Five new families with resistance to thyroid hormone not caused by mutations in the thyroid hormone receptor beta gene

Resistance to thyroid hormone (RTH) is a syndrome of variable tissue hyposensitivity to TH. In 191 families, the RTH phenotype has been linked to mutations located in the ligand-binding or hinge domains of the TH receptor (TR) beta gene. The defective TRbeta molecules interfere with the function of the normal TRs to produce dominantly inherited RTH. Of the 65 families with RTH studied in our laboratory, 59 had mutations in the mutagenic region of the TRbeta gene that encompasses exons 7-10. Isolation of a TRbeta PAC (P1 derived artificial chromosome) clone provided the intronic sequences necessary to amplify and sequence the entire TRbeta gene from genomic DNA. Not a single nucleotide substitution, deletion, or insertion was found in all coding and noncoding TRbeta1- and TRbeta2-specific and common exons of the five families with RTH reported herein. Furthermore, linkage analysis using polymorphic markers excluded involvement of the TRbeta and TRalpha genes in two and three of the five families, respectively. The phenotype of RTH in patients without TRbeta gene defects was not different from that in patients with RTH due to TRbeta gene mutations in terms of clinical presentation and reduced responsiveness of the pituitary and peripheral tissues to TH. However, the degree of thyrotroph hyposensitivity to TH appeared to be among the more severe, similar to that of patients with mutant TRbetas that have more than 50-fold reduction of T3 binding affinity and strong dominant negative effect. In these five families and another with non-TRalpha/non-TRbeta RTH, previously identified in our laboratory, evidence for dominant inheritance was secured in two families, and the appearance of a new defect or recessive inheritance was found in the remaining four families. RTH without a structural TRbeta defect occurs in about 10% of families expressing the classic phenotype of TH hyposensitivity, and TRbeta and TRalpha gene involvement has been excluded in 5%. We postulate that a cofactor that interacts with TR is potentially responsible for the manifestation of RTH in these families. As affected subjects are not infertile, the high prevalence of putative neomutations and the low rate of transmission in this non-TR form of RTH may be due to reduced survival of embryos harboring the defect.     

人甲状腺激素受体相互作用蛋白15与甲状腺激素受体的相互作用

目的　确立人甲状腺激素受体相互作用蛋白 15 (hTRIP15 )与甲状腺激素受体 (TR)的相互作用及其作用方式。方法　应用酵母双杂交技术及半乳糖苷酶定性或定量测定。结果　将构建质粒 (BD TRIP15与AD TR)共转化AH10 9酵母细胞后 ,在高严格度选择的培养基 (SD Leu Trp Ade His) 10天克隆长出 ,相比阳性对照质粒 (BD P5 3 +AD Tag)共转后 3天克隆生长。将构建载体及对照载体转化Y187酵母菌株 ,检测LacZ报告基因的表达 (β gal单位 ) ,阳性对照质粒 (BD P5 3 +AD Tag)为 14 9.5 7± 0 .5 1,BD TRIP15 +AD TR为 3 .2 3± 10 .15 ,加入T3 后为 2 .0 2± 0 .0 8,而阴性对照质粒 (BD P5 3 +AD LamC)仅为 0 .0 2± 0 .0 1β gal单位 ,且T3 呈剂量依赖方式抑制两者的相互作用。结论　hTPIR15能与TR共相互作用 ,T3 呈剂量依赖方式抑制两者间的作用 ,仅在超大剂量时才可完全阻断hTRIP15与TRα间的作用。     

Basal and thyroid hormone receptor auxiliary protein-enhanced binding of thyroid hormone receptor isoforms to native thyroid hormone response elements

There are three known isoforms of the rat thyroid hormone receptor, TR alpha-1, TR beta-1, and TR beta-2. The first two are expressed in all tissues, whereas TR beta-2 appears to be expressed only in the pituitary. The differences in the roles of the three receptor isoforms are unknown, but may involve preferential interaction with different subsets of thyroid hormone-regulated genes in different tissues. We tested the binding of the three TR isoforms to putative thyroid hormone response elements (TREs) from genes that are expressed in the pituitary or other tissues and are regulated by thyroid hormone. In vitro translated 35S-labeled rat TR alpha-1, rat TR beta-2, and human TR beta-1 receptors were bound to a battery of biotinylated synthetic deoxyribonucleotides containing naturally occurring putative TREs from genes expressed either in only pituitary (rat glycoprotein hormone alpha-subunit, TSH beta-subunit, and GH) or in nonpituitary (rat alpha-myosin heavy chain, malic enzyme, and Moloney murine leukemia virus promoter) tissues. All three receptor forms bound to each of the TREs. TR beta-2 did not show preferential binding to TREs of pituitary-specific genes compared to TR beta-1. Additionally, TR alpha-1 had a similar TRE-binding pattern as the TR beta s, except for possibly less binding to rat glycoprotein hormone alpha-subunit TRE. Finally, rat pituitary and liver nuclear extracts enhanced TR binding to TREs, with the greatest enhancement seen with the alpha-subunit TRE. These studies suggest that all TR isoforms bind similarly to native TREs. Also, TR binding to TREs can be differentially enhanced by interactions with nuclear proteins.     

