Methylation of MMP2, TIMP2, MMP9 and TIMP1 in abdominal aortic aneurysm.

OBJECTIVES: Abdominal aortic aneurysm (AAA) and its complications are among the most serious cardiovascular diseases and its occurrence has risen sharply in recent years. The aim of this pilot study is to explore the relationship between the methylation of matrix metalloproteinases and tissue inhibitors of the metalloproteinases genes' promoter region, and abdominal aortic aneurysm (AAA) through the detection of the methylation status of MMP2, TIMP2, TIMP1, and MMP9 genes in peripheral blood. METHODS: The study included 43 males with verified AAA (case group) and 34 healthy males (control group). The methylation status of the genes' promoter region was detected by methylation-specific polymerase chain reaction (MS-PCR). RESULTS: In adominal aortic aneurysm patients, the methylation ratio of MMP2 gene was positive in 9.3 % (4 cases), 2.3 % (1 case) had methylated TIMP2 gene, 7.0 % (3 cases) had methylated TIMP1 gene, while the methylation ratio of MMP9 gene was positive in 93.0 % (40 cases). In the control group, MMP2 gene was found to be methylated in 5.9 % (2 cases), 5.9 % of cases had methylated TIMP2 and TIMP1 genes (2 cases), and MMP9 gene was found to be methylated in 91.2 % (31 cases). CONCLUSION: In our pilot study, we found no association between DNA methylation of gelatinases and their tissue inhibitors, and the development of an abdominal aortic aneurysm (Tab. 2, Fig. 1, Ref. 27).

The Role of Dysregulated MicroRNA Expression in Lung Cancer.

MicroRNAs (miRNAs) are a class of small single-stranded non-protein-coding RNAs that play important regulatory roles in many cellular processes including cell proliferation, differentiation, growth control, and apoptosis. They regulate gene expression on the posttranscriptional level by translational repression, mRNA cleavage, or mRNA degradation in various physiological and pathological processes. In addition, some miRNAs can function as oncogenes or tumor suppressors, so they can regulate several genes that play important roles in tumorigenesis. It was found that miRNAs are directly involved in many types of cancer, including lung cancer. Lung cancer is the leading cause of cancer mortality worldwide with a substantially low survival rate. In this work, we summarize recent findings related to miRNAs mechanisms of action and the role of their dysregulated expression in lung tumorigenesis. We describe the most important miRNAs involved in lung cancer development and targets of their activity. The understanding of the miRNA regulation in cancer may help better understand the molecular mechanisms of tumorigenesis and their importance in cancerous transformation.

Associations of CYP1A2 Polymorphisms with the Risk Haplotypes in Lung Cancer in the Slovak Population.

Phase I enzymes, including cytochrome P450, family 1, subfamily A, and polypeptide 2 (CYP1A2), are involved in the activation of carcinogens to reactive intermediates that are capable of binding covalently to DNA to form DNA adducts, potentially initiating the carcinogenic process. The aim of present study was to investigate the association of CYP1A2 gene polymorphisms and haplotypes with lung cancer risk. A case-control study was carried out on 105 lung cancer patients and 189 controls. To investigate three CYP1A2 polymorphisms: rs2472299, rs2470890, rs11072508 we used a high resolution melting analysis. We found significant allele associations (rs2470890 and rs2422299) with lung cancer risk. We searched for meaningful associations for all variants in the dominant, recessive, and additive genetic models. Genotype associations in the recessive model were of marginal significance for the same single nucleotide polymorphisms. A haplotype analysis included five variants with the frequency higher than 1 %. The haplotype "acc", present with the highest frequency, was associated with increased lung cancer risk (38.7 % vs. 31.5 %; OR 1.38; 95 %CI 0.95-2.01). On the contrary, rare haplotype "gtc" was significantly associated with decreased lung cancer risk in the Slovak population. In conclusion, the present study identified the risk alleles and haploid genotype associations of the CYP1A2 gene in lung cancer.

