ABSTRACT
Overexpression of epidermal growth factor receptor (EGFr) has been demonstrated on many human tumors, and the increase in receptor expression levels has been linked with a poor clinical prognosis. Blocking the interaction of EGFr and the growth factors could lead to the arrest of tumor growth and possibly result in tumor cell death. To this end, using XenoMouse technology, ABX-EGF, a human IgG2 monoclonal antibody (mAb) specific to human EGFr, has been generated. ABX-EGF binds EGFr with high affinity (5x10(-11) M), blocks the binding of both EGF and transforming growth factor-alpha (TGF-alpha) to various EGFr-expressing human carcinoma cell lines, and inhibits EGF-dependent tumor cell activation, including EGFr tyrosine phosphorylation, increased extracellular acidification rate, and cell proliferation. In vivo ABX-EGF prevents completely the formation of human epidermoid carcinoma A431 xenografts in athymic mice. More importantly, administration of ABX-EGF without concomitant chemotherapy results in complete eradication of established tumors. No tumor recurrence was observed for more than 8 months following the last antibody injection, further indicating complete tumor cell elimination by the antibody. Inhibition of human pancreatic, renal, breast and prostate tumor xenografts which express different levels of EGFr by ABX-EGF was also achieved. Tumor expressing more than 17000 EGFr molecules per cell showed significant growth inhibition when treated with ABX-EGF. ABX-EGF had no effect on EGFr-negative tumors. The potency of ABX-EGF in eradicating well-established tumors without concomitant chemotherapy indicates its potential as a monotherapeutic agent for treatment of multiple EGFr-expressing human solid tumors, including those where no effective chemotherapy is available. Utilization of mAbs directed to growth factor receptors as cancer therapeutics has been validated recently by the tumor responses obtained from clinical trials with Herceptin, the humanized anti-HER2 antibody, in patients with HER2 overexpressing metastatic breast cancer. Being a fully human antibody, ABX-EGF is anticipated to exhibit a long serum half-life and minimal immunogenicity with repeated administration, even in immunocompetent patients. These results demonstrate the potent anti-tumor activity of ABX-EGF and its therapeutic potential for the treatment of multiple human solid tumors that overexpress EGFr.
Subject(s)
Antibodies, Monoclonal/therapeutic use , Epidermal Growth Factor/immunology , Neoplasms/drug therapy , Animals , Antibodies, Heterophile/biosynthesis , Antibodies, Heterophile/genetics , Antibodies, Monoclonal/biosynthesis , Antibodies, Monoclonal/genetics , Antineoplastic Agents/therapeutic use , Humans , Mice , PanitumumabABSTRACT
Infections with Streptococcus pneumoniae remain a significant cause of morbidity and mortality. To gain insight into structure-function relationships for human antibodies to pneumococcal capsular polysaccharide (PPS), we studied the response of transgenic mice reconstituted with human immunoglobulin loci, XenoMouse, to PPS antigens in a pneumococcal vaccine. Enzyme-linked immunosorbent assays of sera from mice vaccinated with a 23-valent pneumococcal vaccine revealed that they produced serotype-specific human antibodies, with the greatest response being to the PPS of serotype 3 (PPS 3). Molecular sequence analysis of three monoclonal antibodies (MAbs) to PPS 3 generated from lymphoid cells from mice vaccinated with a 23-valent pneumococcal vaccine or a PPS 3-bovine serum albumin conjugate revealed that they all used heavy-chain immunoglobulin genes from the V(H)3 family, two expressed light chain genes from the human Vkappa1 family, and one expressed a mouse lambda light chain. The protective efficacy of the two MAbs was examined in mice. A 10-microgram dose of both, and a 1-microgram dose of one, significantly prolonged survival from a lethal serotype 3 infection in CBA/N mice. Our data show that XenoMouse mice produced protective, serotype-specific human antibodies to PPS 3, and they lend support to the proposal that these animals represent a useful model to study the human antibody response to PPS antigens.
