Adenylyl cyclase activity of the fission yeast Schizosaccharomyces pombe is not regulated by guanyl nucleotides.

The adenylyl cyclase activity of the fission yeast Schizosaccharomyces pombe is localized to the plasma membrane of the cell. The enzyme utilizes Mn2+/ATP as substrate and free Mn2+ ions as an effector. Unlike the baker yeast Saccharomyces cerevisiae, S. pombe adenylyl cyclase does not utilize Mg2+/ATP as substrate and the activity is not stimulated by guanyl nucleotides. The optimal pH for the S. pombe adenylyl cyclase activity is 6.0. The activity dependence on ATP is cooperative with a Hill coefficient of 1.68 +/- 0.14.

Controlled delivery of a tyrphostin inhibits intimal hyperplasia in a rat carotid artery injury model.

We examined the inhibitory effect of AG-17, a potent inhibitor of protein tyrosine kinase activity on injury-induced vascular SMC proliferation by polymeric-based, periadventitial controlled release implant in the balloon catheter carotid injury model in rats. The AG-17 delivery system was formulated from ethylenevinyl acetate copolymer and the release kinetics as well as drug stability were determined. Polymeric matrices containing 2 or 10% AG-17 were implanted perivascularly in rats following balloon catheter injury. Western blot analysis of explanted arterial segments revealed enhanced tyrosine phosphorylation in injured arteries that was essentially reduced to normal levels in treated arteries. The mean neointima to media ratios were significantly reduced in both 2% (0.79 +/- 0.17, n = 9, P < 0.02) and 10% AG-17 (0.59 +/- 0.09, n = 12, P < 0.001) groups in comparison to the control group (1.38 +/- 0.18, n = 16). The mean areas of the media in the control and the 2% AG-17 group did not differ significantly but a significant reduction of the mean area of the media was observed in 10% AG-17 group. Embedding of the unstable tyrphostin compound, AG-17, in a hydrophobic matrix stabilizes the drug both in vitro and in vivo, and allows delivery-rate modulation as well as protracted site-specific therapy. Perivascular controlled release delivery of the tyrphostin AG-17 inhibits neointimal formation in the rat carotid injury model.

Tyrphostins. 2. Heterocyclic and alpha-substituted benzylidenemalononitrile tyrphostins as potent inhibitors of EGF receptor and ErbB2/neu tyrosine kinases.

We have previously described a novel series of low molecular weight protein tyrosine kinase inhibitors which we named tyrphostins. The characteristic active pharmacophore of these compounds was the hydroxy-cis-benzylidenemalononitrile moiety. In this article we describe three novel groups of tyrphostins: (i) one group has the phenolic moiety of the cis-benzylidenemalononitrile replaced either with other substituted benzenes or with heteroaromatic rings, (ii) another is a series of conformationally constrained derivatives of hydroxy-cis-benzylidenemalononitriles in which the malononitrile moiety is fixed relative to the aromatic ring, and (iii) two groups of compounds in which the position trans to the benzenemalononitrile has been substituted by ketones and amides. Among the novel tyrphostins examined we found inhibitors which discriminate between the highly homologous EGF receptor kinase (HER1) and ErbB2/neu kinase (HER2). These findings may lead to selective tyrosine kinase blockers for the treatment of diseases in which ErbB2/neu is involved.

Alpha-cyanocinnamide derivatives: a new family of non-peptide, non-sulfhydryl inhibitors of Ras farnesylation.

Farnesylation of Ras and other proteins is required for their membrane attachment and normal function. Here we report on the synthesis of alpha-cyanocinnamide derivatives, a new family of farnesyltransferase inhibitors. These compounds are nonpeptidic and do not contain sulfhydryl groups. The most potent compound is a pure competitive inhibitor with respect to the Ras protein and mixed competitive with respect to farnesyl diphosphate. Selectivity studies against geranylgeranyltransferase and biological activities of selected compounds are described.

Synthesis and biological activity of semipeptoid farnesyltransferase inhibitors.

Semipeptoids derived from the Ras farnesyl transferase inhibitor, CVFM, were synthesized by the Simultaneous Multiple Analogue Peptide Synthesis methodology. The semipeptoids were screened for their in vitro inhibition potency towards farnesyl transferase and geranylgeranyl transferase. Structure-activity relationship studies led to a potent and selective inhibitor, HR-11, which blocks Ras farnesylation in vitro with an IC50 of 1.2 nM. The cell permeable methyl ester derivative of HR-11, HR-12, inhibits Ras farnesylation in intact cells with an IC50 of 10 microM and with no detectable inhibition of Rap1A/K-rev geranyl-geranylation.

Suppression of experimental autoimmune encephalomyelitis by tyrphostin AG-556.

Tyrosine kinase blockers from the AG 126/AG-556 tyrphostin family are shown to inhibit the lipopolysaccharide (LPS)-induced production of tumor necrosis factor alpha (TNFalpha), nitric oxide (NO), and prostaglandin E2 (PGE2) in primary rat astrocytes cultures. The tyrphostin AG-556 which was previously shown to be effective against sepsis in mice and dogs also show excellent efficacy in inhibiting experimental autoimmune encephalomyelitis (EAE) in mice. AG-556 does not block the activation of JNK/SAPK and of p38/HOG and therefore seems to act at a target down stream to these kinases which is activated in stress or at a target on an obligatory parallel pathway. These findings together with previous results showing inhibition of sepsis in mice and dogs suggest that protein tyrosine kinase (PTK) blockers of the AG-556 family may be considered in the management of human autoimmune disorders such as multiple sclerosis (MS).