Characterization of the antibody response against NeuGcGM3 ganglioside elicited in non-small cell lung cancer patients immunized with an anti-idiotype antibody

1E10 mAb is an anti-Id murine mAb (Ab2 mAb) specific for an Ab1 mAb that reacts with NeuGc-containing gangliosides, sulfatides, and Ags expressed in some human tumors. In preclinical studies, this Ab2 Ab was able to mimic NeuGc-containing gangliosides only in animals lacking expression of these Ags in normal tissues. In this study, we report on the immune responses elicited in 20 non-small cell lung cancer patients treated with 1 mg of aluminum hydroxide-precipitated 1E10 mAb. In the hyperimmune sera from 16 of 20 patients, a strong specific Ab response of both IgM and IgG isotypes against NeuGcGM3 ganglioside was observed. Patient immune sera were able to induce complement-independent cell death of NeuGcGM3-expressing X63 murine myeloma target cells. Significant immunoreactivity to NeuGcGM3 was still detected after the complete abrogation of the reactivity against 1E10 mAb by the adsorption of patient sera with this Ab. We hypothesize that Id(-)Ag(+) Abs could reflect the activation of an autologous idiotypic cascade into the patients. Both Id(+)Ag(+) and Id(-)Ag(+) fractions were separated by affinity chromatography and characterized. Although IgG isotype Abs were found in both fractions, IgM isotype Abs were found only in the Id(-)Ag(+) fraction. Both Id(+)Ag(+) and Id(-)Ag(+) Abs were able to specifically recognize and induce cell death in NeuGcGM3-expressing X63 myeloma target cells. Patients that developed IgG and/or IgM Abs against NeuGcGM3 showed longer median survival times.     

An anti-idiotype vaccine elicits a specific response to N-glycolyl sialic acid residues of glycoconjugates in melanoma patients

We generated the 1E10 gamma-type anti-idiotype mAb (Ab2) specific to an Ab1 mAb able to react specifically with N-glycolyl-containing gangliosides and with Ags expressed on human melanoma and breast carcinoma cells. This Ab2 mAb induced an Ab response in animal models sharing immunochemically defined idiotopes with the Ab1. The treatment of tumor-bearing mice with 1E10 mAb induced a strong antitumor activity. A clinical trial was conducted in 20 patients with advanced malignant melanoma. Patients were treated with six intradermal injections of aluminum hydroxide-precipitated 1E10 anti-Id mAb given at 2-wk intervals. Sixteen of the 17 patients who received at least four doses of the anti-Id vaccine develop Ab3 Abs capable of inhibiting Ab2 binding to Ab1 (Ab3Id+). In contrast to the incapacity of 1E10 mAb to generate Ab3 Abs with the same antigenic specificity as the Ab1 mAb in mice, a very specific and strong Ab3 response against N-glycolyl-containing gangliosides was induced in 16 patients (Ab3Ag+). No evidence of serious or unexpected adverse effects has been observed in this clinical trial. 1E10 anti-Id vaccine was safe, well tolerated, and immunologically effective, with most patients being able to generate a specific immune response against 1E10 and Neu-glycolyl-GM(3) ganglioside.     

Induction of anti-idiotypic humoral and cellular immune responses by a murine monoclonal antibody recognizing the ovarian carcinoma antigen CA125 encapsulated in biodegradable microspheres

The use of biodegradable poly(dl-lactic-co-glycolic acid) microspheres as a cancer vaccine delivery system for induction of anti-idiotypic responses was investigated using a murine monoclonal antibody B43.13 that recognizes the human ovarian cancer antigen CA125. Immunization of mice with mAb B43.13 encapsulated in poly(dl-lactic-co-glycolic acid) microspheres resulted in enhanced humoral and cellular immune responses compared with mAb B43.13 alone or mAb B43.13 mixed with microspheres. The antibody responses could be further enhanced by the co-encapsulation of mAb B43.13 with monophosphoryl lipid A, a non-toxic adjuvant, in microspheres. Anti-idiotypic humoral responses were shown to result in Ab2 antibodies mimicking the nominal antigen CA125 and Ab3 antibodies recognizing CA125. Further, microsphere delivery of mAb B43.13 also resulted in induction of T cell responses involving T2 cells reactive with mAb B43.13 epitopes and T3 cells recognizing CA125. These results indicate that microsphere delivery of Ab1 can induce both humoral and cellular anti-idiotypic responses relevant to cancer antigens. This raises the possibility of the use of such formulations for anti-idiotypic induction immunotherapy for cancer. Received: 27 August 1997 / Accepted: 24 April 1998     

Molecular characterization of the anti-idiotypic immune response of a relapse-free neuroblastoma patient following antibody therapy: a possible vaccine against tumors of neuroectodermal origin?

