d, H-2ja, H-2p and H-2q murine haplotypes, but also on human HLA-DR antigens. Four determinants of the I-E/Ck antigen (defined by H7-8.26, H10-81.10, H10-93.2 and H8-86.2 antibodies) had a strain distribution analogues to the Ia.7 specificity. However, competitive binding experiments, and the cross-reactivity pattern with Ia-like antigens from other species (e.g. human HLA-DR antigens) indicated that these antibodies detected distinct determinants on the I-E/Ck molecule, thereby subdividing the broad Ia. 7 specificity. Two other determinants (defined by H9-14.8 and H9-15.4 antibodies) had a strain distribution that did not permit a precise assignment to a given Ia antigen, even though preliminary data suggested that they could detect separate determinants on the I-E/Ck product. All these monoclonal antibodies identified membrane antigens with the expected Ia tissue distribution pattern, and most of them could precipitate a molecule containing two chains of 28kD and 35kD, from mouse spleen cell lysates. PMID:6162651

Pierres, M; Kourilsky, F M; Rebouah, J P; Dosseto, M; Caillol, D

1980-12-01

224

Monoclonal antibodies to intermediate filament proteins of human cells: unique and cross- reacting antibodies  

PubMed Central

Monoclonal antibodies were generated against the intermediate filament proteins of different human cells. The reactivity of these antibodies with the different classes of intermediate filament proteins was determined by indirect immunofluorescence on cultured cells, immunologic indentification on SDS polyacrylamide gels (“wester blot” experiments), and immunoperoxidase assays on intact tissues. The following four antibodies are described: (a) an antivimentin antibody generated against human fibroblast cytoskeleton; (b), (c) two antibodies that recognize a 54-kdalton protein in human hepatocellular carcinoma cells; and (d) an antikeratin antibody made to stratum corneum that recognizes proteins of molecular weight 66 kdaltons and 57 kdaltons. The antivimentin antibody reacts with vimentin (58 kdaltons), glial fibrillary acidic protein (GFAP), and keratins from stratum corneum, but does not recognize hepatoma intermediate filaments. In immunofluorescence assays, the antibody reacts with mesenchymal cells and cultured epithelial cells that express vimentin. This antibody decorates the media of blood vessels in tissue sections. One antihepatoma filament antibody reacts only with the 54 kdalton protein of these cells and, in immunofluorescence and immunoperoxidase assays, only recognizes epithelial cells. It reacts with almost all nonsquamous epithelium. The other antihepatoma filament antibody is much less selective, reacting with vimentin, GFAP, and keratin from stratum corneum. This antibody decorates intermediate filaments of both mesenchymal and epithelial cells. The antikeratin antibody recognizes 66-kdalton and 57-kdalton proteins in extracts of stratum corneum and also identifies proteins of similar molecular weights in all cells tested. However, by immunofluorescence, this antibody decorates only the intermediate filaments of epidermoid carcinoma cells. When assayed on tissue sections, the antibody reacts with squamous epithelium and some, but not all, nonsquamous epithelium. Therefore this antistratum corneum antibody and the anti-54-kdalton antibody identify unique epitopes present in the various cytokeratin molecules of epithelial cells. None of the hybridoma antibodies react with neurofilament proteins. The different patterns of reactivity of these antibodies suggest that many of the immunologically distinct intermediate filament proteins contain common antigenic determinants. PMID:6183272

Gown, AM; Vogel, AM

1982-01-01

225

PRODUCTION AND CHARACTERIZATION OF MONOCLONAL ANTIBODIES, APTAMERS AND SINGLE CHAIN ANTIBODIES TO MYCOBACTERIUM AVIUM SUBSP. PARATUBERCULOSIS  

Technology Transfer Automated Retrieval System (TEKTRAN)

Paratuberculosis (MAP) was identified as an unmet need at the 7th International Colloquium on Paratuberculosis in Bilbao, Spain. To fill this gap in Johne’s disease research, monoclonal antibodies (mAbs) against MAP were produced from BALB/c mice immunized with sonicated MAP extracts or recombinant...

226

Mechanisms of Allergen-Antibody Interaction of Cockroach Allergen Bla g 2 with Monoclonal Antibodies  

E-print Network

Mechanisms of Allergen-Antibody Interaction of Cockroach Allergen Bla g 2 with Monoclonal of Molecular Biology, University of Salzburg, Salzburg, Austria Abstract Background: Cockroach allergy-ray crystallography on opposite sites of the quasi-symmetrical cockroach allergen Bla g 2. Methodology

227

Labeling of cerebral amyloid in vivo with a monoclonal antibody.  

PubMed

We assessed the ability of a murine monoclonal antibody to bind selectively to beta-amyloid in the brains of living nonhuman primates. To circumvent the blood-brain barrier, we injected unlabeled antibody 10D5 (murine whole IgG1 and/or Fab fragments) into the cerebrospinal fluid of the cisterna magna in three aged monkeys. A control animal was given an intracisternal injection of nonimmune mouse whole IgG plus Fab. Twenty-four hours later, the animals were perfused and prepared for immunohistochemical detection of bound murine immunoglobulin in brain. All three experimental animals showed selective binding of 10D5 to approximately 5-15% of amyloid deposits in cerebral cortex, primarily near the cortical surface. There was no labeling in the control animal. In vivo-labeled deposits were confirmed to be beta-amyloid by electron microscopy and by in vitro immunohistochemistry in adjacent sections. The animals tolerated the injection well, although some polymorphonuclear leukocytes infiltrated portions of the subarachnoid space and superficial neocortex. These results provide the first demonstration that it may be feasible to selectively direct a tagged monoclonal antibody to beta-amyloid in the brain for therapeutic or diagnostic purposes. With enhancement of labeling efficiency, the method also may be useful for studying the progression of beta-amyloidosis in experimental animals using emission tomography. PMID:8021711

Walker, L C; Price, D L; Voytko, M L; Schenk, D B

1994-07-01

228

Sheep monoclonal antibody fragments generated using a phage display system.  

PubMed

Monoclonal sheep antibodies have great potential for biomedical, veterinary and agricultural purpose. Although conventional sheep monoclonal antibodies can be generated by a modified hybridoma technology, the procedures are not routine. Here, we describe a method to generate recombinant sheep antibody fragments from immunised animals using a modified phage display system. Total RNA from pooled spleens of sheep immunised with the model antigens human serum albumin and conalbumin were used to amplify immunoglobulin V gene repertoires and an efficient two-step cloning method was employed to rapidly construct a phage display single-chain Fv (scFv) library. A total of 14 different scFvs were isolated and characterised. Sequence analysis indicated typical ovine immunoglobulin characteristics. Thirteen Vlambda and 11 VH genes were identified that could be grouped into the sheep Vlambda families I, II, VI and a single VH family. Soluble monomeric scFvs, produced in the periplasm of Escherichia coli, were subjected to affinity measurement via surface plasmon resonance analysis and affinities typical of the secondary immune response were observed. The method described here should be of value for the study of sheep immunology as well as for biorecognition in general. PMID:10699586

Li, Y; Kilpatrick, J; Whitelam, G C

2000-03-01

229

Preparation and identification of monoclonal antibodies against ?-conotoxin MVIIA.  

PubMed

?-Conotoxins MVIIA (?-CTX MVIIA) is a peptide with 25 amino acid residues. It is a selective and reversible N-type voltage-gated calcium channel blocker, which could be used as an analgesic for pain. To date, there are no monoclonal antibodies (MAb) for immunoassay against ?-conotoxin MVIIA. In this study, an MAb against ?-conotoxin MVIIA was prepared. The conotoxin-coding DNA sequence was chemically synthesized and cloned into expression vector pGEX-6p-1 and pET32a (+), respectively. The fusion protein GST-CTX was expressed and purified, and was used to immunize BALB/c mice for preparing the anti-CTX antibody. The spleen cells were fused with SP2/0 myeloma cells after the titer of antiserum was detected and qualified. After being screened by indirect ELISA and cloned by limiting dilution, a hybridoma named 4A12, which produces monoclonal antibody specifically against ?-CTX MVIIA, was successfully obtained. It was found that there are 102 chromosomes in the 4A12 cell, and the subclass for the MAb is IgM. The MAb affinity against ?-CTX MVIIA was 7.33×10(9) L/mol, and the cross-reaction test showed that the MAb specifically bound ?-CTX MVIIA. The MAb could be used as a specific antagonist for ?-CTX MVIIA in the physiological study on the CaV channels in the nervous system. PMID:25171005

Yang, Yanling; Ma, Yanling; Li, Heng; Wang, Shihua; Zhuang, Zhenhong

2014-08-01

230

[Toxic effects and use of therapeutic monoclonal antibodies].  

PubMed

The extensive use of therapeutic monoclonal antibodies in clinic has shown that toxic effects may occur in patients. Examples such as muromonab (anti-CD3) and recently TGN1412 (anti-CD28) clearly establish that prediction of toxic effects for these targeted therapies is still complex. These undesirable effects can be classified in four categories: cytokine release syndrome, auto-immune diseases, organ toxicity and opportunistic infections. Immunogenicity, which is highly variable depending on the degree of humanization, could also potentially lead to adverse effects due to immune-complexes formation. The recent accident observed with the anti-CD28 TGN1412 has led to the conclusion that the relative confidence in the safety of monoclonal antibodies should be revised. In addition, the prediction of non-clinical models is rather limited and extrapolation of animal results to the human situation should be performed with caution. A better knowledge of the cellular target of the antibodies and of their mechanisms of action and the identification of risk factors (immune cell activation, wide expression of the target...) should improve the clinical safety of these products. . PMID:20035692

Pallardy, Marc

2009-12-01

231

Detecting low level sequence variantsin recombinant monoclonal antibodies  

PubMed Central

A systematic analytical approach combining tryptic and chymotryptic peptide mapping with a Mascot Error Tolerant Search (ETS) has been developed to detect and identify low level protein sequence variants, i.e., amino acid substitutions, in recombinant monoclonal antibodies. The reversed-phase HPLC separation with ultraviolet (UV) detection and mass spectral acquisition parameters of the peptide mapping methods were optimized by using a series of model samples that contained low levels (0.5–5.0%) of recombinant humanized anti-HER2 antibody (rhumAb HER2) along with another unrelated recombinant humanized monoclonal antibody (rhumAb A). This systematic approach’s application in protein sequence variant analysis depends upon time and sensitivity constraints. An example of using this approach as a rapid screening assay is described in the first case study. For stable CHO clone selection for an early stage antibody project, comparison of peptide map UV profiles from the top four clone-derived rhumAb B samples quickly detected two sequence variants (M83R at 5% and P274Tat 42% protein levels) from two clones among the four. The second case study described in this work demonstrates how this approach can be applied to late stage antibody projects. A sequence variant, L413Q, present at 0.3% relative to the expected sequence of rhumAb C was identified by a Mascot-ETS for one out of four top producers. The incorporation of this systematic sequence variant analysis into clone selection and the peptide mapping procedure described herein have practical applications for the biotechnology industry, including possible detection of polymorphisms in endogenous proteins. PMID:20400866

Yang, Yi; Strahan, Alex; Li, Charlene; Shen, Amy; Liu, Hongbin; Ouyang, Jun; Katta, Viswanatham; Francissen, Kathleen

2010-01-01

232

Immunosuppression associated with novel chemotherapy agents and monoclonal antibodies.  

PubMed

The introduction of novel agents to the therapeutic armamentarium for oncologic, rheumatologic, and neurologic disorders has resulted in major clinical advances. These agents impact immune function, resulting in a discrete spectrum of infectious complications. Purine analogues and alemtuzumab alter cell-mediated immunity, resulting in opportunistic viral/fungal infections. Herpes zoster incidence increases with bortezomib. Hepatitis B reactivation may occur with rituximab. Cases of progressive multifocal leukoencephalopathy have occurred following monoclonal antibody therapy. Tumor necrosis factor-? inhibitor therapy is complicated by tuberculosis reactivation and fungal infections. We summarize the impact of these therapies on pathogenesis and spectrum of infection complicating their usage. PMID:25352632

Morrison, Vicki A

2014-11-15

233

Preparation of monoclonal antibody against crocin and its characterization.  

PubMed

Three crocin-carrier protein conjugates were synthesized and their hapten numbers were determined by matrix-assisted laser desorption/ionization time of flight mass spectrometry. Three monoclonal antibodies against crocin were produced by hybridomas fused with the splenocytes immunized with crocin hemisuccinate-bovine serum albumin conjugate and HAT-sensitive mouse myeloma cell line, P3-X63-Ag8-653. They were identified as IgG2a and IgG2b possessing lambda light chain, respectively. Their wide reactivities against crocetin glycosides were discussed. PMID:19003338

Xuan, L; Tanaka, H; Xu, Y; Shoyama, Y

1999-01-01

234

Monoclonal antibodies directed against surface molecules of multicell spheroids  

NASA Technical Reports Server (NTRS)

The objective of this project is to generate a library of monoclonal antibodies (MAb's) to surface molecules involved in the cell-cell interactions of mammalian cells grown as multicell spheroids (MCS). MCS are highly organized 3-dimensional multicellular structures which exhibit many characteristics in vivo tissues not found in conventional monolayer or suspension culture. They also provide a functional assay for surface adhesion molecules. In brief, MCS combine the relevance of organized tissues with the accuracy of in vitro methodology. Further, one can manipulate these MCS experimentally to discern important information about their biology.

Martinez, Andrew O.

1993-01-01

235

Internal radiation dosimetry for clinical testing of radiolabeled monoclonal antibodies  

SciTech Connect

In gauging the efficacy of radiolabeled monoclonal antibodies in cancer treatment, it is important to know the amount of radiation energy absorbed by tumors and normal tissue per unit administered activity. This paper describes methods for estimating absorbed doses to human tumors and normal tissues, including intraperitoneal tissue surfaces, red marrow, and the intestinal tract from incorporated radionuclides. These methods use the Medical Internal Radiation Dose (MIRD) scheme; however, they also incorporate enhancements designed to solve specific dosimetry problems encountered during clinical studies, such as patient-specific organ masses obtained from computerized tomography (CT) volumetrics, estimates of the dose to tumor masses within normal organs, and multicellular dosimetry for studying dose inhomogeneities in solid tumors. Realistic estimates of absorbed dose are provided within the short time requirements of physicians so that decisions can be made with regard to patient treatment and procurement of radiolabeled antibodies. Some areas in which further research could improve dose assessment are also discussed. 16 refs., 3 figs.

Fisher, D.R.; Durham, J.S.; Hui, T.E.; Hill, R.L.

1990-11-01

236

Mass spectrometry for the biophysical characterization of therapeutic monoclonal antibodies.  

PubMed

Monoclonal antibodies (mAbs) are powerful therapeutics, and their characterization has drawn considerable attention and urgency. Unlike small-molecule drugs (150-600 Da) that have rigid structures, mAbs (?150 kDa) are engineered proteins that undergo complicated folding and can exist in a number of low-energy structures, posing a challenge for traditional methods in structural biology. Mass spectrometry (MS)-based biophysical characterization approaches can provide structural information, bringing high sensitivity, fast turnaround, and small sample consumption. This review outlines various MS-based strategies for protein biophysical characterization and then reviews how these strategies provide structural information of mAbs at the protein level (intact or top-down approaches), peptide, and residue level (bottom-up approaches), affording information on higher order structure, aggregation, and the nature of antibody complexes. PMID:24291257

Zhang, Hao; Cui, Weidong; Gross, Michael L

2014-01-21

237

Mass Spectrometry for the Biophysical Characterization of Therapeutic Monoclonal Antibodies  

PubMed Central

Monoclonal antibodies (mAbs) are powerful therapeutics, and their characterization has drawn considerable attention and urgency. Unlike small-molecular drugs (150-600 Da) that have rigid structures, mAbs (~150 kDa) are engineered proteins that undergo complicated folding and can exist in a number of low-energy structures, posing a challenge for traditional methods in structural biology. Mass spectrometry (MS)-based biophysical characterization approaches can provide structural information, bringing high sensitivity, fast turnaround, and small sample consumption. This review outlines various MS-based strategies for protein biophysical characterization and then reviews how these strategies provide structural information of mAbs at the protein level (intact or top-down approaches), peptide, and residue level (bottom-up approaches), affording information on higher order structure, aggregation, and the nature of antibody complexes. PMID:24291257

Zhang, Hao; Cui, Weidong; Gross, Michael L.

2014-01-01

238

Cross-reactive monoclonal antibodies for diagnosis of pneumococcal meningitis.  

PubMed Central

A diagnostic test for the detection of Streptococcus pneumoniae meningitis was developed using monoclonal antibodies (MAbs) to phosphocholine (PC) and non-PC determinants of pneumococcal teichoic acids. These MAbs do not recognize other bacteria that commonly cause meningitis. By using a dot blot assay, these MAbs were compared with a polyvalent pneumococcal capsular omniserum and an antiserum made to whole cells for their ability to detect pneumococci in infected spinal fluids. An immunoglobulin M (IgM) anti-PC antibody gave a positive reaction with 16 of 22 (73%) pneumococcal culture-positive spinal fluids. One false-positive result out of 45 pneumococcal culture-negative spinal fluids was also observed. D3114/63, an IgM MAb to non-PC determinants of teichoic acids, detected 15 of 22 of the pneumococcal culture-positive spinal fluids with one false-positive result. IgG2b and IgG3 anti-PC MAbs were less efficient than the IgM anti-PC MAb at detecting pneumococci in spinal fluids. Like the IgM anti-PC MAb, omniserum detected 73% of the culture-positive pneumococcal spinal fluids, with one false-positive result. The use of anti-PC or D3114/63 MAbs instead of a pooled serum such as omniserum has several advantages: (i) use of a single cross-reactive antibody rather than 83 pooled antibodies; (ii) possibility of a higher concentration of reactive antibody, which may increase the sensitivity of the test; (iii) a standardized antibody preparation; (iv) ease of preparation of the antibody; and (v) less expense. PMID:2460493

Waltman, W D; Gray, B; McDaniel, L S; Briles, D E

1988-01-01

239

Human monoclonal IgM antibodies with apoptotic activity isolated from cancer patients  

Microsoft Academic Search

Abstract. Monoclonal antibodies are accepted as ideal adjuvant therapeutic reagents for all kinds of diseases. Polyvalent (cross- linking) and low-mutated IgM antibodies (less immunogenic),are believed to be the most effective weapons,against cancer. The best sources for these types of antibodies are the cancer patients themselves. Using conventional hybridoma technology, not only are fully human monoclonal IgM antibodies isolated, but also

Stephanie Br Andlein; Judith Lorenz; Nele Ruoff; Frank Hensel; Ines Beyer; Justus M Uller; Konrad Neukam; Bertram Illert; Matthias Eck; Hans Konrad M Uller-hermelink; H. Peter Vollmers

240

Active Lymphocytic Myocarditis Treated with Murine OKT3 Monoclonal Antibody  

PubMed Central

This report describes the case of a 33-year-old woman with biopsy-proven, active lymphocytic myocarditis manifested by intractable ventricular tachycardia, nonspecific intraventricular block, and myocardial dysfunction. We treated her successfully with OKT3 monoclonal antibody and antiarrhythmic agents. Immunosuppression is not recommended in patients with infectious or postinfectious myocarditis. However, it may have an important role in autoimmune myocarditis. In the few reports in the medical literature that we were able to find, OKT3 monoclonal antibody was administered early, in the setting of acute, fulminant autoimmune myocarditis. Our patient received OKT3 therapy in a later phase of the disease, when inflammatory infiltrates were accompanied by extensive fibrosis and severe damage of cardiomyocytes. Our patient had concomitant Helicobacter pylori infection and a strong positive family history of gastric cancer, a disease often associated with H. pylori. We discuss the possibility of a causal relationship between H. pylori infection and autoimmune myocarditis. (Tex Heart Inst J 2002;29:113–7) PMID:12075867

Bilinska, Zofia T.; Grzybowski, Jacek; Szajewski, Tomasz; Stepinska, Janina; Michalak, Ewa; Walczak, Ewa; Wagner, Teresa; Kwiatkowska, Barbara; Ruzyllo, Witold

2002-01-01

241

Examination of HER3 targeting in cancer using monoclonal antibodies.  

PubMed

The human EGF receptor (HER/EGFR) family of receptor tyrosine kinases serves as a key target for cancer therapy. Specifically, EGFR and HER2 have been repeatedly targeted because of their genetic aberrations in tumors. The therapeutic potential of targeting HER3 has long been underestimated, due to relatively low expression in tumors and impaired kinase activity. Nevertheless, in addition to serving as a dimerization partner of EGFR and HER2, HER3 acts as a key player in tumor cells' ability to acquire resistance to cancer drugs. In this study, we generated several monoclonal antibodies to HER3. Comparisons of their ability to degrade HER3, decrease downstream signaling, and inhibit growth of cultured cells, as well as recruit immune effector cells, selected an antibody that later emerged as the most potent inhibitor of pancreatic cancer cells grown as tumors in animals. Our data predict that anti-HER3 antibodies able to intercept autocrine and stroma-tumor interactions might strongly inhibit tumor growth, in analogy to the mechanism of action of anti-EGFR antibodies routinely used now to treat colorectal cancer patients. PMID:25564668

Gaborit, Nadčge; Abdul-Hai, Ali; Mancini, Maicol; Lindzen, Moshit; Lavi, Sara; Leitner, Orith; Mounier, Lucile; Chentouf, Myriam; Dunoyer, Sai; Ghosh, Manjusha; Larbouret, Christel; Chardčs, Thierry; Bazin, Hervé; Pčlegrin, André; Sela, Michael; Yarden, Yosef

2015-01-20

242

Generation and characterization of mouse monoclonal antibodies to the myelin-associated glycoprotein (MAG)  

Microsoft Academic Search

A panel of mouse monoclonal antibodies to rat and human myelin-associated glycoprotein (MAG) was developed. Normal mice were unresponsive to rat MAG, and successful immunization with rat MAG was only achieved in autoimmune NZB mice. By contrast, all strains of mice were responsive to human MAG. The monoclonal antibodies developed differ with respect to immunoglobulin type, their specificity for human

Michael J. Dobersen; Jeffrey A. Hammer; Antonio B. Noronha; Tracy D. MacIntosh; Bruce D. Trapp; Roscoe O. Brady; Richard H. Quarles

1985-01-01

243

Monoclonal Antibodies Cross-Reactive with Group A Streptococci and Normal and Psoriatic Human Skin  

Microsoft Academic Search

Infection with group A streptococci has been implicated as a factor capable of exacerbating psoriasis. In order to explore the possibility of cross-reactivity between streptococcal antigens and human skin in this phenomenon, skin from psoriatic patients and control subjects was reacted with 3 monoclonal antibodies against group A streptococci and antibody binding was estimated by the indirect immunofluorecence technique. Monoclonal

Robert A. Swerlick; Madeleine W. Cunningham; Nancy K. Hall

1986-01-01

244

Monoclonal antibodies to cyclodiene insecticides and method for detecting the same  

DOEpatents

Methods are described for making specific monoclonal antibodies useful for detection of cyclodienes in foods and environmental samples. Monoclonal antibodies specifically reactive with cyclodienes can detect accumulated pesticides in food, tissue or environmental samples. Extraction and preparation of organic samples for immunoassay in a polar-nonpolar reaction medium permits detection of halogenated organic ring structures at concentrations in samples.

Stanker, Larry H. (Livermore, CA); Vanderlaan, Martin (Danville, CA); Watkins, Bruce E. (Livermore, CA)

1994-01-01

245

Development and characterization of mouse monoclonal antibodies specific for chicken interleukin 18  

Technology Transfer Automated Retrieval System (TEKTRAN)

Four mouse monoclonal antibodies (mAbs) which are specific for chicken interleukin 18 (chIL18) were produced and characterized by enzyme-linked immunosorbent assay (ELISA), Western blotting, quantitative real-time PCR and neutralization assays. Monoclonal antibodies specific for chIL18 identified a ...

246

Antagonist properties of monoclonal antibodies targeting human CD28  

PubMed Central

Antagonist antibodies targeting CD28 have been proposed as an alternative to the use of CD80/86 antagonists to modulate T cell responses in autoimmunity and transplantation. Advantages would be the blockade of CD28-mediated co-stimulatory signals without impeding the co-inhibitory signals dependent on CD80 interactions with CTLA-4 and PD-L1 that are important for the control of immune responses and for the function of regulatory T cells. Anti-CD28 antibodies are candidate antagonists only if they prevent access to the CD80/86 ligands without simultaneously stimulating CD28 itself, a process that is believed to depend on receptor multimerization. In this study, we evaluated the impact of different formats of a potentially antagonist anti-human CD28 antibody on T cell activation. In particular, we examined the role of valency and of the presence of an Fc domain, two components that might affect receptor multimerization either directly or in the presence of accessory cells expressing Fc receptors. Among monovalent (Fab’, scFv), divalent (Fab’2), monovalent-Fc (Fv-Fc) and divalent-Fc (IgG) formats, only the monovalent formats showed consistent absence of induced CD28 multimerization and absence of associated activation of phosphoinositol-3-kinase, and clear antagonist properties in T cell stimulation assays. In contrast, divalent antibodies showed agonist properties that resulted in cell proliferation and cytokine release in an Fc-independent manner. Conjugation of monovalent antibodies with polyethylene glycol, ?-1-antitrypsin or an Fc domain significantly extended their in vivo half-life without modifying their antagonist properties. In conclusion, these data indicate that monovalency is mandatory for maintaining the antagonistic activity of anti-CD28 monoclonal antibodies. PMID:23221503

Mary, Caroline; Coulon, Flora; Poirier, Nicolas; Dilek, Nahzli; Martinet, Bernard; Blancho, Gilles; Vanhove, Bernard

2013-01-01

247

Identification by monoclonal antibodies and characterization of human platelet caldesmon.  

PubMed

Actin-based gels were prepared from clarified high-salt extracts of human platelets by dialysis against physiological salt buffers. The gel was partially solubilized with 0.3 M KCl. Mice were immunized with the 0.3 M KCl extract of the actin gel, and hybridomas were produced by fusion of spleen cells with myeloma cells. Three hybridomas were generated that secrete antibodies against an 80-kD protein. These monoclonal antibodies stained stress fibers in cultured cells and cross-reacted with proteins in several tissue types, including smooth muscle. The cross-reacting protein in chicken gizzard smooth muscle had an apparent molecular weight of 140,000 and was demonstrated to be caldesmon, a calmodulin and actin-binding protein (Sobue, K., Y. Muramoto, M. Fujita, and S. Kakiuchi, Proc. Natl. Acad. Sci. USA, 78:5652-5655). No proteins of molecular weight greater than 80 kD were detectable in platelets by immunoblotting using the monoclonal antibodies. The 80-kD protein is heat stable and was purified using modifications of the procedure reported by Bretscher for the rapid purification of smooth muscle caldesmon (Bretscher, A., 1985, J. Biol. Chem., 259:12873-12880). The 80-kD protein bound to calmodulin-Sepharose in a Ca++-dependent manner and sedimented with actin filaments, but did not greatly increase the viscosity of F-actin solutions. The actin-binding activity was inhibited by calmodulin in the presence of calcium. Except for the molecular weight difference, the 80-kD platelet protein appears functionally similar to 140-kD smooth muscle caldesmon. We propose that the 80-kD protein is platelet caldesmon. PMID:3517005

Dingus, J; Hwo, S; Bryan, J

1986-05-01

248

Selection of Ceratitis capitata (Diptera: Tephritidae) specific recombinant monoclonal phage display antibodies for prey detection analysis.  

