Monoclonal anti-idiotypic antibodies as probes for common idiotopes shared by anti-"benzo(a)pyrene-like" IgA of cancer patients and rabbit anti-conjugated benzo(a)pyrene antibodies.

Anti-"benzo(a)pyrene [B(a)P]-like" IgA [referred as idiotypic antibodies (Abl)] from cancer patients' sera were found to react with conjugated B(a)P and a monoclonal anti-anti-conjugated B(a)P, internal image of conjugated B(a)P called AIB1 (referred to as Ab2 beta). These IgA were used to raise mouse monoclonal anti-idiotypic antibodies (Ab2). A monoclonal Ab2 called AIK1 was characterized as the internal image of a "B(a)P-like" structure. As shown by competitive experiments, AIK1 inhibited the reaction between Ab1 from a rabbit anti-conjugated B(a)P serum- and its relevant internal image, AIB1. Furthermore, AIB1 inhibited the reaction between AIK1 and anti-"B(a)P-like" IgA from cancer patients' sera. These observations confirmed the cross-reactivity between idiotypic determinants of human anti-"B(a)P-like" IgA and rabbit anti-conjugated B(a)P antibodies (Ab). This result was reinforced by a correlation between the anti-"B(a)P-like" IgA levels found in cancer patients' sera using indirect ELISA method with conjugated B(a)P, AIB1 and AIK1 coated on well-plates.

Identification of a benzo[a]pyrene-like binding protein involved in circulating immune complexes of patients with mammary tumors.

Anti-benzo[a]pyrene (B[a]P)-like autoantibodies (autoAb) have been characterized in sera of patients with epithelial tumors. Circulating immune complexes (CIC) from these sera have been analysed after polyacrylamide gel electrophoresis (PAGE) under non-denaturing conditions. Immunoblotting was performed using a monoclonal antiidiotypic antibody (Ab), internal image of conjugated B[a]P called AIB1 and anti-human immunoglobulins (Ig). An immunoreactivity was seen only with AIB1 Ab, suggesting the presence of a 'B[a]P-like' binding protein. Additional studies showed that this immunoreactivity is not associated with an 18- to 20-kDa protein previously identified in the same CIC.

Chemically induced sarcomas in Sprague-Dawley rats: dose effects on autoantibody levels and tumor progression.

We previously reported that a single injection of 2 mg benzo[a]pyrene (B[a]P) induced seric anti-phosphatidylinositol (PtdIns) autoantibodies (autoAb) and highly malignant sarcomas with 100% efficiency in Sprague-Dawley (SD) female rats. To evaluate the effects of lower doses, we administered a single dose (from 0.125 to 1 mg) of B[a]P into such rats. In all cases, we noted significantly high levels of anti-PtdIns autoAb, with clinically palpable tumors appearing around Day 100. Tumors grew more slowly than they did in the previous model (2 mg B[a]P). Both anti-PtdIns autoAb levels and first appearance of the tumor seemed to be independent of carcinogen doses. Conversely, tumor evolution seemed to depend on B[a]P doses.

Curative effects on rat sarcomas obtained after a treatment combining two monoclonal antibodies.

The effects of a treatment involving two monoclonal antibodies (Ab) were evaluated on benzo(a)pyrene [B(a)P]-induced malignant sarcomas in Sprague-Dawley female rats. These Ab were, respectively, an anti-anti-conjugated B(a)P AB, an internal image of conjugated B(a)P, called AIB1, and an anti-conjugated L-DOPA Ab. They were biweekly injected into animals with small clinically palpable tumors. Anti-'phosphatidylinositol-like' autoantibody (autoAb) levels which were significantly higher in B(a)P-treated rat sera were decreased after Ab treatment. Tumor growth was slowed down compared with that of controls and animal survival was increased. This treatment was more efficient than that involving AIB1 alone. The anti-conjugated L-DOPA Ab may play a role in neovascularization, which is known to be critical for tumor growth.

[Effects of monoclonal anti-idiotypic antibody, internal image of benzo(a)pyrene on sarcoma in rats]. / Effets d'un anticorps monoclonal anti-idiotypique, image interne du benzo(a)pyrène sur des sarcomes de rats.

The effects of a monoclonal anti-idiotypic antibody (Ab), internal image of conjugated benzo(a)pyrene [B(a)P], called AIB1, on B(a)P-induced malignant sarcomas in Sprague-Dawley (SD) female rats were evaluated at different times after B(a)P-administration. As previously described, circulating anti-"phosphatidylinositol (PtdIns)-like" autoantibody levels were found to be significantly higher in B(a)P-treated rat sera. They were decreased after AIB1 Ab-treatment. Furthermore, the tumor growth was slowed down compared with that of controls and animal survival was slightly increased. These results showed the relationships between metabolic pathways involving possible "B(a)P-like" endogenous ligand/cytosolic receptor(s) and the PtdIns.

A monoclonal anti-idiotypic antibody with an internal image of a "phosphatidylinositol-like" structure derived from anti-"phosphatidylinositol-like" IgG from the sera of patients with proliferative malignancies.

