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1.
Cell ; 175(1): 101-116.e25, 2018 09 20.
Artigo em Inglês | MEDLINE | ID: mdl-30220459

RESUMO

IDH1 mutations are common in low-grade gliomas and secondary glioblastomas and cause overproduction of (R)-2HG. (R)-2HG modulates the activity of many enzymes, including some that are linked to transformation and some that are probably bystanders. Although prior work on (R)-2HG targets focused on 2OG-dependent dioxygenases, we found that (R)-2HG potently inhibits the 2OG-dependent transaminases BCAT1 and BCAT2, likely as a bystander effect, thereby decreasing glutamate levels and increasing dependence on glutaminase for the biosynthesis of glutamate and one of its products, glutathione. Inhibiting glutaminase specifically sensitized IDH mutant glioma cells to oxidative stress in vitro and to radiation in vitro and in vivo. These findings highlight the complementary roles for BCATs and glutaminase in glutamate biosynthesis, explain the sensitivity of IDH mutant cells to glutaminase inhibitors, and suggest a strategy for maximizing the effectiveness of such inhibitors against IDH mutant gliomas.


Assuntos
Glioma/metabolismo , Ácido Glutâmico/biossíntese , Transaminases/fisiologia , Linhagem Celular Tumoral , Glioma/fisiopatologia , Ácido Glutâmico/efeitos dos fármacos , Glutaratos/metabolismo , Glutaratos/farmacologia , Homeostase/efeitos dos fármacos , Humanos , Isocitrato Desidrogenase/genética , Isocitrato Desidrogenase/fisiologia , Antígenos de Histocompatibilidade Menor/genética , Antígenos de Histocompatibilidade Menor/fisiologia , Mutação , Oxirredução/efeitos dos fármacos , Proteínas da Gravidez/genética , Proteínas da Gravidez/fisiologia , Transaminases/antagonistas & inibidores , Transaminases/genética
2.
Cell ; 164(5): 884-95, 2016 Feb 25.
Artigo em Inglês | MEDLINE | ID: mdl-26919427

RESUMO

Ischemic preconditioning is the phenomenon whereby brief periods of sublethal ischemia protect against a subsequent, more prolonged, ischemic insult. In remote ischemic preconditioning (RIPC), ischemia to one organ protects others organs at a distance. We created mouse models to ask if inhibition of the alpha-ketoglutarate (αKG)-dependent dioxygenase Egln1, which senses oxygen and regulates the hypoxia-inducible factor (HIF) transcription factor, could suffice to mediate local and remote ischemic preconditioning. Using somatic gene deletion and a pharmacological inhibitor, we found that inhibiting Egln1 systemically or in skeletal muscles protects mice against myocardial ischemia-reperfusion (I/R) injury. Parabiosis experiments confirmed that RIPC in this latter model was mediated by a secreted factor. Egln1 loss causes accumulation of circulating αKG, which drives hepatic production and secretion of kynurenic acid (KYNA) that is necessary and sufficient to mediate cardiac ischemic protection in this setting.


Assuntos
Prolina Dioxigenases do Fator Induzível por Hipóxia/antagonistas & inibidores , Precondicionamento Isquêmico , Ácidos Cetoglutáricos/metabolismo , Animais , Isquemia/prevenção & controle , Ácido Cinurênico/metabolismo , Fígado/metabolismo , Camundongos , Modelos Animais , Traumatismo por Reperfusão Miocárdica/prevenção & controle , Parabiose
3.
Genes Dev ; 33(23-24): 1718-1738, 2019 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-31727771

RESUMO

More than 90% of small cell lung cancers (SCLCs) harbor loss-of-function mutations in the tumor suppressor gene RB1 The canonical function of the RB1 gene product, pRB, is to repress the E2F transcription factor family, but pRB also functions to regulate cellular differentiation in part through its binding to the histone demethylase KDM5A (also known as RBP2 or JARID1A). We show that KDM5A promotes SCLC proliferation and SCLC's neuroendocrine differentiation phenotype in part by sustaining expression of the neuroendocrine transcription factor ASCL1. Mechanistically, we found that KDM5A sustains ASCL1 levels and neuroendocrine differentiation by repressing NOTCH2 and NOTCH target genes. To test the role of KDM5A in SCLC tumorigenesis in vivo, we developed a CRISPR/Cas9-based mouse model of SCLC by delivering an adenovirus (or an adeno-associated virus [AAV]) that expresses Cre recombinase and sgRNAs targeting Rb1, Tp53, and Rbl2 into the lungs of Lox-Stop-Lox Cas9 mice. Coinclusion of a KDM5A sgRNA decreased SCLC tumorigenesis and metastasis, and the SCLCs that formed despite the absence of KDM5A had higher NOTCH activity compared to KDM5A+/+ SCLCs. This work establishes a role for KDM5A in SCLC tumorigenesis and suggests that KDM5 inhibitors should be explored as treatments for SCLC.