PubMed

Several recombinant antibodies against the Mediterranean fruit fly, Ceratitis capitata (Wiedemann) (Diptera: Tephritidae), one of the most important pests in agriculture worldwide, were selected for the first time from a commercial phage display library of human scFv antibodies. The specificity and sensitivity of the selected recombinant antibodies were compared with that of a rabbit polyclonal serum raised in parallel using a wide range of arthropod species as controls. The selected recombinant monoclonal antibodies had a similar or greater specificity when compared with classical monoclonal antibodies. The selected recombinant antibodies were successfully used to detect the target antigen in the gut of predators and the scFv antibodies were sequenced and compared. These results demonstrate the potential for recombinant scFv antibodies to be used as an alternative to the classical monoclonal antibodies or even molecular probes in the post-mortem analysis studies of generalist predators. PMID:23272105

Monzó, César; Urbaneja, Alberto; Ximénez-Embún, Miguel; García-Fernández, Julia; García, José Luis; Castańera, Pedro

2012-01-01

249

Selection of Ceratitis capitata (Diptera: Tephritidae) Specific Recombinant Monoclonal Phage Display Antibodies for Prey Detection Analysis  

PubMed Central

Several recombinant antibodies against the Mediterranean fruit fly, Ceratitis capitata (Wiedemann) (Diptera: Tephritidae), one of the most important pests in agriculture worldwide, were selected for the first time from a commercial phage display library of human scFv antibodies. The specificity and sensitivity of the selected recombinant antibodies were compared with that of a rabbit polyclonal serum raised in parallel using a wide range of arthropod species as controls. The selected recombinant monoclonal antibodies had a similar or greater specificity when compared with classical monoclonal antibodies. The selected recombinant antibodies were successfully used to detect the target antigen in the gut of predators and the scFv antibodies were sequenced and compared. These results demonstrate the potential for recombinant scFv antibodies to be used as an alternative to the classical monoclonal antibodies or even molecular probes in the post-mortem analysis studies of generalist predators. PMID:23272105

Monzó, César; Urbaneja, Alberto; Ximénez-Embún, Miguel; García-Fernández, Julia; García, José Luis; Castańera, Pedro

2012-01-01

250

The first constant domain (C H1 and C L) of an antibody used as heterodimerization domain for bispecific miniantibodies  

Microsoft Academic Search

Bispecific miniantibodies were constructed by genetically fusing the CH1 domain of an IgG1 to the C-terminus of a single-chain Fv fragment (scFv-425), specific for the EGF receptor, and fusing the CL domain of a kappa light chain to the C-terminus of a scFv specific for CD2 (scFv-M1). An efficient dicistronic gene arrangement for functional expression in Escherichia coli was constructed.

Kristian M Müller; Katja M Arndt; Wolfgang Strittmatter; Andreas Plückthun

1998-01-01

251

Monoclonal antibodies in the treatment of lymphoid malignancies.  

PubMed

Rituximab (Rit) was the first monoclonal antibody approved for therapeutic use in cancer patients. Rit is a chimeric mouse/human monoclonal antibody, consisting of the human IgG1 and k constant Fc region, and a mouse variable Fab region specific against the B-cell antigen CD20. Rit exerts its antilymphoma activity through many different mechanisms. Binding of antibody to CD20 antigen, provokes apoptosis through downstream signals that lead to caspase-3 activation. Complement activation by the Fc portion of the antibody results in complement-dependent cytotoxicity. However, the most effective mechanism of action seems to be antigen-dependent cellular cytotoxicity. Effector cytotoxic cells such as natural killer cells (NK) are activated after binding to the Fc portion of the anti-CD20 molecule. Activated NK cells kill the coated lymphoma cells with the use of granzyme-perforin system. More recently, pre-clinical data support the concept that Rituximab can provoke a vaccination-like effect. Finally in-vitro experiments and clinical trials have shown that co-administration of the antibody with cytotoxics confers a strong synergistic effect. The relative contribution of these mechanisms in vivo and in different lymphoma subtypes is not well known and remains to be further evaluated. Among the different histological groups, follicular lymphoma (FL) has been proven to be the most sensitive to Rit when used as a single agent, with overall response rates of 80% and 50% in untreated and previously treated patients, respectively. Moreover, Rit in combination with chemotherapy is superior to chemotherapy alone in terms of response rate and event-free survival, while early data indicate a significant prolongation in overall survival as well. Similarly, the addition of Rit to standard chemotherapy improves the disease-free and overall survival of patients with diffuse large B-cell lymphoma. There is no doubt that Rit represents one of the greatest achievements of biotechnology engineering. However, we need to understand better the mechanisms of its action as well as the mechanisms of resistance to Rit, in order to design more effective treatment modalities. PMID:17935974

Tsirigotis, Panagiotis; Economopoulos, Theofanis

2008-02-01

252

Generation and characterization of new HER2 monoclonal antibodies.  

PubMed

Antibodies are among the most commonly used research and diagnostic tools. Antibody type and clonality are important in any assay as they can influence epitope detection. HER2 oncoprotein is overexpressed or undergoes gene amplification in approximately 30% of invasive breast carcinomas and 20% of gastric adenocarcinomas. Overexpression of HER2 is primarily detected by immunohistochemistry (IHC) on neoplastic tissue sections. We produced five murine hybridoma clones secreting monoclonal antibodies (MAbs) against HER2 protein. For hybridoma production, spleen cells from BALB/c mice immunized with a recombinant fragment of the extracellular portion of HER2 (rHER2) were fused to SP2/O-Ag14 cells, selected in HAT medium and screened by indirect ELISA. MAbs secreted were characterized according to isotypes, functional affinity constants, reaction with the native protein in MCF-7 cells by indirect immunofluorescence and in tissue sections from HER2 positive breast cancer specimens by IHC. Two MAbs were IgG2b and three were IgG1, and their affinity constants ranged from 6×10(7) to 1×10(9)M(-1). All MAbs reacted with the native protein and two stained strongly the membrane of neoplastic cells overexpressing HER2. These two MAbs could be useful in assaying HER2 overexpression in human tissues for research and possibly diagnostic purposes after a proper large-scale validation study. PMID:22901624

Vasconcellos, Flávia Aleixo; Aleixo, Pedro Bandeira; Stone, Simone Cardozo; Conceiçăo, Fabricio Rochedo; Dellagostin, Odir Antônio; Aleixo, José Antonio Guimarăes

2013-04-01

253

Molecular mechanisms of resistance to the EGFR monoclonal antibody cetuximab  

PubMed Central

The epidermal growth factor receptor (EGFR) is a receptor tyrosine kinase belonging to the HER family of receptor tyrosine kinases. Receptor activation upon ligand binding leads to down stream activation of the PI3K/AKT, RAS/RAF/MEK/ERK and PLC?/PKC pathways that influence cell proliferation, survival and the metastatic potential of tumor cells. Increased activation by gene amplification, protein overexpression or mutations of the EGFR has been identified as an etiological factor in a number of human epithelial cancers (e.g., NSCLC, CRC, glioblastoma and breast cancer). Therefore, targeting the EGFR has been intensely pursued as a cancer treatment strategy over the last two decades. To date, five EGFR inhibitors, including three small molecule tyrosine kinase inhibitors (TKIs) and two monoclonal antibodies have gained FDA approval for use in oncology. Both approaches to targeting the EGFR have shown clinical promise and the anti-EGFR antibody cetuximab is used to treat HNSCC and CRC. Despite clinical gains arising from use of cetuximab, both intrinsic resistance and the development of acquired resistance are now well recognized. In this review we focus on the biology of the EGFR, the role of EGFR in human cancer, the development of antibody-based anti-EGFR therapies and a summary of their clinical successes. Further, we provide an in depth discussion of described molecular mechanisms of resistance to cetuximab and potential strategies to circumvent this resistance. PMID:21293176

Brand, Toni M; Iida, Mari

2011-01-01

254

A thermal-cycling method for disaggregating monoclonal antibody oligomers.  

PubMed

Non-native oligomeric forms of biopharmaceutical proteins are therapeutically inactive, and potentially toxic and immunogenic, and therefore undesirable in pharmaceutical formulations. Immunoglobulin G class of antibodies are known to form stable nonnative oligomers through Fab-Fab interactions. In this paper, we investigate thermal-cycling as a technique for disaggregating antibody oligomers. Aggregate containing monoclonal antibody (mAb) samples were exposed to rapid heating and cooling cycles in a thermal-cycler. The heating phase of the thermal-cycle resulted in partial unfolding of the Fab domain, leading to the release of monomer from the oligomer complexes, whereas the rapid cooling that followed led to refolding and minimized the probability of protein reaggregation. The extent of mAb oligomer disaggregation was determined by size-exclusion chromatography and hydrophobic interaction membrane chromatography, whereas protein refolding was assessed by circular dichroism spectroscopy. The thermal-cycling technique in addition to being suitable for disaggregating protein oligomer samples could also potentially be useful for studying the mechanisms of protein aggregation and disaggregation. PMID:24549731

Sadavarte, Rahul H; Ghosh, Raja

2014-03-01

255

The safety and side effects of monoclonal antibodies.  

PubMed

Monoclonal antibodies (mAbs) are now established as targeted therapies for malignancies, transplant rejection, autoimmune and infectious diseases, as well as a range of new indications. However, administration of mAbs carries the risk of immune reactions such as acute anaphylaxis, serum sickness and the generation of antibodies. In addition, there are numerous adverse effects of mAbs that are related to their specific targets, including infections and cancer, autoimmune disease, and organ-specific adverse events such as cardiotoxicity. In March 2006, a life-threatening cytokine release syndrome occurred during a first-in-human study with TGN1412 (a CD28-specific superagonist mAb), resulting in a range of recommendations to improve the safety of initial human clinical studies with mAbs. Here, we review some of the adverse effects encountered with mAb therapies, and discuss advances in preclinical testing and antibody technology aimed at minimizing the risk of these events. PMID:20305665

Hansel, Trevor T; Kropshofer, Harald; Singer, Thomas; Mitchell, Jane A; George, Andrew J T

2010-04-01

256

100% human monoclonal antibodies in oncology: hype or breakthrough?  

PubMed

Targeted therapies have dramatically modified treatment strategies in oncology since the early 2000's, especially for treating digestive cancers. These new biotherapies such as anti-VEGF (bevacizumab) or anti-EGFR (cetuximab) monoclonal antibodies have given oncologists new opportunities to use innovative treatment schedules or combinations with cytotoxics. Consequently, significant improvements in response rates, with trends to longer progression-free survival and/or overall survival have been achieved in patients with metastatic colorectal cancer (mCRC). Panitumumab is a novel, 100% human, anti-EGFR1 (HER1) antibody that has been approved in late 2007 for use as monotherapy in mCRC patients resistant to standard chemotherapy, provided that their tumor express EGFR and display wild-type K-Ras status. Panitumumab has been recently further approved in combination with chemotherapy in mCRC patients. However, owing to the fact that its mechanism of action for targeting EGFR is similar to that of chimeric cetuximab, picturing the specificities in pharmacological and pharmacokinetic properties of this 100% human antibody could help the oncologists to better define their strategies at the bedside. PMID:22978333

Benay, Stephan; Fanciullino, Raphaelle; Mercier, Cedric; Iliadis, Athanassios; Ciccolini, Joseph; Lacarelle, Bruno

2012-01-01

257

Monoclonal Antibodies to the S1 Spike and Membrane Proteins of Avian Infectious Bronchitis Coronavirus Strain Massachusetts M41  

Microsoft Academic Search

SUMMARY We have established four murine hybridoma cell lines which secrete specific antibody to avian infectious bronchitis virus (IBV) strain Massachusetts M41. Two monoclonal antibodies reacted with the spike protein and two reacted with the membrane protein. The specificity of the monoclonal antibodies for the external structural proteins was detected by immunoprecipitations using radiolabelled virus. The reactions of the monoclonal

A. P. ADRIAN MOCKETT; DAVID CAVANAGH; T. D. K. Brown

1984-01-01

258

[Specific monoclonal antibodies to the Mycoplasma-type pathogenic agent of grapevine flavescence dorée].  

PubMed

Three hybridomas producing monoclonal antibodies specific for the mycoplasma-like organism (MLO), pathogenic agent of grapevine flavescence dorée, were obtained after fusion between spleen cells from a mouse immunized against flavescence dorée MLO and Sp2/O-Ag-14 mouse myeloma cells. These hybridomas were selected in an indirect sandwich ELISA in which antibodies from two different animal species were used (rabbit and mouse). This assay is convenient for anti-MLO monoclonal antibody screening, because of its sensitivity, specificity and good preservation of antigens. The three monoclonal antibodies were examined for reactivity towards the MLO causal agents of 15 plant yellows. None of the three were implicated in a serological relationship between the MLO of these plant yellows and flavescence dorée-MLO. Reactivity of the three monoclonal antibodies was checked towards two other isolates of flavescence dorée. One of the three antibodies detected the wild isolates. PMID:2799068

Schwartz, Y; Boudon-Padieu, E; Grange, J; Meignoz, R; Caudwell, A

1989-01-01

259

Immunoelectron microscopic localization of estrogen receptor with monoclonal estrophilin antibodies.  

PubMed

The recent production of a series of monoclonal estrophilin (estrogen receptor) antibodies recognizing estrogen receptor derived from a wide variety of animals and target tissues permits the development of immunoelectron microscopic techniques for identifying estrogen receptor. We have determined suitable conditions for the ultrastructural localization of estrogen receptor in tissue sections. Localization of receptor was observed in the euchromatin, but not in the marginated heterochromatin or nucleoli of epithelial and stromal nuclei of human endometrium. Competition studies indicate that only estrogen receptor specifically inhibits nuclear staining. The absence of any specific cytoplasmic localization at the electron-microscopic level is consistent with earlier light-microscopic observations and suggests that the majority of the cellular pool of estrophilin exists in the nucleus of hormone-responsive cells. PMID:4020102

Press, M F; Nousek-Goebl, N A; Greene, G L

1985-09-01

260

Infectious Complications Associated with Monoclonal Antibodies and Related Small Molecules  

PubMed Central

Summary: Biologics are increasingly becoming part of routine disease management. As more agents are developed, the challenge of keeping track of indications and side effects is growing. While biologics represent a milestone in targeted and specific therapy, they are not without drawbacks, and the judicious use of these “magic bullets” is essential if their full potential is to be realized. Infectious complications in particular are not an uncommon side effect of therapy, whether as a direct consequence of the agent or because of the underlying disease process. With this in mind, we have reviewed and summarized the risks of infection and the infectious disease-related complications for all FDA-approved monoclonal antibodies and some related small molecules, and we discuss the probable mechanisms involved in immunosuppression as well as recommendations for prophylaxis and treatment of specific disease entities. PMID:19366915

Salvana, Edsel Maurice T.; Salata, Robert A.

2009-01-01

261

Novel CD20 monoclonal antibodies for lymphoma therapy  

PubMed Central

Rituximab (RTX), a monoclonal antibody (mAb) against CD20, has been widely used for lymphoma therapy. RTX in combination with cyclophosphamide /doxorubicin /vincristine /prednisone (R-CHOP) remains the standard frontline regimen for diffuse large B-cell lymphoma. However, suboptimal response and /or resistance to rituximab have remained a challenge in the therapy of B-cell non-Hodgkin’s lymphoma (NHL). Novel agents are under active clinical trials. This review will summarize the latest development in new mAbs against CD20, which include second-generation mAbs, ofatumumab, veltuzumab (IMMU-106), ocrelizumab (PRO70769), and third-generation mAbs, AME-133v (ocaratuzumab), PRO131921 and GA101 (obinutumumab). PMID:23057966

2012-01-01

262

Monoclonal antibody-induced cytokine-release syndrome.  

PubMed

Monoclonal antibodies (mAbs) are widely used in anti-inflammatory and tumor therapy. Although effective, mAbs can cause a variety of adverse effects. An important toxicity seen with a few mAbs is cytokine-release syndrome (CRS). These mAbs include: alemtuzumab, muromonab-CD3, rituximab, tosituzumab, CP-870,893, LO-CD2a/BTI-322 and TGN1412. By contrast, over 30 mAbs used clinically are not associated with CRS. In this review, the clinical aspects of CRS, the mAbs associated with CRS, the cytokines involved and putative mechanisms mediating cytokine release will be discussed. This will be followed by a discussion of the poor predictive value of studies in animals and the prospects for creating in vitro screens. Finally, approaches to decreasing the probability of CRS, decreasing the severity or treating CRS, should it occur, will be described. PMID:20477639

Bugelski, Peter J; Achuthanandam, Ram; Capocasale, Renold J; Treacy, George; Bouman-Thio, Esther

2009-09-01

263

Monoclonal antibodies and the transformation of blood typing.  

PubMed

Today, when monoclonal antibodies (mAbs) have become one of the most important classes of therapeutic drugs, it is easy to forget how much they have transformed our healthcare in other ways. One of the first clinical areas, as this paper shows, where mAbs made their mark was in the field of blood typing. The adoption of mAbs for this purpose was done with little public fanfare or funding. Nonetheless, it radically transformed the accuracy and cost of blood typing and shifted the procedure away from a dependence on reagents made from human blood donated by volunteers. This paper argues that the development of mAbs as reagents for blood typing laid the foundation for the first large-scale production of mAbs thereby paving the way to the advent of mAb diagnostics and therapeutics. PMID:25484059

Marks, Lara

2014-11-01

264

Neutralization of Yellow Fever Virus Studied Using Monoclonal and Polyclonal Antibodies  

Microsoft Academic Search

SUMMARY Monoclonal and polyclonal antibodies with known specificity for either the 54K envelope glycoprotein or the 48K non-structural glycoprotein of yellow fever (YF) virus-infected cells were studied in plaque reduction neutralization tests. Viruses employed in the tests comprised wild-type and vaccine strains of YF and a selection of other flaviviruses. Of 17 monoclonal antibodies examined, six of the 54K-specific antibodies

A. Buckley; E. A. Gould

1985-01-01

265

Monoclonal antibodies against rabbit mammary prolactin receptors. Specific antibodies to the hormone binding domain  

SciTech Connect

Three monoclonal antibodies (M110, A82, and A917) were obtained by fusing myeloma cells and spleen cells from mice immunized with partially purified rabbit mammary gland prolactin (PRL) receptors. All 3 antibodies were capable of complete inhibition of SVI-ovine prolactin (oPRL) binding to rabbit mammary PRL receptors in either particulate or soluble form. M110 showed slightly greater potency than oPRL in competing for SVI-oPRL binding. These antibodies also inhibited PRL binding to microsomal fractions from rabbit liver, kidney, adrenal, ovary, and pig mammary gland, although A82 showed poor inhibition in pig mammary gland. There was no cross-reaction of any of the 3 monoclonal antibodies (mAbs) for the other species tested: human (T-47D breast cancer cells) and rat (liver, ovary). In order to confirm that these antibodies are specific to the binding domain, antibodies were purified, iodinated, and binding characteristics were investigated. SVI-M110 and SVI-A82 binding was completely inhibited by lactogenic hormones, whereas nonlactogenic hormones did not cross-react. Competition of 125I-M110 by oPRL was comparable to that of SVI-oPRL by unlabeled oPRL, while SVI-A917 binding was only partially competed (30-60%) by lactogenic hormones. Tissue and species specificity of labeled antibody binding paralleled results of binding inhibition experiments using 125I-oPRL. In addition, A82 and A917 completely inhibited 125I-M110 binding. In contrast, 125I-A82 binding was stimulated by A917 and 125I-A917 binding was stimulated by A82.

Katoh, M.; Djiane, J.; Kelly, P.A.

1985-09-25

266

Detection of Rickettsia prowazekii in Body Lice and Their Feces by Using Monoclonal Antibodies  

PubMed Central

In order to identify Rickettsia prowazekii in lice, we developed a panel of 29 representative monoclonal antibodies selected from 187 positive hybridomas made by fusing splenocytes of immunized mice with SP2/0-Ag14 myeloma cells. Immunoblotting revealed that 15 monoclonal antibodies reacted with the lipopolysaccharide-like (LPS-L) antigen and 14 reacted with the epitopes of a 120-kDa protein. Only typhus group rickettsiae reacted with the monoclonal antibodies against LPS-L. R. felis, a recently identified rickettsial species, did not react with these monoclonal antibodies, confirming that it is not antigenically related to the typhus group. Monoclonal antibodies against the 120-kDa protein were highly specific for R. prowazekii. We successfully applied a selected monoclonal antibody against the 120-kDa protein to detect by immunofluorescence assay R. prowazekii in smears from 56 wild and laboratory lice, as well as in 10 samples of louse feces infected or not infected with the organism. We have developed a simple, practical, and specific diagnostic assay for clinical specimens and large-scale epidemiological surveys with a sensitivity of 91%. These monoclonal antibodies could be added to the rickettsial diagnostic panel and be used to differentiate R. prowazekii from other rickettsial species. PMID:12202579

Fang, Rong; Houhamdi, Linda; Raoult, Didier

2002-01-01

267

The history of monoclonal antibody development – Progress, remaining challenges and future innovations  

PubMed Central

As medicine progresses into a new era of personalised therapy, the use of monoclonal antibodies to treat a wide range of diseases lies at the heart of this new forefront. Since the licencing of the first monoclonal antibody for clinical use 30 years ago, the monoclonal antibody industry has expanded exponentially and is now valued at billions of dollars. With major advances in genetic sequencing and biomedical research, much research into monoclonal antibodies now focuses on identifying new targets for development and maximising their efficacy for use in clinical practice. However, a balance has to be struck with regards to reducing numbers of side-effects and overall economic cost, which arguably somewhat blighted their early clinical and commercial successes. Nowadays, there are approximately 30 monoclonal antibodies that have been approved for use in clinical practice with many more currently being tested in clinical trials. Some of the current major limitations include: the use of inefficient models for generation, a lack of efficacy and issues of cost-effectiveness. Some of the current research focuses on ways to improve the efficacy of existing monoclonal antibodies through optimising their effects and the addition of beneficial modifications. This review will focus on the history of monoclonal antibody development – how it has increasingly moved away from using laborious animal models to a more effective phage display system, some of the major drawbacks from a clinical and economical point of view and future innovations that are currently being researched to maximise their effectiveness for future clinical use.

Liu, Justin K.H.

2014-01-01

268

The history of monoclonal antibody development - Progress, remaining challenges and future innovations.  

PubMed

As medicine progresses into a new era of personalised therapy, the use of monoclonal antibodies to treat a wide range of diseases lies at the heart of this new forefront. Since the licencing of the first monoclonal antibody for clinical use 30 years ago, the monoclonal antibody industry has expanded exponentially and is now valued at billions of dollars. With major advances in genetic sequencing and biomedical research, much research into monoclonal antibodies now focuses on identifying new targets for development and maximising their efficacy for use in clinical practice. However, a balance has to be struck with regards to reducing numbers of side-effects and overall economic cost, which arguably somewhat blighted their early clinical and commercial successes. Nowadays, there are approximately 30 monoclonal antibodies that have been approved for use in clinical practice with many more currently being tested in clinical trials. Some of the current major limitations include: the use of inefficient models for generation, a lack of efficacy and issues of cost-effectiveness. Some of the current research focuses on ways to improve the efficacy of existing monoclonal antibodies through optimising their effects and the addition of beneficial modifications. This review will focus on the history of monoclonal antibody development - how it has increasingly moved away from using laborious animal models to a more effective phage display system, some of the major drawbacks from a clinical and economical point of view and future innovations that are currently being researched to maximise their effectiveness for future clinical use. PMID:25568796

Liu, Justin K H

2014-12-01

269

Monoclonal antibody-based therapies for microbial diseases  

PubMed Central

The monoclonal antibody (mAb) revolution that currently provides many new options for the treatment of neoplastic and inflammatory diseases has largely bypassed the field of infectious diseases. Only one mAb is licensed for use against an infectious disease, although there are many in various stages of development. This situation is peculiar given that serum therapy was one of the first effective treatments for microbial diseases and that specific antibodies have numerous antimicrobial properties. The underdevelopment and underutilization of mAb therapies for microbial diseases has various complex explanations that include the current availability of antimicrobial drugs, small markets, high costs and microbial antigenic variation. However, there are signs that the climate for mAb therapeutics in infectious diseases is changing given increasing antibiotic drug resistance, the emergence of new pathogenic microbes for which no therapy is available, and development of mAb cocktail formulations. Currently, the major hurdle for the widespread introduction of mAb therapies for microbial diseases is economic, given the high costs of immunoglobulin preparations and relatively small markets. Despite these obstacles there are numerous opportunities for mAb development against microbial diseases and the development of radioimmunotherapy provides new options for enhancing the magic bullet. Hence, there is cautious optimism that the years ahead will see more mAbs in clinical use against microbial diseases. PMID:20006139

Saylor, Carolyn; Dadachova, Ekaterina; Casadevall, Arturo

2009-01-01

270

Molecular mimicry by antiidiotypic monoclonal antibody to gonadotropin releasing hormone.  

PubMed

Antipeptide and antiidiotypic antibodies to several receptors are known to mimic their respective ligands in transducing signals on binding their receptors. In our attempts to study gonadotropin releasing hormone receptor, antipeptide and antiidiotypic monoclonal antibodies specific to the receptor were established earlier. The antipeptide mAb F1G4 was to a synthetic peptide corresponding to the extracellular domain of human GnRH receptor and the antiidiotypic mAb 4D10C1 was to the idiotype of a GnRH specific mAb. Here we report the physiological effects of the two mAbs on binding the receptor, as investigated using in vitro cultures of (a) human term placental villi and (b) rat pituitaries. The mAb 4D10C1 exerted a dose-dependent release of human chorionic gonadotropin in cultures of human term placental villi as well as luteinising and follicle stimulating hormones in cultures of rat pituitaries. PMID:10484676

Rajeshwari, K; Karande, A A

1999-01-01

271

A monoclonal antibody specifically reactive with Ewing's sarcoma.  

PubMed Central

We have developed a mouse monoclonal antibody 5C11 (IgG2a) against cell surface antigen of Ewing's sarcoma (ES). 5C11 specifically reacted with ESs but not with other small round cell tumours in childhood, i.e. neuroblastomas, primitive neuroectodermal tumours (PNETs), rhabdomyosarcomas and malignant lymphomas. 5C11 did not react with any other tumours in children except for hepatoblastomas. No reactivity has been identified in normal tissues with the exception of fetal hepatocytes. Immunoelectron microscopically, 5C11 reactive antigen was located on cell membrane of ES cells. Biochemically, 5C11 immunoprecipitated a cell surface protein having molecular weight of 81,000 Da. 5C11 is the first antibody which can clearly distinguish ES from neurogenic tumours, especially from PNETs which were recently reported to have common features to ESs regarding chromosal abnormality and proto-oncogene expression but show evident differentiation into neurogenic direction. The results strongly indicate the usefulness of 5C11 not only for diagnostic purpose when no specific marker is available but also for studying the histogenesis of ES. In addition, no reactivity in normal tissue implies its potential application as a therapeutic reagent when the management of ES patients is still a great problem in clinical field. Images Figure 1 Figure 2 Figure 4 PMID:2605097

Hara, S.; Ishii, E.; Tanaka, S.; Yokoyama, J.; Katsumata, K.; Fujimoto, J.; Hata, J.

1989-01-01

272

Macrophages eliminate circulating tumor cells after monoclonal antibody therapy.  

PubMed

The use of monoclonal antibodies (mAbs) as therapeutic tools has increased dramatically in the last decade and is now one of the mainstream strategies to treat cancer. Nonetheless, it is still not completely understood how mAbs mediate tumor cell elimination or the effector cells that are involved. Using intravital microscopy, we found that antibody-dependent phagocytosis (ADPh) by macrophages is a prominent mechanism for removal of tumor cells from the circulation in a murine tumor cell opsonization model. Tumor cells were rapidly recognized and arrested by liver macrophages (Kupffer cells). In the absence of mAbs, Kupffer cells sampled tumor cells; however, this sampling was not sufficient for elimination. By contrast, antitumor mAb treatment resulted in rapid phagocytosis of tumor cells by Kupffer cells that was dependent on the high-affinity IgG-binding Fc receptor (Fc?RI) and the low-affinity IgG-binding Fc receptor (Fc?RIV). Uptake and intracellular degradation were independent of reactive oxygen or nitrogen species production. Importantly, ADPh prevented the development of liver metastases. Tumor cell capture and therapeutic efficacy were lost after Kupffer cell depletion. Our data indicate that macrophages play a prominent role in mAb-mediated eradication of tumor cells. These findings may help to optimize mAb therapeutic strategies for patients with cancer by helping us to aim to enhance macrophage recruitment and activity. PMID:24430180

Gül, Nuray; Babes, Liane; Siegmund, Kerstin; Korthouwer, Rianne; Bögels, Marijn; Braster, Rens; Vidarsson, Gestur; ten Hagen, Timo L M; Kubes, Paul; van Egmond, Marjolein

2014-02-01

273

Defining process design space for monoclonal antibody cell culture.  

PubMed

The concept of design space has been taking root as a foundation of in-process control strategies for biopharmaceutical manufacturing processes. During mapping of the process design space, the multidimensional combination of operational variables is studied to quantify the impact on process performance in terms of productivity and product quality. An efficient methodology to map the design space for a monoclonal antibody cell culture process is described. A failure modes and effects analysis (FMEA) was used as the basis for the process characterization exercise. This was followed by an integrated study of the inoculum stage of the process which includes progressive shake flask and seed bioreactor steps. The operating conditions for the seed bioreactor were studied in an integrated fashion with the production bioreactor using a two stage design of experiments (DOE) methodology to enable optimization of operating conditions. A two level Resolution IV design was followed by a central composite design (CCD). These experiments enabled identification of the edge of failure and classification of the operational parameters as non-key, key or critical. In addition, the models generated from the data provide further insight into balancing productivity of the cell culture process with product quality considerations. Finally, process and product-related impurity clearance was evaluated by studies linking the upstream process with downstream purification. Production bioreactor parameters that directly influence antibody charge variants and glycosylation in CHO systems were identified. PMID:20589669

Abu-Absi, Susan Fugett; Yang, LiYing; Thompson, Patrick; Jiang, Canping; Kandula, Sunitha; Schilling, Bernhard; Shukla, Abhinav A

2010-08-15

274

Characterization of Zaire ebolavirus glycoprotein-specific monoclonal antibodies.  

