RESUMEN
BACKGROUND: Sodium-glucose cotransporter 2 inhibitors (SGLT-2i) are glucose-lowering agents used for the treatment of type 2 diabetes mellitus, which also improve heart failure and decrease the risk of cardiovascular complications. Epicardial adipose tissue (EAT) dysfunction was suggested to contribute to the development of heart failure. We aimed to elucidate a possible role of changes in EAT metabolic and inflammatory profile in the beneficial cardioprotective effects of SGLT-2i in subjects with severe heart failure. METHODS: 26 subjects with severe heart failure, with reduced ejection fraction, treated with SGLT-2i versus 26 subjects without treatment, matched for age (54.0 ± 2.1 vs. 55.3 ± 2.1 years, n.s.), body mass index (27.8 ± 0.9 vs. 28.8 ± 1.0 kg/m2, n.s.) and left ventricular ejection fraction (20.7 ± 0.5 vs. 23.2 ± 1.7%, n.s.), who were scheduled for heart transplantation or mechanical support implantation, were included in the study. A complex metabolomic and gene expression analysis of EAT obtained during surgery was performed. RESULTS: SGLT-2i ameliorated inflammation, as evidenced by the improved gene expression profile of pro-inflammatory genes in adipose tissue and decreased infiltration of immune cells into EAT. Enrichment of ether lipids with oleic acid noted on metabolomic analysis suggests a reduced disposition to ferroptosis, potentially further contributing to decreased oxidative stress in EAT of SGLT-2i treated subjects. CONCLUSIONS: Our results show decreased inflammation in EAT of patients with severe heart failure treated by SGLT-2i, as compared to patients with heart failure without this therapy. Modulation of EAT inflammatory and metabolic status could represent a novel mechanism behind SGLT-2i-associated cardioprotective effects in patients with heart failure.
Asunto(s)
Tejido Adiposo , Insuficiencia Cardíaca , Mediadores de Inflamación , Pericardio , Índice de Severidad de la Enfermedad , Inhibidores del Cotransportador de Sodio-Glucosa 2 , Humanos , Inhibidores del Cotransportador de Sodio-Glucosa 2/uso terapéutico , Inhibidores del Cotransportador de Sodio-Glucosa 2/farmacología , Inhibidores del Cotransportador de Sodio-Glucosa 2/efectos adversos , Insuficiencia Cardíaca/metabolismo , Insuficiencia Cardíaca/fisiopatología , Insuficiencia Cardíaca/tratamiento farmacológico , Persona de Mediana Edad , Masculino , Femenino , Pericardio/metabolismo , Pericardio/efectos de los fármacos , Tejido Adiposo/efectos de los fármacos , Tejido Adiposo/metabolismo , Resultado del Tratamiento , Mediadores de Inflamación/metabolismo , Volumen Sistólico/efectos de los fármacos , Antiinflamatorios/uso terapéutico , Antiinflamatorios/farmacología , Función Ventricular Izquierda/efectos de los fármacos , Diabetes Mellitus Tipo 2/tratamiento farmacológico , Diabetes Mellitus Tipo 2/metabolismo , Diabetes Mellitus Tipo 2/diagnóstico , Metabolómica , Biomarcadores/sangre , Tejido Adiposo EpicárdicoRESUMEN
(1) Background: empagliflozin, sodium-glucose co-transporter 2 (SGLT-2) inhibitor, is an effective antidiabetic agent with strong cardio- and nephroprotective properties. The mechanisms behind its cardio- and nephroprotection are still not fully clarified. (2) Methods: we used male hereditary hypertriglyceridemic (hHTG) rats, a non-obese model of dyslipidaemia, insulin resistance, and endothelial dysfunction fed standard diet with or without empagliflozin for six weeks to explore the molecular mechanisms of empagliflozin effects. Nuclear magnetic resonance (NMR)-based metabolomics; quantitative PCR of relevant genes involved in lipid and glucose metabolism, or senescence; glucose and palmitic acid oxidation in isolated tissues and cell lines of adipocytes and hepatocytes were used. (3) Results: empagliflozin inhibited weight gain and decreased adipose tissue weight, fasting blood glucose, and triglycerides and increased HDL-cholesterol. It also improved insulin sensitivity in white fat. NMR spectroscopy identified higher plasma concentrations of ketone bodies, ketogenic amino acid leucine and decreased levels of pyruvate and alanine. In the liver, adipose tissue and kidney, empagliflozin up-regulated expression of genes involved in gluconeogenesis and down-regulated expression of genes involved in lipogenesis along with reduction of markers of inflammation, oxidative stress and cell senescence. (4) Conclusion: multiple positive effects of empagliflozin, including reduced cell senescence and oxidative stress, could contribute to its long-term cardio- and nephroprotective actions.
