RESUMEN

Overcoming temozolomide (TMZ)-resistance is a major challenge in glioblastoma therapy. Therefore, identifying the key molecular player in chemo-resistance becomes urgent. We previously reported the downregulation of PDCD10 in primary glioblastoma patients and its tumor suppressor-like function in glioblastoma cells. Here, we demonstrate that the loss of PDCD10 causes a significant TMZ-resistance during treatment and promotes a rapid regrowth of tumor cells after treatment. PDCD10 knockdown upregulated MGMT, a key enzyme mediating chemo-resistance in glioblastoma, accompanied by increased expression of DNA mismatch repair genes, and enabled tumor cells to evade TMZ-induced cell-cycle arrest. These findings were confirmed in independent models of PDCD10 overexpressing cells. Furthermore, PDCD10 downregulation led to the dedifferentiation of glioblastoma cells, as evidenced by increased clonogenic growth, the upregulation of glioblastoma stem cell (GSC) markers, and enhanced neurosphere formation capacity. GSCs derived from PDCD10 knockdown cells displayed stronger TMZ-resistance and regrowth potency, compared to their parental counterparts, indicating that PDCD10-induced stemness may independently contribute to tumor malignancy. These data provide evidence for a dual role of PDCD10 in tumor suppression by controlling both chemo-resistance and dedifferentiation, and highlight PDCD10 as a potential prognostic marker and target for combination therapy with TMZ in glioblastoma.

Asunto(s)

Proteínas Reguladoras de la Apoptosis , Resistencia a Antineoplásicos , Glioblastoma , Temozolomida , Humanos , Glioblastoma/patología , Glioblastoma/genética , Glioblastoma/metabolismo , Glioblastoma/tratamiento farmacológico , Temozolomida/farmacología , Resistencia a Antineoplásicos/genética , Resistencia a Antineoplásicos/efectos de los fármacos , Línea Celular Tumoral , Proteínas Reguladoras de la Apoptosis/metabolismo , Proteínas Reguladoras de la Apoptosis/genética , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Células Madre Neoplásicas/metabolismo , Células Madre Neoplásicas/patología , Células Madre Neoplásicas/efectos de los fármacos , Neoplasias Encefálicas/patología , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/tratamiento farmacológico , Proteínas de la Membrana/metabolismo , Proteínas de la Membrana/genética , Proteínas Proto-Oncogénicas/metabolismo , Proteínas Proto-Oncogénicas/genética , Proliferación Celular/efectos de los fármacos , Metilasas de Modificación del ADN/metabolismo , Metilasas de Modificación del ADN/genética , Proteínas Supresoras de Tumor/metabolismo , Proteínas Supresoras de Tumor/genética , Enzimas Reparadoras del ADN/metabolismo , Enzimas Reparadoras del ADN/genética

RESUMEN

Glioblastoma (GBM) is one of the most aggressive malignant tumors of the brain. We queried PubMed for articles about molecular predictor markers in GBM. This scoping review aims to analyze the most important outcome predictors in patients with GBM and to compare these factors in terms of absolute months of survival benefit and percentages. Performing a gross total resection for patients with GBM undergoing optimal chemo- and radiotherapy provides a significant benefit in overall survival compared to those patients who received a subtotal or partial resection. However, compared to IDH-Wildtype GBMs, patients with IDH-Mutant 1/2 GBMs have an increased survival. MGMT promoter methylation status is another strong outcome predictor for patients with GBM. In the reviewed literature, patients with methylated MGMT promoter lived approximately 50% to 90% longer than those with an unmethylated MGMT gene promoter. Moreover, KPS is an important predictor of survival and quality of life, demonstrating that we should refrain from aggressive surgery in important brain areas. As new therapies (such as TTFs) emerge, we are optimistic that the overall median survival will increase, even for IDH-Wildtype GBMs. In conclusion, molecular profiles are stronger outcome predictors than the extent of neurosurgical resection for GBM.

Asunto(s)

Neoplasias Encefálicas , Glioblastoma , Imagen por Resonancia Magnética , Humanos , Glioblastoma/genética , Glioblastoma/cirugía , Glioblastoma/diagnóstico por imagen , Glioblastoma/patología , Glioblastoma/metabolismo , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/cirugía , Neoplasias Encefálicas/patología , Neoplasias Encefálicas/diagnóstico por imagen , Imagen por Resonancia Magnética/métodos , Metilasas de Modificación del ADN/genética , Metilasas de Modificación del ADN/metabolismo , Enzimas Reparadoras del ADN/genética , Enzimas Reparadoras del ADN/metabolismo , Biomarcadores de Tumor/genética , Proteínas Supresoras de Tumor/genética , Proteínas Supresoras de Tumor/metabolismo , Pronóstico , Metilación de ADN , Isocitrato Deshidrogenasa/genética , Mutación , Regiones Promotoras Genéticas

RESUMEN

Temozolomide (TMZ) resistance is a major challenge in the treatment of glioblastoma (GBM). Tumour reproductive cells (TRCs) have been implicated in the development of chemotherapy resistance. By culturing DBTRG cells in three-dimensional soft fibrin gels to enrich GBM TRCs and performing RNA-seq analysis, the expression of stanniocalcin-1 (STC), a gene encoding a secreted glycoprotein, was found to be upregulated in TRCs. Meanwhile, the viability of TMZ-treated TRC cells was significantly higher than that of TMZ-treated 2D cells. Analysis of clinical data from CGGA (Chinese Glioma Genome Atlas) database showed that high expression of STC1 was closely associated with poor prognosis, glioma grade and resistance to TMZ treatment, suggesting that STC1 may be involved in TMZ drug resistance. The expression of STC1 in tissues and cells was examined, as well as the effect of STC1 on GBM cell proliferation and TMZ-induced DNA damage. The results showed that overexpression of STC1 promoted and knockdown of STC1 inhibited TMZ-induced DNA damage. These results were validated in an intracranial tumour model. These data revealed that STC1 exerts regulatory functions on MGMT expression in GBM, and provides a rationale for targeting STC1 to overcome TMZ resistance.

