Ó³»­´«Ã½

Idenya Bala-Mehta, a senior majoring in Molecular Biology and Biochemistry at Wesleyan University.

Metastasis is the leading cause of cancer-related deaths and the genes that control its progression are poorly understood, hindering therapy development. Unbiased loss-of-function CRISPR screens are a powerful way of discovering genetic drivers of aggressive metastatic behavior, providing a promising tool for the identification of novel therapeutic targets. Looking back, I realize that BSRP was about so much more than learning science. It was a summer research program oriented around the importance of community, friendship, and mentorship. This experience has not only prepared me for a future career in the life sciences, but also deepened my understanding of the values that will guide me both personally and professionally. In order to identify relevant metastatic dependencies associated with organ-specific microenvironments, CRISPR screens need to be executed in vivo and in a large pool of cancer cell lines. This approach is complex, time consuming and resource intensive. Moreover, its feasibility has never been formally assessed. Here, we outline two essential onboarding activities to optimize the screening model. First, we determine cancer cell metastatic propensity using a xenograft assay and a combined analysis of BLI and UMI sequencing. Next, we establish if each cancer model is amenable to genetic screening by examining the knockout efficacy of the Cas12 system in vitro, targeting CD47, a non-essential membrane protein. Visible morphological alterations were witnessed in the liver, kidneys, ovaries, and lungs from In vivo xenografts injected with the lung-derived cancer cell line, NCIH1915. BLI imaging also illustrated evidence of tumor expansion, suggesting that NCIH1915 is able to survive and colonize different microenvironments in-vivo. Additionally, Flow cytometry analysis on NICIH1915 cells stably expressing Cas12 and sgRNAs for CD47 indicate a profound shift in signal compared to a non-knockout control, demonstrating high Cas12 editing efficiency. Results from these quality control experiments suggest that NCIH1915 is ready to be used in the context of a complete screen. 

Project: Essential quality controls for large-scale, in-vivo CRISPR screens to study metastasis

Mentor: Andrea Clocchiatti, Cancer Program