Subject(s)
Antibodies, Bacterial/biosynthesis , Complementarity Determining Regions , Genes, Immunoglobulin , Polysaccharides, Bacterial/immunology , Streptococcus pneumoniae/immunology , Animals , Antibodies, Monoclonal/immunology , Antibody Specificity , Complement Activation , Female , Humans , Hybridomas/immunology , Immunoglobulin Variable Region/genetics , Immunoglobulin Variable Region/immunology , Male , Mice , Mice, Inbred CBA , Mice, Transgenic , Structure-Activity Relationship , Time Factors , VaccinationABSTRACT
Interleukin-8 (IL-8) is a potent chemotactic cytokine implicated in the pathogenesis of a number of inflammatory disease states. Agents that block the binding of IL-8 to its receptor have been shown to block inflammation in animal models of disease. This suggests that drugs specifically targeting IL-8 may prove efficacious in treating multiple human diseases. To this end, we developed a panel of fully human anti-IL-8 monoclonal antibodies (mAbs). These human antibodies were generated from XenoMouse strains, mice created by introducing megabase-size unrearranged human immunoglobulin heavy and kappa light chain loci into a mouse genome in which the corresponding endogenous loci have been inactivated. From the panel of more than 50 mAbs, two antibodies, K4.3 and K2.2, were further characterized and evaluated for their specificity, productivity, affinity, and biological activity. Both K4.3 and K2.2 bind human IL-8 with high affinity (Kd of K4.3 = 2.1x10(10) M; Kd of K2.2 = 2.5x10(-10) M). In vitro, in addition to blocking IL-8 binding to human neutrophils, K4.3 and K2.2 blocked a number of IL-8-dependent cellular functions including neutrophil activation, up-regulation of the cell adhesion receptor CD11b/CD18, and neutrophil chemotaxis, suggesting that the fully human anti-IL-8 mAbs derived from XenoMouse strains are potent anti-inflammatory agents. This was further supported by in vivo studies in which K4.3 and K2.2 significantly inhibited IL-8-induced skin inflammation in rabbits. A pharmacokinetic study in Cynomolgus monkeys demonstrated that the alpha phase half-life is 9.4 h and the beta phase 10.9 days, typical of human mAbs in monkeys. These data support advancing a fully human anti-IL-8 mAb into clinical trials to treat inflammatory diseases.
Subject(s)
Anti-Inflammatory Agents, Non-Steroidal/immunology , Antibodies, Monoclonal/immunology , Immunization, Passive , Inflammation/therapy , Interleukin-8/immunology , Neutrophils/immunology , Animals , Anti-Inflammatory Agents, Non-Steroidal/therapeutic use , Antibodies, Monoclonal/pharmacokinetics , Antibodies, Monoclonal/therapeutic use , Antibody Specificity , Drug Eruptions/etiology , Drug Eruptions/immunology , Drug Eruptions/prevention & control , Genes, Immunoglobulin , Humans , Immunoglobulin Heavy Chains/genetics , Immunoglobulin kappa-Chains/genetics , Interleukin-8/toxicity , Macaca fascicularis , Mice , Mice, Knockout , Mice, Transgenic , Rabbits , Recombinant Proteins/immunologyABSTRACT
A fully human IgG2kappa monoclonal antibody (MAb), E7.6.3, specific to the human epidermal growth factor (EGF) receptor (EGFr) was generated from human antibody-producing XenoMouse strains engineered to be deficient in mouse antibody production and to contain the majority of the human antibody gene repertoire on megabase-sized fragments from the human heavy and kappa light chain loci. The E7.6.3 MAb exhibits high affinity (KD = 5 x 10(-11) M) to the receptor, blocks completely the binding of both EGF and transforming growth factor alpha (TGF-a) to various EGFr-expressing human carcinoma cell lines, and abolishes EGF-dependent cell activation, including EGFr tyrosine phosphorylation, increased extracellular acidification rate, and cell proliferation. The antibody (0.2 mg i.p. twice a week for 3 weeks) prevents completely the formation of human epidermoid carcinoma A431 xenografts in athymic mice. More importantly, the administration of E7.6.3 without concomitant chemotherapy results in complete eradication of established tumors as large as 1.2 cm3. Tumor eradication of A431 xenografts was achieved in nearly all of the mice treated with total E7.6.3 doses as low as 3 mg, administered over the course of 3 weeks, and a total dose of 0.6 mg led to tumor elimination in 65% of the mice. No tumor recurrence was observed for more than 8 months after the last antibody injection, which further indicated complete tumor cell elimination by the antibody. The potency of E7.6.3 in eradicating well-established tumors without concomitant chemotherapy indicates its potential as a monotherapeutic agent for the treatment of multiple EGFr-expressing human solid tumors, including those for which no effective chemotherapy is available. Being a fully human antibody, E7.6.3 is expected to exhibit minimal immunogenicity and a longer half-life as compared with mouse or mouse-derivatized MAbs, thus allowing repeated antibody administration, including in immunocompetent patients. These results suggest E7.6.3 as a good candidate for assessing the full therapeutic potential of anti-EGFr antibody in the therapy of multiple patient populations with EGFr-expressing solid tumors.
