Pilot dose-finding trial on interferon alpha in combination with ribavirin for the treatment of chronic hepatitis C in patients not responding to interferon alone

M. Puoti G.P. Cadeol v. Putzolu M.A. Forleo M.C. Barn? G. Cristinil S. Rossi A. Spinetti S. Zaltron 6. Zanini E. Quiros-Roldan G. Paraninfol F, Gargiulo2 G. Carosi

Background, Effectiveness of combination therapy with standard inalpha doses and ribavirin is far from being demonstrated in patients with hepatitis C non responders to interferon alpha monotherapy Recent kinetic studies revealed that these doses may be suboptima/. Aims. To find the criteria for optimisation of the interferon dose, to be used in combination with ribavirin in patients with hepatitis C non responders to interferon alpha monotherapy Patients. Sixty-three patients enrolled in a pilot controlled trial were treated for 6 months with ribavirin [ 1000-1200 mg daily] and were randomised to concurrently receive interferon alpha 2b for 6 months at: 3 Million Units thrice weekly [group A I21 patients]], 5 MU thrice weekly [group B [2 I patients]] and 5 million units daily [group C 121 patients]]. Results. A sustained virological response was observed in: I patient from group A (5%], 2 patients from group 6 [S%] and 8 patients from group C [38%; p--O.02 vs group A; p=O. 03 vs group B]. Side-effects were not significantly different between the 3 groups. Multivariate analysis showed that infection by hepatitis C virus genotypes 2 or 3 and interferon alpha dosage of 5 ,million units daily were independent predictors of sustained response. Conclusions. These results suggest that higher interferon doses administered daily in combination with ribavirin could be more effective in those patients with hepatitis C who had not responded to interferon alone. terferon

Digest

Liver

Key uuords:

Dis 2001;33:163-72 Hepatitis

C Virus;

Hepatitis

C Virus

kinetics;

Hepatitis

C Virus-RNA

From Clinic for Infectious and Tropical Diseases, 2 Department of Microbiology, I I Division of infectious Diseases, Erescia Hospital, Brescia, Italy. Address for correspondence Dr. M, Puoti, Ciinica di Malattie In fettive e Tropicali, Piazrale Spedali Civili I, 25123 Erescia, Italy. Fax: -f-39-D30-30306’9. E-mail: [email protected] Submitted October I I, 2000. Accepted December 22, 2000.

Hepatitis C Virus (HCV) infects nearly 170 million people worldwide and causes a chronic disease that remains asymptomatic and mild for a prolonged period, but may eventually lead to cirrhosis and/or hepatocellular carcinoma in a subgroup of patients estimated to account for 20-30% of chronically infected individuals ‘. For many years, interferon monotherapy conducted with doses of 3 Million Units (MU) administered three times a week for 6-12 months has been the standard treatment for chronic hepatitis C. It has been associated with 1 of 3 treatment outcomes: sustained virological response, end-of-treatment response with subsequent relapse, or no response *. Most of the patients treated with interferon monotherapy showed no response ‘. The advent of interferon alfa-ribavirin combination for either initial therapy or re-treatment of previous relapse has significantly increased the number of patients who achieve a sustained response 3-5.In patients who have failed to respond to interferon alpha monotherapy, there are no clear

data to indicate that re-treatment will be beneficial ‘m2. In fact, there is firm evidence that re-treatment of nonresponders with standard regimens of interferon alpha monotherapy is of no clinical value 1-27-9.On the other hand, the sustained response rates achieved following combination therapy with standard interferon alpha doses and ribavirin are too low to consider this treatment as cost effective * i”-lg. Recent studies on HCV viral kinetics during interferon treatment have shown that: the reduction in serum levels during the first 24-48 hours is dose-dependent, being more than 50% at doses of interferon between only5andlOMU 2o, there is a rebound of viral replication during the day free from interferon when it is administered intermittently 21 and early viral clearance predicts interferon response 2223. These observations taken together would suggest that actual dosing regimens are inadequate and that higher doses of interferon eventually administered daily result in an increased rate of sustained response. The results of two comparative controlled trials on interferon alpha monotherapy in naive patients support the greater efficacy of higher doses administered three times per week and of daily interferon administration, respectively 2425, other comparative trials failed to show an increased efficacy of higher doses of interferon monotherapy 2627. No controlled interferon dose-finding trials have been performed with interferon and ribavirin in combination and, therefore, it still needs to be assessed whether higher efficacy of combination treatment can be achieved either by increasing the single dose of interferon alpha or by increasing the single dose and the frequency of interferon administration. In addition, the tolerability of these more intensive interferon schedules in combination with ribavirin is still unknown. We report here the results of a randomized pilot trial of re-treatment with interferon alpha in combination with ribavirin in 63 patients not responding to previous interferon monotherapy. The aim of the study was to compare the efficacy and safety of standard interferon doses of 3 MU three times a week with those achieved by either increasing the single doses from 3 to 5 MU or increasing both the single doses and their frequency to 5 MU daily.

