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Human recombinant B cell activating factor (BAFF) is secreted as 3-mers, which can associate to form 60-mers in culture supernatants. However, the presence of BAFF multimers in humans is still debated and it is incompletely understood how BAFF multimers activate the B cells. Here, we demonstrate that BAFF can exist as 60-mers or higher order multimers in human plasma. In vitro, BAFF 60-mer strongly induced the transcriptome of B cells which was partly attenuated by antagonism using a soluble fragment of BAFF receptor 3. Furthermore, compared to BAFF 3-mer, BAFF 60-mer strongly induced a transient classical and prolonged alternate NF-κB signaling, glucose oxidation by both aerobic glycolysis and oxidative phosphorylation, and succinate utilization by mitochondria. BAFF antagonism selectively attenuated classical NF-κB signaling and glucose oxidation. Altogether, our results suggest critical roles of BAFF 60-mer and its BAFF receptor 3 binding site in hyperactivation of B cells.
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Macroautophagy (MA) regulates cellular quality control and energy balance. For example, loss of MA in aP2-positive adipocytes converts white adipose tissue (WAT) into brown adipose tissue (BAT)-like, enhancing BAT function and thereby insulin sensitivity. However, whether MA regulates early BAT development is unknown. We report that deleting Atg7 in myogenic Myf5+ progenitors inhibits MA in Myf5-cell-derived BAT and muscle. Knock out (KO) mice have defective BAT differentiation and function. Surprisingly, their body temperature is higher due to WAT lipolysis-driven increases in fatty acid oxidation in 'Beige' cells in inguinal WAT, BAT and muscle. KO mice also present impaired muscle differentiation, reduced muscle mass and glucose intolerance. Our studies show that ATG7 in Myf5+ progenitors is required to maintain energy and glucose homeostasis through effects on BAT and muscle development. Decreased MA in myogenic progenitors with age and/or overnutrition might contribute to the metabolic defects and sarcopenia observed in these conditions.
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Tejido Adiposo Pardo/metabolismo , Autofagia , Metabolismo Energético , Glucosa/metabolismo , Homeostasis , Músculo Esquelético/metabolismo , Factor 5 Regulador Miogénico/metabolismo , Tejido Adiposo Pardo/crecimiento & desarrollo , Animales , Proteína 7 Relacionada con la Autofagia , Diferenciación Celular , Ácidos Grasos/metabolismo , Ratones , Proteínas Asociadas a Microtúbulos/genética , Proteínas Asociadas a Microtúbulos/metabolismo , Músculo Esquelético/crecimiento & desarrollo , Factor 5 Regulador Miogénico/genética , Células Madre/citología , Células Madre/metabolismoRESUMEN
Autophagy degrades cytoplasmic contents to achieve cellular homeostasis. We show that selective loss of autophagy in hypothalamic proopiomelanocortin (POMC) neurons decreases α-melanocyte-stimulating hormone (MSH) levels, promoting adiposity, impairing lipolysis and altering glucose homeostasis. Ageing reduces hypothalamic autophagy and α-MSH levels, and aged-mice phenocopy, the adiposity and lipolytic defect observed in POMC neuron autophagy-null mice. Intraperitoneal isoproterenol restores lipolysis in both models, demonstrating normal adipocyte catecholamine responsiveness. We propose that an unconventional, autophagosome-mediated form of secretion in POMC neurons controls energy balance by regulating α-MSH production. Modulating hypothalamic autophagy might have implications for preventing obesity and metabolic syndrome of ageing.
