SHORT COMMUNICATION
JOURNAL OF INTERFERON & CYTOKINE RESEARCH Volume 00, Number 00, 2014 ª Mary Ann Liebert, Inc. DOI: 10.1089/jir.2014.0111
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Post-Transplant Course of Hepatitis C After Living Donor Liver Transplantation in Association with Polymorphisms near IFNL3 Kristin G. Monaghan,1,* Humberto C. Gonzalez,2 Albert M. Levin,3,4 Marwan S. Abouljoud,5 and Stuart C. Gordon 6
Donor genotype for polymorphisms near IFNL3 influences hepatitis C virus (HCV) therapy responsiveness. This relationship has not been studied in a sample of HCV-infected living donor liver transplantation (LDLT) recipients in the United States (US). We investigated the association of donor and recipient genotypes near the IFNL3 gene at a large US liver transplant center. Recipient homozygosity for rs12979860 C was associated with increased sustained virologic response (SVR) in antiviral treatment-experienced patients pretransplant (P = 0.055). Consistently, donor homozygosity for rs12979860 C was also associated with increased SVR in patients who received post-transplant antiviral therapy (P = 0.048). Transplantation of an rs12979860 CC graft confers a favorable posttransplant antiviral response among HCV-positive recipients in an LDLT setting. Recipients with the favorable rs12979860 genotype receiving antiviral therapy before transplant are also more likely to achieve SVR. The effect of genotype status in the era of direct-acting antiviral agents will require future study.
Most recently, simeprevir an NS3/4A protease inhibitor with Peg IFN/RBV and sofosbuvir a nucleotide analog NS5B polymerase inhibitor with or without PegIFN/RBV based on the genotype have been approved, providing higher SVR rates. Recurrence of HCV infection after liver transplantation of viremic patients is universal. Recurrent HCV is associated with graft loss, cirrhosis, and death in up to a third of patients at 5 years, leading to a worse outcome when compared to other causes of transplantation (Sallie and others 1994; Aytaman and others 2010). There is currently no standard of care treatment for HCV in the postliver transplant setting, but PegIFN/RBV has been classically used when follow-up liver biopsies demonstrate fibrosis progression (Garcia-Retortillo and Forns 2004). SVR rates in this setting are low, as treatment is characterized by dose reductions, intolerances, and side effects that lead to frequent discontinuation of therapy. When SVR is achieved, there is improved liver histology and survival (Picciotto and others 2007). Early experience with living donor liver transplantation (LDLT) in the setting of chronic HCV infection suggested a less successful outcome than from deceased donors (Takada and others 2006), whereas a recent US experience indicates that donor type does not affect the probability of advanced fibrosis and graft survival in HCV-infected recipients (Terrault and others 2014).
Introduction
H
epatitis C infection is a common blood-borne disease, affecting 2%–3% of the world population. In developed countries, including the United States, the prevalence is less than 2%. Up to 85% of individuals with an acute HCV infection develop chronic hepatitis C. Among chronically infected individuals, 20% develop cirrhosis and 1%–5% will develop hepatocellular carcinoma (HCC). Overall, 5%–7% of HCV patients ultimately die as a result of the infection. HCV infection is the most common reason for liver transplantation in the United States (CDC 2014; WHO 2013). For many years, the standard of care therapy for HCV was the combined use of peginterferon (PegIFN) and ribavirin (RBV). Treatment duration varied according to the HCV genotype. Sustained virologic response (SVR), defined as continued aviremia for 24 weeks after the end of treatment, was usually obtained in 40%–50% for HCV genotype 1 (most common) and 80% for HCV genotype 2 and 3. SVR is associated with virological cure and with improved morbidity and mortality (Ghany and others 2011). In 2011, the direct-acting antiviral agents (DAA) boceprevir and telaprevir were approved for combined use with PegIFN/RBV for the treatment of HCV type 1 becoming the new standard of care (Ghany and others 2011).
1 Medical Genetics, 3Public Health Sciences, 4Center for Bioinformatics, 5Transplant Surgery, 6Gastroenterology and Hepatology, Henry Ford Health System, Detroit, Michigan. 2 Department of Transplant Surgery, University of Tennessee/Methodist University Transplant Institute, Memphis, Tennessee. *Current affiliation: GeneDx, Gaithersburg, Maryland.