Selection of patients Entry criteria were as follows: age between 18 and 60 years, diagnosis of hepatitis C, confirmed histologically no more than 24 months earlier, without confounding aetiologies such as hepatitis B, autoimmune disorders, alcohol abuse, metabolic disorders and human

immunodeficiency virus positivity; one or more previous treatments with 3 MU of interferon alpha given three times weekly for six months or with 6 MU given three times weekly for at least 3 months; documented evidence of non-response in previous cycle(s) as defined by absence of both return to normal of alanine aminotransferase (ALT) and HCV-RNA negativisation during previous treatment; no major side-effects during the previous interferon cycle(s); ALT levels >1.5 times the upper normal limit and a positive HCV-RNA test at the time of inclusion in the study; no evidence of decompensated liver disease or significant cytopenia (leukocyte count ~3000 mm3; platelet count
scale for grading acute and subacute drug toxicities; therapy was discontinued after life threatening events (grade IV). In cases of severe adverse events (grade III toxicity) other than anaemia and flu-like symptoms, the single dose of interferon alpha-2b was reduced to 1.5 MU in group A and 3 MU in groups B and C, and the dose of ribavirin was reduced to 600 mg per day. In those patients in whom haemoglobin concentrations fell below 10 g/dl, only the dose of ribavirin was reduced to 600 mg per day and was totally withdrawn if the concentration fell below 8.5 g/dl. Only the dose of interferon was halved for grade III flu-like symptoms, for grade III leukocytopenia (i.e., less than 1000 white blood cells per mm3) and grade III thrombocytopenia (i.e., less than 50,000 platelets per mm3). The patients were evaluated as outpatients at weeks 1, 2, 4 and then every four weeks during treatment and for a further 24 weeks after stopping treatment. Biochemical and haematological tests were performed on each occasion. Serum HCV-RNA levels were determined in samples collected at baseline and at weeks 4, 12, 24, 36 and 48 and frozen at -70°C until assayed. Serum HCV-RNA was measured by a quantitative reverse-transcription polymerase chain reaction assay with a sensitivity of 1000 copies/ml (Amplicor HCV Monitor 2.0, Roche Diagnostics, Basel, Switzerland) at baseline and at 4 weeks and with a qualitative reverse transcription polymerase chain reaction assay with a sensitivity of 100 copies/ml (Amplicor HCV 2.0) at 12, 24, 36 and 48 weeks. When baseline HCVRNA levels were higher than 1 million copies per ml both samples collected at baseline and after 4 weeks were diluted 1: 10 with human plasma according to the manufacturer’s instructions. HCV genotyping was performed by a Line Probe assay (InnoLIPA, Innogenetits, Ghent, Belgium). Histology Liver biopsies performed no more than 2 years before entry into the study were re-evaluated by a single pathologist who was unaware of treatment results. The degree of hepatic inflammation and fibrosis was graded using the Knodell Histologic Activity Index **. The inflammation grading score was obtained by combining the scores of the first three components of the index: portal, periportal and lobular inflammation. Main outcome measures The primary end point was a sustained response, defined as the absence of serum HCV-RNA in both samples collected 12 and 24 weeks after the completion of the treatment and the presence of ALT levels below the normal upper limit in all samples collected monthly after stopping treatment. The secondary end point was an end-of-treatment response, defined as the absence of

HCV-RNA and return to normal of ALT in the sample collected at the end of treatment. Statistical analysis We planned to assess the results of treatment with an intention-to-treat analysis. The sample size of the study was calculated by assuming that 9% of patients randomised to the standard interferon alpha-2b schedule and a minimum of 50% randomised to the other two schedules would have a sustained virological response. Based on these assumptions, it was calculated that 19 patients should be enrolled in each group for a significant difference to be detected with a type I error alpha of 0.05 and a power of 0.8. Values of age, weight, estimated duration of infection, serum ALT: histological inflammation score and cumulative interferon dose assumed before entry into the study are expressed as median with interquartile ranges. Differences between median values of samples of these quantitative variables were compared using the Kruskal Wallis analysis of variance (ANOVA), the Wilcoxon rank sum test and Mann Whitney test when indicated. Differences in demographic criteria, cirrhosis, the number of patients with high viraemia and infected by specific HCV genotype and the number of patients that developed sustained and end of treatment response were assessed using the x2 test (Mantel-Haenszel), and Fisher exact test when appropriate. The relationship between pre-treatment variables and sustained response was also examined by stepwise logistic-regression analysis. Variables that were associated with a sustained response at ~~0.20 on univariate analysis were entered in a stepwise logistic regression to assess their association when taking into account possible confounders. All p values are two-tailed; a p value of less than 0.05 was considered as significant. EPI INFO software v. 6.0 (CDC, Atlanta, GA, USA) and EGRET software (Statistics and Epidemiology Research, Seattle, WA, USA) were used for the statistical analysis.

