Genetic determinants of sex hormone-binding globulin and myoma nodule volume in patients with uterine myoma

Uterine myoma is a common benign pelvic neoplasm in women, predominantly composed of myometrial smooth muscle cells and fibrous connective tissue [1, 2]. The prevalence of uterine myoma increases with age, peaking in women older than 40 years [3]. This condition affects women of all ethnic groups but is most frequently reported in Black women [4]. Uterine myoma substantially impairs quality of life, particularly through menstrual disturbances such as heavy, prolonged, or irregular bleeding, chronic pelvic pain, dyspareunia, and related symptoms [5].

Sex hormones play an important role in initiating myoma development [6–10]. The activity of sex hormones in the female body is largely mediated by sex hormone-binding globulin (SHBG) [11], which plays a key role in regulating the bioavailability of sex hormones in target tissues and cells [11]. Only a single study by Wang et al. examined the correlations between the genetic determinants of SHBG and uterine myoma, yielding highly inconsistent results [12]. Using Mendelian randomization, the authors found no association between SHBG and uterine myoma in a meta-analysis of GWAS data from two cohorts, FinnGen and FibroGENE, yet identified such an association (p=0.016) within the FibroGENE cohort alone [12]. No other experimental genetic studies have been published on this topic. This near-complete lack of data on the role of SHBG genetic determinants in myoma formation underscores the need for further research in this area.

This study aimed to investigate the association between polymorphisms in genes related to SHBG levels and myoma nodule volume.

Materials and methods

The study group comprised 329 patients with uterine myomas (continuous sampling). Patients were recruited by physicians from the Gynecology Department of the Perinatal Center at St. Ioasaph Belgorod Regional Clinical Hospital between 2008 and 2013. The inclusion criteria were as follows: 1) a confirmed diagnosis of uterine myoma (by ultrasound examination or following hysterectomy); 2) Russian ethnicity; 3) birthplace and residence in the Central Chernozem region of Russia; 4) absence of close biological relatedness among participants; and 5) voluntary informed consent to participate. The exclusion criteria were as follows: 1) presence of oncological disease; 2) severe somatic or autoimmune disease; 3) non-Russian ethnicity; 4) birthplace or residence outside the Central Chernozem region of Russia; and 5) refusal to participate.

The myoma (uterine) nodule volume was calculated using the formula: V (cm³)= (A + B + C) × 0.4186, where A, B, and C represent the longitudinal, anteroposterior, and transverse radii of the myoma nodule (uterus), respectively (cm), and the coefficient 0.4186 corresponds to π4/3, used to calculate the volume of ellipsoidal structures [13]. The total combined nodule volume was calculated for patients with multiple nodules.

Eight polymorphic gene loci associated with SHBG levels according to GWAS data were selected for analysis [14–17] (coordinates given for GRCh38): rs17496332 [NC_000001.11:g.107003753A>G] PRMT6, rs780093 [NC_000002.12:g.27519736T>C] GCKR, rs3779195 [NC_000007.14:g.98364050T>A] BAIAP2L1, rs440837 [NC_000008.11:g.80549739A>G] ZBTB10, rs7910927 [NC_000010.11:g.63379150T>G] JMJD1C, rs4149056 [NC_000012.12:g.21178615T>C] SLCO1B1, rs8023580 [NC_000015.10:g.96165062T>C] NR2F2, and rs12150660 [NC_000017.11:g.7618597G>T] SHBG. Genotyping of the study loci (DNA samples at concentrations of 10–20 ng/μL) was performed using a CFX-96 thermocycler, applying the TaqMan probe method with genotyping kits developed specifically for this purpose by TestGen LLC [18, 19]. Quality control of the resulting genotype data was performed by assessing the Hardy–Weinberg equilibrium (concordance between observed and expected genotype distributions) [20].

Statistical analysis

Statistical analyses were performed using STATISTICA software. The distribution of the continuous variable (myoma nodule volume) was assessed for normality using the Shapiro–Wilk test [21]. As the distribution of this parameter deviated from normal, it was described using the median (Me) and interquartile range (Q1 and Q3). Associations between the single-nucleotide polymorphisms (SNPs) under study and myoma nodule volume were analyzed using transformed values of this parameter (owing to its non-normal distribution), with adjustment for covariates, including age, body mass index, coexistence of uterine myoma with other hyperplastic conditions (endometrial hyperplasia, endometriosis), and myoma nodule localization. Calculations were performed using gPLINK (linear regression method) under four genetic models: allelic, additive, dominant, and recessive [22]. A permutation test was used to correct for multiple comparisons, with p_perm<0.05 considered statistically significant [23].