Assuntos
Diferenciação Celular/genética , Células Neuroendócrinas/citologia , Receptores Notch/fisiologia , Proteína 2 de Ligação ao Retinoblastoma/metabolismo , Transdução de Sinais/genética , Carcinoma de Pequenas Células do Pulmão/enzimologia , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos , Linhagem Celular , Transformação Celular Neoplásica/genética , Modelos Animais de Doenças , Regulação Neoplásica da Expressão Gênica/genética , Histona Desmetilases/metabolismo , Humanos , Técnicas In Vitro , Camundongos , Células Neuroendócrinas/patologia , Carcinoma de Pequenas Células do Pulmão/fisiopatologia
5.
Proc Natl Acad Sci U S A ; 119(21): e2114324119, 2022 05 24.
Artigo em Inglês | MEDLINE | ID: mdl-35584120

RESUMO

Antiandrogen strategies remain the prostate cancer treatment backbone, but drug resistance develops. We show that androgen blockade in prostate cancer leads to derepression of retroelements (REs) followed by a double-stranded RNA (dsRNA)-stimulated interferon response that blocks tumor growth. A forward genetic approach identified H3K9 trimethylation (H3K9me3) as an essential epigenetic adaptation to antiandrogens, which enabled transcriptional silencing of REs that otherwise stimulate interferon signaling and glucocorticoid receptor expression. Elevated expression of terminal H3K9me3 writers was associated with poor patient hormonal therapy outcomes. Forced expression of H3K9me3 writers conferred resistance, whereas inhibiting H3K9-trimethylation writers and readers restored RE expression, blocking antiandrogen resistance. Our work reveals a drug resistance axis that integrates multiple cellular signaling elements and identifies potential pharmacologic vulnerabilities.


Assuntos
Antagonistas de Receptores de Andrógenos , Neoplasias de Próstata Resistentes à Castração , Antagonistas de Androgênios/farmacologia , Antagonistas de Androgênios/uso terapêutico , Antagonistas de Receptores de Andrógenos/farmacologia , Androgênios/farmacologia , Metilação de DNA , Resistencia a Medicamentos Antineoplásicos , Inativação Gênica , Humanos , Interferons , Masculino , Metilação , Nitrilas/uso terapêutico , Neoplasias de Próstata Resistentes à Castração/patologia , Receptores Androgênicos/genética , Receptores Androgênicos/metabolismo
6.
J Physiol ; 601(3): 567-606, 2023 02.
Artigo em Inglês | MEDLINE | ID: mdl-36533558

RESUMO

Nocturnal hypoxaemia, which is common in chronic obstructive pulmonary disease (COPD) patients, is associated with skeletal muscle loss or sarcopenia, which contributes to adverse clinical outcomes. In COPD, we have defined this as prolonged intermittent hypoxia (PIH) because the duration of hypoxia in skeletal muscle occurs through the duration of sleep followed by normoxia during the day, in contrast to recurrent brief hypoxic episodes during obstructive sleep apnoea (OSA). Adaptive cellular responses to PIH are not known. Responses to PIH induced by three cycles of 8 h hypoxia followed by 16 h normoxia were compared to those during chronic hypoxia (CH) or normoxia for 72 h in murine C2C12 and human inducible pluripotent stem cell-derived differentiated myotubes. RNA sequencing followed by downstream analyses were complemented by experimental validation of responses that included both unique and shared perturbations in ribosomal and mitochondrial function during PIH and CH. A sarcopenic phenotype characterized by decreased myotube diameter and protein synthesis, and increased phosphorylation of eIF2α (Ser51) by eIF2α kinase, and of GCN-2 (general controlled non-derepressed-2), occurred during both PIH and CH. Mitochondrial oxidative dysfunction, disrupted supercomplex assembly, lower activity of Complexes I, III, IV and V, and reduced intermediary metabolite concentrations occurred during PIH and CH. Decreased mitochondrial fission occurred during CH. Physiological relevance was established in skeletal muscle of mice with COPD that had increased phosphorylation of eIF2α, lower protein synthesis and mitochondrial oxidative dysfunction. Molecular and metabolic responses with PIH suggest an adaptive exhaustion with failure to restore homeostasis during normoxia. KEY POINTS: Sarcopenia or skeletal muscle loss is one of the most frequent complications that contributes to mortality and morbidity in patients with chronic obstructive pulmonary disease (COPD). Unlike chronic hypoxia, prolonged intermittent hypoxia is a frequent, underappreciated and clinically relevant model of hypoxia in patients with COPD. We developed a novel, in vitro myotube model of prolonged intermittent hypoxia with molecular and metabolic perturbations, mitochondrial oxidative dysfunction, and consequent sarcopenic phenotype. In vivo studies in skeletal muscle from a mouse model of COPD shared responses with our myotube model, establishing the pathophysiological relevance of our studies. These data lay the foundation for translational studies in human COPD to target prolonged, nocturnal hypoxaemia to prevent sarcopenia in these patients.


