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Clearance of Hepatitis C Virus after Newly Acquired Infection in IDUs
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The Journal of Infectious Diseases Oct 1, 2004;190:1270-1274
Marianne Jauncey,1,2 Joanne M. Micallef,3 Stuart Gilmour,2,a Janaki Amin,3 Peter A. White,4 William Rawlinson,5 John M. Kaldor,3 Ingrid van Beek,2 and Gregory J. Dore3
1New South Wales Public Health Officer Training Program, New South Wales Department of Health, 2Kirketon Road Centre, South Eastern Sydney Area Health Service, 3National Centre in HIV Epidemiology and Clinical Research and 4School of Biotechnology and Biomolecular Sciences, Faculty of Science, University of New South Wales, and 5Virology Division, Department of Microbiology, Prince of Wales Hospital, Sydney, New South Wales, Australia
SUMMARY: A retrospective cohort of injection drug users with newly acquired hepatitis C virus (HCV) infection was established to examine viral clearance. Newly acquired HCV infection was defined by anti-HCV antibody seroconversion within a 2-year interval. Stored serum samples were tested for HCV RNA, with viral clearance defined as >2 consecutive negative HCV RNA test results after infection. Ninety-nine cases of HCV infection were identified; 57 had >HCV RNA test results after infection. Viral clearance occurred in 24 (42%) cases, with Kaplan-Meier estimated probabilities of 23%, 38%, and 40% at 6, 12, and 24 months, respectively.
For each person identified as having newly acquired HCV infection, all available stored serum samples from the time of the last negative HCV antibody test result were retrieved from storage and were tested retrospectively for the presence of HCV RNA. Quantitation of HCV RNA was performed on all specimens by use of the VERSANT HCV RNA 3.0 Assay (HCV 3.0 bDNA Assay; Bayer Diagnostics). The assay has lower and upper detection limits of 3.2 × 103 and 4.0 × 107 copies/mL, respectively. Specimens that yielded a value below the detection limit were tested for HCV RNA by use of nested reverse-transcriptase polymerase chain reaction, as described elsewhere. Validity of retrospective HCV RNA testing was assessed in 22 specimens for which clinical qualitative HCV RNA testing had been conducted at the time of sampling. There was concordance for 13 of 14 positive and 8 of 8 negative clinical specimens obtained during the period 1994--2001 (95% of all validation specimens). The single discordant specimen was positive for HCV RNA by clinical testing in August 2000 but was negative by retrospective testing.
INTRODUCTION
Estimates of viral clearance following acute hepatitis C virus (HCV) infection range from 10% to 50%. The small sample size and selected populations in most studies of acute HCV infection have limited the estimation of viral clearance. In general, studies have been based on cases of acute clinical hepatitis C or posttransfusion HCV infection that may not be more broadly representative of acute HCV infection. Progression to chronic HCV infection has generally been defined by the persistence of HCV RNA for at least 6 months; however, recent evidence suggests that spontaneous viral clearance may extend to 2 years. Consequently, uncertainty remains regarding the rate, time course, and predictors of viral clearance following acute HCV infection.
In Australia, >90% of new HCV infections are attributed to injection drug use, and the estimated incidence of HCV increased from 11,000 cases in 1997 to 16,000 cases in 2001. The ongoing collection and storage of serum samples obtained from a large cohort of injection drug users (IDUs) attending a primary care clinic in Sydney provided the opportunity to conduct an investigation of the natural history of newly acquired HCV infection. Specific aims were to determine the rate, time course, and predictors of viral clearance.
Viral clearance after newly acquired HCV infection was the primary outcome variable. The date of HCV infection was estimated as the midpoint between the last negative HCV antibody test result and the first evidence of HCV infection—that is, either a positive HCV antibody test result, an indeterminate HCV antibody test result (if followed by a positive test result), or a positive HCV RNA test result, whichever occurred earliest. Viral clearance was defined as 2 consecutive undetectable HCV RNA test results after the estimated date of infection, with the midpoint between the last positive HCV RNA test result and the first negative HCV RNA test result as the estimated date of viral clearance. In cases with no positive HCV RNA test result, the date of viral clearance was estimated as the midpoint between the estimated date of infection and the first negative HCV RNA test result. Cases in which at least 2 HCV RNA test results were not available were excluded from the analyses of viral clearance.
