Next Generation CRISPR Phenotypic Screens for Identification of Novel Therapeutic Targets in Rhabdomyosarcoma

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Eleanor Chen, MD, PhD,  University of Washington
Recipient of the: $50,000 Dr. Richard and Valerie Aronsohn Memorial Research Award

Rhabdomyosarcoma is the most common pediatric soft tissue sarcoma. Survival rate remains poor for patients with relapsed or metastatic disease. There remains an urgent need for novel therapeutic targets. Our previous work with histone deacetylases (HDACs), key epigenetic modifiers in normal and neoplastic cells, has demonstrated the essential role of HDAC3 in suppressing RMS tumor growth. Through gene expression and proteomic profiling of HDAC3-deficient RMS cells, we have identified candidate genes functioning downstream of or interacting with HDAC3. To characterize the candidate genes, we have developed an effective high-throughput CRISPR-mediated gene targeting strategy, which allows for efficient phenotypic readout from targeted gene disruption both in vitro and in vivo. A set of 8 gene targets with oncogenic function in RMS has been identified from initial targeting of 50+ candidate genes linked to HDAC3 function. We proposed to complete targeting of the identified gene target set in a panel of RMS cell lines and characterize their effects on human xenografts in vivo. As the HDACs may have redundant functions in RMS, we have also developed a multiplex CRISPR-based gene targeting strategy, which allows for identification of redundant key HDACs essential for RMS tumor growth and progression by simultaneously targeting 2 class I and class II HDAC genes of all combinations. In all, we propose to take advantage of novel CRISPR genome-engineering platforms to interrogate functions of HDACs and HDAC3-linked candidate genes in RMS cell lines and xenograft mouse models. Novel therapeutic targets identified from this study will likely improve survival of RMS patients with advanced disease.