Genetic insights into non-obstructive azoospermia: Identifying novel variants and enhancing clinical outcomes

Introduction & Objectives: Non-obstructive azoospermia (NOA), defined by the complete absence of spermatozoa in the ejaculate, is a highly heterogeneous condition, manifesting in a wide array of testicular phenotypes. Spermatogenesis, the complex and tightly regulated biological process by which diploid spermatogonial stem cells (SSCs) differentiate into mature haploid spermatozoa, is orchestrated by a network of over 2,000 genes. These genes are integral to the coordination of cellular differentiation, meiosis, and spermiogenesis. A striking 73% of NOA cases remain genetically unexplained, underscoring the limitations of existing diagnostic methodologies and the necessity for more comprehensive genomic analyses. The primary objective of this study is to identify novel genes and variants implicated in NOA, further refining our understanding of the genetic landscape that contributes to the condition. By improving the genotype-phenotype correlation, this research aims to enhance the accuracy of prognostic assessments for successful sperm retrieval during testicular sperm extraction (TESE).

Materials & Methods: Whole-exome sequencing (WES) was performed on 63 individuals affected by NOA, originating from 43 distinct families. Among these, 20 families exhibited a strong familial predisposition to NOA. All individuals had previously tested negative for conventional genetic screening. Bioinformatics analysis employed a targeted approach, prioritizing known causal or candidate genes based on evidence from human genetic studies, animal models, or functional assays that have previously demonstrated their involvement in key processes such as spermatogenesis and testicular function (Fig. 1).

Results: In this analysis, putative causative genetic defects were identified in 21 distinct genes among a cohort of 33 individuals, representing 57% of the total population studied. Notably, three genes previously recognized as causative factors in disorders associated with spermatogenesis arrest are MEIOB, USP26, and TERB1. Additionally, the genes DMRT1, RBBP7, RBMXL3, RAI1, ADAD2, HUWE1, VCX3A, SPO11, and ESR1 have been reported in a limited number of studies. Furthermore, ten additional genes—FTHL17, SATL1, STS, CENPI, ZCCHC13, HIRIP3, DHX37, NAP1L3, CEP85, and ABCD1—had not been described in man. Defects were identified in genes associated with germ cell development, meiosis, and DNA repair pathways, as well as in genes involved in post-meiotic maturation and meiotic processes (Fig.2).

Conclusions: Interestingly, all individuals with defects in meiotic genes exhibited unsuccessful sperm retrieval, underscoring the critical importance of genetic diagnosis prior to testicular sperm extraction (TESE). This strategy could effectively identify individuals with diminished or absent potential for successful sperm retrieval.