1
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2
Pharmacogenetic analysis can be used to assist physicians in determining the proper medication and dose for therapeutic regimens. With respect to HCV treatment, the favorable alleles for the genetic markers, rs12979860 (C) and rs8099917 (T), have been shown to be associated with spontaneous HCV clearance and favorable response to PegIFN/RBV therapy. The unfavorable alleles (rs12979860 T and rs8099917 G) are associated with decreased likelihood of SVR, unfavorable response to antiviral therapy, HCV-associated liver cirrhosis, and HCC (Ge and others 2009; Suppiah and others 2009; Tanaka and others 2009; Thomas and others 2009; Rauch and others 2010; Fabris and others 2011). These markers are located on chromosome 19q13.13 near the IFNL3 gene (formerly known as IL28B). This gene encodes a cytokine with immunomodulatory activity. Its expression is induced by viral infection. IFNL3 is a member of the lambda interferon gene family. Lambda interferons have been shown to inhibit HCV replication in vitro and are hypothesized to initiate an antiviral response through the JAKSTAT pathway and play an important role in Toll-like receptor-induced antiviral and immune defense in the intestinal epithelium (Gad and others 2009; Li and others 2009; Chinnaswamy 2014; GeneCards 2014). IFNL3 genotyping is available on a clinical basis in North America and Europe. The recipient and donor liver rs12979860 genotype has been shown to be associated with response to HCV antiviral therapy in patients after orthotopic liver transplantation (OLT) (Charlton and others 2011). The rs8099917 genotype has been shown to be associated with SVR in recipient–donor pairs undergoing LDLT for HCV-related liver disease in Japan (Fukuhara and others 2010). The association between both of these markers and the response to PegIFN/RBV-based therapy among the LDLT HCV patient population in the United States has not been previously characterized. We retrospectively investigated the association of donor and recipient rs12979860 and rs8099917 genotypes with SVR at a large US liver transplant center.
MONAGHAN ET AL.
Results Donor ages ranged from 20 to 51 (mean 36.8 years). Ten were male (56%) and 8 were female. Seventeen donors were white and 1 was African American. Six of the recipient–donor pairs were unrelated and 12 were related (all first-degree relationships). Recipient characteristics are summarized in Table 1. Recipient ages ranged from 40 to 69 (mean 53.6 years). Three recipients had a diagnosis of HCC. The majority of patients had HCV type 1. The HCV genotype was not available for 1 recipient (now deceased) who had negative HCV viral counts and whose records were no longer available to us for review. Among the patients who received antiviral therapy, 5 achieved SVR. The 1 patient with ongoing therapy was not responding to treatment at the time of submission of this article and has no significant log drop in viral titer. Four patients required retransplantation for reasons, including allograph failure, hepatic artery thrombosis, and recurrent cholestatic hepatitis C. Among white recipients (n = 15) and donors (n = 17), the rs12979860 favorable C allele (regardless of zygosity) frequency was 0.57 and 0.65, compared with a frequency of 0.73 for the Centre d’Etude du Polymorphisme Humain (CEPH) Utah Americans of western and northern European descent (CEU) subjects in the 1000 genomes project. The rs8099917 favorable T allele frequency was 0.77 and 0.74 for white recipients and donors, respectively. The comparable value in the CEU subjects was 0.84. Using the CEU data for both SNPs, the estimated pairwise linkage disequilibrium r2 measure between the 2 SNPs was 0.47. Due to the small number of African American and Hispanic donors and recipients, the allele frequency was not calculated for these groups. SNP associations with SVR status are displayed in Table 2. Before transplant, 11 patients received antiviral therapy (Table 2). Of these, 3 were homozygous for rs12979860 C. Two (67%) of these individuals achieved SVR before transplant in comparison with none of the 8 nonhomozygous subjects (P = 0.055). Recipient rs8099917 T homozygosity was
Materials and Methods Twenty-two consecutive patients underwent LDLT for HCV-related cirrhosis and HCC between 2000 and 2012 at the Henry Ford Health System in Detroit, MI (ClinicalTrails.gov: NCT 01429155). Of these 22 patients, 18 recipient–donor pairs and 1 recipient only were available for genotyping. Residual extracted DNA from the liver donors had been routinely stored at the Henry Ford Immunology laboratory. Liver explants (recipients) fixed in paraffin were identified in the Pathology laboratory. DNA was extracted from donor whole blood and paraffin-fixed livers of recipients using standard methods. Genotyping of the rs12979860 (C/T) and rs8099917 (T/G) single-nucleotide polymorphisms (SNPs) was performed using a previously described single-nucleotide extension (SNE) assay (Melis and others 2011). SNE results were confirmed for 5 DNA samples of various genotypes using Sanger sequencing. Given prior findings, a recessive genetic model was used to test for recipient and donor genotype associations between each SNP (rs12979860 C and rs8099917 T) and SVR status. Due to the sample size, Fisher’s exact tests were used to evaluate the significance of these associations. This study was approved by the HFHS Institutional Review Board.
Table 1. Summary of Recipient (n = 19) Characteristics n Gender Male Female Race/Ethnicity White African American Hispanic HCV genotype Type 1 Type 3 Unknown Antiviral therapy (pegIFN/RBV) None Pretransplant only Post-transplant only Pre- and post-transplanta
12 7 15 1 3 15 3 1 5 3 3 8
a Includes the 1 patient still undergoing therapy at the time of the data analysis. HCV, hepatitis C virus.
