Anti-idiotypic Antibody Response Research Articles

The Role of Regulatory B Cell-Like Malignant Cells and Treg Cells in the Mouse Model of BCL1 Tumor Dormancy

Cancer dormancy is a clinical state in which residual tumor cells persist for long periods of time but do not cause detectable disease. In the mouse B cell lymphoma model (BCL1), dormancy can be induced and maintained by immunizing mice with a soluble form of the IgM expressed on the surface of the tumor cells. Immunization induces an anti-idiotype antibody response that maintains dormancy. Mice with dormant tumor have low numbers of BCL1 cells in their spleens that divide and are killed at the same rate. When the anti-Id antibodies wane, the tumor cells grow rapidly and kill the host. Spleens from tumor-bearing mice contain both effector (CD4+ and CD8+) and regulatory T cells (Tregs). In other tumor models, it has been reported that Tregs promote tumor progression by preventing effector cells from killing the tumor. In this report, we demonstrate that the tumor site with rapidly dividing BCL1 cells has fewer Tregs than the tumor site harboring dormant BCL1 cells. In both cases, the Tregs were equally suppressive in vitro. In spleens from mice with actively growing tumor, CD8+ but not CD4+ T cells were virtually absent. In vitro analysis demonstrated a tumor-mediated elimination of CD8+ T cells that was contact dependent and involved the caspase-3 pathway. Most importantly, we found that the BCL1 cells expressed characteristics of B10 regulatory B cells, i.e., they were CD1dhiCD5+ and secreted high levels of IL-10. These BCL1 tumor cells can inhibit anti-tumor immune responses by depleting CD8+ effector T cells.

Immunological and histological evaluation of clinical samples from psoriasis patients treated with anti-CD6 itolizumab

Psoriasis is a chronic inflammatory disease with a prevalence of approximately 2–3% in the general population. The majority of diagnosed patients have plaque psoriasis, and about 20% have moderate-to-severe disease. Itolizumab, a new monoclonal antibody specific for the CD6 molecule mainly expressed on T lymphocytes, has demonstrated to inhibit in vitro ligand-induced proliferation and pro-inflammatory cytokine production. We assessed the immunological and histopathological effect of the antibody using clinical samples taken from 26 patients with moderate-to-severe psoriasis included in a clinical trial. The precursor frequency of lymphocytes activated with anti-CD2/CD3/CD28 beads, as well as the number of interferon (IFN)-γ-secreting T cells after stimulation, were measured at different time points of the study. Serum cytokine levels and anti-idiotypic antibody response to itolizumab were also evaluated. Additionally, lymphocyte infiltration and epidermis hyperplasia were studied in five patients. A significant reduction in T cell proliferation capacity and number of IFN-γ-producing T cells was found in treated patients. Serum levels of interleukin-6, tumor necrosis factor and IFN-γ showed an overall trend toward reduction. No anti-idiotypic antibody response was detected. A significant reduction in the epidermis hyperplasia was observed in analyzed patients. These results support the relevance of the CD6 molecule as a therapeutic target for the treatment of this disease.

Targeted DNA vaccines eliciting crossreactive anti-idiotypic antibody responses against human B cell malignancies in mice.

BackgroundTherapeutic idiotypic (Id) vaccination is an experimental treatment for selected B cell malignancies. A broader use of Id-based vaccination, however, is hampered by the complexity and costs due to the individualized production of protein vaccines. These limitations may be overcome by targeted DNA vaccines encoding stereotyped immunoglobulin V regions of B cell malignancies. We have here investigated whether such vaccines might elicit cross-reactive immune responses thus offering the possibility to immunize subsets of patients with the same vaccine.MethodsFusion vaccines targeting patient Id to mouse Major Histocompatibility Complex (MHC) class II molecules (chimeric mouse/human) or chemokine receptors (fully human) on antigen-presenting cells (APC) were genetically constructed for two Chronic Lymphocytic Leukemia (CLL) patients and one prototypic stereotyped B-cell receptor (BCR) commonly expressed by Hepatitis C Virus (HCV)-associated Non Hodgkin Lymphoma (NHL). The A20 murine B lymphoma cells were engineered to express prototypic HCV-associated B cell lymphoma BCR. Anti-Id antibody responses were studied against stereotyped and non-stereotyped BCRs on CLL patients’ cells as well as transfected A20 cells.ResultsDNA vaccination of mice with Id vaccines that target APC elicited increased amounts of antibodies specific for the patient’s Id as compared with non targeted control vaccines. Anti–Id antibodies cross-reacted between CLL cells with closely related BCR. A20 cells engineered to express patients’ V regions were not tumorigenic in mice, preventing tumor challenge experiments.ConclusionsThese findings provide experimental support for use of APC-targeted fusion Id DNA vaccines for the treatment of B cell lymphoma and CLL that express stereotyped BCRs.

