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Sistemas CRISPR-Cas , Edición Génica , Edición Génica/métodos , Animales , Humanos , Sistemas CRISPR-Cas/genéticaRESUMEN
BACKGROUND: Native bone marrow (BM) mesenchymal stem/stromal cells (BM-MSCs) participate in generating and shaping the skeleton and BM throughout the lifespan. Moreover, BM-MSCs regulate hematopoiesis by contributing to the hematopoietic stem cell niche in providing critical cytokines, chemokines and extracellular matrix components. However, BM-MSCs contain a heterogeneous cell population that remains ill-defined. Although studies on the taxonomy of native BM-MSCs in mice have just started to emerge, the taxonomy of native human BM-MSCs remains unelucidated. METHODS: By using single-cell RNA sequencing (scRNA-seq), we aimed to define a proper taxonomy for native human BM non-hematopoietic subsets including endothelial cells (ECs) and mural cells (MCs) but with a focal point on MSCs. To this end, transcriptomic scRNA-seq data were generated from 5 distinct BM donors and were analyzed together with other transcriptomic data and with computational biology analyses at different levels to identify, characterize and classify distinct native cell subsets with relevant biomarkers. RESULTS: We could ascribe novel specific biomarkers to ECs, MCs and MSCs. Unlike ECs and MCs, MSCs exhibited an adipogenic transcriptomic pattern while co-expressing genes related to hematopoiesis support and multilineage commitment potential. Furthermore, by a comparative analysis of scRNA-seq of BM cells from humans and mice, we identified core genes conserved in both species. Notably, we identified MARCKS, CXCL12, PDGFRA, and LEPR together with adipogenic factors as archetypal biomarkers of native MSCs within BM. In addition, our data suggest some complex gene nodes regulating critical biological functions of native BM-MSCs together with a preferential commitment toward an adipocyte lineage. CONCLUSIONS: Overall, our taxonomy for native BM non-hematopoietic compartment provides an explicit depiction of gene expression in human ECs, MCs and MSCs at single-cell resolution. This analysis helps enhance our understanding of the phenotype and the complexity of biological functions of native human BM-MSCs.
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Células Endoteliales , Células Madre Mesenquimatosas , Humanos , Animales , Ratones , Células de la Médula Ósea , Biomarcadores , Análisis de Secuencia de ARNRESUMEN
Attaining personalized healthy aging requires accurate monitoring of physiological changes and identifying subclinical markers that predict accelerated or delayed aging. Classic biostatistical methods most rely on supervised variables to estimate physiological aging and do not capture the full complexity of inter-parameter interactions. Machine learning (ML) is promising, but its black box nature eludes direct understanding, substantially limiting physician confidence and clinical usage. Using a broad population dataset from the National Health and Nutrition Examination Survey (NHANES) study including routine biological variables and after selection of XGBoost as the most appropriate algorithm, we created an innovative explainable ML framework to determine a Personalized physiological age (PPA). PPA predicted both chronic disease and mortality independently of chronological age. Twenty-six variables were sufficient to predict PPA. Using SHapley Additive exPlanations (SHAP), we implemented a precise quantitative associated metric for each variable explaining physiological (i.e., accelerated or delayed) deviations from age-specific normative data. Among the variables, glycated hemoglobin (HbA1c) displays a major relative weight in the estimation of PPA. Finally, clustering profiles of identical contextualized explanations reveal different aging trajectories opening opportunities to specific clinical follow-up. These data show that PPA is a robust, quantitative and explainable ML-based metric that monitors personalized health status. Our approach also provides a complete framework applicable to different datasets or variables, allowing precision physiological age estimation.
