Protective effects of D-Trp6-luteinising hormone-releasing hormone microcapsules against cyclophosphamide-induced gonadotoxicity in female rats.

The possible protective effect of an agonist of luteinising hormone-releasing hormone (LH-RH) against the ovarian damage caused by cyclophosphamide was investigated in rats. D-Trp6-LH-RH microcapsules were injected once a month for 3 months, in a dose calculated to release 25 micrograms day-1. Control animals received the injection vehicle. Sixty days after the first injection of microcapsules, cyclophosphamide was given at a loading dose of 50 mg kg-1 followed by 5 mg kg-1 day-1 for 30 days, while the treatment with D-Trp6-LH-RH was continued. When the ovaries were examined 3 months and 5 months after discontinuation of treatment, a significant reduction in the total number of follicles (P less than 0.01) was found in non-pretreated animals given cyclophosphamide. This reduction affected mainly follicles larger than 100 microns. An irreversible disintegration and destruction of granulosa cells was also observed in this group. In animals pretreated with D-Trp6-LH-RH, administration of cyclophosphamide caused no reduction in the number and diameter of follicles. Thus, the treatment with D-Trp6-LH-RH microcapsules before and during chemotherapy prevented the ovarian injury inflicted by cyclophosphamide. The suppression of gonadal function by LH-RH analogues could be possibly utilised for the protection of the ovaries against damage caused by cytotoxic drugs

Inhibition of growth of OV-1063 human epithelial ovarian cancers and c- jun and c- fos oncogene expression by bombesin antagonists

Receptors for bombesin are present on human ovarian cancers and bombesin-like peptides could function as growth factors in this carcinoma. Therefore, we investigated the effects of bombesin/gastrin-releasing peptide (GRP) antagonists RC-3940-II and RC-3095 on the growth of human ovarian carcinoma cell line OV-1063, xenografted into nude mice. Treatment with RC-3940-II at doses of 10 μg and 20 μg per day s.c. decreased tumour volume by 60.9% (P< 0.05) and 73.5% (P< 0.05) respectively, after 25 days, compared to controls. RC-3095 at a dose of 20 μg per day reduced the volume of OV-1063 tumours by 47.7% (P = 0.15). In comparison, luteinizing hormone-releasing hormone (LH-RH) antagonist Cetrorelix at a dose of 100 μg per day caused a 64.2% inhibition (P< 0.05). RT-PCR analysis showed that OV-1063 tumours expressed mRNA for bombesin receptor subtypes BRS-1, BRS-2, and BRS-3. In OV-1063 cells cultured in vitro, GRP(14–27) induced the expression of mRNA for c- jun and c- fos oncogenes in a time-dependent manner. Antagonist RC-3940-II inhibited the stimulatory effect of GRP(14–27) on c- jun and c- fos in vitro. In vivo, the levels of c- jun and c- fos mRNA in OV-1063 tumours were decreased by 43% (P< 0.05) and 45% (P = 0.05) respectively, after treatment with RC-3940-II at 20 μg per day. Exposure of OV-1063, UCI-107 and ES-2 ovarian carcinoma cells to RC-3940-II at 1 μM concentration for 24 h in vitro, extended the latency period for the development of palpable tumours in nude mice. Our results indicate that antagonists of bombesin/GRP inhibit the growth of OV-1063 ovarian cancers by mechanisms that probably involve the downregulation of c- jun and c- fos proto-oncogenes. © 2000 Cancer Research Campaig

Targeted cytotoxic analogue of bombesin/ gastrin-releasing peptide inhibits the growth of H-69 human small-cell lung carcinoma in nude mice