HEPATITIS C LIVING DONOR LIVER TRANSPLANT AND IL28-B
Table 2.
3
Donor and Recipient Genotype Associations with Pre- and Post-Transplant SVR Pretransplant antiviral
SNP
Genotype source
rs12979860
Donor Recipient
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rs8099917
Donor Recipient
SVR count Genotype CC TT/CT CC TT/CT TT GG/GT TT GG/GT
Yes
No
SVR% by genotype
1 1 2 0 1 1 2 0
3 5 1 8 4 4 4 5
25 17 67 0 20 20 33 0
Post-transplant antiviral SVR count Pa
Yes
No
SVR% by genotype
1.000
3 0 0 3 3 0 0 3
1 5 2 4 2 4 5 2
75 0 0 43 60 0 0 60
0.055 1.000 0.455
Pa 0.048 0.500 0.167 0.167
Eight of the subjects received antivirals, both pre- and post-transplantation, and contributed data to both pre- and post-transplant SVR outcome analyses. Three patients were only evaluated for pretransplant SVR, and another 3 were only evaluated for post-transplant SVR. a Each P value was calculated using Fisher’s exact test. SNP, single-nucleotide polymorphism; SVR, sustained virologic response.
not associated with SVR before transplantation (P = 0.455), although those with a favorable genotype had a higher proportion achieving SVR (33% vs. 0%; Table 2). Post-transplant, 11 patients received therapy (Table 2), 8 of whom were also treated pretransplant (ie, 3 were only treated post-transplant). For 1 LDLT pair, the donor was not available for genotyping. Another patient with unfavorable donor genotypes at both SNPs is still receiving therapy, but at the time of submission of this article had a poor response to therapy. These 2 patients were removed from the analysis, leaving a post-transplant sample of 9 subjects. Of these, 4 of the donors were homozygous for rs12979860 C. Three (75%) patients with these homozygous donor genotypes achieved SVR in comparison with none of the 5 recipients whose donors were nonhomozygous (P = 0.048). SNP rs8099917 T showed a similar trend, with 60% of T allele homozygotes experiencing SVR (Table 2), but this result did not achieve statistical significance (P = 0.167). Favorable recipient genotypes at either SNP were not associated with post-transplant SVR (Table 2). Five of the 19 recipients were excluded from the analysis to determine HCV association between the genotype and SVR because they did not receive antiviral therapy. None of these patients had spontaneous viral clearance.
Conclusions The present report validates the use of an individual patient’s pharmacogenetics profile to guide decisions regarding prevention, diagnosis, and treatment of disease. The presence of a favorable homozygous genotype for both SNPs was associated with an increased rate of SVR following PegIFN/RBV therapy. Knowledge of a patient’s genotype to predict success of therapy, therefore, can be helpful due to the cost of the therapy and high discontinuation rate due to adverse side effects. In patients whose genotype predicts a lower chance of SVR with antiviral therapy, perhaps a shorter duration of treatment or alternative medication may be considered (Pacanowski and others 2012). Genotype analysis for rs12979860 and rs8099917 SNPs near the gene commonly known as IL28B is currently available on a clinical basis. The IL28B rs12979860 genotype information is included in the pharmacogenomics section of the package insert labels for peginterferon alfa-2b and the DAAs, boceprevir and telaprevir.
The recipient and donor liver rs12979860 genotype has been shown to be associated with response to HCV antiviral therapy in patients after OLT (Charlton and others 2011). Among HCVinfected patients who underwent OLT and received antiviral therapy, the presence of a favorable rs12979860 CC donor or recipient genotype was associated with SVR. SVR was lowest when the genotypes of the donor and recipient were unfavorable, increased if either the donor or recipient genotype was favorable, and most likely with both a favorable donor and recipient homozygous genotype. LDLT provides an opportunity to test for the IFNL3 genotype before surgery. A recent study in Japan demonstrated an association between the rs8099917 genotype and SVR in recipient–donor pairs undergoing LDLT for HCV-related liver disease (Fukuhara and others 2010). The highest SVR rate was observed in recipient–donor pairs, where both recipient and donor were homozygous for the favorable (T) allele. When 1 member of the donor–recipient pair had a homozygous favorable genotype and the other member had a nonfavorable genotype, the rate of SVR was lower (Fukuhara 2012, personal communication). A potential limitation of the current study is the small number of patients investigated. Based on this small number of LDLT recipient–donor pairs in the United States, our results suggest that transplantation of an rs12979860 CCpositive graft confers a favorable post-transplant antiviral response among HCV-positive recipients in an LDLT setting. While the donor status was associated with the highest likelihood of SVR post-transplant, a favorable recipient genotype at rs12979860 was also beneficial before surgery. Our data supports prior reports among orthotopic liver transplants suggesting that donor IL28B status affects IFN-based therapy response. The value of studying a living donor population in the United States validates previous results in the deceased donor setting; additionally, such pretransplant testing of the donor might influence the donor selection process. Given the potential clinical utility of these findings, larger studies are warranted to confirm these results. Such studies could involve a collaborative effort between multiple institutions, the analysis of additional SNPs, patients of other races/ethnic backgrounds, and in particular the role of these SNPs as they relate to the newer DAA medications that will soon replace IFN-based HCV therapies.
