Targeted long-read sequencing identifies missing pathogenic variant in unsolved 11[beta]-hydroxylase deficiency

• Published in: BMC endocrine disorders Vol. 24; no. 1
• Main Authors: Liu, Jidong, Tian, Huihui, Jin, Xinchen, Wang, Yanxiang, Zhang, Zhenhong, Li, Mengxue, Dai, Lulu, Zhang, Xiaoli, Jiang, Ling
• Format: Journal Article
• Language: English
• Published: BioMed Central Ltd 14.10.2024
• Subjects: ACTH; Corticosteroids; Genes; Genetic disorders; Genetic screening; Hydroxyprogesterone; Hypertension; Intermedin; Medical colleges; Progesterone; Testosterone; China
• ISSN: 1472-6823; 1472-6823
• DOI: 10.1186/s12902-024-01748-5

Background 11[beta]-hydroxylase deficiency (11[beta]-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 11[beta]-OHD is facing challenges. The study aimed to clarify the underlying genetic causes of two siblings of a Chinese family with 11[beta]-OHD. Methods 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. Results 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. Conclusions Our study suggests that the integrated long-range PCR-based NGS and T-LRS seem to be the most reliable and accurate method for 11[beta]-OHD genetic diagnosis and carrier sequencing. Keywords: 11[beta]-hydroxylase, Congenital adrenal hyperplasia, CYP11B1, Long-read sequencing