| Background and AimsSeveral practice guidelines on chronic hepatitis B (CHB) have been established by some national or regional associations based on the medical evidences and have been updated periodically since2000.As indicated in Chinese guideline (update2010), the overall objectives of CHB treatment are:To suppress hepatitis B virus replication to the most extent, reduce liver cell inflammation, necrosis and liver fibrosis, delay and reduce hepatic decompensation, liver cirrhosis, hepatocellular carcinoma and their complications, thereby improving the quality of life and prolong survival time. CHB require long-term antiviral therapy, which is determined by the characteristics of HBV and the disease, Arbitrary withdrawal of antivirals can bring more serious clinical consequences; however, the importance of long-term treatment and adverse sequences of arbitrary withdrawal do not necessarily mean that the antivirals cannot stop in any occasion.In fact, proper discontinuation of antiviral treatment is one of the patient’s demands. A survey shows that:fewer patients willing to accept a long-term antiviral therapy, long-term adherence to treatment is poor, longer treatment causes higher economic pressures. Antiviral therapy is an etiological treatment, and a finite treatment course or discontinuation of treatment might be possible. The antiviral treatment durations were defined in the guidelines from American Association for the Study of Liver Diseases (AASLD), the European Association for the Study of the Liver (EASL), the Asian Pacific Association for the Study of the Liver (APASL) and Chinese CHB guideline.Although antiviral cessation criteria were proposed by the guidelines, such as a certain consolidation period after HBeAg seroconversion in HBeAg-positive CHB patients or achievement of HBsAg loss in HBeAg-negative CHB patients, but evidence-based medicine data is still insufficient. Meanwhile, we also found a subgroup of HBeAg-positive CHB patients whose HBeAb remained negative despite of HBeAg loss or HBsAg loss (measured with Abbott reagents) in our clinical practice. The cessation criteria in this subgroup also warrants exploration.In order to improve the treatment efficacy of nucleos(t)ide analogues(NA), reduce relapse after discontinuation, provide a reliable reference for clinicians and EBM data for guidelines. A prospective follow-up study on the durability of efficacy in the HBeAg-positive and HBeAg-negative CHB who met certain cessation criteria (see AASLD, EASL, APASL and Chinese CHB guidelines) after NA withdrawal were conducted since2001to evaluate the off-treatment durability of NA treatment and explore the predicting factor for the relapse.Material and MethodsPatientsThis prospective cohort study included279CHB patients who discontinued NA.①118patients in the HBeAg seroconversion group,82males,36females, aged5to52years (median23); Among them,90were given with lamivudine,13with adefovir dipivoxil,13with telbivudine, and2with entecavir. Cessation criteria were as follows:the total withdrawal of treatment≥18months, HBV DNA<103copies/ml, ALT normalization and HBeAg seroconversion maintained≥6months.②HBeAg loss group enrolled78patients,53males and25females, aged15to70years (median31); Among them,55patients with lamivudine,14with adefovir dipivoxil,6patients with telbivudine,3patients with entecavir. Cessation criteria:the total withdrawal of treatment≥18months, HBV-DNA<103copies/ml, ALT normalization and HBeAg loss maintained for≥6months.③HBeAg-negative group enrolled83patients,67males and16females, aged6to65(median34) years of age;67patients with lamivudine,14patients with adefovir, two patients with entecavir. Cessation criteria:total treatment≥24months, HBV-DNA<103copies/ml, ALT normalization maintained≥18months. Among relapsers after LAM withdrawal,24patients were retreated with LAM (100mg/day); and another112patients with initial lamivudine treatment well matched with the relapsers were selected as historical control to compare the incidence of YMDD motif mutation (rtM204I/V).MethodsThe detecting parameters and follow-up(A) HBV DNA quantification was performed by real time PCR (reagents by PG biotech, Shenzhen, China; detection limit of1000copies/ml) using ABi7000real-time PCRthermocycler.(B) HBV serological markers (hepatitis B virus marker, HBVM)quantitative detection was performed with chemiluminescent microparticle immunoassay (CMIA) with Abbott ARCHITECT i2000immunoassay analyzer and the original reagents. HBeAg≤1.0S/CO was defined as negative, anti-HBe≤1.0S/CO as positive. In the present study, the negative HBeAg plus anti-HBe<0.5S/CO is defined as HBeAg seroconversion in HBeAg-positive patients; anti-HBe negative or≥0.5S/CO is defined as HBeAg loss.