RESUMO
Availability of the essential amino acid methionine affects cellular metabolism and growth, and dietary methionine restriction has been implicated as a cancer therapeutic strategy. Nevertheless, how liver cancer cells respond to methionine deprivation and underlying mechanisms remain unclear. Here we find that human liver cancer cells undergo irreversible cell cycle arrest upon methionine deprivation in vitro. Blocking methionine adenosyl transferase 2A (MAT2A)-dependent methionine catabolism induces cell cycle arrest and DNA damage in liver cancer cells, resulting in cellular senescence. A pharmacological screen further identified GSK3 inhibitors as senolytics that selectively kill MAT2A-inhibited senescent liver cancer cells. Importantly, combined treatment with MAT2A and GSK3 inhibitors therapeutically blunts liver tumor growth in vitro and in vivo across multiple models. Together, methionine catabolism is essential for liver tumor growth, and its inhibition can be exploited as an improved pro-senescence strategy for combination with senolytic agents to treat liver cancer.
Assuntos
Quinase 3 da Glicogênio Sintase , Neoplasias Hepáticas , Humanos , S-Adenosilmetionina/metabolismo , S-Adenosilmetionina/farmacologia , Neoplasias Hepáticas/tratamento farmacológico , Neoplasias Hepáticas/genética , Metionina/farmacologia , Metionina Adenosiltransferase/metabolismoAssuntos
Asfixia Neonatal , Hipotermia Induzida , Hipóxia-Isquemia Encefálica , Humanos , Recém-Nascido , Criança , Prognóstico , Hipóxia-Isquemia Encefálica/terapia , Modelos Estatísticos , Asfixia Neonatal/terapia , Deficiências do Desenvolvimento/diagnóstico , Deficiências do Desenvolvimento/etiologiaRESUMO
Hepatocellular carcinoma (HCC) is a typically fatal malignancy exhibiting genetic heterogeneity and limited therapy responses. We demonstrate here that HCCs consistently repress urea cycle gene expression and thereby become auxotrophic for exogenous arginine. Surprisingly, arginine import is uniquely dependent on the cationic amino acid transporter SLC7A1, whose inhibition slows HCC cell growth in vitro and in vivo. Moreover, arginine deprivation engages an integrated stress response that promotes HCC cell-cycle arrest and quiescence, dependent on the general control nonderepressible 2 (GCN2) kinase. Inhibiting GCN2 in arginine-deprived HCC cells promotes a senescent phenotype instead, rendering these cells vulnerable to senolytic compounds. Preclinical models confirm that combined dietary arginine deprivation, GCN2 inhibition, and senotherapy promote HCC cell apoptosis and tumor regression. These data suggest novel strategies to treat human liver cancers through targeting SLC7A1 and/or a combination of arginine restriction, inhibition of GCN2, and senolytic agents.
Assuntos
Carcinoma Hepatocelular , Neoplasias Hepáticas , Arginina/metabolismo , Arginina/farmacologia , Carcinoma Hepatocelular/metabolismo , Humanos , Neoplasias Hepáticas/metabolismo , Proteínas Serina-Treonina Quinases , SenoterapiaRESUMO
Despite the development of metabolism-based therapy for a variety of malignancies, resistance to single-agent treatment is common due to the metabolic plasticity of cancer cells. Improved understanding of how malignant cells rewire metabolic pathways can guide the rational selection of combination therapy to circumvent drug resistance. Here, we show that human T-ALL cells shift their metabolism from oxidative decarboxylation to reductive carboxylation when the TCA cycle is disrupted. The α-ketoglutarate dehydrogenase complex (KGDHC) in the TCA cycle regulates oxidative decarboxylation by converting α-ketoglutarate (α-KG) to succinyl-CoA, while isocitrate dehydrogenase (IDH) 1 and 2 govern reductive carboxylation. Metabolomics flux analysis of T-ALL reveals enhanced reductive carboxylation upon genetic depletion of the E2 subunit of KGDHC, dihydrolipoamide-succinyl transferase (DLST), mimicking pharmacological inhibition of the complex. Mechanistically, KGDHC dysfunction causes increased demethylation of nuclear DNA by α-KG-dependent dioxygenases (e.g., TET demethylases), leading to increased production of both IDH1 and 2. Consequently, dual pharmacologic inhibition of the TCA cycle and TET demethylases demonstrates additive efficacy in reducing the tumor burden in zebrafish xenografts. These findings provide mechanistic insights into how T-ALL develops resistance to drugs targeting the TCA cycle and therapeutic strategies to overcome this resistance.
