A randomized controlled trial of amantadine plus interferon-α2a vs. interferon-α2a alone in naive patients with chronic hepatitis C randomized according to the early virological response to interferon-α2a monotherapy
Professor M. Angelico, Gastroenterology and Hepatology Unit, University of Rome ‘Tor Vergata’, S. Giovanni Calibita Fatebenefratelli Hospital, Isola Tiberina, 39, 00186 Rome, Italy.
Background : An early virological response to interferon-α treatment is a strong predictor of sustained response, but it has never been exploited to stratify patients in clinical trials.
Aim : To evaluate the efficacy of amantadine plus interferon-α compared with interferon-α alone in naive patients with chronic hepatitis C who were randomized on the basis of the early virological response to interferon-α.
Methods : One hundred and eighty-one patients received recombinant interferon-α2a (3 MU three times weekly) for 2 months and 164 were evaluated for early (i.e. month 2) virological response. Hepatitis C virus (HCV) RNA-negative patients (n = 66) were randomized to receive 3 MU of interferon-α three times weekly, with or without amantadine (200 mg/day); HCV RNA-positive patients (n = 98) were randomized to receive 6 MU of interferon-α three times weekly, with or without amantadine (200 mg/day). HCV RNA-positive patients at 6 months discontinued treatment, and all others completed 12 months.
Results : At month 6, HCV RNA-negative patients made up 54.2% of the interferon + amantadine group and 42.0% of the monotherapy group (P = 0.07). At month 12, HCV RNA-negative patients made up 38.5% of the interferon + amantadine group and 28.4% of the monotherapy group (N.S.). The sustained virological response rates were 21.6% and 20.9%, respectively (N.S.).
Conclusion : The addition of amantadine does not enhance the sustained virological response to interferon-α in naive patients with chronic hepatitis C; however, an additive effect of amantadine occurs in the first 6 months, mainly in patients without an early response to monotherapy. Early response to interferon-α is a strong predictor of sustained virological response.
Chronic hepatitis C is a leading cause of chronic liver disease in Western countries. Hepatitis C virus (HCV)-related liver disease is a major public health issue because of its propensity to progress to chronic hepatitis, cirrhosis and hepatocellular carcinoma. The current standard of care for patients with chronic hepatitis C is the combination of pegylated interferon-α and ribavirin. Unfortunately, approximately 45% of patients do not exhibit a sustained response.1, 2 In addition, this therapy frequently causes side-effects necessitating dose reductions or discontinuation of therapy. Thus, there is an obvious need for alternative and/or additional forms of treatment.
In recent years, a favourable role of amantadine in the treatment of chronic hepatitis C has been suggested. Amantadine is a tricyclic amine with anti-viral activity, which was developed in the 1960s and registered in many countries for the prevention of influenza A infection and the treatment of Parkinson's disease.3 Although the anti-viral mechanism of action of amantadine in chronic hepatitis C remains unclear, it may be related to the fact that amantadine can achieve high concentrations in the liver,3 and may block events occurring in late viral uncoating or in early transcription.4 So far, several clinical investigations in patients with chronic hepatitis C have shown that amantadine, when used as monotherapy, has no significant effects on the levels of HCV RNA, although a potential anti-inflammatory activity has been observed.5–10 Controversial results have been reported when amantadine is used in combination with standard interferon-α in naive patients,11–15 and conflicting data have also been observed on the efficacy of combined treatment in patients who have failed or relapsed after a previous interferon-α course.16–20 Finally, triple therapy based on a combination of interferon-α, ribavirin and amantadine seems to show improved efficacy when compared with the standard combination of interferon-α and ribavirin in the ‘difficult-to-treat’ group of non-responders to previous treatment,21, 22 but this finding awaits confirmation in larger trials.
Several studies have shown that the assessment of early viral clearance during anti-viral treatment for hepatitis C is a strong predictor of the later occurrence of a sustained virological response, therefore allowing, in principle, the optimization of the strategy of long-term therapy.23–26 However, to our knowledge, this concept has never been exploited to stratify or randomize patients in clinical trials. Accordingly, the aim of this study was to evaluate the efficacy of the combination of interferon-α and amantadine compared with interferon-α alone in a cohort of naive patients with chronic hepatitis C who were stratified and randomized on the basis of the early virological response to an initial short course of interferon-α monotherapy.
