Antiemetic effect of a potent and selective neurokinin-1 receptor antagonist, FK886, on cisplatin-induced acute and delayed emesis in ferrets.

The antiemetic effect of a potent and selective neurokinin-1 (NK1) receptor antagonist, FK886 ([3,5-bis(trifluoromethyl)phenyl][(2R)-2-(3-hydroxy-4-methylbenzyl)-4-{2-[(2S)-2-(methoxymethyl)morpholin-4-yl]ethyl}piperazin-1-yl]methanone dihydrochloride), on cisplatin-induced acute and delayed emesis in ferrets was studied. Intravenous administration of FK886 dose-dependently inhibited cisplatin (10 mg/kg)-induced acute emesis with a minimum effective dose (MED) of 0.32 mg/kg. In the same study, oral FK886 administered 8 h prior to cisplatin also dose-dependently inhibited the acute emesis during the 4-h observation period with an MED of 3.2 mg/kg. Further, when given by repeated oral administration of ≥1.6 mg/kg at 12-h intervals, the first dose being administered 1 min before cisplatin, FK886 significantly decreased the number of emetic responses in cisplatin (5 mg/kg)-induced delayed emesis. In the same study, oral FK886 (3.2 mg/kg) repeatedly administrated at 12-h intervals, the first dose being administered 36 h post cisplatin, also significantly attenuated the delayed emesis. Pharmacokinetic data in ferrets showed that plasma FK886 reached a maximum concentration within 0.5 h of administration, suggesting rapid oral absorption. In addition, rapid brain penetration of FK886 was suggested by complete and near complete inhibition of GR73632- and copper sulfate-induced emesis, respectively, by low-dose intravenous FK886 administered shortly before the emetogens. These results suggest that FK886 is an orally available NK1 receptor antagonist which is effective against both the acute and delayed emesis induced by cisplatin. Because of its therapeutic efficacy on the delayed emesis and rapid brain distribution after oral administration, FK886 may have potential as an antiemetic agent that can be used for interventional treatment of chemotherapy-induced delayed emesis.

Antiemetic effects of a potent and selective neurokinin-1 receptor antagonist, FK886, on cisplatin- and apomorphine-induced emesis in dogs.

The antiemetic properties of a novel neurokinin-1 (NK1) receptor antagonist, FK886 ([3,5-bis(trifluoromethyl)phenyl][(2R)-2-(3-hydroxy-4-methylbenzyl)-4-{2-[(2S)-2-(methoxymethyl)morpholin-4-yl]ethyl}piperazin-1-yl]methanone dihydrochloride), were studied in dog models of cisplatin- and apomorphine-induced emesis. Intravenously administered FK886 (0.32-1 mg/kg) significantly inhibited cisplatin-induced acute emesis during the 5-h observation period. Nearly complete inhibition was observed at 1 mg/kg. At an equivalent dose range, orally administered FK886 also significantly inhibited emesis, indicating good oral absorption. Similarly, FK886 inhibited apomorphine-induced emetic responses effectively following both intravenous and oral administration. The effects were long lasting, with 1.6 mg/kg of FK886 completely blocking apomorphine-induced retching and vomiting after a 12-h pretreatment period. Furthermore, FK886 showed rapid onset of antiemetic activity after oral administration. At doses of 0.32 mg/kg or more, a pretreatment time of 0.5 h was sufficient for complete inhibition of apomorphine-induced emetic responses. This fast onset after oral administration was supported by pharmacokinetic data, which demonstrated plasma levels of FK886 after oral administration reached levels similar to those 30 min after intravenous administration. These results suggest that FK886 has excellent antiemetic properties in dogs, and that its rapid-onset and long-lasting properties might make it a promising antiemetic agent.

Pharmacological properties of FK886, a new, centrally active neurokinin-1 receptor antagonist.

