Supplementary Material for: Clinical Consequences of Variable Results in the Measurement of Free Thyroid Hormones: Unusual Presentation of a Family with a Novel Variant in the <b><i>THRB</i></b> Gene Causing Resistance to Thyroid Hormone Syndrome

Introduction: Resistance to thyroid hormone β (RTHβ) is an inherited syndrome caused by dominant negative variants in the THRB gene (NM_000461.5). The clinical picture of RTHβ is variable, and patients harboring the same variant may display different degrees of disease severity. Case Presentation: A 30-year-old man presented with thyrotoxicosis and central hyperthyroidism and was found to have a novel variant in the exon 10 of THRB gene (c.C1282G, p.L428V), located within the third hot spot region of the C-terminal of the receptor. Surprisingly, the same variant was found in two other relatives with an apparent normal thyroid function at initial screening. After exclusion of a TSH-secreting adenoma and serum interference in the proband, and the finding that exogenous levothyroxine failed to suppress the TSH in the brother affected by nodular goiter, relatives’ thyroid function tests (TFTs) were reassessed with additional analytical method revealing biochemical features consistent with RTHβ in all carriers of the p.L428V variant. Functional studies showed a slightly impaired in vitro transcriptional activity of p.L428V. Interestingly‚ the expression of the human p.L428V thyroid hormone receptor beta in the zebrafish embryo background generated a phenotype consistent with RTHβ. Conclusion: Variable results of TFTs on some immunoassays can be a cause of RTHβ diagnostic delay, but the genotype-phenotype correlation in this family and functional studies support p.L428V as a novel THRB variant expanding the spectrum of gene variants causing RTHβ. In vivo, rather than in vitro, functional assays may be required to demonstrate the dominant negative action of THRB variants.引言：甲状腺激素β抵抗症（Resistance to thyroid hormone β, RTHβ）是一种由THRB基因（NM_000461.5）上的显性负效变异引发的遗传性综合征。RTHβ的临床表现具有异质性，携带同一变异的患者其疾病严重程度可存在显著差异。 病例报告：一名30岁男性因甲状腺毒症及中枢性甲状腺功能亢进症就诊，检测发现其THRB基因第10外显子存在一处新型变异（c.C1282G, p.L428V），该变异位于受体羧基端的第三个热点区域内。令人意外的是，在初始筛查中甲状腺功能看似正常的另外两名亲属体内，也检测到了同一变异。在排除先证者的促甲状腺激素（TSH）分泌腺瘤及血清干扰，且确认罹患结节性甲状腺肿的兄长对外源性左甲状腺素无法产生TSH抑制反应后，研究人员采用额外的检测方法对亲属的甲状腺功能检测（TFTs）进行了复检，结果显示所有携带p.L428V变异的亲属均呈现出与RTHβ相符的生化特征。功能研究显示，p.L428V变异体的体外转录活性存在轻度受损。有趣的是，在斑马鱼胚胎模型中表达人类p.L428V甲状腺激素β受体，可诱导出与RTHβ相符的表型。 结论：部分免疫检测方法的甲状腺功能检测结果存在异质性，可能是导致RTHβ诊断延迟的原因之一；但本家系的基因型-表型关联分析及功能研究证实，p.L428V是一种新型THRB变异，拓宽了致病RTHβ的基因变异谱。相较于体外实验，体内功能试验或许更适用于验证THRB变异的显性负效作用。