The Methylation of the p53 Targets the Genes MIR-203, MIR-129-2, MIR-34A and MIR-34B/C in the Tumor Tissue of Diffuse Large B-Cell Lymphoma.

The regulation of oncogenes by microRNA is a focus of medical research. hsa-miR-203, hsa-mir-129, hsa-miR-34a, hsa-miR-34b and hsa-miR-34c are oncosuppressive microRNAs that mediate the antitumor activity of p53. We seek to evaluate the frequencies, co-occurrence and clinical significance of the methylation of the MIR-203, MIR-129-2, MIR-34A and MIR-34B/C genes in the tumor tissue of diffuse large B-cell lymphoma (DLBCL). The methylation was assessed in 73 samples of DLBCL and in 11 samples of lymph nodes of reactive follicular hyperplasia by Methyl-Specific Polymerase Chain Reaction (MS-PCR) and Methylation-Sensitive High-Resolution-Melting (MS-HRM) methods. All four studied genes were not methylated in the tissue of reactive lymphatic nodes. The methylation frequencies of the MIR-129-2, MIR-203, MIR-34A and MIR-34B/C genes in lymphoma tissue were 67%, 66%, 27% and 62%, respectively. Co-occurrence of MIR-203, MIR-129-2 and MIR-34B/C genes methylation, as well as the methylation of MIR-34B/C and MIR-34A pair genes were detected. The MIR-34A gene methylation was associated with increased International Prognostic Index (IPI) (p = 0.002), whereas the MIR-34B/C (p = 0.026) and MIR-203 (p = 0.011) genes' methylation was connected with Ki-67 expression level in tumor tissue at more than 45%. We found an increasing frequency of detection of MIR-34A gene methylation in the group of patients with the Germinal-Center B-cell like (GCB-like) subtype of DLBCL (p = 0.046). There was a trend towards a decrease in the remission frequency after the first line of therapy (p = 0.060) and deterioration in overall survival (OS) (p = 0.162) in patients with DLBCL with methylation of the MIR-34A promoter. The methylation of the MIR-34A, MIR-34B/C, MIR-129-2 and MIR-203 genes in DLBCL is tumor-specific and occurs in combination. The methylation of the studied genes may be a potential differential diagnostic biomarker to distinguish between lymphoma and reactive lymph nodes, while its independent predictive value has not been confirmed yet.

The rs78378222 prevalence and the copy loss of the protective allele A in the tumor tissue of diffuse large B-cell lymphoma.

BACKGROUND: Rare single nucleotide polymorphisms (SNPs) are likely to be a crucial genetic factor for human diseases, including cancer. rs78378222 is rare SNP in 3'-untranslated region (UTR) of TP53 gene leading to disturbance of 3'-end mRNA processing. The frequency of rs78378222 varies in several studied populations. The meta-analysis of 34 genome-wide association studies indicated that rs78378222 was significantly associated with an increased risk of cancer overall. Bioinformatic analysis indicates that somatic loss of the protective A allele of rs78378222 occurs in the tumor tissue of some malignant. The goal of the current study is to document the rs78378222 prevalence and evaluate the copy loss status of the protective allele A in the tumor tissue of patients with diffuse large B-cell lymphoma (DLBCL). METHODS: Total DNA was isolated from FFPE-samples and peripheral blood of patients with DLBCL and comparable in age and sex controls. rs78378222 genotyping was performed by the PCR-RFLP method using restriction endonuclease HindIII. Direct Sanger's sequencing was used to confirm the presence of C allele of the rs78378222. The search for TP53 gene mutations was carried out by Sanger's direct sequencing method, according to the IARC protocol. RESULTS: The result of genotyping of 136 DNA samples from DLBCL tumor tissue suggested that frequency of the rs78378222 was 11/136 (8.1%). Rare allele C frequency was 11/272 (4.2%). A total of 5/11 DLBCL rs78378222 heterozygous samples had the heterozygosity loss in the TP53 gene. Only one of these cases was combined with TP53 gene mutations which have proven oncogenic potential-p.Arg196Gln, other four cases have not mutations in the coding regions of gene. CONCLUSIONS: At the stages of DLBCL initiation or progression a loss of the protective allele A of rs78378222 occurs. Further efforts are needed to study possible molecular mechanisms underlying somatic alterations in DLBCL in this region of the TP53 3'-UTR as well as functional studies to illustrate how the presents of rs78378222 may affect tumor progression of lymphoma.