The pharmacological properties of the novel neurokinin-1 (NK(1)) receptor antagonist FK886, ([3,5-bis(trifluoromethyl)phenyl][(2R)-2-(3-hydroxy-4-methylbenzyl)-4-{2-[(2S)-2-(methoxymethyl)morpholin-4-yl]ethyl}piperazin-1-yl]methanone dihydrochloride), were studied. FK886 potently inhibited the binding of [(125)I]Bolton-Hunter-labeled substance P ([(125)I]BH-SP; 100 pM) to human NK(1) receptors expressed in Chinese hamster ovary (CHO) cells (IC(50)=0.70 nM). It also possessed high affinities for dog, ferret, gerbil and guinea pig NK(1) receptors, but not for rat NK(1) receptor. FK886 was highly selective for the NK(1) receptor, with 250- and >20000-fold selectivity for human NK(1) over NK(2) and NK(3), respectively. Further, it did not inhibit radioligand binding at 54 different sites, including receptors, ion channels and transporters. FK886 inhibited substance P (3.2 nM)-induced inositol phosphate formation in human NK(1) receptor-expressing CHO cells (IC(50)=1.4 nM) without stimulating NK(1) receptors. The antagonism exerted by FK886 against human NK(1) receptor was insurmountable in saturation binding experiments, with both the affinity and B(max) of [(125)I]BH-SP being significantly reduced. After intravenous administration, FK886 (0.01-0.1 mg/kg) dose-dependently inhibited the foot-tapping behavior induced by intracerebroventricular administration of a selective NK(1) receptor agonist, GR73632 (10 pmol), in gerbils, with significant inhibition being observed at doses of 0.032-0.1 mg/kg, indicating excellent brain penetration. The brain penetration of FK886 was further demonstrated by the cerebral distribution of radioactivity after intravenous injection of radiolabeled FK886. Taken together, these results demonstrate that FK886 is a potent, highly selective and centrally active, insurmountable antagonist of the NK(1) receptor, and suggest that FK886 antagonizes various NK(1) receptor-mediated biological effects in the central nervous system.

Ameliorating effect of FK614, a novel nonthiazolidinedione peroxisome proliferator-activated receptor gamma agonist, on insulin resistance in Zucker fatty rat.

Effect of 3-(2,4-dichlorobenzyl)-2-methyl-N-(pentylsulfonyl)-3H-benzimidazole-5-carboxamide (FK614), a novel nonthiazolidinedione peroxisome proliferator-activated receptor (PPAR) gamma agonist, on glucose tolerance and insulin resistance in peripheral tissues and in liver using Zucker fatty rats (genetically obese and insulin-resistant) was evaluated and compared to other insulin sensitizers. FK614 (0.32, 1 and 3.2 mg/kg), two thiazolidinedione PPAR gamma agonists, rosiglitazone (0.1, 0.32, 1 and 3.2 mg/kg) and pioglitazone (1, 3.2 and 10 mg/kg), and a biguanide, metformin (320 and 1000 mg/kg), were orally administered to Zucker fatty rats once a day for 14 days. Zucker fatty rats treated with FK614 and rosiglitazone were subjected to evaluation by oral glucose tolerance test. Ameliorating effect of each compound on peripheral and hepatic insulin resistance was evaluated using a euglycemic-hyperinsulineamic clamp procedure. FK614 and rosiglitazone dose-dependently improved impaired glucose tolerance in Zucker fatty rats. In addition, FK614 dose-dependently ameliorated peripheral and hepatic insulin resistance in Zucker fatty rats, with the degree of its effect in peripheral tissues almost equivalent to that in liver when compared at each dose tested. Similar data indicating ameliorating effects on insulin resistance was obtained for rosiglitazone and pioglitazone. Metformin showed less potent effects than other insulin sensitizers and its effect in liver tended to be greater than that in peripheral tissues. These findings suggest clinical potential for FK614 as a treatment of type 2 diabetes, acting by ameliorating insulin resistance both in peripheral tissues and liver.

Marker genes to predict sensitivity to FK228, a histone deacetylase inhibitor.

