RESUMEN
Non-small cell lung cancers (NSCLCs) harboring KEAP1 mutations are often resistant to immunotherapy. Here, we show that KEAP1 targets EMSY for ubiquitin-mediated degradation to regulate homologous recombination repair (HRR) and anti-tumor immunity. Loss of KEAP1 in NSCLC induces stabilization of EMSY, producing a BRCAness phenotype, i.e., HRR defects and sensitivity to PARP inhibitors. Defective HRR contributes to a high tumor mutational burden that, in turn, is expected to prompt an innate immune response. Notably, EMSY accumulation suppresses the type I interferon response and impairs innate immune signaling, fostering cancer immune evasion. Activation of the type I interferon response in the tumor microenvironment using a STING agonist results in the engagement of innate and adaptive immune signaling and impairs the growth of KEAP1-mutant tumors. Our results suggest that targeting PARP and STING pathways, individually or in combination, represents a therapeutic strategy in NSCLC patients harboring alterations in KEAP1.
Asunto(s)
Carcinoma de Pulmón de Células no Pequeñas/inmunología , Interferón Tipo I/metabolismo , Neoplasias Pulmonares/inmunología , Proteínas de Neoplasias/metabolismo , Proteínas Nucleares/metabolismo , Reparación del ADN por Recombinación/genética , Proteínas Represoras/metabolismo , Escape del Tumor/genética , Animales , Carcinoma de Pulmón de Células no Pequeñas/genética , Carcinoma de Pulmón de Células no Pequeñas/metabolismo , Carcinoma de Pulmón de Células no Pequeñas/patología , Línea Celular Tumoral , Femenino , Células HEK293 , Humanos , Inmunidad Innata/genética , Proteína 1 Asociada A ECH Tipo Kelch/genética , Proteína 1 Asociada A ECH Tipo Kelch/metabolismo , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patología , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos NOD , Mutación , Proteínas de Neoplasias/genética , Proteínas Nucleares/genética , Proteínas Represoras/genética , Transducción de Señal/genética , Microambiente Tumoral/genética , Microambiente Tumoral/inmunología , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Repair of damaged DNA is essential for maintaining genome integrity and for preventing genome-instability-associated diseases, such as cancer. By combining proximity labeling with quantitative mass spectrometry, we generated high-resolution interaction neighborhood maps of the endogenously expressed DNA repair factors 53BP1, BRCA1, and MDC1. Our spatially resolved interaction maps reveal rich network intricacies, identify shared and bait-specific interaction modules, and implicate previously concealed regulators in this process. We identified a novel vertebrate-specific protein complex, shieldin, comprising REV7 plus three previously uncharacterized proteins, RINN1 (CTC-534A2.2), RINN2 (FAM35A), and RINN3 (C20ORF196). Recruitment of shieldin to DSBs, via the ATM-RNF8-RNF168-53BP1-RIF1 axis, promotes NHEJ-dependent repair of intrachromosomal breaks, immunoglobulin class-switch recombination (CSR), and fusion of unprotected telomeres. Shieldin functions as a downstream effector of 53BP1-RIF1 in restraining DNA end resection and in sensitizing BRCA1-deficient cells to PARP inhibitors. These findings have implications for understanding cancer-associated PARPi resistance and the evolution of antibody CSR in higher vertebrates.
