Hypermethylation of the BRCA2 gene promoter and its co-hypermethylation with the BRCA1 gene promoter in patients with breast cancer.

BACKGROUND: The BRCA2 gene is an important tumour suppressor in breast cancer, and alterations in BRCA2 may lead to cancer progression. The aim of the study was to investigate the association of hypermethylation of the BRCA2 gene promoter and its co-hypermethylation with the BRCA1 gene promoter with the development and course of breast cancer in women. METHODS: This study included 74 women with breast cancer (tumour tissue samples and peripheral blood) and 62 women without oncological pathology (peripheral blood) - control group. RESULTS: Hypermethylation of the BRCA2 gene was significantly more frequently detected in the tumour tissue of women with breast cancer compared to their peripheral blood and peripheral blood of control subjects (p= 0.0006 and p= 0.00001, respectively). Hypermethylation of BRCA2 was more frequently detected in patients with breast cancer over the age of 50 and in patients with higher Ki67 expression levels (p= 0.045 and p= 0.045, respectively). There was a high frequency of unmethylated BRCA1 and BRCA2 gene combination in women of the control group compared to women with breast cancer, both in blood samples and tumour tissue samples (p= 0.014 and p= 0.00001, respectively). CONCLUSION: Our study confirms the hypothesis that BRCA2 hypermethylation plays an important role in the pathogenesis of breast cancer and the importance of assessing its co-hypermethylation with BRCA1 in predicting the course of the disease.

Association of the C3953T (rs1143634) variant of the interleukin 1 beta gene with the features of a complicated course of COVID-19-associated pneumonia.

BACKGROUND: The pro-inflammatory cytokine IL-1 plays an important role in severe COVID-19. A change in IL-1 production may be associated with a mutation in the IL1Β gene. Our study analyzed the impact of the IL1Β gene variants (rs1143634) on disease progression in patients with severe COVID-19 pneumonia, taking into account treatment strategies. METHODS AND RESULTS: The study enrolled 117 patients with severe COVID-19 pneumonia. The IL1Β gene variants were identified using the polymerase chain reaction-restriction fragment length polymorphism method. In the group of patients, the following genotype frequencies were found based on the investigated rs1143634 variant of the IL1Β gene: CC-65.8%, CT-28.2%, and TT-6.0%. Our results showed that the group of patients with the T allele of the IL1Β gene had higher leukocyte counts (p = 0.040) and more pronounced lymphopenia (p = 0.007). It was determined that patients carrying the T allele stayed on ventilators significantly longer (p = 0.049) and required longer treatment with corticosteroids (p = 0.045). CONCLUSION: Identifying variants of the IL1Β gene can be used as a predictive tool for assessing the severity of COVID-19 pneumonia and tailoring personalized treatment strategies. Further research with a larger patient cohort is required to validate these findings.

Combination of 15q24 Microdeletion Syndrome and Metabolic Imbalance in a Patient with Atypical Autism.

Autistic spectrum disorders (ASD) in children are becoming increasingly common, reaching epidemic proportions. Among the various causes contributing to the development of ASD, the leading place belongs to both chromosomal pathologies and genetic syndromes and their consequence - metabolic imbalance or severe metabolic disorders. Depending on the degree of metabolic pathway damage, certain phenotypes of ASD are formed. A deletion of ~3.1 Mb of chromosome 15q24 was detected in the examined 2-year-old boy with a "mild phenotype" of autism without an obvious delay in mental development. A wide range of additional studies included genetic testing of folate metabolism genes and analysis of metabolites of the methylation cycle and detection of antibodies to folic acid alpha receptors. A heterozygous variant of the MTHFR gene (rs1801133), moderate hyperhomocysteinemia, hypermethylation, and an increased titer of antibodies to alpha receptors of folic acid were revealed in the patient. This clinical case indicates the need for a multifaceted clinical and laboratory examination in children with ASD to identify the metabolic phenotype and prescribe personalized treatment. A personalized treatment strategy will improve the cognitive functions, psycho-emotional state, and social adaptation of individuals with ASD in the long term."

