EVALUATION OF THE IN-VITRO AND IN-VIVO ACTIVITY OF THE L-CIT(6), D,L-CIT(6) AND D-CIT(6) FORMS OF THE LH-RH-ANTAGONIST CETRORELIX (SB-75)

The objective of this study was to examine the in vivo and in vitro go nadotropin-inhibiting potencies, edematogenic activities and the recep tor binding affinities of the D-Cit(6), D,L-Cit(6) and L-Cit(6) forms of the LH-RH antagonist Cetrorelix (SB-75) -Phe(4Cl)(2),D-Pal(3)(3),D- Cit(6),D-Ala(10)]LH-RH. In order to demonstrate the suppressive effect s of two different diastereomers of SB-75 and their racemic mixture on LH and FSH release, [D-Cit(6)] SB-75 was injected subcutaneously in d oses of 2.5 and 10 mu g/rat, [D,L-Cit(6)]-SB-75 in doses of 5 and 20 m u g/rat and [L-Cit(6)] SB-75 in doses of 12.5 and 50 mu g/rat to castr ated male rats. Two hours after administration, there was no differenc e in LH levels between rats injected with the L-form and control anima ls, indicating a low activity and/or a rapid enzymatic degradation of this peptide. The (1:1) diastereomeric mixture was only about half as potent in suppression of LH release compared to [D-Cit(6)] SB-75. Seru m FSH levels were suppressed significantly (p<0.01) for more than 48 h after the administration of 10 mu g [D-Cit(6)] SB-75 and 20 mu g of [ D,L-Cit(6)] SB-75, respectively. [D-Cit(6)] SB-75 administered at a do se of 2 mu g/rat induced 100% inhibition of ovulation, while 4 mu g/ra t of the D,L-Cit(6) peptide were necessary to produce the same effect. [L-Cit(6)] SB-75 given at a high dose of 40 mu g/rat produced only 14 % inhibition of ovulation. The D-Cit(6) form of SB-75 produced skin le sions with a much smaller diameter than the L-isomer, and was about 34 times less edematogenic. [D-Cit(6)] SB-75 was bound more powerfully t o high-affinity pituitary LH-RH receptors than either D,L-Cit(6) or L- Cit(6) analogues. In vitro assays based on the superfusion of disperse d rat pituitary cells on a column, followed by radioimmunoassay for LH , also demonstrated a lower inhibitory activity for the L-Cit(6) analo gue, but the differences between D-, D,L- and L-citrulline analogues w ere smaller than in vivo. Our results indicate that the LH-RH antagoni st [D-Cit(6)] SB-75 is more effective in suppression of gonadotropin r elease in vivo and in vitro, less edematogenic and possesses higher bi nding affinity to pituitary LH-RH receptors than the D,L- and L-citrul line decapeptide analogues. (C) Munksgaard 1995.