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The ATR-CHK1 DNA damage response pathway becomes activated by the exposure of RPA-coated single-stranded DNA (ssDNA) that forms as an intermediate during DNA damage and repair, and as a part of the replication stress response. Here, we identify ZNF827 as a component of the ATR-CHK1 kinase pathway. We demonstrate that ZNF827 is a ssDNA binding protein that associates with RPA through concurrent binding to ssDNA intermediates. These interactions are dependent on two clusters of C2H2 zinc finger motifs within ZNF827. We find that ZNF827 accumulates at stalled forks and DNA damage sites, where it activates ATR and promotes the engagement of homologous recombination-mediated DNA repair. Additionally, we demonstrate that ZNF827 depletion inhibits replication initiation and sensitizes cancer cells to the topoisomerase inhibitor topotecan, revealing ZNF827 as a therapeutic target within the DNA damage response pathway.
Assuntos
Proteínas Quinases , Transdução de Sinais , Proteínas Quinases/metabolismo , Fosforilação , Proteína de Replicação A/metabolismo , Proteínas Mutadas de Ataxia Telangiectasia/metabolismo , Proteínas de Ligação a DNA/metabolismo , Replicação do DNA , Dano ao DNA , DNA de Cadeia Simples , Reparo do DNARESUMO
Alternative lengthening of telomeres (ALT) is a homologous recombination-based telomere maintenance mechanism. It is active in approximately 10-15% of cancers. We present a DNA-fiber protocol, combining YOYO-1 staining of genomic DNA, telomere fluorescence in situ hybridization (FISH), and EdU labeling of nascent DNA, to measure telomere extension events in ALT cancer cells. The protocol can be used to delineate ALT-mediated telomere extension. For complete details on the use and execution of this protocol, please refer to Barroso-Gonzalez et al. (2021).
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Neoplasias , Imagem Individual de Molécula , DNA/genética , Hibridização in Situ Fluorescente/métodos , Neoplasias/genética , Telômero/genética , Homeostase do Telômero/genéticaRESUMO
Alternative lengthening of telomeres (ALT) is a telomere-elongation mechanism observed in â¼15% of cancer subtypes. Current models indicate that ALT is mediated by homology-directed repair mechanisms. By disrupting MSH6 gene expression, we show that the deficiency of MutSα (MSH2/MSH6) DNA mismatch repair complex causes striking telomere hyperextension. Mechanistically, we show MutSα is specifically recruited to telomeres in ALT cells by associating with the proliferating-cell nuclear antigen (PCNA) subunit of the ALT telomere replisome. We also provide evidence that MutSα counteracts Bloom (BLM) helicase, which adopts a crucial role in stabilizing hyper-extended telomeres and maintaining the survival of MutSα-deficient ALT cancer cells. Lastly, we propose a model in which MutSα deficiency impairs heteroduplex rejection, leading to premature initiation of telomere DNA synthesis that coincides with an accumulation of telomere variant repeats (TVRs). These findings provide evidence that the MutSα DNA mismatch repair complex acts to restrain unwarranted ALT.
Assuntos
DNA de Neoplasias/metabolismo , Proteínas de Ligação a DNA/metabolismo , Proteína 2 Homóloga a MutS/metabolismo , Neoplasias/enzimologia , Ácidos Nucleicos Heteroduplexes/metabolismo , Homeostase do Telômero , Telômero/metabolismo , Linhagem Celular Tumoral , Reparo de Erro de Pareamento de DNA , DNA de Neoplasias/genética , Proteínas de Ligação a DNA/genética , Instabilidade Genômica , Células HeLa , Humanos , Modelos Genéticos , Proteína 2 Homóloga a MutS/genética , Neoplasias/genética , Neoplasias/patologia , Conformação de Ácido Nucleico , Ácidos Nucleicos Heteroduplexes/genética , RecQ Helicases/genética , RecQ Helicases/metabolismo , Telômero/genéticaRESUMO
BACKGROUND: Prostate cancer (PC) and its treatments lead to significant acute, chronic, or latent adverse effects that result in declines in patients' physical functions, quality of life and reduced sense of masculinities. Robust evidence shows that physical activity (PA) can improve many health outcomes in men with PC; however, less is known about the facilitators, preferences, and barriers to PA engagement in this population. The purpose of this scoping review is to document the nature and extent of literature related to these aspects of PA participation among men with PC. METHODS: We conducted a scoping review of PA among men with PC. Databases searched included Medline, CINAHL, Embase, Rehabilitation & Sports Medicine Source, and SportDiscus from inception to June 30, 2020. Multiple reviewers were used in all screening and data abstractions. RESULTS: The search yielded 2788 individual citations after duplicates were removed. Following title and abstract screening, 129 underwent full-text review, and 46 articles were included. Quantitative data related to our research question showed that structured group exercise was the most commonly reported facilitator/preference among men with PC, whereas treatment-related effects and lack of time are the most common barriers. In terms of qualitative data, the most prominent theme noted related to masculinities and gender-specific needs within the context of having PC. CONCLUSION: Men with PC have unique facilitators and barriers concerning PA. More work is needed from the research and clinical practice perspectives to enable this population to engage and remain in regular PA.
