Department of Endocrinology and Metabolism, Cheeloo College of Medicine, Qilu Hospital, Shandong University, Jinan, 250012, P.R. China.
Institute of Endocrine and Metabolic Diseases of Shandong University, Jinan, Shandong Province, 250012, China.
BMC Endocr Disord. 2024 Oct 14;24(1):215. doi: 10.1186/s12902-024-01748-5.
11β-hydroxylase deficiency (11β-OHD), caused by homozygosity or compound heterozygosity CYP11B1 variants, is the second most common cause of congenital adrenal hyperplasia (CAH). Due to the high degree of sequence identity between CYP11B1 and CYP11B2, chimeric genes, and complex structural variants (SVs), the conventional approach to gene testing for 11β-OHD is facing challenges. The study aimed to clarify the underlying genetic causes of two siblings of a Chinese family with 11β-OHD.
Peripheral blood samples and clinical information were collected from subjects and their family members. Sex steroid concentrations were measured using LC-MS/MS. Long-range PCR-based next-generation sequencing (NGS), PCR assay and target long-read sequencing were used to detect the pathogenic variants.
Early onset hypertension, increased serum levels of adrenocorticotropin (ACTH), progesterone, testosterone, and decreased cortisol and potassium were detected in both affected siblings. Long-range PCR-based NGS identified a heterozygous missense variant (NM_000497.4:c.281 C > T, p.P94> L) in CYP11B1 gene in the two siblings. PCR detected no chimeric CYP11B2/CYP11B1 gene. We finally identified a second pathogenic variant in CYP11B1 gene via target long-read sequencing (T-LRS). This novel variant was a deletion-insertion variant and located chr8:143957269-143,957,579 (hg19) with the insertion of 'ACAG' (NM_000497.4:c.954 + 78_980delinsACAG), which was in trans with CYP11B1: c.281 C > T.
Our study suggests that the integrated long-range PCR-based NGS and T-LRS seem to be the most reliable and accurate method for 11β-OHD genetic diagnosis and carrier sequencing.
11β-羟化酶缺乏症（11β-OHD）由 CYP11B1 基因的纯合子或复合杂合子变异引起，是先天性肾上腺皮质增生症（CAH）的第二大常见病因。由于 CYP11B1 和 CYP11B2 之间具有高度的序列同一性、嵌合基因和复杂的结构变异（SV），传统的 11β-OHD 基因检测方法面临挑战。本研究旨在阐明一个中国家系中两例 11β-OHD 患者的遗传病因。
收集受检者及其家系成员的外周血样本和临床资料。采用 LC-MS/MS 检测性激素浓度。采用长片段 PCR 为基础的下一代测序（NGS）、PCR 检测和靶向长读测序检测致病变异。
两名受影响的同胞均有早发高血压、血清促肾上腺皮质激素（ACTH）、孕酮、睾酮水平升高，皮质醇和血钾水平降低。长片段 PCR 为基础的 NGS 在 CYP11B1 基因中发现了两例同胞均携带的杂合错义变异（NM_000497.4:c.281 C>T，p.P94>L）。PCR 未检测到 CYP11B2/CYP11B1 嵌合基因。我们最终通过靶向长读测序（T-LRS）发现 CYP11B1 基因的第二个致病变异。该新变异是缺失-插入变异，位于 chr8:143957269-143957579（hg19），插入“ACAG”（NM_000497.4:c.954+78_delinsACAG），与 CYP11B1:c.281 C>T 呈反式。
本研究表明，长片段 PCR 为基础的 NGS 和 T-LRS 的综合应用似乎是 11β-OHD 基因诊断和携带者测序最可靠和准确的方法。
