Endocrine Abstracts

Introduction: According to the free hormone transport hypothesis, only free testosterone (FT) is considered biologically active. Due to low circulating levels and technical challenges of direct FT measurements, calculators have been developed to estimate FT from serum total T, sex hormone-binding globulin (SHBG) and albumin levels. However, genetic polymorphisms altering SHBG’s binding affinity or capacity might result in calculation imprecisions with consequent incorrect diagnosis in these individuals.

Objective: Investigating the prevalence of SHBG single-nucleotide polymorphisms (SNPs) in healthy men and evaluating differences in SHBG, total T, calculated FT (cFT) and measured FT (mFT) levels in SHBG SNP-carriers.

Methods: Population-based sibling-pair study, comprising 999 healthy men aged 24-46 years in whom genotyping was performed (LGC Genomics) for SNPs suggested to affect binding affinity and/or concentration of SHBG (rs6258, rs6259, rs146779355, rs145273466, rs368589266, rs143269613, rs373769356, rs143521188). SHBG and total T concentrations were measured by immunoassay and LC-MS/MS, respectively. FT levels (cFT; using the Vermeulen-formula) and free T-ratio (FT/total T) were calculated. In a subset of participants (145, 10 and 40 individuals for WT/WT, WT/rs6258 and WT/rs6259, respectively) FT was measured directly using LC-MS/MS after equilibrium dialysis (mFT). The difference between cFT and mFT was calculated and expressed as percentage of mFT (delta%). Parameters were compared by t-test or Mann-Whitney test depending on normality; method-comparison was performed using Passing-Bablok regression.

Results: 11/971 (1%) and 0/971 participants were hetero- and homozygote for rs6258; 135/681 (20%) and 9/681 (1%) were hetero- and homozygote for rs6259, respectively. No other SNPs were detected. Versus wild-type, heterozygote rs6258 carriers had lower SHBG (26.5 nmol/l vs 38.5 nmol/l) and total T levels (468.8 ng/dl vs 583.9 ng/dl) but higher free T-ratio (2.3% vs 2.0% for cFT; 2.2% vs 1.7% for mFT). Heterozygote rs6259 carriers had higher SHBG levels (42.8 nmol/l vs 38.5 nmol/l) and lower free T-ratio (1.9% vs 2.0% for cFT) than non-carriers. Comparing cFT and mFT showed a significant difference in heterozygote rs6258 carriers (delta% 5.1% vs 15.4% in wild-type). No other differences nor differences in homozygote rs6259 carriers vs wild-type were observed.

Conclusion: Genetic polymorphisms suggested to affect SHBG concentration or steroid-hormone binding affinity are relatively rare in this population-based cohort of healthy men. Although carriers did present with different SHBG levels and free T-ratios compared to non-carriers, our results do not disprove the calculators’ accuracy in men heterozygous for rs6258 or rs6259, despite in vitro experiments suggesting a 1.8 reduced binding affinity and reduced clearance rate respectively compared to wild-type SHBG.