RESUMO
High-risk neuroblastoma remains therapeutically challenging to treat, and the mechanisms promoting disease aggression are poorly understood. Here, we show that elevated expression of dihydrolipoamide S-succinyltransferase (DLST) predicts poor treatment outcome and aggressive disease in patients with neuroblastoma. DLST is an E2 component of the α-ketoglutarate (αKG) dehydrogenase complex, which governs the entry of glutamine into the tricarboxylic acid cycle (TCA) for oxidative decarboxylation. During this irreversible step, αKG is converted into succinyl-CoA, producing NADH for oxidative phosphorylation (OXPHOS). Utilizing a zebrafish model of MYCN-driven neuroblastoma, we demonstrate that even modest increases in DLST expression promote tumor aggression, while monoallelic dlst loss impedes disease initiation and progression. DLST depletion in human MYCN-amplified neuroblastoma cells minimally affected glutamine anaplerosis and did not alter TCA cycle metabolites other than αKG. However, DLST loss significantly suppressed NADH production and impaired OXPHOS, leading to growth arrest and apoptosis of neuroblastoma cells. In addition, multiple inhibitors targeting the electron transport chain, including the potent IACS-010759 that is currently in clinical testing for other cancers, efficiently reduced neuroblastoma proliferation in vitro. IACS-010759 also suppressed tumor growth in zebrafish and mouse xenograft models of high-risk neuroblastoma. Together, these results demonstrate that DLST promotes neuroblastoma aggression and unveils OXPHOS as an essential contributor to high-risk neuroblastoma. SIGNIFICANCE: These findings demonstrate a novel role for DLST in neuroblastoma aggression and identify the OXPHOS inhibitor IACS-010759 as a potential therapeutic strategy for this deadly disease.
Assuntos
Aciltransferases/metabolismo , Neoplasias Encefálicas/metabolismo , Neuroblastoma/metabolismo , Fosforilação Oxidativa , Animais , Apoptose , Linhagem Celular Tumoral , Colágeno/química , Modelos Animais de Doenças , Combinação de Medicamentos , Feminino , Perfilação da Expressão Gênica , Células HEK293 , Humanos , Concentração Inibidora 50 , Complexo Cetoglutarato Desidrogenase/metabolismo , Laminina/química , Camundongos , Camundongos Endogâmicos BALB C , Invasividade Neoplásica , Transplante de Neoplasias , Oxigênio/metabolismo , Proteoglicanas/química , Interferência de RNA , Risco , Smegmamorpha , Resultado do Tratamento , Ácidos Tricarboxílicos/metabolismo , Peixe-ZebraRESUMO
BACKGROUND: Pediatric patients undergoing heart transplant have a number of factors predisposing them to become fluid-overloaded, including capillary leak syndrome. Capillary leak and FO are associated with organ injury and may influence both short- and long-term outcomes. This study aimed to 1) determine the extent, timing, and predictors of post-operative FO and 2) investigate the association of FO with clinically important outcomes. METHODS: Between 2000 and 2012, 70 children less than 6 years old had a heart transplant at our institution. This was a secondary analysis of data from an ongoing prospective cohort study. RESULTS: FO, defined as cumulative fluid balance greater than 10% of body weight in the first 5 post-operative days, occurred in 16/70 patients (23%); 7 of these had more than 20% FO. Shorter donor ischemic time and longer cardiopulmonary bypass time were independently associated with increased risk of FO. FO >20% was a statistically significant independent predictor of mortality (P = .005), ventilation time, and PICU length of stay. There was no statistically significant association between identified neurodevelopment domains and FO. CONCLUSIONS: Our single-center experience demonstrates that FO was common after pediatric heart transplant and was associated with worse clinical outcomes. FO is a potentially modifiable factor, and research is needed to better determine risk factors and whether intervention to reduce FO can improve outcomes in pediatric heart transplant patients.