Clinical significance of determining the hypermethylation of the RUNX3 gene promoter and its cohypermethylation with the BRCA1 gene for patients with breast cancer.

PURPOSE: The aim of this study was to assess the clinical significance of RUNX3 gene hypermethylation in the pathogenetic mechanisms of breast cancer in women, taking into account its cohypermethylation with the BRCA1 gene. METHODS: This study included 74 women with newly diagnosed breast cancer (samples from female primary breast carcinomas and paired peripheral blood samples) and 62 women without oncological pathology-control group (peripheral blood samples). Epigenetic testing for hypermethylation status studying was performed in all samples on freshly collected material with the addition of a preservative before the storage and DNA isolation. RESULTS: Hypermethylation of the RUNX3 gene promoter region was detected in 71.6% samples of breast cancer tissue and in 35.13% samples of blood. The RUNX3 gene promoter region hypermethylation was significantly higher among breast cancer patients compared to the control group. The frequency of cohypermethylation in RUNX3 and BRCA1 genes was significantly increased in breast cancer tissues compared to the blood of patients. CONCLUSION: A significantly increased frequency of the hypermethylation of the RUNX3 gene promoter region and its cohypermethylation with the BRCA1 gene promoter region was found in tumor tissue and blood samples from patients with breast cancer, in contrast to the control group. The identified differences indicate the importance of further investigations of suppressor genes cohypermethylation in patients with breast cancer. Further large-scale studies are needed to find out whether the detected hypermethylation and cohypermethylation of the RUNX3 gene promoter region will have an impact on the treatment strategy in patients.

NOS3 (rs61722009) gene variants testing in prediction of COVID-19 pneumonia severity.

BACKGROUND: There is a hypothesis that a sufficient level of endothelial nitric oxide synthase is important for reliable protection against COVID-19. Theoretical ideas about the NOS3 gene demonstrated that it can have an effect on links of the complications pathogenesis in COVID-associated pneumonia. We determined the goal - to investigate the association of the NOS3 gene variants with the occurrence of the disease and its clinical course in patients of the intensive care unit. METHODS: The study group included 117 patients with a diagnosis of severe "viral COVID-19 pneumonia". Determination of NOS3 gene variants was performed using the PCR method. The probability of differences in the quantitative results were determined using ANOVA or Kruskal-Wallis test (depend of normality of studied parameters). RESULTS: Our results indicate that the presence of the NOS3 gene 4a allele increase the risk of complicated COVID-19-associated pneumonia (χ2 = 18.84, p = 0.00001, OR = 3.53 (1.95-6.39)). It was showed, that carriers of the 4aa genotype had a significantly higher ratio of SpO2/FiO2 on the first and second days after hospitalization (p = 0.017 and p = 0.03, respectively). Patients with the 4aa genotype also had the acid-base imbalances, as showed by indicators of base deficiency and standard bicarbonate, which were beyond the reference values. Potassium and sodium concentrations on the first and second day after hospitalization were also significantly lower in patients with 4aa genotype (p = 0.009 and p = 0.048, respectively), for whom, in the same time, the concentrations of C-reactive protein and total bilirubin were significantly higher (p = 0.002 and p = 0.033, respectively). CONCLUSIONS: Our results confirmed that the rs61722009 variant of the NOS3 gene is associated with an increased risk of severe СOVID-19-associated pneumonia and its adverse clinical course with potential progression of kidney and liver damage, and occurrence risk of systemic inflammatory response syndrome. These results require further research for the new metabolic strategy formation, in order to prevent the severe COVID-19 associated pneumonia and its complications.

SFTPB (rs11130866) and NR3C1 (rs41423247) gene variants as potential clinical biomarkers for personalized treatment strategy selection in patients with severe COVID-19 pneumonia.