For the molecular genetic markers rs17496332 [A/G] PRMT6 (chromosome 1) and rs440837 [A/G] ZBTB10 (chromosome 8), which were associated with myoma nodule volume, and for loci in strong linkage disequilibrium with them (threshold r² ≥ 0.80), functional significance was evaluated by in silico analysis [24] using HaploReg (accessed January 09, 2026) [25] and GTEx Portal (accessed 26.09.2026) [26].

Results and discussion

The medical, biological, clinical, and anamnestic characteristics of the study patients with uterine myoma are presented in Table 1. The mean age of patients diagnosed with uterine myoma was 43.69 (7.58) years (range, 24–74 years), and the mean body mass index was 28.18 (5.19) kg/m² (range, 17–49 kg/m²). Among the women included in the analysis, 45.90% had isolated uterine myoma, whereas the remaining 54.10% had concomitant uterine pathology. Most patients (77.51%) had a single myoma nodule. In more than half of the women, the myoma nodules were intramural (51.98%) and larger than 4 cm in diameter (57.14%).

Among patients with uterine myoma, the genotype distributions of all eight analyzed single-nucleotide polymorphisms (SNPs) were in Hardy–Weinberg equilibrium (p>0.05) (Table 2). Associations between two of the eight SHBG candidate gene polymorphic loci and myoma nodule volume were identified (Table 2). The minor G allele of the rs17496332 [A/G] polymorphism in PRMT6 (1p) was associated with a smaller myoma nodule volume under the allelic (β=-0.131, p=0.019, p_perm =0.018), additive (β=-0.135, p=0.015, p_perm =0.014), and recessive (β=-0.328, p=0.003, p_perm =0.004) genetic models (Table 2). In women with uterine myoma carrying the rs17496332 PRMT6 GG genotype, the myoma nodule volume was 20.82 (4.02; 63.48) cm³, which was 2.7–3.1 times lower than in individuals with the AG and AA genotypes (55.79 [16.31; 139.22] and 65.30 [10.13; 138.27] cm³, respectively) (Table 2).

The rs440837 [A/G] polymorphism in ZBTB10 (8p) was also associated with myoma nodule volume under the dominant model (for the G allele: β=-0.142, p=0.050, p_perm=0.050) (Table 2). Among women with uterine myoma carrying the rs440837 ZBTB10 GG and AG genotypes, the median myoma nodule volumes were 43.07 (14.68; 77.00) cm³ and 38.49 (9.16; 107.78) cm³, respectively, representing the smallest volumes compared with patients carrying the AA genotype, for whom the corresponding value was 57.91 (12.69; 147.18) cm³ (Table 2).

According to the in-silico analysis performed using HaploReg and GTEx Portal, the rs17496332 [A/G] variant in PRMT6 is functionally important. Polymorphic loci in linkage disequilibrium with this variant (14 SNPs) also demonstrated substantial functional relevance. Specifically, nearly all of these SNPs (93.33%) determine interactions between the genomic region encompassing PRMT6 (29–60 kb upstream of PRMT6) and 59 transcription factors (TFs). Notably, two loci, rs111232683 and rs4914939, among the 13 SNPs in linkage disequilibrium with rs17496332 [A/G] PRMT6, were involved in regulating DNA interactions with the greatest numbers of TFs (21 and 15 TFs, respectively). More than half of the evaluated SNPs (60.00%) influenced PRMT6 expression across more than 20 tissues and organs, including those involved in both SHBG production (liver) and the pathophysiology of uterine myoma (thyroid gland, adrenal glands, brain, adipose tissue, blood, etc.). Importantly, according to our in silico data, the G allele of rs17496332 was associated with reduced PRMT6 expression in virtually all examined tissues and organs: liver (NES=-0.49, p=1.7×10⁻³⁷), thyroid gland (NES=-0.29, p=2.0×10⁻²⁶), adrenal glands (NES=-0.35, p=1.3×10⁻¹¹), brain (hypothalamus; NES=-0.21, p=0.000016), visceral adipose tissue (NES=-0.28, p=2.7×10⁻¹⁴), subcutaneous adipose tissue (NES=-0.32, p=1.3×10⁻¹⁹), and blood (NES=-0.31, p=4.7×10⁻³⁰). Genome-wide association study (GWAS) findings reported by Coviello A.D. et al. demonstrated an association between the rs17496332 [A/G] polymorphism in PRMT6 and circulating SHBG levels: the common A allele was associated with lower SHBG levels (β=-0.028, p=1×10⁻¹¹), whereas the minor G allele was associated with higher SHBG concentrations [14]. Thus, the G allele of rs17496332 is associated with higher SHBG levels (according to the GWAS data of Coviello A.D. et al. [14]) and a smaller myoma nodule volume in patients with uterine myoma (our data: β=-0.135 to -0.328, p_perm=0.004–0.018).

