Early addition of ribavirin to interferon in chronic hepatitis C not responsive to interferon monotherapy

Journal of Hepatology2000; 33: 463468 Printed in Denmark AN rights reserved Mmksgaard Copenhagen

Copyright 0 European Association for the Study of the Liver 2000 Journal of Hepatology ISSN 0168-8278

Early addition of ribavirin to interferon in chronic hepatitis C not responsive to interferon monotherapy Antonio

Bellobuono’,

Luca Mondazzi’, Silvana Tempini r, Federico Lina Furione2 and Gaetano IdCot

Chiodo2, Enrico Magliano2,

‘Department of Hepatology, S. Giuseppe Hospital and =Department of Microbiology, Niguarda Hospital, Milan, Italy

Background/Aim: Persistence of HCV-RNA in serum early in treatment is a strong predictor of failure of a-interferon therapy for chronic hepatitis C. Therefore, we compared the efficacy of ribavirin addition to a-interferon with a doubling of the dosage of ainterferon in case of lack of early virological response to a-interferon therapy. Methods: Sixty patients were administered interferon a2b at the dosage of 3 million units 3 times a week. After the first 4 weeks of therapy, serum HCV-RNA was evaluated. The patients with negative HCV-RNA test received the same treatment for a further 11 months, while those with detectable HCV-RNA were randomized to receive either the same dosage of a-interferon plus ribavirin (1000 mglday) or double dosage of a-interferon (6 million units tiw) for 11 months. We considered sustained response to be the maintenance of normal alanine aminotransferase and negativity at HCV-RNA test-

ing until the end of a 6-month post-treatment follow-up. Results: After the first 4 weeks of treatment, 12 (20%) patients showed virological response and 48 patients (80%) remained positive on HCV-RNA testing. Sustained response was observed in 5/12 (42%) patients with early virological response, in 10124 (42%) patients without early virological response who were administered ribavirin and a-interferon, and in only 1124 (4%) patients who were administered the double dosage of a-interferon (p=O.O06). Conclusions: This study shows the efficacy of the addition of ribavirin to a-interferon and the lack of efficacy of doubling the dosage of a-interferon in patients without clearance of hepatitis C virus early on in treatment. Key words: Antiviral therapy; Hepatitis C virus; Viral hepatitis.

See Editorial, pages 482-484 and Articles, pages 448455

A

(ol-IFN) provides effective therapy in some cases of chronic hepatitis C, but many patients either do not respond or show reactivation of the disease after the end of treatment (l-3). Furthermore, a-IFN is costly and not free of side effects (4). Therefore, a thorough search for predictive factors of long-term response to therapy has been made, in order to avoid potentially severe side effects in patients with a negligible chance of cure and to make the best use of this expensive drug. LPHA-INTERFERON

Received 22 July 1999; revised 10 February: accepted 23 February 2000

Correspondence: Gaetano Idto, Department of Hepatology, S. Giuseppe Hospital, Via S. Vittore no. 12,20123 Milan, Italy. Tel: 2 85994497. Fax: 2 85994245. e-mail: [email protected]

and 456462

Before the start of treatment, the best predictors of sustained response to (x-IFN are: absence of cirrhosis, low serum levels of HCV-RNA and non-l HCV genotype (5-7). With the exception of the case of decompensated cirrhosis, though, the pre-treatment prognostic factors so far identified cannot be used to decide whether or not to start treatment, and all patients without contraindications to (r-IFN are offered this therapy. In contrast, persistence of HCV-RNA in serum early on in therapy has been demonstrated to be a strong predictor of treatment failure and this evidence represents a sound basis for early change of treatment in early non-responders (8-l 3). One possible option is the use of ribavirin in combination with (r-IFN. Ribavirin is a synthetic, purine nucleoside analogue with a broad spectrum of activity 463

A. Bellobuono

et al.

against RNA and DNA viruses (14). In interferon-naive chronic hepatitis C, the combination of ribavirin and a-IFN proved more effective than or-IFN alone in inducing sustained clearance of HCV (15,16). The combination of ribavirin and a-IFN proved superior to retreatment with a-IFN monotherapy also in patients with previous relapse to cr-IFN alone (17-20). Another possible “rescue” treatment for early nonresponders is increasing the dosage of LX-IFN. Although previous studies on this change of treatment gave unsatisfactory results (21-24), in no case was the increased dosage of cx-IFN administered as early as at the first month of treatment on the basis of lack of early virological response to therapy. Therefore, in a prospective trial we compared the efficacy of adding ribavirin to the initial dose of a-IFN, with doubling the dosage of (r-IFN in patients with persistence of HCV-RNA in serum at the end of the 4th week of therapy.

