Identification of genetic determinants of ATR inhibitor sensitivity in ARID1A mutant ovarian clear cell carcinoma
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Ovarian clear cell carcinoma (OCCC) is a gynaecological cancer of unmet need characterized by ARID1A and PPP2R1A mutations. Prior work identified a synthetic lethality between ARID1A tumour suppressor mutations in OCCC and inhibition of the ATR kinase, information that has led to phase II proof-of-concept clinical trials assessing an ATR inhibitor (ATRi) in ARID1A mutant cancers. Using a genome-wide CRISPR-Cas9 mutagenesis screen in an ARID1A mutant OCCC tumour cell line and subsequent genome-wide CRISPR-Cas9 mutagenesis and CRISPR-dCas9 interference screens, I identify multiple protein phosphatase 2A (PP2A) subunit-coding genes as determinants of ATRi sensitivity. Analysis of a cohort of OCCCs indicated that over approximately one half possessed ARID1A mutations as well as mutations in the PP2A scaffolding subunit coding gene, PPP2R1A. CRISPR-prime editing of PPP2R1A demonstrated that cancer-associated hotspot p.R183 missense mutations cause ATRi sensitivity in OCCC cells, even in the presence of ARID1A mutations. The synthetic lethality between PPP2R1A p.R183 missense mutation and ATRi operated in both in vitro and in vivo and was characterized by a decrease in cells in replicating S phase of the cell cycle, an increase in cells entering mitosis with sub-4n genomic content and the accumulation of 53BP1 bodies. Mechanistically, this synthetic lethal effect was dependent upon the kinase WNK1, which itself exhibits increased phosphorylation in PPP2R1A mutant OCCC cells. Depletion of WNK1 restored the replicating S phase population normally depleted by ATRi and reversed ATRi sensitivity in PPP2R1A p.R183 mutant OCCC cells. TOV21G PPP2R1A p.R183 mutant cells were also observed to be more sensitive to ATRi than PPP2R1A wild-type cells in an in vivo model system of OCCC. Together with the data presented here, the co-occurrence of PPP2R1A and ARID1A mutations in OCCC suggests that in addition to ARID1A, PPP2R1A status should be assessed as a biomarker of ATRi response in on-going clinical trials.
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Institute of Cancer Research (University Of London)