Cytokine xxx (2015) xxx–xxx

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Cytokine
journal homepage: www.journals.elsevier.com/cytokine

Assessment of interleukin 28B genotype as a predictor of response

to combined therapy with pegylated interferon plus ribavirin
in HCV infected Egyptian patients
Mona M. Fathy a,⇑, Mohamed E. Abo Taleb b, Mohamed S. El Hawary b, Mona I. Nabih b, Wael M. Aref b,
Manal M. Makhlouf a
a
Clinical and Chemical Pathology, Faculty of Medicine, Cairo University, Cairo, Egypt
b
Department of Internal Medicine, Faculty of Medicine, Cairo University, Cairo, Egypt

a r t i c l e i n f o a b s t r a c t

Article history: Background and aim: Single nucleotide polymorphisms (SNPs) of interleukin 28B (IL28B) gene is associ-
Received 1 November 2014 ated with spontaneous clearance and variable response to combined therapy with pegylated interferon
Received in revised form 24 April 2015 (PEG-IFN) and ribavirin (RBV) in chronic hepatitis C virus (HCV) infected patients. This study aimed at
Accepted 3 May 2015
assessing the value of IL28B rs8099917 gene polymorphism in predicting sustained virological response
Available online xxxx
(SVR) among HCV infected Egyptian patients treated with PEG-IFN and RBV.
Methods: Our study was conducted on 153 chronic HCV infected patients treated with PEG-IFN and RBV.
Keywords:
Genotyping of rs8099917 near the IL-28B gene was performed by Real Time PCR using Taq-Man probe assay.
Interleukin 28B
Genotype
Results: The overall SVR was achieved in 49.6% of patients. Patients with TT genotype showed significantly
Real Time PCR higher SVR rate than minor allele (TG/GG) carriers (74% vs. 26%, P = 0.004). Logistic regression analysis
Interferonk revealed that TT carriers had 2.8 higher chance for SVR achievement than G allele carriers TG/GG
(OR = 2.8, 95% CI = 1.4–5.6, P = 0.004). Younger age, male sex and low activity grading were significant pre-
dictors of SVR (P = 0.003, P = <0.001 and P < 0.001 respectively). High pretreatment AST levels and advanced
liver fibrosis were negative predictors of SVR (P = 0.04 and P < 0.001 respectively).
Conclusion: IL28B genotype is a significant pre-treatment predictor of response to PEG-IFN/RBV in HCV
infected Egyptian patients.
Ó 2015 Elsevier Ltd. All rights reserved.

1. Introduction polymorphisms (SNPs) in rs8099917 and rs12979860 near IL28B

Hepatitis C virus (HCV) is a major cause of chronic liver disease,
with more than 200 million infected individuals worldwide [1].
Genotype 4 (HCV-4) is the most frequent cause of chronic hepatitis
C in the Middle-East, and sub-Saharan Africa and recently it has
spread to Southern Europe as well as other western countries [2–
4]. Egypt is the country with the highest prevalence of HCV world-
wide (15%), where HCV-4 represents 90% of all HCV cases [5].
Combined pegylated interferon and ribavirin therapy
(PEG-IFN/RBN) are still the most widely used therapies for HCV-4
[6]. However, the rate of sustained virological response (SVR) with
this regimen is around 50% [7]. The long-term response to therapy
is influenced by many host and viral factors [8]. Recent evidence
has indicated that two host genetic single nucleotide
⇑ Corresponding author at: Clinical and Chemical Pathology Department, Faculty
of Medicine, Cairo University, Kasr-Al-Eini St, Cairo, Egypt. Tel.: +20 0111 6549 370.
E-mail address: muna_fathy@hotmail.com (M.M. Fathy).
gene, on chromosome 19, might be predictors of virological
response in patients with HCV treated with PEG-IFN/RBN [9–12].
IL28B encodes the antiviral cytokine IFN-k3, which belongs to
type III IFN-k family consisting of IL29/IFN-k1, IL28A/IFN-k2, and
IL28B. Type I IFNs (IFN-a,-ß) and III IFNs induce transcription of
IFN-stimulated genes (ISGs) by activating the Janus kinase-signal
transducer and activator of transcription (JAK-STAT) pathway
through different cell surface receptors in order to mediate their
potent antiviral effects [13]. Studies have revealed an association
between IL28B genotype and expression levels of intrahepatic
ISGs. In addition, the innate immune system: Toll-like receptor 3
and retinoic acid-inducible gene I signaling pathways of IFN-b
induction has an essential role in host antiviral defense against
HCV infection [14]. Asahina et al. [15] showed that the intrahepatic
genes expressions involving innate immunity were strongly associ-
ated with IL28B genotype and response to PEG-IFN/RBV.
Certain subset of dendritic cells (DCs) within human peripheral
blood mononuclear cells could recognize HCV and produce large

