This study examined genetic variants of IL28B rs12979860 as predictors of interferon-based therapy outcomes in people with hepatitis C virus (HCV) in Pakistan. The study found that HCV subgenotype 3a was most prevalent and had higher response rates to therapy than subtype 1a. Carriage of the IL28B rs12979860 CC genotype was associated with higher sustained virological response rates to therapy. In contrast, the IL28B rs8099917 polymorphism was not associated with therapy outcomes. The study concludes that HCV genotype and IL28B rs12979860 can help predict the effectiveness of interferon-plus-ribavirin combination therapy for HCV in Pakistan.

1. 1
Genetic variant of IL28B rs12979860, as predictive
Marker of interferon-based therapy in Pakistani
Population
HCV was discovered as major cause of hepatitis associated with blood transfusion in 1989.
(1) It is a major health burden affecting about 200 million people and persistent HCV infection
lead to liver inflammation,, decompensated liver.
(2)The standard treatment of HCVG1 infection is a combination of pegylated interferon alpha 2a
or 2b (PEG-IFN), ribavirin and protease inhibitor.
(3)HCV treatment is costly and have serious side effects means the treatment duration and the
success rate are mainly dependent on the HCV genotypes.
(4)The treatment response to combinational therapy of PEG-IFN and ribavirin for HCVG 2 and 3
is about 80% and approximately 40–52% for genotype 1.
(5)Two new oral antiviral agents both drugs were administered in combination with PEG-IFN and
ribavirin.
(6)The distribution of HCV genotypes varies from region to region within a country.
(7)The treatment response to interferon and ribavirin in Pakistan for HCVG1 and 4 is low,
however, it is high for HCVG3.
(8)There is a strong association of both host and viral factors with the viral clearance.
(9)Ethnic differences also contribute in natural clearance of HCV infection suggesting the
involvement of host genetics.
(10)IL28B, also known as interferon k (IFN-k3), belongs to type III IFNs. The three members of
this family: IFN-k1, IFN-k2 and IFN-k3, are also known as IL29, IL28A and IL28B respectively.
(11)Antiviral activity of IFN-k is less effective than IFN-a, however, IFN-k possesses a critical
role in the host immune system against viruses in particular.
(12)(GWAS) Genome-wide association studies have dis-closed some important SNPs in vicinity
of IL28B gene that are strongly linked with spontaneous clearance of HCV infection and treatment
response.
(21)The two major SNPs of the IL28B genes, rs12979860 and rs8099917, are acknowledged as
the strongest predictor of natural clearance and treatment response.

2. 2
Genotyping of IL28B
About 5 mL of venous blood sample was collected from each subject into EDTA tube. Genomic DNA was
extracted using Genome DNA extraction kit according to manufacturer’s instructions and quantified using
Nanodrop. AS-PCR and RFLP-PCR were employed for the genetic testing of IL28B rs12979860
polymorphism whereas for IL28B rs8099917, only RFLP-PCR was used. Both of these techniques (AS-
PCR and RFLP-PCR) are regularly used in the study of SNPs and proved to be dependable. The PCR
reaction was done.
RESULTS
NR group have higher mean viral titer than the SVR group. The majority of patients were infected with
HCV sub-genotype 3a and sub-genotype 3b. The response rates to interferon plus ribavirin treatments were
higher in HCV sub-genotypes and 3b compared to HCV sub-genotype 1a.
Distribution of IL28B genotypes
The prevalence of IL28B genetic variants rs12979860 and rs8099917 was evaluated in CHC patients and
healthy controls. The allelic distributions of the two SNPs were according to Hardy–Weinberg
equilibrium. Pearson chi-square analysis of the data showed that there was no significant variation
in distribution of IL28B rs12979860 genotypes in CHC patients and health controls. The
results of AS-PCR of IL28B rs12979860 polymorphism were completely matched with RFLP-
PCR analysis. Statistical analysis of the data showed that the prevalence of IL28B rs8099917
genotypes in both groups was non-significant.
Influence of IL28B SNPs on interferon-based therapy
The Pearson chi-square test revealed a positive association between CHC patients with genotype
IL28B rs12979860CC and treatment. The treatment response of IL28B rs12979860CT and TT
genotypes in SVRs and NRs groups was statistically insignificant.There was no association of any
genotype of IL28B rs8099917 with interferon-based therapy of HCV infection. The distribution
of treatment-favorable IL28B rs8099917TT genotype in SVRs and NRs was 36% and 24%
respectively, but was not statistically significant (p = 0.264).
DISCUSSION
HCV genotypes and baseline viral loads were involved in defining the treatment option and

3. 3
duration. GWAS have demonstrated genetic mutations in IL28B region that are strongly correlated
with natural clearance and treatment outcomes of HCV infection. This study focuses on evaluating
the potential use of HCV genotypes, viral load and IL28B genetic mutations as potential predictive
markers in interferon-based therapy for HCV infection.
Pakistan is located in South Asia. It is the sixth most populated country in the world with four
provinces. Khyber Pakhtunkhwa and Sindh are the two provinces that were selected for this study.
These results are in concordance with the previous studies i-e the HCV sub-genotype 3a as the
most prevalent HCV genotype with the highest response to interferon-based therapy.
Age is an important factor that influences the treatment response to combinational therapy for
HCV infection. CHC patients with lower age are more likely to attain higher response to
combinational therapy than older CHC patients.
The common IL28B rs12979860 genotype was CT followed by CC and TT in both patient and
healthy control groups. There were no significant variations in distributions of the IL28B
rs12979860 genotypes in both groups. However, it was observed that, IL28B rs12979860CC
genotype favors interferon-based therapy for HCV infection (p = 0.019).
IL28B rs8099917 polymorphism, the major genotype was TG followed by GG and TT. There was
no association of IL28B rs8099917TT genotype with the persistent treatment response to HCV
infection (p = 0.264). IL28B rs8099917 is also worldwide studied and known for its positive
association with response to interferon-based therapy against HCV genotype 1 and 4. The
association of this particular SNP is more likely to be dependent on HCV genotype.
Conclusion
HCV genotypes and IL28B rs12979860 are predictive markers for the efficiency of interferon plus
ribavirin combinational therapy of HCV infection. So recommendations are to the test for these
biological markers before start of interferon-based therapy in Pakistani population.
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