Abstract 3246: Pharmacologic targeting of the Hedgehog (HH) signal transduction pathway significantly suppresses growth of malignant pleural mesothelioma (MPM) and non-small cell lung cancer (NSCLC) cells in vitro.

Abstract

Abstract OBJECTIVE. The HH signal transduction pathway plays crucial roles in organ morphogenesis and is active during embryonic development but quiescent in terminally differentiated cells. Binding of HH ligands to PTCH1 relieves its inhibitory activity on the pathway activator SMO leading to pathway activation and upregulation of target gene expression including the transcription factor Gli1 that serves as marker of pathway activation. It is reactivated in many solid cancers including NSCLC as we have previously demonstrated and proven to be the key oncogenic driver in tumors such as skin basal cell carcinoma. The aim of this study is to determine if HH pathway regulates cell growth and survival of MPM cells and to evaluate the efficacy of pathway blockade using SMO antagonists (SMO inhibitor GDC0449 - GDC or the antifungal drug itraconazole -ITRA) or Gli inhibitors (GANT61 or the anti-leukemia drug arsenic trioxide - ATO) in suppressing the viability of not only NSCLC but also MPM cells. METHODS. Expression of main components of HH pathway in 8 MPM cells was assessed by qRT-PCR. Suppression of HH/SMO-mediated signaling by these pathway antagonists was determined by LightII cells. Selective knockdown of SMO to inhibit HH signaling was achieved by siRNA in 3 MPM cells H2373, Gardner and H2452. Cell viability following treatments with GDC, ITRA, GANT61or ATO were evaluated in a panel of 8 MPM cells and 8 NSCLC cells using MTT assay. Apoptosis was determined by annexinV/PE staining and flow cytometry. RESULTS. All MPM cells, similar to previously demonstrated in NSCLC cells, express varying levels of HH ligands, PTCH1, SMO with readily detectable Gli1 observed in 7/8 of MPM cells by qRT-PCR. SMO siRNA mediate 2- to 3-fold reduction of SMO and also of Gli1 gene expression by qRT-PCR, indicating significant HH pathway blockade. This was associated with 40±6% to 70±8% reduction of cell viability (p<0.01 versus non-target siRNA controls). All 4 pathway antagonists completely blocked HH activation in LightII reporter cells. Treating MPM or NSCLC cells with HH inhibitors resulted in 1.5- to 4-fold reduction of Gli1 expression. GANT61, ATO, ITRA and to a lesser degree GDC significantly suppressed viability of all cancer cells at low microM concentrations. More importantly, ITRA, ATO, GANT61 induce significant apoptosis (ranging from 30% to 60%) in representative MPM or NSCLC cells. CONCLUSIONS. HH signaling is active in MPM and regulates cell viability. ATO and ITRA are as effective as the prototypic SMO inhibitor GDC and the Gli inhibitor GANT61 in suppressing HH signaling in MPM and NSCLC cells. Pharmaceuticals that are FDA-approved for other indications but recently found to have anti-HH activity such as ATO or ITRA may be repurposed to treat HH-dependent MPM or NSCLC cells. Citation Format: Min You, Javier Varona Santos, David J. Robbins, Niramol Savaraj, Dao M. Nguyen. Pharmacologic targeting of the Hedgehog (HH) signal transduction pathway significantly suppresses growth of malignant pleural mesothelioma (MPM) and non-small cell lung cancer (NSCLC) cells in vitro. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3246. doi:10.1158/1538-7445.AM2013-3246