In this study, we detected genes sensitive to an histone deacetylase inhibitor, FK228 [(E)-(1S,4S,10S,21R)-7-[(Z)-ethylidene]-4,21-diisopropyl-2-oxa-12,13-dithia-5,8,20,23-tetraazabicyclo-[8,7,6]-tricos-16-ene-3,6,9,19,22-pentanone; FR901228, depsipeptide] in vitro and identified marker genes to predict sensitivity to FK228 in vivo using Affymetrix GeneChip. Three percent of genes (205/7070) were sensitive to FK228 in vitro, 105 and 100 genes, were up- and down-regulated, respectively, by FK228. Commonly up-regulated genes included p21(WAF1/Cip1), interleukin-8 (IL-8), histone family, JunB, caspase 9, mitogen-activated protein kinase phosphatase 1 (MKP-1) and mitogen-activated protein kinase (MAPK) family, and commonly down-regulated genes included cyclin A and MAPK family. One percent of genes (76/7070) showed native differences in patterns of expression, when FK228-sensitive (PC-3 prostate and SC-6-JCK (SC-6) stomach) and FK228-resistant (ACHN and A-498 renal) tumors implanted in BALB/c nu/nu mice were compared. Twenty-seven and forty nine of those genes were expressed at high or low levels, respectively, in FK228-sensitive tumors. Caspase 9 and MKP-1 genes showed distinct differences in patterns of expression between FK228-sensitive and resistant tumors and have been known to have roles in apoptosis and chromatin remodeling. The expression of caspase 9 gene was higher in FK228-sensitive tumors and the expression of MKP-1 gene was higher in FK228-resistant tumors. Caspase 9 and MKP-1 genes in the other FK228-sensitive tumors had the same patterns of expression as they did in PC-3 and SC-6 tumors. Our results present profiles of gene expression related to FK228 and marker genes to predict sensitivity to FK228, such as caspase 9 and MKP-1 genes.

FR177391, a new anti-hyperlipidemic agent from Serratia. II. Pharmacological activity of FR177391.

The pharmacological effect of FR177391, isolated from Serratia liquefaciens No. 1821, was studied in normal animals and various types of animal models of hypertriglyceridemia. Treatment of normal mice with FR177391 resulted in an increase in heparin-releasable lipoprotein lipase (LPL) activity in the blood and epididymal fat tissue. FR177391 treatment decreased triglyceride (TG) and increased high-density lipoprotein cholesterol in the blood in normal rats following 7 days treatment, suggesting potent LPL activating properties of FR177391. Both Triton WR1339-induced severe and fructose-induced mild hypertriglyceridemia in rats were attenuated by FR177391 treatment. Severely elevated levels of TG in db/db mice, an insulin resistant diabetic animal model, also significantly decreased from 14 days of treatment with FR177391. FR177391 treatment for 9 days caused a decrease in the elevated levels of TG in mice induced by intraperitoneal inoculation of murine lymphoma EL-4. Overall, this study demonstrated that FR177391 can be possibly a LPL activating agent and that FR177391 treatment improved hypertriglyceridemia in various rat and mouse animal models. These results suggest that FR177391 is a promising candidate compound for the management of hypertriglyceridemia.

Possible involvement of enhanced intestinal microsomal triglyceride transfer protein (MTP) gene expression in acceleration of lipid absorption by a western-type diet in apolipoprotein E knockout mice.

Evidence has been accumulating that triglyceride (TG)-rich lipoproteins are atherogenic. Microsomal TG transfer protein (MTP) is essential for the synthesis of both chylomicron in the intestine and very low density lipoprotein in the liver. To investigate whether a western-type diet, a so-called atherogenic diet, alters intestinal lipid absorption via change in intestinal MTP expression, the effects of two different diet regimes in apolipoprotein-E knockout (apoE KO) mice were examined. Male apoE KO mice aged 6 weeks were fed a western-type diet or a chow diet for 5 weeks. Then, measurement of plasma TG levels after oral fat-loading and analysis of jejunal MTP gene expression were performed. Both the maximum level and the 0-8 h area under the curve (AUC) of the increase in TG levels in the western-type diet-fed mice were almost three times greater than those in the chow diet-fed mice. MTP gene expression, determined by reverse transcriptase-polymerase chain reaction (RT-PCR), was obviously enhanced in the western-type diet-fed mice compared to the chow diet-fed mice. These results suggest that the enhancement of intestinal MTP gene expression is involved in the accelerated lipid absorption in the western-type diet-fed mice.

Acute and chronic effects of FR-149175, a beta 3-adrenergic receptor agonist, on energy expenditure in Zucker fatty rats.