Subject(s)
Antibodies, Monoclonal/therapeutic use , ErbB Receptors/immunology , Immunoglobulin G/therapeutic use , Neoplasms, Experimental/therapy , Animals , Antibody Affinity , Humans , Immunotherapy , Male , Mice , Mice, Inbred BALB C , Mice, Nude , Neoplasm Transplantation , Neoplasms, Experimental/pathology , Neoplasms, Experimental/prevention & control , Transplantation, HeterologousABSTRACT
The last two years have seen a renaissance of monoclonal antibodies for the treatment of disease. Of the eight antibodies currently approved for human therapy, two are for the treatment of cancer. In large part, the revival of antibodies has been driven by technology developments geared toward making antibodies less likely to elicit an anti-antibody response in humans. The development of transgenic mice, XenoMouse animals, capable of making fully human antibodies offers new opportunities for generating antibodies of therapeutic quality. Recently, this technology has been applied to the generation of a fully human antibody to the epidermal growth factor receptor. A description of the development of this antibody serves to illustrate the power and ease of use of XenoMouse technology.
Subject(s)
Antibodies, Monoclonal/genetics , Neoplasms/therapy , Amino Acid Sequence , Animals , Antibodies, Monoclonal/chemistry , Antibodies, Monoclonal/therapeutic use , ErbB Receptors/immunology , Humans , Immunoglobulin Heavy Chains/chemistry , Immunoglobulin Heavy Chains/genetics , Immunoglobulin Variable Region/chemistry , Immunoglobulin Variable Region/genetics , Mice , Mice, Transgenic , Molecular Sequence Data , Recombinant Fusion Proteins/biosynthesis , Recombinant Fusion Proteins/therapeutic useABSTRACT
We constructed two megabase-sized YACs containing large contiguous fragments of the human heavy and kappa (kappa) light chain immunoglobulin (Ig) loci in nearly germline configuration, including approximately 66 VH and 32 V kappa genes. We introduced these YACs into Ig-inactivated mice and observed human antibody production which closely resembled that seen in humans in all respects, including gene rearrangement, assembly, and repertoire. Diverse Ig gene usage together with somatic hypermutation enables the mice to generate high affinity fully human antibodies to multiple antigens, including human proteins. Our results underscore the importance of the large Ig fragments with multiple V genes for restoration of a normal humoral immune response. These mice are likely to be a valuable tool for the generation of therapeutic antibodies.
