RESUMEN
AIMS: Primary mixed liver cancers (PLCs), combined hepatocellular-cholangiocellular (cHCC-CC) and intermediate-cell carcinomas are rare tumours characterised by different molecular mechanisms. Nestin is a marker of progenitor cells with a promising application in human tumours. The aims of the present paper are (i) to determine the expression of Nestin in mixed PLCs; and (ii) to correlate the PLC immunoprofile with the gene expression in each tumour component. METHODS AND RESULTS: We selected 28 mixed PLCs, 13 (46.4%) cHCC-CC and 15 (53.6%) intermediate-cell carcinomas. The immunohistochemistry panel consisted of keratin 7, keratin 19, CD56 and Nestin. Next-generation sequencing analysis was performed on 17 cases (27 specimens) using a multi-gene custom panel. The differentiated HCC and CC components of cHCC-CC were negative for Nestin in all cases. The intermediate areas of cHCC-CC were immunoreactive for Nestin in 92.3% of cases, for CD56 in 76.9% and for K7/K19 in all cases. The immunoprofile of the intermediate-cell carcinomas showed 73.3% of cases positive for Nestin and 66.7% for CD56. TP53 and TERT were the most frequently mutated genes (31.3% and 17.6% of samples, respectively). Mutations were also found in IDH1, IDH2, PIK3CA and NRAS genes. Intermediate and HCC areas of cHCC-CC seemed to share the same mutational profile, and both harboured different mutations than the CC component. CONCLUSIONS: According to our preliminary data, Nestin was not expressed by hepatocellular or cholangiocellular-cell components, but was expressed by most of the intermediate cells in PLCs, and therefore could be considered in the differential diagnosis of PLCs, together with mutational profile.
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Biomarcadores de Tumor/metabolismo , Carcinoma Hepatocelular/metabolismo , Colangiocarcinoma/metabolismo , Neoplasias Hepáticas/metabolismo , Nestina/metabolismo , Adulto , Anciano , Anciano de 80 o más Años , Biomarcadores de Tumor/genética , Carcinoma Hepatocelular/diagnóstico , Carcinoma Hepatocelular/patología , Colangiocarcinoma/diagnóstico , Colangiocarcinoma/patología , Diagnóstico Diferencial , Femenino , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Inmunohistoquímica , Inmunofenotipificación , Hígado/metabolismo , Hígado/patología , Neoplasias Hepáticas/diagnóstico , Neoplasias Hepáticas/patología , Masculino , Persona de Mediana Edad , Mutación , Nestina/genética , Estudios Retrospectivos , Análisis de Secuencia de ADNRESUMEN
Pancreatic adenocarcinoma (PDAC) is the most frequent histological type of malignancy in the pancreas. Extracellular matrix (ECM), plays a critical role during the process of human carcinogenesis and the possible diversity in matricellular proteins composition of ECM may have a significant impact on the clinical course of PDAC. Aim of this paper was to evaluate the expression of three matricellular proteins, including Periostin (POSTN), Tenascin (TNS) and Osteopontin (OPN), in PDAC from long-survival (LS) and non-long survival (NLS) patients. A total of 30 PDAC were analyzed, 15 from patients that survived more than 60 months after surgery (LS) and 15 that died from the disease within 24 (NLS). RNA was extracted and OPN, TNS and POSTN mRNA levels were evaluated by qRT-PCR. LS and NLS samples showed the same type of POSTN and TN isoforms. On the contrary, OPN seems to be preferentially expressed in NLS PDAC. Moreover, OPNb and OPNc isoforms were expressed exclusively in NLS samples. In conclusion, Our data led to hypothesize a possible relationship between the expression of different isoforms of each of these proteins and the clinical outcome of patients with PDAC.
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Adenocarcinoma , Moléculas de Adhesión Celular/biosíntesis , Proteínas de Neoplasias/biosíntesis , Osteopontina/biosíntesis , Neoplasias Pancreáticas , Tenascina/biosíntesis , Adenocarcinoma/metabolismo , Adenocarcinoma/mortalidad , Adenocarcinoma/cirugía , Adulto , Supervivencia sin Enfermedad , Femenino , Humanos , Masculino , Persona de Mediana Edad , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/mortalidad , Neoplasias Pancreáticas/cirugía , Proyectos Piloto , Isoformas de Proteínas/biosíntesis , Tasa de SupervivenciaRESUMEN
Increased glucose metabolism and reprogramming toward aerobic glycolysis are a hallmark of cancer cells, meeting their metabolic needs for sustained cell proliferation. Metabolic reprogramming is usually considered as a downstream consequence of tumor development and oncogene activation; growing evidence indicates, however, that metabolism on its turn can support oncogenic signaling to foster tumor malignancy. Here, we explored how glucose metabolism regulates gene transcription and found an unexpected link with YAP/TAZ, key transcription factors regulating organ growth, tumor cell proliferation and aggressiveness. When cells actively incorporate glucose and route it through glycolysis, YAP/TAZ are fully active; when glucose metabolism is blocked, or glycolysis is reduced, YAP/TAZ transcriptional activity is decreased. Accordingly, glycolysis is required to sustain YAP/TAZ pro-tumorigenic functions, and YAP/TAZ are required for the full deployment of glucose growth-promoting activity. Mechanistically we found that phosphofructokinase (PFK1), the enzyme regulating the first committed step of glycolysis, binds the YAP/TAZ transcriptional cofactors TEADs and promotes their functional and biochemical cooperation with YAP/TAZ. Strikingly, this regulation is conserved in Drosophila, where phosphofructokinase is required for tissue overgrowth promoted by Yki, the fly homologue of YAP. Moreover, gene expression regulated by glucose metabolism in breast cancer cells is strongly associated in a large dataset of primary human mammary tumors with YAP/TAZ activation and with the progression toward more advanced and malignant stages. These findings suggest that aerobic glycolysis endows cancer cells with particular metabolic properties and at the same time sustains transcription factors with potent pro-tumorigenic activities such as YAP/TAZ.