Neuroblastoma treatment with chimeric antidisialoganglioside GD2 Ab ch14.18 showed objective antitumor responses. Production of anti-idiotypic Abs (Ab2) against ch14.18 (Ab1) in some cases was positively correlated with a more favorable prognosis. According to Jerne's network theory, a subset of anti-idiotypic Abs (Ab2beta) carries an "internal image" of the Ag and induces Abs (Ab3) against the original Ag. The molecular origin of an anti-idiotypic Ab response in tumor patients was not investigated previously. To clone anti-idiotypic Abs, B cells of a ch14.18-treated neuroblastoma patient with Ab2 serum reactivity were used to construct Ab phage display libraries. After repeated biopannings on ch14.18 and its murine relative, anti-GD2 mAb 14G2a, we selected 40 highly specific clones. Sequence analysis revealed at least 10 of 40 clones with different Ig genes. Identities to putative germline genes ranged between 94.90 and 100% for V(H) and between 93.90 and 99.60% for V(L). An overall high rate of replacement mutations suggested a strong Ag-driven maturation of the anti-idiotypic Abs. Two clones that were analyzed further, GK2 and GK8, inhibited binding of ch14.18 to GD2 just as the patient's serum did. GK8 alone inhibited >80% of the patient's anti-idiotypic serum Abs in binding to ch14.18. Rabbits vaccinated with GK8 or GK2 (weaker) produced Ab3 against the original target Ag GD2. GK8 may be useful as a tumor vaccine for GD2-positive [corrected] tumors.     

Induction of delayed-type hypersensitivity responses by monoclonal anti-idiotypic antibodies to tumor cells expressing carcinoembryonic antigen and tumor-associated glycoprotein-72

The use of anti-idiotypic antibodies as immunogens represents one potential approach to active specific immunotherapy of cancer. Two panels of syngeneic monoclonal anti-idiotypic antibodies were generated. One panel was directed against mAb CC49 and the other to mAb COL-1. mAb CC49 recognizes the pancarcinoma antigen (Ag), tumor-associated glycoprotein-72 (TAG-72), and mAb COL-1 recognizes carcinoembryonic antigen (CEA). Seven anti-idiotypic (AI) antibodies (Ab2) designated AI49-1–7 were generated that recognize the variable region of mAb CC49. These mAb were shown to inhibit the interaction of mAb CC49 (Ab1) with TAG-72 (Ag). Five anti-idiotypic antibodies designated CAI-1–5 were also generated to the anti-CEA mAb, COL-1 (Ab1). These Ab2 were shown to inhibit the interaction between COL-1 (Ab1) and CEA (Ag). Immunization of mice, rats, and rabbits with Ab2 directed against CC49 or COL-1 could not elicit specific Ab3 humoral immune responses, i.e., antibody selectively reactive with their respective target antigens. However, immunization of mice with the CC49 anti-idiotypic antibody (Ab2), designated AI49-3, could induce a delayed-type hypersensitivity response (DTH) specific for tumor cells that express TAG-72. Similarly, immunization of mice with an anti-idiotypic antibody directed against COL-1, designated CAI-1, could induce specific DTH cell-mediated immune responses to murine tumor cells that express human CEA on their surface. These results thus demonstrate that while some anti-idiotype mAb may not be potent immunogens in eliciting Ab3 humoral responses, they are capable of eliciting specific cellular immune responses against human carcinoma-associated antigens. This type of mAb may ultimately be useful in active immunotherapy protocols for human carcinoma.Some of the studies described in this paper were in partial fulfillment of requirements for the completion of Dr. Irvine's dissertation at the George Washington University     