http://dx.doi.org/10.1016/j.cyto.2015.05.003
1043-4666/Ó 2015 Elsevier Ltd. All rights reserved.

Please cite this article in press as: Fathy MM et al. Assessment of interleukin 28B genotype as a predictor of response to combined therapy with pegylated
interferon plus ribavirin in HCV infected Egyptian patients. Cytokine (2015), http://dx.doi.org/10.1016/j.cyto.2015.05.003
2 M.M. Fathy et al. / Cytokine xxx (2015) xxx–xxx
amounts of IFN-ks. The ability of production of IFN-k3 was superior
in subjects with a favorable IL28B genotype [16]. Moreover, IFN-a
directly affected DC function and significantly increased IFN-k pro-
duction [17]. Based on these findings, it is tempting to speculate
that exogenous IFN-a would increase IFN-k production by DCs
and/or HCV-infected hepatocytes during IFN-a therapy, and this
could provide a potential explanation to IL28B genetic variants as
a predictor to the outcome of IFN-a therapy.
The objective of this study was to evaluate the predictive value
of the IL-28B rs8099917 gene polymorphism for treatment
response in HCV infected Egyptian patients treated with combined
therapy of pegylated interferon (PEG-IFN) and ribavirin (RBV).
2. Patients and methods
The current prospective study was conducted on 153 chronic
HCV infected Egyptian patients who attended the National
Hepatology and Tropical Medicine Research Institute for receiving
HCV treatment, from May 2011 till May 2012. Patients were sub-
jected to thorough clinical assessment and routine pre-treatment
work up including: complete blood picture (CBC), liver and kidney
function tests, fasting blood sugar (FBS), thyroid stimulating hor-
mone (TSH), antinuclear antibodies (ANA) titre, Hepatitis B surface
antigen (HBsAg), Hepatitis B core antibody (HbcAb), anti-HCV anti-
body, quantitative PCR for HCV-RNA, anti-Schistosomal antibodies,
alpha fetoprotein (AFP) and human immunodeficiency virus (HIV).
Patients were also subjected to abdominal ultrasonography and
histopathological examination of percutaneous liver biopsy.
Eligible patients had HCV with persistent (>6 months) eleva-
tion of ALT 1.5 times above the upper normal limit, detectable
HCV antibodies, positive PCR for HCV-RNA and with histological
evidence of chronic hepatitis. Patients were 18–65 years old and
not previously treated with interferon based therapy. Patients
were excluded if they had decompensated liver disease, HBV or
HIV infection, active Schistosomiasis, hemoglobin (Hb) < 13 g/dL
(for men) and <12 g/dL (for women), white blood cell count
(WBC) < 3000/mm3, neutrophil count < 1500/mm3, platelet
count < 100,000/mm3, serum creatinine above upper limit of nor-
mal, ANA titre > 1/160 or TSH out of normal range (0.5–5 mlU/L).
Also, patients with poorly controlled diabetes mellitus, hyperten-
sion or psychiatric diseases were excluded.
Sixty-eight patients (44.4%) received PEG-IFN-a 2a (180 lg/
week), while 85 patients (55.6%) received PEG-IFN-a 2b (1.5 lg/kg/
week) subcutaneously in combination with oral ribavirin according
to body weight (<60 kg: 800 mg/day; 60–75 kg: 1000 mg/day; and
>75 kg: 1400 mg/day). The end point of the study was the achieve-
ment of SVR (negative HCV RNA at 24 weeks after completion of
treatment). The duration of treatment was adjusted according to
their response which was determined using quantitative PCR for
HCV RNA at the 12th and 24th weeks of treatment. Patients who
had undetectable viraemia or decrease of viral load by more than
two log at the 12th week from start of treatment continued treat-
ment, otherwise treatment was discontinued. Patients with any
detectable viraemia at 24th week from start of treatment also
stopped treatment. The duration of treatment was 48 weeks.
Detection of serum HCV RNA by PCR was performed at the end
of treatment and 24 weeks after the end of treatment for SVR.
All the patients gave an informed consent for the collection
and storage of serum samples and liver biopsy and for testing
of their DNA for research purposes consistent with the current
