Gut Microbiome and colorectal cancer: discovery of bacterial changes with metagenomics application in Turkish population.

BACKGROUND: Colorectal cancer (CRC) is the 3rd most common cancer in the world and colonic carcinogenesis is a multifactorial disease that involves environmental and genetic factors. Gut microbiota plays a critical role in the regulation of intestinal homeostasis. Increasing evidence shows that the gut microbiome plays a role in CRC development and may be a biomarker for early diagnosis. OBJECTIVE: This study aimed to determine the clinical prognostic significance of gut microbiota in CRC patients in the Turkish population by metagenomic analysis and to determine the microbial composition in tumor tissue biopsy samples. METHODS: Tissue biopsies were taken from the participants with sterile forceps during colonoscopy and stored at -80 °C. Then, DNA isolation was performed from the tissue samples and the V3-V4 region of the 16 S rRNA gene was sequenced on the Illumina MiSeq platform. Quality control of the obtained sequence data was performed. Operational taxonomic units (OTUs) were classified according to the Greengenes database. Alpha diversity (Shannon index) and beta diversity (Bray-Curtis distance) analyses were performed. The most common bacterial species in CRC patients and healthy controls were determined and whether there were statistically significant differences between the groups was tested. RESULTS: A total of 40 individuals, 13 CRC patients and 20 healthy control individuals were included in our metagenomic study. The mean age of the patients was 64.83 and BMI was 25.85. In CRC patients, the level of Bacteroidetes at the phylum taxonomy was significantly increased (p = 0.04), the level of Clostridia at the class taxonomy was increased (p = 0.23), and the level of Enterococcus at the genus taxonomy was significantly increased (p = 0.01). When CRC patients were compared with the control group, significant increases were detected in the species of Gemmiger formicilis (p = 0.15), Prevotella copri (p = 0.02) and Ruminococcus bromii (p = 0.001) at the species taxonomy. CONCLUSIONS: Metagenomic analysis of intestinal microbiota composition in CRC patients provides important data for determining the treatment options for these patients. The results of this study suggest that it may be beneficial in terms of early diagnosis, poor prognosis and survival rates in CRC patients. In addition, this metagenomic study is the first study on the colon microbiome associated with CRC mucosa in the Turkish population.

Artificial intelligence model with deep learning in nonalcoholic fatty liver disease diagnosis: genetic based artificial neural networks.

Nonalcoholic fatty liver disease (NAFLD) is one of the most common causes of chronic liver disease in the world. The NAFLD spectrum includes simple steatosis, steatohepatitis, fibrosis, cirrhosis, and hepatocellular carcinoma (HCC). Genetic, nutritional factors, obesity, insulin resistance, gut microbiota are among the risk factors for NAFLD. The genetic variant Patatin-like phospholipase domain-containing protein 3 (PNPLA3) plays an important role in the development of a number of liver diseases ranging from steatosis, chronic hepatitis, cirrhosis and HCC. Due to the increase in the prevalence of NAFLD, new models are being developed with machine learning, deep learning, artificial neural network (ANN) algorithms in the field of artificial intelligence (AI) to determine low-cost, noninvasive diagnostic methods. Models developed with ANN from AI modules are important in order to examine biochemical and genomic information in detail in the diagnosis of NAFLD. The aim of this study is to develop a simple ANN model using biochemical and genotypic parameters in the diagnosis of NAFLD. A total of 300 patients followed up with the diagnosis of NAFLD and 100 controls were included in the study. The data set was divided into two as training and test set. Genotyping of PNPLA3 (CC, CG, GG) as genomic analysis was performed with real time PCR device. The algorithm used for the diagnosis of NAFLD was designed using age, body mass index (BMI), mean platelet volume (MPV), insulin resistance (IR), alanine aminotransferase (ALT), genotype PNPLA3 (CC, CG, GG) parameters. MLP Classifier algorithm from ANN was used in the development of the model. ANN algorithms are used in python programming language. Statistical analyzes were made in SPSS program. Percent accuracy, area under the ROC curve, confusion matrix, Positive (PPV) and Negative Predicted Value (NPV) values, precision, recall, and f1-score results were determined. The accuracy percentage was determined as 0.979 in the train set and 0.970 in the test set. The Log Loss value was set to 0.09. The developed neural network achieved an accuracy percentage of 97.0% during testing, with an area under the ROC curve value of 0.95. We think that the ANN model developed with genomic and biochemical parameters can be used as a cost-effective, noninvasive new predictive diagnostic model in clinical practice in the diagnosis of NAFLD.