One hundred and eighty-one patients (122 males and 59 females), aged 18–65 years (median age, 40 years), were included in the study. Patients were recruited from 14 hospitals in central Italy between January 2000 and December 2000. HCV infection was community acquired in 91 patients. Documented risk factors were present in the remaining 90 patients, including blood transfusions in 14 patients and drug abuse in 55 patients. Patients were considered to be eligible if they met the following inclusion criteria: age between 18 and 65 years; presence of anti-HCV antibodies detected by a third-generation enzyme-linked immunoabsorbent assay; positive serum HCV RNA by standard polymerase chain reaction; persistent elevation of serum alanine transaminase (≥ 1.5 times the upper limit of normal) during the 12 months prior to the study; and a histological diagnosis of chronic hepatitis on a liver biopsy sample taken in the preceding 6 months. Patients were excluded if they had the following: presence of hepatitis B surface antigen in serum or markers of human immunodeficiency virus infection; recent or active alcohol and/or drug abuse; platelet count below 70 000/mL or leucocyte count below 3000/mL; histological evidence of cirrhosis; and evidence of autoimmune or genetic liver diseases, malignancies or renal, haematological, cardiopulmonary, neurological and other gastrointestinal diseases.
This was an open-label, randomized controlled trial, with blind evaluation, in naive patients with chronic HCV infection. All patients fulfilling the inclusion/exclusion criteria received an initial treatment course of 3 MU of recombinant interferon-α2a, given subcutaneously three times weekly for 2 months. Patients were then divided into two groups according to the serum HCV RNA status determined by a polymerase chain reaction assay with a detection limit of 500 IU/mL (HCV RNA-negative or HCV RNA-positive). Patients in each group were randomly assigned to receive interferon-α alone or interferon-α plus amantadine (200 mg/day). In addition, in the HCV RNA-positive group, the dose of interferon-α was increased to 6 MU three times weekly. At the end of month 6 of treatment, the HCV RNA status was re-assessed. All HCV RNA-positive patients were withdrawn from therapy, whereas HCV RNA-negative patients continued treatment until month 12 according to their initial randomization.
Randomization was carried out using the method of sealed envelopes and a computer-generated randomization list. During the study, patients underwent routine clinical and biochemical examinations to assess treatment efficacy and safety at weeks 2, 4, 6 and 8 and every month thereafter. Compliance to treatment was assessed by written records. Non-compliance was defined as an unjustified interruption of treatment for more than 15 days, or a failure to attend two consecutive scheduled visits. The primary end-point of the study was to determine whether the addition of amantadine to standard interferon-α increased the rate of sustained virological response, conventionally defined as the absence of measurable serum HCV RNA at month 6 after treatment completion. Secondary end-points were to define the predictive value of an early response to interferon-α monotherapy and whether stratification and randomization according to the early response to interferon-α allowed better short-term and sustained treatment efficacy to be achieved. The results were evaluated by intention-to-treat analysis.
All patients who met the entry criteria provided written informed consent. The study protocol, which conformed with the ethical guidelines of the 1975 Declaration of Helsinki, was approved by the Ethical Committee of the University of Rome ‘Tor Vergata’ and the committees of collaborating centres.
Quantitative HCV RNA measurement and the assessment of HCV genotypes were performed in a single central laboratory on serum samples stored at − 70 °C until analysis. Serum HCV RNA concentrations were measured at baseline, at months 2, 6 and 12 of treatment and at month 6 after treatment completion using a quantitative multicycle reversed transcribed polymerase chain reaction method, with a lower limit of sensitivity of 500 copies/mL (Amplicor HCV Monitor, Roche, Basle, Switzerland). All patients were also tested with a qualitative HCV RNA assay (detection limit, 50 IU/mL) at the end of follow-up for the evaluation of the sustained virological response, which was confirmed in all cases.