Asunto(s)
Reparación del ADN por Unión de Extremidades/efectos de los fármacos , Proteínas de Unión al ADN/metabolismo , Inhibidores de Poli(ADP-Ribosa) Polimerasas/farmacología , Proteínas Adaptadoras Transductoras de Señales , Proteína BRCA1/antagonistas & inhibidores , Proteína BRCA1/genética , Proteína BRCA1/metabolismo , Proteínas de Ciclo Celular , Línea Celular Tumoral , Roturas del ADN de Doble Cadena , Proteínas de Unión al ADN/antagonistas & inhibidores , Proteínas de Unión al ADN/genética , Humanos , Cambio de Clase de Inmunoglobulina/efectos de los fármacos , Proteínas Mad2/antagonistas & inhibidores , Proteínas Mad2/genética , Proteínas Mad2/metabolismo , Mutagénesis Sitio-Dirigida , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Interferencia de ARN , ARN Interferente Pequeño/metabolismo , Proteínas de Unión a Telómeros/antagonistas & inhibidores , Proteínas de Unión a Telómeros/genética , Proteínas de Unión a Telómeros/metabolismo , Transactivadores/genética , Transactivadores/metabolismo , Proteína 1 de Unión al Supresor Tumoral P53/antagonistas & inhibidores , Proteína 1 de Unión al Supresor Tumoral P53/genética , Proteína 1 de Unión al Supresor Tumoral P53/metabolismo , Ubiquitina-Proteína Ligasas/antagonistas & inhibidores , Ubiquitina-Proteína Ligasas/genética , Ubiquitina-Proteína Ligasas/metabolismoRESUMEN
Tumor suppressor BRCA2 functions in homology-directed repair (HDR), the protection of stalled replication forks, and the suppression of replicative gaps, but their relative contributions to genome integrity and chemotherapy response are under scrutiny. Here, we report that mouse and human cells require a RAD51 filament stabilization motif in BRCA2 for fork protection and gap suppression but not HDR. In mice, the loss of fork protection/gap suppression does not compromise genome stability or shorten tumor latency. By contrast, HDR deficiency increases spontaneous and replication stress-induced chromosome aberrations and tumor predisposition. Unlike with HDR, fork protection/gap suppression defects are also observed in Brca2 heterozygous cells, likely due to reduced RAD51 stabilization at stalled forks/gaps. Gaps arise from PRIMPOL activity, which is associated with 5-hydroxymethyl-2'-deoxyuridine sensitivity due to the formation of SMUG1-generated abasic sites and is exacerbated by poly(ADP-ribose) polymerase (PARP) inhibition. However, HDR proficiency has the major role in mitigating sensitivity to chemotherapeutics, including PARP inhibitors.
Asunto(s)
Proteína BRCA2 , Replicación del ADN , Recombinasa Rad51 , Animales , Humanos , Ratones , Proteína BRCA2/metabolismo , Reparación del ADN , Inestabilidad Genómica , Genómica , Recombinasa Rad51/genética , Recombinasa Rad51/metabolismo , Reparación del ADN por RecombinaciónRESUMEN
POLQ is a key effector of DSB repair by microhomology-mediated end-joining (MMEJ) and is overexpressed in many cancers. POLQ inhibitors confer synthetic lethality in HR and Shieldin-deficient cancer cells, which has been proposed to reflect a critical dependence on the DSB repair pathway by MMEJ. Whether POLQ also operates independent of MMEJ remains unexplored. Here, we show that POLQ-deficient cells accumulate post-replicative ssDNA gaps upon BRCA1/2 loss or PARP inhibitor treatment. Biochemically, cooperation between POLQ helicase and polymerase activities promotes RPA displacement and ssDNA-gap fill-in, respectively. POLQ is also capable of microhomology-mediated gap skipping (MMGS), which generates deletions during gap repair that resemble the genomic scars prevalent in POLQ overexpressing cancers. Our findings implicate POLQ in mutagenic post-replicative gap sealing, which could drive genome evolution in cancer and whose loss places a critical dependency on HR for gap protection and repair and cellular viability.
Asunto(s)
Roturas del ADN de Doble Cadena , Neoplasias , Humanos , Replicación del ADN/genética , Inestabilidad Genómica , ADN de Cadena Simple/genética , Mutaciones Letales Sintéticas , Reparación del ADN por Unión de Extremidades , Neoplasias/genéticaRESUMEN
PARP1 rapidly detects DNA strand break damage and allosterically signals break detection to the PARP1 catalytic domain to activate poly(ADP-ribose) production from NAD+. PARP1 activation is characterized by dynamic changes in the structure of a regulatory helical domain (HD); yet, there are limited insights into the specific contributions that the HD makes to PARP1 allostery. Here, we have determined crystal structures of PARP1 in isolated active states that display specific HD conformations. These captured snapshots and biochemical analysis illustrate HD contributions to PARP1 multi-domain and high-affinity interaction with DNA damage, provide novel insights into the mechanics of PARP1 allostery, and indicate how HD active conformations correspond to alterations in the catalytic region that reveal the active site to NAD+. Our work deepens the understanding of PARP1 catalytic activation, the dynamics of the binding site of PARP inhibitor compounds, and the mechanisms regulating PARP1 retention on DNA damage.
