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BACKGROUND: Recessive dystrophic epidermolysis bullosa (RDEB) is a blistering disease caused by mutations in the gene encoding type VII collagen (C7). RDEB is associated with fibrosis, which is responsible for severe complications. The phenotypic variability observed in RDEB siblings suggests that epigenetic modifications contribute to disease severity. Identifying epigenetic changes may help to uncover molecular mechanisms underlying RDEB pathogenesis and new therapeutic targets. OBJECTIVES: To investigate histone acetylation in RDEB skin and to explore histone deacetylase inhibitors (HDACis) as therapeutic molecules capable of counteracting fibrosis and disease progression in RDEB mice. METHODS: Acetylated histone levels were detected in human skin by immunofluorescence and in RDEB fibroblasts by ELISA. The effects of Givinostat and valproic acid (VPA) on RDEB fibroblast fibrotic behaviour were assessed by collagen-gel contraction assay, Western blot and immunocytofluorescence for α-smooth muscle actin, ELISA for released transforming growth factor-ß1 (TGF-ß1). RNA-seq was performed in HDACi- and vehicle-treated RDEB fibroblasts. VPA was systemically administered to RDEB mice, and effects on overt phenotype were monitored. Fibrosis was investigated in the skin using histological and immunofluorescence analyses. Eye and tongue defects were examined microscopically. Mass spectrometry proteomics was performed on skin protein extracts from VPA-treated RDEB and control mice. RESULTS: Histone acetylation decreases in RDEB skin and primary fibroblasts. RDEB fibroblasts treated with HDACis lowered fibrotic traits including contractility, TGF-ß1 release, and proliferation. VPA administration to RDEB mice mitigated severe manifestations affecting eyes and paws. These effects were associated with fibrosis inhibition. Proteomic analysis of mouse skin revealed that VPA almost normalised protein sets involved in protein synthesis and immune response, processes linked to the increased susceptibility to cancer and bacterial infections observed in RDEB patients. CONCLUSIONS: Dysregulated histone acetylation contributes to RDEB pathogenesis by facilitating the progression of fibrosis. Repurposing of HDACi could be considered for disease-modifying treatments of RDEB.
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BACKGROUND & AIMS: About 15% of intrahepatic cholangiocarcinomas (iCCAs) express fibroblast growth factor receptor 2 (FGFR2) fusion proteins (FFs), usually alongside mutational inactivation of TP53, CDKN2A or BAP1. In FFs, FGFR2 residues 1-768 fuse to sequences encoded by a diverse array of partner genes (>60) causing oncogenic FF activation. While FGFR-specific tyrosine kinase inhibitors (F-TKI) provide clinical benefit in FF+ iCCA, responses are partial and/or limited by resistance mechanisms, such as the V565F substitution in the FGFR2 gatekeeper residue. Improving on FF targeting in iCCA therefore remains a critical unmet need. Herein, we aimed to generate a murine model of FF-driven iCCA and use this to uncover actionable FF-associated dependencies. METHODS: Four iCCA FFs carrying different fusion sequences were expressed in Tp53-/- mouse liver organoids. Tumorigenic properties of genetically modified liver organoids were assessed by transplantation into immuno-deficient mice. Cellular models derived from neoplastic lesions were exploited for pre-clinical studies. RESULTS: Transplantation of FF-expressing liver organoids yielded tumors diagnosed as CCA based on histological, phenotypic and transcriptomic analyses. The penetrance of this tumorigenic phenotype was influenced by FF identity. Tumor organoids and 2D cell lines derived from CCA lesions were addicted to FF signaling via Ras-Erk, regardless of FF identity or V565F mutation. Dual blockade of FF and the Ras-Erk pathway by concomitant pharmacological inhibition of FFs and Mek1/2 provided greater therapeutic efficacy than single agent F-TKI in vitro and in vivo. CONCLUSIONS: FF-driven iCCA pathogenesis was successfully modeled on a Tp53-/- murine background, revealing biological heterogeneity among structurally different FFs. Double blockade of FF-ERK signaling deserves consideration for precision-based approaches against human FF+ iCCA. LAY SUMMARY: Intrahepatic cholangiocarcinoma (iCCA) is a rare cancer that is difficult to treat. A subtype of iCCA is caused by genomic alterations that generate oncogenic drivers known as FGFR2 fusions. Patients with FGFR2 fusions respond to FGFR inhibitors, but clinical responses are often of modest duration. We used animal and cellular models to show that FGFR2 fusions require the activity of a downstream effector named Mek1/2. We found that dual blockade of FGFR2 fusions and Mek1/2 was more effective than isolated inhibition of FGFR2 fusions, pointing to the potential clinical utility of dual FGFR2-MEK1/2 blockade in patients with iCCA.
