New perspectives in the genetic diagnosis of male infertility

features two interesting articles on reproductive health. Sengun et al (1) present their findings of novel mutations in the gene coding for FK506 binding protein-like ( FKBPL ) associated with male infertility, while Bilić et al (2) discuss the ben-efits of ovarian tissue cryopreservation. In spite of tremen-dous advances in the field, various aspects of reproductive health, particularly infertility, still necessitate further study and development of novel diagnostic and therapeutic ap-proaches.

The current issue of the Croatian Medical Journal features two interesting articles on reproductive health. Sengun et al (1) present their findings of novel mutations in the gene coding for FK506 binding protein-like (FKBPL) associated with male infertility, while Bilić et al (2) discuss the benefits of ovarian tissue cryopreservation. In spite of tremendous advances in the field, various aspects of reproductive health, particularly infertility, still necessitate further study and development of novel diagnostic and therapeutic approaches.
Infertility, defined by the World Health Organization as the failure to achieve conception after 12-24 months of unprotected sexual intercourse (3), affects about one out of six couples of childbearing age in Western countries (4). Male factor infertility is identified in about half of the cases (5) and most of the times presents as spermatogenesis failure (SPGF). Spermatogenesis is a 74-day-long process involving up to 2000 genes. Among them, 600 to 900 are exclusively expressed by the male germ-line (6)(7)(8). Despite a comprehensive diagnostic workup, in the majority of cases the etiology of SPGF remains elusive. A monocentric study on 1737 infertile patients reported idiopathic oligozoospermia in ~ 75% of cases (9). Classic genetic screening can identify the cause of male infertility in only 5% of unselected patients and in 20% of patients with non-obstructive azoospermia (NOA) (10). In the remaining cases, the etiology of infertility is elusive.
In recent years, several mutations of genes involved in different steps of spermatogenesis, such as spermatogonia proliferation or meiotic division of primary spermatocytes, have been reported capable of interfering with murine spermatogenesis (11). Interestingly, many of them have been validated in infertile patients, as shown in case reports or case series. Thus, the inclusion of a comprehensive gene panel in the diagnostic algorithm of male infertility could raise the diagnostic yields (11). The first study addressing this issue analyzed a panel of three genes: NR5A1, DMRT1, and TEX11, in a cohort of 80 patients with NOA. NR5A1 (9q33.3) encodes for a transcription factor involved in the regulation of genes playing a role in steroidogenesis. Its mutation has been associated with a wide spectrum of phenotypes, ranging from 46,XY partial and complete gonadal dysgenesis, to hypospadias, micropenis, anorchia, and, in otherwise healthy men, oligozoospermia or NOA. DMRT1 (9p24.3) encodes for a testis-specific transcription factor acting in testis differentiation, whose mutations have been reported in patients with NOA. Finally, TEX11 (Xq13.1) encodes for a meiosis-specific factor, involved in double-strand breaks DNA repair, which may play a role in NOA pathogenesis due to meiotic arrest (11). Out of 80 patients with NOA investigated for NR5A1, DMRT1, and TEX11 gene mutations, the authors reported pathogenic variants in four, thus raising the diagnostic yield by 25% (10). A subsequent study has developed a gene panel including 15 genes (Table 1) involved in germ-cell proliferation and meiotic division. Interestingly, pathogenic mutations of NR5A1 and TEX11 genes were reported in 3/25 patients, increasing the diagnostic rate by 12%. Noteworthy, 11 likely pathogenic variants meriting functional analysis or segregation studies were also observed.
These data highlight the importance of designing a comprehensive and accurate gene panel to be used in patients with otherwise unexplained SPGF. Furthermore, next-generation sequencing makes this analysis widely accessible from both an economic and a geographic point of view. An appropriate gene panel for SPGF could facilitate the identification of the genetic cause of infertility (when present), but in the future it may also represent a diagnostic test predicting sperm recovery after testicular sperm extraction (TESE). Accordingly, a study matching the results of testicular histology with those of genetic testing has been published and others are currently ongoing. Interestingly, mutations (eg, deletions, missense, stop-gain) of specific genes involved in meiosis (eg, M1AP, ADAD2, TERB1, SHOC1, MSH4, RAD21L1, TEX14, DMRT1, TEX11, SYCE1, MEIOB, MEI1, STAG3-a) have been reported in patients with meiotic arrest (13,14). If validated, these gene variations may be included in a pre-TESE prognostic gene panel, which may help to determine the chance of sperm recovery. Finally, another challenge in this field is to understand the implications of the transmission of these gene mutations to the offspring.
In conclusion, the evidence favoring the inclusion of SPGF monogenic mutation assessment in the diagnostic workup of male infertility is accumulating. Despite this, a comprehensive panel has not yet been validated, although some gene mutations are more frequently present than others (eg, NR5A1 or TEX11), and others have recently been discovered. Since more than 2000 genes are involved in spermatogenesis, we are still far from a comprehensive view of the monogenic etiology of SPGF. However, in the near future, this evidence is likely to practically affect the diagnostic workup and decision-making algorithms of male infertility.