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Mutation Analysis of Protein Kinase Binding Domain of HCV NS5A Gene Isolated from Patients with Chronic Hepatitis C



I. Halil Kilic, M. Meltem Koruk, M. Ozaslan, I. Didem Karagoz, Yasemin Zer, Mehmet Koruk, Ceyda Uyar and Izzettin Guler
 
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ABSTRACT

The majority of hepatitis C virus (HCV) isolates are resistant to antiviral therapy. Although, the molecular mechanisms of resistance against to antiviral therapy were investigated in such studies in vivo and/or in vitro and the significant results were obtained, the resistance problem has not been solved yet. The aim of this study was to investigate the relationship between therapy and response by detecting the mutations in ISDR2209-2248 and PKR-BD2209-2274 site among HCV positive patients in Gaziantep. Fifty nine patients (25 men and 34 women) with chronic hepatitis C diagnosed at Gastroenterology Department of Gaziantep University Sahinbey Research Hospital between 2009-2010 years were included. The 59 HCV RNA positive patients: 34 women and 25 men were included. In ISDR2209-2248 site in PKR-BD2209-2274 site, 25 of 89 amino-acid substitutions (28.09%) of the sustained virologic responder patients; 12 of 78 amino-acid substitutions (15.38%) in the non-responder patients were determined. The significant relation between the number of mutation in interferon sensitivity determining region- ISDR2209-2248 and protein kinase binding domain- PKR-BD2209-2274 of NS5A gene and the response to interferon-alpha+ribavirin combination therapy was not determined (p>0.05). The number of mutation in this region was not significant in predicting the response of chronic hepatitis C patients to the treatment. The evaluation of age, individual immunization, nutritional situations, psychological situation of people infected with HCV and other gene regions would be useful.

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  How to cite this article:

I. Halil Kilic, M. Meltem Koruk, M. Ozaslan, I. Didem Karagoz, Yasemin Zer, Mehmet Koruk, Ceyda Uyar and Izzettin Guler, 2012. Mutation Analysis of Protein Kinase Binding Domain of HCV NS5A Gene Isolated from Patients with Chronic Hepatitis C. International Journal of Pharmacology, 8: 519-526.

DOI: 10.3923/ijp.2012.519.526

URL: https://scialert.net/abstract/?doi=ijp.2012.519.526
 
Received: April 20, 2012; Accepted: August 15, 2012; Published: September 06, 2012



INTRODUCTION

Hepatitis C virus infection is a major public health problem in Turkey as well as in Australia, Italy, Romania, Mongolia (Dore et al., 2003; Raffaele et al., 2001; Kmieciak et al., 2006; Gheorghe et al., 2008). Approximately 80% of the hepatitis C infections become chronic and result in important complications such as cirrhosis and hepatocellular carcinoma (Aman et al., 2012). According to the estimations of World Health Organization (WHO); HCV infection prevalence is around 3% although it varies among country (Giannini and Brechot, 2003). Today, combination of Pegylated Interferon Alpha (PEG-IFN) and a nucleoside analogue Ribavirin is used for the treatment of chronic HCV infection. After treatment during 24-48 weeks Sustained Virologic Response (SVR) was determined in 54-56% cases (Manns et al., 2001; Fried et al., 2002). The majority of HCV isolates are resistant to antiviral therapy. The molecular mechanism of resistance to antiviral therapy have been investigated in many studies in vivo and in vitro and significant results were obtained but the molecular basis of the resistance has not been clarified yet (Pawlotsky, 2000). HCV is a unique known member of the Hepacivirus genus of Flaviviridae. It is single stranded and 50 nm positive-sense RNA virus and has six major genotypes (Brass et al., 2006).

HCV genotype 1b is the most resistant and the worst responder group to treatment (Hermida et al., 1997). There are various studies with different results about mutations of Interferon Sensitivity Determining Region (ISDR) of NS5A gene of HCV. For instance, it was reported that although mutations in this region were associated with response to treatment in Japanese studies, no relationship between mutations and treatment was determined in some studies in Europe (Enomoto et al., 1995; Zeuzem et al., 1997). In our country, studies at Ankara University and Dokuz Eylul University have shown that ISDR2209-2248 mutations of NS5A gene were not associated with response to interferon treatment (Aygun, 2003; Aslan et al., 2004). These contradictory results related to ISDR2209-2248 of HCV NS5A gene mutations show us that this issue is still controversial. However, some studies about PKR-BD2209-2274 (Protein Kinase Binding Domain) region located in the same gene region as well as ISDR2209-2248 indicated to relate mutations of this region and IFN-therapy (Macquillan et al., 2004).

