RESUMEN
The energetic costs of duplicating chromatin are large and therefore likely depend on nutrient sensing checkpoints and metabolic inputs. By studying chromatin modifiers regulated by epithelial growth factor, we identified histone acetyltransferase 1 (HAT1) as an induced gene that enhances proliferation through coordinating histone production, acetylation, and glucose metabolism. In addition to its canonical role as a cytoplasmic histone H4 acetyltransferase, we isolated a HAT1-containing complex bound specifically at promoters of H4 genes. HAT1-dependent transcription of H4 genes required an acetate-sensitive promoter element. HAT1 expression was critical for S-phase progression and maintenance of H3 lysine 9 acetylation at proliferation-associated genes, including histone genes. Therefore, these data describe a feedforward circuit whereby HAT1 captures acetyl groups on nascent histones and drives H4 production by chromatin binding to support chromatin replication and acetylation. These findings have important implications for human disease, since high HAT1 levels associate with poor outcomes across multiple cancer types.
Asunto(s)
Histona Acetiltransferasas/metabolismo , Histonas/metabolismo , Proteínas de Neoplasias/metabolismo , Neoplasias/metabolismo , Regiones Promotoras Genéticas , Fase S , Transcripción Genética , Células A549 , Acetilación , Animales , Cromatina/genética , Cromatina/metabolismo , Femenino , Histona Acetiltransferasas/genética , Histonas/genética , Humanos , Ratones , Ratones Endogámicos BALB C , Proteínas de Neoplasias/genética , Neoplasias/genéticaRESUMEN
Loss of estrogen receptor (ER) pathway activity promotes breast cancer progression, yet how this occurs remains poorly understood. Here, we show that serine starvation, a metabolic stress often found in breast cancer, represses estrogen receptor alpha (ERα) signaling by reprogramming glucose metabolism and epigenetics. Using isotope tracing and time-resolved metabolomic analyses, we demonstrate that serine is required to maintain glucose flux through glycolysis and the TCA cycle to support acetyl-CoA generation for histone acetylation. Consequently, limiting serine depletes histone H3 lysine 27 acetylation (H3K27ac), particularly at the promoter region of ER pathway genes including the gene encoding ERα, ESR1. Mechanistically, serine starvation impairs acetyl-CoA-dependent gene expression by inhibiting the entry of glycolytic carbon into the TCA cycle and down-regulating the mitochondrial citrate exporter SLC25A1, a critical enzyme in the production of nucleocytosolic acetyl-CoA from glucose. Consistent with this model, total H3K27ac and ERα expression are suppressed by SLC25A1 inhibition and restored by acetate, an alternate source of acetyl-CoA, in serine-free conditions. We thus uncover an unexpected role for serine in sustaining ER signaling through the regulation of acetyl-CoA metabolism.
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Receptor alfa de Estrógeno , Histonas , Acetilcoenzima A , Receptor alfa de Estrógeno/genética , Histonas/genética , Receptores de Estrógenos , GlucosaRESUMEN
Intrinsic immune responses autonomously inhibit viral replication and spread. One pathway that restricts viral infection in plants and insects is RNA interference (RNAi), which targets and degrades viral RNA to limit infection. To identify additional genes involved in intrinsic antiviral immunity, we screened Drosophila cells for modulators of viral infection using an RNAi library. We identified Ars2 as a key component of Drosophila antiviral immunity. Loss of Ars2 in cells, or in flies, increases susceptibility to RNA viruses. Consistent with its antiviral properties, we found that Ars2 physically interacts with Dcr-2, modulates its activity in vitro, and is required for siRNA-mediated silencing. Furthermore, we show that Ars2 plays an essential role in miRNA-mediated silencing, interacting with the Microprocessor and stabilizing pri-miRNAs. The identification of Ars2 as a player in these small RNA pathways provides new insight into the biogenesis of small RNAs that may be extended to other systems.