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Bacterias Aerobias/metabolismo , Factores de Transcripción/metabolismo , Aciltransferasas , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Animales , Línea Celular Tumoral , Inmunoprecipitación de Cromatina , Drosophila , Glucólisis/genética , Glucólisis/fisiología , Humanos , Inmunoprecipitación , Fosfoproteínas/metabolismo , Reacción en Cadena en Tiempo Real de la Polimerasa , Proteínas Señalizadoras YAPRESUMEN
Thyroid cancer is the most common endocrine malignancy. Knowledge of the molecular pathology of thyroid tumours originating from follicular cells has greatly advanced in the past several years. Common molecular alterations, such as BRAF p.V600E, RAS point mutations, and fusion oncogenes (RET-PTC being the prototypical example), have been, respectively, associated with conventional papillary carcinoma, follicular-patterned tumours (follicular adenoma, follicular carcinoma, and the follicular variant of papillary carcinoma/non-invasive follicular thyroid neoplasm with papillary-like nuclear features), and with papillary carcinomas from young patients and arising after exposure to ionising radiation, respectively. The remarkable correlation between genotype and phenotype shows how specific, mutually exclusive molecular changes can promote tumour development and initiate a multistep tumorigenic process that is characterised by aberrant activation of mitogen-activated protein kinase and phosphoinositide 3-kinase-PTEN-AKT signalling. Molecular alterations are becoming useful biomarkers for diagnosis and risk stratification, and as potential treatment targets for aggressive forms of thyroid carcinoma. What follows is a review of the principal genetic alterations of thyroid tumours originating from follicular cells and of their clinicopathological relevance.
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Adenocarcinoma Folicular/genética , Adenocarcinoma Folicular/patología , Células Epiteliales Tiroideas/patología , Neoplasias de la Tiroides/genética , Neoplasias de la Tiroides/patología , HumanosRESUMEN
AIM: To identify patients with recurrent glioblastoma after temozolomide (TMZ) concurrent with and adjuvant to radiotherapy who could benefit from TMZ rechallenge at the time of disease progression. METHODS: We retrospectively evaluated 106 glioblastoma patients who had nonprogressive disease at first magnetic resonance imaging after completion of TMZ concurrent with and adjuvant to radiotherapy, a treatment-free interval (TFI) of at least 8 weeks and received TMZ rechallenge or a nitrosourea at the time of progression. RESULTS: In patients with TFI ≥5 months, median survival was 17.7 and 11.6 months and median progression-free survival was 8.1 and 5.8 months in the TMZ and nitrosourea group, respectively. Longer TFI was associated with reduced risk for death (p = 0.002) and for disease progression (p = 0.005). CONCLUSION: TFI ≥5 months represents a predictor of retained TMZ sensitivity.
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Neoplasias Encefálicas/tratamiento farmacológico , Glioblastoma/tratamiento farmacológico , Recurrencia Local de Neoplasia/tratamiento farmacológico , Adulto , Anciano , Neoplasias Encefálicas/diagnóstico por imagen , Neoplasias Encefálicas/patología , Dacarbazina/administración & dosificación , Dacarbazina/efectos adversos , Dacarbazina/análogos & derivados , Supervivencia sin Enfermedad , Femenino , Glioblastoma/diagnóstico por imagen , Glioblastoma/patología , Humanos , Imagen por Resonancia Magnética , Masculino , Persona de Mediana Edad , Recurrencia Local de Neoplasia/diagnóstico por imagen , Recurrencia Local de Neoplasia/patología , Temozolomida , Resultado del TratamientoRESUMEN
AIM: To evaluate relevance of clinical and molecular factors in adult low-grade gliomas (LGG) and to correlate with survival. METHODS: We reviewed records from adult LGG patients from 1991 to 2015 who received surgery and had sufficient tissue to molecular biomarkers characterization. RESULTS: 213 consecutive LGG patients were included: 17.4% were low-risk, according to Radiation Therapy Oncology Group (RTOG) risk assessment. IDH 1/2 mutation, 1p/19q co-deletion, MGMT methylation were found in 93, 50.8 and 65.3% of patients. Median follow-up was 98.3 months. In univariate analysis, overall survival was influenced by extent of resection (p = 0.011), IDH mutation (p < 0.001), 1p/19q co-deletion (p = 0.015) and MGMT methylation (p = 0.013). In multivariate analysis, RTOG clinical risk (p = 0.006), IDH mutation (p < 0.001) and 1p/19q co-deletion (p = 0.035) correlated with overall survival. RTOG clinical risk (p = 0.006), IDH mutation (p < 0.001) and 1p/19q co-deletion (p = 0.035) correlated with overall survival. CONCLUSION: Both clinical and molecular factors are essential to determine prognosis and treatment strategies.