Polymorphisms of Selected DNA Repair Genes and Lung Cancer in Chromium Exposure.

Chromium is a well-known mutagen and carcinogen involved in lung cancer development. DNA repair genes play an important role in the elimination of genetic changes caused by chromium exposure. In the present study, we investigated the polymorphisms of the following DNA repair genes: XRCC3, participating in the homologous recombination repair, and hMLH1 and hMSH2, functioning in the mismatch repair. We focused on the risk the polymorphisms present in the development of lung cancer regarding the exposure to chromium. We analyzed 106 individuals; 45 patients exposed to chromium with diagnosed lung cancer and 61 healthy controls. Genotypes were determined by a PCR-RFLP method. We unravelled a potential for increased risk of lung cancer development in the hMLH1 (rs1800734) AA genotype in the recessive model. In conclusion, gene polymorphisms in the DNA repair genes underscores the risk of lung cancer development in chromium exposed individuals.

Mismatch repair gene polymorphisms and association with lung cancer development.

hMLH1 and hMSH2 are two of the main members of the mismatch repair (MMR) genes family. Polymorphism of MMR genes is associated with a risk of developing sporadic and hereditary tumors. In the present case-control study, we investigated the promoter polymorphisms of selected mismatch repair genes: hMLH1 (rs1800734) and hMSH2 (rs2303425), and the risk they present regarding the development of lung cancer in the Slovak population. The study included 422 lung cancer cases, 511 controls for hMLH1 gene and 486 controls for hMSH2 gene. Polymorphism was investigated by a PCR-RFLP method. The risk of cancer development was evaluated in both dominant and recessive genetic models. The evaluation of rs1800734 polymorphism in patients in the dominant model showed a significantly decreased risk of lung cancer in the presence of at least one variant allele A (genotype GA and AA) (OR=1.40; 95% CI=1.08-1.82; p=0.01). These findings were equally strong expressed in women (OR=2.00; 95% CI=1.23-3.25; p=0.006). The results for rs2303425 polymorphism revealed an increased risk of lung cancer for variant genotype CC (OR=2.28; 95% CI=1.12-4.63; p=0.024) in the recessive model. A combination of rs1800734 and rs2303425 polymorphisms was shown to be risky for genotype GGCC; OR=3.08; 95% CI=1.09-8.72; p=0.03. The risk appeared even greater in female gender; (OR=11.56; 95% CI=1.33-100.36, 1.26-94.66; p=0.005. We conclude that the genotype of mismatch repair genes underscores the risk of lung cancer development in the Slovak population.

Polymorphisms of DNA repair genes and lung cancer in chromium exposure.

Chromium is a well known carcinogen involved in the lung cancer development. Polymorphism of some of the DNA repair genes may be associated with elevated risk of cancerous transformation. In the present study, we investigated the polymorphisms of the following selected members of the base and nucleotide excision repair genes: XPC (Lys939Gln), XPD (Lys751Gln), XRCC1(Arg399Gln), and hOGG1(Ser326Ser), and the risk they present toward the development of lung cancer, with emphasis on the effect of chromium exposure. We analyzed 119 individuals; 50 patients exposed to chromium with diagnosed lung cancer and 69 healthy controls. Genotypes were determined by a PCR-RFLP method. We found a significantly increased risk of lung cancer development in XPD genotype Lys/Gln (OR=1.94; 95% CI=1.10-3.43; p=0.015) and in the gene combinations: XPD Lys/Gln+XPC Lys/Gln (OR=6.5; 95% CI=1.53-27.49; p=0.009) and XPD Lys/Gln+XPC Gln/Gln(OR=5.2; 95% CI=1.07-25.32; p=0.04). In conclusion, gene polymorphisms in the DNA repair genes may underscore the risk of lung cancer development in the chromium-exposed individuals.

