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1.
Blood ; 2020 Jun 12.
Artigo em Inglês | MEDLINE | ID: mdl-32530039

RESUMO

Cytokine storm syndromes (CSS) are severe hyperinflammatory conditions characterized by excessive immune system activation leading to organ damage and death. Hemophagocytic lymphohistiocytosis (HLH), a disease often associated with inherited defects in cell-mediated cytotoxicity, serves as a prototypical CSS for which the five-year survival is only 60%. Frontline therapy for HLH consists of the glucocorticoid dexamethasone (DEX) and the chemotherapeutic agent etoposide. Many patients, however, are refractory to this treatment or relapse after an initial response. Notably, many cytokines that are elevated in HLH activate the JAK/STAT pathway, and the JAK1/2 inhibitor ruxolitinib (RUX) has shown efficacy in murine HLH models and humans with refractory disease. We recently reported that cytokine-induced JAK/STAT signaling mediates DEX resistance in T cell acute lymphoblastic leukemia (T-ALL) cells, and that this could be effectively reversed by RUX. Based on these findings, we hypothesized that cytokine-mediated JAK/STAT signaling might similarly contribute to DEX resistance in HLH and that RUX treatment would overcome this phenomenon. Using ex vivo assays, a murine model of HLH, and primary patient samples, we demonstrate that the hypercytokinemia of HLH reduces the apoptotic potential of CD8 T cells leading to relative DEX resistance. Upon exposure to RUX, this apoptotic potential is restored, thereby sensitizing CD8 T cells to DEX-induced apoptosis in vitro and significantly reducing tissue immunopathology and HLH disease manifestations in vivo. Our findings provide rationale for combining DEX and RUX to enhance the lymphotoxic effects of DEX and thus improve the outcomes for patients with HLH and related CSS.

2.
Vet Pathol ; 57(3): 445-456, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32202225

RESUMO

Pediatric patients receiving solid organ transplants may develop lymphoproliferative diseases, including graft-versus-host disease (GvHD) and posttransplant lymphoproliferative diseases (PTLDs). We characterized lesions in 11 clinically ill NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ (NSG) mice that received pediatric-patient-derived solid tumors (PDXs) and developed immunodeficiency-associated lymphoproliferations comparable to GvHD and PTLDs over a period of 46 to 283 days after implantation. Lymphoproliferations were diffusely positive for human-specific biomarkers, including NUMA1, CD45, and CD43, but lacked immunoreactivity for murine CD45. Human immune cells were CD3-positive, with subsets having immunoreactivity for CD4 and CD8 as well as PAX5, CD79a, and IRF4, resulting from populations of human T and B cells present within the xenotransplants. Tissues and organs infiltrated included mucocutaneous zones (oral cavity and perigenital and perianal regions), haired skin, tongue, esophagus, forestomach, thyroid, salivary glands, lungs, liver, kidneys, spleen, lymph nodes, bone marrow, and brain. In 4 of 5 mice with PTLD, Epstein-Barr virus (EBV)-encoded small RNAs (EBERs) were detected by in situ hybridization in PAX5+ human B cells associated with the PDX (n = 1/4) or with engrafted human immune cells at other anatomic locations (n = 4/11). One of the 4 mice had an EBV-associated human large B-cell lymphoma. NSG mice receiving xenotransplants can develop combinations of GvHD, EBV-driven PTLD, and B-cell lymphoma similar to those occurring in human pediatric patients. Therefore, pediatric xenotransplants should undergo histopathologic and immunohistochemical assessment upon collection to ensure that the specimen is not a lymphoma and does not contain lymphoma cells because these neoplasms can morphologically mimic small round blue cell pediatric solid tumors.

3.
Mol Cell ; 77(6): 1206-1221.e7, 2020 03 19.
Artigo em Inglês | MEDLINE | ID: mdl-31980388

RESUMO

Alternative polyadenylation (APA) contributes to transcriptome complexity by generating mRNA isoforms with varying 3' UTR lengths. APA leading to 3' UTR shortening (3' US) is a common feature of most cancer cells; however, the molecular mechanisms are not understood. Here, we describe a widespread mechanism promoting 3' US in cancer through ubiquitination of the mRNA 3' end processing complex protein, PCF11, by the cancer-specific MAGE-A11-HUWE1 ubiquitin ligase. MAGE-A11 is normally expressed only in the male germline but is frequently re-activated in cancers. MAGE-A11 is necessary for cancer cell viability and is sufficient to drive tumorigenesis. Screening for targets of MAGE-A11 revealed that it ubiquitinates PCF11, resulting in loss of CFIm25 from the mRNA 3' end processing complex. This leads to APA of many transcripts affecting core oncogenic and tumor suppressors, including cyclin D2 and PTEN. These findings provide insights into the molecular mechanisms driving APA in cancer and suggest therapeutic strategies.