Characteristics of the patients A total of 63 patients were included in the study and 21 were randomized to each treatment group. The three treatment groups were well matched (Table I). Most patients had previously received a cumulative interferon dose of more than 432 MU, i.e., 6 MU three times a week for 24 weeks. The proportion of patients with high HCV-RNA levels, with unfavourable genotypes and with advanced fibrosis were similar in all three groups. Of the patients enrolled in the study, 75% were infected by HCV genotype 1 or 4 reflecting epidemi-

PM-mm-p

-

Table I. Baseline

characteristics

of the patients,

-.mww-v-

--

Characteristic _m_ll_____JY-IY_____~~~~*-~~~~

-~-_^1-~~~-___11-~~~~-~.~~--.~~~~~-"--~~~~~--~.~~~." Group A Group G -----w"--w J___-I_1XY_~_~--~~~~-~~-",-~----~",",..~"~~-,-~.~~,-~~.~

All

Group

C

Treatments 3 MIU 3 xweek l-I.2 g

5 MIL. 3 x week

5 MILI 1 x day

I-1.2

l-I.2

63

21

21

21

52 [82%1

18 f86%1

17 (81%1

17 [61%1

Age - yr Median [Interquartile range1

43 (32-541

43 (32.5-541

43 (34.5-55.51

42 (31~51.51

Weight - kg median [interquartile range1

70 (63-801

&i23

70 [63-5-801

72 (64-77.51

12 (6201

12 [6-211

IO [9-l 6.51

19.5~250.5I

Interferon

alpha 2b sc:

Ribavirin Number

OS:

dose frequency

daily dose

cn.1

Male sex - n. [%I

Estimated duration of infection [interquartile range1 Source

of infection

- yr median

drug use

Unknown Genotype-n. ;

(IO%1

8 (13%1

3 [14%1

3 (14%)

2

18 (29%)

5 (24%)

5 [24%1

8 (38%)

37 [58%1

13 I62%1

13 [62%1

11 [52%1

11 [67%1 43 (17%1

154 [19%1 (71%1

15 5 (24%) [71%1

132 [IO%1 (62%1

4 (6%) 5 (8%)

1 (5%) 1 (5%)

1 [5%1 1 [5%1

3 (14%) 3 (14%)

[%I

3 4 Serum

g

- n. [%I

Transfusion Injection

g

HCV-RNA

> 1 x10" Many

copies/mln.

septa"

CirrhosisB

(%I

- n. (%I - n. [%I

Grading median [interquartile range1 Serum alanine aminotrasferase [interquartile range1 Previous

median

44 [70%1

15 [71%1

14 [70%1

15 (71%)

27 (43%)

IO [48%1

IO [48%1

7 (33%1

6 (10%)

3 [14%1

1 [5%1

6.5

6

[5-71

(3-71

6.5 [5-91[5-71

107 (69-1631

102 180-1741

107 (83-1511

109 [65-1561

42 (67%)

12 [57%1

14 [67%1

16 (76%)

573

468 (333-8501

2

[IO%1 6.5

therapy

Single course Cumuiative median [interquartile

n. [%I

Interferon

dose 549

range1

[324-882)

~~. ~-~~~-~~~~ There were no significant differences between the two groups. to Knodell 21 5 Fibrosis score 4 according to Knodeil. -~“_-____11_1_1---~-~~--~--~--~-,~.--~,~-----~~~~~~.~~~~~..~~--~“~,~~

(346-8681

549 ~24610081

a_11~sg____*y_/l_l~m-~~~~-,~~~--,..~~-~~-~~-,,~,,-,~‘~"",~ Because

ofrounding-off;

ology of HCV genotypes among non responders to interferon monotherapy in Northern Italy 7. All patients started combination treatment after an interferon-free interval of at least 6 months. Four patients stopped treatment before the sixth month (1 group A, 1 group B, 2 group C). All patients were followed up for 6 months after stopping therapy.

percentages

may not total

,-., 100. “Fibrosis

score

3 or 4 according I-,. Ilil--illlf(l

Vivologic response Virologic response rates observed during the study in the three treatment groups are shown in Figure 1. All patients treated with 5 MU of interferon alpha daily showed a decrease of at least one log of HCV-RNA levels after four weeks of treatment; the proportion being significantly higher than that found in the con-

100%

L7 Group: A IFNa 3 MW

90% 70%

80%

60% 50% 40%

40%

30% 20%

30%

10%

20%

0% HCV-RNA HCV-RNA< HCV-RNA< HCV-RNAc HCV-RNA< ,M) copiesat 4nn*n&n” “+ rnn n,.d”c a, decrease of 1 1000 24 weeks 48 week lag at 4 copWml at 4 12weeks weeks weeks

fig. 1. Percentages of treated patients achieving virological end poinr during the study, stratified according to treatment groups and inte feron a . 2b dosage. il p~O.01 vs group A, Mantel-Haenszel x2 tes “p~O.01 vs group 3, Mantel-Haenszel x2 test; Q p=O.O2 vs group by Fisher exact test; h p=O.O3 vs group B Mantel-Haenszel x2 tes IFNcL: interferon alpha 2b; MU million units; Tw: thrice weekly.

