Presentation Authors: Willy Chertman, Diana M Lopategui, Anthony J Griswold, Himanshu Arora*, Ranjith Ramasamy, MIAMI, FL
Introduction: The pathologic enlargement of veins of the pampiniform plexus within the spermatic cord, or varicocele, is a common cause of impaired semen parameters. Varicocele is prevalent among adolescent males (15%), nevertheless only a small proportion (1%) of men with varicoceles have impaired semen parameters. Yet, varicocele remains the most common correctable cause of male infertility. We investigated genetic variants as possible causes of varicocele with decreased sperm count in two brothers, using targeted exon/whole-exome sequencing (WES).
Methods: We extracted DNA from the blood of 2 affected brothers (with severe oligospermia) and 1 unaffected brother (normozoospermia) using QIAamp DNA Blood Maxi Kit. Extracted DNA was used for the whole exome sequencing (WES). All 3 brothers had left grade II varicocele. Two of them had decreased sperm count ( < 1 million/cc) while the other brother had normal count (>20 million/cc) on at least 2 semen analyses. The raw sequence data were aligned to the reference genome (hg19) and filtered for quality control including coverage and genotype quality. Potentially pathogenic variants were identified by the following method: homozygous variants shared by the two brothers with varicocele and severe oligospermia but not by the brother with varicocele and normal sperm count, rare (below 0.05 allele frequency) according to ExAC and the 1000 Genomes database, and missense, nonsense, or frameshift variants. Sanger sequencing was performed to confirm the variant in the 2 affected brothers and determine segregation in the parents.
Results: A premature stop codon alteration (nonsense) was identified on Chromosome X (37028866 C>T) in the gene FAM47C. The affected brothers were found to be hemizygous for the variant, while the mother was a heterozygous carrier. The unaffected brother and the father both carried the reference allele. Sanger sequencing confirmed the variant. The gene FAM47C has been previously identified with male infertility in a copy-number variant that included the FAM47C gene and is also highly expressed in testis tissue, though its function is unknown .
Conclusions: Using WES we were able to identify a nonsense mutation in FAM47C that segregates with the oligospermia phenotype. The mutation was present in the severely oligospermic brothers and not in the normozoospermic brother despite all 3 brothers having a similar varicocele clinical grade. Because varicocele is prevalent (15%) in the population, identifying men that would have impaired spermatogenesis using approaches like WES can be paradigm shifting.
Source of Funding: Supported by the American Urological Association Research Scholar Award and Stanley Glaser Award to RR.