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Daño del ADN , NAD , Dominio Catalítico , Reparación del ADN , NAD/metabolismo , Poli(ADP-Ribosa) Polimerasa-1/metabolismo , Poli Adenosina Difosfato Ribosa , Inhibidores de Poli(ADP-Ribosa) Polimerasas/farmacologíaRESUMEN
The 53BP1-RIF1-shieldin pathway maintains genome stability by suppressing nucleolytic degradation of DNA ends at double-strand breaks (DSBs). Although RIF1 interacts with damaged chromatin via phospho-53BP1 and facilitates recruitment of the shieldin complex to DSBs, it is unclear whether other regulatory cues contribute to this response. Here, we implicate methylation of histone H3 at lysine 4 by SETD1A-BOD1L in the recruitment of RIF1 to DSBs. Compromising SETD1A or BOD1L expression or deregulating H3K4 methylation allows uncontrolled resection of DNA ends, impairs end-joining of dysfunctional telomeres, and abrogates class switch recombination. Moreover, defects in RIF1 localization to DSBs are evident in patient cells bearing loss-of-function mutations in SETD1A. Loss of SETD1A-dependent RIF1 recruitment in BRCA1-deficient cells restores homologous recombination and leads to resistance to poly(ADP-ribose)polymerase inhibition, reinforcing the clinical relevance of these observations. Mechanistically, RIF1 binds directly to methylated H3K4, facilitating its recruitment to, or stabilization at, DSBs.
Asunto(s)
Roturas del ADN de Doble Cadena , Proteínas de Unión a Telómeros , Proteína BRCA1/genética , ADN/metabolismo , Reparación del ADN por Unión de Extremidades , Reparación del ADN , N-Metiltransferasa de Histona-Lisina/genética , N-Metiltransferasa de Histona-Lisina/metabolismo , Humanos , Metilación , Proteínas de Unión a Telómeros/genética , Proteínas de Unión a Telómeros/metabolismo , Proteína 1 de Unión al Supresor Tumoral P53/genética , Proteína 1 de Unión al Supresor Tumoral P53/metabolismoRESUMEN
Mammalian DNA base excision repair (BER) is accelerated by poly(ADP-ribose) polymerases (PARPs) and the scaffold protein XRCC1. PARPs are sensors that detect single-strand break intermediates, but the critical role of XRCC1 during BER is unknown. Here, we show that protein complexes containing DNA polymerase ß and DNA ligase III that are assembled by XRCC1 prevent excessive engagement and activity of PARP1 during BER. As a result, PARP1 becomes "trapped" on BER intermediates in XRCC1-deficient cells in a manner similar to that induced by PARP inhibitors, including in patient fibroblasts from XRCC1-mutated disease. This excessive PARP1 engagement and trapping renders BER intermediates inaccessible to enzymes such as DNA polymerase ß and impedes their repair. Consequently, PARP1 deletion rescues BER and resistance to base damage in XRCC1-/- cells. These data reveal excessive PARP1 engagement during BER as a threat to genome integrity and identify XRCC1 as an "anti-trapper" that prevents toxic PARP1 activity.
Asunto(s)
Reparación del ADN/genética , ADN/genética , Poli(ADP-Ribosa) Polimerasa-1/metabolismo , Proteína 1 de Reparación por Escisión del Grupo de Complementación Cruzada de las Lesiones por Rayos X/metabolismo , Animales , Línea Celular , Roturas del ADN de Cadena Simple , Daño del ADN/efectos de los fármacos , Daño del ADN/genética , ADN Ligasa (ATP)/metabolismo , ADN Polimerasa beta/metabolismo , Reparación del ADN/efectos de los fármacos , Proteínas de Unión al ADN/metabolismo , Fibroblastos/efectos de los fármacos , Fibroblastos/metabolismo , Humanos , Inhibidores de Poli(ADP-Ribosa) Polimerasas/farmacología , Poli(ADP-Ribosa) Polimerasas/metabolismo , Unión Proteica/efectos de los fármacosRESUMEN
In addition to its role as an electron transporter, mitochondrial nicotinamide adenine dinucleotide (NAD+) is an important co-factor for enzymatic reactions, including ADP-ribosylation. Although mitochondria harbor the most intra-cellular NAD+, mitochondrial ADP-ribosylation remains poorly understood. Here we provide evidence for mitochondrial ADP-ribosylation, which was identified using various methodologies including immunofluorescence, western blot, and mass spectrometry. We show that mitochondrial ADP-ribosylation reversibly increases in response to respiratory chain inhibition. Conversely, H2O2-induced oxidative stress reciprocally induces nuclear and reduces mitochondrial ADP-ribosylation. Elevated mitochondrial ADP-ribosylation, in turn, dampens H2O2-triggered nuclear ADP-ribosylation and increases MMS-induced ARTD1 chromatin retention. Interestingly, co-treatment of cells with the mitochondrial uncoupler FCCP decreases PARP inhibitor efficacy. Together, our results suggest that mitochondrial ADP-ribosylation is a dynamic cellular process that impacts nuclear ADP-ribosylation and provide evidence for a NAD+-mediated mitochondrial-nuclear crosstalk.
