Ovarian Cancer-associated Mutations Disable Catalytic Activity of CDK12, a Kinase That Promotes Homologous Recombination Repair and Resistance to Cisplatin and Poly(ADP-ribose) Polymerase Inhibitors

Joshi, Poorval; Sutor, Shari L.; Huntoon, Catherine J.; Karnitz, Larry M.

doi:10.1074/jbc.m114.551143

Cited by 175 publications

(194 citation statements)

References 33 publications

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“…The DDR and the AR pathways have both been linked with mechanisms of cisplatin resistance in ovarian cancer cells [41,42,44,58,59]. It is conceivable that these two pathways work concertedly to reduce the susceptibility of ovarian cancer cells to cisplatin.…”

Section: Discussionmentioning

confidence: 99%

NRF2 inhibition causes repression of ATM and ATR expression leading to aberrant DNA Damage Response

Khalil¹,

Deeni²

2015

Biodiscovery

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Nuclear factor (erythroid-derived 2)-like 2 (NRF2) is a master regulator of the antioxidant response (AR) pathway and functions as a transcription factor for basal and oxidative stress-induced expression of a battery of detoxification enzymes and cytoprotective genes. Recent evidence has also demonstrated a role of NRF2 in driving resistance of numerous cancers to chemotherapeutic agents. ATM and ATR are serine/threonine kinases that are activated following DNA damage and function as central components of DNA Damage Response (DDR) pathway. Activities of these kinases cause cell cycle arrest and activate DNA repair signals leading to cytoprotection against genotoxic agents. In this study, we elucidated the roles of ATM-and ATR-dependent DDR and NRF2-mediated AR pathways in promoting cytoprotection following cisplatin challenge in ovarian cancer cell line models. We also determined whether these pathways were inter-dependent for full activation following genotoxic insults and as such demonstrated crosstalk in their signaling mechanism to elicit cytoprotective pathways. Treatment with cisplatin caused NRF2 induction and generation of reactive oxygen species (ROS) that caused cytotoxicity in ovarian cancer cells. This was attenuated by the ROS scavenger N-Acetyl cysteine, implicating NRF2 function in cytoprotection against cisplatin. Treatment with retinoic acid (RA) caused down regulation of NRF2, disruption of AR pathway, significant accumulation of ROS and enhanced cisplatin cytotoxicity. Interestingly, RA treatment also led to repression of total ATM and ATR proteins and aberrant DDR activation following cisplatin challenge. In order to determine whether the RA induced ATM and ATR repression was dependent on NRF2 inhibition, we silenced NRF2 using SiRNA. This caused transcriptional repression of both ATM and ATR expression as determined by their promoter driven luciferase assays. Thus, NRF2 inhibition led to DDR suppression by down-regulating ATM and ATR that led to enhanced cytotoxicity. These findings demonstrate mechanism of crosstalk between the AR and the DDR pathways and extend the scope of NRF2 in promoting cancer therapeutic resistance.

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Section: Discussionmentioning

confidence: 99%

NRF2 inhibition causes repression of ATM and ATR expression leading to aberrant DNA Damage Response

Khalil¹,

Deeni²

2015

Biodiscovery

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“…Effect of CDK12 knockdown in cellular models has been consistently associated with HRD (3,5,7,8). HRD in breast and ovarian cancers is mostly related to BRCA1/2 inactivation (9-11).…”

Section: Cdk12 Td-plus Phenotype and Genomic Hrd Hallmarksmentioning

confidence: 99%

“…CDK12 encodes a serine/threonine kinase involved in the regulation of RNA polymerase II (RNA pol II) and mRNA processing (2)(3)(4). CDK12 attracted particular attention as cells inactivated for the CDK12 display hypersensitivity to DNA-damaging agents and to PARP1/2 inhibitors (5,6). This effect of CDK12 inactivation was associated with homologous recombination (HR) deficiency (HRD) due to decreased expression observed for some HR genes, such as BRCA1, FANCI, or FANCD2 (3,5,7,8).…”

Section: Introductionmentioning

confidence: 99%

“…CDK12 attracted particular attention as cells inactivated for the CDK12 display hypersensitivity to DNA-damaging agents and to PARP1/2 inhibitors (5,6). This effect of CDK12 inactivation was associated with homologous recombination (HR) deficiency (HRD) due to decreased expression observed for some HR genes, such as BRCA1, FANCI, or FANCD2 (3,5,7,8). HRD due to inactivation of BRCA1 or BRCA2, the two major genes implicated in ovarian cancer, leads to the large-scale chromosomal instability, readily observed in nearly half of HGS-OvCa (9)(10)(11).…”

Section: Introductionmentioning

confidence: 99%

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Ovarian Cancers Harboring Inactivating Mutations in CDK12 Display a Distinct Genomic Instability Pattern Characterized by Large Tandem Duplications

et al. 2016

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CDK12 is a recurrently mutated gene in serous ovarian carcinoma, whose downregulation is associated with impaired expression of DNA damage repair genes and subsequent hypersensitivity to DNA-damaging agents and PARP1/2 inhibitors. In this study, we investigated the genomic landscape associated with CDK12 inactivation in patients with serous ovarian carcinoma. We show that CDK12 loss was consistently associated with a particular genomic instability pattern characterized by hundreds of tandem duplications of up to 10 megabases (Mb) in size. Tandem duplications were characterized by a bimodal ($0.3 and $3 Mb) size distribution and overlapping microhomology at the breakpoints.This genomic instability, denoted as the CDK12 TD-plus phenotype, is remarkably distinct from other alteration patterns described in breast and ovarian cancers. The CDK12 TD-plus phenotype was associated with a greater than 10% gain in genomic content and occurred at a 3% to 4% rate in The Cancer Genome Atlas-derived and in-house cohorts of patients with serous ovarian carcinoma. Moreover, CDK12-inactivating mutations together with the TD-plus phenotype were also observed in prostate cancers. Our finding provides new insight toward deciphering the function of CDK12 in genome maintenance and oncogenesis. Cancer Res; 76(7); 1882-91. Ó2016 AACR.