Assuntos
Exercício Físico , Neoplasias da Próstata/fisiopatologia , Neoplasias da Próstata/psicologia , Qualidade de Vida , Humanos , MasculinoRESUMO
Sarcoidosis is a non-infective granulomatous disorder of unknown aetiology, with cutaneous involvement affecting up to 30% of patients. Drug-induced sarcoidosis has been reported secondary to modern melanoma therapies including immune-checkpoint inhibitors and first generation BRAF inhibitors such as vemurafenib and dabrafenib. Herein, we report a case of cutaneous micropapular sarcoidosis that first developed on immune-checkpoint inhibition with ipilimumab and nivolumab for metastatic melanoma, which was exacerbated and further complicated by pityriasis rubra pilaris-like palmar plaques upon transition to a next-generation BRAF-dimerisation inhibitor. Both the micropapular eruption and palmar plaques rapidly resolved after cessation of the novel BRAF-inhibitor and concurrent commencement of hydroxychloroquine. It is unclear how inhibition of BRAF-dimerisation results in granuloma formation, though upregulation of TH1/TH17 T-cells and impairment of T-reg cells may be responsible. Clinicians should be aware of the potential for exacerbation of sarcoidosis when transitioning from immune-checkpoint inhibitors to these novel BRAF-dimerisation inhibitors, particularly as their uptake in treating cancers increases beyond clinical trials. Further studies are required to assess whether these next-generation agents can trigger sarcoidosis de-novo, or simply exacerbate pre-existing sarcoidosis.
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BACKGROUND: Despite the completion of numerous phase II studies, a standard of care treatment has yet to be defined for metastatic uveal melanoma (mUM). To determine benchmarks of progression free survival (PFS) and overall survival (OS), we carried out a meta-analysis using individual patient level trial data. METHODS: Individual patient variables and survival outcomes were requested from 29 trials published from 2000 to 2016. Univariable and multivariable analysis were carried out for prognostic factors. The variability between trial arms and between therapeutic agents on PFS and OS was investigated. RESULTS: OS data were available for 912 patients. The median PFS was 3.3 months (95% CI 2.9-3.6) and 6-month PFS rate was 27% (95% CI 24-30). Univariable analysis showed male sex, elevated (i.e. > versus ≤ upper limit of normal) lactate dehydrogenase (LDH), elevated alkaline phosphatase (ALP) and diameter of the largest liver metastasis (≥3 cm versus <3 cm) to be substantially associated with shorter PFS. Multivariable analysis showed male sex, elevated LDH and elevated ALP were substantially associated with shorter PFS. The most substantial factors associated with 6-month PFS rate, on both univariable and multivariable analysis were elevated LDH and ALP. The median OS was 10.2 months (95% CI 9.5-11.0) and 1 year OS was 43% (95% CI 40-47). The most substantial prognostic factors for shorter OS by univariable and multivariable analysis were elevated LDH and elevated ALP. Patients treated with liver directed treatments had statistically significant longer PFS and OS. CONCLUSION: Benchmarks of 6-month PFS and 1-year OS rates were determined accounting for prognostic factors. These may be used to facilitate future trial design and stratification in mUM.
