Discovery and functional characterization of a novel small molecule inhibitor of the intracellular phosphatase, SHIP2.

BACKGROUND AND PURPOSE: The lipid phosphatase known as SH2 domain-containing inositol 5'-phosphatase 2 (SHIP2) plays an important role in the regulation of the intracellular insulin signalling pathway. Recent studies have suggested that inhibition of SHIP2 could produce significant benefits in treatment of type 2 diabetes. However, there were no small molecule SHIP2 inhibitors and we, therefore, aimed to identify this type of compound. EXPERIMENTAL APPROACH: The phosphatase assay with malachite green was used for high-throughput screening. The pharmacological profiles of suitable compounds were further characterized in phosphatase assays, cellular assays and oral administration in normal and diabetic (db/db) mice. KEY RESULTS: During high-throughput screening, AS1949490 was identified as a potent SHIP2 inhibitor (IC(50)= 0.62 microM for SHIP2). This compound was also selective for SHIP2 relative to other intracellular phosphatases. In L6 myotubes, AS1949490 increased the phosphorylation of Akt, glucose consumption and glucose uptake. In FAO hepatocytes, AS1949490 suppressed gluconeogenesis. Acute administration of AS1949490 inhibited the expression of gluconeogenic genes in the livers of normal mice. Chronic treatment of diabetic db/db mice with AS1949490 significantly lowered the plasma glucose level and improved glucose intolerance. These in vivo effects were based in part on the activation of intracellular insulin signalling pathways in the liver. CONCLUSIONS AND IMPLICATIONS: This is the first report of a small molecule inhibitor of SHIP2. This compound will help to elucidate the physiological functions of SHIP2 and its involvement in various diseases, such as type 2 diabetes.

FR226807: a potent and selective phosphodiesterase type 5 inhibitor.

We describe the pharmacological characteristics of a novel phosphodiesterase type 5 inhibitor FR226807, N-(3,4-dimethoxybenzyl)-2-[[(1R)-2-hydroxy-1-methylethyl]amino]-5-nitrobenzamide. FR226807 inhibited phosphodiesterase type 5 isolated from human platelets with an IC(50) value of 1.1 nM. FR226807 also inhibited phosphodiesterase type 6 with an IC(50) of 20 nM; however, the IC(50) value for phosphodiesterase type 6 was 18-fold higher than that for phosphodiesterase type 5. The IC(50) values of FR226807 for other phosphodiesterases (phosphodiesterase type 1, phosphodiesterase type 2, phosphodiesterase type 3, and phosphodiesterase type 4) were 1000-fold higher than that for phosphodiesterase type 5. FR226807 increased the cyclic guanosine monophosphate (cGMP) content in corpus cavernosum isolated from rabbit, an effect associated with relaxation of the muscle. FR226807 enhanced the relaxation response induced by electrical field stimulation of corpus cavernosum isolated from the rabbit. In an anesthetized dog model for the evaluation of erectile function, intravenous administration of FR226807 prolonged the time to return to 75% of maximal intracavernosal pressure after cessation of electrical stimulation of the pelvic nerve. In summary, FR226807 is a potent and highly selective phosphodiesterase type 5 inhibitor with an augmentative effect on penile erection and will be useful for the treatment of erectile dysfunction.

Reciprocal regulation of cyclic GMP content by cyclic GMP-phosphodiesterase and guanylate cyclase in SHR with CsA-induced nephrotoxicity.

1. The effect of the immunosuppressant drug, cyclosporin A (CsA), on the nitric oxide (NO)-cyclic GMP pathway was examined in spontaneous hypertensive rats (SHR). 2. CsA (50 mg kg(-1)) treatment for 14 days induced typical CsA nephrotoxicity, which was characterized by morphological changes in the glomerulus and proximal tubule as well as an abnormality of creatinine clearance, FENa and BUN. 3. CsA significantly decreased both NOS activity in the kidney and NOx contents in urine, but significantly increased cyclic GMP content in the kidney. 4. A marked change in two kinds of enzyme, which contribute towards the increase in cyclic GMP in tissue, namely, a decrease in cyclic GMP-phosphodiesterase activity and increase in guanylate cyclase activity, was observed in the kidney treated with CsA. 5. In the isolated perfused kidney, a decreased in perfusion pressure induced by SNP in the kidney isolated from CsA group was significantly greater than that of control. 6. There seem to exist a reciprocal mechanism to maintain cyclic GMP content via both a decrease in cyclic GMP degradation and an increase in synthesis of cyclic GMP in the kidney treated with CsA. This mechanism is likely to be playing an important role to regulate the homeostasis in the kidney with CsA nephrotoxicity.

Synthesis and characterization of orally active nonpeptide vasopressin V2 receptor antagonists.

The present study was undertaken to evaluate whether a novel series of 2,6-diaza-5-oxobicyclo[5.4.0]undeca-1(7),8,10-triene derivatives exhibited antagonistic activity for vasopressin V1 and V2 receptors. Most of these compounds were synthesized and showed a high affinity potential for V2 receptor and low to moderate affinity potential for V1 receptor. The most potent and V2-selective compound, N-[4-[2,6-diaza-6-[2-(4-methylpiperazinyl)-2-oxoethyl] -5- oxobicyclo[5.4.0]undeca-1(7),8,10-trien-2-yl]-carbonyl]pheny l][2-(4- methylphenyl)phenyl]-formamide (11b), exhibited IC50's of 2.9 nM for the V2 receptor and 200 nM for the V1 receptor, respectively. When administered orally to rat, 11b showed an approximately 18-fold increased urine volume in comparison with control rat.