AB0055 Adalimumab elicits a restricted anti-idiotypic antibody response in autoimmune patients resulting in functional neutralization

BackgroundWorldwide millions of patients are treated with monoclonal antibodies. These biological therapeutics can be immunogenic, resulting in antibody formation which lead to loss of response. Fully human biologics such as...

Adalimumab elicits a restricted anti-idiotypic antibody response in autoimmune patients resulting in functional neutralisation

ObjectivesMillions of patients worldwide are treated with therapeutic monoclonal antibodies. These biological therapeutics can be immunogenic, resulting in anti-drug antibody formation which leads to loss of response. Fully human biological...

Immune monitoring of phase II trial of hu14.18-IL2 for patients with metastatic melanoma.

e19047 Background: Phase I testing of the hu14.18-IL2 immunocytokine (IC) in patients (pts) with melanoma showed immune activation, reversible toxicities, and a maximal tolerated dose (MTD) of 7.5 mg/m2/day. Methods: In this phase II study, 14 metastatic melanoma pts were scheduled to receive hu14.18-IL2 at 6 mg/m2/day as 4-hour intravenous infusions on days 1, 2 and 3 of each 28 day cycle. Pts with stable disease (SD) or regression following cycle 2 could receive 2 additional treatment cycles. The primary objective was to evaluate anti-tumor activity and response duration. Secondary objectives evaluated adverse events and immunologic activation. Results: All pts received 2 cycles of treatment. One pt had a partial response (PR) [1 PR of 14 pts = response rate of 7.1%; confidence interval 0.2%-33.9%] and 4 pts had SD and received cycles 3 and 4. The PR and SD responses lasted 3-4 months. All toxicities were reversible. Analysis of serum IC for all pts showed a mean value of 1.42 ug/ml on C1D1 and 1.38 ug/ml on C1D3. Pts had a peripheral blood lymphocytosis on day 8 and increased C-reactive protein (CRP). A significant increase in sIL2R levels was seen in all courses from baseline to day 3 (p<0.001 for both courses 1 and 2). Infusion of the IC into these pts enabled their serum to mediate antibody dependent cell-mediated cytotoxicity using the circulating IC. Eight pts developed a “positive” anti-idiotype antibody against hu14.18-IL2 based on a bridging assay, and 13 developed a detectible anti-idiotypic antibody based on a binding inhibition assay. There was no significant effect of this anti-idiotypic antibody response on the in vivo level of hu14.18-IL2. While one PR in 14 pts met protocol criteria to proceed to stage 2 and enter 16 additional pts, we suspended stage 2 due to limited availability of hu14.18-IL2 at that time and the brief duration of PR and SD. Conclusions: We conclude that subsequent testing of hu14.18-IL2 should involve melanoma pts with minimal residual disease based on compelling preclinical data as well as the confirmed immune activation with some antitumor activity in this study. In addition, the demonstrated immunological activation of hu14.18-IL2 with manageable toxicities suggests the potential for combination treatments.