Using an enzyme-linked immunosorbent assay (ELISA) with phosphatidylinositol (PtdIns) coated on well-plates, high levels of anti-"PtdIns-like" autoantibodies (autoAbs) have been previously described in sera of cancer patients. These anti-"PtdIns-like" autoAbs were purified and injected into BALB/c mice. A monoclonal anti-idiotypic antibody (Ab2), internal image of human endogenous "PtdIns-like" structure called AIPI, was selected. Then the immunological binding in sera of cancer patients was evaluated on AIPI coated on well-plates. Using this indirect ELISA method, we found a statistically highly significant immunological binding in sera of patients with epithelial tumors, which correlated with that previously found with the PtdIns molecule coated on well-plates. Moreover, AIPI mimics an endogenous structure closely associated with PtdIns specifically encountered in epithelial proliferative disease.

Curative effects of all-trans-retinoic acid on rat sarcomas.

The effects of all-trans-retinoic acid (all-trans-RA) on benzo(a)pyrene [B(a)P]-induced malignant sarcomas in Sprague-Dawley female rats were evaluated. Ninety-eight days after B(a)P administration, all-trans-RA was daily injected to animals with or without clinically palpable tumors. The growth of tumors was slowed down compared with controls. Magnetic resonance imaging analyses showed that all-trans-RA-treated rat tumors presented early necrotic areas. Animal survival was slightly increased. Anti-phosphatidylinositol autoantibody levels which were significantly higher in B(a)P-treated rat sera were not modified by all-trans-RA treatment.

Anti-phosphoinositide auto-antibodies in sera of cancer patients: isotypic and immunochemical characterization.

High levels of circulating anti-phosphatidylinositol (PtdIns) auto-antibodies (auto-Ab) have been previously found in sera of patients with malignant tumors. Using our ELISA test, a high statistical level of immunological binding directed against PtdIns was found in a large series of sera from patients with proliferative pathologies. In contrast to tumor markers, anti-PtdIns auto-Ab did not vary whatever the histological grades, TMN classification and patient's age. These anti-PtdIns auto-Ab were immunoglobulins (Ig) of G class. Their specificity was evaluated by competition experiments in ELISA and found to be rather high. An increase of these circulating auto-Ab reflected possible disturbances in PtdIns turnover and appearance of endogenous neo-antigen with PtdIns structure.

Identification and characterization of a specific autoantiphosphatidylinositol immune response during the time course of benzo(a)pyrene-induced malignant tumors in female Sprague-Dawley rats.

High levels of anti-phosphatidylinositol (PtdIns) autoantibodies (autoAb) have been previously described in sera of cancer patients and in plasma of dimethylbenzanthracene-treated female Sprague-Dawley rats. The presence of anti-PtdIns autoAb was tested in a model of highly malignant sarcomas induced by a single dose of benzo(a)pyrene [B(a)P] diluted in sesame oil and injected in female Sprague-Dawley rats. Significantly elevated levels of anti-PtdIns autoAb were found in sera of B(a)P-treated rats 40 days after B(a)P administration, whereas no significant levels of anti-PtdIns autoAb were noted in oil- or benzo(e)pyrene-treated rats. After day 60, autoantibody levels plateaued in B(a)P-treated rats, and highly malignant sarcomas appeared with 100% efficiency around day 100. Anti-PtdIns autoAb avidity and specificity were found to be high.

Autoanti-phosphatidylinositide antibodies specifically inhibit noradrenaline effects on Ca2+ and Cl- channels in rat portal vein myocytes.

High levels of circulating autoantibodies (auto-Ab) directed against phosphatidylinositides have been identified in the sera of patients with malignant tumors. These polyclonal autoantibodies had higher avidity and specificity for phosphatidylinositol (PtdIns) than for the other phosphatidylinositides. Effects of the auto-Ab were studied in smooth muscle myocytes in the PtdIns-involving transduction mechanism triggered by activation of alpha 1-adrenoceptors. Noradrenaline activated a Ca(2+)-dependent Cl- current through the Ca(2+)-releasing action of inositol 1,4,5-trisphosphate (InsP3) and enhanced the Ca2+ channel current through a diacylglycerol and protein kinase C-dependent mechanism. External applications of auto-Ab (0.03-0.3 mg/ml) were without effect on noradrenaline-induced responses whereas intracellular applications (0.0004-0.012 mg/ml) inhibited both Cl- current activation and Ca2+ channel current stimulation. Intracellular applications of IgG from healthy donors had no effect on noradrenaline-induced responses. When anti-PtdIns Ab were preincubated with PtdIns the inhibition of the noradrenaline-induced responses on Ca2+ and Cl- channels was not observed. Autoanti-PtdIns Ab inhibited also the acetylcholine-activated Cl- current, confirming that the acetylcholine response was mediated through the phosphatidylinositol breakdown. In contrast, the autoanti-PtdIns Ab were ineffective against the transduction pathway after beta-adrenoceptor activation. Therefore, these results suggest that the biological effect of autoanti-PtdIns Ab results from a specific binding to membrane PtdIns or PtdIns metabolites and thereby prevented InsP3 and diacylglycerol production. These autoanti-PtdIns Ab appear to be a new specific tool to identify the role of phosphatidylinositides in intracellular transduction processes.