HCV genotypes were determined after hybridization of biotin-labelled polymerase chain reaction products to oligonucleotide probes bound in strips on nitrocellulose membranes (Inno-LiPA, Innogenetics, Brussels, Belgium). The HCV genotypes were designated according to the nomenclature proposed by Simmonds.27
A percutaneous liver biopsy was obtained within 6 months of starting treatment. Tissue samples were evaluated by two pathologists who were blind to the treatment groups and response to treatment. The histological grading (0–18) and staging (0–6) were scored according to Ishak et al.28
Statistical analysis was performed using SPSS statistical software V8.0 for Windows (SPSS Inc., Chicago, IL, USA). Data were expressed as the mean ± standard deviation (s.d.) for continuous variables. Student's t-test for unpaired data was used for the comparison of mean values. Proportions and categorical variables were tested by the chi-squared test and the two-tailed Fisher exact method when appropriate. All P values were two-tailed; a P value of less than 0.05 was considered to indicate statistical significance. Multivariate analyses were carried out using stepwise logistic regression models which considered the dichotomous response variables (presence or absence of sustained or early virological response) as dependent variables. The following were entered as independent variables: age (< 40 years or ≥ 40 years), gender, baseline serum HCV RNA and alanine transaminase values (below or above median), early virological response, HCV genotype (1 vs. non-1), histological staging (0–1 vs. 2–6) and grading, previous hepatitis B virus infection, habitual alcohol intake (abstainers vs. drinkers), estimated time from infection and treatment arm (interferon-α plus amantadine vs. interferon-α alone). Intention-to-treat analysis was used for all measures of efficacy. Patients who missed the examination at the end of the 18-month follow-up visit were considered as non-responders. In relation to the primary end-point of the study, the required sample size was calculated on the assumption that interferon-α monotherapy would determine a sustained virological response rate of 15% and that the addition of amantadine to standard interferon-α, to be clinically significant, would increase that rate by 50% (i.e. to 22.5%). With these assumptions, using a two-sided test with a significance level of 0.05 and a power of 90%, a sample size of 85 patients in each treatment group was estimated as adequate.
Participant flow and baseline data
One hundred and eighty-one patients were enrolled and started the initial 2-month treatment course with interferon-α monotherapy (3 MU three times weekly). Seventeen patients (9.4%) dropped out within this period, prior to the assessment of viraemia. At month 2, serum HCV RNA was negative in 66 patients (40%) and positive in 98 patients (60%). These findings were taken as the ‘early response’. The demographic and baseline clinical data of the patients according to the early virological response are reported in Table 1. Predictors of the early response were investigated by multiple regression analysis which included all baseline clinical and virological characteristics as independent variables. This showed that only three variables were strong independent predictors of an early response to interferon-α monotherapy, namely the presence of low basal HCV RNA levels (P = 0.0000), a low histological grading score (P < 0.031) and a non-1 HCV genotype (P < 0.0006).
Table 1. Baseline characteristics of patients according to the early virological response (at month 2) to interferon-α monotherapy
|Patients randomized (n)||66||98|| |
|Mean age (years)||39 ± 14||41 ± 11|| |
|Gender (% male)||71||69|| |
|Duration of infection (years)||4.6 ± 4.3||4.5 ± 4.4|| |
|Alcohol intake (g/day)||24.2 ± 35.6||26.2 ± 40.5|| |
|Alcohol intake > 50 g/day (%)||13.6||10.2|| |
|HbcAb positive (%)||35||45|| |
|Primary mode of transmission (n)|
| Intravenous drug use||23||32|| |
| Blood transfusion|| 2||12|| |
| Community acquired||38||53|| |
|Histological grading (score)||4.5 ± 2.7||5.5 ± 3.3|| |
|Histological staging (score)||1.3 ± 1.1||1.7 ± 1.2||0.0001|
|HCV RNA (copies/mL × 1000)||530 ± 510||901 ± 724||0.002|
|Genotype 1 (%)||40.9||65.3|| |
|ALT (mU/mL)||122 ± 73||113 ± 67|| |
The remaining 164 patients were randomized according to the presence or absence of an early response. Of the HCV RNA-negative patients (early responders), 33 were randomized to receive interferon-α plus amantadine and 33 to receive interferon-α alone; of the HCV RNA-positive patients (early non-responders), 50 received interferon-α plus amantadine and 48 received interferon-α alone. In addition, in all early non-responders, the interferon-α dose was increased from 3 to 6 MU three times weekly. Overall, 83 patients were randomized to receive the combination treatment and 81 to receive monotherapy.