study.
2.1. Serological tests for chronic hepatitis markers
Anti-HCV using the third generation enzyme immunoassay
(Abbot Laboratories, North Chicago, IL), HBsAg using Abbott
Please cite this article in press as: Fathy MM et al. Assessment of interleukin 28B genotype as a predictor of response to combined therapy with pegylated
interferon plus ribavirin in HCV infected Egyptian patients. Cytokine (2015), http://dx.doi.org/10.1016/j.cyto.2015.05.003
Monoclonal EIA and anti-HBc (IgG and IgM) using the Abbott
CORZYME commercial assay (Abbott Laboratory, Ludwigshafen,
Germany).
2.2. Detection of HCV-RNA
HCV-RNA was assessed, using real time quantitative PCR
Technique (Applied Biosystems, Foster city, CA, USA),
Pretreatment then at 12th, 24th and 48th weeks of treatment
and at 24 weeks after end of treatment for all patients to detect
the viral load. Lower limit of detection was 50 IU/ml.
2.3. Genotyping of IL-28B (rs8099917) by Real time PCR technique
Extraction of genomic DNA was done using QIAamp DNA blood
Mini kit (Qiagen, Hilden, Germany) by silica-gel spin columns.
Real-time PCR allelic discrimination was designed using Taq-Man
SNP Genotyping Assays [18] (Applied Biosystems, Foster City, CA,
USA) and performed on Step One™ Real Time PCR System
(Applied Biosystems), using the fluorogenic 5‘nuclease with
TaqMan minor groove binder (MGB) probes to define IL-28B gene
SNP (rs8099917), (assay ID: C_559715_20). The final volume of
each reaction was 25 ll, consisting of 12.5 ll TaqMan Universal
PCR Master Mix (2), 1.25 ll assay mix (20) contained primers
and probes, 5 ll genomic DNA, and 6.25 ll nuclease free water.
Negative control (no DNA template) was also kept to ensure that
there was no amplification of contaminating DNA. The amplifica-
tion reactions were carried out with initial hold step at 95 °C for
10 min followed by 40 cycles of three-step PCR: denaturation at
92 °C for 15 s, annealing at 60 °C for 30 s and extension at 60 °C
for 30 s. The fluorescence signal increases when the probe with
the exact sequence match binds to the single stranded template
DNA and is digested by the 5‘nuclease activity of AmpliTaq-Gold
DNA polymerase. Digestion of the probe releases the fluorescent
reporter dye from the quencher dye.
serum levels of IL28 or IFN k were not measured in this study.
2.4. Liver biopsy for histological assessment
Trans-abdominal ultrasound guided liver biopsy was done for
all patients prior to starting treatment for HCV to assess grade of
inflammation (activity) and stage of fibrosis according to
METAVIR scoring system [19]. The stage of liver fibrosis was scored
F0 (no fibrosis), F1 (mild fibrosis: portal fibrosis without septa), F2
(moderate fibrosis: few septa), F3 (severe fibrosis: numerous septa
without cirrhosis) and F4 (cirrhosis). Inflammation was graded A0
(no activity), A1 (mild activity), A2 (moderate activity), A3 (severe
activity).
3. Statistical analysis
Quantitative data were presented as mean ± SD for normally
distributed data and as medians and interquartile ranges for
skewed data. Differences among groups were compared by
Student’s t test, ANOVA test, Kruskal–Wallis analysis of variance,
and/or Mann–Whitney test, for normally distributed and skewed
data respectively. All tests were two-tailed and considered signifi-
cant at P < 0.05. The Hardy–Weinberg equilibrium was tested by a
goodness-of-fit v2 test to see genotyping error and population
stratification, P < 0.05 was considered significant deviation. To
assess the relative contributions of predictors of SVR, stepwise
logistic regression analysis was applied, Odds ratio (OR) were cal-
culated and given within the 95% confidence interval (CI). All sta-
tistical calculations were done using computer programs SPSS
 M.M. Fathy et al. / Cytokine xxx (2015) xxx–xxx 3

(Statistical Package for the Social Science; SPSS Inc., Chicago, IL, Table 2
USA) version 15 for Microsoft Windows. Univariate analysis of factors associated with SVR.