BACKGROUND: Exploring the pathogenetic mechanisms behind severe lung damage in COVID-19 is crucial. In this study, we decided to focus on two molecular markers that affect surfactant metabolism and lung development: the surfactant protein B (SFTPB) and the glucocorticoid receptor (NR3C1) genes. The aim of our study was to determine the effect of SFTPB (rs11130866) and NR3C1 (rs41423247) gene variants on the course of the disease in patients with COVID-19, and the treatment measures they required. METHODS: The study group included 58 patients with a diagnosis of severe "viral COVID-19 pneumonia." Determination of SFTPB and NR3C1 gene variants was performed using the PCR-RFLP method. RESULTS: Our results indicate that the presence of the SFTPB gene CC genotype increases the risk of developing acute respiratory distress syndrome in patients with COVID-19 (χ2 = 4.03, p = 0.045, OR = 3.90 [1.19-12.78]). However, patients with the SFTPB gene TT genotype required respiratory support for a shorter period of time. Patients with the NR3C1 gene CC genotype underwent a longer glucocorticoid therapy. Moreover, for patients with the CC genotype, a longer stay in the intensive care unit was detected before lethal outcome. CONCLUSIONS: The obtained results confirm the influence of the SFTPB (rs11130866) and NR3C1 (rs41423247) gene variants on the therapy, course, and severity of the disease in patients with COVID-19. Of course, these results require further study, analysis, and larger, complex, systematic research.

Hyperhomocysteinemia in men and women of married couples with reproductive disorders. What is the difference?

Hyperhomocysteinemia (HHcy) is an autosomal recessive inherited metabolic disease caused by variations in folate metabolism genes, characterized by impaired methionine metabolism and accumulation of homocysteine (Hcy) in the blood serum. It was shown that men usually have higher plasma Hcy levels than women, but have not yet assessed the leading factors of these differences, which is important for the development of personalized protocols for the prevention of folate metabolism disorders in couples with reproductive disorders. This study aimed to analyze the effect of intergenic and gene-factor interactions on the risk of developing HHcy in men and women of married couples with reproductive disorders. In our study were involved 206 married Caucasian couples (206 males and 206 females) from central regions of Ukraine with early pregnancy losses in the anamnesis. We found that the incidence of HHcy in men was significantly higher than in women. Gender differences in folic acid and vitamin B12 levels were identified. The best predictors of HHcy in men (MTRR (A66G), MTHFR (C677T), MTR (A2756G), vitamin B12 level) and in women (MTHFR (C677T), MTR (A2756G), vitamin B12 level) were selected by binary logistic regression. There was no significant difference in the distribution of genotypes by the studied gene variants when comparing men and women with HHcy. Our findings demonstrate that there is a gender difference in the development of HHcy. This difference is caused by intergenic interaction and by environmental factors, in particular, nutrition and vitamins consumption.

Betaine consumption as a new clinical approach to treatment and prophylaxis of folate-related pathologies.

The most important pathway in the development of folate-related pathologies is an increase in the level of homocysteine (HC). HC, a cytotoxic and neurotoxic amino acid (when its level is ≥12 µmol/L), is 1 of the most widely studied compounds in cardiology, neurobiology, oncology, and embryology for the last 20 years. Given its toxicity, the processes of endogenous detoxification of HC are of particular interest to medicine. To date, the most studied pathway is that of remethylation (the conversion of HC to methionine), with the participation of B12- and B9-dependent methionine synthase. Less studied is remethylation with the participation of the choline derivatives betaine and betaine-HC-S-methyltransferase (BHMT). Therefore, the aim of this review was to conduct a theoretical analysis of available information regarding the contribution of betaine metabolism, its enzyme, and its genetic polymorphism to folate metabolism disturbances, and the development of folate-related pathologies. This review emphasizes the potential clinical significance of 2 factors that can influence the remethylation reaction of HC: the use of betaine and identifying the BHMT gene variants and their impact on the risk for developing certain folate-related pathologies, and treatment options. Moreover, with a high level of methylation of the BHMT gene and in the presence of its low-function variants (eg, rs3733890), it is necessary to use betaine as an additional methyl donor, especially during folate therapy. More clinical research is needed to identify the effects of the different BHMT gene variants on the individual risk for folate-related pathologies to better assess the clinical significance, the need for genetic testing, and betaine consumption.

