Oğuz Üsküdar

No association of pre-microRNA-146a rs2910164 polymorphism and risk of hepatocellular carcinoma development in Turkish population: A case-control study

AIMnMicroRNAs (miRNAs) are an abundant class of small non-protein coding RNAs with posttranscriptional regulatory functions as tumor suppressors and oncogenes. Aberrant expression and structural alteration of miRNAs are considered to participate in tumorigenesis and cancer development. It has been suggested that the presence of single nucleotide polymorphisms in precursor miRNAs (pre-miRNAs) can alter miRNA processing, expression, and/or binding to target mRNA and represent another type of genetic variability that can contribute to the susceptibility of human cancers. A G/C polymorphism (rs2910164), which is located in the sequence of miR-146a precursor, results in a change from G:U to C:U in its stem region.nnnMETHODSnTo determine the association of the miR-146a rs2910164 polymorphism on the risk of hepatocellular carcinoma (HCC) development in Turkish population, a hospital-based case-control study was designed consisting of 222 subjects with HCC and 222 cancer-free control subjects matched on age, gender, smoking and alcohol status. The genotype frequency of miR-146a rs2910164 polymorphism was determined by using a polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) assay.nnnRESULTSnNo statistically significant differences were found in the allele or genotype distributions of the miR-146a rs2910164 polymorphism among HCC and cancer-free control subjects (p>0.05).nnnCONCLUSIONnOur results demonstrate that the miR-146a rs2910164 polymorphism has no 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.

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.

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

BACKGROUND AND OBJECTIVEnThe hepatitis C virus (HCV) which infects 3% of the worlds 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 individuals 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.nnnMETHODSnGenotypes 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).nnnRESULTSnAllele 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.nnnCONCLUSIONSnIn 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.

Higher Hepatis C virus concentration in platelets than in plasma in a patient with ITP

To the Editor Thrombocytopenia is a frequent feature in patients with liver cirrhosis. It has been mainly attributed to the sequestration of platelets by an enlarged spleen due to portal hypertension which is named as ‘‘hypersplenism’’ [1]. Mechanisms other than hypersplenism such as reduced thrombopoetin or bone marrow suppression also contribute this thrombocytopenia [2]. A 55 year old male was admitted to the hospital with a diagnosis of severe thrombocytopenia. Platelet counts were 4500 m/l and peripheral smear was compatible with it. Patient’s history revealed right nephrectomy twelve years ago due to chronic pyelonephritis and blood transfusion. HCV hepatitis was diagnosed 8 years ago and liver biopsy showed chronic hepatitis with hepatitis activity index 9 and fibrosis score stage 3–4. Patient was treated with ribavirin plus interferon for 6 months but viral eradication could not be achieved. Physical examination revealed splenomegaly without other signs of decompansation. Neither petechiael nor purpuric lesions were observed. Laboratory studies revealed AST: 104 U/l, ALT: 60 U/l, albumin: 3.2 g/dl, creatinine: 0.8 mg, INR: 1.2, WBC: 4500, hemoglobin: 12.2 g/dl. HCV viral load was 816 000 IU/ml with genotype 1. ANA was negative but antithrombocyte antibody was positive. Bone marrow aspiration and biopsy showed increased number of megacaryocytes. (Figure 1) Thrombocytes were filtered in aphaeresis unit and thrombocyte HCV viral load was measured as 5 560 000 IU/Ml which was seven times higher than serum level. Patient did not respond to 14 days steroid (1 mg/kg) treatment and intravenous immunoglobulin (1 g/kg, for 2 days) was administered. Ten days after immunoglobulin administration platelet counts increased up to 300 000/ml. (Figure 2) The degree of thrombocytopenia in HCV patients appears to be significantly greater than other forms of liver disease [3]. Other than aforementioned mechanisms hepatitis C virus can also cause thrombocytopenia as a result of bone marrow suppression resulting from HCV itself or aberrations of the immune system

C-Reactive Protein and Platelet-Lymphocyte Ratio as Potential Tumor Markers in Low-Alpha-Fetoprotein Hepatocellular Carcinoma

The hepatocellular carcinoma (HCC) tumor marker alpha-fetoprotein (AFP) is only elevated in about half of the HCC patients, limiting its usefulness in following the effects of therapy or screening. New markers are needed. It has been previously noted that the inflammation markers C-reactive protein (CRP) and platelet-lymphocyte ratio (PLR) are prognostically important and may reflect HCC aggressiveness. We therefore examined these 2 markers in a low-AFP HCC cohort and found that for HCCs > 2 cm, both markers significantly rise with an increasing maximum tumor diameter (MTD). We calculated the sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV), and Youden index value for each marker, and their area-under-the-curve values for each MTD group. Patients were dichotomized into 2 groups based on the CRP and PLR from the receiver-operating characteristic curve analysis. In the logistic regression models of the 4 different MTD patient groups, CRP and PLR levels were statistically significant to estimate MTD in univariate logistic regression models of MTD groups > 2 cm. CRP and PLR were then combined, and the combination was statistically significant to estimate MTD groups of 3-, 4-, and 5-cm cutoffs. CRP and PLR thus have potential as tumor markers for low-AFP HCC patients, and possibly for screening.