4.
Vet Pathol ; 57(1): 160-171, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31736441

RESUMO

The NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ strain (NOD scid gamma, NSG) is a severely immunodeficient inbred laboratory mouse used for preclinical studies because it is amenable to engraftment with human cells. Combining scid and Il2rgnull mutations results in severe immunodeficiency by impairing the maturation, survival, and functionality of interleukin 2-dependent immune cells, including T, B, and natural killer lymphocytes. While NSG mice are reportedly resistant to developing spontaneous lymphomas/leukemias, there are reports of hematopoietic cancers developing. In this study, we characterized the immunophenotype of spontaneous lymphoma/leukemia in 12 NSG mice (20 to 38 weeks old). The mice had a combination of grossly enlarged thymus, spleen, or lymph nodes and variable histologic involvement of the bone marrow and other tissues. All 12 lymphomas were diffusely CD3, TDT, and CD4 positive, and 11 of 12 were also positive for CD8, which together was consistent with precursor T-cell lymphoblastic lymphoma/leukemia (pre-T-LBL). A subset of NSG tissues from all mice and neoplastic lymphocytes from 8 of 12 cases had strong immunoreactivity for retroviral p30 core protein, suggesting an association with a viral infection. These data highlight that NSG mice may develop T-cell lymphoma at low frequency, necessitating the recognition of this spontaneously arising disease when interpreting studies.

5.
Nat Commun ; 10(1): 3718, 2019 08 16.
Artigo em Inglês | MEDLINE | ID: mdl-31420543

RESUMO

High throughput omics approaches provide an unprecedented opportunity for dissecting molecular mechanisms in cancer biology. Here we present deep profiling of whole proteome, phosphoproteome and transcriptome in two high-grade glioma (HGG) mouse models driven by mutated RTK oncogenes, PDGFRA and NTRK1, analyzing 13,860 proteins and 30,431 phosphosites by mass spectrometry. Systems biology approaches identify numerous master regulators, including 41 kinases and 23 transcription factors. Pathway activity computation and mouse survival indicate the NTRK1 mutation induces a higher activation of AKT downstream targets including MYC and JUN, drives a positive feedback loop to up-regulate multiple other RTKs, and confers higher oncogenic potency than the PDGFRA mutation. A mini-gRNA library CRISPR-Cas9 validation screening shows 56% of tested master regulators are important for the viability of NTRK-driven HGG cells, including TFs (Myc and Jun) and metabolic kinases (AMPKa1 and AMPKa2), confirming the validity of the multiomics integrative approaches, and providing novel tumor vulnerabilities.


Assuntos
Neoplasias Encefálicas/genética , Perfilação da Expressão Gênica , Glioma/genética , Proteômica , Proteínas Quinases Ativadas por AMP/metabolismo , Animais , Neoplasias Encefálicas/metabolismo , Modelos Animais de Doenças , Retroalimentação Fisiológica , Glioma/metabolismo , Camundongos , Mutação , Proteína Oncogênica p65(gag-jun)/metabolismo , Fosfopeptídeos/metabolismo , Fosfoproteínas/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas Proto-Oncogênicas c-myc/metabolismo , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/genética , Receptor trkA/genética , Transdução de Sinais , Biologia de Sistemas , Regulação para Cima
6.
Sci Transl Med ; 11(506)2019 08 21.
Artigo em Inglês | MEDLINE | ID: mdl-31434755

RESUMO

In ß-thalassemia, accumulated free α-globin forms intracellular precipitates that impair erythroid cell maturation and viability. Protein quality control systems mitigate ß-thalassemia pathophysiology by degrading toxic free α-globin, although the associated mechanisms are poorly understood. We show that loss of the autophagy-activating Unc-51-like kinase 1 (Ulk1) gene in ß-thalassemic mice reduces autophagic clearance of α-globin in red blood cell precursors and exacerbates disease phenotypes, whereas inactivation of the canonical autophagy-related 5 (Atg5) gene has relatively minor effects. Systemic treatment with the mTORC1 inhibitor rapamycin reduces α-globin precipitates and lessens pathologies in ß-thalassemic mice via an ULK1-dependent pathway. Similarly, rapamycin reduces free α-globin accumulation in erythroblasts derived from CD34+ cells of ß-thalassemic individuals. Our findings define a drug-regulatable pathway for ameliorating ß-thalassemia.