Subject(s)
Antibody Formation , Genes, Immunoglobulin , Transgenes , Amino Acid Sequence , Animals , Antibodies, Monoclonal/biosynthesis , Antibodies, Monoclonal/genetics , Antibodies, Monoclonal/immunology , Antibody Affinity , Antibody Diversity , B-Lymphocytes/cytology , B-Lymphocytes/immunology , Chromosomes, Artificial, Yeast/genetics , ErbB Receptors/immunology , Gene Rearrangement, B-Lymphocyte , Humans , Hybridomas/immunology , Immunoglobulin Heavy Chains/biosynthesis , Immunoglobulin Heavy Chains/genetics , Immunoglobulin kappa-Chains/biosynthesis , Immunoglobulin kappa-Chains/genetics , Interleukin-8/immunology , Mice , Mice, Inbred C57BL , Mice, Knockout , Mice, Transgenic , Molecular Sequence Data , Species Specificity , Tumor Necrosis Factor-alpha/immunologyABSTRACT
IVA039.1 is a bifunctional antibody with specificity for the murine IL-2 receptor and vinca alkaloids. Biodistribution studies show that IVA039.1 can target and deliver vinca alkaloids to tissues that contain IL-2 receptor positive cells. Vinca alkaloids are lymphocytotoxic. Therapy of diabetic mice with IVA039.1 plus vincristine results in a significant decrease in the glucose levels of diabetic compared to untreated mice. The therapeutic effect of IVA039.1 plus vincristine therapy was additive but surprisingly not synergistic. The binding of IVA039.1 to vincristine has moderate affinity with a slow off rate. In vitro studies suggest that, when bound to IVA039.1, the vincristine is inactivated. We attribute the lack of an enhanced therapeutic response to bifunctional antibody therapy using IVA039.1 plus vincristine to the inaccessibility of the drug to the target cells.
Subject(s)
Antibodies, Bispecific/immunology , Antibodies, Bispecific/therapeutic use , Diabetes Mellitus, Experimental/chemically induced , Diabetes Mellitus, Experimental/therapy , Receptors, Interleukin-2/immunology , Streptozocin/toxicity , Vinca Alkaloids/immunology , Vinca Alkaloids/therapeutic use , Animals , Cell Line , Male , Mice , Mice, Inbred Strains , Vincristine/antagonists & inhibitors , Vincristine/immunology , Vincristine/therapeutic useABSTRACT
Inadvertent oncolytic overdoses occur rarely, but can have serious consequences. We have investigated the possibility of using an antibody, 27.8.1A, reactive with vinca alkaloids, as a means of reducing the toxicity associated with overdose situations. In vitro cytotoxicity of a vinca derivative, 4-desacetyl- vinblastine-3-carbox-hydrazide (DAVLBHYD), with and without the addition of 27.8.1A, was determined. Using CCRF-CEM, a human acute lymphoblastic leukemia cell line, as a target in this assay, we observed a greater than 90% increase in cell viability using 100 micrograms/ml 27.8.1A with a 0.1 microgram/ml concentration of DAVLBHYD. 27.8.1A had no effect on cell viability when doxorubicin was used as a control drug in this assay. Similarly, the addition of an irrelevant antibody. EGFrL11, had no effect on the toxicity of DAVLBHYD. In an in vivo survival experiment, nude mice were injected with a toxic dose of DAVLBHYD and subsequently given four doses of 27.8.1A. All anti vinca antibody-treated mice survived, in contrast to the untreated group or irrelevant antibody-treated group in which only 25% and 10% of the mice survived, respectively.