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Neoplasias Encefálicas/genética , Neoplasias Encefálicas/mortalidad , Glioma/genética , Glioma/mortalidad , Adolescente , Adulto , Anciano , Neoplasias Encefálicas/terapia , Deleción Cromosómica , Cromosomas Humanos Par 1 , Estudios de Cohortes , Metilasas de Modificación del ADN/genética , Enzimas Reparadoras del ADN/genética , Femenino , Glioma/terapia , Humanos , Isocitrato Deshidrogenasa/genética , Masculino , Persona de Mediana Edad , Análisis Multivariante , Mutación , Procedimientos Neuroquirúrgicos , Pronóstico , Factores de Riesgo , Proteínas Supresoras de Tumor/genéticaRESUMEN
BACKGROUND: MGMT methylation status represents a powerful prognostic factor in newly diagnosed glioblastoma (GBM). Recently, its role in recurrent tumors has also been suggested; however, few data investigating the stability of this biomarker during the clinical course of the disease are available. In this study, we evaluated the rate of change of MGMT methylation status between diagnosis and first recurrence in patients who received tumor resection for recurrent GBM. METHODS: We included patients who received temozolomide concurrent with and adjuvant to radiotherapy after diagnosis of GBM and had a second surgery performed at least 3 months after radiotherapy completion. Other eligibility criteria were age ≥18 years and Eastern Cooperative Oncology Group performance status 0-2. We evaluated the MGMT methylation status by methylation-specific polymerase chain reaction. RESULTS: From our institutional data warehouse, 295 patients with recurrent GBM who underwent second surgery were evaluated. MGMT methylation status at both first and second surgery was available for 108 patients. MGMT was methylated in both surgeries in 38 patients (35.2%), while it was unmethylated in 43 patients (39.8%). We found a significant concordance between the first and the second MGMT methylation assessments (K = 0.500, p < .001), MGMT methylation being stable in 75% of the cases. CONCLUSION: MGMT methylation presents relative stability during the clinical course of GBM. The Oncologist 2017;22:432-437 IMPLICATIONS FOR PRACTICE: MGMT methylation is a prognostic factor in newly diagnosed glioblastoma. In this study, we evaluated the rate of change of MGMT methylation during the clinical course of the disease, and we found a significant concordance between the first and the second MGMT methylation assessments, with MGMT methylation being stable in 75% of the cases. Thus, re-testing this biomarker at recurrence does not provide further information for clinicians. MGMT methylation at first surgery, extent of resection at second surgery, and time between first and second surgery are significantly correlated with overall survival. Age and extent of resection are correlated with post-progression survival.
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Metilación de ADN/genética , Metilasas de Modificación del ADN/genética , Enzimas Reparadoras del ADN/genética , Glioblastoma/genética , Pronóstico , Proteínas Supresoras de Tumor/genética , Adolescente , Adulto , Anciano , Biomarcadores de Tumor/genética , Dacarbazina/administración & dosificación , Dacarbazina/análogos & derivados , Supervivencia sin Enfermedad , Femenino , Glioblastoma/diagnóstico , Glioblastoma/patología , Glioblastoma/cirugía , Humanos , Masculino , Persona de Mediana Edad , Recurrencia Local de Neoplasia/genética , Recurrencia Local de Neoplasia/patología , Regiones Promotoras Genéticas , TemozolomidaRESUMEN
According to the 2016 World Health Organization (WHO) classification of tumors of the central nervous system, assessment of exon 4 mutations in isocitrate dehydrogenase 1 or 2 genes (IDH1 or IDH2) is an essential step in the characterization of gliomas. The p.R132H mutation is the most frequent alteration in IDH genes, however other non-canonical IDH mutations can be identified. The aim of this study is to investigate in depth the prevalence of non-R132H IDH ("non-canonical") mutations in brain tumors classified according to the 2016 WHO scheme and their clonal distribution in neoplastic cells. A total of 288 consecutive cases of brain gliomas (grade II-IV) were analyzed for exon 4 IDH1 and IDH2 mutations. IDH1 and IDH2 analysis was performed using next generation sequencing. Non-canonical IDH mutations were identified in 13/52 (25.0%) grade II gliomas (astrocytomas: 8/31, 25.8%; oligodendrogliomas: 5/21, 23.8%) and in 5/40 (12.5%) grade III gliomas (astrocytomas: 3/25, 12.0%; oligodendrogliomas: 2/15, 13.3%). They were not identified in 196 grade IV gliomas (192 glioblastomas, 4 gliosarcomas). In the large majority (>80%) of tumors IDH mutations, both IDH1-R132H and the non-canonical ones, were present in the large majority (>80%) of neoplastic cells. Our data highlight the importance of investigating not only the IDH1-R132H mutation but also the non-canonical ones. These mutations are clonally distributed, with proportions of mutated neoplastic cells overlapping with those of p.R132H, a finding consistent with their driver role in gliomagenesis.
