IMMUNONEUTRALIZATION OF HETERODIMERIC FSH USING FSH BETA-SUBUNIT AS THE IMMUNOGEN

PROBLEM: Immunization with beta subunit of gonadotropin may elicit antibody formation to endogenous heterodimeric gonadotropin and result in reproductive unresponsiveness. The objectives of this study were to determine if antibodies produced in rats following immunization with human follicle stimulating hormone beta-subunit (hFSH-beta) could bind to and immunoneutralize heterodimeric FSH. and to elucidate the immunoneutralizing epitope. METHOD: Mature female Sprague-Dawley rats received subcutaneous injections of 10 mug of hFSH-beta emulsified in complete Freund's adjuvant, while control animals received only adjuvant. Animals received 10 mug hFSH-beta booster injections emulsified in incomplete Freund's adjuvant 2, 4, 11, and 21.5 wk after the initial immunization. RESULTS: Immunization with hFSH-beta produced appreciable antibody titers against human FSH (hFSH) as measured in enzyme-linked immunosorbant assays (ELISA) of immunized rat sera. A more modest titer to rat (rFSH) and no antibody response to rat Luteinizing Hormone (rLH) was observed, thus confirming the specificity of the immune response. Titers against hFSH increased throughout the study. Rat anti-hFSH-beta sera was tested to determine its ability to inhibit binding (immunoneutralizing) of I-125-hFSH to the FSH receptor. Continued immunization resulted in all animals producing immunoneutralizing antibodies. Immunization of rats also resulted in disrupted estrous cycles, but only animals with subsequent titers high enough to completely block binding of FSH to its receptor failed to conceive. In order to assess the immunoneutralizing epitope, antisera were tested in a peptide ELISA. Peptides used in the ELISA corresponded to amino acids that spanned the entire hFSHbeta sequence. It was found that antibodies from all rats immunized with hFSHbeta bound to amino acids within hFSH-beta 33-53. CONCLUSIONS: Collectively, these data suggest that amino acids within hFSH-beta 33-53 are necessary but not sufficient to confer immunocontraception. Amino acids within this linear sequence appear in a variety of epitopes of hFSH-beta and hFSH, only some of which are immunoneutralizing.