The aim of this study was to investigate the relationship between therapy and response to HCV infection by detecting the mutations in ISDR2209-2248 and PKR-BD2209-2274 region among HCV positive patients in Gaziantep.

MATERIALS AND METHODS

Patients: Fifty nine patients (25 men and 34 women) with chronic hepatitis C whose positive anti-HCV anticore and HCV RNA diagnosed by ELISA and also detected quantitative HCV RNA levels at Gastroenterology Department of Gaziantep University Sahinbey Research Hospital between January 2009 and January 2010 years were included.

Before treatment, quantitative HCV RNA values and biochemical values of patients were determined. These patients were treated with combination of peg-interferon alpha-2a or peg-interferon alpha 2b once for every day and ribavirin for a week at Gastroenterology Department. Patients were classified according to number of mutations in the ISDR and PKRBD regions of NS5A of HCV gene; Wild type (no mutation), Intermediate type (1-3 mutations) and Mutant Type (4 and more mutations) (Enomoto et al., 1995).

RNA extraction: Ten microliter of peripheral venous blood samples were taken from each patient. HCV RNA was obtained with HCV RNA extraction kit (RocheHigh Pure Viral RNA Kit, Germany) as directed by the manufacturer. It was added 400 μL binding buffer supplemented with Poly (A) to 200 μL serum or plasma and mixed well and then transferred to high pure filter tube assembly. After then, it was centrifuged for 15 sec at 8,000 xg and discarded the flow through and collected into the collection tube. Five hundred microliter inhibitor removal buffer was added to the upper reservoir and centrifuged for 1 min at 8,000 xg and discarded the flow through and into the collection tube 450 μL Washing Buffer was added to the upper reservoir twice. After centrifugation for 1 min at 8,000 xg and then centrifuged repeatedly at max speed for 10 sec and discarded the flow through. And 50 μL Elution Buffer added to the upper reservoir into the collection tube and centrifuged for 1 min at 8,000 xg.

RT-PCR: It was performed RT-PCR procedure according to Saiz et al. (1998) with slight modification. Twenty microliter reaction mixture containing 4 μL Buffer, 5 μL dNTP (Fermantas), 0.05 μL AMV RT, 4 μL water with DEPC, 0.05 μL RNase inhibitor, 2 μL R3 primer (Metabion, Germany) (5’-GCAATGGGCACCCGTGTACC-3’) and pure RNA was prepared. This mixture was incubated for 1 h at 42°C in Minicycler MJ Research (United States of America) device.

Amplification of PKRBD (2209-2274) of HCV NS5A gene: Nested PCR was performed for amplification and it was studied in a 50 μL reaction mixture containing 5 μL Buffer, 5 μL dNTP, 3 μL MgCI2, 2 μL Primer F3 (5’-GGGCATGACCACTGACAACGT-3’), 2 μL Primer R3 (5’-CAATGGGCACCCGTGTACC-3’), 0.25 μL Taq polymerase, 5 μL cDNA and 27.5 μL distilled water. Samples were subjected to denaturation by 1 cycle at 95°C for 5 min; followed by 30 cycles at 95°C for 1 min, at 55°C for 1 min and at 72°C for 2 min (annealing) and a final step by 1 cycle at 72°C for 7 min (elongation) in Minicycler MJ Research (United States of America) device.

In the second round of nested PCR was performed in a 50 μL mixture containing 5 μL Buffer, 5 μL dNTP, 3 μL MgCI2, 2 μL Primer F4 (5’-GCAGTGCTCACTTCCATGC TCAC-3’), 2 μL Primer R4 (5’-GGACTCTAGCAGTGGA GGGTTGTA-3’), 0.25 μL taq polymerase, 1 μL DNA and 31.75 μL distilled water. The second PCR was conducted in the same way. The size, purity and approximate obtained yield of the DNA were verified by direct observation of 1% agarose gel.

