PROJECT SUMMARY Large scale transcriptional analysis with RNA-sequencing has revealed the abundant expression of repetitive element RNAs in cancers compared to normal tissues. Moreover, many of these repeat RNAs are capable of retrotransposition through reverse transcription (RT), which leads to alterations in the cancer genome. Our preclinical cell line and mouse models have demonstrated the ability of these endogenous RT to be blocked by nucleoside reverse transcriptase inhibitors (NRTIs) commonly used for viral infections. Interestingly, we have found anti-cancer effects of NRTIs in multiple colorectal cancer (CRC) cell lines grown in 3D tumor spheres and in mouse xenografts in vivo. This led to the initial of a Phase II clinical trial of the NRTI 3TC in metastatic CRC patients where pre- and post-treatment biopsies are being obtained for correlative science research. The Ting lab has expertise in cell line and animal models to study NRTI effects in CRC and the sequencing and RNA in situ hybridization methods to quantify repeat elements in these systems. The Greenbaum lab has developed computational tools to characterize repeat RNA expression and their effects on innate immunity. The Park lab has pioneered the tools to study DNA retrotransposition events in cancer genomes. The combined expertise and tools generated to study repeat RNAs by the Ting, Greenbaum, and Park labs uniquely positions us to execute the goals of this project. We plan to: 1) Understand the effects of 3TC on cellular response and repeat RNA expression in CRC model systems 2) Characterize the effect of 3TC on repeat retrotransposition in xenograft models 3) Evaluate repeat RNA expression, retrotransposition, and the tumor immune microenvironment from patient biopsies obtained from CRC patients treated with 3TC