Evaluation of possible role of the hTERT gene rs2853669 polymorphism in the development of colorectal cancer as a genetic risk factor.

Colorectal cancer (CRC) is the second deadliest malignancy. Human telomerase reverse transcriptase (hTERT) gene has been identified as one of the potential cancer susceptibility genes. We evaluated the relationship between the risk of CRC and CRC's clinicopathological features of the hTERT rs2853669 (A > G/T > C, by the chain direction) polymorphism in Turkish population. The rs2853669 polymorphism was investigated with the LightCycler 96 device in 100 CRC patients and 327 controls. We found that the rs2853669 polymorphism AG/GG genotypes in genetic models reduced the risk of CRC. However, there was no significant relationship between rs2853669 polymorphism and clinicopathological features of CRC in studied population. The results of this study showed that the risk of colorectal cancer is significantly reduced in the individuals having the G (C) allele. Our recommendation is to analyze the hTERT gene expression by studying the hTERT promoter mutations with this polymorphism in colorectal cancer.

Metagenomic identification of gut microbiota distribution on the colonic mucosal biopsy samples in patients with non-alcoholic fatty liver disease.

BACKGROUND: Non-alcoholic fatty liver disease (NAFLD) is known to be the most common liver disease in the world, and there are currently no approved pharmacological treatments to prevent or treat this condition. In addition to being associated with an increased risk of hepatocellular carcinoma and cirrhosis, NAFLD has now become the leading cause of liver failure-associated transplantation. The 16S rRNA gene which conserved regions can serve as universal primer binding sites for PCR amplification of gene fragments, while hypervariable regions contain significant sequence diversity useful for prokaryotic identification purposes. 16S rRNA gene sequences can be use by researchers to identify prokaryotic taxonomy found in clinical samples. As a result of increasing microbiota studies with developing technological developments, the role of intestinal microbiota in the pathogenesis of NAFLD is revealed in an important way. In this study, it was aimed to determine the clinical prognostic importance of gut microbiota in the pathogenesis of NAFLD and to determine the microbial composition with intestinal mucosal biopsy samples in NAFLD patients. MATERIAL AND METHOD: We included 20 patients diagnosed with NAFLD as a result of liver function tests, histological, ultrasonographic, biopsy evidence and 20 normal control groups created under exclusion criteria in this study. The healthy control group of the same age and gender as the patients were determined to be equal, and the age, gender, BMI, insulin resistance, AST, ALT levels of the individuals were recorded for analysis. Intestinal mucosal biopsy samples were taken from the individuals included in the study under sterile conditions. Microbial results were obtained as a result of 16S rRNA amplicon metagenomic processes. The region of approximately 1500 bp covering the V1-V9 region of the 16S rRNA gene was targeted to detect microbial diversity. The amplified regions were sequenced using next-generation sequencing. Operational Taxonomic Unit (OTU) value was obtained with bioinformatics software with the obtained sequence data. The analysis of the recorded parameters was done with the SPSS.19 statistical program. RESULTS: In the designed study, 16 phyla, 28 class, 56 order, 128 family, 415 genera, 1041 species microorganisms were analyzed taxonomically in a total of 40 individuals. In our study, Intestinal microbial diversity is lower in NAFLD patients compared to control group individuals. In addition, gram-negative bacteria were found to be more dominant in NAFLD patients. As a phylum, Proteobacteria increased in NAFLD group, Bacteroidetes and Actinobacteria in control group, while Firmicutes had equal distribution in both groups. BMI OR = 6.37, 95 %CI (0.39-0.40) p value was 0.001 in laboratory data, whereas Proteobacteria OR = 1.754, 95% CI (0.901-3.416), p value 0.05 in microbial profile. CONCLUSION: The 16S rRNA metagenomic study of intestinal microbiota using colonic mucosal biopsy samples in NAFLD disease was the first study in the Turkish population, and important data were obtained for other studies. In the data obtained, we think Proteobacteria, Ruminococcaceae, Escherichia coli and Bacilli are very important in both diagnostic and treatment options as a microbial profile in NAFLD.