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Proteínas de Drosophila/metabolismo , Drosophila/genética , Drosophila/inmunología , Complejo Proteico Nuclear de Unión a la Caperuza/metabolismo , Interferencia de ARN , Vesiculovirus/inmunología , Animales , Drosophila/virología , Proteínas de Drosophila/química , Proteínas de Drosophila/genética , MicroARNs/genética , ARN Bicatenario/metabolismo , ARN Interferente Pequeño/genética , Receptores del Ligando Inductor de Apoptosis Relacionado con TNF/metabolismo , Vesiculovirus/genéticaRESUMEN
Here we identify a component of the nuclear RNA cap-binding complex (CBC), Ars2, that is important for miRNA biogenesis and critical for cell proliferation. Unlike other components of the CBC, Ars2 expression is linked to the proliferative state of the cell. Deletion of Ars2 is developmentally lethal, and deletion in adult mice led to bone marrow failure whereas parenchymal organs composed of nonproliferating cells were unaffected. Depletion of Ars2 or CBP80 from proliferating cells impaired miRNA-mediated repression and led to alterations in primary miRNA processing in the nucleus. Ars2 depletion also reduced the levels of several miRNAs, including miR-21, let-7, and miR-155, that are implicated in cellular transformation. These findings provide evidence for a role for Ars2 in RNA interference regulation during cell proliferation.
Asunto(s)
Proliferación Celular , Complejo Proteico Nuclear de Unión a la Caperuza/metabolismo , Proteínas Nucleares/metabolismo , Interferencia de ARN , Animales , Arsénico/toxicidad , Línea Celular , Guanosina/análogos & derivados , Guanosina/metabolismo , Humanos , Ratones , MicroARNsRESUMEN
Ars2 is a component of the nuclear cap-binding complex that contributes to microRNA biogenesis and is required for cellular proliferation. Here, we expand on the repertoire of Ars2-dependent microRNAs and determine that Ars2 regulates a number of mRNAs, the largest defined subset of which code for histones. Histone mRNAs are unique among mammalian mRNAs because they are not normally polyadenylated but, rather, are cleaved following a 3' stem loop. A significant reduction in correctly processed histone mRNAs was observed following Ars2 depletion, concurrent with an increase in polyadenylated histone transcripts. Furthermore, Ars2 physically associated with histone mRNAs and the noncoding RNA 7SK. Knockdown of 7SK led to an enhanced ratio of cleaved to polyadenylated histone transcripts, an effect dependent on Ars2. Together, the data demonstrate that Ars2 contributes to histone mRNA 3' end formation and expression and these functional properties of Ars2 are negatively regulated by interaction with 7SK RNA.
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Histonas/genética , Proteínas Nucleares/fisiología , Procesamiento de Término de ARN 3' , ARN Mensajero/metabolismo , Células HeLa , Humanos , Metiltransferasas/antagonistas & inhibidores , Metiltransferasas/fisiología , MicroARNs/metabolismo , Proteínas Nucleares/antagonistas & inhibidores , Proteínas Nucleares/genética , Interferencia de ARN , ARN Interferente PequeñoRESUMEN
Cellular proliferation depends on the integration of mitogenic stimuli with environmental conditions. Increasing evidence suggests that microRNAs play a regulatory role in this integration. Here we show that during periods of cellular quiescence, mature microRNAs are stabilized and stored in Argonaute protein complexes that can be activated by mitogenic stimulation to repress mitogen-stimulated targets, thus influencing subsequent cellular responses. In quiescent cells, the majority of microRNAs exist in low molecular weight, Argonaute protein-containing complexes devoid of essential components of the RNA-induced silencing complex (RISC). For at least 3 wk, this pool of Argonaute-associated microRNAs is stable and can be recruited into RISC complexes subsequent to mitogenic stimulation. Using several model systems, we demonstrate that stable Argonaute protein-associated small RNAs are capable of repressing mitogen-induced transcripts. Therefore, mature microRNAs may represent a previously unappreciated form of cellular memory that allows cells to retain posttranscriptional regulatory information over extended periods of cellular quiescence.
