sermorelin and Colonic-Neoplasms

Treatment of colon cancer with an antagonist of growth hormone-releasing hormone (GHRH), JMR-132, results in a cell cycle arrest in S-phase of the tumor cells. Thus, we investigated the effect of JMR-132 in combination with S-phase-specific cytotoxic agents, 5-FU, irinotecan and cisplatin on the in vitro and in vivo growth of HT-29, HCT-116 and HCT-15 human colon cancer cell lines. In vitro, every compound inhibited proliferation of HCT-116 cells in a dose-dependent manner. Treatment with JMR-132 (5 μM) combined with 5-FU (1.25 μM), irinotecan (1.25 μM) or cisplatin (1.25 μM) resulted in an additive growth inhibition of HCT-116 cells in vitro as shown by MTS assay. Cell cycle analyses revealed that treatment of HCT-116 cells with JMR-132 was accompanied by a cell cycle arrest in S-phase. Combination treatment using JMR-132 plus a cytotoxic drug led to a significant increase of the sub-G 1 fraction, suggesting apoptosis. In vivo, daily treatment with GHRH antagonist JMR-132 decreased the tumor volume by 40-55% (p < 0.001) of HT-29, HCT-116 and HCT-15 tumors xenografted into athymic nude mice. Combined treatment with JMR-132 plus chemotherapeutic agents 5-FU, irinotecan or cisplatin resulted in an additive tumor growth suppression of HT-29, HCT-116 and HCT-15 xenografts to 56-85%. Our observations indicate that JMR-132 enhances the antiproliferative effect of S-phase-specific cytotoxic drugs by causing accumulation of tumor cells in S-phase.

Topics: Animals; Antineoplastic Agents; Camptothecin; Cell Line, Tumor; Cell Proliferation; Cisplatin; Colonic Neoplasms; Fluorouracil; Growth Hormone-Releasing Hormone; HCT116 Cells; HT29 Cells; Humans; Irinotecan; Mice; Mice, Nude; S Phase Cell Cycle Checkpoints; Sermorelin; Transplantation, Heterologous

We investigated the mechanisms of inhibitory effect of growth hormone-releasing hormone (GHRH) antagonist JMR-132 on the growth of HT29, HCT-116 and HCT-15 human colon cancer cells in vitro and in vivo. High-affinity binding sites for GHRH and mRNA for GHRH and splice variant-1 (SV1) of the GHRH receptor were found in all three cell lines tested. Proliferation of HT-29, HCT-116 and HCT-15 cells was significantly inhibited in vitro by JMR-132. Time course studies revealed that the treatment of human HCT-116 colon cancer cells with 10 muM GHRH antagonist JMR-132 causes a significant DNA damage as shown by an increase in olive tail moment (OTM) and loss of inner mitochondrial membrane potential (Delta Psi m). Western blotting demonstrated a time-dependent increase in protein levels of phospho-p53 (Ser46), Bax, cleaved caspase-9, -3, cleavage of poly(ADP-ribose)polymerase (PARP) and a decrease in Bcl-2 levels. An augmentation in cell cycle checkpoint protein p21(Waf1/Cip1) was accompanied by a cell cycle arrest in S-phase. DNA fragmentation visualized by the comet assay and the number of apoptotic cells increased time dependently as determined by flow cytometric annexinV and PI staining assays. In vivo, JMR-132 decreased the volume of HT-29, HCT-116 and HCT-15 tumors xenografted into athymic mice up to 75% (p < 0.05) and extended tumor doubling time (p < 0.001). Our observations suggest that GHRH antagonist JMR-132 exerts its antiproliferative effect on experimental colon cancer cells through p21(Waf1/Cip1) mediated S-phase arrest along with apoptosis involving the intrinsic pathway.

Topics: Animals; Apoptosis; bcl-2-Associated X Protein; Caspase 3; Caspase 9; Cell Line, Tumor; Colonic Neoplasms; Cyclin-Dependent Kinase Inhibitor p21; DNA Damage; DNA Fragmentation; Growth Hormone-Releasing Hormone; HCT116 Cells; Humans; Male; Membrane Potential, Mitochondrial; Mice; Mice, Nude; Poly(ADP-ribose) Polymerases; Proto-Oncogene Proteins c-bcl-2; S Phase; Sermorelin; Transplantation, Heterologous; Tumor Suppressor Protein p53

Insulin-like growth factors (IGFs) I and II are implicated in progression of various tumours including colorectal carcinomas. To interfere with the production of IGFs, we treated male nude mice bearing xenografts of HT-29 human colon cancer with various potent growth hormone-releasing hormone (GH-RH) antagonists. Twice daily injections of antagonist MZ-4-71, 10 microg intraperitoneally or 5 microg subcutaneously (s.c.) resulted in a significant 43-45% inhibition of tumour growth. Longer acting GH-RH antagonists, MZ-5-156 and JV-1-36 given once daily at doses of 20 microg s.c. produced a 43-58% decrease in volume and weight of cancers. Histological analyses of HT-29 cancers demonstrated that both a decreased cell proliferation and an increased apoptosis contributed to tumour inhibition. GH-RH antagonists did not change serum IGF-I or IGF-II levels, but significantly decreased IGF-II concentration and reduced mRNA expression for IGF-II in tumours. In vitro studies showed that HT-29 cells produced and secreted IGF-II into the medium, and addition of MZ-5-156 dose-dependently decreased IGF-II production by about 40% as well as proliferation of HT-29 cells. Our studies demonstrate that GH-RH antagonists inhibit growth of HT-29 human colon cancers in vivo and in vitro. The effect of GH-RH antagonists may be mediated through a reduced production and secretion of IGF-II by cancer cells.

Topics: Animals; Cell Division; Colonic Neoplasms; Growth Hormone-Releasing Hormone; HT29 Cells; Humans; Insulin-Like Growth Factor I; Insulin-Like Growth Factor II; Male; Mice; Mice, Nude; Neoplasm Proteins; Nucleolus Organizer Region; RNA, Messenger; Sermorelin; Transplantation, Heterologous