7.
Nat Commun ; 10(1): 3623, 2019 08 09.
Artigo em Inglês | MEDLINE | ID: mdl-31399583

RESUMO

Coordinated regulation of the lysosomal and autophagic systems ensures basal catabolism and normal cell physiology, and failure of either system causes disease. Here we describe an epigenetic rheostat orchestrated by c-MYC and histone deacetylases that inhibits lysosomal and autophagic biogenesis by concomitantly repressing the expression of the transcription factors MiT/TFE and FOXH1, and that of lysosomal and autophagy genes. Inhibition of histone deacetylases abates c-MYC binding to the promoters of lysosomal and autophagy genes, granting promoter occupancy to the MiT/TFE members, TFEB and TFE3, and/or the autophagy regulator FOXH1. In pluripotent stem cells and cancer, suppression of lysosomal and autophagic function is directly downstream of c-MYC overexpression and may represent a hallmark of malignant transformation. We propose that, by determining the fate of these catabolic systems, this hierarchical switch regulates the adaptive response of cells to pathological and physiological cues that could be exploited therapeutically.


Assuntos
Autofagia/fisiologia , Epigênese Genética , Lisossomos/metabolismo , Biogênese de Organelas , Politetrafluoretileno/metabolismo , Proteínas Proto-Oncogênicas c-myc/metabolismo , Autofagia/genética , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/metabolismo , Sítios de Ligação , Linhagem Celular Tumoral , Neoplasias do Colo/genética , Neoplasias do Colo/metabolismo , Neoplasias do Colo/patologia , Fatores de Transcrição Forkhead/metabolismo , Regulação Neoplásica da Expressão Gênica , Histona Desacetilase 2/metabolismo , Histona Desacetilases/metabolismo , Humanos , Regiões Promotoras Genéticas , Proteínas Proto-Oncogênicas c-myc/genética , Células-Tronco , Transcrição Genética
8.
Sci Adv ; 5(5): eaav4832, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-31149633

RESUMO

Ensuring robust gamete production even in the face of environmental stress is of utmost importance for species survival, especially in mammals that have low reproductive rates. Here, we describe a family of genes called melanoma antigens (MAGEs) that evolved in eutherian mammals and are normally restricted to expression in the testis (http://MAGE.stjude.org) but are often aberrantly activated in cancer. Depletion of Mage-a genes disrupts spermatogonial stem cell maintenance and impairs repopulation efficiency in vivo. Exposure of Mage-a knockout mice to genotoxic stress or long-term starvation that mimics famine in nature causes defects in spermatogenesis, decreased testis weights, diminished sperm production, and reduced fertility. Last, human MAGE-As are activated in many cancers where they promote fuel switching and growth of cells. These results suggest that mammalian-specific MAGE genes have evolved to protect the male germline against environmental stress, ensure reproductive success under non-optimal conditions, and are hijacked by cancer cells.