Materials and Methods Patients Sixty consecutive, interferon-naive, HCV-RNA positive patients affected by biopsy-proven chronic hepatitis C were recruited into this study between June and November 1996. Patients’ ages ranged between 26 and 59. median 44. The male/female ratio was approximately 2:l. All patients had increased serum alanine aminotransferase (ALT) levels (more than 1.5 times the upper limit of the normal range) on at least three determinations during the 6 months before therapy. They were all positive for anti-HCV antibodies in a second-generation enzyme-linked immunosorbent assay (ELISA) test (HCV-ELISA, Ortho Diagnostic Systems, Raritan, NJ, USA), and they had histological confirmation of chronic hepatitis obtained no more than 1 year prior to the start of the study. The histological samples were evaluated according to standard criteria (25). Chronic hepatitis activity (grade) was subdivided into the following categories: a) minimal chronic hepatitis (score of grading: l--3), b) mild chronic hepatitis (score: 48) c) moderate chronic hepatitis (score: 9912) and d) severe chronic hepatitis (score: 13.-18). Chronic hepatitis stage, determined by the extent of fibrosis, architectural disturbance and development of cirrhosis, was subdivided into four categories: a) no fibrosis (score 0). b) mild to moderate fibrosis (score l-3), c) severe fibrosis with marked bridging (score 4-5) and d) probable or definite cirrhosis (score 6). In all cases we could rule out by means of the clinical history and appropriate tests: hepatitis A virus, hepatitis B virus, cytomegalovirus, Epstein-Barr virus infection, alcohol abuse (consumption greater than 40 g of alcohol per day), autoimmune hepatitis. primary biliary cirrhosis, sclerosing cholangitis, hemochromatosis, Wilson’s disease, drug-induced liver disease and alpha-I-antitrypsin deficiency. Exclusion criteria for a-IFN therapy were as follows: decompensated cirrhosis, previous treatment with a-IFN, steroids or immunomodulating agents, HIV positivity, drug addiction. psychosis, malignancy or other serious illnesses. Treatment und followup This study protocol conformed to the ethical guidelines of the 1975 Declaration of Helsinki as reflected in a priori approval by the human research committee of our institution, and informed consent in writing was obtained from each patient before starting treatment. The patients were administered 3 million units (MU) of recombinant a2b IFN 3 times a week for 4 weeks as initial treatment. After the first

464

4 weeks of therapy, serum HCV-RNA and transaminase levels were evaluated. The patients with negative serum HCV-RNA test (“inhouse” nested reverse transcription polymerase chain reaction) received the same treatment for a further 11 months. Independently of transaminase levels, the patients with detectable serum HCV-RNA after the first 4 weeks of n-IFN therapy were randomized to receive either the same dosage of n-IFN plus ribavirin (1000 mgiday) or double dosage of a-IFN (6 MU tiw) for 11 months, In these patients. the time lag between HCVRNA testing and the randomization to either “rescue” treatment ranged between 8 and 13 days. Ribavirin was provided for compassionate use (Schering Plough S.p.A., Milan, Italy) and informed consent was obtained from each patient before the start of treatment. Appropriate precautions were taken with the administration of ribavirin, including prevention of pregnancy and monitoring of hemolytic anemia. Biochemical parameters were monitored monthly during the treatment and for 6 months after the discontinuation of a-IFN. During the treatment, HCV-RNA testing was repeated every 3 months in the patients with early clearance of the virus from serum and every month in the patients without this response. During the follow-up, serum HCV-RNA testing was repeated every 3 months in all patients. All patients completed the follow-up. We defined the responses to treatment as follows: early virological response (EVR): loss of serum HCV-RNA after the initial month of therapy; end-therapy response (ETR): steady normalization of transaminase levels and negativity at serum HCV-RNA testing until the end of treatment: sustained response (SR): normal transaminase values and negativity at serum HCV-RNA testing maintained for at least 6 months after the end of therapy. HCV- RNA detection, yuant~ficuiion und !,ping Serum HCV-RNA was detected by ‘?n-house” nested reverse transcription polymerase chain reaction (RTPCR) with primers from the 5’ untranslated region of the HCV genome. The test had a detection limit of less than 100 copies/ml (I 3). Serum HCV-RNA was quantified by using a second-generation branched DNA (bDNA) assay (26) (Q uantiplex. Chiron Corporation, Emeryville, CA, USA) performed on sera collected before the start of treatment and at the end of the 4th week of therapy, stored frozen at -80°C. HCV genotyping was performed according to the method described by Okamoto et al. (2728) which uses PCR amplification of core gene sequences with universal and five type-specific primers to generate DNA fragments with a size specific for five HCV types which are identified by Roman numbers (I to V). HCV genotypes I. II. 111. IV and V according to Okamoto correspond to genotypes 1a. 1b, Za. 2b and 3a, respectively, as classified by Simmonds (29.30). The type 2a specific primer was modified with respect to the original method (.5’-CCC CC4 TGA AGG GCG AGA AC-3’). because of small nucleotide differences found between type 3a Italian and Japanese isolates. Genotyping fidelity was confirmed by cloning and sequencing of core region sequences from reprcscntative clones classified according to the modified procedure (3 1). Statistical analysis Statistical analy-sis was performed by using the unpaired Student’s ttest, the x2 test or Fisher’s exact test as appropriate. ~~0.05 was considered to be significant. Results are presented as mean+SD.

