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IL28B single nucleotide polymorphisms in the treatment of hepatitis C

  • Christian M. Lange
    Affiliations
    Klinikum der J.W. Goethe-Universität Frankfurt am Main, Medizinische Klinik 1, Theodor-Stern-Kai 7, 60590 Frankfurt am Main, Germany

    Centre Hospitalier Universitaire Vaudois, University of Lausanne, Rue Bugnon 46, CH-1010 Lausanne, Switzerland
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  • Stefan Zeuzem
    Correspondence
    Corresponding author. Tel.: +49 69 6301 5455; fax: +49 69 6301 6448.
    Affiliations
    Klinikum der J.W. Goethe-Universität Frankfurt am Main, Medizinische Klinik 1, Theodor-Stern-Kai 7, 60590 Frankfurt am Main, Germany
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Open AccessPublished:March 25, 2011DOI:https://doi.org/10.1016/j.jhep.2011.03.006

      Summary

      Recent genome-wide association studies (GWAS) have identified genetic variations near the IL28B gene which are strongly associated with spontaneous and treatment-induced clearance of hepatitis C virus (HCV) infection. Protective IL28B variations are strongly associated with on-treatment viral kinetics and approximately 2-fold increased sustained virologic response (SVR) rates in HCV genotype 1 and 4 patients. In HCV genotype 1 patients, IL28B variations were shown to be the strongest pre-treatment predictor of virologic response. In the treatment of HCV genotype 2 and 3 infected patients, IL28B variations play only a minor role. Preliminary data indicate that IL28B variations are also associated with treatment outcome of regimens, including directly acting antiviral (DAA) agents, though their impact seems to be attenuated compared to standard treatment. Here, we review these important findings and discuss possible implications for clinical decision making in the treatment of HCV infection.

      Abbreviations:

      HCV (hepatitis C virus), HIV (human immunodeficiency virus), SVR (sustained virologic response), RVR (rapid virologic response), EVR (early virologic response), peg (pegylated), IFN (interferon), IL28B (interleukin 28B), IL10 (interleukin-10), STAT (signal transducers and activators of transcription), JAK (janus kinase), ALT (alanine aminotransferase), AST (aspartate aminotransferase), LDL (low-density lipoprotein cholesterol), DAA (directly acting antiviral agents)

      Keywords

      Genome-wide association studies (GWAS) and implications for personalized medicine

      According to various estimations, the sequence of the human genome differs in approximately 0.1% from person to person [
      • Cirulli E.T.
      • Goldstein D.B.
      Uncovering the roles of rare variants in common disease through whole-genome sequencing.
      ,
      • Feero W.G.
      • Guttmacher A.E.
      • Collins F.S.
      Genomic medicine – an updated primer.
      ,
      • Manolio T.A.
      Genomewide association studies and assessment of the risk of disease.
      ]. These inter-individual genetic variations may for example influence regulatory sequences and thereby gene expression, they may impact the splicing of gene products, alter the sequence of non-coding RNAs, or they may directly result in changes of protein-sequences and function [
      • Feero W.G.
      • Guttmacher A.E.
      • Collins F.S.
      Genomic medicine – an updated primer.
      ]. Base-pair variations at a given location with a minor allele frequency of >1% within a population are called single nucleotide polymorphisms (SNPs), in contrast to so called rare variations which are present at lower frequencies [
      • Cirulli E.T.
      • Goldstein D.B.
      Uncovering the roles of rare variants in common disease through whole-genome sequencing.
      ,
      • Feero W.G.
      • Guttmacher A.E.
      • Collins F.S.
      Genomic medicine – an updated primer.
      ]. After the entire sequencing of the human genome, in particular the HapMap Project provided the basis for the conduction of GWAS by drawing a detailed genome-wide map of the organization of SNPs in haplotypes [
      • InternationalHapMapConsortium
      A haplotype map of the human genome.
      ]. Haplotypes are a series of variations/SNPs that tend to be inherited together, and their knowledge allows the selection of marker SNPs throughout the genome for GWAS. As a consequence, SNPs identified in GWAS frequently are not causal variants, but are likely to correlate with a causal variant within a haplotype [
      • Feero W.G.
      • Guttmacher A.E.
      • Collins F.S.
      Genomic medicine – an updated primer.
      ,
      • Goldstein D.B.
      Common genetic variation and human traits.
      ,
      • McCarthy M.I.
      • Abecasis G.R.
      • Cardon L.R.
      • Goldstein D.B.
      • Little J.
      • Ioannidis J.P.
      • et al.
      Genome-wide association studies for complex traits: consensus, uncertainty and challenges.
      ]. In the last years, numerous GWAS have been conducted (listed in http://www.genome.gov/26525384), and many SNPs were associated with complex diseases such as diabetes mellitus, atherosclerosis, or various types of cancer [
      • Feero W.G.
      • Guttmacher A.E.
      • Collins F.S.
      Genomic medicine – an updated primer.
      ,
      • Goldstein D.B.
      Common genetic variation and human traits.
      ,
      • McCarthy M.I.
      • Abecasis G.R.
      • Cardon L.R.
      • Goldstein D.B.
      • Little J.
      • Ioannidis J.P.
      • et al.
      Genome-wide association studies for complex traits: consensus, uncertainty and challenges.
      ]. However, most identified SNPs explained only little of the estimated genetic basis of a disease, and whole-genome sequencing may uncover rare variations with major impact at an individual level. In these regards, the strong association of IL28B variations with spontaneous and treatment-induced clearance in HCV infected patients is an exciting exception. The random approach of GWAS also has a high potential to identify unexpected pathways involved in pathogenesis. In addition, many disease-associated SNPs identified by GWAS are not located within protein-coding regions of the genome (some were even far away from any known coding region), which highlight the regulatory importance of non-coding DNA regions [
      • Manolio T.A.
      Genomewide association studies and assessment of the risk of disease.
      ,
      • Cheng J.
      • Kapranov P.
      • Drenkow J.
      • Dike S.
      • Brubaker S.
      • Patel S.
      • et al.
      Transcriptional maps of 10 human chromosomes at 5-nucleotide resolution.
      ]. However, GWAS can assess only part of genetic (and no epigenetic) variations between individuals, and important inter-individual differences such as copy-number variations, alternative splicing variations, or deletions may be only uncovered by next-generation technologies like whole-genome or RNA sequencing [
      • Cirulli E.T.
      • Goldstein D.B.
      Uncovering the roles of rare variants in common disease through whole-genome sequencing.
      ,
      • Cheng J.
      • Kapranov P.
      • Drenkow J.
      • Dike S.
      • Brubaker S.
      • Patel S.
      • et al.
      Transcriptional maps of 10 human chromosomes at 5-nucleotide resolution.
      ,
      • Kidd J.M.
      • Cooper G.M.
      • Donahue W.F.
      • Hayden H.S.
      • Sampas N.
      • Graves T.
      • et al.
      Mapping and sequencing of structural variation from eight human genomes.
      ,
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      • Turner E.H.
      • Robertson P.D.
      • Flygare S.D.
      • Bigham A.W.
      • Lee C.
      • et al.
      Targeted capture and massively parallel sequencing of 12 human exomes.
      ,
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      • Ishikawa S.
      • Fitch K.R.
      • Feuk L.
      • Perry G.H.
      • Andrews T.D.
      • et al.
      Global variation in copy number in the human genome.
      ].

      Biology of interferon-λ (IFN-λ)

      The IL28B gene encodes for interferon-λ3 (IFN-λ3), which constitutes the IFN-λ family together with IFN-λ1 (encoded by IL29) and IFN-λ2 (encoded by IL28A). The IL28A, IL28B, and IL29 genes are located on chromosome 19 in close proximity [
      • Ank N.
      • Paludan S.R.
      Type III IFNs: new layers of complexity in innate antiviral immunity.
      ]. Interferons (IFNs) are categorized into three different families, type 1 IFNs (mainly IFN-α, -β), type 2 IFNs (only IFN-γ), and type 3 IFNs (IFN-λ1–3) [
      • Kawai T.
      • Akira S.
      Innate immune recognition of viral infection.
      ]. Due to their molecular structure, type 3 IFNs belong to the interleukin-10 (IL-10) superfamily, but functionally they are closely related to type 1 IFNs which play a major role in antiviral immunity [
      • Gad H.H.
      • Dellgren C.
      • Hamming O.J.
      • Vends S.
      • Paludan S.R.
      • Hartmann R.
      Interferon-lambda is functionally an interferon but structurally related to the interleukin-10 family.
      ]. Viral infection is sensed in cells by pattern recognition receptors such as toll-like receptors (TLR) or retinoic acid-inducible gene I (RIG-I)-like-helicases, which lead to a signal cascade inducing the interferon response factors 3 or 7 (IRF3, IRF7) [
      • Kawai T.
      • Akira S.
      Innate immune recognition of viral infection.
      ]. Importantly, IFN-λ2/3 and IFN-α expression is induced by IRF7, and IFN-λ1 and IFN-β expression is induced by IRF3 and IRF7 [
      • Li M.
      • Liu X.
      • Zhou Y.
      • Su S.B.
      Interferon-lambdas: the modulators of antivirus, antitumor, and immune responses.
      ,
      • Onoguchi K.
      • Yoneyama M.
      • Takemura A.
      • Akira S.
      • Taniguchi T.
      • Namiki H.
      • et al.
      Viral infections activate types I and III interferon genes through a common mechanism.
      ]. via signaling through the JAK-STAT pathway, both IFN-α/β and IFN-λ1–3 induce a large number of widely overlapping interferon-stimulated genes (ISGs), which orchestrate an antiviral cellular state, Fig. 1 [
      • Ank N.
      • West H.
      • Bartholdy C.
      • Eriksson K.
      • Thomsen A.R.
      • Paludan S.R.
      Lambda interferon (IFN-lambda), a type III IFN, is induced by viruses and IFNs and displays potent antiviral activity against select virus infections in vivo.
      ,
      • Doyle S.E.
      • Schreckhise H.
      • Khuu-Duong K.
      • Henderson K.
      • Rosler R.
      • Storey H.
      • et al.
      Interleukin-29 uses a type 1 interferon-like program to promote antiviral responses in human hepatocytes.
      ,

      Zhang L, Jilg N, Shao RX, Lin W, Fusco DN, Zhao H, et al. IL28B inhibits Hepatitis C virus replication through the JAK-STAT pathway. J Hepatol 2010, epub ahead of print.