Asunto(s)
ADP-Ribosilación , Núcleo Celular/enzimología , Mitocondrias/enzimología , NAD/metabolismo , Poli(ADP-Ribosa) Polimerasa-1/metabolismo , ADP-Ribosilación/efectos de los fármacos , Animales , Antimicina A/análogos & derivados , Antimicina A/farmacología , Línea Celular , Línea Celular Tumoral , Núcleo Celular/efectos de los fármacos , Núcleo Celular/genética , Cromatina/química , Cromatina/metabolismo , Transporte de Electrón/efectos de los fármacos , Células HeLa , Humanos , Peróxido de Hidrógeno/farmacología , Metacrilatos/farmacología , Ratones , Ratones Endogámicos C57BL , Mitocondrias/efectos de los fármacos , Mitocondrias/genética , Mioblastos/citología , Mioblastos/efectos de los fármacos , Mioblastos/enzimología , Oligomicinas/farmacología , Osteoblastos/citología , Osteoblastos/efectos de los fármacos , Osteoblastos/enzimología , Poli(ADP-Ribosa) Polimerasa-1/genética , Rotenona/farmacología , Tiazoles/farmacologíaRESUMEN
The DNA damage response (DDR) fulfils essential roles to preserve genome integrity. Targeting the DDR in tumors has had remarkable success over the last decade, exemplified by the licensing of PARP inhibitors for cancer therapy. Recent studies suggest that the application of DDR inhibitors impacts on cellular innate and adaptive immune responses, wherein key DNA repair factors have roles in limiting chronic inflammatory signaling. Antitumor immunity plays an emerging part in cancer therapy, and extensive efforts have led to the development of immune checkpoint inhibitors overcoming immune suppressive signals in tumors. Here, we review the current understanding of the molecular mechanisms underlying DNA damage-triggered immune responses, including cytosolic DNA sensing via the cGAS/STING pathway. We highlight the implications of DDR components for therapeutic outcomes of immune checkpoint inhibitors or their use as biomarkers. Finally, we discuss the rationale for novel combinations of DDR inhibitors with antagonists of immune checkpoints and current hindrances limiting their broader therapeutic applications.
Asunto(s)
Reparación del ADN/fisiología , Inmunidad Celular/genética , Inmunoterapia , Neoplasias/terapia , Inmunidad Adaptativa/genética , Daño del ADN/inmunología , Receptores con Dominio Discoidina/antagonistas & inhibidores , Humanos , Inhibidores de Puntos de Control Inmunológico/uso terapéutico , Neoplasias/tratamiento farmacológico , Inhibidores de Poli(ADP-Ribosa) Polimerasas/farmacología , Inhibidores de Poli(ADP-Ribosa) Polimerasas/uso terapéutico , Inhibidores de Proteínas Quinasas/uso terapéuticoRESUMEN
The anti-tumor potency of poly(ADP-ribose) polymerase (PARP) inhibitors (PARPis) has been linked to trapping of PARP1 on damaged chromatin. However, little is known about their impact on PARP2, an isoform with overlapping functions at DNA lesions. Whether the release of PARP1/2 from DNA lesions is actively catalyzed by molecular machines is also not known. We found that PARPis robustly trap PARP2 and that the helicase ALC1 (CHD1L) is strictly required for PARP2 release. Catalytic inactivation of ALC1 quantitatively traps PARP2 but not PARP1. ALC1 manipulation impacts the response to single-strand DNA breaks through PARP2 trapping, potentiates PARPi-induced cancer cell killing, and mediates synthetic lethality upon BRCA deficiency. The chromatin remodeler ALC1 actively drives PARP2 turnover from DNA lesions, and PARP2 contributes to the cellular responses of PARPi. This suggests that disrupting the ATP-fueled remodeling forces of ALC1 might enable therapies that selectively target the DNA repair functions of PARPs in cancer.
