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Diagnóstico Diferencial , Leiomiosarcoma/diagnóstico , Liposarcoma/diagnóstico , Proteínas Proto-Oncogénicas c-mdm2/genética , Neoplasias Uterinas/diagnóstico , Femenino , Amplificación de Genes , Humanos , Leiomiosarcoma/genética , Leiomiosarcoma/secundario , Liposarcoma/genética , Persona de Mediana Edad , Neoplasias Uterinas/genética , Neoplasias Uterinas/patologíaRESUMEN
Pancreatic undifferentiated carcinoma is a heterogeneous group of neoplasms, including pleomorphic giant cell, sarcomatoid, round cell, and rhabdoid carcinomas, the molecular profiles of which have so far been insufficiently characterized. We studied 14 undifferentiated carcinomas with prominent rhabdoid cells, occurring as advanced tumors in seven females and seven males aged 44-96 years (mean: 65 years). Histologically, 10 tumors qualified as pleomorphic giant cell and 4 as monomorphic anaplastic carcinomas. A glandular component, either in the primary or in the metastases, was seen in 5 out of 14 tumors (4 out of 10 pleomorphic giant cell and 1 out of 4 monomorphic anaplastic subtypes, respectively). Osteoclast-like giant cells were absent. Immunohistochemistry revealed coexpression of cytokeratin and vimentin, and loss of membranous ß-catenin and E-cadherin staining in the majority of cases. Nuclear SMARCB1 (INI1) expression was lost in 4 out of 14 cases (28%), representing all 4 tumors of the monomorphic anaplastic subtype. FISH and mutation testing of KRAS revealed KRAS amplification in 5 out of 13 (38%) and exon 2 mutations in 6 out of 11 (54%) successfully analyzed cases. A strong correlation was found between KRAS alterations (mutation and/or copy number changes) and intact SMARCB1 expression (7 out of 8; 87%). On the other hand, loss of SMARCB1 expression correlated with the absence of KRAS alterations (3 out of 5 cases; 60%). The results suggest that rhabdoid phenotype in pancreatic undifferentiated rhabdoid carcinomas has a heterogeneous genetic background. SMARCB1 loss is restricted to the anaplastic monomorphic subtype and correlates with the absence of KRAS alterations, whereas the pleomorphic giant cell subtype is characterized by KRAS alterations and intact SMARCB1 expression. Recognition and appropriate subtyping of these rare variants might become necessary for future therapeutic strategies.
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Proteínas Cromosómicas no Histona/biosíntesis , Proteínas de Unión al ADN/biosíntesis , Neoplasias Pancreáticas/patología , Proteínas Proto-Oncogénicas/genética , Tumor Rabdoide/patología , Factores de Transcripción/biosíntesis , Proteínas ras/genética , Adulto , Anciano , Anciano de 80 o más Años , Biomarcadores de Tumor/análisis , Análisis Mutacional de ADN , Femenino , Humanos , Inmunohistoquímica , Hibridación Fluorescente in Situ , Masculino , Persona de Mediana Edad , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/metabolismo , Proteínas Proto-Oncogénicas p21(ras) , Tumor Rabdoide/genética , Tumor Rabdoide/metabolismo , Proteína SMARCB1RESUMEN
OBJECTIVES: Recurrent gene fusions of anaplastic lymphoma receptor tyrosine kinase (ALK) and echinoderm microtubule-associated protein-like 4 (EML4) have been recently identified in â¼5% of non-small cell lung cancers (NSCLCs) and are targets for selective tyrosine kinase inhibitors. While fluorescent in situ hybridization (FISH) is the current gold standard for detection of EML4-ALK rearrangements, several limitations exist including high costs, time-consuming evaluation and somewhat equivocal interpretation of results. In contrast, targeted massive parallel sequencing has been introduced as a powerful method for simultaneous and sensitive detection of multiple somatic mutations even in limited biopsies, and is currently evolving as the method of choice for molecular diagnostic work-up of NSCLCs. MATERIALS AND METHODS: We developed a novel approach for indirect