Abstract LB-108: Racotumomab antitumor activity in the murine model of non small cell lung carcinoma 3LL is related with the level of expression on NGcGM3

Abstract N-Glycolylneuraminic acid (NGc) is a sialic acid molecule found in most mammalian cells. It is usually found as terminal constituents of different membrane glycoconjugates such as the GM3 ganglioside (NGcGM3) and has been described as tumor antigen in non small cell lung cancer (NSCLC) in humans. However, most mouse cancer cell lines, such as 3LL NSCLC, are negative for the expression of this antigen although mice somatic cells frequently express it. It was reported that minor component of the ganglioside fraction derived from primary tumors of 3LL is NGcGM3. Interestingly in metastatic lesions the presence of NGcGM3 reach the 73.4% of total lipid bound sialic acid. Anti idiotypic antibodies (Ab2) are proposed as therapeutic vaccines when they bear the internal image of tumor antigens. Racotumomab is a gamma type anti idiotypic antibody that is able to trigger a strong anti anti idiotypic antibody (Ab3) response. These Ab3 antibodies have the same antigenic specificity than the Ab1 (P3) but they are substantially different from the internal image of it. Additionally, since it was demonstrated that immunization with Racotumomab elicited an anti NGcGM3 antibody response in humans, the question about to what extent the presence of the vaccine target antigen in the tumor decides the effect of the active treatment remains open. In this work, we evaluated the association of NGcGM3 expression in tumor cells in relation to the antitumor activity of the vaccine. Evaluation of Racotumomab antitumor activity over 3LL primary tumor showed no difference in tumor growth rate between treated and control mice. Moreover, the vaccine showed no antitumor activity in the metastatic dissemination in C57BL/6 mice treated with iv 3LL cells. However, when Racotumomab activity was evaluated in a spontaneous metastasis model it was observed a reduction in lung metastatic nodules in treated group compared to the control. The amount of NGcGM3 in 3LL cells can be increased by preincubation in vitro with purified NGc. Using this NGcGM3 positive metastasis model, we observed that Racotumomab was able to develop an active antitumor activity. The number of experimental metastases was reduced significantly between control and treated group (3.5 (1-18) vs 0.5(1-10), control vs Racotumomab. p<0.05, Kruskal-Wallis test). In a preliminary study of the therapeutic combination or Racotumomab with chemotherapy, mice were injected with 3LL cells and treated with Racotumomab, Pemetrexed or both. The combination of the vaccine and Pemetrexed show a similar antitumor effect than Racotumomab alone. Our results suggest a close association between the expression of NGcGM3 and the antitumor activity of Racotumomab. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr LB-108. doi:1538-7445.AM2012-LB-108

Inhibition of Interleukin-6 (IL-6) Reverses Anemia In Patients with Advanced Non Small Cell Lung Cancer (NSCLC): Results of a Phase II, Randomized, Double-Blind, Placebo-Controlled Trial