Recently, we developed a powerful cytotoxic analogue of bombesin AN-215, in which the bombesin-like carrier peptide Gln–Trp–Ala–Val–Gly–His–Leu–Ψ(CH2-NH)–Leu–NH2 (RC-3094) is conjugated to a potent derivative of doxorubicin, 2-pyrrolinodoxorubicin (AN-201). Small-cell lung carcinomas (SCLCs) are known to express high levels of bombesin receptors. We evaluated whether these receptors could be used for targeting cytotoxic bombesin analogue to H-69 SCLC cells. H-69 cells were xenografted into male nude mice, which then received an intravenous injection of AN-215, cytotoxic radical AN-201, the carrier peptide RC-3094 alone or unconjugated mixture of RC-3094 and AN-201. The levels of mRNA for bombesin receptor subtypes were evaluated by reverse transcription-polymerase chain reaction. In vitro, both the analogue AN-215 and the radical AN-201 showed strong antiproliferative effects on H-69 cells, AN-215 requiring more time to exert its action at 10–8M concentration than AN-201. In vivo, the growth of H-69 SCLC tumours was significantly inhibited by the treatment with 200 nmol kg–1 of AN-215, while equimolar doses of the cytotoxic radical AN-201 or the mixture of AN-201 and the carrier peptide were toxic and produced only a minor tumour inhibition as compared with control groups. mRNA for bombesin receptor subtypes 2 (BRS-2) and 3 (BRS-3) was detected in H-69 tumours. The mRNA levels for BRS-3, but not for BRS-2, were lower in the AN-215-treated tumours as compared with controls. Our results demonstrate that the cytotoxic bombesin analogue AN-215 could be considered for targeted therapy of tumours, such as SCLC, that express bombesin receptors. © 1999 Cancer Research Campaig

[Phe4]somatostatin: a potent, selective inhibitor of growth hormone release.

[Phe4]Somatostatin was twice as active as somatostatin (SS) in suppressing rat growth hormone release in vitro but had only weak activity toward inhibition of insulin and glucagon release in vivo. The ability of this analogue to inhibit growth hormone release more actively than SS was confirmed in vivo by two separately designed bioassays. Further structure/activity studies of position 4 were carried out with [Glu4]SS, [Thr4]SS, and des-Lys4-SS, all of which had negligible inhibiting activity in the pituitary and pancreas. In this context the strikingly selective activity of [Phe4]SS suggests a fundamental difference in the SS receptors of pituitary and pancreas and the normal side-chain basicity of position 4 appears to be more important for action in pancreas than in pituitary. [Phe4]SS has properties that may be useful in the development of agents for the treatment of acromegaly or other disorders associated with increased growth hormone levels

Synergistic effects of bombesin and epidermal growth factor on cancers.

Bombesin and gastrin-releasing peptide act as autocrine mitogens in various cancers. Bombesin antagonist RC-3095 inhibited growth in some cancers and slowed the progression of premalignant lesions, possibly by down-regulating epidermal growth factor (EGF) receptors. Since the EGF receptor mitogen response involves tyrosine kinase stimulation, we tested the hypotheses that bombesin stimulates, and RC-3095 inhibits, phosphorylation; EGF and bombesin promote the phosphorylation of the same substrates; and EGF and bombesin act synergistically on phosphorylation. Therefore, in vitro assays for phosphorylation were performed in the presence or absence of EGF, bombesin, RC-3095, and combinations in samples derived from tumor, tissue surrounding tumor, cell lines, and normal and transforming tissue derived from the 9,10-dimethyl-1,2-benzanthracene-induced squamous cell lesions of the hamster cheek pouch. Bombesin increased, and RC-3095 decreased, phosphorylation in these samples. In the human hepatoma sample and surrounding tissue, these ligands altered the phosphorylation of the same substrates affected by EGF. EGF and bombesin stimulated phosphorylation synergistically in the hamster samples and the hepatoma. Bombesin-induced phosphorylation was greater in tissue surrounding the hepatoma, whereas RC-3095 was more effective in inhibiting phosphorylation in the hepatoma itself. This cancer, therefore, could be endogenously stimulated by gastrin-releasing peptide. These observations support the hypothesis that bombesin stimulates growth of tissues and tumors by amplifying the phosphorylation response to EGF. The growth inhibitory response to RC-3095, or other bombesin analogues, of individual tumors may be prognosed by in vitro phosphorylation assays using the samples from the patient's tumor