9.
Blood ; 134(2): 147-159, 2019 07 11.
Artigo em Inglês | MEDLINE | ID: mdl-31015190

RESUMO

Hemophagocytic lymphohistiocytosis (HLH) is an often-fatal disorder characterized by the overactivation of T cells and macrophages that excessively produce proinflammatory cytokines, including interferon-γ (IFN-γ). Previously, we reported that the JAK inhibitor ruxolitinib dampens T-cell activation and lessens inflammation in a model of HLH in which perforin-deficient (Prf1 -/-) mice are infected with lymphocytic choriomeningitis virus (LCMV). Ruxolitinib inhibits signaling downstream of IFN-γ, as well as several other JAK-dependent cytokines. As a consequence, it remained unclear whether ruxolitinib was exerting its beneficial effects in HLH by inhibiting IFN-γ signaling or by targeting signaling initiated by other proinflammatory cytokines. To address this question, we compared the effects of ruxolitinib with those obtained using an IFN-γ-neutralizing antibody (αIFN-γ) in 2 murine HLH models. In both models, ruxolitinib and αIFN-γ reduced inflammation-associated anemia, indicating that ruxolitinib operates in an IFN-γ-dependent manner to reverse this HLH manifestation. In contrast, the number and activation status of T cells and neutrophils, as well as their infiltration into tissues, were significantly reduced following treatment with ruxolitinib, but they remained unchanged or were increased following treatment with αIFN-γ. Notably, despite discontinuation of ruxolitinib, LCMV-infected Prf1 -/- mice exhibited enhanced survival compared with mice in which αIFN-γ was discontinued. This protective effect could be mimicked by transient treatment with αIFN-γ and a neutrophil-depleting antibody. Thus, ruxolitinib operates through IFN-γ-dependent and -independent mechanisms to dampen HLH by targeting the deleterious effects of T cells and neutrophils, with the latter representing an unappreciated and understudied cell type that contributes to HLH pathogenesis.


Assuntos
Linfo-Histiocitose Hemofagocítica/imunologia , Neutrófilos/efeitos dos fármacos , Pirazóis/farmacologia , Linfócitos T/efeitos dos fármacos , Animais , Modelos Animais de Doenças , Ativação Linfocitária/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos C57BL
10.
Cancer Res ; 79(12): 3112-3124, 2019 06 15.
Artigo em Inglês | MEDLINE | ID: mdl-31015228

RESUMO

The cell membrane glycolipid GD2 is expressed by multiple solid tumors, including 88% of osteosarcomas and 98% of neuroblastomas. However, osteosarcomas are highly heterogeneous, with many tumors exhibiting GD2 expression on <50% of the individual cells, while some tumors are essentially GD2-negative. Anti-GD2 immunotherapy is the current standard of care for high-risk neuroblastoma, but its application to recurrent osteosarcomas, for which no effective therapies exist, has been extremely limited. This is, in part, because the standard assays to measure GD2 expression in these heterogeneous tumors are not quantitative and are subject to tissue availability and sampling bias. To address these limitations, we evaluated a novel, sensitive radiotracer [64Cu]Cu-Bn-NOTA-hu14.18K322A to detect GD2 expression in osteosarcomas (six patient-derived xenografts and one cell line) in vivo using positron emission tomography (PET). Tumor uptake of the radiolabeled, humanized anti-GD2 antibody [64Cu]Cu-Bn-NOTA-hu14.18K322A was 7-fold higher in modestly GD2-expressing osteosarcomas (32% GD2-positive cells) than in a GD2-negative tumor (9.8% vs. 1.3% of the injected dose per cc, respectively). This radiotracer also identified lesions as small as 29 mm3 in a 34% GD2-positive model of metastatic osteosarcoma of the lung. Radiolabeled antibody accumulation in patient-derived xenografts correlated with GD2 expression as measured by flow cytometry (Pearson r = 0.88, P = 0.01), distinguishing moderately GD2-expressing osteosarcomas (32%-69% GD2-positive cells) from high GD2 expressors (>99%, P < 0.05). These results support the utility of GD2 imaging with PET to measure GD2 expression in osteosarcoma and thus maximize the clinical impact of anti-GD2 immunotherapy. SIGNIFICANCE: In situ assessment of all GD2-positive osteosarcoma sites with a novel PET radiotracer could significantly impact anti-GD2 immunotherapy patient selection and enable noninvasive probing of correlations between target expression and therapeutic response.


Assuntos
Anticorpos Monoclonais/farmacologia , Neoplasias Ósseas/patologia , Gangliosídeos/antagonistas & inibidores , Neoplasias Pulmonares/secundário , Recidiva Local de Neoplasia/patologia , Osteossarcoma/patologia , Tomografia por Emissão de Pósitrons/métodos , Animais , Apoptose , Neoplasias Ósseas/tratamento farmacológico , Neoplasias Ósseas/imunologia , Neoplasias Ósseas/metabolismo , Proliferação de Células , Gangliosídeos/imunologia , Humanos , Imunoterapia , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/imunologia , Neoplasias Pulmonares/metabolismo , Camundongos , Camundongos Nus , Recidiva Local de Neoplasia/tratamento farmacológico , Recidiva Local de Neoplasia/imunologia , Recidiva Local de Neoplasia/metabolismo , Osteossarcoma/tratamento farmacológico , Osteossarcoma/imunologia , Osteossarcoma/metabolismo , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de Xenoenxerto
11.
J Clin Invest ; 129(6): 2251-2265, 2019 04 29.
Artigo em Inglês | MEDLINE | ID: mdl-31033479

