Basic Science: Concise Communications
The transforming growth factor-β high-producer genotype is associated with response to hepatitis C virus-specific therapy in HIV-positive patients with acute hepatitis C
Nattermann, Jacoba; Vogel, Martina; Nischalke, Hans Dietera; Danta, Markb; Ahlenstiel, Goloa; Michalk, Monikaa; Sauerbruch, Tilmana; Rockstroh, Jürgen Ka; Spengler, Ulricha; for the Kompetenznetz HIV/AIDS
aDepartment of Internal Medicine I, University of Bonn, Bonn, Germany
bFaculty of Medicine UNSW, St Vincent's Clinical School, Sydney, Australia.
Correspondence to J. Nattermann, Department of Internal Medicine I, University of Bonn, Sigmund Freud Straße 25, D-53105 Bonn, Germany. Tel: +49 228 287 15383; fax: +49 228 287 14611; e-mail: email@example.com
Background: Coinfection with the hepatitis C virus (HCV) in HIV-positive patients is an emerging health problem. The factors affecting response to HCV-specific therapy are poorly understood but may involve host genetic factors. HCV NS5A-induced inhibition of transforming growth factor-β signaling has been suggested as a potential mechanism involved in HCV pathogenesis. Transforming growth factor-β, a multifunctional cytokine, displays gene polymorphisms (transforming growth factor-β codon 10T/C and codon 25G/C) associated with differential cytokine secretion. Here, we studied whether transforming growth factor-β gene polymorphisms affect treatment response in HCV/HIV coinfection.
Methods: Transforming growth factor-β genotypes were determined in 60 HIV-positive patients with acute hepatitis C treated with pegylated interferon-α. Patients were classified into those with a high-producer genotype and others with non-high-producer genotypes. Rates of sustained virological responses were compared between high-producer and non-high-producer patients. As a control, 100 healthy, 201 HIV(+)/HCV(−), and 148 HCV(+)/HIV(−) subjects were studied.
Results: Transforming growth factor-β genotype distribution did not differ significantly between the groups. In HIV/HCV coinfection carriers of the transforming growth factor-β high-producer genotype had significantly higher sustained virological response rates than patients with a transforming growth factor-β non-high-producer genotype (75 vs. 41.7%; P = 0.039). In a forward-conditional stepwise regression model, transforming growth factor-β high-producer genotype was confirmed as an independent positive predictor for sustained virological response in interferon-α therapy (odds ratio, 4.4; 95% confidence interval, 1.5–13.4; P = 0.009).
Conclusion: Response rates to interferon-α therapy are enhanced in acute HCV-infected HIV-positive patients carrying the transforming growth factor-β ‘high-producer’ genotype. This finding may indicate that a transforming growth factor-β ‘high-producer’ state can partially compensate HCV NS5A-induced inhibition of transforming growth factor-β signaling.
In Europe and the USA, about 30% of HIV-positive individuals are coinfected with the hepatitis C virus (HCV) , and currently outbreaks of acute hepatitis C in HIV-infected men have been reported. HCV/HIV coinfection is characterized by more rapid progression toward severe liver disease [2–4] resulting in a higher overall mortality  than in individuals with HCV monoinfection [1,6]. Therefore, HCV coinfection has become a major health problem in HIV-infected patients, especially after the dramatic decline in morbidity and mortality due to opportunistic diseases following the introduction of HAART in 1996. Previous historical studies suggest that individuals with concomitant HIV infection are far less likely to spontaneously clear HCV than patients with HCV monoinfection. It is currently recommended that individuals who are acutely infected with HCV should undergo monitoring of HCV viral load levels to determine whether spontaneous clearance is likely or whether the opportunity for early treatment should be considered .
At the moment, combination of pegylated interferon-α with ribavirin represents the backbone of HCV-specific therapy. With interferon-based combination therapy sustained viral response (SVR) is, however, achieved in only about 40% of chronically infected patients  in clinical studies and may be even lower in clinical practice .
HCV genotype and HCV viral load are major determinants of response to treatment in HCV infection. Nevertheless, host genetic factors have also been shown to influence the natural course of infection as well as response to treatment.
Cytokines play a key role in regulating antiviral immune responses and a variety of cytokine gene polymorphisms have been shown to affect natural course of and treatment response in HCV monoinfection. Transforming growth factor (TGF)-β is a multifunctional cytokine that is involved in control of cell growth, differentiation, and apoptosis of cells. In addition, TGF-β has a major regulatory role in hepatic fibrosis and cirrhosis. Importantly, the TGF-β gene is polymorphic (codon 10 and 25) leading to differences in cytokine production. Thus, patients can be classified in high, intermediate, or low producers of TGF-β according to their TGF-β genotype.
