Histone deacetylase inhibitors (HDACis) are a promising new class of anticancer drugs. Two HDACis are licensed by the United States FDA for the treatment of advanced cutaneous T-cell lymphoma. Although clinical trials with HDACis as monotherapy in solid tumors have not been successful, HDACi shows promise in combination therapy for patients with recurrent metastatic lung cancer, suggesting that the full therapeutic potential of current HDACis will probably be best realized through a combination with other anticancer agents. However, currently there is little information to direct the clinical use of HDACi in combination with other agents. As colorectal cancer (CRC) remains an essentially incurable disease, it is critical to identify mechanism of HDACi resistance that can lead to combination strategies that increase HDACi therapeutic potential in CRC. The overall goal of this project is to uncover the HDAC mechanism responsible for the limited efficacy of HDACi treatment in colorectal cancer. There are 18 HDACs in humans and these different HDACs are not redundant in their function. In CRC, one of the most cancer relevant HDACs is HDAC2. We have found that in CRC cells, HDAC2 knockdown enhances the anti-tumor effect of SAHA, whereas overexpression of HDAC2 confers resistance to SAHA treatment. These findings strongly suggest that HDAC2 influences HDACi sensitivity and contributes to refractoriness to HDACi therapy in CRC cells. But the underlying mechanism by which HDAC2 works remains undefined. Clinical evidence suggests that HDACi refractoriness may involve mechanisms independent of deacetylase inhibition. Intriguingly, we have discovered that HDAC2 possesses a deacetylase-independent sumoylation-promoting activity and promotes sumoylation of Gsk3 (a key component of Wnt signaling). Based on our preliminary results, we hypothesize that HDAC2 sumoylation-promoting activity contributes to HDACi refractoriness in CRC cells by upregulating the Wnt/-catenin pathway through enhancing Gsk3 sumoylation. In this proposal, we will determine the role of the Wnt/-catenin pathway in HDAC2-mediated refractoriness in HDACi therapy. We will then validate the role of HDAC2 sumoylation-promoting activity in HDACi sensitivity of CRC cells. Finally, we will dissect how HDAC2 upregulates the Wnt/-catenin pathway through enhancing Gsk3 sumoylation. The successful completion of this proposed study could reveal a possible cause of insensitivity to HDACi treatment in CRC.