Clinical therapies for both obesity and obese non-insulin-dependent diabetes mellitus require maintenance of reduced body weight after the initial successful reduction resulting from calorie control, exercise, or medication. Although beta(3)-adrenergic receptor (beta(3)-AR) agonists have been shown to stimulate whole body energy expenditure and lipid mobilization, whether stimulatory effects on oxygen consumption and lipolysis are influenced by chronic exposure to agonists has not been fully characterized. We therefore examined the acute and chronic effects of FR-149175, a selective beta(3)-AR agonist, on whole body oxygen consumption in genetically obese Zucker fatty rats. Chronic treatment with FR-149175 caused a decrease in both body weight gain and white fat pad weight at doses that induced lipolysis in acute treatment (1 and 3.2 mg/kg p.o.). Single administration of FR-149175 (0.1, 1, and 3.2 mg/kg p.o.) dose dependently increased whole body oxygen consumption. Repetitive administration did not cause attenuation of the thermogenic response at lower doses (0.1 and 1 mg/kg 2 times daily), whereas the highest dose (3.2 mg/kg 2 times daily) induced a progressive increase in oxygen consumption. PCR analyses of retroperitoneal white adipose tissue indicated little or no change in beta(3)-AR mRNA levels. Uncoupling protein 1 gene expression increased at 1 mg/kg, and drastic upregulation was detected at 3.2 mg/kg. FR-149175 also increased HSL mRNA levels in a dose-related manner, whereas there was no effect on genes involved in beta-oxidation. These results support that the thermogenic effect of beta(3)-AR agonists is not attenuated by chronic exposure to agonists.

Phosphodiesterase type 5 inhibition ameliorates nephrotoxicity induced by cyclosporin A in spontaneous hypertensive rats.

Our recent study suggests that there is a reciprocal mechanism to maintain cGMP content, via both a decrease in cGMP degradation (decrease in cGMP-phosphodiesterase activity) and an increase in synthesis of cGMP (increase in guanylate cyclase activity) in the kidney of cyclosporin A-treated rats. We undertook this study to clarify the role of cGMP-phosphodiesterase in cyclosporin A nephrotoxicity by evaluating N-(3,4-dimethoxybenzyl)-2-[[(1R)-2-hydroxy-1-methylethyl]amino]-5-nitrobenzamide (FR226807), a phosphodiesterase type 5 inhibitor, in an animal model. Male spontaneous hypertensive rats (SHR) were treated with cyclosporin A (50 mg/kg) for 2 weeks or with cyclosporin A and FR226807 (3.2 mg/kg or 10 mg/kg) for 2 weeks. Cyclosporin A-treated rats showed renal dysfunction and histological change compared with vehicle-treated rats. Administration of FR226807 improved the renal dysfunction (increase in serum creatinine and fractional excretion of sodium, and decrease in creatinine clearance) as well as the pathological changes (tubular vacuolization) induced by cyclosporin A in SHR. At the molecular level, administration of FR226807 resulted in a further increase in cGMP content in the kidney, aorta and platelets from cyclosporin A-treated rats. Our present study demonstrates that cGMP-phosphodiesterase plays an important role in the cyclosporin A nephrotoxicity and also suggests that further inhibition of cGMP-phosphodiesterase is a potential pharmacological target for preventing cyclosporin A nephrotoxicity.

Contribution of tachykinin receptor subtypes to micturition reflex in guinea pigs.

The aim of the present study was to determine the role of tachykinin in the micturition reflex in guinea pigs. We investigated the effects of tachykinin NK(1) receptor antagonists, GR205171 ([2-methoxy-5-(5-trifluoromethyl-tetrazol-1-yl)-benzyl]-(2S-phenyl-piperidin-3S-yl)-amine), CP99994 ((+), (2R, 3R)-3-(2-methoxybenzyl-amino)-2-phenylpiperidine) and FK888 (N(2)-[(4R)-4-hydroxy-1-(1-methyl-1H-indol-3-yl) carbonyl-L-prolyl]-N-methyl-N-phenylmethyl-3-(2-naphthyl)-L-alaninamide), the tachykinin NK(2) receptor antagonist, SR48968 ((+)-N-methyl-[4-(4-acetylamino-4-phenyl piperidino)-2-(3, 4-dichloro-phenyl)butyl] benzamide), and the tachykinin NK(3) receptor antagonist, SB223412 ((S)-(-)-N-(alpha-ethylbenzyl)-3-hydroxy-2-phenylquinoline-4-carboxamide) on rhythmic bladder contraction. GR205171 and CP99994 but not SR48968 or SB223412 reduced bladder contraction frequency. FK888 inhibited the frequency very slightly at the highest dose tested. The distribution of tachykinin NK(1) receptor antagonists to the central nervous system after intravenous administration was examined using an ex vivo binding assay. GR205171 was distributed to the brain and spinal cord, but the tachykinin NK(1) receptor antagonist, FK888, was not. These results suggest that tachykinin NK(1) receptors, which are located in the central nervous system, play an important role in micturition in guinea pigs.