4
MONAGHAN ET AL.
Acknowledgment Genotyping studies were paid for through the Henry Ford Gastroenterology Hepatology Research Fund.
Author Disclosure Statement No competing financial interests exist.
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References Aytaman A, Kaufman M, Terrault NA. 2010. Management of posttransplant hepatitis C infection. Curr Opin Organ Transplant 15:301–309. Centers for Disease Control and Prevention. Chapter 3, Infectious diseases related to travel: Hepatitis C. COC health information for international travel 2014. Available at wwwnc.cdc.gov/ travel/yellowbook/2014/chapter-3-infectious-diseases-relatedto-travel/hepatitis-C. Accessed 12/2013. Charlton MR, Thompson A, Veldt BJ, Watt K, Tillmann H, Poterucha JJ, Heimbach JK, Goldstein D, McHutchinson J. 2011. Interleukin-28B Polymorphisms are associated with histological recurrence and treatment response following liver transplantation in patients with hepatitis C virus infection. Hepatology 53:317–324. Chinnaswamy S. 2014. Genetic variants at the IFNL3 locus and their association with hepatitis C Virus infections reveal novel insights into host-virus interactions. J Interferon Cytokine Res 34:479–497. Fabris C, Falleti E, Cussigh A, Bitetto D, Fontanini E, Bignulin S, Cmet S, Fornasiere E, Fumolo E, Fangazio S, Cerutti A, Minisini R, Pirisi M, Toniutto P. 2011. IL-28B rs12979860 C/T allele distribution in patients with liver cirrhosis: role in the course of chronic viral hepatitis and the development of HCC. J Hepatol 54:716–722. Fukuhara T, Taketomi A, Motomura T, Okano S, Ninomiya A, Abe T, Uchiyama H, Soejima Y, Shirabe K, Matsuura Y, Maehara Y. 2010. Variants in IL28B in liver recipients and donors correlate with response to peg-interferon and ribavirin therapy for recurrent hepatitis C. Gastroenterology 139:1577–1585. Gad HH, Dellgren C, Hamming OJ, Vends S, Paludan SR, Hartmann R. 2009. Interferon-lambda is functionally an interferon but structurally related to the interleukin-10 family. J Biol Chem 284:20869–20875. Garcia-Retortillo M, Forns X. 2004. Prevention and treatment of hepatitis C virus recurrence after liver transplantation. J Hepatol 41:2–10. Ge D, Fellay J, Thompson AJ, Simon JS, Shianna KV, Urban TJ, Heinzen EL, Qui P, Bertelsen AH, Muir AJ, Sulkowski M, McHutchinson JG, Goldstein DB. 2009. Genetic variation in IL28B predicts hepatitis C treatment-induced viral clearance. Nature 461:399–401. GeneCards. 2014. Interferon, Lambda 3 (IFNL3) Gene. Available at www.genecards.org. Accessed 8/26/14. Ghany MG, Nelson DR, Strader DB, Thomas DL, Seeff LB. 2011. An update on treatment of genotype 1 chronic hepatitis C virus infection: 2011 practice guideline by the American association for the study of liver diseases. Hepatology 54: 1433–1444. Li M, Liu X, Zhou Y, Su SB. 2009. Interferon-as: the modulators of antivirus, antitumor, and immune responses. J Leukoc Biol 86:23–32. Melis R, Fauron C, McMillin G, Lyon E, Shirts B, Hubley LM, Slev PR. 2011. Simultaneous genotyping of rs12979860 and rs8099917 variants near the IL28B locus associated with HCV clearance and treatment response. J Mol Diagn 13:446–451.