Asunto(s)
Tejido Adiposo/metabolismo , Compuestos de Bencidrilo/administración & dosificación , Senescencia Celular/efectos de los fármacos , Gluconeogénesis/efectos de los fármacos , Glucósidos/administración & dosificación , Hipertrigliceridemia/tratamiento farmacológico , Hipertrigliceridemia/metabolismo , Hipoglucemiantes/administración & dosificación , Riñón/metabolismo , Lipogénesis/efectos de los fármacos , Hígado/metabolismo , Estrés Oxidativo/efectos de los fármacos , Inhibidores del Cotransportador de Sodio-Glucosa 2/administración & dosificación , Células 3T3-L1 , Administración Oral , Animales , Supervivencia Celular/efectos de los fármacos , Modelos Animales de Enfermedad , Regulación hacia Abajo/efectos de los fármacos , Dislipidemias/tratamiento farmacológico , Gluconeogénesis/genética , Células Hep G2 , Humanos , Resistencia a la Insulina , Lipogénesis/genética , Masculino , Ratones , Ratas , Resultado del Tratamiento , Regulación hacia Arriba/efectos de los fármacos , Aumento de Peso/efectos de los fármacosRESUMEN
Mitochondria and oxidative phosphorylation (OXPHOS) are emerging as intriguing targets for the efficient elimination of cancer cells. The specificity of this approach is aided by the capacity of non-proliferating non-cancerous cells to withstand oxidative insult induced by OXPHOS inhibition. Recently we discovered that mitochondrial targeting can also be employed to eliminate senescent cells, where it breaks the interplay between OXPHOS and ATP transporters that appear important for the maintenance of mitochondrial morphology and viability in the senescent setting. Hence, mitochondria/OXPHOS directed pharmacological interventions show promise in several clinically-relevant scenarios that call for selective removal of cancer and senescent cells.
Asunto(s)
Senescencia Celular , Mitocondrias/metabolismo , Neoplasias/patología , Adenosina Difosfato/metabolismo , Adenosina Trifosfato/metabolismo , Transporte Biológico , Muerte Celular , Proliferación Celular , Humanos , Neoplasias/metabolismo , Fosforilación Oxidativa , Especies Reactivas de Oxígeno/metabolismoRESUMEN
Tumor suppressor PML is induced under viral and genotoxic stresses by interferons and JAK-STAT signaling. However, the mechanism responsible for its cell type-specific regulation under non-stimulated conditions is poorly understood. To analyze the variation of PML expression, we utilized three human cell types, BJ fibroblasts and HeLa and U2OS cell lines, each with a distinct PML expression pattern. Analysis of JAK-STAT signaling in the three cell lines revealed differences in levels of activated STAT3 but not STAT1 correlating with PML mRNA and protein levels. RNAi-mediated knockdown of STAT3 decreased PML expression; both STAT3 level/activity and PML expression relied on IL6 secreted into culture media. We mapped the IL6-responsive sequence to an ISRE(-595/-628) element of the PML promoter. The PI3K/Akt/NFκB branch of IL6 signaling showed also cell-type dependence, being highest in BJ, intermediate in HeLa, and lowest in U2OS cells and correlated with IL6 secretion. RNAi-mediated knockdown of NEMO (NF-κ-B essential modulator), a key component of NFκB activation, suppressed NFκB targets LMP2 and IRF1 together with STAT3 and PML. Combined knockdown of STAT3 and NEMO did not further promote PML suppression, and it can be bypassed by exogenous IL6, indicating the NF-κB pathway acts upstream of JAK-STAT3 through induction of IL6. Our results indicate that the cell type-specific activity of IL6 signaling pathways governs PML expression under unperturbed growth conditions. As IL6 is induced in response to various viral and genotoxic stresses, this cytokine may regulate autocrine/paracrine induction of PML under these pathophysiological states as part of tissue adaptation to local stress.