Antitumor efficacy of FK228, a novel histone deacetylase inhibitor, depends on the effect on expression of angiogenesis factors.

UNLABELLED: It has been recently demonstrated that histone deacetylase inhibitors inhibit angiogenesis, but their mechanism of action has not been characterized well. In this study, we examined the in vitro and in vivo effects of FK228 [(E)-(1S,4S,10S,21R)-7-[(Z)-ethylidene]-4,21-diisopropyl-2-oxa-12,13-dithia-5,8,20,23-tetraazabicyclo-[8,7,6]-tricos-16-ene-3,6,9,19,22-pentanone; FR901228, depsipeptide], an HDAC inhibitor, on the expression of angiogenesis factors in FK228-sensitive PC-3 prostate and FK228-resistant ACHN renal cancer cells. FK228 suppressed the expression of VEGF mRNA in PC-3 cells, but not in ACHN cells. FK228 also suppressed the expression of basic fibroblast growth factor (bFGF) mRNA in both PC-3 and ACHN cells. Under conditions of hypoxia, FK228 suppressed the expression of VEGF mRNA without modulating the expression of hypoxia-inducible factor-1 alpha mRNA in PC-3 cells. FK228 induced the highest acetylation of histone H3 and H4 in the P2 region of the VEGF promoter, which includes the hypoxia-inducible factor-1 alpha binding site that plays an important role in regulating the expression of VEGF gene. Moreover, FK228 reduced the amount of VEGF and bFGF protein, and their mRNA levels in PC-3 xenograft implanted in nude mice, but did not reduce them in ACHN xenograft. IN CONCLUSION: (i) FK228 showed a suppressive effect on the expression of angiogenesis factors, such as VEGF and bFGF, in PC-3 xenograft but not in ACHN xenograft, which suggests that the effect on the expression of angiogenesis factors is important for the antitumor efficacy of FK228; (ii) FK228 caused histone acetylation of the VEGF promoter regions, which may contribute to the suppression of VEGF gene expression.

Enzyme activities of the nitric oxide-cGMP pathway in corpus cavernosum isolated from middle-aged rats.

Cyclic guanosine-3',5'-monophosphate (cGMP)-mediated mechanisms play an important role in vasodilation and blood pressure regulation. We investigated basal activity of the nitric oxide (NO)-cGMP signal transduction pathway in corpus cavernosum from both middle-aged and young rats, and the electrical field stimulation-induced relaxation in the organ was also evaluated. In middle-aged rats, nitric oxide synthase (NOS) and cGMP-phosphodiesterase activities were significantly decreased; however, guanylate cyclase activity was similar. cGMP concentration, a secondary messenger of NO, remained almost the same level as compared with young rats. These results suggest that decrease in cGMP-phosphodiesterase activity is likely to account for the maintenance of cGMP concentration. In isolated corpus cavernosum from middle-aged rats, electrical field stimulation-induced relaxation was partially impaired. These results suggest that downregulation of the NOS and cGMP-phosphodiesterase activities are early events in the pathogenesis of erectile dysfunction.

Effects of FK228, a novel histone deacetylase inhibitor, on tumor growth and expression of p21 and c-myc genes in vivo.

In this study, we examined the effects of FK228 (FR901228, depsipeptide) on tumor growth and expression of p21 and c-myc genes in vivo. FK228 induced the expression of p21 mRNA and decreased c-myc mRNA in tumor xenograft sensitive to FK228. However, FK228 did not sufficiently modulate the expression of p21 mRNA and increased the expression of c-myc in tumor xenograft less sensitive to FK228. The modulation of p21 and/or c-myc genes may be critical for the marked antitumor activity of FK228 in vivo.

Effects of FK317, a novel anti-cancer agent, on survival of mice bearing B16BL6 melanoma and Lewis lung carcinoma.