      ]. However, IFN-α/β and IFN-λ1–3 engage completely different transmembrane receptors, the IFN-α receptor (IFNAR) complex, and the heterodimeric IL28-Rα/IL-10R2 receptor complex, respectively [
      • Kotenko S.V.
      • Gallagher G.
      • Baurin V.V.
      • Lewis-Antes A.
      • Shen M.
      • Shah N.K.
      • et al.
      IFN-lambdas mediate antiviral protection through a distinct class II cytokine receptor complex.
      ]. Though type 1 and type 3 IFN-signalings converge in the JAK-STAT pathway, their binding to different receptors may result in different kinetics of ISG expression [
      • Marcello T.
      • Grakoui A.
      • Barba-Spaeth G.
      • Machlin E.S.
      • Kotenko S.V.
      • MacDonald M.R.
      • et al.
      Interferons alpha and lambda inhibit hepatitis C virus replication with distinct signal transduction and gene regulation kinetics.
      ]. Moreover, tissue distributions of the IFNAR and the IL28-Rα/IL-10R2 receptor complexes differ significantly [
      • Li M.
      • Liu X.
      • Zhou Y.
      • Su S.B.
      Interferon-lambdas: the modulators of antivirus, antitumor, and immune responses.
      ]. In humans, IL28-Rα/IL-10R2 expression is restricted to hepatocytes, epithelial cells, and plasmacytoid dendritic cells, whereas IFNAR is broadly expressed in numerous tissues. The cellular sources of IFN-λ are predominantly plasmacytoid dendritic cells, probably macrophages (including Kupffer cells), and potentially other cell types like liver sinusoidal endothelial cells. In contrast to humans, mouse hepatocytes do not respond to IFN-λ signaling [
      • Li M.
      • Liu X.
      • Zhou Y.
      • Su S.B.
      Interferon-lambdas: the modulators of antivirus, antitumor, and immune responses.
      ].
      Figure thumbnail gr1
      Fig. 1IFN-α/β and IFN-λ signaling pathways. IFN-α/β and IFN-λ are both induced by IRF7/IRF3, and they are signaling through the JAK/STAT pathway to induce a large number of widely overlapping interferon-stimulated genes (ISGs). However, IFN-α/β and IFN-λ engage completely different receptors, which also differ significantly in their tissue distribution. Plasmacytoid dendritic cells are a main cellular source of IFN-λ, whereas IFN-α/β is broadly expressed. Please note that IFN-α/β and IFN-λ can also act in a paracrine manner, not just in the autocrine way illustrated in this Figure. IL28B genetic variations may alter for example expression, stability, or receptor binding of IFN-λ3. IFN, interferon; MAVS, mitochondrial anti-viral signaling protein; RIG-I, retinoic acid-inducible gene I; TRIF, Toll-IL-1 receptor domain-containing adaptor inducing IFN-β; IRF, interferon-response factor; TLR, toll-like receptor; IFNAR, IFN-α receptor complex; Jak, janus kinase; TYK, tyrosine kinase; STAT, signal transducers and activators of transcription; ISG, interferon-stimulated genes.
      It is important to keep in mind that nearly all IL28B variations associated with spontaneous and treatment-induced clearance are not located within coding regions, but only in close proximity of the IL28B gene. By fine mapping strategies only, a non-synonymous coding variation (rs8103142) in exon 2 of IL28B was identified, but thus far no functional differences between the resulting IFN-λ3 variants (Lys70Arg) could be shown [
      • Urban T.J.
      • Thompson A.J.
      • Bradrick S.S.
      • Fellay J.
      • Schuppan D.
      • Cronin K.D.
      • et al.
      IL28B genotype is associated with differential expression of intrahepatic interferon-stimulated genes in patients with chronic hepatitis C.
      ]. Moreover, data on expression levels of INF-λ3 in patients with different IL28B genotypes remain conflicting, and only some studies found higher INF-λ3 mRNA or protein levels in the liver or blood of patients with good-response IL28B alleles [
      • Suppiah V.
      • Moldovan M.
      • Ahlenstiel G.
      • Berg T.
      • Weltman M.
      • Abate M.L.
      • et al.
      IL28B is associated with response to chronic hepatitis C interferon-alpha and ribavirin therapy.
      ,
      • Urban T.J.
      • Thompson A.J.
      • Bradrick S.S.
      • Fellay J.
      • Schuppan D.
      • Cronin K.D.
      • et al.
      IL28B genotype is associated with differential expression of intrahepatic interferon-stimulated genes in patients with chronic hepatitis C.
      ,
      • Fukuhara T.
      • Taketomi A.
      • Motomura T.
      • Okano S.
      • Ninomiya A.
      • Abe T.
      • et al.
      Variants in IL28B in liver recipients and donors correlate with response to peg-interferon and ribavirin therapy for recurrent hepatitis C.
      ,
      • Honda M.
      • Sakai A.
      • Yamashita T.
      • Nakamoto Y.
      • Mizukoshi E.
      • Sakai Y.
      • et al.
      Hepatic ISG expression is associated with genetic variation in interleukin 28B and the outcome of IFN therapy for chronic hepatitis C.
      ,
      • Langhans B.
      • Kupfer B.
      • Braunschweiger I.
      • Arndt S.
      • Schulte W.
      • Nischalke H.D.
      • et al.
      Interferon-lambda serum levels in hepatitis C.
      ]. Thus, the highly suggestive link between IL28B variations and IFN-λ3 signaling remains to be proven.

      IL28B SNPs and spontaneous clearance of HCV infection

      Spontaneous clearance of HCV occurs in only 15–50% of all HCV infected individuals, while the majority of patients develop a chronic infection. Thomas et al. found that IL28B rs12979860 is strongly associated with the chance to clear HCV spontaneously in populations of African or European ancestry, with an approximately three times higher clearance rate in individuals with the rs12979860 genotype C/C versus C/T, T/T [
      • Thomas D.L.
      • Thio C.L.
      • Martin M.P.
      • Qi Y.
      • Ge D.
      • O’Huigin C.
      • et al.
      Genetic variation in IL28B and spontaneous clearance of hepatitis C virus.
      ]. The same IL28B alleles were associated in the very first GWAS on treatment response with SVR versus non-SVR, respectively [
      • Ge D.
      • Fellay J.
      • Thompson A.J.
      • Simon J.S.
      • Shianna K.V.
      • Urban T.J.
      • et al.
      Genetic variation in IL28B predicts hepatitis C treatment-induced viral clearance.
      ]. In the Swiss hepatitis C cohort, the association between IL28B variations (top hit rs809917) and spontaneous clearance reached genome-wide significance [
      • Rauch A.
      • Kutalik Z.
      • Descombes P.
      • Cai T.
      • Di Iulio J.
      • Mueller T.
      • et al.
      Genetic variation in IL28B is associated with chronic hepatitis C and treatment failure: a genome-wide association study.
      ]. In the homogeneous German anti-D cohort, comprising exclusively women infected with HCV genotype 1b, rs12979860 C/C versus C/T, T/T was not only associated with spontaneous clearance, but also with jaundice, providing a link to the clinical observation that patients with symptomatic acute hepatitis C are more likely to clear the virus than those with silent disease [
      • Tillmann H.L.
      • Thompson A.J.
      • Patel K.
      • Wiese M.
      • Tenckhoff H.
      • Nischalke H.D.
      • et al.
      A polymorphism near IL28B is associated with spontaneous clearance of acute hepatitis C virus and jaundice.
      ]. Intriguingly, the study by Thomas et al. also characterized the frequencies of rs12979860 C versus T alleles in several world-wide populations [
      • Thomas D.L.
      • Thio C.L.
      • Martin M.P.
      • Qi Y.
      • Ge D.
      • O’Huigin C.
      • et al.
      Genetic variation in IL28B and spontaneous clearance of hepatitis C virus.
      ]. The highest frequencies of the protective C allele were observed in East Asia, frequencies were intermediate in Europe, and lowest in Africa. This geographic distribution of C/T allele frequencies largely parallel reported rates of spontaneous and treatment induced clearance in these areas.
      Interestingly, several [
      • Rauch A.
      • Kutalik Z.
      • Descombes P.
      • Cai T.
      • Di Iulio J.
      • Mueller T.
      • et al.
      Genetic variation in IL28B is associated with chronic hepatitis C and treatment failure: a genome-wide association study.
      ,
      • McCarthy J.J.
      • Li J.H.
      • Thompson A.
      • Suchindran S.
      • Lao X.Q.
      • Patel K.
      • et al.
      Replicated association between an IL28B gene variant and a sustained response to pegylated interferon and ribavirin.
      ,
      • Montes-Cano M.A.
      • Garcia-Lozano J.R.
      • Abad-Molina C.
      • Romero-Gomez M.
      • Barroso N.
      • Aguilar-Reina J.
      • et al.
      Interleukin-28B genetic variants and hepatitis virus infection by different viral genotypes.
      ,
      • Sarrazin C.
      • Susser S.
      • Doehring A.
      • Lange C.M.
      • Muller T.
      • Schlecker C.
      • et al.
      Importance of IL28B gene polymorphisms in hepatitis C virus genotype 2 and 3 infected patients.
      ,
      • Lindh M.
      • Lagging M.
      • Norkrans G.
      • Hellstrand K.
      Observed and calculated interleukin-28B genotype frequencies in hepatitis C virus infection.
      ] but not all [
      • Mangia A.
      • Thompson A.J.
      • Santoro R.
      • Piazzolla V.
      • Tillmann H.L.
      • Patel K.
      • et al.
      An IL28B polymorphism determines treatment response of hepatitis C virus genotype 2 or 3 patients who do not achieve a rapid virologic response.
      ] studies reported lower IL28B rs12979860 C versus T allele frequencies in patients infected with HCV genotypes 1 versus 2 or 3. For example, in a large German cohort the rs12979860 C/C genotype was found in 42.7% of HCV genotype 2 and 3 patients, in 33.9% of HCV genotype 1 patients, and in 49% of uninfected control individuals [
      • Sarrazin C.
      • Susser S.
      • Doehring A.
      • Lange C.M.
      • Muller T.
      • Schlecker C.
      • et al.
      Importance of IL28B gene polymorphisms in hepatitis C virus genotype 2 and 3 infected patients.
      ]. The frequency of the protective C allele in HCV genotype 2/3 patients, which is close to that of uninfected individuals, allows speculating that protective IL28B variations provide a more substantial advantage in acute HCV genotype 1 versus 2/3 infection, or that spontaneous clearance rates are higher in HCV genotype 1 infection. Appropriate studies are required to address this issue.

