Yufei Shi

Low rate of ret proto-oncogene activation (PTC/retTPC) in papillary thyroid carcinomas from saudi arabia

Background. The ret proto‐oncogene activation (PTC/retTPC oncogene) in thyroid papillary carcinoma has been reported in different populations with different frequencies. Thyroid papillary carcinoma appears to behave more aggressively in the Persian Gulf region than elsewhere. In the current study, the frequency of PTC/retTPC oncogene in thyroid tumors from Saudi Arabia was investigated.

High frequency of somatic mitochondrial DNA mutations in human thyroid carcinomas and complex I respiratory defect in thyroid cancer cell lines.

Significant progress has been made to elucidate the molecular mechanisms that determine thyroid tumor development and progression. However, most investigations have mainly focused on the genetic alterations of nuclear DNA. The potential role of mitochondrial DNA (mtDNA) mutations in thyroid tumorigenesis is not well defined. In the present study, we investigated the frequency of mtDNA mutations in 24 thyroid tumor specimens (19 primary papillary thyroid carcinomas (PTC), one follicular thyroid carcinoma, and four multinodular hyperplasias) and four thyroid cancer cell lines by sequencing the entire coding regions of mitochondrial genome. Among the 19 PTC samples tested, seven (36.8%) had somatic mutations. Somatic mtDNA mutations were also detected in one of four multinodular hyperplasias examined. All the thyroid tumor cell lines carried sequence variations that change amino acid and have not been reported previously as normal sequence variants. Flow cytometry analysis of mitochondria respiratory function in the thyroid tumor cell lines revealed a severe defect in mitochondrial complex I activity. The majority of the mutations was involved in genes located in the complex I of the mitochondrial genome. The mutations were either A → G or C → T transitions, often resulting in a change of a moderately or highly conserved amino acid of their corresponding protein. These data suggest that mtDNA mutations may play an important role in the thyroid tumorigenesis. Given that mtDNA mutation is present in the benign multinodular hyperplasia, it might be involved in the early stage of tumor development.

Inverse association between cyclin D1 overexpression and retinoblastoma gene mutation in thyroid carcinomas

Cyclin D1 plays a key role in the regulation of the G1/S transition through the cell cycle. Deregulation of cyclin D1, most often leading to overexpression of the gene, has been reported in many tumor types. It has been suggested that cyclin D1 overexpression could be an alternative mechanism for pRb inactivation. We have previously found Rb gene mutations in 55% of malignant thyroid tumors. In the present study, we examined the cyclin D1 gene expression and amplification in 24 tumor samples (two of them are benign goiters) randomly selected from the same series of thyroid tumors, to see whether cyclin D1 over-expression is present in those specimens without Rb gene mutations. We found a four- to fivefold increase in cyclin D1 expression in 7 of 22 thyroid carcinomas as compared with that in benign nodular goiters. Six of them were found in carcinomas without Rb gene mutations. Among the remaining 15 thyroid carcinoma samples, 11 were found previously to have Rb gene mutations. The association between increased cyclin D1 expression and absence of Rb mutation is statistically significant (p < 0.05). We found no evidence of the cyclin D1 gene amplification or rearrangement to account for such an increase in cyclin D1 expression. We conclude that cyclin D1 overexpression may be relevant to thyroid carcinogenesis. Two mechanisms may be involved in the inactivation of pRb: one is through Rb gene mutations, and the other is by cyclin D1 overexpression.

Expression of thyrotrophin receptor gene in thyroid carcinoma is associated with a good prognosis

OBJECTIVE The clinical course of thyroid carcinoma is very variable. It is well known that thyroid carcinomas of similar histology can behave differently in terms of local invasion and distant metastases: there is no reliable way to predict the disease course with confidence. In the present study we compared the TSH receptor and c‐myc mRNA levels in different stages of thyroid carcinomas to identify whether they are useful markers for thyroid tumour biological behaviour and prognosis.