RESUMO

Systemic lupus erythematosus (SLE) is a complex autoimmune disease with genetic and environmental contributions. Hallmarks of the disease are the appearance of immune complexes (IC) containing autoreactive Abs and TLR-activating nucleic acids, whose deposition in kidney glomeruli is suspected to promote tissue injury and glomerulonephritis (GN). Here, using a mouse model based on the human SLE susceptibility locus TNFAIP3-interacting protein 1 (TNIP1, also known as ABIN1), we investigated the pathogenesis of GN. We found that GN was driven by TLRs but, remarkably, proceeded independently of ICs. Rather, disease in 3 different mouse models and patients with SLE was characterized by glomerular accumulation of patrolling monocytes (PMos), a cell type with an emerging key function in vascular inflammation. Consistent with such function in GN, monocyte-specific deletion of ABIN1 promoted kidney disease, whereas selective elimination of PMos provided protection. In contrast to GN, PMo elimination did not protect from reduced survival or disease symptoms such as IC generation and splenomegaly, suggesting that GN and other inflammatory processes are governed by distinct pathogenic mechanisms. These data identify TLR-activated PMos as the principal component of an intravascular process that contributes to glomerular inflammation and kidney injury.


Assuntos
Nefrite Lúpica/imunologia , Monócitos/imunologia , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/imunologia , Animais , Modelos Animais de Doenças , Humanos , Inflamação/genética , Inflamação/imunologia , Inflamação/patologia , Nefrite Lúpica/genética , Nefrite Lúpica/patologia , Camundongos , Camundongos Knockout , Monócitos/patologia , Receptores Toll-Like/genética , Receptores Toll-Like/imunologia
12.
Oncotarget ; 10(12): 1235-1237, 2019 Feb 08.
Artigo em Inglês | MEDLINE | ID: mdl-30815226

RESUMO

Internal translational initiation of the mRNA encoding the Arf tumor suppressor yields an N-terminally truncated small Arf protein (smArf) that lacks amino acid residues required for Mdm2 binding and p53 activation. Here, we report that female, but not male, mice engineered to produce only smArf in lieu of the full-length Arf protein retain residual, sexually dimorphic tumor suppressive activity.

13.
J Clin Invest ; 128(11): 5073-5082, 2018 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-30295646

RESUMO

Arteriolar endothelial cell-expressed (EC-expressed) α-globin binds endothelial NOS (eNOS) and degrades its enzymatic product, NO, via dioxygenation, thereby lessening the vasodilatory effects of NO on nearby vascular smooth muscle. Although this reaction potentially affects vascular physiology, the mechanisms that regulate α-globin expression and dioxygenase activity in ECs are unknown. Without ß-globin, α-globin is unstable and cytotoxic, particularly in its oxidized form, which is generated by dioxygenation and recycled via endogenous reductases. We show that the molecular chaperone α-hemoglobin-stabilizing protein (AHSP) promotes arteriolar α-globin expression in vivo and facilitates its reduction by eNOS. In Ahsp-/- mice, EC α-globin was decreased by 70%. Ahsp-/- and Hba1-/- mice exhibited similar evidence of increased vascular NO signaling, including arteriolar dilation, blunted α1-adrenergic vasoconstriction, and reduced blood pressure. Purified α-globin bound eNOS or AHSP, but not both together. In ECs in culture, eNOS or AHSP enhanced α-globin expression posttranscriptionally. However, only AHSP prevented oxidized α-globin precipitation in solution. Finally, eNOS reduced AHSP-bound α-globin approximately 6-fold faster than did the major erythrocyte hemoglobin reductases (cytochrome B5 reductase plus cytochrome B5). Our data support a model whereby redox-sensitive shuttling of EC α-globin between AHSP and eNOS regulates EC NO degradation and vascular tone.