      Implications for the treatment of acute hepatitis C

      Early treatment of acute hepatitis C with 24 weeks of monotherapy with (pegylated) IFN-α results in HCV eradication in the majority of patients. However, the optimal time-point for treatment initiation is still under debate, since late treatment initiation may reduce the chance of HCV clearance, whereas very early treatment initiation will lead to treatment of a number of patients who would have cleared HCV spontaneously [
      • Sarrazin C.
      • Berg T.
      • Ross R.S.
      • Schirmacher P.
      • Wedemeyer H.
      • Neumann U.
      • et al.
      Prophylaxis, diagnosis and therapy of hepatitis C virus (HCV) infection: the German guidelines on the management of HCV infection.
      ]. Due to significant side-effects and costs of IFN-α based therapy, IL28B genetic testing may help to identify those individuals with low chance of spontaneous clearance for early treatment initiation. Whether IL28B SNPs also impact treatment response in acute hepatitis C has not been comprehensively investigated. However, a study in approximately 50 patients with recent HCV infection (up to 24 months after sero-conversion) found no effect of IL28B variations on the outcome of IFN-α based therapy [
      • Grebely J.
      • Petoumenos K.
      • Hellard M.
      • Matthews G.V.
      • Suppiah V.
      • Applegate T.
      • et al.
      Potential role for interleukin-28B genotype in treatment decision-making in recent hepatitis C virus infection.
      ]. In addition, no significant impact of IL28B variations on the outcome of treatment of acute hepatitis C in human immunodeficiency virus (HIV) coinfected patients was observed [
      • Nattermann J.
      • Vogel M.
      • Nischalke H.D.
      • Danta M.
      • Mauss S.
      • Stellbrink H.J.
      • et al.
      Genetic Variation in IL28B and Treatment-Induced Clearance of Hepatitis C Virus in HIV-Positive Patients With Acute and Chronic Hepatitis C.
      ].

      IL28B SNPs and treatment-induced clearance of chronic HCV genotype 1 infection

      In 2009 and 2010, four independent GWAS on response to treatment of chronic hepatitis C with pegIFN-α and ribavirin were published [
      • Ge D.
      • Fellay J.
      • Thompson A.J.
      • Simon J.S.
      • Shianna K.V.
      • Urban T.J.
      • et al.
      Genetic variation in IL28B predicts hepatitis C treatment-induced viral clearance.
      ,
      • Rauch A.
      • Kutalik Z.
      • Descombes P.
      • Cai T.
      • Di Iulio J.
      • Mueller T.
      • et al.
      Genetic variation in IL28B is associated with chronic hepatitis C and treatment failure: a genome-wide association study.
      ,
      • Suppiah V.
      • Moldovan M.
      • Ahlenstiel G.
      • Berg T.
      • Weltman M.
      • Abate M.L.
      • et al.
      IL28B is associated with response to chronic hepatitis C interferon-alpha and ribavirin therapy.
      ,
      • Tanaka Y.
      • Nishida N.
      • Sugiyama M.
      • Kurosaki M.
      • Matsuura K.
      • Sakamoto N.
      • et al.
      Genome-wide association of IL28B with response to pegylated interferon-alpha and ribavirin therapy for chronic hepatitis C.
      ]. In each of these GWAS, only SNPs around the IL28B gene reached genome-wide significance for the association with treatment outcome. All identified IL28B SNPs correlate with each other and can, therefore, be clustered in haplotypes. Consistently, rs12979860 C (good-response allele) versus T (poor-response allele) and rs809917 T (good-response allele) versus G (poor-response allele) showed the strongest association with SVR of thus far characterized SNPs. Both SNPs are in strong linkage disequilibrium, but allele frequencies in particular of rs809917 differ somewhat between world-wide populations. Therefore, the predictive power of both SNPs may vary between different cohorts, as for example rs809917 was only a weak predictor of SVR in African–American patients [
      • Ge D.
      • Fellay J.
      • Thompson A.J.
      • Simon J.S.
      • Shianna K.V.
      • Urban T.J.
      • et al.
      Genetic variation in IL28B predicts hepatitis C treatment-induced viral clearance.
      ].
      In the largest GWAS including more than 1000 HCV genotype 1 patients, rs12979860 C/C versus T/T was associated with a more than 2-fold higher chance to achieve a SVR [
      • Ge D.
      • Fellay J.
      • Thompson A.J.
      • Simon J.S.
      • Shianna K.V.
      • Urban T.J.
      • et al.
      Genetic variation in IL28B predicts hepatitis C treatment-induced viral clearance.
      ]. SVR rates according to the heterozygous genotype rs12979860 C/T were only slightly better compared to rs12979860 T/T. Importantly, these findings were comparable in patients of European–American, African–American, and Hispanic ancestry, though SVR rates in African–American patients generally were lower compared to European–Americans. However, the IL28B genetic background was a stronger predictor of SVR than ancestry, and different SVR rates between these ethnic groups were explained to approximately 50% by lower rs12979860 C/C frequencies in African–Americans. In the three other GWAS, comparable associations between IL28B variations and treatment-response were observed in populations of Asian, European, and European–Australian ancestry, (Table 1) [
      • Rauch A.
      • Kutalik Z.
      • Descombes P.
      • Cai T.
      • Di Iulio J.
      • Mueller T.
      • et al.
      Genetic variation in IL28B is associated with chronic hepatitis C and treatment failure: a genome-wide association study.
      ,
      • Suppiah V.
      • Moldovan M.
      • Ahlenstiel G.
      • Berg T.
      • Weltman M.
      • Abate M.L.
      • et al.
      IL28B is associated with response to chronic hepatitis C interferon-alpha and ribavirin therapy.
      ,
      • Tanaka Y.
      • Nishida N.
      • Sugiyama M.
      • Kurosaki M.
      • Matsuura K.
      • Sakamoto N.
      • et al.
      Genome-wide association of IL28B with response to pegylated interferon-alpha and ribavirin therapy for chronic hepatitis C.
      ].
      Table 1Characteristics of four pivotal GWAS on treatment-induced HCV clearance.
      ∗These studies included replication cohorts. In the present Table, only numbers and results for the initial GWAS are shown.
      ∗∗Additional patients were assessed for spontaneous HCV clearance.
      ∗∗∗Patients were excluded who received <80% of recommended dose for pegIFN-α/ribavirin throughout treatment period (Ge et al.) or during the first 12 weeks (Tanaka et al., Rauch et al.).
      #Different platforms were used for GWAS. Therefore, not all SNPs were equally represented in each GWAS. Only SNPs that reached genome-wide significance are shown.
      ##SNPs were also associated with spontaneous clearance in this study.
      Several studies revealed comparable associations between IL28B variations and treatment-induced HCV clearance in patients co-infected with HIV [
      • Rauch A.
      • Kutalik Z.
      • Descombes P.
      • Cai T.
      • Di Iulio J.
      • Mueller T.
      • et al.
      Genetic variation in IL28B is associated with chronic hepatitis C and treatment failure: a genome-wide association study.
      ,
      • Nattermann J.
      • Vogel M.
      • Nischalke H.D.
      • Danta M.
      • Mauss S.
      • Stellbrink H.J.
      • et al.
      Genetic Variation in IL28B and Treatment-Induced Clearance of Hepatitis C Virus in HIV-Positive Patients With Acute and Chronic Hepatitis C.
      ,
      • Pineda J.A.
      • Caruz A.
      • Rivero A.
      • Neukam K.
      • Salas I.
      • Camacho A.
      • et al.
      Prediction of response to pegylated interferon plus ribavirin by IL28B gene variation in patients coinfected with HIV and hepatitis C virus.
      ,
      • Rallon N.I.
      • Naggie S.
      • Benito J.M.
      • Medrano J.
      • Restrepo C.
      • Goldstein D.
      • et al.
      Association of a single nucleotide polymorphism near the interleukin-28B gene with response to hepatitis C therapy in HIV/hepatitis C virus-coinfected patients.
      ]. For example, one study in HIV co-infected patients reported 2–3-fold higher SVR rates for HCV genotypes 1 and 4 according to rs12979860 C/C versus C/T, T/T, but no difference for HCV genotype 3 according to rs12979860 C/C versus C/T, T/T [
      • Rallon N.I.
      • Naggie S.
      • Benito J.M.
      • Medrano J.
      • Restrepo C.
      • Goldstein D.
      • et al.
      Association of a single nucleotide polymorphism near the interleukin-28B gene with response to hepatitis C therapy in HIV/hepatitis C virus-coinfected patients.
      ].

