Characterization of the antitumor effects of the selective farnesyl protein transferase inhibitor R115777 in vivo and in vitro.

R115777 [(B)-6-[amino(4-chlorophenyl)(1-methyl-1H-imidazol-5-yl)-methyl]-4-(3-chlorophenyl)-1-methyl-2(1H)-quinolinone] is a potent and selective inhibitor of farnesyl protein transferase with significant antitumor effects in vivo subsequent to oral administration in mice. In vitro, using isolated human farnesyl protein transferase, R115777 competitively inhibited the farnesylation of lamin B and K-RasB peptide substrates, with IC50s of 0.86 nM and 7.9 nM, respectively. In a panel of 53 human tumor cell lines tested for growth inhibition, approximately 75% were found to be sensitive to R115777. The majority of sensitive cell lines had a wild-type ras gene. Tumor cell lines bearing H-ras or N-ras mutations were among the most sensitive of the cell lines tested, with responses observed at nanomolar concentrations of R115777. Tumor cell lines bearing mutant K-ras genes required higher concentrations for inhibition of cell growth, with 50% of the cell lines resistant to R115777 up to concentrations of 500 nM. Inhibition of H-Ras, N-Ras, and lamin B protein processing was observed at concentrations of R115777 that inhibited cell proliferation. However, inhibition of K-RasB protein-processing could not be detected. Oral administration b.i.d. of R115777 to nude mice bearing s.c. tumors at doses ranging from 6.25-100 mg/kg inhibited the growth of tumors bearing mutant H-ras, mutant K-ras, and wild-type ras genes. Histological evaluations revealed heterogeneity in tumor responses to R115777. In LoVo human colon tumors, treatment with R115777 produced a prominent antiangiogenic response. In CAPAN-2 human pancreatic tumors, an antiproilferative response predominated, whereas in C32 human melanoma, marked induction of apoptosis was observed. The heterogeneity of histological changes associated with antitumor effects suggested that R115777, and possibly farnesyl protein transferase inhibitors as a class, alter processes of transformation related to tumor-host interactions in addition to inhibiting tumor-cell proliferation.

Release of spermidine from the rat cortex following permanent middle cerebral artery occlusion.

We have studied the effects of middle cerebral artery (MCA) occlusion in rats on polyamine efflux in the parietal cortex using the microdialysis technique. Dialysis probe implantation itself provoked a delayed, prolonged and vigorous release of spermidine and putrescine. Spermidine release returned to stable baseline levels within 48 hours. Putrescine release also returned to lower levels within this time period but putrescine levels in the dialysate fluctuated dramatically in individual animals. Because of the underlying effects of the dialysis probe (likely a reflection of traumatic cerebral damage and stimulation of polyamine metabolism and release within the immediate vicinity of the dialysis probe), MCA occlusion was performed 48 hours after probe implantation. MCA occlusion persistently (5/5 animals) resulted in a significant increase in cortical spermidine efflux, although the onset, magnitude and duration of this increased release was variable. Putrescine efflux was significantly increased in 2/5 animals with MCA occlusion but the increase in release was similar to the spontaneous fluctuations observed in control animals. Spermine was not detectable in cortical dialysates of control or MCA occluded groups. Spermidine, but not spermine or putrescine is consistently released from the parietal cortex following permanent focal ischaemia and may contribute to ischaemic neuropathology either through its effects at the N-methyl-D-aspartate (NMDA) receptor or via direct, and as yet uncharacterised, neurotoxic effects.

Neuroprotective efficacy of N omega-nitro-L-arginine after focal cerebral ischemia in the mouse and inhibition of cortical nitric oxide synthase.

The neuroprotective effects of various doses of N omega-nitro-L-arginine have been correlated with the degree of N omega-nitro-L-arginine-induced inhibition of cortical nitric oxide synthase activity measured ex vivo. Following focal cerebral ischemia induced by permanent occlusion of middle cerebral artery in the mouse, repeated administration of 1 mg/kg i.p. of N omega-nitro-L-arginine (beginning 5 min after surgery) reproducibly decreased by 66-76% the infarct volume measured at 6 days post-occlusion. This dose of N omega-nitro-L-arginine decreased cortical nitric oxide (NO) synthase activity by 70-73%. The neuroprotective efficacy of N omega-nitro-L-arginine increased dose-dependently over the range of doses of 0.1-1 mg/kg. Within this dose range of N omega-nitro-L-arginine, there was a good parallelism between the extent of inhibition of cortical NO synthase activity measured ex vivo and the degree of neuroprotection. However, higher doses of N omega-nitro-L-arginine (3 and 10 mg/kg i.p.), which inhibited NO synthase activity more effectively (up to 94%) failed to significantly reduce the infarct size. Repeated administrations of increasing doses of L-arginine (up to 30 mg/kg i.p.) with a low dose of N omega-nitro-L-arginine (1 mg/kg i.p.) caused a dose-dependent reduction in the neuroprotective efficacy of N omega-nitro-L-arginine while the extent of NO synthase inhibition measured ex vivo did not decrease significantly.(ABSTRACT TRUNCATED AT 250 WORDS)

Lack of neuroprotective effect of some sigma ligands in a model of focal cerebral ischemia in the mouse.

The potential neuroprotective effect of seven sigma ligands has been evaluated in a mouse model of focal cerebral ischemia (induced by coagulation of the left middle cerebral artery) and compared to that of known N-methyl-D-aspartate (NMDA) antagonists. When given after the induction of cerebral ischemia, the NMDA receptor antagonists dizocilpine and CGS 19755 and the mixed NMDA antagonist/sigma ligand eliprodil (SL 82.0715) afforded very substantial protection against cortical infarction (92, 44 and 72%, at the doses of 1, 10 and 10 mg/kg, i.p., for dizocilpine, CGS 19755 and eliprodil, respectively). In contrast, none of the sigma ligands investigated--DTG, DMTG, GBR 12909, (+)- and (-)-3-PPP (up to 10 mg/kg), BMY 14802 (up to 30 mg/kg), except haloperidol at a high dose (3 mg/kg)--had neuroprotective effects.

Functional, behavioral, and histological changes induced by transient global cerebral ischemia in rats: effects of cinnarizine and flunarizine.

Temporary cerebral ischemia (15 min) produced by "four-vessel occlusion" in the rat causes neurological disorders, changes in behavior (locomotor hyperactivity), and neuronal damage in the neocortex, striatum, and especially the CA1 zone of the hippocampus. We have studied the effects of two calcium overload blockers, flunarizine (50 mg/kg p.o. twice a day) and cinnarizine (100 mg/kg p.o. twice a day), on these alterations. Cinnarizine markedly improved the functional abnormalities of ischemia but had little or no effect upon the neuronal damage. In contrast, flunarizine provided far greater neuronal protection but with less obvious effects upon behavioral parameters. However, there was evidence of sedation 2 h after treating animals with this dose of flunarizine that might have masked any positive effect of the drug on behavior. We conclude that under the present experimental conditions, there is no correlation between the early and late behavioral changes observed following a temporary cerebral ischemic episode and the histological damage observed in certain vulnerable neurons, particularly in the hippocampus, 72 h after the insult.