A ssociative recognition is a basic regulatory mechanism of the immune response and refers to the phenomenon in which immunity to one determinant of a multideterminant antigen enhances the response to other determinants. The cooperative effect is mediated by a determinant recognized by T helper cells (Th) and has been demonstrated for B cell responses (T-B cooperation) (1) and cytotoxic T lymphocytes responses (Th-cytotoxic T lymphocyte cooperation) (2). A similar form of associative recognition between two Th cell determinants (Th-Th cooperation) has not been described yet but could prove valuable to responses against poorly immunogenic antigens such as tumor antigens. The immune response against tumor antigens is hindered, among other factors, by down-regulation of MHC molecules, self-tolerance, and functional hierarchy in the immunogenicity of T cell determinants. It is now clear that T cell determinants of protein antigens can be categorized into dominant, subdominant, and cryptic to reflect a different degree of immunogenicity in vivo (3). Consequently, as T cells reactive with dominant determinants of tumor antigens are eliminated in the thymus during negative selection, the adult immune repertoire is composed mainly of precursor cells with moderate avidity for subdominant and cryptic determinants. Thus, methods to heighten the response of CD4 and CD8 T lymphocytes of the residual repertoire against poorly immunogenic determinants are needed to develop better immunogens against cancer.MUC-1, a glycosylated molecule of high molecular weight expressed in malignant tumors of epithelial origin (4, 5), induces both CD8 (6-8) and CD4 T lymphocytes (9, 10) against an antigenic core consisting of a tandem repeat of 20 amino acid residues. For these reasons, MUC-1 is a useful model antigen for targeted manipulations of the immune response. Specific T cell responses against MUC-1 were induced in C57BL͞6 mice using a model of epitope-based genetic vaccination, somatic transgene immunization (11). This approach is based on the inoculation into the spleen of adult mice of plasmid DNA comprising an Ig heavy (H) chain gene controlled by a B cell-specific promoter to target resident B cells (12). The use of Ig genes modified in the complementaritydetermining regions (CDR) to code for heterologous epitopes (13) allows one to direct the in vivo synthesis and secretion by B cells of transgenic Ig, which immunize the host against the B and T cell epitopes expressed in their V region (14,15). Unlike conventional DNA vaccination (16), which induces Th1 responses (17), somatic transgene immunization activates Th0 cells that produce IL-2, IFN-␥, and IL-4 (15). One advantage of somatic transgene immunization is that heterologous epitopes can be assembled on the Ig V region in various assortment and combination to maximize immunogenicity but also to provide easy tools to study the reciprocal regulation between distinct antigenic determinants during the immune response in vivo.Here, we report that an immunologically silent Th cell d...

Interferons (IFN) are biological molecules with antiviral, antiproliferative, and immunomodulatory actions. Plasmapheresis (PP) combined with IFN therapy in 24 multiple sclerosis (MS) patients was associated with a rapid increase in detectable IFN levels. We describe the presence of a detectable factor in the serum of MS patients which decreases the efficacy of IFN therapy in these patients. We call this factor "interferon inhibitor factor" (IIF). Standard IFN assay indicates inhibition of WISH cell protection by IFN owing to the presence of IIF in patient's serum. Similar results were also obtained with human fibroblast and human leukocyte IFNs. The best results were obtained with an IFN mixture; results with 1:20 diluted patient's sera showed elevation of 120% greater than 1:10 dilution. With 1:40 dilution, an elevation of 1,041% was noticed. The IIF from patient sera collected during PP was purified and characterized. Native gel electrophoresis of IIF indicates a single protein band; further analysis on SDS gels indicates two bands at the 200 and 21-kD range. ELISA failed to reveal the presence of any anti-IFN antibodies. This study demonstrates the presence of IIF in MS patients' sera which are removed from the circulation by PP. Removal of IIF from circulation was associated with increased IFN levels and clinical improvement as measured by Kurtzke's disability status scale (KDSS).

The rationale for the use of interferon (IFN) in the treatment of multiple sclerosis (MS) is based on its recognized antiviral and immunomodulating actions. The pathogenesis of MS is believed to be due to an immunologic response in a genetically predisposed individual, localized within the central nervous system white matter, and triggered by exposure to an environmental agent such as a virus. Based on our personal experience we find that the efficacy of IFN therapy is hampered in MS patients by the presence of an interferon inhibitor factor (IIF) in the patients' sera which we have isolated and characterized. When plasmapheresis (PP) was done on 24 MS patients with intermittent 3-day administration of IFN-alpha and human leukocyte IFN, marked increase of IFN in 18 patients and modest increase in three patients correlated with clinical improvement. Three clinical nonresponders showed no increase in IFN levels following therapy. The ability to remove IIF and lymphokine inhibitor factor (LIF) by PP may explain the successful treatment of our patients. We describe the evaluation of helper T cells, suppressor T cells, HLADR antigen, natural killer cells, and monocyte/macrophage cell populations by flow cytometry before and after PP. A significant increase in these immune-competent cells correlated with marked improvement in Kurtzke disability status scale in 13 patients, while eight stabilized. Patients showing progression of the disease either showed decrease or no change in these parameters after therapy. Encouraging results from this pilot study suggest that PP combined with immunomodulatory regimens of IFN may be an effective therapy for MS.

A human IgG1.k monoclonal antibody (MAb) designated GMA1 was developed by fusing pooled lymph node lymphocytes from cancer patients with the human lymphoblastoid cell line, SHFP-1. The GMA1 MAb reacted with several melanoma and neuroblastoma cell lines. Normal tissue derived from human brain and tumor-cell lines derived from colon, ovary, and breast were not reactive. FACS analysis performed using live cells demonstrated that the antibody recognizes a cell-surface antigen. Enzyme immunoassay (EIA) and thin layer chromatography (TLC) immunostaining with purified gangliosides indicated that the antibody has specificity for the major tumor associated gangliosides GD3, GM3, and GD2. GMA1 heavy and light chain genes were isolated by RT-PCR and a recombinant derivative of this human antibody was expressed in Chinese hamster ovary (CHO) cells. High-level antibody synthesis and secretion was achieved using a vector designed to maximize expression. FACS analysis and TLC immunostaining indicated recombinant GMA1 reacted with human tumor cell lines and gangliosides GD3, GM3, GD2 in a manner similar to the antibody produced by the hybridoma cell line, demonstrating that the specificity of the antibody was not altered during molecular cloning.

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