      IL28B SNPs and established predictors of virologic response

      Thompson et al. performed an intention-to-treat analysis of 1671 patients of the IDEAL study and of additional 67 patients of another clinical trial (adherent patients of this cohort were reported in the GWAS by Ge et al.) [
      • Thompson A.J.
      • Muir A.J.
      • Sulkowski M.S.
      • Ge D.
      • Fellay J.
      • Shianna K.V.
      • et al.
      Interleukin-28B polymorphism improves viral kinetics and is the strongest pretreatment predictor of sustained virologic response in genotype 1 hepatitis C virus.
      ]. In these HCV genotype 1 patients, IL28B rs12979860 was the strongest pre-treatment predictor of virologic response and explained approximately 15% of inter-individual variability of SVR. In contrast, other significant pre-treatment predictors of treatment outcome (hepatic fibrosis stage, baseline viral load, fasting glucose level, body mass index (BMI), ethnic background) each accounted for not more than 5% of variability of SVR.
      In this study, Thompson et al. also demonstrated that the rs12979860 genotype is strongly associated with on-treatment viral kinetics. At treatment week 2, Caucasian patients with the rs12979860 C/C, C/T, and T/T genotype had experienced a median 2.6, 0.9, and 0.6 log10 HCV RNA decline, respectively. In African–American patients, similar tendencies were observed, but viral load reductions were slightly lower in all rs12979860 groups. Despite ongoing decreases, differences in HCV RNA serum concentration were of similar magnitude at treatment week 4 and 12, compared to treatment week 2. Thus, IL28B variations are associated with (very) early on-treatment viral kinetics, which lead to increased rapid virologic response (RVR) and early virologic response (EVR) rates. When comparing the predictive value of rs12979860 and RVR, Thompson et al. found that the rs12979860 genotype had a higher sensitivity and a higher negative predictive value of SVR, but RVR had the highest positive predictive value and a better specificity of SVR. In addition, patients with RVR achieved high SVR rates independently of the rs12979860 background. In contrast, the rs12979860 genotype was strongly associated with SVR in patients who did not achieve a RVR. However, it is important to keep in mind that RVR is rare in HCV genotype 1 patients, and RVR itself is associated with rs12979860 C/C.
      The findings by Thompson et al. were extended by another study that analyzed the influence of IL28B variations on viral kinetics during the very first days of treatment with pegIFN-α and ribavirin [

      Bochud PY, Bibert S, Negro F, Haagmans B, Soulier A, Ferrari C, et al. IL28B polymorphisms predict reduction of HCV RNA from the first day of therapy in chronic hepatitis C. J Hepatol 2011;epub ahead of print.

      ]. In this study, IL28B variations were strongly associated with differences in the 1st phase HCV RNA decline between day 1 and 4 of therapy. IL28B variations were also associated with the 2nd phase decline of HCV RNA from the second throughout the fourth week of therapy, but this association was no longer significant after adjustment for the 1st phase decline. Thus, associations of IL28B variations with RVR and SVR appear to be related to a strong impact on very early viral kinetics during treatment, which reflects the antiviral effectiveness of IFN-α. Whether IL28B variations are also associated with residual differences in the 2nd phase HCV RNA decline is unclear at the moment.

      IL28B SNPs in patients infected with HCV genotypes 2, 3, and 4

      HCV genotype 1 patients with a good-response IL28B genetic background still achieve somewhat lower SVR rates than HCV genotype 2 and 3 patients in general. Thus, it is obvious that the HCV genotype will remain an important independent pre-treatment predictor of virologic response. Directly acting antiviral (DAA) agents will improve treatment options for HCV genotype 1 patients in the very near future, whereas the development of agents effective against other HCV genotypes is at an earlier stage [
      • Lange C.M.
      • Sarrazin C.
      • Zeuzem S.
      Review article: specifically targeted anti-viral therapy for hepatitis C – a new era in therapy.
      ]. Thus, the impact of IL28B variations on the outcome of standard therapy in HCV non-genotype 1 infected patients requires attention.
      Several studies clearly showed that associations between IL28B variations and RVR and SVR are comparable between HCV genotype 1 and 4 patients [
      • Rauch A.
      • Kutalik Z.
      • Descombes P.
      • Cai T.
      • Di Iulio J.
      • Mueller T.
      • et al.
      Genetic variation in IL28B is associated with chronic hepatitis C and treatment failure: a genome-wide association study.
      ,
      • Rallon N.I.
      • Naggie S.
      • Benito J.M.
      • Medrano J.
      • Restrepo C.
      • Goldstein D.
      • et al.
      Association of a single nucleotide polymorphism near the interleukin-28B gene with response to hepatitis C therapy in HIV/hepatitis C virus-coinfected patients.
      ,
      • Stattermayer A.F.
      • Stauber R.
      • Hofer H.
      • Rutter K.
      • Beinhardt S.
      • Scherzer T.M.
      • et al.
      Impact of IL28B Genotype on the Early and Sustained Virologic Response in Treatment-Naive Patients With Chronic Hepatitis C.
      ]. In contrast, results from studies in HCV genotype 2 and 3 patients remain conflicting. Mangia et al. analyzed the role of IL28B rs12979860 in treatment response in a large cohort of HCV genotype 2 and 3 patients who were treated with pegIFN-α and ribavirin either for 24 weeks (standard treatment), or for 12 or 24 weeks, according to whether they achieved an RVR [
      • Mangia A.
      • Thompson A.J.
      • Santoro R.
      • Piazzolla V.
      • Tillmann H.L.
      • Patel K.
      • et al.
      An IL28B polymorphism determines treatment response of hepatitis C virus genotype 2 or 3 patients who do not achieve a rapid virologic response.
      ]. Rs12979860 was neither associated with RVR and SVR in the standard treatment arm, nor with SVR in patients who achieved a RVR and who were treated for 12 weeks only. However, in patients who did not achieve an RVR and who received 24 weeks of treatment, rs12979860 was significantly associated with SVR (SVR rates 87%, 67%, and 29% for rs12979860 C/C, C/T, and T/T, respectively). In contrast to Mangia et al., Sarrazin et al. found an association of rs12979860 with RVR in HCV genotype 2 and 3 patients, and also with SVR in those patients who did achieve a RVR [
      • Sarrazin C.
      • Susser S.
      • Doehring A.
      • Lange C.M.
      • Muller T.
      • Schlecker C.
      • et al.
      Importance of IL28B gene polymorphisms in hepatitis C virus genotype 2 and 3 infected patients.
      ]. In the latter study, the proportion and absolute number of HCV genotype 3 versus 2 patients was much higher compared to the Mangia study, but treatment regimens were more heterogeneous. Several other studies yielded mixed results, though most studies failed to show a significant association of IL28B variations with SVR [
      • Rauch A.
      • Kutalik Z.
      • Descombes P.
      • Cai T.
      • Di Iulio J.
      • Mueller T.
      • et al.
      Genetic variation in IL28B is associated with chronic hepatitis C and treatment failure: a genome-wide association study.
      ,
      • McCarthy J.J.
      • Li J.H.
      • Thompson A.
      • Suchindran S.
      • Lao X.Q.
      • Patel K.
      • et al.
      Replicated association between an IL28B gene variant and a sustained response to pegylated interferon and ribavirin.
      ,
      • Nattermann J.
      • Vogel M.
      • Nischalke H.D.
      • Danta M.
      • Mauss S.
      • Stellbrink H.J.
      • et al.
      Genetic Variation in IL28B and Treatment-Induced Clearance of Hepatitis C Virus in HIV-Positive Patients With Acute and Chronic Hepatitis C.
      ,
      • Kawaoka T.
      • Hayes C.N.
      • Ohishi W.
      • Ochi H.
      • Maekawa T.
      • Abe H.
      • et al.
      Predictive value of the IL28B polymorphism on the effect of interferon therapy in chronic hepatitis C patients with genotypes 2a and 2b.
      ,
      • Moghaddam A.
      • Melum E.
      • Reinton N.
      • Ring-Larsen H.
      • Verbaan V.
      • Bjøro K.
      • et al.
      L28B genetic variation and treatment response in patients with HCV genotype 3 infection.
      ,
      • Scherzer T.M.
      • Hofer H.
      • Staettermayer A.F.
      • Rutter K.
      • Beinhardt S.
      • Steindl-Munda P.
      • et al.
      Early virologic response and IL28B polymorphisms in patients with chronic hepatitis C genotype 3 treated with peginterferon alfa-2a and ribavirin.
      ].