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Algoritmos , Estado de Salud , Encuestas Nutricionales , Aprendizaje AutomáticoRESUMEN
Macrophages (MΦ) are increasingly recognized as HIV-1 target cells involved in the pathogenesis and persistence of infection. Paradoxically, in vitro infection assays suggest that virus isolates are mostly T-cell-tropic and rarely MΦ-tropic. The latter are assumed to emerge under CD4+ T-cell paucity in tissues such as the brain or at late stage when the CD4 T-cell count declines. However, assays to qualify HIV-1 tropism use cell-free viral particles and may not fully reflect the conditions of in vivo MΦ infection through cell-to-cell viral transfer. Here, we investigated the capacity of viruses expressing primary envelope glycoproteins (Envs) with CCR5 and/or CXCR4 usage from different stages of infection, including transmitted/founder Envs, to infect MΦ by a cell-free mode and through cell-to-cell transfer from infected CD4+ T cells. The results show that most viruses were unable to enter MΦ as cell-free particles, in agreement with the current view that non-M-tropic viruses inefficiently use CD4 and/or CCR5 or CXCR4 entry receptors on MΦ. In contrast, all viruses could be effectively cell-to-cell transferred to MΦ from infected CD4+ T cells. We further showed that viral transfer proceeded through Env-dependent cell-cell fusion of infected T cells with MΦ targets, leading to the formation of productively infected multinucleated giant cells. Compared to cell-free infection, infected T-cell/MΦ contacts showed enhanced interactions of R5 M- and non-M-tropic Envs with CD4 and CCR5, resulting in a reduced dependence on receptor expression levels on MΦ for viral entry. Altogether, our results show that virus cell-to-cell transfer overcomes the entry block of isolates initially defined as non-macrophage-tropic, indicating that HIV-1 has a more prevalent tropism for MΦ than initially suggested. This sheds light into the role of this route of virus cell-to-cell transfer to MΦ in CD4+ T cell rich tissues for HIV-1 transmission, dissemination and formation of tissue viral reservoirs.
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Infecciones por VIH , VIH-1 , Antígenos CD4/metabolismo , Linfocitos T CD4-Positivos , Infecciones por VIH/metabolismo , VIH-1/metabolismo , Humanos , Macrófagos/metabolismo , Receptores CCR5/metabolismo , Internalización del VirusRESUMEN
The consistency of cerebrospinal fluid amyloid-ß (Aß)42/40 ratio and Aß42 has not been assessed in the AT(N) classification system. We analyzed the classification changes of the dichotomized amyloid status (A+/A-) in 363 patients tested for Alzheimer's disease biomarkers after Aß42 was superseded by the Aß42/40 ratio. The consistency of Aß42 and the Aß42/40 ratio was very low. Notably, the proportions of "false" A+T-patients were considerable (74-91%) and corresponded mostly to patients not clinically diagnosed with Alzheimer's disease. Our results suggest that the interchangeability of Aß42/40 ratio and Aß42 is limited for classifying patients in clinical setting using the AT(N) scheme.
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Péptidos beta-Amiloides/líquido cefalorraquídeo , Biomarcadores/líquido cefalorraquídeo , Disfunción Cognitiva/líquido cefalorraquídeo , Fragmentos de Péptidos/líquido cefalorraquídeo , Terminología como Asunto , Anciano , Enfermedad de Alzheimer/clasificación , Humanos , Proteínas tau/líquido cefalorraquídeoRESUMEN
Autophagy has been associated with oncogenesis with one of its emerging key functions being its contribution to the metabolism of tumors. Therefore, deciphering the mechanisms of how autophagy supports tumor cell metabolism is essential. Here, we demonstrate that the inhibition of autophagy induces an accumulation of lipid droplets (LD) due to a decrease in fatty acid ß-oxidation, that leads to a reduction of oxidative phosphorylation (OxPHOS) in acute myeloid leukemia (AML), but not in normal cells. Thus, the autophagic process participates in lipid catabolism that supports OxPHOS in AML cells. Interestingly, the inhibition of OxPHOS leads to LD accumulation with the concomitant inhibition of autophagy. Mechanistically, we show that the disruption of mitochondria-endoplasmic reticulum (ER) contact sites (MERCs) phenocopies OxPHOS inhibition. Altogether, our data establish that mitochondria, through the regulation of MERCs, controls autophagy that, in turn finely tunes lipid degradation to fuel OxPHOS supporting proliferation and growth in leukemia.