Asunto(s)

Neoplasias Encefálicas , Resistencia a Antineoplásicos , Regulación Neoplásica de la Expresión Génica , Glioblastoma , Glicoproteínas , Temozolomida , Animales , Femenino , Humanos , Masculino , Ratones , Antineoplásicos Alquilantes/farmacología , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/tratamiento farmacológico , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patología , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Daño del ADN , Metilasas de Modificación del ADN/metabolismo , Metilasas de Modificación del ADN/genética , Enzimas Reparadoras del ADN/genética , Enzimas Reparadoras del ADN/metabolismo , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Glioblastoma/genética , Glioblastoma/metabolismo , Glioblastoma/tratamiento farmacológico , Glioblastoma/patología , Glicoproteínas/metabolismo , Glicoproteínas/genética , Temozolomida/farmacología , Proteínas Supresoras de Tumor/genética , Proteínas Supresoras de Tumor/metabolismo

RESUMEN

It is well known how the precise localization of glioblastoma multiforme (GBM) predicts the direction of tumor spread in the surrounding neuronal structures. The aim of the present review is to reveal the lateralization of GBM by evaluating the anatomical regions where it is frequently located as well as the main molecular alterations observed in different brain regions. According to the literature, the precise or most frequent lateralization of GBM has yet to be determined. However, it can be said that GBM is more frequently observed in the frontal lobe. Tractus and fascicles involved in GBM appear to be focused on the corticospinal tract, superior longitudinal I, II and III fascicles, arcuate fascicle long segment, frontal strait tract, and inferior fronto­occipital fasciculus. Considering the anatomical features of GBM and its brain involvement, it is logical that the main brain regions involved are the frontal­temporal­parietal­occipital lobes, respectively. Although tumor volumes are higher in the right hemisphere, it has been determined that the prognosis of patients diagnosed with cancer in the left hemisphere is worse, probably reflecting the anatomical distribution of some detrimental alterations such as TP53 mutations, PTEN loss, EGFR amplification, and MGMT promoter methylation. There are theories stating that the right hemisphere is less exposed to external influences in its development as it is responsible for the functions necessary for survival while tumors in the left hemisphere may be more aggressive. To shed light on specific anatomical and molecular features of GBM in different brain regions, the present review article is aimed at describing the main lateralization pathways as well as gene mutations or epigenetic modifications associated with the development of brain tumors.

Asunto(s)

Neoplasias Encefálicas , Glioblastoma , Humanos , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/patología , Glioblastoma/genética , Glioblastoma/patología , Glioblastoma/metabolismo , Receptores ErbB/genética , Receptores ErbB/metabolismo , Mutación , Glioma/genética , Glioma/patología , Glioma/metabolismo , Proteína p53 Supresora de Tumor/genética , Proteína p53 Supresora de Tumor/metabolismo , Fosfohidrolasa PTEN/genética , Fosfohidrolasa PTEN/metabolismo , Proteínas Supresoras de Tumor/genética , Proteínas Supresoras de Tumor/metabolismo , Pronóstico , Enzimas Reparadoras del ADN/genética , Enzimas Reparadoras del ADN/metabolismo , Metilasas de Modificación del ADN/genética , Metilasas de Modificación del ADN/metabolismo

RESUMEN

Ubiquitination is a crucial posttranslational modification required for the proper repair of DNA double-strand breaks (DSBs) induced by ionizing radiation (IR). DSBs are mainly repaired through homologous recombination (HR) when template DNA is present and nonhomologous end joining (NHEJ) in its absence. In addition, microhomology-mediated end joining (MMEJ) and single-strand annealing (SSA) provide backup DSBs repair pathways. However, the mechanisms controlling their use remain poorly understood. By using a high-resolution CRISPR screen of the ubiquitin system after IR, we systematically uncover genes required for cell survival and elucidate a critical role of the E3 ubiquitin ligase SCFcyclin F in cell cycle-dependent DSB repair. We show that SCFcyclin F-mediated EXO1 degradation prevents DNA end resection in mitosis, allowing MMEJ to take place. Moreover, we identify a conserved cyclin F recognition motif, distinct from the one used by other cyclins, with broad implications in cyclin specificity for cell cycle control.

Asunto(s)

Ciclo Celular , Ciclinas , Roturas del ADN de Doble Cadena , Reparación del ADN , Exodesoxirribonucleasas , Humanos , Ciclo Celular/genética , Exodesoxirribonucleasas/metabolismo , Exodesoxirribonucleasas/genética , Ciclinas/metabolismo , Ciclinas/genética , Enzimas Reparadoras del ADN/metabolismo , Enzimas Reparadoras del ADN/genética , Reparación del ADN por Unión de Extremidades , Ubiquitinación , Radiación Ionizante

