Delineating the role of 5’ untranslated regions in prostate cancer clonality

About This Grant

ABSTRACT Prostate cancer is a highly heterogeneous disease even within individual patients, which is one of the primary reasons it is the second leading cause of cancer-related deaths in men in the United States. While much is known about the genetic and epigenetic drivers of clonal heterogeneity, nothing is known about how untranslated regions modulate clonal growth within a population of prostate cancer cells. To address this question, we conducted an unbiased 5’ UTR tiled CRISPR screen across 283 genes implicated in mRNA translation, DNA transcription, kinase activity, protein degradation, RNA binding, chromatin binding, and cancer. Remarkably, 1,569 out of 27,104 single guide RNAs increased or decreased clonal growth in our pooled screen, demonstrating that there are regulons of 5’ UTR regions that can differentially expand or contract clones within a population of cancer cells. Conducting two additional screens, we observed that there are differential clonal dependencies on specific regions of 5’ UTRs that are prostate cancer subtype or treatment dependent. Lastly, validating 41 individual sgRNAs, we found that most of the mutations we introduced (35/41 = 85.4%) caused no change in mRNA levels, but profoundly impacted protein abundance. Thus, our unbiased preliminary work has uncovered a critical role for distinct 5’ UTR regions that modulate protein synthesis and clonal growth in prostate cancer. Based on this work, we hypothesize that 5’ UTR elements are a new layer of gene regulation that control clonal growth dynamics in prostate cancer. The major goals of this study are to: 1) determine at nucleotide resolution how distinct 5’ UTR regions titrate their protein levels to influence prostate cancer clonal growth; and 2) elucidate how 5’ UTRs shape clonal growth within prostate cancer subtypes and in response to therapeutics. To this end, we will conduct the following Aims: Aim1: Delineate how oncogenic 5’ UTR elements modulate mRNA translation and clonal growth in prostate cancer; Aim 2: Determine how 5’ UTR elements impact human prostate cancer clonal expansion and the therapeutic response. This work will require murine models because they more accurately replicate the tumor microenvironment which will have a substantial impact on how we translate these findings into people. Completion of this study will provide the first mechanistic insights how 5’ UTRs regulate clonal growth in prostate cancer. At a mechanistic level, we will determine how functional regions fold, regulate translation initiation rates, and interface with trans-acting factors. At a patient level, we will delineate the subtype-specific dependencies of 5’ UTR clonal drivers and inhibitors across the spectrum of human mCRPC in vitro and in vivo. At a therapeutic level, we will uncover if 5’ UTR regions of KDM5 genes can be genetically or therapeutically targeted to enhance the efficacy of enzalutamide. Together, this work has the potential to delineate a new mechanistic basis for clonal growth in prostate cancer with therapeutic implications.