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Colangiocarcinoma/etiología , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/antagonistas & inhibidores , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/genética , Proteína p53 Supresora de Tumor/efectos de los fármacos , Análisis de Varianza , Animales , Línea Celular/metabolismo , Colangiocarcinoma/genética , Modelos Animales de Enfermedad , Ratones , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/metabolismo , Transducción de Señal/efectos de los fármacosRESUMEN
Hepatitis C virus (HCV) causes B-cell lymphoproliferative disorders (LPDs) expressing stereotyped B-cell receptors (BCRs) endowed with rheumatoid factor (RF) activity and putatively recognizing the HCV E2 protein. To further untangle the shaping and function of these BCRs, we analyzed immunoglobulin gene rearrangements of monoclonal B cells from 13 patients with HCV-associated LPDs and correlated their features with the clinical outcomes of antiviral therapy. While only two patients shared a stereotyped heavy-chain complementarity determining region 3 (CDR3) sequence, two kappa chain CDR3 stereotyped sequences accounted for 77% of BCRs. Light chains were enriched in sequences homologous to anti-HCV E2 antibodies compared with heavy chains (7/13 vs. 0/13; p = 0.005). Anti-HCV E2 homology was uniquely associated (7/7 vs. 0/6; p = 0.0006) with a stereotyped CDR3 sequence encoded by IGKV3-20/3D-20 gene(s) accounting for 54% of BCRs. An IGKV3-15/IGKJ1-encoded stereotyped sequence homologous to WA RF accounted for 23% of BCRs. LPDs expressing KCDR3s homologous to anti-HCV E2 antibodies responded more frequently to the eradication of HCV by antiviral therapy (6/6 vs. 1/6; p = 0.015). These findings, although limited by the small sample size, suggest that a stereotyped KCDR3 may predominantly shape anti-HCV specificity of BCRs, possibly providing a signature that may help identifying bona fide HCV-dependent LPDs.
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Cadenas Ligeras de Inmunoglobulina/genética , Receptores de Antígenos de Linfocitos B/genética , Anciano , Secuencia de Aminoácidos/genética , Linfocitos B/metabolismo , Regiones Determinantes de Complementariedad/genética , Femenino , Hepacivirus/genética , Hepacivirus/patogenicidad , Hepatitis C/complicaciones , Humanos , Cadenas Ligeras de Inmunoglobulina/metabolismo , Región Variable de Inmunoglobulina/genética , Región Variable de Inmunoglobulina/inmunología , Trastornos Linfoproliferativos , Masculino , Persona de Mediana Edad , Receptores de Antígenos de Linfocitos B/metabolismo , Factor Reumatoide/inmunologíaRESUMEN
Sézary syndrome (SS) is an incurable leukemic variant of cutaneous T-cell lymphoma characterized by recurrent chromosomal alterations, among which, chromosome 10q deletion is very frequent. In this study, we investigated the PTEN status, on locus 10q23, in 44 SS patients; our findings show that PTEN is deleted in 36% of SS cases, whereas PTEN downregulation is observed in almost all of the samples evaluated by quantitative reverse-transcriptase polymerase chain reaction and Western blotting analysis. Neither DNA sequence mutation nor promoter hypermethylation were found at the PTEN locus, but we demonstrate that PTEN level can be also reduced by a group of miRs previously found upregulated and of prognostic relevance in SS; particularly, miR-21, miR-106b, and miR-486 were able to control PTEN abundance either in vitro or in vivo. Finally, because reduced PTEN activates the PI3/AKT-mediated pathway of cell growth and survival, we demonstrate that PTEN deficiency is associated with activated AKT in skin resident but not circulating SS cells, suggesting that the cutaneous milieu may strongly contribute to the SS cell growth. To our knowledge, this is the first study fully exploring the PTEN status in a large cohort of SS patients, unveiling potential elements of clinical utility in this malignancy.