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Autofagia/genética , Hipotálamo/metabolismo , Lipólisis/genética , Neuronas/metabolismo , Proopiomelanocortina/metabolismo , Adiposidad/genética , Envejecimiento/genética , Envejecimiento/metabolismo , Animales , Proteína 7 Relacionada con la Autofagia , Resistencia a la Insulina/genética , Masculino , Ratones , Ratones Noqueados , Proteínas Asociadas a Microtúbulos/genética , Proopiomelanocortina/genética , alfa-MSH/metabolismoRESUMEN
PURPOSE: To explore the potential of testosterone therapy in managing cytopenias in myelodysplastic neoplasm and investigate the link between hypogonadism and hematologic malignancies. METHODS: A case of a patient with intermediate-risk myelodysplastic neoplasm and hypogonadism treated with testosterone replacement therapy is presented. Testosterone, prostate specific antigen, and erythropoietin levels were checked prior to therapy initiation and 3 months after. Blood counts were monitored over time. This is followed by a literature review of testosterone use in myelodysplastic neoplasm and the prevalence of hypogonadism in hematologic malignancies. RESULTS: The patient showed sustained improvement in anemia with testosterone therapy and reported subjective improvement in his weakness and fatigue. This improvement occurred even in the setting of an undetectable follow up erythropoietin level. His repeat prostate specific antigen levels remained low, while testosterone levels showed marked improvement. The literature review demonstrated positive response rates for testosterone in treating myelodysplastic neoplasm-related cytopenias, and showed a higher incidence of hypogonadism in hematologic malignancies. CONCLUSION: Our review suggests that the use of testosterone in low and intermediate-risk myelodysplastic neoplasm is underexplored and poses to have significant potential as a future therapeutic agent, after careful consideration of risks and benefits. In addition, the incidence of hypogonadism in myelodysplastic neoplasm and its potential impact on exacerbating cytopenias in myelodysplastic neoplasm warrants further investigation.
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Hipogonadismo , Síndromes Mielodisplásicos , Testosterona , Humanos , Testosterona/uso terapéutico , Masculino , Síndromes Mielodisplásicos/tratamiento farmacológico , Síndromes Mielodisplásicos/complicaciones , Hipogonadismo/tratamiento farmacológico , Terapia de Reemplazo de Hormonas/métodos , Anemia/tratamiento farmacológico , Anemia/etiología , Anciano , Persona de Mediana Edad , CitopeniaRESUMEN
The role of splicing dysregulation in cancer is underscored by splicing factor mutations; however, its impact in the absence of such rare mutations is poorly understood. To reveal complex patient subtypes and putative regulators of pathogenic splicing in Acute Myeloid Leukemia (AML), we developed a new approach called OncoSplice. Among diverse new subtypes, OncoSplice identified a biphasic poor prognosis signature that partially phenocopies U2AF1-mutant splicing, impacting thousands of genes in over 40% of adult and pediatric AML cases. U2AF1-like splicing co-opted a healthy circadian splicing program, was stable over time and induced a leukemia stem cell (LSC) program. Pharmacological inhibition of the implicated U2AF1-like splicing regulator, PRMT5, rescued leukemia mis-splicing and inhibited leukemic cell growth. Genetic deletion of IRAK4, a common target of U2AF1-like and PRMT5 treated cells, blocked leukemia development in xenograft models and induced differentiation. These analyses reveal a new prognostic alternative-splicing mechanism in malignancy, independent of splicing-factor mutations.
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While therapies targeting CD19 by antibodies, chimeric antigen receptor T cells (CAR-T), and T cell engagers have improved the response rates in B cell malignancies, the emergence of resistant cell populations with low CD19 expression can lead to relapsed disease. We developed an in vitro model of adaptive resistance facilitated by chronic exposure of leukemia cells to a CD19 immunotoxin. Single-cell RNA-Seq (scRNA-Seq) showed an increase in transcriptionally distinct CD19lo populations among resistant cells. Mass cytometry demonstrated that CD22 was also decreased in these CD19lo-resistant cells. An assay for transposase-accessible chromatin with sequencing (ATAC-Seq) showed decreased chromatin accessibility at promoters of both CD19 and CD22 in the resistant cell populations. Combined loss of both CD19 and CD22 antigens was validated in samples from pediatric and young adult patients with B cell acute lymphoblastic leukemia (B-ALL) that relapsed after CD19 CAR-T-targeted therapy. Functionally, resistant cells were characterized by slower growth and lower basal levels of MEK activation. CD19lo resistant cells exhibited preserved B cell receptor signaling and were more sensitive to both Bruton's tyrosine kinase (BTK) and MEK inhibition. These data demonstrate that resistance to CD19 immunotherapies can result in decreased expression of both CD19 and CD22 and can result in dependency on BTK pathways.