Assuntos
Células Endoteliais/metabolismo , Modelos Cardiovasculares , Chaperonas Moleculares/metabolismo , Contração Muscular , Transdução de Sinais , alfa-Globinas/metabolismo , Animais , Arteríolas , Células Endoteliais/citologia , Camundongos , Camundongos Knockout , Chaperonas Moleculares/genética , Óxido Nítrico/genética , Óxido Nítrico/metabolismo , Óxido Nítrico Sintase Tipo III/genética , alfa-Globinas/genética
14.
Sci Rep ; 8(1): 11034, 2018 07 23.
Artigo em Inglês | MEDLINE | ID: mdl-30038294

RESUMO

Respiratory syncytial virus (RSV) infection is the most frequent cause of hospitalization in infants and young children worldwide. Although mucosal RSV vaccines can reduce RSV disease burden, little is known about mucosal immune response capabilities in children. Neonatal or adult mice were infected with RSV; a subset of neonatal mice received interferon alpha (IFN-α) (intranasal) prior to RSV infection. B cells, B cell activating factor (BAFF) and IgA were measured by flow cytometry. RSV specific IgA was measured in nasal washes. Nasal associated lymphoid tissue (NALT) and lungs were stained for BAFF and IgA. Herein, we show in a mouse model of RSV infection that IFN-α plays a dual role as an antiviral and immune modulator and age-related differences in IgA production upon RSV infection can be overcome by IFN-α administration. IFN-α administration before RSV infection in neonatal mice increased RSV-specific IgA production in the nasal mucosa and induced expression of the B-cell activating factor BAFF in NALT. These findings are important, as mucosal antibodies at the infection site, and not serum antibodies, have been shown to protect human adults from experimental RSV infection.


Assuntos
Imunoglobulina A/imunologia , Imunoglobulina A/metabolismo , Interferon Tipo I/metabolismo , Infecções por Vírus Respiratório Sincicial/imunologia , Infecções por Vírus Respiratório Sincicial/metabolismo , Animais , Fator Ativador de Células B/metabolismo , Citometria de Fluxo , Camundongos , Camundongos Endogâmicos BALB C , Palivizumab/uso terapêutico , Reação em Cadeia da Polimerase em Tempo Real , Infecções por Vírus Respiratório Sincicial/tratamento farmacológico
15.
Oncotarget ; 9(43): 27087-27091, 2018 Jun 05.
Artigo em Inglês | MEDLINE | ID: mdl-29930752

RESUMO

Osteosarcoma (OS) cell lines are widely used in understanding the biological functions of cancer, identification and validation of therapeutic targets, as well as in vitro or in vivo preclinical drug screening. Here we report there is a frequent loss-of-function of polycomb repressive complex 2 (PRC2) in OS cell lines but it is rare in tumor samples based on genomic sequencing data, western blotting and immunohistochemistry analysis of H3K27me3. U2OS and 143B cell lines have a complete loss of function of PRC2 and several others have partial loss. In OS tumor tissues, only 1 out of 14 has low expression of H3K27me3. Kaplan-Meier analysis indicates that high EZH2, the component of PRC2, is associated with poor metastasis-free survival. Our observations are to raise the alarm that particular caution should be taken when using OS cell line models to study the disease, functional genomics, therapeutic target validation, drug screening, and epigenetic studies. Nevertheless, these cell lines will become useful biological tools to dissect the functions of PRC2.

16.
J Exp Med ; 215(6): 1519-1529, 2018 06 04.
Artigo em Inglês | MEDLINE | ID: mdl-29793924

RESUMO

Pyroptosis is an inflammasome-induced lytic cell death mode, the physiological role of which in chronic inflammatory diseases is unknown. Familial Mediterranean Fever (FMF) is the most common monogenic autoinflammatory disease worldwide, affecting an estimated 150,000 patients. The disease is caused by missense mutations in Mefv that activate the Pyrin inflammasome, but the pathophysiologic mechanisms driving autoinflammation in FMF are incompletely understood. Here, we show that Clostridium difficile infection of FMF knock-in macrophages that express a chimeric FMF-associated MefvV726A Pyrin elicited pyroptosis and gasdermin D (GSDMD)-mediated interleukin (IL)-1ß secretion. Importantly, in vivo GSDMD deletion abolished spontaneous autoinflammatory disease. GSDMD-deficient FMF knock-in mice were fully protected from the runted growth, anemia, systemic inflammatory cytokine production, neutrophilia, and tissue damage that characterize this autoinflammatory disease model. Overall, this work identifies pyroptosis as a critical mechanism of IL-1ß-dependent autoinflammation in FMF and highlights GSDMD inhibition as a potential antiinflammatory strategy in inflammasome-driven diseases.