Pacanowski M, Amur S, Zineh I. 2012. New genetic discoveries and treatment for hepatitis C. JAMA 307:1921–1922. Picciotto FP, Tritto G, Lanza AG, Addario L, De Luca M, Di Costanzo GG, Lampasi F, Tartaglione MT, Marsilia GM, Calise F, Cuomo O, Ascione A. 2007. Sustained virological response to antiviral therapy reduces mortality in HCV reinfection after liver transplantation. J Hepatol 46:459–465. Rauch A, Kutalik Z, Descombes P Cai T, Iulio JD, Mueller T, Bochud M, Battegay M, Bernasconi E, Borovicka J, Colombo S, Cerny A, Dufour J-F, Furrer H, Gunthard HF, Heim M, Hirschel B, Malinverni R, Maradpour D, Mullhaupt B, Witteck A, Beckmann JS, Berg T, Bergmann S, Negro F, Telenti A, Bochud P-Y. 2010. Genetic variation in IL28B is associated with chronic hepatitis C and treatment failure: a genome-wide association study. Gastroenterology 138:1338–1345. Sallie R, Cohen AT, Tibbs CJ, Portmann BC, Rayner A, O’Grady JG, Tan KC, Williams R. 1994. Recurrence of hepatitis C following orthotopic liver transplantation: a polymerase chain reaction and histological study. J Hepatol 21:536–542. Suppiah V, Moldovan M, Ahlenstiel G, Berg T, Weltman M, Abate ML, Bassendine M, Spengler U, Dore GJ, Powell E, Riordan S, Sheridan D, Smedile A, Fragomeli V, Muller T, Bahlo M, Stewart GJ, Booth DR, George J. 2009. IL28B is associated with response to chronic hepatitis C interferon-a and ribavirin therapy. Nat Genet 41:1100–1104. Takada Y, Haga H, Ito T, Nabeshima M, Ogawa K, Kasahara M, Oike F, Ueda M, Egawa H, and Tanaka K. 2006. Clinical outcomes of living donor liver transplantation for hepatitis C virus (HCV)-positive patients. Transplantation 81:350–354. Tanaka Y, Nishida N, Sugiyama M, Kurosaki M, Matsuura K, Sakamoto N, Nakagawa M, Korenga M, Hino K, Hige S, Ito Y, Mita E, Tanaka E, Mochida S, Murawaki Y, Honda M, Sakai A, Hiasa Y, Nishiguchi S, Koide A, Sakaida I, Imamura M, Ito K, Yano K, Masaki N, Sugauchi F, Izumi N, Tokunaga K, Mizokami M. 2009. Genome-wide association of IL28B with response to pegylated interferon-a and ribavirin therapy for chronic hepatitis C. Nat Genet 41:1105–1111. Terrault NA, Stravitz RT, Lok ASF, Everson GT, Brown RS, Kulik LM, Olthoff KM, Saab S, Adeyi O, Argo CK, Everhart JR, Rodrigo DR; the A2ALL Study Group. 2014. Hepatitis C disease severity in living versus deceased donor liver transplant recipients: an extended observation study. Hepatology 59:1311–1319. Thomas DL, Thio CL, Martin MP Qi Y, Ge D, O’hUigin C, Kidd J, Kidd K, Khakoo SI, Alexander G, Goedert JJ, Kirk GD, Donfield SM, Rosen HR, Tobler LH, Busch MP, McHutchinson JG, Goldstein DB, Carrington M. 2009. Genetic variation in IL28B and spontaneous clearance of hepatitis C virus. Nature 461:798–802. World Health Organization (WHO). 2013. Global alert and response, hepatitis C. Available at www.who.int/csr/disease/ hepatitis/whocdscsrlyo2003/en/index4.html. Accessed 12/2013.
Address correspondence to: Dr. Humberto C. Gonzalez Department of Transplant Surgery University of Tennessee/Methodist University Transplant Institute 1211 Union Ave. Suite 340 Memphis, TN 38104 E-mail: [email protected] Received 3 July 2014/Accepted 10 September 2014
JOURNAL OF INTERFERON & CYTOKINE RESEARCH Volume 00, Number 00, 2014 ª Mary Ann Liebert, Inc. DOI: 10.1089/jir.2014.0111
Journal of Interferon & Cytokine Research Downloaded from online.liebertpub.com by Univ Braunschweig on 11/17/14. For personal use only.
Post-Transplant Course of Hepatitis C After Living Donor Liver Transplantation in Association with Polymorphisms near IFNL3 Kristin G. Monaghan,1,* Humberto C. Gonzalez,2 Albert M. Levin,3,4 Marwan S. Abouljoud,5 and Stuart C. Gordon 6
Donor genotype for polymorphisms near IFNL3 influences hepatitis C virus (HCV) therapy responsiveness. This relationship has not been studied in a sample of HCV-infected living donor liver transplantation (LDLT) recipients in the United States (US). We investigated the association of donor and recipient genotypes near the IFNL3 gene at a large US liver transplant center. Recipient homozygosity for rs12979860 C was associated with increased sustained virologic response (SVR) in antiviral treatment-experienced patients pretransplant (P = 0.055). Consistently, donor homozygosity for rs12979860 C was also associated with increased SVR in patients who received post-transplant antiviral therapy (P = 0.048). Transplantation of an rs12979860 CC graft confers a favorable posttransplant antiviral response among HCV-positive recipients in an LDLT setting. Recipients with the favorable rs12979860 genotype receiving antiviral therapy before transplant are also more likely to achieve SVR. The effect of genotype status in the era of direct-acting antiviral agents will require future study.
