Synthesis, Stability and Direct Antiproliferative Effect of New Cysteine Modified GnRH Analogs

It is demonstrated that gonadotropin-releasing hormone (GnRH) analogs can directly inhibit the proliferation of reproductive tissue cancer cells, but the poor pharmacokinetic properties still restrict their application in treating hormone-dependent diseases. Modifications in position 6 and 10 of natural GnRH can improve the metabolic stability. In order to study the effect of incorporation Cys 6 substitution with C-terminal Pro 9 -NHEt modification and dimerization of linear peptides on metabolic stability and antiproliferative activity of GnRH analogs, two new GnRH analogs [ l -Cys 6 , desGly 10 , Pro 9 -NHEt]-GnRH ( 1 ) and [ d -Cys 6 , desGly 10 , Pro 9 -NHEt]-GnRH ( 2 ), and their corresponding dimer derivatives ([ l -Cys 6 , desGly 10 , Pro 9 -NHEt]-GnRH) 2 ( 3 ) and ([ d -Cys 6 , desGly 10 , Pro 9 -NHEt]-GnRH) 2 ( 4 ) were synthesized. Incubation of these analogs with human serum was carried out to evaluate their metabolic stability, and direct growth inhibitory effect of the two dimer derivatives 3 and 4 on MCF-7 human breast cancer cell line was examined by MTT assay. The metabolic stability of dimer derivatives 3 and 4 was remarkably improved in comparison with natural GnRH. The d -Cys 6 substituted dimer derivative 4 exhibited higher inhibitory effect (29.6–39.7% growth reduction) on cell growth than its corresponding counterpart 3 (21.8–26.2% growth reduction) at concentration range of 50, 100 and 200 μΜ. The cell growth inhibition of leuprolide was 16.4–27.2% at the tested concentrations. The dimer derivative 4 was the most stable and active GnRH analog in this study and has the potential for future preclinical investigations as promising antitumor drug candidate.