Chimeric human/murine monoclonal IgM antibodies to HIV-1 Nef antigen expressed on chronically infected cells

Human IgM antibody (Ab) to gangliosides induced cytolysis of HIV-1-infected cells by homologous human complement. We expected that any human IgM Ab reactive with HIV-1 infected cells could cause complement-mediated cytolysis. The trans-chromosome mouse (TC mouse) contains human chromosomes harboring genes responsible for immunoglobulin production. Spleen cells from TC mice immunized with recombinant Nef were fused with mouse myeloma cells to generate hybridomas, and we selected those that produced human mu-chain-positive Abs reactive with Nef fixed on an ELISA plate. However, the L-chain of the monoclonal Abs (mAbs) were murine lambda in type and were chimeric, and we could not succeed in obtaining mAb with human mu- and human kappa-chains. The chimeric mAbs reacted with the HIV-1 infected cells as seen with flow cytometric analysis, and the surface expression of Nef was also detectable on chronically infected OM10.1 cells which had no detectable gp120. However, although the reaction of the chimeric IgM mAb with HIV-1-infected MOLT4 cells induced C3 deposition on cell surfaces on incubation with fresh human serum, the cells remained unlysed, as determined by 51Cr release assay. The amount of Nef antigen on the cells might not have been high enough to overcome the function of HRF20 (CD59) that restricts formation of membrane attack complexes of homologous complement. However, combination of anti-Nef IgM mAb with other IgM mAbs reactive with the surface of HIV-1-infected cells may induce a synergistic effect in complement mediated cytolysis.     

Reactivity and regulatory properties of human anti-idiotypic antibodies induced by T cell vaccination

Immunization with irradiated autoreactive T cells (T cell vaccination) induces anti-idiotypic T cell responses that preferentially recognize complementarity-determining region 3 sequences, contributing to clonal depletion of autoreactive T cells. However, it remains unknown whether T cell vaccination elicits anti-idiotypic humoral responses and whether the anti-idiotypic Abs play a similar role in the regulatory mechanism induced by T cell vaccination. In this study we examined the occurrence, the reactivity pattern, and the regulatory role of anti-idiotypic Abs elicited by T cell vaccination in patients with multiple sclerosis. We demonstrated for the first time that B cells producing anti-idiotypic Abs could be isolated from vaccinated patients. These EBV-transformed B cell lines were selected for specific reactivity to a 20-mer TCR peptide incorporating a common complementarity-determining region 3 sequence of the immunizing T cell clones. The resulting anti-idiotypic Abs were found to react with the original immunizing T cell clones and exhibit an inhibitory effect on their proliferation. The findings suggest that anti-idiotypic Ab responses can be induced by T cell vaccination in humans and that their regulatory properties are likely to contribute to the suppression of myelin basic protein-reactive T cells in vaccinated patients. The study has important implications in our understanding of the regulatory role of the anti-idiotypic humoral responses induced by T cell vaccination.     

Induction of an anti-human type II collagen response by monoclonal anti-idiotypic antibody

Several distinct epitopes on human type II collagen were defined by using mAb. The presence of species-specific and species-nonspecific (common) epitopes was thus clarified. Anti-idiotypic mAb (Ab2) was developed against one of the antibodies (Ab1) reactive with species-specific epitopes. Thus Ab2 was demonstrated to recognize an idiotope expressed on the Ag-binding site (paratope) of Ab1, since the binding of Ab1 to human type II collagen was blocked by Ab2, and the binding of Ab2 to Ab1 was inhibited by soluble human type II collagen, but not by murine and bovine type II collagens. DBA/1 mice immunized with Ab2 coupled to keyhole limpet hemocyanin produced an antibody (Ab3) specifically reactive with human type II collagen. It was also demonstrated that Ab3 expressed an idiotype similar to that of Ab1. These findings indicate that anti-idiotypic antibody directed against mAb to human type II collagen mimics a species-specific epitope on human type II collagen. The anti-idiotypic antibody bearing internal image of type II collagen will open the way to isolation of the arthritogenic epitope on type II collagen.     

