Functional Characterization and Genotype-Phenotype Correlation of Androgen Receptor Variants in Patients With...

Androgen insensitivity syndrome (AIS) is a 46,XY disorder of sex development caused by pathogenic variants in the androgen receptor (AR) gene. This study is aimed at characterizing the clinical features and investigating the functional consequences of AR variants identified in a Chinese AIS cohort. A systematic literature review was conducted to compare existing functional assays and pathogenic mechanisms. Multiple in silico tools were applied to predict the functional impact of AR variants, followed by in vitro functional analyses of missense variants in MLTC-1 cells. Protein expression was evaluated by Western blotting, subcellular localization by immunofluorescence, and transcriptional activity using a pGL4.11-(ARE)₃-TATA-luc dual-luciferase reporter assay. Among reported AR missense variants, only 28% (141/504) had undergone functional evaluation, comprising 274 assays, mainly ligand-binding and transcriptional activity analyses. In the present cohort, 33 patients with AIS harboring AR variants were analyzed, including 15 patients with complete AIS and 18 patients with partial AIS. Seventeen missense variants were further evaluated using in vitro functional assays. Reduced AR protein expression was observed in 82% (14/17) of variants, impaired nuclear localization in 59% (10/17), and reduced transcriptional activity in 53% (9/17), whereas increased transcriptional activity was observed in 12% (2/17) of variants. Nuclear localization was positively correlated with transcriptional activity, and residual transcriptional activity was positively associated with EMS scores. These findings demonstrate that AR variants cause AIS through heterogeneous molecular mechanisms and that residual AR function contributes to phenotypic variability.