Ten patients (5.5%) dropped out between months 3 and 6 of treatment. At re-assessment of the HCV RNA status at month 6, 75 (41.4%) were HCV RNA positive and, according to the protocol, stopped treatment; 79 patients (43.6%) were HCV RNA negative and therefore continued their assigned treatment without change. Sixty-eight (37.5%) of these 79 patients completed 12 months of treatment, whereas an additional 11 (6%) dropped out between months 6 and 12. No other patient dropped out until the end of follow-up. The demographic and baseline clinical data of the patients treated with or without amantadine are summarized in Table 2. The two groups were comparable with respect to gender, risk factors, baseline alanine transaminase levels, pre-treatment serum HCV RNA titre, distribution of HCV genotypes and histological findings.
Table 2. Demographic and baseline clinical data of patients randomized to receive interferon-α monotherapy or interferon-α plus amantadine
|Patients randomized (n)||81||83|
|Mean age (years)||41 ± 12||39 ± 13|
|Gender (% male)||66||73|
|Duration of infection (years)||4.5 ± 4.4||4.5 ± 4.2|
|Alcohol intake (g/day)||21.9 ± 32.9||28.9 ± 43|
|Alcohol intake > 50 g/day (%)||9.9||13.3|
|HbcAb positive (%)||37||44|
|Primary mode of transmission (n)|
| Intravenous drug use||20||35|
| Blood transfusion||10|| 4|
| Community acquired||48||43|
|Histological grading (score)||5.1 ± 3.2||5.0 ± 3.1|
|Histological staging (score)||1.5 ± 1.1||1.7 ± 1.3|
|HCV RNA (copies/mL × 1000)||738 ± 585||766 ± 747|
|Genotype 1 (%)||56.8||54.2|
|ALT (mU/mL)||110 ± 66||123 ± 73|
Drop-outs and side-effects
A total of 38 patients (21%) dropped out (for various reasons specified below) during the entire study period. The type, frequency and entity of side-effects and laboratory abnormalities were similar in patients treated with or without amantadine. Overall, adverse events were mild or moderate and consisted mainly of flu-like syndrome, fatigue, irritability, depression and insomnia. None of the patients had clinically significant neutropenia, anaemia or decreased platelet count. Only 11 patients (6.0%) discontinued therapy due to treatment-related side-effects, seven after randomization: five treated with interferon-α alone (three due to thyroid diseases and one each due to severe depression and decompensated diabetes) and two treated with amantadine plus interferon-α (one due to thyroid disease and one due to severe depression). All side-effects were reversible after the cessation of therapy. The remaining 27 drop-outs were due to poor compliance.
Virological response after randomization
The virological response (defined as undetectable serum HCV RNA) was evaluated throughout the study (Table 3). At month 6 of treatment, 45 of the 83 patients (54.2%) in the interferon-α and amantadine combination group showed negative HCV RNA vs. 34 of the 81 patients (42%) in the interferon-α monotherapy group, a difference close to statistical significance (P = 0.07). This was entirely due to the larger number of patients with undetectable HCV RNA at month 6 among those who did not exhibit an early response(Table 4). At month 12, patients with an end-of-treatment virological response included 32 (38.5%) in the interferon-α and amantadine group and 23 (28.4%) in the interferon-α monotherapy group (P = N.S.). At month 6 after treatment, patients with a sustained virological response included 18 (21.6%) in the interferon-α and amantadine group and 17 (20.9%) in the interferon-α monotherapy group. The percentages of patients exhibiting a breakthrough during treatment or a relapse after treatment withdrawal were 10.8% and 14.4%, respectively, in the interferon-α and amantadine group, and 4.9% and 7.4%, respectively, in the interferon-α monotherapy group.
