A GWAS in Idiopathic/Unexplained Infertile Men Detects a Genomic Region Determining Follicle-Stimulating Hormone Levels.

CONTEXT: Approximately 70% of infertile men are diagnosed with idiopathic (abnormal semen parameters) or unexplained (normozoospermia) infertility, with the common feature of lacking etiologic factors. Follicle-stimulating hormone (FSH) is essential for initiation and maintenance of spermatogenesis. Certain single-nucleotide variations (SNVs; formerly single-nucleotide polymorphisms [SNPs]) (ie, FSHB c.-211G > T, FSHR c.2039A > G) are associated with FSH, testicular volume, and spermatogenesis. It is unknown to what extent other variants are associated with FSH levels and therewith resemble causative factors for infertility. OBJECTIVE: We aimed to identify further genetic determinants modulating FSH levels in a cohort of men presenting with idiopathic or unexplained infertility. METHODS: We retrospectively (2010-2018) selected 1900 men with idiopathic/unexplained infertility. In the discovery study (n = 760), a genome-wide association study (GWAS) was performed (Infinium PsychArrays) in association with FSH values (Illumina GenomeStudio, v2.0). Minor allele frequencies (MAFs) were analyzed for the discovery and an independent normozoospermic cohort. In the validation study (n = 1140), TaqMan SNV polymerase chain reaction was conducted for rs11031005 and rs10835638 in association with andrological parameters. RESULTS: Imputation revealed 9 SNVs in high linkage disequilibrium, with genome-wide significance (P < 4.28e-07) at the FSHB locus 11p.14.1 being associated with FSH. The 9 SNVs accounted for up to a 4.65% variance in FSH level. In the oligozoospermic subgroup, this was increased up to 6.95% and the MAF was enhanced compared to an independent cohort of normozoospermic men. By validation, a significant association for rs11031005/rs10835638 with FSH (P = 4.71e-06/5.55e-07) and FSH/luteinizing hormone ratio (P = 2.08e-12/6.4e-12) was evident. CONCLUSIONS: This GWAS delineates the polymorphic FSHB genomic region as the main determinant of FSH levels in men with unexplained or idiopathic infertility. Given the essential role of FSH, molecular detection of one of the identified SNVs that causes lowered FSH and therewith decreases spermatogenesis could resolve the idiopathic/unexplained origin by this etiologic factor.

HLA-G and HLA-E Immune Checkpoints Are Widely Expressed in Ewing Sarcoma but Have Limited Functional Impact on the Effector Functions of Antigen-Specific CAR T Cells.

Immune-inhibitory barriers in the tumor microenvironment of solid cancers counteract effective T cell therapies. Based on our finding that Ewing sarcomas (EwS) respond to chimeric antigen receptor (CAR) gene-modified effector cells through upregulation of human leukocyte antigen G (HLA-G), we hypothesized that nonclassical HLA molecules, HLA-G and HLA-E, contribute to immune escape of EwS. Here, we demonstrate that HLA-G isotype G1 expression on EwS cells does not directly impair cytolysis by GD2-specific CAR T cells (CART), whereas HLA-G1 on myeloid bystander cells reduces CART degranulation responses against EwS cells. HLA-E was induced in EwS cells by IFN-γ stimulation in vitro and by GD2-specific CART treatment in vivo and was detected on tumor cells or infiltrating myeloid cells in a majority of human EwS biopsies. Interaction of HLA-E-positive EwS cells with GD2-specific CART induced upregulation of HLA-E receptor NKG2A. However, HLA-E expressed by EwS tumor cells or by myeloid bystander cells both failed to reduce antitumor effector functions of CART. We conclude that non-classical HLA molecules are expressed in EwS under inflammatory conditions, but have limited functional impact on antigen-specific T cells, arguing against a relevant therapeutic benefit from combining CART therapy with HLA-G or HLA-E checkpoint blockade in this cancer.

Does the FSHB c.-211G>T polymorphism impact Sertoli cell number and the spermatogenic potential in infertile patients?

BACKGROUND: A genetic variant within the FSHB gene can deviate FSH action on spermatogenesis. The c.-211G>T FSHB single nucleotide polymorphism impacts FSHB transcription and biosynthesis due to interference with the LHX3 transcription factor binding. This SNP was previously shown to be strongly associated with lowered testicular volume, reduced sperm counts, and decreased FSH levels in patients carrying one or two T-alleles. OBJECTIVE: To determine the impact of the SNP FSHB c.-211G>T on Sertoli cell (SC) number, Sertoli cell workload (SCWL) and thereby spermatogenic potential. MATERIAL AND METHODS: Testicular biopsies of 31 azoospermic, homozygous T patients (26 non-obstructive azoospermia (NOA), and five obstructive azoospermia (OA)) were matched to patients with GG genotype. Marker proteins for SC (SOX9), spermatogonia (MAGE A4), and round spermatids (CREM) were used for semi-automatical quantification by immunofluorescence. SCWL (number of germ cells served by one SC) was determined and an unbiased clustering on the patient groups performed. RESULTS: Quantification of SC number in NOA patients did not yield significant differences when stratified by FSHB genotype. SC numbers are also not significantly different between FSHB genotypes for the OA patient group and between NOA and OA groups. SCWL in the NOA patient cohort is significantly reduced when compared to the OA control patients; however, in neither group an effect of the genotype could be observed. The cluster analysis of the whole study cohort yielded two groups only, namely NOA and OA, and no clustering according to the FSHB genotype. DISCUSSION AND CONCLUSION: The FSHB c.-211G>T polymorphism does not affect SC numbers or SCWL, thereby in principle maintaining the spermatogenic potential. The previously observed clinical phenotype for the FSHB genotype might therefore be caused by a hypo-stimulated spermatogenesis and not due to a decreased SC number.

Pharmacogenetics of FSH Action in the Male.

Male infertility is a major contributor to couple infertility, however in most cases it remains "idiopathic" and putative treatment regimens are lacking. This leads to a scenario in which intra-cytoplasmic spermatozoa injection (ICSI) is widely used in idiopathic male infertility, though the treatment burden is high for the couple and it entails considerable costs and risks. Given the crucial role of the Follicle-stimulating hormone (FSH) for spermatogenesis, FSH has been used empirically to improve semen parameters, but the response to FSH varied strongly among treated infertile men. Single nucleotide polymorphisms (SNPs) within FSH ligand/receptor genes (FSHB/FSHR), significantly influencing reproductive parameters in men, represent promising candidates to serve as pharmacogenetic markers to improve prediction of response to FSH. Consequently, several FSH-based pharmacogenetic studies have been conducted within the last years with unfortunately wide divergence concerning selection criteria, treatment and primary endpoints. In this review we therefore outline the current knowledge on single nucleotide polymorphisms (SNPs) in the FSH and FSH receptor genes and their putative functional effects. We compile and critically assess the previously performed pharmacogenetic studies in the male and propose a putative strategy that might allow identifying patients who could benefit from FSH treatment.