Can Nucleos(t)ide Analogue (NA) Therapy Ever be Stopped in HBeAg-Negative Chronic Hepatitis B?

Transcription

1 DOI /s HEPATITIS B (P MARTIN AND P LAMPERTICO, SECTION EDITORS) Can Nucleos(t)ide Analogue (NA) Therapy Ever be Stopped in HBeAg-Negative Chronic Hepatitis B? Stephanos J. Hadziyannis & Dimitrios Vassilopoulos & Vassilios Sevastianos & Emilia Hadziyannis # Springer Science+Business Media New York 2014 Abstract In this review, evidence is presented that the majority of HBeAg-negative CHB patients previously treated with lamivudine and subsequently with adefovir and entecavir, experience virological and clinical relapse if these therapies were discontinued. Thus current treatment guidelines both of AASLD and EASL recommend indefinite duration of NA treatment in such patients. However, the recommendations of APASL are different, advising a stop of NA treatment if HBV-DNA is negative in three consecutive assays at least 6 months apart. It is clear that the duration of posttreatment sustained remission is variable and its final outcome unpredictable. However, fluctuations of post-treatment HBV- S. J. Hadziyannis (*): V. Sevastianos : E. Hadziyannis Liver Unit and its Molecular Biology Laboratory at Evgenidion Hospital, National and Kapodistrian University of Athens, Athens, Greece hadziyannis@ath.forthnet.gr V. Sevastianos vsevastianos@gmail.com E. Hadziyannis emhadzi@med.uoa.gr D. Vassilopoulos: E. Hadziyannis Second Department of Medicine at Hippokration Hospital, National and Kapodistrian University of Athens, Athens, Greece D. Vassilopoulos dvassilop@med.uoa.gr S. J. Hadziyannis 4 Dirrachiou Str., Athens 15669, Greece D. Vassilopoulos 114 Vas Sophias Ave., Athens 11527, Greece V. Sevastianos Ipsilantou 45, Athens 10676, Greece E. Hadziyannis 63 Evrou Str., Athens 11527, Greece DNA levels are not uncommon and HBeAg-negative CHB patients may experience long periods of transient HBV-DNA undetectability. Data on the frequency of post-treatment HBsAg loss are promising but still limited. The need for prospective, multinational, multicenter studies is stressed. Keywords Chronic hepatitis B. HBsAg quantification. Long term therapy. Nucleos/tide analogues. Interferon. Treatment discontinuation Introduction In patients with hepatitis B e antigen (HBeAg) negative chronic hepatitis B (CHB) treated with nucleos(t)ide analogues (NAs), discontinuation of therapy of 1 2 years duration is followed by virological and biochemical relapse in the vast majority [1 3]. Yet, in 2004 in a study of withdrawal of effective lamivudine treatment of 2-year duration in Canadian-Chinese patients with HBeAg-negative CHB, a sustained clinical and virological response was reported at post-treatment month 18 in up to 50 % of patients [4]. Similar results have been reported in subsequent studies of lamivudine discontinuation in HBeAg-negative CHB patients, conducted mostly in the early and middle 2000s [5 8]. However, because of frequent post-treatment clinical relapses, ALT flares [5, 9], occasional liver decompensation in patients with underlying cirrhosis and even because of some reports of fulminant hepatitis after stopping NA therapy [10, 11], physicians and NA treated CHB patients have been reluctant to attempt discontinuation of NA therapy. On the other hand, both in early and subsequent studies, retreatment of patients who had relapsed after discontinuation of NA therapy was always effective resulting in rapid complete responses [4, 12, 13, 14]. At the same time, because of high rates of development of HBV mutants resistant to lamivudine and less so to adefovir

2 [15], long term antiviral therapy was also complicated in clinical practice by virological and biochemical breakthroughs [16, 17]. However, with the development of the current first line NAs Entecavir and Tenofovir, of high anti-hbv potency and high barrier to HBV resistance [15], long-term/indefinite NA treatment of HBeAg-negative CHB achieves the goal of virological and biochemical efficacy without HBV resistance. Actually, this treatment paradigm represents a standard of care currently recommended by the AASLD and EASL Treatment Guidelines [18, 19]. Conversely, the Treatment Guidelines of the Asian Pacific Association for the Study of the Liver (APASL) both of 2008 [20] and 2012 [21] are different, recommending that NA treatment of HBeAg-negative patients with CHB can be discontinued if undetectable HBV-DNA has been documented on three occasions 6 months apart. In the current clinical practice of Western countries, discontinuation of NA therapy in HBeAg-negative CHB is practiced only in patients who achieve the closest to cure endpoint of HBsAg loss, preferably either following the development of anti-hbs or after a consolidation period of 12 months [22 24]. However, in a prospective study of discontinuation of effective long-term ADV treatment applied for 4 or 5 years to Mediterranean patients with HBeAg-negative CHB genotype D, unexpectedly high rates of sustained virological remission and of HBsAg loss were achieved [13, 25]. Moreover, in small cohorts of Western patients with HBeAgnegative CHB, sustained virological responses and HBsAg clearance have