Genome-wide and high-density CRISPR-Cas9 screens identify point mutations in PARP1 causing PARP inhibitor resistance

Pettitt, Stephen J.; Krastev, Dragomir B.; Brandsma, Inger; Dréan, Amy; Song, Feifei; Aleksandrov, Radoslav; Harrell, Maria I.; Menon, Malini; Brough, Rachel; Campbell, James; Frankum, Jessica; Ranes, Michael; Pemberton, Helen; Rafiq, Rumana; Fenwick, Kerry; Swain, Amanda; Guettler, Sebastian; Lee, Jungmin; Swisher, Elizabeth M.; Stoynov, Stoyno; Yusa, Kosuke; Ashworth, Alan; Lord, Christopher J.

doi:10.1038/s41467-018-03917-2

Cited by 342 publications

(289 citation statements)

References 57 publications

(95 reference statements)

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“…It has been well-established in previous studies that PARP1 itself is required for the cytotoxicity of PARPi. [13][14][15] In line with this, we found that PARP1 was ranked among the most significant of hits in our screen, as well ( Figure 1D).…”

Section: Genome-wide Crispr Screens To Identify Genetic Determinants supporting

confidence: 85%

Identification of regulators of poly-ADP-ribose polymerase (PARP) inhibitor response through complementary CRISPR knockout and activation screens

Clements

Hale

Tolman

et al. 2019

Preprint

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Inhibitors of poly-ADP-ribose polymerase 1 (PARPi) are highly effective in killing cells deficient in the homologous recombination (HR) DNA repair pathway, such as those lacking BRCA2. In light of this, PARPi have been utilized in recent years to treat BRCA2-mutant tumors, with many patients deriving impressive clinical benefit. However, positive response to PARPi is not universal, even among patients with HRdeficient tumors. Here, we present the results of three genome-wide CRISPR knockout and activation screens which provide an unbiased look at genetic determinants of PARPi response in wildtype or BRCA2-knockout cells. Strikingly, we reveal that depletion of the histone acetyltransferase TIP60, a top hit from our screens, robustly reverses the PARPi sensitivity caused by BRCA2 deficiency. Mechanistically, we show that TIP60 depletion rewires double strand break repair in BRCA2-deficient cells by promoting 53BP1 binding to double strand breaks to suppress end resection. Our work provides a comprehensive set of putative biomarkers that serve to better understand and predict PARPi response, and identifies a novel pathway of PARPi resistance in BRCA2-deficient cells.

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Section: Genome-wide Crispr Screens To Identify Genetic Determinants supporting

confidence: 85%

Identification of regulators of poly-ADP-ribose polymerase (PARP) inhibitor response through complementary CRISPR knockout and activation screens

Clements

Hale

Tolman

et al. 2019

Preprint

View full text Add to dashboard Cite

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“…However, similar to our findings, a recent report showed that CRISPR/Cas9-mediated PARP1 KO provided PARPi resistance in SUM149PT, but not MDA-MB-436, cells. The authors also demonstrated that PARP1 KO-induced PARPi resistance in SUM149PT cells was dependent on residual activity provided by the BRCA1-Δ11q protein (Pettitt et al, 2018). …”

Section: Discussionmentioning

confidence: 99%

BRCA1 Mutation-Specific Responses to 53BP1 Loss-Induced Homologous Recombination and PARP Inhibitor Resistance