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Neoplasias Encefálicas/genética , Glioma/genética , Isocitrato Deshidrogenasa/genética , Mutación , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Neoplasias Encefálicas/clasificación , Neoplasias Encefálicas/epidemiología , Neoplasias Encefálicas/patología , Estudios de Cohortes , Exones , Femenino , Glioma/clasificación , Glioma/epidemiología , Glioma/patología , Humanos , Italia , Masculino , Persona de Mediana Edad , Clasificación del Tumor , Prevalencia , Organización Mundial de la Salud , Adulto JovenRESUMEN
BACKGROUND: The most appropriate management of recurrent glioblastoma is still controversial. In particular, the role of surgery at recurrence remains uncertain. PATIENTS & METHODS: From our Institutional data warehouse we analyzed 270 consecutive patients who received second surgery for recurrent glioblastoma, to assess survival after second surgery, and to evaluate prognostic factors. RESULTS: Complete resection was found in 128 (47.4%) and partial resection in 142 patients (52.6%). Median survival from second surgery was 11.4 months (95% CI: 10.0-12.7). Multivariate analysis showed that age (p = 0.001), MGMT methylation (p = 0.021) and extent of surgery (p < 0.001) are associated with better survival. CONCLUSION: A complete resection should be the goal for second resection and younger age and MGMT methylation status might be considered in the selection of patients.
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Neoplasias Encefálicas/patología , Neoplasias Encefálicas/cirugía , Glioblastoma/patología , Glioblastoma/cirugía , Adolescente , Adulto , Anciano , Biomarcadores de Tumor , Neoplasias Encefálicas/mortalidad , Terapia Combinada , Metilación de ADN , Metilasas de Modificación del ADN/genética , Enzimas Reparadoras del ADN/genética , Manejo de la Enfermedad , Femenino , Glioblastoma/mortalidad , Humanos , Masculino , Persona de Mediana Edad , Recurrencia Local de Neoplasia , Pronóstico , Retratamiento , Tomografía Computarizada por Rayos X , Resultado del Tratamiento , Proteínas Supresoras de Tumor/genética , Adulto JovenRESUMEN
Studies from single institutions have analyzed BRAF in papillary microcarcinomas, sometimes with contradictory results. Most of them have provided limited integration of histological and clinical data. To obtain a comprehensive picture of BRAF V600E-mutated microcarcinomas and to evaluate the role of BRAF testing in risk stratification we performed a retrospective multicenter analysis integrating microscopical, pathological, and clinical information. Three hundred and sixty-five samples from 300 patients treated at six medical institutions covering different geographical regions of Italy were analyzed with central review of all cases. BRAF V600E statistical analysis was conducted on 298 microcarcinomas from 264 patients after exclusion of those that did not meet the required criteria. BRAF V600E was identified in 145/298 tumors (49%) including the following subtypes: 35/37 (95%, P<0.0001) tall cell and 72/114 (64%, P<0.0001) classic; conversely 94/129 follicular variant papillary microcarcinomas (73%, P<0.0001) were BRAF wild type. BRAF V600E-mutated microcarcinomas were characterized by markedly infiltrative contours (P<0.0001) with elongated strings of neoplastic cells departing from the tumor, and by intraglandular tumor spread (P<0.0001), typically within 5 mm of the tumor border. Multivariate analysis correlated BRAF V600E with specific microscopic features (nuclear grooves, optically clear nuclei, tall cells within the tumor, and tumor fibrosis), aggressive growth pattern (infiltrative tumor border, extension into extrathyroidal tissues, and intraglandular tumor spread), higher American Thyroid Association recurrence risk group, and non-incidental tumor discovery. The following showed the strongest link to BRAF V600E: tall cell subtype, many neoplastic cells with nuclear grooves or with optically clear nuclei, infiltrative growth, intraglandular tumor spread, and a tumor discovery that was non-incidental. BRAF V600E-mutated microcarcinomas represent a distinct biological subtype. The mutation is associated with conventional clinico-pathological features considered to be adverse prognostic factors for papillary microcarcinoma, for which it could be regarded as a surrogate marker. BRAF analysis may be useful to identify tumors (BRAF wild type) that have negligible clinical risk.