Hepatitis B Reactivation in Patients Treated with Direct-Acting Antivirals for Hepatitis C.

INTRODUCTION: There is limited research about HBV reactivation (HBVr) due to direct-acting antivirals (DAA) for HCV and most are limited by short duration of follow-up, small sample size, and absence of baseline HBV DNA. We aimed to determine the incidence and clinical course of HBVr in HBsAg and/or anti-HBcIgG positive patients treated with DAA for HCV. METHODS: Seven centers retrospectively analyzed their database on HCV patients treated with DAA between 2015 and 2019. Patients with HBV coinfection or resolved HBV infection were enrolled. Serum transaminases, HBsAg, HBeAg, and HBV DNA were followed every 4 weeks during DAA treatment and every 12 weeks 1 year after treatment. Entecavir or tenofovir disoproxil fumarate was started in case of HBVr. The development of HBVr, HBV flare, liver failure, and mortality were determined. RESULTS: 852 patients received DAA treatment for HCV. Among them, 35 (4.1%) had HBV coinfection and 246 (28.9%) had resolved HBV infection. 257 patients (53.3% male, mean age: 63 ± 9) constituted the study group (29 with coinfection and 228 with resolved infection). Three patients with coinfection were HBV DNA positive. HBVr developed in 10 (34.5%) HBsAg positive patients, either during (n = 3) or 12-48 weeks after finishing DAA treatment. HBV flare and acute liver failure developed in 1 patient (3.4%), each. Two patients with resolved infection developed HBVr (0.87%) and one (0.44%) had HBV flare. Overall, none of the patients died or underwent liver transplantation due to HBVr. CONCLUSION: Patients with HBV/HCV coinfection have a high risk of HBVr after DAA treatment and should receive antiviral prophylaxis. Patients with resolved infection have a low risk of HBVr and can be monitored by serial ALT measurements.

Paraneoplastic syndrome frequency and prognostic effect in hepatocellular carcinoma patients.

OBJECTIVE: Symptomatic hepatocellular carcinoma (HCC) patients may generally display constitutional symptoms such as abdominal pain, weight loss, anorexia and localized mass, or atypical clinical features of paraneoplastic syndrome (PNS) such as hypercholesterolemia, hypercalcemia, hypoglycemia, erythrocytosis and thrombocytosis. The most common PNS in HCC is hypercholesterolemia, hypercalcemia, hypoglycemia and erythrocytosis. The aim of this study isto evaluate the relationship of PNS in HCC patients. MATERIAL AND METHOD: In this study, the data of 534 patients who were followed up with the diagnosis of HCC between January 2010 and December 2020 in the Gastroenterology clinic were evaluated retrospectively. Clinical data, age, gender, complete blood count of patients with and without PNS, liver biochemistry, alpha-fetoprotein (AFP) level, hepatitis B surface antigen, anti-hepatitis B virus, Child-Pugh score, model for end-stage liver disease score, tumor volume, portal vein thrombosis, liver biopsy histology and radiologic images were taken from the hospital data system and analyzed. RESULTS: Out of the 534 HCC patients, 120 (22.3%) were PNS-positive patients. There was a significant difference between the ages of PNS-positive and PNS-negative patients, and PNS-positive patients were older (64.60±12.97) (P=0.02). PNS-positive HCC was determined as hypoglycemia 5.8%, hypercalcemia 6.3%, erythrocytosis 3.9%, hypercholesterolemia 2.4% and thrombocytosis 3.9%. AFP level (22908 ± 60 ng/ml) and tumor diameter (>10 cm) were higher in the PNS-positive group. Multivariate analysis showed that stage C according to Child-Pugh score and tumor diameter >10 cm were independent predictors of poor prognosis, whereas PNS erythrocytosis and thrombocytosis were independent predictors of better prognosis. CONCLUSION: In PNS-positive HCC patients, hypoglycemia and hypercalcemia were associated with poor prognosis according to Child-Pugh score, whereas erythrocytosis and thrombocytosis were associated with good prognosis.