been reported after stopping long term NA therapy, particularly if HBsAg titers were <500 IU/mL [26 28]. The topic of discontinuation of NA therapy in HBeAgnegative CHB has been approached more extensively in Asian patients and a number of relevant studies, mostly retrospective and observational ones, has appeared in the literature during the last three years [12, 14, 29, 30, 31, 32, 33, 34, 35, 36, 37]. It is the purpose of this article to review critically the evidence on withdrawal of long term NA therapy in HBeAg-negative CHB under the provocative title proposed by the editors of Current Hepatitis Reports on whether NA therapy can ever be stopped in these patients. Discontinuation of NA therapy in HBeAg-positive patients will not be included in this article. Current international guidelines suggest that finite duration of treatment with NAs in HBeAg-positive CHB is a reasonable option and NAs can be stopped after HBeAg seroconversion followed by an additional 6 12 months consolidation period [18, 19]. However high rates of relapse have been observed mostly in Asian patients [38, 39, 40] and several aspects of this topic are still under study [41, 42]. The Rrationale for Stopping Effective Long-term NA Therapies in HBeAg-negative CHB It must be stressed a priori that contrary to the clinical importance of this topic, current worldwide data are few, fragmentary, restricted to small cohorts of patients, and mostly retrospective. Although current therapies with first line NAs in HBeAgnegative CHB even of very long duration do induce sustained biochemical and virological responses, this favorable effect uncommonly leads to HBsAg clearance [33, 43, 44]. Thus, the annual rate of HBsAg loss is less than 2.5 % and on the basis of HBsAg kinetics it has been calculated that in order to achieve this goal the duration of NA therapy has to be expanded for 2 3 and possibly more decades [45 50]. Also, in treatment naïve HBeAg negative patients who eventually cleared HBsAg, a slow decline of HBsAg has been documented and the best predictor of seroclearance 3 years later were HBsAg levels <200 IU/mL and 0.5 log annual reduction [51]. The total annual cost of effective first line NA therapies is quite high representing a considerable financial burden on the health systems particularly of poor countries of high and intermediate HBV endemicity. In several parts of the world patients treated with NAs, particularly with the most effective first line ones, are partly (or not at all) reimbursed for the cost of treatment [12, 14]. Finally, although side effects of NAs may be few, yet their very long-term safety is not proven [31, 35, 52]. In light of this it becomes reasonable to question whether after withdrawal of long term NA treatment, in HBeAgnegative CHB that has achieved virological and biochemical remission, can its efficacy become a sustained one that may even progress to HBsAg loss? If this proves to be the case at least in selected patients, then discontinuation of long-term NA therapy would be absolutely justifiable. After all no therapy should be continued for any longer than necessary [23]. What is the Evidence and Expectations for Favorable Outcomes of HBeAg-Negative CHB Following Withdrawal of long term NA Treatment Discontinuation of long-term therapy with NAs including lamivudine [4, 7, 31, 36 ], adefovir dipivoxil [13, 35] and entecavir [12, 53 55] has been attempted in HBeAg-negative CHB patients under treatment maintained remission aiming at sustained clinical and virological response terminating in HBsAg clearance. Reappearance of HBV DNA in serum at levels quantifiable by real time PCR assays has been reported to be universal but in several patients it was transient and lower than 2000 IU/mL [12, 13 ]. Virological

3 relapses with HBV DNA levels 2000 IU/mL do occur during the first year of post-treatment follow-up at rates higher than 70 % while clinical relapses, defined as HBV DNA levels 2000 IU/ml with increases in ALT levels 2 times the ULN have been reported in almost 50 % of patients. In most studies of withdrawal of NA therapy in HBeAg-negative CHB, patients who have experienced clinical relapses have been retreated immediately without further monitoring of the relapse neither for confirmation nor for evaluation of possible evolution to spontaneous remission. However, both virological and clinical relapses occurring after discontinuation of longterm NA therapy in HBeAg-negative CHB can be shortlived and with spontaneous return of ALT levels to normal and of serum HBV DNA to <2000 IU/ml or to non-detectability. It is noteworthy that in a study of CHB patients infected with HBV genotype D approximately 50 % of clinical relapses, that occurred after stopping long term NA therapy, resolved spontaneously and have also been followed by subsequent loss of HBsAg [13 ]. An example of such outcome of posttreatment clinical relapses is depicted in Fig. 1. A favorable spontaneous outcome of post-treatment relapses appears to be linked to a restored activity of the host immune system against HBV that might be attributed to the reduced HBsAg levels occurring during the preceding period of long NA