RESUMEN

OBJECTIVES: High-grade gliomas (HGGs) are highly malignant, aggressive, and have a high incidence and mortality rate. The aim of this study was to investigate survival outcomes and prognostic factors in patients with HGGs. METHODS: In this retrospective study, a total of 159 patients with histologically confirmed HGGs were included. The recruitment period was from January 2011 to December 2019. We evaluated patient demographic data, tumor characteristics, treatment methods, immunocytochemistry results, overall survival (OS) time, and progression-free survival (PFS) time using Kaplan-<>Meier survival analysis with log-rank testing. Additionally, we employed Cox regression analysis to identify independent factors associated with survival outcomes. RESULTS: Kaplan-Meier survival analysis revealed that the 1-, 2-, and 5-years OS rates were 81.8%, 50.3%, and 12.6%, respectively. Similarly, the 1-, 2-, and 5-years PFS rates were 50.9%, 22.4%, and 3.1%, respectively. The median OS duration was 35.0 months. The univariate analysis indicated that postoperative pathological classification, grade, and age were significantly associated with patient outcomes (p < 0.01). Among the patients, 147 received concurrent chemoradiotherapy, while 12 did not. The immunohistochemical markers of ki-67, MGMT, IDH1R132H, and p53 demonstrated statistically significant differences in their prognostic impact (p = 0.001, p = 0.020, p = 0.003, and p = 0.021, respectively). In conclusion, we found that grades, age, pathological classification, ki-67, MGMT, and IDH1R132H expression were statistically significantly associated with PFS (p < 0.01, p = 0.004, p = 0.003, p = 0.001, p = 0.036, and p = 0.028). Additionally, immunohistochemical expressions of TRIB3 and AURKA were significantly higher in patients with shorter survival (p = 0.015 and p = 0.023). CONCLUSIONS: Tumor grade and the use of concurrent chemoradiotherapy after surgery were independent prognostic factors that significantly influenced patient survival. Additionally, tumor grade and MGMT expression were found to be independent factors affecting progression-free survival (PFS). Notably, the expression of TRIB3 and AURKA was higher in patients with poor survival outcomes.

Asunto(s)

Neoplasias Encefálicas , Glioma , Clasificación del Tumor , Humanos , Femenino , Masculino , Glioma/mortalidad , Glioma/patología , Glioma/terapia , Glioma/metabolismo , Estudios Retrospectivos , Persona de Mediana Edad , Adulto , Pronóstico , Anciano , Neoplasias Encefálicas/mortalidad , Neoplasias Encefálicas/terapia , Neoplasias Encefálicas/patología , Neoplasias Encefálicas/metabolismo , Quimioradioterapia , Adulto Joven , Estimación de Kaplan-Meier , Biomarcadores de Tumor/metabolismo , Supervivencia sin Progresión , Isocitrato Deshidrogenasa/genética , Isocitrato Deshidrogenasa/metabolismo , Metilasas de Modificación del ADN/genética , Metilasas de Modificación del ADN/metabolismo , Análisis Multivariante , Proteínas Supresoras de Tumor/metabolismo , Tasa de Supervivencia , Adolescente , Enzimas Reparadoras del ADN/metabolismo , Enzimas Reparadoras del ADN/genética , Enzimas Reparadoras del ADN/análisis

RESUMEN

Senataxin is an evolutionarily conserved DNA/RNA helicase, whose dysfunctions are linked to neurodegeneration and cancer. A main activity of this protein is the removal of R-loops, which are nucleic acid structures capable to promote DNA damage and replication stress. Here we found that Senataxin deficiency causes the release of damaged DNA into extranuclear bodies, called micronuclei, triggering the massive recruitment of cGAS, the apical sensor of the innate immunity pathway, and the downstream stimulation of interferon genes. Such cGAS-positive micronuclei are characterized by defective membrane envelope and are particularly abundant in cycling cells lacking Senataxin, but not after exposure to a DNA breaking agent or in absence of the tumor suppressor BRCA1 protein, a partner of Senataxin in R-loop removal. Micronuclei with a discontinuous membrane are normally cleared by autophagy, a process that we show is impaired in Senataxin-deficient cells. The formation of Senataxin-dependent inflamed micronuclei is promoted by the persistence of nuclear R-loops stimulated by the DSIF transcription elongation complex and the engagement of EXO1 nuclease activity on nuclear DNA. Coherently, high levels of EXO1 result in poor prognosis in a subset of tumors lacking Senataxin expression. Hence, R-loop homeostasis impairment, together with autophagy failure and unscheduled EXO1 activity, elicits innate immune response through micronuclei formation in cells lacking Senataxin.

Asunto(s)

Autofagia , Daño del ADN , ADN Helicasas , Inflamación , Enzimas Multifuncionales , Nucleotidiltransferasas , Estructuras R-Loop , ARN Helicasas , Humanos , Autofagia/genética , Proteína BRCA1/metabolismo , Proteína BRCA1/genética , Proteína BRCA1/deficiencia , ADN Helicasas/metabolismo , ADN Helicasas/genética , ADN Helicasas/deficiencia , Enzimas Reparadoras del ADN/metabolismo , Enzimas Reparadoras del ADN/genética , Enzimas Reparadoras del ADN/deficiencia , Exodesoxirribonucleasas/metabolismo , Exodesoxirribonucleasas/genética , Inmunidad Innata , Inflamación/patología , Inflamación/metabolismo , Inflamación/genética , Enzimas Multifuncionales/metabolismo , Enzimas Multifuncionales/genética , Nucleotidiltransferasas/metabolismo , Nucleotidiltransferasas/genética , Fosfoproteínas , ARN Helicasas/metabolismo , ARN Helicasas/genética

RESUMEN

Cockayne syndrome (CS) is a premature ageing condition characterized by microcephaly, growth failure, and neurodegeneration. It is caused by mutations in ERCC6 or ERCC8 encoding for Cockayne syndrome B (CSB) and A (CSA) proteins, respectively. CSA and CSB have well-characterized roles in transcription-coupled nucleotide excision repair, responsible for removing bulky DNA lesions, including those caused by UV irradiation. Here, we report that CSA dysfunction causes defects in the nuclear envelope (NE) integrity. NE dysfunction is characteristic of progeroid disorders caused by a mutation in NE proteins, such as Hutchinson-Gilford progeria syndrome. However, it has never been reported in Cockayne syndrome. We observed CSA dysfunction affected LEMD2 incorporation at the NE and increased actin stress fibers that contributed to enhanced mechanical stress to the NE. Altogether, these led to NE abnormalities associated with the activation of the cGAS/STING pathway. Targeting the linker of the nucleoskeleton and cytoskeleton complex was sufficient to rescue these phenotypes. This work reveals NE dysfunction in a progeroid syndrome caused by mutations in a DNA damage repair protein, reinforcing the connection between NE deregulation and ageing.

