Abstract
PKB/AKT kinase is the core component of the PI3K/AKT signaling pathway which is frequently hyperactivated in human cancers. We designed and synthesized a series of 2-pyrimidyl-5-amidothiazole compounds based on the ATP binding site of AKT, and the most potent compound DC120 was identified to inhibit AKT activity in vitro with EC50 of 153nM by a FRET-based Z'-Lyte assay. Then the anti-tumor effect of DC120 was tested on human CNE2 and MDA-MB-453 cell lines and CNE2 xenograft model. The results showed that DC120 could obviously inhibit the proliferation of CNE2 and MDA-MB-453 cells via induction of apoptosis, with the evidence of increasing sub-G1 and AnnexinV-positive cells, characteristic morphologic changes of apoptosis in the nucleus, and cleaved caspase-3. Further study showed that MDA-MB-453 cells transfected with constitutively activated AKT1 were more sensitive to DC120, while CNE2 cells with knock-down of AKT1 expression by shRNA were more resistant to DC120. More importantly, DC120 partially attenuated the phosphorylation levels of FKHR, FKHRL1, GSK-β and mTOR in a dose-dependent and time-dependent fashion, and led to an increase in the nuclear accumulation of exogenous FKHR in cancer cells. And DC120 at 20 mg/kg/qd inhibited the CNE2 xenograft tumor growth with the T/C value of 38.1%, accompanied by increasing TUNEL-positve cells in tumor sample. Additionally, DC120 induced a feedback loop to activate MAPKs pathway, and treatment with MEK inhibitor U0126 and DC120 synergistically induced cancer cell apoptosis. These data provide validation for the development of DC120 to treat cancers displaying elevated levels of AKT.
- Received December 18, 2011.
- Revision received April 26, 2012.
- Accepted April 26, 2012.
- The American Society for Pharmacology and Experimental Therapeutics