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Regulación Neoplásica de la Expresión Génica , Proteínas de Neoplasias/fisiología , Fosfohidrolasa PTEN/fisiología , Síndrome de Sézary/metabolismo , Anciano , Anciano de 80 o más Años , Cromosomas Humanos Par 10/ultraestructura , Metilación de ADN , Análisis Mutacional de ADN , Regulación hacia Abajo , Femenino , Eliminación de Gen , Dosificación de Gen , Humanos , Masculino , MicroARNs/genética , Persona de Mediana Edad , Proteínas de Neoplasias/análisis , Proteínas de Neoplasias/genética , Fosfohidrolasa PTEN/análisis , Fosfohidrolasa PTEN/genética , Fosfatidilinositol 3-Quinasas/fisiología , Proteínas Proto-Oncogénicas c-akt/fisiología , ARN Mensajero/biosíntesis , ARN Neoplásico/biosíntesis , Análisis de Secuencia de ADN , Síndrome de Sézary/genética , Transducción de Señal , Piel/metabolismo , Piel/patologíaRESUMEN
A clonal population of B cells expressing a V(H) 1-69-encoded idiotype accumulates in hepatitis C virus (HCV) associated mixed cryoglobulinemia (MC). These cells are phenotypically heterogeneous, resembling either typical marginal zone (MZ) B cells (IgM(+) IgD(+) CD27(+) CD21(+) ) or the exhausted CD21(low) B cells that accumulate in HIV infection or in common variable immunodeficiency. We show that both the MZ-like and the CD21(low) V(H) 1-69(+) B cells of MC patients are functionally exhausted, since they fail to respond to TLR and BCR ligands. The proliferative defect of V(H) 1-69(+) B cells can be overcome by co-stimulation of TLR9 and BCR in the presence of interleukin(IL)-2 and IL-10. The MZ-like V(H) 1-69(+) B cells do not express the inhibitory receptors distinctive of CD21(low) B cells, but display constitutive activation of extracellular signal regulated kinase (ERK) and attenuated BCR/ERK signaling. These cells also express abundant transcripts of Stra13 (DEC1, Bhlhb2, Sharp2, Clast5), a basic helix-loop-helix transcription factor that acts as a powerful negative regulator of B-cell proliferation and homeostasis. Our findings suggest that MZ B cells activated by HCV undergo functional exhaustion associated with BCR signaling defects and overexpression of a key antiproliferative gene, and may subsequently become terminally spent CD21(low) B cells. Premature exhaustion may serve to prevent the outgrowth of chronically stimulated MZ B cells.
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Linfocitos B/inmunología , Crioglobulinemia/inmunología , Proteínas de Unión al ADN/fisiología , Hepatitis C/complicaciones , Proteínas Nucleares/fisiología , Receptores de Complemento 3d/análisis , Adulto , Anciano , Anciano de 80 o más Años , Proteínas de Unión al ADN/análisis , Femenino , Humanos , Activación de Linfocitos , Masculino , Persona de Mediana Edad , Proteínas Nucleares/análisis , Fenotipo , Receptores de Antígenos de Linfocitos B/fisiología , Transducción de Señal , Receptor Toll-Like 9/fisiologíaRESUMEN
Sézary syndrome (SS) is an aggressive variant of cutaneous t-cell lymphoma characterized by the accumulation of neoplastic CD4+ lymphocytes-the SS cells-mainly in blood, lymph nodes, and skin. The tumor spread pattern of SS makes this lymphoma a unique model of disease that allows a concurrent blood and skin sampling for analysis. This review summarizes the recent studies highlighting the transcriptional programs triggered by the crosstalk between SS cells and blood-skin microenvironments. Emerging data proved that skin-derived SS cells show consistently higher activation/proliferation rates, mainly driven by T-cell receptor signaling with respect to matched blood SS cells that instead appear quiescent. Biochemical analyses also demonstrated an hyperactivation of PI3K/AKT/mTOR, a targetable pathway by multiple inhibitors currently in clinical trials, in skin SS cells compared with a paired blood counterpart. These results indicated that active and quiescent SS cells coexist in this lymphoma, and that they could be respectively treated with different therapeutics. Finally, this review underlines the more recent discoveries into the heterogeneity of circulating SS cells, highlighting a series of novel markers that could improve the diagnosis and that represent novel therapeutic targets (GPR15, PTPN13, KLRB1, and ITGB1) as well as new genetic markers (PD-1 and CD39) able to stratify SS patients for disease aggressiveness.