      IL28B SNPs and treatment with directly acting antiviral agents

      With the expected approval of the HCV NS3-4A protease inhibitors telaprevir and boceprevir in 2011, numerous patients with HCV genotype 1 infection will likely be subjected to triple therapy regimens, including telaprevir or boceprevir in combination with pegIFN-alfa and ribavirin [
      • Bacon B.R.
      • Gordon S.C.
      • Lawitz E.
      • Marcellin P.
      • Vierling J.M.
      • Zeuzem S.
      • et al.
      HCV respond-2 final results: high sustained virologic response among genotype 1 previous non-responders and relapsers to peginterferon/ribavirin when re-treated with boceprevir plus pegintron (pegintron alfa-2b)/ribavirin.
      ,
      • Hezode C.
      • Forestier N.
      • Dusheiko G.
      • Ferenci P.
      • Pol S.
      • Goeser T.
      • et al.
      Telaprevir and peginterferon with or without ribavirin for chronic HCV infection.
      ,
      • Jacobson I.M.
      • McHutchison J.G.
      • Dusheiko G.
      • Di Bisceglie A.M.
      • Reddy K.R.
      • Bzowej N.H.
      • et al.
      Telaprevir in combination with peginterferon and ribavirin in genotype 1 HCV treatmnet-naive patients: final results of phase 3 ADVANCE study.
      ,
      • Kwo P.Y.
      • Lawitz E.J.
      • McCone J.
      • Schiff E.R.
      • Vierling J.M.
      • Pound D.
      • et al.
      Efficacy of boceprevir, an NS3 protease inhibitor, in combination with peginterferon alfa-2b and ribavirin in treatment-naive patients with genotype 1 hepatitis C infection (SPRINT-1): an open-label, randomised, multicentre phase 2 trial.
      ,
      • McHutchison J.G.
      • Everson G.T.
      • Gordon S.C.
      • Jacobson I.M.
      • Sulkowski M.
      • Kauffman R.
      • et al.
      Telaprevir with peginterferon and ribavirin for chronic HCV genotype 1 infection.
      ,
      • McHutchison J.G.
      • Manns M.P.
      • Muir A.J.
      • Terrault N.A.
      • Jacobson I.M.
      • Afdhal N.H.
      • et al.
      Telaprevir for previously treated chronic HCV infection.
      ,
      • Poordad E.
      • McCone J.
      • Bacon B.
      • Bruno S.
      • Manns M.P.
      • Sulkowski M.
      • et al.
      Boceprevir combined with peginterferon alfa-2B/ribavirin for treatment-naive patients with hepatitis C virus genotype (G) 1: SPRINT-2 final results.
      ,
      • Sherman K.E.
      • Flamm S.L.
      • Afdhal N.H.
      • Nelson D.R.
      • Sulkowski M.S.
      • Everson G.T.
      • et al.
      Telaprevir in combination with peginterferon alfa-2a and ribavirin for 24 or 48 weeks in treatment-naive genotype 1 HCV patients who achieved an extended rapid viral response: final results of phase 3 ILLUMINATE study.
      ]. Other directly acting antiviral (DAA) agents in advanced clinical development include HCV NS5B polymerase inhibitors, HCV NS5A inhibitors, or the cyclophilin A inhibitor Debio-025 (alisporivir) [
      • Lange C.M.
      • Sarrazin C.
      • Zeuzem S.
      Review article: specifically targeted anti-viral therapy for hepatitis C – a new era in therapy.
      ,
      • Sarrazin C.
      • Zeuzem S.
      Resistance to direct antiviral agents in patients with hepatitis C virus infection.
      ]. Preliminary data on the role of IL28B variations in predicting response to triple-therapy regimens are available. A Japanese study analyzed 72 HCV genotype 1 patients who were either treated for 12 weeks with telaprevir plus pegIFN-α and ribavirin (n = 20), or for 12 weeks with telaprevir plus pegIFN-α and ribavirin followed by additional 12 weeks of pegIFN-α and ribavirin only (n = 52) [
      • Akuta N.
      • Suzuki F.
      • Hirakawa M.
      • Kawamura Y.
      • Yatsuji H.
      • Sezaki H.
      • et al.
      Amino acid substitution in hepatitis C virus core region and genetic variation near the interleukin 28B gene predict viral response to telaprevir with peginterferon and ribavirin.
      ]. Overall SVR rates after 12 and 24 weeks of total therapy were 45% and 67%, respectively. In this study, both rs809917 T/T versus G/T, G/G and rs12979860 C/C versus C/T, T/T were associated with a more than 2-fold higher chance to achieve SVR after triple therapy. In contrast, on-treatment viral kinetics during treatment with the HCV protease inhibitor TMC435 in combination with pegIFN-α and ribavirin were only slightly influenced by IL28B variations [
      • Fried M.W.
      • Buti M.
      • Dore G.J.
      • Ferenci P.
      • Jacobson I.M.
      • Marcellin P.
      • et al.
      Efficacy and safety of TMC435 in combination with peginterferon-alfa-2a and ribavirin in treatment-naive genotype-1 HCV patients: 24-week interim results from the PILLAR study.
      ].
      Theoretically, the finding that IL28B genetic variations are strongly associated with differences in early on-treatment viral kinetics during standard treatment suggests a higher risk of resistance development to DAA agents in patients with poor-response IL28B alleles. Since DAA agents differ significantly in their barrier to resistance development, the impact of IL28B SNPs on treatment–response may vary among DAA agents (Fig. 2) [
      • Sarrazin C.
      • Zeuzem S.
      Resistance to direct antiviral agents in patients with hepatitis C virus infection.
      ]. Indeed, a small clinical trial found no significant association of IL28B SNPs with RVR rates during treatment with the nucleotide–analog HCV polymerase inhibitor PSI-7977 in combination with pegIFN-α and ribavirin [
      • McHutchison J.G.
      • Goldstein D.B.
      • Shianna K.V.
      • Lawitz E.
      • Lalezari J.P.
      • Rodriguez-Torres M.
      • et al.
      IL28B SNP geographical distribution and antiviral responses in a 28-day phase 2a trial of PSI-7977 daily dosing plus peg-IFN/RBV.
      ]. In contrast, the rs12979860 genotype was significantly associated with on-treatment virologic response rates during combination therapy with the non-nucleotide HCV polymerase inhibitor ANA598 plus pegIFN-α and ribavirin (RVR rates 80% versus 31% for C/C versus C/T, T/T, respectively) [
      • Muir A.J.
      • Lawitz E.
      • Rodriguez-Torres M.
      • Rustgi V.K.
      • Hassanein M.T.
      • Appelman J.R.
      • et al.
      IL28B polymorphisms and kinetics of antiviral activity for ANA598 in combination with pegylated interferon-alfa2a plus ribavrin in treatment-naive genotype-1 chronic HCV patients.
      ]. Overall, IL28B variations seem to play an attenuated role during triple therapy, but additional research is required before final conclusions can be drawn.
      Figure thumbnail gr2
      Fig. 2Features of emerging directly acting antiviral (DAA) agents for the treatment of chronic hepatitis C. DAA agents differ significantly with respect to HCV genotype coverage, antiviral activity, and barrier to resistance. The risk of resistance development increases with time of active HCV replication during exposure to DAA agents, and may therefore be higher in patients with poor-response IL28B genotype. Intensive research is required to define the precise value of IL28B variations in predicting treatment response to regiments including DAA agents.