trol group and in the group of patients treated with 5 MU of interferon alpha three times per week [S out of 21 (38%) in each group; 95% Confidence Interval (CI) 17-59%; ~~0.01 x2 test]. The proportion of patients with HCV-RNA levels below 1000 copies per ml did not differ in the three groups The proportion of patients with serum HCV-RNA levels below 100 copies per ml at the 12th week of treatment was significantly higher among those treated with 5 MU of interferon alpha per day (~~0.01 x2 test). In accordance with conventional end points defined in the National Institutes of Health Consensus Development Conference 23, we found that 11 out of 21 patients (52%; 95% CI 31-73%) treated with standard doses of interferon alpha 2b showed a virologic response at the end of treatment. This was identical to findings in the control group for patients treated with 5 MU three times per week (1 l/21; 52%; 95% CI 3173%) and slightly, but not significantly, higher in patients treated with 5 MU daily (15/21; 71%; 95% CI 52-90%). One out of the 21 patients (5%; 95% CI O10%) in the control group treated with 3 MU three times per week showed a virological response at the end of the follow-up (sustained response); this rate was lower than, but not significantly different from the rate found in patients treated with 5 MU three times per week (2/21, 10%; 95% CI 3-15%). Sustained response occurred in S/21 patients treated with 5 MU per day (38%; 95% CI 17-59%); it was significantly higher than in the control group (p=O.O2 by two tailed Fisher exact test) and higher than in pa-

10% CVX

“I”

Normal ALT at 4 weeks

Normal ALT at 12 weeks

Normal ALT at 24 weeks

Normal ALT at 48 weeks -L___

Fig. 2. Percentages of treated patients in whom ALT returned to normal during the study, stratified according to treatment groups and interferon a 2b dosage. @ p=O.O2 vs group A by Fisher exact test; n p=O.O3 vs. group B Mantel-Haenszel x2 test. IFNa: Interferon alpha 2b; MU: million units; TW: thrice weekly.

tients treated with 5 MU three times per week, just less than the level of statistical significance (p=O.O3 Mantel-Haenszel x2 test). Biochemical response Biochemical response rates observed during the study are set out in Figure 2. No difference in the proportion of biochemical response rates was observed between the three treatment groups during or at the end of the treatment. Biochemical response at the end of treatment and at the end of follow-up was observed only in patients with a virologic response at the same time points. Variables associated with sustained virologic response A comparison of baseline characteristics between patients with and without a sustained response is shown in Table II. Univariate analysis showed that the treatment schedule of 5 MU of interferon alpha 2b per day was the strongest predictor of response. Age was significantly lower and HCV genotypes 2 and 3 were significantly more frequent in patients with a sustained response. A sustained response was unrelated to sex, body weight, estimated duration of infection, source of infection, HCV-RNA levels, stage of fibrosis, grade of necroinflammmatory activity, ALT levels, number of previous cycles of interferon or cumulative dose of interferon administered prior to randomisation. Stepwise logistic regression analysis revealed that greater efficacy was associated with daily administra-

---

Table II. Characteristics of the study population stratified according to nccurrence of sustained response. Characteristic -_l___m.--_l---~=s‘-Number fn.1

Sustained response Yf!S No -m-v11 52

Male sex - n. [%I

8 [73%1

44 Em%1

Age - yr median (Interquartile range1

32’ [30-491

44.5 (33-551

Weight - kg median [interquartile range1

16&l

(637-0803

Likelihood

Estimated duration of infection - yrs median [interquartile range1

11 (6-161

12 B-21 1

Source of infection - n. (%I Transfusion Injection drug use Unknown

1 El%1 5 [45%1 5 (45%1

7 (I 3%1 13 f25%1 32 [61%1

Genotype - n. (%I 1 or4 2 or 3

5 f45%1§ 6 [55%1

43 (83%1 9 [17%1

Serum HCV-RNA >I x 1Oa copies/ml n. Many septa - n. [%I Grading - median iinterquartile range1

7 [64%1 2 f’l8%1 6 (5-71

37 f71%1 25 [48%1

Serum alanine aminotrasferase median (interquartile range1

109 WI-291 I

104 @O-l 631

f5S7l

Previous therapy Single course n. [%I Cumulative interferon dose median [interquartile range1

9 [82%1

33 [63%1

390 (234-7381

561 (324-9271

Treatment 5 MU per day n. (%I 5 MU per single dose n. [%I

8 [73%1# 13 [25%1

10 [91%1 32 [61%1

--M--____p-

p-O.05 p~O.01

~-~~

Mann Whitney test; § 0.02 two tailed Fisher exact test

two tailed

Fisher

exact

test

----c

*

3n of increased doses of interferon alpha 2b and with infection by genotypes 2 or 3 (Table III). Some variables observed during treatment showed an association with a sustained response. A decrease in HCV-RNA levels of at least one log was significantly associated with a sustained response: it occurred in all sustained responders and in 26 out of 52 (50%) patients without a sustained response (p
w-I*.v.-”

Table III. Sustained response predictors by stapwise logistic regression analysis in patients retreated with IFN-cr and ribavirin in combination for chronic hepatitis C. ~-~~~-~ --ms-_l_l*_--e.-w-e--i-ll_l Characteristic OR 95% Cl P .mem--k-“------*-* Treatment with 5 MU/day IFN CLSC 6.6 ?.7-42.8 0.009 Genotype 2 or 3 7 1.4 - 34.4 0.016 ~~~~~~,~ Ratio Statistics

6.18 p=O. 013; Abbreviations:

see list.