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Proteínas Argonautas/metabolismo , Proliferación Celular , Regulación de la Expresión Génica/fisiología , Sustancias Macromoleculares/metabolismo , MicroARNs/metabolismo , Animales , Secuencia de Bases , Northern Blotting , Western Blotting , Cromatografía en Gel , Regulación de la Expresión Génica/genética , Secuenciación de Nucleótidos de Alto Rendimiento , Luciferasas , Ratones , Ratones Noqueados , Datos de Secuencia Molecular , Interferencia de ARN , ARN Interferente Pequeño/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
BACKGROUND: The treatment of differentiated thyroid cancer refractory to radioactive iodine (RAI) had been hampered by few effective therapies. Recently, tyrosine kinase inhibitors (TKIs) have shown activity in this disease. Clinical guidance on the use of these agents in RAI-refractory thyroid cancer is warranted. MATERIALS AND METHODS: Molecular mutations found in RAI-refractory thyroid cancer are summarized. Recent phase II and III clinical trial data for TKIs axitinib, lenvatinib, motesanib, pazopanib, sorafenib, sunitinib, and vandetinib are reviewed including efficacy and side effect profiles. Molecular targets and potencies of these agents are compared. Inhibitors of BRAF, mammalian target of rapamycin, and MEK are considered. RESULTS: Routine testing for molecular alterations prior to therapy is not yet recommended. TKIs produce progression-free survival of approximately 1 year (range: 7.7-19.6 months) and partial response rates of up to 50% by Response Evaluation Criteria in Solid Tumors. Pazopanib and lenvatinib are the most active agents. The majority of patients experienced tumor shrinkage with TKIs. Common adverse toxicities affect dermatologic, gastrointestinal, and cardiovascular systems. CONCLUSION: Multiple TKIs have activity in RAI-refractory differentiated thyroid cancer. Selection of a targeted agent should depend on disease trajectory, side effect profile, and goals of therapy.
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Inhibidores de Proteínas Quinasas/uso terapéutico , Neoplasias de la Tiroides/tratamiento farmacológico , Neoplasias de la Tiroides/genética , Supervivencia sin Enfermedad , Humanos , Indoles/uso terapéutico , Radioisótopos de Yodo/efectos adversos , Quinasa 1 de Quinasa de Quinasa MAP/genética , Niacinamida/análogos & derivados , Niacinamida/genética , Niacinamida/uso terapéutico , Oligonucleótidos , Compuestos de Fenilurea/uso terapéutico , Proteínas Proto-Oncogénicas B-raf/genética , Sorafenib , Neoplasias de la Tiroides/patologíaRESUMEN
Citrate is a critical metabolite required to support both mitochondrial bioenergetics and cytosolic macromolecular synthesis. When cells proliferate under normoxic conditions, glucose provides the acetyl-CoA that condenses with oxaloacetate to support citrate production. Tricarboxylic acid (TCA) cycle anaplerosis is maintained primarily by glutamine. Here we report that some hypoxic cells are able to maintain cell proliferation despite a profound reduction in glucose-dependent citrate production. In these hypoxic cells, glutamine becomes a major source of citrate. Glutamine-derived α-ketoglutarate is reductively carboxylated by the NADPH-linked mitochondrial isocitrate dehydrogenase (IDH2) to form isocitrate, which can then be isomerized to citrate. The increased IDH2-dependent carboxylation of glutamine-derived α-ketoglutarate in hypoxia is associated with a concomitant increased synthesis of 2-hydroxyglutarate (2HG) in cells with wild-type IDH1 and IDH2. When either starved of glutamine or rendered IDH2-deficient by RNAi, hypoxic cells are unable to proliferate. The reductive carboxylation of glutamine is part of the metabolic reprogramming associated with hypoxia-inducible factor 1 (HIF1), as constitutive activation of HIF1 recapitulates the preferential reductive metabolism of glutamine-derived α-ketoglutarate even in normoxic conditions. These data support a role for glutamine carboxylation in maintaining citrate synthesis and cell growth under hypoxic conditions.