      Implications for the treatment of chronic hepatitis C

      IL28B SNPs provide a strong predictive value for the outcome of standard therapy in the difficult to treat HCV genotype 1 and 4 patients. Due to the availability of this information before treatment initiation, IL28B SNPs will likely enrich future decision-making in the management of chronic hepatitis C. Before IL28B genotypes can be definitely included in treatment recommendations, analyses of appropriate randomized controlled clinical trials are mandatory, in which treatment arms are compared after stratification of patients according to IL28B genotype. IL28B SNPs were shown to be the strongest pretreatment-predictor of treatment outcome in HCV genotype 1 patients. Nevertheless, it was estimated that IL28B variations account for “only” about 15% of inter-individual variability of SVR, a finding which supports the importance of additional predictors of treatment response, possibly including more recently established predictors such as vitamin D deficiency, interferon-γ-inducible protein-10 (IP-10) serum levels, or steatosis/insulin resistance [
      • Zeuzem S.
      • Berg T.
      • Moeller B.
      • Hinrichsen H.
      • Mauss S.
      • Wedemeyer H.
      • et al.
      Expert opinion on the treatment of patients with chronic hepatitis C.
      ,
      • Berg T.
      • Sarrazin C.
      • Herrmann E.
      • Hinrichsen H.
      • Gerlach T.
      • Zachoval R.
      • et al.
      Prediction of treatment outcome in patients with chronic hepatitis C: significance of baseline parameters and viral dynamics during therapy.
      ,
      • Thompson A.J.
      • Muir A.J.
      • Sulkowski M.S.
      • Ge D.
      • Fellay J.
      • Shianna K.V.
      • et al.
      Interleukin-28B polymorphism improves viral kinetics and is the strongest pretreatment predictor of sustained virologic response in genotype 1 hepatitis C virus.
      ,
      • Bitetto D.
      • Fattovich G.
      • Fabris C.
      • Ceriani E.
      • Falleti E.
      • Fornasiere E.
      • et al.
      Complementary role of vitamin D deficiency and the IL-28B rs12979860 C/T polymorphism in predicting antiviral response in chronic hepatitis C.
      ,
      • Lange C.M.
      • Bojunga J.
      • Ramos-Lopez E.
      • von Wagner M.
      • Hassler A.
      • Vermehren J.
      • et al.
      Vitamin D deficiency and a CYP27B1–1260 promoter polymorphism are associated with chronic hepatitis C and poor response to interferon-alfa based therapy.
      ,
      • Lange C.M.
      • von Wagner M.
      • Bojunga J.
      • Berg T.
      • Farnik H.
      • Hassler A.
      • et al.
      Serum lipids in European chronic HCV genotype 1 patients during and after treatment with pegylated interferon-alpha-2a and ribavirin.
      ,
      • Petta S.
      • Camma C.
      • Scazzone C.
      • Tripodo C.
      • Di Marco V.
      • Bono A.
      • et al.
      Low vitamin D serum level is related to severe fibrosis and low responsiveness to interferon-based therapy in genotype 1 chronic hepatitis C.
      ,
      • Lagging M.
      • Askarieh G.
      • Negro F.
      • Bibert S.
      • Söderholm J.
      • Westin J.
      • et al.
      ]. In this context, models for the prediction of SVR which include a variety of parameters in combination with IL28B genotype might be of value, since they were shown to achieve higher accuracy in predicting treatment response compared to the IL28B genotype alone [
      • Bitetto D.
      • Fattovich G.
      • Fabris C.
      • Ceriani E.
      • Falleti E.
      • Fornasiere E.
      • et al.
      Complementary role of vitamin D deficiency and the IL-28B rs12979860 C/T polymorphism in predicting antiviral response in chronic hepatitis C.
      ,
      • Medrano J.
      • Neukam K.
      • Rallon N.
      • Rivero A.
      • Resino S.
      • Naggie S.
      • et al.
      Modeling the probability of sustained virological response to Therapy with pegylated interferon plus ribavirin in patients coinfected with hepatitis C virus and HIV.
      ,
      • Darling J.M.
      • Aerssens J.
      • Fanning G.
      • McHutchison J.G.
      • Goldstein D.B.
      • Thompson A.J.
      • et al.
      Quantitation of pretreatment serum interferon-gamma-inducible protein-10 improves the predictive value of an IL28B gene polymorphism for hepatitis C treatment response.
      ,
      • Lagging M.
      • Romero A.I.
      • Westin J.
      • Norkrans G.
      • Dhillon A.P.
      • Pawlotsky J.M.
      • et al.
      IP-10 predicts viral response and therapeutic outcome in difficult-to-treat patients with HCV genotype 1 infection.
      ]. In addition, the role of other genetic factors that influence spontaneous or treatment-induced HCV clearance, such as genes encoding the natural killer (NK) cell receptor KIR2DL3 and its ligand, human leukocyte antigen C group 1 (HLA-C1), should be re-analyzed in relationship to IL28B genotype [
      • Askar M.
      • Avery R.
      • Corey R.
      • Lopez R.
      • Thomas D.
      • Pidwell D.
      • et al.
      Lack of killer immunoglobulin-like receptor 2DS2 (KIR2DS2) and KIR2DL2 is associated with poor responses to therapy of recurrent hepatitis C virus in liver transplant recipients.
      ,
      • Carneiro V.L.
      • Lemaire D.C.
      • Bendicho M.T.
      • Souza S.L.
      • Cavalcante L.N.
      • Angelo A.L.
      • et al.
      Natural killer cell receptor and HLA-C gene polymorphisms among patients with hepatitis C: a comparison between sustained virological responders and non-responders.
      ,
      • Khakoo S.I.
      • Thio C.L.
      • Martin M.P.
      • Brooks C.R.
      • Gao X.
      • Astemborski J.
      • et al.
      HLA and NK cell inhibitory receptor genes in resolving hepatitis C virus infection.
      ]. Thus, clinical decisions surely should not be based exclusively on IL28B genotyping, and a poor-response IL28B genetic background alone should never be an argument to withhold therapy from a patient [
      • Afdhal N.
      • Mchutchison J.G.
      • Zeuzem S.
      • Mangia A.
      • Pawlotsky J.M.
      • Murray J.C.
      • et al.
      Hepatitis C pharmacogenetics: State of the art in 2010.
      ].
      At present, it is one of the most relevant questions in the management of chronic HCV genotype 1 infection whether early standard therapy is advisable in a given patient, or whether treatment should be postponed in anticipation of the approval of DAA agents. Diagnosing a good-response IL28B genotype may substantially back up immediate initiation of standard therapy in a patient (Fig. 3). In contrast, postponement of therapy appears to be a reasonable strategy in patients with a poor-response IL28B genetic background and mild to moderate liver fibrosis. This strategy may be further supported by preliminary data showing that poor-response IL28B genotypes might be associated with decelerated liver fibrosis progression (see below). Triple therapy with pegIFN-α and ribavirin in combination with DAA agents in general is substantially more effective in HCV genotype 1 patients compared to standard therapy [
      • Bacon B.R.
      • Gordon S.C.
      • Lawitz E.
      • Marcellin P.
      • Vierling J.M.
      • Zeuzem S.
      • et al.
      HCV respond-2 final results: high sustained virologic response among genotype 1 previous non-responders and relapsers to peginterferon/ribavirin when re-treated with boceprevir plus pegintron (pegintron alfa-2b)/ribavirin.
      ,
      • Hezode C.
      • Forestier N.
      • Dusheiko G.
      • Ferenci P.
      • Pol S.
      • Goeser T.
      • et al.
      Telaprevir and peginterferon with or without ribavirin for chronic HCV infection.
      ,
      • Jacobson I.M.
      • McHutchison J.G.
      • Dusheiko G.
      • Di Bisceglie A.M.
      • Reddy K.R.
      • Bzowej N.H.
      • et al.
      Telaprevir in combination with peginterferon and ribavirin in genotype 1 HCV treatmnet-naive patients: final results of phase 3 ADVANCE study.
      ,
      • Kwo P.Y.
      • Lawitz E.J.
      • McCone J.
      • Schiff E.R.
      • Vierling J.M.
      • Pound D.
      • et al.
      Efficacy of boceprevir, an NS3 protease inhibitor, in combination with peginterferon alfa-2b and ribavirin in treatment-naive patients with genotype 1 hepatitis C infection (SPRINT-1): an open-label, randomised, multicentre phase 2 trial.
      ,
      • McHutchison J.G.
      • Everson G.T.
      • Gordon S.C.
      • Jacobson I.M.
      • Sulkowski M.
      • Kauffman R.
      • et al.
      Telaprevir with peginterferon and ribavirin for chronic HCV genotype 1 infection.
      ,
      • McHutchison J.G.
      • Manns M.P.
      • Muir A.J.
      • Terrault N.A.
      • Jacobson I.M.
      • Afdhal N.H.
      • et al.
      Telaprevir for previously treated chronic HCV infection.
      ,
      • Poordad E.
      • McCone J.
      • Bacon B.
      • Bruno S.
      • Manns M.P.
      • Sulkowski M.
      • et al.
      Boceprevir combined with peginterferon alfa-2B/ribavirin for treatment-naive patients with hepatitis C virus genotype (G) 1: SPRINT-2 final results.
      ,
      • Sherman K.E.
      • Flamm S.L.
      • Afdhal N.H.
      • Nelson D.R.
      • Sulkowski M.S.
      • Everson G.T.
      • et al.
      Telaprevir in combination with peginterferon alfa-2a and ribavirin for 24 or 48 weeks in treatment-naive genotype 1 HCV patients who achieved an extended rapid viral response: final results of phase 3 ILLUMINATE study.
      ]. However, triple therapy is also burdened with additional significant side effects and cost. Thus, IL28B SNPs may still be used after approval of DAA agents to select patients in whom standard therapy might still be justified. Most patients with good-response IL28B genotypes will have a high chance of cure with both regimens, and factors such as preference of short treatment duration, risk of anemia, or presence of skin disorders may be crucial for individual decisions.
      Figure thumbnail gr3
      Fig. 3Possible treatment algorithms for chronic HCV genotype 1 infected patients according to IL28B alleles. Instead of rs12979860 C/C versus C/T, T/T, other IL28B SNPs may be used. The suggested treatment algorithms are of hypothetical nature, and randomized controlled clinical studies are required, in which patients are stratified according to IL28B genotypes. Moreover, IL28B SNPs should not be used as exclusive predictors of treatment response, and treatment algorithms may be modified according to other predictors of virologic response. TVR, telaprevir; BOC, boceprevir; eRVR, extended rapid virologic response (HCV RNA below limit of detection at weeks 4 and 12 during treatment).
      IL28B variations may also significantly support individualizing treatment durations (Fig. 3). In this context, it is important that IL28B variations provide additional, discrete information to on-treatment viral kinetics. Though RVR has a higher specificity and positive predictive value for SVR than a good-response IL28B genotype, IL28B SNPs exhibit a better negative-predictive value and sensitivity for SVR [
      • Thompson A.J.
      • Muir A.J.
      • Sulkowski M.S.
      • Ge D.
      • Fellay J.
      • Shianna K.V.
      • et al.
      Interleukin-28B polymorphism improves viral kinetics and is the strongest pretreatment predictor of sustained virologic response in genotype 1 hepatitis C virus.
      ]. In addition, IL28B SNPs are strongly associated with SVR in patients who do not achieve RVR [
      • Thompson A.J.
      • Muir A.J.
      • Sulkowski M.S.
      • Ge D.
      • Fellay J.
      • Shianna K.V.
      • et al.
      Interleukin-28B polymorphism improves viral kinetics and is the strongest pretreatment predictor of sustained virologic response in genotype 1 hepatitis C virus.
      ]. Sarrazin et al. retrospectively evaluated the importance of IL28B SNPs in completely individualized treatment durations (24–72 weeks) with pegIFN-α and ribavirin in HCV genotype 1 patients [
      • Sarrazin C.
      • Schwendy S.
      • Moeller B.
      • Dikopoulos N.
      • Buggisch P.
      • Encke J.
      • et al.
      Completely individualized treatment durations with peginterferon-alfa-2b and ribavrin in HCV genotype 1-infected patients and importance of IL28B genotype (INDIV-2 study).
      ]. Duration of treatment was defined by a response-guided approach according to the first time when HCV RNA fell below the limit of detection. Importantly, SVR rates were consistently higher in patients with IL28B rs12979860 genotype C/C (85% SVR), compared to C/T (58% SVR) and T/T (46% SVR). Thus, patients with good-response IL28B genotypes may be optimal candidates for individualized durations of standard treatment with pegIFN-α and ribavirin. At present, the role of IL28B variations in response-guided approaches during triple-therapy regimens is unclear, though one might speculate that 12–24 weeks of triple therapy may be sufficient to achieve high SVR rates in patients with good-response IL28B genotypes.
      Apparently, HCV genotype 1 (and 4) patients with poor-response IL28B alleles should be considered as difficult to cure patients. Traditional approaches such as prolonged therapy with pegIFN-α and ribavirin are of limited efficacy [
      • Farnik H.
      • Lange C.M.
      • Sarrazin C.
      • Kronenberger B.
      • Zeuzem S.
      • Herrmann E.
      Meta-analysis shows extended therapy improves response of patients with chronic hepatitis C virus genotype 1 infection.
      ]. In contrast, phase II and III clinical trials have evidenced an enormous potential of telaprevir or boceprevir in combination with pegIFN-α and ribavirin in previous partial non-responders and relapsers to standard therapy [
      • Bacon B.R.
      • Gordon S.C.
      • Lawitz E.
      • Marcellin P.
      • Vierling J.M.
      • Zeuzem S.
      • et al.
      HCV respond-2 final results: high sustained virologic response among genotype 1 previous non-responders and relapsers to peginterferon/ribavirin when re-treated with boceprevir plus pegintron (pegintron alfa-2b)/ribavirin.
      ,
      • McHutchison J.G.
      • Manns M.P.
      • Muir A.J.
      • Terrault N.A.
      • Jacobson I.M.
      • Afdhal N.H.
      • et al.
      Telaprevir for previously treated chronic HCV infection.
      ]. These cohorts are presumably enriched with patients carrying poor-response IL28B alleles. It is, therefore, likely that patients with poor-response IL28B alleles will strongly benefit from triple therapy regimens. Though preliminary data indicate an attenuated association of IL28B variations with response to triple therapy as well, longer treatment durations (48 weeks) may be required in patients with poor-response IL28B alleles. Unfortunately, treatment of prior null-responders to standard therapy, which in most cases will carry poor-response IL28B alleles, will be still unsatisfactory with telaprevir-based triple therapy (boceprevir was not evaluated in null-responders) [
      • McHutchison J.G.
      • Manns M.P.
      • Muir A.J.
      • Terrault N.A.
      • Jacobson I.M.
      • Afdhal N.H.
      • et al.
      Telaprevir for previously treated chronic HCV infection.
      ]. Therefore, alternative treatment strategies such as quadruple therapy or treatment with DAA agents with a higher genetic barrier to resistance may be optimal in subgroups of patients with poor-response IL28B alleles, and in particular in prior null-responders to standard therapy (Fig. 3).
      In HCV genotype 2 and 3 patients, IL28B variations in general are only weakly associated with SVR. However, a poor-response IL28B genotype might indicate a need for prolonged therapy of 48 weeks in HCV genotype 2 and 3 patients who do not attain an RVR [
      • Mangia A.
      • Thompson A.J.
      • Santoro R.
      • Piazzolla V.
      • Tillmann H.L.
      • Patel K.
      • et al.
      An IL28B polymorphism determines treatment response of hepatitis C virus genotype 2 or 3 patients who do not achieve a rapid virologic response.
      ]. Whether a good-response IL28B genotype in HCV genotype 2 and 3 patients may be an argument of shortened treatment duration (e.g. 12 weeks) is unclear at the moment.