PubMed

Zaire ebolavirus (ZEBOV) can be transmitted by human-to-human contact and causes acute haemorrhagic fever with case fatality rates up to 90%. There are no effective therapeutic or prophylactic treatments available. The sole transmembrane glycoprotein (GP) is the key target for developing neutralizing antibodies. In this study, recombinant VSV?G/ZEBOVGP was used to generate monoclonal antibodies (MAbs) against the ZEBOV GP. A total of 8 MAbs were produced using traditional hybridoma cell fusion technology, and then characterized by ELISA using ZEBOV VLPs, Western blotting, an immunofluorescence assay, and immunoprecipitation. All 8 MAbs worked in IFA and IP, suggesting that they are all conformational MAbs, however six of them recognized linearized epitopes by WB. ELISA results demonstrated that one MAb bound to a secreted GP (sGP 1-295aa); three bind to a part of the mucin domain (333-458aa); three MAbs recognized epitopes on the C-terminal domain of GP1 (296-501aa); and one bound to full length GP (VLPs/GP1,2 ?Tm). Using a mouse model these MAbs were evaluated for their therapeutic capacity during a lethal infection. All 8 MAb improved survival rates by 33%-100% against a high dose lethal challenge with mouse-adapted ZEBOV. This work has important implications for further development of vaccines and immunotherapies for ZEBOV infection. PMID:21925951

Qiu, Xiangguo; Alimonti, Judie B; Melito, P Leno; Fernando, Lisa; Ströher, Ute; Jones, Steven M

2011-11-01

275

An antifibrin monoclonal antibody useful in immunoscintigraphic detection of thrombi  

SciTech Connect

Balb/c mice were immunized with human plasmin-generated fibrinogen degradation product Y. Spleen cells were fused with P3X63-Ag8.653 myeloma cells. A clone (Y22) was found that produces monoclonal antibodies (MoAbs) with a strong reactivity with human fibrin and only a weak reactivity with fibrinogen in an enzyme immunoassay (EIA). Y22 also reacts with fibrin of rabbits, rats, sheep, and dogs. The antibodies are of the IgG1 kappa-type and appear to be directed against a conformation-dependent epitope in the D-domain of fibrin. Experiments with {sup 99m}Tc-labeled Y22 in vitro show that Y22 binds rapidly to forming clots. {sup 99}mTc-Y22 also binds to preformed plasma clots in a plasma milieu, even in the presence of high concentrations of heparin. Clot localization experiments in rabbits and rats confirm the high fibrin specificity and the potential of {sup 99m}Tc-Y22 for thrombus imaging in vivo.

Wasser, M.N.; Koppert, P.W.; Arndt, J.W.; Emeis, J.J.; Feitsma, R.I.; Pauwels, E.K.; Nieuwenhuizen, W. (Gaubius Institute TNO, Leiden (Netherlands))

1989-08-01

276

Monoclonal antibodies: Pharmacokinetics as a basis for new dosage regimens?  

PubMed

Complete monoclonal IgG antibodies which are in use in clinical practice share some pharmacological properties resulting in high concentrations in plasma. This fact is reflected in their low volumes of distribution, which can also be correlated with a high molecular weight and water solubility. This feature allows a novel approach to be applied to the dosing schedule for this group of drugs with fixed doses being used instead of the initially developed weight- or body surface-adjusted dosing schedules. In addition, the development of a new formulation containing hyaluronidase allows a subcutaneous route of administration to be used, because hyaluronidase creates a space in the subcutaneous tissue that helps antibody absorption. This method requires higher doses, but has allowed testing the feasibility of administering a fixed dose, with no individual dose adjustments based on weight or body surface. Moreover, loading doses are not needed, because the first dose results, within 3 weeks, in minimum concentrations that are higher than effective concentrations. PMID:24903270

Azanza, J-R; Sádaba, B; Gómez-Guiu, A

2014-06-01

277

Radiolabeled monoclonal antibodies for imaging and therapy: Potential, problems, and prospects: Scientific highlights  

SciTech Connect

This meeting focused on areas of research on radiolabeled monoclonal antibodies. Topics covered included the production, purification, and fragmentation of monoclonal antibodies and immunochemistry of hybridomas; the production and the chemistry of radionuclides; the radiohalogenation and radiometal labeling techniques; the in-vivo pharmacokinetics of radiolabeled antibodies; the considerations of immunoreactivity of radiolabeled preparations; the instrumentation and imaging techniques as applied to radioimmunodetection; the radiation dosimetry in diagnostic and therapeutic use of labeled antibodies; the radioimmunoscintigraphy and radioimmunotherapy studies; and perspectives and directions for future research. Tutorial as well as scientific lectures describing the latest research data on the above topics were presented. Three workshop panels were convened on ''Methods for Determining Immunoreactivity of Radiolabeled Monoclonal Antibodies - Problems and Pitfalls,'' Radiobiological and Dosimetric Considerations for Immunotherapy with Labeled Antibodies,'' and ''The Human Anti-Mouse Antibody Response in Patients.''

Srivastava, S.C.; Buraggi, G.L.

1986-01-01

278

Structure of solid tumors and their vasculature: Implications for therapy with monoclonal antibodies  

SciTech Connect

Delivery of monoclonal antibodies to solid tumors is a vexing problem that must be solved if these antibodies are to realize their promise in therapy. Such success as has been achieved with monoclonal antibodies is attributable to the local hyperpermeability of the tumor vasculature, a property that favors antibody extravasation at tumor sites and that is mediated by a tumor-secreted vascular permeability factor. However, leaky tumor blood vessels are generally some distance removed from target tumor cells, separated by stroma and by other tumor cells that together represent significant barriers to penetration by extravasated monoclonal antibodies. For this reason, alternative approaches may be attractive. These include the use of antibody-linked cytotoxins, which are able to kill tumor cells without immediate contact, and direction of antibodies against nontumor cell targets, for example, antigens unique to the tumor vascular endothelium or to tumor stroma. 50 refs.

Dvorak, H.F.; Nagy, J.A.; Dvorak, A.M. (Beth Israel Hospital, Boston, MA (USA))

1991-03-01

279

Melioidosis in a patient on monoclonal antibody therapy for psoriatic arthritis.  

PubMed

Melioidosis is caused by the environmental bacterium Burkholderia pseudomallei and can present with severe sepsis. Predisposing risk factors are present in 80% of cases. Monoclonal antibodies are increasingly prescribed for varied medical conditions. This report describes the first known case of melioidosis in a patient whose only risk factor for disease is treatment with a monoclonal antibody. Prescribers of monoclonal antibodies and other immunosuppressants should ensure that their patients are aware of the potential risk of melioidosis prior to travel and the precautions that should be taken. PMID:25442759

Commons, R J; Grivas, R; Currie, B J

2014-12-01

280

Monoclonal antibodies neutralizing the haemolytic activity of box jellyfish (Chironex fleckeri) tentacle extracts.  

PubMed

1. Three monoclonal antibodies have been produced which neutralize in vitro the haemolytic activity present in tentacle extracts of the box jellyfish (Chironex fleckeri). 2. Two of these monoclonal antibodies bound specifically to a component of relative molecular mass 50,000 in tentacle extract on Western blots. 3. This binding only occurred when the extracts were electrophoresed under non-reducing conditions. 4. The third monoclonal antibody did not display binding to Western blots of tentacle extract under any of our experimental conditions. PMID:8104761

Collins, S P; Comis, A; Marshall, M; Hartwick, R F; Howden, M E

1993-09-01

281

Kinetic analysis of the multistep aggregation mechanism of monoclonal antibodies.  

PubMed

We investigate by kinetic analysis the aggregation mechanism of two monoclonal antibodies belonging to the IgG1 and IgG2 subclass under thermal stress. For each IgG, we apply a combination of size exclusion chromatography and light scattering techniques to resolve the time evolution of the monomer, dimer, and trimer concentrations, as well as the average molecular weight and the average hydrodynamic radius of the aggregate distribution. By combining the detailed experimental characterization with a theoretical kinetic model based on population balance equations, we extract relevant information on the contribution of the individual elementary steps on the global aggregation process. The analysis shows that the two molecules follow different aggregation pathways under the same operating conditions. In particular, while the monomer depletion of the IgG1 is found to be rate-limited by monomeric conformational changes, bimolecular collision is identified as the rate-limiting step in the IgG2 aggregation process. The measurement of the microscopic rate constants by kinetic analysis allows the quantification of the protein-protein interaction potentials expressed in terms of the Fuchs stability ratio (W). It is found that the antibody solutions exhibit large W values, which are several orders of magnitude larger than the values computed in the frame of the DLVO theory. This indicates that, besides net electrostatic repulsion, additional effects delay the aggregation kinetics of the antibody solutions with respect to diffusion-limited conditions. These effects likely include the limited efficiency of the collision events due to the presence of a limited number of specific aggregation-prone patches on the heterogeneous protein surface, and the contribution of additional repulsive non-DLVO forces to the protein-protein interaction potential, such as hydration forces. PMID:25119992

Nicoud, Lucrčce; Arosio, Paolo; Sozo, Margaux; Yates, Andrew; Norrant, Edith; Morbidelli, Massimo

2014-09-11

282

Development of new staining technology "eastern blotting" using monoclonal antibody.  

PubMed

Ginsenosides contained in Panax species were separated by silica gel TLC blotted to a polyvinylidene difluoride (PVDF) membrane which was dipped in a sodium periodide (NaIO(4)) solution and reacted with protein, preparing a ginsenoside-protein conjugate for binding a ginsenoside on a PVDF membrane. The blotted spots were stained by anti-ginsenoside-Rb1 monoclonal antibody (MAb) and anti-ginsenoside-Rg1MAb, respectively. The newly established immunostaining method, eastern blotting was applied for the determination of ginsenosides possessing protopanaxadiol and/or protopanaxatriol. Double staining of eastern blotting for ginsenosides using anti-ginsenoside-Rb1 MAb and anti-ginsenoside-Rg1 MAb promoted complete identification of ginsenosides in Panax species. This technique has been devised for the chromatographic separation and identification of ginsenosides using polyethersulfone (PES) membrane. It caused an acceptable separation of ginsenoside-Rb1, -Rc and -Rd in various ginseng extracts. Newly developed technique is quite simple and applies for immunoassay system. Ginsenosides separated using a PES membrane were directly treated with a NaIO(4) solution and then reacted with bovine serum albumin (BSA) for making a ginsenoside-protein conjugate. After the blocking, anti-ginsenoside-Rb1 MAb recognized a ginsenoside on a PES membrane and then a sec-ond antibody labeled with enzyme reacted to the first antibody. Finally a substrate was oxidized with the enzyme and de-veloped the staining of ginsenosides. The staining spots of ginsenosides on membrane were quantitatively evaluated by NIH Image indicating at least 62.5 ng of each ginsenoside-Rb1, -Rc and -Rd were detected with clarity. The determination range of three ginsenosides was from 0.125 to 2.0 µg of direct amount on PES membrane. PMID:21143134

Morinaga, Osamu; Shoyama, Yukihiro

2011-03-01

283

Phase I clinical trial of the bispecific antibody MDX-H210 (anti-FcgammaRI x anti-HER-2/neu) in combination with Filgrastim (G-CSF) for treatment of advanced breast cancer.  

PubMed

A phase I study of the bispecific antibody MDX-H210 in combination with granulocyte colony-stimulating factor (G-CSF) was performed in stage IV breast carcinoma patients, overexpressing HER-2/neu. MDX-H210, constructed by crosslinking antigen binding fragments (F(ab') fragments) of monoclonal antibody (mAb) H22 to Fc gamma receptor I (FcgammaRI), and mAb 520C9 to HER-2/neu, respectively, mediates the lysis of tumour cells in vitro, and in human FcgammaRI transgenic mouse models. The proto-oncogene HER-2/neu is overexpressed in approximately 30% of breast cancer patients, and represents a promising target for antibody-based immunotherapy. Fc gamma receptor I (CD64) is an effective trigger molecule, which is expressed on monocytes/macrophages, immature dendritic cells, and G-CSF-primed polymorphonuclear cells (PMN). Patients received G-CSF (Filgrastim) for 8 consecutive days, and cohorts of three patients were treated on day 4 with escalating, single doses of MDX-H210. A total of 30 patients were included, and treatment was generally well tolerated, without reaching dose-limiting toxicity. Side effects consisted mainly of fever and short periods of chills, which were timely related to elevated plasma levels of interleukin 6 and tumour necrosis factor alpha. In the last two cohorts, MDX-H210 plasma levels exceeded 1 microg ml(-1), and on circulating myeloid cells >50% saturation of FcgammaRI was found until day 4. These effector cells were highly effective in antibody-dependent cell-mediated cytotoxicity. Immunohistochemical analyses of tumour biopsies in individual patients documented infiltration of monocytes and PMN after MDX-H210 infusion. Although the clinical course of the disease was not altered by the single dose of MDX-H210, a favourable toxicity profile--even at high doses--and remarkable biological effects were seen when combined with G-CSF. Therefore, the combination of G-CSF and MDX-H210 should be evaluated in further immunotherapeutical strategies. PMID:14676800

Repp, R; van Ojik, H H; Valerius, T; Groenewegen, G; Wieland, G; Oetzel, C; Stockmeyer, B; Becker, W; Eisenhut, M; Steininger, H; Deo, Y M; Blijham, G H; Kalden, J R; van de Winkel, J G J; Gramatzki, M

2003-12-15

284

Different rhodopsin monoclonal antibodies reveal different binding patterns on developing and adult rat retina  

Microsoft Academic Search

We used a battery of 10 monoclonal antibodies directed against different identified peptide sequences within the carboxyl, transmembrane loop, and amino terminal regions of rhodopsin to label retinas from early postnatal and adult rats. Intensity of label, age of initial appearance of staining, and distribution of label varied depending on the antibody. Most antibodies showed detectable labeling at postnatal day

DAVID HICKS; COLIN J. BARNSTABLE

1987-01-01

285

Development of Oligodendrocytes and Schwann Cells Studied with a Monoclonal Antibody against Galactocerebroside  

Microsoft Academic Search

We have generated a hybridoma cell line secreting a monoclonal antibody that specifically binds to the surfaces of oligodendrocytes and Schwann cells, the cells involved in myelin formation in the central and peripheral nervous systems, respectively. Binding studies using purified sphingolipids showed that this antibody reacts strongly with galactocerebroside (GalC), the major galactosphingolipid of myelin. The antibody was used in

B. Ranscht; P. A. Clapshaw; J. Price; M. Noble; W. Seifert

1982-01-01

286

Effective lysis of lymphoma cells with a stabilised bispecific single-chain Fv antibody against CD19 and FcgammaRIII (CD16).  

PubMed

A recombinant bispecific single-chain fragment variable antibody (bsscFv), directed against the B-cell antigen CD19 and the low affinity Fc-receptor FcgammaRIII (CD16), was designed for use in the treatment of patients with leukaemias and lymphomas. The Fc-portions of whole antibodies were deliberately eliminated in this construct to avoid undesired effector functions. A stabilised bsscFv, ds[CD19 x CD16], was generated, in which disulphide bonds bridging the respective variable light (VL) and variable heavy (VH) chains were introduced into both component single-chain (sc)Fvs. After production in 293T cells and chromatographic purification, ds[CD19 x CD16] specifically and simultaneously bound both antigens. The serum stability of ds[CD19 x CD16] was increased more than threefold when compared with the unstabilised counterpart, while other biological properties were not affected by these mutations. In antibody-dependent cellular cytotoxicity experiments, ds[CD19 x CD16] mediated specific lysis of both CD19-positive malignant human B-lymphoid cell lines and primary tumour cells from patients with B-cell chronic lymphocytic leukaemia or B-cell acute lymphoblastic leukaemia. Natural killer cells, mononuclear cells (MNCs) from healthy donors and, in some instances, MNCs isolated from patients after allogeneic stem cell transplantation, were used as effectors. Thus, ds[CD19 x CD16] holds promise for the treatment of CD19(+) B-lineage malignancies. PMID:16029450

Bruenke, Joerg; Barbin, Karin; Kunert, Susanne; Lang, Peter; Pfeiffer, Matthias; Stieglmaier, Kristin; Niethammer, Dietrich; Stockmeyer, Bernhard; Peipp, Matthias; Repp, Roland; Valerius, Thomas; Fey, Georg H

2005-07-01

287

T-cell activation and cytokine production via a bispecific single-chain antibody fragment targeted to blood-stage malaria parasites.  

PubMed

A novel bispecific single-chain antibody fragment (biscFv) has been constructed to address the possibility of a new approach to malaria therapeutic drug development. The biscFv consists of 2 different single-chain antibody fragments linked by a flexible peptide linker (Gly(4)-Ser)(3). Of the 2 scFv fragments, one is directed against a conserved epitope of the 19-kDa C-terminal fragment of the major surface protein of human malignant malaria parasite, Plasmodium falciparum, and the other is directed against the CD3 antigen of human T cells. The biscFv expressed by a recombinant baculovirus retained the antigen-binding properties of the corresponding univalent single-chain antibody fragments and formed a bridge between P falciparum and T cells. In cooperation with T cells, the biscFv specifically induced not only interferon gamma and tumor necrosis factor alpha, but also a significant increase of merozoite phagocytosis and growth inhibition of P falciparum in vitro. Thus, the biscFv possesses highly selective malaria-targeting properties and stimulates T cells to induce cytokines, presumably resulting in activation of macrophages, neutrophils, and natural killer cells, and parasite killing in vivo. PMID:12411309

Yoshida, Shigeto; Kobayashi, Tominari; Matsuoka, Hiroyuki; Seki, Chisato; Gosnell, William L; Chang, Sandra P; Ishii, Akira

2003-03-15

288

Rearranging the domain order of a diabody-based IgG-like bispecific antibody enhances its antitumor activity and improves its degradation resistance and pharmacokinetics.  

PubMed

One approach to creating more beneficial therapeutic antibodies is to develop bispecific antibodies (bsAbs), particularly IgG-like formats with tetravalency, which may provide several advantages such as multivalent binding to each target antigen. Although the effects of configuration and antibody-fragment type on the function of IgG-like bsAbs have been studied, there have been only a few detailed studies of the influence of the variable fragment domain order. Here, we prepared four types of hEx3-scDb-Fc, IgG-like bsAbs, built from a single-chain hEx3-Db (humanized bispecific diabody [bsDb] that targets epidermal growth factor receptor and CD3), to investigate the influence of domain order and fusion manner on the function of a bsDb with an Fc fusion format. Higher cytotoxicities were observed with hEx3-scDb-Fcs with a variable light domain (VL)-variable heavy domain (VH) order (hEx3-scDb-Fc-LHs) compared with a VH-VL order, indicating that differences in the Fc fusion manner do not affect bsDb activity. In addition, flow cytometry suggested that the higher cytotoxicities of hEx3-scDb-Fc-LH may be attributable to structural superiority in cross-linking. Interestingly, enhanced degradation resistance and prolonged in vivo half-life were also observed with hEx3-scDb-Fc-LH. hEx3-scDb-Fc-LH and its IgG2 variant exhibited intense in vivo antitumor effects, suggesting that Fc-mediated effector functions are dispensable for effective anti-tumor activities, which may cause fewer side effects. Our results show that merely rearranging the domain order of IgG-like bsAbs can enhance not only their antitumor activity, but also their degradation resistance and in vivo half-life, and that hEx3-scDb-Fc-LHs are potent candidates for next-generation therapeutic antibodies. PMID:24983585

Shimomura, Ippei; Konno, Shota; Ito, Akiko; Masakari, Yosuke; Orimo, Ryota; Taki, Shintaro; Arai, Kyoko; Ogata, Hiromi; Okada, Mai; Furumoto, Shozo; Onitsuka, Masayoshi; Omasa, Takeshi; Hayashi, Hiroki; Katayose, Yu; Unno, Michiaki; Kudo, Toshio; Umetsu, Mitsuo; Kumagai, Izumi; Asano, Ryutaro

2014-07-01

289

Rearranging the domain order of a diabody-based IgG-like bispecific antibody enhances its antitumor activity and improves its degradation resistance and pharmacokinetics.  

PubMed

One approach to creating more beneficial therapeutic antibodies is to develop bispecific antibodies (bsAbs), particularly IgG-like formats with tetravalency, which may provide several advantages such as multivalent binding to each target antigen. Although the effects of configuration and antibody-fragment type on the function of IgG-like bsAbs have been studied, there have been only a few detailed studies of the influence of the variable fragment domain order. Here, we prepared four types of hEx3-scDb-Fc, IgG-like bsAbs, built from a single-chain hEx3-Db (humanized bispecific diabody [bsDb] that targets epidermal growth factor receptor and CD3), to investigate the influence of domain order and fusion manner on the function of a bsDb with an Fc fusion format. Higher cytotoxicities were observed with hEx3-scDb-Fcs with a variable light domain (VL)-variable heavy domain (VH) order (hEx3-scDb-Fc-LHs) compared with a VH-VL order, indicating that differences in the Fc fusion manner do not affect bsDb activity. In addition, flow cytometry suggested that the higher cytotoxicities of hEx3-scDb-Fc-LH may be attributable to structural superiority in cross-linking. Interestingly, enhanced degradation resistance and prolonged in vivo half-life were also observed with hEx3-scDb-Fc-LH. hEx3-scDb-Fc-LH and its IgG2 variant exhibited intense in vivo antitumor effects, suggesting that Fc-mediated effector functions are dispensable for effective anti-tumor activities, which may cause fewer side effects. Our results show that merely rearranging the domain order of IgG-like bsAbs can enhance not only their antitumor activity, but also their degradation resistance and in vivo half-life, and that hEx3-scDb-Fc-LHs are potent candidates for next-generation therapeutic antibodies. PMID:25517309

Asano, Ryutaro; Shimomura, Ippei; Konno, Shota; Ito, Akiko; Masakari, Yosuke; Orimo, Ryota; Taki, Shintaro; Arai, Kyoko; Ogata, Hiromi; Okada, Mai; Furumoto, Shozo; Onitsuka, Masayoshi; Omasa, Takeshi; Hayashi, Hiroki; Katayose, Yu; Unno, Michiaki; Kudo, Toshio; Umetsu, Mitsuo; Kumagai, Izumi

2014-09-01

290

Bispecific antibodies directed to CD4 domain 2 and HIV envelope exhibit exceptional breadth and picomolar potency against HIV-1  

PubMed Central

In the absence of an effective HIV-1 vaccine, passive immunization using broadly neutralizing Abs or Ab-like molecules could provide an alternative to the daily administration of oral antiretroviral agents that has recently shown promise as preexposure prophylaxis. Currently, no single broadly neutralizing Ab (bNAb) or combination of bNAbs neutralizes all HIV-1 strains at practically achievable concentrations in vivo. To address this problem, we created bispecific Abs that combine the HIV-1 inhibitory activity of ibalizumab (iMab), a humanized mAb directed to domain 2 of human CD4, with that of anti-gp120 bNAbs. These bispecific bNAbs (BibNAbs) exploit iMab’s potent anti–HIV-1 activity and demonstrated clinical efficacy and safety to anchor and thereby concentrate a second broadly neutralizing agent at the site of viral entry. Two BibNabs, PG9-iMab and PG16-iMab, exhibit exceptional breadth and potency, neutralizing 100% of the 118 viruses tested at low picomolar concentrations, including viruses resistant to both parental mAbs. The enhanced potency of these BibNAbs was entirely dependent on CD4 anchoring, not on membrane anchoring per se, and required optimal Ab geometry and linker length. We propose that iMab-based BibNAbs, such as PG9-iMab and PG16-iMab, are promising candidates for passive immunization to prevent HIV-1 infection. PMID:23878231

Pace, Craig S.; Song, Ruijiang; Ochsenbauer, Christina; Andrews, Chasity D.; Franco, David; Yu, Jian; Oren, Deena A.; Seaman, Michael S.; Ho, David D.

2013-01-01

291

Will immunogenicity limit the use, efficacy, and future development of therapeutic monoclonal antibodies?  

PubMed Central

While monoclonal antibodies show promise for use in the treatment of a variety of disease states, including cancer, autoimmune disease, and allograft rejection, generation of anti-antibody responses still remains a problem. For example, 50% of the patients who receive OKT3 produce blocking antibodies that interfere with its binding to T cells, thus decreasing the therapeutic effect (51). HAMA responses have also interfered with tumor imaging (39,40) and radioimmunotherapy (56). The generation of an anti-antibody response is dependent on many factors. These include the dose of antibody, the number of injections of antibody, the immunogenicity of the antibody, the form of the antibody, and the immunocompetence of the recipient. Predictably, both the number of injections of antibody and the dosage are influential in the generation of an anti-antibody response. It is apparent that human antibodies, chimeric antibodies, and mouse Fab fragments are much less likely to induce anti-antibody responses than intact mouse monoclonal antibodies or mouse F(ab')2 fragments when one injection is administered. Injections of human or chimeric antibodies appears to reduce immunogenicity, but the probability that anti-antibody responses can still be induced on multiple injections must be considered and appropriately evaluated. Several areas demand extensive investigation to enhance the clinical utility of monoclonal antibodies. First, results of thorough clinical trials with human or chimeric antibodies need to be evaluated for the induction of anti-antibodies after multiple injections of antibodies. Second, less immunogenic forms of antibodies (Fab, Fv) need to be studied for their clinical efficacies and for their abilities to induce anti-antibody responses. PMID:8556470

Kuus-Reichel, K; Grauer, L S; Karavodin, L M; Knott, C; Krusemeier, M; Kay, N E

1994-01-01

292

Localization of a tumor cell adhesion domain of laminin by a monoclonal antibody  

SciTech Connect

Monoclonal antibodies were prepared to localize the domain(s) of laminin to which tumor cells adhere. Rat Y3-Ag 1.2.3 myeloma cells were fused with spleen cells from a rat immunized with a purified 440-kDa fragment of chymotrypsin-digested laminin. Three monoclonal antibodies (AL-1 to AL-3) that bound to intact laminin in a solid-phase radioimmunoassay were chosen for further analysis. The epitopes recognized by these antibodies were characterized by radioimmunoassays, immunoblotting, radioimmunoprecipitation, and immunoaffinity chromatography. In cell adhesion assays, monoclonal antibody AL-2 inhibited the highly metastatic melanoma cell line, K-1735-M4, to both intact laminin and the 440-kDa fragment of laminin. Electron microscopic examination of laminin-monoclonal antibody interactions showed that monoclonal antibody AL-2 reacted with the long arm of laminin directly below the cross-region. Two monoclonal antibodies that failed to inhibit tumor cell adhesion to laminin reacted with epitopes on the lateral short arms or cross-region of laminin as seen by electron microscopy. These results suggest that a new tumor cell binding domain of laminin may be located close to the cross-region on the long arm of laminin.

Skubitz, A.P.N.; Charonis, A.S.; Tsilibary, E.C.; Furcht, L.T. (Univ. of Minnesota, Minneapolis (United States))

1987-12-01

293

Monoclonal rat antibodies directed against Toxoplasma gondii suitable for studying tachyzoite-bradyzoite interconversion in vivo.  

PubMed Central

We previously reported the in vitro analysis of stage differentiation of Toxoplasma gondii in murine bone marrow-derived macrophages. The purpose of this study was to generate monoclonal rat antibodies that might be suitable for investigating tachyzoite-bradyzoite interconversion in vivo with the murine model. Immunization of Fischer rats with cysts of T. gondii NTE resulted in the generation of seven monoclonal antibodies of the immunoglobulin G2a, G2b, or M isotype, which were further characterized by the immunoblot technique, immunofluorescence assay, immunohistology, and immunoelectron microscopy. Immunoblots demonstrated specific reactivity of five monoclonal antibodies with proteins with molecular masses of 40, 52, 55, 60, 64, 65, and 115 kDa. One antibody (CC2) appeared to recognize a differently expressed antigen depending on the parasite stage, reacting with a 40-kDa molecule in tachyzoites and a 115-kDa antigen in bradyzoites and oocysts. Several other monoclonal antibodies were shown to be stage specific and to react in immunofluorescence assays or in immunoblots with either tachyzoites or bradyzoites. Kinetics of stage conversion in vitro could be monitored by immunofluorescence with two of these monoclonal antibodies. Preliminary immunohistological investigations of tissue sections from infected mice demonstrated the possible usefulness of these monoclonal antibodies for future in vivo studies on stage differentiation of T. gondii in the murine system. PMID:8548532

Gross, U; Bormuth, H; Gaissmaier, C; Dittrich, C; Krenn, V; Bohne, W; Ferguson, D J

1995-01-01

294

The application of monoclonal antibody panels to characterize pestivirus isolates from ruminants in Great Britain  

Microsoft Academic Search

Summary Monoclonal antibodies were prepared against bovine virus diarrhoea virus and hog cholera virus. They were used to test 101 field isolates of ruminant pestivirus in a simple binding assay using an indirect immunoperoxidase label on fixed cell cultures. The monoclonals were divided into three panels: (1) pestivirus group specific, (2) hog cholera specific, (3) selectively reactive with ruminant pestiviruses.