Asunto(s)
Proliferación Celular , Citratos/metabolismo , Isocitrato Deshidrogenasa/metabolismo , Ácidos Cetoglutáricos/metabolismo , Ácidos Carboxílicos/metabolismo , Hipoxia de la Célula , Línea Celular Tumoral , Supervivencia Celular , Ciclo del Ácido Cítrico , Cromatografía de Gases y Espectrometría de Masas , Glucosa/metabolismo , Glutamina/metabolismo , Humanos , Factor 1 Inducible por Hipoxia/genética , Factor 1 Inducible por Hipoxia/metabolismo , Immunoblotting , Isocitrato Deshidrogenasa/genética , Neoplasias/genética , Neoplasias/metabolismo , Neoplasias/patología , Oxidación-Reducción , Interferencia de ARNRESUMEN
HAT1, also known as Histone acetyltransferase 1, plays a crucial role in chromatin synthesis by stabilizing and acetylating nascent H4 before nucleosome assembly. It is required for tumor growth in various systems, making it a potential target for cancer treatment. To facilitate the identification of compounds that can inhibit HAT1 enzymatic activity, we have devised an acetyl-click assay for rapid screening. In this simple assay, we employ recombinant HAT1/Rbap46, which is purified from activated human cells. The method utilizes the acetyl-CoA analog 4-pentynoyl-CoA (4P) in a click-chemistry approach. This involves the enzymatic transfer of an alkyne handle through a HAT1-dependent acylation reaction to a biotinylated H4 N-terminal peptide. The captured peptide is then immobilized on neutravidin plates, followed by click-chemistry functionalization with biotin-azide. Subsequently, streptavidin-peroxidase recruitment is employed to oxidize amplex red, resulting in a quantitative fluorescent output. By introducing chemical inhibitors during the acylation reaction, we can quantify enzymatic inhibition based on a reduction of the fluorescence signal. Importantly, this reaction is scalable, allowing for high throughput screening of potential inhibitors for HAT1 enzymatic activity.
Asunto(s)
Química Clic , Histonas , Humanos , Histonas/metabolismo , Acetilación , Histona Acetiltransferasas/metabolismo , PéptidosRESUMEN
The short-chain fatty acids (SCFA) propionate and butyrate are produced in large amounts by microbial metabolism and have been identified as unique acyl lysine histone marks. In order to better understand the function of these modifications we used ChIP-seq to map the genome-wide location of four short-chain acyl histone marks H3K18pr/bu and H4K12pr/bu in treated and untreated colorectal cancer (CRC) and normal cells, as well as in mouse intestines in vivo . We correlate these marks with open chromatin regions along with gene expression to access the function of the target regions. Our data demonstrate that propionate and butyrate act as promoters of growth, differentiation as well as ion transport. We propose a mechanism involving direct modification of specific genomic regions, resulting in increased chromatin accessibility, and in case of butyrate, opposing effects on the proliferation of normal versus CRC cells.
RESUMEN
The short-chain fatty acids (SCFA) propionate and butyrate are produced in large amounts by microbial metabolism and have been identified as unique acyl lysine histone marks. In order to better understand the function of these modifications we used ChIP-seq to map the genome-wide location of four short-chain acyl histone marks H3K18pr/bu and H4K12pr/bu in treated and untreated colorectal cancer (CRC) and normal cells, as well as in mouse intestines in vivo. We correlate these marks with open chromatin regions along with gene expression to access the function of the target regions. Our data demonstrate that propionate and butyrate act as promoters of growth, differentiation as well as ion transport. We propose a mechanism involving direct modification of specific genomic regions, resulting in increased chromatin accessibility, and in case of butyrate, opposing effects on the proliferation of normal versus CRC cells.
RESUMEN
The functions of non-coding regulatory elements (NCREs), which constitute a major fraction of the human genome, have not been systematically studied. Here we report a method involving libraries of paired single-guide RNAs targeting both ends of an NCRE as a screening system for the Cas9-mediated deletion of thousands of NCREs genome-wide to study their functions in distinct biological contexts. By using K562 and 293T cell lines and human embryonic stem cells, we show that NCREs can have redundant functions, and that many ultra-conserved elements have silencer activity and play essential roles in cell growth and in cellular responses to drugs (notably, the ultra-conserved element PAX6_Tarzan may be critical for heart development, as removing it from human embryonic stem cells led to defects in cardiomyocyte differentiation). The high-throughput screen, which is compatible with single-cell sequencing, may allow for the identification of druggable NCREs.