The effects of FK317 (11-acetyl-8-carbamoyloxymethyl-4-formyl-6-methoxy-14- oxa-1,11-diazatraacylo[7.4.1.0(2.7).0(10.2)]-tetradeca-2,4,6-trien-9-yl acetate), a novel anti-cancer agent, and mitomycin C (MMC) on survival time of mice bearing B16BL6 melanoma and Lewis lung carcinoma (LLC), induced by intravenous inoculation of the tumor, were investigated. Treatment with FK317 resulted in a significant prolongation of survival time in both tumor models. Four of ten mice bearing B16BL6 were disease-free following FK317 treatment. In contrast, MMC was not effective in prolonging survival time. Overall, this study demonstrated that FK317 shows more potent survival extension in mice bearing B16BL6 and LLC than MMC, suggesting that FK317 may have therapeutic utility for cancer chemotherapy.

Effects of FK228, a novel histone deacetylase inhibitor, on human lymphoma U-937 cells in vitro and in vivo.

FK228 [(E)-(1S,4S,10S,21R)-7-[(Z)-ethylidene]-4,21-diisopropyl-2-oxa-12,13-dithia-5,8,20,23-tetraazabicyclo-[8,7,6]-tricos-16-ene-3,6,9,19,22-pentanone; FR901228, depsipeptide] is a novel histone deacetylase inhibitor that shows therapeutic efficacy in Phase I trials of patients with malignant lymphoma. However, its mechanism of action has not been characterized. In this study, we examined the in vitro and in vivo effects of FK228 on human lymphoma U-937 cells. FK228 very strongly inhibited the growth of U-937 cells with an IC(50) value of 5.92 nM. In a scid mouse lymphoma model, mice treated with FK228 once or twice a week survived longer than control mice, with median survival times of 30.5 (0.56 mg/kg) and 33 days (0.32 mg/kg), respectively (vs. 20 days in control mice). Remarkably, 2 out of 12 mice treated with FK228 (0.56 mg/kg once or twice a week) survived past the observation period of 60 days. The apoptotic population of U-937 cells time-dependently increased to 37.7% after 48 hr of treatment with FK228. In addition, FK228 induced G1 and G2/M arrest and the differentiation of U-937 cells to the CD11b(+)/CD14(+) phenotype. Expression of p21(WAF1/Cip1) and gelsolin mRNA increased up to 654- and 152-fold, respectively, after 24hr of treatment with FK228. FK228 caused histone acetylation in p21(WAF1/Cip1) promoter regions, including the Sp1-binding sites. In conclusion, (i) FK228 prolonged the survival time of scid mice in a lymphoma model, and (ii) the beneficial effects of FK228 on human lymphoma may be exerted through the induction of apoptosis, cell cycle arrest, and differentiation via the modulation of gene expression by histone acetylation.

Resiniferatoxin antagonizes cisplatin-induced emesis in dogs and ferrets.

We evaluated the antiemetic activity of resiniferatoxin, an ultrapotent capsaicin analogue, on cisplatin- and apomorphine-induced emesis in dogs, and on cisplatin-induced acute and delayed emesis in ferrets. In the dog, resiniferatoxin (10 microg/kg, s.c.) 30 min before the injection of cisplatin markedly prevented acute emesis induced by cisplatin. When animals were given resiniferatoxin (10 microg/kg, s.c.) 24 h prior to cisplatin, the emesis was still inhibited, but not significantly. Resiniferatoxin (10 microg/kg, s.c.) 30 min before the administration of apomorphine also significantly reduced the emetic responses induced by apomorphine in dogs. In the ferret, resiniferatoxin (10 microg/kg, s.c.) 30 min prior to cisplatin completely inhibited acute emesis caused by cisplatin (10 mg/kg, i.p.). When ferrets were given resiniferatoxin (10 microg/kg, s.c.) 16 h prior to cisplatin, the emesis was still significantly inhibited. Cisplatin (5 mg/kg, i.p.) induced both acute (0-24 h) and delayed (24-72 h) phase emesis, and a single injection of resiniferatoxin (10 microg/kg, s.c.) at 36 h after cisplatin significantly reduced subsequent emetic responses during the 36-72 h period. These results suggest that resiniferatoxin-related vanilloids may be useful drugs against both acute and delayed emesis induced by cancer chemotherapy.