Study question: What is the long-term outcome of individuals born with bilateral testicular regression (BTR) in relation to its underlying etiology?

Summary answer: Statural growth and pubertal development are adequate with incremental doses of testosterone replacement therapy (TRT); however, penile growth is often suboptimal, especially in those with a suspected genetic etiology (i.e. heterozygous DHX37 variants) or a micropenis at birth.

What is known already: BTR is a rare and poorly understood condition. Although a vascular origin has been postulated, heterozygous missense variants in DHX37 have been attributed to the phenotype as well. How these various etiologies impact the clinical phenotype, gonadal histology and outcome of BTR remains unclear.

Study design size duration: For this cross-sectional study, individuals with BTR were recruited in eight Belgian pediatric endocrinology departments, between December 2019 and December 2022. A physical exam was performed cross-sectionally in all 17 end-pubertal participants and a quality of care questionnaire was completed by 11 of them. Exome-based panel testing of 241 genes involved in gonadal development and spermatogenesis was performed along with a retrospective analysis of presentation and management. A centralized histological review of gonadal rests was done for 10 participants.

Participants/materials setting methods: A total of 35 participants (33 with male, 1 with female, and 1 with non-binary gender identity) were recruited at a mean age of 15.0 ± 5.7 years.

Main results and the role of chance: The median age at presentation was 1.2 years [0-14 years]. Maternal gestational complications were common (38.2%), with a notably high incidence of monozygotic twin pregnancies (8.8%). Heterozygous (likely) pathogenic missense variants in DHX37 (p.Arg334Trp and p.Arg308Gln) were found in three participants. No other (likely) pathogenic variants were found. All three participants with a DHX37 variant had a microphallus at birth (leading to female sex assignment in one), while only six of the remaining 31 participants without a DHX37 variant (19.4%) had a microphallus at birth (information regarding one participant was missing). Testosterone therapy during infancy to increase penile growth was more effective in those without versus those with a DHX37 variant. The three participants with a DHX37 variant developed a male, female, and non-binary gender identity, respectively; all other participants identified as males. TRT in incremental doses had been initiated in 25 participants (median age at start was 12.4 years). Final height was within the target height range in all end-pubertal participants; however, 5 out of 11 participants (45.5%), for whom stretched penile length (SPL) was measured, had a micropenis (mean adult SPL: 9.6 ± 2.5). Of the 11 participants who completed the questionnaire, five (45.5%) reported suboptimal understanding of the goals and effects of TRT at the time of puberty induction. Furthermore, only 6 (54.5%) and 5 (45.5%) of these 11 participants indicated that they were well informed about the risks and potential side effects of TRT, respectively. Histological analysis of two participants with DHX37 variants suggested early disruption of gonadal development due to the presence of Müllerian remnants in both and undifferentiated gonadal tissue in one. In eight other analyzed participants, no gonadal remnants were found, in line with the BTR diagnosis.

Limitations reasons for caution: The limitations of this study include the relatively small sample size (n = 35) and the few individuals with DHX37 variants (n = 3). Furthermore, data on the SPL were often missing, due to this being undocumented or refused by participants.

Wider implications of the findings: TRT provides adequate statural growth, even when initiated in late adolescence, thus providing time for physicians to explore the patients' gender identity if needed. However, sufficient and understandable information regarding the effects and side effects of TRT is required throughout the management of these patients. SPL remains suboptimal in many individuals and could be improved by TRT during infancy to mimic the physiological mini-puberty. An environmental origin in some participants is supported by the high incidence of gestational complications (38.2%) and by the three monozygotic twin pregnancies discordant for the BTR phenotype. Individuals with a heterozygous DHX37 variant have a more severe phenotype with severely restricted penile growth until adulthood. Histological analysis confirmed DHX37 as a gonadal development, rather than a BTR-related, gene.

Study funding/competing interests: Funding was provided by the Belgian Society for Pediatric Endocrinology and Diabetology (BESPEED) and by Ghent University Hospital under the NucleUZ Grant (E.D.B.). M.C. and E.D.B. are supported by an FWO senior clinical investigator grant (1801018N and 1802220N, respectively). The authors report no conflicts of interest.

Trial registration number: N/A.