Assuntos
Doença Pulmonar Obstrutiva Crônica , Sarcopenia , Humanos , Camundongos , Animais , Sarcopenia/metabolismo , Proteostase , Músculo Esquelético/metabolismo , Hipóxia/metabolismo , Doença Pulmonar Obstrutiva Crônica/complicações
7.
Nature ; 539(7627): 107-111, 2016 11 03.
Artigo em Inglês | MEDLINE | ID: mdl-27595393

RESUMO

Clear cell renal cell carcinoma, the most common form of kidney cancer, is usually linked to inactivation of the pVHL tumour suppressor protein and consequent accumulation of the HIF-2α transcription factor (also known as EPAS1). Here we show that a small molecule (PT2399) that directly inhibits HIF-2α causes tumour regression in preclinical mouse models of primary and metastatic pVHL-defective clear cell renal cell carcinoma in an on-target fashion. pVHL-defective clear cell renal cell carcinoma cell lines display unexpectedly variable sensitivity to PT2399, however, suggesting the need for predictive biomarkers to be developed to use this approach optimally in the clinic.


Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/antagonistas & inibidores , Carcinoma de Células Renais/tratamento farmacológico , Carcinoma de Células Renais/patologia , Indanos/farmacologia , Indanos/uso terapêutico , Neoplasias Renais/tratamento farmacológico , Neoplasias Renais/patologia , Sulfonas/farmacologia , Sulfonas/uso terapêutico , Animais , Biomarcadores Farmacológicos , Carcinoma de Células Renais/genética , Carcinoma de Células Renais/metabolismo , Linhagem Celular Tumoral , Modelos Animais de Doenças , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Neoplasias Renais/genética , Neoplasias Renais/metabolismo , Camundongos , Modelos Biológicos , Metástase Neoplásica/tratamento farmacológico , Metástase Neoplásica/patologia , Transcrição Gênica/efeitos dos fármacos , Proteína Supressora de Tumor Von Hippel-Lindau/genética , Proteína Supressora de Tumor Von Hippel-Lindau/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
8.
Genes Dev ; 28(13): 1429-44, 2014 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-24990963

RESUMO

The three EglN prolyl hydroxylases (EglN1, EglN2, and EglN3) regulate the stability of the HIF transcription factor. We recently showed that loss of EglN2, however, also leads to down-regulation of Cyclin D1 and decreased cell proliferation in a HIF-independent manner. Here we report that EglN2 can hydroxylate FOXO3a on two specific prolyl residues in vitro and in vivo. Hydroxylation of these sites prevents the binding of USP9x deubiquitinase, thereby promoting the proteasomal degradation of FOXO3a. FOXO transcription factors can repress Cyclin D1 transcription. Failure to hydroxylate FOXO3a promotes its accumulation in cells, which in turn suppresses Cyclin D1 expression. These findings provide new insights into post-transcriptional control of FOXO3a and provide a new avenue for pharmacologically altering Cyclin D1 activity.


Assuntos
Fatores de Transcrição Forkhead/metabolismo , Regulação Neoplásica da Expressão Gênica , Prolina Dioxigenases do Fator Induzível por Hipóxia/metabolismo , Ubiquitina Tiolesterase/metabolismo , Animais , Linhagem Celular , Células Cultivadas , Ciclina D1/genética , Proteína Forkhead Box O3 , Hidroxilação , Células MCF-7 , Camundongos , Ligação Proteica , Estabilidade Proteica
9.
Semin Cancer Biol ; 67(Pt 2): 34-42, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-32209418