S. Edwards; J. J. Sands; J. W. Harkness

1988-01-01

295

Comparison of the growth and monoclonal antibody production of suspended mammalian cells in three perfusion systems  

E-print Network

The purpose of this thesis was to provide a broad survey of bioprocess options for typical drug production vehicles in the biotechnology industry. This goal was accomplished by comparing the growth and monoclonal antibody ...

Hufford, Kathy (Kathy E.)

2007-01-01

296

78 FR 7438 - Prospective Grant of Exclusive License: Development of Human Monoclonal Antibodies Against DR4  

Federal Register 2010, 2011, 2012, 2013, 2014

...DEPARTMENT OF HEALTH AND HUMAN SERVICES National Institutes of Health...Grant of Exclusive License: Development of Human Monoclonal Antibodies Against DR4 AGENCY...Institutes of Health, Department of Health and Human Services, is contemplating the grant...

2013-02-01

297

Specific monoclonal antibodies against embryonic chicken adipose tissue cell membranes and their application against developing adipocytes  

E-print Network

Use of anti-adipocyte monoclonal antibodies (MAb) for reduction of fat mass in chickens was investigated in two experiments. Reduction of adipose tissue mass was obtained only in Experiment II. In Experiment I, 132 fertilized broiler chicken eggs...

Corcoran, Melinda Valdez

2002-01-01

298

Phase separation in solutions of monoclonal antibodies and the effect of human serum albumin  

E-print Network

We report the observation of liquid-liquid phase separation in a solution of human monoclonal antibody, IgG2, and the effects of human serum albumin, a major blood protein, on this phase separation. We find a significant ...

Wang, Ying

299

Tactical planning optimization for campaign scheduling of active pharmaceutical ingredient production based on monoclonal antibodies  

E-print Network

Monoclonal Antibodies (mAb's) are the fastest growing segment in the biopharmaceutical industry. They are used today as therapeutics and diagnostics for several medical applications, including various types of cancer, ...

Assia, Shai

2014-01-01

300

Monoclonal Antibody 14F7, Which Recognizes a Stage-Specific Immature Oligodendrocyte  

E-print Network

Monoclonal Antibody 14F7, Which Recognizes a Stage-Specific Immature Oligodendrocyte Surface Molecule, Inhibits Oligodendrocyte Differentiation Mediated in Co-Culture With Astrocytes Kazunori Institute of Medical Science, Tokyo, Japan Cells at an intermediate stage of oligodendrocyte lineage

Kawato, Suguru

301

Non-antigen-contacting region of an asymmetric bispecific antibody to factors IXa/X significantly affects factor VIII-mimetic activity.  

PubMed

While antibody engineering improves the properties of therapeutic antibodies, optimization of regions that do not contact antigens has been mainly focused on modifying the effector functions and pharmacokinetics of antibodies. We recently reported an asymmetric anti-FIXa/FX bispecific IgG4 antibody, ACE910, which mimics the cofactor function of FVIII by placing the two factors into spatial proximity for the treatment of hemophilia A. During the optimization process, we found that the activity was significantly affected by IgG subclass and by modifications to the inter-chain disulfide bonds, upper hinge region, elbow hinge region, and Fc glycan, even though these regions were unlikely to come into direct contact with the antigens. Of these non-antigen-contacting regions, the tertiary structure determined by the inter-chain disulfide bonds was found to strongly affect the FVIII-mimetic activity. Interestingly, IgG4-like disulfide bonds between Cys131 in the heavy chain and Cys114 in the light chain, and disulfide bonds between the two heavy chains at the hinge region were indispensable for the high FVIII-mimetic activity. Moreover, proline mutations in the upper hinge region and removal of the Fc glycan enhanced the FVIII-mimetic activity, suggesting that flexibility of the upper hinge region and the Fc portion structure are important for the FVIII-mimetic activity. This study suggests that these non-antigen-contacting regions can be engineered to improve the biological activity of IgG antibodies with functions similar to ACE910, such as placing two antigens into spatial proximity, retargeting effector cells to target cells, or co-ligating two identical or different antigens on the same cell. PMID:25524207

Sampei, Zenjiro; Igawa, Tomoyuki; Soeda, Tetsuhiro; Funaki, Miho; Yoshihashi, Kazutaka; Kitazawa, Takehisa; Muto, Atsushi; Kojima, Tetsuo; Nakamura, Satoshi; Hattori, Kunihiro

2015-01-01

302

Assignment of a fibronectin gene to human chromosome 2 using monoclonal antibodies  

SciTech Connect

The locus coding for the presumed structural gene for fibronectin has been mapped to human chromosome 2 using human-mouse somatic cell hybrids. The assignment of fibronectin has been made by testing man-mouse somatic cell hybrids with two anti-human fibronectin monoclonal antibodies which recognize different antigenic determinants of human, but not mouse, fibronectin. Both monoclonal antibodies demonstrate a highly concordant association between the presence of two different human fibronectin antigens and human chromosome 2.

Koch, G.A. (Roswell Park Memorial Inst., Buffalo, NY); Schoen, R.C.; Klebe, R.J.; Shows, T.B.

1982-10-01

303

Morphogenesis of the atrichous isorhiza, a type of nematocyst, in Hydra observed with a monoclonal antibody  

Microsoft Academic Search

We produced a monoclonal antibody, AE03, which recognized mucous granules in the basal disk gland cells in Hydra and the secreted mucus with which they stick onto substrate. AE03 also recognized atrichous isorhizas, one of the four types\\u000a of nematocyst present in tentacles, and their nematoblasts present in the body column. With this monoclonal antibody, we could\\u000a observe the detailed

H. Amano; Osamu Koizumi; Y. Kobayakawa

1997-01-01

304

The use of monoclonal antibody for the immunodiagnosis of Fasciola gigantica infection in cattle  

Microsoft Academic Search

Antigens that were specific to Fasciola gigantica were obtained from the whole worm homogenate of the parasite by immunoaffinity chromatography in cyanogen bromide-activated sepharose 4B columns and used for the production of monoclonal antibodies. The F. gigantica-specific monoclonal antibody was labelled with horseradish peroxidase and used for the detection of circulating antigen by the direct ELISA method in the sera

B. O. Fagbemi; O. A. Aderibigbe; E. E. Guobadia

1997-01-01

305

Assignment of a fibronectin gene to human chromosome 2 using monoclonal antibodies  

Microsoft Academic Search

The locus coding for the presumed structural gene for fibronectin has been mapped to human chromosome 2 using human-mouse somatic cell hybrids. The assignment of fibronectin has been made by testing man-mouse somatic cell hybrids with two anti-human fibronectin monoclonal antibodies which recognize different antigenic determinants of human, but not mouse, fibronectin. Both monoclonal antibodies demonstrate a highly concordant association

G. A. Koch; R. C. Schoen; R. J. Klebe; T. B. Shows

1982-01-01

306

A monoclonal antibody against GBM heparan sulfate induces an acute selective proteinuria in rats  

Microsoft Academic Search

A monoclonal antibody against GBM heparan sulfate induces an acute selective proteinuria in rats. After immunization of mice with partially-purified heparan sulfate proteoglycan (HSPG) isolated from rat glomeruli, a monoclonal antibody (mAb JM-403) was obtained, which was directed against heparan sulfate (HS), the glycosaminoglycan side chain of HSPG. In ELISA it reacted with isolated human glomerular basement membrane (GBM) HSPG,

Jacob van den Born; Lambert P W J van den Heuvel; Marinka A H Bakker; Jacques H Veerkamp; Karel J M Assmann; Jo H M Berden

1992-01-01

307

Altered anionic GBM components in monoclonal antibody against slit diaphragm-injected proteinuric rats  

Microsoft Academic Search

Altered anionic GBM components in monoclonal antibody against slit diaphragm injected proteinuric rats.BackgroundWe previously reported that monoclonal antibody (mAb) 5-1-6 bound to renal filtration slits induces massive proteinuria without causing ultrastructural changes in the glomerulus. This study evaluated the underlying mechanisms of the increase in glomerular permeability.MethodsThe distribution of endogenous albumin and IgG in the glomerular basement membrane (GBM) was

Yoshihide Fujigaki; Mitsumasa Nagase; Sumi Hidaka; Katsuyuki Matsui; Mikiko Shirai; Hitonari Nosaka; Hiroshi Kawachi; Fujio Shimizu; Akira Hishida

1998-01-01

308

Directed Selection of Recombinant Human Monoclonal Antibodies to Herpes Simplex Virus Glycoproteins from Phage Display Libraries  

NASA Astrophysics Data System (ADS)

Human monoclonal antibodies have considerable potential in the prophylaxis and treatment of viral disease. However, only a few such antibodies suitable for clinical use have been produced to date. We have previously shown that large panels of human recombinant monoclonal antibodies against a plethora of infectious agents, including herpes simplex virus types 1 and 2, can be established from phage display libraries. Here we demonstrate that facile cloning of recombinant Fab fragments against specific viral proteins in their native conformation can be accomplished by panning phage display libraries against viral glycoproteins "captured" from infected cell extracts by specific monoclonal antibodies immobilized on ELISA plates. We have tested this strategy by isolating six neutralizing recombinant antibodies specific for herpes simplex glycoprotein gD or gB, some of which are against conformationally sensitive epitopes. By using defined monoclonal antibodies for the antigen-capture step, this method can be used for the isolation of antibodies to specific regions and epitopes within the target viral protein. For instance, monoclonal antibodies to a nonneutralizing epitope can be used in the capture step to clone antibodies to neutralizing epitopes, or antibodies to a neutralizing epitope can be used to clone antibodies to a different neutralizing epitope. Furthermore, by using capturing antibodies to more immunodominant epitopes, one can direct the cloning to less immunogenic ones. This method should be of value in generating antibodies to be used both in the prophylaxis and treatment of viral infections and in the characterization of the mechanisms of antibody protective actions at the molecular level.

Sanna, Pietro Paolo; Williamson, R. Anthony; de Logu, Alessandro; Bloom, Floyd E.; Burton, Dennis R.

1995-07-01

309

Directed selection of recombinant human monoclonal antibodies to herpes simplex virus glycoproteins from phage display libraries.  

PubMed Central

Human monoclonal antibodies have considerable potential in the prophylaxis and treatment of viral disease. However, only a few such antibodies suitable for clinical use have been produced to date. We have previously shown that large panels of human recombinant monoclonal antibodies against a plethora of infectious agents, including herpes simplex virus types 1 and 2, can be established from phage display libraries. Here we demonstrate that facile cloning of recombinant Fab fragments against specific viral proteins in their native conformation can be accomplished by panning phage display libraries against viral glycoproteins "captured" from infected cell extracts by specific monoclonal antibodies immobilized on ELISA plates. We have tested this strategy by isolating six neutralizing recombinant antibodies specific for herpes simplex glycoprotein gD or gB, some of which are against conformationally sensitive epitopes. By using defined monoclonal antibodies for the antigen-capture step, this method can be used for the isolation of antibodies to specific regions and epitopes within the target viral protein. For instance, monoclonal antibodies to a nonneutralizing epitope can be used in the capture step to clone antibodies to neutralizing epitopes, or antibodies to a neutralizing epitope can be used to clone antibodies to a different neutralizing epitope. Furthermore, by using capturing antibodies to more immunodominant epitopes, one can direct the cloning to less immunogenic ones. This method should be of value in generating antibodies to be used both in the prophylaxis and treatment of viral infections and in the characterization of the mechanisms of antibody protective actions at the molecular level. Images Fig. 2 Fig. 3 PMID:7604009

Sanna, P P; Williamson, R A; De Logu, A; Bloom, F E; Burton, D R

1995-01-01

310

A monoclonal antibody specific for mouse dendritic cells.  

PubMed Central

Dendritic cells (DC) are a small subpopulation of lymphoid cells with distinctive cytologic features, surface properties, and functions. This report describes the DC-specific antibody (Ab) secreted by clone 33DI. Rat spleen cells immune to mouse DC were fused to the P3U myeloma. Hybrid culture supernatants were screened simultaneously against DC, a macrophage (M phi) cell line, and mitogen-stimulated lymphoblasts. 33DI Ab specifically killed 80-90% of DC from spleen and lymph node, but no other leukocytes, including Ia+ and Ia- M phi (Ia, I-region-associated antigen,). Quantitative binding studies with 5H-labeled 33D1 Ab showed that DC had an average of 14,000 binding sites per cell. Binding to DC was inhibited with Fab fragment of 33D1 Ab but not with a panel of other monoclonal antibodies, including anti-Ia Ab. Adherence and flotation procedures that enriched for DC enriched for 3H-labeled 33D1 Ab binding in parallel. 33D1 antigen was not detectable on: M phi from spleen, peritoneal cavity, and blood; three M phi cell lines; lymphocytes; granulocytes; platelets; and erythroid cells. DC continued to express the 33D1 antigen after 4 days in culture, whereas M phi and lymphocytes did not acquire it. Quantitative and autoradiographic studies confirmed that spleen and lymph node suspensions contain less than 1% DC. We conclude that 33D1 Ab detects a stable and specific DC antigen and can be used to monitor DC content in complex lymphoid mixtures. Images PMID:6948298

Nussenzweig, M C; Steinman, R M; Witmer, M D; Gutchinov, B

1982-01-01

311

A monoclonal antibody specific for mouse dendritic cells.  

PubMed

Dendritic cells (DC) are a small subpopulation of lymphoid cells with distinctive cytologic features, surface properties, and functions. This report describes the DC-specific antibody (Ab) secreted by clone 33DI. Rat spleen cells immune to mouse DC were fused to the P3U myeloma. Hybrid culture supernatants were screened simultaneously against DC, a macrophage (M phi) cell line, and mitogen-stimulated lymphoblasts. 33DI Ab specifically killed 80-90% of DC from spleen and lymph node, but no other leukocytes, including Ia+ and Ia- M phi (Ia, I-region-associated antigen,). Quantitative binding studies with 5H-labeled 33D1 Ab showed that DC had an average of 14,000 binding sites per cell. Binding to DC was inhibited with Fab fragment of 33D1 Ab but not with a panel of other monoclonal antibodies, including anti-Ia Ab. Adherence and flotation procedures that enriched for DC enriched for 3H-labeled 33D1 Ab binding in parallel. 33D1 antigen was not detectable on: M phi from spleen, peritoneal cavity, and blood; three M phi cell lines; lymphocytes; granulocytes; platelets; and erythroid cells. DC continued to express the 33D1 antigen after 4 days in culture, whereas M phi and lymphocytes did not acquire it. Quantitative and autoradiographic studies confirmed that spleen and lymph node suspensions contain less than 1% DC. We conclude that 33D1 Ab detects a stable and specific DC antigen and can be used to monitor DC content in complex lymphoid mixtures. PMID:6948298

Nussenzweig, M C; Steinman, R M; Witmer, M D; Gutchinov, B

1982-01-01

312

Polyelectrolyte Nanogels Decorated with Monoclonal Antibody for Targeted Drug Delivery  

PubMed Central

Novel surface-functionalized cross-linked nanogels were developed as a platform to allow conjugation of monoclonal antibodies (mAb) for targeted drug delivery. Well-defined diblock copolymers of poly(ethylene glycol)-b-poly(methacrylic acid) (PEG-b-PMA) with PEG terminal aldehyde functionality were synthesized by atom transfer radical polymerization (ATRP) and characterized by GPC and 1H NMR. These copolymers were used to prepare nanogels via condensation of PEG-b-PMA with Ca2+ ions into micelle-like aggregates, cross-linking of the PMA/Ca2+ cores and removal of Ca2+ ions. The resulting nanogels represent highly swollen spherical polyelectrolyte particles with free terminal aldehyde functionalities at the nonionic PEG chains. A reductive amination reaction between aldehyde groups and amino groups of mAb resulted in effective conjugation to the nanogels of mAb CC49 against tumor-associated glycoprotein 72 (TAG-72). The mAb retained the binding affinity to bovine submaxillary mucin after conjugation as shown by surface plasmon resonance (SPR). Therefore, aldehyde functionalized nanogels can be linked to mAb using a simple, one-step approach. They may have potential for targeted delivery of diagnostic and therapeutic agents to tumors. PMID:21503276

Nukolova, Nataliya V.; Yang, Zigang; Kim, Jong Oh; Kabanov, Alexander V.; Bronich, Tatiana K.

2010-01-01

313

Are monoclonal antibodies a safe treatment for cancer during pregnancy?  

PubMed

Monoclonal antibodies (mAbs) are the cornerstone of the treatment of several types of tumors, but their use in pregnant women is not clearly defined. Here, we report and analyze all available data on mAb treatment in pregnant cancer patients. A literature search was performed from 2000 until January 2013 and all articles addressing safety of mAbs during pregnancy were reviewed. We found very few data on the use of bevacizumab in pregnant women. However, owing to its antiangiogenic effects and possible consequences on fetal development, it should be avoided during pregnancy. Trastuzumab administration has been associated with an elevated incidence of oligohydramnios and poor neonatal outcomes, particularly when prescribed after the first trimester for repeated infusions, and therefore it is not recommended. Rituximab does not seem to be teratogenic, but a transient prolonged neutropenia in the newborns was reported, without major infectious consequences in most cases. Few data are available about other mAbs, and hence their use during pregnancy remains discouraged. PMID:23829624

Sarno, Maria Anna; Mancari, Rosanna; Azim, Hatem A; Colombo, Nicoletta; Peccatori, Fedro A

2013-07-01

314

Production of a Chaetomium globosum Enolase Monoclonal Antibody  

PubMed Central

Chaetomium globosum is a hydrophilic fungal species and a contaminant of water-damaged building materials in North America. Methods to detect Chaetomium species include subjective identification of ascospores, viable culture, or molecular-based detection methods. In this study, we describe the production and initial characterization of a monoclonal antibody (MAb) for C. globosum enolase. MAb 1C7, a murine IgG1 isotype MAb, was produced and reacted with recombinant C. globosum enolase (rCgEno) in an enzyme-linked immunosorbent assay and with a putative C. globosum enolase in a Western blot. Epitope mapping showed MAb 1C7 specific reactivity to an enolase decapeptide, LTYEELANLY, that is highly conserved within the fungal class Sordariomycetes. Cross-reactivity studies showed MAb 1C7 reactivity to C. atrobrunneum but not C. indicum. MAb 1C7 did not react with enolase from Aspergillus fumigatus, which is divergent in only two amino acids within this epitope. The results of this study suggest potential utility of MAb 1C7 in Western blot applications for the detection of Chaetomium and other Sordariomycetes species. PMID:25495488

Nayak, Ajay P.; Lemons, Angela R.; Rittenour, William R.; Hettick, Justin M.; Beezhold, Donald H.

2014-01-01

315

Monoclonal antibodies for the detection of Puccinia striiformis urediniospores.  

PubMed

The fungal pathogen Pst causes yellow rust disease in wheat plants leading to crop losses. The organism spreads by releasing wind-dispersed urediniospores from infected plants. In this study a library of novel monoclonal antibodies (mAbs) was developed against Pst urediniospores. Nine mAb-producing cell lines were cloned and their cross-reactivities characterised against a panel of airborne fungal spores representing genera commonly found in the same environment as Pst. Two specific mAbs were used to develop a competitive ELISA (Pst mAb4) and a subtractive inhibition ELISA (Pst mAb8). Standard curves for both assays had good intra- and interday reproducibility. The subtractive inhibition ELISA had greater sensitivity with a detection limit of 1.5 x 10(5) spores ml(-1). Cross-reactivity studies of Pst mAb8 in the subtractive inhibition ELISA, showed reaction with other Puccinia spores only, suggesting that common epitopes exist within this genus. The biosensor-compatible Pst mAb8 assay principle developed in this study has the potential to be implemented in future 'label-free' in-the-field systems for Pst detection. PMID:17350244

Skottrup, Peter; Frřkiaer, Hanne; Hearty, Stephen; O'Kennedy, Richard; Hejgaard, Jřrn; Nicolaisen, Mogens; Justesen, Annemarie F

2007-03-01

316

Production of a Chaetomium globosum Enolase Monoclonal Antibody.  

PubMed

Chaetomium globosum is a hydrophilic fungal species and a contaminant of water-damaged building materials in North America. Methods to detect Chaetomium species include subjective identification of ascospores, viable culture, or molecular-based detection methods. In this study, we describe the production and initial characterization of a monoclonal antibody (MAb) for C. globosum enolase. MAb 1C7, a murine IgG1 isotype MAb, was produced and reacted with recombinant C. globosum enolase (rCgEno) in an enzyme-linked immunosorbent assay and with a putative C. globosum enolase in a Western blot. Epitope mapping showed MAb 1C7 specific reactivity to an enolase decapeptide, LTYEELANLY, that is highly conserved within the fungal class Sordariomycetes. Cross-reactivity studies showed MAb 1C7 reactivity to C. atrobrunneum but not C. indicum. MAb 1C7 did not react with enolase from Aspergillus fumigatus, which is divergent in only two amino acids within this epitope. The results of this study suggest potential utility of MAb 1C7 in Western blot applications for the detection of Chaetomium and other Sordariomycetes species. PMID:25495488

Green, Brett J; Nayak, Ajay P; Lemons, Angela R; Rittenour, William R; Hettick, Justin M; Beezhold, Donald H

2014-12-01

317

Development of a monoclonal antibody specific to cooked mammalian meats.  

PubMed

Detection of species adulteration in ground meat products is important for consumer protection and food-labeling law enforcement. This study was conducted to develop monoclonal antibodies (MAbs) that can be used in an enzyme-linked immunosorbent assay (ELISA) for rapid detection of any cooked mammalian meats in cooked poultry products. Soluble muscle proteins extracted from cooked pork (heated at 100 degrees C for 15 min) were used as the antigen to immunized mice for developing the MAb. One that was developed, MAb 2F8 (IgG2b class), strongly reacted with cooked meat of five mammalian species (beef cattle, hogs, sheep, horse, and deer) but did not react with any cooked poultry (chicken, turkey, and duck) or raw meats. At least 0.5% by weight of pork, beef, lamb, and horse meats in a chicken-based mixture could not detect using the indirect ELISA with MAb 2F8. The MAb 2F8 is useful in a single initial screening test to detect the presence of five nonpoultry meat adulterants in cooked poultry products. PMID:9709213

Hsieh, Y H; Sheu, S C; Bridgman, R C

1998-04-01

318

Adverse events to monoclonal antibodies used for cancer therapy  

PubMed Central

Fifteen monoclonal antibodies (mAbs) are currently registered and approved for the treatment of a range of different cancers. These mAbs are specific for a limited number of targets (9 in all). Four of these molecules are indeed directed against the B-lymphocyte antigen CD20; 3 against human epidermal growth factor receptor 2 (HER2 or ErbB2), 2 against the epidermal growth factor receptor (EGFR), and 1 each against epithelial cell adhesion molecule (EpCAM), CD30, CD52, vascular endothelial growth factor (VEGF), tumor necrosis factor (ligand) superfamily, member 11 (TNFSF11, best known as RANKL), and cytotoxic T lymphocyte-associated protein 4 (CTLA4). Collectively, the mAbs provoke a wide variety of systemic and cutaneous adverse events including the full range of true hypersensitivities: Type I immediate reactions (anaphylaxis, urticaria); Type II reactions (immune thrombocytopenia, neutopenia, hemolytic anemia); Type III responses (vasculitis, serum sickness; some pulmonary adverse events); and Type IV delayed mucocutaneous reactions as well as infusion reactions/cytokine release syndrome (IRs/CRS), tumor lysis syndrome (TLS), progressive multifocal leukoencephalopathy (PML) and cardiac events. Although the term “hypersensitivity” is widely used, no common definition has been adopted within and between disciplines and the requirement of an immunological basis for a true hypersensitivity reaction is sometimes overlooked. Consequently, some drug-induced adverse events are sometimes incorrectly described as “hypersensitivities” while others that should be described are not. PMID:24251081

Baldo, Brian A

2013-01-01

319

Role of cosolutes in the aggregation kinetics of monoclonal antibodies.  

PubMed

We propose a general strategy based on kinetic analysis to investigate how cosolutes affect the aggregation behavior of therapeutic proteins. We apply this approach to study the impact of NaCl and sorbitol on the aggregation kinetics of two monoclonal antibodies, an IgG1 and an IgG2. By using a combination of size exclusion chromatography and light scattering techniques, we study the impact of the cosolutes on the monomer depletion, as well as on the formation of dimers, trimers, and larger aggregates. We analyze these macroscopic effects in the frame of a kinetic model based on Smoluchowski's population balance equations modified to account for nucleation events. By comparing experimental data with model simulations, we discriminate the effect of cosolutes on the elementary steps which contribute to the global aggregation process. In the case of the IgG1, it is found that NaCl accelerates the kinetics of aggregation by promoting specifically aggregation events, while sorbitol delays the kinetics of aggregation by specifically inhibiting protein unfolding. In the case of the IgG2, whose monomer depletion kinetics is limited by dimer formation, NaCl and sorbitol are found respectively to accelerate and inhibit conformational changes and aggregation events to the same extent. PMID:25243487

Nicoud, Lucrčce; Sozo, Margaux; Arosio, Paolo; Yates, Andrew; Norrant, Edith; Morbidelli, Massimo

2014-10-16

320

Her2 Monoclonal Antibodies, Antibody Drug Conjugates, and Site Specific Antibody Conjugate Methods  

Cancer.gov

Antibody drug conjugates (ADC) can demonstrate high efficacy as cancer therapeutics, however, much more can be done to improve their efficacy and safety profile. Site-specific antibody drug conjugation is a promising way to do this.

321

Supraagonistic, bispecific single-chain antibody purified from the serum of cloned, transgenic cows induces T-cell-mediated killing of glioblastoma cells in vitro and in vivo.  

PubMed

Here we characterize the antitumor activity of a recombinant bispecific single-chain antibody isolated from the serum of cloned transgenic cows. The antibody, termed r28M, is directed to a melanoma-associated proteoglycan, also expressed on glioblastoma cells, and to human CD28. Bound to tumor cells, r28M induced exceedingly efficient supraagonistic T-cell activation via the CD28 molecule without an additional stimulus via the TCR/CD3 complex. In vitro, T cells and NK cells contributed to tumor cell killing after r28M-mediated activation of peripheral blood mononuclear cells. However, NK activity depended on T-cell-derived cytokines. In vivo, r28M markedly inhibited the growth of human glioblastoma cells in nude mice. The serum half-life of the protein after i.v. injection was approximately 6 hr. Thus, r28M is unique not only in inducing supraagonistic CD28-mediated T-cell activation against tumor cells in vitro and in vivo, it also meets 2 additional requirements that are critical for clinical application: a relatively long serum half-life and the possibility of obtaining large amounts of active material from cloned transgenic livestock. PMID:16003729

Grosse-Hovest, Ludger; Wick, Wolfgang; Minoia, Rosa; Weller, Michael; Rammensee, Hans-Georg; Brem, Gottfried; Jung, Gundram

2005-12-20

322

Exceptionally Potent and Broadly Cross-Reactive, Bispecific Multivalent HIV-1 Inhibitors Based on Single Human CD4 and Antibody Domains  

PubMed Central

Soluble forms of the human immunodeficiency virus type 1 (HIV-1) primary receptor CD4 (soluble CD4 [sCD4]) have been extensively characterized for a quarter of a century as promising HIV-1 inhibitors, but they have not been clinically successful. By combining a protein cavity-filling strategy and the power of library technology, we identified an engineered cavity-altered single-domain sCD4 (mD1.22) with a unique combination of excellent properties, including broad and potent neutralizing activity, high specificity, stability, solubility, and affinity for the HIV-1 envelope glycoprotein gp120, and small molecular size. To further improve its neutralizing potency and breadth, we generated bispecific multivalent fusion proteins of mD1.22 with another potent HIV-1 inhibitor, an antibody domain (m36.4) that targets the coreceptor-binding site on gp120. The fusion proteins neutralized all HIV-1 isolates tested, with potencies about 10-, 50-, and 200-fold higher than those of the broadly neutralizing antibody VRC01, the U.S. FDA-approved peptide inhibitor T20, and the clinically tested sCD4-Fc fusion protein CD4-Ig, respectively. In addition, they exhibited higher stability and specificity and a lower aggregation propensity than CD4-Ig. Therefore, mD1.22 and related fusion proteins could be useful for HIV-1 prevention and therapy, including eradication of the virus. PMID:24198429

Feng, Yang; Prabakaran, Ponraj; Ying, Tianlei; Wang, Yanping; Sun, Jianping; Macedo, Camila D. S.; Zhu, Zhongyu; He, Yuxian; Polonis, Victoria R.

