Interference with PDK1-Akt survival signaling pathway by UCN-01 (7-hydroxystaurosporine)

Abstract

3-Phosphoinositide-dependent protein kinase-1 (PDK1) plays a central role in activating the AGC subfamily of protein kinases. In particular, PDK1 plays an important role in the regulation of Akt/PKB survival pathway by phosphorylating Akt on Thr 308. Here we show that UCN-01 (7-hydroxystaurosporine), a drug now in clinical trials and with a unique ®ngerprint pattern, induced dephosphorylation and inactivation of Akt, resulting in the turno of the survival signals and the induction of apoptosis. Further analysis revealed that UCN-01-mediated Akt inactivation was caused by inhibiting upstream Akt kinase PDK1 (IC 50 =33 nM) both in vitro and from cells, but not by suppressing Akt itself or phosphatidylinositide-3-OH kinase. UCN-01-induced PDK1 inhibition was also observed in in vivo murine and human tumor xenografts. Overexpression of active form of Akt diminished the cytotoxic eects of UCN-01, suggesting that UCN-01 may in part exert its cytotoxicity by inhibiting PDK1-Akt survival pathway. Because UCN-01 has already proved to have potent anti-tumor activity in vivo, PDK1-Akt survival pathway is a new, attractive target for cancer chemotherapy.

Potentiation of the anticancer effect of valproic acid, an antiepileptic agent with histone deacetylase inhibitory activity, by the kinase inhibitor Staurosporine or its clinically relevant analogue UCN-01

“…Staurosporine profoundly inhibited ERK1/2 activation and at the same time mediated phosphorylation of Akt in cultured thoracic cancer cells within the similar time interval. This effect of STP on Akt phosphorylation was surprising, given the fact that its closely related analogue UCN-01 suppressed Akt phosphorylation (Sato et al, 2002;Amornphimoltham et al, 2004;Kondapaka et al, 2004; and also our own observation). This was totally unexpected but very reproducible in many independent experiments with our cell lines and the molecular basis of this discrepancy was unclear.…”

“…Whereas less than 20% of cultured thoracic cancer cells exposed to either CC, STP or UCN-01 alone or TSA alone were apoptotic, 55% to more than 90% of cells treated with the respective drug combinations had undergone apoptosis ( Figure 1B). However, the supra-additive induction of apoptosis by UCN-01 or STP in combination with TSA may also be attributable to their function as negative modulators of Akt and MEK/ERK1/2 activation via inhibition of PDK1 (Sato et al, 2002), in addition to being PKC inhibitors. To further investigate the effect of STP or UCN-01 on MAPK ERK1/2 or Akt activation in cultured thoracic cancer cells, dose -response and time-course studies were performed on cells treated with STP (100 -400 nM) or UCN-01 (250 -1000 nM) and harvested at 1 or 24 h after the onset of drug exposure ( Figure 2).…”

“…Staurosporine and UCN-01, in addition to being PKC inhibitors, have also been shown to inhibit the kinase activity of PDK1, leading to the suppression of Akt phosphorylation and activation among other downstream targets (Sato et al, 2002). Sato et al (2004) have also demonstrated that PDK1 may directly phosphorylate and activate MEK and ERK1/2.…”

“…More recent studies have identified these compounds as kinase inhibitors with a broader range of molecular targets in vivo, including chk1/2 and PDK1, a kinase that act directly upstream of Akt and MAP/extracellular signal-regulated kinase (ERK) kinase (MEK) (Sato et al, 2002). Exposure of cultured cancer cells to UCN-01 (250 -1000 nM) resulted in complete abrogation of phosphorylation at Ser473 and Thr308 residues of Akt, both of which are essential for full activation of Akt kinase activity (Amornphimoltham et al, 2004;Kondapaka et al, 2004).…”

Clinical anticancer drug development: targeting the cyclin-dependent kinases

“…(Wang et al, 1996;Busby et al, 2000;Yu et al, 2002). UCN-01 also abrogates S-phase arrest in CHO cells treated with cisplatin, promoting the induction of apoptosis (Bunch and Eastman, 1997), and may target Akt signalling by inhibiting PDK1 (IC 50 ¼ 33 nM) (Sato et al, 2002). Frequent dosing is required to optimise the antitumour activity of UCN-01 , and 72 h of drug exposure is required to achieve growth inhibition.…”