The modifier role of RET-G691S polymorphism in hereditary medullary thyroid carcinoma: functional characterization and expression/penetrance studies

Colombo, Carla; Minna, Emanuela; Rizzetti, Maria Grazia; Romeo, Paola; Lecis, Daniele; Persani, Luca; Mondellini, Piera; Greco, Angela; Fugazzola, Laura; Borrello, Maria Grazia

doi:10.1186/s13023-015-0231-z

Cited by 25 publications

(23 citation statements)

References 30 publications

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“…Mutations in this gene are associated with the disorders multiple endocrine neoplasia (MEN) type IIA and IIB and medullary thyroid carcinoma ( Krampitz and Norton, 2014 ). In line with previous findings in other endocrine tumour entities ( Rotondi et al , 2009 ; Schulte et al , 2010 ; Colombo et al , 2015 ; Yeganeh et al , 2015 ), we determined RET variants in ∼9% (6 out of 70) of pulmonary NET patients. By the way, the activating p.M918T RET mutation was previously assessed in one metastatic SCLC specimen ( Dabir et al , 2014 ).…”

Section: Discussionsupporting

confidence: 85%

Mutational analysis of pulmonary tumours with neuroendocrine features using targeted massive parallel sequencing: a comparison of a neglected tumour group

et al. 2015

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Background:Lung cancer is the leading cause of cancer-related deaths worldwide. The typical and atypical carcinoid (TC and AC), the large-cell neuroendocrine carcinoma (LCNEC) and the small-cell lung cancers (SCLC) are subgroups of pulmonary tumours that show neuroendocrine differentiations. With the rising impact of molecular pathology in routine diagnostics the interest for reliable biomarkers, which can help to differentiate these subgroups and may enable a more personalised treatment of patients, grows.Methods:A collective of 70 formalin-fixed, paraffin-embedded (FFPE) pulmonary neuroendocrine tumours (17 TCs, 17 ACs, 19 LCNECs and 17 SCLCs) was used to identify biomarkers by high-throughput sequencing. Using the Illumina TruSeq Amplicon-Cancer Panel on the MiSeq instrument, the samples were screened for alterations in 221 mutation hot spots of 48 tumour-relevant genes.Results:After filtering >26 000 detected variants by applying strict algorithms, a total of 130 mutations were found in 29 genes and 49 patients. Mutations in JAK3, NRAS, RB1 and VHL1 were exclusively found in SCLCs, whereas the FGFR2 mutation was detected in LCNEC only. KIT, PTEN, HNF1A and SMO were altered in ACs. The SMAD4 mutation corresponded to the TC subtype. We prove that the frequency of mutations increased with the malignancy of tumour type. Interestingly, four out of five ATM-mutated patients showed an additional alteration in TP53, which was by far the most frequently altered gene (28 out of 130; 22%). We found correlations between tumour type and IASLC grade for ATM- (P=0.022; P=0.008) and TP53-mutated patients (P<0.001). Both mutated genes were also associated with lymph node invasion and distant metastasis (P⩽0.005). Furthermore, PIK3CA-mutated patients with high-grade tumours showed a reduced overall survival (P=0.040) and the mutation frequency of APC and ATM in high-grade neuroendocrine lung cancer patients was associated with progression-free survival (PFS) (P=0.020).Conclusions:The implementation of high-throughput sequencing for the analysis of the neuroendocrine lung tumours has revealed that, even if these tumours encompass several subtypes with varying clinical aggressiveness, they share a number of molecular features. An improved understanding of the biology of neuroendocrine tumours will offer the opportunity for novel approaches in clinical management, resulting in a better prognosis and prediction of therapeutic response.

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Section: Discussionsupporting

confidence: 85%

Mutational analysis of pulmonary tumours with neuroendocrine features using targeted massive parallel sequencing: a comparison of a neglected tumour group

et al. 2015

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“…Total protein extraction, SDS PAGE and Western blot analyses were performed as previously described [ 69 ]. The primary antibody c-MYC (#5605), pAkt (Ser473, #4060), mTOR (#2983), pmTOR (Ser2448, #5536), S6 (#2217) and pS6 (Ser235/236, #4858) are from Cell Signaling Technology (Cell Signaling Technology Inc., MA, USA); MIF (ab175189) from Abcam (Abcam Inc., Cambridge, UK); AKT specific for AKT1 (#610861) from BD Transduction Laboratories (BD Biosciences, NJ, USA); Actin (A2066) from Sigma-Aldrich and GAPDH (#sc-32233) from Santa Cruz Biotechnology (Santa Cruz Biotechnology Inc., CA, USA).…”