Asunto(s)
Interleucina-6/metabolismo , Leucemia Promielocítica Aguda/metabolismo , Proteínas Nucleares/metabolismo , Transducción de Señal , Factores de Transcripción/metabolismo , Proteínas Supresoras de Tumor/metabolismo , Secuencia de Bases , Inmunoprecipitación de Cromatina , Cartilla de ADN , Técnica del Anticuerpo Fluorescente Indirecta , Células HeLa , Humanos , Quinasas Janus/metabolismo , Leucemia Promielocítica Aguda/patología , Proteína de la Leucemia Promielocítica , Reacción en Cadena en Tiempo Real de la Polimerasa , Factores de Transcripción STAT/metabolismoRESUMEN
Myokines represent important regulators of muscle metabolism. Our study aimed to explore the effects of a cyclical ketogenic reduction diet (CKD) vs. a nutritionally balanced reduction diet (RD) combined with regular resistance/aerobic training in healthy young males on serum concentrations of myokines and their potential role in changes in physical fitness. Twenty-five subjects undergoing regular resistance/aerobic training were randomized to the CKD (n = 13) or RD (n = 12) groups. Anthropometric and spiroergometric parameters, muscle strength, biochemical parameters, and serum concentrations of myokines and cytokines were assessed at baseline and after 8 weeks of intervention. Both diets reduced body weight, body fat, and BMI. Muscle strength and endurance performance were improved only by RD. Increased musclin (32.9 pg/mL vs. 74.5 pg/mL, p = 0.028) and decreased osteonectin levels (562 pg/mL vs. 511 pg/mL, p = 0.023) were observed in RD but not in the CKD group. In contrast, decreased levels of FGF21 (181 pg/mL vs. 86.4 pg/mL, p = 0.003) were found in the CKD group only. Other tested myokines and cytokines were not significantly affected by the intervention. Our data suggest that changes in systemic osteonectin and musclin levels could contribute to improved muscle strength and endurance performance and partially explain the differential effects of CKD and RD on physical fitness.
Asunto(s)
Dieta Cetogénica , Insuficiencia Renal Crónica , Entrenamiento de Fuerza , Masculino , Humanos , Osteonectina , Fuerza Muscular/fisiología , Dieta Reductora , Citocinas , Composición Corporal/fisiologíaRESUMEN
Background: Mitochondria present an emerging target for cancer treatment. We have investigated the effect of mitochondrially targeted tamoxifen (MitoTam), a first-in-class anti-cancer agent, in patients with solid metastatic tumours. Methods: MitoTam was tested in an open-label, single-centre (Department of Oncology, General Faculty Hospital, Charles University, Czech Republic), phase I/Ib trial in metastatic patients with various malignancies and terminated oncological therapies. In total, 75 patients were enrolled between May 23, 2018 and July 22, 2020. Phase I evaluated escalating doses of MitoTam in two therapeutic regimens using the 3 + 3 design to establish drug safety and maximum tolerated dose (MTD). In phase Ib, three dosing regimens were applied over 8 and 6 weeks to evaluate long-term toxicity of MitoTam as the primary objective and its anti-cancer effect as a secondary objective. This trial was registered with the European Medicines Agency under EudraCT 2017-004441-25. Findings: In total, 37 patients were enrolled into phase I and 38 into phase Ib. In phase I, the initial application of MitoTam via peripheral vein indicated high risk of thrombophlebitis, which was avoided by central vein administration. The highest dose with acceptable side effects was 5.0 mg/kg. The prevailing adverse effects (AEs) in phase I were neutropenia (30%), anaemia (30%) and fever/hyperthermia (30%), and in phase Ib fever/hyperthermia (58%) together with anaemia (26%) and neutropenia (16%). Serious AEs were mostly related to thromboembolic (TE) complications that affected 5% and 13% of patients in phase I and Ib, respectively. The only statistically significant AE related to MitoTam treatment was anaemia in phase Ib (p = 0.004). Of the tested regimens weekly dosing with 3.0 mg/kg for 6 weeks afforded the best safety profile with almost all being grade 1 (G1) AEs. Altogether, five fatalities occurred during the study, two of them meeting criteria for Suspected Unexpected Serious Adverse Events Reporting (SUSAR) (G4 thrombocytopenia and G5 stroke). MitoTam showed benefit evaluated as clinical benefit rate (CBR) in 37% patients with the largest effect in renal cell carcinoma (RCC) where four out of six patients reached disease stabilisation (SD), one reached partial response (PR) so that in total, five out of six (83%) patients showed CBR. Interpretation: In this study, the MTD was established as 5.0 mg/kg and the recommended dose of MitoTam as 3.0 mg/kg given once per week via central vein with recommended preventive anti-coagulation therapy. The prevailing toxicity included haematological AEs, hyperthermia/fever and TE complications. One fatal stroke and non-fatal G4 thrombocytopenia were recorded. MitoTam showed high efficacy against RCC. Funding: Smart Brain Ltd. Translation: For the Czech translation of the abstract see Supplementary Materials section.