2014-01-01

323

Anti-factor IXa/X bispecific antibody ACE910 prevents joint bleeds in a long-term primate model of acquired hemophilia A  

PubMed Central

ACE910 is a humanized anti-factor IXa/X bispecific antibody mimicking the function of factor VIII (FVIII). We previously demonstrated in nonhuman primates that a single IV dose of ACE910 exerted hemostatic activity against hemophilic bleeds artificially induced in muscles and subcutis, and that a subcutaneous (SC) dose of ACE910 showed a 3-week half-life and nearly 100% bioavailability, offering support for effective prophylaxis for hemophilia A by user-friendly SC dosing. However, there was no direct evidence that such SC dosing of ACE910 would prevent spontaneous bleeds occurring in daily life. In this study, we newly established a long-term primate model of acquired hemophilia A by multiple IV injections of an anti-primate FVIII neutralizing antibody engineered in mouse-monkey chimeric form to reduce its antigenicity. The monkeys in the control group exhibited various spontaneous bleeding symptoms as well as continuous prolongation of activated partial thromboplastin time; notably, all exhibited joint bleeds, which are a hallmark of hemophilia. Weekly SC doses of ACE910 (initial 3.97 mg/kg followed by 1 mg/kg) significantly prevented these bleeding symptoms; notably, no joint bleeding symptoms were observed. ACE910 is expected to prevent spontaneous bleeds and joint damage in hemophilia A patients even with weekly SC dosing, although appropriate clinical investigation is required. PMID:25274508

Yoshihashi, Kazutaka; Takeda, Minako; Kitazawa, Takehisa; Soeda, Tetsuhiro; Igawa, Tomoyuki; Sampei, Zenjiro; Kuramochi, Taichi; Sakamoto, Akihisa; Haraya, Kenta; Adachi, Kenji; Kawabe, Yoshiki; Nogami, Keiji; Shima, Midori; Hattori, Kunihiro

2014-01-01

324

Exceptionally potent and broadly cross-reactive, bispecific multivalent HIV-1 inhibitors based on single human CD4 and antibody domains.  

PubMed

Soluble forms of the human immunodeficiency virus type 1 (HIV-1) primary receptor CD4 (soluble CD4 [sCD4]) have been extensively characterized for a quarter of a century as promising HIV-1 inhibitors, but they have not been clinically successful. By combining a protein cavity-filling strategy and the power of library technology, we identified an engineered cavity-altered single-domain sCD4 (mD1.22) with a unique combination of excellent properties, including broad and potent neutralizing activity, high specificity, stability, solubility, and affinity for the HIV-1 envelope glycoprotein gp120, and small molecular size. To further improve its neutralizing potency and breadth, we generated bispecific multivalent fusion proteins of mD1.22 with another potent HIV-1 inhibitor, an antibody domain (m36.4) that targets the coreceptor-binding site on gp120. The fusion proteins neutralized all HIV-1 isolates tested, with potencies about 10-, 50-, and 200-fold higher than those of the broadly neutralizing antibody VRC01, the U.S. FDA-approved peptide inhibitor T20, and the clinically tested sCD4-Fc fusion protein CD4-Ig, respectively. In addition, they exhibited higher stability and specificity and a lower aggregation propensity than CD4-Ig. Therefore, mD1.22 and related fusion proteins could be useful for HIV-1 prevention and therapy, including eradication of the virus. PMID:24198429

Chen, Weizao; Feng, Yang; Prabakaran, Ponraj; Ying, Tianlei; Wang, Yanping; Sun, Jianping; Macedo, Camila D S; Zhu, Zhongyu; He, Yuxian; Polonis, Victoria R; Dimitrov, Dimiter S

2014-01-01

325

Alkaline cation-exchange chromatography for the reduction of aggregate and a mis-formed disulfide variant in a bispecific antibody purification process.  

PubMed

During the purification development of a bispecific antibody, cation-exchange chromatography was screened for its ability to separate a prominently expressed (>12%) mis-formed disulfide bond variant, termed MAb-diabody, and aggregate from the product of interest. The influence of pH, product load (g of product per liter of resin) and linear velocity on the separations were evaluated for the strong cation-exchange resins SP Sepharose HP and POROS(®) HS50. Cation-exchange chromatography is commonly operated distant to the isoelectric point of a molecule, generally leading to acidic conditions for antibody purification. However, the results herein demonstrated improved removal of MAb-diabody with increasing pH, resulting in reduction of MAb-diabody content greater than 12-fold when operating near the alkaline pI of the product. This approach was successful over a range of linear velocities and g/L of resin loading. Aggregate removal was less affected by pH and was effectively reduced from 10.9% to less than 3% for each condition. Furthermore, this method was successfully scaled to a 60cm diameter column using SP Sepharose HP resin. PMID:25462105

Hall, Troii; Wilson, Joseph J; Brownlee, Tammy J; Swartling, James R; Langan, Sarah E; Lambooy, Peter K

2015-01-15

326

Monoclonal antibody capture enzyme-linked immunosorbent assay for detection of bovine enteric coronavirus.  

PubMed

Monoclonal antibodies reactive with three different viral polypeptides were evaluated singly and in combination as the capture antibody(s) in an enzyme-linked immunosorbent assay system for the detection of bovine enteric coronavirus. Similar levels of sensitivity were found for all combinations tested. A sensitive, highly specific, and reproducible assay for the detection of bovine enteric coronavirus was developed, using a mixture of two of these monoclonal antibodies reactive with antigenic components either external or internal to the virion. These monoclonal antibodies were bound indirectly to 96-well plates via rabbit anti-mouse immunoglobulin. After sample application and incubation, virus was detected by using rabbit anti-coronavirus peroxidase conjugate followed by enzyme substrate and chromagen. Fecal samples from a single herd of cows were screened for the presence of coronavirus by this assay. Five percent of clinically normal cows were found to be shedding coronavirus. PMID:6325490

Crouch, C F; Raybould, T J; Acres, S D

1984-03-01

327

Antigen-Specific Monoclonal Antibodies Isolated from B Cells Expressing Constitutively Active STAT5  

PubMed Central

Background Fully human monoclonal antibodies directed against specific pathogens have a high therapeutic potential, but are difficult to generate. Methodology/Principal Findings Memory B cells were immortalized by expressing an inducible active mutant of the transcription factor Signal Transducer and Activator of Transcription 5 (STAT5). Active STAT5 inhibits the differentiation of B cells while increasing their replicative life span. We obtained cloned B cell lines, which produced antibodies in the presence of interleukin 21 after turning off STAT5. We used this method to obtain monoclonal antibodies against the model antigen tetanus toxin. Conclusions/Significance Here we describe a novel and relatively simple method of immortalizing antigen-specific human B cells for isolation of human monoclonal antibodies. These results show that STAT5 overexpression can be employed to isolate antigen specific antibodies from human memory B cells. PMID:21525985

Scheeren, Ferenc A.; van Geelen, Caroline M. M.; Yasuda, Etsuko; Spits, Hergen; Beaumont, Tim

2011-01-01

328

Perfusion of tumor-bearing kidneys as a model for scintigraphic screening of monoclonal antibodies  

SciTech Connect

Tumor-bearing human kidneys were used in an ex vivo perfusion model to screen monoclonal antibodies, recognizing renal cell carcinoma-associated antigens for diagnostic potential in vivo. Perfusion of tumor-bearing kidneys with /sup 99m/Tc-labeled G250 and RC38 antibody resulted in visualization of the tumor, whereas perfusion with two other monoclonal antibodies, RC2 and RC4, did not lead to tumor visualization. Uptake of radiolabel in normal kidney tissue was low for G250 and RC38 antibody. Tumor-to-kidney tissue ratios after perfusion with G250 and RC38 antibody were 2.7 and 2.2, respectively. After rinsing for 3 hr with unlabeled perfusion fluid the tumor-to-kidney tissue ratios increased to 8.6 for G250 antibody and to 2.7 for RC38 antibody. We conclude that perfusion of tumor-bearing human kidneys with radiolabeled monoclonal antibodies is a relatively simple way to evaluate renal cell carcinoma associated monoclonal antibodies as diagnostic agents in vivo.

van Dijk, J.; Oosterwijk, E.; van Kroonenburgh, M.J.; Jonas, U.; Fleuren, G.J.; Pauwels, E.K.; Warnaar, S.O.

1988-06-01

329

Two monoclonal antibodies raised against different epitopes of chloroplast fructose-1. 6-bisphosphatase (FBPase)  

SciTech Connect

Two monoclonal antibodies (GR-BP5 and GR-BP8) were obtained by fusion of spleen cells of mice immunized against pea photosynthetic FBPase with cells of myeloma NSI. Both mAbs showed by double immunodiffusion a {chi} light chain, and the GR-BP8 secreted an IgM. By Western-blotting and immunoprecipitation of the in vivo labelled pea FBPase, GR-BP5 and GR-BP8 showed specificity for the chloroplast enzyme. Competition binding of the {sup 125}I-labelled mAbs against pea FBPase showed specific binding sites to different epitopes of the enzyme molecule. Cross reaction assays between both monoclonal antibodies and pea and spinach chloroplast FBPases showed a 90-100% homology in the corresponding epitopes of both enzymes. Preliminary assays showed a moderate inhibition of FBPase by GR-BP5 monoclonal antibody, but a weak enhancement by the GR-BP8 monoclonal one.

Hermoso, R.; Fonolla, J.; Lopez-Gorge, J. (Estacion Experimental del Zaidin, Granada (Spain)); Ruiz-Cabello, F.; Garrido, F. (C.S. Virgen de las Nieves, Granada (Spain))

1990-05-01

330

Characterization of cross-reactive norovirus-specific monoclonal antibodies.  

PubMed

Noroviruses (NoVs) commonly cause acute gastroenteritis outbreaks. Broadly reactive diagnostic assays are essential for rapid detection of NoV infections. We previously generated a panel of broadly reactive monoclonal antibodies (MAbs). We characterized MAb reactivities by use of virus-like particles (VLPs) from 16 different NoV genotypes (6 from genogroup I [GI], 9 from GII, and 1 from GIV) coating a microtiter plate (direct enzyme-linked immunosorbent assay [ELISA]) and by Western blotting. MAbs were genotype specific or recognized multiple genotypes within a genogroup and between genogroups. We next applied surface plasmon resonance (SPR) analysis to measure MAb dissociation constants (Kd) as a surrogate for binding affinity; a Kd level of <10 nM was regarded as indicating strong binding. Some MAbs did not interact with the VLPs by SPR analysis. To further assess this lack of MAb-VLP interaction, the MAbs were evaluated for the ability to identify NoV VLPs in a capture ELISA. Those MAbs for which a Kd could not be measured by SPR analysis also failed to capture the NoV VLPs; in contrast, those with a measurable Kd gave a positive signal in the capture ELISA. Thus, some broadly cross-reactive epitopes in the VP1 protruding domain may be partially masked on intact particles. One MAb, NV23, was able to detect genogroup I, II, and IV VLPs from 16 genotypes tested by sandwich ELISA, and it successfully detected NoVs in stool samples positive by real-time reverse transcription-PCR when the threshold cycle (CT) value was <31. Biochemical analyses of MAb reactivity, including SPR analysis, identified NV23 as a broadly reactive ligand for application in norovirus diagnostic assays. PMID:25428247

Kou, Baijun; Crawford, Sue E; Ajami, Nadim J; Czakó, Rita; Neill, Frederick H; Tanaka, Tomoyuki N; Kitamoto, Noritoshi; Palzkill, Timothy G; Estes, Mary K; Atmar, Robert L

2015-02-01

331

Mapping Broadly Reactive Norovirus Genogroup I and II Monoclonal Antibodies.  

PubMed

Noroviruses are responsible for most acute nonbacterial epidemic outbreaks of gastroenteritis worldwide. To develop cross-reactive monoclonal antibodies (MAbs) for rapid identification of genogroup I and II (GI and GII) noroviruses (NoVs) in field specimens, mice were immunized with baculovirus-expressed recombinant virus-like particles (VLPs) corresponding to NoVs. Nine MAbs against the capsid protein were identified that detected both GI and GII NoV VLPs. These MAbs were tested in competition enzyme-linked immunosorbent assays (ELISAs) to identify common epitope reactivities to GI and GII VLPs. Patterns of competitive reactivity placed these MAbs into two epitope groups (groups 1 and 2). Epitopes for MAbs NV23 and NS22 (group 1) and MAb F120 (group 2) were mapped to a continuous region in the C-terminal P1 subdomain of the capsid protein. This domain is within regions previously defined to contain cross-reactive epitopes in GI and GII viruses, suggesting that common epitopes are clustered within the P1 domain of the capsid protein. Further characterization in an accompanying paper (B. Kou et al., Clin Vaccine Immunol 22:160-167, 2015, http://dx.doi.org/10.1128/CVI.00519-14) revealed that MAb NV23 (epitope group 1) is able to detect GI and GII viruses in stool. Inclusion of the GI and GII cross-reactive MAb NV23 in antigen detection assays may facilitate the identification of GI and GII human noroviruses in stool samples as causative agents of outbreaks and sporadic cases of gastroenteritis worldwide. PMID:25428246

Crawford, Sue E; Ajami, Nadim; Parker, Tracy Dewese; Kitamoto, Noritoshi; Natori, Katsuro; Takeda, Naokazu; Tanaka, Tomoyuki; Kou, Baijun; Atmar, Robert L; Estes, Mary K

2015-02-01

332

Safety and immunotoxicity assessment of immunomodulatory monoclonal antibodies  

PubMed Central

Most therapeutic monoclonal antibodies (mAbs) licensed for human use or in clinical development are indicated for treatment of patients with cancer and inflammatory/autoimmune disease and as such, are designed to directly interact with the immune system. A major hurdle for the development and early clinical investigation of many of these immunomodulatory mAbs is their inherent risk for adverse immune-mediated drug reactions in humans such as infusion reactions, cytokine storms, immunosuppression and autoimmunity. A thorough understanding of the immunopharmacology of a mAb in humans and animals is required to both anticipate the clinical risk of adverse immunotoxicological events and to select a safe starting dose for first-in-human (FIH) clinical studies. This review summarizes the most common adverse immunotoxicological events occurring in humans with immunomodulatory mAbs and outlines non-clinical strategies to define their immunopharmacology and assess their immunotoxic potential, as well as reduce the risk of immunotoxicity through rational mAb design. Tests to assess the relative risk of mAb candidates for cytokine release syndrome, innate immune system (dendritic cell) activation and immunogenicity in humans are also described. The importance of selecting a relevant and sensitive toxicity species for human safety assessment in which the immunopharmacology of the mAb is similar to that expected in humans is highlighted, as is the importance of understanding the limitations of the species selected for human safety assessment and supplementation of in vivo safety assessment with appropriate in vitro human assays. A tiered approach to assess effects on immune status, immune function and risk of infection and cancer, governed by the mechanism of action and structural features of the mAb, is described. Finally, the use of immunopharmacology and immunotoxicity data in determining a minimum anticipated biologic effect Level (MABEL) and in the selection of safe human starting dose is discussed. PMID:20421713

Morton, Laura Dill; Spindeldreher, Sebastian; Kiessling, Andrea; Allenspach, Roy; Hey, Adam; Muller, Patrick Y; Frings, Werner; Sims, Jennifer

2010-01-01

333

Response of a concentrated monoclonal antibody formulation to high shear.  

PubMed

There is concern that shear could cause protein unfolding or aggregation during commercial biopharmaceutical production. In this work we exposed two concentrated immunoglobulin-G1 (IgG1) monoclonal antibody (mAb, at >100 mg/mL) formulations to shear rates between 20,000 and 250,000 s(-1) for between 5 min and 30 ms using a parallel-plate and capillary rheometer, respectively. The maximum shear and force exposures were far in excess of those expected during normal processing operations (20,000 s(-1) and 0.06 pN, respectively). We used multiple characterization techniques to determine if there was any detectable aggregation. We found that shear alone did not cause aggregation, but that prolonged exposure to shear in the stainless steel parallel-plate rheometer caused a very minor reversible aggregation (<0.3%). Additionally, shear did not alter aggregate populations in formulations containing 17% preformed heat-induced aggregates of a mAb. We calculate that the forces applied to a protein by production shear exposures (<0.06 pN) are small when compared with the 140 pN force expected at the air-water interface or the 20-150 pN forces required to mechanically unfold proteins described in the atomic force microscope (AFM) literature. Therefore, we suggest that in many cases, air-bubble entrainment, adsorption to solid surfaces (with possible shear synergy), contamination by particulates, or pump cavitation stresses could be much more important causes of aggregation than shear exposure during production. PMID:19370772

Bee, Jared S; Stevenson, Jennifer L; Mehta, Bhavya; Svitel, Juraj; Pollastrini, Joey; Platz, Robert; Freund, Erwin; Carpenter, John F; Randolph, Theodore W

2009-08-01

334

Subtractive immunization yields monoclonal antibodies that specifically inhibit metastasis  

PubMed Central

Subtractive immunization allowed the isolation and characterization of monoclonal antibodies that specifically inhibit metastasis but not proliferation of highly metastatic human tumor cells. The tolerizing agent cyclophosphamide was used to suppress the immune system in mice to dominant immunodeterminants present on a non-metastatic variant (M-) of the human epidermoid carcinoma cell line (HEp3). Mice were then inoculated with a highly metastatic variant (M+) of HEp3 to enhance an immune response to antigenic determinants present on metastatic cells. Hybridomas were generated and screened by ELISA for differential reactivity to M+ HEp3 over M- HEp3 cells. This experimental approach, termed subtractive immunization (S.I.), was compared to a control immunization protocol, which eliminated the cyclophosphamide treatment. The S.I. protocol resulted in an eight-fold increase in the proportion of mAbs that react with molecules enriched on the surface of the M+ HEp3 cells. Two of the mAbs derived from the S.I. protocol, designated DM12-4 and 1A5, were purified and examined for their effect in a metastasis model system in which chick embryos are transplanted with primary HEp3 tumors. Purified mAbs DM12-4 and 1A5, inoculated i.v. into the embryos, inhibited spontaneous metastasis of HEp3 cells by 86 and 90%, respectively. The mAbs are specifically anti-metastatic in that they have no effect on the growth of HEp3 cells in vitro nor did they inhibit primary tumor growth in vivo. The mAbs recognize M+ HEp3 cell surface molecules of 55 kD and 29 kD, respectively. These data demonstrate that the S.I. protocol can be used for the development of unique mAbs that are reactive with antigenic determinants whose expression is elevated on metastatic human tumor cells and which function mechanistically in the metastatic cascade. PMID:8376467

1993-01-01

335

PRODUCTION OF MONOCLONAL ANTIBODIES TO 'LEGIONELLA PNEUMOPHILA' SEROGROUPS 1 AND 6  

EPA Science Inventory

To better define the surface antigens of Legionella pneumophila for clinical and experimental purposes, were produced monoclonal antibodies to L. pneumophila serogroups 1 and 6. Two hybridomas were produced in serogroup 1. One antibody, LP-I-17, recognized a serogroup-common anti...

336

Comparability assessments of process and product changes made during development of two different monoclonal antibodies  

Microsoft Academic Search

To assess the impact of manufacturing changes on antibody structure and function during the course of product development, three comparability studies were performed for each of two different IgG1 monoclonal antibody product candidates. Comparability study #1 evaluated the effect of changing the cell line and bulk drug substance manufacturing process for cell culture and purification. Results indicated that these process

Anthony Lubiniecki; David B. Volkin; Marcia Federici; Michael D. Bond; Michael L. Nedved; Linda Hendricks; Promod Mehndiratta; Mark Bruner; Sudhir Burman; Paul DalMonte; Jane Kline; Alex Ni; Mark E. Panek; Bill Pikounis; Gordon Powers; Omid Vafa; Rich Siegel

2011-01-01

337

Prophylaxis and therapy of influenza pneumonia in mice by intratracheal instillation of monoclonal antibody  

SciTech Connect

This study on passive immunity dealt principally with the following topics: pathogenesis of the pneumonia produced by influenza virus (PR8) in CF-1 mice; the distribution and retention of monoclonal antibody instilled intratracheally (IT) into the lung; and prophylaxis and therapy of influenza pneumonia using specific monoclonal antibody (IgG 2a/k anti-HA). The fate of a single 50 ul bolus of antibody instilled IT was determined by monitoring the activity of /sup 125/I-labelled monoclonal IgG in the lungs and by lavage recovery of functional antibody.Antibody was demonstrated in high concentrations for the first 3 days and was present in the lungs for a period of 7 days. For prophylaxis several trials indicated that monoclonal antibody provided significant protection from lethal effects of the virus. Antibody given to clinically ill mice on day 3 produced a highly significant reduction in mortality (P < 0.001) when compared to control mice. The treatment reversed the weight loss and apparently arrested the development of lesions in most of the mice within 2 days following antibody administration.

Ratcliffe, D.R.

1985-01-01

338

Different antigen expression on Wolffian and Müllerian cells in rat embryos as detected by monoclonal antibodies  

Microsoft Academic Search

The Wolffian duct and the developing Müllerian duct of 14 and 15 day old rat embryos were examined with the monoclonal antibodies GZ1 and GZ2. These antibodies react with antigens situated in the cell membrane of Wolffian cells; they do not react with Müllerian cells. This different antigen expression confirms the current opinion that these cells are of different types.

C. Dohr; T. Tarmann; H. Schiechl

1987-01-01

339

A hybridoma secreting a neutralising monoclonal antibody to egg drop syndrome 1976 virus  

Microsoft Academic Search

A hybridoma (HPRS\\/AM\\/1) secreting neutralising antibody to Egg drop syndrome 1976 virus strain D61 was established. This monoclonal antibody also neutralised two other strains tested thereby showing that all the strains possess at least one common antigenic determinant which is important in infection. However, this epitope was not involved in haemagglutination. The antigen was identified in the nuclei of infected

A. P. A. Mockett; T. D. K. Brown; P. J. Davis

1984-01-01

340

Development and Characterization of Monoclonal Antibodies and Aptamers Against Major Antigens of Mycobacterium avium subsp. paratuberculosis  

Technology Transfer Automated Retrieval System (TEKTRAN)

Specific antibodies, available in unlimited quantities, have not been produced against Mycobacterium avium subsp. paratuberculosis, the bacterium that causes Johne’s disease (JD). To fill this gap in JD research, monoclonal antibodies (mAbs) against M. avium subsp. paratuberculosis were produced fr...

341

Production of monoclonal antibody against a glucosyltransferase of Streptococcus mutans 6715.  

PubMed Central

A mouse hybrid cell line secreted monoclonal antibody which reacted specifically with Streptococcus mutans 6715 (serotype g) glucosyltransferase (GTase)-synthesizing water-insoluble glucan and inhibited with enzyme reaction. The antibody was cross-reactive with GTase of serotype d but not with GTase of other serotypes of S. mutans when an enzyme-linked immunosorbent assay was used. Images PMID:6223885

Furuta, T; Nisizawa, T; Chiba, J; Hamada, S

1983-01-01

342

A Mechanistic Perspective of Monoclonal Antibodies in Cancer Therapy Beyond Target-Related Effects  

Microsoft Academic Search

Several monoclonal antibodies are now in clinical use for cancer therapy, and many others are currently undergo- ing clinical evaluation. These agents offer unique specific- ity against key molecular targets on tumor cells or in the tumor microenvironment. The clinical efficacy of mono- clonal antibodies is generally attributed to target-specific mechanisms resulting from neutralizing or inhibiting a growthfactororreceptorthatdrivescellproliferationand tumor growth.

SCOTT E. STROME; EDWARD A. SAUSVILLE

343

A Monoclonal Antibody Against Kinesin Inhibits Both Anterograde and Retrograde Fast Axonal Transport in Squid Axoplasm  

Microsoft Academic Search

One of our monoclonal antibodies against the heavy chain of bovine kinesin (H2) also recognized the heavy chain of squid kinesin. The immunofluorescence pattern of H2 in axoplasm was similar to that seen in mammalian cells with antibodies specific for kinesin light and heavy chains, indicating that squid kinesin is also concentrated on membrane-bounded organelles. Although kinesin is assumed to

Scott T. Brady; K. Kevin Pfister; George S. Bloom

1990-01-01

344

A Spectrum of Monoclonal Antibodies Reactive with Human Mammary Tumor Cells  

NASA Astrophysics Data System (ADS)

Splenic lymphocytes of mice, immunized with membrane-enriched fractions of metastatic human mammary carcinoma tissues, were fused with the NS-1 non-immunoglobulin-secreting murine myeloma cell line. This resulted in the generation of hybridoma cultures secreting immunoglobulins reactive in solid-phase radioimmunoassays with extracts of metastatic mammary carcinoma cells from involved livers, but not with extracts of apparently normal human liver. As a result of further screening of immunoglobulin reactivities and double cloning of cultures, 11 monoclonal antibodies were chosen that demonstrated reactivities with human mammary tumor cells and not with apparently normal human tissues. These monoclonal antibodies could be placed into at least five major groups on the basis of their differential binding to the surface of various live human mammary tumor cells in culture, to extracts of mammary tumor tissues, or to tissue sections of mammary tumor cells studied by the immunoperoxidase technique. Whereas a spectrum of reactivities to mammary tumors was observed with the 11 monoclonal antibodies, no reactivity was observed to apparently normal cells of the following human tissues: breast, lymph node, lung, skin, testis, kidney, thymus, bone marrow, spleen, uterus, thyroid, intestine, liver, bladder, tonsils, stomach, prostate, and salivary gland. Several of the antibodies also demonstrated a ``pancarcinoma'' reactivity, showing binding to selected non-breast carcinomas. None of the monoclonal antibodies showed binding to purified ferritin or carcinoembryonic antigen. Monoclonal antibodies of all five major groups, however, demonstrated binding to human metastatic mammary carcinoma cells both in axillary lymph nodes and at distal sites.

Colcher, D.; Horan Hand, P.; Nuti, M.; Schlom, J.

1981-05-01

345

Immunodiagnosis of human cysticercosis (Taenia solium) with antigens purified by monoclonal antibodies.  

PubMed Central

Monoclonal antibodies were generated from mice immunized with scolex protein antigen of Cysticercus cellulosae. Three monoclonal antibodies specific for cysticercal antigens, which did not show any cross-reactivity with Taenia solium or Taenia saginata antigens, were selected. Each monoclonal antibody coupled to Sepharose could purify one antigen, which appeared as a single band on polyacrylamide gel electrophoresis. When antigens purified by monoclonal antibodies were used to detect antibody in serum samples taken from patients with cysticercosis, taeniasis, and other parasitic infections in an enzyme-linked immunosorbent assay, cross-reactivity was observed until a serum dilution of 1:128 was reached. Since serum samples from unexposed subjects showed positive reactions until a dilution of 1:64 was reached, we chose a discriminative dilution (1:128) above which no cross-reaction was observed. The percent positive serum samples from cysticercosis patients was 100% by the enzyme-linked immunosorbent assay with any of the antigens purified by monoclonal antibodies. Images PMID:3611310

Nascimento, E; Tavares, C A; Lopes, J D

1987-01-01

346

Simple diagnosis of Encephalitozoon sp. microsporidial infections by using a panspecific antiexospore monoclonal antibody.  