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Carcinoma Papilar/genética , Carcinoma Papilar/patología , Mutación , Proteínas Proto-Oncogénicas B-raf/genética , Neoplasias de la Tiroides/genética , Neoplasias de la Tiroides/patología , Adulto , Carcinoma Papilar/mortalidad , Femenino , Humanos , Inmunohistoquímica , Estimación de Kaplan-Meier , Masculino , Persona de Mediana Edad , Pronóstico , Estudios Retrospectivos , Factores de Riesgo , Neoplasias de la Tiroides/mortalidadRESUMEN
The proteasome is the core of the ubiquitin-proteasome system and is involved in synaptic protein metabolism. The incorporation of three inducible immuno-subunits into the proteasome results in the generation of the so-called immunoproteasome, which is endowed of pathophysiological functions related to immunity and inflammation. In healthy human brain, the expression of the key catalytic ß5i subunit of the immunoproteasome is almost absent, while it is induced in the epileptogenic foci surgically resected from patients with pharmaco-resistant seizures, including temporal lobe epilepsy. We show here that the ß5i immuno-subunit is induced in experimental epilepsy, and its selective pharmacological inhibition significantly prevents, or delays, 4-aminopyridine-induced seizure-like events in acute rat hippocampal/entorhinal cortex slices. These effects are stronger in slices from epileptic vs normal rats, likely due to the more prominent ß5i subunit expression in neurons and glia cells of diseased tissue. ß5i subunit is transcriptionally induced in epileptogenic tissue likely by Toll-like receptor 4 signaling activation, and independently on promoter methylation. The recent availability of selective ß5i subunit inhibitors opens up novel therapeutic opportunities for seizure inhibition in drug-resistant epilepsies.
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Epilepsia/enzimología , Complejo de la Endopetidasa Proteasomal/metabolismo , Animales , Modelos Animales de Enfermedad , Corteza Entorrinal/fisiopatología , Epilepsia/fisiopatología , Hipocampo/fisiopatología , Masculino , Oligopéptidos/farmacología , Inhibidores de Proteasoma/farmacología , Subunidades de Proteína/metabolismo , ARN Mensajero/metabolismo , Ratas , Ratas WistarRESUMEN
Genetic analyses in Drosophila epithelia have suggested that the phenomenon of "cell competition" could participate in organ homeostasis. It has been speculated that competition between different cell populations within a growing organ might play a role as either tumor promoter or tumor suppressor, depending on the cellular context. The evolutionarily conserved Hippo (Hpo) signaling pathway regulates organ size and prevents hyperplastic disease from flies to humans by restricting the activity of the transcriptional cofactor Yorkie (yki). Recent data indicate also that mutations in several Hpo pathway members provide cells with a competitive advantage by unknown mechanisms. Here we provide insight into the mechanism by which the Hpo pathway is linked to cell competition, by identifying dMyc as a target gene of the Hpo pathway, transcriptionally upregulated by the activity of Yki with different binding partners. We show that the cell-autonomous upregulation of dMyc is required for the supercompetitive behavior of Yki-expressing cells and Hpo pathway mutant cells, whereas the relative levels of dMyc between Hpo pathway mutant cells and wild-type neighboring cells are critical for determining whether cell competition promotes a tumor-suppressing or tumor-inducing behavior. All together, these data provide a paradigmatic example of cooperation between tumor suppressor genes and oncogenes in tumorigenesis and suggest a dual role for cell competition during tumor progression depending on the output of the genetic interactions occurring between confronted cells.
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Proteínas de Unión al ADN/metabolismo , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/citología , Drosophila melanogaster/metabolismo , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Mutación/genética , Proteínas Nucleares/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Transducción de Señal , Transactivadores/metabolismo , Factores de Transcripción/metabolismo , Animales , Secuencia de Bases , Proliferación Celular , Células Clonales , Proteínas de Unión al ADN/genética , Proteínas de Drosophila/genética , Drosophila melanogaster/genética , Genes de Insecto/genética , Datos de Secuencia Molecular , Factores de Transcripción/genética , Transcripción Genética , Regulación hacia Arriba/genética , Proteínas Señalizadoras YAPRESUMEN
Gaucher Disease (GD), the most common lysosomal disorder, arises from mutations in the GBA1 gene and is characterized by a wide spectrum of phenotypes, ranging from mild hematological and visceral involvement to severe neurological disease. Neuronopathic patients display dramatic neuronal loss and increased neuroinflammation, whose molecular basis are still unclear. Using a combination of Drosophila dGBA1b loss-of-function models and GD patient-derived iPSCs differentiated towards neuronal precursors and mature neurons we showed that different GD- tissues and neuronal cells display an impairment of growth mechanisms with an increased cell death and reduced proliferation. These phenotypes are coupled with the downregulation of several Hippo transcriptional targets, mainly involved in cells and tissue growth, and YAP exclusion from nuclei. Interestingly, Hippo knock-down in the GBA-KO flies rescues the proliferative defect, suggesting that targeting the Hippo pathway can be a promising therapeutic approach to neuronopathic GD.