RESUMEN
Mitochondrially targeted anticancer drugs (mitocans) that disrupt the energy-producing systems of cancer are emerging as new potential therapeutics. Mitochondrially targeted tamoxifen (MitoTam), an inhibitor of mitochondrial respiration respiratory complex I, is a first-in-class mitocan that was tested in the phase I/Ib MitoTam-01 trial of patients with metastatic cancer. MitoTam exhibited a manageable safety profile and efficacy; among 37% (14/38) of responders, the efficacy was greatest in patients with metastatic renal cell carcinoma (RCC) with a clinical benefit rate of 83% (5/6) of patients. This can be explained by the preferential accumulation of MitoTam in the kidney tissue in preclinical studies. Here we report the mechanism of action and safety profile of MitoTam in a case series of RCC patients. All six patients were males with a median age of 69 years, who had previously received at least three lines of palliative systemic therapy and suffered progressive disease before starting MitoTam. We recorded stable disease in four, partial response in one, and progressive disease (PD) in one patient. The histological subtype matched clear cell RCC (ccRCC) in the five responders and claro-cellular carcinoma with sarcomatoid features in the non-responder. The number of circulating tumor cells (CTCs) was evaluated longitudinally to monitor disease dynamics. Beside the decreased number of CTCs after MitoTam administration, we observed a significant decrease of the mitochondrial network mass in enriched CTCs. Two patients had long-term clinical responses to MitoTam, of 50 and 36 weeks. Both patients discontinued treatment due to adverse events, not PD. Two patients who completed the trial in November 2019 and May 2020 are still alive without subsequent anticancer therapy. The toxicity of MitoTam increased with the dosage but was manageable. The efficacy of MitoTam in pretreated ccRCC patients is linked to the novel mechanism of action of this first-in-class mitochondrially targeted drug.
RESUMEN
Repetitive sequences are among the most unstable regions in the eukaryotic genome and defects in their maintenance correlate with premature aging and cancer development. Promyelocytic leukemia protein (PML) induces accumulation of proteins at distinct nuclear sites, thereby affecting a plethora of processes including DNA repair or maintenance of telomeres. Doxorubicin, the broadly used chemotherapeutic compound, induces formation of PML-nucleolar associations (PNAs). Nevertheless, molecular factors affecting formation of PNAs are still largely unknown. Here we show that PNAs can accumulate ribosomal DNA (rDNA) and, after restoration of RNA polymerase I activity, these structures transfer a fraction of rDNA outside the nucleolus. Mutagenesis of PML isoforms revealed that this process depends on the SUMO-interacting motif and adjacent serine-rich region, and is enhanced by exon8b present exclusively in PML IV isoform. Moreover, we demonstrate that PNAs formation is also regulated by p14ARF/p53 tumor suppressors and casein kinase 2. Our data elucidate how PML nucleolar compartment is assembled, bring the first evidence of PML interacting with rDNA, and show the PML-dependent translocation of rDNA away from the nucleolus.
Asunto(s)
Proteínas Nucleares , Ácidos Nucleicos de Péptidos , ADN Ribosómico/genética , Doxorrubicina/farmacología , Proteínas Nucleares/metabolismo , Isoformas de Proteínas/metabolismo , Proteínas Supresoras de Tumor/genética , Proteínas Supresoras de Tumor/metabolismoRESUMEN
Type 2 diabetes mellitus represents a major health problem with increasing prevalence worldwide. Limited efficacy of current therapies has prompted a search for novel therapeutic options. Here we show that treatment of pre-diabetic mice with mitochondrially targeted tamoxifen, a potential anti-cancer agent with senolytic activity, improves glucose tolerance and reduces body weight with most pronounced reduction of visceral adipose tissue due to reduced food intake, suppressed adipogenesis and elimination of senescent cells. Glucose-lowering effect of mitochondrially targeted tamoxifen is linked to improvement of type 2 diabetes mellitus-related hormones profile and is accompanied by reduced lipid accumulation in liver. Lower senescent cell burden in various tissues, as well as its inhibitory effect on pre-adipocyte differentiation, results in lower level of circulating inflammatory mediators that typically enhance metabolic dysfunction. Targeting senescence with mitochodrially targeted tamoxifen thus represents an approach to the treatment of type 2 diabetes mellitus and its related comorbidities, promising a complex impact on senescence-related pathologies in aging population of patients with type 2 diabetes mellitus with potential translation into the clinic.