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Antígenos CD19 , Leucemia-Linfoma Linfoblástico de Células Precursoras , Lectina 2 Similar a Ig de Unión al Ácido Siálico , Niño , Humanos , Adulto Joven , Agammaglobulinemia Tirosina Quinasa , Antígenos CD19/genética , Cromatina , Inmunoterapia Adoptiva , Quinasas de Proteína Quinasa Activadas por Mitógenos , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/terapia , Receptores Quiméricos de Antígenos , Lectina 2 Similar a Ig de Unión al Ácido Siálico/genéticaRESUMEN
Dysregulation of the innate immune system and inflammatory-related pathways has been implicated in hematopoietic defects in the bone marrow microenvironment and associated with aging, clonal hematopoiesis, myelodysplastic syndromes (MDS), and acute myeloid leukemia (AML). As the innate immune system and its pathway regulators have been implicated in the pathogenesis of MDS/AML, novel approaches targeting these pathways have shown promising results. Variability in expression of Toll like receptors (TLRs), abnormal levels of MyD88 and subsequent activation of NF-κß, dysregulated IL1-receptor associated kinases (IRAK), alterations in TGF-ß and SMAD signaling, high levels of S100A8/A9 have all been implicated in pathogenesis of MDS/AML. In this review we not only discuss the interplay of various innate immune pathways in MDS pathogenesis but also focus on potential therapeutic targets from recent clinical trials including the use of monoclonal antibodies and small molecule inhibitors against these pathways.
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Background: Cancer patients show increased morbidity with COVID-19 and need effective immunization strategies. Many healthcare regulatory agencies recommend administering 'booster' doses of COVID-19 vaccines beyond the standard two-dose series, for this group of patients. Therefore, studying the efficacy of these additional vaccine doses against SARS-CoV-2 and variants of concern is of utmost importance in this immunocompromised patient population. Methods: We conducted a prospective single arm clinical trial enrolling patients with cancer that had received two doses of mRNA or one dose of AD26.CoV2.S vaccine and administered a third dose of mRNA vaccine. We further enrolled patients that had no or low responses to three mRNA COVID vaccines and assessed the efficacy of a fourth dose of mRNA vaccine. Efficacy was assessed by changes in anti-spike antibody, T-cell activity, and neutralization activity, which were again assessed at baseline and 4 weeks. Results: We demonstrate that a third dose of COVID-19 vaccine leads to seroconversion in 57% of patients that were seronegative after primary vaccination series. The immune response is durable as assessed by anti-SARS-CoV-2 (anti-S) antibody titers, T-cell activity, and neutralization activity against wild-type (WT) SARS-CoV2 and BA1.1.529 at 6 months of follow-up. A subset of severely immunocompromised hematologic malignancy patients that were unable to mount an adequate immune response (titer <1000 AU/mL) after the third dose and were treated with a fourth dose in a prospective clinical trial which led to adequate immune boost in 67% of patients. Low baseline IgM levels and CD19 counts were associated with inadequate seroconversion. Booster doses induced limited neutralization activity against the Omicron variant. Conclusions: These results indicate that third dose of COVID vaccine induces durable immunity in cancer patients and an additional dose can further stimulate immunity in a subset of patients with inadequate response. Funding: Leukemia Lymphoma Society, National Cancer Institute. Clinical trial number: NCT05016622.
People with cancer have a higher risk of death or severe complications from COVID-19. As a result, vaccinating cancer patients against COVID-19 is critical. But patients with cancer, particularly blood or lymphatic system cancers, are less likely to develop protective immunity after COVID-19 vaccination. Immune suppression caused by cancer or cancer therapies may explain the poor vaccine response. Booster doses of the vaccine may improve the vaccine response in patients with cancer. But limited information is available about how well booster doses protect patients with cancer against COVID-19. Thakkar et al. show that a third dose of a COVID-19 vaccine can induce a protective immune response in half of the patients with cancer with no immunity after the first two doses. In the experiments, Thakkar et al. tracked the immune reaction to COVID-19 booster shots in 106 cancer patients. A third booster dose protected patients for up to four to six months and reduced breakthrough infection rates to low levels. Eighteen patients with blood cancers and severe immune suppression had an inadequate immune response after three doses of the vaccine; a fourth dose boosted the immune response for two-thirds of them, which for some included neutralization of variants such as Omicron. The experiments show that booster doses can increase COVID-19 vaccine protection for patients with cancer, even those who do not respond to the initial vaccine series. Thakkar et al. also show that pre-vaccine levels of two molecules linked to the immune system, (immunoglobin M and the CD19 antigen) predicted the patients' vaccine response, which might help physicians identify which individuals would benefit from booster doses.