AbstractAbstract 640 Background:ALD518 is a humanized, desialyated anti-IL-6 antibody being developed for the treatment of cancer-related anemia, cachexia and fatigue. The primary objective of the study was to determine the efficacy and safety of ALD518 in patients with advanced NSCLC. Secondary objectives examined hematologic parameters. Methods:124 patients with NSCLC, ECOG 0–3, weight loss in the preceding 3 months of >5% body weight, hemoglobin (Hb) >7g/dL, and C-reactive protein (CRP) >10mg/L were dosed. Patients were randomized to 1 of 4 groups (n~30/group). Placebo or ALD518 80mg, 160mg, or 320mg was administered intravenously every 8 weeks. Pts were followed up for 24 weeks. Data included hematologic parameters, clinical chemistry, CRP, D-dimer, lean body mass and adverse events (AEs). Quality of life data included the FACIT-F, FACT-L, and FAACT questionnaires. Data presented in this abstract relates to the safety and hematology results. Results:29 pts completed the study treatments and evaluations, 38 failed to complete every visit, 52 died of progressive disease, and 5 withdrew because of adverse events. There were no dose limiting toxicities (DLTs), infusion reactions, or anti-idiotypic antibody responses to ALD518 observed in the study. 84 pts had serious AEs of which 1 was deemed to be possibly related to administration of ALD518 (rectal hemorrhage). The majority of the serious adverse events were due to progression of the NSCLC. Six patients had a CTC grade 4 change in laboratory safety data during the study. Four patients experienced a grade 4 hypercalcemia: 1 (3.6%), 2 (6.1%), and 1 (3.2%) in the ALD518 80mg, 160mg and placebo groups, respectively, and there was 1 patient with grade 4 GGT elevation (placebo) and 1 patient with grade 4 hypokalemia (ALD518 160mg). There were no treatment related differences in vitals sign or 12-lead ECG data.The mean (±SD) values for Hb, hematocrit (Hct), mean corpuscular Hb (MCH) and platelet counts are listed below:38/93 pts treated with ALD518 and 10/31 given placebo had a pre-dose Hb =< 11g/dL. 24 of these pts on ALD518 and 7 of these pts on placebo remained in the study at week 4. 14/24 pts on ALD518 and 0/7 on placebo had raised their Hb from =< 11g/dL to >= 12g/dL. Conclusions:ALD518 increased Hb, Hct, MCH in NSCLC pts and raised Hb to >= 12g/dL in 58% of pts with a Hb =< 11g/dL at baseline. There was also a modest fall in platelet count observed in patients treated with ALD518 but no patients had a CTC grade 4 thrombocytopenia and only one patient (ALD518 160mg group) had a grade 3 thrombocytopenia at one time point. There were no major safety signals related to the administration of ALD518. Further study of ALD518 as a novel non-erythropoietic stimulating agent for cancer-related anemia is warranted. Disclosures:Schuster:Alder Biopharmaceuticals Inc: Honoraria. Rigas: Alder Biopharmaceuticals inc: Honoraria. Smith:Alder Biopharmaceuticals Inc: Employment.

Immunogenicity of the hu14.18-IL2 immunocytokine molecule in adults with melanoma and children with neuroblastoma.

Immunocytokine (IC) hu14.18-IL2 is a fusion protein of humanized antidisialoganglioside (GD2) antibody (hu14.18) and interleukin (IL)-2. Sixty-one melanoma and neuroblastoma patients received IC in phase I/Ib studies. Patient sera were examined in ELISA to determine if an anti-IC antibody response occurred during treatment. Serum was assayed for anti-idiotypic antibody (anti-id Ab) based on ability to bridge biotinylated hu14.18 to plate-bound hu14.18 and ability to inhibit binding of hu14.18 to GD2 antigen and/or murine anti-idiotypic antibody. ELISA was also used to detect antibodies to the Fc-IL2 end of hu14.18-IL2. Thirty-two patients (52%) developed an anti-idiotypic antibody response (absorbance, >0.7) in the bridge ELISA. Twelve patients (20%) had an intermediate response, whereas 17 patients (28%) were negative (adsorbance, <0.3). The development of antibody to hu14.18-IL2 detected in the bridge ELISA was not related to the dose of hu14.18-IL2. Twenty of 33 adult patients (61%) demonstrated an anti-idiotypic antibody response based on binding inhibition ELISA. The anti-idiotypic response was inversely correlated (P < 0.002) with IC measured during the second course of treatment, indicating that development of anti-idiotypic antibodies interfered with detection of circulating hu14.18-IL2. All patients developed some inhibitory activity in the binding inhibition assay designed to detect antibodies to the Fc-IL2 region of the IC. There was a positive correlation between the peak serum level of IC in course 1 and the anti-Fc-IL2 response. Patients treated with hu14.18-IL2 developed anti-idiotypic antibodies and anti Fc-IL2 antibodies. No association was seen between development of anti-IC antibodies and clinical toxicity.

Anti-idiotype antibody response after vaccination correlates with better overall survival in follicular lymphoma

AbstractPrevious studies demonstrated that vaccination-induced tumor-specific immune response is associated with superior clinical outcome in patients with follicular lymphoma. Here, we investigated whether this positive correlation extends to overall survival (OS). We analyzed 91 untreated patients who received CVP chemotherapy (cyclophosphamide, vincristine, and prednisone) followed by idiotype vaccination. Idiotype proteins were produced either by the hybridoma method or by expression of recombinant idiotype-encoding sequences in mammalian or plant-based expression systems. We found that achieving a complete response/complete response unconfirmed (CR/CRu) to CVP and making an anti-idiotype antibody are 2 independent factors that each correlated with longer OS at 10 years (89% vs 68% with or without a CR/CRu, P = .024; 90% vs 69% with or without tumor-specific antibody production; P = .027). In the subset of patients who received hybridoma-generated vaccines, we found that anti-idiotype production was even more highly associated with superior OS (P < .002); this was the case even in patients with a partial response (PR) to CVP (P < .001).