Asunto(s)
Sistemas CRISPR-Cas , ARN Guía de Sistemas CRISPR-Cas , Humanos , ARN Guía de Sistemas CRISPR-Cas/genética , Células K562 , Sistemas CRISPR-Cas/genética , Células HEK293 , Genoma Humano/genética , Diferenciación Celular/genética , Miocitos Cardíacos/metabolismo , ARN no Traducido/genética , Células Madre Embrionarias Humanas/metabolismo , Factor de Transcripción PAX6/genética , Factor de Transcripción PAX6/metabolismo , Biblioteca de GenesRESUMEN
HAT1 is a central regulator of chromatin synthesis that acetylates nascent histone H4. To ascertain whether targeting HAT1 is a viable anticancer treatment strategy, we sought to identify small-molecule inhibitors of HAT1 by developing a high-throughput HAT1 acetyl-click assay. Screening of small-molecule libraries led to the discovery of multiple riboflavin analogs that inhibited HAT1 enzymatic activity. Compounds were refined by synthesis and testing of over 70 analogs, which yielded structure-activity relationships. The isoalloxazine core was required for enzymatic inhibition, whereas modifications of the ribityl side chain improved enzymatic potency and cellular growth suppression. One compound (JG-2016 [24a]) showed relative specificity toward HAT1 compared to other acetyltransferases, suppressed the growth of human cancer cell lines, impaired enzymatic activity in cellulo, and interfered with tumor growth. This is the first report of a small-molecule inhibitor of the HAT1 enzyme complex and represents a step toward targeting this pathway for cancer therapy.
Asunto(s)
Histonas , Neoplasias , Humanos , Histonas/metabolismo , Histona Acetiltransferasas/metabolismo , Cromatina , Línea Celular , AcetilaciónRESUMEN
The Warburg effect is the major metabolic hallmark of cancer. According to Warburg himself, the consequence of the Warburg effect is cell dedifferentiation. Therefore, reversing the Warburg effect might be an approach to restore cell differentiation in cancer. In this study, we used a mitochondrial uncoupler, niclosamide ethanolamine (NEN), to activate mitochondrial respiration, which induced neural differentiation in neuroblastoma cells. NEN treatment increased the NAD+/NADH and pyruvate/lactate ratios and also the α-ketoglutarate/2-hydroxyglutarate (2-HG) ratio. Consequently, NEN treatment induced promoter CpG island demethylation and epigenetic landscape remodeling, activating the neural differentiation program. In addition, NEN treatment upregulated p53 but downregulated N-Myc and ß-catenin signaling in neuroblastoma cells. Importantly, even under hypoxia, NEN treatment remained effective in inhibiting 2-HG generation, promoting DNA demethylation, and suppressing hypoxia-inducible factor signaling. Dietary NEN intervention reduced tumor growth rate, 2-HG levels, and expression of N-Myc and ß-catenin in tumors in an orthotopic neuroblastoma mouse model. Integrative analysis indicated that NEN treatment upregulated favorable prognosis genes and downregulated unfavorable prognosis genes, which were defined using multiple neuroblastoma patient datasets. Altogether, these results suggest that mitochondrial uncoupling is an effective metabolic and epigenetic therapy for reversing the Warburg effect and inducing differentiation in neuroblastoma. SIGNIFICANCE: Targeting cancer metabolism using the mitochondrial uncoupler niclosamide ethanolamine leads to methylome reprogramming and differentiation in neuroblastoma, providing a therapeutic opportunity to reverse the Warburg effect and suppress tumor growth. See related commentary by Byrne and Bell, p.167.