Most recently, simeprevir an NS3/4A protease inhibitor with Peg IFN/RBV and sofosbuvir a nucleotide analog NS5B polymerase inhibitor with or without PegIFN/RBV based on the genotype have been approved, providing higher SVR rates. Recurrence of HCV infection after liver transplantation of viremic patients is universal. Recurrent HCV is associated with graft loss, cirrhosis, and death in up to a third of patients at 5 years, leading to a worse outcome when compared to other causes of transplantation (Sallie and others 1994; Aytaman and others 2010). There is currently no standard of care treatment for HCV in the postliver transplant setting, but PegIFN/RBV has been classically used when follow-up liver biopsies demonstrate fibrosis progression (Garcia-Retortillo and Forns 2004). SVR rates in this setting are low, as treatment is characterized by dose reductions, intolerances, and side effects that lead to frequent discontinuation of therapy. When SVR is achieved, there is improved liver histology and survival (Picciotto and others 2007). Early experience with living donor liver transplantation (LDLT) in the setting of chronic HCV infection suggested a less successful outcome than from deceased donors (Takada and others 2006), whereas a recent US experience indicates that donor type does not affect the probability of advanced fibrosis and graft survival in HCV-infected recipients (Terrault and others 2014).
Introduction
H
epatitis C infection is a common blood-borne disease, affecting 2%–3% of the world population. In developed countries, including the United States, the prevalence is less than 2%. Up to 85% of individuals with an acute HCV infection develop chronic hepatitis C. Among chronically infected individuals, 20% develop cirrhosis and 1%–5% will develop hepatocellular carcinoma (HCC). Overall, 5%–7% of HCV patients ultimately die as a result of the infection. HCV infection is the most common reason for liver transplantation in the United States (CDC 2014; WHO 2013). For many years, the standard of care therapy for HCV was the combined use of peginterferon (PegIFN) and ribavirin (RBV). Treatment duration varied according to the HCV genotype. Sustained virologic response (SVR), defined as continued aviremia for 24 weeks after the end of treatment, was usually obtained in 40%–50% for HCV genotype 1 (most common) and 80% for HCV genotype 2 and 3. SVR is associated with virological cure and with improved morbidity and mortality (Ghany and others 2011). In 2011, the direct-acting antiviral agents (DAA) boceprevir and telaprevir were approved for combined use with PegIFN/RBV for the treatment of HCV type 1 becoming the new standard of care (Ghany and others 2011).
1 Medical Genetics, 3Public Health Sciences, 4Center for Bioinformatics, 5Transplant Surgery, 6Gastroenterology and Hepatology, Henry Ford Health System, Detroit, Michigan. 2 Department of Transplant Surgery, University of Tennessee/Methodist University Transplant Institute, Memphis, Tennessee. *Current affiliation: GeneDx, Gaithersburg, Maryland.
1
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2
Pharmacogenetic analysis can be used to assist physicians in determining the proper medication and dose for therapeutic regimens. With respect to HCV treatment, the favorable alleles for the genetic markers, rs12979860 (C) and rs8099917 (T), have been shown to be associated with spontaneous HCV clearance and favorable response to PegIFN/RBV therapy. The unfavorable alleles (rs12979860 T and rs8099917 G) are associated with decreased likelihood of SVR, unfavorable response to antiviral therapy, HCV-associated liver cirrhosis, and HCC (Ge and others 2009; Suppiah and others 2009; Tanaka and others 2009; Thomas and others 2009; Rauch and others 2010; Fabris and others 2011). These markers are located on chromosome 19q13.13 near the IFNL3 gene (formerly known as IL28B). This gene encodes a cytokine with immunomodulatory activity. Its expression is induced by viral infection. IFNL3 is a member of the lambda interferon gene family. Lambda interferons have been shown to inhibit HCV replication in vitro and are hypothesized to initiate an antiviral response through the JAKSTAT pathway and play an important role in Toll-like receptor-induced antiviral and immune defense in the intestinal epithelium (Gad and others 2009; Li and others 2009; Chinnaswamy 2014; GeneCards 2014). IFNL3 genotyping is available on a clinical basis in North America and Europe. The recipient and donor liver rs12979860 genotype has been shown to be associated with response to HCV antiviral therapy in patients after orthotopic liver transplantation (OLT) (Charlton and others 2011). The rs8099917 genotype has been shown to be associated with SVR in recipient–donor pairs undergoing LDLT for HCV-related liver disease in Japan (Fukuhara and others 2010). The association between both of these markers and the response to PegIFN/RBV-based therapy among the LDLT HCV patient population in the United States has not been previously characterized. We retrospectively investigated the association of donor and recipient rs12979860 and rs8099917 genotypes with SVR at a large US liver transplant center.