Macroscopic Portal Vein Thrombosis in HCC Patients

Macroscopic portal vein invasion (PVT) by hepatocellular carcinoma (HCC) in the liver is one of the most important negative prognostic factors for HCC patients. The characteristics of a large cohort of such patients were examined. We found that the percent of patients with PVT significantly increased with increasing maximum tumor diameter (MTD), from 13.7% with tumors of MTD <5cm to 56.4% with tumors of MTD >10cm. There were similar numbers of HCC patients with very large tumors with and without PVT. Thus, MTD alone was insufficient to explain the presence of PVT, as were high AFP levels, since less than 50% of high AFP patients had PVT. However, the percent of patients with PVT was also found to significantly increase with increasing blood alpha-fetoprotein (AFP) levels and tumor multifocality. A logistic regression model that included these 3 factors together showed an odds ratio of 17.9 for the combination of MTD>5.0cm plus tumor multifocality plus elevated AFP, compared to low levels of these 3 parameters. The presence or absence of macroscopic PVT may therefore represent different HCC aggressiveness phenotypes, as judged by a significant increase in tumor multifocality and AFP levels in the PVT positive patients. Factors in addition to MTD and AFP must also contribute to PVT development.

Characteristics of Hepatocellular Carcinoma Aggressiveness Factors in Turkish Patients

A large cohort of hepatocellular carcinoma (HCC) patients from several collaborating Turkish institutions were examined for the tumor parameters of maximum diameter (MTD), portal vein thrombosis (PVT), and α-fetoprotein (AFP) levels. A relationship was found between MTD and blood platelet levels. Patients with large ≥5 cm tumors who had normal platelet levels had significantly larger tumors, higher percent of PVT, and significantly lower blood total bilirubin and liver cirrhosis than similar ≥5 cm tumor patients having thrombocytopenia. A comparison of patients with and without PVT showed significantly larger tumors, greater multifocality, blood AFP, and C-reactive protein levels, and, interestingly, lower HDL levels in the patients with PVT. Fifty-eight percent of the total cohort had AFP levels ≤100 IU/mL (and 42.1% had values ≤20 IU/mL). These patients had significantly smaller tumors, less tumor multifocality and percent PVT, lower total bilirubin, and less cirrhosis. There was considerable geographic heterogeneity within Turkey in the patterns of HCC presentation, with areas of higher and lower hepatitis B virus, hepatitis D virus, cirrhosis, and tumor aggressiveness parameters. Turkish patients thus have distinct patterns of presentation, but the biological relationships between MTD and both platelets and bilirubin levels are similar to the relationships that have been reported in other ethnic patient groups.

HCC with low- and normal serum alpha-fetoprotein levels

A large database of 1773 HCC patients in Turkey was examined. 41.9% had alpha-fetoprotein (AFP) levels <20 IU/ml and an additional 16.123% had values between 20-100 IU/ml. This 58% of the cohort (<100 IU/ml AFP levels) was examined in detail. 66% of patients with small (<5 cm) HCCs had low AFP, compared to 49% of patients with larger (>5 cm) HCCs. The mean diameter (MTD) of larger MTD, low AFP tumors was 8.4cm. Therefore, factors other than AFP must contribute to HCC tumor growth. Larger tumors in low AFP patients had both higher platelet levels and increased PVT percent. Linear regression analysis for both MTD and multifocality showed that platelet numbers and presence of PVT were significant variables; whereas for PVT, significant variables were albumin, alkaline phosphatase and MTD. Comparisons between patients with AFP levels <20, 20-<100, 100-<1000 and >1000 IU/ml showed the most significant tumor finding was an increase in PVT percent between each group, and to a lesser extent, MTD. Thus, low- or normal-AFP HCCs constitute the majority of patients and have slightly lower MTD and much lower PVT percent than HCCs associated with elevated blood AFP levels. New, non-AFP markers are thus needed, especially for small HCCs.