RESUMO

Inactivation of the von Hippel Lindau tumor suppressor protein (pVHL) is a hallmark of clear cell Renal Cell Carcinoma (ccRCC), which is the most common form of kidney cancer in adults. In complex with Elongin B/C, pVHL functions as the substrate recognition subunit of a ubiquitin ligase, perhaps best known to target the hypoxia inducible factor (HIF) transcription factor for ubiquitin-dependent proteolysis. Beyond kidney cancer, the pseudo-hypoxic state caused due to chronic HIF activation in pVHL-deficient cells has become a biological model to study hypoxia's profound effects on tumor angiogenesis, metabolism, and epigenetics. However, a number of HIF-independent substrates of pVHL, which function in a broad range of biological pathways, have also been discovered. Independently, the development of high-throughput chemical and genetic screening strategies have enabled the identification of novel, HIF-independent, targetable dependencies in ccRCC. In this review we summarize the history of pVHL and HIF mediated oxygen sensing, discuss the current status of this field, and identify critical challenges that need to be overcome. The confluence of historical discovery, development of unbiased screening strategies, and the evolution of medicinal chemistry has allowed us to begin therapeutically targeting vulnerabilities that emerge due to pVHL loss in ccRCC. Ongoing mechanistic studies on the biological consequences of pVHL loss, therefore, are likely to become the cornerstones of modern therapeutics in renal cancer.


Assuntos
Neoplasias Renais/metabolismo , Terapia de Alvo Molecular/métodos , Proteína Supressora de Tumor Von Hippel-Lindau/genética , Proteína Supressora de Tumor Von Hippel-Lindau/metabolismo , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Epigênese Genética , Humanos , Neoplasias Renais/tratamento farmacológico , Neoplasias Renais/genética , Neoplasias Experimentais/genética , Neoplasias Experimentais/patologia , Oxigênio/metabolismo , Doença de von Hippel-Lindau/etiologia , Doença de von Hippel-Lindau/genética
10.
Cell Rep ; 43(1): 113575, 2024 01 23.
Artigo em Inglês | MEDLINE | ID: mdl-38181788

RESUMO

Progression of prostate cancer depends on androgen receptor, which is usually activated by androgens. Therefore, a mainstay treatment is androgen deprivation therapy. Unfortunately, despite initial treatment response, resistance nearly always develops, and disease progresses to castration-resistant prostate cancer (CRPC), which remains driven by non-gonadal androgens synthesized in prostate cancer tissues. 3ß-Hydroxysteroid dehydrogenase/Δ5-->4 isomerase 1 (3ßHSD1) catalyzes the rate-limiting step in androgen synthesis. However, how 3ßHSD1, especially the "adrenal-permissive" 3ßHSD1(367T) that permits tumor synthesis of androgen from dehydroepiandrosterone (DHEA), is regulated at the protein level is not well understood. Here, we investigate how hypoxia regulates 3ßHSD1(367T) protein levels. Our results show that, in vitro, hypoxia stabilizes 3ßHSD1 protein by suppressing autophagy. Autophagy inhibition promotes 3ßHSD1-dependent tumor progression. Hypoxia represses transcription of autophagy-related (ATG) genes by decreasing histone acetylation. Inhibiting deacetylase (HDAC) restores ATG gene transcription under hypoxia. Therefore, HDAC inhibition may be a therapeutic target for hypoxic tumor cells.


Assuntos
Neoplasias de Próstata Resistentes à Castração , Neoplasias da Próstata , Masculino , Humanos , Androgênios/metabolismo , Neoplasias da Próstata/patologia , Antagonistas de Androgênios/uso terapêutico , Receptores Androgênicos/metabolismo , Neoplasias de Próstata Resistentes à Castração/genética , Linhagem Celular Tumoral
11.
Neuro Oncol ; 26(8): 1388-1401, 2024 Aug 05.
Artigo em Inglês | MEDLINE | ID: mdl-38456228

RESUMO

BACKGROUND: Hypoxia is associated with poor prognosis in many cancers including glioblastoma (GBM). Glioma stem-like cells (GSCs) often reside in hypoxic regions and serve as reservoirs for disease progression. Long non-coding RNAs (lncRNAs) have been implicated in GBM. However, the lncRNAs that modulate GSC adaptations to hypoxia are poorly understood. Identification of these lncRNAs may provide new therapeutic strategies to target GSCs under hypoxia. METHODS: lncRNAs induced by hypoxia in GSCs were identified by RNA-seq. Lung cancer-associated transcript-1 (LUCAT1) expression was assessed by qPCR, RNA-seq, Northern blot, single molecule FISH in GSCs, and interrogated in IvyGAP, The Cancer Genome Atlas, and CGGA databases. LUCAT1 was depleted by shRNA, CRISPR/Cas9, and CRISPR/Cas13d. RNA-seq, Western blot, immunohistochemistry, co-IP, ChIP, ChIP-seq, RNA immunoprecipitation, and proximity ligation assay were performed to investigate mechanisms of action of LUCAT1. GSC viability, limiting dilution assay, and tumorigenic potential in orthotopic GBM xenograft models were performed to assess the functional consequences of depleting LUCAT1. RESULTS: A new isoform of Lucat1 is induced by Hypoxia inducible factor 1 alpha (HIF1α) and Nuclear factor erythroid 2-related factor 2 (NRF2) in GSCs under hypoxia. LUCAT1 is highly expressed in hypoxic regions in GBM. Mechanistically, LUCAT1 formed a complex with HIF1α and its co-activator CBP to regulate HIF1α target gene expression and GSC adaptation to hypoxia. Depletion of LUCAT1 impaired GSC self-renewal. Silencing LUCAT1 decreased tumor growth and prolonged mouse survival in GBM xenograft models. CONCLUSIONS: A HIF1α-LUCAT1 axis forms a positive feedback loop to amplify HIF1α signaling in GSCs under hypoxia. LUCAT1 promotes GSC self-renewal and GBM tumor growth. LUCAT1 is a potential therapeutic target in GBM.