Section: Methodsmentioning

confidence: 99%

miR-451a is underexpressed and targets AKT/mTOR pathway in papillary thyroid carcinoma

et al. 2016

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Papillary Thyroid Carcinoma (PTC) is the most frequent thyroid cancer. Although several PTC-specific miRNA profiles have been reported, only few upregulated miRNAs are broadly recognized, while less consistent data are available about downregulated miRNAs. In this study we investigated miRNA deregulation in PTC by miRNA microarray, analysis of a public dataset from The Cancer Genome Atlas (TCGA), literature review and meta-analysis based on a univocal miRNA identifier derived from miRBase v21. A list of 18 miRNAs differentially expressed between PTC and normal thyroid was identified and validated in the TCGA dataset. Furthermore, we compared our signature with miRNA profiles derived from 15 studies selected from literature. Then, to select possibly functionally relevant miRNA, we integrated our miRNA signature with those from two in vitro cell models based on the PTC-driving oncogene RET/PTC1. Through this strategy, we identified commonly deregulated miRNAs, including miR-451a, which emerged also by our meta-analysis as the most frequently reported downregulated miRNA. We showed that lower expression of miR-451a correlates with aggressive clinical-pathological features of PTC as tall cell variant, advanced stage and extrathyroid extension. In addition, we demonstrated that ectopic expression of miR-451a impairs proliferation and migration of two PTC-derived cell lines, reduces the protein levels of its recognized targets MIF, c-MYC and AKT1 and attenuates AKT/mTOR pathway activation.Overall, our study provide both an updated overview of miRNA deregulation in PTC and the first functional evidence that miR-451a exerts tumor suppressor functions in this neoplasia.

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“…Study on a larger cohort of Javenese HSCR patients needs to be performed to confirm this hypothesis. 14,16 On the other hand, one cannot exclude that it is the combination of two variant alleles of the same gene (not in cis) that give the effect or that it is the effect of variants in different genes that give rise to the HSCR phenotype. In silico analysis using online software proved useful, especially when we are dealing with many variants to be functionally studied.…”

Section: Ednrb Variantsmentioning

confidence: 99%

“…The rest of those variants (35%) were believed to be non-causative variants. [13][14][15][16] It can therefore not be excluded that part of the reported mutations merely are benign, rare polymorphic variants.…”

Section: Introductionmentioning

confidence: 99%

RET and EDNRB mutation screening in patients with Hirschsprung disease: Functional studies and its implications for genetic counseling

et al. 2015

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Hirschsprung disease (HSCR) is a major cause of chronic constipation in children. HSCR can be caused by germline mutations in RET and EDNRB. Defining causality of the mutations identified is difficult and almost exclusively based on in silico predictions. Therefore, the reported frequency of pathogenic mutations might be overestimated. We combined mutation analysis with functional assays to determine the frequencies of proven pathogenic RET and EDNRB mutations in HSCR. We sequenced RET and EDNRB in 57 HSCR patients. The identified RET-coding variants were introduced into RET constructs and these were transfected into HEK293 cells to determine RET phosphorylation and activation via ERK. An exon trap experiment was performed to check a possible splice-site mutation. We identified eight rare RET-coding variants, one possible splice-site variant, but no rare EDNRB variants. Western blotting showed that three coding variants p. (Pr270Leu) (Tyr1062Cys)) resulted in reduced ERK activation. Splice-site assays on c.1880-11A4G could not confirm its pathogenicity. Our data suggest that indeed almost half of the identified rare variants are proven pathogenic and that, hence, functional studies are essential for proper genetic counseling.

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The modifier role of RET-G691S polymorphism in hereditary medullary thyroid carcinoma: functional characterization and expression/penetrance studies

Cited by 25 publications

References 30 publications

Mutational analysis of pulmonary tumours with neuroendocrine features using targeted massive parallel sequencing: a comparison of a neglected tumour group

Mutational analysis of pulmonary tumours with neuroendocrine features using targeted massive parallel sequencing: a comparison of a neglected tumour group

miR-451a is underexpressed and targets AKT/mTOR pathway in papillary thyroid carcinoma

RET and EDNRB mutation screening in patients with Hirschsprung disease: Functional studies and its implications for genetic counseling

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