treatment [43, 56 58]. However, this important point on the outcome of posttreatment relapses has not been evaluated in most of the relevant studies and on the basis of such relapses, the outcome of treatment discontinuation has been considered, a priori, as failure [39 ]. Thus, NA treatment has been promptly reinitiated. In this context it is appropriate to refer to the guidelines of the Japan Society of Hepatology published recently in English[59 ] which stress that after discontinuation of treatment, transient abnormalities in the ALT or the HBV DNA level may be observed in approximately two-thirds of patients who would finally achieve the inactive carrier state. Therefore, even if the ALT or HBV DNA levels show mild elevations, it is possible to follow up without retreatment EOT HBsAg (-) ALT (IU/L) HBV-DNA c/ml Months ALTMonthsHBV DNA Fig. 1 Withdrawal of ADV treatment of 5 years duration in an HBeAg negative patient with chronic hepatitis B (first arrow in the left) followed by virological relapse and subsequent ALT increase (clinical relapse). The patient was followed at 2 week intervals without retreatment. Note that spontaneous resolution of the relapse occurred and HBsAg was cleared later. HBV DNA was measured in copies/ml

4 For how long Should NAs be Continued before their Withdrawal is Attempted, what are the Possible Outcomes after Stopping Therapy and what Variables can be Applied as Predictors of Favorable Post-treatment Outcomes Studies of discontinuation of various NAs in HBeAg-negative CHB have been summarized in Table 1. In most of them the duration of treatment exceeds 2 years but no clear cut correlation between longer than 2-year duration of treatment and sustained response rates following its discontinuation has been documented. In the majority of the studies, summarized in Table 1, the APASL Guidelines have been adopted but with varying consolidation periods extending up to 96 weeks. In the study by Jeng et al. [12 ] it was shown that the longer the duration of the consolidation period the higher the posttreatment rate of sustained response, but this was true only for non cirrhotic patients who harbored HBV DNA levels higher than IU/mL. Similarly, in the study of Kim et al. lower baseline HBV DNA and longer total duration of treatment, including the period of consolidation, tended to be associated with lower relapse rates [35]. In a prospective study of discontinuation of 4 or 5 years of adefovir treatment [13 ], 54.5 % (18 of 33) of patients achieved a sustained response and 72 % of them (13 of 18) finally cleared HBsAg. All patients with HBsAg levels 100 IU/ml at the end of treatment (EOT) cleared HBsAg and a similar finding has been reported by Chan et al. [36 ] with HBsAg loss of 72 %, 5 years after stopping therapy among patients with HBsAg <100 IU/ml at EOT. From the data shown in Table 1 and particularly from the studies by Jeng et al. [12 ] and Kim et al. [35], it appears that rates of relapse are lower after withdrawal of ETV treatment compared to LAM and LTD. Similarly, sustained response rates have been reported to be higher and relapse rates lower in patients with baseline HBV DNA levels IU/mL [12 ]. In practically all studies, NA treatment was discontinued after achieving biochemical and virological remission. Although the duration of treatment was variable, it exceeded 2 years in all studies. However, the duration of post-treatment follow up was limited in many of them to only 12 months, a period during which the vast majority of relapses appear to occur [59 ]. However, sustained responses during the first year of follow up may subsequently evolve into virological and clinical relapses as this was clearly shown in the study by Liu et al. [31 ] with the total relapse rate increasing from 43.6 % at month 12 to 55 % at year 3. On the other hand, both virological and clinical relapses can be resolved spontaneously and terminate in HBsAg loss, a finding reported in 14 of 18 such untreated occasions [13 ]. These aspects have been evaluated only in a few studies and this has to be stressed since together with several other variables such as previous treatment, HBV genotypes, age and sex of included patients, HBsAg levels at baseline, their decline under therapy and the values reached at end of treatment [13, 27, 36, 60] may have an important impact on post treatment outcomes and may also prove to be useful predictors of favorable posttreatment outcomes. Particularly, as already stated above, restoration of the host immune activity against HBV may explain the very high levels of anti-hbc IgM and of serum IP-10 during most post-treatment clinical relapses terminating spontaneously in sustained virological remission and HBsAg loss [25]. This may be more frequently encountered in younger age groups with presumed shorter duration of chronic HBV infection as well as with significant decline in HBsAg titers that appear to contribute to restoration of impaired anti-hbv immune function [13, 31, 36, 56, 57]. What Criteria Should be Applied to Discontinuation of NA Treatment and how Strict and Frequent Should the Post-treatment Monitoring be? As already stated, discontinuation of