ESPE Abstracts

Background: Severe Ovarian Dysgenesis (OD), a rare heterogeneous XX disorder of Sex Development (XX-DSD) presents clinically with primary amenorrhea, hypergonadotrophic hypogonadism and infertility. The genetic basis of OD remains unknown in about 70% of cases. To identify novel causes of OD, we study patients in which known genes have been excluded.

Methods: Whole-exome-sequencing was performed on DNA extracted from peripheral lymphocytes of a 16y old female from consanguineous Israeli-Arab family who presented with lack of spontaneous pubertal development primary amenorrhea and hypergonadotrophic hypogonadism. Imaging studies detected a small uterus while no ovaries could be identified. DNA damage response (DDR) and chromosomal stability were tested using Mitomycin C (MMC) in chromosomal breakage assay.

Results: A homozygous missense mutation; c.C1972T, p.R658W was identified in the FAN1 gene, a DDR pathway gene. Segregation was consistent with recessive inheritance. Structural analysis revealed that R658 acts as an anchoring point of DNA upon FAN1 homo-dimerization and stabilize DNA bifurcation into single strand. As such, R658W destabilizes FAN1 interaction with the DNA and prevents its binding to FAN1 dimers. Chromosomal breakage assays revealed significantly higher number of DNA breaks in patient-derived leukocytes compared to control. DNA breaks increase occurred following DNA damage induction with MMC in a dose dependent manner. Induction of breaks with 150nM MMC resulted in: 1.63±0.28 breaks per cell (bpc) in the patient vs. only 0.70±0.31 in controls (P=0.003). Induction of breaks with 300nM resulted in 4.00±0.55 bpc in patient vs. 1.87±0.29 in controls (P=0.0005).

Conclusions: The novel homozygous missense mutation in FAN1 is accompanied by impaired DDR and suggests FAN1 as a novel genetic etiology for ovarian dysgenesis in humans. This further expands the crucial role of DDR pathway in normal ovariogenesis, and indicates the importance of the chromosomal breakage assay in the differential diagnosis for OD. The implication on routine clinical cancer surveillance in the patient and family members is under further studies.