Assuntos
Neoplasias Encefálicas , Regulação Neoplásica da Expressão Gênica , Glioblastoma , Subunidade alfa do Fator 1 Induzível por Hipóxia , RNA Longo não Codificante , Humanos , RNA Longo não Codificante/genética , Glioblastoma/metabolismo , Glioblastoma/patologia , Glioblastoma/genética , Animais , Camundongos , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patologia , Neoplasias Encefálicas/genética , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Progressão da Doença , Ensaios Antitumorais Modelo de Xenoenxerto , Proliferação de Células , Células Tumorais Cultivadas , Camundongos Nus , Linhagem Celular Tumoral , Biomarcadores Tumorais/metabolismo , Biomarcadores Tumorais/genética , Prognóstico , Apoptose
12.
Cell Rep ; 43(1): 113661, 2024 01 23.
Artigo em Inglês | MEDLINE | ID: mdl-38175754

RESUMO

Myeloid-derived suppressor cells (MDSCs) impair antitumor immune responses. Identifying regulatory circuits during MDSC development may bring new opportunities for therapeutic interventions. We report that the V-domain suppressor of T cell activation (VISTA) functions as a key enabler of MDSC differentiation. VISTA deficiency reduced STAT3 activation and STAT3-dependent production of polyamines, which causally impaired mitochondrial respiration and MDSC expansion. In both mixed bone marrow (BM) chimera mice and myeloid-specific VISTA conditional knockout mice, VISTA deficiency significantly reduced tumor-associated MDSCs but expanded monocyte-derived dendritic cells (DCs) and enhanced T cell-mediated tumor control. Correlated expression of VISTA and arginase-1 (ARG1), a key enzyme supporting polyamine biosynthesis, was observed in multiple human cancer types. In human endometrial cancer, co-expression of VISTA and ARG1 on tumor-associated myeloid cells is associated with poor survival. Taken together, these findings unveil the VISTA/polyamine axis as a central regulator of MDSC differentiation and warrant therapeutically targeting this axis for cancer immunotherapy.


Assuntos
Células Supressoras Mieloides , Neoplasias , Animais , Humanos , Camundongos , Camundongos Knockout , Células Mieloides/metabolismo , Neoplasias/patologia , Poliaminas/metabolismo , Fator de Transcrição STAT3/metabolismo , Linfócitos T
13.
Hematol Oncol Clin North Am ; 37(5): 827-840, 2023 10.
Artigo em Inglês | MEDLINE | ID: mdl-37246090

RESUMO

The treatment of advanced renal cell carcinoma (RCC) has changed dramatically with immune checkpoint inhibitors, yet most patients do not have durable responses. There is consequently a tremendous need for novel therapeutic development. RCC, and particularly the most common histology clear cell RCC, is an immunobiologically and metabolically distinct tumor. An improved understanding of RCC-specific biology will be necessary for the successful identification of new treatment targets for this disease. In this review, we discuss the current understanding of RCC immune pathways and metabolic dysregulation, with a focus on topics important for future clinical development.


Assuntos
Carcinoma de Células Renais , Neoplasias Renais , Humanos , Carcinoma de Células Renais/genética , Carcinoma de Células Renais/terapia , Neoplasias Renais/genética , Neoplasias Renais/terapia , Inibidores de Checkpoint Imunológico/uso terapêutico , Redes e Vias Metabólicas
14.
J Clin Invest ; 133(22)2023 11 15.
Artigo em Inglês | MEDLINE | ID: mdl-37966114

