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02612 PRODUCTION OF A RECOMBINANT HUMAN ANTI-SPERM ANTIBODY, RASA: IMPLICATIONS AS A SPERMICIDAL AGENT Fralix, Kimberly Nonoxynol-9 (N-9), the most widely used spermicide, has recently been shown to offer no protection against HIV/STI transmission and, in fact, increase the risk of infection when used frequently. As an alternative to N-9, a mouse monoclonal antibody, S19, which agglutinates sperm and exhibits signifi-cant contraceptive effects in laboratory tests, has been identified. A second generation of S19 was made as a single chain variable region antibody (ScFv) called RASA (Recombinant Anti-Sperm Antibody). To develop RASA into a spermicidal candidate, two objectives must be achieved: 1) establish a large-scale expression system; and 2) humanize RASA to prevent human anti-mouse antibody (HAMA) responses. Various expression systems for large-scale production were tested, including E. coli (bacteria), P. pastoris (yeast), N. tabacum (tobacco), and a cell-free system (Roche). Both yeast and tobacco systems produced reasonable yields of soluble protein and proteins were easily extractable. Indirect immunoflourescence staining showed that RASA expressed in tobacco bound to human sperm in a manner similar to S19. Additionally, RASA was humanized by site-directed mutagenesis of amino acids in the variable region framework to resemble those found in human IgG using overlap extension PCR. Humanized RASA will be expressed in yeast/tobacco systems and evaluated for sperm agglutinat-ing abilities. Conceivably, RASA can replace spermicides like N-9 to reduce the incidence of HIV/STI transmission to women who are already at high risk and be combined with microbicidal agents provid-ing dual protection to women wishing to protect themselves against disease and unplanned pregnancy. Dr. Kimberly D. Fralix |
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