Prognostic Value of International Normalized Ratio-to-Albumin Ratio and Ferritin Level in Chronic Liver Patients with Hepatocellular Carcinoma.

OBJECTIVE: Hepatocellular carcinoma (HCC) is the sixth among the most common cancers and the fourth among cancer-related causes of death in the world. In the evaluation of liver function in HCC patients, parameters such as albumin-bilirubin, prothrombin time-international normalized ratio (PT-INR) to albumin ratio (PTAR) are used among new methods other than Child-Pugh and MELD scores. Biomarkers are widely used in clinical practice in cases such as diagnosing various diseases, evaluating treatment response and predicting prognosis. We aimed to evaluate the prognostic role of serum ferritin and INR/albumin ratio in patients with chronic liver disease who develop HCC. METHODS: This retrospective study included 534 patients who were followed up with the diagnosis of HCC between 2009 and 2020. The patients with HCC etiology were evaluated in 3 groups (chronic hepatitis B group, chronic hepatitis C group, and other group). When comparing serum ferritin level and prothromin time-international normalized ratio to albumin ratio with Child Pugh score (CTP) in chronic liver patients with HCC, liver functional reserve and its role in predicting prognosis were investigated. RESULTS: The serum ferritin level was 226 ± 334 in the CTP A group, 239 ± 302 in the CTP B group, and 678 ± 966 in the CTP C group, and the p value was 0.001. The PTAR CTP group was 0.35 ± 0.10, the CTP B group was 0.50 ± 0.26, the CTP C group was 1.18 ± 6.01, and the p value was 0.001. Multivariant analysis results showed that ferritin hazard ratio is 1.00, 95% CI 0.99-1.00, and p value was 0.09, and PTAR hazard ratio is 1.38, 95% CI 2.37-8.00, and p value was 0.49. The etiological distribution of HCC was determined as HBV (61.6%), HCV (19.9%), and other etiologies (18.5%). Significant values were determined for age, gender, glucose, GGT, T. cholesterol, and tumor diameter parameters according to etiological distribution. CONCLUSIONS: Serum ferritin level and PTAR score increased in proportion to the severity of liver disease and were associated with poor prognosis. We think that high serum ferritin and PTAR score is a prognostic biomarker in predicting the synthesis function of the liver and mortality in critically ill patients with cirrhosis.

Assessment of the Relationship between Ulcerative Colitis and Forkhead Box P3 Polymorphisms.

BACKGROUND/AIMS: Ulcerative colitis (UC) is a chronic disease that does not have a definitive treatment and causes repetitive inflammation of the colon and impaired quality of life. The FOXP3 gene codes FOXP3 protein responsible for development and function of regulatory T (Treg) cells. The rs2232365 A/G and the rs3761548 A/C polymorphisms of FOXP3 gene were indicated to be associated with inflammation-related diseases such as ulcerative colitis. The effectiveness of Treg cells, which act as immune-suppressors in the control of inflammation, can be affected by these polymorphisms. The aim of the present study was to evaluate the association between these polymorphisms with ulcerative colitis. MATERIALS AND METHODS: The current study researched the FOXP3 gene polymorphisms in 146 patients with UC and in 292 healthy individuals by a real-time polymerase chain reaction (RT-PCR). RESULTS: The patients with rs2232365 G allele had a 1.44-fold higher UC risk than patients carrying other allele (P=0.013), and had significantly a 2.56-fold higher risk for extent of UC (P=0.001). Contrary, rs3761548 polymorphism didn't reach statistically significant in any analysis. CONCLUSION: This is the first study to reveal the relationship of the rs2232365 and the rs3761548 polymorphisms with ulcerative colitis in Caucasian population. The rs2232365 has an important effect on the risk of UC. The current study suggests that these polymorphisms should be explored together with the FOXP3 expression and FOXP3+ Treg cell count in blood and colon tissue of UC patients to clarify the exact effect of FOXP3 polymorphisms on UC risk.