negative and poor positive predictor of sustained response in this series (100% negative predictive value and 30% positive predictive value). HCV-RNA levels below 1000 copies/ml after 1 month were not associated with a sustained response occurring in 6/11 (54%) patients with and in 16/52 (31%) without a sustained response; return to normal of ALT after 4 and 12 weeks and HCV-RNA levels below 100 copies after 12 weeks were strongly associated with a sustained response occurring, respectively, in 82%, 91% and 73% of patients with and in 38%, 58% and 36% of patients without a sustained response (p respectively co.01 by Mantel Haenszel test, 0.04 by Fisher exact test). However, both variables were poor positive and inaccurate negative predictors of sustained response: 3 l%, 25% and 30% positive predictive value and 94%, 97% and 92% negative predictive value, respectively, for return to normal of ALT at 4 and 12 weeks and for HCV-RNA below 100 copies/ml at 12 weeks. Safety Haemoglobin concentrations decreased to less than log per decilitre, requiring a reduction in the dose of ribavirin in 40% of patients: 8 in group A, 8 in group B and 9 in group C; anaemia improved after reduction of the ribavirin dose; the decrease in ribavirin dose was not associated with a lower rate of sustained response. The single interferon dose was reduced to 3 MU due to severe (grade III toxicity) flu-like symptoms in 2 patients from group B and in 4 patients from group C, two out of four showed a sustained response. Flu-like symptoms improved with reduction of the interferon dose. The decrease in haemoglobin, leukocyte and platelet counts was dose dependent. Haemoglobin median nadir was 11.6 g/d1 (interquartile range 10.7-12.5) in group A, 11.7 (11-12.6) in group B and 11.2 (10.312) in group C. (~~0.01 by Kruskal Wallis one-way ANOVA test). Nadir leukocyte counts were 3.1~10-~/1 (2.4-3.8) in group A, 2.8 (2.4-3.8) in group B and 2.6 (2.2-2.9) in group C (p=O.O2 by Kruskal Wallis oneway ANOVA test). Nadir platelet counts were 153x10. 9/1 (121-191) in group A, 153 (115-166) in group B and 115 (88-158) in group C (p
one-way ANOVA test). One patient in group C stopped treatment at the second month due to onset of severe muco-cutaneous lesions that did not improve with a reduction in both ribavirin and interferon doses, but re-

Table IV.

Side-effects and their grade according to treatment group.

W-V-IGroup A

Treatments: Interferon 2b SC: dose frequency Aibavirin dose per OS:

Group6

Group

C

3MU 5MU 5 MU 3 x week 3 x week 1 x day I-1.2 g daily I-1.2 g daily l-l.2 g daily 21

21

21

Fatiaua or asthenie WHO grade I WHO grad-IF!II

16 [76%1 15 1

18 [&%I 16 2

21 [lOO%l 17 a

Fever WHO grade I WHO grade II WHO grade Ill

16 f76%1 5 11 0

19 [90%1 8

21 [lOO%l 7 IO 4

Headache WHO grade I WHO grade II WHO grade HI

10 [48%1

9 [43%1 5 4 0

8 f38%1

Arthralgia WHO grade I WHO grade 110

4 [I 9%1 4 2

4 119%) E

3 [14%1 3

Anorexia WHO grade I

3 f14%1 3

3 f14%1 3

4 [I 9%1 4

Insomniairritability WHO grade I

2 UO%l 2

4 f19%1 4

3 [14%1 3

Nausea WHO grade I

3 [14%1 1

00 0

5 (24%15 5

Epigastric pain WHO grade I

2 [IO%1 2

1 Ki%l 1

2 [I O%l 2

1 l5%1

NLlflnber

Side-effect and grade:

z 1

Itching, cutaneous lesions WHO grade I WHO grade III

:

; 0

i 0

A

2 [IO%1 1 1

1 [5%1

0

0

Hypothyroidism

2 [IO%1

0

0

Depression WHO grade I WHO grade II

0 I:

1 ti%1 0 1

1 ci%l 0 1

::

:

1 @%I 1

Neuropathy

Hair loss WHO grade II No significant

difference

wes observedbetween

the treatmentgroups.

solved spontaneously after treatment was stopped. One patient in group A developed thyroiditis at the third month of treatment with an increase in anti thyroid peroxysome antibodies and an increase in thyreotrope hormone levels in excess of upper normal limits; the anti-thyroid antibody titre increased even after reducing the interferon dose; treatment was stopped and thyroid function recovered spontaneously after 3 months. A similar picture occurred in another patient in group A at the sixth month of treatment. Treatment was stopped at the end of the sixth month, according to treatment protocol, and the thyroid disorder resolved spontaneously in 2 months. Two other patients, one each from groups B and C dropped out voluntarily due to subjective intolerance to grade II flu-like symptoms. One patient in group A showed mild neuropathy (grade I toxicity) at the sixth month confirmed by electrophysiological study with grade I symptoms that resolved spontaneously after treatment was stopped at the sixth month. More than 75% of treated patients showed at least one side-effect as shown in Table IV The severity and the proportion of patients showing side-effects did not differ between the treatment groups, although side-effects tended to be more frequent and more severe in group C. The numbers of patients complaining of severe (grade III) toxicities were 5/21 in group C (24%; 95% CI: 6-42%), l/21 in group A (5%; 95% CI: O-14%) and 2/21 in group B (10%; 95% CI: O-22%); (~~0.05 group C vs group A; group C vs group B; group C vs groups A and B; two tailed Fisher exact test).