Anti-Idiotype Antibody Response after Vaccination Correlates with Better Overall Survival in Follicular Lymphoma.

Purpose Immunoglobulin idiotype (Id) is a clonal marker expressed by B cells. Lymphoma B cells express a unique Id which serves as an ideal target for immunotherapy. It has been shown that induction of anti-Id immune response through vaccination is associated with many clinical benefits, including tumor regression, molecular remission, and prolonged progression free survival (PFS). However, it is unknown if induced immune response is an independent predictor for overall survival (OS). Patients and Methods We analyzed 91 patients who were uniformly treated with CVP followed by idiotype vaccination as the first-line therapy. The induction of anti-Id immune response was evaluated and correlated with OS. Univariate and multivariate analyses were applied to select independent factors that predicted OS.Results: Those patients who achieved a CR/CRu to CVP chemotherapy or produced anti-Id antibody after vaccination had longer survival than those who did not. At 10 years, OS was 90% and 68% for patients with or without a CR/CRu after CVP (p = 0.024); 90% and 69% for patients with or without tumor-specific antibody production, respectively (p = 0.027). In contrast, generation of anti-Id cellular response did not correlate with OS. Furthermore, CR/CRu to CVP and generation of anti-Id antibody response were the only two independent factors among those tested that were correlated with longer survival.Conclusion: Generation of anti-Id antibody after active immunotherapy is an independent predictor for better OS in follicular lymphoma.

Generation and Characterization of an Anti-Idiotype Monoclonal Antibody Related to GM3(NeuGc) Ganglioside

The 14F7 monoclonal antibody (MAb), IgG1 isotype, which reacts specifically to GM3(NeuGc) ganglioside induced a specific IgG anti-idiotypic antibody (Ab2) response in syngeneic mice when it was administered coupled with KLH and in the presence of Freund's adjuvant. Spleen cells from these mice were used in somatic-cell hybridization experiments using the murine myeloma cell line P3-X63-Ag8 653 as fusion partner. An IgG1 Ab2 MAb was selected. This Ab2 MAb, called 4G9, was able to block the binding of 14F7 MAb to GM3(NeuGc) ganglioside and developed a strong IgG anti-anti-idiotypic antibody (Ab3) response, when injected into syngeneic mice. These Ab3 antibodies were characterized to bear 14F7 MAb idiotopes, but did not have the same specificity as 14F7 MAb. In the other hand, a very specific anti-NeuGc-containing ganglioside response was generated in chickens immunized with this Ab2 MAb, thus behaving, in this species as an "internal image" antibody.

Chimeric anti-N-glycolyl-ganglioside and its anti-idiotypic MAbs: immunodominance of their variable regions.

P3 monoclonal antibody (MAb) is a murine IgM that specifically recognizes N-glycolyl (NeuGc)-gangliosides and sulfatides. It also reacts with antigens expressed in human breast tumors and melanoma. In syngeneic model, P3 MAb is able to elicit a strong anti-idiotypic (Ab2) antibody response, even in the absence of adjuvants or carrier proteins. 1E10 MAb is an anti-idiotypic antibody specific for P3 MAb that has demonstrated anti-tumoral effects in syngeneic and allogeneic animals. Here we report the construction of the human IgG(1) chimeric P3 and 1E10 antibodies, and the evaluation of the maintenance of the main properties of the murine MAbs. Chimeric P3 antibody specifically reacted with GM3(NeuGc) and GM2(NeuGc) gangliosides, and not with their acetylated variants. Also, it strongly recognized the anti-idiotypic 1E10 MAb. Chimeric 1E10 antibody specifically reacted with P3 MAb. Upon immunization of Balb/c mice with both chimeric antibodies, we were able to demonstrate the immunodominance of their variable regions. The anti-idiotypic response induced by both antibodies was strong and in most of the mice was even significantly higher than the anti-isotypic response, despite the fact that 70% of the chimeric molecule is xenogenic with respect to the animal model.