Thyroid hormone receptor beta mutations in the 'hot-spot region' are rare events in thyroid carcinomas

Thyroid cancer constitutes the most frequent endocrine neoplasia. Targeted expression of rearranged during transfection (RET)/papillary thyroid carcinoma (PTC) and V600E V-raf murine sarcoma viral oncogene homolog B1 (BRAF) to the thyroid glands of transgenic mice results in tumours similar to those of human PTC, providing evidence for the involvement of these oncogenes in PTC. Kato et al. developed a mouse model that mimics the full spectrum of the human follicular form of thyroid cancer (FTC). FTC rapidly develops in these mice through introduction of the thyroid hormone receptor beta (THRB)(PV) mutant on the background of the inactivated THRB wt locus. Our aim was to verify if, in the context of human follicular thyroid carcinogenesis, THRB acted as a tumour suppressor gene. We screened for mutations of the THRB gene in the hot-spot region, spanning exons 7-10, in 51 thyroid tumours and six thyroid cancer cell lines by PCR and direct sequencing. We did not find mutations in any of the tumours or cell lines analysed. Our findings suggest that, in contrast to the findings on the THRB-mutant transgenic mice, THRB gene mutations are not a relevant mechanism for human thyroid carcinogenesis.     

Two somatic TSH receptor mutations in a patient with toxic metastasising follicular thyroid carcinoma and non-functional lung metastases

In a 59-year-old patient, thyroid follicular cancer was diagnosed in two right-sided toxic thyroid nodules, which had presented clinically as unilateral thyroid autonomy. In addition, the patient had histologically proven lung metastases of thyroid cancer; however, these failed to exhibit iodine uptake and were resistant to radioiodine treatment. The functional activity of the thyroid nodules prompted us to screen for TSH receptor (TSHR) mutations, and the histological diagnosis of follicular carcinoma led us to search for the PAX8-PPARgamma1 rearrangement and mutations in the ras genes. Each thyroid nodule harboured a different TSHR mutation (large nodule, Asp633Tyr; small nodule, Phe631Ile). Presence of both mutations in one sample suggestive of local invasion of a thyroid carcinoma could not be demonstrated, although several specimens from different nodule locations were screened. Only the wild-type TSHR sequence was identified in the histologically normal left thyroid lobe, and no genetic alterations were found in the other investigated genes. No TSHR mutations were detected in the pulmonary metastases. This is the first case report of a patient with toxic follicular thyroid carcinoma harbouring two different TSHR mutations and presenting with non-functional lung metastases.     

Germline mutations in lung cancer

Genetic testing for alterations in oncogenic driver genes has become essential and standard in the clinical practice of the treatment of lung cancer. Germline mutations potentially predisposing patients to lung cancer are rare; however, with the introduction of next-generation sequencing in the clinical practice of lung cancer, the identification of potentially predisposing germline abnormalities is becoming more common. In addition, liquid biopsy, which analyzes cell-free DNA in blood, increases the possibility of detecting these germline mutations. In this review, we summarize the germline mutations detected in lung cancer patients and briefly describe the future perspectives.     

Three Japanese patients from two families with generalized resistance to thyroid hormone with mutations in exon 9 of the thyroid hormone receptor beta gene

Resistance to thyroid hormone (RTH) is a genetic disorder caused by mutations in the thyroid hormone receptor (TR) beta gene. The mutations are clustered in two regions: exon 9 and exon 10. To date, only one patient with an exon 9 mutation has been reported in Japan. We herein report three patients from two Japanese families with RTH and mutations in exon 9. A 52-year-old woman and her 18-year-old daughter, both with inappropriate secretion of TSH (SITSH) were diagnosed simultaneously with generalized RTH. Molecular analysis revealed a G345D mutation. An 11-year-old girl with SITSH, whose only manifestation was a goiter, had an R338W mutation, which is frequently associated with pituitary RTH. Thus, RTH with mutations in exon 9 of the TR beta gene is not so rare in Japan.     

Germline and somatic mutations in exon 15 of the APC gene and K-ras mutations in duodenal adenomas in patients with familial adenomatous polyposis.

BACKGROUND: In sporadic colorectal adenomas mutations in the adenomatous polyposis gene (APC) are among the first gene aberrations to appear. In familial adenomatous polyposis (FAP) the patients already have a germline mutation in the APC gene. To investigate the natural history of duodenal adenomas in FAP patients, we examined germline and somatic mutations of the APC gene and K-ras mutations in these lesions. METHODS: Frozen sections from 54 duodenal polyps from 31 FAP patients were used to histologically verify the presence of adenomatous growth in the mucosa; the rest of each biopsy specimen was processed for DNA extraction. APC exon 15 was investigated with the protein truncation test (PTT), using four overlapping polymerase chain reaction (PCR) fragments, and samples showing an APC mutation were thereafter sequenced. The adenomas were examined for K-ras mutations by use of a combination of the 'enriched PCR method' and temporal temperature gradient electrophoresis. RESULTS: APC germline mutations in exon 15 were found in 19 of 31 (61%) patients, whereas somatic mutations were localized to 12 of 54 (22%) duodenal adenomas. In seven adenomas both the germline and the somatic mutations were found, whereas five small adenomas showed somatic mutations only. There was no tendency for more mutations to be detected in large and severely dysplastic adenomas compared with small and mildly dysplastic ones. K-ras mutations were found in four (7%) duodenal adenomas. CONCLUSIONS: The low rate of somatic APC and K-ras mutations in duodenal adenomas may indicate another neoplastic pathway than in FAP adenomas of the large bowel, or that a modifier gene is cosegregating with the disease.