Asunto(s)
Roturas del ADN de Cadena Simple , ADN Helicasas/metabolismo , Proteínas de Unión al ADN/metabolismo , Neoplasias/enzimología , Inhibidores de Poli(ADP-Ribosa) Polimerasas/metabolismo , Poli(ADP-Ribosa) Polimerasas/metabolismo , Proteínas Proto-Oncogénicas/metabolismo , Línea Celular Tumoral , ADN Helicasas/genética , Proteínas de Unión al ADN/genética , Humanos , Neoplasias/genética , Neoplasias/patología , Poli(ADP-Ribosa) Polimerasa-1/genética , Poli(ADP-Ribosa) Polimerasa-1/metabolismo , Poli(ADP-Ribosa) Polimerasas/genética , Proteínas Proto-Oncogénicas/genéticaRESUMEN
ADP-ribosylation (ADPRylation) is a posttranslational modification of proteins discovered nearly six decades ago, but many important questions remain regarding its molecular functions and biological roles, as well as the activity of the ADP-ribose (ADPR) transferase enzymes (PARP family members) that catalyze it. Growing evidence indicates that PARP-mediated ADPRylation events are key regulators of the protein biosynthetic pathway, leading from rDNA transcription and ribosome biogenesis to mRNA synthesis, processing, and translation. In this review we describe the role of PARP proteins and ADPRylation in all facets of this pathway. PARP-1 and its enzymatic activity are key regulators of rDNA transcription, which is a critical step in ribosome biogenesis. An emerging role of PARPs in alternative splicing of mRNAs, as well as direct ADPRylation of mRNAs, highlight the role of PARP members in RNA processing. Furthermore, PARP activity, stimulated by cellular stresses, such as viral infections and ER stress, leads to the regulation of mRNA stability and protein synthesis through posttranscriptional mechanisms. Dysregulation of PARP activity in these processes can promote disease states. Collectively, these results highlight the importance of PARP family members and ADPRylation in gene regulation, mRNA processing, and protein abundance. Future studies in these areas will yield new insights into the fundamental mechanisms and a broader utility for PARP-targeted therapeutic agents.
Asunto(s)
ADP-Ribosilación/fisiología , Expresión Génica/fisiología , Poli(ADP-Ribosa) Polimerasas/metabolismo , Biosíntesis de Proteínas/fisiología , Proteostasis/fisiología , Animales , Humanos , Procesamiento Proteico-Postraduccional , ARN/metabolismoRESUMEN
SLFN11 sensitizes cancer cells to a broad range of DNA-targeted therapies. Here we show that, in response to replication stress induced by camptothecin, SLFN11 tightly binds chromatin at stressed replication foci via RPA1 together with the replication helicase subunit MCM3. Unlike ATR, SLFN11 neither interferes with the loading of CDC45 and PCNA nor inhibits the initiation of DNA replication but selectively blocks fork progression while inducing chromatin opening across replication initiation sites. The ATPase domain of SLFN11 is required for chromatin opening, replication block, and cell death but not for the tight binding of SLFN11 to chromatin. Replication stress by the CHK1 inhibitor Prexasertib also recruits SLFN11 to nascent replicating DNA together with CDC45 and PCNA. We conclude that SLFN11 is recruited to stressed replication forks carrying extended RPA filaments where it blocks replication by changing chromatin structure across replication sites.
Asunto(s)
Proteínas Nucleares/genética , Proteínas de la Ataxia Telangiectasia Mutada/metabolismo , Camptotecina , Proteínas de Ciclo Celular/metabolismo , Línea Celular Tumoral , Cromatina/metabolismo , Daño del ADN , ADN Helicasas/metabolismo , Replicación del ADN/genética , Replicación del ADN/fisiología , ADN de Cadena Simple/genética , ADN de Cadena Simple/metabolismo , Humanos , Proteínas de Mantenimiento de Minicromosoma/metabolismo , Proteínas Nucleares/metabolismo , Pirazinas , Pirazoles , Proteína de Replicación A/metabolismoRESUMEN
Ovarian cancer is the most lethal gynecological malignancy and is often associated with both DNA repair deficiency and extensive metabolic reprogramming. While still emerging, the interplay between these pathways can affect ovarian cancer phenotypes, including therapeutic resistance to the DNA damaging agents that are standard-of-care for this tumor type. In this review, we will discuss what is currently known about cellular metabolic rewiring in ovarian cancer that may impact DNA damage and repair in addition to highlighting how specific DNA repair proteins also promote metabolic changes. We will also discuss relevant data from other cancers that could be used to inform ovarian cancer therapeutic strategies. Changes in the choice of DNA repair mechanism adopted by ovarian cancer are a major factor in promoting therapeutic resistance. Therefore, the impact of metabolic reprogramming on DNA repair mechanisms in ovarian cancer has major clinical implications for targeted combination therapies for the treatment of this devastating disease.