Immunological properties of a single-chain fragment of the anti-idiotypic antibody ACA125

Vaccination with anti-idiotypic antibodies has been described as a promising concept for treatment of several malignant diseases. The murine monoclonal anti-idiotypic antibody ACA125 imitates a specific epitope of the tumor-associated antigen CA125 expressed by 80% of ovarian carcinomas. In the first clinical trial it could be shown that mAb ACA125 is able to elicit anti-anti-idiotypic antibodies (Ab3) with anti-CA125 specificity in patients with advanced ovarian cancer. In order to improve the capabilities of anti-idiotype vaccines we generated a genetically engineered single-chain fragment (scFv) ACA125 composed of heavy- and light-chain variable regions connected by a flexible linker. The antigenicity of scFv ACA125 was demonstrated by immunizing rats i.p. with scFv or complete mAb in complete/incomplete Freund's adjuvants (CFA/IFA) or precipitated by aluminium hydroxide. Negative control groups included applications of irrelevant mouse IgG or adjuvants alone. Anti-anti-idiotypic antibodies (Ab3) directed against the mAb ACA125 as well as specific anti-CA125 antibodies (Ab1′) could be detected in all animals treated with scFv in CFA/IFA. Nevertheless, antibody titers were lower than when the complete mAb ACA125 was used. Suprisingly, an increase of specificity could not be observed in scFv-immunized animals, which had been expected because of the lack of heavy- and light-chain constant regions that could raise rather unspecific anti-isotypic and anti-allotypic rat anti-(mouse Ig) antibodies (RAMA). In contrast, the RAMA responses detected in these rats were even stronger than those following immunization with complete mAb ACA125. In conclusion, the anti-idiotypic scFv ACA125 alone cannot improve the immunogenic features of the corresponding mAb, but provides a useful tool for the further development of genetic vaccines. Received: 20 January 2000 / Accepted: 24 April 2000     

Protective immunization against group B meningococci using anti-idiotypic mimics of the capsular polysaccharide

Use of the serogroup B meningococcal capsular polysaccharide (MenB CP) as a vaccine is hampered by the presence of epitopes that cross-react with human polysialic acid. As non-cross-reactive, protective capsular epitopes have also been described, we set out to develop protein mimics of one of such epitopes using as a template a highly protective mAb (mAb Seam 3) raised against a chemically modified form of the MenB CP (N-Pr MenB CP). Using phage display, anti-idiotypic single-chain Ab fragments (scFvs) were obtained from spleen cells of mice immunized with the Seam 3 mAb. Two Seam 3-specific scFvs competed with N-Pr MenB CP for binding to either mAb Seam 3 or rabbit Abs present in typing sera. Moreover, in mice and rabbits the scFvs elicited the production of Abs reacting with both N-Pr MenB CP and whole meningococci, but not with human polysialic acid. These scFv-induced Ab responses were boostable and of the Th1 type, as shown by a predominance of IgG2a. In addition, passive immunization with sera from scFv-immunized animals partially protected neonatal mice from experimental infection with group B meningococci. In conclusion, we have produced anti-idiotypic scFvs that mimic a protective MenB CP epitope and may be useful in the development of an alternative group B meningococcal vaccine.     

Antibodies against tumor cell glycolipids and proteins, but not mucins, mediate complement-dependent cytotoxicity

One of several effector mechanisms thought to contribute to Ab efficacy against cancer is complement-dependent cytotoxicity (CDC). Serological analysis of a series of clinical trials conducted over a 10-year period suggested that six vaccines containing different glycolipids induced Abs mediating CDC whereas four vaccines containing carbohydrate or peptide epitopes carried almost exclusively by mucin molecules induced Abs that did not mediate CDC. To explore this further, we have now compared cell surface reactivity using flow cytometry assays (FACS), complement-fixing ability, and CDC activity of a panel of mAbs and immune sera from these trials on the same two tumor cell lines. Abs against glycolipids GM2, globo H and Lewis Y, protein KSA (epithelial cell adhesion molecule, also known as EpCAM) and mucin Ags Tn, sialylated Tn, Thomsen Friedenreich (TF), and MUC1 all reacted comparably by FACS with tumor cells expressing these Ags. Compared with the strong complement binding and CDC with Abs against glycolipids and KSA, complement binding was diminished with Abs against mucin Ags and no CDC was detected. A major difference between these two groups of Ags is proximity to the cell membrane. Glycolipids and globular glycoproteins extend less than 100 A from the cell membrane while mucins extend up to 5000 A. Although complement activation at sites remote from the cell membrane has long been known as a mechanism for resistance from complement lysis in bacteria, it is identified here for the first time as a factor which may contribute to resistance from CDC against cancer cells.     

