Supplementary Material for: Hyperfunctioning Papillary Thyroid Carcinoma with a <b><i>BRAF</i></b> Mutation: The First Case Report and a Literature Review

<b><i>Introduction:</i></b> Hyperfunctioning papillary thyroid carcinoma (PTC) is rare and consequently, little information on its molecular etiology is available. Although <i>BRAF</i> V600E (<i>BRAF</i> c.1799T&gt;A, p.V600E) is a prominent oncogene in PTC, its mutation has not yet been reported in hyperfunctioning PTC. <b><i>Case Presentation:</i></b> Ultrasonography detected a 26-mm nodule in the right lobe of the thyroid gland of a 48-year-old man. Thyroid function tests indicated that he was hyperthyroid with a TSH level of 0.01 mIU/L (reference range: 0.05–5.00) and a free thyroxine level of 23.2 pmol/L (reference range: 11.6–21.9). TSHR autoantibodies were &lt;0.8 IU/L (reference value: &lt;2.0 IU/L). The ^99mTc thyroid scintigram revealed a round, right-sided focus of tracer uptake by the nodule with a decreased uptake in the remainder of the gland. The patient underwent total thyroidectomy because fine-needle aspiration cytology revealed a malignancy. The histopathological diagnosis was conventional PTC. Subsequent mutational analysis of <i>BRAF</i> (exon 15), <i>TSHR</i> (exons 1–10), <i>GNAS</i> (exons 7–10), <i>EZH1</i> (exon 16), <i>KRAS</i>, <i>NRAS</i>, <i>HRAS</i> (codons 12, 13, and 61), and <i>TERT</i> promoter (C250T and C228T) identified a heterozygous point mutation in <i>BRAF</i> V600E in a tumor tissue sample. In addition, we identified a <i>TSHR</i> D727E polymorphism (<i>TSHR</i> c.2181C&gt;G, p.D727E) in both the tumor and the surrounding normal thyroid tissue. <b><i>Discussion and Conclusions:</i></b> We report a case of hyperfunctioning PTC with a <i>BRAF</i> V600E mutation for the first time. Our literature search yielded 16 cases of hyperfunctioning thyroid carcinoma in which a mutational analysis was conducted. We identified <i>TSHR</i> mutations in 13 of these cases. One case revealed a combination of <i>TSHR</i> and <i>KRAS</i> mutations; the other case revealed a <i>TSHR</i> mutation with a <i>PAX8/PPARG</i> rearrangement. These findings suggest that the concomitant activation of oncogenes (in addition to constitutive activation of the TSHR-cyclic AMP cascade) are associated with the malignant phenotype in hyperfunctioning thyroid nodules.<b><i>引言：</i></b> 高功能性乳头状甲状腺癌（papillary thyroid carcinoma, PTC）较为罕见，目前关于其分子病因学的研究资料相对匮乏。尽管BRAF V600E突变（BRAF c.1799T>A, p.V600E）是乳头状甲状腺癌中重要的致癌基因突变，但尚未见该突变在高功能性乳头状甲状腺癌中被报道的案例。<b><i>病例报告：</i></b> 一名48岁男性患者，经超声检查于甲状腺右叶发现一枚直径26mm的结节。甲状腺功能检测结果提示患者存在甲状腺功能亢进：促甲状腺激素（TSH）水平为0.01 mIU/L（参考范围：0.05~5.00 mIU/L），游离甲状腺素水平为23.2 pmol/L（参考范围：11.6~21.9 pmol/L）；促甲状腺激素受体（TSHR）抗体水平<0.8 IU/L（参考值：<2.0 IU/L）。锝-99m（⁹⁹ᵐTc）甲状腺闪烁扫描显示，结节部位呈圆形，右侧甲状腺区域显像剂摄取增强，其余甲状腺组织显像剂摄取减少。因细针穿刺细胞学检查提示恶性病变，患者接受了甲状腺全切除术。术后组织病理学诊断为经典型乳头状甲状腺癌。后续针对BRAF（第15外显子）、TSHR（第1~10外显子）、GNAS（第7~10外显子）、EZH1（第16外显子）、KRAS、NRAS、HRAS（12、13、61号密码子）以及TERT启动子（C250T和C228T位点）进行突变检测，结果在肿瘤组织样本中检出BRAF V600E杂合点突变。此外，在肿瘤组织及周围正常甲状腺组织中均检出TSHR D727E多态性（TSHR c.2181C>G, p.D727E）。<b><i>讨论与结论：</i></b> 本研究首次报道一例携带BRAF V600E突变的高功能性乳头状甲状腺癌病例。经文献检索，共找到16例接受过突变检测的高功能性甲状腺癌病例，其中13例检出TSHR基因突变。1例同时存在TSHR与KRAS基因突变，另有1例检出TSHR基因突变并伴随PAX8/PPARG重排。上述结果提示，除促甲状腺激素受体-环磷酸腺苷（cAMP）级联通路的组成性激活外，癌基因的协同激活与高功能性甲状腺结节的恶性表型密切相关。