Asunto(s)
Diferenciación Celular , Epigenoma , Neuroblastoma , Efecto Warburg en Oncología , Animales , Ratones , beta Catenina/genética , Diferenciación Celular/genética , Línea Celular Tumoral , Epigenoma/genética , Epigenoma/fisiología , Etanolamina/farmacología , Etanolamina/uso terapéutico , Etanolaminas/uso terapéutico , Hipoxia/tratamiento farmacológico , Neuroblastoma/genética , Neuroblastoma/patología , Niclosamida/farmacología , Efecto Warburg en Oncología/efectos de los fármacos , Mitocondrias/efectos de los fármacos , Mitocondrias/fisiologíaRESUMEN
Tumor cells are metabolically reprogrammed to fuel cell proliferation. Most transformed cells take up high levels of glucose and produce ATP through aerobic glycolysis. In cells exhibiting aerobic glycolysis, a significant fraction of glucose carbon is also directed into de novo lipogenesis and nucleotide biosynthesis. The glucose-responsive transcription factor carbohydrate responsive element binding protein (ChREBP) was previously shown to be important for redirecting glucose metabolism in support of lipogenesis in nonproliferating hepatocytes. However, whether it plays a more generalized role in reprogramming metabolism during cell proliferation has not been examined. Here, we demonstrated that the expression of ChREBP can be induced in response to mitogenic stimulation and that the induction of ChREBP is required for efficient cell proliferation. Suppression of ChREBP resulted in diminished aerobic glycolysis, de novo lipogenesis, and nucleotide biosynthesis, but stimulated mitochondrial respiration, suggesting a metabolic switch from aerobic glycolysis to oxidative phosphorylation. Cells in which ChREBP was suppressed by RNAi exhibited p53 activation and cell cycle arrest. In vivo, suppression of ChREBP led to a p53-dependent reduction in tumor growth. These results demonstrate that ChREBP plays a key role both in redirecting glucose metabolism to anabolic pathways and suppressing p53 activity.
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Anabolizantes/metabolismo , Factores de Transcripción Básicos con Cremalleras de Leucinas y Motivos Hélice-Asa-Hélice/fisiología , Proliferación Celular , Glucosa/farmacología , Animales , Secuencia de Bases , Ciclo Celular , Línea Celular Tumoral , Neoplasias Colorrectales/metabolismo , Neoplasias Colorrectales/patología , Humanos , Masculino , Ratones , Ratones Desnudos , ARN Interferente Pequeño , Trasplante Heterólogo , Proteína p53 Supresora de Tumor/metabolismoRESUMEN
Talazoparib, a PARP inhibitor, is active in germline BRCA1 and BRCA2 (gBRCA1/2)-mutant advanced breast cancer, but its activity beyond gBRCA1/2 is poorly understood. We conducted Talazoparib Beyond BRCA ( NCT02401347 ), an open-label phase II trial, to evaluate talazoparib in patients with pretreated advanced HER2-negative breast cancer (n = 13) or other solid tumors (n = 7) with mutations in homologous recombination (HR) pathway genes other than BRCA1 and BRCA2. In patients with breast cancer, four patients had a Response Evaluation Criteria in Solid Tumors (RECIST) partial response (overall response rate, 31%), and three additional patients had stable disease of ≥6 months (clinical benefit rate, 54%). All patients with germline mutations in PALB2 (gPALB2; encoding partner and localizer of BRCA2) had treatment-associated tumor regression. Tumor or plasma circulating tumor DNA (ctDNA) HR deficiency (HRD) scores were correlated with treatment outcomes and were increased in all gPALB2 tumors. In addition, a gPALB2-associated mutational signature was associated with tumor response. Thus, talazoparib has been demonstrated to have efficacy in patients with advanced breast cancer who have gPALB2 mutations, showing activity in the context of HR pathway gene mutations beyond gBRCA1/2.
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Neoplasias de la Mama , ADN Tumoral Circulante , Humanos , Femenino , Inhibidores de Poli(ADP-Ribosa) Polimerasas/uso terapéutico , Recombinación Homóloga , Neoplasias de la Mama/tratamiento farmacológico , Mutación , Proteína BRCA1/genética , Proteína BRCA2/genéticaRESUMEN