Elimination of the immunogenicity of therapeutic antibodies

The immunogenicity of therapeutic Abs limits their long-term use. The processes of complementarity-determining region grafting, resurfacing, and hyperchimerization diminish mAb immunogenicity by reducing the number of foreign residues. However, this does not prevent anti-idiotypic and anti-allotypic responses following repeated administration of cell-binding Abs. Classical studies have demonstrated that monomeric human IgG is profoundly tolerogenic in a number of species. If cell-binding Abs could be converted into monomeric non-cell-binding tolerogens, then it should be possible to pretolerize patients to the therapeutic cell-binding form. We demonstrate that non-cell-binding minimal mutants of the anti-CD52 Ab CAMPATH-1H lose immunogenicity and can tolerize to the "wild-type" Ab in CD52-expressing transgenic mice. This finding could have utility in the long-term administration of therapeutic proteins to humans.     

Vaccination with a human high molecular weight melanoma-associated antigen mimotope induces a humoral response inhibiting melanoma cell growth in vitro

Peptide mimics of a conformational epitope that is recognized by a mAb with antitumor activity are promising candidates for formulations of anticancer vaccines. These mimotope vaccines are able to induce a polyclonal Ab response focused to the determinant of the mAb. Such attempts at cancer immunotherapy are of special interest for malignant melanoma that is highly resistant to chemotherapy and radiotherapy. In this study, we describe for the first time the design and immunogenicity of a vaccine containing a mimotope of the human high m.w. melanoma-associated Ag (HMW-MAA) and the biological potential of the induced Abs. Mimotopes were selected from a pVIII-9mer phage display peptide library with the anti-HMW-MAA mAb 225.28S. The mimotope vaccine was then generated by coupling the most suitable candidate mimotope to tetanus toxoid as an immunogenic carrier. Immunization of rabbits with this vaccine induced a specific humoral immune response directed toward the epitope recognized by the mAb 225.28S on the native HMW-MAA. The induced Abs inhibited the in vitro growth of the melanoma cell line 518A2 up to 62%. In addition, the Abs mediated 26% lysis of 518A2 cells in Ab-dependent cellular cytotoxicity. Our results indicate a possible application of this mimotope vaccine as a novel immunotherapeutic agent for the treatment of malignant melanoma.     

Induction of the immune response to Legionella pneumophila cytolysin by monoclonal anti-idiotypic antibodies

A panel of mAb (IgG1, IgG3, IgM) against Legionella pneumophila cytolysin (CL)-protease of 37 kDa was obtained. Subtyping of L. pneumophila strains of serogroup 1 by using mAb against CL (mAb-CL) was carried out. The results of comparative analysis of the specificity of mAb-CL and the panel of mAb kindly provided by Dr. J. M. Barbaree (Centers for Disease Control, Atlanta, GA) allowed us to recommend mAb-CL to be used as a diagnostic tool to reveal the pathogenicity of L. pneumophila strains of serogroup 1. Hybridomas were also raised in a syngenic system which produced anti-idiotypic mAb (mAb2) against anti-CL mAb B6/1. The Ab2 belonged to Ab2 gamma type: 1) Ab2 reacted with B6/1 Id only, 2) Ab2 inhibited the interaction of B6/1 Ab1 with CL, and 3) CL inhibited the reaction of Ab2 with Ab1. The use of Ab2 allowed us to show that B6/1 Id is expressed in 4 to 32% of serum antibodies during the primary and secondary immune responses of BALB/c mice to CL. Ab2 induced the production of anti-anti-idiotypic antibodies (Ab3) in BALB/c mice, and some of them reacted with CL. Thus, we have demonstrated the possibility of inducing an antibody response to CL (one of the main L. pneumophila pathogenic factors) in intact syngenic mice with anti-idiotypic antibodies.     