Serum levels of inflammatory markers CRP, ESR and albumin in relation to survival for patients with hepatocellular carcinoma.

INTRODUCTION: Hepatocellular carcinoma is associated with several chronic inflammatory conditions. It is increasingly understood that the inflammation may be part of the carcinogenic process and prognostically important. OBJECTIVE: To evaluate the serum levels of three inflammation markers in relation to survival in HCC patients. METHODS: We retrospectively examined the serum levels of CRP, albumin and ESR, both singly and in combination, in relation to patient survival. RESULTS: Survival worsened with increase in CRP or ESR or decrease in albumin levels. Combinations of CRP plus albumin or CRP plus ESR were associated with an even greater range of survival (3-fold), together with significant differences in maximum tumor diameter (PVT) and percent of patients with portal vein thrombosis (PVT). The triplet of CRP plus albumin plus ESR was associated with a sevenfold difference in survival, comparing low vs high parameter levels. These significant differences were found in patients with small or large tumors. CONCLUSIONS: Combinations of CRP with albumin or ESR or all three parameters together significantly related to differences in survival and to differences in MTD and percent PVT, in patients with both small and large size HCCs.

Association between chronic hepatitis B virus infection and HLA-DP gene polymorphisms in the Turkish population.

AIM: Hepatitis B virus (HBV) affects approximately 360 million people worldwide. 10-15% of patients with chronic HBV develop liver cirrhosis (LC), liver failure and hepatocellular carcinoma (HCC). Chronic HBV infection or HBV clearance is influenced by both viral and host factors. In genome-wide association studies (GWAS), the human leukocyte antigen (HLA) gene polymorphisms rs3077 and rs9277535 were identified to be associated with chronic hepatitis B. HLA genes have been linked to immune response to infectious agents. Genetic variants in HLA genes influence HLA mRNA expression which might also affect antigen presentation. We evaluated the association between HLA gene polymorphisms and the risk for persistent HBV infection. METHODS: In the current study, HLA gene polymorphisms were investigated in a case-control study of 294 chronic HBV patients and 234 persons with HBV natural clearance by using a real-time polymerase chain reaction (RT-PCR). RESULTS: The results showed that rs9277535 allele frequency is associated with HBV infection in the Turkish subjects examined (P=0.048). However, no association was found for rs3077. Additionally, the AG haplotype block showed a protective effect against the risk of persistent HBV infection (for the rs3077A/rs9277535G, OR=0.52; 95% 0.34-0.80, P=0.003). CONCLUSIONS: Our results, for the first time, demonstrate that HLA-DPB1 gene rs9277535A allele has a major effect on the risk of persistent HBV infection. We suggest that further independent studies are necessary to clarify the association of these polymorphisms with persistence or natural clearance of HBV infection in Caucasian populations.

Polymorphisms in human telomerase reverse transcriptase (hTERT) gene and susceptibility to gastric cancer in a Turkish population: Hospital-based case-control study.