(C) HBV genetic resistance mutation rtM204I/V detection:restriction fragment length polymorphism technique after nested PCR (nt-PCR-RFLP) were used to detect the mutation.(D) Follow ups after discontinuation were performed at1,2,3,4,6,9and12months after treatment, and then every6months thereafter.Conventional physical examination, liver function and HBV DNA quantification were performed in every visit. HBVM quantification was performed immediately if relapse founded, or it was performed every12months. Relapse was defined as HBV-DNA≥104copies/ml (confirmed by another test2weeks apart) and/or ALT elevated. Relapse was the primary endpoint of this study. For non-relapsers (all were followed up for≥12months), last follow-up for the study endpoint.(E) For the relapsers retreated with lamivudine, follow-up visits were made every3months and liver function and HBV-DNA were tested, the total follow-ups duration were≥2years. The YMDD motif mutation (rtM204I/V)was tested if relapse was observed (definition see above) or HBV-DNA≥104copies/ml lasted more than6months. The appearance of YMDD motif mutation (rtM204I/V) was the primary endpoint in this section.Statistical analysisAll data were collected and input into SPSS for windows13.0statistical software to organize, analyze data. Measurement data were tested with t test or Wilcoxon test; count data using x2test or Fisher’s exact test. The cumulative relapse rate and M2041/V (YMDD) mutation rate were calculated and tested using Kaplan-Meier method and Log-rank test.Factors related with off-treatment efficacy were explored using Cox proportional hazards model (Proportional Hazard Model). P<0.05was considered statistically significant.ResultTreatment duration and follow-up timeThe treatment durations were18to99months (median28.5),18to108months (median42), and24to87months (median30) months in HBeAg seroconversion group, HBeAg loss and HBeAg-negative group respectively. The median follow-up time (excluding relapsers) in HBeAg seroconversion group, HBeAg loss group and HBeAg-negative group was48(12to144),48(12to129), and60(12to144) months, respectively.Cumulative relapse rate and relapse time after discontinuationThe cumulative relapse rates in HBeAg seroconversion group, HBeAg loss group and HBeAg-negative was16.9%,39.7%and47.0%within1year;18.8%,43.0%and54.1%within3years;24.1%,43.0%, and61.1%within5years;24.1%,43.0%and64.3%within10years. HBeAg-negative group had the highest cumulative relapse rate, followed by the HBeAg toss group and HBeAg seroconversion group (log-rank test (Log-rank test) x2=30.133, P<.001.25patients in HBeAg seroconversion group relapsed, the relapse time ranged from2months to5years, the cumulative relapse relapse were44.4%of total number of cases (11cases),60.0%(15cases),80.0%(20cases) and88.0%(22cases) at6,12and24months after treatment. 33patients in HBeAg loss group relapsed. the relapse time ranged from1months to36months.the cumulative relapse relapse69.7%respectively of the total number of cases (23cases),84.8%(28cases),93.9%(31cases) and96.7%(32cases) at6,12and24months after treatment.49patients in HBeAg loss group relapsed, the relapse time ranged from1months to8years.The cumulative relapse relapse46.9%respectively of the total number of cases (23cases),53.1%(26cases),79.6%(39cases) and87.6%(43cases) at6,12and24months after treatment.Factors related with off-treatment efficacy durability of NA treatment(A) Univariate analysisIn HBeAg seroconversion group,12variables were analyzed and the results showed that age at withdrawal (t=4.251, P<0.001) and the treatment time (z=2.257, P=0.024) differedbetween relapsers and non-relapsers. In HBeAg loss group, only age at withdrawal diffed between relapsers and non-relapsers (t=2.208, P=0.030); In HBeAg-negative group,11variables were analyzed and only HBVDNA loss time between relapsers and non-relapsers was statistically different (z=2.257, P=0.024). Age at withdrawal of non-relapsers was30.4±13.4years and that of relapsers was35.0±11.2years; but the difference was not statistically significant (t=1.679,P=0.092).