In vitro study | ZM 336372 shows 10-fold selectivity over B- Raf. ZM 336372 weakly inhibits SAPK2a/p38α and SAPK2b/p38β with IC50 of 2 μM, and is selective over 17 other protein kinases including PKA, PKC, AMPK, p42 MAPK, MKK1, SAPK1/JNK and CDK1 even at the concentration of up to 50 μM. ZM 336372 does not prevent constitutive as well as growth factor or phorbol ester induced activation of MKKl or p42 MAPK/ERK2. Moreover, ZM 336372 dose not reverse the phenotype of Ras- or Raf-transformed cell lines. ZM 336372 treatment induces >100 activation of c-Raf and the B- Raf isoform but it does not trigger any activation of MKKI or p42 MAPK/ERKP or induce any increase in the GTP-loading of Ras, suggesting that a feedback control loop exists by which Raf isoforms suppress their own activation such that inhibition is always counterbalanced by reactivation. ZM 336372-induced activation of c-Raf is not prevented by inhibition of the MAPK cascade, protein kinase C or phosphatidylinositide 3-kinase. ZM 336372 (1 μM) abolishes the upregulation of eNOS after treatment with hydrogen peroxide. ZM 336372 treatment in carcinoid tumor cells results in progressive phosphorylation of Raf-1 mitogen-activated protein kinase 1/2, and extracellular signal-regulated kinase 1/2, and causes a significant reduction of bioactive hormone levels as well as the transcription factor, human achaete-scute homologue-1. Furthermore ZM 336372 treatment leads to a marked suppression of cellular proliferation and induction of the cell cycle inhibitors p21 and p18. ZM 336372 inhibits the proliferation of pheochromocytoma cells, and suppresses NE vasoactive peptide production. ZM 336372 treatment in HepG2 induces the suppression of proliferation in a dose-dependent manner, suppression of hormone secretion and up-regulation of cell cycle inhibitors. ZM 336372 also induced tolerance in pancreatic adenocarcinoma cell lines by inhibiting glycogen synthesis of GSK-3β through phosphation of GSK-3β at Ser 9. The selectivity of ZM 336372 for C- Raf was 10 times that for B- Raf. ZM 336372 weakly inhibits both SAPK2a/p38α and SAPK2b/p38β with an IC50 of 2 μm and is more selective than PKA,PKC,AMPK at C- Raf even at concentrations as high as 50 μm, 17 other protein kinases, including p42 MAPK,MKK1,SAPK1/JNK, and Cdk1. ZM 336372 did not prevent growth factor or phorbol ester-induced MKKl or p42 MAPK/ERK2 activation. Furthermore, ZM 336372 did not reverse the phenotype of the Ras-or Raf-transformed cell lines. ZM 336372 treatment induced> 100 activation of the c-Raf and B- Raf subtypes, but it did not cause activation of MKKI or p42 MAPK/ERKP, nor did it induce an increase in GTP-loaded Ras, it was shown that a feedback control loop exists by the Raf isoforms inhibiting their own activation, so the inhibitory effect is always counteracted by reactivation. ZM 336372 induced c-Raf activation not blocked by MAPK cascade activation, inhibition of protein kinase C or phosphatidylinositol 3-kinase. ZM 336372 (1 μm) eliminates the upregulation of eNOS after hydrogen peroxide treatment. ZM 336372 treatment, leading to Raf-1, mitogen-activated protein kinase 1/2, and extracellular signal-regulated kinase 1/2 continuous phosphorylation in benign tumor cells, and caused a significant reduction in the levels of biologically active hormones, transcription factors, and human sessile congener -1. In addition, ZM 336372 treatment significantly inhibited cell proliferation and induced cell cycle inhibitors p21 and p18. ZM 336372 inhibits the proliferation of pheochromocytoma cells and inhibits NE vasoactive peptide production. ZM 336372 treatment induced dose-dependent inhibition of proliferation, inhibition of hormone secretion, and upregulation of cell cycle inhibitors in hepg2. In pancreatic cancer cell lines, ZM 336372 also inhibits glycogen synthase kinase -3 β by phosphorylation of GSK-3β on Ser 9, thereby inducing apoptosis. |