Table 3. Virological response rates in patients treated with interferon-α monotherapy or interferon-α plus amantadine
|Responders at month 6 (%)||42 |
(n = 34)
(n = 45)
|End-of-treatment responders (%)||28.4 |
(n = 23)
(n = 32)
|Sustained responders (%)||20.9 |
(n = 17)
(n = 18)
|Breakthrough (%)||4.9 |
(n = 4)
(n = 9)
|Relapsers (%)||7.4 |
(n = 6)
(n = 12)
|Drop-outs (%)||12.3 |
(n = 10)
(n = 11)
Table 4. Virological response rates in patients treated with interferon-α monotherapy or interferon-α plus amantadine after randomization according to HCV RNA status at month 2 (early virological response)
|Responders at month 6 (%)||75.7 |
(n = 25)
(n = 23)
(n = 20)
(n = 11)
(n = 18)
(n = 16)
(n = 14)
(n = 7)
|Sustained responders (%)||39.4 |
(n = 13)
(n = 12)
(n = 5)
(n = 5)
Predictors of sustained virological response
When combining all the different treatment groups, an early response to the initial course of interferon-α monotherapy appeared to be a strong predictor of sustained virological response at univariate analysis. The percentages of patients with a sustained response were 37.9% (25/66) in early responders and 10.2% (10/98) in early non-responders to interferon-α monotherapy (P < 0.0001). This finding was confirmed using multiple logistic regression analysis, which showed that an early response to interferon-α monotherapy (P = 0.0004) was a highly significant positive predictor of sustained virological response, independent of a number of variables (age, gender, baseline serum viral load and alanine transaminase levels, HCV genotype, histological grading and staging, past hepatitis B virus infection, estimated duration of infection, habitual alcohol intake and treatment with or without amantadine). Multiple regression also identified infection with a non-1 HCV genotype (P = 0.0299) and an estimated shorter time from infection (P = 0.034) as two additional independent predictors of sustained virological response, whereas treatment with or without amantadine appeared to be of no value (Table 5).
Table 5. Multiple logistic regression analysis of factors associated with a sustained virological response *
|Genotype (1 vs. non-1)||0.9498||0.4374||0.0299||0.3868||0.1641||0.9117|
|Estimated time from infection (years)||− 0.1308||0.0619||0.0348||1.1397||1.0094||1.2868|
|Early viral response (negative/positive)||− 1.5756||0.4407||0.0004||4.8334||2.0376||11.4654|
Patients achieving normal serum alanine transaminase levels at month 6 included 45 of the 83 (54.2%) in the interferon-α and amantadine group and 43 of the 81 (53%) in the interferon-α monotherapy group. At month 12 (end of treatment), patients with normal alanine transaminase included 28 (33.7%) in the interferon-α and amantadine group and 22 (27.2%) in the interferon-α monotherapy group. At month 18, there were 17 (20.5%) sustained biochemical responders in the interferon-α and amantadine group and 16 (19.8%) in the interferon-α alone group. None of these figures achieved statistical significance.
Recent studies have investigated the potential efficacy of the combination of interferon-α and amantadine as first-line treatment for newly diagnosed patients with chronic hepatitis C.11–15 As conflicting results have been reported, the clinical relevance of the addition of amantadine to interferon-α is unknown and remains under debate. In all published trials related to this issue, patients have been randomly allocated to receive interferon-α (6 MU three times weekly) or the same interferon-α schedule plus amantadine for 6 months, followed, in most studies, by a reduction of the interferon-α dose to 3 MU three times weekly for a further 6 months.11–15 In no case has allocation to treatment with amantadine taken into consideration the early individual response to therapy — a known predictor of sustained response — which may perhaps explain the contradictory results.
The present study is the first randomized controlled trial evaluating the efficacy and safety of the combination of interferon-α plus amantadine compared with interferon-α monotherapy in naive patients with chronic hepatitis C who were stratified and randomized according to the early virological response to an initial short course of interferon-α monotherapy. Patients with (i.e. with undetectable serum HCV RNA at month 2) or without (i.e. with measurable serum HCV RNA at month 2) an early response were randomly assigned to continue interferon-α monotherapy or to add to their treatment 100 mg b.d. of amantadine. This approach enables a better balance to be achieved between treatment arms, avoiding the possible effects of confounding factors, including different HCV genotypes, which are known to have a strong influence on the early and late response to treatment (as also confirmed in the present study). In addition, given the predictive value of the early response, it was possible to identify the more resistant or ‘difficult-to-treat’ patients and offer them a higher dose of interferon-α (6 MU three times weekly vs. the conventional dose of 3 MU three times weekly). However, this therapeutic strategy, which was based on the recommendations of an ad hoc committee of the Italian Association for the Study of the Liver available at the time of study planning, was disappointing, as a marginal number (10.2%) of patients without an early response responded after interferon-α augmentation, regardless of amantadine co-treatment. Moreover, in the present study, the efficacy of amantadine was assessed in a cohort of patients in whom random allocation to treatment was tailored according to the most important predictor of sustained response, the early virological response 26 (as also clearly shown in this study). Finally, now that the predictive value of a decrease in 2 logs after 12 weeks has been established as the best definition of an early response,29 it is clear that our definition (less than 500 IU/mL after 2 months) provides an even more stringent and accurate approach.