long term NA therapy of HBeAg-negative CHB patients in biochemical and virological remission is expected to be followed by virological and clinical relapses in varying frequency depending on multiple factors. Virological relapses may either subside spontaneously or evolve into clinical relapses with increase in ALT levels of varying severity [13 ]. Accordingly strict monitoring of all patients is mandatory after stopping NA therapy. The vast majority of post-treatment relapses occur during the first year after withdrawal of treatment. Recently published guidelines for reduction of risk from discontinuation of nucleoside/nucleotide analogs in patients with chronic hepatitis B recommend that patients who discontinue treatment should be followed up by blood tests every 2 weeks up to post-treatment week 16 and every 4 weeks thereafter [59 ]. Although prompt NA retreatment of clinical relapses has been widely documented to be effective and safe securing a rapid virological and biochemical remission without progression of the underlying liver disease [4, 12, 13, 59 ], this may not always be the case and therefore the risk for hepatic decompensation in cirrhotic patients should be considered [39 ]. For safety reasons, withdrawal of effective NA therapy should not be encouraged in patients with cirrhosis even in those with compensated disease [13, 39 ]. At the same time it is noteworthy that patients with cirrhosis harbor relative lower baseline HBV DNA levels compared to non-cirrhotic ones and that among such patients there are fewer relapses and more frequent sustained responses following discontinuation of NA treatment [12, 35].

5 Table 1 Relapse rates at month 6 and 12 after withdrawal of NA therapy longer than 2 year duration, in HBeAg negative patients with chronic hepatitis B. Sustained virological response rates, when available, have also been entered. Criteria and definitions of virological and clinical response differed between the reviewed studies while data on long term post-treatment HBsAg loss have been reported in studies [13, 31, 32, 36, 55] ranging from 8.5 % to 42 % in 5 years of follow-up Author Year NA Treatment duration Patients effectively treated Follow-up duration (months) Post-treatment relapse Month 6 Month 12 SVR at end of follow-up n A. Viral B. Clinical A. Viral B. Clinical Fung et al. [4] 2004 LAM 2 years % 12 % 50 % 18 % 50 % Chan et al.[ 7] 2007 LAM 2 years % 74 % 26 % Liu et al. [31 ] 2011 LAM 2 years % 26 % 44 % 44 % 44 % Liang et al. [34] 2011 LAM,ADV,ENT 32±8 months ±6 40 % 40 % 47 %^ 47 %^ Chan et al. [36 ] 2011 LAM 34± 53 47± % 23 months Ha et al. [32 ] 2012 ADV 2 years % 54 % 61 % 61 % 36 % Hadziyannis et al ADV 4 or 5 years % 91 % 91 % 76 % 55 % [13 ] He et al.[33 ] 2013 LAM, ADV, ENT, % 30 % 70 % # LdT weeks Kim et al. [35] 2013 LAM,ADV, ENT 38± 45 20±11 49 % 36 % 73 % 53 % 27 % 10 months Jeng et al. [12 ] 2013 ENT 2 years % 56% 45% 64% LAM 2 years % LdT 2 years % Chen et al. [55] 2013 ETV weeks % 7 % 47 % 31 % 43 % LAM lamivudine, ADV adefovir, ETV entecavir, LdT telbivudine, SVR sustained virologic response ^ Mean time to relapse was 36 HBV DNA >200 IU/mL # Ranging from 46 % in LAM to 100 % in LdT treated What Further Studies are Needed Most studies of discontinuation of NA therapy in patients with HBeAg-negative CHB reviewed in the table are retrospective and observational ones, with small number of included patients, with significant differences between them concerning duration of treatment, specific NA, duration of post-treatment follow-up, and post-treatment outcomes. In most of them virological and clinical relapse rates at various points in time (mostly at 6 and 12 months after stopping therapy) have been evaluated with significant differences in the definition of relapses. Moreover, there is limited information on the long-term outcome both of virological and clinical relapses if these are left untreated and on the rates of spontaneous resolution as well as on the possibility of subsequent long-term loss of HBsAg. However, sustained post-treatment clinical and virological remission has been reported and documented now to be followed by frequent HBsAg loss particularly in patients who achieve significant decline in serum HBsAg concentration with long-term NA treatment and specifically among those who reach levels below 500 IU/mL and even lower [13, 25, 27, 61]. Moreover, several other variables such as age and sex of included patients, host genetic and immunological factors and reactivity and viral factors as specific HBV genotype, amount of cccdna in the liver, and number and pattern of HBsAg-positive hepatocytes may also play important additional roles [25]. All these variables should be taken into account in properly designed multinational, multicenter prospective studies aiming at elucidating the special characteristics of HBeAgnegative patients who will be eligible for safe withdrawal of long-term NA therapy terminating to HBsAg clearance. Despite a certain amount of pessimism expressed in a recent editorial [39 ], a prospective research approach of NA withdrawal is alreadyonitswaybynihandadditionalprospectivestudies appear to also be running in Europe.