SVR n = 76 No SVR P-
n = 77 value
4. Results
Age (years), mean (SD) 40 (10) 46 (9) <0.001
Age > 45 years n (%) 31 (41) 50 (65) 0.003
4.1. Patients’ profiles and virological responses
Gender male n (%) 53 (70) 31 (40) <0.001
Female n (%) 23 (30) 46 (60)
The distribution of IL-28B (rs8099917) genotypes among all Body Mass Index (kg/m2), mean (SD) 26 (3) 28 (4) 0.001
patients were in concordance with Hardy–Weinberg equilibrium, Body Mass Index > 25 kg/m2, n (%) 50 (66) 59 (77) 0.13
P = 0.46. TSH mlU/L, median (25th-75th)* 1.5 (1–2.3) 1.5 (1.9–2.5) 0.97
Hemoglobin g/dl, mean (SD) 13.7 (1.4) 13.4 (1.3) 0.07
The baseline demographic, biochemical and virological features
WBC (103/mm3), mean (SD) 6.6 (1.9) 5.8 (1.7) 0.004
of the patients are shown in Table 1. The rates of SVR, <2 log Platelet count (103/mm3), mean (SD) 193 (49) 210 (67) 0.06
decrease in viral load at 12th wk, detectable viremia at 24th wk, AST U/L, median (25th–75th)* 40 (34–55) 57 (34–79) <0.001
relapse after end of treatment response were 49.7%, 16.3%, 17% ALT U/L, median (25th–75th)* 44 (32–60) 63 (35–84) <0.001
Albumin g/dl, mean (SD) 3.9 (0.3) 4.0 (0.3) 0.25
and 17% respectively.
Prothrombin conc.%, mean (SD) 85 (9) 85 (10) 0.88
Baseline HCV RNA103IU/ml, median 237 (77– 284 (66– 0.93
4.2. Factors associated with SVR (25th–75th)* 630) 763)
Baseline HCV RNA > 400,000 IU/ml, n (%) 27 (36) 31 (40) 0.28
Univariate analysis showed that younger age, male sex, lower Activity grading: n (%)
BMI, WBC count, low pretreatment levels of AST and ALT, low activ- A1 46 (61) 24 (31)
A2 30 (39) 51 (66) <0.001
ity grade, less advanced liver fibrosis, and IL-28B (rs8099917) geno-
A3 0 (0) 2 (3)
type all were significantly associated with higher SVR rate (Table 2).
Fibrosis staging: n (%)
Patients with TT genotype had a significantly higher rate of SVR
F1 55 (72) 30 (39)
than G allele carriers (74% vs. 26%, P = 0.004) (Table 2). F2 12 (16) 22 (29) <0.001
Stepwise logistic regression analysis revealed that younger age, F3 9 (12) 25 (32)
male sex, low activity grading and carriers of the favorable TT Type of interferon: n (%)
genotype were significant predictive factors for achievement of a 2a (pegasys) 36 (47) 32 (42) 0.47
SVR (Table 3). TT genotype carriers had 2.8 higher chance for a 2b (peg-interon) 40 (53) 45 (58)
achievement of SVR than G allele carriers (OR = 2.8, 95% CI 1.4– IL-28B (rs8099917) genotype:
5.6, P = 0.004) (Table 3). High pretreatment AST levels and (TT)/(TG + GG), n (%) 56 (74)/20 38 (49)/39 0.004
advanced fibrosis were negative predictors of SVR (OR = 0.5, (26) (51)

