Error-free repair of DNA double-strand breaks (DSBs) is achieved by homologous recombination (HR), and BRCA1 is an important factor for this repair pathway. In the absence of BRCA1-mediated HR, the ...administration of PARP inhibitors induces synthetic lethality of tumour cells of patients with breast or ovarian cancers. Despite the benefit of this tailored therapy, drug resistance can occur by HR restoration. Genetic reversion of BRCA1-inactivating mutations can be the underlying mechanism of drug resistance, but this does not explain resistance in all cases. In particular, little is known about BRCA1-independent restoration of HR. Here we show that loss of REV7 (also known as MAD2L2) in mouse and human cell lines re-establishes CTIP-dependent end resection of DSBs in BRCA1-deficient cells, leading to HR restoration and PARP inhibitor resistance, which is reversed by ATM kinase inhibition. REV7 is recruited to DSBs in a manner dependent on the H2AX-MDC1-RNF8-RNF168-53BP1 chromatin pathway, and seems to block HR and promote end joining in addition to its regulatory role in DNA damage tolerance. Finally, we establish that REV7 blocks DSB resection to promote non-homologous end-joining during immunoglobulin class switch recombination. Our results reveal an unexpected crucial function of REV7 downstream of 53BP1 in coordinating pathological DSB repair pathway choices in BRCA1-deficient cells.
Celotno besedilo
Dostopno za:
DOBA, IJS, IZUM, KILJ, KISLJ, NUK, PILJ, PNG, SAZU, SBMB, SIK, UILJ, UKNU, UL, UM, UPUK
Tumours with specific DNA repair defects can be completely dependent on back-up DNA repair pathways for their survival. This dependence can be exploited therapeutically to induce synthetic lethality ...in tumour cells. For instance, homologous recombination (HR)-deficient tumours can be effectively targeted by DNA double-strand break-inducing agents. However, not all HR-defective tumours respond equally well to this type of therapy. Tumour cells may acquire resistance by invoking biochemical mechanisms that reduce drug action or by acquiring additional alterations in DNA damage response pathways. A thorough understanding of these processes is important for predicting treatment response and for the development of novel treatment strategies that prevent the emergence of therapy-resistant tumours.
Celotno besedilo
Dostopno za:
DOBA, IJS, IZUM, KILJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
Mutations in homologous recombination (HR) genes BRCA1 and BRCA2 predispose to tumorigenesis. HR-deficient cancers are hypersensitive to Poly (ADP ribose)-polymerase (PARP) inhibitors, but can ...acquire resistance and relapse. Mechanistic understanding how PARP inhibition induces cytotoxicity in HR-deficient cancer cells is incomplete. Here we find PARP inhibition to compromise replication fork stability in HR-deficient cancer cells, leading to mitotic DNA damage and consequent chromatin bridges and lagging chromosomes in anaphase, frequently leading to cytokinesis failure, multinucleation and cell death. PARP-inhibitor-induced multinucleated cells fail clonogenic outgrowth, and high percentages of multinucleated cells are found in vivo in remnants of PARP inhibitor-treated Brca2
;p53
and Brca1
;p53
mammary mouse tumours, suggesting that mitotic progression promotes PARP-inhibitor-induced cell death. Indeed, enforced mitotic bypass through EMI1 depletion abrogates PARP-inhibitor-induced cytotoxicity. These findings provide insight into the cytotoxic effects of PARP inhibition, and point at combination therapies to potentiate PARP inhibitor treatment of HR-deficient tumours.
Hereditary breast cancers are frequently caused by germline
BRCA1 mutations. The
BRCA1
C61G
mutation in the BRCA1 RING domain is a common pathogenic missense variant, which reduces BRCA1/BARD1 ...heterodimerization and abrogates its ubiquitin ligase activity. To investigate the role of BRCA1 RING function in tumor suppression and therapy response, we introduced the
Brca1
C61G
mutation in a conditional mouse model for BRCA1-associated breast cancer. In contrast to BRCA1-deficient mammary carcinomas, tumors carrying the
Brca1
C61G
mutation responded poorly to platinum drugs and PARP inhibition and rapidly developed resistance while retaining the
Brca1
C61G
mutation. These findings point to hypomorphic activity of the BRCA1-C61G protein that, although unable to prevent tumor development, affects response to therapy.
► BRCA1 RING function is dispensable for therapy resistance ► BRCA1 RING mutant tumors have an altered DNA damage response ► Cisplatin-resistant tumors do not show genetic reversion of the
Brca1
C61G
mutation ► Functional assays for HRD may be more predictive than genomic classifiers
PARP inhibition is synthetic lethal with defective DNA repair via homologous recombination. Phase I and II clinical trials show that PARP inhibitors are effective at well-tolerated doses and have ...antitumor activity for BRCA1- and BRCA2-associated cancers. However, not all patients respond equally well and tumors may eventually become resistant. Thus far, the only resistance mechanism that has been found in human tumors is genetic reversion that corrects or bypasses the original BRCA1- or BRCA2-inactivating mutation. However, data from fundamental and preclinical research suggest that resistance to PARP inhibitors may be induced by additional mechanisms involving hypomorphic activity of mutant BRCA1 alleles, upregulation of drug efflux pumps, and rewiring of the DNA damage response. Preclinical models will be instrumental to develop methods for adequate patient stratification, as well as treatment strategies that prevent or counteract resistance to PARP inhibitors.