Sequence analysis: The amplified PCR products were sequenced by BigDye® Terminator v3.1 Cycle Sequencing Kit (Applied Biosystem, USA) as directed by the manufacturer within ABI 3130XL Genetic Analyzer. It was performed in a 20 μL reaction followed mixture: 8 μL big dye, 1-100 ng PCR template, 3.2 pmol primers and deionized water. HCV-genotyping was performed by the blast search program of the sequences with the HCV genotype gene bank. The phylogenetic analysis was used to differentiate the separate clusters of the identified HCV genotypes. Following sequences were used: AB056525 for 1b, AB056524 for 1b, HM042084 for 3b, HM042083 for 3b, FJ896370 for 1a, FJ896369 for 1a, AB056550 for 1c, AB056546 for 1c, AB056569 for 2a, AB056564 for 2a and AF339212 for 3a (GENBANK). PKR-BD2209-2274 and ISDR2209-2248 of HCV NS5A gene were aligned to the following reference sequence: HCV-CR (GenBank) for genotype 1b. For mutation and phylogenetic analysis MEGA 4.1 program was used.

Statistical analysis: For statistical analysis Chi-square (χ2) test at SPSS 17.0 program was used (Yuan et al., 2010). The p-value of 0.05 or less was considered to represent statistical significance.

RESULTS

Twenty nine of 59 patients with chronic hepatitis C had sustained virologic responder against to interferon-ribavirin combination therapy while 30 patients had non-responder. Amino acid changes of PKR-BD2209-2274 and ISDR2209-2248 of HCV NS5A gene were determined by comparison with sequences of HCV genotype 1b of fifty-nine patients and the NS5A-CR (GeneBank) Reference Sequence (Fig. 1). The response of patients to IFN therapy by calculated mutation frequencies in ISDR2209-2248 and PKR-BD2209-2274 site of HCV isolates was shown at Table 1. In addition, changes in amio-acids caused by mutations were determined by based on universal genetic code. Some findings of the mutation and changes in the amino-acids were presented all together in Fig. 2-5. The most common amino-acid changes was 2270 codon in PKR-BD2209-2274 region (from CTT (Leu) to GTT (Val)) (Fig. 2). Some amino-acid changes of PKR-BD2209-2274 of HCV NS5A gene isolated from patients with chronic hepatitis C was 2217 codon (from ACC (Thr) to GCC (Ala)) (Fig. 3). Most amino acid mutations was detected on patients No. 29 (9 amino acid changes). Three of nine amino acid changes was 2223, 2224 and 2225 codon (Fig. 4) Two of amino acid changes was 2251 and 2253 codon (respectively; from GTA (Val) to ATA (Ile), from CTG (Leu) toATG (Met)). In our study, 24 amino-acids changes in PKR-BD2209-2274 and 39 amino-acids changes in ISDR2209-2248 were detected. Taking into consideration of mutation frequencies in ISDR2209-2248 site of HCV isolates from patients with chronic HCV infection, 17 of 29 sustained virologic responder patients (58.6%) were wild type, 11 (37.6%) were intermediate type and only 1 (3.4%) was mutant type. And also 21 of non-responder patients to IFN combination therapy (70%) and 17 of responder patients (58.6%) were wild type. In addition, mutation frequencies in ISDR2209-2248 site of HCV isolates from non-responder patients was lower than from responder patients.

To PKR-BD2209-2274 site of HCV isolates, 21 of 29 sustained virologic responder patients to IFN combination therapy (72.4%) were intermediate type and the rest of them were mutant type. Twenty two of 30 non-responder patients were intermediate type and the rest were mutant type. Wild type PKR-BD2209-2274 site was not determined among responder and also non-responder patients. The differences between two groups was not significant (p>0.05).