P = 0.04; OR: 0.24, P < 0.001 respectively) (Table 3). Bold means Statistically significant (< 0.05).
*
Skewed data were presented as median and interquartile range (25th–75th).
Table 1
Baseline demographic, biochemical and virological characteristics of all patients.
4.3. Characteristics of patients with different IL-B28 genotypes
Age (years), mean (SD) 43 (10)
Gender male n (%) 69 (45)
Analysis of the basic demographic, biochemical and virological
Female n (%) 84 (55)
TSH mlU/L, median (25th–75th)a 1.5 (1–2.5) features revealed that patients with TT genotype had significantly
Hemoglobin g/dl, mean (SD) 13.5 (1.3) lower pretreatment AST and ALT levels than G allele carriers (42 vs.
WBC (103/mm3), mean (SD) 6.2 (1.8) 59 U/L P = 0.004 and 44 vs. 60 U/L P = 0.005 respectively), signifi-
Platelet count (103/mm3), mean (SD) 201 (59) cantly lower degree of activity (A1 54% vs. 32.3% P = 0.029) and sig-
AST U/L, mean (SD) 53 (28)
ALT U/L, mean (SD) 57 (20)
nificantly higher SVR rate (60% vs. 34% P = 0.002) (Fig. 1). Other
Albumin g/dl, mean (SD) 4.0 (0.3) factors did not show significant association with different IL-28B
Prothrombin conc.%, mean (SD) 86 (9.5) genotypes (Table 4).
Baseline HCV RNA103 IU/ml, median (25th–75th)a 260 (72–696)
Activity grading: n (%)
A1(mild) 70 (45.7)
5. Discussion
A2 (moderate) 81 (53)
A3 (severe) 2 (1.3)
Cytokines are among the predominant mechanisms of host
Fibrosis staging: n (%)
F1 (mild) 85 (56) defense against viral infections. Polymorphisms in the regulatory
F2 (moderate) 34 (22) regions of the cytokine genes may influence the expression of these
F3 (severe) 34 (22) genes. In HCV infection, secretion of inappropriate amounts of
IL-28B (rs8099917) genotype: n (%) cytokines may be associated with progression to chronic disease
Wild type (TT) 94 (62)
Minor allele heterozygous (TG) 54 (35) Table 3
Minor allele homozygous (GG) 5 (3) Logistic regression analysis of factors associated with treatment response.

Type of interferon: n (%) Variables ORa 95% CIb P-value

a 2a (pegasys) 68 (44)
Age <45/>45 2.6 1.3–5.1 0.003
a 2b (peg-interon) 85 (56)
Gender: male/female 3.42 1.7–6.6 <0.001
Response to treatment: n (%) AST >40/<40 U/L 0.5 0.26–0.97 0.04
Sustained virological response (SVR) positive 76 (49.7) Activity grading (A1/A2 + A3) 3.3 1.7–6.5 <0.001
SVR negative: 77 (50.3) Advanced liver Fibrosis (F3/F1 + F2) 0.24 0.12–0.48 <0.001
<2 log decrease in viral load at 12 th wk 25 (16.3) IL-28B(rs8099917) (TT/TG + GG) 2.8 1.4–5.6 0.004
Detected vireamia at 24th wk 26 (17)
Relapse after end of treatment response 26 (17) Bold means Statistically significant (< 0.05).
a
OR: Odds Ratio.
a b
Skewed data were presented as median and interquartile range (25th–75th). CI: Confidence Interval.

Please cite this article in press as: Fathy MM et al. Assessment of interleukin 28B genotype as a predictor of response to combined therapy with pegylated
interferon plus ribavirin in HCV infected Egyptian patients. Cytokine (2015), http://dx.doi.org/10.1016/j.cyto.2015.05.003
4 M.M. Fathy et al. / Cytokine xxx (2015) xxx–xxx

