Sphingosine-1-phosphate and CD44v6 regulate phospho-STAT3 induced drug resistance
Development of resistance to chemotherapeutic agents, such as OXAliplatin (OXA) is a major obstacle in the treatment of colorectal cancers (CRCs).The overall goal of this study is to define the mechanisms of drug resistance to begin to understand the key mechanisms of OXA-resistance underlying the clinical behavior of CRC. The signaling cell surface receptor, CD44 variant 6 (HyaR), which is specifically overexpressed by epithelial tumor cells correlates with resistance to drug-induced apoptosis. Accumulating evidence suggests that the lipid-raft micro domain consists of a dynamic association between cholesterol and sphingolipid that regulates drug resistance. Sphingosine-kinase 1 (SK1)-stimulated sphingosine-1-phosphate (S1P) interacts with specific cell surface G protein-coupled receptors (S1PR (1-5)) and functions as an oncogene in tumorigenesis. Our preliminary studies suggest that there are two endocytic caveolin (CAV1) or clathrin routes for HyaR recruitment in OXA-resistant versus OXA-sensitive cells and also demonstrate that OXA preferentially regulates HyaR recruitment through the CAV1 endocytic pathway. Our preliminary studies also indicate that SK1 induces expression of HyaR. In addition, we also found that S1PR2 is mainly responsible for HyaR-mediated resistance to OXA-induced apoptosis. Therefore, we have focused the studies proposed in this application on the lipid raft affiliated S1PR- HyaR complex that mediates OXA-resistance through activation of STAT3-BCL2 pathway. There are no pharmacological inhibitors for HyaR. Thus, we highlight a novel gene silencing opportunity to target HyaR and S1PR2 protein. Our preliminary data also demonstrate that blockade of S1PR2 and HyaR inhibited growth of subcutaneous tumors derived from OXA-resistant HT29R cells. Therefore, one goal of the proposed studies is to target SK1 and as well the v6 sites of HyaR using SK1shRNA, or HyaRshRNA, or their combination to overcome drug resistance and suppress tumor growth. We propose one aim. Aim 1: Test the prediction that SK1-induced S1P/S1PR2 interaction promotes CRC resistance to OXA by promoting HyaR- STAT3-BCL2 signaling via CAV1-microdomain. Studies to address Aim 1A will delineate the role of SK1 induced S1PR2-HyaR signaling complex in CAV1-lipid raft mechanism resulting in STAT3 activation, BCL2 expression and OXA resistance. Studies to address the Aim 1B will further provide proof-of concept using a mouse spleen --> liver metastases model that mimics the human CRC that inhibition of the SK1/S1PR2/HyaR pathway by inhibiting SK1 activity and/or HyaR expression/activity will sensitize the CRC tumors to low dose of OXA induced apoptosis. Taken together, the results of our proposed studies, will take us from an initial finding of CAV1-dependent HyaRS1PR2 regulation of OXA-resistance to testing in a preclinical model. Understanding the role of CRC cellresponses involved in resistance to apoptosis is needed for design of future therapeutics using HyaR-shRNA, and/or SK1shRNA plus low-dose OXA.