DISCUSSION

Hepatitis C virus has been a major public health problem in our country like as in all over the world. Current therapy for HCV infection has been a combination of pegylated interferon alpha- ribavirin and response of patients to therapy have been affected by several host and viral factors (Davis and Lau, 1997; Ferenci, 2004; Gonzalez and Keeffe, 2011). Major effective factor for therapy has appeared HCV genotype (Shiratori et al., 1997; Martinot-Peignoux et al., 1998). We investigated whether PKR-BD2209-2274 and ISDR2209-2248 of NS5A gene of HCV has been effective viral factors for response to therapy or not. In our study, the most common change was 2218. amino-acid in ISDR2209-2248 of NS5A gene of HCV (from CAT (His) to CGT (Arg/R)). The finding was correlated with the result of Enomoto et al. (1995). The most common change was 2270. Amino-acid in PKR-BD2209-2274 region (from CTT (Leu) to GTT (Val)). The frequency of non-responder patients with wild type ISDR2209-2248 sequence was higher than responder patients. However, the frequency of non-responder patients with intermediate type ISDR2209-2248 sequence was higher than responder patients. None of those patients were detected with wild type PKR-BD2209-2274 sequence. There was no significance in the number of amino-acid changes in PKR-BD2209-2274 sequence between non-responder patients and responder patients (p>0.05). Our result was consistent with the finding of Kmieciak et al. (2006). Initially Enomoto et al. (1995) suggested that there was a strong relationship between the frequency of mutations in ISDR2209-2248 and response to treatment. Seventeen (44.7%) of thirty-eight patients with wild type ISDR2209-2248 sequence were responder and 21 (55.3%) of the patients were non-responder.

Table 1: Comparison of the number of mutations in ISDR and PKR-BD with treatment response of patients
Image for - Mutation Analysis of Protein Kinase Binding Domain of HCV NS5A Gene Isolated 
  from Patients with Chronic Hepatitis C
SVR: Sustained virological responder, NR: Non-responder

Image for - Mutation Analysis of Protein Kinase Binding Domain of HCV NS5A Gene Isolated 
  from Patients with Chronic Hepatitis C
Fig. 1: Amino acid changes of PKR-BD and ISDR of HCV NS5A gene and reference sequence (SVR: Sustained virologic response, NR: Non responders)

Image for - Mutation Analysis of Protein Kinase Binding Domain of HCV NS5A Gene Isolated 
  from Patients with Chronic Hepatitis C
Fig. 2: L2270V mutation (from CTT (Leu) to GTT (Val))

Image for - Mutation Analysis of Protein Kinase Binding Domain of HCV NS5A Gene Isolated 
  from Patients with Chronic Hepatitis C
Fig. 3: T2217A mutation (from ACC (Thr) to GCC (Ala))

Image for - Mutation Analysis of Protein Kinase Binding Domain of HCV NS5A Gene Isolated 
  from Patients with Chronic Hepatitis C
Fig. 4: A2223L, A2224A and L2225P mutations, respectively; from GCC (Ala) to CTT (Leu), from GAC (Asp) to GCC (Ala), from CTC (Leu) to TTC (Phe)

Image for - Mutation Analysis of Protein Kinase Binding Domain of HCV NS5A Gene Isolated 
  from Patients with Chronic Hepatitis C
Fig. 5: V2251I and L2253M mutations, respectively; from GTA (Val) to ATA (Ile), from CTG (Leu) to ATG (Met)

As evaluation ISDR2209-2248 site of responder patients, the frequency of intermediate type was higher than rate of wild type which seems to support that the positive relationship between sustained virologic response and the frequency of mutation. Japanese studies confirmed generally that there was a strong relationship between response to treatment and ISDR2209-2248 mutations (Enomoto et al., 1995; Kurosaki et al., 2011). However, European and Asian studies were differed with them (Zeuzem et al., 1997; Hu et al., 2002). Several studies were performed the relationship between response to treatment and mutations in ISDR2209-2248 by different groups of Japan, Europe and America (Hofgartner et al., 1997; Torres-Puente et al., 2008; El-Shamy et al., 2011).

Zeuzem et al. (1997) showed that no association between European NS5A2209-2248 of HCV-1b isolates and initial-sustain response to IFN-alpha therapy. They also found no correlation between the number of amino acid changes in the NS5A2209-2248 region and the initial decline of serum HCV-RNA (Zeuzem et al., 1997).

We studied the same issue in 59 Turkish patients chronically infected with subtype HCV-1b and we obtained similar result with Zeuzem and colleagues (Zeuzem et al., 1997). We found no significant relationship between mutation number of ISDR2209-2248 of NS5A gene of HCV-1b isolates and response to IFN-alpha therapy (p>0.05).

