Abstract 748: Novel selective orally bioavailable small molecule PAK4 allosteric modulators display antitumor activity and induce apoptosis in vitro and in vivo
Authors: Senapedis William , Baloglu Erkan , Pursell Natalie , Crochiere Marsha , McCauley Dilara , Ellis Joel , Kashyap Trinayan , Klebanov Boris , Carlson Robert , Kalid Ori , Kauffman Michael , Shacham Sharon , Landesman Yosef 
Published: May 2015

P21-Activated Kinase 4 (PAK4) is a member of the PAK family of proteins that regulate cell survival, cell division and apoptosis. The six members of the PAK family are divided into two groups; Group I (PAK1, 2, 3) and Group II (PAK4, 5, 6), based upon their sequence homology and regulatory mechanisms. PAK4 is a member of the group II family of PAKs and is amplified or mutated in many cancer types. PAK4 is also a key downstream effector of the K-Ras pathway.

Methods: Flow cytometry, CellTiter AQueous One assay (MTS) and colony formation assays were used to determine compound effects on cell cycle distribution, proliferation and viability. Immunoblots were used to measure effects of compounds on protein steady state levels and phosphorylation. The breast cancer cell line, MDA-MB-468, and the mantle cell lymphoma line, Z-138, were used in xenograft models in mice to test the in vivo efficacy of these compounds.

Results: We have identified selective, orally bioavailable small molecule PAK4 allosteric modulators, which demonstrated anti-tumor activity in a variety of cancer cell lines (IC50 values = 0.005 - 1 μM). Treatment of cancer cells with these small molecules resulted in the reduction of PAK4 steady state levels and reduced phosphorylation of key growth signaling proteins such as Akt, ERK1/2, p90RSK, β-catenin, cofilin, p21, and cyclin D1. These allosteric modulators induced apoptosis through the activation of caspases 3 and 8 and subsequent cleavage of PARP. After 48 hours of compound treatment, it was found that the cancer cells were arrested at the G1 and G2 phases of the cell cycle. In addition, we have observed anti-tumor activity against Z-138 and MDA-MB-468 xenografts in mice at a daily oral dose of 10 mg/kg in the absence of any clinical signs of toxicity up to 200 mg/kg daily dose.

Conclusions: PAK4 represents a novel anti-cancer target as a major downstream effector of Ras oncogene. We have identified selective, orally-bioavailable small molecule PAK4 allosteric modulators with anti-tumor activity both in vitro and in vivo. These compounds inactivate PAK4 by directly inducing PAK4 destabilization. This represents a novel mechanism of the protein kinase inactivation involving degradation of PAK4 rather than direct inhibition of the kinase activity. Moreover, these allosteric modulators induce tumor cell growth arrest and apoptosis. Based on the in vitro and in vivo activity, these PAK4 allosteric modulators show promising results for the treatment of a wide variety of cancers.

Citation Format: William Senapedis, Erkan Baloglu, Natalie Pursell, Marsha Crochiere, Dilara McCauley, Joel Ellis, Trinayan Kashyap, Boris Klebanov, Robert Carlson, Ori Kalid, Michael Kauffman, Sharon Shacham, Yosef Landesman. Novel selective orally bioavailable small molecule PAK4 allosteric modulators display antitumor activity and induce apoptosis in vitro and in vivo. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 748. doi:10.1158/1538-7445.AM2014-748