Aberrant BRAF splicing as an alternative mechanism for oncogenic B-Raf activation in thyroid carcinoma†

Activating BRAF mutations have recently been reported in 28–83% of papillary thyroid carcinomas (PTCs). However, it is not known whether aberrant BRAF splicing occurs in thyroid carcinoma. To investigate aberrant BRAF splicing and its association with BRAF mutation in thyroid tumours, we studied aberrant BRAF splicing and BRAF mutation from 68 thyroid tumours. BRAFV600E mutation was detected in 20 of 43 PTCs and all three anaplastic thyroid carcinomas (ATCs). There is a higher frequency of BRAF mutation in PTC patients with stage III and IV tumours compared with stage I and II. Novel BRAF splicing variants were detected in 12 PTCs, three follicular variants of PTC (FVPTCs), and one ATC, as well as in two thyroid carcinoma cell lines, ARO and NPA. These variants did not have the N‐terminal auto‐inhibitory domain of wild‐type B‐Raf, resulting in an in‐frame truncated protein that contained only the C‐terminal kinase domain and caused constitutive activation of B‐Raf. These variants were significantly associated with advanced disease stage and BRAFV600E mutation (p < 0.001, Fisher exact test). Furthermore, expression of these variants in NIH3T3 and CHO cells could activate the MAP kinase signalling pathway, transform them in vitro, and induce tumours in nude mice. These data suggest that BRAF splicing variants may function as an alternative mechanism for oncogenic B‐Raf activation. Combination of the BRAFV600E mutation and its splicing variants may contribute towards disease progression to poorly differentiated thyroid carcinoma. Copyright

Association of mitochondrial DNA transversion mutations with familial medullary thyroid carcinoma/multiple endocrine neoplasia type 2 syndrome.

Medullary thyroid carcinoma (MTC) is a malignant tumour of the calcitonin-secreting parafollicular C cells of the thyroid, and occurs sporadically or as a component of the multiple endocrine neoplasia (MEN) type 2/familial medullary thyroid carcinoma (FMTC) syndromes. In the present study, we investigated the frequency of mtDNA mutations in 26 MTC tumour specimens (13 sporadic and 13 familial MTC) and their matched normal tissues by sequencing the entire coding regions of mitochondrial genome. Nonsynonymous mutations were detected in 20 MTC samples (76.9%): nine out of 13 sporadic MTC (69.2%) and 11 out of 13 (84.6%) familial MTC/MEN2. Both transition and transversion types of mutations were found in the samples. Interestingly, 76.2% (16/21) of transversion mutations were found in FMTC/MEN2 patients, whereas 66.7% (12/18) of transition mutations were in sporadic MTC. Synonymous mutations were found in 12 MTC samples. In total, we identified 27 transversion mutations (21 nonsynonymous and six synonymous) in MTC. Of them, 22 (81.5%) were from FMTC/MEN2, and five (18.5%) were from sporadic MTC. The association of transversion mutation with familial MTC/MEN2 was statistically significant (P=0.0015, binomial test). Majority of the mutations were involved in the genes located in the complex I of the mitochondrial genome, and were often resulting in a change of a moderately or highly conserved amino acid of their corresponding protein. Mitochondrial respiratory function was also compromised in a TT cell line, which carries mtDNA mutation at nt 4917 and 11 720, and in peripheral lymphocytes of MTC patients with mtDNA mutations. These data suggest that mtDNA mutation may be involved in MTC tumourigenesis and progression. Given that mtDNA mutation spectra are different between sporadic and familial MTC, different mechanisms of oxidative DNA damage may occur in the disease process.

