Genetics of 21-OH Deficiency and Genotype-Phenotype Correlation: Experience of the Hellenic National Referral Center.

21-hydroxylase deficiency (21-OHD) represents the most common form of congenital adrenal hyperplasia (CAH) due to CYP21A2 gene pathogenic variants. Τhe aim of this study was the identification of CYP21A2 variants in 500 subjects of Greek origin with a suspicion of 21-OHD and, by using the existing hormonal assessment and genotypes of the 500 subjects tested, to identify a biomarker that could differentiate between the heterozygotes and the cases with no pathogenic variants identified. Five hundred subjects with clinical suspicion of 21-OHD underwent CYP21A2 gene sequencing and Multiplex Ligation Dependent Probe Amplification (MLPA). Genetic diagnosis was achieved in 27.4% of the subjects tested, most of which presented with the non-classic form (NC) of 21-OHD. Heterozygotes accounted for 42.6% of cases, whereas no pathogenic variants were identified in 27% of cases. De novo aberrations, duplications, and five novel variants were also identified. Statistical analysis revealed that the difference between the basal and 60' post-ACTH stimulation 17-hydroxyprogesterone concentrations (Δ17-OHP^60-0) could be a potential biomarker (p < 0.05) distinguishing the heterozygotes from the cases with no pathogenic variants identified, although no clear cut-off value could be set. Further analysis revealed overlapping clinical manifestations among all the subjects tested. The presented phenotypic traits of the subjects tested and the inability to identify a discriminative biochemical marker highlight the importance of comprehensive CYP21A2 genotyping to ascertain the correct genetic diagnosis and proper genetic counselling.