hCG beta residues 94-96 alter LH activity without appearing to make key receptor contacts.

The ability of human chorionic gonadotropin (hCG) to distinguish lutropin (LHR) and follitropin (FSHR) receptors is controlled principally by beta-subunit residues 94-117. To learn how residues 94-96 (Arg-Arg-Ser) influence LHR binding, we studied the effects of replacing them on the LH and FSH activities of a bifunctional hCG analog in which residues 101-109 were derived from FSH. Analogs containing 1-3 arginines and no aspartates at residues 94-96 bound LHR with 25-400% the potency of hCG. When residues 94-96 were neutral or contained 1-3 aspartates, LHR binding was reduced 6-100 fold but remained at least ten-fold greater than the negative control analog containing residues 94-117 derived from FSH. Residues 94-96 had little influence on FSHR binding. These observations support a model [Moyle et al. (1995) J. Biol. Chem. 270:20,020] in which residues 94-96 influence LHR binding specificity primarily through an effect on hormone conformation rather than by direct participation in essential high affinity receptor contacts.

Membrane disrupting lytic peptide conjugates destroy hormone dependent and independent breast cancer cells in vitro and in vivo.

We have prepared conjugates of a membrane disrupting lytic peptide (hecate) and a 15-amino acid segment of the beta-chain of CG and hecate and the decapeptide, luteinizing hormone releasing hormone (LHRH). We have tested the concept that these conjugates will target breast cancer cells expressing LH/CG or LHRH receptors. In previous studies, we were able to destroy prostate cancers in vitro and in vivo with lytic peptide conjugates. Hecate, hecate-betaCG and LHRH-hecate were added to cultures of the human breast cancer cell lines MCF-7 and MDA-MB-435S. Hecate and its conjugates showed concentration dependent toxicity to both cell lines. The lytic peptide alone showed similar EC50 values for both cell lines; however, there was a significant difference between the EC50 values when the conjugates were tested. The hormone dependent MCF-7 cell line was less sensitive to the betaCG conjugate than to the LHRH conjugate; the reverse was found for the hormone independent MDA-MB-435S cells. Removal of steroids decreased the sensitivity of MCF-7 cells to both lytic peptide conjugates and this sensitivity could be restored by adding estradiol. Activation of protein kinase C further increased the sensitivity to the drug. MDA-MB-435S xenografts were established in intact female athymic nude mice, which were treated once a week for 3 weeks with hecate-betaCG via the lateral tail vein. The ability of hecate-betaCG to destroy xenografts of human breast cancer cells (MDA-MB-435S) in nude mice was demonstrated for the first time. We conclude that hecate-betaCG and LHRH-hecate conjugates could serve as useful drugs for the treatment of breast cancer.