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COVID-19 , Neoplasias , Humanos , Vacunas contra la COVID-19 , Ad26COVS1 , Estudios Prospectivos , ARN Viral , COVID-19/prevención & control , SARS-CoV-2 , Neoplasias/terapia , Inmunidad , Anticuerpos AntiviralesRESUMEN
Malignancies can become reliant on glutamine as an alternative energy source and as a facilitator of aberrant DNA methylation, thus implicating glutaminase (GLS) as a potential therapeutic target. We demonstrate preclinical synergy of telaglenastat (CB-839), a selective GLS inhibitor, when combined with azacytidine (AZA), in vitro and in vivo, followed by a phase Ib/II study of the combination in patients with advanced MDS. Treatment with telaglenastat/AZA led to an ORR of 70% with CR/mCRs in 53% patients and a median overall survival of 11.6 months. scRNAseq and flow cytometry demonstrated a myeloid differentiation program at the stem cell level in clinical responders. Expression of non-canonical glutamine transporter, SLC38A1, was found to be overexpressed in MDS stem cells; was associated with clinical responses to telaglenastat/AZA and predictive of worse prognosis in a large MDS cohort. These data demonstrate the safety and efficacy of a combined metabolic and epigenetic approach in MDS.
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Advanced pancreatic ductal adenocarcinoma (PDAC) is usually an incurable malignancy that needs newer therapeutic targets. Interleukin-1 receptor accessory protein (IL1RAP) is an innate immune mediator that regulates activation of pro-inflammatory and mitogenic signaling pathways. Immunohistochemistry on tissue microarrays demonstrated expression of IL1RAP in majority of human PDAC specimens and in murine pancreatic tumors from K-RasG122D/p53R172H/PDXCre (KPC) mice. Single cell RNA-Seq analysis of human primary pre-neoplastic lesions and adenocarcinoma specimens indicated that overexpression occurs during carcinogenesis. IL1RAP overexpression was associated with worse overall survival. IL1RAP knockdown significantly reduced cell viability, invasiveness, and clonogenic growth in pancreatic cancer cell lines. Inhibition of the downstream interleukin-1 receptor-associated kinase 4 (IRAK4) using two pharmacologic inhibitors, CA-4948 and PF06650833, resulted in reduced growth in pancreatic cancer cell lines and in xenograft models.
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Adenocarcinoma , Carcinoma Ductal Pancreático , Proteína Accesoria del Receptor de Interleucina-1 , Neoplasias Pancreáticas , Adenocarcinoma/patología , Animales , Carcinogénesis , Carcinoma Ductal Pancreático/patología , Línea Celular Tumoral , Humanos , Inmunidad Innata , Proteína Accesoria del Receptor de Interleucina-1/metabolismo , Ratones , Neoplasias Pancreáticas/patología , Neoplasias PancreáticasRESUMEN
Background: Mutations in the SF3B1 splicing factor are commonly seen in myelodysplastic syndromes (MDS) and acute myeloid leukemia (AML), yet the specific oncogenic pathways activated by mis-splicing have not been fully elucidated. Inflammatory immune pathways have been shown to play roles in the pathogenesis of MDS, though the exact mechanisms of their activation in splicing mutant cases are not well understood. Methods: RNA-seq data from SF3B1 mutant samples was analyzed and functional roles of interleukin-1 receptor-associated kinase 4 (IRAK4) isoforms were determined. Efficacy of IRAK4 inhibition was evaluated in preclinical models of MDS/AML. Results: RNA-seq splicing analysis of SF3B1 mutant MDS samples revealed retention of full-length exon 6 of IRAK4, a critical downstream mediator that links the Myddosome to inflammatory NF-kB activation. Exon 6 retention leads to a longer isoform, encoding a protein (IRAK4-long) that contains the entire death domain and kinase domain, leading to maximal activation of NF-kB. Cells with wild-type SF3B1 contain smaller IRAK4 isoforms that are targeted for proteasomal degradation. Expression of IRAK4-long in SF3B1 mutant cells induces TRAF6 activation leading to K63-linked ubiquitination of CDK2, associated with a block in hematopoietic differentiation. Inhibition of IRAK4 with CA-4948, leads to reduction in NF-kB activation, inflammatory cytokine production, enhanced myeloid differentiation in vitro and reduced leukemic growth in xenograft models. Conclusions: SF3B1 mutation leads to expression of a therapeutically targetable, longer, oncogenic IRAK4 isoform in AML/MDS models. Funding: This work was supported by Cincinnati Children's Hospital Research Foundation, Leukemia Lymphoma Society, and National Institute of Health (R35HL135787, RO1HL111103, RO1DK102759, RO1HL114582), Gabrielle's Angel Foundation for Cancer Research, and Edward P. Evans Foundation grants to DTS. AV is supported by Edward P. Evans Foundation, National Institute of Health (R01HL150832, R01HL139487, R01CA275007), Leukemia and Lymphoma Society, Curis and a gift from the Jane and Myles P. Dempsey family. AP and JB are supported by Blood Cancer UK (grants 13042 and 19004). GC is supported by a training grant from NYSTEM. We acknowledge support of this research from The Einstein Training Program in Stem Cell Research from the Empire State Stem Cell Fund through New York State Department of Health Contract C34874GG. MS is supported by a National Institute of Health Research Training and Career Development Grant (F31HL132420).