detection of EML4-ALK rearrangements based on 454 massive parallel sequencing after reverse transcription and subsequent multiplex amplification (multiplex ALK RNA-seq) which takes advantage of unbalanced expression of the 5' and 3' ALK mRNA regions. Two lung cancer cell lines and a selected series of 32 NSCLC samples including 11 cases with EML4-ALK rearrangement were analyzed with this novel approach in comparison to ALK FISH, ALK qRT-PCR and EML4-ALK RT-PCR. RESULTS: The H2228 cell line with known EML4-ALK rearrangement showed 171 and 729 reads for 5' and 3' ALK regions, respectively, demonstrating a clearly unbalanced expression pattern. In contrast, the H1299 cell line with ALK wildtype status displayed no reads for both ALK regions. Considering a threshold of 100 reads for 3' ALK region as indirect indicator of EML4-ALK rearrangement, there was 100% concordance between the novel multiplex ALK RNA-seq approach and ALK FISH among all 32 NSCLC samples. CONCLUSION: Multiplex ALK RNA-seq is a sensitive and specific method for indirect detection of EML4-ALK rearrangements, and can be easily implemented in panel based molecular diagnostic work-up of NSCLCs by massive parallel sequencing.
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Carcinoma de Pulmón de Células no Pequeñas/genética , Secuenciación de Nucleótidos de Alto Rendimiento/métodos , Neoplasias Pulmonares/genética , Proteínas de Fusión Oncogénica/análisis , Proteínas de Fusión Oncogénica/genética , Adulto , Anciano , Anciano de 80 o más Años , Femenino , Humanos , Masculino , Persona de Mediana Edad , Reacción en Cadena de la Polimerasa Multiplex/métodos , Sensibilidad y Especificidad , Análisis de Secuencia de ARN/métodosRESUMEN
BACKGROUND: Typical Williams-Beuren syndrome (WBS) is commonly caused by a ~1.5 Mb - ~1.8 Mb heterozygous deletion of contiguous genes at chromosome region 7q11.23. The majority of WBS cases occurs sporadically but few familial cases of autosomal dominant inheritance have been reported. Recent data demonstrated the existence of the paracentric inversion polymorphism at the WBS critical region in 7q11.23 in some of the progenitors transmitting the chromosome which shows the deletion in the affected child. In parents having a child affected by WBS the prevalence of such a structural variant has been reported to be much higher (~25- ~30%) than in the general population (~1- ~6%). However, in these previously reported studies only a limited number of randomly selected patients and non transmitting parents of WBS patients were used as controls, but without specification of any clinical data. Therefore we have undertaken a German population-based molecular cytogenetic investigation. We evaluated the incidence of the paracentric inversion polymorphism at 7q11.23 analyzing interphase nuclei of lymphocytes using a three color fluorescence in situ hybridization (FISH) probe. RESULTS: FISH analysis was carried out on couples with a child affected by WBS as compared to a population sample composed of different normal individuals: Control group I: couples with two healthy children, control group II: couples with fertility problems, planning ICSI and control group III: couples with two healthy children and one child with a chromosome aberration, not involving region 7q11.23. The three color FISH assay showed that the frequency of the paracentric inversion polymorphism at 7q11.23 in couples with a child affected by WBS was 20.8% (5 out of 24 pairs) as compared to 8.3% (2 out of 24 pairs, control group I), 25% (4 out of 16 pairs, control group II) and 9.1% (1 out of 11 pairs, control group III), respectively (total 7 out of 51 pairs, 13.8%). The frequencies differed between the groups, but this was statistically not significant (p > 0.05, Fisher's test). CONCLUSION: Our results do not support the hypothesis that the paracentric inversion polymorphism at 7q11.23 is a major predisposing factor for the WBS deletion.