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Enfermedad de Gaucher , Humanos , Enfermedad de Gaucher/genética , Enfermedad de Gaucher/metabolismo , Enfermedad de Gaucher/terapia , Glucosilceramidasa/genética , Glucosilceramidasa/metabolismo , Vía de Señalización Hippo , Neuronas/metabolismo , Proliferación CelularRESUMEN
PURPOSE: Stratifying patients defective in mismatch repair (dMMR) with high microsatellite instability (MSI-H) in colorectal cancer (CRC) is of increasing relevance and may provide a more tailored approach to CRC adjuvant therapy. Here, we describe the discovery of a new MSI marker for colorectal cancer located in the 3'-untranslated region (3'UTR, T20 mononucleotide repeat) of the metallothionein 1X gene (MT1XT20). METHODS: We studied 340 consecutive CRCs using three multiplexed polymerase chain reactions amplifying BAT25, BAT26, TGFBR2, MybT22, BAT40, MT1XT20, NR21, NR24, CAT25, D2S123, D5S346, D17S250, D18S58, CSF1PO, D7S820, and D18S51. Fragments length was evaluated by automated capillary electrophoresis. RESULTS: Based on the NCI/ICG-HNPCC criteria for MSI classification, 40 CRCs were found to be MSI-high (11.8%), 46 (13.5%) CRCs were MSI-low, and 254 CRCs (74.7%) were stable (MSS). MT1XT20 showed very high sensitivity (97.3%) comparable to BAT26 (97.5%) and CAT25 (97.1%) and the best specificity (100%) as well as MybT22 and CAT25. Indeed, MT1XT20 instability was detected in 36 out of 37 cases (97.3%) of MSI-high colorectal cancers, whereas no MT1XT20 alterations were observed in 254 MSS or in 46 MSI-low cases. On the contrary, BAT40 was found to be unstable in 8/46 MSI-low cases, BAT25 in 6/46, BAT26 4/46, NR21 1/46, and NR24 in 1/45. CONCLUSIONS: Our results suggest that MT1XT20 represents a sensitive and specific marker for MSI testing and could be included in a complete set of MSI markers for the confident identification of familial or sporadic dMMR patients in CRCs.
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Biomarcadores de Tumor/genética , Neoplasias Colorrectales/genética , Metalotioneína/genética , Inestabilidad de Microsatélites , Secuencias Repetidas en Tándem/genética , Regiones no Traducidas 3' , Adulto , Anciano , Anciano de 80 o más Años , Reparación de la Incompatibilidad de ADN , Femenino , Marcadores Genéticos , Humanos , Masculino , Reacción en Cadena de la Polimerasa , Sensibilidad y EspecificidadRESUMEN
BACKGROUND: Genetic studies in Drosophila melanogaster reveal an important role for Myc in controlling growth. Similar studies have also shown how components of the insulin and target of rapamycin (TOR) pathways are key regulators of growth. Despite a few suggestions that Myc transcriptional activity lies downstream of these pathways, a molecular mechanism linking these signaling pathways to Myc has not been clearly described. Using biochemical and genetic approaches we tried to identify novel mechanisms that control Myc activity upon activation of insulin and TOR signaling pathways. RESULTS: Our biochemical studies show that insulin induces Myc protein accumulation in Drosophila S2 cells, which correlates with a decrease in the activity of glycogen synthase kinase 3-beta (GSK3ß ) a kinase that is responsible for Myc protein degradation. Induction of Myc by insulin is inhibited by the presence of the TOR inhibitor rapamycin, suggesting that insulin-induced Myc protein accumulation depends on the activation of TOR complex 1. Treatment with amino acids that directly activate the TOR pathway results in Myc protein accumulation, which also depends on the ability of S6K kinase to inhibit GSK3ß activity. Myc upregulation by insulin and TOR pathways is a mechanism conserved in cells from the wing imaginal disc, where expression of Dp110 and Rheb also induces Myc protein accumulation, while inhibition of insulin and TOR pathways result in the opposite effect. Our functional analysis, aimed at quantifying the relative contribution of Myc to ommatidial growth downstream of insulin and TOR pathways, revealed that Myc activity is necessary to sustain the proliferation of cells from the ommatidia upon Dp110 expression, while its contribution downstream of TOR is significant to control the size of the ommatidia. CONCLUSIONS: Our study presents novel evidence that Myc activity acts downstream of insulin and TOR pathways to control growth in Drosophila. At the biochemical level we found that both these pathways converge at GSK3ß to control Myc protein stability, while our genetic analysis shows that insulin and TOR pathways have different requirements for Myc activity during development of the eye, suggesting that Myc might be differentially induced by these pathways during growth or proliferation of cells that make up the ommatidia.