Subject(s)
Antibodies, Monoclonal/pharmacology , Antineoplastic Agents/toxicity , Precursor Cell Lymphoblastic Leukemia-Lymphoma/drug therapy , Vinblastine/analogs & derivatives , Animals , Antineoplastic Agents/immunology , Antineoplastic Agents/pharmacokinetics , Body Weight/drug effects , Cell Survival/drug effects , Drug Interactions , Female , Humans , Inactivation, Metabolic , Kinetics , Mice , Mice, Inbred Strains , Tumor Cells, Cultured , Vinblastine/immunology , Vinblastine/pharmacokinetics , Vinblastine/toxicityABSTRACT
The autoreactive T cell plays a pivotal role in the pathogenesis of type I diabetes in humans and in rodent animal models. Elimination or attenuation of these cells may provide a means to treat the disease. The use of antibodies directed to T cells has shown varying degrees of effectiveness in the treatment of autoimmune disease. The use of a bifunctional antibody directed to T cells with a cytolytic agent may provide an additional level of therapeutic efficacy compared to anti-T-cell antibodies alone. To test this hypothesis, we prepared a bifunctional antibody (IVA039.1) with specificity for the mouse interleukin-2 (IL-2) receptor and vinca alkaloids. The antibody was derived from the fusion of vinca immune spleen cells with PC61 5.3, a hybridoma that produces rat anti-mouse IL-2 receptor antibody. IVA039.1 was purified by affinity chromatography through Protein A and anti-vinca affinity columns followed by TSK-DEAE high-pressure liquid chromatography (HPLC). Bifunctionality of the antibody was confirmed by fluorescence-activated cell sorting (FACS) analysis, enzyme-linked immunoadsorbent assay (ELISA) and a cell assay designed to measure simultaneously both IL-2 receptor and vinca reactivities. The biodistribution of IVA039.1 was determined in normal and streptozotocin-complete Freund's adjuvant (CFA) induced diabetic mice. Enhanced uptake of IVA039.1 was observed in the pancreata, spleens, and lymph nodes of diabetic compared to normal mice. These data suggest that bifunctional antibodies that can deliver cytolytic agents to T cells may be appropriate candidates for the treatment of diabetes and other autoimmune diseases.
Subject(s)
Antibodies, Bispecific/immunology , Receptors, Interleukin-2/immunology , Vindesine/immunology , Animals , Antibodies, Bispecific/biosynthesis , Antibodies, Bispecific/metabolism , Antibodies, Monoclonal/immunology , Antibody Specificity , Diabetes Mellitus, Experimental/immunology , Enzyme-Linked Immunosorbent Assay , Female , Flow Cytometry , Hybridomas/immunology , Immunoenzyme Techniques , Male , Mice , Mice, Inbred A , Mice, Inbred BALB C , Mice, Inbred Strains , Mice, Nude , Tissue DistributionABSTRACT
A panel of four murine monoclonal antibodies apparently directed against three distinct epitopes of carcinoembryonic antigen (CEA) was conjugated via oxidized carbohydrate groups to 4-desacetylvinblastine-3-carboxyhydrazide. The resulting antibody-vinca conjugates were evaluated for antitumor activity against 2-9-day-established LS174T human colorectal carcinoma xenografts. The antibodies (immunoglobulin G, IgG) employed in this study were 11.285.14 (IgG1), 14.95.55 (IgG2a), CEM231 (IgG1), ZCE025 (IgG1). Additive immunofluorescence studies indicated that CEM231 and ZCE025 recognized the same or a closely related epitope(s) on CEA which was distinct from the two epitopes bound by 11.285.14 and 14.95.55. The in vivo antitumor efficacy studies demonstrated that chemoimmunoconjugates prepared from 14.95.55 and ZCE025 were more active than the conjugates constructed from the 11.285.14 and CEM231 antibodies. The 14.95.55 and ZCE025 immunoconjugates were also more efficacious than free drug or drug conjugated to irrelevant murine IgG. The presence of increased carbohydrate content on the light chain of ZCE025 may have been responsible for the ability to construct ZCE025-vinca conjugates with about twice the drug content (approximately 10 mol of vinca/mol of IgG) than was achieved with the other antibodies. The highly conjugated form of ZCE025 demonstrated similar efficacy but was much less toxic than a ZCE025 conjugate containing 5 mol of vinca/mol of IgG. These data indicated that significant differences existed in the ability of monoclonal antibodies to target a cytotoxic agent for effective antitumor activity even when the immunoconjugates recognized the same antigen or even the same or closely related antigen epitope(s). Furthermore, these differences could not have been identified without extensive in vivo evaluation for antitumor efficacy.