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Autofagia/fisiología , Retículo Endoplásmico/genética , Retículo Endoplásmico/metabolismo , Leucemia Mieloide Aguda/metabolismo , Leucemia/metabolismo , Mitocondrias/metabolismo , Animales , Autofagia/genética , Línea Celular Tumoral , Proliferación Celular/genética , Proliferación Celular/fisiología , Citometría de Flujo , Humanos , Leucemia/genética , Leucemia Mieloide Aguda/patología , Metabolismo de los Lípidos/genética , Metabolismo de los Lípidos/fisiología , Lipogénesis/genética , Lipogénesis/fisiología , Ratones , Mitocondrias/genética , Oxidación-Reducción , Fosforilación OxidativaRESUMEN
Human adipose-derived stem/stromal cells (hASCs) can differentiate into specialized cell types and thereby contribute to tissue regeneration. As such, hASCs have drawn increasing attention in cell therapy and regenerative medicine, not to mention the ease to isolate them from donors. Culture conditions are critical for expanding hASCs while maintaining optimal therapeutic capabilities. Here, we identified a role for transforming growth factor ß1 (TGFß1) in culture medium in influencing the fate of hASCs during in vitro cell expansion. Human ASCs obtained after expansion in standard culture medium (Standard-hASCs) and in endothelial cell growth medium 2 (EGM2-hASCs) were characterized by high-throughput transcriptional studies, gene set enrichment analysis and functional properties. EGM2-hASCs exhibited enhanced multipotency capabilities and an immature phenotype compared with Standard-hASCs. Moreover, the adipogenic potential of EGM2-hASCs was enhanced, including toward beige adipogenesis, compared with Standard-hASCs. In these conditions, TGFß1 acts as a critical factor affecting the immaturity and multipotency of Standard-hASCs, as suggested by small mother of decapentaplegic homolog 3 (SMAD3) nuclear localization and phosphorylation in Standard-hASCs vs EGM2-hASCs. Finally, the typical priming of Standard-hASCs into osteoblast, chondroblast, and vascular smooth muscle cell (VSMC) lineages was counteracted by pharmacological inhibition of the TGFß1 receptor, which allowed retention of SMAD3 into the cytoplasm and a decrease in expression of osteoblast and VSMC lineage markers. Overall, the TGFß1 pathway appears critical in influencing the commitment of hASCs toward osteoblast, chondroblast, and VSMC lineages, thus reducing their adipogenic potential. These effects can be counteracted by using EGM2 culture medium or chemical inhibition of the TGFß1 pathway.
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Adipocitos Beige/metabolismo , Adipocitos Blancos/metabolismo , Tejido Adiposo/metabolismo , Células del Estroma/metabolismo , Factor de Crecimiento Transformador beta1/metabolismo , Proliferación Celular , Células Cultivadas , Medios de Cultivo , HumanosRESUMEN
Agriculture has benefited from various conventional techniques for plant breeding, including chemical- or radiation-induced mutagenesis, and to some extent from transgenesis. Genome editing techniques are likely to allow straightforward, cost-effective and efficient gene-specific modifications for identified genetic traits associated to agronomic interest. As for previous plant breeding techniques, genome editing techniques need an appraisal for unintended effects. Hence, an evaluation of potential specific risks associated with genome editing must be considered. The Scientific Committee of the High Council for biotechnology (HCB), using a broad theoretical and literature-based approach, identified three categories of points to consider in terms of hazards in health and environment, as compared to conventional breeding: (1) technical unintended effects related to effector persistence as well as risks associated with off-target modifications or other unintended genome modifications, (2) risks arising from the desired trait and its novelty in the plant, and (3) risks associated with the potential modification of plant breeding practices, owing to efficacy and technical ease-of-use of genome editing (acceleration), be it for single traits or for combined modifications (multiplex genome editing). Due to novelty, HCB also envisions the need for specific risk assessment and management.
