Whole exome sequencing analysis of susceptibility loci in transgender individuals.

Background: Genetic factors contributing to sex-associated dimorphic brain development may also underlie gender identity-related anxiety disorders.

Aim: To establish a high-throughput whole-exome sequencing (WES) and bioinformatics pipeline for identifying rare variants in sex-dimorphic neural pathways and explore their association with gender identity-related anxiety.

Methods: Peripheral genomic DNA was collected from 23 patients (13 Assigned male at birth (AMAB), 10 Assigned female at birth (AFAB)) presenting with gender identity-related anxiety at Shanghai Mental Health Centre between March 2020 and February 2022. WES libraries were prepared and sequenced to an average depth of 100×. Raw reads underwent stringent quality control, alignment, variant calling, and annotation against public databases (gnomAD, ClinVar). Rare (minor allele frequency [MAF] < 1%) high-confidence variants were filtered to focus on exonic, splice-site, and insertion and deletion (indel) events. Candidate genes were subjected to Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment and Gene Ontology (GO) analyses to identify overrepresented neural development pathways, with particular emphasis on estrogen receptor-mediated signaling.

Outcome: A total of 479 rare, potentially pathogenic variants across 19 estrogen receptor-mediated neurodevelopmental genes were identified for further validation.

Results: After quality control, 266 265 high-confidence variants were retained; of 217 757 exomic calls, 48 508 (22.3%) were exonic (49.8% nonsynonymous), including 4.1% splice-site, 225 missense, 27 nonsense, 43 frameshift, and various indels. KEGG analysis highlighted significant enrichment in axon guidance signaling, while GO terms pointed to neuronal projection and synaptic assembly. Nineteen genes within the estrogen receptor pathway harbored rare deleterious variants, suggesting disruptions in sex hormone-driven neural differentiation.

Clinical translation: This WES-based framework enables the identification of novel candidate loci for diagnostic panels and may inform personalized interventions for gender identity-related anxiety.

Strengths and limitations: This study leveraged high-depth whole-exome sequencing, stringent bioinformatics filtering, and a pathway-focused approach to pinpoint rare variants in sex-dimorphic neurodevelopmental genes; however, its small sample size, lack of functional validation, potential population stratification bias, and cross-sectional design limit causal inference.

Conclusion: Our integrated WES and bioinformatics pipeline uncovers rare variants in estrogen receptor-mediated neurodevelopmental genes, providing new insights into the genetic architecture of sex-dimorphic brain development and its role in gender identity-related anxiety.

Study registration: The article was registered at the ISRCTN Registry https://www.isrctn.com/ISRCTN18336816 (no. 18336816) under an observational study record.