The overall result of this study is negative, as we found no statistically significant differences in the rate of sustained virological or biochemical response between naive HCV patients treated with interferon-α plus amantadine or interferon-α monotherapy. The only, almost significant, difference emerging from our data was a trend towards a higher proportion of patients showing apparent serum HCV RNA clearance at month 6 and at the end of treatment with combination therapy compared with monotherapy, although almost identical biochemical responses were observed in both groups. This suggests, however, that amantadine may have a transient mild role in improving the anti-viral efficacy of interferon-α against HCV whilst on therapy, although this effect is completely lost after discontinuation of treatment. These disappointing results confirm those of other recent controlled randomized trials of interferon-α and amantadine combination therapy in naive patients with chronic hepatitis C,11, 13–15 which also failed to show any effectiveness of the addition of amantadine to conventional treatment with interferon-α. In the study by Zeuzem et al.,11 in which naive patients were randomized to receive amantadine or placebo at entry and were treated with a higher interferon-α dosing schedule than in our study, no different virological response rates were observed at any time during or after treatment. However, in two other trials, a higher rate of on-treatment virological response was reported in patients treated with the combination of interferon-α and amantadine,13, 14 but this was not associated with an improved sustained response. Negative results have also been reported recently by Helbling et al.15 in another large, double-blind, randomized controlled trial, in which no significantly increased sustained virological response was observed with the interferon-α plus amantadine combination, although in a multivariate analysis amantadine reached a level of significance as an independent predictor of sustained response.
Our findings, as well as those mentioned above, are in contrast with those of Mangia et al.,12 who, in an open-label trial, reported an enhanced efficacy of interferon-α plus amantadine combination therapy in the initial treatment of HCV patients. These authors found a sustained virological response in 29.3% of patients treated with the combination and in 16.8% of those treated with interferon-α alone, a difference which was statistically significant. However, it is important to note that, in this trial, patients received a double interferon-α dosing schedule (6 MU three times weekly) compared with the standard dose of 3 MU three times weekly. However, in the present study, in the sub-group of 98 patients without an early response to treatment, in whom the interferon-α dose was increased to 6 MU three times weekly, we were unable to show any difference between patients treated with or without amantadine.
The most important finding of the current study is the evidence that the HCV RNA status determined after 2 months of standard interferon-α monotherapy is a strong independent predictor of sustained virological response, with an approximately four-fold greater chance of cure for those patients who have undetectable serum HCV RNA compared with those with measurable viraemia. This supports the clinical usefulness of the evaluation of the early virological response to initial standard interferon-α therapy in order to identify ‘difficult-to-treat’ patients who may require more aggressive, individualized forms of treatment. This concept will most probably apply also to the current standard of treatment based on pegylated interferon-α and ribavirin.
The combination of interferon-α plus amantadine was safe and well tolerated and no major side-effects due to amantadine administration were observed. Discontinuation of therapy for adverse events was similar in both treatment groups. We were unable to confirm the reported improvement in fatigue and other interferon-α-related side-effects in patients treated with amantadine.11, 20
In conclusion, although amantadine may result in a slight increase in the initial anti-viral efficacy of combination treatment, our findings do not support its clinical usefulness in combination with standard interferon-α in the treatment of naive patients with chronic hepatitis C.
This study was partially supported by the National Health Service and by grant no. 20902003 from the University of Rome ‘Tor Vergata’, Department of Public Health.
Other investigators of the SMIEC 1 group included O. Sorbello, G. D'Amico, D. Di Giammartino, A. Di Nicola, M. Chircu, A. Liuti and P. Stoppini.