Abstract P70: Exploring cancer stem cell regulators and its mechanisms using CRISPR-activation based screening

Abstract

Cancer stem cells (CSCs) potentially possess multiple mechanisms of resistance to therapies and contribute to cancer initiation and progression. However, effective therapies that target CSCs have not yet been developed, as underlying mechanisms of stemness maintance, differentiation induction and drug resistantce remain unclear. In this study, we aimed to establish a research platform for CSCs and to identify the underlying molecular mechanisms. We conducted a genome-wide gain-of-function screening using a CRISPR activation (CRISPR-a) library to explore candidate regulators of CSCs. We used the PEO1-NS cell line, which lacks CD133 expression and stemness that PEO1-SL, the counterpart maintaining stemness. A CRISPR-a library covering 23,430 human genes was introduced into the cells, allowing the expression of one gene per cell. Subsequently, cells in which CD133 expression was induced were isolated using a cell sorter, and the sgRNA sequences present in these cells were analyzed by Illumina sequencing. As a result, 13 candidate genes potentially involved in CSC regulation were identified. These includ genes related to energy metabolism, such as mitochondrial function and glycolysis, and were presumed to act through MYC regulation. For the validation study, we evaluated the GI50 of cisplatin in cells overexpressing each candidate gene. Several genes significantly reduced drug sensitivity. The candidate genes identified in this study may include key regulators involved in CSC maintenance and function via CD133 expression. Since changes in energy metabolism and mitochondrial dynamics are closely linked to CSC traits, our findings provide insight into the molecular basis regulating these processes. This study may offer a useful research platform for CSC research to identify regulatory mechanisms and develop novel therapiutic aveneus.