In accordance with the meta-analysis study based on geographical varieties; an obvious positive relationship between ISDR2209-2248 sequence types and sustained virologic response was shown in European patients. However, the rate of sustained virologic response in Japanese patients infected with mutant ISDR2209-2248 type HCV was more than in European patients. The rate of sustained virologic response increased as the number of mutation in ISDR2209-2248 increased. However, it was suggested that this relationship was more apparent in Japanese patients (Pascu et al., 2004).

In a study at our country, Aslan and colleagues reported that the mutation in ISDR2209-2248 of NS5A gene was not associated with response to interferon treatment in Turkish patients with chronic hepatitis C virus genotype lb infection (Aslan et al., 2004). In Spain, De Rueda et al. (2008) investigated that relationship between mutation of E2-PePHD, NS5A-PKRBD, NS5A-ISDR and NS5A-V3 of HCV Genotype 1 and PEGinterferon+ribavirin therapy. They reported that the presence of >4 mutations in the PKR-BD2209-2274 region was associated with SVR and early virologic responses. In terms of this region, when intermediate type and mutant type were evaluated according to sustained virologic response, no statistically significant relationship had been detected (De Rueda et al., 2008). In our study, wild type PKR-BD2209-2274 sequence in responder and also non-responder patients was not determined. In the same region, patients with intermediate type PKR-BD2209-2274 sequence (72.9%) was more than patients with mutant type PKR-BD2209-2274 sequence (27.1%).

Murphy et al. (2002) reported that no relationship between the frequencies of mutations in PKR-BD2209-2274 and in intermediate type as correlation with us (Murphy et al., 2002). In contrary to our result, it was reported that the difference between the frequency of mutation in PKR-BD2209-2274 and in intermediate type was not significant as compared with sustained virologic responder and non-responder patients (Gale et al., 1998; Christoph et al., 2000). While our findings related to this region do not indicate any parallelism to the studies of Gale et al. (1998) and Christoph et al. (2000), they indicate conformity with the study of Murphy et al. (2002).

CONCLUSION

In conclusion, any relationship was not found between PKR-BD2209-2274 of NS5A gene of HCV and response to interferon therapy. Thus, it was detected that the number of mutation in this region was not significant in predicting the response of chronic hepatitis C patients to the treatment. In addition to these regions, evaluation of age, individual immunization, nutritional situations, psychologic situation of people infected with HCV and other gene regions would be useful.

REFERENCES

  1. Aman, W., M. Shaymaa, S. Gamal and M.A. Shaker, 2012. Current status and future directions in the management of chronic hepatitis C. Virol. J., vol 9,
    CrossRef  |  


  2. Aslan, N., A.M. Bozdayi, H. Cetinkaya, M. Sarioglu and C. Turkay et al., 2004. The mutations in ISDR of NS5A gene are not associated with response to interferon treatment in Turkish patients with chronic hepatitis C virus genotype lb infection. Turk. J. Gastroenterol., 15: 21-26.
    Direct Link  |  


  3. Aygun, N., 2003. Interferon + ribavirin in the treatment of chronic hepatitis C patients with virological nonresponders to interferon sensitivity determining genotype 1b virus genome (ISDR) to determine the series of mutations/Myth of national public access system. Masters Thesis, Health Sciences Institute, Department of Medical Biology and Genetics, Dokuz Eylul University, Izmir, Turkey.


  4. Brass, V., D. Moradpour and H.E. Blum, 2006. Molecular virology of Hepatitis C Virus (HCV): 2006 Update. Int. J. Med. Sci., 3: 29-34.
    Direct Link  |  


  5. Dore, G.J., M. Law, M. MacDonald and J.M. Kaldor, 2003. Epidemiology of hepatitis C virus infection in Australia. J. Clin. Virol., 26: 171-184.
    PubMed  |  


  6. El-Shamy, A., I. Shoji, T. Saito, H. Watanabe and Y.H. Ide et al., 2011. Sequence heterogeneity of NS5A and core proteins of hepatitis C virus and virological responses to pegylated-interferon/ribavirin combination therapy. Microbiol. Immunol., 55: 418-426.
    CrossRef  |  


