Antitumour effect of a gonadotropin-releasing-hormone antagonist (MI-1544) and its conjugate on human breast cancer cells and their xenografts

Abstract

Our gonadotropin-releasing hormone (GnRH) antagonist analogue MI-1544 ([Ac-D-Trp1,3,D-Cpa2,D-Lys6,D-Ala10]GnRH) was developed as a potential contraceptive material, because it decreased the luteinizing hormone level without unfavourable side-effects. The antagonist was covalently bound to poly[Lys-(Ac-Glu0.96-DL-Ala3.1)] (AcEAK)-a branched polypeptide having a polylysine backbone--resulting in a MI-1544-AcEAK conjugate. According to our in vitro experiments the MI-1544 induced a 33%-35% decrease in cell numbers of MCF-7 and MDA-MB-231 human breast cancer cell lines at a dose of 30 microM. The biodegradable polymeric carrier, AcEAK, alone inhibited cell proliferation by only 13%-15%, while the MI-1544-AcEAK conjugate, applied at the same dose, was capable of producing 45%-50% inhibition of cell proliferation. Our in vivo experiments using immunosuppressed mice showed that MI-1544, applied twice daily s.c., inhibited the growth of oestrogensensitive and -insensitive xenografts by 65% and 30% respectively. This effect was potentiated (70%) in both types of xenografts by the presence of the polymeric carrier in the conjugate; however, the carrier by itself did not cause tumour growth inhibition. The polymeric polypeptide carrier is supposed to increase the stability of the GnRH antagonist and to prevent the rapid excretion of the covalently bound peptide molecule. The antagonist and its conjugate may have various direct and indirect effects on breast cancer cells and, as a consequence, the new GnRH antagonist conjugates are suitable for treating an extended range of breast cancers.

Gonadotropin-releasing hormone analogue conjugates with strong selective antitumor activity

“…The experimental procedure for the in vivo antitumor experiments is described in ref. 17. Female CBA͞Ca HRIJ-T6 and BDF 1 mice weighing 19-22 g were immunosuppressed by thymectomy, whole-body irradiation, and syngeneic bone marrow transplantation.…”

“…Human MCF-7 and MDA-MB-231 breast, PC3 and LNCaP prostate, and Ishikawa endometrium tumor cell lines (American Type Culture Collection) with GnRH receptors (16,17) were maintained in Dulbecco's modified Eagle's minimal essential medium (DMEM; GIBCO͞BRL) supplemented with 10% fetal calf serum (GIBCO). PC3 cells were cultured in RPMI medium 1640 (GIBCO).…”

“…GnRH receptor genes were found in MDA-MB-231 (10), MCF-7 breast (11), LNCaP (12) and PC3 prostate, Ishikawa endometrial (13,14), and ovarian (15) cancer cell lines. These cell lines contain high-and low-affinity binding sites for GnRH (16,17). The high-affinity GnRH-binding sites are commonly regarded as the same as the GnRH receptor of the pituitary gland.…”

Drug targeting by macromolecules without recognition unit?

“…The solution conformation, in vitro cytotoxicity, biodistribution and antitumour properties were characterized. We found pronounced antitumour activity of polyanionic conjugate of a GnRH antagonist, Ac-[D-Trp 1-3 , D-Cpa 2 , D-Lys 6 , D-Ala 10 ]-GnRH-poly[Lys(Ac-Glu i -DL-Ala m )] (Ac-EAK; Mez} o o et al, 1996a,b;Vincze et al, 1994) and of a conjugate in which Dau is attached to an amphoteric polypeptide, poly[Lys(-Glu i -DL-Ala m )] (EAK; Gaál and . The marked effect of some polycationic derivatives of MTX was also described on L. donovani-infected mice as outlined below (Kóczán et al, 2002).…”

“…2). Conjugates of Dau (Hudecz et al, 1992;Gaál and Hudecz, 1998), MTX Kóczán et al, 2002), GnRH antagonist (Mez} o o et al, 1996a;Vincze et al, 1994), amiloride (Pató et al, 1999), boron compounds (Mez} o o et al, 1996b) and several radionuclides (Pimm et al, 1995;Perkins et al, 1998) have been synthesised. The solution conformation, in vitro cytotoxicity, biodistribution and antitumour properties were characterized.…”