Asunto(s)

Síndrome de Cockayne , Enzimas Reparadoras del ADN , Reparación del ADN , Membrana Nuclear , Proteínas de Unión a Poli-ADP-Ribosa , Membrana Nuclear/metabolismo , Humanos , Síndrome de Cockayne/genética , Síndrome de Cockayne/metabolismo , Enzimas Reparadoras del ADN/genética , Enzimas Reparadoras del ADN/metabolismo , Proteínas de Unión a Poli-ADP-Ribosa/metabolismo , Proteínas de Unión a Poli-ADP-Ribosa/genética , Daño del ADN/genética , ADN Helicasas/genética , ADN Helicasas/metabolismo , Mutación , Proteínas de la Membrana/metabolismo , Proteínas de la Membrana/genética , Progeria/genética , Progeria/metabolismo , Proteínas Nucleares/metabolismo , Proteínas Nucleares/genética , Factores de Transcripción

RESUMEN

DNA damage has been implicated in the stimulation of the type 1 interferon (T1IFN) response. Here, we show that downregulation of the DNA repair protein, polynucleotide kinase/phosphatase (PNKP), in a variety of cell lines causes robust phosphorylation of STAT1, upregulation of interferon-stimulated genes and persistent accumulation of cytosolic DNA, all of which are indicators for the activation of the T1IFN response. Furthermore, this did not require damage induction by ionizing radiation. Instead, our data revealed that production of reactive oxygen species (ROS) synergises with PNKP loss to potentiate the T1IFN response, and that loss of PNKP significantly compromises mitochondrial DNA (mtDNA) integrity. Depletion of mtDNA or treatment of PNKP-depleted cells with ROS scavengers abrogated the T1IFN response, implicating mtDNA as a significant source of the cytosolic DNA required to potentiate the T1IFN response. The STING signalling pathway is responsible for the observed increase in the pro-inflammatory gene signature in PNKP-depleted cells. While the response was dependent on ZBP1, cGAS only contributed to the response in some cell lines. Our data have implications for cancer therapy, since PNKP inhibitors would have the potential to stimulate the immune response, and also to the neurological disorders associated with PNKP mutation.

Asunto(s)

Enzimas Reparadoras del ADN , ADN Mitocondrial , Interferón Tipo I , Fosfotransferasas (Aceptor de Grupo Alcohol) , Radiación Ionizante , Especies Reactivas de Oxígeno , Humanos , Interferón Tipo I/metabolismo , Interferón Tipo I/genética , Fosfotransferasas (Aceptor de Grupo Alcohol)/genética , Fosfotransferasas (Aceptor de Grupo Alcohol)/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Enzimas Reparadoras del ADN/metabolismo , Enzimas Reparadoras del ADN/genética , ADN Mitocondrial/genética , ADN Mitocondrial/metabolismo , Reparación del ADN , Factor de Transcripción STAT1/metabolismo , Factor de Transcripción STAT1/genética , Daño del ADN , Línea Celular , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Transducción de Señal , Nucleotidiltransferasas/metabolismo , Nucleotidiltransferasas/genética , Fosforilación , Citosol/metabolismo , Línea Celular Tumoral , Proteínas de Unión al ARN/metabolismo , Proteínas de Unión al ARN/genética

RESUMEN

Cockayne Syndrome B (CSB) is a hereditary multiorgan syndrome which-through largely unknown mechanisms-can affect the brain where it clinically presents with microcephaly, intellectual disability and demyelination. Using human induced pluripotent stem cell (hiPSC)-derived neural 3D models generated from CSB patient-derived and isogenic control lines, we here provide explanations for these three major neuropathological phenotypes. In our models, CSB deficiency is associated with (i) impaired cellular migration due to defective autophagy as an explanation for clinical microcephaly; (ii) altered neuronal network functionality and neurotransmitter GABA levels, which is suggestive of a disturbed GABA switch that likely impairs brain circuit formation and ultimately causes intellectual disability; and (iii) impaired oligodendrocyte maturation as a possible cause of the demyelination observed in children with CSB. Of note, the impaired migration and oligodendrocyte maturation could both be partially rescued by pharmacological HDAC inhibition.

Asunto(s)

Síndrome de Cockayne , Células Madre Pluripotentes Inducidas , Oligodendroglía , Humanos , Células Madre Pluripotentes Inducidas/metabolismo , Células Madre Pluripotentes Inducidas/citología , Células Madre Pluripotentes Inducidas/patología , Síndrome de Cockayne/genética , Síndrome de Cockayne/metabolismo , Síndrome de Cockayne/patología , Oligodendroglía/metabolismo , Oligodendroglía/citología , Movimiento Celular , Enzimas Reparadoras del ADN/metabolismo , Enzimas Reparadoras del ADN/genética , Neuronas/metabolismo , Neuronas/patología , Autofagia , Encéfalo/metabolismo , Encéfalo/patología , Proteínas de Unión a Poli-ADP-Ribosa/metabolismo , Proteínas de Unión a Poli-ADP-Ribosa/genética , Ácido gamma-Aminobutírico/metabolismo , ADN Helicasas/metabolismo , ADN Helicasas/genética , Microcefalia/patología , Microcefalia/metabolismo , Microcefalia/genética , Enfermedades Desmielinizantes/patología , Enfermedades Desmielinizantes/metabolismo , Diferenciación Celular

Asunto(s)