Anti-idiotypic DNA vaccines for lymphoma immunotherapy require the presence of both variable region genes for tumor protection.

Vaccination with immunogenic formulations of lymphoma-derived immunoglobulin can elicit strong anti-idiotypic immune responses which have proved effective in murine B cell tumor challenge experiments and suggested possible benefits in recent human clinical trials. Naked plasmid DNA vaccines encoding the Id determinants as scFv fragments provide the most promising alternative to protein immunization. With this approach the addition of an immunogenic domain linked to the scFv has proved essential for the induction of a protective immune response. In this study we have produced a scFv gene construct linked to the CH3 exon of the human IgG1 constant region and tested its efficacy in inducing protective immunity against the mouse BCL1 lymphoma. We have also generated a second construct in which the BCL1 VL gene was deleted to investigate whether the VH region domain contains sufficient antigenic determinants for a protective immune response. Both constructs induced anti-idiotypic antibodies that specifically reacted with the BCL1 IgM protein in ELISA and with BCL1 tumor cells in flow cytometry assays. Protection against tumor challenge was fully achieved with the complete scFv construct whereas immunization with the construct lacking the VL gene resulted in only a slight prolongation of the survival. We therefore conclude that a plasmid DNA vaccine containing the VH and VL genes of the lymphoma Ig linked to the human IgG1 CH3 exon is highly effective in inducing a protective immune response in the BCL1 model. We also demonstrated that VH gene immunization can induce strong anti-idiotypic antibody responses.

PHARMACOKINETIC AND PHARMACODYNAMIC EVALUATION OF A TWO DOSE DACLIZUMAB REGIMEN IN LIVER TRANSPLANTATION

100 Daclizumab (Zenapax®) is a humanized mAb directed to the α-chain of the IL-2 receptor (CD25). 1st pharmacokinetics and pharmacodynamics of a two dose regimen were characterized in an open-label, single center phase I/II trial in liver transplant patients. Methods: 26 de novo liver transplant patients received two doses of daclizumab intravenously at day 0, 1mg/kg and day 4, 0.5 mg/kg in addition to the microemulsion of cyclosporine and steroids. Serial blood samples were collected over 12 weeks for measurement of daclizumab serum and ascites concentrations (idiotype-specific ELISA) and the formation of anti-idiotypic antibodies. Flow cytometry (CD3, Cd4, CD8, CD20, CD25) was performed in parallel in all patients. The daily blodd loss and ascites fluid drainage were documented up to two weeks posttransplant. Absolute lymphocyte counts and differential blood pictures were performed by standard techniques. Results: (1) Peak serum concentrations after the first dose were 13.8 ± 5.8 μg/ml and 8.9 ± 4.1 μg/ml. (2) Concentrations higher than 1 μg/ml were measured until day 14 posttransplant (1.2 ± 1.1 μg/ml). (3) There were no relevant changes in CD3+, CD4+, CD8+ and CD20+ lymphocytes during exposure to daclizumab; whereas, CD25+ lymphocytes were not detectable during treatment (1.9 ± 0.9%) compared to pretransplant (21.4 ± 8.8%) and post-exposure (16 ± 14.1%). (4) Daclizumab demonstrated a concentration-effect relation, whereby concentrations exceeding 1 μg/ml were associated with CD25 suppression. (5) Daclizumab concentrations in ascites ranged from 2 to 6 μg/ml, but did not had an influence on serum levels. (6) No anti-idiotypic antibody response was seen. (7) 1/26 patients had a mild acute rejection on day 18 posttransplant and no side effects were seen after administration of daclizumab. Conclusions: Daclizumab was well tolerated in liver transplant recipients. Drainage of ascites fluid and postoperative bleeding contribute to the total body clearance of daclizumab, however there was no influence on the efficacy of CD25 coating. Therefore, daclizumab could be administered safely in a two dose regimen on day 0 and 4 after liver transplantation, achieving receptor saturation for 21 days.

