Enhancement of antigonadotropin response to the β-subunit of ovine luteinizing hormone by carrier conjugation and combination with the β-subunit of human chorionic gonadotropin

Abstract

Previous studies have demonstrated the control of fertility in monkeys on immunization with the beta subunit of ovine luteinizing hormone (beta-oLH) along with Freund's complete adjuvant (CFA). An eventual birth control vaccine would demand immunization without CFA. Beta-oLH linked to tetanus toxoid (TT) adsorbed on alum-induced antibody response in bonnet monkeys with detoxified sodium phthalyl derivative of Salmonella enteritidis lipopolysaccharide added only in the first injection. The antibodies reacted with both LH and human chorionic gonadotropin (hCG). A higher immune response was obtained with the use of a conjugate of beta-oLH and beta-hCG linked to a common carrier (TT) or with a mixture of beta-hCG/TT and beta-oLH/TT. In both cases, the antibodies were of high affinity with Ka ranging from 5 X 10(9) to 6 X 10(10) M-1. The antibodies were devoid of reactivity with human thyroid-stimulating hormone and human follicle-stimulating hormone. Monkeys with titers above 120 ng/ml hCG binding capacity (greater than 60 ng/ml in most cases) were protected from becoming pregnant during repeated mating with males of proven fertility in ovulatory cycles.The potential for immunizing against gonadotropins without using Freund's complete adjuvant was explored in bonnet monkeys by using tetanus toxoid (TT) as carrier and Salmonella lipopolysaccharide (LPS) as adjuvant. Pure hCG beta subunit and or sheep LH beta subunit was coupled with TT by employing N-succinimidyl-pyridyl-dithio-propionate reagent. Fertile female bonnet monkeys were injected with 50 mcg gonadotropin equivalent monthly. 1 mg sodium phthalyl derivative of LPS was added to the 1st injection. Animals with low titers were also given a booster on Day 145 with Leiras adjuvant. 3 of 5 monkeys immunized with ovine beta-LH subunit bonded to TT had strong responses, and 2 produced high antibody titers after a booster with Leiras adjuvant. A 2nd group of 3 monkeys treated with both ovine beta LH and beta hCG conjugated to a common carrier, TT, showed high titers, between 750 and 1300 ng/ml, which were sustained for nearly a year. Scathard analysis indicated that the combined antigens raised antibodies of high affinity, with Ka values ranging from 5 x 109 to 6 x 1010 per M. There were no cross reactions with either human FSH or TSH. 2 of the monkeys immunized against the combined antigens remained infertile for 6 and 3 cycles respectively, or until their antibody titers fell to 35 and 5 Monkeys in the 1st group also were infertile for several cycles before their antibody levels fell below 120 ng/ml against hCG.