DNA repair genes polymorphism and lung cancer risk with the emphasis to sex differences.

Polymorphisms in nucleotide and base excision repair genes are associated with the variability in the risk of developing lung cancer. In the present study, we investigated the polymorphisms of following selected DNA repair genes: XPC (Lys939Gln), XPD (Lys751Gln), hOGG1 (Ser326Cys) and XRCC1 (Arg399Gln), and the risks they present towards the development of lung cancer with the emphasis to gender differences within the Slovak population. We analyzed 761 individuals comprising 382 patients with diagnosed lung cancer and 379 healthy controls. Genotypes were determined by polymerase chain reaction/restriction fragment length polymorphism method. We found out statistically significant increased risk for lung cancer development between genders. Female carrying XPC Gln/Gln, XPC Lys/Gln+Gln/Gln and XRCC1 Arg/Gln, XRCC1 Arg/Gln+Gln/Gln genotypes had significantly increased risk of lung cancer corresponding to OR = 2.06; p = 0.04, OR = 1.66; p = 0.04 and OR = 1.62; p = 0.04, OR = 1.69; p = 0.02 respectively. In total, significantly increased risk of developing lung cancer was found in the following combinations of genotypes: XPD Lys/Gln+XPC Lys/Lys (OR = 1.62; p = 0.04), XRCC1 Gln/Gln+hOGG1 Ser/Ser (OR = 2.14; p = 0.02). After stratification for genders, the following combinations of genotype were found to be significant in male: XPD Lys/Gln+XPC Lys/Lys (OR = 1.87; p = 0.03), XRCC1 Arg/Gln+XPC Lys/Lys (OR = 4.52; p = 0.0007), XRCC1 Arg/Gln+XPC Lys/Gln (OR = 5.44; p < 0.0001). In female, different combinations of the following genotypes were found to be significant: XRCC1 Arg/Gln+hOGG1 Ser/Ser (OR = 1.98; p = 0.04), XRCC1 Gln/Gln+hOGG1 Ser/Ser (OR = 3.75; p = 0.02), XRCC1 Arg/Gln+XPC Lys/Gln (OR = 2.40; p = 0.04), XRCC1 Arg/Gln+XPC Gln/Gln (OR = 3.03; p = 0.04). We found out decreased cancer risk in genotype combinations between female patients and healthy controls: XPD Lys/Lys+XPC Lys/Gln (OR = 0.45; p = 0.02), XPD Lys/Gln+XPC Lys/Lys (OR = 0.32; p = 0.005), XPD Lys/Gln+XPC Lys/Gln (OR = 0.48; p = 0.02). Our results did not show any difference between pooled smokers and non-smokers in observed gene polymorphisms in the association to the lung cancer risk. However, gender stratification indicated the possible effect of heterozygous constitution of hOGG1 gene (Ser/Cys) on lung cancer risk in female non-smokers (OR = 0.20; p = 0.01) and heterozygous constitution of XPC gene (Lys/Gln) in male smokers (OR = 2.70; p = 0.01).

Expression of Ki-67, Bcl-2, Survivin and p53 Proteins in Patients with Pulmonary Carcinoma.

Apoptosis is the fundamental process necessary for eliminating damaged or mutated cells. Alterations in the apoptotic pathway appear to be key events in cancer development and progression. Bcl-2 is the key member of the Bcl-2 family of apoptosis regulator proteins with anti-apoptotic effects. Survivin acts as an inhibitor of apoptosis as well and has been implicated in both inhibition of apoptosis and mitosis regulation. p53 is one of the tumor suppressor proteins, prevents tumor formation through cell cycle blocking and eliminates damaged cells via the activation of apoptosis. The Ki-67 protein is a cellular marker for proliferation. To investigate the possible interactions of the aforementioned proteins, we examined their expression in 76 patients with diagnosed lung cancer using immunohistochemical visualisation. Ki-67 protein was expressed in the cancer cells of all patients with small cell lung cancer (SCLC). We found a negative correlation between survivin and p53 expression. A decreased intensity of survivin expression and fewer cells positive for survivin (66.7%) in SCLC in comparison with other lung cancer types (98.0%) was detected. Reversely, expression of Bcl-2 was found in more than 90% of cases with SCLC. We hypothesize that high expression and intensity of Bcl-2 protein could be a factor behind a bad prognosis in SCLC.