Vaccine-induced CD4+ T cell responses to MAGE-3 protein in lung cancer patients

MAGE-3 is the most commonly expressed cancer testis Ag and thus represents a prime target for cancer vaccines, despite infrequent natural occurrence of MAGE-3-specific immune responses in vivo. We report in this study the successful induction of Ab, CD8(+), and CD4(+) T cells in nonsmall cell lung cancer patients vaccinated with MAGE-3 recombinant protein. Two cohorts were analyzed: one receiving MAGE-3 protein alone, and one receiving MAGE-3 protein with adjuvant AS02B. Of nine patients in the first cohort, three developed marginal Ab titers and another one had a CD8(+) T cell response to HLA-A2-restricted peptide MAGE-3 271-279. In contrast, of eight patients from the second cohort vaccinated with MAGE-3 protein and adjuvant, seven developed high-titered Abs to MAGE-3, and four had a strong concomitant CD4(+) T cell response to HLA-DP4-restricted peptide 243-258. One patient simultaneously developed CD8(+) T cells to HLA-A1-restricted peptide 168-176. The novel monitoring methodology used in this MAGE-3 study establishes that protein vaccination induces clear CD4(+) T cell responses that correlate with Ab production. This development provides the framework for further evaluating integrated immune responses in vaccine settings and for optimizing these responses for clinical benefit.     

Polymorphonuclear granulocytes induce antibody-dependent apoptosis in human breast cancer cells

Recent studies in HER-2/neu-targeted immunotherapy demonstrated that polymorphonuclear neutrophils (PMN) mediated Ab-dependent cellular cytotoxicity against HER-2/neu-positive breast cancer cell lines. However, the mechanism of cell death remained unclear. We used several assays to analyze the induction of apoptosis in the breast cancer cell line SK-BR-3 via PMN-dependent Ab-dependent cellular cytotoxicity. In the presence of the HER-2/neu Ab 520C9 and PMN from healthy donors, apoptosis occurred as detected by annexin V binding and disappearance of euploid SK-BR-3 nuclei, which can be differentiated from PMN nuclei by their increased DNA contents. Apoptosis induction was observed with E:T cell ratios as low as 10:1. Laser scanning fluorescence microscopy of TUNEL tumor cells or staining for cleaved cytokeratin-18 further confirmed apoptosis of the SK-BR-3 breast cancer cells. Killing via 520C9 was dependent on the interaction with FcR on PMN, because 1) F(ab')(2) fragments of 520C9 mediated no cytotoxicity, 2) target cell death was influenced by a biallelic polymorphism of FcgammaRIIa on the effector cells, and 3) a bispecific Ab against HER-2/neu and the IgA receptor (FcalphaRI) expressed on effector cells significantly induced apoptosis. Thus, PMN induce Ab-dependent apoptosis against human breast cancer cells targeted with HER-2/neu-directed mAbs or FcR directed bispecific Abs.     

High idiotypic connectivity of the VH7183-encoded antibodies directed to a murine embryonic carbohydrate antigen, Lewis Y, as ascertained by syngenic anti-idiotype monoclonal antibodies

The Lewis Y Ag is a carbohydrate Ag which is closely related to a well-known murine embryonic Ag, the stage-specific embryonic Ag-1 (SSEA-1), in its biochemical structure. It is expressed at the surface of murine embryonic cells as well as many murine cancer cells. For the analysis of idiotopes carried by the anti-Lewis Y antibodies, we generated two syngenic anti-idiotypic mAb, Id-A1 and Id-B4 (both BALB/c IgG1), which are directed to the idiotypic determinants carried by the anti-Lewis Y mAb, AH-6 (BALB/c IgM). Both Id-A1 and Id-B4 (Ab2) recognized paratope-related idiotopes carried by the AH-6 antibody (Ab1); they specifically inhibited the binding of AH-6 to the Lewis Y Ag. The high idiotypic connectivity of anti-Lewis Y antibodies was noted; the polyclonal anti-idiotype antibody, produced in the sera of BALB/c mice by immunizing AH-6 antibody, cross-reacted with several anti-Lewis Y mAb which has been established in different laboratories. Id-B4 and Id-A1 seem to represent such cross-reactive anti-idiotypic antibodies. Id-A1 recognized an idiotope carried by two out of six panel Ab1 mAb directed to the Lewis Y Ag. Id-B4 reacted with four out of the six panel antibodies, and was considered to recognize a recurrent idiotope of anti-Lewis Y antibodies which occurs more commonly than the idiotope recognized by Id-A1. All of the anti-Lewis Y antibodies which carry idiotopes that react with Id-A1 or Id-B4 were encoded by the VH genes of the VH7183 family; the most D-J proximal VH gene family in BALB/c mice, which is known to be preferentially expressed in embryonic B cells. Immunization of BALB/c mice with keyhole limpet hemocyanin-conjugated Id-B4 and/or Id-A1 induced a significant titer of anti-Lewis Y antibodies (Ab1-like Ab3) in the sera.     