Genes contain blocks of code that tell cells how to make each part of a protein. Between these blocks are sections of linking DNA, which cells remove when they are preparing to use their genes. Scientists call this process 'splicing'. Cells can splice some genes in more than one way, allowing them to make different proteins from the same genetic code. Mutations that affect the splicing process can change the way cells make their proteins, leading to disease. For example, the myelodysplastic syndromes are a group of blood cancers often caused by mutations in splicing proteins, such as SF3B1. The disorder stops blood cells from maturing and causes abnormal inflammation. So far, the link between splicing, blood cell immaturity, inflammation and cancer is not clear. To find out more, Choudhary, Pellagatti et al. looked at the spliced genetic code from people with myelodysplastic syndromes. Mutations in the splicing protein SF3B1 changed the way cells spliced an important signalling molecule known as IRAK4. Affected cells cut out less genetic code and made a longer version of this signalling protein, named IRAK4-Long. This altered protein activated inflammation and stopped blood cells from maturing. Blocking IRAK4-Long reversed the effects. It also reduced tumour formation in mice carrying affected human cells. The molecule used to block IRAK4, CA-4948 also known as Emavusertib is currently being evaluated in clinical trials for myelodysplastic syndromes and other types of blood cancer. The work of Choudhary, Pellagatti et al. could help scientists to design genetic tests to predict which patients might benefit from this treatment.
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Leucemia Mieloide Aguda , Síndromes Mielodisplásicos , Fosfoproteínas/metabolismo , Factores de Empalme de ARN/metabolismo , Niño , Humanos , Quinasas Asociadas a Receptores de Interleucina-1/genética , Quinasas Asociadas a Receptores de Interleucina-1/metabolismo , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/patología , Mutación , Síndromes Mielodisplásicos/metabolismo , FN-kappa B/genética , FN-kappa B/metabolismo , Isoformas de Proteínas/metabolismo , Empalme del ARNRESUMEN
The terrorist attacks on the World Trade Center (WTC) created an unprecedented environmental exposure to aerosolized dust, gases and potential carcinogens. Clonal hematopoiesis (CH) is defined as the acquisition of somatic mutations in blood cells and is associated with smoking and exposure to genotoxic stimuli. Here we show that deep targeted sequencing of blood samples identified a significantly higher proportion of WTC-exposed first responders with CH (10%; 48 out of 481) when compared with non-WTC-exposed firefighters (6.7%; 17 out of 255; odds ratio, 3.14; 95% confidence interval, 1.64-6.03; P = 0.0006) after controlling for age, sex and race/ethnicity. The frequency of somatic mutations in WTC-exposed first responders showed an age-related increase and predominantly affected DNMT3A, TET2 and other CH-associated genes. Exposure of lymphoblastoid cells to WTC particulate matter led to dysregulation of DNA replication at common fragile sites in vitro. Moreover, mice treated with WTC particulate matter developed an increased burden of mutations in hematopoietic stem and progenitor cell compartments. In summary, the high burden of CH in WTC-exposed first responders provides a rationale for enhanced screening and preventative efforts in this population.