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Productos Agrícolas/genética , Edición Génica/métodos , Genoma de Planta/genética , Plantas Modificadas Genéticamente/genética , ADN/genética , Ingeniería Genética/tendencias , Humanos , Fenotipo , Fitomejoramiento , Plantas Modificadas Genéticamente/crecimiento & desarrolloRESUMEN
INTRODUCTION: The transcriptional activity of the UGT1A1 gene is modulated by a variable number of repetitions of the dinucleotide (TA) within its promoter region. By comparison to the most common allele (TA)6 (UGT1A1*1), decreased activity is observed with increasing TA repetitions. The aim of this study was to determine whether the presence of the variant allele UGT1A1*28, harbouring seven TA repetitions, (TA)7, in the homozygous state, is associated with precancerous colonic lesions and/or with specific colorectal cancer characteristics. MATERIAL AND METHODS: All patients treated for colorectal cancer in a tertiary care centre, between January 2009 and December 2013, who had routine UGT1A1 genotyping for irinotecan dose-adjustment were included. Data were retrospectively collected. RESULTS: 292 patients were enrolled, including 23 UGT1A1*28/*28 homozygous (7.9%), 137 wild type homozygous (46.9%) and 132 heterozygous (45.2%). There were no significant differences in phenotypic colonic characteristics between homozygous and heterozygous patients carrying the UGT1A1*28 allele as compared to *1/*1 homozygous. Patients treated with aspirin were significantly more common in the UGT1A1*28/*28 homozygous group than in the other groups (7/23 (30.4%) compared to 22/269 (8.2%), pâ¯=â¯0.001). CONCLUSION: Dinucleotide polymorphism in the promoter region of the UGT1A1 gene is not associated with a specific colonic phenotype in patients with sporadic colorectal cancer.
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Neoplasias Colorrectales/genética , Glucuronosiltransferasa/genética , Regiones Promotoras Genéticas , Adulto , Anciano , Anciano de 80 o más Años , Alelos , Femenino , Francia , Predisposición Genética a la Enfermedad , Heterocigoto , Homocigoto , Humanos , Irinotecán , Masculino , Persona de Mediana Edad , Fenotipo , Polimorfismo Genético , Estudios Retrospectivos , Centros de Atención TerciariaRESUMEN
AIM: Hepatocellular carcinoma (HCC) is a common outcome of chronic hepatitis C virus (HCV) infection and constitutes the main burden of this disease. The molecular mechanisms underlying the development of HCC are multiple and might involve certain microRNA (miR). As discordant results have been reported concerning the detection of expression of miR-152 and miR-122 in HCC, our aim was to measure the levels of both miRs in serum and liver samples. METHODS: We analyzed miR-152 and miR-122 expression by reverse transcription-quantitative polymerase chain reaction in a serum cohort from 14 HCV-infected patients who developed HCC, 20 HCV+ patients without HCC, and 19 control patients. We also studied miR-152 and miR-122 in an independent tissue cohort from 11 normal livers, and from paired HCC and non-tumor adjacent livers of 11 HCV-infected patients and 12 non-infected patients. RESULTS: In serum samples, higher levels of miR-122 were found in non-HCC HCV+ compared to HCC HCV+ and control groups, whereas miR-152 was detectable in a lower range in HCC HCV+ compared to non-HCC HCV+ and control groups. We found higher signals for miR-122 and miR-152 in non-tumor liver and HCC tissues compared to control tissues. Hepatocellular carcinoma etiology had no detectable influence on miR-122 expression, whereas miR-152 was increased in HCV+ tissue samples. CONCLUSIONS: Detection of low values of circulating miR-152 is a potentially interesting marker of hepatocarcinogenesis in HCV+ patients, in contrast to miR-122, which varies according to hepatocyte damage.