Assuntos
Ensaios Clínicos como Assunto/normas , Neoplasias Hepáticas/tratamento farmacológico , Melanoma/tratamento farmacológico , Projetos de Pesquisa/estatística & dados numéricos , Neoplasias Uveais/tratamento farmacológico , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Fosfatase Alcalina/sangue , Benchmarking , Conjuntos de Dados como Assunto , Feminino , Humanos , Estimativa de Kaplan-Meier , L-Lactato Desidrogenase/sangue , Neoplasias Hepáticas/sangue , Neoplasias Hepáticas/mortalidade , Neoplasias Hepáticas/secundário , Masculino , Melanoma/sangue , Melanoma/mortalidade , Melanoma/patologia , Pessoa de Meia-Idade , Prognóstico , Intervalo Livre de Progressão , Estudos Prospectivos , Fatores Sexuais , Fatores de Tempo , Neoplasias Uveais/sangue , Neoplasias Uveais/mortalidade , Neoplasias Uveais/patologia , Adulto JovemRESUMO
The collapse of stalled replication forks is a major driver of genomic instability. Several committed mechanisms exist to resolve replication stress. These pathways are particularly pertinent at telomeres. Cancer cells that use Alternative Lengthening of Telomeres (ALT) display heightened levels of telomere-specific replication stress, and co-opt stalled replication forks as substrates for break-induced telomere synthesis. FANCM is a DNA translocase that can form independent functional interactions with the BLM-TOP3A-RMI (BTR) complex and the Fanconi anemia (FA) core complex. Here, we demonstrate that FANCM depletion provokes ALT activity, evident by increased break-induced telomere synthesis, and the induction of ALT biomarkers. FANCM-mediated attenuation of ALT requires its inherent DNA translocase activity and interaction with the BTR complex, but does not require the FA core complex, indicative of FANCM functioning to restrain excessive ALT activity by ameliorating replication stress at telomeres. Synthetic inhibition of FANCM-BTR complex formation is selectively toxic to ALT cancer cells.
Assuntos
Proteínas de Transporte/metabolismo , DNA Helicases/metabolismo , DNA Topoisomerases Tipo I/metabolismo , Proteínas de Ligação a DNA/metabolismo , Neoplasias/metabolismo , Proteínas Nucleares/metabolismo , RecQ Helicases/metabolismo , Homeostase do Telômero , Telômero/metabolismo , Linhagem Celular Tumoral , Replicação do DNA , Células HCT116 , Células HEK293 , Células HeLa , HumanosRESUMO
BACKGROUND: Pembrolizumab demonstrated robust antitumor activity and safety in the phase Ib KEYNOTE-001 study (NCT01295827) of advanced melanoma. Five-year outcomes in all patients and treatment-naive patients are reported herein. Patients whose disease progressed following initial response and who received a second course of pembrolizumab were also analyzed. PATIENTS AND METHODS: Patients aged ≥18 years with previously treated or treatment-naive advanced/metastatic melanoma received pembrolizumab 2 mg/kg every 3 weeks, 10 mg/kg every 3 weeks, or 10 mg/kg every 2 weeks until disease progression, intolerable toxicity, or patient/investigator decision to withdraw. Kaplan-Meier estimates of overall survival (OS) and progression-free survival (PFS) were calculated. Objective response rate and PFS were based on immune-related response criteria by investigator assessment (data cut-off, September 1, 2017). RESULTS: KEYNOTE-001 enrolled 655 patients with melanoma; median follow-up was 55 months. Estimated 5-year OS was 34% in all patients and 41% in treatment-naive patients; median OS was 23.8 months (95% CI, 20.2-30.4) and 38.6 months (95% CI, 27.2-not reached), respectively. Estimated 5-year PFS rates were 21% in all patients and 29% in treatment-naive patients; median PFS was 8.3 months (95% CI, 5.8-11.1) and 16.9 months (95% CI, 9.3-35.5), respectively. Median response duration was not reached; 73% of all responses and 82% of treatment-naive responses were ongoing at data cut-off; the longest response was ongoing at 66 months. Four patients [all with prior response of complete response (CR)] whose disease progressed during observation subsequently received second-course pembrolizumab. One patient each achieved CR and partial response (after data cut-off). Treatment-related AEs (TRAEs) occurred in 86% of patients and resulted in study discontinuation in 7.8%; 17% experienced grade 3/4 TRAE. CONCLUSIONS: This 5-year analysis of KEYNOTE-001 represents the longest follow-up for pembrolizumab to date and confirms the durable antitumor activity and tolerability of pembrolizumab in advanced melanoma. CLINICAL TRIAL REGISTRY: ClinicalTrials.gov, NCT01295827.
