Growth hormone responses to growth hormone-releasing hormone (1-29)-NH2 and a D-Ala2 analog in normal men

Synthetic analogs of growth hormone-releasing hormone, GHRH(1-29)-NH2 and D-Ala2 GHRH(1-29)-NH2 were administered as a bolus intravenous injection to five normal men in a dose range of 0.015 to 0.5 micrograms/kg body weight. Vehicle only was administered in a control study. Peak responses to GHRH analogs occurred at 15 or 30 min. An increase in the integrated plasma growth hormone (GH) response was observed at each dose. The dose-response curve of GHRH(1-29)-NH2 indicated that it has a similar molar potency to GHRH(1-40) and GHRH(1-44). The potency of D-Ala2 GHRH(1-29)-NH2 was approximately twice that of GHRH(1-29)-NH2. Neither analog affected blood levels of PRL, TSH, LH, FSH, ACTH, insulin, glucagon, glucose, cortisol, free thyroxine, and free triiodothyronine. No side effects were noted other than transient flushing with the highest dose administered. The findings demonstrate GHRH(1-29)-NH2 and its D-Ala2 analog are potent stimulators of GH release and have potential application in clinical medicine.     

The use of growth hormone-releasing hormone in the diagnosis and treatment of short stature

We have assessed the role of growth hormone-releasing hormone (GHRH) as a diagnostic test in 40 children and young adults with growth hormone deficiency (GHD), principally using the GHRH(1-29)NH2 analogue. Following 200 micrograms GHRH as an acute intravenous bolus, serum GH rose to normal or just subnormal levels in 13 out of 17 children with structural lesions, and in 8 of 14 patients with idiopathic GHD or panhypopituitarism. Of 9 children (mean age 12 years) with GHD following treatment with cranial irradiation for nonendocrine tumours, all responded acutely to GHRH. 12- and 24-hour infusions with GHRH(1-29)NH2, and 1- and 2-week treatments with twice-daily subcutaneous GHRH(1-29)NH2, showed persistent stimulation of GH release. It is concluded that many children with GHD of diverse aetiology will respond both acutely and chronically to treatment with GHRH.     

Effect of long-term GHRH and somatostatin administration on GH release and body weight in prepubertal female rats

In order to find a chronic GHRH administration capable of stimulating growth rate without depleting pituitary GH content, prepubertal female rats were subcutaneously (sc) treated with GHRH (1-29)-NH2 and somatostatin (SS). In experiment 1, the rats received sc injections of GHRH and cyclic natural SS for 19 days. In the second study, female rats were continuously treated during 21 days with GHRH, using a slow release pellet, alone or combined with one daily injection of long acting SS (octreotide). In experiment 1, body weight was significantly increased when GHRH was administered at the highest daily dosage (1200 microg/day), accompanied by an slight increment in pituitary GH content. Hypothalamic SS concentrations decreased when GHRH or SS were administered alone whereas the combined treatment with both peptides did not modify this parameter, which suggests the existence of a balance between the chronic actions of both peptides on hypothalamus. In experiment 2, the continuous infusion of GHRH increased plasma GH levels and tended to enhance pituitary GH content. Nevertheless, GHRH effect was not effective enough to increase body weight. By adding one daily injection of SS both GHRH effects on the pituitary gland were abolished. Our study indicates that female rats retain responsiveness to chronic GHRH and SS treatments at both pituitary and hypothalamic levels.     

Subcutaneous treatment with growth hormone-releasing hormone for short stature

In the present study we report the effects of therapy with growth hormone-releasing factor (1-29)NH2 (GRF) on growth rate, plasma levels of insulin growth factor I (IGF-I) and growth hormone (GH) secretion in 11 children who were selected solely on the basis of their short stature and normal GH secretion on standard provocative tests. All children received GRF for 6 months (5 micrograms/kg body weight subcutaneously) each evening. The 24-hour GH secretory profile was studied before and after 6 months of treatment. Simultaneously, GH secretory responses to single intravenous bolus GRF (1.5 micrograms/kg body weight) were also studied before, during, and 6 months off therapy with GRF(1-29)NH2. Plasma levels of IGF-I were measured before, during (1, 2 and 6 months), and after 6 months off therapy with GRF. Statural growth was measured at 3-month intervals. The peak plasma GH level in response to GRF was 56.04 +/- (SD) 24.46 ng/ml before treatment, and similar results were found after therapy. The 24-hour GH secretory profile did not show differences before, during, and after treatment. Comparably, no differences were found in GH pulse frequency, pulse amplitude, pulse height, pulse increment, pulse area and total area before, and 6 months off therapy with GRF. The increments in serum IGF-I achieved were not significantly different at all intervals studied. All patients increased growth velocities (mean +/- SD, cm/year) in response to GRF therapy. Our results demonstrate that GRF administration was effective in accelerating growth velocity in 11 children without GH deficiency.     