Results After the first 4 weeks of treatment, 12 (20%) patients showed EVR while the other 48 patients (80%) remained positive at serum HCV-RNA testing. Transaminase levels were normal in all patients with EVR and in 18 out of the 48 patients (37.5%) without EVR. Table 1 shows the main baseline features of the patients

Early addition of ribavirinto interferon TABLE

1

Baseline features

of the patients

Age (mean 2 SD) Sex (M/F) Grade of hepatitis Minimal Mild Moderate Severe Stage of hepatitis No fibrosis Mild to moderate fibrosis Marked bridging fibrosis Probable or definite cirrhosis HCV-genotype 1 Serum HCV-RNA titer* Mean? SD > 1 X lo6 copies/ml

With EVR 12 (20%)

Without EVR p 48 (80%)

41*11 1.6

44?13 1.4

NS NS NS

2 (16.7%) 4 (33.3%) 6 (50%) 0

9 15 22 2

(18.7%) (31.2%) (45.8%) (4.2%)

4 7 1 0 3

12 34 2 0 28

(25%) (70.8%) (4.2%)

(33.3%) (58.3%) (8.3%) (25%)

0.9kO.6 1 (8.3%)

NS

(58.3%)

3.320.8 42 (85%)

0.04
* MEq/ml.

TABLE

2

Main features of treatment

of the patients

without

Age (mean?SD) Sex (M/F) Grade of hepatitis Minimal Mild Moderate Severe Stage of hepatitis No fibrosis Mild to moderate fibrosis Marked bridging fibrosis Probable or definite cirrhosis HCV-genotype 1 Serum HCV-RNA titer* Mean&SD * MEq/ml; treatment.

test performed

EVR in relation

to the schedule

a-IFN+ ribavirin 24 (50%)

a-IFN: 6 MU tiw 24 (50%)

P

45-1-11 1.3

44?12 1.5

NS NS NS

3 (12.5%) 8 (33.3%) 13 (54.2%) 0 7 15 2 0 13

(29.2%) (62.5%) (8.3%) (54.2%)

0.9kO.7

6 7 9 2

(25%) (29.2%) (37.5%) (8.3%)

5 (20.8%) 19 (79.2%) 0 0 15 (62.5%) 0.820.6

at the time of randomization

NS

NS NS to either

with EVR in comparison to the patients without EVR. The pre-treatment viral load was significantly lower in the patients with EVR and they also showed a lower prevalence of HCV genotype 1 in comparison to the patients without EVR. In contrast, no difference was observed between the two groups as far as age, sex and grade and stage of hepatitis were concerned. The patients with EVR were kept on the initial treatment schedule. In this group, we observed ETR in 8/12 (67%) patients and SR in 912 (42%) patients (Fig. 1). The patients without EVR were randomized