6 Conclusions 1. Five years after discontinuation of effective long-term NA therapy in HBeAg-negative CHB patients, a significant proportion of them, accounting for >50 % in a prospective study, may remain in sustained remission and two thirds of those who achieve SVR may also be expected to experience subsequent HBsAg loss over time, an outcome that represents the one closest to cure of chronic HBV infection. 2. Such favorable outcomes appear to increase in frequency if NA treatment is discontinued at a point in time when HBsAg levels are lower than 500 IU/mL and preferably lower than 200 IU/mL. 3. Early post-treatment increases in serum ALT activity and even flares preceded by re-detectability of HBV DNA in serum represent immune mediated reactions, are usually short-lived and should be first followed carefully for spontaneous host induced resolution rather than being immediately retreated by NAs. 4. Close monitoring of those patients who discontinue long term NA therapy is mandatory to secure the safety of such a new treatment paradigm in HBeAg-negative CHB. 5. For the time being, the above remarks and conclusions on safe discontinuation of NA therapy in HBeAg-negative CHB refer only to non-cirrhotic patients and should not be extrapolated to cirrhotic ones as well. 6. Large prospective multinational and multicenter studies of this topic are required to confirm or reject the above preliminary concluding remarks. Compliance with Ethics Guidelines Conflict of Interest Vassilios Sevastianos declares no conflicts of interest. Stephanos Hadziyannis reports grants from Janssen, personal fees from Gilead, Novartis, Pfizer, outside the submitted work. Dimitrios Vassilopoulos reports personal fees from Abbott, Actelion, Bristol-Myers Squibb, MSD/Schering-Plough, Novartis, Roche, UCB, Wyeth/Pfizer, outside the submitted work. Emilia Hadziyannis reports personal fees from Roche Diagnostics, outside the submitted work. Human and Animal Rights and Informed Consent This article does not contain any studies with human or animal subjects performed by any of the authors. References Papers of particular interest, published recently, have been highlighted as: Of importance Of major importance 1. Tassopoulos NC, Volpes R, Pastore G, Heathcote J, Buti M, Goldin RD, et al. Efficacy of lamivudine in patients with hepatitis B e antigen-negative/hepatitis B virus DNA-positive (precore mutant) chronic hepatitis B. Lamivudine Precore Mutant Study Group Hepatology. 1999;29: Honkoop P, de Man RA, Niesters HG, Zondervan PE, Schalm SW. Acute exacerbation of chronic hepatitis B virus infection after withdrawal of lamivudine therapy. Hepatology. 2000;32: Santantonio T, Mazzola M, Iacovazzi T, Miglietta A, Guastadisegni A, Pastore G. Long-term follow-up of patients with anti-hbe/hbv DNA-positive chronic hepatitis B treated for 12 months with lamivudine. J Hepatol. 2000;32: Fung SK, Wong F, Hussain M, Lok AS. Sustained response after a 2-year course of lamivudine treatment of hepatitis B e antigennegative chronic hepatitis B. J Viral Hepat. 2004;11: Kim JK, Hwang SG, Park H, Choi HY, Cho HJ, Ko KH, et al. Rim KS: [Clinical outcomes after discontinuation of Lamivudine in chronic hepatitis B patients with Lamivudine resistant HBV mutant]. Korean J Hepatol. 2005;11: Chien RN, Liaw YF. Short-term lamivudine therapy in HBeAgnegative chronic active hepatitis B in Taiwan. Antivir Ther. 2006;11: Chan HL, Wang H, Niu J, Chim AM, Sung JJ. Two-year lamivudine treatment for hepatitis B e antigen-negative chronic hepatitis B: a double-blind, placebo-controlled trial. Antivir Ther. 2007;12: Huang YH, Wu JC, Chang TT, Sheen IJ, Lee PC, Huo TI, et al. Analysis of clinical, biochemical and viral factors associated with early relapse after lamivudine treatment for hepatitis B e antigennegative chronic hepatitis B patients in Taiwan. J Viral Hepat. 