RESUMEN
Sézary syndrome (SS) is a rare and aggressive variant of cutaneous T-cell lymphoma. It is characterized by the copresence of CD4+ neoplastic lymphocytes, named Sezary cells, mainly in the blood, lymph nodes, and skin where they induce chronic inflammation that in turn impairs the patient's QOL and fuels neoplastic cells. SS is not readily cured, but immunotherapy is becoming an effective option for this lymphoma. In this study, we investigated, in a large cohort of patients with SS, the expression and function of the immune checkpoint molecule CD39, which degrades proinflammatory extracellular adenosine triphosphate. We showed that the SNP rs10748643 A/G within the ENTPD1 gene coding for the CD39 protein controls its expression level. Patients carrying the A/GâG/G genotype showed a significantly higher frequency of clonal CD4+CD39+ SS cells than those carrying the A/A genotype. Different from other cancers, high CD39 expression correlates with a better prognosis. Comparing primary G/G with A/A lymphoma cells, we observed that G/G SS cells have a higher ability to degrade adenosine triphosphate, increased apoptotic susceptibility, and upon activation, reduced IL-2 production. Accordingly, CD39 enzymatic inhibition enhances SS cell viability and IL-2 production on activation. These results strongly suggest a special caution for SS treatment with therapeutic inhibitors of CD39.
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Apirasa , Síndrome de Sézary , Neoplasias Cutáneas , Humanos , Adenosina Trifosfato/metabolismo , Apirasa/genética , Supervivencia Celular/genética , Proteínas de Punto de Control Inmunitario , Interleucina-2/genética , Linfocitos/metabolismo , Pronóstico , Calidad de Vida , Síndrome de Sézary/genética , Síndrome de Sézary/metabolismo , Neoplasias Cutáneas/genética , Neoplasias Cutáneas/metabolismo , Linfocitos T ReguladoresRESUMEN
The phosphoinositide 3-kinase(PI3K)/protein kinase B (AKT)/ mammalian target of rapamycin (mTOR) pathway is hyperactivated in many tumors, as well as in cutaneous T-cell lymphoma (CTCL), which includes the mycosis fungoides and the aggressive variant known as Sezary syndrome (SS). TORC1 signaling is activated in SS cells by cytokines and chemokines, which are overexpressed in SS tissues. Furthermore, the recurrent copy number variation of genes belonging to this cascade, such as PTEN, LKB1, and P70S6K, contributes to the hyperactivation of the pathway. The aim of this study was to investigate the therapeutic potential of mTOR inhibitors in CTCL. We compared the efficacy of three rapalogs (rapamycin, temsirolimus, and everolimus) and the dual-mTOR/PI3K inhibitor PF-04691502 (hereinafter PF-502) in four CTCL cell lines. PF-502 was revealed to be the most effective inhibitor of cell growth. Interestingly, PF-502 also exerted its antitumor activity in patient-derived CTCL cells and in a xenograft mouse model, where it induced significant apoptosis and increased survival of treated mice. Furthermore, we found an inverse correlation between PTEN gene expression and the ability of PF-502 to induce apoptosis in SS cells. Our data strongly support the therapeutic potential of dual PI3K/mTOR inhibitors in CTCL.
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Antineoplásicos/uso terapéutico , Linfoma Cutáneo de Células T/tratamiento farmacológico , Inhibidores de Proteínas Quinasas/uso terapéutico , Piridonas/uso terapéutico , Pirimidinas/uso terapéutico , Linfocitos T/inmunología , Animales , Línea Celular , Evaluación Preclínica de Medicamentos , Everolimus/uso terapéutico , Femenino , Humanos , Diana Mecanicista del Complejo 1 de la Rapamicina/metabolismo , Ratones , Ratones Desnudos , Terapia Molecular Dirigida , Fosfatidilinositol 3-Quinasas/metabolismo , Transducción de Señal , Sirolimus/análogos & derivados , Sirolimus/uso terapéutico , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Memory B cells (MBCs) epitomize the adaptation of the immune system to the environment. We identify two MBC subsets in peripheral blood, CD27dull and CD27bright MBCs, whose frequency changes with age. Heavy chain variable region (VH) usage, somatic mutation frequency replacement-to-silent ratio, and CDR3 property changes, reflecting consecutive selection of highly antigen-specific, low cross-reactive antibody variants, all demonstrate that CD27dull and CD27bright MBCs represent sequential MBC developmental stages, and stringent antigen-driven pressure selects CD27dull into the CD27bright MBC pool. Dynamics of human MBCs are exploited in pregnancy, when 50% of maternal MBCs are lost and CD27dull MBCs transit to the more differentiated CD27bright stage. In the postpartum period, the maternal MBC pool is replenished by the expansion of persistent CD27dull clones. Thus, the stability and flexibility of human B cell memory is ensured by CD27dull MBCs that expand and differentiate in response to change.