Erosion of telomeres, tandem nucleotide repeats (TTAGGG)n that cap the end of eukaryotic chromosomes, has been related with carcinogenesis. The human telomerase reverse transcriptase (hTERT) gene is encoded the rate-limiting catalytic subunit of the telomerase complexes, which is essential for the protection of telomeric DNA length and chromosomal stability. The purpose of this study was to examine the effect of four functional single nucleotide polymorphisms (SNPs) of hTERT (rs2736109 G>A, rs2735940 T>C, rs2853669 A>G and rs2736100 T>G) on susceptibility to gastric cancer (GC) in Turkish population. The genotype frequency of hTERT rs2736109 G>A, rs2735940 T>C, rs2853669 A>G and rs2736100 T>G polymorphisms were determined by using a polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and TaqMan methods in 104 subjects with GC and 209 healthy control subjects. We found that hTERT rs2736109 G>A (AA+AG vs. GG OR=1.68 95% CI=1.01-2.81, P=0.04), rs2735940 T>C (CC vs. CT+TT: OR=2.53 95% CI=1.01-6.13, P=0.03), and rs2736100 T>G (TT vs. TG+GG: OR=2.27 95% CI=1.23-4.17, P=0.006) polymorphisms were associated with risk of GC. In the haplotype analysis, hTERT Grs2736109/Trs2735940/Ars2853669/Grs2736100 haplotype was also related with an increased risk of GC (OR=1.75; 95% CI: 1.05-2.93, P=0.03). Because this is the first study regarding the hTERT rs2736109 G>A, rs2735940 T>C, rs2853669 A>G and rs2736100 T>G polymorphisms and the risk of GC susceptibility in the literature, further independent studies are needed to verify our results in a larger sample sizes, as well as in patients of different populations.

A functional HOTAIR rs920778 polymorphism does not contributes to gastric cancer in a Turkish population: a case-control study.

An aberrant up-regulation of HOX transcript antisense intergenic RNA (HOTAIR), a long non-coding RNA (lncRNA), is associated with human cancers including gastric cancer (GC) and worse clinicopathological features. A naturally occurring functional single nucleotide polymorphism (SNP) rs920,778 (C→T) in the intronic enhancer of HOTAIR gene has been demonstrated to affect HOTAIR expression and cancer susceptibility. To investigate the association of the HOTAIR rs920778 polymorphism on the risk of GC susceptibility in Turkish population, a hospital-based case-control study was carried out consisting of 104 GC and 209 healthy control subjects matched on age and gender. The genotype frequency of HOTAIR rs920778 polymorphism was determined by using TaqMan Real-Time Polymerase Chain Reaction. No statistically significant differences were found in the allele or genotype distributions of the HOTAIR rs920778 polymorphism among GC and healthy control subjects (P > 0.05). Our results demonstrate that the HOTAIR rs920778 polymorphism has not been in any major role in genetic susceptibility to gastric carcinogenesis, at least in the population studied here. Independent studies are needed to validate our findings in a larger series, as well as in patients of different ethnic origins.

Relationship between IL28B gene rs8099917 polymorphism and SVR in Turkish patients with hepatitis C virus genotype 1.

BACKGROUND AND OBJECTIVE: The hepatitis C virus (HCV) which infects 3% of the world's population is a global challenge. Recently, genome-wide association studies (GWAS) have identified that the IL28B gene rs8099917 polymorphism was associated with the response to the pegylated-interferon alpha/ribavirin (PegIFNα/RBV) combination therapy in patients infected with HCV genotype 1. IL28B gene rs8099917 polymorphism should be determined before beginning treatment of HCV-infected patients to predict an individual's response. The aims of this study were to analyze the correlation between IL28B gene rs8099917 (T/G) polymorphism and PegIFNα/RBV therapy outcome in the Turkish population. METHODS: Genotypes of the IL28B gene rs8099917 (T/G) single nucleotide polymorphism (SNP) were determined in 308 patients with HCV infection by using a polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) assay. One group consisted of 148 patients with a sustained virological response (SVR), whereas the second group consisted of 160 nonresponders (non-SVR). RESULTS: Allele and genotype associations of IL28B gene rs8099917 polymorphism with a sustained virological response were observed in comparisons between the SVR and non-SVR groups (P<0.001). In addition, the characteristics of the subjects did not differ between these two groups except for age and fibrosis stage (P<0.05). Additionally, neither SVR nor rs80999917 genotypes were associated by HCV RNA levels. CONCLUSIONS: In conclusion, the rs8099917 polymorphism was thus found strongly associated with a sustained virological response to therapy in Turkish patients infected with HCV genotype 1. Consequently, we suggest determining IL28B gene rs8099917 polymorphism of patients with HCV genotype 1 before onset of treatment.