90 When studied in different ethnic populations, it has been shown

80
Percent of response (%)

* 66% that allele distributions of IL28B SNPs are different between races
70 60% and ethnic backgrounds [21]. This is thought to partially explain
*
60 49.7% 50.3%
the difference in response rates to current antiviral therapy
40%
50 34%
between races and different HCV genotypes. Suppiah et al. [9] and
40 SVR
Tanaka et al. [11], showed that rs8099917 was the polymorphism
30 No SVR
most strongly associated with SVR in Australian and Japanese
20
chronic HCV patients respectively, all infected with HCV genotype
10
1. Rauch et al. [22] conducted a GWAS on Caucasians infected with
0
All (n=153) TT (n=94) TG/GG (n=59) viral genotypes 1–4, and found that the rs8099917 minor allele was
associated with both progression to chronicity and failure to
Fig. 1. Association between IL-28B (rs8099917) genotype and treatment response. respond to treatment in patients infected with genotypes 1 and 4,
⁄
P = 0.002 the rate of SVR was significantly higher and the rate of No SVR was but not with genotype 2 or 3. Further studies observed that the
significantly lower in patients with TT genotype compared to those with minor
allele carriers (TG/GG).
effect of the IL28B genotype on SVR was weaker in patients infected
with genotype 2 or 3 than in genotype 1 [22,23].
or resistance to IFN treatment [20]. Recent Genome Wide In the present study, 76 out of 153 patients (49.6%) achieved
Association Studies (GWAS) have revealed a strong association of SVR which was comparable to previously reported studies
two SNPs rs8099917 (T/G) and rs12979860(C/T), which are located [7,24,25]. The rs8099917 TT genotype was present in 94 patients
upstream of the IL28B gene (that encodes for IFN-k3) with both (62%), TG genotype was present in 54 patients (35%) and GG geno-
spontaneous clearance and treatment outcomes of PEG-IFN-a plus type was present in only 5 patients (3%). Similar frequencies of
RBV therapy [9–12]. According to previous studies [13,20], exoge- rs8099917 genotype distribution were observed by previous stud-
nous IFN-a increases IFN-k production by HCV-infected hepato- ies [24–28]. Carriers of the rs8099917 TT genotype (major allele)
cytes during IFN-a therapy. IFN-k3 induces antiviral activity by showed a higher rate of SVR compared to TG + GG (minor allele
itself and through the Janus kinase-signal transducer and activator carriers) (74% vs. 26%, P = 0.004). Further, logistic stepwise regres-
of transcription (JAK-STAT) complex, which induces IFN Stimulated sion analysis showed that TT genotype carriers had 2.8 higher
Genes (ISGs) that also have antiviral activity against HCV. The chance for achievement of SVR compared to G allele carriers
amounts of IFN-k produced on HCV-infected human hepatocytes (OR = 2.8, 95% CI 1.4–5.6, P = 0.004).
were larger in liver with a favorable IL28B genotype [13]. These results confirm the previously reported findings con-
veyed by Suppiah et al. [9], Tanaka et al. [11] and Rauch et al.
Table 4 [22] in various populations around the world in a large cohort of
Basic demographic, biochemical and virological features with different IL-28B HCV genotype-1 mono-infected patients. Stättermayer et al. [8]
genotypes. studied the rs8099917 SNP in HCV-4 infected patients and found
TT Genotype TG/GG P- that the major allele (T/T) was associated with higher SVR rate.
(n = 94) Genotype value They also observed that low base line viral load had significant
(n = 59) effects on the SVR rate. Similarly, Antaki el al. [29] found that a
Age (years), mean (SD) 42 (10) 44 (10) 0.12 SVR was achieved in 26% of rs8099917 TG/GG carriers compared
Gender male n (%) 56 (60) 28 (48) 0.14 with 60% of TT carriers (P < 0.0001) in HCV-4 infected patients.
Female n (%) 38 (40) 31 (52)
Similar results were observed by Ragheb et al. [25] and Derbala
Body Mass Index (kg/m2), 27 (3.6) 28 (4) 0.26
mean (SD) et al. [26] both studies carried on Egyptian patients with HCV-4.
Body Mass Index > 25 kg/m2, 65 (69) 44 (75) 0.47 Various viral and host factors have been identified as significant
n (%) determinants of the outcome of IFN-based treatments. In the present
TSH mlU/L, median (25th–75th) 1.6 (1.1–2.4) 1.4 (0.8–2.3) 0.18 study, on univariate analysis, male sex, younger age, BMI, WBC, AST,
Hemoglobin g/dl, mean (SD) 13.7 (1.5) 13 (1.1) 0.13
WBC (103/mm3), mean (SD) 6.1 (1.7) 6.4 (1.9) 0.39
ALT, activity and fibrosis scores and IL-28B rs8099917(TT) genotype
Platelet count (103/mm3), 196 (49) 210 (72) 0.13 were significantly associated with SVR (p < 0.05 in all).
mean (SD) Stättermayer et al. [8] and Kurosaki et al. [30] showed that
AST U/L, median (25th–75th) 42 (32–57) 59 (37–73) 0.004 younger patients had better SVR rates, however there was no dif-
ALT U/L, median (25th–75th) 44 (30–66) 60 (30–83) 0.005
ference in response to treatment between males and females.
Albumin g/dl, mean (SD) 4.0 (0.38) 4.0 (0.4) 0.87
Prothrombin conc.%, mean (SD) 85 (10) 87 (8) 0.27 Obesity is a predictor of disease progression in patients with
Baseline HCV RNA 103IU/ml, 276 (85–718) 179 (62–761) 0.40 chronic HCV; a high BMI has been significantly associated with
median (25th–75th) fibrosis progression [31]. In accordance to previous studies
Baseline HCV RNA > 400,000 IU/ 37 (39) 21 (36) 0.85 [32,33], we observed that lower BMI was significantly associated
ml, n (%)
with SVR (P = 0.001).
Activity grading: n (%) In the present study, lower baseline AST and ALT were associ-
A1 51 (54) 19 (32.3)
A2 42 (44.9) 39 (66) 0.029
ated with SVR (P < 0.001). This was in accordance with previous
A3 1 (1.1) 1 (1.7) studies which demonstrated that the lower baseline AST reflects
Fibrosis staging: n (%)
less severe baseline histopathological parameters in sustained
F1 59 (63) 26 (44) responders [23,34].
F2 17 (18) 17 (29) 0.074 Contrary to previous reports [8,24,35], pre-treatment viral load
F3 18 (19) 16 (27) was not found to be associated with SVR in our study. However, it
Type of interferon: n (%) is well known that viral load fluctuates and a single reading of HCV
a 2a (pegasys) 38 (40) 30 (51) 0.207 quantification may not reflect the actual viral load at the time of
a 2b (peg-interon) 56 (60) 29 (49)
treatment, especially if it was assessed at varying intervals from
Response to treatment: n (%) the onset of treatment. It has also been reported that the differ-
SVR, n (%) 56 (60) 20 (34) 0.002
ences in interferon response could be secondary to either a differ-
No SVR, n (%) 38 (40) 39 (66)
ence in the viral virulence and/or replication rate among different
Bold means Statistically significant (< 0.05). HCV genotypes and not the absolute viral load [36].