Methylation of promoter region of BRCA1 gene versus pathogenic variants of gene: risk factor or clinical marker of breast cancer.

BACKGROUND: In this study, we compared the contribution of pathogenic variants of the BRCA1/2 genes (5382insC, 185delAG, 6174delT, 4153delA, T300G) and hypermethylation of the BRCA1 gene promoter region to the risk of breast cancer and clinical features in women. METHODS: This study enrolled 74 women (tumor tissue, blood) with newly diagnosed breast cancer and 62 women (blood) without oncological pathology (control group). Molecular genetic testing of samples and determination of hypermethylation status were performed on freshly collected material with the addition of a preservative before the procedure of DNA isolation. RESULTS: Hypermethylation of the BRCA1 gene promoter in women is a risk breast cancer factor (χ2 = 19.10, p = 0.001, OR = 16.25 (3.67-71.92)) and is more common than major pathogenic variants in the BRCA1/2 genes. The patients with the BRCA1 gene promoter hypermethylation were more likely to be diagnosed with late-stage metastatic cancer (χ2 = 4.31, p = 0.038, OR = 4.04 (1.19-13.65)). Hypermethylation of the BRCA1 gene promoter was predominant in tumor tissue among BC patients without family history compared to patients with cancer in relatives. CONCLUSION: We proved that hypermethylation of the BRCA1 gene promoter is a risk factor for breast cancer and possibly an early biological marker of clinical onset, as its presence contributed to rapid disease progression with metastasis. The high frequency of hypermethylation in the examined breast cancer patients may be a consequence of environmental factors pressure on the risk of the disease development. Further large-scale studies are needed for the clinical application of the results.

The analysis of using a panel of the most common variants in the PAH gene for the newborn screening in Ukraine.

Phenylketonuria (PKU) is hyperphenylalaninemia that develops due to a deficiency of the phenylalanine hydroxylase enzyme (PAH). Identification of variants in the PAH gene is necessary for verification of the diagnosis, choice of treatment tactics, detection of heterozygous carriers. The aim of the study was to analyze the effectiveness of identification of selected pathological variants in the PAH gene during the newborn screening program. This study relied on the results of the examination of 257 patients (138 boys and 119 girls) with hyperphenylalaninemia from different regions of Ukraine. Genotyping was performed on nine pathogenic variants in PAH gene: I65T, R261Q, G272*, R252W, R261*, R408W, IVS12 + 1G > A, Y414C, IVS10-11G > A. According to the results of the study, variants R408W (AF = 52.7%), R252W (AF = 3.5%) and Y414C (AF = 1.8%) were the most common. More than half of the examined patients (51.7%) had a compound genotype with a major variant of R408W in one allele. Approximately a quarter of the examined patients (26.8%) had the R408W/R408W genotype. In 12.1% of patients, the applied panel of variants of the РАН gene did not allow us to identify the pathogenic variant in any allele. We conclude that the selected panel allowed us to identify the presence of variants in 87.9% of patients with PKU. The panel of genetic testing in the PAH gene for the newborns that we used for the study allows accurate prediction of some phenotypes for therapy planning. But in-depth analysis of pathological gene variants may be necessary for unclear and difficult cases of the disease, and for genetic counseling of patients families.

Contribution of BRCA1 5382insC mutation to triplene-gative and luminal types of breast cancer in Ukraine.