      IL28B SNPs in liver transplanted patients with recurrent hepatitis C

      Recurrent hepatitis C after liver transplantation is almost universal and has a substantial impact on graft and overall survival [
      • Terrault N.A.
      Hepatitis C therapy before and after liver transplantation.
      ]. Thus, prevention or treatment of HCV liver graft reinfection is of major importance. Unfortunately, pegIFN-α and ribavirin are of limited efficacy and poorly tolerated in liver transplanted patients. Employing IL28B SNPs in the management of post-transplant recurrent hepatitis C would be complicated by the genetic chimerism of transplanted patients, in whom liver donor and recipient IL28B genotypes may differ. Intriguingly, several retrospective studies have shown that both liver donor and recipient IL28B variations are associated with response to (peg)IFN-α based therapy of recurrent hepatitis C [
      • Fukuhara T.
      • Taketomi A.
      • Motomura T.
      • Okano S.
      • Ninomiya A.
      • Abe T.
      • et al.
      Variants in IL28B in liver recipients and donors correlate with response to peg-interferon and ribavirin therapy for recurrent hepatitis C.
      ,
      • Charlton M.R.
      • Thompson A.
      • Veldt B.J.
      • Watt K.
      • Tillmann H.
      • Poterucha J.J.
      • et al.
      Interleukin-28B polymorphisms are associated with histological recurrence and treatment response following liver transplantation in patients with hepatitis C virus infection.
      ,

      Lange CM, Moradpour D, Doehring A, Lehr HA, Mullhaupt B, Bibert S, et al. Impact of donor and recipient IL28B rs12979860 genotypes on hepatitis C virus liver graft reinfection. J Hepatol 2010, epub ahead of print.

      ]. In the largest study, both donor and recipient IL28B variations were strong and independent predictors of SVR, with minimal SVR rates in patients with donor and recipient poor-response IL28B genotype, intermediate SVR rates in patients with a good-response IL28B genotype of either the donor or the recipient, and excellent SVR rates in patients with good-response IL28B genotype of both the donor and recipient [
      • Charlton M.R.
      • Thompson A.
      • Veldt B.J.
      • Watt K.
      • Tillmann H.
      • Poterucha J.J.
      • et al.
      Interleukin-28B polymorphisms are associated with histological recurrence and treatment response following liver transplantation in patients with hepatitis C virus infection.
      ]. In contrast to non-transplanted patients, IL28B variations also appear to be important predictors of treatment outcome in HCV genotype 2 and 3 infected patients with recurrent hepatitis C [

      Lange CM, Moradpour D, Doehring A, Lehr HA, Mullhaupt B, Bibert S, et al. Impact of donor and recipient IL28B rs12979860 genotypes on hepatitis C virus liver graft reinfection. J Hepatol 2010, epub ahead of print.