2003;10: Zhang NP, Reijnders JG, Perquin M, Hansen BE, Janssen HL. Frequency and clinical outcomes of flares related to nucleos(t)ide analogue therapy in patients with chronic hepatitis B. J Viral Hepat. 2011;18:e Lim SG, Wai CT, Rajnakova A, Kajiji T, Guan R. Fatal hepatitis B reactivation following discontinuation of nucleoside analogues for chronic hepatitis B. Gut. 2002;51: Wang J, Wang M, Huang Y. Acute liver failure resulting from discontinuation of nucleoside analogues in chronic hepatitis B patients: a report of two cases. Scand J Infect Dis. 2013;45: Jeng WJ, Sheen IS, Chen YC, Hsu CW, Chien RN, Chu CM, et al. Off-therapy durability of response to entecavir therapy in hepatitis B e antigen-negative chronic hepatitis B patients. Hepatology. 2013;58: A retrospective study showing an overall 45% clinical relapse at 1 year of follow up reduced with consolidation therapy and of 29% in patients with low HBV DNA at baseline. 13. Hadziyannis SJ, Sevastianos V, Rapti I, Vassilopoulos D, Hadziyannis E. Sustained responses and loss of HBsAg in HBeAg-negative patients with chronic hepatitis B who stop longterm treatment with adefovir. Gastroenterology. 2012;143: Prospective study with long term follow up of ADV discontinuation with high rates of sustained response and HBsAg loss. 14. Jin YJ, Kim KM, Yoo DJ, Shim JH, Lee HC, Chung YH, et al. Clinical course of chronic hepatitis B patients who were off-treated after lamivudine treatment: analysis of 138 consecutive patients. Virol J. 2012;9: Hadziyannis SJ, Vassilopoulos D, Hadziyannis E. The natural course of chronic hepatitis B virus infection and its management. Adv Pharmacol. 2013;67: Papatheodoridis GV, Dimou E, Laras A, Papadimitropoulos V, Hadziyannis SJ. Course of virologic breakthroughs under longterm lamivudine in HBeAg-negative precore mutant HBV liver disease. Hepatology. 2002;36: Hadziyannis SJ, Papatheodoridis GV, Dimou E, Laras A, Papaioannou C. Efficacy of long-term lamivudine monotherapy in patients with hepatitis B e antigen-negative chronic hepatitis B. Hepatology. 2000;32:

7 18. Lok AS, McMahon BJ. Chronic hepatitis B: update Hepatology. 2009;50: EASL clinical practice guidelines: Management of chronic hepatitis B virus infection. J Hepatol. 2012;57: Liaw YF, Leung N, Kao JH, Piratvisuth T, Gane E, Han KH, et al. Asian-Pacific consensus statement on the management of chronic hepatitis B: a 2008 update. Hepatol Int. 2008;2: Liaw YF, Kao JH, Piratvisuth T, Chan HLY, Chien RN, Liu CJ, et al. Asian-Pacific consensus statement on the management of chronic hepatitis B: a 2012 update. Hepatol Int. 2012;6: Invernicci F, Lampertico P, Loglio A, et al. Nucleos(t)ide analogues can be safely discontinued in chronic hepatitis B patients achieving HBsAg seroclearance. Hepatology. 2012;56:368A 369A. 23. Petersen J, Buti M. Considerations for the long-term treatment of chronic hepatitis B with nucleos(t)ide analogs. Expert Rev Gastroenterol Hepatol. 2012;6: quiz Perez-Cameo C, Pons M, Esteban R. New therapeutic perspectives in HBV: when to stop NAs. Liver Int. 2014;34 Suppl 1: Hadziyannis S, Sevastianos V, Rapti I. Outcome of HBeAgnegative chronic hepatitis B (CHB) 5 years after discontinuation of long term adefovir dipivoxil (ADL) treatment. J Hepatol. 2009;50:S Petersen J, Buggish P, Hinrichsen H, et al. Stopping long-term nucleos(t)ide analogue therapy before HBsAg loss in HBeAg negative CHB patients: follow-up of long term responders. J Hepatol. 2013;58:S Petersen J, Buggish P, Stoehr A, Hinrichsen H, Mauss S, Berg T, et al. Stopping long-term nucleos(t)ide analogue therapy before HBsAg loss or seroconversion in HBeAg negative CHB patients: experience from five referral centers in Germany. Hepatology 2011;54:1033A. 28. Hansen B, Comberg M, Chan HL. High relapse rates in HBeAg negative chronic hepatitis B patients after discontinuation of nucleos(t)ide analogues. Hepatology. 