Genetic Characterization of a Cohort of Italian Patients with Congenital Adrenal Hyperplasia Due to 21-Hydroxylase Deficiency

Introduction Defects in the steroid 21-hydroxylase gene (CYP21A2) cause 21-hydroxylase deficiency (21OHD), the main cause of congenital adrenal hyperplasia (CAH). The disease shows a broad spectrum of clinical forms, ranging from severe or classical (salt wasting, SW, and simple virilizing, SV), to mild late onset or nonclassical (NC). 21OHD affects 1 in 15,000 in its severe classic form and 1 in 200-1000 in its mild NC form. There are many studies reporting the frequency of CYP21A2 pathogenic variants in different populations; however, few of them provide comprehensive information about Italian patients. Here, we present genetic results from a cohort of 245 unrelated Italian individuals with clinical diagnosis of CAH due to 21OHD. Methods A specific polymerase chain reaction (PCR) protocol combined with Sanger sequencing was used for CYP21A2 analysis. The multiplex ligation-dependent probe amplification (MLPA) assay was employed for copy number variation (CNV) determination. Results One hundred fourteen (46.5\%) of the index cases had the NC form, 57 (23.3\%) had the SV form, and 74 (30.2\%) presented the SW form of the disease. The most prevalent variant found in NC patients was the p.Val282Leu (51.3\%), while the most frequent variants in the classical form were p.Ile173Asn (8.6\%) and c.293-13C>G (26.0\%). In our study, the frequency of large rearrangements was 15.3\%, with CAH-X alleles representing 40\% of all DEL/CONV. In addition, 12 alleles carried rare variants, and 1 had a novel variant p.(Arg342Gln). We observed phenotype-genotype correlation in 94.7\% of cases. A complete concordance was observed in Groups 0 (enzyme activity completely impaired) where all patients had the SW form as expected. In Group A (0-1\% residual enzyme activity), 78.4\% of patients had the anticipated SW form while 21.6\% were diagnosed with the SV form. Within Group B (similar to 2\% residual enzyme activity), 93.4\% of patients exhibited SV form and 6.5\% SW disease. Finally, 92.6\% and 7.4\% of patients belonging to Group C (enzyme partially impaired to similar to 20-60\% residual activity) exhibited NC and SV phenotypes, respectively. Conclusion This work, representing a comprehensive genetic study, expanded the CYP21A2 variants spectrum of Italian patients with 21OHD and could be helpful in prenatal diagnosis and genetic counseling.