PURPOSE: The gene BRCA1 plays a key role in DNA repair in breast and ovarian cell lines and this is considered one of target tumor suppressor genes in same line of cancers. The 5382insC mutation is among the most frequently detected in patients (Eastern Europe) with triple-negative breast cancer (TNBC). In Ukraine, there is not enough awareness of necessity to test patients with TNBC for BRCA1 mutations. That is why this group of patients is not well-studied, even through is known the mutation may affect the course of disease. METHODS: The biological samples of 408 female patients were analyzed of the 5382insC mutation in BRCA1. We compared the frequency of the 5382insC mutation in BRCA1 gene observed in Ukraine with known frequencies in other countries. RESULTS: For patients with TNBC, BRCA1 mutations frequency was 11.3%, while in patients with luminal types of breast cancers, the frequency was 2.8%. Prevalence of 5382insC among TNBC patients reported in this study was not different from those in Tunisia, Poland, Russia, and Bulgaria, but was higher than in Australia and Germany. CONCLUSION: The BRCA1 c.5382 mutation rate was recorded for the first time for TNBC patients in a Ukrainian population. The results presented in this study underscore the importance of this genetic testing of mutations in patients with TNBC. Our study supports BRCA1/2 genetic testing for all women diagnosed with TNBC, regardless of the age of onset or family history of cancer and not only for women diagnosed with TNBC at <60y.o., as guidelines recommend.

Modifying effects of TNF-α, IL-6 and VDR genes on the development risk and the course of COVID-19. Pilot study.

OBJECTIVES: COVID-19 continues to range around the world and set morbidity and mortality antirecords. Determining the role of genetic factors in the development of COVID-19 may contribute to the understanding of the pathogenetic mechanisms that lead to the development of complications and fatalities in this disease. The aim of our study was to analyze the effect of TNF-α (rs1800629), IL-6 (rs1800795) and VDR (rs731236 and rs1544410) genes variants on the development risk and the course of COVID-19 in intensive care patients. METHODS: The study group included 31 patients with diagnosis "viral COVID-19 pneumonia". All patients underwent standard daily repeated clinical, instrumental and laboratory examinations. Determination of IL-6, TNF-α, and VDR genes variants was performed using the PCR-RFLP method. RESULTS: It was found a significant increase in the rate of the CC genotype and C allele (38.7 vs. 12.0% and 0.6 vs. 0.4%, respectively) of the IL-6 gene in all patients of the study in comparison with population frequencies. There was a significantly higher rate of heterozygous genotypes TC and GA of the VDR gene in group of died patients. The rs1800629 variant of the TNF-α gene is associated with the need for respiratory support and its longer duration in patients with COVID-19. CONCLUSIONS: The obtained results support a hypothesis about the influence of variants of IL-6, TNF-α and VDR genes on severity of COVID-19. However, in order to draw definite conclusions, further multifaceted research in this area are need.

Association between rankl [RS9594759] and IL10 [RS1800896] Genes polymorphism and deciduous tooth eruption terms in Ukrainians born macrosomic.

OBJECTIVE: The aim: The article deals with analyzing the influence of polymorphic variants of CYP19A1 [rs2414096, rs936306], ESR1 [rs2234693, rs9340799], IL1 [rs1143627], IL6 [rs1800796], IL10 [rs1800896] and RANKL [rs959389] genes on deciduous tooth eruption terms in individuals born macrosomic. PATIENTS AND METHODS: Materials and methods: 171 individuals participated in the multi-stage study (144 macosomic-at-birth individuals and 27 normosomic-at-birth persons). This study included only persons who have preserved information about the timing of deciduous tooth eruption - 159 persons (aged from 4 to 55 years), male and female (male / female ratio was 1.5 / 1). RESULTS: Results and conclusions: The presence of the G allele in CYP19A1 [rs2414096] gene and the -351 A allele in ESR1 [rs9340799] gene were found to be risk factors for fetal macrosomia formation. The research revealed an association of RANKL [rs9594759] gene variants which is a multiplicative model of inheritance and IL-10 [rs1800896], an overdominant model of inheritance, with an increased risk of tooth delay. Besides the variants of RANKL [rs9594759] and IL-10 [rs1800896] genes a multidirectional modifying effect on the timing of tooth eruption in macrosomic-at-birth individuals made the variant of CYP19A1 [rs2414096] gene - a significant dominant and over-dominant model of inheritance. Further analysis of intergenic interactions will facilitate the application of the obtained results in clinical practice by creating a molecular profile of individuals with deviations in the tooth eruption timing.