Target identification with systematic CRISPR-based chemical-genetic profiling in human HAP1 cells

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We proposed to build and apply a genome-wide chemical genomic interaction screening platform in a human HAP1 cell line (a near-haploid cell line derived from a leukemic cell line), to functionally characterize compounds and elucidate their targets in human cells. Our unique and innovative approach will lead to the establishment of a reference human chemical-genetic interaction database for compounds with known modes of action, and the functional annotation of previously uncharacterized compounds from the RIKEN Natural Product Depository (NPDepo) or the University of Tokyo Drug Discovery Initiative (DDI) Core Library. We will biochemically validate roles of a collection of promising tool compounds that can be used to explore diverse aspects of cell biology and exploited as potential leads for drug development. In FY2022, we employed and set up a unique CRISPR-Cas9 gRNA library, the Toronto KnockOut library v3, covering ~18,000 human genes to enable genome-wide scale loss of function screens in mammalian cells. To select bioactive compounds to be examined in the human chemical genomics pipeline, we screened the RIKEN NPDepo and the University of Tokyo DDI Core Library for the compounds that possess strong and highly conserved target predictions based on our integrated yeast chemical-genetic platform.