目的探索二代测序（NGS）技术检测超声引导下细针穿刺活检（US-FNA）单发甲状腺微小结节（TMN）标本中BRAF、RAS、TERT启动子和TP53基因突变情况及其诊断甲状腺微小癌（TMC）的价值。
方法现场招募2018年9月至2021年7月于首都医科大学附属北京同仁医院住院及门诊接受甲状腺超声检查、超声可疑单发TMC且随后进行了US-FNA及NGS的428例TMN患者，最终纳入147例患者。采用NGS技术对US-FNA标本进行BRAF、RAS、TERT启动子和TP53基因突变检测。对比BRAF基因突变组与非基因突变组患者年龄、性别及结节最大径的差异。以术后病理诊断为金标准，通过受试者工作特征（ROC）曲线计算BRAF基因突变诊断TMC的效能。
结果147例患者的年龄［ M（ Q 1， Q 3）］为43.0（32.0，51.0）岁，男37例（25.2%）。147份US-FNA标本中，97份（66.0%）检测出BRAF基因突变，全部为p.V600E点突变；6份（4.1%）检测出RAS基因突变，未检出TERT启动子和TP53基因突变。术后病理证实136例（92.5%）为TMC，病理类型全部为甲状腺微小乳头状癌（PTMC）；11例（7.5%）为良性。其中，136份TMC标本中有97份（71.3%）检测出BRAF基因突变。BRAF基因突变组患者年龄、性别及结节最大径与非BRAF基因突变组差异均无统计学意义（均 P>0.05）。TMN中BRAF基因突变诊断TMC的灵敏度和特异度分别为71.3%及100.0%，ROC曲线下面积（AUC）（95% CI）为0.857（0.789~0.925）；Bethesda Ⅲ~Ⅴ类TMN中BRAF基因突变诊断TMC的灵敏度和特异度分别为63.0%及100.0%，AUC（95% CI）为0.815（0.680~0.950）。
结论NGS技术可成功检测TMN患者US-FNA标本中多基因突变情况，尤其是BRAF基因突变，且BRAF基因突变对于诊断TMC有一定价值。
ObjectiveTo explore the detection of BRAF, RAS, TERT promoter, and TP53 gene mutations in solitary thyroid micronodule (TMN) specimens obtained by ultrasound-guided fine-needle aspiration (US-FNA) using next-generation sequencing (NGS) technology and assess its diagnostic value in thyroid microcarcinomas (TMC).
MethodsOn-site recruitment of 428 patients with single suspicious TMC who underwent thyroid ultrasound examination, US-FNA, and NGS from September 2018 to July 2021 at Beijing Tongren Hospital affiliated to Capital Medical University. A total of 147 patients were finally included. NGS was used to detect mutations in the BRAF, RAS, TERT promoter, and TP53 genes in the US-FNA specimens. Comparisons were made between patients with and without gene mutations in terms of age, gender, and the maximum diameter of nodules. The diagnostic efficiency of BRAF mutation for TMC was calculated using the receiver operating characteristic (ROC) curve, with postoperative pathology as the gold standard.
ResultsThe age [ M ( Q 1, Q 3)] of the 147 patients was 43.0 (32.0, 51.0) years, and 37 were male (25.2%). Among the 147 US-FNA specimens, 97 (66.0%) were detected with BRAF gene mutations, all of which were p.V600E point mutation; 6 (4.1%) were detected with RAS gene mutation, and no TERT promoter or TP53 gene mutations were detected. Postoperative pathology confirmed that 136 cases were TMC, all of which were papillary thyroid microcarcinomas (PTMC); 11 cases (7.5%)were benign. Among 136 TMC samples, BRAF gene mutations were detected in 97 cases (71.3%). There were no statistically significant differences in age, gender, and maximum nodule diameter between patients with and without BRAF gene mutations (all P>0.05). The sensitivity and specificity of BRAF gene mutation in diagnosing TMC were 71.3% and 100.0%, respectively, with an area under the ROC curve (AUC) (95% CI) of 0.857 (0.789-0.925). For nodules classified as Bethesda Ⅲ-Ⅴ, the sensitivity and specificity were 63.0% and 100.0%, respectively, with an AUC (95% CI) of 0.815 (0.680-0.950).
ConclusionsNGS technology can successfully detect multiple gene mutations in US-FNA specimens from TMN patients, especially BRAF gene mutation, and BRAF gene mutation has certain value in diagnosing TMC.
于风霞,牛丽娟,李卫华,等. 二代测序技术检测超声引导下细针穿刺活检甲状腺微小结节标本中多基因突变及其诊断甲状腺微小癌的价值[J]. 中华医学杂志,2024,104(38)：3580-3585.
DOI:10.3760/cma.j.cn112137-20240628-01445版权归中华医学会所有。
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| 项目 | BRAF基因突变组（ n=97） | 非BRAF基因突变组（ n=50） | Z/χ 2值 | P值 |
|---|---|---|---|---|
| 年龄（岁） a | 42.0（31.5，49.5） | 46.5（34.5，51.8） | 1.211 | 0.226 |
| <55岁 b | 82（84.5） | 40（80.0） | 0.481 | 0.488 |
| 男性 b | 26（26.8） | 11（22.0） | 0.404 | 0.525 |
| 结节最大径（mm） a | 8.2（6.2，9.5） | 7.8（5.7，9.6） | 0.020 | 0.984 |
| 结节最大径≤5 mm b | 13（13.4） | 6（12.0） | 0.058 | 0.810 |
注：TMN为甲状腺微小结节； a M（ Q 1， Q 3）； b例（%）
| 项目 | Youden指数 | 灵敏度（%） | 特异度（%） | 阳性预测值（%） | 阴性预测值（%） | 标准误 | AUC（95% CI） |
|---|---|---|---|---|---|---|---|
| TMN中BRAF基因突变 | 0.713 | 71.3 | 100.0 | 100.0 | 22.0 | 0.035 | 0.857（0.789~0.925） |
| Bethesda Ⅲ~Ⅴ类TMN中BRAF基因突变 | 0.630 | 63.0 | 100.0 | 100.0 | 16.7 | 0.069 | 0.815（0.680~0.950） |
注：TMN为甲状腺微小结节；TMC为甲状腺微小癌；AUC为曲线下面积
注：TMN为甲状腺微小结节；TMC为甲状腺微小癌；ROC为受试者工作特征
于风霞：病例收集、数据整理、统计学分析、文章撰写；牛丽娟、应建明、吕宁：研究指导、设计实验、实施研究；李卫华：研究指导、分析与解释数据；朱强：研究指导、设计实验、实施研究、病例收集、论文修改、经费支持
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