Lung adenocarcinoma and squamous cell carcinoma in association with genetic polymorphisms of GSTs in Slovak population.

Slovak Republic belongs to the countries with high incidence of lung cancer. Gene polymorphisms of the glutathione S-transferases (GSTs) may play a role in individual lung cancer susceptibility. In presented case-control study we investigate the incidence of polymorphism of GSTT1, GSTM1, GSTP1 genes and their combinations as possible predictive factors for identification of individuals with increased risk of formation and development of adenocarcinoma (AC) and squamous cell carcinoma (SCC) of lung in Slovak population. The study was conducted on 520 individuals consisting of 118 patients with adenocarcinoma, 112 patients with squamous cell carcinoma and 290 control individuals. GSTT1, GSTM1, GSTP1 gene polymorphisms were assayed by standard PCR and PCR-RFLP technique. The results of this study indicate that the GSTT1null-genotype and combination GSTT1 null and Ile/Val or Val/Val are associated with increased risk of lung adenocarcinoma. A significant association with 2.13 - fold increased risk was observed between lung adenocarcinoma and GSTT1 null genotype (95% CI = 1.29 - 3.51; p= 0.004). Also it was proved 2.83 times statistically higher risk for development of this histological type of lung cancer (95% CI = 1.34 - 6.01; P= 0.005) in combination of GSTT1null and Ile/Val or Val/Val genotypes. GSTT1, GSTM1, GSTP1 polymorphism did not show any significant association with SCC. Our study suggests that genetic make-up in metabolizing genes may increase susceptibility towards lung cancer development.

Microsomal epoxide hydrolase polymorphisms, cigarette smoking and prostate cancer risk in the Slovak population.

Polymorphisms in tobacco carcinogen metabolizing enzymes may generate interindividual variations towards the risk of developing prostate cancer. One of these enzymes is microsomal epoxide hydrolase (EPHX1) which metabolizes polycyclic aromatic hydrocarbons, or PAH, carcinogens found in cigarette smoke. The activity of this enzyme is affected by two polymorphisms, a substitution of Tyr113 by His in exon 3 and a substitution of His139 by Arg in exon 4. The aim of this study was to use a population-based case-control study to investigate whether or not such genetic polymorphisms in EPHX1 gene can modify the relationship between smoking status and the risk of developing prostate cancer. We used restriction fragment length polymorphism, or PCR-RFLP to determine EPHX1 genotypes in subjects comprising 194 patients with histologically verified prostate cancer and 305 healthy individuals as control. We found no overall association between prostate cancer risk and functional polymorphisms of EPHX1 gene in exon 3 and exon 4. We further analysed the association between the EPHX1 genotypes and smoking. Smokers carrying the exon 3 Tyr/Tyr and Tyr/His genotypes were at no significant risk compared to non-smokers with the "rapid" Tyr/Tyr genotype. By contrast, a significant interaction of smoking and the exon 4 polymorphism was present.

[Five year-results of chemoresistance testing in cancer patients]. / Nase pät'rocné výsledky in vitro testovania chemorezistencie u onkologických pacientov.