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Desastres , Socorristas , Ataques Terroristas del 11 de Septiembre , Animales , Hematopoyesis Clonal , Polvo , Humanos , RatonesRESUMEN
OBJECTIVE: Brown adipose tissue (BAT) is specialized in thermogenesis. The conversion of energy into heat in brown adipocytes proceeds via stimulation of ß-adrenergic receptor (ßAR)-dependent signaling and activation of mitochondrial uncoupling protein 1 (UCP1). We have previously demonstrated a functional role for pannexin-1 (Panx1) channels in white adipose tissue; however, it is not known whether Panx1 channels play a role in the regulation of brown adipocyte function. Here, we tested the hypothesis that Panx1 channels are involved in brown adipocyte activation and thermogenesis. METHODS: In an immortalized brown pre-adipocytes cell line, Panx1 currents were measured using patch-clamp electrophysiology. Flow cytometry was used for assessment of dye uptake and luminescence assays for adenosine triphosphate (ATP) release, and cellular temperature measurement was performed using a ratiometric fluorescence thermometer. We used RNA interference and expression plasmids to manipulate expression of wild-type and mutant Panx1. We used previously described adipocyte-specific Panx1 knockout mice (Panx1Adip-/-) and generated brown adipocyte-specific Panx1 knockout mice (Panx1BAT-/-) to study pharmacological or cold-induced thermogenesis. Glucose uptake into brown adipose tissue was quantified by positron emission tomography (PET) analysis of 18F-fluorodeoxyglucose (18F-FDG) content. BAT temperature was measured using an implantable telemetric temperature probe. RESULTS: In brown adipocytes, Panx1 channel activity was induced either by apoptosis-dependent caspase activation or by ß3AR stimulation via a novel mechanism that involves Gßγ subunit binding to Panx1. Inactivation of Panx1 channels in cultured brown adipocytes resulted in inhibition of ß3AR-induced lipolysis, UCP-1 expression, and cellular thermogenesis. In mice, adiponectin-Cre-dependent genetic deletion of Panx1 in all adipose tissue depots resulted in defective ß3AR agonist- or cold-induced thermogenesis in BAT and suppressed beigeing of white adipose tissue. UCP1-Cre-dependent Panx1 deletion specifically in brown adipocytes reduced the capacity for adaptive thermogenesis without affecting beigeing of white adipose tissue and aggravated diet-induced obesity and insulin resistance. CONCLUSIONS: These data demonstrate that Gßγ-dependent Panx1 channel activation is involved in ß3AR-induced thermogenic regulation in brown adipocytes. Identification of Panx1 channels in BAT as novel thermo-regulatory elements downstream of ß3AR activation may have therapeutic implications.
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Tejido Adiposo Pardo/metabolismo , Conexinas/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Termogénesis/fisiología , Adipocitos Marrones/metabolismo , Adiponectina/metabolismo , Tejido Adiposo Pardo/patología , Tejido Adiposo Blanco/metabolismo , Animales , Frío , Conexinas/genética , Fluorodesoxiglucosa F18 , Resistencia a la Insulina , Lipólisis , Ratones , Ratones Noqueados , Proteínas del Tejido Nervioso/genética , Obesidad/metabolismo , Transducción de Señal , Termogénesis/genética , TranscriptomaRESUMEN
The BCL2-inhibitor, Venetoclax (VEN), has shown significant anti-leukemic efficacy in combination with the DNMT-inhibitor, Azacytidine (AZA). To explore the mechanisms underlying the selective sensitivity of mutant leukemia cells to VEN and AZA, we used cell-based isogenic models containing a common leukemia-associated mutation in the epigenetic regulator ASXL1. KBM5 cells with CRISPR/Cas9-mediated correction of the ASXL1G710X mutation showed reduced leukemic growth, increased myeloid differentiation, and decreased HOXA and BCL2 gene expression in vitro compared to uncorrected KBM5 cells. Increased expression of the anti-apoptotic gene, BCL2, was also observed in bone marrow CD34+ cells from ASXL1 mutant MDS patients compared to CD34+ cells from wild-type MDS cases. ATAC-sequencing demonstrated open chromatin at the BCL2 promoter in the ASXL1 mutant KBM5 cells. BH3 profiling demonstrated increased dependence of mutant cells on BCL2. Upon treatment with VEN, mutant cells demonstrated increased growth inhibition. In addition, genome-wide methylome analysis of primary MDS samples and isogenic cell lines demonstrated increased gene-body methylation in ASXL1 mutant cells, with consequently increased sensitivity to AZA. These data mechanistically link the common leukemia-associated mutation ASXL1 to enhanced sensitivity to VEN and AZA via epigenetic upregulation of BCL2 expression and widespread alterations in DNA methylation.