PubMed Central

Microsporidia (phylum Microsproa) have recently become recognized as common opportunistic protozoans in the United States and worldwide, particularly affecting immunodeficient patients. Microsporidian organisms within the genus Encephalitozoon are the cause of nephrologic, ophthalmic, pneumologic, gastroenteric, and systemic infections. However, diagnosis of the small spores by light microscopy is difficult, even with newly developed and improved staining techniques. We have developed an anti-Encephalitozoon species monoclonal antibody-based immunoassay for easy diagnosis. A hybridoma was produced and selected following one main criterion: recognition by immunofluorescence of all known Encephalitozoon spores affecting humans. The selected monoclonal antibody-secreting hybridomas were characterized by enzyme-linked immunosorbent assay, immunofluorescence, Western blot, and immunoelectron microscopy using Encephalitozoon species from fresh and fixed samples from patients and from in vitro cultures. In the immunofluorescence assay, one monoclonal antibody, termed 3B6, strongly recognized Encephalitozoon cuniculi, E. hellem, and E. intestinalis. Monoclonal antibody 3B6 bound to other microsporidia (Nosema and Vairimorpha spp.) without cross-reacting with any other parasite, including Enterocytozoon bieneusi, fungus, or bacterium tested. In immunoelectron microscopy assays, monoclonal antibody 3B6 bound to the exospore of Encephalitozoon species, while in Western blot assays, it recognized three to seven antigens with molecular masses ranging from 34 to 117 kDa. We have developed a sensitive and specific monoclonal antibody-based immunoassay to diagnose common microsporidian infections, particularly with Encephalitozoon species. This is a new tool for identifying spores in bodily fluids and biopsy samples and is an efficient diagnostic test. Additionally, monoclonal antibody 3B6 can serve to assess the prevalence of microsporidial infections in immunodeficient and immunocompetent patients. PMID:9041420

Enriquez, F J; Ditrich, O; Palting, J D; Smith, K

1997-01-01

347

The production and characterization of protective monoclonal antibodies against Fasciola hepatica and Schistosoma mansoni  

E-print Network

Monoclonal Antibody Production ELISA 27 29 Immunofluorescence Assays Western Blot Analysis Biochemical modification of Antigenic Epitopes Affinity Chromatography Passive Protection 34 35 37 39 40 RESULTS 42 Monoclonal Antibody Production...' to ~* (Puerto Rican strain) in ELISA F I~h't' Sp 'f' M 1 1 Antibodies Cross React with Schistosoma mansoni (Puerto Rican strain) in IFA's F ' I ~ht' Sp 't' M* I 1 At'bd'CoR t 'thSht' (Kenyan strain) P ' T f of P t* t' t ~P' I h~t' * gM 1o 1 At bod 1psb...

Hicks, Carolyn Sue

1988-01-01

348

Monoclonal antibodies that distinguish between normal and denervated human acetylcholine receptor.  

PubMed

Ten monoclonal anti-human acetylcholine receptor (AChR) monoclonal antibodies (m.abs) all exhibited high avidity binding to the human AChR. None was able to inhibit alpha-bungarotoxin (alpha-Butx) binding to the receptor. Five distinct but partially overlapping antibody-binding regions were defined by competition experiments. Four antibodies, which competed with each other for one region on denervated human AChR and also bound to human fetal AChR, failed to bind appreciably to normal human AChR in solution, to normal AChR solubilized from 6 other species, or to human endplates in frozen sections. PMID:3082932

Whiting, P J; Vincent, A; Schluep, M; Newsom-Davis, J

1986-05-01

349

A monoclonal antibody detecting the Ly-9.2 (Lgp 100) cell-membrane alloantigen.  

PubMed

A mouse monoclonal cell line (20-1.5) was produced by the cell fusion method and the antibody secreted by this line defined the Ly-9.2 specificity--the reciprocal specificity to that previously identified as the Lgp 100 or the T100 molecule. Although most concentrated on lymph-node cells, the antigen is also found on thymocytes, spleen and bone-marrow cells as well as liver and brain tissue. The monoclonal antibody precipitates a 100,000 molecular moiety from thymocytes. The antigenic specificities appear to be highly immunogeneic and antibodies to these specificities contaminate many antisera. These sera are noncytotoxic as is the case with the monoclonal antibody even though it is of the IgG2a subclass. As with T100 or Lgp 100, the Ly-9 locus appears to be linked to the H-25 locus. PMID:6160098

Hogarth, P M; Craig, J; McKenzie, I F

1980-07-01

350

Virus mutation frequencies can be greatly underestimated by monoclonal antibody neutralization of virions.  

PubMed Central

Monoclonal antibody-resistant mutants have been widely used to estimate virus mutation frequencies. We demonstrate that standard virion neutralization inevitably underestimates monoclonal antibody-resistant mutant genome frequencies of vesicular stomatitis virus, due to phenotypic masking-mixing when wild-type (wt) virions are present in thousandsfold greater numbers. We show that incorporation of antibody into the plaque overlay medium (after virus penetration at 37 degrees C) can provide accurate estimates of genome frequencies of neutral monoclonal antibody-resistant mutant viruses in wt clones. By using this method, we have observed two adjacent G----A base transition frequencies in the I3 epitope to be of the order of 10(-4) in a wt glycine codon. This appears to be slightly lower than the frequencies observed at other sites for total (viable and nonviable) virus genomes when using a direct sequence approach. Images PMID:2479770

Holland, J J; de la Torre, J C; Steinhauer, D A; Clarke, D; Duarte, E; Domingo, E

1989-01-01

351

Characterization of a Novel Single-Chain Bispecific Antibody for Retargeting of T Cells to Tumor Cells via the TCR Co-Receptor CD8  

PubMed Central

There is currently growing interest in retargeting of effector T cells to tumor cells via bispecific antibodies (bsAbs). Usually, bsAbs are directed on the one hand to the CD3 complex of T cells and on the other hand to a molecule expressed on the surface of the target cell. A bsAb-mediated cross-linkage via CD3 leads to an activation of CD8+ T cells and consequently to killing of the target cells. In parallel, CD4+ T cells including TH1, TH2, TH17 cells and even regulatory T cells (Tregs) will be activated as well. Cytokines produced by CD4+ T cells can contribute to severe side effects e. g. life-threatening cytokine storms and, thinking of the immunosupressive function of Tregs, can even be counterproductive. Therefore, we asked whether or not it is feasible to limit retargeting to CD8+ T cells e. g. via targeting of the co-receptor CD8 instead of CD3. In order to test for proof of concept, a novel bsAb with specificity for CD8 and a tumor-associated surface antigen was constructed. Interestingly, we found that pre-activated (but not freshly isolated) CD8+ T cells can be retargeted via CD8-engaging bsAbs leading to an efficient lysis of target cells. PMID:24751697

Koristka, Stefanie; Arndt, Claudia; Cartellieri, Marc; Ehninger, Armin; Ehninger, Gerhard; Bachmann, Michael P.

2014-01-01

352

A phase II study of the bispecific antibody MDX-H210 (anti-HER2 × CD64) with GM-CSF in HER2+ advanced prostate cancer  

PubMed Central

The proto-oncogene HER2 presents a novel therapeutic target. We report results in 25 patients with HER2+ advanced prostate cancer treated with the bispecific antibody MDX-H210 15??g m?2by intravenous infusion plus GM-CSF 5??g?kg?1day?1by subcutaneous injection for 4 days repeated weekly for 6 weeks. Patients with stable disease or better received further cycles of treatment until disease progression or study withdrawal. 1 patient received no treatment and 4 received less than 1 cycle and are included in the toxicity analysis only. Median duration of follow up was 105+ (range 21–188) days. Toxicity was generally NCI-CTG 0–2. There were 2 grade 4 adverse events (heart failure and dyspnoea) and 1 grade 3 event (allergic reaction) resulting in discontinuation of the study medication. There were 9 further grade 3 events not resulting in trial withdrawal. There were no treatment-related deaths. 7/20 (35%) evaluable patients had a >50% PSA response of median duration 128 (range 71–184+) days. 7/12 (58%) patients with evaluable pain had improvements in pain scores. The PSA relative velocity on therapy decreased in 15/18 (83%) assessable patients compared to pre-study. GM-CSF and MDX-H210 is active in hormone refractory prostate carcinoma with acceptable toxicity; further studies are warranted. © 2001 Cancer Research Campaign http://www.bjcancer.com PMID:11461069

James, N D; Atherton, P J; Jones, J; Howie, A J; Tchekmedyian, S; Curnow, R T

2001-01-01

353

Radioimmunoassay for detecting antibodies against murine malarial parasite antigens: monoclonal antibodies recognizing Plasmodium yoelii antigens  

SciTech Connect

A solid-phase radioimmunoassay (SPRIA) in microtiter wells was established for detecting antibodies against Plasmodium yoelii Ag. The SPRIA was found (1) to require as little as 5 ..mu..g of crude parasite Ag per well, (2) to be able to detect 0.5 ng of monoclonal Ab, and (3) to be 10/sup 4/ times more sensitive than the indirect fluorescent Ab staining technique. In a modification of the above assay using intact RBC as an Ag, hyperimmune serum showed significant binding to the surface of erythrocytes of mice infected with P. yoelii parasites but not to RBC of normal mice. Hybridomas were prepared by fusing infected mouse spleen cells with myeloma cells. Using the SPRIA, hybrids secreting Ab against P. yoelii 17XL Ag were detected.

Kim, K.J.; Taylor, D.W.; Evans, C.B.; Asofsky, R.

1980-12-01

354

Chemotherapy Combinations With Monoclonal Antibodies in Non-Hodgkin’s Lymphoma  

PubMed Central

Although the use of monoclonal antibodies as single agents has had a tremendous impact on the care of patients with non-Hodgkin’s lymphoma (NHL), the greatest benefit has been generated by the addition of monoclonal antibodies to conventional cytotoxic chemotherapy. Rituximab is the monoclonal antibody responsible for all clinical improvement noted to date. The addition of rituximab to cyclophosphamide, doxorubicin, vincristine, and prednisone (CHOP) chemotherapy (R-CHOP regimen) improves the response rate, progression-free survival (PFS), and overall survival (OS) in diffuse large B-cell lymphoma (DLBCL). Adding rituximab to CHOP chemotherapy improves response rates and PFS in mantle cell lymphoma (MCL). Finally, the addition of rituximab to a variety of chemotherapy regimens improves the response rates, PFS, and OS in follicular lymphoma (FL). Several other (epratuzumab, bevacizumab, alemtuzumab) monoclonal antibody–chemotherapy combinations are currently under study in NHL. This review will summarize the data supporting the addition of rituximab to chemotherapy in NHL and discuss preliminary data regarding the use of other monoclonal antibodies in combination with chemotherapy. PMID:18381103

Kahl, Brad

2010-01-01

355

Human peripheral blood monocytes display surface antigens recognized by monoclonal antinuclear antibodies  

SciTech Connect

The authors used monoclonal anti-nuclear autoantibodies and indirect immunofluorescence to examine normal human peripheral blood mononuclear leukocytes for the presence of cell surface nuclear antigens. Only one monoclonal anti-histone antibody (MH-2) was found to bind to freshly isolated PBL, staining approximately 10% of large cells. However, after cells were placed into culture for 16-24 h, a high percentage (up to 60%) of large-sized cells were recognized by an anti-DNA (BWD-1) and several different antihistone monoclonal antibodies (BWH-1, MH-1, and MH-2). These antibodies recognize separate antigenic determinants on chromatin and histones extracted from chromatin. The histone antigen-positive cells were viable, and the monoclonal antibodies could be shown to be binding to the cell surface and not to the nucleus. Using monoclonal antibodies specific for monocytes and T cells, and complement-mediated cytotoxicity, the cells bearing histone antigens were shown to be primarily monocytes. The appearance of histone and DNA antigen-positive cells was nearly completely inhibited by the addition of low concentrations of cycloheximide at initiation of the cultures. In contrast, little effect on the percentage of positive cells was detected if cells were exposed to high doses of gamma irradiation before culture. These data further support the existence of cell surface nuclear antigens on selected cell subsets, which may provide insight into the immunopathogenesis of systemic lupus erythematosus and related autoimmune diseases.

Holers, V.M.; Kotzin, B.L.

1985-09-01

356

Preparation and characterization of monoclonal antibodies against native membrane-bound penicillin-binding protein 1B of Escherichia coli.  

PubMed Central

We prepared monoclonal antibodies against penicillin-binding protein 1B (PBP 1B) of Escherichia coli to study the membrane topology, spatial organization, and enzyme activities of this protein. The majority of the antibodies derived with PBP 1B as the immunogen reacted against the carboxy terminus. To obtain monoclonal antibodies recognizing other epitopes, we used PBP 1B lacking the immunodominant carboxy-terminal 65 amino acids as the immunogen. Eighteen monoclonal antibodies directed against membrane-bound PBP 1B were isolated and characterized. The epitopes recognized by those monoclonal antibodies were located with various truncated forms of PBP 1B. We could distinguish four different epitope areas located on different parts of the molecule. Interestingly, we could not isolate monoclonal antibodies against the amino terminus, although they were specifically selected for. This is attributed to its predicted extreme hydrophilicity and flexibility, which could make the amino terminus very sensitive to proteolytic degradation. All antibodies reacted against native PBP 1B in a dot-blot immunobinding assay. One monoclonal antibody also recognized PBP 1B in a completely sodium dodecyl sulfate-denatured form. This suggests that all the other monoclonal antibodies recognize conformational epitopes. These properties make the monoclonal antibodies suitable tools for further studies. Images PMID:2466033

Den Blaauwen, T; Wientjes, F B; Kolk, A H; Spratt, B G; Nanninga, N

1989-01-01

357

Comparison of monoclonal versus polyclonal calretinin antibodies for immunohistochemical diagnosis of malignant mesothelioma.  

PubMed

Of putative specific markers for diffuse malignant mesothelioma, nuclear staining with Zymed polyclonal calretinin antibody has shown the best specificity to date for epithelial diffuse malignant mesothelioma versus adenocarcinoma. We compared specificity and sensitivity of this polyclonal antibody for diagnosis of diffuse malignant mesothelioma with a new monoclonal antibody from DAKO. One hundred eighteen adenocarcinomas and 111 diffuse malignant mesotheliomas-70 epithelial, 22 sarcomatous, and 19 biphasic-were immunostained with calretinin antibodies from Zymed (polyclonal rabbit, prediluted, PAD:DC8) and DAKO(monoclonal mouse, 1:100, clone DAK Calret 1) using manufacturer-recommended procedures. Cases were blinded and assessed for nuclear versus cytoplasmic staining, percent positive cells, and background. Both antibodies showed similar positive predictive values for diffuse malignant mesothelioma by nuclear staining (Zymed=95%; DAKO=97%). False positives in 4 (3.4%) and 2 (1.7%) adenocarcinomas, respectively, stained greater than 10% of cells. Sensitivity for epithelial malignant mesothelioma was slightly less for DAKO antibody (Zymed=80%; DAKO=73%). Neither antibody performed well on sarcomatous malignant mesothelioma (Zymed=2/22; DAKO=1/22). Both antibodies are useful in the diagnosis of epithelial malignant mesothelioma, although monoclonal antibody is slightly less sensitive. PMID:15722797

Granville, Laura A; Younes, Mamoun; Churg, Andrew; Roggli, Victor L; Henderson, Douglas W; Cagle, Philip T

2005-03-01

358

Detection of antibodies to equine arteritis virus by a monoclonal antibody-based blocking ELISA.  

PubMed Central

A potent ELISA antigen was prepared from equine arteritis virus (EAV) by differential centrifugation of EAV-infected cell culture fluid, followed by solubilization of the preparation by Triton X-100 treatment. Using this antigen and a mouse monoclonal antibody against the G(L) protein of EAV, a reliable blocking ELISA (bELISA) was developed for the detection of EAV antibodies in equine sera. The bELISA was evaluated using a total of 837 test serum samples. The relative sensitivity (n = 320) of the bELISA compared to the serum neutralization (SN) test was 99.4%. The bELISA appears to be a highly specific test, the specificity of which did not appear to be adversely affected by previous exposure of horses to non-EAV-containing biologicals. Of 119 serum samples, 21 from horses without any history of exposure to EAV and 98 from racetrack Thoroughbreds, 118 were negative in the SN test and bELISA. One sample was SN-negative but suspicious with the bELISA. Based on testing 465 SN-negative field samples and 52 SN-negative samples from experimental horses, and excluding any sera giving a suspicious reaction, the relative specificity of the bELISA was 97.7%. Samples should be examined undiluted and diluted 1/10 in the bELISA because the testing of sera of high neutralizing antibody titer may be affected by a prozone-like phenomenon. The bELISA is a more rapid and cost-efficient test than the SN test for the detection of EAV antibodies in equine sera. PMID:10680655

Cho, H J; Entz, S C; Deregt, D; Jordan, L T; Timoney, P J; McCollum, W H

2000-01-01

359

Characteristics and performance of a bispecific F (ab'gamma)2 antibody for delivering saporin to a CD7+ human acute T-cell leukaemia cell line.  

PubMed Central

We have investigated the efficacy of a F(ab'gamma)2 bispecific antibody (BsAb) with dual specificity for the CD7 molecule in one Fab arm and for the ribosome inactivating protein (rip) saporin in the other arm, for delivering saporin to the acute T-cell leukaemia cell line HSB-2. Saporin titration experiments revealed that BsAb increased the toxicity of saporin 435-fold for HSB-2 cells, reducing the IC50 for saporin alone from 0.1 mumol to 0.23 nmol when BsAb was included. The rate of protein synthesis inactivation brought about by BsAb-mediated toxin delivery to HSB-2 cells was very similar to that described for conventional immunotoxins (IT's) with a t10 (time taken for a one log inhibition of protein synthesis compared with controls) of 46 h obtained at a saporin concentration of 1 nmol and 226 h at 0.1 nmol. BsAb titration studies demonstrated a clear dose response effect of BsAb concentration on target cell protein synthesis inhibition and cell proliferation. The absolute specificity of toxin delivery was unequivocally demonstrated by a failure of BsAb to deliver an effective dose of saporin to the CD7- cell line HL60 and by the blocking of BsAb-mediated delivery of saporin to HSB-2 cells with an excess of F(ab)2 fragments of the anti-CD7 antibody, HB2. These studies have clearly demonstrated the effectiveness of this BsAb for delivering saporin to a T-ALL cell line utilising CD7 as the target molecule on the cell surface. BsAb's would therefore appear to offer a realistic alternative to IT's for toxin delivery to tumour cells and may even offer certain advantages over conventional IT's for clinical use. PMID:1716453

Flavell, D. J.; Cooper, S.; Morland, B.; Flavell, S. U.

1991-01-01

360

DETECTION OF ROTAVIRUS WITH A NEW POLYCLONAL ANTIBODY ENZYME IMMUNOASSAY (ROTAZYME 2) AND A COMMERCIAL LATEX AGGLUTINATION TEXT (ROTALEX): COMPARISON WITH A MONOCLONAL ANTIBODY ENZYME IMMUNOASSAY  

EPA Science Inventory

A total of 176 human fecal specimens were examined for the presence of rotavirus using four different assays: a monoclonal antibody enzyme immunoassay; the original polyclonal antibody enzyme immunoassay marketed by Abbott Laboratories, Chicago, IL (Rotazyme I); a modification of...

361

Development of monoclonal antibodies against parathyroid hormone: genetic control of the immune response to human PTH  

SciTech Connect

Seventeen monocloanl antibodies against the aminoterminal portion of parathyroid hormone (PTH) were generated by using BALB/c mouse for immunization fully biologically active synthetic human PTH-(1-34) and bovine PTH-(1-84) as immunogens, monoclonal antibody methods, and a solid-phase screening assay. Isotypic analysis of these monoclonal antibodies was performed using affinity purified goat antimouse immunoglobulins specific for IgG heavy chains and ..mu..(IgM). All antibodies were IgM as evidenced by 40 times greater than background activity when 25,000 cpm of /sup 125/I-labelled goat anti-mouse IgM was used as second antibody in a radioimmunoassay.

Nussbaum, S.R.; Lin, C.S.; Potts, J.T. Jr.; Rosenthal, A.S.; Rosenblatt, M.

1985-01-01

362

Monoclonal antibodies against Nereis virens brain homogenates as probes for isolation and in situ detection of new neurosecretions.  

PubMed

The brain of Nereis contains 26 ganglionic nuclei which produce numerous neurosecretions. Only a few of them have been characterized. The production of monoclonal antibodies was adopted as an approach to discover unknown neurosecretions. Monoclonal antibodies produced against Nereis virens brain homogenates were selected using stepwise ELISA tests first with brain homogenates, then with brain neurosecretions. Eight antibodies specific for Nereis neurosecretions were selected. The results are illustrated with one of these monoclonal antibodies which was directed against a major peak after HPLC purification of brain neurosecretions. This antibody was subsequently used for the in situ detection of recognized epitope(s) in the brain and ventral nerve cord cells. PMID:1473354

Delaire-Hesdin, A; Bulet, P; Boilly-Marer, Y; Porchet-Hennere, E; Masson, M

1992-01-01

363

Australian consensus guidelines for the safe handling of monoclonal antibodies for cancer treatment by healthcare personnel.  

PubMed

These consensus guidelines provide recommendations for the safe handling of monoclonal antibodies. Definitive recommendations are given for the minimum safe handling requirements to protect healthcare personnel. The seven recommendations cover: (i) appropriate determinants for evaluating occupational exposure risk; (ii) occupational risk level compared with other hazardous and non-hazardous drugs; (iii) stratification of risk based on healthcare personnel factors; (iv) waste products; (v) interventions and safeguards; (vi) operational and clinical factors and (vii) handling recommendations. The seventh recommendation includes a risk assessment model and flow chart for institutions to consider and evaluate clinical and operational factors unique to individual healthcare services. These guidelines specifically evaluated monoclonal antibodies used in the Australian cancer clinical practice setting; however, the principles may be applicable to monoclonal antibodies used in non-cancer settings. The guidelines are only applicable to parenterally administered agents. PMID:25302720

Alexander, M; King, J; Bajel, A; Doecke, C; Fox, P; Lingaratnam, S; Mellor, J D; Nicholson, L; Roos, I; Saunders, T; Wilkes, J; Zielinski, R; Byrne, J; MacMillan, K; Mollo, A; Kirsa, S; Green, M

2014-10-01

364

Monoclonal antibodies that demonstrate specificity for several types of human lung cancer.  

PubMed Central

Monoclonal antibodies with selectivity for human lung cancer were produced by immunizing BALB/c mice with an established line of human small cell lung cancer (NCI-H69) and fusing the mouse spleen cells to mouse myeloma line X63-Ag8.653. The resulting hybrid cells were initially screened by immunoautoradiography for production of antibodies that would react with NCI-H69 and another small cell lung cancer line (NCI-H128) but not its autologous B-lymphoblastoid line (NCI-H128BL). Stable monoclonal antibody-producing lines were isolated by repeated cloning. Three independently derived monoclonal antibodies, designated 525A5, 534F8, and 538F12, were found to react with three of the major types of human lung cancer (small cell, adenocarcinoma, and squamous carcinoma). They did not react with bronchioloalveolar and large cell lung cancers, myeloma, lymphomas, leukemias, osteogeneic sarcoma, mesothelioma, hypernephroma, malignant melanoma, simian virus 40-transformed human fetal lung cells, skin fibroblast lines, human B-lymphoblastoid lines, human erythrocytes, and rodent cells. Interestingly, these antibodies also bound to three out of three human neuroblastomas and two out of three breast cancers but failed to react with mouse neuroblastoma and rat pheochromocytoma. The monoclonal antibodies reacted with human small cell lung cancer tumors obtained at autopsy, but had insignificant reactions with normal human lung, liver, spleen, and skeletal muscle. We conclude that monoclonal antibodies have been generated that react with common antigenic determinants expressed on several human lung cancer types, neuroblastoma, and some breast cancers, but are not detectable by our current assays on a variety of other human tumors or normal adult human tissues. Such antibodies are of potential clinical and biological importance. PMID:6270685

Cuttitta, F; Rosen, S; Gazdar, A F; Minna, J D

1981-01-01

365

Quantitative explanation for increased affinity shown by mixtures of monoclonal antibodies: importance of a circular complex.  

PubMed

Mixtures of some but not all monoclonal antibodies which bind to separate epitopes on human chorionic gonadotropin (hCG) show an increased affinity for the hormone. To find an explanation for the increase in affinity, we developed a mathematical model which predicts the quantities of intermediates formed when pairs of IgG1 mouse monoclonal antibodies having affinities of approximately 10(8) M-1 for hCG are mixed with the hormone. At low antibody concentrations (i.e. less than 1 nM or 0.15 micrograms/ml) analysis of possible antibody-hormone combinations, including linear and circular chains composed of less than 12 molecules of antibody and 12 molecules of hCG, suggests the increase in affinity is due to formation of a circular complex containing two molecules of antibody and two of hCG. Further, the model predicts that the circular complex will be the major species formed at antibody-antigen equivalence. This prediction is supported by experimental observations on the molecular weight of a new complex formed in the presence of hCG and the mixture of the monoclonal antibodies. In addition, based on experimental values of binding constants for individual antibodies to hCG, the model correctly quantifies the loss in complex observed in the presence of excess hCG antigen. At high antibody concentrations (i.e. greater than 10 nM or 1.5 micrograms/ml) the formation of linear chains of antibody hCG pairs becomes appreciable and contributes to the increase in apparent affinity of the mixture for hCG. These results suggest that the observed affinity of complex mixtures of antibody for antigens containing multiple epitopes calculated from Scatchard plots may not be related to the affinity or avidity of any of the antibody species for a given epitope. PMID:6191209

Moyle, W R; Lin, C; Corson, R L; Ehrlich, P H

1983-04-01

366

Studies on the syngeneic mixed lymphocyte reaction. III. Development of a monoclonal antibody with specificity for autoreactive T cells  

PubMed Central

Monoclonal antibodies with specificity for autoreactive murine T cells have been developed. These antibodies inhibit proliferative response of splenic T cells activated by syngeneic spleen cells. These antibodies have no effect on the proliferative response of T cells activated by allogeneic spleen cells or PHA. The number of splenic T cells that react with these monoclonal antibodies is comparable in several normal mouse strains. PMID:6225824

1983-01-01

367

8th Annual European Antibody Congress 2012  

PubMed Central

The 8th European Antibody Congress (EAC), organized by Terrapin Ltd., was again held in Geneva, Switzerland, following on the tradition established with the 4th EAC. The new agenda format for 2012 included three parallel tracks on: (1) naked antibodies; (2) antibody drug conjugates (ADCs); and (3) bispecific antibodies and alternative scaffolds. The meeting started and closed with three plenary lectures to give common background and to share the final panel discussion and conclusions. The two day event included case studies and networking for nearly 250 delegates who learned of the latest advances and trends in the global development of antibody-based therapeutics. The monoclonal antibody track was focused on understanding the structure-function relationships, optimization of antibody design and developability, and processes that allow better therapeutic candidates to move through the clinic. Discussions on novel target identification and validation were also included. The ADC track was dedicated to evaluation of the ongoing success of the established ADC formats alongside the rise of the next generation drug-conjugates. The bispecific and alternative scaffold track was focused on taking stock of the multitude of bispecific formats being investigated and gaining insight into recent innovations and advancements. Mechanistic understanding, progression into the clinic and the exploration of multispecifics, redirected T cell killing and alternative scaffolds were extensively discussed. In total, nearly 50 speakers provided updates of programs related to antibody research and development on-going in the academic, government and commercial sectors. PMID:23493119

Beck, Alain; Carter, Paul J.; Gerber, Hans-Peter; Lugovskoy, Alexey A.; Wurch, Thierry; Junutula, Jagath R.; Kontermann, Roland E; Mabry, Robert

2013-01-01

368

False positive Legionella pneumophila direct immunofluorescent monoclonal antibody test caused by Bacillus cereus spores.  

PubMed

Direct immunofluorescent monoclonal antibody stain testing for Legionella pneumophila in Oklahoma lake water yielded an unknown bacillus with fluorescence intensity equal to that of L. pneumophila stock strains. The organism in question was identified as Bacillus cereus, a ubiquitous bacterium. When B. cereus cultures were studied, fluorescence was seen in spores but not in vegetative cells. Since a positive immunofluorescent monoclonal antibody test (alone) might be considered by some individuals as unequivocal to very good evidence for the presence of L. pneumophila, this finding emphasizes the importance of confirming positive stain results with cultures whenever possible. PMID:3133155

Flournoy, D J; Belobraydic, K A; Silberg, S L; Lawrence, C H; Guthrie, P J

1988-02-01

369

Monoclonal antibodies to antigens on human neutrophils, activated T lymphocytes, and acute leukemia blast cells  

SciTech Connect

The authors describe the production of two mouse hybridomas secreting monoclonal antibodies to antigenic determinants of the surface membranes of human neutrophils, activated T lymphocytes, and acute leukemic blast cells. The degree of lymphocyte stimulation was estimated from incorporation of /sup 3/H-thymidine with parallel microculture. Monoclonal antibodies of supernatants of hybridoma cultures shown here reacted in both immunofluorescence test and cytotoxicity test with surface membrane antigens on the majority of neutrophils and PHA-activated peripheral blood lymphocytes from healthy subjects, but did not give positive reactions with unactivated lymphocytes, adherent monocytes, erythrocytes, and alloantigen-stimulated lymphocytes.

Miterev, G.Yu.; Burova, G.F.; Puzhitskaya, M.S.; Danilevich, S.V.; Bulycheva, T.I.

1987-11-01

370

Novel monoclonal antibody inhibits tumor growth in breast cancer and angiosarcoma  

Cancer.gov

A monoclonal antibody targeting a protein known as SFPR2 has been shown by researchers at the University of North Carolina and its Lineberger Comprehensive Cancer Center to inhibit tumor growth in pre-clinical models of breast cancer and angiosarcoma. In a paper published in the April 19 issue of Molecular Cancer Therapeutics, a team used a monoclonal antibody to target SFRP2 expressed in cells from triple-negative breast cancer and the aggressive blood-vessel malignancy angiosarcoma, reducing the rate of tumor growth.