Syngeneic anti-idiotypic monoclonal antibodies to an anti-NeuGc-containing ganglioside monoclonal antibody.

An IgM monoclonal antibody (MAb), named P3, has the characteristic to react specifically with a broad battery of N-glycolyl containing-gangliosides and with antigens expressed on breast tumors. When this MAb was administered alone in syngeneic mice, an specific IgG anti-idiotypic antibody (Ab2) response was induced, this Ab2 response was increased when P3 MAb was injected coupled to a carrier protein and in the presence of Freund's adjuvant. Spleen cells from these mice were used in somatic-cell hybridization experiments, using the murine myeloma cell line P3-X63-Ag8.653 as fusion partner. Five Ab2 MAbs specific to P3 MAb were selected. These IgG1 Ab2 MAbs were able to block the binding of P3 MAb to GM3(NeuGc) ganglioside and to a human breast carcinoma cell line. Cross-blocking experiments demonstrated that these Ab2 MAbs are recognizing the same or very close sites on the Abl MAb. The five Ab2 MAbs were injected into syngeneic mice and four of them produced strong anti-anti-idiotypic antibody (Ab3) response. While these Ab2 MAbs were unable to generate Ab3 antibodies with the same antigenic specificity than P3 MAb, three of them induced antibodies bearing P3 MAb idiotopes (Ag-Id+ Ab3). These results demonstrated that these Ab2 MAbs are not "internal image" antibodies, but they could define "regulatory idiotopes."

Edrecolomab (monoclonal antibody 17-1A).

Edrecolomab is a mouse-derived monoclonal IgG2a antibody. It recognises the human tumour-associated antigen CO17-1A which is expressed on the cell surface of a wide variety of tumours and normal epithelial tissue. Edrecolomab is thought to destroy tumour cells by activating an array of endogenous cytotoxic mechanisms, including antibody-dependent cell-mediated cytotoxicity and possibly antibody-dependent complement-mediated cytotoxicity. Edrecolomab may induce antitumour activity indirectly by inducing a host anti-idiotypic antibody response. Adjuvant therapy with edrecolomab (500 mg initial dose followed by four 100 mg infusions administered at 4-weekly intervals) significantly improved survival and reduced the tumour recurrence rate in patients with resected Dukes' stage C colorectal cancer and minimal residual disease. Data from several small clinical trials suggest that edrecolomab given as monotherapy or in combination with other antineoplastic agents has limited efficacy in the treatment of advanced colorectal or pancreatic tumours. However, results from a small phase I study in patients with advanced breast cancer were more promising. Edrecolomab was generally well tolerated in clinical trials. In a postmarketing surveillance study, the most common adverse events associated with edrecolomab were flushing/erythema and gastrointestinal symptoms including diarrhoea, abdominal pain and nausea and vomiting. Because edrecolomab is of murine origin, anaphylactic reactions have developed in some patients treated with the drug.

DNA vaccines against lymphoma: promotion of anti-idiotypic antibody responses induced by single chain Fv genes by fusion to tetanus toxin fragment C.

Idiotypic determinants can act as tumor-associated Ags for B cell lymphoma. Vaccination with idiotypic protein and adjuvant is known to induce specific protection against lymphoma challenge in mice, largely mediated by anti-idiotypic Ab. For facilitating the approach for patients, the V(H) and V(L) genes used to encode the individual idiotypic determinants of each tumor can be obtained by PCR and assembled as single chain Fv (scFv). DNA vaccines containing scFv sequences alone induce low and poorly reproducible levels of anti-idiotypic Ab, likely to be insufficient to suppress tumor in patients. In addition, it may be necessary to break tolerance to Id in tumor bearers. By fusing the gene for fragment C of tetanus toxin to the C terminus of human scFv, we have promoted the anti-scFv Ab response in mice by >50-fold in three of three cases. The induced Abs are mainly against idiotypic determinants, and react specifically with patients' tumor cells, indicating optimal folding of the scFv molecule in the fusion protein. For both antigenic components of the DNA vaccine, the IgG subclass distribution showed a relative increase in IgG2a as compared with vaccination with IgM protein in adjuvant. In patients, the fusion gene should both promote anti-idiotypic Ab and induce Abs against fragment C of tetanus toxin. The latter response would provide a potentially useful comparative measure of the ability of patients to respond to conventional Ag delivered via DNA.