Bioactivity of growth hormone releasing hormone (1-29) analogues after SC injection in man

The bioactivity of growth hormone releasing hormone 1-29 [GHRH(1-29)NH2] has been compared with that of an agonist analogue [Ac-D-Tyr1,D-Ala2]-GHRH(1-29)NH2, in normal male volunteers. Using a submaximal dose of 3 micrograms/kg administered subcutaneously, peak growth hormone (GH) response and area under the GH curve were similar for the native and agonist analogue. In addition, no significant differences were found in peak GHRH(1-29) immunoreactivity, area under the GHRH(1-29) curves or plasma disappearance rates of the two peptides. The results suggest that, in keeping with the relative activities of other "superactive" analogues tested so far, the greatly enhanced activity of [Ac-D-Tyr1,D-Ala2]-GHRH(1-29)NH2 observed in the rat is not found in humans. It is possible that this species difference is due to differences in the interaction of GHRH peptides with the rat and the human somatotroph GHRH receptor.     

Response of growth hormone release to human growth hormone-releasing factor and its analogs in the bovine

Responses of growth hormone (GH) release to synthetic human growth hormone-releasing factor (hGRF)-44-NH2 analogs were determined, and the GH-releasing potency based on dose per kg of body weight (bw) was compared with that of hGRF-44-NH2 in female dairy calves. Four- and 12-month-old calves were injected intravenously with 0.25 microgram of hGRF-44-NH2 or its analogs per kg of bw. Blood samples were collected before, and during 180 min after each injection, and plasma GH concentrations were measured by radioimmunoassay. Areas under the GH response curves for 180 min after injection of hGRF-44-NH2 and its analogs were used as an index of the GH-releasing potency of each peptide. The GH-releasing potency of hGRF(1-26)-NH2 was significantly lower than that of hGRF-44-NH2 (P less than 0.05). On the other hand, hGRF(1-29)-NH2 possessed similar potency to hGRF-44-NH2. [D-Tyr1]-hGRF-44-NH2 showed prolonged GH-releasing activity, though its potency was similar to that of hGRF-44-NH2. Also, [D-Ala2]-hGRF(1-29)-NH2 exhibited prolonged GH-releasing activity, and its potency was 2.5 (P less than 0.05) and twice (P less than 0.05) as great as that of hGRF-44-NH2 and hGRF(1-29)-NH2, respectively. These results demonstrate that the N-terminal 29 amino acid residues of hGRF possess the activity site required for full GH release in vivo, and [D-Ala2]-hGRF(1-29)-NH2 has longer and greater activity, on a dose basis, than hGRF-44-NH2 in the calves.     

The influence of albumin on vitamin D metabolism in fetal chick osteoblast-like cells

Human pancreatic growth hormone releasing factor (1-29)-amide [hpGRF (1-29)-NH2] and the following analogs: [D-Tyr-1]-hpGRF(1-29)-NH2, [D-Ala-2]-hpGRF(1-29)-NH2, [D-Asp-3]-hpGRF(1-29)-NH2, and [N-Ac-Tyr-1]-hpGRF (1-29)-NH2 were synthesized using solid phase methodology and tested for their ability to stimulate growth hormone (GH) secretion in the rat and the pig in vivo. [D-Ala-2]-hpGRF (1-29)-NH2 was approximately 50 times more potent than the parent molecule in eliciting GH secretion in the rat. The other analogs were less active, but all were more potent than the 1-29 amide in the rat. [D-Tyr-1]-hpGRF(1-29)-NH2 was 10 times more potent, [D-Asp-3]-hpGRF(1-29)-NH2 7 times more potent, and the acetylated molecule approximately 12 times more potent than hpGRF(1-29)-NH2.     