Here, we analyzed the impact of the TGF-polymorphism in response to HCV-specific therapy in HIV-positive patients with acute hepatitis C.
Patients and methods
Design and study populations
Sixty HIV-positive white patients with acute hepatitis C were enrolled into this study. Acute hepatitis C was diagnosed when at least two of the following three criteria were fulfilled within the past 4 months prior to the diagnosis of HCV infection: HCV seroconversion; alanine aminotransferase (ALT) more than 350 IU with prior normal aminotransferases, and risk exposure to HCV (modified from ). HCV/HIV coinfected patients were treated with pegylated interferon-α and ribavirin (800 mg/day) for 24 (HCV genotypes 2 and 3) or 48 weeks (genotypes 1 and 4). Patients who were HCV RNA negative 6 months after the end of treatment were classified as sustained virological responders. All other patients were considered to be non-responders.
As a control, 148 HCV monoinfected, 201 HIV monoinfected patients, and 100 healthy individuals were included to analyze the distribution of the TGF-β genotypes.
Informed consent was obtained from each patient prior to inclusion into the study, and the study conformed to the ethical guidelines of the Helsinki declaration as approved by the local ethics committees.
TGF-β genotypes (TGF-β codon 10T/C and codon 25G/C) were analyzed using the cytokine genotyping tray (One Lambda, Canoga Park, California, USA) following the manufacturer's protocol allowing classification into TGF-β high (10T/T 25G/G; 10T/C 25G/G), intermediate (10T/C 25G/C; 10C/G 25G/G; 10T/T 25G/C), and low (10C/C 25G/C; 10C/C 25C/C; 10T/T 26C/C; 10T/C 25C/C) producers.
Cytokine genotype distribution as well as treatment response rates in patients with different cytokine genotypes were analyzed using 2 × 2 contingency tables. For statistical comparisons between the groups, χ2 statistics, Fisher's exact test, and Mann–Whitney U test were used as appropriate. To determine the effect of the TGF-β genotype in comparison to HCV load and genotype, the main established predictors of response, we stratified our patient data as HCV genotype 1 versus non-HCV genotype 1, and HCV RNA 2.0 million copies/ml or less versus more than 2.0 million copies/ml [8,9]. These stratified parameters were analyzed together with TGF-β producer status in a forward-conditional stepwise logistic regression model using SVR as outcome variable. P values less than 0.05 (two-sided) were regarded as significant.
Sixty HIV-positive patients with acute hepatitis C [age, mean (range) 39.2 (25–70) years; CD4 cell count, 498.2 (148–969) cells/μl; HIV load, 26.6 × 103 (50–341 × 103) copies/ml; ALT, 558.4 (24–3089) IU/ml; γ-glutamyl transpeptidase (GT), 396.6 (77–962) IU/ml; HCV load, 6.5 × 106 (4.5 × 103–47 × 106) copies/ml] were enrolled into this study (Table 1). Patients were treated with pegylated interferon-α plus body weight adapted doses of ribavirin for 24 (genotypes 2 and 3) and 48 (genotypes 1 and 4) weeks, respectively. Overall, an end-of-treatment response (ETR) was seen in 76.7% (46/60) and a SVR rate was achieved in 68.3% (41/60) of treated patients.
Distribution of transforming growth factor-β genotypes
Distribution of TGF-β genotypes in HIV/HCV coinfected patients [high producer 48/60 (80.4%), intermediate producer 11/60 (18.3%), low producer 1/60 (1.6%)] was comparable with that seen in healthy individuals [high producer 82/100 (82%), intermediate producer 18 (18%), low producer 0 (0%)], HIV monoinfected patients [high producer 158/201 (78.6%), intermediate producer 43/201 (21.4%), low producer 0 (0%)], and HCV monoinfected subjects [low producer 111/148 (75%), intermediate producer 35/148 (23.6%), low producer 2/148 (1.3%)], respectively. All distributions were in accordance with the Hardy–Weinberg equilibrium. Furthermore, no significant differences could be detected for demographic variables, route of infection, liver enzymes, HCV genotype, HCV or HIV load between the TGF-β genotypes in HCV/HIV coinfected patients (data not shown).
Transforming growth factor-β genotype and treatment response
Regarding the low prevalence of the low producer genotype, data analysis was based on patients with a high-producer genotype in comparison to carriers of other genotypes (non-high producer).
Patients with a high-producer genotype were more likely to achieve an ETR as compared to carriers of a non-high-producer genotype (82.4 versus 58.3%), although this difference failed to reach statistical significance (Fig. 1a, left graph). We, however, found coinfected carriers of a high-producer genotype to have a significantly higher SVR rate than non-high-producer patients (75 versus 41.7%; P = 0.039) (Fig. 1b, left graph).