RESUMO

Half of all men with advanced prostate cancer (PCa) inherit at least 1 copy of an adrenal-permissive HSD3B1 (1245C) allele, which increases levels of 3ß-hydroxysteroid dehydrogenase 1 (3ßHSD1) and promotes intracellular androgen biosynthesis. Germline inheritance of the adrenally permissive allele confers worse outcomes in men with advanced PCa. We investigated whether HSD3B1 (1245C) drives resistance to combined androgen deprivation and radiotherapy. Adrenally permissive 3ßHSD1 enhanced resistance to radiotherapy in PCa cell lines and xenograft models engineered to mimic the human adrenal/gonadal axis during androgen deprivation. The allele-specific effects on radiosensitivity were dependent on availability of DHEA, the substrate for 3ßHSD1. In lines expressing the HSD3B1 (1245C) allele, enhanced expression of DNA damage response (DDR) genes and more rapid DNA double-strand break (DSB) resolution were observed. A correlation between androgen receptor (AR) expression and increased DDR gene expression was confirmed in 680 radical prostatectomy specimens. Treatment with the nonsteroidal antiandrogen enzalutamide reversed the resistant phenotype of HSD3B1 (1245C) PCa in vitro and in vivo. In conclusion, 3ßHSD1 promotes prostate cancer resistance to combined androgen deprivation and radiotherapy by upregulating DNA DSB repair. This work supports prospective validation of early combined androgen blockade for high-risk men harboring the HSD3B1 (1245C) allele.


Assuntos
Neoplasias de Próstata Resistentes à Castração , Neoplasias da Próstata , Humanos , Masculino , Antagonistas de Androgênios/farmacologia , Antagonistas de Androgênios/uso terapêutico , Androgênios/metabolismo , DNA , Genótipo , Hidroxiesteroide Desidrogenases/genética , Complexos Multienzimáticos/genética , Neoplasias da Próstata/genética , Neoplasias da Próstata/radioterapia , Neoplasias da Próstata/metabolismo , Neoplasias de Próstata Resistentes à Castração/genética , Receptores Androgênicos/genética , Receptores Androgênicos/metabolismo
15.
Cancer Res ; 82(13): 2417-2430, 2022 07 05.
Artigo em Inglês | MEDLINE | ID: mdl-35536859

RESUMO

Androgen deprivation therapy suppresses tumor androgen receptor (AR) signaling by depleting circulating testosterone and is a mainstay treatment for advanced prostate cancer. Despite initial treatment response, castration-resistant prostate cancer nearly always develops and remains driven primarily by the androgen axis. Here we investigated how changes in oxygenation affect androgen synthesis. In prostate cancer cells, chronic hypoxia coupled to reoxygenation resulted in efficient metabolism of androgen precursors to produce androgens and activate AR. Hypoxia induced 3ßHSD1, the rate-limiting androgen synthesis regulator, and reoxygenation replenished necessary cofactors, suggesting that hypoxia and reoxygenation both facilitate potent androgen synthesis. The EGLN1/VHL/HIF2α pathway induced 3ßHSD1 expression through direct binding of HIF2α to the 5' regulatory region of HSD3B1 to promote transcription. Overexpression of HIF2α facilitated prostate cancer progression, which largely depended on 3ßHSD1. Inhibition of HIF2α with the small-molecule PT2399 prevented prostate cancer cell proliferation. These results thus identify HIF2α as a regulator of androgen synthesis and potential therapeutic target in prostate cancer. SIGNIFICANCE: Hypoxia followed by reoxygenation in prostate cancer drives androgen deprivation therapy resistance via increasing the rate-limiting enzyme and cofactors for androgen synthesis, revealing HIF2α as a therapeutic target to subvert resistance.


Assuntos
Neoplasias de Próstata Resistentes à Castração , Neoplasias da Próstata , Antagonistas de Androgênios/farmacologia , Antagonistas de Androgênios/uso terapêutico , Androgênios/metabolismo , Linhagem Celular Tumoral , Humanos , Hipóxia , Masculino , Neoplasias da Próstata/metabolismo , Neoplasias de Próstata Resistentes à Castração/patologia , Receptores Androgênicos/metabolismo , Testosterona , Regulação para Cima
16.
Clin Cancer Res ; 28(21): 4689-4701, 2022 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-35776130