Clinical aspects of TP53 gene inactivation in diffuse large B-cell lymphoma.

BACKGROUND: The knowledge about specific mechanisms generating TP53 dysfunction in diffuse large B-cell lymphoma is limited. The aim of the current study was to comprehensively explore TP53 gene variability resulting from somatic mutations, promoter methylation, and allelic imbalance in tumorous tissue of diffuse large B-cell lymphoma (DLBCL). METHODS: DNA samples from 74 patients with DLBCL were used. Genomic DNA was isolated from paraffin blocks of lymph nodes or from extranodal biopsies of tumors by the phenol-chloroform extraction method with guanidine. Analysis of coding sequences of the TP53 gene was based on Sanger's direct sequencing method. The methylation status of the TP53 promoter was analyzed using by methylation-specific PCR on bisulfite-converted DNA. Assessment of the detected mutations was carried out in the IARC TP53 Database and the TP53 UMD mutation database of human cancer. RESULTS: The mutations in regions coding for the DNA-binding domain were prevalent (95%). In the analyzed sample of patients, codons 275, 155, 272, and 212 were hotspots of mutations in the TP53 gene. In addition, functionally significant intron mutations (IVS6-36G > C and IVS5 + 43G > T) were detected. Instances of TP53 promoter methylation were observed only in a few samples of diffuse large B-cell lymphoma tissue. Furthermore, loss of heterozygosity was revealed only in the subgroup of patients with altered status of the gene (mutations were detected in five patients and promoter methylation in one case). CONCLUSIONS: Thus, the results suggest that there are two sequential events in the formation of diffuse large B-cell lymphoma in at least some cases. The first event is mutation or methylation of the TP53 promoter, leading to appearance of a cell with increased risk of malignant transformation. The second event is the loss of an intact allele of the gene; this change is necessary for tumorigenesis. We identified TP53 mutation patterns in a Russian cohort of patients with de novo DLBCL who were treated with R-CHOP and R-CHOP-like regimens and confirmed that TP53 mutation status is a valuable prognostic biomarker.

The polymorphism of dopamine receptor D4 (DRD4) and dopamine transporter (DAT) genes in the men with antisocial behaviour and mixed martial arts fighters.

Objectives: Variable-number tandem repeat (VNTR) polymorphisms of DRD4 and DAT genes were studied in the Russian and Chechen men convicted of crimes, and two control groups comprised of the MMA fighters and a sample of general population. A group of MMA fighters included only the subjects without history of antisocial behaviour. Methods: DNA was isolated by phenol-chloroform extraction from the blood. Genotyping VNTR polymorphisms of the DRD4 and DAT genes were performed by PCR on published methods. Results: Among those convicted of felonies and most grave crimes, carriers of DRD4 long alleles are found more frequently, similarly to the cohort of MMA fighters (lacking criminal record in both paternal lines). The 9/9 DAT genotype carriers are more frequently encountered among the habitual offenders. A frequency of the combination of the DRD4 genotype 4/7 and DAT genotype 10/10 is clearly higher among the convicts of violent crimes and the MMA fighters. One can speculate the presence of a 'controlled aggression' without a predisposition to pathological violence in the MMA fighters. Conclusions: Our study supports the hypothesis of genetic predisposition to different variants of extreme behaviour mediated by genetic determinants involved in the functioning of neuromediator systems including those controlling dopamine pathways.

GLCE rs3865014 (Val597Ile) polymorphism is associated with breast cancer susceptibility and triple-negative breast cancer in Siberian population.