Asunto(s)
Daño del ADN , Reparación del ADN , Neoplasias Ováricas , Humanos , Neoplasias Ováricas/genética , Neoplasias Ováricas/metabolismo , Neoplasias Ováricas/patología , Femenino , AnimalesRESUMEN
DNA double-strand break (DSB) resection, once thought to be a simple enzymatic process, is emerging as a highly complex series of coordinated activities required to maintain genome integrity. Progress in cell biology, biochemistry, and genetics has deciphered the precise resecting activities, the regulatory components, and their ability to properly channel the resected DNA to the appropriate DNA repair pathway. Herein, we review the mechanisms of regulation of DNA resection, with an emphasis on negative regulators that prevent single-strand (ss)DNA accumulation to maintain genome stability. Interest in targeting DNA resection inhibitors is emerging because their inactivation leads to poly(ADP-ribose) polymerase inhibitor (PARPi) resistance. We also present detailed regulation of DNA resection machineries, their analysis by functional assays, and their impact on disease and PARPi resistance.
Asunto(s)
Roturas del ADN de Doble Cadena , Reparación del ADN , ADN , Inhibidores de Poli(ADP-Ribosa) Polimerasas , Poli(ADP-Ribosa) Polimerasas/metabolismoRESUMEN
ADP-ribose is a versatile modification that plays a critical role in diverse cellular processes. The addition of this modification is catalyzed by ADP-ribosyltransferases, among which notable poly(ADP-ribose) polymerase (PARP) enzymes are intimately involved in the maintenance of genome integrity. The role of ADP-ribose modifications during DNA damage repair is of significant interest for the proper development of PARP inhibitors targeted toward the treatment of diseases caused by genomic instability. More specifically, inhibitors promoting PARP persistence on DNA lesions, termed PARP "trapping," is considered a desirable characteristic. In this review, we discuss key classes of proteins involved in ADP-ribose signaling (writers, readers, and erasers) with a focus on those involved in the maintenance of genome integrity. An overview of factors that modulate PARP1 and PARP2 persistence at sites of DNA lesions is also discussed. Finally, we clarify aspects of the PARP trapping model in light of recent studies that characterize the kinetics of PARP1 and PARP2 recruitment at sites of lesions. These findings suggest that PARP trapping could be considered as the continuous recruitment of PARP molecules to sites of lesions, rather than the physical stalling of molecules. Recent studies and novel research tools have elevated the level of understanding of ADP-ribosylation, marking a coming-of-age for this interesting modification.
Asunto(s)
Inestabilidad Genómica , Poli(ADP-Ribosa) Polimerasa-1 , Humanos , Adenosina Difosfato Ribosa , Daño del ADN , Reparación del ADN , Poli(ADP-Ribosa) Polimerasa-1/química , Poli(ADP-Ribosa) Polimerasa-1/genética , Poli(ADP-Ribosa) Polimerasa-1/metabolismo , Inhibidores de Poli(ADP-Ribosa) Polimerasas/farmacología , AnimalesRESUMEN
Metastatic melanoma is still a difficult-to-treat cancer type owing to its frequent resistance mechanisms to targeted and immunotherapy. Therefore, we aimed to unravel novel therapeutic strategies for melanoma patients. Preclinical and clinical studies show that melanoma patients may benefit from a treatment with poly (ADP-ribose) polymerase (PARP) inhibitors (PARPi). In this study, we focus on PARP1 as a potential biomarker to predict the response of melanoma cells to PARPi therapy. We found that melanoma cells with high basal PARP1 expression exhibit significantly increased cell death after PARPi treatment owing to higher PARP1 trapping compared with melanoma cells with low PARP1 expression. In addition, we could demonstrate that PARP1 expression levels are low in nonmalignant skin cells, and metastatic melanomas show considerably higher PARP1 levels compared with primary melanomas. Most strikingly, we found that high PARP1 levels correlate with worse overall survival of late stage metastasized melanoma patients. In conclusion, we show that PARP1 might act as a biomarker to predict the response to PARPi therapy, and that in particular the late stage metastasized melanoma patients are especially sensitive to PARPi therapy owing to elevated PARP1 expression. Our data suggest that the PARPi cytotoxicity primarily will affect the high PARP1 expressing melanoma cells, rather than the low PARP1 expressing nonmalignant skin cells resulting in only low side effects.