Metilasas de Modificación del ADN , Enzimas Reparadoras del ADN , Resistencia a Antineoplásicos , Glioblastoma , Células Madre Neoplásicas , Temozolomida , Regulación hacia Arriba , Glioblastoma/tratamiento farmacológico , Glioblastoma/genética , Glioblastoma/metabolismo , Glioblastoma/patología , Temozolomida/uso terapéutico , Temozolomida/farmacología , Humanos , Células Madre Neoplásicas/efectos de los fármacos , Células Madre Neoplásicas/metabolismo , Metilasas de Modificación del ADN/metabolismo , Metilasas de Modificación del ADN/genética , Enzimas Reparadoras del ADN/genética , Enzimas Reparadoras del ADN/metabolismo , Resistencia a Antineoplásicos/efectos de los fármacos , Resistencia a Antineoplásicos/genética , Regulación hacia Arriba/efectos de los fármacos , Proteínas Supresoras de Tumor/genética , Proteínas Supresoras de Tumor/metabolismo , Antineoplásicos Alquilantes/uso terapéutico , Antineoplásicos Alquilantes/farmacología

RESUMEN

Trypanosoma cruzi is the etiological agent of Chagas disease and a peculiar eukaryote with unique biological characteristics. DNA damage can block RNA polymerase, activating transcription-coupled nucleotide excision repair (TC-NER), a DNA repair pathway specialized in lesions that compromise transcription. If transcriptional stress is unresolved, arrested RNA polymerase can activate programmed cell death. Nonetheless, how this parasite modulates these processes is unknown. Here, we demonstrate that T. cruzi cell death after UV irradiation, a genotoxic agent that generates lesions resolved by TC-NER, depends on active transcription and is signaled mainly by an apoptotic-like pathway. Pre-treated parasites with α-amanitin, a selective RNA polymerase II inhibitor, become resistant to such cell death. Similarly, the gamma pre-irradiated cells are more resistant to UV when the transcription processes are absent. The Cockayne Syndrome B protein (CSB) recognizes blocked RNA polymerase and can initiate TC-NER. Curiously, CSB overexpression increases parasites' cell death shortly after UV exposure. On the other hand, at the same time after irradiation, the single-knockout CSB cells show resistance to the same treatment. UV-induced fast death is signalized by the exposition of phosphatidylserine to the outer layer of the membrane, indicating a cell death mainly by an apoptotic-like pathway. Furthermore, such death is suppressed in WT parasites pre-treated with inhibitors of ataxia telangiectasia and Rad3-related (ATR), a key DDR kinase. Signaling for UV radiation death may be related to R-loops since the overexpression of genes associated with the resolution of these structures suppress it. Together, results suggest that transcription blockage triggered by UV radiation activates an ATR-dependent apoptosis-like mechanism in T. cruzi, with the participation of CSB protein in this process.

Asunto(s)

Proteínas de la Ataxia Telangiectasia Mutada , Daño del ADN , Reparación del ADN , Estructuras R-Loop , Transcripción Genética , Trypanosoma cruzi , Rayos Ultravioleta , Trypanosoma cruzi/metabolismo , Proteínas de la Ataxia Telangiectasia Mutada/metabolismo , Proteínas de Unión a Poli-ADP-Ribosa/metabolismo , Proteínas de Unión a Poli-ADP-Ribosa/genética , Enzimas Reparadoras del ADN/metabolismo , Enzimas Reparadoras del ADN/genética , Proteínas Protozoarias/metabolismo , ADN Helicasas/metabolismo , ADN Helicasas/genética , Muerte Celular , Apoptosis , Humanos

RESUMEN

Glioblastoma (GBM) is a highly aggressive brain tumor associated with poor patient survival. The current standard treatment involves invasive surgery, radiotherapy, and chemotherapy employing temozolomide (TMZ). Resistance to TMZ is, however, a major challenge. Previous work from our group has identified candidate genes linked to TMZ resistance, including genes encoding translesion synthesis (TLS) DNA polymerases iota (PolÉ©) and kappa (Polκ). These specialized enzymes are known for bypassing lesions and tolerating DNA damage. Here, we investigated the roles of PolÉ© and Polκ in TMZ resistance, employing MGMT-deficient U251-MG glioblastoma cells, with knockout of either POLI or POLK genes encoding PolÉ© and Polκ, respectively, and assess their viability and genotoxic stress responses upon subsequent TMZ treatment. Cells lacking either of these polymerases exhibited a significant decrease in viability following TMZ treatment compared to parental counterparts. The restoration of the missing polymerase led to a recovery of cell viability. Furthermore, knockout cells displayed increased cell cycle arrest, mainly in late S-phase, and lower levels of genotoxic stress after TMZ treatment, as assessed by a reduction of γH2AX foci and flow cytometry data. This implies that TMZ treatment does not trigger a significant H2AX phosphorylation response in the absence of these proteins. Interestingly, combining TMZ with Mirin (double-strand break repair pathway inhibitor) further reduced the cell viability and increased DNA damage and γH2AX positive cells in TLS KO cells, but not in parental cells. These findings underscore the crucial roles of PolÉ© and Polκ in conferring TMZ resistance and the potential backup role of homologous recombination in the absence of these TLS polymerases. Targeting these TLS enzymes, along with double-strand break DNA repair inhibition, could, therefore, provide a promising strategy to enhance TMZ's effectiveness in treating GBM.

