Abstract 2805: In vitro characterization of AMG 511, a potent and selective class I PI3K inhibitor for the treatment of cancer
Authors: Caenepeel Sean , Zhang Nancy , Wang Ling , Norman Mark H. , Burgess Terri , Radinsky Robert , Kendall Rick , Freeman Daniel , Hughes Paul E. 
Published: August 2012

Abstract
The PI3K signaling pathway is frequently activated in cancer and has been implicated in many aspects of tumor growth and survival. Inhibition of this pathway represents a potential therapeutic path for the treatment of cancer. This study evaluated the in vitro characteristics of AMG 511, a potent and selective pan class I PI3K inhibitor exhibiting IC50 values of 8, 11, 2, and 6 nM against the PI3K β, α, β, and ≤ isoforms respectively. AMG 511 was shown to be inactive against members of the closely related phosphoinositide 3 kinase related kinases (PIKK) family of kinases and did not inhibit mTOR, hVPS34, PI4Kα or PI4Kα in-vitro (IC50 values &gt; 1 μM). In addition, AMG 511 was inactive against a majority of protein kinases (372) in the human kinome as measured by in-vitro binding assays. AMG 511 inhibited PI3K pathway signaling in U87 MG glioblastoma cells as determined by dose-dependent reduction in AKT S473 phosphorylation (IC50 = 4 nM). AKT inhibition resulted in a concomitant reduction in PRAS40 phosphorylation (IC50 = 23 nM), a downstream effector of AKT. Reduced phosphorylation of mTORC1 substrates p70S6K (IC50 = 30 nM) and S6 (IC50 = 70 nM) but not 4EBP1 (T37/46), was also detected in U87 MG cells, suggesting that upstream blockade of PI3K pathway signaling with AMG 511 treatment leads to a selective reduction in downstream mTORC1 activity. Given the well documented role of mTORC1 in cap-dependent translation we profiled AMG 511 in a methionine-analog incorporation assay in U87 MG cells. However, no significant inhibition of bulk translation was observed following treatment with AMG 511 in U87 MG cells. Treatment of U87 MG cells with AMG 511 revealed a pronounced G1 arrest with a concurrent reduction in BrdU+ cells, detectable within 8 hours of treatment. This anti-proliferative effect was fully reversible by 18 hours following washout. In line with these anti-proliferative effects, reduced cyclin D1 levels and elevated p27 levels were detected within 4 hours of treatment. Minimal cell killing effects were detected with AMG 511 treatment in U87 MG cells as measured by induction of cleaved caspase-3 and DNA content &lt; 2N. AMG 511 was profiled across a large panel of tumor cell lines encompassing several tumor types including breast and lung. A majority of the cell lines tested exhibited sensitivity to AMG 511 with a subset exhibiting evidence for cell death upon treatment with AMG 511. Breast cancer cell lines harboring activating mutations in PI3Kα, loss of PTEN, or amplification of Her2 tended to show greater sensitivity to AMG 511 treatment. In conclusion, AMG 511 is a potent and selective pan class I PI3K inhibitor, capable of inhibiting PI3K signaling and inducing robust anti-proliferative effects via a G1 arrest in many tumor cell lines, with evidence of cell killing in a subset of lines.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2805. doi:1538-7445.AM2012-2805