Subject(s)
Antibodies, Monoclonal/immunology , Antineoplastic Agents/pharmacology , Carcinoembryonic Antigen/immunology , Immunotoxins/toxicity , Vinblastine/analogs & derivatives , Animals , Epitopes , Fluorescence , Fluorescent Antibody Technique , Humans , Mice , Mice, Nude , Neoplasm Transplantation , Transplantation, Heterologous , Tumor Cells, Cultured , Vinblastine/pharmacologyABSTRACT
In addition to the bispecific antibody, the hybrid-hybridoma 28.19.8 secretes antibodies monospecific for carcinoembryonic antigen (CEA) and antibodies monospecific for vinca alkaloids. By exhaustive affinity purification, the immunoglobulins isolated by Protein A chromatography from ascitic fluid have been fractionated into four populations, three of which are immunoreactive. Studies on these fractions by FPLC, SDS-PAGE, and a variety of ELISAs have shown that seven of the ten possible combinations of heavy and light chains exist as immunoreactive forms. The results suggest that the other three inactive combinations are also secreted by the hybrid-hybridoma.
Subject(s)
Antibodies/metabolism , Hybridomas/metabolism , Immunoglobulin Heavy Chains/immunology , Immunoglobulin Light Chains/immunology , Animals , Antibodies/isolation & purification , Antibody Specificity , Carcinoembryonic Antigen/immunology , Chromatography, Affinity , Chromatography, Liquid , Electrophoresis, Polyacrylamide Gel , Enzyme-Linked Immunosorbent Assay , Haptens , Immunoglobulin Heavy Chains/isolation & purification , Immunoglobulin Isotypes/immunology , Immunoglobulin Light Chains/isolation & purification , Mice , Mice, Inbred BALB CABSTRACT
The hybrid-hybrid monoclonal antibody 28-19-8 has specificity for the tumour-associated antigen carcinoembryonic antigen and the vinca alkaloids. This bifunctional antibody has been used to target unmodified vinblastine sulphate to well-established MAWI human tumour xenografts implanted in nude mice. The highly significant suppression of tumour growth achieved throughout treatment has also been sustained for over 2 months after the withdrawal of treatment. Histological examination of excised tumours from treated animals has shown profound changes in their morphology when compared with tumours from control animals. Cells in tumours that had started to grow again after withdrawal of therapy were shown still to express carcinoembryonic antigen, the target antigen recognised by the bispecific antibody.
Subject(s)
Adenocarcinoma/therapy , Antibodies, Monoclonal/therapeutic use , Immunotoxins/administration & dosage , Vinblastine/administration & dosage , Adenocarcinoma/immunology , Adenocarcinoma/pathology , Animals , Antibodies, Monoclonal/isolation & purification , Carcinoembryonic Antigen/immunology , Humans , Immunization Schedule , Immunoenzyme Techniques , Mice , Mice, Nude , Neoplasm TransplantationABSTRACT
A method has been developed to allow the direct coupling of the cytotoxic vinca alkaloid 4-desacetylvinblastine-3-carbohydrazide (DAVLB hydrazide) to a variety of murine monoclonal antibodies directed against human solid tumors. Periodate oxidation of carbohydrate residues on the antibodies, followed by reaction with DAVLB hydrazide in aqueous acid affords, in most cases, conjugates with conjugation ratios of 4-6 vincas per antibody in high yield without significantly impairing antigen binding or solubility. The outcome of the conjugation reaction is highly dependent on the concentration of, and time of exposure of the protein to, the oxidant. These conjugates exhibit potent antitumor activity in vivo against a number of human solid tumor-nude mouse xenografts, with efficacy and safety increased over unconjugated DAVLB hydrazide. This antitumor activity is also superior to that of similarly prepared but nontarget tumor binding antibody-DAVLB hydrazide conjugates. MoAb-DAVLB hydrazide conjugates release DAVLB hydrazide in solution in a temperature- and pH-dependent manner. Hydrolytic release of unmodified DAVLB hydrazide from tumor-localized MoAb-DAVLB hydrazide conjugates in vivo may be an important factor in their antitumor activity.
