Induction of therapeutic T-cell immunity by tumor targeting with soluble recombinant B7-immunoglobulin costimulatory molecules.

Tumor targeting with immunomodulatory molecules is an attractive strategy to enhance the host's antitumor response. Expression of CD80 (B7-1) and CD86 (B7-2) costimulatory molecules in tumor cells has proven to be an efficient way to enhance their immunogenicity. Here, we studied the effects of tumor targeting with biotinylated recombinant soluble B7-1- and B7-2 immunoglobulin G molecules (bio-B7-IgG) using a pretargeting approach based on the sequential use of a biotinylated antitumor monoclonal antibody and avidin. Mouse RMA T-lymphoma cells bearing either bio-B7-1-IgG or bio-B7-2-IgG on their surface prime in vitro naive CD8+ CTLs, which are highly effective in adoptive immunotherapy, and induce therapeutic immunity when injected in tumor-bearing animals. In vivo targeting of established RMA tumors with bio-B7-IgG either cures tumor-bearing mice or significantly prolongs their survival. The antitumor response induced by targeted bio-B7-IgG depends on both CD4+ and CD8+ T cells. Moreover, tumor targeting with bio-B7-IgG in vivo is critical for both expansion in lymphoid organs and mobilization into the tumor of tumor-specific CD8+ CTLs. When targeting is performed on poorly immunogenic TS/A mammary adenocarcinoma, only bio-B7-1-IgG primes naive CTLs in vitro and cures or significantly prolongs the survival of tumor-bearing mice in vivo, confirming that the two costimulatory molecules are not redundant with this tumor. Altogether, these data suggest that tumor avidination and targeting with soluble bio-B7-IgG may represent a promising strategy to enhance the antitumor response in the host.

Production of mouse monoclonal antibodies directed against the oolemma of human and hamster oocytes by intra-splenic injection of oocytes.

PROBLEM: To develop an additional approach for the study of oolemmal surface moieties involved in gamete interactions, we decided to obtain monoclonal antibodies by intrasplenic injection of human and hamster oocytes in Balb/c mice. METHOD: Two Balb/c males were injected three times intrasplenically at 15-day intervalS with approximately 40 zona-free hamster and 3-5 zona-free human oocytes. After the third injection, spleen cells were fused and hybridomas developed. We used a novel screening system based upon the use of sections of frozen human and hamster eggs, tested by means of indirect immunofluorescence. The antibodies that we produced were evaluated for their ability to interfere with the zona-free hamster eggs penetration by human spermatozoa. The B2B5 antibody was also developed as ascitic fluid and further characterized. RESULTS: Seven antibodies reactive with hamster oocytes were produced. Six of them also reacted with human oolemmas. The binding was confined to the oolemma, and no staining of the zona nor the cytoplasm was present. One of these antibodies reduced the penetration of zona-free hamster eggs by human spermatozoa. This antibody, B2B5, an IgM kappa, was confirmed to interact with the oolemma by means of indirect immunofluorescence of fresh eggs and Covasphere binding. B2B5 did not react with other human or hamster tissues except capacitated human spermatozoa. The reactivity with the oolemma of hamster oocytes was not lost after egg penetration by human sperm. CONCLUSIONS: Intrasplenic immunization using zona-free human and hamster oocytes allows the production of anti-oolemma antibodies. A system of screening based upon the use of sections of frozen eggs also allows an easy and quick scoring of many supernatants. B2B5 monoclonal anti-oolemma antibody deserves further studies in that is able to interfere with fertilization and its antigen appears to be confined to the gametes surface.