2012;56:412A. 29. Tsuge M, Murakami E, Imamura M, Abe H, Miki D, Hiraga N, et al. Serum HBV RNA and HBeAg are useful markers for the safe discontinuation of nucleotide analogue treatments in chronic hepatitis B patients. J Gastroenterol. 2013;48: Matsumoto A, Tanaka E, Suzuki Y, Kobayashi M, Tanaka Y, Shinkai N, et al. Combination of hepatitis B viral antigens and DNA for prediction of relapse after discontinuation of nucleos(t)ide analogs in patients with chronic hepatitis B. Hepatol Res. 2012;42: LiuF,WangL,LiXY,LiuYD,WangJB,ZhangZH,etal.Poor durability of lamivudine effectiveness despite stringent cessation criteria: a prospective clinical study in hepatitis B e antigennegative chronic hepatitis B patients. J Gastroenterol Hepatol. 2011;26: Prospective study with long term post lamivudine treatment follow up. 32. Ha M, Zhang G, Diao S, Lin M, Sun L, She H, et al. A prospective clinical study in hepatitis B e antigen-negative chronic hepatitis B patients with stringent cessation criteria for adefovir. Arch Virol. 2012;157: Prospective study of discontinuation of ADV with 5 years follow up. The majority of the relapses occurred during the firt year. Low rates of HBsAg seroconversion. 33. He D, Guo S, Chen W, Chen X, Yan G, Wang J, et al. Long-term outcomes after nucleos(t)ide analogues discontinuation in chronic hepatitis B patients with HBeAg-negative. BMC Infect Dis. 2013;13:458. Long term consolidation period of 96 weeks and low relapse rate <30%. 34. Liang Y, Jiang J, Su M, Liu Z, Guo W, Huang X, et al. Predictors of relapse in chronic hepatitis B after discontinuation of anti-viral therapy. Aliment Pharmacol Ther. 2011;34: Kim YJ, Kim K, Hwang SH, Kim SS, Lee D, Cheong JY, et al. Durability after discontinuation of nucleos(t)ide therapy in chronic HBeAg negative hepatitis patients. Clin Mol Hepatol. 2013;19: Chan HL, Wong GL, Chim AM, Chan HY, Chu SH, Wong VW. Prediction of off-treatment response to lamivudine by serum hepatitis B surface antigen quantification in hepatitis B e antigennegative patients. Antivir Ther. 2011;16: On the predictive value of HBsAg levels under therapy for sustained post treatment response and HBsAg loss. 37. Kim HY, Choi JY, Park CH, Jang JW, Kim CW, Bae SH, et al. Outcome after discontinuing antiviral agents during pregnancy in women infected with hepatitis B virus. J Clin Virol. 2013;56: Tseng TC, Liu CJ, Su TH, Yang HC, Wang CC, Chen CL, et al. Young chronic hepatitis B patients with nucleos(t)ide analogueinduced hepatitis B e antigen seroconversion have a higher risk of HBV reactivation. J Infect Dis. 2012;206: Reijnders JG, Janssen HL. Relapse of chronic hepatitis B after discontinuation of nucleos(t)ide analogs: is the glass half full or half empty? Hepatology. 2013;58: Critical approach of the topic of discontinuation of long tern NA treatment in CHB. 40. Chaung KT, Ha NB, Trinh HN, Garcia RT, Nguyen HA, Nguyen KK, et al. High frequency of recurrent viremia after hepatitis B e antigen seroconversion and consolidation therapy. J Clin Gastroenterol. 2012;46: Reijnders JG, Perquin MJ, Zhang N, Hansen BE, Janssen HL. Nucleos(t)ide analogues only induce temporary hepatitis B e antigen seroconversion in most patients with chronic hepatitis B. Gastroenterology. 2010;139: Kao JH. HBeAg-positive chronic hepatitis B: why do I treat my patients with pegylated interferon? Liver Int. 2014;34 Suppl 1: Seto WK, Wong DK, Fung J, Huang FY, Lai CL, Yuen MF. Reduction of hepatitis B surface antigen levels and hepatitis B surface antigen seroclearance in chronic hepatitis B patients receiving 10 years of nucleoside analogue therapy. Hepatology. 