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“…6 Additionally, we have recently demonstrated that dinaciclib is the most potent inhibitor of CDK12 known thus far, and therefore a transcriptional regulator of HR genes. [7][8][9][10] We comprehensively examined the combination of dinaciclib and PARP inhibitors, such as veliparib and olaparib, in preclinical models of triple-negative breast cancer (TNBC), including BRCA wild-type cells, BRCA-mutated cells, and patient-derived xenograft (PDX) models of primary and acquired resistance to PARP inhibition. 10 First, pathway analyses demonstrated that the genes downregulated by dinaciclib in BRCA wild-type TNBC cells were significantly enriched for those involved in HR repair and DNA damage-sensing, including BRCA1 and RAD51.…”

mentioning

confidence: 99%

Sensitizing HR-proficient cancers to PARP inhibitors

Lim

Johnson

Geyer

et al. 2017

Molecular & Cellular Oncology

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Homologous recombination (HR) is a major DNA repair pathway that is deficient in a small subset of cancers. The best characterized of these are cancers associated with a germline or acquired mutation in the breast cancer 1 and 2 tumor suppressor genes (BRCA1 and BRCA2, respectively). HR-deficient cells rely on non-HR DNA repair pathways that are dependent on poly (ADP-ribose) polymerase (PARP), so that inhibition of PARP is an Achilles' heel in these cancers. PARP inhibitors have emerged as an effective therapeutic strategy in HR-deficient tumors, 1 and PARP inhibitors are currently FDA approved for use in BRCA1/2 germline mutation associated advanced ovarian and breast cancers.In spite of the promise that this class of therapies holds, it is increasingly apparent that tumors may possess intrinsic and/or acquired resistance mechanisms involving restoration of HR, presenting a pressing clinical problem.2-5 Thus, strategies targeting HR may potentially confer or restore PARP inhibitor sensitivity. One such strategy involves the drug dinaciclib, an inhibitor of cyclin-dependent kinases (CDKs) 1, 2, 5, and 9. 6 Additionally, we have recently demonstrated that dinaciclib is the most potent inhibitor of CDK12 known thus far, and therefore a transcriptional regulator of HR genes. 7-10We comprehensively examined the combination of dinaciclib and PARP inhibitors, such as veliparib and olaparib, in preclinical models of triple-negative breast cancer (TNBC), including BRCA wild-type cells, BRCA-mutated cells, and patient-derived xenograft (PDX) models of primary and acquired resistance to PARP inhibition. 10 First, pathway analyses demonstrated that the genes downregulated by dinaciclib in BRCA wild-type TNBC cells were significantly enriched for those involved in HR repair and DNA damage-sensing, including BRCA1 and RAD51. Consequently, dinaciclib disrupted HR repair in cell-based assays, and the IC50 of veliparib was consistently reduced in the presence of dinaciclib in a panel of BRCA wild-type TNBC cell lines. These results were phenocopied by CRISPR/Cas9-mediated knockout of CDK12 in TNBC cells. Importantly, dinaciclib had minimal effects on the cell cycle of breast cancer cells. These data point to a pivotal role of dinaciclib in disrupting HR, mediated through inhibition of CDK12.In light of this, we hypothesized that BRCA-mutated TNBC with acquired PARP inhibitor resistance might be resensitized to PARP inhibition with dinaciclib. Using a BRCA1-mutated cell line with acquired resistance after exposure to increasing PARP inhibitor concentrations in vitro, and an in vivo TNBC PDX model derived from a BRCA2 carrier, who had progressed on olaparib and cisplatin, we demonstrated that the combined dinaciclib and veliparib therapy restored tumor growth inhibition.We next evaluated this strategy in BRCA1-mutated TNBC cell lines with de novo PARP inhibitor resistance, and a TNBC PDX model derived from an 185delAG BRCA1 carrier, whose tumor demonstrated primary resistance to olaparib. Despite BRCA1 mutation, these cell lines...

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Ovarian Cancer-associated Mutations Disable Catalytic Activity of CDK12, a Kinase That Promotes Homologous Recombination Repair and Resistance to Cisplatin and Poly(ADP-ribose) Polymerase Inhibitors

Cited by 175 publications

References 33 publications

NRF2 inhibition causes repression of ATM and ATR expression leading to aberrant DNA Damage Response

NRF2 inhibition causes repression of ATM and ATR expression leading to aberrant DNA Damage Response

Ovarian Cancers Harboring Inactivating Mutations in CDK12 Display a Distinct Genomic Instability Pattern Characterized by Large Tandem Duplications

Sensitizing HR-proficient cancers to PARP inhibitors

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