ceive either the same dosage of (r-IFN in association with ribavirin (24 patients) or the double dosage of CZIFN (24 patients). Table 2 shows the main features of these patients, including HCV-RNA load as assessed at the time of randomization to either treatment. The two groups were comparable as regards age, sex, grade and stage of hepatitis, genotype distribution and HCVRNA levels. In the group of patients randomized to receive interferon and ribavirin, ETR was achieved in 15/24 (62%) cases and SR in lo/24 (42O/o)cases. In the group of patients who received the double dosage of a-IFN, ETR was achieved in 4/24 (17%) cases and SR in only l/24 (4%) cases. Fig. 1 reassumes the results of treatment in all groups of patients. In the patients without EVR, the addition of ribavirin was significantly more effective than doubling the dosage of aIFN in inducing ETR (p=O.O03) and SR (p=O.O06). In the patients without EVR who were administered ribavirin and a-IFN therapy, ETR and SR were very similar to those observed in the patients with early clearance of the virus from serum. When transaminase levels were considered, we found that addition of ribavirin induced SR in 5/10 (50%) patients with normal values and in 504 (35.7%) patients with persistence of abnormal transaminase values at the end of the 4th week of a-IFN therapy (p: NS). Table 3 shows the rates of HCV-RNA clearance and of ALT normalization in the three groups of treatment. In the 10 patients without EVR who were administered ribavirin in addition to a-IFN and showed SR, serum HCV-RNA testing became negative and ALT levels were normal by the end of the third month of treatment (second month of combination therapy). When we considered separately the patients infected with genotype 1 or genotype non-l who were adminis-

to re-

50%

0% IFN 3 MU

Ftibavirin addition

Increased IFN dosage

Fig. 1. Response to treatment. ETR induced by ribavirin addition vs increased a-IFN dosage: p=O.O03; SR induced by ribavirin addition vs increased a-IFN dosage: p=O.O06.

465

A. Bellobuono et ul. TABLE

3

ALT normality

and HCV-RNA

negativity

during

treatment

and follow-up Month

EVR a-IFN 3 MU tiw (12 patients) Normal ALT Negative HCV-RNA Non-EVR Ribavirin addition (24 patients) Normal ALT Negative HCV-RNA Non-EVR a-IFN 6 MU tiw (24 patients) Normal ALT Negative HCV-RNA

1

2

3

6

9

12 12

12 12

11 11

10 10

10 0

13 6

15 12

8 0

10 1

10 8

tered ribavirin in addition to a-IFN, we found the responses to treatment reported in Fig. 2. In the group of patients who underwent combination treatment we also evaluated the correlations between SR and the following variables: 1) baseline

50%

0% Genotype

non-l

Genotype

1

Fig. 2. Response to a-IFN+rihuvirin in rehtion to HCV genotype. SR in genotype non-l vs genotype I: p=O.O.55.

50%

0% U= 2 MEq/ml

+3

+6

10 9

9 8

6 6

5 5

16 15

16 15

16 15

13 13

10 10

8 7

7 4

7 4

4 2

2 1

serum HCV-RNA titer; 2) serum HCV-RNA titer at the time of ribavirin addition; 3) variation in serum HCV-RNA concentration between baseline and the time of ribavirin addition. Baseline serum HCV-RNA titre proved significantly lower in the patients with SR than in those without SR: 1.97k1.57 rs 3.8621.92 MEq/ml, respectively (~~0.018). Also serum HCVRNA titer at the start of combination treatment was lower in the patients with SR than in those without SR: 0.4250.25 vs 1.4850.79 MEq/ml, respectively (j~O.001). In contrast, the variation in HCV-RNA titer between baseline and the time of ribavirin addition did not differ between these two groups of patients: 1.55k1.74 vs 2.4922.16 (p=NS). When we considered separately the patients infected with baseline HCV-RNA concentrations 22 or >2 MEq/ml who were administered the combination therapy, we found the responses to treatment reported in Fig. 3. The treatment could be completed in all cases. The change of therapy with addition of ribavirin to (r-IFN was associated with some hemolysis in all patients, but only two cases required a reduction in the dosage of ribavirin (to 800 mg and to 600 mg/day, respectively) because of anemia. In the group of ribavirin and a-IFN a fall in hemoglobin greater than 2 g/d1 (15/24: 62.5%) and pruritus (8/24: 33.3%) were significantly more frequent (j~O.001 andp=0.03, respectively) than in the patients who were administered the increased dosage of a-IFN (6124: 25% and 2/24: 8.4% respectively). In all cases the side effects were reversible after cessation of treatment or reduction in the dosage of ribavirin.

>2MEq/ml

Fig. 3. Response to a-IFN+ribavirin in relution to baseline viral loud. SR in baseline HCV-RNA concentration 52 vs >2 MEqlml: p=O.134.