When patients were stratified according to HCV genotypes, patients with genotype 1 infection had a lower SVR rate as compared to patients infected with non-HCV genotype 1 infection (66.7 versus 76.9.6%; P = n.s.).
Importantly, an effect of the TGF-β genotypes on treatment outcome was only seen in the subgroup of patients with HCV genotypes 1 (P = 0.013), whereas SVR rates were not significantly affected by the TGF-β genotype in patients infected with other HCV genotypes (Fig. 1b, right graph).
In addition, we analyzed our data in a stepwise forward-conditional regression model (Table 2). In the model the TGF-β high-producer genotype remained a strong predictor for SVR therapy (odds ratio 5.6; 95% confidence interval CI 1.3–25.7; P = 0.02), whereas HCV load and age were removed from the final regression model.
Response to HCV-specific therapy depends on both viral and host factors. Among the host factors, cytokine gene polymorphisms have been shown to affect the response to antiviral therapy. For instance, the tumor necrosis factor-α promoter polymorphism at position −308  and the interferon (IFN)-γ promoter single nucleotide polymorphism (SNP) −764G/C  have been shown to predict response to combination therapy in HCV infection.
Here, we studied the impact of the TGF-β gene polymorphism in response to HCV-specific treatment in HIV(+) patients with acute hepatitis C and identified the TGF-β high-producer genotype as an independent predictor of SVR. Performing a subgroup analysis, we found that the association between the TGF-β genotype and treatment response was restricted to patients with a HCV genotype 1 infection. This observation, however, needs to be balanced against the relatively small number of patients infected with a genotype other than genotype 1 in our study. Thus, we cannot exclude with certainty that the TGF-β genotype also affects treatment response in HIV-positive patients infected with HCV genotypes other than genotype 1.
TGF-β is a profibrinogenic cytokine, which has been shown to be involved in development of fibrosis in hepatocytes of patients with chronic viral hepatitis. In addition, TGF-β has been implicated in suppression of immune responses and downmodulation of growth promoting activities. Therefore, the identification of the TGF-β high-producer genotype as a predictor of response to HCV therapy is primarily counterintuitive. Supportive data are, however, obtained by Tambur et al. , who reported the TGF-β high-producer genotype to be strongly associated with the absence of HCV recurrence following liver transplantation. Data obtained by Erard et al.  imply a dual function of TGF-β. While TGF-β alone suppresses immune responses, simultaneous presentation together with interleukin (IL)-4 favored differentiation of CD8 cells toward the type 1 response. Namely, addition of both TGF-β and IL-4 has been shown to induce IFN-γ secretion by CD8-positive cell cultures and to enhance CD8 cytotoxicity. Accordingly, McKeirl et al. reported that TGF-β can downregulate HIV-1 viral replication in vitro. With respect to hepatitis C, HCV NS5A-mediated suppression of TGF-β signaling has been suggested as a step in HCV pathogenesis. Thus, the TGF-β high-producer genotype might partially compensate for the HCV-induced inhibition of TGF-β signaling thereby enabling the generation of a type 1 response. In this context it is important to note that strong antiviral type 1 immune responses have been proposed to be associated with viral clearance both during the acute phase of primary HCV infection and antiviral therapy of chronic hepatitis [14–16].
Whether TGF-β polymorphisms also affects spontaneous elimination of HCV in HIV-positive subjects remains to be clarified. Kimura et al.  recently showed that the −509CC genotype and the −509C allele were significantly associated with higher HCV clearance rates and with lower transcriptional activity. Nevertheless, our finding that HCV/HIV coinfected subjects displayed a similar TGF-β genotype distribution to that of healthy controls and HIV monoinfected subjects, argues against a significant impact on the natural course of HCV infection. Accordingly, Suzuki et al.  did not find any significant differences regarding TGF-β genotype distributions between HCV monoinfected patients and healthy control subjects. Likewise, Barrett et al.  found no association between TGF-β polymorphisms and natural course of hepatitis C monoinfection whereas Gewaltig et al.  proposed an association of polymorphisms of the TGF-β gene with the rate of progression of HCV-induced liver fibrosis. Thus, further studies are needed to clarify this issue.
Our data, however, support the importance of genetic factors for the response to HCV-specific therapy in HIV-infected patients with acute hepatitis C.
This work was supported by the Hector-Stiftung (M42).
1. Rockstroh JK, Spengler U. HIV and hepatitis C virus coinfection. Lancet Infect Dis 2004; 4:437–444.
2. Giordano TP, Kramer JR, Souchek J, Richardson P, El-Serag HB. Cirrhosis and hepatocellular carcinoma in HIV-infected veterans with and without the hepatitis C virus: a cohort study, 1992-2001. Arch Intern Med 2004; 164:2349–2354.