BACKGROUNDS: Translational medicine is a medical field encompassing basic research and development of new diagnostic and therapeutic strategies for clinical practice. The present scientific paper focuses on our previous experience in the field of chemoresistance testing in patients with oncological diseases. MATERIAL AND METHODS: Since 2005, we sampled 71 patients with a leukaemia (AML, ALL and CML) and 92 patients with a solid tumour (lung and gastrointestinal tract cancer). Malignant cell in vitro drug resistance testing was carried out using cytotoxic methyl-thiazol tetrazolium (MTT) assay. RESULTS: Based on the LC50 (lethal concentration of a drug killing 50% of cell population), we found that patients with acute myeloblastic leukaemia exhibit a greater degree of resistance than patients with acute lymphoblastic leukaemia. In patients with bronchogenic carcinomas, primary resistance to cisplatin was identified in 28% of tested samples, paclitaxel 36%, vincristine 50%, etoposide 56%, vinorelbine 57%, topotecan 62%, gemcitabine 77% and dacarbazine 86%. CONCLUSION: In vitro tests with gastrointestinal tract cancers also suggested high effectiveness of cisplatin (with the exception of gastric carcinoma) that was comparable with 5-fluorouracil. Even though the MTT assay has some limitations (insufficient number of vital cells, possible contamination by non-malignant cells, etc.), this in vitro method proved very effective in testing malignant cell resistance to clinically used cytostatics.

Lung cancer incidence and survival in chromium exposed individuals with respect to expression of anti-apoptotic protein survivin and tumor suppressor P53 protein.

OBJECTIVE: Workers chronically exposed to hexavalent chromium have elevated risk of lung cancer. Our study investigates the incidence of lung cancer types, age at onset of the disease, and survival time among chromium exposed workers with respect to the expression of anti-apoptotic p53 and pro-apoptotic survivin proteins. MATERIAL AND METHODS: 67 chromium exposed workers and 104 male controls diagnosed with lung cancer were analyzed. The mean exposure time among workers was 16.7 ±10.0(SD) years (range 1- 41 years). To investigate the possible regulation of survivin by p53 we examined the expression of both proteins using immohistochemical visualization. RESULTS: Chromium exposure significantly decreases the age of onset of the disease by 3.5 years (62.2 ±9.1 in the exposed group vs. 65.7 ±10.5 years in controls; P=0.018). Small cell lung carcinoma (SCLC) amounted for 25.4% of all cases in chromium exposed workers and for 16.3% in non-exposed individuals. The mean survival time in the exposed group was 9.0 ±12.7 vs. 12.1 ±21.9 months in controls, but this difference was not significant. Survivin was predominantly expressed in both cell nucleus and cytoplasm, whereas p53 was expressed in the nucleus. There was a negative correlation between survivin and p53 expression. A decreased intensity of expression and fewer cells positive for survivin was detected in SCLC compared with other types of lung cancer. p53 was expressed in 94.1% and survivin in 79.6% of the samples analyzed. CONCLUSION: The study calls attention to decreased expression of survivin, as opposed to p53, in small cell lung carcinoma.

Polymorphisms of biotransforming enzymes (GSTs) and their association with colorectal cancer in the Slovak population.

The aim of present study was to summarize the results of a case-control study focused on genetic polymorphisms of selected Phase II metabolizing enzymes (GSTM1, T1, P1) and to investigate the association of these polymorphisms with the colorectal cancer risk among the Slovak population. A case-control study with 183 colorectal cancer cases and 422 controls was conducted. DNA was extracted from peripheral blood leukocytes, and the polymorphisms of GSTM1, GSTT1 and GSTP1 enzymes were determined by PCR-based methods. Association between specific genotypes and the development of colorectal cancer were examined using logistic regression analysis to calculate odds ratios (OR) and 95% confidence intervals (CI). The GSTP1 val/val genotype (OR=2.1, 95%CI: 1.1 - 4.0, chi2 = 0.28 and P = 0.0025) was associated with an elevated risk. The statistically significant correlation was found also for the combined genotypes of GSTM1 null and GSTP1 valine homozygosity (OR = 2.7, 95% CI: 1.1-6.1, chi2 = 4.5 and P = 0.03). The genotype of certain metabolising enzymes affects the risk for colorectal cancer. This effect is also important when certain allelic combinations are studied. In the near future, individual risk assessment may be reached by further increasing the number of studies of polymorphisms, combining them with the traditional epidemiological risk factor.