Asunto(s)

Metilasas de Modificación del ADN , ADN Polimerasa iota , Enzimas Reparadoras del ADN , ADN Polimerasa Dirigida por ADN , Resistencia a Antineoplásicos , Glioblastoma , Temozolomida , Temozolomida/farmacología , Humanos , Glioblastoma/genética , Glioblastoma/tratamiento farmacológico , Glioblastoma/metabolismo , Glioblastoma/patología , ADN Polimerasa Dirigida por ADN/metabolismo , ADN Polimerasa Dirigida por ADN/genética , Línea Celular Tumoral , Metilasas de Modificación del ADN/metabolismo , Metilasas de Modificación del ADN/genética , Enzimas Reparadoras del ADN/metabolismo , Enzimas Reparadoras del ADN/genética , Proteínas Supresoras de Tumor/metabolismo , Proteínas Supresoras de Tumor/genética , Proteínas Supresoras de Tumor/deficiencia , Antineoplásicos Alquilantes/farmacología , Antineoplásicos Alquilantes/uso terapéutico , Daño del ADN , Supervivencia Celular/efectos de los fármacos , Neoplasias Encefálicas/tratamiento farmacológico , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patología , Reparación del ADN , Técnicas de Inactivación de Genes

RESUMEN

Glioblastoma (GBM) presents a daunting challenge due to its resistance to temozolomide (TMZ), a hurdle exacerbated by the proneural-to-mesenchymal transition (PMT) from a proneural (PN) to a mesenchymal (MES) phenotype. TAGLN2 is prominently expressed in GBM, particularly in the MES subtype compared to low-grade glioma (LGG) and the PN subtype. Our research reveals TAGLN2's involvement in PMT and TMZ resistance through a series of in vitro and in vivo experiments. TAGLN2 knockdown can restrain proliferation and invasion, trigger DNA damage and apoptosis, and heighten TMZ sensitivity in GBM cells. Conversely, elevating TAGLN2 levels amplifies resistance to TMZ in cellular and intracranial xenograft mouse models. We demonstrate the interaction relationship between TAGLN2 and ERK1/2 through co-immunoprecipitation (Co-IP) and liquid chromatography-tandem mass spectrometry (LC-MS/MS) spectrometry analysis. Knockdown of TAGLN2 results in a decrease in the expression of p-ERK1/2, whereas overexpression of TAGLN2 leads to an increase in p-ERK1/2 expression within the nucleus. Subsequently, the regulatory role of TAGLN2 in the expression and control of MGMT has been demonstrated. Finally, the regulation of TAGLN2 by NF-κB has been validated through chromatin immunoprecipitation and ChIP-PCR assays. In conclusion, our results confirm that TAGLN2 exerts its biological functions by interacting with the ERK/MGMT axis and being regulated by NF-κB, thereby facilitating the acquisition of promoting PMT and increased resistance to TMZ therapy in glioblastoma. These results provide valuable insights for the advancement of targeted therapeutic approaches to overcome TMZ resistance in clinical treatments.

Asunto(s)

Antineoplásicos Alquilantes , Neoplasias Encefálicas , Resistencia a Antineoplásicos , Glioblastoma , Temozolomida , Animales , Humanos , Ratones , Antineoplásicos Alquilantes/farmacología , Apoptosis/efectos de los fármacos , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/patología , Neoplasias Encefálicas/tratamiento farmacológico , Neoplasias Encefálicas/metabolismo , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Metilasas de Modificación del ADN/metabolismo , Metilasas de Modificación del ADN/genética , Enzimas Reparadoras del ADN/genética , Enzimas Reparadoras del ADN/metabolismo , Transición Epitelial-Mesenquimal/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Glioblastoma/patología , Glioblastoma/genética , Glioblastoma/tratamiento farmacológico , Glioblastoma/metabolismo , Ratones Desnudos , Proteínas de Microfilamentos/genética , Proteínas de Microfilamentos/metabolismo , Temozolomida/farmacología , Proteínas Supresoras de Tumor/genética , Proteínas Supresoras de Tumor/metabolismo , Ensayos Antitumor por Modelo de Xenoinjerto

RESUMEN

Background.Glioblastoma Multiforme (GBM) is an aggressive form of malignant brain tumor with a generally poor prognosis.O6-methylguanine-DNA methyltransferase (MGMT) promoter methylation has been shown to be a predictive bio-marker for resistance to treatment of GBM, but it is invasive and time-consuming to determine methylation status. There has been effort to predict the MGMT methylation status through analyzing MRI scans using machine learning, which only requires pre-operative scans that are already part of standard-of-care for GBM patients.Purpose.To improve the performance of conventional transfer learning in the identification of MGMT promoter methylation status, we developed a 3D SpotTune network with adaptive fine-tuning capability. Using the pretrained weights of MedicalNet with the SpotTune network, we compared its performance with a randomly initialized network for different combinations of MR modalities.Methods.Using a ResNet50 as the base network, three categories of networks are created: (1) A 3D SpotTune network to process volumetric MR images, (2) a network with randomly initialized weights, and (3) a network pre-trained on MedicalNet. These three networks are trained and evaluated using a public GBM dataset provided by the University of Pennsylvania. The MRI scans from 240 patients are used, with 11 different modalities corresponding to a set of perfusion, diffusion, and structural scans. The performance is evaluated using 5-fold cross validation with a hold-out testing dataset.Results.The SpotTune network showed better performance than the randomly initialized network. The best performing SpotTune model achieved an area under the Receiver Operating Characteristic curve (AUC), average precision of the precision-recall curve (AP), sensitivity, and specificity values of 0.6604, 0.6179, 0.6667, and 0.6061 respectively.Conclusions.SpotTune enables transfer learning to be adaptive to individual patients, resulting in improved performance in predicting MGMT promoter methylation status in GBM using equivalent MRI modalities as compared to a randomly initialized network.