Asunto(s)
Diabetes Mellitus Experimental , Diabetes Mellitus Tipo 2 , Anciano , Animales , Diabetes Mellitus Experimental/complicaciones , Diabetes Mellitus Experimental/tratamiento farmacológico , Diabetes Mellitus Tipo 2/complicaciones , Diabetes Mellitus Tipo 2/tratamiento farmacológico , Glucosa/metabolismo , Humanos , Ratones , Obesidad/complicaciones , Obesidad/tratamiento farmacológico , Obesidad/metabolismo , Tamoxifeno/farmacología , Tamoxifeno/uso terapéuticoRESUMEN
BACKGROUND: Pheochromocytoma and paraganglioma (PPGL) are neuroendocrine tumors with frequent mutations in genes linked to the tricarboxylic acid cycle. However, no pathogenic variant has been found to date in succinyl-CoA ligase (SUCL), an enzyme that provides substrate for succinate dehydrogenase (SDH; mitochondrial complex II [CII]), a known tumor suppressor in PPGL. METHODS: A cohort of 352 patients with apparently sporadic PPGL underwent genetic testing using a panel of 54 genes developed at the National Institutes of Health, including the SUCLG2 subunit of SUCL. Gene deletion, succinate levels, and protein levels were assessed in tumors where possible. To confirm the possible mechanism, we used a progenitor cell line, hPheo1, derived from a human pheochromocytoma, and ablated and re-expressed SUCLG2. RESULTS: We describe 8 germline variants in the guanosine triphosphate-binding domain of SUCLG2 in 15 patients (15 of 352, 4.3%) with apparently sporadic PPGL. Analysis of SUCLG2-mutated tumors and SUCLG2-deficient hPheo1 cells revealed absence of SUCLG2 protein, decrease in the level of the SDHB subunit of SDH, and faulty assembly of the complex II, resulting in aberrant respiration and elevated succinate accumulation. CONCLUSIONS: Our study suggests SUCLG2 as a novel candidate gene in the genetic landscape of PPGL. Large-scale sequencing may uncover additional cases harboring SUCLG2 variants and provide more detailed information about their prevalence and penetrance.
Asunto(s)
Neoplasias de las Glándulas Suprarrenales , Paraganglioma , Feocromocitoma , Neoplasias de las Glándulas Suprarrenales/genética , Neoplasias de las Glándulas Suprarrenales/patología , Mutación de Línea Germinal , Humanos , Paraganglioma/genética , Paraganglioma/patología , Feocromocitoma/genética , Feocromocitoma/patología , Succinato Deshidrogenasa/genética , Succinato Deshidrogenasa/metabolismoRESUMEN
Mitochondrial oxidative phosphorylation (OXPHOS) generates ATP, but OXPHOS also supports biosynthesis during proliferation. In contrast, the role of OXPHOS during quiescence, beyond ATP production, is not well understood. Using mouse models of inducible OXPHOS deficiency in all cell types or specifically in the vascular endothelium that negligibly relies on OXPHOS-derived ATP, we show that selectively during quiescence OXPHOS provides oxidative stress resistance by supporting macroautophagy/autophagy. Mechanistically, OXPHOS constitutively generates low levels of endogenous ROS that induce autophagy via attenuation of ATG4B activity, which provides protection from ROS insult. Physiologically, the OXPHOS-autophagy system (i) protects healthy tissue from toxicity of ROS-based anticancer therapy, and (ii) provides ROS resistance in the endothelium, ameliorating systemic LPS-induced inflammation as well as inflammatory bowel disease. Hence, cells acquired mitochondria during evolution to profit from oxidative metabolism, but also built in an autophagy-based ROS-induced protective mechanism to guard against oxidative stress associated with OXPHOS function during quiescence.Abbreviations: AMPK: AMP-activated protein kinase; AOX: alternative oxidase; Baf A: bafilomycin A1; CI, respiratory complexes I; DCF-DA: 2',7'-dichlordihydrofluorescein diacetate; DHE: dihydroethidium; DSS: dextran sodium sulfate; ΔΨmi: mitochondrial inner membrane potential; EdU: 5-ethynyl-2'-deoxyuridine; ETC: electron transport chain; FA: formaldehyde; HUVEC; human umbilical cord endothelial cells; IBD: inflammatory bowel disease; LC3B: microtubule associated protein 1 light chain 3 beta; LPS: lipopolysaccharide; MEFs: mouse embryonic fibroblasts; MTORC1: mechanistic target of rapamycin kinase complex 1; mtDNA: mitochondrial DNA; NAC: N-acetyl cysteine; OXPHOS: oxidative phosphorylation; PCs: proliferating cells; PE: phosphatidylethanolamine; PEITC: phenethyl isothiocyanate; QCs: quiescent cells; ROS: reactive oxygen species; PLA2: phospholipase A2, WB: western blot.
Asunto(s)
Autofagia , Enfermedades Inflamatorias del Intestino , Proteínas Quinasas Activadas por AMP/metabolismo , Adenosina Trifosfato/metabolismo , Animales , Cisteína/metabolismo , ADN Mitocondrial/metabolismo , Dextranos/metabolismo , Células Endoteliales/metabolismo , Fibroblastos/metabolismo , Formaldehído/metabolismo , Humanos , Enfermedades Inflamatorias del Intestino/metabolismo , Isotiocianatos , Lipopolisacáridos/metabolismo , Diana Mecanicista del Complejo 1 de la Rapamicina/metabolismo , Ratones , Proteínas Asociadas a Microtúbulos/metabolismo , Mitocondrias/metabolismo , Fosfatidiletanolaminas/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Respiración , SirolimusRESUMEN
Dihydroorotate dehydrogenase (DHODH) is an enzyme of the de novo pyrimidine synthesis pathway that provides nucleotides for RNA/DNA synthesis essential for proliferation. In mammalian cells, DHODH is localized in mitochondria, linked to the respiratory chain via the coenzyme Q pool. Here we discuss the role of DHODH in the oxidative phosphorylation system and in the initiation and progression of cancer. We summarize recent findings on DHODH biology, the progress made in the development of new, specific inhibitors of DHODH intended for cancer therapy, and the mechanistic insights into the consequences of DHODH inhibition.