Subject(s)
Antibodies, Monoclonal/chemical synthesis , Immunotoxins/chemical synthesis , Vinblastine/analogs & derivatives , Animals , Antibodies, Monoclonal/therapeutic use , Drug Design , Humans , Immunotoxins/therapeutic use , Mice , Mice, Nude , Neoplasms, Experimental/drug therapy , Structure-Activity Relationship , Vinblastine/chemical synthesis , Vinblastine/therapeutic useABSTRACT
A method is described that allows the simultaneous visualization and relative assessment of both the antibody and drug components of monoclonal antibody-drug conjugates at the target cell membrane. The antibody is detected by a fluorescein-conjugated anti-mouse immunoglobulin serum while the drug is visualized by rhodamine avidin or phycoerythrin-streptavidin binding to a biotinylated anti-Vinca alkaloid monoclonal antibody. This technique was effective in demonstrating the cell surface localization of a monoclonal antibody-Vinca alkaloid conjugate to human lung adenocarcinoma cells grown in vitro and was also used to demonstrate targeting of the conjugate in vivo to the membranes of these same tumor cells grown as a nude mouse xenograft. This method was also utilized to help elucidate the mechanism of action of monoclonal antibody-drug conjugates.
Subject(s)
Immunotoxins/metabolism , Antibodies, Monoclonal/metabolism , Cell Membrane/metabolism , Endocytosis , Flow Cytometry , Fluorescent Antibody Technique , Humans , Immunotoxins/pharmacology , Tumor Cells, Cultured/metabolismABSTRACT
The functional properties of a hybrid-hybrid monoclonal antibody (MAb) recognising both CEA and Vinca alkaloids have been explored in vivo in nude mice xenografted with MAWI, a human colorectal tumour. The hybrid-hybrid MAb localises specifically onto CEA-expressing tumour tissue and, furthermore, is able to target Vinca alkaloids to tumour. Under the influence of the hybrid-hybrid MAb a profound change in the bio-distribution patterns of the Vinca alkaloids is observed. Therapeutic data produced in this in vivo model indicates that treatment with Vinca alkaloids in conjunction with hybrid-hybrid MAb is significantly more effective in suppressing tumour growth of established tumour xenografts than the Vincas when given as free drug.
Subject(s)
Antibodies, Monoclonal/administration & dosage , Carcinoembryonic Antigen/immunology , Neoplasms, Experimental/drug therapy , Vinca Alkaloids/administration & dosage , Animals , Mice , Vinca Alkaloids/immunologyABSTRACT
Recent developments of hybridoma technology have allowed us to prepare a bispecific monoclonal antibody recognising both the tumour-associated antigen carcinoembryonic antigen (CEA) and the cytostatic vinca alkaloids. The yields of the hybrid-hybrid 28.19.8 monoclonal after affinity chromatography purification are close to 50% of the total Ig produced. The hybrid-hybrid has a molecular weight ca. 150,000 daltons. The heavy chains of the hybrid-hybrid are a gamma 1 heavy chain from the parental anti-CEA monoclonal and a gamma 2a heavy chain from the anti-vinca alkaloid donor lymphocytes.
Subject(s)
Antibodies, Monoclonal/immunology , Carcinoembryonic Antigen/immunology , Hybridomas , Vinca Alkaloids/immunology , Animals , Antibody Affinity , Antibody Specificity , Antibody-Dependent Cell Cytotoxicity , Mice , Molecular WeightABSTRACT
By using a bispecific monoclonal antibody recognising both carcinoembryonic antigen (CEA) and the cytostatic vinca alkaloid drugs we have been able to show specific tumour localisation of vinca alkaloids. In vitro studies with sections of human colorectal tumours have demonstrated that the hybrid-hybrid 28.19.8 monoclonal is able to specifically localise vindesine to cells expressing CEA. Furthermore, the hybrid-hybrid 28.19.8 localises in vivo preferentially to tumour tissues in nude mice bearing the MAWI human xenograft tumour. This tumour-bound hybrid-hybrid monoclonal antibody induces profound changes in the bio-distribution of vinca alkaloid drugs, targeting them specifically to tumour tissues.