(B) Multivariate analysisThe HBeAg seroconversion group when118cases of chronic hepatitis B HBeAg withdrawal age, sex, time of administration, ALT levels at baseline treatment, AST levels, LgHBVDNA quantitative values, HBV-DNA disappearance time, HBVDNA disappear duration, HBeAg conversion duration, previous antiviral treatment history, family history of hepatitis B antivirals and12factors into the Cox proportional hazards regression model for multivariate analysis, get the best regression model. The results showed that when age and withdrawal relapse after withdrawal regression coefficient B is0.080,95%CI1.045-1.124; administration time and relapse after treatment, the regression coefficient B is-0.098,95%CI0.833-0.988.When the group of78cases of chronic hepatitis B HBeAg loss withdrawal age of 12factors into the Cox proportional hazards regression model for multivariate analysis, was the best regression model. The results showed that when age and stopping relapse of withdrawal, regression coefficient B is0.034,95%confidence interval,1.0055-1.066.The83cases when HBeAg negative chronic hepatitis B withdrawal age, sex, time of administration, ALT treatment baseline levels, AST levels, LgHBVDNA quantitative values, HBV-DNA disappearance time, HBVDNA disappear duration, previous treatment history of anti-virus, anti-hepatitis B viral drugs and family history of11factors into the Cox proportional hazards regression model for multivariate analysis, get the best regression model. The results showed that when age and withdrawal relapse after withdrawal, regression coefficient B of0.029,95%CI1.003-1.056; time HBVDNA seroconversion occurred with relapse after treatment, the regression coefficient is0.087B,95%confidence interval0.833-0.988.Stratified analysis on the impact factors of off-treatment durabilityThree patients were univariate and multivariate analyzes were performed to prove, age and other factors withdrawal when the predictors of relapse after withdrawal, the withdrawal of the age when stratified as≤20years old,21to30years old,21to30years old and>40years4groups.When HBeAg conversion with withdrawal age group increases, the cumulative relapse rate increased after treatment, especially in the age group over30years of cumulative relapse rate increased significantly, five years after discontinuation (60months) cumulative relapse rate of50%or more, and30-year-old age group and the cumulative relapse rate is low, the cumulative5-year relapse rate after treatment is about10%. By using receiver operating characteristic curve (ROC), calculate the area under the ROC curve (area under the ROC curve, AUC) to determine the optimal age of the critical point. When the critical point of age (cut-off value) was30.5years old, the largest area under the ROC curve of0.77, can be obtained as the best30years of age boundaries. The30-year-old as the age of the line is divided into two groups to calculate the cumulative relapse rate after treatment,>30age group the cumulative relapse rate after discontinuation of1,2-3,4, and5-year cumulative relapse rate was approximately35%,40%,45%and55%;<30years of age and the cumulative relapse rate of about10%.HBeAg disappears when the same group with the withdrawal age increases, the cumulative relapse rate increased after treatment, especially in the age group over40years of cumulative relapse rate, six months after stopping the cumulative relapse rate of over50%. Through the use of ROC, calculate AUC. When the critical point of age (cut-off value) was26.5years old, the largest area under the ROC curve was0.62, obtained26years as the best age boundaries. The26-year-old as the age of the line is divided into two groups, calculated the cumulative relapse rate after treatment, the results showed that>26age group the cumulative relapse rate after stopping a cumulative relapse rate of nearly50%after3years and no relapse.<26years age group the cumulative relapse rate is low, after stopping a cumulative relapse rate of about25%, and no relapse after1year.HBeAg-negative group showed that the age group over21years cumulative relapse rate is high. When the critical point of age (cut-off value) was20.5years old, the largest area under the ROC curve was0.59, obtained20-year-old as the best age boundaries. The20-year-old as the age of the line is divided into two groups, calculated the cumulative relapse rate after treatment, the results showed that the cumulative relapse rate after discontinuation, although HBeAg-negative group;<20years age group, but lower cumulative relapse rate after stopping1-year cumulative relapse rate of less than30%, no relapse after1year. And>20age group the cumulative relapse rate after stopping a cumulative relapse rate of51%,2to3years after stopping nearly60%, more than four-year cumulative relapse rate of about70%, and no relapse after8years.HBsAg loss and HBsAg seroconversion during treatment and follow upWhen stopping, HBsAg disappears in9cases, in HBeAg seroconversion group, disappeared group and HBeAg-negative group,3cases,4cases and2cases, respectively; among HBsAg seroconversion have3cases, in HBeAg seroconversion group, disappeared group and HBeAg-negative group were1case,1 case and1cases.In follow-up withdrawal, HBsAg disappears accumulated to29cases in HBeAg