Assuntos
Transplante de Coração , Hospitalização/estatística & dados numéricos , Complicações Pós-Operatórias/epidemiologia , Desequilíbrio Hidroeletrolítico/epidemiologia , Criança , Pré-Escolar , Feminino , Humanos , Lactente , Recém-Nascido , Masculino , Prognóstico , Estudos Prospectivos , Fatores de Tempo , Resultado do TratamentoRESUMO
A tumor is not simply a group of cancer cells, but rather a heterogeneous collection of infiltrating and resident host cells, secreted factors and extracellular matrix. Tumor cells stimulate significant molecular, cellular and physical changes within their host tissues to support tumor growth and progression. An emerging tumor microenvironment is a complex and continuously evolving entity. The composition of the tumor microenvironment varies between tumor types, but hallmark features include immune cells, stromal cells, blood vessels, and extracellular matrix. It is believed that the "tumor microenvironment is not just a silent bystander, but rather an active promoter of cancer progression" (Truffi et al., 2020). Early in tumor growth, a dynamic and reciprocal relationship develops between cancer cells and components of the tumor microenvironment that supports cancer cell survival, local invasion and metastatic dissemination. To overcome a hypoxic and acidic microenvironment, the tumor microenvironment coordinates a program that promotes angiogenesis to restore oxygen and nutrient supply and remove metabolic waste. Tumors become infiltrated with diverse adaptive and innate immune cells that can perform both pro- and anti- tumorigenic functions (Figure 1). An expanding literature on the tumor microenvironment has identified new targets within it for therapeutic intervention.
Assuntos
Carcinogênese/patologia , Matriz Extracelular/metabolismo , Neoplasias/patologia , Neovascularização Patológica , Microambiente Tumoral/imunologia , Carcinogênese/imunologia , Humanos , Neoplasias/irrigação sanguínea , Neoplasias/imunologiaRESUMO
Two studies in this issue position the stabilization of the transcription factor BACH1 as a critical node in the metastasis of lung cancer and propose two new therapeutic approaches for this leading cause of cancer-related deaths (Lignitto et al., 2019; Wiel et al., 2019).
Assuntos
Neoplasias Pulmonares , Fator 2 Relacionado a NF-E2 , Fatores de Transcrição de Zíper de Leucina Básica , Regulação Neoplásica da Expressão Gênica , HumanosRESUMO
The tricarboxylic acid (TCA) cycle is a central route for oxidative phosphorylation in cells, and fulfills their bioenergetic, biosynthetic, and redox balance requirements. Despite early dogma that cancer cells bypass the TCA cycle and primarily utilize aerobic glycolysis, emerging evidence demonstrates that certain cancer cells, especially those with deregulated oncogene and tumor suppressor expression, rely heavily on the TCA cycle for energy production and macromolecule synthesis. As the field progresses, the importance of aberrant TCA cycle function in tumorigenesis and the potentials of applying small molecule inhibitors to perturb the enhanced cycle function for cancer treatment start to evolve. In this review, we summarize current knowledge about the fuels feeding the cycle, effects of oncogenes and tumor suppressors on fuel and cycle usage, common genetic alterations and deregulation of cycle enzymes, and potential therapeutic opportunities for targeting the TCA cycle in cancer cells. With the application of advanced technology and in vivo model organism studies, it is our hope that studies of this previously overlooked biochemical hub will provide fresh insights into cancer metabolism and tumorigenesis, subsequently revealing vulnerabilities for therapeutic interventions in various cancer types.
Assuntos
Ciclo do Ácido Cítrico/efeitos dos fármacos , Terapia de Alvo Molecular/métodos , Neoplasias/tratamento farmacológico , Neoplasias/metabolismo , Animais , Humanos , Neoplasias/genética , Neoplasias/patologia , Oncogenes/genética , Proteínas Supressoras de Tumor/metabolismoAssuntos
Azepinas/farmacologia , Caseína Quinase II/antagonistas & inibidores , Naftiridinas/farmacologia , Proteínas de Neoplasias/antagonistas & inibidores , Proteínas de Neoplasias/metabolismo , Leucemia-Linfoma Linfoblástico de Células T Precursoras/metabolismo , Receptor Notch1/imunologia , Triazóis/farmacologia , Azepinas/agonistas , Caseína Quinase II/genética , Caseína Quinase II/metabolismo , Linhagem Celular Tumoral , Sinergismo Farmacológico , Feminino , Humanos , Masculino , Naftiridinas/agonistas , Proteínas de Neoplasias/genética , Fenazinas , Leucemia-Linfoma Linfoblástico de Células T Precursoras/tratamento farmacológico , Leucemia-Linfoma Linfoblástico de Células T Precursoras/genética , Leucemia-Linfoma Linfoblástico de Células T Precursoras/patologia , Estabilidade Proteica , Receptor Notch1/genética , Triazóis/agonistasRESUMO
The zebrafish represents a revolutionary tool in large-scale genetic and small-molecule screens for gene and drug discovery. Transgenic zebrafish are often utilized in these screens. Many transgenic fish lines are maintained in the heterozygous state due to the lethality associated with homozygosity; thus, their progeny must be sorted to ensure a population expressing the transgene of interest for use in screens. Sorting transgenic embryos under a fluorescence microscope is very labor-intensive and demands fine-tuned motor skills. Here we report an efficient transgenic method of utilizing pigmentation rescue of nacre mutant fish for accurate naked-eye identification of both mosaic founders and stable transgenic zebrafish. This was accomplished by co-injecting two constructs with the I-SceI meganuclease enzyme into pigmentless nacre embryos: I-SceI-mitfa:mitfa-I-SceI to rescue the pigmentation and I-SceI-zpromoter:gene-of-interest-I-SceI to express the gene of interest under a zebrafish promoter (zpromoter). Pigmentation rescue reliably predicted transgene integration. Compared with other transgenic techniques, our approach significantly increases the overall percentage of founders and facilitates accurate naked-eye identification of stable transgenic fish, greatly reducing laborious fluorescence microscope sorting and PCR genotyping. Thus, this approach is ideal for generating transgenic fish for large-scale screens.