Subject(s)
Antibodies, Monoclonal/administration & dosage , Carcinoembryonic Antigen/immunology , Colonic Neoplasms/drug therapy , Vinca Alkaloids/immunology , Vindesine/administration & dosage , Animals , Humans , Mice , Mice, Nude , Organ Culture TechniquesABSTRACT
Unmodified vinblastine (VLB) targeted through one of the antigen combining sites of the hybrid-hybrid 28.19.8 monoclonal is potentially more effective in suppressing the growth of established MAWI tumour xenografts implanted on nude mice than free VLB in the absence of the targeting agent, presumably due to an increased local drug concentration. Our efficacy results in this study suggest that drug, specifically removed from the circulation by hybrid-hybrid antibody previously located to the tumour mass, can be made available in a pharmacologically active from. Histological analysis of the treated tumours revealed dramatic changes in the tumour organisation with only a few surviving tumour cells with altered morphology.
Subject(s)
Antibodies, Monoclonal/administration & dosage , Colonic Neoplasms/drug therapy , Vinblastine/administration & dosage , Animals , Carcinoembryonic Antigen/immunology , Evaluation Studies as Topic , Mice , Mice, Nude , Vinblastine/therapeutic use , Vinca Alkaloids/immunologyABSTRACT
The anticancer alkaloid vindesine (VDS) was conjugated to four mouse monoclonal antibodies recognizing human tumor-associated antigens. The antibodies were 96.5 (antimelanoma, IgG2a); 791T/36 (antiosteogenic sarcoma, IgG2b); 11.285.14, and 14.95.55 (anticarcinoembryonic antigen, IgG1 and IgG2a respectively). Conjugates VDS-96.5 and VDS-791T/36 were tested in vitro and shown to be specifically cytotoxic for target cells expressing the appropriate antigen. The in vivo effects of the antibodies and conjugates were tested against human tumor xenografts in athymic or immunodeprived mice using multiple treatments. Conjugate VDS-96.5 retarded the initial growth of a melanoma xenograft, whereas free antibody was without effect. Similarly, VDS-791T/36 but not free antibody retarded the growth of osteogenic sarcoma 791T. The most marked antitumor effects observed were those obtained with VDS conjugates of the anti-CEA antibodies against a colorectal tumor xenograft. Antibody 14.95.55 suppressed tumor growth both alone and as a VDS conjugate, whereas 11.285.14 produced only a slight effect alone but an almost complete and lasting suppression of tumor growth as a VDS conjugate. Free VDS had little effect at nontoxic levels. Acute studies showed that VDS-11.285.14 conjugate was considerably less toxic than free VDS in Balb/c mice.
Subject(s)
Antibodies, Monoclonal/therapeutic use , Antineoplastic Agents/administration & dosage , Neoplasms, Experimental/therapy , Vinblastine/analogs & derivatives , Animals , Antineoplastic Agents/toxicity , Melanoma/therapy , Mice , Mice, Nude , Vinblastine/administration & dosage , Vinblastine/toxicity , VindesineABSTRACT
Low serum sulphydryl (SH) levels are a feature of active rheumatoid arthritis (RA). We have investigated whether a similar blockade of membrane SH groups on mononuclear cells modifies the function of these cells in this disease. Using pokeweed mitogen stimulated IgG synthesis as the assay system, we have found that the accessory cell function of peripheral blood monocytes is totally dependent on free SH groups on the cell surface. Monocytes from patients with active RA display poor accessory cell function when compared with healthy monocytes or with cells from patients treated with D-penicillamine. The poor function of the rheumatoid accessory cells may be corrected in vitro by 2-mercaptoethanol (2-ME). Addition of 2-ME to the culture system also enhances IgG synthesis by rheumatoid mononuclear cells to normal levels. We suggest that surface SH-dependent mechanisms are responsible, at least in part, for the depressed mononuclear cell functions of rheumatoid cells in vitro and may explain some effects of D-penicillamine therapy in rheumatoid patients.