371

Cytokine-induced killer cells targeted by the novel bispecific antibody CD19xCD5 (HD37xT5.16) efficiently lyse B-lymphoma cells  

Microsoft Academic Search

Due to their dual binding capacity, bispecific antibodies (bsAb) can be used to cross-link cytotoxic effector cells with malignant\\u000a targets and may thereby improve adoptive immunotherapy. In this study, the development and preclinical testing of the quadroma-derived\\u000a bsAb HD37xT5.16 of the specificity CD19xCD5 is reported. Effector cells used were a population of ex vivo expanded and activated\\u000a T cells called

Freddy Tita-Nwa; Gerhard Moldenhauer; Markus Herbst; Christian Kleist; Anthony D. Ho; Martin Kornacker

2007-01-01

372

Characterization of monoclonal antibodies to a crystal protein of Bacillus thuringiensis subsp. kurstaki.  

PubMed

Ten monoclonal antibodies were produced against a k-1-type crystal protein of Bacillus thuringiensis subsp. kurstaki. Eight of the antibodies belong to the immunoglobulin G1 (IgG1) subclass, with pI values ranging from 5.5 to 8.6, one could be assigned to the IgG2b subclass, and one could be assigned to the IgM class. Competitive antibody-binding assays and analysis of antibody specificity indicated that the 10 antibodies recognized at least nine distinct antigenic determinants. Eight antibodies bound to both protoxin and toxin, whereas the other two interacted with protoxin only. One antibody completely inhibited the biological activity of the delta-endotoxin, five antibodies reduced it by 15 to 82%, and four antibodies did not affect it at all. Based on cross-reaction studies, homologies and differences in the crystal protein structures of different B. thuringiensis subspecies were revealed. All of the monoclonal antibodies strongly cross-reacted with crystal proteins from strains of B. thuringiensis subsp. tolworthi, B. thuringiensis subsp. galleriae, B. thuringiensis subsp. dendrolimus, B. thuringiensis subsp. sotto, and B. thuringiensis subsp. subtoxicus. Some antibodies interacted only weakly with crystal proteins from strains of B. thuringiensis subsp. morrisoni and B. thuringiensis subsp. entomocidus, and some of these did not interact with B. thuringiensis subsp. kenyae and B. thuringiensis subsp. darmstadiensis. No cross-reaction was found with the parasporal inclusion protein of B. thuringiensis subsp. israelensis. Studies with the monoclonal antibodies also disclosed that crystal proteins from strains of the same subspecies can exhibit substantial differences in antigenic structure. In particular, striking strain-specific differences in the protoxins of B. thuringiensis subsp. kurstaki and B. thuringiensis subsp. thuringiensis were observed. PMID:3759236

Huber-Lukac, M; Jaquet, F; Luethy, P; Huetter, R; Braun, D G

1986-10-01

373

Discovery of lung cancer biomarkers by profiling the plasma proteome with monoclonal antibody libraries.  

PubMed

A challenge in the treatment of lung cancer is the lack of early diagnostics. Here, we describe the application of monoclonal antibody proteomics for discovery of a panel of biomarkers for early detection (stage I) of non-small cell lung cancer (NSCLC). We produced large monoclonal antibody libraries directed against the natural form of protein antigens present in the plasma of NSCLC patients. Plasma biomarkers associated with the presence of lung cancer were detected via high throughput ELISA. Differential profiling of plasma proteomes of four clinical cohorts, totaling 301 patients with lung cancer and 235 healthy controls, identified 13 lung cancer-associated (p < 0.05) monoclonal antibodies. The monoclonal antibodies recognize five different cognate proteins identified using immunoprecipitation followed by mass spectrometry. Four of the five antigens were present in non-small cell lung cancer cells in situ. The approach is capable of generating independent antibodies against different epitopes of the same proteins, allowing fast translation to multiplexed sandwich assays. Based on these results, we have verified in two independent clinical collections a panel of five biomarkers for classifying patient disease status with a diagnostics performance of 77% sensitivity and 87% specificity. Combining CYFRA, an established cancer marker, with the panel resulted in a performance of 83% sensitivity at 95% specificity for stage I NSCLC. PMID:21947365

Guergova-Kuras, Mariana; Kurucz, István; Hempel, William; Tardieu, Nadčge; Kádas, János; Malderez-Bloes, Carole; Jullien, Anne; Kieffer, Yann; Hincapie, Marina; Guttman, András; Csánky, Eszter; Dezso, Balázs; Karger, Barry L; Takács, László

2011-12-01

374

Monoclonal antibodies to type X collagen. Biosynthetic studies using an antibody to the amino-terminal domain.  

PubMed

Monoclonal antibodies to chick type X collagen have been used to study the structure, biosynthesis, and location of type X in cartilage. The antibodies were produced by injecting purified type X collagen into female SJL/J mice and then fusing their spleen cells with Sp2/0 myeloma cells. Hybridoma culture supernatants were screened for antibodies to type X collagen by enzyme-linked immunosorbent assay and Western blots. Positive supernatants did not cross-react with other collagen types (I, II, IX, XI) or with fibronectin. Three monoclonal antibodies were chosen for further characterization. Two of them (1A6 and 6F6) recognize a pepsin-sensitive domain of type X collagen. Rotary shadowing showed that 1A6 and 6F6 both recognize the same end of type X, probably the aminoterminal non-triple helical domain. Amino acid sequencing of the intact protein and of the epitope-containing peptide confirmed that the antibody recognition sites for 1A6 and 6F6 are within the amino-terminal domain. Monoclonal antibody 2B3 reacts with the pepsinized (45 kDa) and weakly with the nonpepsinized (59 kDa) forms of type X collagen. The monoclonal antibodies were used for immunolocalization of type X in hypertrophic chondrocytes and reacted only with tissue samples from areas undergoing endochondral ossification, e.g. growth plate and fracture callus. Antibody 6F6, when coupled to Sepharose, selectively binds to type X collagen from cell and organ cultures. In a pulse-chase experiment, no processing of the 59-kDa form of type X could be detected. Two components with molecular masses of approximately 70 and 85 kDa, arising from a disulfide-bonded aggregate, were synthesized by both the permanent and calcifying cartilage organ cultures but did not react with the antibody, suggesting that these proteins are not related to type X. In summary, the pulse-chase results and the immune precipitation with monoclonal antibody 6F6 did not detect biosynthetic precursors larger than 59 kDa or proteolytically processed forms of type X. PMID:2826450

Summers, T A; Irwin, M H; Mayne, R; Balian, G

1988-01-01

375

Hybridoma-derived monoclonal immunoglobulin M antibodies to Legionella pneumophila serogroup 1 with diagnostic potential.  

PubMed Central

Mouse hybridomas were isolated by fusing P3-X63-Ag 8.653 myeloma cells with spleen cells from mice that had been repeatedly immunized with Legionella pneumophila serogroup 1 organisms. In one fusion, three independent hybridoma cultures which secreted antibodies that reacted with the immunizing strain in the indirect immunofluorescent-antibody test were selected for cloning. Representative continuously growing clones, one of each hybridoma, which remained stable in producing high-titer antibodies were examined in detail. Extensive specificity tests revealed that these hybridoma-derived monoclonal antibodies were specifically directed against L. pneumophila serogroup 1 organisms and showed no cross-reactions in the indirect immunofluorescent-antibody test either with the other known serogroups of L. pneumophila or with other unrelated bacterial species. The three monoclonal antibodies F4/CB5/K18, F/4CB5/K104, and F4/JD3.8/K101 belonged to the immunoglobulin M class and were capable of agglutinating serogroup 1 organisms of L. pneumophila exquisitely. These monoclonal antibodies against L. pneumophila with defined fine specificity should enable purification and subsequent analysis of the corresponding antigenic determinant(s) and can also be used for the preparation of unlimited supplies of standard diagnostic reagents for the identification of L. pneumophila in the tissues and body fluids. PMID:6874913

Sethi, K K; Drüeke, V; Brandis, H

1983-01-01

376

Effect of kinase inhibitors on the therapeutic properties of monoclonal antibodies.  

PubMed

Targeted therapies of malignancies currently consist of therapeutic monoclonal antibodies and small molecule kinase inhibitors. The combination of these novel agents raises the issue of potential antagonisms. We evaluated the potential effect of 4 kinase inhibitors, including the Bruton tyrosine kinase inhibitor ibrutinib, and 3 PI3K inhibitors idelalisib, NVP-BEZ235 and LY294002, on the effects of the 3 monoclonal antibodies, rituximab and obinutuzumab (directed against CD20) and trastuzumab (directed against HER2). We found that ibrutinib potently inhibits antibody-dependent cell-mediated cytotoxicity exerted by all antibodies, with a 50% inhibitory concentration of 0.2 microM for trastuzumab, 0.5 microM for rituximab and 2 microM for obinutuzumab, suggesting a lesser effect in combination with obinutuzumab than with rituximab. The 4 kinase inhibitors were found to inhibit phagocytosis by fresh human neutrophils, as well as antibody-dependent cellular phagocytosis induced by the 3 antibodies. Conversely co-administration of ibrutinib with rituximab, obinutuzumab or trastuzumab did not demonstrate any inhibitory effect of ibrutinib in vivo in murine xenograft models. In conclusion, some kinase inhibitors, in particular, ibrutinib, are likely to exert inhibitory effects on innate immune cells. However, these effects do not compromise the antitumor activity of monoclonal antibodies in vivo in the models that were evaluated. PMID:25523586

Duong, Minh Ngoc; Matera, Eva-Laure; Mathé, Doriane; Evesque, Anne; Valsesia-Wittmann, Sandrine; Clémenceau, Béatrice; Dumontet, Charles

2015-01-01

377

Multiple signaling pathways induced by hexavalent, monospecific, anti-CD20 and hexavalent, bispecific, anti-CD20/CD22 humanized antibodies correlate with enhanced toxicity to B-cell lymphomas and leukemias  

PubMed Central

We have generated hexavalent antibodies (HexAbs) comprising 6 Fabs tethered to one Fc of human IgG1. Three such constructs, 20-20, a monospecific HexAb comprising 6 Fabs of veltuzumab (humanized anti-CD20 immunoglobulin G1? [IgG1?]), 20-22, a bispecific HexAb comprising veltuzumab and 4 Fabs of epratuzumab (humanized anti-CD22 IgG1?), and 22-20, a bispecific HexAb comprising epratuzumab and 4 Fabs of veltuzumab, were previously shown to inhibit pro-liferation of several lymphoma cell lines at nanomolar concentrations in the absence of a crosslinking antibody. We now report an in-depth analysis of the apoptotic and survival signals induced by the 3 HexAbs in Burkitt lymphomas and provide in vitro cytotoxicity data for additional lymphoma cell lines and also chronic lymphocytic leukemia patient specimens. Among the key findings are the significant increase in the levels of phosphorylated p38 and phosphatase and tensin homolog deleted on chromosome 10 (PTEN) by all 3 HexAbs and the notable differences in the signaling events triggered by the HexAbs from those incurred by crosslinking veltuzumab or rituximab with a secondary antibody. Thus, the greatly enhanced direct toxicity of these HexAbs correlates with their ability to alter the basal expression of various intracellular proteins involved in regulating cell growth, survival, and apoptosis, with the net outcome leading to cell death. PMID:20628151

Gupta, Pankaj; Goldenberg, David M.; Rossi, Edmund A.

2010-01-01

378

Evaluation of monoclonal antibody-based capture enzyme immunoassays for detection of specific antibodies to measles virus.  

PubMed Central

Monoclonal antibodies to the hemagglutinin protein, fusion protein, phosphoprotein, matrix protein, and nucleoprotein of measles virus were evaluated as detector antibodies in capture enzyme immunoassays (EIAs) for the detection of specific serum immunoglobulin G (IgG), IgA, and IgM antibodies to measles virus. A pool of monoclonal antibodies to hemagglutinin protein and nucleoprotein proved optimal and was further evaluated. Specific IgM was detected in 97% of adolescents with clinical measles, 97% of infants 3 weeks postvaccination, and less than 1% of normal serum specimens. Specific IgA antibodies were found in 97% of adolescents with clinical measles, 97% of infants 3 weeks postvaccination, and less than 1% of normal serum specimens. Specific IgA antibodies were found in 97% of clinical measles cases and vaccinees, in 26% of healthy persons, and in 36% of infants 8 months postvaccination; consequently, IgA antibodies were not a useful indicator of recent measles infection. A significant increase in IgG antibodies between paired specimens was detected in 92% of clinical cases and all vaccinees. Only 59% of infant specimens had persistent IgG antibodies as detected by capture EIA at 8 months postvaccination, whereas all specimens had antibodies as detected by hemagglutination inhibition and plaque neutralization. An alternative indirect EIA, in which antigen was directly absorbed to the solid phase, was more sensitive than the capture design, detecting IgG antibodies in all infants postvaccination. When standardized with a microneutralization assay for the detection of persistent antibodies, the indirect IgG EIA gave predictive values for positive and negative tests exceeding 90%. Our capture IgM and indirect IgG EIAs provide a practical combination of serologic tests for the determination of acute measles virus infection and past exposure to measles virus or vaccine, respectively. PMID:1885743

Erdman, D D; Anderson, L J; Adams, D R; Stewart, J A; Markowitz, L E; Bellini, W J

1991-01-01

379

An epithelial cell adhesion molecule- and CD3-bispecific antibody plus activated T-cells can eradicate chemoresistant cancer stem-like pancreatic carcinoma cells in vitro.  

PubMed

Cancer stem-like properties of various types of cancer, including pancreatic cancer, one of the most aggressive types, correlate with metastasis, invasion, and therapeutic resistance. More importantly, chemoresistance in cancer stem-like cells (CSLCs) is a critical problem for eradication of pancreatic cancer. Several cell surface markers, such as CD44 and epithelial cell adhesion molecule (EpCAM), are molecular targets on CSLCs of pancreatic carcinoma. In this study, we investigated whether catumaxomab, a clinical-grade bi-specific antibody that binds to both EpCAM on tumor cells and CD3 on T-cells, combined with activated T-cells can eliminate chemoresistant pancreatic CSLCs in vitro. Firstly, we established a CSLC line (MU-PK1) from human pancreatic carcinoma cells derived from a patient with chemoresistant and disseminated pancreatic cancer. These CSLCs were almost completely resistant to gemcitabine-mediated cytotoxicity up to a concentration of 10 ?g/ml. The cells expressed high levels of CSLC markers (CD44 and EpCAM) and had significantly higher capacities for sphere formation, invasion, and aldehyde dehydrogenase-1 expression, which are associated with cancer stemness properties. We found that pre-treatment with catumaxomab and subsequent addition of interleukin-2/OKT3 activated autologous T-cells eliminated CSLCs during a short incubation period. Moreover, when MU-PK1 cells were cultured under hypoxic conditions, the CSLCs became more aggressive. However, the combination of cytokine-activated killer T-cells with catumaxomab successfully lysed almost all these cells. In conclusion, catumaxomab combined with activated T-cells may be a potent therapeutic modality to eradicate chemoresistant pancreatic CSLCs. PMID:25075094

Umebayashi, Masayo; Kiyota, Akifumi; Koya, Norihiro; Tanaka, Hiroto; Onishi, Hideya; Katano, Mitsuo; Morisaki, Takashi

2014-08-01

380

A dimeric bispecific miniantibody combines two specificities with avidity  

Microsoft Academic Search

Bispecific antibodies extend the capabilities of nature and might be applied in immunotherapy and biotechnology. By fusing the gene of a single-chain Fv (scFv) fragment to a helical dimerization domain, followed by a second scFv fragment of different specificity, we were able to express a functional protein in E. coli, which is bispecific and has two valencies for each specificity.

Kristian M. Müller; Katja M. Arndt; Andreas Plückthun

1998-01-01

381

Human Monoclonal Antibodies Against a Plethora of Viral Pathogens From Single Combinatorial Libraries  

NASA Astrophysics Data System (ADS)

Conventional antibody generation usually requires active immunization with antigen immediately prior to the preparation procedure. Combinatorial antibody library technology offers the possibility of cloning a range of antibody specificities at a single point in time and then accessing these specificities at will. Here we show that human monoclonal antibody Fab fragments against a plethora of infectious agents can be readily derived from a single library. Further examination of a number of libraries shows that whenever antibody against a pathogen can be detected in the serum of the donor, then specific antibodies can be derived from the corresponding library. We describe the generation of human Fab fragments against herpes simplex virus types 1 and 2, human cytomegalovirus, varicella zoster virus, rubella, human immunodeficiency virus type 1, and respiratory syncytial virus. The antibodies are shown to be highly specific and a number are effective in neutralizing virus in vitro.

Williamson, R. Anthony; Burioni, Roberto; Sanna, Pietro P.; Partridge, Lynda J.; Barbas, Carlos F., III; Burton, Dennis R.

1993-05-01

382

Human monoclonal antibodies against a plethora of viral pathogens from single combinatorial libraries.  

PubMed Central

Conventional antibody generation usually requires active immunization with antigen immediately prior to the preparation procedure. Combinatorial antibody library technology offers the possibility of cloning a range of antibody specificities at a single point in time and then accessing these specificities at will. Here we show that human monoclonal antibody Fab fragments against a plethora of infectious agents can be readily derived from a single library. Further examination of a number of libraries shows that whenever antibody against a pathogen can be detected in the serum of the donor, then specific antibodies can be derived from the corresponding library. We describe the generation of human Fab fragments against herpes simplex virus types 1 and 2, human cytomegalovirus, varicella zoster virus, rubella, human immunodeficiency virus type 1, and respiratory syncytial virus. The antibodies are shown to be highly specific and a number are effective in neutralizing virus in vitro. Images Fig. 1 Fig. 2 PMID:7683424

Williamson, R A; Burioni, R; Sanna, P P; Partridge, L J; Barbas, C F; Burton, D R

1993-01-01

383

Monoclonal antibodies to human hemoglobin S and cell lines for the production thereof  

DOEpatents

The present invention provides monoclonal antibodies specific to and distinguish between hemoglobin S and hemoglobin A and methods for their production and use. These antibodies are capable of distinguishing between two hemoglobin types which differ from each other by only a single amino acid residue. The antibodies produced according to the present method are useful as immunofluorescent markers to enumerate circulating red blood cells which have the property of altered expression of the hemoglobin gene due to somatic mutation in stem cells. Such a measurement is contemplated as an assay for in vivo cellular somatic mutations in humans. Since the monoclonal antibodies produced in accordance with the instant invention exhibit a high degree of specificity to and greater affinity for hemoglobin S, they are suitable for labeling human red blood cells for flow cytometric detection of hemoglobin genotype.

Jensen, Ronald H. (Livermore, CA); Vanderlaan, Martin (San Ramon, CA); Bigbee, William L. (Livermore, CA); Stanker, Larry H. (Livermore, CA); Branscomb, Elbert W. (Walnut Creek, CA); Grabske, Robert J. (Berkeley, CA)

1988-01-01

384

Monoclonal antibodies to human hemoglobin S and cell lines for the production thereof  

DOEpatents

The present invention provides monoclonal antibodies specific to and distinguishing between hemoglobin S and hemoglobin A and methods for their production and use. These antibodies are capable of distinguishing between two hemoglobin types which differ from each other by only a single amino acid residue. The antibodies produced according to the present method are useful as immunofluorescent markers to enumerate circulating red blood cells which have the property of altered expression of the hemoglobin gene due to somatic mutation in stem cells. Such a measurement is contemplated as an assay for in vivo cellular somatic mutations in humans. Since the monoclonal antibodies produced in accordance with the instant invention exhibit a high degree of specificity to and greater affinity for hemoglobin S, they are suitable for labeling human red blood cells for flow cytometric detection of hemoglobin genotype. 4 figs.

Jensen, R.H.; Vanderlaan, M.; Bigbee, W.L.; Stanker, L.H.; Branscomb, E.W.; Grabske, R.J.

1984-11-29

385

Enhancing effects of retinoic acid on monoclonal antibody production of human-human hybridomas.  

PubMed

The effects of retinoids on the production of monoclonal antibody of human-human hybridomas were examined. IgG antibody secretion of a hybridoma CLNH11 was enhanced up to about two- to fourfold by retinoic acid (RA) at concentrations ranging from 10(-9) to 10(-5) M, where RA had little effect on the growth rate and saturation density of the cell. Among other retinoids, retinol magnified the antibody production as well as RA. Retinal and retinyl acetate had weak effects. Retinyl palmitate showed no effect. RA also enhanced the production of monoclonal antibodies from other human-human hybridomas: SLNF10, IgG-producing; CoLNE10, IgA-producing; TOS/H8, IgM-producing. RA and human hybridomas provide a defined system to study the effects of retinoids on immune responses at a molecular level. PMID:2015633

Aotsuka, Y; Naito, M

1991-04-01

386

Mechanisms of G-CSF- or GM-CSF-stimulated tumor cell killing by Fc receptor-directed bispecific antibodies  

Microsoft Academic Search

Studies with gene-modified mice have recently reinforced the importance of Fc receptor-mediated effector mechanisms for the therapeutic efficacy of rituxan and herceptin — two clinically approved antibodies for the treatment of tumor patients. We investigated Fc receptor-dependent tumor cell killing by mononuclear and granulocytic effector cells — comparing human IgG1 antibodies against CD20 or HER-2\\/neu with their respective Fc?RI (CD64)-,

Bernhard Stockmeyer; David Elsässer; Michael Dechant; Roland Repp; Martin Gramatzki; Martin J Glennie; Jan G. J van de Winkel; Thomas Valerius

2001-01-01

387

Labeling and use of monoclonal antibodies in immunofluorescence: protocols for cytoskeletal and nuclear antigens.  

PubMed

Antibodies are widely used to target and label specifically extra- or intracellular antigens within cells and tissues. Most protocols follow an indirect approach implying the successive incubation with primary and secondary antibodies. In these protocols the primary antibodies are specifically targeted against the antigen in question and are normally not labeled. The secondary antibodies come from a different species and are in contrast fluorescently labeled. The idea is that the primary antibodies specifically bind to their targets but cannot be visualized directly. Only binding of the secondary (fluorescent) antibodies to the constant region of the primary antibodies allows consecutively the visualization in a fluorescent microscope.Primary antibodies can be either of monoclonal (normally produced in mouse) or of polyclonal origin (normally produced in rabbit, goat, sheep, or donkey). Using (primary) monoclonal antibodies has the clear advantage that all antibodies used are identical in origin and behavior and should thus give a more clear-cut labeling result. On the other hand the demands towards labeling protocols might be concomitantly higher: Binding of primary antibodies will only occur if fixation and labeling protocols preserve the antigen sufficiently to keep its specific and unique target structure available. One could imagine that for polyclonal antibodies this demand is slightly lower as there is a pool of antibodies with varying specificities against multiple parts of their target antigens. Certain fractions of this pool might thus tolerate a larger variety of conditions, and consequently a larger variety of protocols might still result in successful labeling.Each step in a labeling protocol can be decisive for the outcome of an experiment especially if monoclonal antibodies are used. Especially critical are choice of buffer and fixation and permeabilization parameters of the protocol.In this chapter we discuss and detail proven protocols using monoclonal antibodies for two key targets within a cell, namely, (a) the cytoskeleton with emphasis on microtubules (Cramer and Mitchison, J Cell Biol 31:179-189, 1995; Traub et al., Biol Cell 90:319-337, 1998; Desai et al., Methods Cell Biol 61:385-412, 1999) and (b) the nuclear pore complex (Davis and Blobel, Cell 45:699-709, 1986; Kimura et al., Mol Cell Biol 23:1304-1315, 2003). The two protocols which differ substantially in using either a chemical fixation/permeabilization approach versus a one-step coagulation protocol are chosen to offer the reader tools to design their own procedures and adapt them accordingly to fit their individual experimental setup and the biological question asked. PMID:24515489

Bauer, Christoph R

2014-01-01

388

Inhibition of kinesin-driven microtubule motility by monoclonal antibodies to kinesin heavy chains  

PubMed Central

We have prepared and characterized seven mouse monoclonal antibodies (SUK 1-7) to the 130-kD heavy chain of sea urchin egg kinesin. On immunoblots, SUK 3 and SUK 4 cross-reacted with Drosophila embryo 116- kD heavy chains, and SUK 4, SUK 5, SUK 6, and SUK 7 bound to the 120-kD heavy chains of bovine brain kinesin. Three out of seven monoclonal antikinesins (SUK 4, SUK 6, and SUK 7) caused a dose-dependent inhibition of sea urchin egg kinesin-induced microtubule translocation, whereas the other four monoclonal antibodies had no detectable effect on this motility. The inhibitory monoclonal antibodies (SUK 4, SUK 6, and SUK 7) appear to bind to spatially related sites on an ATP- sensitive microtubule binding 45-kD chymotryptic fragment of the 130-kD heavy chain, whereas SUK 2 binds to a spatially distinct site. None of the monoclonal antikinesins inhibited the microtubule activated MgATPase activity of kinesin, suggesting that SUK 4, SUK 6, and SUK 7 uncouple this MgATPase activity from motility. PMID:2974459

1988-01-01

389

Monoclonal antibodies to human beta-interferon produced by adoptive transfer in irradiated mice  

SciTech Connect

Three murine anti-human beta-interferon (IFN-beta) monoclonal antibodies have been isolated following adoptive transfer of immune spleen cells. Adoptive transfer was used to increase the specific efficiency of the fusion. These antibodies have been used to define two epitopes on IFN-beta; the antiviral, antiproliferative, and immunomodulatory effects of IFN-beta are associated with one of these epitopes.

Thompson, K.J.; Dawson, K.M.; Davies, J.A.; Powell-Jones, C.H.; Buckham, S.; O'Neill, G.J.; McCullagh, K.G.

1986-02-01

390

The analysis with monoclonal antibodies of the heterogeneity of Ia glycoproteins on chronic lymphocytic leukemia cells  

SciTech Connect

The accessible Ia molecules on the surface of chronic lymphocytic leukemia (CLL) cells were quantitated in the cellular radioimmunoassay with saturating concentrations of monoclonal antibodies. Monoclonal antibody 21w4, like DA/2 antibody, recognizes monomorphic determinants of human Ia antigens.The amount of 21w4 or DA/2 bound to CLL cells derived from eight patients (varying from 2.6 to 13.9 x 10/sup 5/ molecules/cell) appears to be the maximum observed with the antibodies studied. Two other antibodies, 18d5 and 21r5, although also directed at nonpolymorphic Ia determinants, bind differentially to CLL cells, with the ratios of 21r5/21w4 and 18d5/21w4 varying from 0.08 to 0.90. Sequential immunoprecipitation studies have established that the four epitopes 18d5, 21r5, 21w4, and DA/2 were present on the same molecules. All Ia molecules express 21w4 and DA/2 epitopes, whereas only certain subsets of Ia molecules carry accessible 21r5 or 18d5 epitopes. Competitive binding studies showed that the epitopes recognized by the four monoclonal antibodies were different. Monoclonal antibodies 21r5 and 21w4 did not inhibit each other's binding. Furthermore, binding of 21w4 to CLL cells potentiated the binding of /sup 125/I-21r5 IgG to the same cells, suggesting that binding of 21w4 antibody induces a conformational change in the molecule that renders 21r5 epitopes more accessible.

Addis, J.B. (Hospital for Sick Children, Toronto, Canada); Tisch, R.; Falk, J.A.; Letarte, M.

1982-11-01

391

In vivo Therapy with Monoclonal Anti-I-A Antibody Suppresses Immune Responses to Acetylcholine Receptor  

NASA Astrophysics Data System (ADS)

A monoclonal antibody to I-A gene products of the immune response gene complex attenuates both humoral and cellular responses to acetylcholine receptor and appears to suppress clinical manifestations of experimental autoimmune myasthenia gravis. This demonstrates that use of antibodies against immune response gene products that are associated with susceptibility to disease may be feasible for therapy in autoimmune conditions such as myasthenia gravis.