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Antineoplásicos/farmacología , Azacitidina/farmacología , Compuestos Bicíclicos Heterocíclicos con Puentes/farmacología , Leucemia Mielógena Crónica BCR-ABL Positiva/genética , Proteínas Represoras/genética , Sulfonamidas/farmacología , Línea Celular Tumoral , Epigénesis Genética/efectos de los fármacos , Regulación Leucémica de la Expresión Génica/efectos de los fármacos , Humanos , Leucemia Mielógena Crónica BCR-ABL Positiva/tratamiento farmacológico , Mutación/efectos de los fármacos , Mutación Puntual/efectos de los fármacosRESUMEN
Innate immune signaling through the NLRP3 inflammasome is activated by multiple diabetes-related stressors, but whether targeting the inflammasome is beneficial for diabetes is still unclear. Nucleoside reverse-transcriptase inhibitors (NRTI), drugs approved to treat HIV-1 and hepatitis B infections, also block inflammasome activation. Here, we show, by analyzing five health insurance databases, that the adjusted risk of incident diabetes is 33% lower in patients with NRTI exposure among 128,861 patients with HIV-1 or hepatitis B (adjusted hazard ratio for NRTI exposure, 0.673; 95% confidence interval, 0.638 to 0.710; P < 0.0001; 95% prediction interval, 0.618 to 0.734). Meanwhile, an NRTI, lamivudine, improves insulin sensitivity and reduces inflammasome activation in diabetic and insulin resistance-induced human cells, as well as in mice fed with high-fat chow; mechanistically, inflammasome-activating short interspersed nuclear element (SINE) transcripts are elevated, whereas SINE-catabolizing DICER1 is reduced, in diabetic cells and mice. These data suggest the possibility of repurposing an approved class of drugs for prevention of diabetes.
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Diabetes Mellitus Tipo 2/tratamiento farmacológico , Reposicionamiento de Medicamentos , Inflamasomas/efectos de los fármacos , Resistencia a la Insulina , Inhibidores de la Transcriptasa Inversa/farmacología , Adipocitos/metabolismo , Animales , Supervivencia Celular , ARN Helicasas DEAD-box/metabolismo , Diabetes Mellitus Tipo 2/prevención & control , Dieta Alta en Grasa/efectos adversos , VIH-1/efectos de los fármacos , Hepatitis B , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Células Musculares/metabolismo , Ribonucleasa III/metabolismoRESUMEN
Even though pancreatic ductal adenocarcinoma (PDAC) is associated with fibrotic stroma, the molecular pathways regulating the formation of cancer associated fibroblasts (CAFs) are not well elucidated. An epigenomic analysis of patient-derived and de-novo generated CAFs demonstrated widespread loss of cytosine methylation that was associated with overexpression of various inflammatory transcripts including CXCR4. Co-culture of neoplastic cells with CAFs led to increased invasiveness that was abrogated by inhibition of CXCR4. Metabolite tracing revealed that lactate produced by neoplastic cells leads to increased production of alpha-ketoglutarate (aKG) within mesenchymal stem cells (MSCs). In turn, aKG mediated activation of the demethylase TET enzyme led to decreased cytosine methylation and increased hydroxymethylation during de novo differentiation of MSCs to CAF. Co-injection of neoplastic cells with TET-deficient MSCs inhibited tumor growth in vivo. Thus, in PDAC, a tumor-mediated lactate flux is associated with widespread epigenomic reprogramming that is seen during CAF formation.