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Linfocitos B/inmunología , Memoria Inmunológica , Miembro 7 de la Superfamilia de Receptores de Factores de Necrosis Tumoral/metabolismo , Adulto , Anciano , Anciano de 80 o más Años , Niño , Preescolar , Femenino , Perfilación de la Expresión Génica , Humanos , Cambio de Clase de Inmunoglobulina/genética , Región Variable de Inmunoglobulina/genética , Memoria Inmunológica/genética , Lactante , Recién Nacido , Persona de Mediana Edad , Modelos Inmunológicos , Embarazo , Hipermutación Somática de Inmunoglobulina/genética , Donantes de Tejidos , Transcripción GenéticaRESUMEN
Sezary syndrome (SS) is a primary cutaneous T-cell lymphoma (CTCL) characterized by erythroderma, lymphadenopathy and leukemic involvement of the peripheral blood. The high relapse rates and a poor prognosis complicate its clinical course and treatment. The phenotypic characterization and genomic/transcriptomic approaches revealed high heterogeneity of Sezary cells, identifying a wide spectrum of biomarkers implicated in the development of this lymphoma. In this context, we discuss the major malignancy-related biomarkers reported in the literature for the diagnosis, prognosis and staging of SS. The hope for a single reliable diagnostic marker appears increasingly unrealistic, but the discovery of multiple potential biomarkers, with pathogenetic implications, paves the road to promising personalized therapies in SS.
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Biomarcadores/química , Síndrome de Sézary/tratamiento farmacológico , Humanos , Pronóstico , Síndrome de Sézary/patologíaRESUMEN
Sezary Syndrome is an aggressive T-cell Lymphoma involving blood, skin and lymphonodes Involvement of the CXCR4-SDF1 has been previously shown. We here present evidence also of the involvement of B-arrestin a downstream regulator of CXCR4, that is depleted and downregulated as well as a potential functional role for this depletion.
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Linfoma de Células T/genética , Síndrome de Sézary/genética , Neoplasias Cutáneas/genética , Arrestina beta 2/genética , Arrestina beta 2/fisiología , Células Cultivadas , Estudios de Cohortes , Variaciones en el Número de Copia de ADN , Eliminación de Gen , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Linfoma de Células T/patología , Invasividad Neoplásica , ARN Interferente Pequeño/farmacología , Receptores CXCR4/genética , Síndrome de Sézary/patología , Neoplasias Cutáneas/patología , Arrestina beta 2/antagonistas & inhibidoresRESUMEN
Sézary syndrome (SS) is a rare and aggressive variant of Cutaneous T-Cell Lymphoma characterized by neoplastic distribution mainly involving blood, skin, and lymph-node. Although a role of the skin microenvironment in SS pathogenesis has long been hypothesized, its function in vivo is poorly characterized. To deepen this aspect, here we compared skin to blood-derived SS cells concurrently obtained from SS patients highlighting a greater proliferation-index and a PI3K/AKT/mTORC1 pathway activation level, particularly of mTOR protein, in skin-derived-SS cells. We proved that SDF-1 and CCL21 chemokines, both overexpressed in SS tissues, induce mTORC1 signaling activation, cell proliferation and Ki67 up-regulation in a SS-derived cell line and primary-SS cells. In a cohort of 43 SS cases, we observed recurrent copy number variations (CNV) of members belonging to this cascade, namely: loss of LKB1 (48%), PTEN (39%) and PDCD4 (35%) and gains of P70S6K (30%). These alterations represent druggable targets unraveling new therapeutic treatments as metformin here evaluated in vitro. Moreover, CNV of PTEN, PDCD4, and P70S6K, evaluated individually or in combination, are associated with reduced survival of SS patients. These data shed light on effects in vivo of skin-SS cells interaction underlying the prognostic and therapeutic relevance of mTORC1 pathway in SS.
