Aims: Pathogenic variants in the CYP21A2 gene are related to the classic and non-classic forms of congenital adrenal hyperplasia (CAH). However, the role of CAH carrier status in the clinical presentation of polycystic ovarian syndrome (PCOS) is still unclear. Moreover, the possible associations of different CYP21A2 gene polymorphisms with metabolic and reproductive abnormalities in PCOS have not been investigated. Therefore, the present study aims to examine the prevalence of the most common CYP21A2 pathogenic variant IVS2-13A/C>G (c.293-13A/C>G) in Eastern European women with PCOS and to evaluate the associations between common intron 2 genetic polymorphisms and the clinical symptoms of the patients. Methods: Sixty consecutively recruited women with PCOS were genotyped for the CYP21A2 intron 2 IVS2-13A/C>G genetic variant. Additionally, CYP21A2 intron 2 polymorphic variants rs6453 (c.293-44G>T), rs6451 (c.293-67C>A/G), rs369651496 (c.293-104del), and rs6474 (c.308G>A/p.R103L) were tested and described. The clinical and hormonal characteristics were compared in women with PCOS and with polymorphic and wild-type genotypes. Results: The heterozygous CYP21A2 pathogenic variant IVS2-13A/C>G was found in one of the investigated PCOS patients (1.67%) with a non-hyperandrogenic type of PCOS. The presence of the rs6453 (c.293-44G>T) T-allele was associated with increased levels of DHEAS (15.18 vs. 9.14 µmol/L, p = 0.003) compared to the wild-type genotype in the investigated group. The rs6451 (c.293-67C>A/G) minor alleles were associated with an earlier age of menarche in the patients (12.0 vs. 13.0 years, p = 0.007). The polymorphic rs369651496 minor 6G allele was related to a better lipid profile in the women with PCOS, while the rs6474 variant modulated the blood pressure of the patients. Conclusions: The presence of CYP21A2 genetic minor alleles of rs6467 (IVS2-13A/C, c.293-13A/C), rs6453 (c.293-44G>T), rs6451 (c.293-67C>A/G), rs369651496 (c.293-104del), and rs6474 (c.308G>A/p.R103L) might modulate the adrenal androgens, age of menarche, and metabolic features in women with PCOS. Further studies on 21-hydroxylase genetic variants (pathogenic and polymorphisms) in different ethnic groups might help reveal the influence of adrenal steroidogenesis on PCOS development, clinical manifestations, and lifelong cardiovascular risks.

The present study aimed to investigate the possible associations between the melatonin receptor 1B (MTNR1b) genetic polymorphisms and the presence of reproductive disturbances due to spontaneous abortion and implantation failure leading to pregnancy loss. The patients group included 35 women presenting with one or more miscarriages and/or implantation failure. Another 36 healthy women without history of implantation failure or miscarriages and with at least one birth served as a control group. Genomic DNA was extracted and genotyping for the MTNR1b polymorphisms rs1562444, rs10830962 and rs10830963 was performed by PCR-RFLP analysis. The genotype distribution of the investigated MTNR1b polymorphisms in patients with reproductive failure did not differ significantly to the group of women with at least one birth (p > 0.05 for all). Nevertheless, the presence of a haplotype rs1562444 AA rs10830962 GG was significantly less common in women with miscarriages or implantation failure than in women with at least one birth (2.9% vs. 22.2%, p = 0.028). We suggest that the haplotype might be protective against implantation failure and spontaneous miscarriage. Additionally, rs1562444 AA genotype was not found in women with recurrent pregnancy loss. Studies in larger populations are necessary to reveal the proper associations between different MTNR1b haplotypes, ovarian function and reproductive failure.

Germline mutations affecting STK11 (LRG_319) are profoundly studied in relation to Peutz-Jeghers syndrome, predisposing to the development of various cancers at multiple sites. Though somatic mutations in STK11 are found to be present in several cancers, limited data on its involvement in sporadic breast cancer are available. The present study aims to evaluate the frequency and spectrum of genetic alterations in STK11 in a group of Bulgarian patients with sporadic breast cancer. A total of 73 tumor and 22 corresponding blood specimens derived from the patients, and 10 blood samples from clinically healthy controls were analyzed. High Resolution Melting analysis followed by Sanger sequencing and bioinformatic prediction tools were utilized. Seven patients (9.58%) harbored STK11 alterations, only two (2.74%) of which are exonic: one nonsense c.322A>T; p.K108X (deleterious) and one missense c.440G>A; p.Arg147His (of unknown significance). Two intronic variants were also observed: c.290+36G>T and c. *16+18C>A (novel). To our knowledge the results represent the first data indicating presence of STK11 alterations in patients with sporadic breast cancer. The limited number of the detected deleterious mutations indicates that mutational inactivation of the gene is a rare event and probably plays a minor role in sporadic breast carcinogenesis.

Background and Clinical Significance: Anti-Müllerian hormone (AMH) is a dimeric glycoprotein secreted from the granulosa cells of the preantral and small antral follicles, which has entered routine clinical practice as a valuable tool for the diagnosis of different ovarian disorders. Increased AMH levels have been recommended as a criterion for polycystic ovary syndrome (PCOS). However, its widespread use remains limited due to analytical diversity and contradictory age-specific thresholds, among other factors that modulate AMH levels. Case Presentation: Herein, we present a rare case series of women with increased AMH levels. The difficulties in the differential diagnosis of patients with elevated AMH levels, because of PCOS combined with pituitary dysfunction, increased ovarian volume, or granulosa cell tumors (GCTs), are discussed. Conclusions: The presented rare cases of increased AMH emphasize the important role of AMH as a diagnostic marker in women with hypogonadotropic hypogonadism and granulosa cell tumors. On the other hand, it is still unknown if increased AMH produced by unusually enlarged or supernumerary ovaries should be considered as actual PCOS cases or as a specific subgroup. Additionally, the unusual case of GCTs with pronounced AMH and LH increase but normal steroids supports the pathophysiological role of AMH for the development of neuroendocrine dysfunction. Moreover, it suggests that GCTs should be considered in the differential diagnosis of chronic anovulation even in women with normal ovarian steroid production in case of unusually high AMH levels for the age. Further studies are needed to explain PCOS heterogeneity and to ensure proper differential diagnosis for every affected woman.