ENAapplication/xmlftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR111/024/SRR11185024/SRR11185024.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR111/027/SRR11185027/SRR11185027.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR111/025/SRR11185025/SRR11185025.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR111/026/SRR11185026/SRR11185026.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR111/022/SRR11185022/SRR11185022.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR111/023/SRR11185023/SRR11185023.fastq.gzprimaryOK200GenomicsMIThttps://www.ebi.ac.uk/ena/browser/view/PRJNA608860Mus musculusApproximately 20-30% of human lung adenocarcinomas (LUAD) harbor loss-of-function (LOF) mutations in Kelch-like ECH Associated-Protein 1 (KEAP1), which lead to hyperactivation of the antioxidant program downstream from the nuclear factor, erythroid 2-like 2 (NRF2) transcription factor and correlates with poor prognosis1–3. We previously showed that Keap1 mutation accelerates KRAS-driven LUAD and produces a marked dependency on glutaminolysis4. To extend the investigation of genetic dependencies in the context of Keap1 mutation, we performed a druggable genome CRISPR-Cas9 screen in Keap1-mutant cells. This analysis uncovered a Keap1-mutant-specific dependency on solute carrier family 33 member 1 (Slc33a1), an endomembrane-associated protein with roles in autophagy regulation5, as well as a series of functionally-related genes implicated in the unfolded protein response. Targeted genetic and biochemical experiments using mouse and human Keap1-mutant tumor lines, as well as preclinical genetically-engineered mouse models (GEMMs) of LUAD, validate Slc33a1 as a robust Keap1-mutant-specific dependency. Furthermore, unbiased genome-wide CRISPR screening identified additional genes related to Slc33a1 dependency. Overall, our study provides a strong rationale for stratification of patients harboring KEAP1-mutant or NRF2-hyperactivated tumors as likely responders to targeted SLC33A1 inhibition and underscores the value of integrating functional genetic approaches with GEMMs to identify and validate genotype-specific therapeutic targets. Overall design: Keap1-mutant (n = 2), WT (n = 2), and WT plus sgSlc33a1 transduced (n = 2) samples were jointly analyzed to derive a murine signature of Slc33a1-mutant gene expression changesENAKLHL19, dKEAP1, AT2R1, KEAP1, AT-1, SPG42., INRF2, INrf2, dkeap1, CG3962, AT1BR, AI788741, AG2S, keap1, Acatn, mKIAA0132, results, Genotypes, AT1, Inrf2, AT2R1A, Genogroups, DmelCG3962, AT2R1B, Genogroup, HAT1R, KIAA0132, ACATN, AT1R, AI315656, AGTR1A, AGTR1B, dKeap1, AT1AR, AT1B, CCHLND, D630022N01RikKLHL19, dKEAP1, AT2R1, KEAP1, AT-1, SPG42., INRF2, INrf2, dkeap1, CG3962, AT1BR, AI788741, AG2S, keap1, Acatn, mKIAA0132, results, Genotypes, AT1, Inrf2, AT2R1A, Genogroups, DmelCG3962, AT2R1B, Genogroup, HAT1R, KIAA0132, ACATN, AT1R, AI315656, AGTR1A, AGTR1B, dKeap1, AT1AR, AT1B, CCHLND, D630022N01RikfalseKeap1 deficiency results in a genotype-specific dependency on Slc33a1Keap1 deficiency results in a genotype-specific dependency on Slc33a12025-09-242020-04-17PRJNA608860GSE14594510090