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Proteínas de Drosophila/metabolismo , Drosophila melanogaster/metabolismo , Glucógeno Sintasa Quinasa 3/metabolismo , Insulina/metabolismo , Proteínas Proto-Oncogénicas c-myc/genética , Transducción de Señal , Serina-Treonina Quinasas TOR/metabolismo , Animales , Línea Celular , Proteínas de Drosophila/genética , Drosophila melanogaster/genética , Drosophila melanogaster/crecimiento & desarrollo , Ojo/crecimiento & desarrollo , Ojo/metabolismo , Regulación del Desarrollo de la Expresión Génica , Glucógeno Sintasa Quinasa 3 beta , Discos Imaginales/crecimiento & desarrollo , Discos Imaginales/metabolismo , Estabilidad Proteica , Proteínas Proto-Oncogénicas c-myc/metabolismo , Alas de Animales/crecimiento & desarrollo , Alas de Animales/metabolismoRESUMEN
AIMS: In metastatic colorectal carcinomas (mCRC), RAS/RAF genes mutations are first tested to determine the eligibility for anti-EGFR (Epidermal Growth Factor Receptor) therapy in combination with conventional cytotoxic agents. Recent advancements in next-generation sequencing (NGS) have highlighted the potential of multi-gene panels. This multi-gene analysis may provide useful information for the molecular characterisation of mCRC, other than the status of RAS/RAF genes. Aim of this study was to evaluate the feasibility of two NGS custom multi-gene panels in the characterisation of CRC cases and evaluating the relevance of PIK3CA mutation in a routine cohort of consecutive CRC cases. METHODS: A total of 961 formalin-fixed and paraffin-embedded specimens from two medical centres (Bologna and Naples) were analysed using two lab-developed NGS multi-gene panels. RESULTS: KRAS mutations (56.2%) were the more frequent alterations observed in our cohort. Intriguingly, PIK3CA mutations were more frequent (16.8%) than variants observed in the other two genes nowadays analysed in CRC clinical practice (NRAS and BRAF, 4.2% and 9.6%, respectively). Moreover, in more than 10% of samples, coexistent mutations were detected in our cohort of CRC. CONCLUSIONS: Our study demonstrates the feasibility and efficacy of lab-developed targeted multi-gene NGS panels in the clinical practice of CRC. Moreover, the data lead to hypothesise that PIK3CA mutations, together with those of RAS/BRAF, worth to be further investigated in clinical CRC specimens.
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Neoplasias Colorrectales , Proteínas Proto-Oncogénicas B-raf , Fosfatidilinositol 3-Quinasa Clase I/genética , Neoplasias Colorrectales/patología , Genes ras , Humanos , Mutación , Proteínas Proto-Oncogénicas B-raf/genética , Proteínas Proto-Oncogénicas p21(ras)/genéticaRESUMEN
We report on a fetus presenting with an increased nuchal translucency, in which chorionic villus sampling led to the diagnosis of mosaic trisomy 8. Ultrasound scan performed at 15(+6) weeks revealed bilateral cleft lip and palate, flat facial profile, and arrhinia. Pregnancy was terminated at 16(+6); postmortem examination showed additional findings including hypospadias, bilateral renal dysplasia, and focal portal fibrosis of the liver. In order to confirm the presence of trisomy 8, FISH analysis was performed in abnormal renal and hepatic tissue, which, unexpectedly, showed a higher fraction of cells with only one fluorescent probe signal (43% and 23%, respectively), if compared with normal fetal liver and kidney (3-10%). This finding is consistent with the survival in this fetus of a monosomic cell line after mitotic non-disjunction, which is in contrast with what is generally thought about mosaic trisomy genesis. We hypothesize that the possible persistence of the monosomic cell line, in addition to the variable distribution of aneuploid cells in the body tissues, could explain the high heterogeneity of mosaic trisomy 8 phenotype.
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Monosomía/genética , Mosaicismo , Trisomía/diagnóstico , Cariotipo Anormal , Aborto Inducido , Adulto , Autopsia , Muestra de la Vellosidad Coriónica , Cromosomas Humanos Par 8/genética , Fisura del Paladar/diagnóstico por imagen , Fisura del Paladar/patología , Femenino , Feto/patología , Humanos , Hipospadias/diagnóstico , Hipospadias/genética , Hipospadias/patología , Hibridación Fluorescente in Situ , Riñón/patología , Hígado/patología , Masculino , Monosomía/diagnóstico , Monosomía/patología , Medida de Translucencia Nucal , Embarazo , Trisomía/genética , Trisomía/patologíaRESUMEN
Mutations in the superoxide dismutase-1 (SOD1) gene occur in some forms of familial amyotrophic lateral sclerosis (ALS). To date about 150 mutations are known to involve this gene. Here we describe a novel missense mutation in exon 5 of the SOD1 gene in an Italian family with two members affected by ALS. Sequencing of the SOD1 gene was performed on 11 members of the family and 75 healthy controls. Electron microscopy was also performed on one ALS patient. We identified a heterozygous mutation in codon 137 leading to substitution of threonine by alanine. Further studies are needed to clarify the role of this alteration in ALS aetiopathogensis; nevertheless, T137A seems to represent a new missense mutation of the SOD1 gene in ALS patients.