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Anticorpos Monoclonais Humanizados/administração & dosagem , Antineoplásicos Imunológicos/administração & dosagem , Melanoma/tratamento farmacológico , Neoplasias Cutâneas/tratamento farmacológico , Adulto , Anticorpos Monoclonais Humanizados/efeitos adversos , Antineoplásicos Imunológicos/efeitos adversos , Esquema de Medicação , Seguimentos , Humanos , Estimativa de Kaplan-Meier , Melanoma/mortalidade , Melanoma/patologia , Critérios de Avaliação de Resposta em Tumores Sólidos , Neoplasias Cutâneas/mortalidade , Neoplasias Cutâneas/patologiaRESUMO
BACKGROUND: Immunotherapy (IO) agents can cause late-onset immune-related adverse events (irAEs). In phase I trials, observation for dose-limiting toxicities (DLTs) is typically limited to the first cycle. The incidence of delayed-onset DLTs and their potential impact on dose determination have not been fully elucidated. PATIENTS AND METHODS: Consecutive patients enrolled in early phase IO trials at Princess Margaret Cancer Centre between August 2012 and September 2016 were retrospectively reviewed, applying trial-specific definitions for DLTs. A clinically significant AE (csAE) was defined as a treatment-related adverse event requiring corticosteroids, hormone replacement, IO delay or discontinuation. RESULTS: A total of 352 consecutive trial enrolments in 21 early phase clinical trials were included. Two-hundred seventy-eight patients (79%) received monotherapy and 74 (21%) received combination IO. Two hundred sixty (74%) patients experienced irAEs. There were two protocol-defined DLTs. Twenty (5.7%) patients had 24 csAEs qualifying as DLTs except for occurrence after the protocol-specified DLT period. One-hundred and six (10%) of irAEs were csAEs, including endocrine (26%), respiratory (14%), gastrointestinal (11%), general (10%), dermatological (8%), hepatic (8%), musculoskeletal (6%), pancreatic (6%), haematological, metabolic, neurological, cardiac (each 2%), infective and ocular (each 1%) events. The highest risk of first-onset csAE was during the first 4 weeks compared with the period from 4 weeks to end of treatment (odds ratio 3.13, 95% confidence interval 1.95-5.02). The median time to first onset csAE was significantly shorter with combination than monotherapy IO (32 vs. 146 days, P < 0.001). CONCLUSIONS: In our series of early phase IO trials, the risk of csAE was highest during the initial 4 weeks on IO treatment, supporting the use of the conventional DLT period for dose escalation decision. However, there were 24 clinically significant late-onset DLTs in 5.7% of patients. Combination IO was associated with greater risk of and also earlier onset for csAE, which may need to be considered for early phase trial design.
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Antineoplásicos Imunológicos/efeitos adversos , Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos/diagnóstico , Imunoterapia/efeitos adversos , Neoplasias/tratamento farmacológico , Medição de Risco/métodos , Adulto , Idoso , Idoso de 80 Anos ou mais , Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos/etiologia , Feminino , Seguimentos , Humanos , Masculino , Dose Máxima Tolerável , Pessoa de Meia-Idade , Neoplasias/imunologia , Neoplasias/patologia , Prognóstico , Estudos Retrospectivos , Adulto JovemRESUMO
BACKGROUND: The use and detailed costs of services provided for people with advanced melanoma (amel) are not well known. We conducted an analysis to determine the use of health care services and the associated costs delineated by relevant attributable costs, which we defined for subjects in the province of Ontario. METHODS: Through the Ontario Cancer Data Linkage Project, a cohort of amel patients with diagnoses between 31 August 2005 and 2012 (follow-up to 2013) and with valid International Classification of Diseases (9th revision, Clinical Modification) 172 codes and histology codes was identified. A cohort of individuals with amel having a combination of at least 1 palliative, 1 medical oncology, and 1 hospitalization code was generated. The health system services used by this population were clustered into hospitalization, palliation, physician medical visits, medication, homecare, laboratory, diagnostics, and other resources. Overall rates of use and disaggregated costs were determined by phase of care for the entire cohort. RESULTS: The mean age for the 2748 individuals in the cohort was 67 years. The greater proportion of the patients were men (65.6%) and were more than 65 years of age (>50%). In this advanced cohort, fewer than 45% of patients were alive 3 years after the malignant melanoma diagnosis. The average annual cost per patient over the time horizon was $6,551. At $15,830, year 1 after diagnosis was the most expensive, followed by year 2, at $8,166. CONCLUSIONS: Our data provide a baseline for the costs associated with amel treatment. Future studies will include newer agents and comparative effectiveness research for personalized therapies.