Effect of substance P/bombesin antagonists on the release of growth hormone by GHRP and GHRH.

The substance P(SP)/bombesin (Bn) antagonists [DArg1DTrp7,9Leu11] SP(P-7482), [DArg1-DPro2DTrp7,9Leu11]SP (P-7483), [DArg1DPhe5DTrp7,9Leu11]SP(P-7492), and the growth hormone releasing hormone (GHRH) antagonist [DArg2Ala8,9,15]GHRH(1-29)(DC21-366) were tested for their in vitro effects on the release of growth hormone (GH) in the presence of GHRH and growth hormone releasing peptide, HisDTrpAlaTrpDPheLysNH2(GHRP). P-7492, P-7483, and P-7482 decreased, dose-dependently, the release of GH by GHRP (IC50 = 0.2 microM, 0.85 microM, and 6 microM, respectively). These antagonists had only a 10-15% inhibitory effect on the stimulated GH release of GHRH even at high dosage. DC21-366 decreased the stimulated release of GH by GHRH (IC50 = 0.16 microM) but not by GHRP. Neither SP nor Bn had GH releasing or inhibitory effects in this system.     

Hypothalamic-pituitary somatotropic function in prepubertal hypothyroid rats: effect of growth hormone replacement therapy.

The effect of thyroid hormone deficiency and growth hormone (GH) treatment on hypothalamic GH-releasing hormone (GHRH)/somatostatin (SS) concentrations, GHRH/SS mRNA levels, and plasma GH and somatomedin-C (IGF-I) concentrations were studied in 28- and 35-day-old rats made hypothyroid by giving dams propylthiouracil in the drinking water since the day of parturition. Hypothyroid rats, at both 28 and 35 days of life, had decreased hypothalamic GHRH content and increased GHRH mRNA levels, unaltered SS content and SS mRNA levels, and reduced plasma GH and IGF-I concentrations. Treatment of hypothyroid rats with GH for 14 days completely restored hypothalamic GHRH content and reversed the increase in GHRH mRNA, but did not alter plasma IGF-I concentrations. These data indicate that, in hypothyroid rats, the changes in hypothalamic GHRH content and gene expression are due to the GH deficiency ensuing from the hypothyroid state. Failure of the GH treatment to increase plasma IGF-I indicates that the feedback regulation on GHRH neurons is operated by circulating GH and/or perhaps tissue but not plasma IGF-I concentrations. Presence of low plasma IGF-I concentrations would be directly related to thyroid hormone deficiency.     

Potentiating effects of GHRH analogs on the response to chemotherapy

Growth hormone releasing hormone (GHRH) from hypothalamus nominatively stimulates growth hormone release from adenohypophysis. GHRH is also produced by cancers, acting as an autocrine/paracrine growth factor. This growth factor function is seen in lymphoma, melanoma, colorectal, liver, lung, breast, prostate, kidney, bladder cancers. Pituitary type GHRH receptors and their splice variants are also expressed in these malignancies. Synthetic antagonists of the GHRH receptor inhibit proliferation of cancers. Besides direct inhibitory effects on tumors, GHRH antagonists also enhance cytotoxic chemotherapy. GHRH antagonists potentiate docetaxel effects on growth of H460 non-small cell lung cancer (NSCLC) and MX-1 breast cancer plus suppressive action of doxorubicin on MX-1 and HCC1806 breast cancer. We investigated mechanisms of antagonists on tumor growth, inflammatory signaling, doxorubicin response, expression of drug resistance genes, and efflux pump function. Triple negative breast cancer cell xenografted into nude mice were treated with GHRH antagonist, doxorubicin, or their combination. The combination reduced tumor growth, inflammatory gene expression, drug-resistance gene expression, cancer stem-cell marker expression, and efflux-pump function. Thus, antagonists increased the efficacy of doxorubicin in HCC1806 and MX-1 tumors. Growth inhibition of H460 NSCLC by GHRH antagonists induced marked downregulation in expression of prosurvival proteins K-Ras, COX-2, and pAKT. In HT-29, HCT-116 and HCT-15 colorectal cancer lines, GHRH antagonist treatment caused cellular arrest in S-phase of cell cycle, potentiated inhibition of in vitro proliferation and in vivo growth produced by S-phase specific cytotoxic agents, 5-FU, irinotecan and cisplatin. This enhancement of cytotoxic therapy by GHRH antagonists should have clinical applications.     