RESULTS
Viral clearance occurred in 42% (24/57) of IDUs. Estimated viral clearance by Kaplan-Meier survival analysis was 23%, 38%, and 40% at 6, 12, and 24 months, respectively. Median time to viral clearance was 5.9 months (IQR, 1.4--11.2 months). There were no baseline demographic, clinical, or behavioral factors significantly associated with viral clearance. IDUs who experienced ALT normalization were more likely to have experienced viral clearance; however, this did not reach significance (odds ratio, 1.90; 95% confidence interval, 0.77--4.65). Estimates of viral clearance at 6, 12, and 24 months were 27%, 42%, and 45% for IDUs who experienced HCV antibody seroconversion in 1992--1996 (n = 30), compared with 19%, 34%, and 34% for IDUs who experienced seroconversion in 1997--2002 (n = 27), with no significant difference in the overall time to clearance (P = .30).
For 24 of these 57 IDUs, all specimens tested within 1 year of the estimated time of infection were HCV RNA negative. Because of concerns regarding potential false-positive HCV antibody seroconversion, an analysis of IDUs with detectable HCV RNA within 1 year of the estimated time of infection (33/57 IDUs) was performed. This analysis showed that 24% (8/33) of IDUs had viral clearance, with Kaplan-Meier estimates of 6%, 23%, and 26% at 6, 12, and 24 months, respectively. There were no significant differences in baseline characteristics or predictors of clearance between IDUs who were and were not HCV RNA positive within 1 year of the estimated time of infection.
AUTHOR DISCUSSION
Among a large cohort of IDUs with newly acquired HCV infection, estimates of viral clearance at 2 years were 26%--40%, depending on the criteria used for case inclusion. Although estimated viral clearance rates varied according to the strictness of the case definition, there were no baseline demographic, clinical, or behavioral factors associated with clearance. The vast majority of clearances occurred within the initial 12 months after the estimated time of infection.
The present study has several limitations. We relied on retrospective HCV RNA testing of serum samples stored at -20°C. Storage conditions are known to affect the stability of HCV RNA, and degradation of HCV RNA occurs with freeze-thaw cycles and storage temperatures below -70°C. A validation based on 22 stored specimens found 95% concordance between the retrospective and concurrent test results, and estimated clearance rates did not differ significantly by time period of the stored specimens. Testing intervals for HCV RNA varied with some wide intervals, potentially leading to overestimation of the time to viral clearance. However, the HCV RNA testing interval was >9 months for only 25% of samples. Viral clearance was defined as >2 consecutive negative HCV RNA test results with a sensitive qualitative HCV RNA assay, but low-level HCV viremia with relapse has been described early during HCV infection. We have reported preliminary findings on HCV reinfection in this cohort. Three of the 33 HCV viremic IDUs at baseline had a further positive HCV RNA test result following a series of negative HCV RNA test results. These IDUs had a minimum of 4 consecutive negative HCV RNA test results over the course of 12, 19, and 26 months, and 2 had a change in HCV genotype, indicating probable cases of reinfection. Detection of clinical symptoms related to acute hepatitis relied on a review of medical notes; therefore, ascertainment bias may explain the absence of an association between the presence of these symptoms and viral clearance.
As in a previous study of a Baltimore cohort of IDUs, we have used a case definition for newly acquired HCV infection based on HCV antibody seroconversion. In contrast, most other studies have monitored subjects with either acute clinical hepatitis or posttransfusion HCV infection. To assess viral clearance, the majority of these studies have required the detection of HCV RNA at baseline. We, therefore, performed analyses of the subgroup of IDUs with confirmed HCV RNA detection occurring near the estimated time of HCV infection. Within this subgroup, viral clearance was estimated at 〜26% at 2 years, higher than the 15% estimate at 3 years from the Baltimore cohort of IDUs but similar to the 25% preliminary estimate from a meta-analysis that we have recently conducted on studies of acute and newly acquired HCV infection. When HCV RNA was not detected at baseline, estimates of viral clearance at 2 years were higher (〜40%).
Predictors of viral clearance in previous studies have included female sex, ethnicity, symptomatic presentation, absence of HIV infection, rapid decline in HCV RNA load, and the strength and pattern of HCV-specific CD4 and CD8 cell responses. The lack of significant demographic predictors of viral clearance within our cohort may relate to the relatively homogenous nature of the study population. The vast majority of IDUs were young, white, HIV antibody negative, and had asymptomatic HCV infection. Furthermore, despite our study population being one of the largest cohorts of subjects with acute or newly acquired HCV infection yet described, the sample size may have limited the detection of significant predictors of viral clearance.
We have demonstrated that at least 1 in 4 IDUs with newly acquired HCV infection undergoes viral clearance. The inclusion of IDUs who had experienced HCV antibody seroconversion, rather than acute clinical hepatitis alone, makes our study more broadly representative of newly acquired HCV infection than most previous studies have been.
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