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Hybridoma Technology (hybridoma + technology)
Selected AbstractsLBY135, a novel anti-DR5 agonistic antibody induces tumor cell,specific cytotoxic activity in human colon tumor cell lines and xenografts,DRUG DEVELOPMENT RESEARCH, Issue 2 2008Jing Li Abstract TRAIL (TNF-related apoptosis-inducing ligand) induces apoptosis on binding to DR4 and DR5 receptors on the surface of tumor cells. These receptors are of particular interest in the development of cancer therapeutics as they preferentially mediate tumor cell apoptosis. We have generated a chimeric anti-DR5 agonistic antibody, LBY135, from its murine parental antibody, LCR211, identified using hybridoma technology. Both LCR211 and LBY135 specifically bind to DR5 with nanomolar affinity, mimic TRAIL to induce cell death in tumor cells, and have little effect on non-transformed cells in vitro. The anti-DR5 antibody reduced viability in 45% of a panel of 40 human colon cancer cell lines with IC50 values of 20,nM or less. In vivo, using human colorectal tumor xenograft mouse models, LCR211 induced tumor regression and showed enhanced efficacy when combined with 5-FU. Both in vitro evaluation of ADCC (antibody-dependent cell-mediated cytotoxicity) and CDC (complement-dependent cytotoxicity), and in vivo studies using a non-functional DR5 specific antibody or SCID-Beige mice, suggested ADCC and CDC are unlikely to be the mechanism to ablate tumors in vivo. LBY135 and LCR211 appear to mediate cell death and tumor regression mainly through apoptosis, as demonstrated by the activation of caspase 3, caspase 8, M30, and TUNEL assay. In addition, the discovery of synergy between cross-linked LBY135 and TRAIL not only revealed the unique epitope of LBY135, but also demonstrated an additional mechanism of action for LBY135 in vivo. LBY135 demonstrates promise as a novel therapeutic for cancer treatment and is currently in Phase I clinical trials. Drug Dev Res 69: 69,82, 2008. © 2008 Wiley-Liss, Inc. [source] Monoclonal antibodies: a morphing landscape for therapeuticsDRUG DEVELOPMENT RESEARCH, Issue 10 2006Nicholas C. Nicolaides Abstract The concept of using antibodies as therapeutics to cure human diseases was postulated nearly 100 years ago by Paul Ehrlich and subsequently enabled by the discovery of hybridoma technology by Kohler and Milstein in 1975. While the use of monoclonal antibodies (mAbs) as drugs that can specifically target a disease-associated antigen is compelling, it has taken a quarter century for these molecules to be adopted as bona fide therapeutic agents. Despite their slow pursuit in drug development during the pioneering years, it is now estimated that there are nearly 500 mAb-based therapies in development. Major factors that have influenced the acceptance of monoclonal antibodies as therapeutics include their drug safety profiles, technological advancements for facilitating mAb discovery and development, and market success. Early on, it was demonstrated that antibodies could elicit clinical benefit by antagonizing a specific antigen without the common side effects that are prevalent with small chemical entities due to their nonspecific effects on homeostatic biochemical pathways. In addition, the significant technological advances that the biotechnology industry has established for developing and producing monoclonal antibodies at commercial scale in a more efficient and cost-effective manner has broadly enabled their use as therapeutics. However, despite the beneficial pharmacologic advantages and technological advances, it has been the sheer market success that monoclonal antibody products have achieved over the past few years that has propelled their vast pursuit by the biopharmaceutical industry in light of their value-creating potential. Here we provide an overview of the monoclonal antibody industry and discuss evolving technologies and strategies that are being pursued to overcome challenges in the changing marketplace. Drug Dev. Res. 67:781,789, 2006. © 2007 Wiley-Liss, Inc. [source] Design and engineering human forms of monoclonal antibodiesDRUG DEVELOPMENT RESEARCH, Issue 3 2004Manuel L. Penichet Abstract The antibody molecule has multiple properties that make it a key component of the immune response. These include its ability to recognize a vast array of different foreign substrates and to interact with and activate the host effector systems. Antibodies with defined specificities may serve as "magic bullets" for the diagnosis and therapy of multiple diseases. With the development of the hybridoma technology, it was possible to produce rodent (mouse or rat) monoclonal antibodies that are the product of a single clone of antibody producing cells and have only one antigen binding specificity. However, the therapeutic use of rodent monoclonals antibodies in humans is limited by their immunogenicity, short circulating half-life, and inability to efficiently trigger human effector mechanisms. However, it proved difficult to produce human monoclonal antibodies using the same methods. To address these problems genetic engineering and expression systems have instead been used to produce chimeric, humanized, and totally human antibodies as well as antibodies with novel structures and functional properties. In addition, the use of yeast and human artificial chromosome vectors for animal transgenesis has allowed the development of animal models that produce antigen specific antibodies that are totally human. As a consequence, recombinant antibody-based therapies are now used to treat a variety of clinical conditions including infectious diseases, inflammatory disorders, and cancer. This article summarizes and compares different strategies for designing and engineering human antibodies and their derivatives. Drug Dev. Res. 61:121,136, 2004. © 2004 Wiley-Liss, Inc. [source] Two novel monoclonal