Fifty per cent of all chronic hepatitis C patients treated with interferon alpha during the last ten years were non-responders I 2; currently, there are no clinical data indicating that re-treatment of these patients will be beneficial. Many of the interferon non-responders had advanced fibrosis, therefore effective treatment strategies for these patients are of considerable clinical importance. New treatment strategies proposed for these patients include re-treatment with ribavirin, modifying the interferon dose from standard (3 MU three times a week) to higher doses (5 MU three times a week), even administered more frequently (5 MU once a day). In this pilot study on the re-treatment of previous non-responders to interferon monotherapy, we have demonstrated that, by increasing both the dose and the frequency of interferon alpha, it is possible to significantly increase the rate of sustained response to combination treatment, with only a slight increase in side-effects. Indeed, in the study population, high doses of interferon alpha ad-

ministered daily were significantly more effective than the standard doses; in addition, pooling the results obtained with different interferon doses in 63 non-responders, use of higher daily doses of interferon was associated with a sustained response irrespective of other factors. This study, though limited, shows an internal and external validity: we used strict selection criteria and validated definitions of treatment outcome, the three treatment groups were comparable and their characteristics were similar to those observed in non-responders, with the exception of the percentage of subjects with cirrhosis, which was slightly lower than in a previous series 7. However, cirrhosis has not been associated with a reduced response rate to interferon and ribavirin combination re-treatment of non-responders 6 and 42% of the study population showed severe fibrosis (stage 3 according to Knodell’s classification). It has been hypothesised that patients with a partial viral response during the initial course of treatment, i.e., l-2 log inhibition of HCV-RNA levels, might be more likely to respond than those in whom the viral levels do not change. We have no data on HCV-RNA quantitative assay during the previous cycle(s) of interferon treatment in our patients; however, transient negativisation of HCV-RNA during previous cycle(s), which was associated with a higher rate of response to a second treatment cycle in non-responders 29 was considered as an exclusion criterion. In this study, 38% (95% CI 17-59%) of treated patients showed a sustained response after treatment with high doses of interferon administered daily in combination with ribavirin. This response rate seems to be remarkably higher than reported with thrice weekly interferon administration in previous controlled studies published as full papers: cumulatively, in these studies, 278 patients were treated with interferon alpha administered three times per week in combination with ribavirin and only 24 out of 278 (9%; 95% CI 6-12%) showed a sustained response 2 l”-19. Our pilot study included too few patients to allow comparison with previously published series stratified by HCV genotype; however, 4 out of 5 of the patients infected by favourable HCV genotypes (2 and 3) and 4 out of 16 infected by unfavourable HCV genotypes showed a sustained response to the most intensive treatment strategy. This response rate seems to be higher than rates reported by a meta-analysis in non-responders to interferon monotherapy re-treated with combination therapy: 18% in patients with favourable and 6% in those with unfavourable genotypes 6. A higher daily dose of interferon alpha induced a more rapid and potent inhibition of viral replication than observed with a standard dose or increased dose with thrice weekly administration. This rapid and potent in-

hibition was significantly associated with a sustained response. Given the high turnover rates of HCV with rapid generation of viral diversity, 2o21 23 it is tempting to suggest that this more potent and earlier inhibition might have reduced the opportunity for viral escapes from antiviral therapy and from the host immune response stimulated by both interferon and ribavirin. In addition, the longer suppression of HCV replication due to this earlier inhibition obtained with high daily doses, might have allowed better elimination of infected hepatocytes by anti-HCV-specific cytotoxic T lymphocytes that seem to be enhanced by ribavirin 30. These hypotheses could explain why the rate of sustained response observed in this pilot study is so high. However, further studies on HCV quasispecies evolution and half-life of infected hepatocytes during combination treatment are needed to support these hypotheses. We treated our patients for 6 months only. An analysis of databases of international randomised trials 4 5 has demonstrated that it is possible to increase the response rates in patients with more than 2 out of 5 unfavourable prognostic factors (high fibrosis, older age, high viraemia, male sex or unfavourable HCV genotype) prolonging treatment for more than 6 months 3’. It is debatable whether treating patients for a longer period (i.e., 48 weeks for those infected by genotype 1) would have resulted in an even higher rate of sustained-response. Further studies are also needed to determine whether prolonging daily interferon doses beyond 6 months would be cost effective. It must be pointed out, however, that this study was aimed to demonstrate the feasibility of using an intensive dosage of interferon in combination with ribavirin, as suggested by recent data on hepatitis C viral kinetics. Therefore, these results should be confirmed in larger controlled trials, currently under way, before intensive interferon dosage can be extensively used in the re-treatment of non-responders to interferon monotherapy. In addition, other dose-finding trials need to be performed to determine the minimal effective daily dose (3 or 5 MU) and to compare daily doses of interferon alpha with once weekly administration of long-acting interferons such as pegylated interferon alpha 2b and 2a. Daily doses of interferon in combination with ribavirin, if confirmed as cost effective, should not be limited to interferon monotherapy nonresponders only. At present, most patients with chronic hepatitis C do not show sustained response 3-6even with interferon alpha and ribavirin combination therapy. Therefore, cost-effectiveness of the daily high-dose interferon schedule used in this trial, should be evaluated also in controlled trials conducted in selected groups of patients, naive to anti-HCV therapy, with poor pre-treatment probabilities of response 31.