et al. 2018

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SUMMARY BRCA1 functions in homologous recombination (HR) both up- and downstream of DNA end resection. However, in cells with 53BP1 gene knockout (KO), BRCA1 is dispensable for the initiation of resection, but whether BRCA1 activity is entirely redundant after end resection is unclear. Here, we found that 53bp1 KO rescued the embryonic viability of a Brea1ΔC/ΔC mouse model that harbors a stop codon in the coiled-coil domain. However, Brca1ΔC/ΔC;53bp1−/− mice were susceptible to tumor formation, lacked Rad51 foci, and were sensitive to PARP inhibitor (PARPi) treatment, indicative of suboptimal HR. Furthermore, BRCA1 mutant cancer cell lines were dependent on truncated BRCA1 proteins that retained the ability to interact with PALB2 for 53BP1 KO induced RAD51 foci and PARPi resistance. Our data suggest that the overall efficiency of 53BP1 loss of function induced HR may be BRCA1 mutation dependent. In the setting of 53BP1 KO, hypomorphic BRCA1 proteins are active downstream of end resection, promoting RAD51 loading and PARPi resistance.

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“…Molecular alterations leading to therapeutic resistance include, for example, small insertions/ deletions that result in frameshift mutations and synthesis of truncated proteins (e.g., inherited founder mutation BRCA1 185delAG ; ref. 83); secondary BRCA reversion mutations that reinstate HR proficiency through restoration of the open reading frame and BRCA reexpression (84); exon 11 deletion splice variants that produce truncated, hypomorphic proteins (85); or point mutations in PARP1 that alter PARP trapping (86). In addition to genomic alterations, epigenetic changes such as loss of BRCA1 promoter hypermethylation via BRCA1 locus fusion rearrangements, with subsequent BRCA1 reexpression, have also been described after acquired resistance to DNA-damaging drugs, including platinum or olaparib (87).…”

Section: Germline Brca-associated Tnbcmentioning

confidence: 99%

Insights into Molecular Classifications of Triple-Negative Breast Cancer: Improving Patient Selection for Treatment

2019

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Triple-negative breast cancer (TNBC) remains the most challenging breast cancer subtype to treat. To date, therapies directed to specifi c molecular targets have rarely achieved clinically meaningful improvements in outcomes of patients with TNBC, and chemotherapy remains the standard of care. Here, we seek to review the most recent efforts to classify TNBC based on the comprehensive profi ling of tumors for cellular composition and molecular features. Technologic advances allow for tumor characterization at ever-increasing depth, generating data that, if integrated with clinical-pathologic features, may help improve risk stratifi cation of patients, guide treatment decisions and surveillance, and help identify new targets for drug development.Signifi cance: TNBC is characterized by higher rates of relapse, greater metastatic potential, and shorter overall survival compared with other major breast cancer subtypes. The identifi cation of biomarkers that can help guide treatment decisions in TNBC remains a clinically unmet need. Understanding the mechanisms that drive resistance is key to the design of novel therapeutic strategies to help prevent the development of metastatic disease and, ultimately, to improve survival in this patient population. TNBC AND INTRINSIC BREAST CANCER SUBTYPESEarly transcriptomic profi ling of breast cancer using microarrays classifi ed tumors into fi ve intrinsic subtypes: luminal-A, luminal-B, HER2-enriched, basal-like, and a normal breast-like group ( 5,6 ). Although all intrinsic subtypes can be found within immunohistochemically defi ned triplenegative disease, basal-like tumors exhibit the greatest overlap with TNBC. Between 50% and 75% of TNBC have basal phenotype, and approximately 80% of basal-like tumors are ER-negative/HER2-negative ( Fig. 2 ; refs. 7, 8 ). Characterization Research.on August 1, 2020.

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Genome-wide and high-density CRISPR-Cas9 screens identify point mutations in PARP1 causing PARP inhibitor resistance

Cited by 342 publications

References 57 publications

Identification of regulators of poly-ADP-ribose polymerase (PARP) inhibitor response through complementary CRISPR knockout and activation screens

Identification of regulators of poly-ADP-ribose polymerase (PARP) inhibitor response through complementary CRISPR knockout and activation screens

BRCA1 Mutation-Specific Responses to 53BP1 Loss-Induced Homologous Recombination and PARP Inhibitor Resistance

Insights into Molecular Classifications of Triple-Negative Breast Cancer: Improving Patient Selection for Treatment

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