Abstract 1: Mcl-1 downregulation plays a crucial role in the synergistic anticancer activities between PI-3 kinase inhibitors and histone deacetylase inhibitors in primary NSCLC tumors in vitro and in vivo

Proceedings: AACR 102nd Annual Meeting 2011‐‐ Apr 2‐6, 2011; Orlando, FL Interactions between the PI-3 kinase inhibitors and histone deacetylase inhibitors (HDACi) were examined in vivo using SCID mice bearing patient-derived primary NSCLC xenografts which contain the “hot spot” mutation PIK3CA \[E545K (G1633A)\] (K-Ras and EGFR wild-type), and in vitro using the primary cell cultures derived directly from the in vivo xenografts. Co-treatment of primary cell cultures to marginally toxic concentrations of GDC-0941 (1.0 uM) and HDACi LBH589 (2.5 nM) induced a striking increase of apoptosis, as evidenced by Annex V positive staining, cytochrome c release and caspase 3 cleavage. Similar effects were observed using other HDACis (e.g. SAHA or MS275). The combination was associated with profound Akt inactivation, GSK-3β activation and Mcl-1 attenuation without affecting other Bcl-2 family members. Although LBH589 induced p21CIP1/WAF1 expression, GDC-0941 blocked LBH589-mediated p21CIP1/WAF1 accumulation. Ectopic expression of the constitutively active (myristolated) Akt, dominant negative GSK-3β (K85A), Mcl-1, or p21CIP1/WAF1 significantly blocked apoptosis induced by this combination. Inactivation of GSK-3β by either shRNA knockdown or its pharmacological inhibitor markedly diminished GDC-0941/LBH589-induced cell death, and abrogated the Mcl-1 and p21CIP1/WAF1 downregulation seen with the combination, indicating that GSK-3β modulation is the process upstream of Mcl-1 and p21. However, overexpression of either Mcl-1 or p21CIP1/WAF1 did not demonstrate an interaction with each other under the co-treatment, suggesting an independent functional cascade for each Mcl-1 and p21CIP1/WAF1. The downregulation of Mcl-1 and p21CIP1/WAF1 was reversed by proteasome inhibitor, indicating a proteasome-dependent degradation of Mcl-1 and p21CIP1/WAF1. Lastly, the combination of GDC-0941/LBH589 in vivo inhibited the growth of primary tumor xenografts more efficiently than either agent alone. In vivo downregulation of Mcl-1 and p21CIP1/WAF1 was also observed, consistent with the modulation of molecular events presented in vitro. Taken together, our results demonstrate that GDC-0941 potentiates HDACi-mediated apoptosis through PI3K/Akt/GSK-3β pathway, and subsequently attenuates Mcl-1 and abrogates p21CIP1/WAF1 induction in primary NSCLC xenografts in vivo and the primary cell cultures in vitro, thereby suggesting a novel therapeutic approach for NSCLC. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1. doi:10.1158/1538-7445.AM2011-1