BACKGROUND: Despite significant progress in cancer immunotherapy in recent years, resistance to existing immune checkpoint therapies (ICT) is common. V-domain Ig suppressor of T cell activation (VISTA), a predominantly myeloid immune checkpoint regulator, represents a promising therapeutic target due to its role in suppressing proinflammatory antitumor responses in myeloid-enriched tumor microenvironments. However, uncertainty around the cognate VISTA ligand has made the development of effective anti-VISTA antibodies challenging. The expression of VISTA on normal immune cell subtypes argues for a neutralizing non-depleting antibody, however, previous reported anti-VISTA antibodies use IgG1 Fc isotypes that deplete VISTA+ cells by antibody dependent cellular cytotoxicity/complement dependent cytotoxicity and these antibodies have shown fast serum clearance and immune toxicities. METHOD: Here we used a rational antibody discovery approach to develop the first Fc-independent anti-VISTA antibody, HMBD-002, that binds a computationally predicted functional epitope within the C-C-loop, distinct from other known anti-VISTA antibodies. This epitope is species-conserved allowing robust in vitro and in vivo testing of HMBD-002 in human and murine models of immune activation and cancer including humanized mouse models. RESULTS: We demonstrate here that blockade by HMBD-002 inhibits VISTA binding to potential partners, including V-Set and Immunoglobulin domain containing 3, to reduce myeloid-derived suppression of T cell activity and prevent neutrophil migration. Analysis of immune cell milieu suggests that HMBD-002 treatment stimulates a proinflammatory phenotype characterized by a Th1/Th17 response, recapitulating a phenotype previously noted in VISTA knockout models. This mechanism of action is further supported by immune-competent syngenic and humanized mouse models of colorectal, breast and lung cancer where neutralizing VISTA, without depleting VISTA expressing cells, significantly inhibited tumor growth while decreasing infiltration of suppressive myeloid cells and increasing T cell activity. Finally, we did not observe either the fast serum clearance or immune toxicities that have been reported for IgG1 antibodies. CONCLUSION: In conclusion, we have shown that VISTA-induced immune suppression can be reversed by blockade of the functional C-C' loop region of VISTA with a first-in-class rationally targeted and non-depleting IgG4 isotype anti-VISTA antibody, HMBD-002. This antibody represents a highly promising novel therapy in the VISTA-suppressed ICT non-responder population.
Asunto(s)
Terapia de Inmunosupresión/métodos , Activación de Linfocitos/inmunología , Neoplasias/inmunología , Receptores Fc/metabolismo , Animales , Modelos Animales de Enfermedad , Humanos , Ratones , Microambiente TumoralRESUMEN
Although alterations in chromatin structure are known to exist in tumors, how these alterations relate to molecular phenotypes in cancer remains to be demonstrated. Multi-omics profiling of human tumors can provide insight into how alterations in chromatin structure are propagated through the pathway of gene expression to result in malignant protein expression. We applied multi-omics profiling of chromatin accessibility, RNA abundance, and protein abundance to 36 human thyroid cancer primary tumors, metastases, and patient-match normal tissue. Through quantification of chromatin accessibility associated with active transcription units and global protein expression, we identify a local chromatin structure that is highly correlated with coordinated RNA and protein expression. In particular, we identify enhancers located within gene-bodies as predictive of correlated RNA and protein expression, that is independent of overall transcriptional activity. To demonstrate the generalizability of these findings we also identify similar results in an independent cohort of human breast cancers. Taken together, these analyses suggest that local enhancers, rather than distal enhancers, are likely most predictive of cancer gene expression phenotypes. This allows for identification of potential targets for cancer therapeutic approaches and reinforces the utility of multi-omics profiling as a methodology to understand human disease.
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Neoplasias de la Mama/genética , Cromatina/metabolismo , Regulación Neoplásica de la Expresión Génica , Cáncer Papilar Tiroideo/genética , Neoplasias de la Tiroides/genética , Neoplasias de la Mama/patología , Secuenciación de Inmunoprecipitación de Cromatina , Estudios de Cohortes , Conjuntos de Datos como Asunto , Elementos de Facilitación Genéticos , Epigénesis Genética , Femenino , Redes Reguladoras de Genes , Humanos , Masculino , Regiones Promotoras Genéticas , Proteómica , ARN/metabolismo , RNA-Seq , Cáncer Papilar Tiroideo/patología , Cáncer Papilar Tiroideo/cirugía , Glándula Tiroides/patología , Glándula Tiroides/cirugía , Neoplasias de la Tiroides/patología , Neoplasias de la Tiroides/cirugía , Tiroidectomía , Factores de Transcripción/metabolismoRESUMEN