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All tumors have in common to reactivate a telomere maintenance mechanism to allow for unlimited proliferation. On the other hand, genetic instability found in some tumors can result from the loss of telomeres. Here, we measured telomere length in colorectal cancers (CRCs) using TRF (Telomere Restriction Fragment) analysis. Telomeric DNA content was also quantified as the ratio of total telomeric (TTAGGG) sequences over that of the invariable Alu sequences. In most of the 125 CRCs analyzed, there was a significant diminution in telomere length compared with that in control healthy tissue. Only 34 tumors exhibited no telomere erosion and, in some cases, a slight telomere lengthening. Telomere length did not correlate with age, gender, tumor stage, tumor localization or stage of tumor differentiation. In addition, while telomere length did not correlate with the presence of a mutation in BRAF (V-raf murine sarcoma viral oncogene homolog B), PIK3CA (phosphatidylinositol 3-kinase catalytic subunit), or MSI status, it was significantly associated with the occurrence of a mutation in KRAS. Interestingly, we found that the shorter the telomeres in healthy tissue of a patient, the larger an increase in telomere length in the tumor. Our study points to the existence of two types of CRCs based on telomere length and reveals that telomere length in healthy tissue might influence telomere maintenance mechanisms in the tumor.
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Fosfatidilinositol 3-Quinasa Clase I/genética , Neoplasias Colorrectales/genética , Proteínas Proto-Oncogénicas B-raf/genética , Proteínas Proto-Oncogénicas p21(ras)/genética , Telómero/genética , Biomarcadores de Tumor/genética , Neoplasias Colorrectales/patología , Femenino , Humanos , Masculino , Inestabilidad de Microsatélites , Mutación , Patología Molecular , Homeostasis del Telómero/genéticaRESUMEN
Since the advent of monoclonal antibodies against epidermal growth factor receptor (EGFR) in colorectal cancer therapy, the determination of RAS mutational status is needed for therapeutic decision-making. Most prevalent in colorectal cancer are KRAS exon 2 mutations (40% prevalence); lower prevalence is observed for KRAS exon 3 and 4 mutations (6%) and NRAS exon 2, 3, and 4 mutations (5%). The Idylla™ KRAS Mutation Test on the molecular diagnostics Idylla™ platform is a simple (<2 minutes hands-on time), highly reliable, and rapid (approximately 2 hours turnaround time) in vitro diagnostic sample-to-result solution. This test enables qualitative detection of 21 mutations in codons 12, 13, 59, 61, 117, and 146 of the KRAS oncogene being clinically relevant according to the latest clinical guidelines. Here, the performance of the Idylla™ KRAS Mutation Assay, for Research Use Only, was assessed on archived formalin-fixed paraffin-embedded (FFPE) tissue sections by comparing its results with the results previously obtained by routine reference approaches for KRAS genotyping. In case of discordance, samples were assessed further by additional methods. Among the 374 colorectal cancer FFPE samples tested, the overall concordance between the Idylla™ KRAS Mutation Assay and the confirmed reference routine test results was found to be 98.9%. The Idylla™ KRAS Mutation Assay enabled detection of 5 additional KRAS-mutated samples not detected previously with reference methods. As conclusion the Idylla™ KRAS Mutation Test can be applied as routine tool in any clinical setting, without needing molecular infrastructure or expertise, to guide the personalized treatment of colorectal cancer patients.