Please cite this article in press as: Fathy MM et al. Assessment of interleukin 28B genotype as a predictor of response to combined therapy with pegylated
interferon plus ribavirin in HCV infected Egyptian patients. Cytokine (2015), http://dx.doi.org/10.1016/j.cyto.2015.05.003
 M.M. Fathy et al. / Cytokine xxx (2015) xxx–xxx
By studying the relation between the degree of inflammation
and fibrosis stage according to the baseline liver biopsy and
response to therapy, patients with mild activity (A1) showed a
higher SVR rate than patients with moderate to severe activity
(A2&A3) (P < 0.001) and patients with mild fibrosis (F1) showed a
higher SVR rate than patients with moderate to severe fibrosis
(F2&F3) (P < 0.001). The presence of advanced liver fibrosis and cir-
rhosis has been recognized to be associated with lower response
rates to IFN-based treatment [8,22,26].
Interestingly, in the present study, patients carrying the favor-
able TT genotype had significantly lower pretreatment levels of
AST and ALT (42 vs. 59 U/L, P = 0.004, 44 vs. 60 U/L P = 0.005
respectively), also they had higher rates of low activity grading
(A1) (54% vs. 32%, P = 0.029). However, the gene polymorphism
had no association with the baseline HCV viral load which might
influence the treatment response. Yu et al. [37] found that among
HCV-2 infected patients, those with favorable TT genotype had sig-
nificant lower levels of HCV RNA. The exact mechanism underlying
genetic association with viral load remains unclear.
Our findings suggest that the IL28B genotype is a useful base-
line predictor of virological response which should be used for
selecting the treatment decision: whether to treat patients with
combined PEG-IFN/RBV or to wait for more effective future therapy
including direct acting antiviral drugs. Decisions of treatment may
be weighted between the possibility of a response against potential
risk of adverse events and the cost of the therapy, or disease pro-
gression while waiting for future therapy. Further studies are
needed on a larger group of patients to validate the results of our
study and prove the definite role of IL28B gene polymorphism in
decision taking for treatment of patient with chronic HCV.
6. Conclusion
IL-28B gene polymorphism is a significant pre-treatment pre-
dictor of response to combined therapy of PEG-IFN/RBV in chroni-
cally HCV infected Egyptian patients. Thus, analysis of IL-28B
genotype might be used to guide treatment decisions for these
patients particularly in the context of merging therapies and
direct-acting antiviral agents.
Acknowledgements
This study was partially funded by Cairo University with the
Real-time PCR kit for IL28B genotyping.
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