Despite the fact that Otto H. Warburg discovered the Warburg effect almost one hundred years ago, why cancer cells waste most of the glucose carbon as lactate remains an enigma. Warburg proposed a connection between the Warburg effect and cell dedifferentiation. Hypoxia is a common tumor microenvironmental stress that induces the Warburg effect and blocks tumor cell differentiation. The underlying mechanism by which this occurs is poorly understood, and no effective therapeutic strategy has been developed to overcome this resistance to differentiation. Using a neuroblastoma differentiation model, we discovered that hypoxia repressed cell differentiation through reducing cellular acetyl-CoA levels, leading to reduction of global histone acetylation and chromatin accessibility. The metabolic switch triggering this global histone hypoacetylation was the induction of pyruvate dehydrogenase kinases (PDK1 and PDK3). Inhibition of PDKs using dichloroacetate (DCA) restored acetyl-CoA generation and histone acetylation under hypoxia. Knocking down PDK1 induced neuroblastoma cell differentiation, highlighting the critical role of PDK1 in cell fate control. Importantly, acetate or glycerol triacetate (GTA) supplementation restored differentiation markers expression and neuron differentiation under hypoxia. Moreover, ATAC-Seq analysis demonstrated that hypoxia treatment significantly reduced chromatin accessibility at RAR/RXR binding sites, which can be restored by acetate supplementation. In addition, hypoxia-induced histone hypermethylation by increasing 2-hydroxyglutarate (2HG) and reducing α-ketoglutarate (αKG). αKG supplementation reduced histone hypermethylation upon hypoxia, but did not restore histone acetylation or differentiation markers expression. Together, these findings suggest that diverting pyruvate flux away from acetyl-CoA generation to lactate production is the key mechanism that Warburg effect drives dedifferentiation and tumorigenesis. We propose that combining differentiation therapy with acetate/GTA supplementation might represent an effective therapy against neuroblastoma.
Asunto(s)
Acetatos/farmacología , Antineoplásicos/farmacología , Ensamble y Desensamble de Cromatina/efectos de los fármacos , Neuroblastoma/tratamiento farmacológico , Neurogénesis/efectos de los fármacos , Efecto Warburg en Oncología/efectos de los fármacos , Acetilación , Animales , Línea Celular Tumoral , Regulación Neoplásica de la Expresión Génica , Histonas/metabolismo , Humanos , Masculino , Ratones , Neuroblastoma/genética , Neuroblastoma/metabolismo , Neuroblastoma/patología , Proyección Neuronal/efectos de los fármacos , Piruvato Deshidrogenasa Quinasa Acetil-Transferidora/genética , Piruvato Deshidrogenasa Quinasa Acetil-Transferidora/metabolismo , Transducción de Señal , Carga Tumoral/efectos de los fármacos , Hipoxia Tumoral , Microambiente Tumoral , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
PURPOSE: Patients with triple-negative breast cancer (TNBC) with homologous recombination deficient tumors achieve significantly higher pathologic complete response (pCR) rates when treated with neoadjuvant platinum-based therapy. Tumor-infiltrating lymphocytes (TIL) are prognostic and predictive of chemotherapy benefit in early stage TNBC. The relationship between TILs, BRCA1/2 mutation status, and homologous recombination deficiency (HRD) status in TNBC remains unclear. EXPERIMENTAL DESIGN: We performed a pooled analysis of five phase II studies that included patients with TNBC treated with neoadjuvant platinum-based chemotherapy to evaluate the association of TILs with HRD status (Myriad Genetics) and tumor BRCA1/2 mutation status. Furthermore, the relationship between pathologic response assessed using the residual cancer burden (RCB) index and HRD status with adjustment for TILs was evaluated. RESULTS: Among 161 patients, stromal TIL (sTIL) density was not significantly associated with HRD status (P = 0.107) or tumor BRCA1/2 mutation status (P = 0.391). In multivariate analyses, sTIL density [OR, 1.23; 95% confidence interval (CI), 0.94-1.61; P = 0.139] was not associated with pCR, but was associated with RCB 0/I status (OR 1.62; 95% CI, 1.20-2.28; P = 0.001). HRD was significantly associated with both pCR (OR 12.09; 95% CI, 4.11-44.29; P = 7.82 × 10-7) and RCB 0/I (OR 10.22; 95% CI, 4.11-28.75; P = 1.09 × 10-7) in these models. CONCLUSIONS: In patients with TNBC treated with neoadjuvant platinum-based therapy, TIL density was not significantly associated with either tumor BRCA1/2 mutation status or HRD status. In this pooled analysis, HRD and sTIL density were independently associated with treatment response, with HRD status being the strongest predictor.