From the Cyclooxygenase-2 Inhibitor Celecoxib to a Novel Class of 3-Phosphoinositide-Dependent Protein Kinase-1 Inhibitors

Abstract

The blockade of Akt activation through the inhibition of 3-phosphoinositide-dependent kinase-1 (PDK-1) represents a major signaling mechanism whereby celecoxib mediates apoptosis. Celecoxib, however, is a weak PDK-1 inhibitor (IC 50 , 48 M), requiring at least 30 M to exhibit discernable effects on the growth of tumor cells in vitro. Here, we report the structure-based optimization of celecoxib to develop PDK-1 inhibitors with greater potency in enzyme inhibition and growth inhibition. Kinetics of PDK-1 inhibition by celecoxib with respect to ATP suggest that celecoxib derivatives inhibit PDK-1 by competing with ATP for binding, a mechanism reminiscent to that of many kinase inhibitors. Structureactivity analysis together with molecular modeling was used to generate compounds that were tested for their potency in inhibiting PDK-1 kinase activity and in inducing apoptosis in PC-3 prostate cancer cells. Docking of potent compounds into the ATP-binding site of PDK-1 was performed for lead optimization, leading to two compounds, OSU-03012 and OSU-03013, with IC 50 values in PDK-1 inhibition and apoptosis induction in the low M range. Exposure of PC-3 cells to these agents led to Akt dephosphorylation and inhibition of p70 S6 kinase activity. Moreover, overexpression of constitutively active forms of PDK-1 and Akt partially protected OSU-03012-induced apoptosis. Screening in a panel of 60 cell lines and more extensive testing in PC-3 cells indicated that the mean concentration for total growth inhibition was ϳ3 M for both agents. Considering the conserved role of PDK-1/Akt signaling in promoting tumorigenesis, these celecoxib analogs are of translational relevance for cancer prevention and therapy.

Celecoxib derivatives induce apoptosis via the disruption of mitochondrial membrane potential and activation of caspase 9

“…We next compared these inhibition patterns with the recently published IC 50 for PDK1 by the derivatives. 32 (Fig. 3b) paralleled IC 50 of PDK1 (rsϭ0.89, pϽ0.01 and rsϭ0.71, pϽ0.05, respectively).…”

“…8,31,32 LY294002, a PI3K inhibitor; PD98059, a MAPK/ Erk kinase inhibitor; Ac-DEVD-CHO (DEVD) and Ac-IETD-CHO (IETD) reversible inhibitors of Caspases 3 and 8, respectively, and Z-LE(OMe)HD(OMe)-FMK (LEHD), an irreversible inhibitor of caspase 9, were purchased from Alexis Corporation (San Diego, CA). Cyclosporin A (CsA) was obtained from Sigma Chemical Co. (St. Louis, MO).…”

“…8,[31][32][33] We observed that the exposure of the premalignant and malignant epithelial cell lines to these derivatives leads to both the accumulation of cells in the G 2 /M phase of the cell cycle and the induction of apoptosis with much greater potency than celecoxib. Since derivative-induced apoptosis in the human PC-3 prostate cancer cell line involved PI3K/PDK1/Akt and MAPK/ Erk1/2 signaling pathways, we were interested in determining whether these pathways were also involved in malignant human oral epithelial cell line apoptosis.…”

“…8,29,30 This has led to the design of celecoxib derivatives that optimize the apoptosis-inducing potency, separate from the COX inhibiting activity of celecoxib. 8,[31][32][33] Recent studies with the first and second generation derivatives of celecoxib in the PC-3 human prostate cell line suggests that apoptosis is mediated through the blockade of Akt activation by derivative inhibition of 3-phosphoinositide dependent kinase-1 (PDK1). 32,33 These data suggests that celecoxib induced apoptosis may proceed through multiple mitochondrial dependent signaling pathways involving phosphatidylinositol 3-Kinase (PI3K)/PDK1/Akt, mitogen-activated protein kinase/extracellular signal-regulated kinase (MAPK/ Erk), caspase-9/Apaf and other yet undefined upstream signals.…”

“…8,[31][32][33] Recent studies with the first and second generation derivatives of celecoxib in the PC-3 human prostate cell line suggests that apoptosis is mediated through the blockade of Akt activation by derivative inhibition of 3-phosphoinositide dependent kinase-1 (PDK1). 32,33 These data suggests that celecoxib induced apoptosis may proceed through multiple mitochondrial dependent signaling pathways involving phosphatidylinositol 3-Kinase (PI3K)/PDK1/Akt, mitogen-activated protein kinase/extracellular signal-regulated kinase (MAPK/ Erk), caspase-9/Apaf and other yet undefined upstream signals. Multiple targets for inducing apoptosis via the mitochondrial pathway makes these compounds clinically attractive for treating or sensitizing different cancer cell types.…”