seroconversion group, disappeared group and HBeAg-negative group were9cases,10cases and10cases; in which HBsAg seroconversion has accumulated to19cases for HBeAg seroconversion group7cases,6cases of disappearance group, HBeAg-negative group6cases. Discontinuation of treatment and follow-up of patients with HBsAg loss occurred in the no stopping relapse.Resistance mutation in lamivudine retreated patients and naive patients24elapse after discontinuation of lamivudine treatment of chronic hepatitis B patients with a history of112patients with previously untreated control patients with M204I/V(YMDD) mutation cumulative incidence after treatment1and2years, respectively45.8%,17.9%and50.0%,32.6%. Discontinuation of treatment of chronic hepatitis B relapse in patients with M204I/V(YMDD) mutation cumulative incidence in previously untreated patients (P=0.017).Conclusion and SignificanceFirst, The cumulative relapse rates after discontinuation in various groups are different. HBeAg-negative group has the highest cumulative relapse rate, followed by HBeAg loss group and HBeAg seroconversion group. The cumulative relapse rates within10years were64.3%,43.0%and24.1%, respectively.Second, The relapse time in different groups has their own characteristics, which should be noted in the follow-up after treatment. HBeAg conversion group relapsed in two months after stopping to60months, no relapse after5years; withdrawal from3to6months, relapse accounted for about45%to60.0%, withdrawal1-2years, relapse accounted to80%to90.0%. HBeAg loss group relapsed after discontinuation of one month to36months, no relapse after3years; relapse mainly within one year after the withdrawal, while stopping three months, relapse accounted for nearly70.0%, withdrawal1year, relapse accounted for nearly94%. HBeAg-negative group relapsed after stopping early, the first months of relapse in8cases, accounting for16.3%of total recurrent cases; withdrawal from3to6months, relapse accounted for about50.0%; relapse can occur within as long as eight years.Third, Related factors with off-treatment durability of antiviral drug.For HBeAg seroconversion group, univariate analysis and multivariate analysis showed that when age and total treatment duration were independent predictors of relapse after treatment. Patients with older age and short treatment duration are more likely to relapse.For HBeAg loss group, univariate analysis and multivariate analysis showed that the age at withdrawal was an independent predictor of relapse after treatment. When the older withdrawal relapse rate, is a risk lactor of relapse after treatment.For HBeAg-negative group, multivariate analysis showed that the age and HBVDNA loss time were independent predictors of relapse after treatment. Older age and late HBV-DNA loss are high risk factors of relapse after treatment.Fourth, Age as independent predictors of off-treatment efficacy, and the cutoff value o fage using ROC analysis.The cutoff value of age in HBeAg seroconversion group is30years, the cumulative relapse rate in patients>30years after discontinuation was approximately35%,40%,45%and55%at of1,2-3,4and5years; in patients≤30years old the cumulative relapse rate of about10%. When considering NA withdrawal, age at withdrawal must be taken into account. Patients with older than30years should prefer extended treatment rather than N A withdrawal.The cutoff value of age in HBeAg loss group is26years, the cumulative relapse rate in patients>26years after discontinuation was approximately50%within1year; in patients≤26years old the cumulative relapse rate of about25%. When considering NA withdrawal, age at withdrawal must be taken into account. NA withdrawal could be considered in patients with≤26years, and should be carefully followed up.The cutoff value of age in HBeAg-negative group is20years, the cumulative relapse rate in patients≤20years after discontinuation was less than30%within1year; in patients>20years old the cumulative relapse rate of about51%within1year, and up to60%within2-3years. NA withdrawal could be considered in patients with≤20years, and should be carefully followed up.Fifth, No relapses were observed in all patients who experienced HBsAg loss in follow up. For patients with high risk of relapse in HBeAg-negative group and HBeAg-positive group, NA can be stopped when HBsAg disappear.Sixth, Lamivudine re-treatment is still effective for relapsers after lamivudine withdrawal; but resistance mutations are significantly more frequent than untreated patients. Therefore, NA with low resistance rates or interferon should be preferred when considering re-treatment in spite of lamivudine. |
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