RESUMO
The twenty-three Fanconi anemia (FA) proteins cooperate in the FA/BRCA pathway to repair DNA interstrand cross-links (ICLs). The cell division cycle and apoptosis regulator 1 (CCAR1) protein is also a regulator of ICL repair, though its possible function in the FA/BRCA pathway remains unknown. Here, we demonstrate that CCAR1 plays a unique upstream role in the FA/BRCA pathway and is required for FANCA protein expression in human cells. Interestingly, CCAR1 co-immunoprecipitates with FANCA pre-mRNA and is required for FANCA mRNA processing. Loss of CCAR1 results in retention of a poison exon in the FANCA transcript, thereby leading to reduced FANCA protein expression. A unique domain of CCAR1, the EF hand domain, is required for interaction with the U2AF heterodimer of the spliceosome and for excision of the poison exon. Taken together, CCAR1 is a splicing modulator required for normal splicing of the FANCA mRNA and other mRNAs involved in various cellular pathways.
Assuntos
Proteínas Reguladoras de Apoptose , Proteínas de Ciclo Celular , Proteína do Grupo de Complementação A da Anemia de Fanconi , Anemia de Fanconi , Splicing de RNA , Fator de Processamento U2AF , Humanos , Proteína BRCA1/metabolismo , Proteína BRCA1/genética , Proteína BRCA2/metabolismo , Proteína BRCA2/genética , Reparo do DNA , Endodesoxirribonucleases , Éxons , Anemia de Fanconi/genética , Anemia de Fanconi/metabolismo , Proteína do Grupo de Complementação A da Anemia de Fanconi/genética , Proteína do Grupo de Complementação A da Anemia de Fanconi/metabolismo , Células HEK293 , Células HeLa , Ligação Proteica , Precursores de RNA/metabolismo , Precursores de RNA/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Transdução de Sinais , Spliceossomos/metabolismo , Spliceossomos/genética , Fator de Processamento U2AF/metabolismo , Fator de Processamento U2AF/genética , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Proteínas Reguladoras de Apoptose/genética , Proteínas Reguladoras de Apoptose/metabolismoRESUMO
Recent studies suggest that PARP inhibitors and POLQ inhibitors confer synthetic lethality in BRCA1-deficient tumors by accumulation of single-stranded DNA (ssDNA) gaps at replication forks. Loss of USP1, a deubiquitinating enzyme, is also synthetic lethal with BRCA1 deficiency, and USP1 inhibitors are now undergoing clinical development for these cancers. Here, we show that USP1 inhibitors also promote the accumulation of ssDNA gaps during replication in BRCA1-deficient cells, and this phenotype correlates with the drug sensitivity. USP1 inhibition increased monoubiquitinated PCNA at replication forks, mediated by the ubiquitin ligase RAD18, and knockdown of RAD18 caused USP1 inhibitor resistance and suppression of ssDNA gaps. USP1 inhibition overcame PARP inhibitor resistance in a BRCA1-mutated xenograft model and induced ssDNA gaps. Furthermore, USP1 inhibition was synergistic with PARP and POLQ inhibition in BRCA1-mutant cells, with enhanced ssDNA gap accumulation. Finally, in patient-derived ovarian tumor organoids, sensitivity to USP1 inhibition alone or in combination correlated with the accumulation of ssDNA gaps. Assessment of ssDNA gaps in ovarian tumor organoids therefore represents a rapid approach for predicting response to USP1 inhibition in ongoing clinical trials.
RESUMO
DNA damage response (DDR) mechanisms are critical to maintenance of overall genomic stability, and their dysfunction can contribute to oncogenesis. Significant advances in our understanding of DDR pathways have raised the possibility of developing therapies that exploit these processes. In this expert-driven consensus review, we examine mechanisms of response to DNA damage, progress in development of DDR inhibitors in IDH-wild-type glioblastoma and IDH-mutant gliomas, and other important considerations such as biomarker development, preclinical models, combination therapies, mechanisms of resistance and clinical trial design considerations.
Assuntos
Neoplasias Encefálicas , Dano ao DNA , Humanos , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/terapia , Neoplasias Encefálicas/patologia , Pesquisa Translacional Biomédica , Animais , Reparo do DNA , Glioma/genética , Glioma/terapia , Glioma/patologia , Consenso , MutaçãoRESUMO
PURPOSE: The multicenter, open-label, randomized phase 2 NCI-9944 study (NCT02595892) demonstrated that addition of ATR inhibitor (ATRi) berzosertib to gemcitabine increased progression-free survival (PFS) compared to gemcitabine alone (hazard ratio [HR]=0.57, one-sided log-rank P = .044, which met the one-sided significance level of 0.1 used for sample size calculation). METHODS: We report here the final overall survival (OS) analysis and biomarker correlations (ATM expression by immunohistochemistry, mutational signature 3 and a genomic biomarker of replication stress) along with post-hoc exploratory analyses to adjust for crossover from gemcitabine to gemcitabine/berzosertib. RESULTS: At the data cutoff of January 27, 2023 (>30 months of additional follow-up from the primary analysis), median OS was 59.4 weeks with gemcitabine/berzosertib versus 43.0 weeks with gemcitabine alone (HR 0.79, 90% CI 0.52 to 1.2, one-sided log-rank P = .18). An OS benefit with addition of berzosertib to gemcitabine was suggested in patients stratified into the platinum-free interval ≤3 months (N = 26) subgroup (HR, 0.48, 90% CI 0.22 to 1.01, one-sided log-rank P =.04) and in patients with ATM-negative/low (N = 24) tumors (HR, 0.50, 90% CI 0.23 to 1.08, one-sided log-rank P = .06). CONCLUSION: The results of this follow-up analysis continue to support the promise of combined gemcitabine/ATRi therapy in platinum resistant ovarian cancer, an active area of investigation with several ongoing clinical trials.