Asunto(s)
Melanoma , Poli(ADP-Ribosa) Polimerasa-1 , Inhibidores de Poli(ADP-Ribosa) Polimerasas , Neoplasias Cutáneas , Femenino , Humanos , Masculino , Biomarcadores de Tumor/metabolismo , Biomarcadores de Tumor/genética , Línea Celular Tumoral , Resistencia a Antineoplásicos/genética , Melanoma/tratamiento farmacológico , Melanoma/mortalidad , Melanoma/genética , Melanoma/patología , Melanoma/metabolismo , Metástasis de la Neoplasia , Poli(ADP-Ribosa) Polimerasa-1/metabolismo , Inhibidores de Poli(ADP-Ribosa) Polimerasas/uso terapéutico , Inhibidores de Poli(ADP-Ribosa) Polimerasas/farmacología , Poli(ADP-Ribosa) Polimerasas/metabolismo , Pronóstico , Neoplasias Cutáneas/tratamiento farmacológico , Neoplasias Cutáneas/mortalidad , Neoplasias Cutáneas/patología , Neoplasias Cutáneas/genética , Neoplasias Cutáneas/metabolismoRESUMEN
BACKGROUND: Poly ADP-ribose polymerase (PARP) inhibitors are approved for the treatment of some men with advanced prostate cancer. Rare but serious side effects include myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML). The impact of PARP inhibitors on clonal hematopoiesis (CH), a potential precursor lesion associated with MDS and AML, is incompletely understood in prostate cancer. We hypothesized that PARP inhibitors would increase CH prevalence and abundance. METHODS: We prospectively enrolled participants with advanced prostate cancer treated with PARP inhibitors. The presence of CH was assessed from leukocytes using an ultra-deep error-corrected dual unique molecular identifiers sequencing method targeting 49 genes most commonly mutated in CH and myeloid malignancies. Variant allele frequencies (VAF) of ≥0.5% were considered clinically significant. Blood samples were collected before and after PARP inhibitor treatment. RESULTS: Ten men were enrolled; mean age of 67 years. Six patients had Gleason 7 disease, and four had Gleason ≥8 disease at diagnosis. Nine had localized disease at diagnosis, and eight had prior treatment with radiation. The mean time between pre- and post-treatment blood samples was 11 months (range 2.6-31 months). Six patients (60%) had CH identified prior to PARP inhibitor treatment, three with multiple clones. Of 11 CH clones identified in follow-up, 5 (45%) appeared or increased after treatment. DNMT3A, TET2, and PPM1D were the most common CH alterations observed. The largest post-treatment increase involved the PPM1D gene. CONCLUSION: CH alterations are frequently found after treatment with PARP inhibitors in patients with prostate cancer and this may be one mechanism by which PARP inhibitors lead to increased risk of MDS/AML.
Asunto(s)
Hematopoyesis Clonal , Inhibidores de Poli(ADP-Ribosa) Polimerasas , Neoplasias de la Próstata , Humanos , Masculino , Inhibidores de Poli(ADP-Ribosa) Polimerasas/efectos adversos , Inhibidores de Poli(ADP-Ribosa) Polimerasas/uso terapéutico , Neoplasias de la Próstata/tratamiento farmacológico , Neoplasias de la Próstata/genética , Neoplasias de la Próstata/patología , Anciano , Persona de Mediana Edad , Hematopoyesis Clonal/genética , Estudios Prospectivos , Progresión de la Enfermedad , Prevalencia , Anciano de 80 o más Años , Proteínas de Unión al ADN , DioxigenasasRESUMEN
BACKGROUND: Resistance to therapies that target homologous recombination deficiency (HRD) in breast cancer limits their overall effectiveness. Multiple, preclinically validated, mechanisms of resistance have been proposed, but their existence and relative frequency in clinical disease are unclear, as is how to target resistance. PATIENTS AND METHODS: Longitudinal mutation and methylation profiling of circulating tumour (ct)DNA was carried out in 47 patients with metastatic BRCA1-, BRCA2- or PALB2-mutant breast cancer treated with HRD-targeted therapy who developed progressive disease-18 patients had primary resistance and 29 exhibited response followed by resistance. ctDNA isolated at multiple time points in the patient treatment course (before, on-treatment and at progression) was sequenced using a novel >750-gene intron/exon targeted sequencing panel. Where available, matched tumour biopsies were whole exome and RNA sequenced and also used to assess nuclear RAD51. RESULTS: BRCA1/2 reversion mutations were present in 60% of patients and were the most prevalent form of resistance. In 10 cases, reversions were detected in ctDNA before clinical progression. Two new reversion-based mechanisms were identified: (i) intragenic BRCA1/2 deletions with intronic breakpoints; and (ii) intragenic BRCA1/2 secondary mutations that formed novel splice acceptor sites, the latter being confirmed by in vitro minigene reporter assays. When seen before commencing subsequent treatment, reversions were associated with significantly shorter time to progression. Tumours with reversions retained HRD mutational signatures but had functional homologous recombination based on RAD51 status. Although less frequent than reversions, nonreversion mechanisms [loss-of-function (LoF) mutations in TP53BP1, RIF1 or PAXIP1] were evident in patients with acquired resistance and occasionally coexisted with reversions, challenging the notion that singular resistance mechanisms emerge in each patient. CONCLUSIONS: These observations map the prevalence of candidate drivers of resistance across time in a clinical setting, information with implications for clinical management and trial design in HRD breast cancers.