RESUMO

PURPOSE: Advanced/metastatic forms of clear-cell renal cell carcinomas (ccRCC) have limited therapeutic options. Genome-wide genetic screens have identified cellular dependencies in many cancers. Using the Broad Institute/Novartis combined short hairpin RNA (shRNA) dataset, and cross-validation with the CRISPR/Cas9 DepMap (21Q3) dataset, we sought therapeutically actionable dependencies in kidney lineage cancers. EXPERIMENTAL DESIGN: We identified preferential genetic dependencies in kidney cancer cells versus other lineages. BCL2L1, which encodes the BCL-XL antiapoptotic protein, scored as the top actionable dependency. We validated this finding using genetic and pharmacologic tools in a panel of ccRCC cell lines. Select BCL-XL-dependent (versus independent) cell lines were then transcriptionally profiled to identify biomarkers and mechanistic drivers of BCL-XL dependence. Cell-based studies (in vitro and in vivo) and clinical validations were used to address physiologic relevance. RESULTS: Inactivation of BCL-XL, but not BCL-2, led to fitness defects in renal cancer cells, and sensitized them to chemotherapeutics. Transcriptomic profiling identified a "BCL-XL dependency" signature, including an elevated mesenchymal gene signature. A mesenchymal state was both necessary and sufficient to confer increased BCL-XL dependence. The "BCL-XL dependency" signature was observed in approximately 30% of human ccRCCs, which were also associated with worse clinical outcomes. Finally, an orally bioavailable BCL-XL inhibitor, A-1331852, showed antitumor efficacy in vivo. CONCLUSIONS: Our studies uncovered an unexpected link between cell state and BCL-XL dependence in ccRCC. Therapeutic agents that specifically target BCL-XL are available. Our work justifies testing the utility of BCL-XL blockade to target, likely, a clinically aggressive subset of human kidney cancers. See related commentary by Wang et al., p. 4600.


Assuntos
Carcinoma de Células Renais , Neoplasias Renais , Humanos , Carcinoma de Células Renais/tratamento farmacológico , Carcinoma de Células Renais/genética , Linhagem Celular Tumoral , Proteína bcl-X/genética , Proteína bcl-X/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Apoptose/genética , Neoplasias Renais/tratamento farmacológico , Neoplasias Renais/genética , RNA Interferente Pequeno
17.
Proc Natl Acad Sci U S A ; 105(16): 6103-8, 2008 Apr 22.
Artigo em Inglês | MEDLINE | ID: mdl-18413597

RESUMO

Adenovirus E1A drives oncogenesis by targeting key regulatory pathways that are critical for cellular growth control. The interaction of E1A with p400 is essential for many E1A activities, but the downstream target of this interaction is unknown. Here, we present evidence that the oncoprotein transcription factor Myc is the target of this interaction. We show that E1A stabilizes Myc protein via p400 and promotes the coassociation of Myc and p400 at Myc target genes, leading to their transcriptional induction. We also show that E1A requires Myc for its ability to activate Myc-dependent gene expression and induce apoptosis, and that forced expression of Myc is sufficient to rescue the activity of an E1A-mutant defective in p400 binding. Together, these findings establish that Myc, via p400, is an essential downstream target of E1A.


Assuntos
Proteínas E1A de Adenovirus/metabolismo , Transformação Celular Viral , DNA Helicases/metabolismo , Proteínas de Ligação a DNA/metabolismo , Proteínas Proto-Oncogênicas c-myc/metabolismo , Proteínas E1A de Adenovirus/genética , Linhagem Celular , Imunoprecipitação da Cromatina , Humanos , Proteínas Proto-Oncogênicas c-myc/genética
18.
Exp Cell Res ; 315(10): 1772-8, 2009 Jun 10.
Artigo em Inglês | MEDLINE | ID: mdl-19331831

RESUMO

Mechanisms that couple protein turnover to cell cycle progression are critical for coordinating the events of cell duplication and division. Despite the importance of cell cycle-regulated proteolysis, however, technologies to measure this phenomenon are limited, and typically involve monitoring cells that are released back into the cell cycle after synchronization. We describe here the use of laser scanning cytometry (LSC), a technical merger between fluorescence microscopy and flow cytometry, to determine cell cycle-dependent changes in protein stability in unperturbed, asynchronous, cultures of mammalian cells. In this method, the ability of the LSC to accurately measure whole cell fluorescence is employed, together with RNA fluorescence in situ hybridization and immunofluorescence, to relate abundance of a particular RNA and protein in a cell to its point at the cell cycle. Parallel monitoring of RNA and protein levels is used, together with protein synthesis inhibitors, to reveal cell cycle-specific changes in protein turnover. We demonstrate the viability of this method by analyzing the proteolysis of two prominent human oncoproteins, Myc and Cyclin E, and argue that this LSC-based approach offers several practical advantages over traditional cell synchronization methods.


