The PI3K pathway plays an important role in regulating cancer cell proliferation, growth, survival and metabolism. The serine/threonine kinase Akt, a central node of the PI3K pathway, is frequently activated in a significant proportion of human solid tumors, making Akt an attractive target for therapeutic intervention. Akt inhibitors, which target the signaling pathway downstream of some of the most important growth factors and their tyrosine kinase receptors such as HER2, IGF1R, EGFR, and c-MET, should have broad utility against a wide array of human tumors. Utilizing a traditional compound screening approach, we have identified Akt inhibitors that block both the activation and kinase activity of the enzyme. Medicinal chemistry exploration of the structure activity relationships of these leads resulted in potent and selective Akt compounds including MK-2206. MK-2206 inhibits Akt isozymes 1, 2, and 3 with in vitro IC50 values of 8, 12, and 65 nM, respectively. MK-2206 is an allosteric inhibitor requiring the presence of the Pleckstrin homology domain for activity, and therefore is highly selective against Akt exhibiting no inhibitory activities against over 250 protein kinases when tested at 1\#956;M. In multiple cancer cell lines, MK-2206 inhibited Akt1 kinase activity (IC50 \#8776; 20 nM), and blocked Akt2 and Akt3 activities with 2- to 6-fold less potency. MK-2206 inhibited auto-phosphorylation of both Akt T308 and S473, as well as prevented Akt-mediated phosphorylation of down-stream signaling molecules, including TSC2, PRAS40 and ribosomal S6 proteins. MK-2206 exhibited potent anti-proliferative activity against a number of cancer cell lines harboring one or more of the following genetic defects: 1) gene amplification resulting in constitutive activation of upstream receptor tyrosine kinases such as HER2, 2) a PI3KCA activating mutation, 3) inactivation of tumor suppressor PTEN, and 4) amplification and mutation of Akt itself. In addition, activation of the Ras pathway tended to predict non-response to MK-2206. In nude mice bearing A2780 ovarian cancer xenografts, a single oral dose of MK-2206 at 240 mg/kg resulted in sustained inhibition (>70 %) of phospho-Akt1/2 (T308 and S473) in the tumors. In the same tumor model, MK-2206 inhibited tumor growth by \#8776; 60% when administrated orally at 240 mg/kg per day three times a week. We have further evaluated the effect of MK-2206 in combination with chemotherapeutic agents or receptor tyrosine kinase inhibitors. Additive or synergistic interactions were seen when MK-2206 was combined with cytotoxic agents with different mode of action including topoisomerase inhibitors (doxorubicin and camptothecin), anti-metabolites (gemcitabine and 5-FU), anti-microtubule (docetaxcel), and DNA cross linkers (carboplatin). Akt inhibition sensitized tumor cells in induction of apoptosis by these agents. In vivo, MK-2206 enhanced anti-tumor efficacy of docetaxel, gemcitabine and carboplatin in nude mice xenograft tumor models. In vitro synergistic interaction was also observed when MK-2206 was combined with EGFR inhibitor erlotinib in non-small cell lung cancer cell lines or combined with dual EGFR-Her2 inhibitor lapatinib in breast cancer cell lines. Co-treatment of MK-2206 enhanced anti-tumor activity of erlotinib or lapatinib and led to tumor regression in lung and breast cancer models. The biochemical mechanism underlying synergistic interactions between MK-2206 and these agents was investigated. MK-2206 is generally well-tolerated in preclinical studies. Mechanism-related pharmacodynamic changes in blood glucose and insulin were seen in animals treated with MK-2206. Both hyperglycemia and hyperinsulinemia were mild and transient, recovering to baseline upon completion of the treatment. These preclinical results support further clinical development of MK-2206 in humans. MK-2206 is currently being studies in two Phase I trials, one in healthy volunteers (HV) and one in cancer patients. In the first-in-human HV trial, twenty-four healthy, male subjects participated in this Phase I randomized, double-blind, placebo-controlled, sequential-panel, multiple-period, rising single oral dose study. Eight (8) subjects were assigned to each of 3 panels (Panels A, B, and C) where in each treatment period in a panel the same 6 subjects received MK-2206 and 2 subjects received placebo after an overnight fast. The volunteers were administered single doses from 0.25 to 100 mg and blood samples were collected predose and at prespecified postdose time points for pharmacokinetic and pharmacodynamic (whole blood inhibition of phospho Akt) assays. Single doses of MK-2206, up to 100 mg, were found to be generally well tolerated. No serious clinical or laboratory adverse experience was reported. The most commonly reported adverse experiences were headache, common cold, and diarrhea. One subject was discontinued from the study due to the clinical adverse experience of blurry vision which resolved. There were no clinically meaningful changes in laboratory safety tests or ECG evaluations. No clinically significant hyperglycemia or hyperinsulinemia was seen in these subjects. Preliminary pharmacokinetic results found that orally administered MK-2206 was readily absorbed with a median Tmax of 6 to 8 hours. The median half-life was 55 to 78 hours. AUC0-\#8734; and Cmax displayed dose proportional behavior from 2-mg to 100-mg. Preliminary pharmacodynamic results found that single doses of 40-, 80- and 100-mg MK-2206 inhibited Akt in whole blood to a greater extent than placebo. Maximum Akt inhibition occurred at 6 hours postdose for both the 80- and 100-mg doses with mean plasma concentrations of >65 nM. There was evidence of Akt inhibition from 2 through 24 hours. In conclusion, MK-2206 was generally well tolerated following single dose administration to healthy subjects. MK-2206 displays dose proportional pharmacokinetics with clear evidence of Akt inhibition. Clinical development of MK-2206 in cancer patients is ongoing with a focus on tumors harboring PI3K pathway activation events.

Citation Information: In: Proc Am Assoc Cancer Res; 2009 Apr 18-22; Denver, CO. Philadelphia (PA): AACR; 2009. Abstract nr DDT01-1.

100th AACR Annual Meeting-- Apr 18-22, 2009; Denver, CO