MONAGHAN ET AL.
Results Donor ages ranged from 20 to 51 (mean 36.8 years). Ten were male (56%) and 8 were female. Seventeen donors were white and 1 was African American. Six of the recipient–donor pairs were unrelated and 12 were related (all first-degree relationships). Recipient characteristics are summarized in Table 1. Recipient ages ranged from 40 to 69 (mean 53.6 years). Three recipients had a diagnosis of HCC. The majority of patients had HCV type 1. The HCV genotype was not available for 1 recipient (now deceased) who had negative HCV viral counts and whose records were no longer available to us for review. Among the patients who received antiviral therapy, 5 achieved SVR. The 1 patient with ongoing therapy was not responding to treatment at the time of submission of this article and has no significant log drop in viral titer. Four patients required retransplantation for reasons, including allograph failure, hepatic artery thrombosis, and recurrent cholestatic hepatitis C. Among white recipients (n = 15) and donors (n = 17), the rs12979860 favorable C allele (regardless of zygosity) frequency was 0.57 and 0.65, compared with a frequency of 0.73 for the Centre d’Etude du Polymorphisme Humain (CEPH) Utah Americans of western and northern European descent (CEU) subjects in the 1000 genomes project. The rs8099917 favorable T allele frequency was 0.77 and 0.74 for white recipients and donors, respectively. The comparable value in the CEU subjects was 0.84. Using the CEU data for both SNPs, the estimated pairwise linkage disequilibrium r2 measure between the 2 SNPs was 0.47. Due to the small number of African American and Hispanic donors and recipients, the allele frequency was not calculated for these groups. SNP associations with SVR status are displayed in Table 2. Before transplant, 11 patients received antiviral therapy (Table 2). Of these, 3 were homozygous for rs12979860 C. Two (67%) of these individuals achieved SVR before transplant in comparison with none of the 8 nonhomozygous subjects (P = 0.055). Recipient rs8099917 T homozygosity was
Materials and Methods Twenty-two consecutive patients underwent LDLT for HCV-related cirrhosis and HCC between 2000 and 2012 at the Henry Ford Health System in Detroit, MI (ClinicalTrails.gov: NCT 01429155). Of these 22 patients, 18 recipient–donor pairs and 1 recipient only were available for genotyping. Residual extracted DNA from the liver donors had been routinely stored at the Henry Ford Immunology laboratory. Liver explants (recipients) fixed in paraffin were identified in the Pathology laboratory. DNA was extracted from donor whole blood and paraffin-fixed livers of recipients using standard methods. Genotyping of the rs12979860 (C/T) and rs8099917 (T/G) single-nucleotide polymorphisms (SNPs) was performed using a previously described single-nucleotide extension (SNE) assay (Melis and others 2011). SNE results were confirmed for 5 DNA samples of various genotypes using Sanger sequencing. Given prior findings, a recessive genetic model was used to test for recipient and donor genotype associations between each SNP (rs12979860 C and rs8099917 T) and SVR status. Due to the sample size, Fisher’s exact tests were used to evaluate the significance of these associations. This study was approved by the HFHS Institutional Review Board.
Table 1. Summary of Recipient (n = 19) Characteristics n Gender Male Female Race/Ethnicity White African American Hispanic HCV genotype Type 1 Type 3 Unknown Antiviral therapy (pegIFN/RBV) None Pretransplant only Post-transplant only Pre- and post-transplanta
12 7 15 1 3 15 3 1 5 3 3 8
a Includes the 1 patient still undergoing therapy at the time of the data analysis. HCV, hepatitis C virus.
HEPATITIS C LIVING DONOR LIVER TRANSPLANT AND IL28-B
Table 2.
3
Donor and Recipient Genotype Associations with Pre- and Post-Transplant SVR Pretransplant antiviral
SNP
Genotype source
rs12979860
Donor Recipient
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rs8099917
Donor Recipient
SVR count Genotype CC TT/CT CC TT/CT TT GG/GT TT GG/GT
Yes
No
SVR% by genotype
1 1 2 0 1 1 2 0
3 5 1 8 4 4 4 5
25 17 67 0 20 20 33 0
Post-transplant antiviral SVR count Pa
Yes
No
SVR% by genotype
1.000
3 0 0 3 3 0 0 3
1 5 2 4 2 4 5 2
75 0 0 43 60 0 0 60
0.055 1.000 0.455
Pa 0.048 0.500 0.167 0.167
Eight of the subjects received antivirals, both pre- and post-transplantation, and contributed data to both pre- and post-transplant SVR outcome analyses. Three patients were only evaluated for pretransplant SVR, and another 3 were only evaluated for post-transplant SVR. a Each P value was calculated using Fisher’s exact test. SNP, single-nucleotide polymorphism; SVR, sustained virologic response.