3. Pineda JA, Romero-Gómez M, Díaz-García F, Girón-González JA, Montero JL, Torre-Cisneros J, et al. HIV coinfection shortens the survival of patients with hepatitis C virus-related decompensated cirrhosis. Hepatology 2005; 41:779–789.
4. Kramer JR, Giordano TP, Souchek J, El-Serag HB. Hepatitis C coinfection increases the risk of fulminant hepatic failure in patients with HIV in the HAART era. J Hepatol 2005; 42:309–314.
5. El-Serag HB, Giordano TP, Kramer J, Richardson P, Souchek J. Survival in hepatitis C and HIV co-infection: a cohort study of hospitalized veterans. Clin Gastroenterol Hepatol 2005; 3:175–183.
6. Qurishi N, Kreuzberg C, Lüchters G, Effenberger W, Kupfer B, Sauerbruch T, et al. Effect of antiretroviral therapy on liver-related mortality in patients with HIV and hepatitis C virus coinfection. Lancet 2003; 362:1708–1713.
7. Brejt N, Gilleece Y, Fisher M. Acute hepatitis C: changing epidemiology and association with HIV infection. J HIV Ther 2007; 12:3–6.
8. Falck-Ytter Y, Kale H, Mullen KD, Sarbah SA, Sorescu L, McCullough AJ. Surprisingly small effect of antiviral treatment in patients with hepatitis C. Ann Intern Med 2002; 136:288–292.
9. Jaeckel E, Cornberg M, Wedemeyer H, Santantonio T, Mayer J, Zankel M, et al. Treatment of acute hepatitis C with interferon alfa-2b. N Engl J Med 2001; 345:1452–1457.
10. Dai CY, Chuang WL, Chang WY, Chen SC, Lee LP, Hsieh MY, et al. Tumor necrosis factor-alpha promoter polymorphism at position −308 predicts response to combination therapy in hepatitis C virus infection. J Infect Dis 2006; 193:98–101.
11. Huang Y, Yang H, Borg BB, Su X, Rhodes SL, Yang K, et al. A functional SNP of interferon-gamma gene is important for interferon-alpha-induced and spontaneous recovery from hepatitis C virus infection. Proc Natl Acad Sci USA 2007; 104:985–990.
12. Tambur AR, Ortegel JW, Ben-Ari Z, Shabtai E, Klein T, Michowiz R, et al. Role of cytokine gene polymorphism in hepatitis C recurrence and allograft rejection among liver transplant recipients. Transplantation 2001; 71:1475–1480.
13. Erard F, Garcia-Sanz JA, Moriggl R, Wild MT. Presence or absence of TGF-beta determines IL-4-induced generation of type 1 or type 2 CD8 T cell subsets. J Immunol 1999; 162:209–214.
14. Missale G, Bertoni R, Lamonaca V, Valli A, Massari M, Mori C, et al. Different clinical behaviors of acute hepatitis C virus infection are associated with different vigor of the antiviral cell-mediated immune response. J Clin Invest 1996; 98:706–714.
15. Diepolder HM, Zachoval R, Hoffmann RM, Wierenga EA, Santantonio T, Jung MC, et al. Possible mechanism involving T-lymphocyte response to nonstructural protein 3 in viral clearance in acute hepatitis C virus infection. Lancet 1995; 346:1006–1007.
16. Spengler U, Nattermann J. Immunopathogenesis in hepatitis C virus cirrhosis. Clin Sci (Lond) 2007; 112:141–155.
17. Kimura T, Saito T, Yoshimura M, Yixuan S, Baba M, Ji G, et al. Association of transforming growth factor-beta 1 functional polymorphisms with natural clearance of hepatitis C virus. J Infect Dis 2006; 193:1371–1374.
18. Suzuki S, Tanaka Y, Orito E, Sugauchi F, Hasegawa I, Sakurai M, et al. Transforming growth factor-beta-1 genetic polymorphism in Japanese patients with chronic hepatitis C virus infection. J Gastroenterol Hepatol 2003; 18:1139–1143.
19. Barrett S, Collins M, Kenny C, Ryan E, Keane CO, Crowe J. Polymorphisms in tumour necrosis factor-alpha, transforming growth factor-beta, interleukin-10, interleukin-6, interferon-gamma, and outcome of hepatitis C virus infection. J Med Virol 2003; 71:212–218.
20. Gewaltig J, Mangasser-Stephan K, Gartung C, Biesterfeld S, Gressner AM. Association of polymorphisms of the transforming growth factor-beta1 gene with the rate of progression of HCV-induced liver fibrosis. Clin Chim Acta 2002; 316:83–94.
gene polymorphism; hepatitis C virus; HCV/HIV coinfection; HIV; therapy; transforming growth factor-β
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