Waldor, Matthew K.; Sriram, Subramaniam; McDevitt, Hugh O.; Steinman, Lawrence

1983-05-01

392

Monoclonal Antibody Defining a Stage-Specific Mouse Embryonic Antigen (SSEA-1)  

Microsoft Academic Search

A monoclonal antibody derived by fusion of mouse myeloma cells with spleen cells from a mouse immunized with F9 teratocarcinoma cells is described. This antibody, which reacts with embryonal carcinoma cells of mouse and human origin and with some preimplantation stage mouse embryos, defines an embryonic stage-specific antigen. This stage-specific antigen (SSEA-1) is first detected on blastomeres of 8-cell stage

Davor Solter; Barbara B. Knowles

1978-01-01

393

The production and characterization of anti-bovine CD14 monoclonal antibodies  

Microsoft Academic Search

To characterize further the chemical and biological properties of bovine soluble (bos) CD14, a panel of ten murine monoclonal antibodies (mAb) reactive with recombinant (r) bosCD14 were produced. A sandwich ELISA, using murine mAb and rabbit polyclonal antibodies reactive with rbosCD14 was developed. All the mAb were reactive by ELISA with baculovirus-derived rbosCD14 and they recognized rbosCD14 (40 kDa) by

Eun J. SOHNa; Max J. PAAPEb; Robert R. Petersa; Raymond H. Fetterer; Neil C. TALBOTd; Douglas D. BANNERMANb

2004-01-01

394

The role of monoclonal antibodies and the recombinant DNA technology in studying autoantibody production.  

PubMed

The hybridoma technology has made it possible to sample the B-cell repertoire and to generate monoclonal antibodies which can be analyzed for their specificity and idiotypy. Using the recombinant DNA technology, the structure of the genes which encode those antibodies can be analyzed. The knowledge gained from the application of these techniques has made it possible to pose specific questions about the origins of autoantibodies. PMID:3093101

Scharff, M D; DePinho, R A; Behar, S; Beychok, C; Shin, S U; French, D

1986-04-15

395

Monoclonal Antibody against Babesia equi: Characterization and Potential Application of Antigen for Serodiagnosis  

Microsoft Academic Search

Monoclonal antibody (MAb) BEG3 was produced against Babesia equi parasites to define a species-specific antigen for diagnostic use. The MAb reacted with single, paired, and Maltese cross forms of B. equi, and no reaction was observed with this MAb on acetone-fixed Babesia caballi, Babesia ovata ,o rBabesia microti parasites in the indirect immunofluorescent antibody test. Confocal laser and immunoelectron microscopic

ABGAANDORJIIN AVARZED; IKUO IGARASHI; DANIEL T. DE WAAL; SATORU KAWAI; YUKIO OOMORI; NOBORU INOUE; YOSHIYUKI MAKI; YOSHITAKA OMATA; ATSUSHI SAITO; HIDEYUKI NAGASAWA; YUTAKA TOYODA; NAOYOSHI SUZUKI

1998-01-01

396

Subunit-specific monoclonal antibodies to tarantula hemocyanin, and a common epitope shared with calliphorin  

Microsoft Academic Search

Three murine hybridoma cell lines secreting IgG1 antibodies to 4×6 tarantula (Eurypelma californicum) hemocyanin were isolated, and the monoclonal antibodies Ec-7, Ec-8 and Ec-24 characterized by immunoblotting, immunoelectrophoresis and ELISA. WholeEurypelma hemocyanin, and the isolated subunitsa tog served as probes. For the subunits a novel, quick purification scheme on FPLC combined with immuno-affinity chromatography was established.

Jfirgen Markl; Stefanie Winter

1989-01-01

397

Monoclonal antibody-based enzyme immunoassays for the sensitive detection of s-triazines in water  

Microsoft Academic Search

Immunoassays in pesticide residue analysis significantly profit from the monoclonal antibody (mAb) technology because a sufficient supply of standardized antibodies can be provided. For the production of atrazine-specific mAbs hybridoma cells were produced by fusion of mouse myeloma cells and spleen cells from mice which were immunized with 4-chloro-6-ethylamino- 1,3,5-triazine-2-(6-aminohexanoic acid) coupled to keyhole limped hemocyanin. After screening with a

Bertold Hock; Thomas Giersch; Karl Kramer

1993-01-01

398

Reversal of lethal digoxin toxicity in guinea pigs using monoclonal antibodies and Fab fragments.  

PubMed

To extend the potential application of digoxin-specific immunoglobulin (Ig) (Fab) fragments for the reversal of advanced digitalis intoxication, monoclonal digoxin-specific antibodies were obtained by fusion of myeloma cell lines with spleen cells from mice immunized with a digoxin-serum albumin conjugate. The monoclonal antibody from the cell line designated Dig 26-10 had high affinity (KA = 5 X 10(9) M-1) and specificity for digoxin and was tested for its efficacy in the reversal of advanced, otherwise lethal digoxin toxicity in guinea pigs given a loading dose of 500 micrograms of digoxin per kg b.wt. i.v. followed by continuous infusion of digoxin at 10 (IgG-treated group) or 50 (Fab-treated group) microgram/kg/min. Control animals given nonspecific rabbit or mouse Igs after the onset of digoxin-toxic ventricular arrhythmias all died. Administration of monoclonal digoxin-specific antibody as intact IgG in doses stoichiometrically equivalent to the digoxin dose fully reversed digoxin toxicity in six of eight animals and prolonged survival somewhat in the remaining two animals. Fab fragments from the Dig 26-10 monoclonal antibody were even more effective, with rapid reversal (mean time 7 min) of all arrhythmias and survival of all animals so treated. We conclude that murine monoclonal antibodies and their Fab fragments are capable of reversing advanced and otherwise lethal digoxin-induced arrhythmias in a guinea-pig experimental model and offer potential advantages over polyclonal antibodies in the management of this clinically important problem. PMID:6707937

Lechat, P; Mudgett-Hunter, M; Margolies, M N; Haber, E; Smith, T W

1984-04-01

399

Bispecific minibodies targeting HER2/neu and CD16 exhibit improved tumor lysis when placed in a divalent tumor antigen binding format.  

PubMed

Unconjugated monoclonal antibodies have emerged as important therapeutic agents for selected malignancies. One mechanism by which antibodies can exert cytotoxic effects is antibody-dependent cellular cytotoxicity (ADCC). In an effort to increase the efficiency of ADCC at tumor sites, we have focused on the construction of bispecific antibodies specific for the tumor antigen HER2/neu and the Fc gamma RIII-activating receptor (CD16) found on NK cells, mononuclear phagocytes, and neutrophils. Here, we describe the production of bispecific minibodies in two distinct binding formats. The parent minibody was constructed such that the IgG1 C(H)3 constant domain serves as the oligomerization domain and is attached to an anti-CD16 and an anti-HER2/ neu single-chain Fv via 19- and 29-amino acid linkers, respectively. This molecule can be expressed in mammalian cells from a dicistronic vector and has been purified using sequential affinity purification techniques. Analysis by surface plasmon resonance shows that the bispecific minibody can bind to HER2/neu and CD16, both individually and simultaneously. Furthermore, cytotoxicity studies show that the minibody can induce significant tumor cell lysis at a concentration as low as 20 nm. A trimeric, bispecific minibody (TriBi) that binds dimerically to HER2/neu and monomerically to CD16 induces equivalent cytotoxicity at lower antibody concentrations than either the parent minibody or the corresponding single-chain dimer. Both minibody constructs are stable in mouse and human serum for up to 72 h at 37 degrees C. These minibodies have the potential to target solid tumors and promote tumor lysis by natural killer cells and mononuclear phagocytes. PMID:15471859

Shahied, Lillian S; Tang, Yong; Alpaugh, R Katherine; Somer, Robert; Greenspon, Dana; Weiner, Louis M

2004-12-24

400

Preparation of monoclonal antibody of anti-feline calicivirus and establishment of double-antibody sandwich enzyme-linked immunosorbent assay detecting method.  

PubMed

This study aimed to prepare monoclonal antibody of feline calicivirus (FCV) and identify its basic biological characteristics. Saturated ammonium sulfate precipitation, combined differential centrifugation, and cesium chloride density gradient centrifugation were used for the purification of FCV. The purified FCV was used as antigen to immunize BALB/c mice. The hybridoma lines of anti-FCV monoclonal antibodies were established using cell fusion and hybridoma screening techniques. The subtypes of the monoclonal antibody were identified. The results showed that 3 strains of hybridoma cell lines stably secreted anti-FCV monoclonal antibody; they were named as D8, E5, and H4. The D8 and E5 were IgM subtype antibodies, and H4 was IgG2b subtype antibody. The monoclonal antibody obtained shared no cross-reactivity with feline parvovirus, canine parvovirus, and canine distemper virus. According to the different recognition sites of 2 monoclonal antibodies E5 and H4 to the FCV, they were used to coat microtiter plates and prepare 2 enzyme-labeled secondary antibodies to establish double-antibody sandwich enzyme-linked immunosorbent assay detecting method. PMID:25222237

Yuan, B; Ai, C-X; Yuan, L; Gao, W; Hu, J-P; Chen, J; Ren, W-Z

2014-01-01

401

Isolation and characterization of monoclonal antibodies directed against plant plasma membrane and cell wall epitopes: identification of a monoclonal antibody that recognizes extensin and analysis of the process of epitope biosynthesis in plant tissues and cell cultures  

Microsoft Academic Search

Membranes from tobacco cell suspension cultures were used as antigens for the preparation of monoclonal antibodies. Use of solid phase and indirect immunofluorescence assays led to the identification of hybridomas producing antibodies directed against cell surface epitopes. One of these monoclonal antibodies (ll.D2) was found to recognize a molecular species which on two-dimensional analysis (using nonequilib- rium pH-gradient electrophoresis and

David J. Meyer; Claudio L. Afonso; David W. Galbraith

1988-01-01

402

The clinical application of monoclonal antibodies in chronic lymphocytic leukemia  

PubMed Central

Chronic lymphocytic leukemia (CLL) represents the most prevalent adult leukemia. Treatment with chemotherapy over the past 3 decades has been palliative. The introduction of therapeutic antibodies has increased the number of treatment options for this disease. Despite this increase, our true understanding of the mechanism of action of antibody therapy in CLL remains limited. Rituximab, a CD20 antibody, is currently widely used in combination-based strategies for both previously untreated symptomatic CLL and as salvage therapy. Recent data suggest that the addition of rituximab to fludarabine with or without cyclophosphamide prolongs survival in younger patients with CLL. Other improved CD20 antibodies with promising clinical activity, including ofatumumab and GA-101, are coming forward. Alemtuzumab, a CD52 antibody, likewise has demonstrated benefit in both symptomatic, previously untreated CLL and in patients with relapsed disease but has less selectivity. Development of other therapeutic antibodies targeting alternative B-cell–specific antigens in CLL has been less successful, although many promising candidate antibodies and/or small modular immune pharmaceuticals (SMIPs) are coming forward. In addition, recent efforts to combine currently applied therapeutic antibodies with other biologic and targeted therapies with efficacy in CLL offers the potential to move toward alternative non–chemotherapy-based treatment approaches. PMID:20610811

Jaglowski, Samantha M.; Alinari, Lapo; Lapalombella, Rosa; Muthusamy, Natarajan

2010-01-01

403

B cell responses to HIV and the development of human monoclonal antibodies.  

PubMed Central

In this review B cell responses in HIV-infected individuals are summarized together with the techniques used to date to produce human monoclonals to HIV and the properties of these antibodies. Profound disturbances in B cell responses are apparent both in vivo and in vitro. While there is evidence in vivo of marked polyclonal B cell activation, primary and secondary antibody responses are impaired. Similarly these cells exhibit spontaneous immunoglobulin secretion upon in vitro culture but do not readily respond to B cell mitogens and recall antigens including HIV. Furthermore, certain of these defects can be reproduced in normal B cells in vitro by incubation with HIV or HIV coded peptides. Individuals infected with HIV develop antibodies to HIV structural proteins (e.g. p17, p24, gp41 and gp120) and regulatory proteins (e.g. vif, nef, RT). Autoantibodies against a number of immunologically important molecules are also frequently observed. The anti-HIV antibodies are predominantly of the IgG1 isotype and exhibit a variety of effects on the virus in vitro. To date, using conventional immortalization strategies, an appreciable number of human monoclonals to HIV have been developed. These have been specific for gp41, gp120 and gag with antibodies of the former specificity predominating. The majority of these antibodies have been of the IgG1 isotype. Only a small number of the antibodies neutralize virus in vitro and most of these react with gp120. The neutralizing antibodies recognize conformational and carbohydrate epitopes or epitopes in amino acid positions 306-322. The predominant epitopes recognized by the anti-gp41 antibodies were in amino acid positions 579-620 and 644-662. A high percentage (congruent to 25%) of these antibodies enhance viral growth in vitro. The problems relating to the production of human monoclonals to HIV are discussed together with strategies that could be used in the future. PMID:1572084

Boyd, J E; James, K

1992-01-01

404

Prevalence, specificity and functionality of anti-ganglioside antibodies in neuropathy associated with IgM monoclonal gammopathy.  

PubMed

IgM antibodies against gangliosides and their complexes were studied in sera from 54 patients with polyneuropathy and IgM monoclonal gammopathy (IgM-PNP) without anti-MAG antibodies. Anti-ganglioside antibodies were found in 19 (35%) patients. Five (9%) patients had antibodies against ganglioside complexes. IgM antibodies against gangliosides activated complement in vitro. Light chain usage was restricted to kappa or lambda in most, but not all patients. In conclusion, anti-ganglioside antibodies in IgM-PNP are common, display pathogenic properties and do not always arise from a monoclonal B cell proliferation. PMID:24529728

Stork, Abraham C J; Jacobs, Bart C; Tio-Gillen, Anne P; Eurelings, Marijke; Jansen, Marc D; van den Berg, Leonard H; Notermans, Nicolette C; van der Pol, W-Ludo

2014-03-15

405

Development and Characterization of Monoclonal Antibodies to Detect Klotho  

PubMed Central

Although antibodies are commercially available to allow investigation into the biology of the age-regulating protein Klotho, problems with antibody specificity and application functionality are significant barriers to progress. Chief among these limitations is the inability of current tools to allow in vivo validation of binding partners originally identified through transfection of tagged proteins. To overcome this barrier, we generated a series of hybridoma cell lines by immunizing rats with a GST-KL1 fusion protein. Purified antibodies generated from these cell lines differentially detect human or mouse Klotho protein via Western blot, immunocyto/histochemistry, and immunoprecipitation. Specificity of antibody binding to Klotho was confirmed by mass spectrometry following immunoprecipitation. With this confidence in antibody specificity, co-immunoprecipitation was utilized to validate the interaction of Klotho/FGFR and Klotho/wnt7a in mouse kidney lysates. PMID:25513981

Maltare, Astha; Nietz, Angela K.; Laszczyk, Ann M.; Dunn, Taylor S.; Ballestas, Mary E.; Accavitti-Loper, Mary Ann

2014-01-01

406

Monoclonal Antibody Identification of Subpopulations of Cerebral Cortical Neurons Affected in Alzheimer disease  

Microsoft Academic Search

Neuronal degeneration is one of the hallmarks of Alzheimer disease (AD). Given the paucity of molecular markers available for the identification of neuronal subtypes, the specificity of neuronal loss within the cerebral cortex has been difficult to evaluate. With a panel of four monoclonal antibodies (mAbs) applied to central nervous system tissues from AD patients, we have immunocytochemically identified a

Carol A. Miller; Maria Rudnicka; David R. Hinton; Janet C. Blanks; Mark Kozlowski

1987-01-01

407

DEVELOPMENT OF A MONOCLONAL ANTIBODY BASED ELISA FOR THE BETA ADRENERGIC AGONIST ZILPATEROL.  

Technology Transfer Automated Retrieval System (TEKTRAN)

South Africa and Mexico, but neither the EU, USA, nor any Asian country have approved zilpaterol for use as a growth promoter in cattle, making development of a screening assay desirable. Monoclonal antibodies were generated in mice by immunization with zilpaterol-butyrate-KLH, and the same hapten ...

408

Parainfluenza Virus Type 2 Haemagglutinin-Neuraminidase Glycoprotein Characterized with Monoclonal Antibodies  

Microsoft Academic Search

SUMMARY Thirteen monoclonal antibodies (MAbs) were prepared against human parain- fluenza virus type 2 (PIV2). These MAbs reacted with the haemagglutinin- neuraminidase glycoprotein with an Mr of 84K. The MAbs defined one antigenic site which could be divided.into five epitopes. A correlation between haemagglutination inhibition (HI) and neutralization activity could be seen although one MAb, which recognized a distinct epitope,

ROBERT RYDBECK; A. Love; C. Orvell

1988-01-01

409

A new xenogenic monoclonal antibody detecting class I H-2 molecules  

Microsoft Academic Search

In an attempt to produce monoclonal antibodies which recognize surface antigens of splenic lymphocytes of NZB mice, female Lewis rats were injected intraperitoneally with 10 v NZB spleen cells three times at weekly intervals and boosted 1-2 months later with the same dose. Three days after the last injection, the recipient spleen cells were hybridized in the presence of polyethylene

Vijaya Manohar; Elinor M. Brown; William M. Leiserson; Thomas M. Chused

1982-01-01

410

Synthesis and self-assembly of a functional monoclonal antibody in transgenic Nicotiana tabacum  

Microsoft Academic Search

Immunoglobulin light and heavy chains are synthesized in mammalian cells as precursors containing a signal peptide. Processing and assembling result in formation of active antibodies. Chimeric genes have been made containing the coding sequence of the barley a-amylase signal peptide which has been fused to cDNAs coding for either the mature light or the mature heavy chain of a monoclonal

Sigrun Hippe; Fritz Kreuzaler; Jeff Schell

1990-01-01

411

Detection of Helicobacter pylori in Gastric Aspirates Using a Monoclonal Antibody-Based Test  

PubMed Central

Background/Aims The objective of this study was to evaluate a monoclonal antibody-based test to detect Helicobacter pylori-specific antigen in gastric aspirates from humans. Methods Sixty-one volunteers were enrolled in the study. All of the subjects underwent a 13C-urea breath test (UBT) before esophagogastroduodenoscopy. Gastric aspirates were analyzed for pH and ammonia and used for polymerase chain reaction (PCR), culture, and monoclonal antibody-based detection of H. pylori. Multiple biopsies of the gastric antrum and body were obtained for a rapid urease test (RUT) and histological evaluation. Results Thirty-six subjects were H. pylori-positive and 25 were H. pylori-negative according to the UBT results. Compared with the H. pylori-negative subjects, H. pylori-positive subjects had a higher pH (4.77±1.77 vs 3.49±1.30, p<0.05) and ammonia level (1,130.9±767.4 vs 184.2±126.3, p<0.0001). The sensitivities and specificities of the PCR test, RUT, culture test, and monoclonal antibody-based test were 100% and 72%, 89% and 100%, 47% and 100%, and 78% and 100%, respectively. Conclusions The monoclonal antibody-based test for diagnosing H. pylori infection in gastric aspirates has increased sensitivity compared with the culture test and specificity as high as that of the RUT. The test may be useful as an additive test for examining gastric aspirates. PMID:23423538

Kim, Ho Dong; Kim, Do Hyun; Park, Hyeuk; Kim, Woo Jong; Ahn, Yong Soo; Lee, Young Jik; Park, Sun Mi; Seo, Eun Seon; Park, Chul; Kim, Yang Ho; Kim, Hyung Rag; Joo, Young Eun

2013-01-01

412

Advances in the use of monoclonal antibodies in the therapy of chronic lymphocytic leukemia  

Microsoft Academic Search

Monoclonal antibody (moAb)-based therapies are evolving as an integrated component in the treatment of chronic lymphocytic leukemia (CLL). Advantages such as different mechanisms of action (compared with those of chemotherapy), no or minimal stem cell toxicity, as well as the absence of hair loss and delayed nausea may result in a rapidly increasing usage of these agents in different phases

Jeanette Lundin; Anders Österborg

2004-01-01

413

Monoclonal antibodies to synthetic pyrethroids and method for detecting the same  

DOEpatents

Methods are described for making specific monoclonal antibodies which may be used in a sensitive immunoassay for detection of synthetic pyrethroids in foods and environmental samples. Appropriate sample preparation and enzyme amplification of the immunoassay for this widely-used class of pesticides permits detection at low levels in laboratory and field tested samples.

Stanker, Larry H. (Livermore, CA); Vanderlaan, Martin (Danville, CA); Watkins, Bruce E. (Livermore, CA); Van Emon, Jeanette M. (Henderson, NV); Bigbee, Carolyn L. (Livermore, CA)

1992-01-01

414

Preclinical safety testing of monoclonal antibodies: the significance of species relevance  

Microsoft Academic Search

Selecting a pharmacologically relevant animal species for testing the safety and toxicity of novel monoclonal antibody (mAb) therapies to support clinical testing can be challenging. Frequently, the species of choice is the primate. With the increased number of mAbs in the pharmaceutical pipeline, this has significant implications for primate use, and so raises several important scientific, ethical and economic issues.

Nick Pullen; Mark Graham; Ian Ragan; Kathryn Chapman

2007-01-01

415

Production and characterization of monoclonal antibodies against different epitopes of Ebola virus antigens  

Microsoft Academic Search

Ebola virus (EBOV) causes hemorrhagic fever in humans and nonhuman primates with up to 90% mortality rate. In this study, Ebola virus like particles (EVLPs) and the aglycosyl subfragment of glycoprotein (GP(1) subfragment D) were used to generate monoclonal antibodies (MAbs) against different epitopes of the viral antigens. Such MAbs could be useful in diagnostics and potential therapeutics of viral

Soraya Shahhosseini; Dipankar Das; Xiangguo Qiu; Heinz Feldmann; Steven M. Jones; Mavanur R Suresh

2007-01-01

416

An analysis of the properties of monoclonal antibodies directed to epitopes on influenza virus hemagglutinin  

Microsoft Academic Search

Summary Monoclonal antibodies (MAbs) specific for the hemagglutinin (HA) of the H3 subtype of influenza A virus were grouped according to their inability to bind to particular MAb-selected neutralization escape mutants of the virus having an amino acid substitution in one of the five postulated antigenic sites on the molecule. Additional residues critical to the binding of the MAbs were

L. E. Brown; J. M. Murray; D. O. White; D. C. Jackson

1990-01-01

417

Monoclonal antibody specific for human colon fibroblast-derived T-PA  

SciTech Connect

This patent describes a murine-derived hybridoma cell line capable of producing monoclonal antibody against human colon fibroblast-derived tissue plasminogen activator and the cell line selected from the group consisting of cell lines 63-4 (ATCC HB 9155), 54-2 (ATCC HB 9157) or 79-7 (ATCC HB 9156).

Schaumann, J.P.; Olander, J.V.; Harakas, N.K.; Feder, J

1989-05-23

418

Phytochrome in photosynthetically competent plants characterization by monoclonal antibodies: Progress report  

SciTech Connect

Monoclonal antibodies to oat and to pea phytochrome have been characterized and immunopurified. In addition a fully automated, microcomputer-based, dual wavelength spectrophotometer was designed and constructed. An ELISA that is sensitive to subfemtomol levels of phytochrome has been developed. 1 fig.

Pratt, L.H.

1983-08-01

419

INITIAL CHARACTERIZATION OF MONOCLONAL ANTIBODIES AGAINST THE FUNGAL HEMOLYSIN STACHYLYSIN FROM STACHYBOTRYS CHARTARUM  

EPA Science Inventory

Stachybotrys chartarum is known to produce the hemolysin stachylysin and its detection in human serum has been proposed as a biomarker for exposure to the fungus. In this study we report the initial characterization of monoclonal antibodies (mAbs) against stachylysin and the dev...

420

ANTIGEN DETECTION WITH MONOCLONAL ANTIBODIES FOR THE DIAGNOSIS OF ADENOVIRUS GASTROENTERITIS  

EPA Science Inventory

The authors have developed a monoclonal antibody-based enzyme immunoassay (EIA) for direct detection of enteric adenoviruses in stool specimens from patients with gastroenteritis. Tests specific for each of the enteric adenoviruses, adenovirus type 40 (Ad40) and type 41 (Ad41) we...

421

Production, characterization and application of monoclonal antibody to spherulocytes: A subpopulation of coelomocytes of Apostichopus japonicus  

Technology Transfer Automated Retrieval System (TEKTRAN)

One monoclonal antibody (mAb 3F6) against coelomocytes of sea cucumber Apostichopus japonicus was developed by immunization of Balb/C mice. Analyzed by indirect immunofluorescence assay test (IIFAT), immunocytochemical assay (ICA),Western blotting and fluorescence-activated cell sorter (FACS), mAb 3...

422

Ebola GP-Specific Monoclonal Antibodies Protect Mice and Guinea Pigs from Lethal Ebola Virus Infection  

Microsoft Academic Search

Ebola virus (EBOV) causes acute hemorrhagic fever in humans and non-human primates with mortality rates up to 90%. So far there are no effective treatments available. This study evaluates the protective efficacy of 8 monoclonal antibodies (MAbs) against Ebola glycoprotein in mice and guinea pigs. Immunocompetent mice or guinea pigs were given MAbs i.p. in various doses individually or as

Xiangguo Qiu; Lisa Fernando; P. Leno Melito; Jonathan Audet; Heinz Feldmann; Gary Kobinger; Judie B. Alimonti; Steven M. Jones

2012-01-01

423

Protective Efficacy of Neutralizing Monoclonal Antibodies in a Nonhuman Primate Model of Ebola Hemorrhagic Fever  

Microsoft Academic Search

Ebola virus (EBOV) is the causative agent of severe hemorrhagic fever in primates, with human case fatality rates up to 90%. Today, there is neither a licensed vaccine nor a treatment available for Ebola hemorrhagic fever (EHF). Single monoclonal antibodies (MAbs) specific for Zaire ebolavirus (ZEBOV) have been successfully used in passive immunization experiments in rodent models, but have failed

Andrea Marzi; Reiko Yoshida; Hiroko Miyamoto; Mari Ishijima; Yasuhiko Suzuki; Megumi Higuchi; Yukie Matsuyama; Manabu Igarashi; Eri Nakayama; Makoto Kuroda; Masayuki Saijo; Friederike Feldmann; Douglas Brining; Heinz Feldmann; Ayato Takada

2012-01-01

424

Drug Insight: antiangiogenic therapies for gastrointestinal cancers—focus on monoclonal antibodies  

Microsoft Academic Search

Tumor angiogenesis is strongly induced by vascular endothelial growth factor (VEGF), which is overexpressed in most human gastrointestinal cancers. VEGF overexpression is known to be associated with poor prognosis and survival in patients with various solid tumors. The humanized monoclonal anti-VEGF antibody bevacizumab (Avastin®, Genentech Inc., South San Francisco, CA) is a prototypic antiangiogenic compound, and has proven therapeutic benefit

Anke Reinacher-Schick; Michael Pohl; Wolff Schmiegel

2008-01-01

425

Influence of cocktails of labeled monoclonal antibodies on the localization of antibodies in human tumor xenografts.  

PubMed

In order to evaluate the usefulness of cocktails of labeled monoclonal antibodies (MoAbs) recognizing different antigen molecules to localize human cancer xenografts, we have compared the potential of three MoAbs recognizing representative cancer-associated CA 19-9, 17-1A and CEA antigens when administered alone or in combination. Specific binding of radioiodinated F(ab')2 fragments of these three MoAbs was observed to human colorectal cancer cell lines SW1116, LS180 and Co-3. The percentage of in vitro cell binding of a cocktail of any two MoAbs to cancer cells was equal to the average of those obtained with the two MoAbs alone. The three MoAbs were preferentially localized in tumor tissues xenografted in nude mice. When cocktails of any two MoAbs were used, the obtained tumor-to-normal tissue ratios and percent of injected dose per gram of tumor were between the levels obtained for each MoAb when administered alone, in all three tumors transplanted in nude mice. These data suggest that, although cocktails of labeled MoAbs recognizing different antigens may extend the spectrum of tumor specificities, their use does not improve the tumor localization ability of MoAb-conjugates. PMID:2112530

Watanabe, Y; Endo, K; Saga, T; Koizumi, M; Sakahara, H; Nakai, T; Hosono, M; Yao, Z S; Kuroki, M; Matsuoka, Y

1990-03-01

426

Mice are actively immunized after passive monoclonal antibody prophylaxis and ricin toxin challenge. (Reannouncement with new availability information)  

SciTech Connect

Mice passively immunized by a protective, anti-ricin A-chain monoclonal antibody, then challenged intravenously with ricin, were protected from a subsequent ricin challenge, and were actively immunized. Two significant advantages accrued from this experiment: the monoclonal antibody neutralized the toxicity of the ricin immunogen, and active immunization was achieved with very low antigen load (approx. 0.5 micrograms/mouse). We ruled out the possibility that residual monoclonal antibody provided the protection by using three independent criteria. There was significant (four orders of magnitude) enhancement of the immune response in the presence of the monoclonal antibody; control immunizations of mice with ricin A-chain, ricin B-chain or either chain with the monoclonal antibody did not induce active immunity; and the active immunization could not be replicated when protective goat polyclonal antibody was substituted for the monoclonal antibody. Because high titers were achieved rapidly without any adjuvant, we are currently investigating haptenized ricin to determine if anti-hapten monoclonal antibodies can be produced by this refined procedure.

Lemley, P.V.; Wright, D.C.

1992-12-31