Asunto(s)

Neoplasias Encefálicas , Metilación de ADN , Metilasas de Modificación del ADN , Enzimas Reparadoras del ADN , Glioblastoma , Imagen por Resonancia Magnética , Regiones Promotoras Genéticas , Proteínas Supresoras de Tumor , Humanos , Glioblastoma/genética , Glioblastoma/diagnóstico por imagen , Imagen por Resonancia Magnética/métodos , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/diagnóstico por imagen , Metilasas de Modificación del ADN/genética , Metilasas de Modificación del ADN/metabolismo , Proteínas Supresoras de Tumor/genética , Proteínas Supresoras de Tumor/metabolismo , Enzimas Reparadoras del ADN/genética , Enzimas Reparadoras del ADN/metabolismo , Aprendizaje Automático , Curva ROC , Masculino , Femenino , Redes Neurales de la Computación , Adulto , Algoritmos

RESUMEN

DNA damage severely impedes gene transcription by RNA polymerase II (Pol II), causing cellular dysfunction. Transcription-Coupled Nucleotide Excision Repair (TC-NER) specifically removes such transcription-blocking damage. TC-NER initiation relies on the CSB, CSA and UVSSA proteins; loss of any results in complete TC-NER deficiency. Strikingly, UVSSA deficiency results in UV-Sensitive Syndrome (UVSS), with mild cutaneous symptoms, while loss of CSA or CSB activity results in the severe Cockayne Syndrome (CS), characterized by neurodegeneration and premature aging. Thus far the underlying mechanism for these contrasting phenotypes remains unclear. Live-cell imaging approaches reveal that in TC-NER proficient cells, lesion-stalled Pol II is swiftly resolved, while in CSA and CSB knockout (KO) cells, elongating Pol II remains damage-bound, likely obstructing other DNA transacting processes and shielding the damage from alternative repair pathways. In contrast, in UVSSA KO cells, Pol II is cleared from the damage via VCP-mediated proteasomal degradation which is fully dependent on the CRL4CSA ubiquitin ligase activity. This Pol II degradation might provide access for alternative repair mechanisms, such as GG-NER, to remove the damage. Collectively, our data indicate that the inability to clear lesion-stalled Pol II from the chromatin, rather than TC-NER deficiency, causes the severe phenotypes observed in CS.

Asunto(s)

Síndrome de Cockayne , Daño del ADN , ADN Helicasas , Enzimas Reparadoras del ADN , Reparación del ADN , Proteínas de Unión a Poli-ADP-Ribosa , ARN Polimerasa II , Transcripción Genética , ARN Polimerasa II/metabolismo , ARN Polimerasa II/genética , Humanos , Enzimas Reparadoras del ADN/metabolismo , Enzimas Reparadoras del ADN/genética , Proteínas de Unión a Poli-ADP-Ribosa/genética , Proteínas de Unión a Poli-ADP-Ribosa/metabolismo , ADN Helicasas/metabolismo , ADN Helicasas/genética , Síndrome de Cockayne/genética , Síndrome de Cockayne/metabolismo , Factores de Transcripción/metabolismo , Factores de Transcripción/genética , Proteína que Contiene Valosina/metabolismo , Proteína que Contiene Valosina/genética , Proteínas de Ciclo Celular/metabolismo , Proteínas de Ciclo Celular/genética , Adenosina Trifosfatasas/metabolismo , Adenosina Trifosfatasas/genética , Rayos Ultravioleta , Línea Celular , Reparación por Escisión , Proteínas Portadoras

RESUMEN

BACKGROUND: MutT homolog 1 (MTH1) sanitizes oxidized dNTP pools to promote the survival of cancer cells and its expression is frequently upregulated in cancers. Polyubiquitination stabilizes MTH1 to facilitate the proliferation of melanoma cells, suggesting the ubiquitin system controls the stability and function of MTH1. However, whether ubiquitination regulates MTH1 in gastric cancers has not been well defined. This study aims to investigate the interaction between MTH1 and a deubiquitinase, USP9X, in regulating the proliferation, survival, migration, and invasion of gastric cancer cells. METHODS: The interaction between USP9X and MTH1 was evaluated by co-immunoprecipitation (co-IP) in HGC-27 gastric cancer cells. siRNAs were used to interfere with USP9X expression in gastric cancer cell lines HGC-27 and MKN-45. MTT assays were carried out to examine the proliferation, propidium iodide (PI) and 7-AAD staining assays were performed to assess the cell cycle, Annexin V/PI staining assays were conducted to examine the apoptosis, and transwell assays were used to determine the migration and invasion of control, USP9X-deficient, and USP9X-deficient plus MTH1-overexpressing HGC-27 and MKN-45 gastric cancer cells. RESULTS: Co-IP data show that USP9X interacts with and deubiquitinates MTH1. Overexpression of USP9X elevates MTH1 protein level by downregulating its ubiquitination, while knockdown of USP9X has the opposite effect on MTH1. USP9X deficiency in HGC-27 and MKN-45 cells causes decreased proliferation, cell cycle arrest, extra apoptosis, and defective migration and invasion, which could be rescued by excessive MTH1. CONCLUSION: USP9X interacts with and stabilizes MTH1 to promote the proliferation, survival, migration and invasion of gastric cancer cells.

Asunto(s)

Movimiento Celular , Proliferación Celular , Enzimas Reparadoras del ADN , Invasividad Neoplásica , Monoéster Fosfórico Hidrolasas , Neoplasias Gástricas , Ubiquitina Tiolesterasa , Humanos , Apoptosis , Línea Celular Tumoral , Proliferación Celular/genética , Supervivencia Celular , Enzimas Reparadoras del ADN/metabolismo , Enzimas Reparadoras del ADN/genética , Monoéster Fosfórico Hidrolasas/metabolismo , Monoéster Fosfórico Hidrolasas/genética , ARN Interferente Pequeño , Neoplasias Gástricas/patología , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/genética , Ubiquitina Tiolesterasa/metabolismo , Ubiquitina Tiolesterasa/genética , Ubiquitinación , Hidrolasas Nudix/genética , Hidrolasas Nudix/metabolismo

RESUMEN

African swine fever virus (ASFV), as a contagious viral pathogen, is responsible for the occurrence of African swine fever (ASF), a rapidly spreading and highly lethal disease. Since ASFV was introduced into China in 2018, it has been quickly spread to many provinces, which brought great challenges to the pig industry in China. Due to the limited knowledge about the pathogenesis of ASFV, neither vaccines nor antiviral drugs are available. We have found that ASFV infection can induce oxidative stress responses in cells, and DNA repair enzymes play a key role in this process. This study employed RNA interference, RT-qPCR, Western blotting, Hemadsorption (HAD), and flow cytometry to investigate the effects of the inhibitors of DNA repair enzymes OGG1 and MTH1 on ASFV replication and evaluated the anti-ASFV effects of the inhibitors. This study provides reference for the development of anti-viral drugs.