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Diana Mecanicista del Complejo 1 de la Rapamicina/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Síndrome de Sézary/metabolismo , Síndrome de Sézary/patología , Transducción de Señal , Neoplasias Cutáneas/metabolismo , Neoplasias Cutáneas/patología , Biomarcadores , Línea Celular Tumoral , Movimiento Celular/efectos de los fármacos , Proliferación Celular , Variaciones en el Número de Copia de ADN , Humanos , Inmunohistoquímica , Inmunofenotipificación , Metformina/farmacología , Modelos Biológicos , Células Neoplásicas Circulantes/metabolismo , Células Neoplásicas Circulantes/patología , Pronóstico , Síndrome de Sézary/mortalidad , Neoplasias Cutáneas/mortalidadRESUMEN
T Cell Leukemia/Lymphoma 1A is expressed during B-cell differentiation and, when over-expressed, acts as an oncogene in mouse (Tcl1a) and human (TCL1A) B-cell chronic lymphocytic leukemia (B-CLL) and T-cell prolymphocytic leukemia (T-PLL). Furthermore, in the murine system Tcl1a is expressed in the ovary, testis and in pre-implantation embryos, where it plays an important role in blastomere proliferation and in embryonic stem cell (ESC) proliferation and self-renewal. We have also observed that Tcl1-/- adult mice exhibit alopecia and deep ulcerations. This finding has led us to investigate the role of TCL1 in mouse skin and hair follicles. We have found that TCL1 is expressed in the proliferative structure (i.e. the secondary hair germ) and in the stem cell niche (i.e. the bulge) of the hair follicle during regeneration phase and it is constitutively expressed in the basal layer of epidermis where it is required for the correct proliferative-differentiation program of the keratinocytes (KCs). Taking advantage of the murine models we have generated, including the Tcl1-/- and the K14-TCL1 transgenic mouse, we have analysed the function of TCL1 in mouse KCs and the molecular pathways involved. We provide evidence that in the epidermal compartment TCL1 has a role in the regulation of KC proliferation, differentiation, and apoptosis. In particular, the colony-forming efficiency (CFE) and the insulin-like growth factor 1 (IGF1)-induced proliferation are dramatically impaired, while apoptosis is increased, in KCs from Tcl1-/- mice when compared to WT. Moreover, the expression of differentiation markers such as cytokeratin 6 (KRT6), filaggrin (FLG) and involucrin (IVL) are profoundly altered in mutant mice (Tcl1-/-). Importantly, by over-expressing TCL1A in basal KCs of the K14-TCL1 transgenic mouse model, we observed a significant rescue of cell proliferation, differentiation and apoptosis of the mutant phenotype. Finally, we found TCL1 to act, at least in part, via increasing phospho-ERK1/2 and decreasing phospho-P38 MAPK. Hence, our data demonstrate that regulated levels of Tcl1a are necessary for the correct proliferation and differentiation of the interfollicular KCs.