  7. Enomoto, N., I. Sakuma, Y. Asahina, M. Kurosaki and T. Murakami et al., 1995. Comparison of full-length sequences of interferon-sensitive and resistant hepatitis C virus 1b. J. Clin. Invest., 96: 224-230.
    CrossRef  |  Direct Link  |  


  8. Ferenci, P., 2004. Predictors of response to therapy for chronic hepatitis C. Semin Liver Dis., 24: 25-31.
    CrossRef  |  


  9. Fried, M.W., M.L. Shiffman, K.R. Reddy, C. Smith and G. Marinos et al., 2002. Peginterferon alfa-2a plus ribavirin for chronic hepatitis C virus infection. N. Engl. J. Med., 347: 975-982.
    PubMed  |  Direct Link  |  


  10. Gale, M., C.M. Blakely, B. Kwieciszewski, S.L. Tan and M. Dossett et al., 1998. Control of PKR protein kinase by hepatitis C virus nonstructural 5A protein: Molecular mechanisms of kinase regulation. Mol. Cell Biol., 18: 5208-5218.
    Direct Link  |  


  11. Davis, G.L. and J.Y.N. Lau, 1997. Factors predictive of a beneficial response to therapy of hepatitis C. Hepatology, 26: 122S-127S.
    CrossRef  |  


  12. Gheorghe, L., I.E. Csiki, S. Iacob, C. Gheorghe, G. Smira and L. Regep, 2008. The prevalence and risk factors of hepatitis C virus infection in adult population in Romania: A nationwide survey 2006-2008. J. Gastrointestin Liver Dis., 19: 373-379.
    PubMed  |  


  13. Giannini, C. and C. Brechot, 2003. Hepatitis C virus biology. Cell Death Differ., 10: 27-38.
    CrossRef  |  PubMed  |  


  14. Hermida, M., A. Castro, S. Lopez., P. Vega and J. Pedreira, 1997. The efficacy of interferon alfa treatment in chronic hepatitis C in relation to the serum concentration of RNA-HCV and the viral genotype. An. Med. Int., 14: 500-505.
    PubMed  |  


  15. Hofgartner, W.T., S.J. Polyak, D.G. Sullivan, R.L. Carithers Jr. and D.R. Gretch, 1997. Mutations in the NS5A gene of hepatitis C virus in North American patients infected with HCV genotype 1a or 1b. J. Med. Virol., 53: 118-126.
    CrossRef  |  PubMed  |  


  16. Hu, Y., M. Tang, W. Jiang, Y. Wu, Z. Yuan and Y. Wen, 2002. Association between NS5A gene sequence and response to interferon therapy in chronic hepatitis C patients in Shanghai. Zhonghua Shi Yan He Lin Chuang Bing Du Xue Za Zhi, 16: 114-118.
    PubMed  |  Direct Link  |  


  17. Kmieciak, D., L. Kruszyna, P. Migdalski, M. Lacinski, J. Juszczyk and W.H. Trzeciak, 2006. Mutations within protein kinase R-binding domain of NS5A protein of Hepatitis C Virus (HCV) and specificity of HCV antibodies in pretreatment sera of HCV-chronically infected patients and their effect on the result of treatment. Jpn. J. Infect. Dis., 59: 92-99.
    PubMed  |  Direct Link  |  


  18. Kurosaki, M., N. Sakamoto, M. Iwasaki, M. Sakamoto and Y. Suzuki et al., 2011. Sequences in the interferon sensitivity-determining region and core region of hepatitis C virus impact pretreatment prediction of response to PEG-interferon plus ribavirin: Data mining analysis. J. Med. Virol., 83: 445-452.
    CrossRef  |  PubMed  |  


  19. Macquillan, G.C., X. Niu, D. Speers, S. Englihs and G. Garas et al., 2004. Does sequencing the PKRBD of hepatitis C virus NS5A predict therapeutic response to combination therapy in an Australian population? J. Gastroenterol. Hepatol. 19: 551-557.
    CrossRef  |  PubMed  |  


  20. Martinot-Peignoux, M., N. Boyer, M. Pouteau, C. Castelnau and N. Giuily et al., 1998. Predictors of sustained response to alpha interferon therapy in chronic hepatitis C. J. Hepatol., 29: 214-223.
    PubMed  |  