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Mitocondrias/genética , Neoplasias/genética , Fosforilación Oxidativa , Oxidorreductasas actuantes sobre Donantes de Grupo CH-CH/genética , Proliferación Celular/efectos de los fármacos , Dihidroorotato Deshidrogenasa , Transporte de Electrón/genética , Inhibidores Enzimáticos/uso terapéutico , Humanos , Mitocondrias/metabolismo , Neoplasias/patología , Oxidorreductasas actuantes sobre Donantes de Grupo CH-CH/antagonistas & inhibidores , Ubiquinona/análogos & derivados , Ubiquinona/genéticaRESUMEN
Myelodysplastic syndromes (MDS) are preleukemic disorders characterized by clonal growth of mutant hematopoietic stem and progenitor cells. MDS are associated with proinflammatory signaling, dysregulated immune response, and cell death in the bone marrow (BM). Aging, autoinflammation and autoimmunity are crucial features of disease progression, concordant with promoting growth of malignant clones and accumulation of mutations. Suprabasin (SBSN), a recently proposed proto-oncogene of unknown function, physiologically expressed in stratified epithelia, is associated with poor prognosis of several human malignancies. Here, we showed that SBSN is expressed in the BM by myeloid cell subpopulations, including myeloid-derived suppressor cells, and is secreted into BM plasma and peripheral blood of MDS patients. The highest expression of SBSN was present in a patient group with poor prognosis. SBSN levels in the BM correlated positively with blast percentage and negatively with CCL2 chemokine levels and lymphocyte count. In vitro treatment of leukemic cells with interferon-gamma and demethylating agent 5-azacytidine (5-AC) induced SBSN expression. This indicated that aberrant cytokine levels in the BM and epigenetic landscape modifications in MDS patients may underlie ectopic expression of SBSN. Our findings suggest SBSN as a candidate biomarker of high-risk MDS with a possible role in disease progression and therapy resistance.
Asunto(s)
Antígenos de Diferenciación/metabolismo , Médula Ósea/metabolismo , Síndromes Mielodisplásicos/metabolismo , Proteínas de Neoplasias/metabolismo , Antígenos de Diferenciación/sangre , Antígenos de Diferenciación/genética , Azacitidina/farmacología , Biomarcadores/sangre , Biomarcadores/metabolismo , Compartimento Celular/efectos de los fármacos , Línea Celular Tumoral , Quimiocina CCL2/metabolismo , Regulación Neoplásica de la Expresión Génica , Células HEK293 , Humanos , Interferón gamma/farmacología , Leucocitos Mononucleares/metabolismo , Recuento de Linfocitos , Síndromes Mielodisplásicos/sangre , Células Mieloides/efectos de los fármacos , Células Mieloides/metabolismo , Proteínas de Neoplasias/sangre , Proteínas de Neoplasias/genética , Pronóstico , Proto-Oncogenes Mas , ARN Mensajero/genética , ARN Mensajero/metabolismoRESUMEN
p53-mutated tumors often exhibit increased resistance to standard chemotherapy and enhanced metastatic potential. Here we demonstrate that inhibition of dihydroorotate dehydrogenase (DHODH), a key enzyme of the de novo pyrimidine synthesis pathway, effectively decreases proliferation of cancer cells via induction of replication and ribosomal stress in a p53- and checkpoint kinase 1 (Chk1)-dependent manner. Mechanistically, a block in replication and ribosomal biogenesis result in p53 activation paralleled by accumulation of replication forks that activate the ataxia telangiectasia and Rad3-related kinase/Chk1 pathway, both of which lead to cell cycle arrest. Since in the absence of functional p53 the cell cycle arrest fully depends on Chk1, combined DHODH/Chk1 inhibition in p53-dysfunctional cancer cells induces aberrant cell cycle re-entry and erroneous mitosis, resulting in massive cell death. Combined DHODH/Chk1 inhibition effectively suppresses p53-mutated tumors and their metastasis, and therefore presents a promising therapeutic strategy for p53-mutated cancers.