Asunto(s)

Virus de la Fiebre Porcina Africana , ADN Glicosilasas , Monoéster Fosfórico Hidrolasas , Replicación Viral , Virus de la Fiebre Porcina Africana/genética , Virus de la Fiebre Porcina Africana/efectos de los fármacos , Animales , Replicación Viral/efectos de los fármacos , Porcinos , ADN Glicosilasas/metabolismo , ADN Glicosilasas/genética , Monoéster Fosfórico Hidrolasas/genética , Monoéster Fosfórico Hidrolasas/antagonistas & inhibidores , Monoéster Fosfórico Hidrolasas/metabolismo , Enzimas Reparadoras del ADN/genética , Enzimas Reparadoras del ADN/metabolismo , Fiebre Porcina Africana/virología , Antivirales/farmacología , Interferencia de ARN , ARN Interferente Pequeño/genética , Inhibidores Enzimáticos/farmacología , Estrés Oxidativo/efectos de los fármacos , Células Vero

RESUMEN

OBJECTIVE: Analysis of long-lived patients from the group of patients with glioblastomas after using photodynamic therapy in the structure of their complex treatment in order to assess the influence of various factors on their life expectancy. MATERIAL AND METHODS: In total, a single-center, retrospective categorical study analyzed the long-term results of treatment of 63 patients with glioblastoma in the structure of complex treatment including photodynamic therapy. Clinical factors (age, sex, number of cases, preoperative Karnofsky index, location and size of the tumor, radicality of the operation), histological (nuclear polymorphism, mitosis, vascular proliferation, necrosis), immunohistochemical (Ki-67, p53 index) molecular-genetic factors (expression of VEGF, MGMT, IDH, CD34), amount of radiation and chemotherapy were analyzed. RESULTS: In the entire group of patients, there was a direct correlation of life expectancy with MGMT status, IDH status, the number of courses of chemotherapy, the age of the patient, and the severity of the first surgical intervention. CONCLUSION: Clinical features such as age at diagnosis and extent of surgical resection and amount of chemotherapy have predictive value in assessing their effect on life expectancy. Mutations in IDH and MGMT promoter methylation were the most important molecular factors determining long-term patient survival.

Asunto(s)

Neoplasias Encefálicas , Fotoquimioterapia , Humanos , Masculino , Femenino , Persona de Mediana Edad , Fotoquimioterapia/métodos , Neoplasias Encefálicas/tratamiento farmacológico , Neoplasias Encefálicas/mortalidad , Estudios Retrospectivos , Adulto , Anciano , Glioblastoma/tratamiento farmacológico , Glioblastoma/mortalidad , Glioblastoma/genética , Proteínas Supresoras de Tumor/genética , Metilasas de Modificación del ADN/genética , Metilasas de Modificación del ADN/metabolismo , Isocitrato Deshidrogenasa/genética , Enzimas Reparadoras del ADN/genética , Enzimas Reparadoras del ADN/metabolismo , Glioma/tratamiento farmacológico , Glioma/mortalidad , Glioma/genética , Tasa de Supervivencia , Esperanza de Vida

RESUMEN

Mutations in the Cockayne Syndrome group B (CSB) gene cause cancer in mice, but premature aging and severe neurodevelopmental defects in humans. CSB, a member of the SWI/SNF family of chromatin remodelers, plays diverse roles in regulating gene expression and transcription-coupled nucleotide excision repair (TC-NER); however, these functions do not explain the distinct phenotypic differences observed between CSB-deficient mice and humans. During investigating Cockayne Syndrome-associated genome instability, we uncover an intrinsic mechanism that involves elongating RNA polymerase II (RNAPII) undergoing transient pauses at internal T-runs where CSB is required to propel RNAPII forward. Consequently, CSB deficiency retards RNAPII elongation in these regions, and when coupled with G-rich sequences upstream, exacerbates genome instability by promoting R-loop formation. These R-loop prone motifs are notably abundant in relatively long genes related to neuronal functions in the human genome, but less prevalent in the mouse genome. These findings provide mechanistic insights into differential impacts of CSB deficiency on mice versus humans and suggest that the manifestation of the Cockayne Syndrome phenotype in humans results from the progressive evolution of mammalian genomes.

Asunto(s)

Síndrome de Cockayne , ADN Helicasas , Enzimas Reparadoras del ADN , Inestabilidad Genómica , Proteínas de Unión a Poli-ADP-Ribosa , Estructuras R-Loop , ARN Polimerasa II , Síndrome de Cockayne/genética , Síndrome de Cockayne/patología , Síndrome de Cockayne/metabolismo , ARN Polimerasa II/metabolismo , ARN Polimerasa II/genética , Animales , Humanos , Proteínas de Unión a Poli-ADP-Ribosa/genética , Proteínas de Unión a Poli-ADP-Ribosa/metabolismo , Enzimas Reparadoras del ADN/metabolismo , Enzimas Reparadoras del ADN/genética , Ratones , ADN Helicasas/metabolismo , ADN Helicasas/genética , Estructuras R-Loop/genética , Reparación del ADN , Elongación de la Transcripción Genética , Ratones Noqueados