The rs440837 [A/G] polymorphism in ZBTB10 and five loci in strong linkage disequilibrium with it are located within the ZBTB10/RP11-48B3.3/RP11-48B3.4 genomic region and influence DNA interactions with 22 transcription factors (TFs). In the liver, the primary site of sex hormone-binding globulin (SHBG) synthesis, the rs440837 [A/G] variant in ZBTB10 and the linked rs7013042 variant in RP11-48B3.4 are located within DNA regions containing putative promoters/enhancers (H3K4me3/H3K4me1) as well as active promoters/enhancers (H3K9ac/H3K27ac). The protein encoded by ZBTB10 is a key regulator of gene transcription through its modulatory effects on the binding of RNA polymerase II to genomic DNA [27], which may be relevant to the regulation of SHBG synthesis in the liver. According to GWAS data, the rs440837 [A/G] polymorphism in ZBTB10 is associated with circulating SHBG levels: the common A allele is associated with lower SHBG concentrations (β=-0.030, p=3×10⁻⁹), whereas the minor G allele is associated with higher SHBG concentrations [14]. Thus, the G allele of rs440837 [A/G] ZBTB10 is associated with both higher SHBG concentrations (GWAS data from Coviello A.D. et al. [14]) and smaller myoma nodule volume in patients with uterine myoma (our data: β=-0.142).

Notably, SNPs in linkage disequilibrium with rs440837 [A/G] ZBTB10 are also associated with SHBG concentrations at the genome-wide significance level: rs72688090 (D'=0.85; r²=0.33) [28], rs388922 (D'=0.96; r²=0.53), rs575452 (D'=0.71; r²=0.28), and rs117921873 (D'=1.00; r²=0.26) [15]. These findings underscore the important role of the genomic region encompassing rs440837 [A/G] ZBTB10 in the regulation of circulating SHBG concentrations and suggest its potential involvement in the pathophysiology of uterine myoma.

One possible mechanism underlying the association between SHBG-associated gene polymorphisms and myoma nodule volume is the role of SHBG in testosterone transport and sequestration [6]. Current evidence indicates that approximately 80% of circulating testosterone is bound to SHBG, whereas only 1% circulates as the biologically active free fraction [6]. Consequently, by directly regulating the balance between SHBG-bound and free testosterone, SHBG is likely to substantially influence the biological effects of testosterone in uterine myoma. The close relationship between SHBG and testosterone is further supported by the strong negative genetic correlation between SHBG and circulating free testosterone, which reaches -0.75 in women [17, 28]. Wong J.Y. et al. demonstrated that higher bioavailable testosterone levels are associated with an increased risk of uterine myoma (OR=1.33) [7]. In addition, several studies have shown that testosterone is converted to estradiol by aromatase within myoma cells, thereby promoting local hyperestrogenism in myoma tissue and stimulating myoma growth [6].

SHBG also plays an important role in estrogen transport. Approximately 38% of circulating estrogens are bound to SHBG, whereas only 2% circulate as the biologically active free fraction [29]. Through activation of estrogen receptors (ERα and ERβ), estrogens promote myoma cell proliferation [12, 30] by increasing the expression of growth factors involved in the pathogenesis of this disease, including insulin-like growth factor 1 (IGF-1), epidermal growth factor (EGF), and platelet-derived growth factor (PDGF) [10].

Taken together, our findings and the available literature suggest that SHBG-increasing allelic variants, which are associated with lower levels of biologically active testosterone and estrogens, are associated with smaller myoma nodule volume in patients with uterine myoma.

Conclusion

The SHBG candidate gene polymorphisms rs17496332 [A/G] in PRMT6 and rs440837 [A/G] in ZBTB10 are associated with myoma nodule volume in patients with uterine myoma. The G alleles of rs17496332 [A/G] PRMT6 and rs440837 [A/G] ZBTB10 are associated with the smallest myoma nodule volume.