Asunto(s)
Neoplasias de la Mama/metabolismo , Puntos de Control del Ciclo Celular , Proliferación Celular , Pirimidinas/biosíntesis , Ribosomas/metabolismo , Proteína p53 Supresora de Tumor/deficiencia , Animales , Protocolos de Quimioterapia Combinada Antineoplásica/farmacología , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Puntos de Control del Ciclo Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Quinasa 1 Reguladora del Ciclo Celular (Checkpoint 1)/antagonistas & inhibidores , Quinasa 1 Reguladora del Ciclo Celular (Checkpoint 1)/genética , Quinasa 1 Reguladora del Ciclo Celular (Checkpoint 1)/metabolismo , Dihidroorotato Deshidrogenasa , Femenino , Regulación Neoplásica de la Expresión Génica , Genes erbB-2 , Células HCT116 , Humanos , Leflunamida/farmacología , Células MCF-7 , Ratones Endogámicos BALB C , Ratones Endogámicos NOD , Ratones SCID , Ratones Transgénicos , Oxidorreductasas actuantes sobre Donantes de Grupo CH-CH/antagonistas & inhibidores , Oxidorreductasas actuantes sobre Donantes de Grupo CH-CH/genética , Oxidorreductasas actuantes sobre Donantes de Grupo CH-CH/metabolismo , Compuestos de Fenilurea/farmacología , Inhibidores de Proteínas Quinasas/farmacología , Pirazinas/farmacología , Ribosomas/genética , Transducción de Señal , Proteína p53 Supresora de Tumor/genéticaRESUMEN
Diverse stress insults trigger interactions of PML with nucleolus, however, the function of these PML nucleolar associations (PNAs) remains unclear. Here we show that during induction of DNA damage-induced senescence in human non-cancerous cells, PML accumulates at the nucleolar periphery simultaneously with inactivation of RNA polymerase I (RNAP I) and nucleolar segregation. Using time-lapse and high-resolution microscopy, we followed the genesis, structural transitions and destiny of PNAs to show that: 1) the dynamic structural changes of the PML-nucleolar interaction are tightly associated with inactivation and reactivation of RNAP I-mediated transcription, respectively; 2) the PML-nucleolar compartment develops sequentially under stress and, upon stress termination, it culminates in either of two fates: disappearance or persistence; 3) all PNAs stages can associate with DNA damage markers; 4) the persistent, commonly long-lasting PML multi-protein nucleolar structures (PML-NDS) associate with markers of DNA damage, indicating a role of PNAs in persistent DNA damage response characteristic for senescent cells. Given the emerging evidence implicating PML in homologous recombination-directed DNA repair, we propose that PNAs contribute to sequestration and faithful repair of the highly unstable ribosomal DNA repeats, a fundamental process to maintain a precise balance between DNA repair mechanisms, with implications for genomic integrity and aging.
Asunto(s)
Nucléolo Celular/metabolismo , Senescencia Celular , Daño del ADN , Proteína de la Leucemia Promielocítica/metabolismo , Células Cultivadas , Doxorrubicina , Humanos , Imagenología Tridimensional , Estrés FisiológicoRESUMEN
Cellular senescence is a form of cell cycle arrest that limits the proliferative potential of cells, including tumour cells. However, inability of immune cells to subsequently eliminate senescent cells from the organism may lead to tissue damage, inflammation, enhanced carcinogenesis and development of age-related diseases. We found that the anticancer agent mitochondria-targeted tamoxifen (MitoTam), unlike conventional anticancer agents, kills cancer cells without inducing senescence in vitro and in vivo. Surprisingly, it also selectively eliminates both malignant and non-cancerous senescent cells. In naturally aged mice treated with MitoTam for 4 weeks, we observed a significant decrease of senescence markers in all tested organs compared to non-treated animals. Mechanistically, we found that the susceptibility of senescent cells to MitoTam is linked to a very low expression level of adenine nucleotide translocase-2 (ANT2), inherent to the senescent phenotype. Restoration of ANT2 in senescent cells resulted in resistance to MitoTam, while its downregulation in non-senescent cells promoted their MitoTam-triggered elimination. Our study documents a novel, translationally intriguing role for an anticancer agent targeting mitochondria, that may result in a new strategy for the treatment of age-related diseases and senescence-associated pathologies.