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Galectin-3 is a biomarker of fibrosis and atrial remodeling, involved in the mechanisms of initiation and maintenance of atrial fibrillation (AF). We sought to study the accuracy of galectin-3 level in predicting recurrences of AF after ablation. Serum concentrations of galectin-3 were determined in a consecutive series of patients addressed for AF ablation in our center. After a 3-month blanking period, recurrences of atrial arrhythmias were collected during the first year in all patients, using Holter monitoring at 3, 6 months and 12 months. A total of 160 patients were included, with a mean galectin-3 rate was 14.4 ± 5.6 ng/mL. At 12-month, 55 patients (34%) had reexperienced sustained atrial arrhythmia. Only higher galectin-3 level (HR = 1.07 [1.01-1.12], p = 0.02) and larger left atrial diameter (HR = 1.07 [1.03-1.12], p = 0.001) independently predicted recurrence. Patients with both galectin-3 level <15 ng/mL and left atrial diameter <40 millimeters had a 1-year arrhythmia-free survival rate - after a single procedure without anti-arrhythmic drug - of 91%, as compared with 41% in patients with galectin-3 ≥ 15 and left trial diameter ≥40 (p < 0.0001), whether AF was paroxysmal or persistent. Galectin-3 and left atrial diameters, rather than clinical presentation of AF, predict recurrences after ablation.
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BACKGROUND: Deficient mismatch repair (dMMR) colon cancer (CC) is reportedly resistant to 5-fluorouracil (5FU) adjuvant chemotherapy while preliminary data suggest chemosensitivity to oxaliplatin. We assessed the efficacy of fluoropyrimidine with and without oxaliplatin in a large cohort of dMMR CC patients. METHODS: This retrospective multicenter study included all consecutive patients who underwent curative surgical resection for stage II or III dMMR CC between 2000 and 2011. Prognostic factors were analyzed using Cox models, and hazard ratios (HRs) with 95% confidence intervals (CIs) were calculated. All statistical tests were two-sided. RESULTS: A total of 433 dMMR CC patients were included (56.8% stage II, 43.2% stage III). Mean follow-up was 47.0 months. The patients received surgery alone (n = 263) or surgery plus adjuvant chemotherapy consisting of fluoropyrimidine with (n = 119) or without (n = 51) oxaliplatin. Adjuvant chemotherapy was administered to 16.7% of stage II and 69.0% of stage III CC patients. As compared with surgery alone, adjuvant oxaliplatin-based chemotherapy improved disease-free survival (DFS) in multivariable analysis (HR = 0.35, 95% CI = 0.19 to 0.65, P < .001), contrary to adjuvant fluoropyrimidine alone (HR = 0.73, 95% CI = 0.36 to 1.49, P = .38). In the subgroup analysis, the DFS benefit of oxaliplatin-based chemotherapy was statistically significant in multivariable analysis only in stage III (HR = 0.41, 95% CI = 0.19 to 0.87, P = .02). CONCLUSION: This study supports the use of adjuvant chemotherapy with fluoropyrimidine plus oxaliplatin in stage III dMMR CC.
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Antineoplásicos/uso terapéutico , Neoplasias Encefálicas , Neoplasias del Colon/tratamiento farmacológico , Neoplasias del Colon/genética , Neoplasias Colorrectales , Inestabilidad de Microsatélites , Síndromes Neoplásicos Hereditarios , Pirimidinas/uso terapéutico , Adulto , Anciano , Anciano de 80 o más Años , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Quimioterapia Adyuvante , Neoplasias del Colon/patología , Neoplasias del Colon/cirugía , Femenino , Mutación de Línea Germinal , Humanos , Estimación de Kaplan-Meier , Masculino , Persona de Mediana Edad , Estadificación de Neoplasias , Compuestos Organoplatinos/administración & dosificación , Oxaliplatino , Pirimidinas/administración & dosificación , Estudios Retrospectivos , Resultado del TratamientoRESUMEN
RNA delivery is an attractive strategy to achieve transient gene expression in research projects and in cell- or gene-based therapies. Despite significant efforts investigating vector-directed RNA transfer, there is still a requirement for better efficiency of delivery to primary cells and in vivo. Retroviral platforms drive RNA delivery, yet retrovirus RNA-packaging constraints limit gene transfer to two genome-molecules per viral particle. To improve retroviral transfer, we designed a dimerization-independent MS2-driven RNA packaging system using MS2-Coat-retrovirus chimeras. The engineered chimeric particles promoted effective packaging of several types of RNAs and enabled efficient transfer of biologically active RNAs in various cell types, including human CD34(+) and iPS cells. Systemic injection of high-titer particles led to gene expression in mouse liver and transferring Cre-recombinase mRNA in muscle permitted widespread editing at the ROSA26 locus. We could further show that the VLPs were able to activate an osteoblast differentiation pathway by delivering RUNX2- or DLX5-mRNA into primary human bone-marrow mesenchymal-stem cells. Thus, the novel chimeric MS2-lentiviral particles are a versatile tool for a wide range of applications including cellular-programming or genome-editing.