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Esclerosis Amiotrófica Lateral/diagnóstico , Esclerosis Amiotrófica Lateral/genética , Mutación Missense/genética , Superóxido Dismutasa/genética , Adulto , Anciano , Secuencia de Bases , Femenino , Humanos , Italia , Masculino , Persona de Mediana Edad , Datos de Secuencia Molecular , Linaje , Estructura Secundaria de Proteína , Superóxido Dismutasa/química , Superóxido Dismutasa-1RESUMEN
BACKGROUND: Neoplastic overgrowth depends on the cooperation of several mutations ultimately leading to major rearrangements in cellular behaviour. Precancerous cells are often removed by cell death from normal tissues in the early steps of the tumourigenic process, but the molecules responsible for such a fundamental safeguard process remain in part elusive. With the aim to investigate the molecular crosstalk occurring between precancerous and normal cells in vivo, we took advantage of the clonal analysis methods that are available in Drosophila for studying the phenotypes due to lethal giant larvae (lgl) neoplastic mutation induced in different backgrounds and tissues. RESULTS: We observed that lgl mutant cells growing in wild-type imaginal wing discs show poor viability and are eliminated by Jun N-terminal Kinase (JNK)-dependent cell death. Furthermore, they express very low levels of dMyc oncoprotein compared with those found in the surrounding normal tissue. Evidence that this is a cause of lgl mutant cells elimination was obtained by increasing dMyc levels in lgl mutant clones: their overgrowth potential was indeed re-established, with mutant cells overwhelming the neighbouring tissue and forming tumourous masses displaying several cancer hallmarks. Moreover, when lgl mutant clones were induced in backgrounds of slow-dividing cells, they upregulated dMyc, lost apical-basal cell polarity and were able to overgrow. Those phenotypes were abolished by reducing dMyc levels in the mutant clones, thereby confirming its key role in lgl-induced tumourigenesis. Furthermore, we show that the eiger-dependent Intrinsic Tumour Suppressor pathway plays only a minor role in eliminating lgl mutant cells in the wing pouch; lgl-/- clonal death in this region is instead driven mainly by dMyc-induced Cell Competition. CONCLUSIONS: Our results provide the first evidence that dMyc oncoprotein is required in lgl tumour suppressor mutant tissue to promote invasive overgrowth in larval and adult epithelial tissues. Moreover, we show that dMyc abundance inside versus outside the mutant clones plays a key role in driving neoplastic overgrowth.
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Muerte Celular/genética , Transformación Celular Neoplásica/genética , Proteínas de Unión al ADN/metabolismo , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Fenotipo , Factores de Transcripción/metabolismo , Proteínas Supresoras de Tumor/genética , Proteínas Supresoras de Tumor/metabolismo , Animales , Células Epiteliales/metabolismo , Técnica del Anticuerpo Fluorescente , Técnicas de Inactivación de Genes , Mutación/genéticaRESUMEN
BACKGROUND: anaplastic lymphoma kinase (ALK) overexpression and gene alterations have been detected in several malignancies, with prognostic and therapeutic implications. However, few studies investigated the correlation between ALK altered expression and prognosis in patients with glioblastoma (GBM). METHODS: We performed an evaluation of ALK overexpression and structural/quantitative chromosome alterations through immune-histochemical assay (IHC with D5F3 antibody) and fluorescent in situ hybridization (FISH) in patients with isocitrate dehydrogenase (IDH) wild type (wt) GBM. Assuming an ALK overexpression in 20 % of patients we planned a sample of 44 patients to achieve a probability of 90 % to include from 10 % to 30 % of patients with ALK alterations. RESULTS: We evaluated 44 patients with IDH wt GBM, treated in our institution and dead due to GBM progression in 2017. ALK overexpression obtained by a composed score (the product of IHC intensity staining and rate of positive cells) was observed in 19 (43 %) patients. FISH analysis showed that 11 patients (25 %) had gene deletion, 2 patients (4.5 %) had monosomy and one patient (2.3 %) presented polysomy. Only one patient (2.3 %) demonstrated ALK rearrangement. There was no statistical difference in median OS between patients with ALK-positive (mOS = 18.9 months) and ALK-negative IHC (mOS = 18.0 months). CONCLUSION: We identified some rare previously unreported alterations of ALK gene in patients with IDH wt GBM. In these patients, the ALK overexpression does not influences survival.