not associated with SVR before transplantation (P = 0.455), although those with a favorable genotype had a higher proportion achieving SVR (33% vs. 0%; Table 2). Post-transplant, 11 patients received therapy (Table 2), 8 of whom were also treated pretransplant (ie, 3 were only treated post-transplant). For 1 LDLT pair, the donor was not available for genotyping. Another patient with unfavorable donor genotypes at both SNPs is still receiving therapy, but at the time of submission of this article had a poor response to therapy. These 2 patients were removed from the analysis, leaving a post-transplant sample of 9 subjects. Of these, 4 of the donors were homozygous for rs12979860 C. Three (75%) patients with these homozygous donor genotypes achieved SVR in comparison with none of the 5 recipients whose donors were nonhomozygous (P = 0.048). SNP rs8099917 T showed a similar trend, with 60% of T allele homozygotes experiencing SVR (Table 2), but this result did not achieve statistical significance (P = 0.167). Favorable recipient genotypes at either SNP were not associated with post-transplant SVR (Table 2). Five of the 19 recipients were excluded from the analysis to determine HCV association between the genotype and SVR because they did not receive antiviral therapy. None of these patients had spontaneous viral clearance.
Conclusions The present report validates the use of an individual patient’s pharmacogenetics profile to guide decisions regarding prevention, diagnosis, and treatment of disease. The presence of a favorable homozygous genotype for both SNPs was associated with an increased rate of SVR following PegIFN/RBV therapy. Knowledge of a patient’s genotype to predict success of therapy, therefore, can be helpful due to the cost of the therapy and high discontinuation rate due to adverse side effects. In patients whose genotype predicts a lower chance of SVR with antiviral therapy, perhaps a shorter duration of treatment or alternative medication may be considered (Pacanowski and others 2012). Genotype analysis for rs12979860 and rs8099917 SNPs near the gene commonly known as IL28B is currently available on a clinical basis. The IL28B rs12979860 genotype information is included in the pharmacogenomics section of the package insert labels for peginterferon alfa-2b and the DAAs, boceprevir and telaprevir.
The recipient and donor liver rs12979860 genotype has been shown to be associated with response to HCV antiviral therapy in patients after OLT (Charlton and others 2011). Among HCVinfected patients who underwent OLT and received antiviral therapy, the presence of a favorable rs12979860 CC donor or recipient genotype was associated with SVR. SVR was lowest when the genotypes of the donor and recipient were unfavorable, increased if either the donor or recipient genotype was favorable, and most likely with both a favorable donor and recipient homozygous genotype. LDLT provides an opportunity to test for the IFNL3 genotype before surgery. A recent study in Japan demonstrated an association between the rs8099917 genotype and SVR in recipient–donor pairs undergoing LDLT for HCV-related liver disease (Fukuhara and others 2010). The highest SVR rate was observed in recipient–donor pairs, where both recipient and donor were homozygous for the favorable (T) allele. When 1 member of the donor–recipient pair had a homozygous favorable genotype and the other member had a nonfavorable genotype, the rate of SVR was lower (Fukuhara 2012, personal communication). A potential limitation of the current study is the small number of patients investigated. Based on this small number of LDLT recipient–donor pairs in the United States, our results suggest that transplantation of an rs12979860 CCpositive graft confers a favorable post-transplant antiviral response among HCV-positive recipients in an LDLT setting. While the donor status was associated with the highest likelihood of SVR post-transplant, a favorable recipient genotype at rs12979860 was also beneficial before surgery. Our data supports prior reports among orthotopic liver transplants suggesting that donor IL28B status affects IFN-based therapy response. The value of studying a living donor population in the United States validates previous results in the deceased donor setting; additionally, such pretransplant testing of the donor might influence the donor selection process. Given the potential clinical utility of these findings, larger studies are warranted to confirm these results. Such studies could involve a collaborative effort between multiple institutions, the analysis of additional SNPs, patients of other races/ethnic backgrounds, and in particular the role of these SNPs as they relate to the newer DAA medications that will soon replace IFN-based HCV therapies.
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Acknowledgment Genotyping studies were paid for through the Henry Ford Gastroenterology Hepatology Research Fund.
Author Disclosure Statement No competing financial interests exist.
Journal of Interferon & Cytokine Research Downloaded from online.liebertpub.com by Univ Braunschweig on 11/17/14. For personal use only.
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Address correspondence to: Dr. Humberto C. Gonzalez Department of Transplant Surgery University of Tennessee/Methodist University Transplant Institute 1211 Union Ave. Suite 340 Memphis, TN 38104 E-mail: [email protected] Received 3 July 2014/Accepted 10 September 2014