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Procalcitonin (PCT) levels are commonly used for diagnostic guidance in routine bacterial infections. By contrast, little data are currently available regarding PCT in parasitic diseases, and its role in cases of invasive amoebiasis has not yet been described. For this purpose, 35 adult patients with a proven diagnosis of invasive or digestive amoebiasis were included in a 4-year study period. Serum PCT was retrospectively assessed. Results were analysed with regard to the usual inflammatory biomarkers, like C-reactive protein (CRP). PCT was significantly higher in patients with proven invasive amoebiasis than in digestive amoebiasis (mean value: 4.03 µg/L versus 0.07 µg/L, respectively; P < 0.001), but the SD was greater than with CRP, and the effect was less than that demonstrated in bacterial infections. By contrast, PCT was not shown to be elevated during digestive amoebiasis.
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Amebiasis/patología , Calcitonina/sangre , Precursores de Proteínas/sangre , Adulto , Proteína C-Reactiva/análisis , Péptido Relacionado con Gen de Calcitonina , Femenino , Humanos , Masculino , Persona de Mediana Edad , Estudios Retrospectivos , Suero/químicaRESUMEN
To evaluate the diagnostic value of MDM2 status in craniofacial fibro-osseous lesions, we investigated MDM2 expression by immunohistochemistry and analyzed MDM2 amplification by qPCR in 30 cases of ossifying fibroma (including 13 cases of the juvenile variant) and 17 cases of fibrous dysplasia. Two cases of uncommon extragnathic psammomatoid fibrous dysplasia and a mixed control group of 15 cases of low-grade osteosarcoma and 15 cases of well-differentiated/dedifferentiated liposarcoma were included. MDM2 amplification was found in 33% of ossifying fibromas (peak of 69% for the juvenile variant) and in 12% of fibrous dysplasia, in none of which was MDM2 overexpressed. All control cases exhibited MDM2 amplification and overexpression. To investigate possible polysomy of chromosome 12, we studied RASAL1 amplification, a gene telomeric to MDM2 on the long arm of chromosome 12. RASAL1 amplification was reported in all benign fibro-osseous lesions exhibiting MDM2 amplification but not in controls. Simultaneous amplification of these two genes was significantly higher in juvenile ossifying fibromas compared with fibrous dysplasia (P=0.004), non-juvenile ossifying fibromas (P=0.001), and all other benign craniofacial fibro-osseous lesions combined (P=0.0001). Of the nine cases of juvenile ossifying fibroma exhibiting amplification, three were locally invasive and four were recurrent, suggesting aggressive disease. The two cases of extragnathic psammomatoid fibrous dysplasia also showed MDM2 and RASAL1 amplification with no MDM2 overexpression. This large chromosome 12 rearrangement, spanning MDM2 and RASAL1, is the first recurrent molecular abnormality to be reported in juvenile ossifying fibroma. It may represent both a molecular diagnostic marker and a characteristic of more aggressive forms with a higher risk of recurrence. Finally, the presence of this rearrangement in extragnathic psammomatoid fibro-osseous lesions mimicking ossifying fibromas might reflect a common molecular pathway in their pathogenesis and calls into question the classification of such lesions within fibrous dysplasia.