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Fibroblastos Asociados al Cáncer/patología , Reprogramación Celular/efectos de los fármacos , Epigénesis Genética/efectos de los fármacos , Ácido Láctico/farmacología , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patología , 5-Metilcitosina/análogos & derivados , 5-Metilcitosina/metabolismo , Animales , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/patología , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Transformación Celular Neoplásica/efectos de los fármacos , Reprogramación Celular/genética , Metilación de ADN/efectos de los fármacos , Humanos , Ácidos Cetoglutáricos/metabolismo , Células Madre Mesenquimatosas/efectos de los fármacos , Células Madre Mesenquimatosas/metabolismo , Ratones , Invasividad Neoplásica , Receptores CXCR4/metabolismo , Células del Estroma/efectos de los fármacos , Células del Estroma/metabolismo , Transcriptoma/genética , Neoplasias PancreáticasRESUMEN
Autophagy failure is associated with metabolic insufficiency. Although caloric restriction (CR) extends healthspan, its adherence in humans is poor. We established an isocaloric twice-a-day (ITAD) feeding model wherein ITAD-fed mice consume the same food amount as ad libitum controls but at two short windows early and late in the diurnal cycle. We hypothesized that ITAD feeding will provide two intervals of intermeal fasting per circadian period and induce autophagy. We show that ITAD feeding modifies circadian autophagy and glucose/lipid metabolism that correlate with feeding-driven changes in circulating insulin. ITAD feeding decreases adiposity and, unlike CR, enhances muscle mass. ITAD feeding drives energy expenditure, lowers lipid levels, suppresses gluconeogenesis, and prevents age/obesity-associated metabolic defects. Using liver-, adipose-, myogenic-, and proopiomelanocortin neuron-specific autophagy-null mice, we mapped the contribution of tissue-specific autophagy to system-wide benefits of ITAD feeding. Our studies suggest that consuming two meals a day without CR could prevent the metabolic syndrome.
Asunto(s)
Autofagia , Restricción Calórica , Ritmo Circadiano , Ayuno , Síndrome Metabólico/prevención & control , Adiposidad , Factores de Edad , Animales , Glucemia/análisis , Conducta Alimentaria , Femenino , Humanos , Insulina/sangre , Lípidos/análisis , Hígado/metabolismo , Masculino , Ratones , Ratones Noqueados , Desarrollo de MúsculosRESUMEN
The integrative physiology of inter-organ communication in lipophagy regulation is not well understood. Lipophagy and the cytosolic lipases ATGL and HSL contribute to lipid droplet (LD) mobilization; however, whether autophagy proteins engage with lipases to promote lipid utilization remains unknown. Here, we show that cold induces autophagy in proopiomelanocortin (POMC) neurons and activates lipophagy in brown adipose tissue (BAT) and liver in mice. Targeted activation of autophagy in POMC neurons via intra-hypothalamic rapamycin is sufficient to trigger lipid utilization in room temperature-housed mice. Conversely, inhibiting autophagy in POMC neurons or in peripheral tissues or denervating BAT blocks lipid utilization. Unexpectedly, the autophagosome marker LC3 is mechanistically coupled to ATGL-mediated lipolysis. ATGL exhibits LC3-interacting region (LIR) motifs, and mutating a single LIR motif on ATGL displaces ATGL from LD and disrupts lipolysis. Thus, cold-induced activation of central autophagy activates lipophagy and cytosolic lipases in a complementary manner to mediate lipolysis in peripheral tissues.
Asunto(s)
Tejido Adiposo Pardo/metabolismo , Autofagia , Hipotálamo/citología , Lipólisis , Hígado/metabolismo , Adipocitos Marrones/fisiología , Tejido Adiposo Pardo/citología , Tejido Adiposo Pardo/inervación , Secuencia de Aminoácidos , Animales , Frío , Femenino , Lipasa/metabolismo , Gotas Lipídicas/metabolismo , Hígado/citología , Lisosomas/metabolismo , Masculino , Ratones Endogámicos C57BL , Ratones Transgénicos , Proteínas Asociadas a Microtúbulos/metabolismo , Datos de Secuencia Molecular , Neuronas/fisiología , Consumo de Oxígeno , Proopiomelanocortina/metabolismoRESUMEN
Autophagy is a conserved pathway that maintains cellular quality control. Extracellular signal-regulated kinase (ERK) controls various aspects of cell physiology including proliferation. Multiple signalling cascades, including ERK, have been shown to regulate autophagy, however whether autophagy proteins (ATG) regulate cell signalling is unknown. Here we show that growth factor exposure increases the interaction of ERK cascade components with ATG proteins in the cytosol and nucleus. ERK and its upstream kinase MEK localize to the extra-luminal face of autophagosomes. ERK2 interacts with ATG proteins via its substrate-binding domains. Deleting Atg7 or Atg5 or blocking LC3 lipidation or ATG5-ATG12 conjugation decreases ERK phosphorylation. Conversely, increasing LC3-II availability by silencing the cysteine protease ATG4B or acute trehalose exposure increases ERK phosphorylation. Decreased ERK phosphorylation in Atg5â»/â» cells does not occur from overactive phosphatases. Our findings thus reveal an unconventional function of ATG proteins as cellular scaffolds in the regulation of ERK phosphorylation.