The effect of cyproheptadine on plasma growth hormone (GH) and on somatostatin response to GH-releasing hormone in man

Cyproheptadine (CPH)--a putative serotonin antagonist--is known to inhibit growth hormone (GH) response to various pharmacological stimuli, as well as during sleep. To elucidate the possible site at which this drug takes effect, we examined plasma GH and somatostatin response to i.v. GHRH1-44 (1 microgram/kg body wt.) before and after CPH treatment in 10 healthy volunteers. The oral administration of CPH (8-12 mg daily for 5 days; total dose 56 mg) significantly curbed GH response to GHRH as expressed in peak plasma GH values (32.0 +/- 6.1 micrograms/l vs. 12.6 +/- 3.2 micrograms/l; P less than 0.01) and in integrated GH response area (2368 +/- 517 micrograms x l-1 x 2 h vs. 744 +/- 172 micrograms x l-1 x 2 h; P less than 0.01). Plasma somatostatin levels did not change in response to GHRH.     

Evaluation of the biological potency of new agmatine analogs of growth hormone-releasing hormone in the bovine.

Four new growth hormone-releasing hormone (GHRH) analogs with C-terminal agmatine were compared with the parent human GHRH(1-29)NH2 fragment to assess their abilities to increase serum concentrations of growth hormone (GH) in the bovine. The four analogs were: [D-Ala2, Nle27] GHRH(1-28)Agm (JG-73); [desNH2-Tyr1, Ala15, Nle27] GHRH(1-28)Agm (MZ-2-51); [desNH2-Tyr1, Ala15, D-Lys21, Nle27] GHRH(1-28)Agm (MZ-2-75); and [desNH2-Tyr1, D-Lys12,21, Ala15, Nle27] GHRH(1-28)Agm (MZ-2-87). The special characteristic of all four GHRH analogs is that arginine was replaced by agmatine (Agm) in Position 29. Five pregnant Holstein cows received these peptides subcutaneously at the following doses: 0.0156, 0.0625, 0.25, 1, and 4 micrograms/kg body wt. Each cow received each analog-dose combination according to a 5 x 5 Greco-Latin square design repeated for the 5-week treatment. Each cow also received saline vehicle only at the end of the 5-week treatment. Blood samples were collected from 30 min before until 360 min after treatment injection. Total area under the GH response curves for the 6-hr sampling period for each dose of each GHRH analog was compared. There was a linear dose-dependent GH release in response to hGHRH(1-29)NH2 and its four GHRH(1-28)Agm analogs. At the dose of 0.25 micrograms/kg, two GHRH analogs, JG-73 and MZ-2-75, stimulated greater GH release than hGHRH(1-29)NH2 (P less than 0.05). No differences were seen at the two lowest doses, 0.0625 and 0.156 micrograms/kg. When both total area under the GH response curves and GH peak amplitudes for each treatment were averaged for all doses, JG-73 and MZ-2-75 stimulated greater GH release than hGHRH(1-29)NH2 (P less than 0.05). In summary, three GHRH(1-28)Agm analogs, JG-73, MZ-2-75, and MZ-2-51, were found to be 11.8, 11.3, and 6.5 times more potent, respectively, on a weight basis, than hGHRH(1-29)NH2 in stimulating the release of GH in cows.     

Antagonistic analogs of growth hormone releasing hormone (GHRH) inhibit cyclic AMP production of human cancer cell lines in vitro.