Effect of PON1 gene polymorphisms in Turkish patients with hepatocellular carcinoma.

BACKGROUND: Reactive oxygen species (ROS) can oxidize biological molecules that mediate carcinogenesis by causing metabolic malfunction and damage to DNA. Human serum paraoxonases (PON1, PON2 and PON3) play a role in antioxidant defense and protect the cell against ROS. PON1 polymorphisms Q192R and L55M have been shown to be associated with several human cancers, but their association with hepatocellular carcinoma (HCC) has yet to be investigated. METHODS: We performed genotyping analysis using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) assay in a hospital-based case-control study of 217 confirmed HCC patients and 217 age-, gender-, smoking- and alcohol consumption-matched cancer-free controls in Turkish population. RESULTS: Q192R and L55M polymorphisms were in significant linkage disequilibrium (LD) (D' = 0.77). However, allele, genotype and haplotype analysis showed no significant differences between the risks of HCC and PON1 polymorphisms. Moreover, no significant differences were found between clinical findings, clinicopathological features and sex in comparison with the PON1 genotypes in HCC group. CONCLUSION: Our results suggest for the first time that neither the Q192R polymorphism nor the L55M polymorphism has relationship with the risk of developing HCC. Further independent studies are required to clarify the possible role of PON1 gene Q192R and L55M polymorphisms on the risk of developing HCC in a larger series and also in patients of different ethnic origins.

Lack of an association of programmed cell death-1 PD1.3 polymorphism with risk of hepatocellular carcinoma susceptibility in Turkish population: a case-control study.

AIM: The programmed cell death-1 (PD-1) is a potent immunoregulatory molecule which is responsible for the negative regulation of T-cell activation and peripheral tolerance. Recently, overexpression of PD-1 has been reported to contribute to immune system evasion and poor survival of hepatocellular carcinoma (HCC). A common single nucleotide polymorphism in intron 4 of PD-1 gene called PD-1.3 has been reported to influence PD-1 expression, but its association with HCC has yet to be investigated. The aim of the present study was to investigate whether this polymorphism could be involved in the risk of HCC susceptibility. METHODS: The genotype frequency of PD-1.3 polymorphism was determined by using a polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method in 236 subjects with HCC and 236 cancer-free control subjects matched on age, gender, smoking and alcohol status. RESULTS: No statistically significant differences were found in the genotype distributions of the PD-1.3 polymorphism among HCC and cancer-free control subjects (P=0.22). CONCLUSION: Our results demonstrate for the first time that the PD-1.3 polymorphism has not been in any major role in genetic susceptibility to hepatocellular carcinogenesis, at least in the population studied here. Independent studies are needed to validate our findings in a larger series, as well as in patients of different ethnic origins.

Functional polymorphisms of cyclooxygenase-2 gene and risk for hepatocellular carcinoma.

Cyclooxygenase-2 (COX-2) influences carcinogenesis through immune response suppression, apoptosis inhibition, regulation of angiogenesis and tumor cell invasion, and metastasis. It is now well established that COX-2 is overexpressed in many premalignant, malignant, and metastatic cancers, including hepatocellular carcinoma (HCC). DNA sequence variations in the COX-2 gene may lead to altered COX-2 production and/or activity, and so they cause inter-individual differences in the susceptibility to HCC. Functional coding region polymorphisms -1195A>G (rs689466), -765G>C (rs20417), and +8473T>C (rs5275) in the COX-2 gene have recently been shown to be associated with several human cancers but their association with HCC has yet to be investigated. We used hospital-based case-control study to assess the hypothesis that the functional COX-2 variation may affect individual susceptibility to the HCC. COX-2 polymorphisms were investigated in 129 confirmed subjects with HCC and 129 cancer-free control subjects matched on age, gender, smoking, and alcohol consumption using a polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) assay. The distribution of the COX-2 -1195A>G and +8473T>C genotypes were not significantly different between HCC cases and control. However, proportion of the COX-2 -765CC genotype which leads to a 30% reduction of the COX-2 promoter activity was significantly lower in patients with HCC (3.1%) when compared to control subjects (11.6%) (P < 0.05). Logistic regression analyses revealed that the COX-2 -765G>C variant genotype (-765CC) was associated with a significantly decreased risk of HCC compared with the -765GG wild-type homozygotes [P < 0.05, odds ratio (OR) = 0.25, 95% confidence interval (CI) = 0.08-0.79]. Our results suggest for the first time that the -765CC genotype of COX-2 -765G>C polymorphism, causing lower COX-2 gen expression, is a genetic protective factor for HCC. However, because this is the first report concerning the COX-2 -1195A>G, -765G>C, and +8473T>C polymorphisms and the risk of HCC, independent studies are needed to validate our findings.