  21. Pascu, M., P. Martus, M. Hohne, B. Wiedenmann, U. Hopf, E. Scherier and T. Berg, 2004. Sustained virological response in hepatitis C virus type 1b infected patients is predicted by the number of mutations within the NS5A-ISDR: A meta-analysis focused on geographical differences. Gut, 53: 1345-1351.
    CrossRef  |  


  22. Pawlotsky, J.M., 2000. Hepatitis C virus resistance to antiviraltherapy. Hepatology, 32: 889-896.
    CrossRef  |  PubMed  |  


  23. Raffaele, A., M. Valenti, M. Iovenitti, A. Matani and M.L. Bruno et al., 2001. High prevalence of HCV infection among the general population in a rural area of central Italy. Eur. J. Epidemiol., 17: 41-46.
    CrossRef  |  PubMed  |  


  24. De Rueda, P.M., J. Casado, R. Paton, D. Quintero and A. Palacios et al., 2008. Mutations in E2-PePHD, NS5A-PKRBD, NS5A-ISDR and NS5A-V3 of hepatitis C virus genotype 1 and their relationships to pegylated interferon-ribavirin treatment responses. J. Virol., 82: 6644-6653.
    CrossRef  |  


  25. Saiz, J.C., F.X. Lopez-Labrador, S. Ampurdanes, J. Dopazo, X. Forns, J.M. Sanchez-Tapias and J. Rodes, 1998. Theprognosticrelevance of thenonstructural 5A gene interferon sensitivitydeterminingregion is different in infectionswithgenotype 1b and3a isolates of hepatitis C virus. J. Infect. Dis., 177: 839-847.
    PubMed  |  


  26. Christoph, S., T. Berg, J.H. Lee, B. Ruster, B. Kronenberger, W.K. Roth and S. Zeuzem, 2000. Mutations in the protein kinase-binding domain of the NS5A protein in patients infected with hepatitis C virus type 1a are associated with treatment response. J. Infect. Dis., 181: 432-441.
    CrossRef  |  


  27. Shiratori, Y., N. Kato, O. Yokosuka, F. Imazeki and E. Hashimoto et al., 1997. Predictors of the efficacy of interferon therapy in chronic hepatitis C virus infection. Tokyo-Chiba Hepatitis Res. Group. Gastroenterol., 113: 558-566.
    PubMed  |  


  28. Torres-Puente, M., J.M. Cuevas, N. Jimenez-Hernandez, M.A. Bracho and I. Garcia-Robles et al., 2008. Gonzalez-Candelas. Hepatitis C virus and the controversial role of the interferon sensitivity determining region in the response to interferon treatment. J. Med. Virol., 80: 247-253.
    CrossRef  |  


  29. Yuan, H., M. Jain, K.K. Snow, M. Gale, W.M. Lee and The HALT-C Trial Group, 2010. Evolution of hepatitis C virus NS5A region in breakthrough patients during pegylated ınterferon and ribavirin therapy. J. Viral. Hepat., 17: 208-216.
    CrossRef  |  


  30. Zeuzem, S., J.H. Lee and W.K. Roth, 1997. Mutations in the nonstructural 5A gene of European hepatitis C virus isolates and response to interferon Alfa. Hepatology, 25: 740-743.
    CrossRef  |  PubMed  |  


  31. Gonzalez, S.A. and E.B. Keeffe, 2011. IL-28B as a predictor of sustained virologic response in patients with chronic hepatitis C virus ınfection. Gastroenterol. Hepatol., 7: 366-673.
    Direct Link  |  


  32. Murphy, M.D., H.R. Rosen, G.I. Marousek and S. Chou, 2002. Analysis of sequence configurations of the ISDR, PKR-Binding domain and V3 region as predictors of response to induction interferon alfa and Ribavirin therapy in chronic hepatitis C infection. Digestive Dis. Sci., 47: 1195-1205.
    CrossRef  |  PubMed  |  


  33. Manns, M.P., J.G. McHutchison, S.C. Gordon, V.K. Rustgi and M. Shiffman et al., 2001. Peginterferon alfa-2b plus ribavirin compared with interferon alfa-2b plus ribavirin for initial treatment of chronic hepatitis C: A randomised trial. Lancet, 358: 958-965.
    CrossRef  |  PubMed  |  Direct Link  |  


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