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Translocador 2 del Nucleótido Adenina/metabolismo , Antineoplásicos Hormonales/farmacología , Senescencia Celular/efectos de los fármacos , Mitocondrias/efectos de los fármacos , Tamoxifeno/farmacología , Translocador 2 del Nucleótido Adenina/genética , Animales , Apoptosis/efectos de los fármacos , Apoptosis/genética , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/genética , Técnicas de Silenciamiento del Gen , Humanos , Células MCF-7 , Ratones , Ratones Endogámicos NOD , Ratones SCID , Ratones Transgénicos , Mitocondrias/metabolismo , Transfección , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Radiation and chemotherapy represent standard-of-care cancer treatments. However, most patients eventually experience tumour recurrence, treatment failure and metastatic dissemination with fatal consequences. To elucidate the molecular mechanisms of resistance to radio- and chemotherapy, we exposed human cancer cell lines (HeLa, MCF-7 and DU145) to clinically relevant doses of 5-azacytidine or ionizing radiation and compared the transcript profiles of all surviving cell subpopulations, including low-adherent stem-like cells. Stress-mobilized low-adherent cell fractions differed from other survivors in terms of deregulation of hundreds of genes, including those involved in interferon response. Exposure of cancer cells to interferon-gamma but not interferon-beta resulted in the development of a heterogeneous, low-adherent fraction comprising not only apoptotic/necrotic cells but also live cells exhibiting active Notch signalling and expressing stem-cell markers. Chemical inhibition of mitogen-activated protein kinase/ERK kinase (MEK) or siRNA-mediated knockdown of extracellular signal-regulated kinase 1/2 (Erk1/2) and interferon responsible factor 1 (IRF1) prevented mobilization of the surviving low-adherent population, indicating that interferon-gamma-mediated loss of adhesion and anoikis resistance required an active Erk pathway interlinked with interferon signalling by transcription factor IRF1. Notably, a skin-specific protein suprabasin (SBSN), a recently identified oncoprotein, was among the top scoring genes upregulated in surviving low-adherent cancer cells induced by 5-azacytidine or irradiation. SBSN expression required the activity of the MEK/Erk pathway, and siRNA-mediated knockdown of SBSN suppressed the low-adherent fraction in irradiated, interferon-gamma- and 5-azacytidine-treated cells, respectively, implicating SBSN in genotoxic stress-induced phenotypic plasticity and stress resistance. Importantly, SBSN expression was observed in human clinical specimens of colon and ovarian carcinomas, as well as in circulating tumour cells and metastases of the 4T1 mouse model. The association of SBSN expression with progressive stages of cancer development indicates its role in cancer evolution and therapy resistance.
Asunto(s)
Antígenos de Diferenciación/genética , Antineoplásicos/farmacología , Azacitidina/farmacología , Interferones/farmacología , Proteínas de Neoplasias/genética , Neoplasias/tratamiento farmacológico , Animales , Anoicis/efectos de los fármacos , Anoicis/efectos de la radiación , Línea Celular Tumoral , Resistencia a Antineoplásicos , Femenino , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica/efectos de la radiación , Humanos , Ratones , Ratones Endogámicos BALB C , Neoplasias/genética , Neoplasias/radioterapia , Células Madre Neoplásicas/efectos de los fármacos , Células Madre Neoplásicas/metabolismo , Células Madre Neoplásicas/efectos de la radiación , Regulación hacia Arriba/efectos de los fármacos , Regulación hacia Arriba/efectos de la radiaciónRESUMEN
Cancer cells without mitochondrial DNA (mtDNA) do not form tumors unless they reconstitute oxidative phosphorylation (OXPHOS) by mitochondria acquired from host stroma. To understand why functional respiration is crucial for tumorigenesis, we used time-resolved analysis of tumor formation by mtDNA-depleted cells and genetic manipulations of OXPHOS. We show that pyrimidine biosynthesis dependent on respiration-linked dihydroorotate dehydrogenase (DHODH) is required to overcome cell-cycle arrest, while mitochondrial ATP generation is dispensable for tumorigenesis. Latent DHODH in mtDNA-deficient cells is fully activated with restoration of complex III/IV activity and coenzyme Q redox-cycling after mitochondrial transfer, or by introduction of an alternative oxidase. Further, deletion of DHODH interferes with tumor formation in cells with fully functional OXPHOS, while disruption of mitochondrial ATP synthase has little effect. Our results show that DHODH-driven pyrimidine biosynthesis is an essential pathway linking respiration to tumorigenesis, pointing to inhibitors of DHODH as potential anti-cancer agents.
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ADN Mitocondrial/metabolismo , Mitocondrias/metabolismo , Neoplasias/metabolismo , Oxidorreductasas actuantes sobre Donantes de Grupo CH-CH/fisiología , Pirimidinas/metabolismo , Animales , Línea Celular Tumoral , Respiración de la Célula , Dihidroorotato Deshidrogenasa , Humanos , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Fosforilación Oxidativa , Ubiquinona/metabolismoRESUMEN
As a part of cellular pathogen defense, IFNγ triggers induction of NADPH oxidase NOX2, which produces superoxide into phagosomes of immune cells. Recent data show that a similar mechanism can also operate in IFNγ-mediated anticancer control. IFNγ is capable of inducing expression of constitutively active NADPH oxidase NOX4 in tumor cells leading to generation of reactive oxygen species (ROS) damaging DNA, activation of DNA damage response and cell cycle arrest/premature cellular senescence.