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BACKGROUND: Autophagy is a survival mechanism that allows recycling of cellular breakdown products, particularly in stressed cells. Here we evaluate the hypotheses that up-regulation of autophagy is a common mechanism of resistance to chemotherapy, and that drug resistance can be reversed by inhibiting autophagy with a proton pump inhibitor. METHODS: We exposed human PC3, LNCaP and MCF7 cells to seven clinically-used chemotherapy drugs ± pantoprazole, examined the up-regulation of autophagy and the effect on cellular proliferation by Western Blots, MTS assay and colony-forming assay. The distribution of drug effects and of autophagy was quantified in LNCaP tumor sections in relation to blood vessels and hypoxia by immunohistochemistry using γH2AX, cleaved caspase-3 and p62. RESULTS: All anticancer drugs led to up-regulation of autophagy in cultured tumor cells. Pantoprazole inhibited the induction of autophagy in a time- and dose-dependent manner, and sensitized cancer cells to the seven anti-cancer drugs. Treatment of LNCaP xenografts with paclitaxel induced both DNA damage and autophagy; autophagy was inhibited and markers of toxicity were increased by pantoprazole. CONCLUSIONS: Induction of autophagy is a general mechanism associated with resistance to anticancer drugs and that its inhibition is a promising therapeutic strategy to enhance the effects of chemotherapy and improve clinical outcomes.
Assuntos
2-Piridinilmetilsulfinilbenzimidazóis/farmacologia , Antineoplásicos/uso terapêutico , Autofagia/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Inibidores da Bomba de Prótons/farmacologia , Antineoplásicos Fitogênicos/farmacologia , Vasos Sanguíneos/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células , Relação Dose-Resposta a Droga , Humanos , Hipóxia/patologia , Paclitaxel/farmacologia , Pantoprazol , Microambiente Tumoral/efeitos dos fármacos , Ensaio Tumoral de Célula-Tronco , Regulação para Cima/efeitos dos fármacos , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
BACKGROUND: Detailed epidemiology for patients with advanced metastatic melanoma in Canada is not well characterized. We conducted an analysis of patients with this disease in the province of Ontario, with the aim being to study the presentation, disease characteristics and course, and treatment patterns for malignant melanoma. METHODS: In this Canadian observational prospective and retrospective study of patients with malignant melanoma, we used data collected in the Canadian Melanoma Research Network (cmrn) Patient Registry. We identified patients who were seen at 1 of 3 cancer treatment centres between April 2011 and 30 April 2013. Patient data from 2011 and 2012 were collected retrospectively using chart records and existing registry data. Starting January 2013, data were collected prospectively. Variables investigated included age, sex, initial stage, histology, mutation type, time to recurrence, sites of metastases, resectability, and previous therapies. RESULTS: A cohort of 810 patients with melanoma was identified from the cmrn registry. Mean age was 58.7 years, and most patients were men (60% vs. 40%). Factors affecting survival included unresectable or metastatic melanoma, initial stage at diagnosis, presence of brain metastasis, and BRAF mutation status. The proportion of surviving patients decreased with higher initial disease stages. CONCLUSIONS: Using registry data, we were able to determine the detailed epidemiology of patients with melanoma in the Canadian province of Ontario, validating the comprehensive and detailed information that can be obtained from registry data.
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BACKGROUND: Few prognostic models for overall survival (OS) are available for patients with metastatic castration-resistant prostate cancer (mCRPC) treated with recently approved agents. We developed a prognostic index model using readily available clinical and laboratory factors from a phase III trial of abiraterone acetate (hereafter abiraterone) in combination with prednisone in post-docetaxel mCRPC. PATIENTS AND METHODS: Baseline data were available from 762 patients treated with abiraterone-prednisone. Factors were assessed for association with OS through a univariate Cox model and used in a multivariate Cox model with a stepwise procedure to identify those of significance. Data were validated using an independent, external, population-based cohort. RESULTS: Six risk factors individually associated with poor prognosis were included in the final model: lactate dehydrogenase > upper limit of normal (ULN) [hazard ratio (HR) = 2.31], Eastern Cooperative Oncology Group performance status of 2 (HR = 2.19), presence of liver metastases (HR = 2.00), albumin ≤4 g/dl (HR = 1.54), alkaline phosphatase > ULN (HR = 1.38) and time from start of initial androgen-deprivation therapy to start of treatment ≤36 months (HR = 1.30). Patients were categorized into good (n = 369, 46%), intermediate (n = 321, 40%) and poor (n = 107, 13%) prognosis groups based on the number of risk factors and relative HRs. The C-index was 0.70 ± 0.014. The model was validated by the external dataset (n = 286). CONCLUSION: This analysis identified six factors used to model survival in mCRPC and categorized patients into three distinct risk groups. Prognostic stratification with this model could assist clinical practice decisions for follow-up and monitoring, and may aid in clinical trial design. TRIAL REGISTRATION NUMBERS: NCT00638690.