Assuntos
Ciclo Celular , Citometria de Varredura a Laser , Processamento de Proteína Pós-Traducional , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular Tumoral , Ciclina E/metabolismo , Proteínas F-Box/metabolismo , Proteína 7 com Repetições F-Box-WD , Regulação Neoplásica da Expressão Gênica , Humanos , Fosforilação , Proteínas Proto-Oncogênicas c-myc/genética , Proteínas Proto-Oncogênicas c-myc/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ubiquitina-Proteína Ligases/metabolismo
19.
Science ; 363(6432): 1217-1222, 2019 03 15.
Artigo em Inglês | MEDLINE | ID: mdl-30872525

RESUMO

Oxygen sensing is central to metazoan biology and has implications for human disease. Mammalian cells express multiple oxygen-dependent enzymes called 2-oxoglutarate (OG)-dependent dioxygenases (2-OGDDs), but they vary in their oxygen affinities and hence their ability to sense oxygen. The 2-OGDD histone demethylases control histone methylation. Hypoxia increases histone methylation, but whether this reflects direct effects on histone demethylases or indirect effects caused by the hypoxic induction of the HIF (hypoxia-inducible factor) transcription factor or the 2-OG antagonist 2-hydroxyglutarate (2-HG) is unclear. Here, we report that hypoxia promotes histone methylation in a HIF- and 2-HG-independent manner. We found that the H3K27 histone demethylase KDM6A/UTX, but not its paralog KDM6B, is oxygen sensitive. KDM6A loss, like hypoxia, prevented H3K27 demethylation and blocked cellular differentiation. Restoring H3K27 methylation homeostasis in hypoxic cells reversed these effects. Thus, oxygen directly affects chromatin regulators to control cell fate.


Assuntos
Cromatina/metabolismo , Histona Desmetilases/metabolismo , Proteínas Nucleares/metabolismo , Oxigênio/metabolismo , Animais , Hipóxia Celular , Células HEK293 , Histona Desmetilases/genética , Histonas/metabolismo , Humanos , Fator 1 Induzível por Hipóxia/metabolismo , Histona Desmetilases com o Domínio Jumonji/genética , Histona Desmetilases com o Domínio Jumonji/metabolismo , Células MCF-7 , Metilação , Camundongos , Proteínas Nucleares/genética
20.
Cancer Discov ; 9(2): 230-247, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30373918

RESUMO

Small cell lung cancer (SCLC) accounts for 15% of lung cancers and is almost always linked to inactivating RB1 and TP53 mutations. SCLC frequently responds, albeit briefly, to chemotherapy. The canonical function of the RB1 gene product RB1 is to repress the E2F transcription factor family. RB1 also plays both E2F-dependent and E2F-independent mitotic roles. We performed a synthetic lethal CRISPR/Cas9 screen in an RB1 -/- SCLC cell line that conditionally expresses RB1 to identify dependencies that are caused by RB1 loss and discovered that RB1 -/- SCLC cell lines are hyperdependent on multiple proteins linked to chromosomal segregation, including Aurora B kinase. Moreover, we show that an Aurora B kinase inhibitor is efficacious in multiple preclinical SCLC models at concentrations that are well tolerated in mice. These results suggest that RB1 loss is a predictive biomarker for sensitivity to Aurora B kinase inhibitors in SCLC and perhaps other RB1 -/- cancers. SIGNIFICANCE: SCLC is rarely associated with actionable protooncogene mutations. We did a CRISPR/Cas9-based screen that showed that RB1 -/- SCLC are hyperdependent on AURKB, likely because both genes control mitotic fidelity, and confirmed that Aurora B kinase inhibitors are efficacious against RB1 -/- SCLC tumors in mice at nontoxic doses.See related commentary by Dick and Li, p. 169.This article is highlighted in the In This Issue feature, p. 151.


Assuntos
Aurora Quinase B/metabolismo , Proliferação de Células , Genes Supressores de Tumor , Neoplasias Pulmonares/patologia , Mutação , Proteínas de Ligação a Retinoblastoma/metabolismo , Carcinoma de Pequenas Células do Pulmão/patologia , Ubiquitina-Proteína Ligases/metabolismo , Animais , Antineoplásicos/farmacologia , Apoptose , Aurora Quinase B/genética , Sistemas CRISPR-Cas , Segregação de Cromossomos , Resistencia a Medicamentos Antineoplásicos , Feminino , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/metabolismo , Camundongos , Camundongos Nus , Proteínas de Ligação a Retinoblastoma/antagonistas & inibidores , Proteínas de Ligação a Retinoblastoma/genética , Transdução de Sinais , Carcinoma de Pequenas Células do Pulmão/tratamento farmacológico , Carcinoma de Pequenas Células do Pulmão/metabolismo , Células Tumorais Cultivadas , Ubiquitina-Proteína Ligases/antagonistas & inibidores , Ubiquitina-Proteína Ligases/genética , Ensaios Antitumorais Modelo de Xenoenxerto
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