Small molecule inhibitors of protein kinase D inhibit cancer cell growth through inhibition of growth factor-stimulated proliferation and induction of apoptosis.

3256 Protein kinase D (PKD) lies at the crossroads of the DAG and PKC signalling network, which is frequently deregulated in cancer. PKD has been shown to play a role in many of the biological processes that lead to tumor growth, including cell proliferation, survival, epithelial-to-mesenchymal transition and invasion. Thus PKD represents a potential therapeutic target for intervention. siRNA was used to dissect the role of specific PKD isoforms (PKD1, PKD2 and PKD3) in cancer cells. siRNA-mediated depletion of PKD1, PKD2 and PKD3 expression (≈70%) revealed a role for PKD1 in cell proliferation and PKD2 in cellular resistance to apoptosis. Small molecule inhibitors of PKD were then identified by high throughput screening and characterized with respect to their potential as anti-cancer agents. A series of ATP-competitive PKD inhibitors was developed with good physicochemical and in vitro ADME properties. A lead compound was selected (CRT0059359) which had potent (sub nanomolar IC 50 ) and selective activity against PKD in biochemical assays. CRT0059359 also caused potent (low micromolar IC 50 ) inhibition of PKD activity in a variety of cancer cell lines in vitro (e.g. PANC-1, A549, MDA-MB-435), as determined by modulation of phorbol ester-stimulated PKD-Ser916 autophosphorylation. In addition, treatment of PANC-1 cells in vitro with CRT0059359 inhibited growth-factor-stimulated proliferation to basal levels, and caused a 12-fold increase in Caspase 3/7 activity, a biomarker of apoptosis. Pharmacokinetic studies in mice demonstrated that plasma levels of CRT0059359 in excess of those required to inhibit PKD in vitro are achievable; in vivo efficacy studies are being initiated. Collectively, these data validate PKD as a target for therapeutic intervention in cancer and demonstrate the utility of potent PKD inhibitors as anti-cancer agents.