Abstract Precision medicine depends critically on developing treatment strategies that can selectively target cancer cells with minimal adverse effects. Identifying unique transcriptional regulators of oncogenic signaling, and targeting cancer-cell-specific enhancers that may be active only in specific tumor cell lineages, could provide the necessary high specificity, but a scarcity of functionally validated enhancers in cancer cells presents a significant hurdle to this strategy. We address this limitation by carrying out large-scale functional screens for pro-growth enhancers using highly multiplexed CRISPR-based perturbation and sequencing in multiple cancer cell lines. We used this strategy to identify 488 pro-growth enhancers in a colorectal cancer cell line and 22 functional enhancers for the MYC and MYB key oncogenes in an additional nine cancer cell lines. The majority of pro-growth enhancers are accessible and presumably active only in cancer cells but not in normal tissues, and are enriched for elements associated with poor prognosis in colorectal cancer. We further identify master transcriptional regulators and demonstrate that the cancer pro-growth enhancers are modulated by lineage-specific transcription factors acting downstream of growth signaling pathways. Our results uncover context-specific, potentially actionable pro-growth enhancers from cancer cells, yielding insight into altered oncogenic transcription and revealing potential therapeutic targets for cancer treatment.

Discovery and Functional Characterization of Pro-growth Enhancers in Human Cancer Cells