d-Glucuronyl C5-epimerase (GLCE) is one of key enzymes in heparan sulfate biosynthesis and possesses tumour-suppressor function in breast carcinogenesis. Here, we investigated a potential involvement of GLCE polymorphism(s) in breast cancer development in Siberian women population. Comprehensive analysis of SNP databases revealed GLCE rs3865014 (Val597Ile) missense polymorphism as the main significantly present in human populations. According the TaqMan-based SNP assay, allele distributions for the rs3865014 (A>G) were similar in healthy Siberian women (n=136) and cancer patients (n=129) (A0,73:G0,27) and intermediate between the European and Asian populations, while genotype distributions were different, with the increase of AG rate in breast cancer patients (OR=1.76; 95% CI=1.04-1.90; P(Y)=0.035 χ2=4.44). Heterozygous AG genotype was associated with tumour size (OR=3.67, P(Y)=0.004), ER-negative tumours (OR=3.25, P(Y)=0.0028), triple-negative tumours (OR=4.94, P(Y)=0.015) but not menopausal status, PR and HER-2 status, local or distant metastasis. Homozygous GLCE genotypes (AA/GG) were more common for ER+PR+ luminal A breast cancer (OR=0.25, P(Y)=0.031). Loss-of-heterozigosity was identified in 5 of 51 breast tumours and the loss of G allele was associated with the decreased GLCE expression. Epidemiologic data for the GLCE SNP in different racial/ethnic groups demonstrated high AG genotype rates as a risk factor not for breast cancer incidence but for poor prognosis of the disease. The obtained data suggest an involvement of GLCE rs3865014 in breast cancer development. Heterozygous AG genotype might be a risk factor for breast cancer susceptibility in Siberian women and is associated with aggressive ER-negative and triple-negative cancer subtypes.

Association of the genetic markers for myocardial infarction with sudden cardiac death.

OBJECTIVE: Investigate the association of rs17465637 gene MIAF3 (1q41), rs1376251 gene TAS2R50 (12p13), rs4804611 gene ZNF627 (19p13), rs619203 gene ROS1 (6q22), rs1333049 (9p21), rs10757278 (9p21), rs2549513 (16q23), rs499818 (6p24) associated with myocardial infarction available from the international genome-wide studies with sudden cardiac death (SCD) in a case-control study. METHODS: A sample of SCD cases (n=285) was formed using the WHO criteria; the control sample (n=421) was selected according to sex and age. DNA was isolated by phenol-chloroform extraction from the myocardial tissue of SCD cases and blood of control cases. The groups were genotyped for the selected SNPs by real-time PCR using TaqMan probes (Applied Biosystems, United States). RESULTS: No statistically significant differences in the genotype and allelic frequencies of studied single nucleotide polymorphisms between sudden cardiac death cases and control were detectable in general group. By separating the groups of sex and age differences in the genotype frequencies of rs1333049, rs10757278 and rs499818 are statistical significance. Genotypes CC of rs1333049 and GG of rs10757278 are associated with an increased sudden cardiac death risk in men (p=0.019, OR=1.7, 95% CI 1.1-2.8; p=0.011, OR=1.8, 95% CI 1.2-2.8, respectively). Genotype AG of rs499818 is associated with an increased sudden cardiac death risk in the women over 50 years old (p=0.009, OR=2.4, 95% CI 1.3-4.6). CONCLUSION: Polymorphisms rs1333049 and rs10757278 are associated with SCD in men and rs499818 in the women aged over 50 years.

Transient receptor potential melastatin 8 gene polymorphism is associated with cold-induced airway hyperresponsiveness in bronchial asthma.

BACKGROUND AND OBJECTIVE: Cold-induced airway hyperresponsiveness (CAH) is common in bronchial asthma (BA) patients and represents a problem for those living in cold climate. Transient receptor potential melastatin 8 (TRPM8) channel is the main cold temperature sensor in humans that could mediate cold response in asthmatics with CAH. No associations between TRPM8 gene polymorphisms and CAH have been reported. METHODS: The present study involved 123 BA patients. CAH was assessed by 3-min isocapnic (5% CO2 ) cold air (-20°C) hyperventilation challenge. The c.750G > C (rs11562975), c.1256G > A (rs7593557), c.3048C > T (rs11563208) and c.3174C > G (rs11563071) polymorphisms of TRPM8 gene were genotyped by allele-specific polymerase chain reaction (PCR) and PCR with subsequent restriction fragment length polymorphism analysis. RESULTS: GC genotype and C allele carriers of the c.750G > C (rs11562975) polymorphism were more frequently observed to exhibit CAH. The estimated odds ratio for the GC genotype was 3.73 95%CI (1.48; 9.37), P = 0.005. Furthermore, GC heterozygotes had a prominent decrease in forced expiratory volume in 1 s after the challenge as compared to GG homozygotes (-12% (-16; -8.1) vs -6.45% (-11; -2.1), P < 0.001). GC carriers also had a marked reduction in other spirometric parameters. CONCLUSIONS: The GC variant of the TRPM8:c.750G > C (rs11562975) polymorphism is associated with CAH in patients with BA, which suggests a potential role of TRPM8 in CAH development.

