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Targeted long-read sequencing for comprehensive detection of CYP21A2 mutations in patients with 21-hydroxylase deficiency

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Abstract

Background

21-Hydroxylase deficiency (21-OHD) is caused by pathogenic CYP21A2 variations. CYP21A2 is arranged in tandem with its highly homologous pseudogene CYP21A1P; therefore, it is prone to mismatch and rearrangement, producing different types of complex variations. There were few reports on using only one method to detect different CYP21A2 variants simultaneously.

Aims

Targeted long-read sequencing method was used to detect all types of CYP21A2 variants in a series of patients with 21-OHD.

Methods

A total of 59 patients with 21-OHD were enrolled from Peking Union Medical College Hospital. Long-range locus-specific PCR and long-read sequencing (LRS) were performed to detect the pathogenic variants in CYP21A2.

Results

Copy-number variants of CYP21A2 were found in 25.4% of patients, including 5.1% with 3 copies of CYP21A2, 16.9% with 1 copy of CYP21A2, and 3.4% with 0 copy of CYP21A2. The remaining 74.6% of patients had 2 copies of CYP21A2. Pathogenic variants were identified in all 121 alleles of 59 patients. Specifically, single-nucleotide variants and small insertions/deletions (< 50 bp) were detected in 79 alleles, of which conversed from CYP21A1P were detected in 63 alleles, and rare variants were found in the other 16 alleles. Large gene conversions (> 50 bp) from pseudogene were detected in 10 alleles, and different chimeric genes (CYP21A1P/CYP21A2 or TNXA/TNXB) formed by large deletions were detected in 32 alleles. Of all variants, p.I173N was the most common variant (19.0%).

Conclusions

Our study demonstrated that targeted long-read sequencing is a comprehensive method for detecting CYP21A2 variations, which is helpful for genetic diagnosis in 21-OHD patients.

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Data availability

The original contributions presented in the study are included in the article/supplementary files, and further inquiries can be directed to the corresponding author/s.

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Acknowledgements

The authors would like to thank all patients and their family members for participating in the study.

Funding

This work was supported by National High Level Hospital Clinical Research Funding (2022-PUMCH-D-002), CAMS Innovation Fund for Medical Sciences (CIFMS 2021-I2M-1-003), National Natural Science Foundation of China (Grant No. 81971375), Natural Science Foundation of Beijing (Grant No. 7212080), and National High Level Hospital Clinical Research Funding (2022-PUMCH-C-028).

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Authors and Affiliations

  1. Department of Endocrinology, Key Laboratory of Endocrinology, National Health Commission (NHC), Peking Union Medical College Hospital), Peking Union Medical College Hospital, Peking Union Medical College, Chinese Academy of Medical Sciences, Beijing, China

    X. Zhang, Y. Gao, L. Lu, Y. Cao, W. Zhang, B. Sun, X. Wu, A. Tong, S. Chen, X. Wang, J. Mao & M. Nie

  2. State Key Laboratory of Complex, Severe and Rare Diseases, Peking Union Medical College Hospital, Peking Union Medical College, Chinese Academy of Medical Sciences, Beijing, China

    M. Nie

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  1. X. Zhang
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  2. Y. Gao
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  3. L. Lu
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  4. Y. Cao
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  5. W. Zhang
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  6. B. Sun
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  7. X. Wu
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  8. A. Tong
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  9. S. Chen
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  10. X. Wang
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  11. J. Mao
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  12. M. Nie
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Contributions

MN designed and guided the experiments. XW, XW, and JM collected blood samples. SC, AT, and LL collected clinical data. YG, YC, BS, WZ, and XZ carried out the experiment. XZ and MN analyzed the experimental data and wrote the paper. All authors contributed to the article and approved the submitted version.

Corresponding author

Correspondence to M. Nie.

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Conflict of interest

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Research involving human participants and/or animals

The present study was approved by the ethics committee of Peking Union Medical College Hospital.

Informed consent

Informed consent was obtained from all individual participants included in the study.

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Cite this article

Zhang, X., Gao, Y., Lu, L. et al. Targeted long-read sequencing for comprehensive detection of CYP21A2 mutations in patients with 21-hydroxylase deficiency. J Endocrinol Invest 47, 833–841 (2024). https://doi.org/10.1007/s40618-023-02197-y

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  • DOI: https://doi.org/10.1007/s40618-023-02197-y

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