Cost-effective genotyping for classical congenital adrenal hyperplasia (CAH) due to 21-hydroxylase deficiency (21-OHD) in resource-poor settings: multiplex ligation probe amplification (MLPA) with/without sequential next-generation sequencing (NGS)

• Published in: Hormones (Athens, Greece) Vol. 22; no. 2; pp. 311 - 320
• Main Authors: Monteiro, Ana, Pavithran, Praveen V., Puthukulangara, Manuprasad, Bhavani, Nisha, Nampoothiri, Sheela, Yesodharan, Dhanya, Kumaran, Reshma
• Format: Journal Article
• Language: English
• Published: Cham Springer International Publishing 01.06.2023
• Subjects: Adrenal Hyperplasia, Congenital - diagnosis; Adrenal Hyperplasia, Congenital - genetics; Cost-Benefit Analysis; Endocrinology; Genotype; High-Throughput Nucleotide Sequencing; Humans; Medicine; Medicine & Public Health; Metabolic Diseases; Mutation; Original Article; Steroid 21-Hydroxylase - genetics; Variant; Deletion; Duplication; South India; Salt wasting; Simple virilizing
• ISSN: 1109-3099; 2520-8721; 2520-8721
• DOI: 10.1007/s42000-023-00445-7

Purpose Genotyping of classic congenital adrenal hyperplasia (CAH) due to 21-hydroxylase deficiency (21-OHD) is becoming increasingly significant beyond prenatal counseling in the current era of emerging gene therapy/editing technologies. While the knowledge of common variants helps in designing cost-effective genotyping strategies, limited data are currently available from the Indian subcontinent, especially South India, mainly due to financial constraints. The aim of this study is to assess the genotype of individuals with classic CAH from a South Indian cohort in a cost-effective manner. Methods The genotypes of 46 unrelated subjects with classic CAH were studied through initial multiplex ligation-dependent probe amplification (MLPA) using the SALSA MLPA Probe-mix P050 CAH (MRC Holland). Next-generation sequencing (NGS) was done in 10 subjects, as their MLPA was either negative or showed heterozygous variants. Results The common variants observed in our study population of 46 subjects were large deletions (35.8%), intron 2 variant [c.293-13A/C > G] (35.8%), 8 bp del [c.332_339del p.(Gly111Valfs*21)] (7.7%), and R356W [c.1069 C > T p.(Arg357Trp)] (6.6%). MLPA alone detected pathogenic variants in 78.2% of the initial study samples (36/46). Sequential NGS resulted in a 100% detection rate in our study population. Conclusion MLPA appears to be an effective first genotyping modality for this South Indian cohort due to the high prevalence of large deletions and common variants. MLPA as a first initial screening genotyping test with sequential NGS when required may be a cost-effective and highly sensitive approach to CYP21A2 genotyping in our part of the world and in resource-poor settings.