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Proliferación Celular/fisiología , Factor I del Crecimiento Similar a la Insulina/metabolismo , Queratinocitos/metabolismo , Proteínas Proto-Oncogénicas/metabolismo , Animales , Apoptosis/fisiología , Diferenciación Celular/fisiología , Modelos Animales de Enfermedad , Epidermis/metabolismo , Epidermis/patología , Proteínas Filagrina , Regulación Neoplásica de la Expresión Génica/fisiología , Folículo Piloso/metabolismo , Folículo Piloso/patología , Queratinocitos/patología , Leucemia Linfocítica Crónica de Células B/metabolismo , Leucemia Linfocítica Crónica de Células B/patología , Sistema de Señalización de MAP Quinasas/fisiología , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Piel/metabolismo , Piel/patología , Nicho de Células Madre/fisiología , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismoRESUMEN
Cutaneous T-cell lymphoma is a group of incurable extranodal non-Hodgkin lymphomas that develop from the skin-homing CD4+ T cell. Mycosis fungoides and Sézary syndrome are the most common histological subtypes. Although next-generation sequencing data provided significant advances in the comprehension of the genetic basis of this lymphoma, there is not uniform consensus on the identity and prevalence of putative driver genes for this heterogeneous group of tumors. Additional studies may increase the knowledge about the complex genetic etiology characterizing this lymphoma. We used SNP6 arrays and GISTIC algorithm to prioritize a list of focal somatic copy-number alterations in a dataset of multiple sequential samples from 21 Sézary syndrome patients. Our results confirmed a prevalence of significant focal deletions over amplifications: single well-known tumor suppressors, such as TP53, PTEN, and RB1, are targeted by these aberrations. In our cohort, ZEB1 (TCF8, ZFHX1A) spans a deletion having the highest level of significance. In a larger group of 43 patients, we found that ZEB1 is affected by deletions and somatic inactivating mutations in 46.5% of cases; also, we found potentially relevant ZEB1 germline variants. The survival analysis shows a worse clinical course for patients with ZEB1 biallelic inactivation. Multiple abnormal expression signatures were found associated with ZEB1 depletion in Sézary patients we verified that ZEB1 exerts a role in oxidative response of Sézary cells. Our data confirm the importance of deletions in the pathogenesis of cutaneous T-cell lymphoma. The characterization of ZEB1 abnormalities in Sézary syndrome fulfils the criteria of a canonical tumor suppressor gene. Although additional confirmations are needed, our findings suggest, for the first time, that ZEB1 germline variants might contribute to the risk of developing this disease. Also, we provide evidence that ZEB1 activity in Sézary cells, influencing the reactive oxygen species production, affects cell viability and apoptosis.
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Regulación Neoplásica de la Expresión Génica , Predisposición Genética a la Enfermedad , Mutación de Línea Germinal , Síndrome de Sézary/genética , Neoplasias Cutáneas/genética , Homeobox 1 de Unión a la E-Box con Dedos de Zinc/genética , Apoptosis , Línea Celular Tumoral , Supervivencia Celular , Variaciones en el Número de Copia de ADN , Humanos , Análisis de Secuencia por Matrices de Oligonucleótidos , Fosfohidrolasa PTEN/genética , Fosfohidrolasa PTEN/inmunología , Polimorfismo de Nucleótido Simple , Especies Reactivas de Oxígeno/inmunología , Especies Reactivas de Oxígeno/metabolismo , Proteínas de Unión a Retinoblastoma/genética , Proteínas de Unión a Retinoblastoma/inmunología , Síndrome de Sézary/inmunología , Síndrome de Sézary/mortalidad , Síndrome de Sézary/patología , Piel/inmunología , Piel/patología , Neoplasias Cutáneas/inmunología , Neoplasias Cutáneas/mortalidad , Neoplasias Cutáneas/patología , Análisis de Supervivencia , Linfocitos T/inmunología , Linfocitos T/patología , Proteína p53 Supresora de Tumor/genética , Proteína p53 Supresora de Tumor/inmunología , Ubiquitina-Proteína Ligasas/genética , Ubiquitina-Proteína Ligasas/inmunología , Homeobox 1 de Unión a la E-Box con Dedos de Zinc/deficiencia , Homeobox 1 de Unión a la E-Box con Dedos de Zinc/inmunologíaRESUMEN
The transcription factor DEC1/STRA13 (also known as BHLHE40 and SHARP2) is involved in a number of processes including inhibition of cell proliferation and delay of cell cycle, and is a negative regulator of B cell activation and development in mice. We show here that, unlike in mice, DEC1/STRA13 expression is induced in human naïve and memory resting B cells by activation through the B-cell receptor (BCR) or Toll-like receptor 9 (TLR9). siRNA silencing of DEC1/STRA13 increases the capacity of activated B cells to perform a high number of divisions after TLR9 ligation. This identifies DEC1/STRA13 as a critical negative regulator of clonal expansion of activated human B cells. We also show that DEC1/STRA13 is upregulated in human anergic CD21low B cells clonally expanded in patients with HCV-associated mixed cryoglobulinemia, which fail to proliferate in response to BCR or TLR9 ligation. siRNA knockdown of DEC1/STRA13, however, fails to restore responsiveness to stimuli in these cells, although it might improve the proliferative capacity in a subset of anergic cells with less pronounced proliferative defect.