An investigation of the association of the H558R polymorphism of the SCN5A gene with idiopathic cardiac conduction disorders.

AIM: The aim of this study was to investigate the predictive value of the rs1805124 polymorphism of the SCN5A gene with regard to idiopathic cardiac conduction disorders. RESULTS: The AG genotype frequency was significantly higher in patients with an atrioventricular block (61,2%±6,0%) compared with healthy control subjects (34,8%±2,3%), p<0.0001. The AG genotype frequencies among patients with only idiopathic complete right bundle-branch block (CRBBB) (54,2%±5,5%) and those with both CLBBB and LAH (50%±5,1) were significantly higher than in the control group (34,8%±2,3%), p<0.005. CONCLUSIONS: The AG genotype of the H558R (rs1805124) polymorphism of the SCN5A gene is a genetic predictor of idiopathic disorders of atrioventricular and intraventricular conduction.

Prognostic impact of the TP53 rs1625895 polymorphism in DLBCL patients.

This study aimed to clarify the association between the TP53 rs1625895 polymorphism and the efficiency of R-CHOP (rituximab, cyclophosphamide, doxorubicin, vincristine, prednisolone) therapy in 106 patients with diffuse large B cell lymphoma (DLBCL). All patients received six to eight courses of R-CHOP therapy as a first-line treatment. The rs1625895 polymorphism was genotyped by polymerase chain reaction with restriction fragment length polymorphism assay. The G/G genotype of the TP53 rs1625895 polymorphism was shown to be associated with a high probability of R-CHOP therapy failure in DLBCL patients according to the probability of remission as well as 5-year overall and relapse-free survivals.

Crosstalk between the FGFR2 and TP53 genes in breast cancer: data from an association study and epistatic interaction analysis.

To evaluate the potential for gene-gene interaction effects in sporadic breast cancer (BC) risk, we studied combinations of the fibroblast growth factor receptor 2 (FGFR2) rs1219648 and tumor protein 53 (TP53) rs1042522, rs1625895, and rs17878362 polymorphisms in BC patients (n=388) and healthy persons (n=275). In addition to a single-locus effect manifested by the association of FGFR2 rs1219648 and TP53 rs1042522 polymorphisms with high BC risk, depending on menopause status (0.001

CCL5/RANTES gene polymorphisms in Slavonic patients with myocardial infarction.

Coronary artery inflammation is a critical process in the pathogenesis of myocardial infarction (MI). The chemokine CCL5/RANTES (regulated upon activation, normal T cells expressed and secreted) is expressed in advanced atherosclerotic lesions. Functional polymorphisms of the RANTES gene can, therefore, be involved in the pathogenesis of coronary artery disease. We examined the association of polymorphisms in the RANTES gene with myocardial infarction in Slavonic populations of Czech and Russian origin. A total of 467 post-MI patients and 337 control subjects were genotyped for RANTES promoter G-403A (rs2107538) and intron 1.1 T/C (rs2280789) variants by PCR-SSP. Both RANTES genotypes and allele frequencies did not differ between case and control groups. Haplotype-based analysis also failed to reveal an association between MI and investigated markers. Strong linkage disequilibrium was detected between particular RANTES alleles. The data do not support an association between RANTES G-403A polymorphism and MI, as reported previously.

The macrophage migration inhibitory factor (MIF) gene polymorphism in Czech and Russian patients with myocardial infarction.

BACKGROUND: Macrophage migration inhibitory factor (MIF) is a cytokine implicated in early and advanced atherosclerosis. The aim of this study was to investigate whether polymorphism of MIF gene is associated with myocardial infarction (MI). METHODS: Single nucleotide polymorphism (SNP) in MIF gene (-173G/C, rs755622) was investigated in Czech (n=219) and Russian (n=240) MI patients and population control from the same geographical areas (Czech, n=137; Russian, n=174). Further, another SNP (rs1007888) located within the 3' flanking region of the MIF gene was investigated in Czech MI patients and control subjects. RESULTS: There were no significant differences in the distribution of MIF -173G/C genotypes, alleles or carriage rates between case and control groups in either populations. However, the GG genotype of the MIF SNP rs1007888 was associated with MI in Czech female patients (p=0.027). CONCLUSIONS: Taken together with previous reports, our study suggests that particular MIF gene polymorphisms may contribute to MI susceptibility in females.