A Novel G102E Mutation of CYP27B1 in a Large Family with Vitamin D-Dependent Rickets Type 1

CONTEXT Mutations in the CYP27B1 gene, which encodes vitamin D 1alpha-hydroxylase, are the genetic basis for vitamin D-dependent rickets type 1 (VDDR-I). OBJECTIVE The aim of this study was to investigate the CYP27B1 mutation in a large family with VDDR-I and characterize the genotype-phenotype correlation. PATIENTS AND METHODS The index patient was a 23-yr-old female who had a progressive form of rickets and growth retardation since the age of 9 months. Laboratory data showed hypocalcemia, low urine calcium, hypophosphatemia, high serum alkaline phosphatase, elevated PTH, and low serum 1,25-dihydroxyvitamin D(3). Her parents were healthy first-degree cousins, and two of her 12 siblings were affected with similar but milder rickets. Three other siblings were asymptomatic but had biochemical evidence of the disease. The entire coding region of the CYP27B1 gene was sequenced, and the mutation was characterized by functional studies. RESULTS We found a novel biallelic c.305G>A sequence variation at codon 102, changing amino acid from glycine to glutamic acid (G102E) in the patient and five affected siblings, whereas a monoallelic c.305G>A variation was present in the mother and five nonaffected siblings. This variation was not present in 100 population controls. Expression of this mutant in CHO cells revealed an 80% reduction in the 1alpha-hydroxylase activity as compared to wild-type activity. CONCLUSIONS A novel mutation in the CYP27B1 gene was found in patients with VDDR-I. This mutation resulted in a significant reduction in 1alpha-hydroxylase activity. The residual enzymatic activity may account for the mild phenotype presentation in some affected members.

A novel deletion of the MEN1 gene in a large family of multiple endocrine neoplasia type 1 (MEN1) with aggressive phenotype

Context  The MEN1 syndrome is associated with parathyroid, pancreatic and pituitary tumours and is caused by mutations in the MEN1 gene. In general, there is no genotype–phenotype correlation.

The expression of the MDM2 gene, a p53 binding protein, in thyroid carcinogenesis.

Background. The authors previously found p53 mutations in 24% of malignant thyroid tumors, representing a wide staging spectrum. Overexpression of MDM2, most often due to gene amplification, has ben suggested to be ab additional mechanism for abrogation of the p53 function. In the current study. MDM2 gene expression and amplification were examined in a randomly selected subset of these tumors to explore the possibility that wildtype p53 may be inactivated by complexing with MDM2 in specimens without p53 mutations.

Biallelic p.R2223H Mutation in the Thyroglobulin Gene Causes Thyroglobulin Retention and Severe Hypothyroidism with Subsequent Development of Thyroid Carcinoma

CONTEXT Dyshormonogenesis due to genetic defect in thyroglobulin (Tg) synthesis and secretion can lead to congenital hypothyroidism. OBJECTIVES The aim of the study was to analyze the TG gene for the presence of mutations and to study the underlying mechanisms leading to dyshormonogenesis. CASES Two siblings aged 25 and 31 yr presented with recurrent goitrous hypothyroidism with undetectable serum Tg. The older sibling was diagnosed with follicular variant of papillary thyroid carcinoma (FVPTC) at age 21 and metastatic FVPTC 8 yr later. METHODS The entire coding region of TG gene was sequenced. BRAF, RAS, and P53 mutations or PAX8/PPAR-gamma rearrangement were screened in the FVPTC. Tg expression was studied by immunohistochemistry. RESULTS Biallelic c.6725G>A (p.R2223H) and c.6396C>T (p.S2113L) sequence variations were detected in both patients and monoallelic variations in their family members. The c.6396C>T (p.S2113L) sequence variation was found in 14% of 100 population controls, whereas c.6725G>A variation was not present in the controls. Two previously reported polymorphisms (c.2200T>G and c.3082A>G) were present in all the family members. Strong cytoplasmic immunostaining of Tg was observed in the hyperplastic thyroid epithelial cells and weak or no staining in the follicular lumen. Cytoplasmic staining was localized in the endoplasmic reticulum. Reduced staining was found in the FVPTC. Neither RAS, BRAF, or P53 gene mutation nor a PAX8/PPAR-gamma rearrangement was detected in the tumor tissue. CONCLUSIONS Biallelic c.6725G>A (p.R2223H) mutation causes Tg retention in the endoplasmic reticulum, resulting in dyshormonogenesis. Prolonged TSH stimulation may promote malignant transformation and development of thyroid cancer. The c.6396C>T (p.S2113L) is a novel polymorphism.