No association of NAD(P)H: quinone oxidoreductase 1 (NQO1) C609T polymorphism and risk of hepatocellular carcinoma development in Turkish subjects.

NAD(P)H:quinone oxidoreductase 1 (NQO1) is a cytosolic enzyme that catalyzes the two-electron reduction of numerous quinoid compounds into their less toxic form, thus NQO1 protecting cells against oxidative stress. The gene coding for NQO1 has a single nucleotide polymorphism (C-->T) at nucleotide position 609 (proline to serine substitution at position 187 in amino acid sequence (P187S)) (rs1800566) of the NQO1 cDNA which results in very low enzimatic activity, so it would be expected that individuals with the homologous NQO1 C609T polymorphism would have a susceptibility developing cancer. Previous studies of the association between functional NQO1 C609T polymorphism and several human cancers have had mixed findings but association of NQO1 C609T polymorphism with hepatocellular carcinoma (HCC) development has yet to be investigated. In this study, we aim to evaluate the the association of NQO1 C609T with the risk of hepatocellular carcinoma (HCC) development among Turkish population. NQO1 C609T polymorphism was investigated in 167 confirmed subjects with HCC and 167 cancer-free control subjects matched on age, gender, smoking and alcohol consumption by using a polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP) assay. There is no association between the allel or genotype of NQO1 C609T polymorphism and HCC development risk in the Turkish subjects examined (p>0.05). Our result demonstrate for the first time that the NQO1 C609T polymorphism is not a genetic susceptibility factor for HCC in the Turkish population. Independent studies are need to validate our findings in a larger series, as well as in patients of different ethnic origins.

Clinicopathologic features of dual chronic hepatitis B and C infection: a comparison with single hepatitis B, C and delta infections.

UNLABELLED: There is controversial data in the literature about the characteristics or features of dual hepatitis B and C infection. Several studies have reported that the dual infection has a more severe histological picture; faster progression leading to cirrhosis and a higher risk for hepatocellular carcinoma compared with the single infections. These findings have not yet been supported. We assessed the patients with dual hepatitis B and C infection with respect to their different features in our country. METHOD: the chronic hepatitis patients of our clinics were tested, and both HBsAg and anti-HCV positive patients with chronic hepatitis were enrolled to the study. All patients were tested for the biochemical parameters and the presence of HBV-DNA and HCV-RNA. RESULTS: Of the 1950 patients, 51 (2.6%) were both HBsAg and anti-HCV positive and 67 were anti-delta positive. Patients were followed up for 5.4+/-2.1 years. Of the 51 dual hepatitis patients, 6 had no HBV-DNA and HCV-RNA detectable by PCR, 36 were only HCV-RNA positive, 9 were only HBV-DNA positive and 3 were both HBV-DNA and HCV-RNA positive. Dominant infection in (3/4) of the patients was hepatitis C. Clinical and histological properties of the cases with dual Hepatitis B and C infection showed no significant differences compared to the single infections. In conclusion, regarding the prognosis, no significant differences were found between such dual and single infections. Dual infection with hepatitis B virus and delta virus is a significantly more severe condition than the dual infection with hepatitis B and C viruses.