antibodies to VWFA3 inhibit VWF-collagen and VWF-platelet interactionsJOURNAL OF THROMBOSIS AND HAEMOSTASIS, Issue 9 2007Y. ZHAO Summary.,Background:,The interaction of collagen-von Willebrand factor (VWF)-GPIb is essential for platelet adhesion, especially under high shear conditions. VWF, which acts as a bridge between platelets and exposed subendothelium, interacts with collagen through its A3 domain, which is a new target for the antithrombotic agent. Objective:,To develop functional blockers that specifically inhibit VWF-dependent adhesion of platelets to collagen under high shear stress. Methods:,To develop murine antihuman VWF A3 monoclonal antibodies (mAbs) by standard hybridoma technology, and characterize their abilities to block interactions between VWF A3 and collagen as well as platelet function. Results:,Thirty anti-VWF-A3 mAbs were obtained. Among them, two mAbs, designated as SZ-123 and SZ-125, were found to inhibit VWF-collagen type III interaction. SZ-123 and SZ-125 inhibited the binding of purified human VWF (1.5 or 3 ,g mL,1) to human placenta collagen type III (IC50 = 0.07 ± 0.02 and 0.15 ± 0.03 ,g mL,1, respectively) or to calf skin collagen type III (IC50 = 0.48 ± 0.06 and 0.51 ± 0.07 ,g mL,1, respectively) coated on plates. Under flow shear condition (1000 s,1), SZ-123 and SZ-125 inhibited platelet adhesion on human placenta collagen- or calf skin collagen-coated surfaces. Both mAbs also inhibited platelet aggregation induced by ristocetin, botrocetin or bovine plasma. Conclusions:,SZ-123 and SZ-125 inhibited VWF-collagen and VWF-platelet interactions. [source] Detection and characterization of gamete-specific molecules in Mytilus edulis using selective antibody productionMOLECULAR REPRODUCTION & DEVELOPMENT, Issue 1 2009Heiko Stuckas Abstract The mussel Mytilus edulis can be used as model to study the molecular basis of reproductive isolation because this species maintains its species integrity, despite of hybridizing in zones of contact with the closely related species M. trossulus or M. galloprovincialis. This study uses selective antibody production by means of hybridoma technology to identify molecules which are involved in sperm function of M. edulis. Fragmented sperm were injected into mice and 25 hybridoma cell clones were established to obtain monoclonal antibodies (mAb). Five clones were identified producing mAb targeting molecules putatively involved in sperm function based on enzyme immunoassays, dot and Western blotting as well as immunostaining of tissue sections. Specific localization of these mAb targets on sperm and partly also in somatic tissue suggests that all five antibodies bind to different molecules. The targets of the mAb obtained from clone G26-AG8 were identified using mass spectrometry (nano-LC-ESI-MS/MS) as M6 and M7 lysin. These acrosomal proteins have egg vitelline lyses function and are highly similar (76%) which explains the cross reactivity of mAb G26-AG8. Furthermore, M7 lysin was recently shown to be under strong positive selection suggesting a role in interspecific reproductive isolation. This study shows that M6 and M7 lysin are not only found in the sperm acrosome but also in male somatic tissue of the mantle and the posterior adductor muscle, while being completely absent in females. The monoclonal antibody G26-AG8 described here will allow elucidating M7/M6 lysin function in somatic and gonad tissue of adult and developing animals. Mol. Reprod. Dev. 76: 4,10, 2009. © 2008 Wiley-Liss, Inc. [source] Production and partial characterization of mouse monoclonal antibodies recognizing common cytokine receptor gamma chain (,c) of human, mouse and primate origin,APMIS, Issue 10 2001KAROLINA LUNDIN Monoclonal antibodies specific for the common cytokine receptor gamma chain, ,c, were produced using traditional hybridoma technology. Fusion of P3X63-Ag8.653 myeloma cells with splenocytes from Balb/c mice immunized with Spodoptera frugiperda insect cells infected with the recombinant baculovirus VL1392-hIL-2R, resulted in several hybridoma cell clones producing monoclonal ,c -specific antibodies. Four of these antibody-producing clones, IIIC3, IIIE8, IG3 and IF10C5, were further characterized by immunoblotting, flow cytometry and ELISA. Data are presented demonstrating that the generated monoclonal antibodies can identify the extracellular domain of the common cytokine receptor , chain of human and mouse origin, and two of the antibodies recognize ,c of primate origin as well. [source] Method for generation of human hyperdiversified antibody fragment libraryBIOTECHNOLOGY JOURNAL, Issue 1 2007Philippe Mondon Abstract The selection of antibody fragments from libraries using in vitro screening technologies has proven to be a very good alternative to the classical hybridoma technology, and has overcome the laborious process of antibody humanization. However, the complexity of the library is critical in the probability of being able to directly isolate a high affinity antibody specific to a target. We report a method to make hyperdiversified antibody fragment libraries, based on human immunoglobulin variable genes mimicking the somatic hypermutation process. This mutagenesis technology, MutaGenÔ, was used for the first time on the entire variable domain (frameworks and CDRs) of large repertoires of human variable antibody domains. Our MutaGenÔ process uses low-fidelity human polymerases, known as mutases, suggested to be involved in the somatic hypermutation process of immunoglobulin genes. Depending on the mutases used, we generated complementary mutation patterns with randomly distributed mutations. The libraries were generated with an average of 1.8 mutations per 100 amino acids. The hyperdiversified antibody fragment libraries constructed with our process should enable the selection of antibody fragments specific to virtually any target. [source] | |||||||||||||