Identification and immunochemical characterization of IgA in sera of patients with mammary tumors.

It has been reported that several types of human cancer are associated with elevated levels of class-A immunoglobulins (Ig) and IgA-containing immune complexes. Moreover, most previous work has come up against the lack of IgA reactivity for chemically defined antigen (Ag). To overcome this, we first evaluated possible immunological binding in sera of patients with mammary tumors or malignant hematologic diseases and controls on a mouse monoclonal anti-idiotypic antibody (Ab2), internal image of conjugated benzo(a)pyrene (BP) coated on well plates. Using this indirect ELISA, a statistically highly significant immunological binding was found in sera of patients with mammary tumors of every grade, type and size. This immunological binding was linked to the IgA isotype. Second, we performed competition experiments between a BP conjugate coated on well-plates and anti-anti-"BP-like" antibodies (Ab) previously incubated with rabbit idiotypic antibodies (Ab1) directed against conjugated BP. A part of these anti-anti-BP-like Ab, raised in rabbits immunized with human IgA of patients with mammary tumors, recognized the Ag-combining site of polyclonal Ab1, previously developed in a rabbit immunized with BP conjugates. It appears that part of the human Ig from sera of patients with mammary tumors shares common idiotopes with rabbit polyclonal Ab1 raised against conjugated BP.

Visualization of a protein involved in seric immune complexes of patients with epithelial tumors having elevated levels of anti-'benzo[a]pyrene-like' IgA.

Many authors have previously described that several types of human cancer were associated with elevated levels of IgA and IgA-containing immune complexes. We have recently identified and characterized IgA directed against a "benzo[a]pyrene-like' antigen in sera of patients with mammary tumors. Circulating immune complexes from these sera were precipitated and analysed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. A 18-20-kDa protein was found closely associated with high levels of anti-'benzo[a]pyrene-like' IgA in sera of patients having a proliferative pathology.

Increase of auto anti-phosphatidylinositol antibodies in plasma of female rats during the appearance of DMBA-induced malignant mammary tumors.

Using a lipid-adapted ELISA, antiphosphatidylinositol (PtdIns) antibodies (Ab) have been found in sera of cancer patients. With the same procedure, we have checked their possible raising in plasma of female rats during the appearance of 7,12-dimethylbenz[a]anthracene (DMBA)-induced mammary tumors. Twenty days after DMBA administration, the mean anti-PtdIns Ab level was already higher than that of control rats (oil). Immunochemical analysis of the anti-PtdIns Ab site showed a rather high Ab avidity (8 x 10(-9) M, at half-displacement) and high specificity for glyceryl phosphate inositol residues. Other phospholipids (PL) were not recognized by the anti-PtdIns Ab induced by malignant cell transformation.

Autoantibodies directed against lipid membrane components in sera of patients with malignant tumors.

Tumor-associated antigens (Ag) are expressed on neoplastic cells. Using an enzyme-linked immunosorbent assay (ELISA), an attempt was made to evaluate the autoimmune responses directed against lipid membrane components. A comparison was made of autoantibody (autoAb) levels in human sera of 684 patients with malignant tumors and those of 185 controls (healthy subjects and patients suffering from other diseases). A highly significant difference was found between the immunological binding of the groups for only one phospholipids (PL), i.e., phosphatidylinositol (PI). Using ELISA tests and PI-related compounds differing in fatty acid residue types and/or in phosphatidyl group, it was demonstrated that the hydrophilic residue is the immunodominant part recognized by the autoAb detected in sera of cancer patients.

[Autoantibodies directed against membrane phospholipid in serum in patients with malignant tumors]. / Autoanticorps dirigés contre un phospholipide membranaire dans les sérums de malades porteurs de tumeurs malignes.

Modifications of membrane lipids in the levels of fatty acids and phospholipids are associated with malignant tumors. In order to evaluate if these changes are recognized by the immune system, we have attempted to assay the possible presence of autoantibodies directed against the following lipids: phosphatidylinositol (PI), phosphatidylserine, phosphatidylcholine, phosphatidylethanolamine, gangliosides, galactocerebrosides, sphingomyelin, sphingosin, and cardiolipin, in the sera of patients with malignant tumors (n = 324) and from controls [healthy subjects (n = 20) and patients suffering of other diseases (n = 60)]. Using an adaptated immunoenzymatic assay (ELISA method), a highly significant difference (p less than 0.001) was found between the mean absorbances read on the cancer and control group for only one lipid, the PI. Whatsmore, these auto anti-PI were found in all sera (diluted 15,000 times) of patients with malignant tumors, whatever type, grade, or organ localization defined. These data indicated that the immune system recognized the PI antigenic modifications which appear to be linked to the cell transformation. This PI-immunological binding may have a predictive value as we have recently noticed in animals bearing chemically-induced malignant tumors.