Selective inhibition of constitutively elevated oncogenes provides an opportunity to hinder the proliferation of malignant cells while minimizing toxicity to the host. The cell-survival-promoting oncoprotein Akt has attracted much attention because it is abnormally activated in many human malignancies. Here, we report the discovery of a novel phospho-Akt (pAkt) inhibitor, SR13668, which was developed and improved from a naturally occurring anticancer agent, indole-3-carbinol (I3C), as a result of lead-based rational drug design using computational modeling. We demonstrate that SR13668 exhibits potent oral antitumor activity against both estrogen-dependent (MCF-7) and -independent (MDA-MB-231 and MDA-MB-468) breast, prostate (PC-3), and drug-resistant ovarian (SKOV-3) cancer xenografts in nude mouse models and is noncytotoxic and nonmutagenic in Ames tests. Western blotting showed that SR13668 inhibits pAkt in a variety of cancer cells in a time- and dose-dependent manner, and SR13668's growth inhibitory activity is positively correlated with pAkt expression in the cell lines. In our in vivo pharmacodynamic studies of pAkt inhibition, groups of tumor-bearing (MB-468) mice were given a single oral dose (50 mg/kg) of SR13668 or vehicle and sacrificed at various timepoints. SR13668 significantly inhibited pAkt in treated tumors after 24 h of oral treatment, as determined by immunohistochemistry (IHC) using a pAkt (Ser473) antibody. TUNEL analysis of the SR13668 treated tumors revealed that tumor growth inhibition was mediated through a significant increase in apoptosis throughout the tumor. Scoring tumor sections for proliferation after BrdU IHC showed that proliferation of treated tumors was reduced 40% compared to controls. Cell cycle analysis showed that SR13668-treated cancer cells exhibited significantly increased accumulation in G0/G1 phase and decreased in S-phase. Because pAkt has been linked to VEGF expression and signaling, the anti-angiogenic effect of SR13668 was also assessed by the chorioallantoic membrane assay. SR13668 significantly inhibited blood vessel growth in this chick embryo model. Inhibition of PI3K signaling is known to result in hyperinsulinemia and glucose intolerance. Wortmannin has resulted in an exaggerated increase in the blood glucose level. In vivo assessment of glucose metabolism in mice showed no adverse effects on the fasting glucose level after 14 days of oral treatment with SR13668 at doses 10 times higher than were needed for antitumor activity. Results from screening a broad selection of kinase targets indicated that SR13668 is very specific. Collectively, our data show that SR13668 represents a promising approach for cancer therapy, acting on the constitutively activated oncoprotein Akt that is causally involved in the malignant progression of human cancers and apparently sparing normal and non-neoplastic tissues.

[Proc Amer Assoc Cancer Res, Volume 45, 2004]