466

12

Discussion In the group of patients similar to those reported

with EVR, the SR rate was in comparable studies (9, IO).

Early addition of ribavirin to interferon

In the patients without EVR, regardless of ALT behavior, the change of treatment with the addition of ribavirin to a-IFN induced SR in 42% of cases. This SR rate was equal to that observed in the patients with EVR. Therefore, the addition of ribavirin to a-IFN early on in treatment in patients without EVR seemed to restore the chance of SR, making it similar to that of patients with EVR. The SR rate we observed in the cases treated with ribavirin and a-IFN was also very similar to those reported in two multicenter trials in which the same doses of a-IFN and ribavirin were used for a very similar length of time (15,16). When we considered the patients infected with HCV genotype 1 or with the other genotypes separately, we found that the SR rate to the combination of (r-IFN and ribavirin was quite different in the two groups (23% vs 64%, respectively), although the difference did not reach statistical significance, probably because of the small size of the sample. When we considered the patients infected with baseline HCV-RNA concentrations 12 or >2 MEq/ml separately, we found that the SR rate to the combination of ol-IFN and ribavirin was quite different in the two groups (67% vs 27%, respectively). Again, though, the difference did not reach statistical significance, probably because of the small size of the sample. Indeed, in the two multicenter trials mentioned above, the reported SR rates in relation to genotype or to HCV-RNA load were very similar to those observed in the present study, and the differences between these SR rates were statistically relevant (15,16). The similarity of results between the present study, in which ribavirin was added to a-IFN early on in treatment in case of lack of EVR, and the two multicenter studies mentioned above, in which the combination of a-IFN and ribavirin was used from the start of therapy, suggests that the two treatment strategies might have similar efficacy. In case of infection with genotype 2 or 3, independent of the baseline HCV-RNA concentration, a 12month course of a-IFN monotherapy at the dosage of 3 MU tiw induced an SR in 3 1% of patients (15,16). In the patients infected with these HCV genotypes, the combination of a-IFN and ribavirin from the start of treatment, administered for 6 months in accordance with the suggestion in the Consensus Statement of the EASL International Consensus Conference on Hepatitis C held in Paris in 1999 (32), induces SR in 4869% of cases (15,16,33,34). Although the difference in SR rate between the two groups of treatment is relevant, there is a significant chance of cure even without the use of ribavirin. The results of our study suggest that in patients infected with genotype 2 or 3, a course

of cc-IFN monotherapy could be tried, adding ribavirin only in case of lack of early clearance of the virus from serum. In particular, this strategy would be advisable in patients with borderline hemoglobin concentration or not willing to take ribavirin, if not absolutely necessary, because of its potential toxicity (35,36). However, further prospective studies in larger numbers of patients are required before this clinical recommendation is applied to routine clinical use. In case of infection with HCV genotype 1, in contrast, a 12-month course of a-IFN monotherapy at the dosage of 3 MU tiw induces SR in less than 10% of patients (15,16). On the basis of this unsatisfactory SR rate, in the patients infected with genotype 1 the combination of a-IFN and ribavirin should be used from the start of treatment. In case of lack of EVR, the combination of a-IFN and ribavirin was significantly more effective than doubling the dosage of a-IFN. Previous studies evaluated the efficacy of increased ol-IFN doses in non-responders and gave unsatisfactory results. In one of these studies, the increased dose regime was administered on the basis of lack of virological response, but this was evaluated at the end of the third month of therapy (24). In the other studies, the increased dose regime was administered on the basis of lack of normalization of ALT levels, and generally no sooner than 8-12 weeks after the start of treatment (21-23). In the present study, doubling the dosage of ol-IFN in case of persistence of HCV-RNA in serum 4 weeks after the start of treatment, independently of ALT levels, induced sustained response in only 1 out of 24 cases (4.7%). This very poor response rate confirms the negative prognostic value of viral persistence at the end of the 4th week of therapy and demonstrates the lack of efficacy of increasing the dosage of a-IFN, even though the dosage is changed as early as at the end of the first month of therapy, even in patients with normal ALT levels at that time. In conclusion, the present study: a) shows the efficacy of the addition of ribavirin to a-IFN in case of lack of virus clearance after 4 weeks of therapy; b) casts doubt on the need for the use of ribavirin from the start of therapy in patients infected with HCV genotype 2 or 3; c) shows the lack of efficacy of doubling the dosage of a-IFN in patients without early clearance of the virus. However, these initial findings need to be verified in larger prospective studies.

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