Induction of cellular immunity by anti-idiotypic antibodies mimicking GD2 ganglioside

Gangliosides are potentially useful targets for tumor destruction by antibodies. However, the role of gangliosides in T cell-mediated immunity to tumors is unclear. We produced three murine monoclonal anti-idiotypic antibodies (Ab2) against a monoclonal antibody (Ab1) that binds strongly to melanoma-associated GD2 ganglioside and weakly to GD3 ganglioside. All three Ab2 induced anti-anti-idiotypic antibodies (Ab3) with Ab1-like binding specificity to tumor cells and antigen in rabbits. The Ab3 specifically bound to GD2(+) tumor cells and isolated GD2, and shared idiotopes with the Ab1. Two of the three Ab2 induced GD2-specific delayed-type hypersensitivity responses in BALB/c and C57BL/6 mice, but not in C57BL/6/CD4(-/-) mice. Peripheral blood mononuclear cells (PBMC) from a melanoma patient proliferated specifically in response to in vitro stimulation with Ab2. Proliferation was accompanied by Th1-type cytokine production. Our studies demonstrate the induction of ganglioside-specific T cell-dependent immunity by Ab2 in mice. These T cells showed specific reactivity to ganglioside expressed by tumor cells.     

Gangliosides induce cell apoptosis in the cytotoxic line CTLL-2, but not in the promyelocyte leukemia cell line HL-60

Gangliosides induce apoptosis in the cells of the IL-2-dependent cytotoxic mouse line CTLL-2. Upon incubation with gangliosides for 24 h, their effect resulting in appearance of apoptotic cells, falls in a series GM2 > GM3 > GM1 > GD1a > GD1b > GT1b. In the presence of rIL-2, apoptosis induced by GM1 is suppressed, whereas that induced by GM2 is enhanced (the effect of intracellular agent C2-Cer is independent of this cytokine). The GM1-induced apoptosis is cancelled by the caspase I inhibitor. The gangliosides under study are not able to induce apoptosis in the promyelocyte leukemia cell line HL-60. Physiological aspects of the phenomenon found are discussed.     

Anti-idiotype-induced, lipopolysaccharide-specific antibody response to Pseudomonas aeruginosa

Antibody directed to the O-specific polysaccharide (Ps) side chain of Pseudomonas aeruginosa LPS provides immunotype-specific protection against infection by virtue of enhancing opsonophagocytosis. We have developed a syngeneic anti-idiotypic antibody (mAb2) directed to a functionally active monoclonal immunotype 1 Ps-antibody (mAb1). The mAb2 performed as a molecular mimic of Ps as evidenced by 1) blocking of mAb1/mAb2 interaction by Ps, 2) blocking of mAb1/Ps binding by mAb2, 3) cross-species binding of mAb2 to human Ps antibodies from individuals immunized with the same immunotype 1 Ps, and 4) induction of anti-LPS antibody by immunization with mAb2 in syngeneic mice. Our studies thus show that an anti-idiotypic antibody may functionally mimic the O-polysaccharide of P. aeruginosa LPS, and bind to cross-reactive Id present in human Ps antibodies. We have further shown that this anti-idiotypic antibody induces anti-LPS antibody when used as an Ag in syngeneic mice, suggesting that this approach may eventually be used to successfully immunize humans.