In conclusion, we have demonstrated that an aggressive high daily dosage of interferon can be administered in combination with ribavirin in patients who failed to respond to previous interferon alpha monotherapy with a mild but acceptable increase in side-effects and that it is more effective than the standard interferon dose. These results pave the way for larger controlled trials, aimed at introducing into clinical practice a combination of ribavirin with more aggressive interferon schedules for the treatment of the “difficult” patients with chronic hepatitis C. Abbreviations Cl: Confidence Interval; HCV: Hepatitis pha; MU: million units: OR: odds ratio: 95% Confidence Interval

C Virus; IFN: interferon SC: subcutaneously; 95%

alCl:

I

i EASL International Consensus Conference on Hepatitis C. Paris 26-27 February 1999. Consensus statement. J Hepatol 1999;3l(Suppl 1):3-S. 2 Pagliaro L, Peri V, Linea C, Camml C, Giunta M, Magrin S. Natural history of chronic Hepatitis C. ha1 J Gastroenterol Hepatol 1999;31:28-44. 3 MC Hutchinson JG, Gordon S, Schiff ER, Shiffman ML, Lee WM, Rutsgi VK, et al. Interferon alfa-2b monotherapy versus interferon alfa 2b plus ribavirin as initial treatment for chronic hepatitis C: results of US multicenter randomized controlled trial. N Engl J Med 1998; 339:1485-92. 4 Davis GL, Esteban-Mur R, Rutsgi VK, Hoefs J, Gordon S, Trepo C, et al. Recombinant interferon alfa-2b alone or in combination with ribavirin for retreatment of interferon relapse in chronic hepatitis. N Engl J Med 1998;339: 1493-9. 5 Poynard T, Marcellin P, Lee SS, Niederau C, Minuk GS, Ideo G, et al. Randomized trial of interferon alfa-2b plus Ribavirin for 48 weeks or for 24 weeks versus interferon alfa 2b plus placebo for 24 weeks versus interferon alfa-2b with and without ribavirin for treatment of chronic infection with hepatitis C. Lancet 1998:352:1426-32. 6 Schalm SW, Brouwer JT, Bekkering FC, Van Rossum TGJ. New treatment strategies in non-responder patients with chronic hepatitis C. J Hepatol 1999;(Suppl 1):184X ’ Chemello L, Cavalletto L, Donada C, Bonetti P, Casarin P, Urban F, et al. Efficacy of a second cycle of interferon therapy in patients with chronic hepatitis C. Gastroenterology 1997;113:1654-9. 8 Alberti A, Chemello L, Noventa F, Cavalletto L, de Salvo GL Therapy of hepatitis C: retreatment with alpha interferon. Hepatology 1997:26 (Suppl 1): 137S42. 9 Heathcote EJL, Keefe EB, Lee SS, Feinman SV, Tong MJ, Reddy KR, et al. Re-treatment of chronic hepatitis C with consensus interferon. Hepatology 1998:27: 1136-43. lo Schalm SW, Hansen BE, Chemello L, Bellobuono A, Brouwer JT, Weiland 0, et al. Ribavirin enhances efficacv but not the adverse effects of interferon in chronic hepatitis C. Meta-analysis of individual patient data from European centres. J Hepatol 1997:26:961-6. ” Brillanti S, Garson J, Foli M, Whitby K, Deaville R, Masci C, et al. A pilot study of combination therapy with ribavirin plus in-