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Neoplasias Óseas/genética , Cromosomas Humanos Par 12/genética , Fibroma Osificante/genética , Displasia Fibrosa Ósea/genética , Proteínas Activadoras de GTPasa/genética , Proteínas Proto-Oncogénicas c-mdm2/genética , Adolescente , Adulto , Anciano , Neoplasias Óseas/diagnóstico , Niño , Preescolar , Huesos Faciales , Femenino , Fibroma Osificante/diagnóstico , Displasia Fibrosa Ósea/diagnóstico , Reordenamiento Génico , Humanos , Inmunohistoquímica , Lactante , Masculino , Persona de Mediana Edad , Reacción en Cadena de la Polimerasa , Estudios Retrospectivos , Cráneo , Neoplasias Craneales/diagnóstico , Neoplasias Craneales/genética , Adulto JovenRESUMEN
BACKGROUND: Na(V)1.5 voltage-gated sodium channels are abnormally expressed in breast tumours and their expression level is associated with metastatic occurrence and patients' death. In breast cancer cells, Na(V)1.5 activity promotes the proteolytic degradation of the extracellular matrix and enhances cell invasiveness. FINDINGS: In this study, we showed that the extinction of Na(V)1.5 expression in human breast cancer cells almost completely abrogated lung colonisation in immunodepressed mice (NMRI nude). Furthermore, we demonstrated that ranolazine (50 µM) inhibited Na(V)1.5 currents in breast cancer cells and reduced Na(V)1.5-related cancer cell invasiveness in vitro. In vivo, the injection of ranolazine (50 mg/kg/day) significantly reduced lung colonisation by Na(V)1.5-expressing human breast cancer cells. CONCLUSIONS: Taken together, our results demonstrate the importance of Na(V)1.5 in the metastatic colonisation of organs by breast cancer cells and indicate that small molecules interfering with Na(V) activity, such as ranolazine, may represent powerful pharmacological tools to inhibit metastatic development and improve cancer treatments.
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Acetanilidas/farmacología , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/patología , Pulmón/patología , Canal de Sodio Activado por Voltaje NAV1.5/metabolismo , Invasividad Neoplásica/patología , Piperazinas/farmacología , Animales , Neoplasias de la Mama/metabolismo , Línea Celular Tumoral , Movimiento Celular/efectos de los fármacos , Femenino , Humanos , Ratones , Ratones Desnudos , Metástasis de la Neoplasia/tratamiento farmacológico , Metástasis de la Neoplasia/patología , RanolazinaRESUMEN
The classic model of tumor suppression implies that malignant transformation requires full "two-hit" inactivation of a tumor-suppressor gene. However, more recent work in mice has led to the proposal of a "continuum" model that involves more fluid concepts such as gene dosage-sensitivity and tissue specificity. Mutations in the tumor-suppressor gene von Hippel-Lindau (VHL) are associated with a complex spectrum of conditions. Homozygotes or compound heterozygotes for the R200W germline mutation in VHL have Chuvash polycythemia, whereas heterozygous carriers are free of disease. Individuals with classic, heterozygous VHL mutations have VHL disease and are at high risk of multiple tumors (e.g., CNS hemangioblastomas, pheochromocytoma, and renal cell carcinoma). We report here an atypical family bearing two VHL gene mutations in cis (R200W and R161Q), together with phenotypic analysis, structural modeling, functional, and transcriptomic studies of these mutants in comparison with classical mutants involved in the different VHL phenotypes. We demonstrate that the complex pattern of disease manifestations observed in VHL syndrome is perfectly correlated with a gradient of VHL protein (pVHL) dysfunction in hypoxia signaling pathways. Thus, by studying naturally occurring familial mutations, our work validates in humans the "continuum" model of tumor suppression.