Antagonistic analogs of growth hormone-releasing hormone (GHRH) inhibit growth of various human cancers both in vivo and in vitro. GHRH, vasoactive intestinal peptide (VIP), and pituitary adenylate cyclase-activating peptide stimulate cyclic AMP (cAMP) release from various human cancer cell lines in vitro. Thus, in the present study, we investigated the effects of antagonistic analogs of GHRH on the GHRH- and VIP-induced cAMP release from cultured human cancer cells in a superfusion system. Various human cancer cell lines were exposed to human GHRH(1-29)NH2 (2-20 nM) or VIP (0.1-5 nM) repeatedly for 12 min or continuously for 96 min. GHRH antagonist MZ-5-156 at 100 to 200 nM concentration inhibited the GHRH- or VIP-induced cAMP release from mammary (MDA-MB-468), prostatic (PC-3), and pancreatic (SW-1990 and CAPAN-2) cancer cells. These results show that antagonistic analogs of GHRH suppress the stimulatory effects of GHRH and VIP on the cAMP production of various cancer cells. Because cAMP is a potent second messenger controlling many intracellular functions, including the stimulation of cell growth, an inhibition of autocrine/paracrine action of GHRH by the GHRH antagonists may provide the basis for the development of new methods for cancer treatment.     

Changes in the hypothalamic-pituitary somatotropic function of infant hypothyroid rats

The effects of the perturbation of the pituitary-thyroid axis induced during development on the functional activity of the growth hormone (GH) regulatory neuronal systems, GH-releasing hormone (GHRH), and somatostatin (SS) were studied in 14- and 21-day-old rats made hypothyroid by giving dams propylthiouracil in the drinking water since the day of parturition. Infant hypothyroid rats, both at 14 and 21 days of life, had elevated plasma thyroid-stimulating hormone levels and decreased pituitary and plasma GH levels. Simultaneous determination of hypothalamic GHRH/SS-like immunoreactivity (LI) and GHRH/SS mRNA levels did not reveal any difference in 14-day-old hypothyroid rats when compared with age-matched controls. In contrast, 21-day-old hypothyroid rats had decreased GHRH-LI content and a striking rise in GHRH mRNA levels, whereas SS-LI content and SS gene expression remained unaltered. These data indicate that in infant hypothyroid rats, changes in the functional activity of the GHRH neuronal system occur later than changes in GH secretion and are probably dependent on the GH deficiency. The functional activity of SS neurons was apparently unaltered in these hypothyroid rats, pointing to a lesser sensitivity of this system to the perturbation of the pituitary-thyroid axis.     

Effects of acute intravenous injection of two growth hormone-releasing hormones (GHRH 1-40 and 1-29) on serum growth hormone and other pituitary hormones in short children with pulsatile growth hormone secretion

We administered two different growth hormone-releasing hormones (GHRH) to 20 short, prepubertal children who had spontaneous secretion of growth hormone (GH), assessed from 24-hour GH secretion profiles (72 sampling periods of 20 min). We compared one i.v. injection of 1 microgram/kg of GHRH 1-40 with that of GHRH 1-29 regarding serum concentrations of GH, prolactin, luteinizing hormone, follicle-stimulating hormone and IGF-I. The children were allocated to two groups without statistical randomization. Both groups were given both peptides, with at least 1 week in between. The first group started with GHRH 1-40, the other with GHRH 1-29. The peptides both induced an increased serum concentration of GH of the same magnitude: mean maximal peak of 89 +/- 12 mU/l after GHRH 1-40 and 94 +/- 10 mU/l after GHRH 1-29 (n.s.). The mean difference in maximum serum GH concentration in each child after injection was 52 +/- 9 mU/l, range 1-153 mU/l. GHRH 1-29 also induced a short-term, small increase in the concentrations of prolactin (p less than 0.05), luteinizing hormone (p less than 0.01) and follicle-stimulating hormone (p less than 0.05). We conclude that the shorter sequence GHRH 1-29, when given in a dose of 1 microgram/kg, gives a rise in serum concentration of GH similar to that after the native form GHRH 1-40.     

睡眠中间断低氧对大鼠下丘脑-垂体-肾上腺轴和生长激素的影响

目的:探讨睡眠中间断低氧对大鼠下丘脑-垂体-肾上腺轴和生长激素水平的影响.方法:大鼠分别给予吸入空气,持续低氧和间断低氧气体,在1 d,3 d,7 d和30 d后测定下丘脑促肾上腺皮质激素释放激素(CRH)和生长激素释放激素(GHRH)mRNA水平,并测定30d后血浆CRH,GHRH,促肾上腺皮质激素(ACTH)和皮质酮水平,分析其间的变化关系.结果:与对照组比较,在低氧后1 d,3 d,7 d后大鼠下丘脑CRH mRNA升高,GHRH mRNA降低,在30 d后,间断低氧组下丘脑CRH mRNA升高,GHRH mRNA降低,而持续低氧组则接近正常.间断低氧30 d后,血浆CRH、ACTH,皮质酮均升高,GHRH降低,而生长激素没有明显变化.结论:大鼠睡眠中慢性间断低氧可以引起下丘脑-垂体-肾上腺轴激素水平升高,反馈调节紊乱,可引起GHRH分泌抑制.     