terferon alfa for interferon alfa resistant chronic hepatitis C. Gastroenterology 1994;107:812-7. I2 Schvarcz R, Ando Y, Sonnerborg A, Weiland 0. Combination treatment with interferon alfa-2b and ribavirin for chronic hepatitis C in patients who have failed to achieve sustained response to interferon alone: Swedish experience. J Hepatol 1995;23(Suppl 2):17-21. I3 Bellobuono A, Mondazzi L, Tempini S, Silini E, Vicari F, Ideo G. Ribavirin and interferon-a combination therapy vs. interferon-a alone in the re-treatment of chronic hepatitis C: a randomized clinical trial. J Viral Hepat 1997;4:185-91. I4 Sostegni R, Ghisetti V, Pittaluga F, Marchiaro G, Rocca G, Borghesio E, et al. Sequential versus concomitant administration of ribavirin and interferon alfa-n3 in patients with chronic hepatitis C not responding to Interferon alone: results of a randomized controlled trial. Hepatology 1998;28:341-6. I5 Andreone P, Gramenzi A, Cursaro C, Sbolli G, Fiorino S, Di Giammarino L, et al. Interferon-a plus ribavirin in chronic hepatitis C resistant to previous interferon-a course: results of a randomized multicenter trial. J Hepatol 1999;30:788-93. I6 Pol S. Couzigou P, Bourliere M, Abergel A, Combis JM, Larrey D, et al. A randomized trial of ribavirin and interferon-alpha vs. interferon-alpha alone in patients with chronic hepatitis C who were non-responders to a previous treatment. Multicenter Study Group under the coordination of the Necker Hospital, Paris, France. J Hepatol 1999;31:1-7. I7 Scotto G, Fazio V, Tantimonaco G Pilot study of a short course of ribavirin and alpha interferon in the treatment of chronic active hepatitis C not responding to alpha-interferon alone. Ital J Gastroenterol 1996;28:505-11. I8 Barbaro G, Di Lorenzo G, Belloni G, Ferrari L, Paiano A, Del Poggio P, et al. Interferon alpha-2B and ribavirin in combination for patients with chronic hepatitis C who failed to respond to, or relapsed after, interferon alpha therapy: a randomized trial. Am J Med 1999; 107:112-C I9 Bell H, Hellum K, Harthug S, Myrvang B, Ritland S, Maeland A, et al. Treatment with interferon-alpha2a alone or interferonalpha2a plus ribavirin in patients with chronic hepatitis C previously treated with interferon-alpha2a. CONSTRUCT Group. Stand J Gastroenterol 1999;34:194-8. ‘a Neumann AU, Lam NP, Dahari H, Gretch DR, Wiley TE, Layden TJ. Hepatitis C viral dynamics in vivo and the antiviral efficacy of interferon-a therapy. Science 1998;282:103-7. ” Lam NP, Neumann AU, Gretch DR, Wiley TE, Perelson AR, Layden AJ. Dose-dependent acute clearance of hepatitis C genotype 1 virus with interferon alpha. Hebatology 1997:26:226-31.

” Bekkering FC, Brouwer JT, Leroux-Roels G, Van Vlieberghe H, Eleuwat A, Schalm SW Ultrarapid hepatitis C virus clearance by daily high-dose interferon in non-responders to standard therapy. J Hepatol 1998;28:960-4. 23 Zeuzem S, Schmidt JM, Lee JH, von Wagner M, Teuber G, Roth WK. Quantification of the initial decline of serum hepatitis C virus RNA and response to interferon alfa. Hepatology 1998;27:1149-56. 24 Simon DM, Gordon SC, Kaplan MM, Koff RS, Regenstein F, Everson G, et al. Treatment of chronic hepatitis C with interferon alfa n-3: a multicenter, randomized, open label trial. Hepatology 1997;25:445-8. ‘j Bruno R, Debiaggi M, Sacchi P, Maffezzini E, Zara F, Brunetti E, et al. Daily interferon regimen for chronic hepatitis C. A prospective randomised study. Clin Drug Invest 1999; 18: 1 l-6. l6 Tassopoulos NC, Karvountzis G, Touloumi G, Delladetsima JK, Papatheodoridis GV, Katsoulidou A, et al. Comparative efficacy of a high or low dose of interferon alpha 2b in chronic hepatitis

C: a randomized multicenter trial. Am J Gastroenterol 1996;91:1734-8. >’ Lindsay KL, Davis GL, Schiff ER, Bodenheimer HC, Balart LA, Dienstag JL, et al. Response to higher doses of interferon alfa 2b in patients with chronic hepatitis C: a randomized multicenter trial. Hepatology 1996;24:1034-40. 28 Knodell RG, Ishak KG, Black WC, Chen TS, Craig R, Kaplowitz N, et al. Formulation and application of a numerical scoring system for assessing histological activity in asymptomatic chronic active hepatitis. Hepatology 1981;1:431-5. 2y Heathcote EJ, James S, Mullen KD, Hauser SC, Rosenblate H,

Albert DG Jr. Chronic hepatitis C virus patients with breakthroughs during interferon treatment can successfully be retreated with consensus interferon. The Consensus Interferon Study Group. Hepatology 1999;30:562-6. 3o Cramp ME, Ross01 S, Chokshi S, Carucci P, Williams R, Naoumov NV. Hepatitis C virus-specific T-cell reactivity during interferon and ribavirin treatment in chronic hepatitis C. Gastroenterology 2000;118:346-55. 31 Poynard T, MC Hutchinson J, Goodman Z, Ling MH, Albrecht J. Is an “A la Carte” combination Interferon alfa-2b plus ribavirin regimen possible for the first line treatment in patients with chronic hepatitis C? Hepatology 2000;3 1:21 l-8.

33rd EUROPEAN PANCREATIC CLUB The meeting will take place in Toulouse, France, June 13th-16th 2001. The goal is to gain new insights into a broad range of fields, exploring the potential clinical impact of basic research, in relation to pancreatic diseases, diagnosis, therapy and/or prevention. In addition to lectures by invited speakers, selected oral free presentations of original research work and poster sessions are included. A training course will be organised on Wednesday June 13th, supported by the Graduate School of Biology and Health Sciences, MD-PhD Program, University Paul Sabatier, Toulouse, on “Genomics and post genomics developments in biomedical sciences”. Contact: Nicole Vaysse,MD, PhD, INSERM U.531,IFR31, CHU Rangueil, 31403 Toulouse, France; call: +33 (0) 561322402; fax: +33 (0) 561322403; e-mail: nicole.vaysse @rangueil. inserm.fr; or visit: www.e-p-c.ovg.