ABSTRACT
A modified GnRH peptide (CHWSYGLRPG-NH(2)) was conjugated to tetanus toxoid (TT) or diphtheria toxoid (DT) and formulated with Quil A saponin or a sustained release injectible PLGA (poly(lactide-co-glycolide)/triacetin). For the Quil A formulations, two administrations of TT conjugate at 3-weekly intervals were followed by two booster injections with the DT conjugate in entire ram lambs. With the PLGA formulations, only two injections were administered; the first containing TT and the second DT at 6-weekly intervals. Evaluation was carried out by comparing the specific antibody levels produced in relationship to hormone profiles and testicular changes. The Quil A formulation was considered the most effective, as it caused significant reduction in testosterone and follicle stimulating hormone levels, resulting in marked suppression of spermatogenesis.
Subject(s)
Contraception, Immunologic/methods , Gonadotropin-Releasing Hormone/immunology , Sheep/physiology , Spermatogenesis , Vaccines, Contraceptive/administration & dosage , Adjuvants, Immunologic , Animals , Antibodies/blood , Diphtheria Toxoid/administration & dosage , Diphtheria Toxoid/immunology , Enzyme-Linked Immunosorbent Assay , Follicle Stimulating Hormone/blood , Gonadotropin-Releasing Hormone/chemistry , Histocytochemistry , Immunization Schedule , Lactic Acid , Male , Polyglycolic Acid , Polylactic Acid-Polyglycolic Acid Copolymer , Polymers , Quillaja Saponins , Saponins/immunology , Testicular Hormones/blood , Testis/cytology , Testosterone/blood , Tetanus Toxoid/administration & dosage , Tetanus Toxoid/immunology , Vaccines, Conjugate/chemistry , Vaccines, Conjugate/immunologyABSTRACT
PROBLEM: We previously immunoneutralized gonadotrophin releasing hormone (GnRH), using an analogue of GnRH (des-1 GnRH-I), conjugated to tetanus toxoid via a carbodiimide reaction. The castration effect on the reproductive system was not consistent in all the treated animals. Therefore, we examined the possibility that conjugation to the carrier protein via the N- or C-terminal could have an effect on efficacy. METHOD OF STUDY: GnRH analogue sequences were synthesized consisting of an additional cysteine at either terminal and specific conjugation was carried out using a bifunctional linker agent. RESULTS: Conjugation of the monomer through the N-terminal proved to be a highly effective means of causing immunocastration in terms of decreased gonadotrophin and testosterone concentrations and testicular size, whereas conjugation through the C-terminal proved to be ineffective. This was reflected in the ability of the antibodies to bind native GnRH, but not the levels of the anti-GnRH antibodies. CONCLUSION: Immunoneutralization efficacy was attributed to the importance of preserving the GnRH C-terminal.