Insulinhypoglycemia but not arginine infusion stimulates circulating plasma growth hormone-releasing hormone (GHRH) concentrations in children

The effect of insulinhypoglycemia and arginine infusion on circulating concentrations of plasma growth hormone-releasing hormone (GHRH) and growth hormone (GH) has been studied in 24 children (4.4 to 14.3 years). Plasma GH and GHRH concentrations were determined by RIA. Basal plasma GHRH levels were detectable in the plasma of all patients ranging from 6.8 to 27.1 pg/ml. Injection of 0.1 U/kg body wt. insulin i.v. resulted in an increase of plasma GHRH levels (11.1 +/- 1.4 pg/ml vs. 18.8 +/- 2.6 pg/ml; P less than 0.01) preceding that of plasma GH (1.5 +/- 0.4 ng/ml vs. 13.6 +/- 1.3 ng/ml; P less than 0.01). Infusion of 0.5 gm/kg body wt. arginine hydrochloride did increase GH concentrations (2.0 +/- 0.6 ng/ml vs. 13.9 +/- 2.3 ng/ml; P less than 0.01) but did not change circulating plasma GHRH levels. Since the source of peripheral GHRH concentrations is not known the importance of these findings remains to be determined.     

GHRH antagonist MZ-5-156 increases the expression of AMPK in A549 lung cancer cells

AMP-activated protein kinase (AMPK) regulates cellular proliferation, growth and metabolism. Targeted activation of AMPK is considered an important therapeutic strategy for cancer treatment. To evaluate the effect of growth hormone-releasing hormone (GHRH) and its antagonist MZ-5-156 on the phosphorylation of AMPK and other related regulatory intracellular proteins we employed human non-small cell lung cancer cell line A549, which expresses GHRH receptors. Treatment of A549 cells with GHRH antagonist decreased cell proliferation and activated AMPK as well as glycogen synthase kinase (GSK)3β. Furthermore, MZ-5-156 inhibited Akt, the mammalian target of rapamycin (mTOR) and its downstream target eIF4E which controls protein synthesis and cell growth. GHRH(1-29)NH2 counteracted all these effects. HeLa human endometrial cancer cells which do not express any GHRH receptors were used as a negative control and GHRH did not induce the AMPK activation in these cells. Our results demonstrate for the first time that GHRH antagonists can regulate the AMPK metabolic pathway, which is crucial for the growth of non-small cell lung cancer and other major cancers.     

Growth hormone response to a bolus injection of 1-44 growth-hormone-releasing hormone in very short children with intrauterine onset of growth failure

The release of growth hormone (GH) during the 120 min following a bolus venous injection of 1-44 GH-releasing hormone (GHRH) 2 micrograms/kg was studied in 52 prepubertal children aged 8.4 +/- 2.1 years, having a nonfamilial growth deficiency of prenatal onset (-3.26 +/- 1.13 SDS at birth, -3.22 +/- 0.88 SDS at the time of study) and a normal response to conventional GH stimulation tests. GH release reached a peak level of 96.1 +/- 60.2 microU/ml, being significantly higher than that found in 68 non-GH-deficient very short children whose growth failure had a postnatal onset, and not significantly correlated with the response to conventional tests. 26 of the 52 intrauterine growth retardation (IUGR) patients were re-tested with GHRH in similar conditions after 6-12 months of daily subcutaneous injections of GH and 2 days without. They reached at the second test a peak plasma GH level of 91.7 +/- 56.1 microU/ml, not different from their response to the first test. These data could be taken into consideration for long-term studies of the clinical effects of GH in IUGR children with persisting severe growth deficiency.     

GHRH antagonist causes DNA damage leading to p21 mediated cell cycle arrest and apoptosis in human colon cancer cells

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 10μM 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 (?Ψm). 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 p21Waf1/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