Expression of anti-apoptotic protein survivin and tumor suppressor p53 protein in patients with pulmonary carcinoma.

BACKGROUND: Survivin is one of the inhibitors of the apoptosis gene family that has been implicated in both inhibition of apoptosis and mitosis regulation. p53 is one of the tumor suppressor genes; prevents tumor formation through cell cycle blocking and eliminates damaged cells via activation of apoptosis. OBJECTIVE: To investigate the possible regulation of survivin by p53, we examined the expression of both proteins in 67 patients with diagnosed lung cancer using immunohistochemical visualization. RESULTS: Survivin was predominantly expressed in both nucleus and cytoplasm, whereas p53 was expressed in the nucleus. There was a negative correlation between survivin and p53 expression. A decreased intensity of expression and fewer cells positive for survivin in small cell lung cancer in comparison with other lung cancer types were detected. There was no significant difference in the intensity of expression and the number of cells positive for p53 between small cell and non-small cell lung cancer types. CONCLUSION: The present study suggests that survivin expression, as opposed to that of p53, is decreased in small cell lung cancer, which may differentiate this cancer from other lung cancer types other types.

Gene polymorphisms of biotransforming enzymes (GSTs) and their association with lung cancer in the Slovakian population.

OBJECTIVE: The aim of present study was to present the results of a case-control study focused on genetic polymorphisms of selected Phase II metabolizing enzymes (GSTM1, T1, and P1) and to investigate the association of these polymorphisms with lung cancer risk in the Slovakian population. MATERIAL AND METHODS: The study encompassed 160 lung cancer cases and 220 controls. DNA was extracted from peripheral blood leukocytes, and the polymorphisms of GSTM1, GSTT1 and GSTP1 enzymes were determined by PCR-based methods. We determined the genotype distribution of all these genes and their combinations. The association between specific genotypes and the development of lung cancer were examined using logistic regression analysis to calculate odds ratios (OR) and 95% confidence intervals (CI). RESULTS: We found that the GSTM1 null genotype (OR=1.6; 95% CI=1.03-2.4; chi(2)=4.08, and P=0.04) was associated with elevated risk. A significant correlation also was found for the combined genotypes of GSTM1 null and GSTP1 Ile/Val and Val/Val (OR=2.01; 95% CI=1.1-6.1; chi(2)=3.6, and P=0.02) and GSTM1 null and GSTT1 positive (OR=2.00; 95% CI=1.2-3.2; chi(2)=7.3, and P=0.006). CONCLUSIONS: We conclude that the genotype of metabolizing enzymes and allelic combinations underscore the risk for lung cancer. Individual risk assessment may be further improved by increasing the number of polymorphisms studied and combining them with the traditional epidemiological risk factor.

Relationship between antioxidant potential and oxidative damage to lipids, proteins and DNA in aged rats.

Oxidative stress has been implicated to play a major role in aging and age-related diseases. In the present study, we investigated the effects of aging on the total antioxidant capacity, uric acid, lipid peroxidation, total sulfhydryl group content and damage to DNA in adult (6 months), old (15 months) and senescent (26 months) male Wistar rats. The antioxidant capacity, determined by phycoerythrin-based TRAP method (total peroxyl radical-trapping potential) was significantly decreased in the plasma and myocardium of old and senescent rats, whereas plasma level of uric acid was elevated in 26-month-old rats. Age-related decline in plasma and heart antioxidant capacity was accompanied by a significant loss in total sulfhydryl group content, increased lipid peroxidation and higher DNA damage in lymphocytes. Correlations between TRAP and oxidative damage to lipids, proteins and DNA suggest that the decline in antioxidant status may play an important role in age-related accumulation of cell damage caused by reactive oxygen species.