Genotype–phenotype correlation in partial combined 17α-hydroxylase/17,20-lyase deficiency: functional characterization of CYP17A1 variants in ten 46,XY patients

Abstract Background Partial combined 17α-hydroxylase/17,20-lyase deficiency (PC-17OHD) is a rare 17OHD subtype with residual 17α-hydroxylase and 17,20-lyase activities. In 46,XY individuals, genital virilization is highly variable, and genotype–phenotype correlations remain poorly defined. Objective To characterize the clinical and endocrine spectrum of 46,XY PC-17OHD, functionally validate uncharacterized CYP17A1 variants, and establish genotype–phenotype correlations. Methods We enrolled ten 46,XY PC-17OHD patients from 9 families. Clinical phenotypes were characterized, and long-read CYP17A1 sequencing was performed. Splicing effects and enzyme activities were evaluated by minigene assays and HEK293T functional studies, respectively. Variants were classified according to ACMG guidelines. Results All patients presented with the triad of elevated mineralocorticoid precursors, generalized androgen suppression, and preserved cortisol levels. Median age at diagnosis was 17.5 years, with hypertension in 30% (3/10) and hypokalemia in 20% (2/10). Ten distinct variants were identified, including 2 novel splice-site variants (c.666+2T>C, c.1243+2T>G). Haplotype analysis revealed a shared identical haplotype among 5 patients carrying the c.1263G>A allele, indicating a Chinese founder effect. Four uncharacterized variants showed residual 17α-hydroxylase/17,20-lyase activities: p.Trp499Ter (11%/6.3%), p.Ala448Val (22.5%/14.2%), p.Leu433Ser (25.9%/21.7%), and c.666+2T>C (28.9%/22.6%). Notably, 2 patients with identical genotypes (c.985_987delinsAA/c.1263G>A) exhibited drastically divergent phenotypes—hypospadias (raised male) vs complete female external genitalia (raised female)—indicating that genotype alone cannot accurately predict clinical phenotypes in PC-17OHD. Conclusions Residual CYP17A1 activity largely determines genital masculinization, while marked phenotypic heterogeneity among patients with identical genotypes indicates that genotype alone cannot fully predict clinical manifestations. These findings improve molecular diagnosis and individualized care for this disorder.