Asunto(s)
Linfocitos B/inmunología , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Anergia Clonal , Proteínas de Homeodominio/metabolismo , Activación de Linfocitos , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Ciclo Celular , Proliferación Celular , Células Cultivadas , Crioglobulinemia/genética , Crioglobulinemia/inmunología , Crioglobulinemia/patología , Regulación de la Expresión Génica/inmunología , Técnicas de Silenciamiento del Gen , Hepatitis C/genética , Hepatitis C/inmunología , Hepatitis C/patología , Proteínas de Homeodominio/genética , Humanos , ARN Interferente Pequeño/metabolismo , Receptores de Antígenos de Linfocitos B/metabolismo , Transducción de Señal , Receptor Toll-Like 9/metabolismoRESUMEN
Monoclonal marginal zone (MZ) B cells expressing a V(H)1-69-encoded idiotype accumulate in HCV-associated mixed cryoglobulinemia (MC). These cells recognize the E2 protein of HCV and their massive clonal expansion reflects the propensity of MZ B cells to proliferate robustly upon antigenic stimulation by microorganisms, a property that makes them prone to neoplastic transformation. V(H)1-69(+) B cells of MC patients are phenotypically heterogeneous and resemble either mature MZ B cells (IgM(+)CD27(+)CD21(high)) or the unusual CD21(low) B cells that accumulate in other immunological disorders such as common variable immunodeficiency (CVID) or HIV infection. The CD21(low) V(H)1-69(+) B cells of MC patients, like those of CVID and HIV patients, are anergic to BCR and TLR9 stimulation and display deregulation of several anergy-related genes; proliferative anergy is also observed in CD21(high) MZ-like V(H)1-69(+) B cells, that over-express the antiproliferative transcriptional repressor Stra13. Upon evolution to splenic marginal zone lymphoma, MZ-like V(H)1-69(+) B cells down-regulate Stra13 and partially recover their capacity to proliferate in response to TLR9 ligation. Like yin and yang, robust clonal expansion and early proliferative anergy may be viewed as the opposite forces balancing the responses of human MZ B cells to chronic microbial stimuli. Disruption of this balance facilitates autoimmunity and lymphoproliferation.
Asunto(s)
Autoinmunidad , Linfocitos B/citología , Linfocitos B/inmunología , Crioglobulinemia/inmunología , Crioglobulinemia/virología , Hepacivirus/inmunología , Activación de Linfocitos , Crioglobulinemia/genética , Perfilación de la Expresión Génica , Regulación de la Expresión Génica , Hepatitis C/inmunología , Hepatitis C/patología , Humanos , Cadenas Pesadas de Inmunoglobulina/genética , Cadenas Pesadas de Inmunoglobulina/inmunología , Región Variable de Inmunoglobulina/genética , Región Variable de Inmunoglobulina/inmunología , Receptores de Complemento 3d/metabolismo , Bazo/inmunologíaRESUMEN
In this study, we used single nucleotide polymorphism and comparative genomic hybridization array to study DNA copy number changes and loss of heterozygosity for 28 patients affected by Sézary syndrome (SS), a rare form of cutaneous T-cell lymphoma (CTCL). Our data identified, further confirming previous studies, recurrent losses of 17p13.2-p11.2 and 10p12.1-q26.3 occurring in 71% and 68% of cases, respectively; common gains were detected for 17p11.2-q25.3 (64%) and chromosome 8/8q (50%). Moreover, we identified novel genomic lesions recurring in >30% of tumors: loss of 9q13-q21.33 and gain of 10p15.3-10p12.2. Individual chromosomal aberrations did not show a significant correlation with prognosis; however, when more than three recurrent chromosomal alterations (gain or loss) were considered, a statistical association was observed using Kaplan-Meier survival analysis. Integrating mapping and transcriptional data, we were able to identify a total of 113 deregulated transcripts in aberrant chromosomal regions that included cancer-related genes such as members of the NF-kappaB pathway (BAG4, BTRC, NKIRAS2, PSMD3, and TRAF2) that might explain its constitutive activation in CTCL. Matching this list of genes with those discriminating patients with different survival times, we identify several common candidates that might exert critical roles in SS, such as BUB3 and PIP5K1B. Altogether, our study confirms and maps more precisely the regions of gain and loss and, combined to transcriptional profiles, suggests a novel set of genes of potential interest in SS.