2013;58: Fasano M, Lampertico P, Marzano A, Di Marco V, Niro GA, Brancaccio G, et al. HBV DNA suppression and HBsAg clearance in HBeAg negative chronic hepatitis B patients on lamivudine therapy for over 5 years. J Hepatol. 2012;56: Reijnders JG, Rijckborst V, Sonneveld MJ, Scherbeijn SM, Boucher CA, Hansen BE, et al. Kinetics of hepatitis B surface antigen differ between treatment with peginterferon and entecavir. J Hepatol. 2011;54: Manesis EK, Hadziyannis ES, Angelopoulou OP, Hadziyannis SJ. Prediction of treatment-related HBsAg loss in HBeAG-negative chronic hepatitis B: a clue from serum HBsAg levels. Antivir Ther. 2007;12: Hadziyannis E, Hadziyannis SJ. Hepatitis B surface antigen quantification in chronic hepatitis B and its clinical utility. Expert Rev Gastroenterol Hepatol. 2014;8: Boglione L, D Avolio A, Cariti G, Gregori G, Burdino E, Baietto L, et al. Kinetics and prediction of HBsAg loss during therapy with analogues in patients affected by chronic hepatitis B HBeAg negative and genotype D. Liver Int. 2013;33: Chevaliez S, Hezode C, Bahrami S, Grare M, Pawlotsky JM. Longterm hepatitis B surface antigen (HBsAg) kinetics during nucleoside/nucleotide analogue therapy: finite treatment duration unlikely. J Hepatol. 2013;58: Zoutendijk R, Hansen BE, van Vuuren AJ, Boucher CA, Janssen HL. Serum HBsAg decline during long-term potent nucleos(t)ide analogue therapy for chronic hepatitis B and prediction of HBsAg loss. J Infect Dis. 2011;204: Seto WK, Wong DK, Fung J, Hung IF, Fong DY, Yuen JC, et al. A large case-control study on the predictability of hepatitis B surface antigen levels three years before hepatitis B surface antigen seroclearance. Hepatology. 2012;56: Fontana RJ. Side effects of long-term oral antiviral therapy for hepatitis B. Hepatology. 2009;49:S

8 53. Ridruejo E, Marciano S, Galdame O, Reggiardo MV, Munoz AE, Adrover R, et al. Relapse rates in chronic hepatitis B naive patients after discontinuation of antiviral therapy with entecavir. J Viral Hepat doi: /jvh Shouval D, Lai CL, Chang TT, Cheinquer H, Martin P, Carosi G, et al. Relapse of hepatitis B in HBeAg-negative chronic hepatitis B patients who discontinued successful entecavir treatment: the case for continuous antiviral therapy. J Hepatol. 2009;50: Chen C, Lee C, Hung J, et al. The role of hepatitis B surface antigen quantification predict HBV reactivation after discontinuation of entecavir treatment. Hepatology. 2013;58:655A. 56. Bertoletti A, Ferrari C. Innate and adaptive immune responses in chronic hepatitis B virus infections: towards restoration of immune control of viral infection. Gut. 2012;61: Boni C, Laccabue D, Lampertico P, Giuberti T, Vigano M, Schivazappa S, et al. Restored function of HBV-specific T cells after long-term effective therapy with nucleos(t)ide analogues. Gastroenterology. 2012;143: Tseng TC, Liu CJ, Yang HC, Su TH, Wang CC, Chen CL, et al. Determinants of spontaneous surface antigen loss in hepatitis B e antigen-negative patients with a low viral load. Hepatology. 2012;55: Tanaka E, Matsumoto A. Guidelines for avoiding risks resulting from discontinuation of nucleoside/nucleotide analogs in patients with chronic hepatitis B. Hepatol Res. 2014;44:1 8. Comprehensive review and guidelines for safe discontinuation of long term NA therapy in CHB. 60. Seto WK, Liu K, Wong DK, Fung J, Huang FY, Hung IF, et al. Patterns of hepatitis B surface antigen decline and HBV DNA suppression in Asian treatment-experienced chronic hepatitis B patients after three years of tenofovir treatment. J Hepatol. 2013;59: Petersen J, Hansen B, Buggish P, et al. Discontinuation of long-term nucleos(t)ide analogue therapy before HBsAg loss in HBeAg negative CHB patients:follow-up of long-term responders. Hepatology. 2013;58:680A.