Triple-negative breast cancer (TNBC) is a subtype of breast cancer with limited treatment options and poor clinical prognosis. Inhibitors of transcriptional CDKs are currently under thorough ...investigation for application in the treatment of multiple cancer types, including breast cancer. These studies have raised interest in combining these inhibitors, including CDK12/13 inhibitor THZ531, with a variety of other anti-cancer agents. However, the full scope of these potential synergistic interactions of transcriptional CDK inhibitors with kinase inhibitors has not been systematically investigated. Moreover, the mechanisms behind these previously described synergistic interactions remain largely elusive.
Kinase inhibitor combination screenings were performed to identify kinase inhibitors that synergize with CDK7 inhibitor THZ1 and CDK12/13 inhibitor THZ531 in TNBC cell lines. CRISPR-Cas9 knockout screening and transcriptomic evaluation of resistant versus sensitive cell lines were performed to identify genes critical for THZ531 resistance. RNA sequencing analysis after treatment with individual and combined synergistic treatments was performed to gain further insights into the mechanism of this synergy. Kinase inhibitor screening in combination with visualization of ABCG2-substrate pheophorbide A was used to identify kinase inhibitors that inhibit ABCG2. Multiple transcriptional CDK inhibitors were evaluated to extend the significance of the found mechanism to other transcriptional CDK inhibitors.
We show that a very high number of tyrosine kinase inhibitors synergize with the CDK12/13 inhibitor THZ531. Yet, we identified the multidrug transporter ABCG2 as key determinant of THZ531 resistance in TNBC cells. Mechanistically, we demonstrate that most synergistic kinase inhibitors block ABCG2 function, thereby sensitizing cells to transcriptional CDK inhibitors, including THZ531. Accordingly, these kinase inhibitors potentiate the effects of THZ531, disrupting gene expression and increasing intronic polyadenylation.
Overall, this study demonstrates the critical role of ABCG2 in limiting the efficacy of transcriptional CDK inhibitors and identifies multiple kinase inhibitors that disrupt ABCG2 transporter function and thereby synergize with these CDK inhibitors. These findings therefore further facilitate the development of new (combination) therapies targeting transcriptional CDKs and highlight the importance of evaluating the role of ABC transporters in synergistic drug-drug interactions in general.
The tumor suppressor protein BRCA2 is a key component of the homologous recombination pathway of DNA repair, acting as the loader of RAD51 recombinase at sites of double-strand breaks. Here we show ...that BRCA2 associates with telomeres during the S and G2 phases of the cell cycle and facilitates the loading of RAD51 onto telomeres. Conditional deletion of Brca2 and inhibition of Rad51 in mouse embryonic fibroblasts (MEFs), but not inactivation of Brca1, led to shortening of telomeres and accumulation of fragmented telomeric signals--a hallmark of telomere fragility that is associated with replication defects. These findings suggest that BRCA2-mediated homologous recombination reactions contribute to the maintenance of telomere length by facilitating telomere replication and imply that BRCA2 has an essential role in maintaining telomere integrity during unchallenged cell proliferation. Mouse mammary tumors that lacked Brca2 accumulated telomere dysfunction-induced foci. Human breast tumors in which BRCA2 was mutated had shorter telomeres than those in which BRCA1 was mutated, suggesting that the genomic instability in BRCA2-deficient tumors was due in part to telomere dysfunction.
Celotno besedilo
Dostopno za:
DOBA, IJS, IZUM, KILJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
Using genome-wide radiogenetic profiling, we functionally dissect vulnerabilities of cancer cells to ionizing radiation (IR). We identify ERCC6L2 as a major determinant of IR response, together with ...classical DNA damage response genes and members of the recently identified shieldin and CTC1-STN1-TEN1 (CST) complexes. We show that ERCC6L2 contributes to non-homologous end joining (NHEJ), and it may exert this function through interactions with SFPQ. In addition to causing radiosensitivity, ERCC6L2 loss restores DNA end resection and partially rescues homologous recombination (HR) in BRCA1-deficient cells. As a consequence, ERCC6L2 deficiency confers resistance to poly (ADP-ribose) polymerase (PARP) inhibition in tumors deficient for both BRCA1 and p53. Moreover, we show that ERCC6L2 mutations are found in human tumors and correlate with a better overall survival in patients treated with radiotherapy (RT); this finding suggests that ERCC6L2 is a predictive biomarker of RT response.
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•Radiogenetic profiling identifies ERCC6L2 as a major determinant of IR response•Loss of ERCC6L2 restores HR and causes PARPi resistance in BRCA1-deficient cells•ERCC6L2 contributes to NHEJ, possibly through its interaction with SFPQ•Patients with ERCC6L2-mutated UCEC show better survival upon RT
Francica et al. identify ERCC6L2 as an accessory NHEJ gene by using radiogenetic profiling in haploid cells. Loss of ERCC6L2 partially restores HR in BRCA1-deficient cells, and ERCC6L2 may be a useful predictive biomarker of radiotherapy response.
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