Asunto(s)
Antineoplásicos , Neoplasias de la Mama , Femenino , Humanos , Antineoplásicos/farmacología , Antineoplásicos/uso terapéutico , Proteína BRCA1/genética , Proteína BRCA2/genética , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Recombinación Homóloga , Mutación , Inhibidores de Poli(ADP-Ribosa) Polimerasas/farmacología , Inhibidores de Poli(ADP-Ribosa) Polimerasas/uso terapéutico , Proteína 1 de Unión al Supresor Tumoral P53RESUMEN
BACKGROUND: The prospective phase III multi-centre L-MOCA trial (NCT03534453) has demonstrated the encouraging efficacy and manageable safety profile of olaparib maintenance therapy in the Asian (mainly Chinese) patients with platinum-sensitive relapsed ovarian cancer (PSROC). In this study, we report the preplanned exploratory biomarker analysis of the L-MOCA trial, which investigated the effects of homologous recombination deficiency (HRD) and programmed cell death ligand 1 (PD-L1) expression on olaparib efficacy. METHODS: HRD status was determined using the ACTHRD assay, an enrichment-based targeted next-generation sequencing assay. PD-L1 expression was assessed by SP263 immunohistochemistry assay. PD-L1 expression positivity was defined by the PD-L1 expression on ≥ 1% of immune cells. Kaplan-Meier method was utilised to analyse progression-free survival (PFS). RESULTS: This exploratory biomarker analysis included 225 patients and tested HRD status [N = 190; positive, N = 125 (65.8%)], PD-L1 expression [N = 196; positive, N = 56 (28.6%)], and BRCA1/2 mutation status (N = 219). The HRD-positive patients displayed greater median PFS than the HRD-negative patients [17.9 months (95% CI: 14.5-22.1) versus 9.2 months (95% CI: 7.5-13.8)]. PD-L1 was predominantly expressed on immune cells. Positive PD-L1 expression on immune cells was associated with shortened median PFS in the patients with germline BRCA1/2 mutations [14.5 months (95% CI: 7.4-18.2) versus 22.2 months (95% CI: 18.3-NA)]. Conversely, positive PD-L1 expression on immune cells was associated with prolonged median PFS in the patients with wild-type BRCA1/2 [20.9 months (95% CI: 13.9-NA) versus 8.3 months (95% CI: 6.7-13.8)]. CONCLUSIONS: HRD remained an effective biomarker for enhanced olaparib efficacy in the Asian patients with PSROC. Positive PD-L1 expression was associated with decreased olaparib efficacy in the patients with germline BRCA1/2 mutations but associated with improved olaparib efficacy in the patients with wild-type BRCA1/2. TRIAL REGISTRATION: NCT03534453. Registered at May 23, 2018.
Asunto(s)
Antígeno B7-H1 , Biomarcadores de Tumor , Quimioterapia de Mantención , Neoplasias Ováricas , Ftalazinas , Piperazinas , Humanos , Femenino , Ftalazinas/uso terapéutico , Neoplasias Ováricas/tratamiento farmacológico , Neoplasias Ováricas/genética , Piperazinas/uso terapéutico , Biomarcadores de Tumor/genética , Persona de Mediana Edad , Quimioterapia de Mantención/métodos , Anciano , Adulto , Estudios Prospectivos , Recurrencia Local de Neoplasia/tratamiento farmacológico , Proteína BRCA2/genética , Antineoplásicos/uso terapéutico , Proteína BRCA1/genética , Recombinación HomólogaRESUMEN
ADP-ribosyltransferases of the PARP family encompass a group of enzymes with variegated regulatory functions in cells, ranging from DNA damage repair to the control of cell-cycle progression and immune response. Over the years, this knowledge has led to the use of PARP1/2 inhibitors as mainstay pharmaceutical strategies for the treatment of ovarian, pancreatic, prostate and breast cancers, holding mutations in genes encoding for proteins involved in the DNA repair mechanisms (synthetic lethality). Meanwhile, the last decade has witnessed significant progress in comprehending cellular pathways regulated by mono-ADP-ribosylation, with a huge effort in the development of novel selective compounds to inhibit those PARPs endowed with mono-ADP-ribosylation activity. This review focuses on the progress achieved in the cancer field, delving into most recent findings regarding the role of a subset of enzymes - the interferon-stimulated PARPs - in cancer progression.