Assuntos
Proteínas Reguladoras de Apoptose/metabolismo , Febre Familiar do Mediterrâneo/metabolismo , Febre Familiar do Mediterrâneo/patologia , Inflamação/metabolismo , Inflamação/patologia , Animais , Clostridium difficile/fisiologia , Citocinas/biossíntese , Modelos Animais de Doenças , Mediadores da Inflamação/metabolismo , Interleucina-1beta/metabolismo , Macrófagos/metabolismo , Macrófagos/microbiologia , Camundongos , Neutrófilos/patologia , Pirina/metabolismo , Pirina/farmacologia , Piroptose , Baço/patologia , Síndrome de Emaciação/patologia
17.
Cell Rep ; 23(3): 808-822, 2018 Apr 17.
Artigo em Inglês | MEDLINE | ID: mdl-29669286

RESUMO

Tumor cells initiate platelet activation leading to the secretion of bioactive molecules, which promote metastasis. Platelet receptors on tumors have not been well-characterized, resulting in a critical gap in knowledge concerning platelet-promoted metastasis. We identify a direct interaction between platelets and tumor CD97 that stimulates rapid bidirectional signaling. CD97, an adhesion G protein-coupled receptor (GPCR), is an overexpressed tumor antigen in several cancer types. Purified CD97 extracellular domain or tumor cell-associated CD97 stimulated platelet activation. CD97-initiated platelet activation led to granule secretion, including the release of ATP, a mediator of endothelial junction disruption. Lysophosphatidic acid (LPA) derived from platelets induced tumor invasiveness via proximal CD97-LPAR heterodimer signaling, coupling coincident tumor cell migration and vascular permeability to promote transendothelial migration. Consistent with this, CD97 was necessary for tumor cell-induced vascular permeability in vivo and metastasis formation in preclinical models. These findings support targeted blockade of tumor CD97 as an approach to ameliorate metastatic spread.


Assuntos
Antígenos CD/metabolismo , Plaquetas/metabolismo , Receptores Acoplados a Proteínas-G/metabolismo , Transdução de Sinais , Antígenos CD/genética , Plaquetas/citologia , Adesão Celular , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Dimerização , Fator de Crescimento Epidérmico/farmacologia , Transição Epitelial-Mesenquimal , Humanos , Lisofosfolipídeos/farmacologia , Ativação Plaquetária/efeitos dos fármacos , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Receptores Acoplados a Proteínas-G/antagonistas & inibidores , Receptores Acoplados a Proteínas-G/genética , Receptores de Ácidos Lisofosfatídicos/antagonistas & inibidores , Receptores de Ácidos Lisofosfatídicos/metabolismo , Junções Íntimas/metabolismo , Fator de Crescimento Transformador beta/metabolismo
18.
Nat Med ; 24(4): 427-437, 2018 05.
Artigo em Inglês | MEDLINE | ID: mdl-29505030

RESUMO

Spinal bulbar muscular atrophy (SBMA) is a motor neuron disease caused by toxic gain of function of the androgen receptor (AR). Previously, we found that co-regulator binding through the activation function-2 (AF2) domain of AR is essential for pathogenesis, suggesting that AF2 may be a potential drug target for selective modulation of toxic AR activity. We screened previously identified AF2 modulators for their ability to rescue toxicity in a Drosophila model of SBMA. We identified two compounds, tolfenamic acid (TA) and 1-[2-(4-methylphenoxy)ethyl]-2-[(2-phenoxyethyl)sulfanyl]-1H-benzimidazole (MEPB), as top candidates for rescuing lethality, locomotor function and neuromuscular junction defects in SBMA flies. Pharmacokinetic analyses in mice revealed a more favorable bioavailability and tissue retention of MEPB compared with TA in muscle, brain and spinal cord. In a preclinical trial in a new mouse model of SBMA, MEPB treatment yielded a dose-dependent rescue from loss of body weight, rotarod activity and grip strength. In addition, MEPB ameliorated neuronal loss, neurogenic atrophy and testicular atrophy, validating AF2 modulation as a potent androgen-sparing strategy for SBMA therapy.