Assuntos
Técnicas de Transferência de Genes , Fator de Transcrição Associado à Microftalmia/genética , Pigmentação/genética , Regiões Promotoras Genéticas , Proteínas de Peixe-Zebra/genética , Animais , Animais Geneticamente Modificados , Genótipo , Proteínas de Fluorescência Verde/genética , Microscopia de Fluorescência , Peixe-Zebra/genética , Peixe-Zebra/fisiologiaRESUMO
Utilizing ENU mutagenesis, we identified a mutant mouse with elevated platelets. Genetic mapping localized the mutation to an interval on chromosome 19 that encodes the Jak2 tyrosine kinase. We identified a A3056T mutation resulting in a premature stop codon within exon 19 of Jak2 (Jak2(K915X)), resulting in a protein truncation and functionally inactive enzyme. This novel platelet phenotype was also observed in mice bearing a hemizygous targeted disruption of the Jak2 locus (Jak2(+/-)). Timed pregnancy experiments revealed that Jak2(K915X/K915X) and Jak2(-/-) displayed embryonic lethality; however, Jak2(K915X/K915X) embryos were viable an additional two days compared to Jak2(-/-) embryos. Our data suggest that perturbing JAK2 activation may have unexpected consequences in elevation of platelet number and correspondingly, important implications for treatment of hematological disorders with constitutive Jak2 activity.
Assuntos
Plaquetas/citologia , Janus Quinase 2/genética , Fenótipo , Animais , Western Blotting , Mapeamento Cromossômico , Etilnitrosoureia , Fluoruracila , Genótipo , Células HEK293 , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Mutantes , Mutagênese/genética , Fenil-Hidrazinas , Mutação Puntual/genética , Análise de Sequência de DNARESUMO
The ability of random mutagenesis techniques to annotate the mammalian genome can be hampered due to genetic redundancy and compensatory pathways that mask heterozygous mutations under homeostatic conditions. The objective of this study was to devise a pharmacologically sensitized screen using the chemotherapeutic drug, 5-fluorouracil (5FU), to induce cytopenia. 5FU dose was optimized in the 129/SvImJ, C57BL/6J, BALB/cJ, and C3H/HeJ strains of laboratory mice. N-ethyl-N-nitrosourea (ENU) mutagenesis was performed on 129/SvImJ males and phenotypic variants were identified by backcrossing on to the C57BL/6J background. G1 animals were challenged with 100 µg/g 5FU and phenodeviants with altered platelet recovery were monitored. Of 546 G1 animals tested, 15 phenodeviants were identified that displayed increased baseline platelet number, a platelet overshoot, or delayed platelet recovery, thereby demonstrating the utility of this approach for uncovering mutations in megakaryocyte and platelet development. Four G1 mice were selected for further analysis. The phenotypes were heritable in all four strains and genetic mapping identified a chromosome location in two of the three G2 lines tested. In conclusion, our group has developed a sensitized random mutagenesis screen utilizing 5FU and has shown that the strain combination of 129/SvImJ × C57BL/6J is robust for identification of founder lines with defects in megakaryocyte and platelet development.