      ].
      In addition, IL28B variations were associated with parameters reflecting the natural course of recurrent hepatitis C. For example, recipient but not donor rs12979860 C/C versus C/T, T/T genotypes were associated with delayed time to histologic recurrence of hepatitis C and with decelerated fibrosis progression of the liver graft [
      • Charlton M.R.
      • Thompson A.
      • Veldt B.J.
      • Watt K.
      • Tillmann H.
      • Poterucha J.J.
      • et al.
      Interleukin-28B polymorphisms are associated with histological recurrence and treatment response following liver transplantation in patients with hepatitis C virus infection.
      ]. Thus, adequate prospective studies may also document an effect of IL28B variations on survival rates of liver transplanted patients with recurrent hepatitis C. This could theoretically justify an implementation of IL28B genetic testing in the allocation of liver graft to HCV infected patients. Concrete treatment recommendations on the basis of IL28B genotypes in liver transplanted patients cannot be given at the moment. Nevertheless, IL28B genetic testing appears to offer important opportunities in the management of recurrent hepatitis C. For example, alternative treatment strategies such as high-dose silibinin infusions after liver transplantation might be considered in particular for patients with a poor-response IL28B genetic background [
      • Neumann U.P.
      • Biermer M.
      • Eurich D.
      • Neuhaus P.
      • Berg T.
      Successful prevention of hepatitis C virus (HCV) liver graft reinfection by silibinin mono-therapy.
      ].

      IL28B SNPs and the natural course of hepatitis C

      In addition to their striking role in spontaneous and treatment-induced clearance, IL28B genetic variations seem to influence various phenotypes in chronic hepatitis C. Already the first GWAS by Ge postponement found that the same IL28B genotypes which are associated with treatment-induced clearance also correlate with higher baseline viral loads [
      • Ge D.
      • Fellay J.
      • Thompson A.J.
      • Simon J.S.
      • Shianna K.V.
      • Urban T.J.
      • et al.
      Genetic variation in IL28B predicts hepatitis C treatment-induced viral clearance.
      ]. This finding was confirmed by various studies including HCV genotype 1, 2, 3, and 4 patients [
      • Rauch A.
      • Kutalik Z.
      • Descombes P.
      • Cai T.
      • Di Iulio J.
      • Mueller T.
      • et al.
      Genetic variation in IL28B is associated with chronic hepatitis C and treatment failure: a genome-wide association study.
      ,
      • McCarthy J.J.
      • Li J.H.
      • Thompson A.
      • Suchindran S.
      • Lao X.Q.
      • Patel K.
      • et al.
      Replicated association between an IL28B gene variant and a sustained response to pegylated interferon and ribavirin.
      ,
      • Sarrazin C.
      • Susser S.
      • Doehring A.
      • Lange C.M.
      • Muller T.
      • Schlecker C.
      • et al.
      Importance of IL28B gene polymorphisms in hepatitis C virus genotype 2 and 3 infected patients.
      ,
      • Stattermayer A.F.
      • Stauber R.
      • Hofer H.
      • Rutter K.
      • Beinhardt S.
      • Scherzer T.M.
      • et al.
      Impact of IL28B Genotype on the Early and Sustained Virologic Response in Treatment-Naive Patients With Chronic Hepatitis C.
      ]. However, Thompson postponement reported that rs12979860 genotype frequencies were similar when patients were stratified in groups with low or high pretreatment HCV RNA serum concentrations (threshold 600,000 IU/ml) [
      • Thompson A.J.
      • Muir A.J.
      • Sulkowski M.S.
      • Ge D.
      • Fellay J.
      • Shianna K.V.
      • et al.
      Interleukin-28B polymorphism improves viral kinetics and is the strongest pretreatment predictor of sustained virologic response in genotype 1 hepatitis C virus.
      ]. This may explain why both IL28B variations and baseline viral load are independent and reliable predictors of treatment outcome.
      Good-response IL28B variations are also associated with the activity of chronic hepatitis C in terms of more severe necro-inflammation in liver biopsies, as well as with higher alanine and aspartate aminotransferase serum levels [
      • Sarrazin C.
      • Susser S.
      • Doehring A.
      • Lange C.M.
      • Muller T.
      • Schlecker C.
      • et al.
      Importance of IL28B gene polymorphisms in hepatitis C virus genotype 2 and 3 infected patients.
      ,
      • Abe H.
      • Ochi H.
      • Maekawa T.
      • Hayes C.N.
      • Tsuge M.
      • Miki D.
      • et al.
      Common variation of IL28 affects gamma-GTP levels and inflammation of the liver in chronically infected hepatitis C virus patients.
      ]. These findings may reflect a more pronounced immune response against HCV in patients with good-response IL28B variations, which may be true in particular for the adaptive immunity since good-response IL28B variations are associated with lower levels of interferon-stimulated genes (see below). Potential associations of IL28B variations with hepatic fibrosis are less well characterized. In a large GWAS, no association of IL28B variations with the degree of liver fibrosis at baseline was observed [
      • Thompson A.J.
      • Clark P.J.
      • Fellay J.
      • Muir A.J.
      • Tillmann H.L.
      • Patel K.
      • et al.
      IL28B genotype is not associated with advanced hepatic fibrosis in chronic hepatitis C patients enrolled in the IDEAL study.
      ]. However, in the Swiss hepatitis C cohort study, which includes more than 600 untreated patients with paired liver biopsies, “good-response” IL28B variations were associated with more accelerated progression of liver fibrosis (Pierre-Yves Bochud, personal communication).
      Finally, metabolic abnormalities associated with chronic hepatitis C were correlated with IL28B genotypes. A profound association of the good-response rs12979860 C/C genotype with higher low-density lipoprotein (LDL) levels and other serum lipid parameters was observed in HCV infected patients [
      • Li J.H.
      • Lao X.Q.
      • Tillmann H.L.
      • Rowell J.
      • Patel K.
      • Thompson A.
      • et al.
      Interferon-lambda genotype and low serum low-density lipoprotein cholesterol levels in patients with chronic hepatitis C infection.
      ]. In addition, good response IL28B SNPs were associated with a lower incidence of hepatic steatosis in patients with chronic hepatitis C [

      Cai T, Dufour JF, Muellhaupt B, Gerlach T, Heim M, Moradpour D, et al. Viral Genotype-Specific Role of PNPLA3, PPARG, MTTP and IL28B in Hepatitis C Virus-Associated Steatosis. J Hepatol 2011, epub ahead of print.

      ]. Interferons were shown to decrease serum levels of cholesterol and other parameters of lipid metabolism [
      • Lange C.M.
      • von Wagner M.
      • Bojunga J.
      • Berg T.
      • Farnik H.
      • Hassler A.
      • et al.
      Serum lipids in European chronic HCV genotype 1 patients during and after treatment with pegylated interferon-alpha-2a and ribavirin.
      ]. The association of good-response IL28B variations with higher LDL and cholesterol serum levels may, therefore, reflect a lower pre-treatment IFN-(λ) activity in these patients.
      In line, the expression level of interferon-stimulated genes (ISGs) may provide a possible causal link between IL28B variations and such histologic and metabolic features of chronic hepatitis C. Two studies found a strong association of IL28B variations with the expression level of ISGs in micro-arrays of liver specimens [
      • Urban T.J.
      • Thompson A.J.
      • Bradrick S.S.
      • Fellay J.
      • Schuppan D.
      • Cronin K.D.
      • et al.
      IL28B genotype is associated with differential expression of intrahepatic interferon-stimulated genes in patients with chronic hepatitis C.
      ,
      • Honda M.
      • Sakai A.
      • Yamashita T.
      • Nakamoto Y.
      • Mizukoshi E.
      • Sakai Y.
      • et al.
      Hepatic ISG expression is associated with genetic variation in interleukin 28B and the outcome of IFN therapy for chronic hepatitis C.
      ]. However, a more recent study provided evidence that this association may be not causal, but rather explained by a frequent co-incidence of good-response IL28B alleles with low ISG expression levels in those patients who will respond to IFN-α-based therapy [
      • Dill M.T.
      • Duong F.H.
      • Vogt J.E.
      • Bibert S.
      • Bochud P.Y.
      • Terracciano L.
      • et al.
      Interferon-Induced Gene Expression Is a Stronger Predictor of Treatment Response Than IL28B Genotype in Patients With Hepatitis C.
      ].

      Conclusions

      IL28B genetic variations are the strongest established pre-treatment predictor of SVR to standard therapy in HCV genotype 1 patients. Preliminary data indicate that IL28B variations also exhibit an attenuated effect on response to triple therapy including DAA agents. Though IL28B genetic variations are strongly associated with on-treatment viral kinetics, they provide discrete information to RVR and EVR. In view of these facts, IL28B SNPs will likely play an important role in future management of chronic hepatitis C, especially in the selection of appropriate treatment regimens and in the definition of individualized treatment durations. However, detailed analyses of randomized controlled clinical trials in which patients were stratified according to the IL28B genotype are necessary before IL28B genotyping can be included in treatment recommendations. Clearly, clinical decisions should not be made exclusively on the IL28B genotype, and additional predictors of treatment response will maintain importance. The strong effect of genetic variations near the IL28B gene on spontaneous and treatment-induced clearance of HCV also points to an important role of IFN-λ-signaling in HCV infection. This unexpected observation boosted basic research on the role of IFN-λs in the pathogenesis of hepatitis C, as well as the development of pegylated interferon-λ1 (pegIFN-λ1), which is currently in phase II clinical evaluation for the treatment of chronic hepatitis C.

      Conflict of interest

      CM Lange: none.S Zeuzem: Consultancy for Abbott, Achillion, Anadys, BMS, Gilead, Itherx, Merck, Novartis, Pfizer, Roche, Santaris, Tibotec, and Vertex.

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