O-310 Browsing the Male Genome to Unravel Its Reproductive Potential
Authors: Cheung S , Rosenwaks Z , Palermo G.D 
Published: June 2022

Study question

Can whole genome profiling of sperm DNA be used to identify aspects of ART failure and predict embryo developmental competence?

Summary answer

Whole genome profiling of sperm DNA identifies germline gene mutations associated with reproductive failure and helps characterize subtle male factor infertility.

What is known already

A routine semen analysis provides limited information on the characteristics of the male gamete and is unable to predict spermatozoa performance in ART. Therefore, ancillary tests may be used to further assess the male gamete’s reproductive potential. Whole exome sequencing (WES) of the male genome carried out on somatic cells has proven to be a powerful technique capable of identifying the genetic roots of infertility. Here, we aim to preferentially detect germline mutations by exclusively sequencing spermatozoal DNA to identify genes related to the different underlying etiologies of reproductive failure.

Study design, size, duration

Over a 6-year period, 31 consenting couples with negative female infertility workups and normal semen parameters were included in this study. These couples were divided according to whether they reported a successful pregnancy with ART (fertile; n = 10) or not (infertile; n = 21). Sperm aneuploidy assessment by copy number variant (CNV) analysis with WES were carried out on ejaculated spermatozoa. Gene mutation profiles were enlisted and compared between the two patient cohorts to identify genes involved.

Participants/materials, setting, methods

DNA was extracted and amplified from at least 500 spermatozoa (DNA concentration, 760±486 ng/ul; quality, 1.7±0.1 nm) for CNV analyses by WES. Mutations corresponding to the CNV were then annotated and assessed using the CLC Genomic Server 9.0. Genes were considered duplicated or deleted when their read depth was &gt;1.5 or &lt; 0.5 times the median read depth in the control, respectively.

Main results and the role of chance

All couples (n = 31) (maternal age, 37.6±3yrs; paternal age, 39.7±5yrs) had adequate semen parameters (concentration, 59.2±30x106/mL, 44.8±18% motility, normal morphology) and normal peripheral karyotypes.

The fertile cohort (n = 10) underwent 12 ICSI cycles, achieving an 82.6% (57/69) fertilization rate with 10/12 (83.3%) cycles resulting in live births. The infertile cohort (n = 21) underwent 25 ICSI cycles, achieving a 68.4% (91/133) fertilization rate and 6/14 (42.9%) clinical pregnancies, all resulting in pregnancy loss. CNV analysis indicated lower sperm aneuploidy in the fertile (4.0% vs. 8.4%) cohort (P &lt; 0.00001). In both cohorts, mutations associated with sperm–egg fusion (ADAM3A) and acrosomal development (SPACA1, SPATA16) were identified, justifying ICSI utilization.

The infertile cohort included complete fertilization failure, poor early embryo development, implantation failure, or pregnancy loss. Couples with complete fertilization failure (n = 4) had gene deletions (PLCZ1, PIWIL1, ADAM15) indicating sperm-related oocyte-activating deficiency. Those with poor early embryo development (n = 5) had mutations essential for centrosome integrity (HAUS1) and spindle/microtubular stabilization (KIF4A, XRN1). Couples who failed to achieve pregnancy (n = 7) had mutations commonly implicated in embryonic implantation (IL9R) and microtubule/centrosomal integrity (MAP1S). Those with pregnancy losses (n = 5) displayed mutations related to trophoblast development (NLRP7), cell cycle regulation (MARK4, TRIP13, DAB2IP, KIF1C), and a gene linked to recurrent miscarriage (TP53).

Limitations, reasons for caution

Using WES, we were able to identify germline mutations that appear to be involved in various aspects of human reproduction. These findings are new and should be validated in a larger study population. Moreover, although we attempted to control for maternal age, we still cannot exclude confounding female factors.

Wider implications of the findings

Evaluating the sperm genome can help identify elusive genetic factors associated with reproductive competence and help guide treatment options for couples unable to conceive who undergo ART. Therefore, screening spermatozoal DNA may serve as an additional tool in precision medicine to identify and treat subtle male factor infertility.

Trial registration number

N/A