Assuntos
Atrofia Muscular Espinal/patologia , Degeneração Neural/patologia , Receptores Androgênicos/química , Receptores Androgênicos/metabolismo , Animais , Benzimidazóis/farmacologia , Benzimidazóis/uso terapêutico , Proteínas Correpressoras/metabolismo , Modelos Animais de Doenças , Drosophila melanogaster , Células HEK293 , Humanos , Masculino , Camundongos Transgênicos , Atrofia Muscular Espinal/tratamento farmacológico , Degeneração Neural/tratamento farmacológico , Fenótipo , Projetos Piloto , Domínios Proteicos , Expansão das Repetições de Trinucleotídeos/genética , ortoaminobenzoatos/farmacologia , ortoaminobenzoatos/uso terapêutico
19.
BMC Cancer ; 16: 186, 2016 Mar 05.
Artigo em Inglês | MEDLINE | ID: mdl-26944546

RESUMO

BACKGROUND: Intrinsic and acquired resistance to drug therapies remains a challenge for malignant melanoma patients. Intratumoral heterogeneities within the tumor microenvironment contribute additional complexity to the determinants of drug efficacy and acquired resistance. METHODS: We use 3D biomimetic platforms to understand dynamics in extracellular matrix (ECM) biogenesis following pharmaceutical intervention against mitogen-activated protein kinases (MAPK) signaling. We further determined temporal evolution of secreted ECM components by isogenic melanoma cell clones. RESULTS: We found that the cell clones differentially secrete and assemble a myriad of ECM molecules into dense fibrillar and globular networks. We show that cells can modulate their ECM biosynthesis in response to external insults. Fibronectin (FN) is one of the key architectural components, modulating the efficacy of a broad spectrum of drug therapies. Stable cell lines engineered to secrete minimal levels of FN showed a concomitant increase in secretion of Tenascin-C and became sensitive to BRAF(V600E) and ERK inhibition as clonally- derived 3D tumor aggregates. These cells failed to assemble exogenous FN despite maintaining the integrin machinery to facilitate cell- ECM cross-talk. We determined that only clones that increased FN production via p38 MAPK and ß1 integrin survived drug treatment. CONCLUSIONS: These data suggest that tumor cells engineer drug resistance by altering their ECM biosynthesis. Therefore, drug treatment may induce ECM biosynthesis, contributing to de novo resistance.


Assuntos
Matriz Extracelular/metabolismo , Melanoma/metabolismo , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Transdução de Sinais , Animais , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Linhagem Celular Tumoral , Movimento Celular , Sobrevivência Celular , Modelos Animais de Doenças , Resistencia a Medicamentos Antineoplásicos , Proteínas da Matriz Extracelular/metabolismo , Feminino , Fibronectinas/metabolismo , Xenoenxertos , Humanos , Melanoma/tratamento farmacológico , Melanoma/patologia , Proteínas Quinases Ativadas por Mitógeno/antagonistas & inibidores , Metástase Neoplásica , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/uso terapêutico , Tenascina/metabolismo , Microambiente Tumoral
20.
Cell Rep ; 13(10): 2147-58, 2015 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-26628377

RESUMO

Primary prostate cancer almost always has a luminal phenotype. However, little is known about the stem/progenitor properties of transformed cells within tumors. Using the aggressive Pten/Tp53-null mouse model of prostate cancer, we show that two classes of luminal progenitors exist within a tumor. Not only did tumors contain previously described multipotent progenitors, but also a major population of committed luminal progenitors. Luminal cells, sorted directly from tumors or grown as organoids, initiated tumors of adenocarcinoma or multilineage histological phenotypes, which is consistent with luminal and multipotent differentiation potentials, respectively. Moreover, using organoids we show that the ability of luminal-committed progenitors to self-renew is a tumor-specific property, absent in benign luminal cells. Finally, a significant fraction of luminal progenitors survived in vivo castration. In all, these data reveal two luminal tumor populations with different stem/progenitor cell capacities, providing insight into prostate cancer cells that initiate tumors and can influence treatment response.


Assuntos
Adenocarcinoma/patologia , Células-Tronco Neoplásicas/patologia , Neoplasias da Próstata/patologia , Animais , Linhagem da Célula , Separação Celular , Modelos Animais de Doenças , Células Epiteliais/patologia , Citometria de Fluxo , Masculino , Camundongos , Camundongos Endogâmicos NOD , Camundongos Knockout , Camundongos SCID , Organoides , Fenótipo
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