Anti-igG autoantibodies and possible immune regulatory mechanisms in patients with pulmonary tuberculosis

Setting: Anti-Ig antibodies are known to have important clinical and biological implications. Objectives: To determine naturally occurring anti-F(ab′)2γ and anti-Fcγ antibodies in patients with pulmonary tuberculosis (PTB) in relation to various clinical manifestations and human leukocyte antigen (HLA). Design: Antibodies to F(ab′)2 and Fc portions of IgG were detected in the sera of normal healthy individuals ( n = 41), patients with pulmonary tuberculosis ( n = 50) and their household family contacts ( n = 20) using an enzyme immuno assay (EIA) system. Results: As compared to controls (0.110 ± 0.01 optical density [OD]), the levels of anti-F(ab′)2γ were significantly increased in PTB patients (0.998 ± 0.08 OD, P < 0.0001) and in their contacts (0.486 ± 0.04 OD, P<0.001) suggesting that the occurrence of these autoantibodies is related to infection/exposure to Mycobacterium tuberculosis. Anti-F(ab′)2γ antibodies were significantly increased in both sputum positive and negative patients ( P<0.0001) and no deviation was observed between these two groups. The levels of these antibodies were positively correlated with disease severity assessed by chest X-ray. The drug failure patients had higher activity of anti-F(ab′)2γ than drug responders and no impact of anti-tuberculosis chemotherapy was observed. A statistically significant increase of anti-F(ab′)2γ levels (1.25 ± 0.21 OD) was observed in HLA-DR2 positive patients as compared to the DR2 negative groups (1.02 ± 0.09 OD), P<0.01. No deviation was observed in the levels of anti-Fcγ levels between controls and any group of PTB patients. Conclusion: The present data suggests that the elevated levels of anti-F(ab′)2γ antibodies in PTB patients represent an anti-idiotypic antibody response to anti- M. tuberculosis antibody caused by an immune imbalance following M. tuberculosis infection.

Vaccination with syngeneic, lymphoma-derived immunoglobulin idiotype combined with granulocyte/macrophage colony-stimulating factor primes mice for a protective T-cell response.

The idiotype of the Ig expressed by a B-cell malignancy (Id) can serve as a unique tumor-specific antigen and as a model for cancer vaccine development. In murine models of Id vaccination, formulation of syngeneic Id with carrier proteins or adjuvants induces an anti-idiotypic antibody response. However, inducing a potent cell-mediated response to this weak antigen instead would be highly desirable. In the 38C13 lymphoma model, we observed that low doses of free granulocyte/macrophage colony-stimulating factor (GM-CSF) 10,000 units i.p. or locally s.c. daily for 4 days significantly enhanced protective antitumor immunity induced by s.c. Id-keyhole limpet hemocyanin (KLH) immunization. This effect was critically dependent upon effector CD4+ and CD8+ T cells and was not associated with any increased anti-idiotypic antibody production. Lymphocytes from spleens and draining lymph nodes of mice primed with Id-KLH plus GM-CSF, but not with Id-KLH alone, demonstrated significant proliferation to Id in vitro without any biased production of interferon gamma or interleukin 4 protein or mRNA. As a further demonstration of potency, 50% of mice immunized with Id-KLH plus GM-CSF on the same day as challenge with a large s.c. tumor inoculum remained tumor-free at day 80, compared with 17% for Id-KLH alone, when immunization was combined with cyclophosphamide. Taken together, these results demonstrate that GM-CSF can significantly enhance the immunogenicity of a defined self-antigen and that this effect is mediated exclusively by activating the T-cell arm of the immune response.