Author Affiliations: Department of Gastroenterology, Marmara University Faculty of Medicine, Istanbul, Turkey (Yegin EG and Ozdogan OC)

Corresponding Author: Ender G Yegin, MD, Department of Gastroenterology, Marmara University Faculty of Medicine, Fevzi Çakmak mahallesi, Mimar Sinan caddesi, no. 41, Üst Kaynarca, Pendik, Istanbul, Turkey (Tel: +90-216- 625-4545; Email: drendergunes@hotmail.com)

 

© 2014, Hepatobiliary Pancreat Dis Int. All rights reserved.

doi: 10.1016/S1499-3872(14)60303-9

Published online November 10, 2014.

 

Contributors: YEG proposed the study. YEG and OOC performed research. YEG wrote the draft. Both authors contributed to the design and interpretation of the study and to further draft. YEG is the guarantor.

Funding: None.

Ethical approval: This study was approved by Central Research Ethics Commitee.

Competing interest: No benefits in any form have been received or will be received from a commercial party related directly or indirectly to the subject of this article.

BACKGROUND: The definition of partial virological response (PVR) was proposed because of its clinical relevance. PVR relates to subsequent therapeutic failure which results in the modification of the regimen. Whether this rationale can be applied to all nucleotide analogues (NA) is not clear. This study was undertaken to analyze PVR influence on therapeutic outcomes during lamivudine, entecavir or tenofovir monotherapy in NA-naive patients with chronic hepatitis B in routine clinical practice.

 

METHODS: We retrospectively analyzed 150 NA-naive patients with chronic hepatitis B. These subjects received lamivudine, entecavir or tenofovir monotherapy between February 2001 and July 2013.

 

RESULTS: Sixty-nine patients were treated with lamivudine, 35 with entecavir, and 46 with tenofovir. The median therapeutic duration was 19.5 (6-147) months. PVR rates at 24 weeks were similar among three NAs (lamivudine 33.3%, entecavir 35.0%, tenofovir 32.4%, P=0.981). For all three NAs, patients with a higher baseline viral load or HBeAg-positive status had a higher serum viral positive rate tested by polymerase chain reaction at week 24 and 48. Cumulative probability of virological breakthrough (VBR) for patients treated with lamivudine was 67% at 5 years, and PVR at 24 weeks was the independent risk factor for VBR (HR: 3.09; 95% CI: 1.09-8.74; P=0.034); also lamivudine treated patients older than 50 years seemed to have a tendency for VBR (HR: 2.80; 95% CI: 0.99-8.18; P=0.052). A majority of entecavir and tenofovir partial responders achieved and maintained virological response with prolonged monotherapy, except one entecavir treated patient who experienced VBR due to resistance mutations.

 

CONCLUSIONS: Management strategy for lamivudine treatment should include adaptation of regimen according to PVR as an on-treatment response parameter due to its relation with unacceptably high VBR probability. Similar conclusion should not be directly related to entecavir or tenofovir treatment.

 

(Hepatobiliary Pancreat Dis Int 2014;13:602-611)

 

KEY WORDS: partial virological response; lamivudine; entecavir; tenofovir; therapeutic outcome

Chronic hepatitis B (CHB) virus infection is a global health problem. An estimated 400 million people worldwide are chronically infected, causing approximately 1 million deaths each year from end-stage liver disease or hepatocellular carcinoma (HCC).^[1] The risk of developing cirrhosis or HCC is thought to be correlated with increasing HBV DNA levels, and the effective suppression of HBV DNA levels has been found to reduce the risk of these complications.^[2-6] Therefore, prolonged effective suppression of viral replication is considered the primary goal of CHB treatment.^[7]

 

Lamivudine, which was introduced in 1998, revolutionized the management of CHB. Lamivudine has been an extensively studied and well-established antiviral with proven efficiency, safety, and relatively low cost.^{[8, 9]} During the past decade, adefovir dipivoxil (2002), entecavir (2005), telbivudine (2006), and tenofovir disoproxil fumarate (2008) have been licensed globally. Lamivudine has been used as a comparator for these newer nucleotide analogues (NA) in clinical studies. Because complete eradication of HBV infection is a seldomly achieved goal, the treatment focuses on suppressing viral replication in a sustained manner. Maintenance of viral suppression is known to be a determinant of therapeutic outcomes like biochemical, serological or histological responses,^{[10, 11]} but requires long-term, sometimes indefinite treatment. Unfortunately, long-term treatment with NAs can be limited with an increasing risk of developing resistance, which may lead to virological breakthrough (VBR), biochemical breakthrough (BBR), and sometimes hepatic decompensation,^[12] and also may reverse the histological benefit achieved.^[13]

 

Lamivudine therapy carries a high risk of emergence of resistant mutants with rates approaching to 70% after four years of therapy.^[14-16] Despite the recommendation against the use of lamivudine as the first-line therapy with potent drugs like entecavir and tenofovir in current guidelines, lamivudine is still a widely prescribed drug in Asian, sub-Saharan African and Alaskan populations.^[17] Therefore, measures should be taken to optimize the application of available therapies, and predictive factors determining the outcomes of the therapies are of great importance.

 

A relationship between suboptimal responses at 24 weeks and increasing risk of resistance in the subsequent months for lamivudine and telbivudine, and a similar relationship at 48 weeks for adefovir were reported.^[11,18-22] Thus, a new definition, "partial virological response" (PVR), i.e. a decrease in HBV DNA of more than 1 lg IU/mL but detectable HBV DNA after at least 6 months of therapy, was proposed. The treatment with a more potent drug in patients receiving lamivudine or telbivudine with a PVR at 24 weeks, and in those receiving adefovir with a PVR at 48 weeks has been emphasized in guidelines.^[23] However, there are not enough data on the therapeutic outcomes of patients with PVR for NAs like entecavir and tenofovir with potent activity and low risk of resistance. The scarce evidence supporting the recommendation of treatment with entecavir and tenofovir in guidelines reflects the uncertainty of optimal management of these patients.

 

Therefore, the present study aimed to analyze the PVR and subsequent breakthrough outcomes during treatment with lamivudine, entecavir or tenofovir in routine clinical practice, and to assess the long-term efficacy of these NA(s) with regard to PVR.

 

 

A total of 150 adult NA-naive CHB patients treated with lamivudine, entecavir or tenofovir between February 2001 and July 2013 in Hepatology Clinics at Marmara University Faculty of Medicine Hospital in Istanbul, Turkey, were retrospectively identified. Laboratory and clinical data were collected by an extensive review of medical records. Eligibility criteria for inclusion in the analysis were: (a) presence of hepatitis B surface antigen (HBsAg) positivity for at least 6 months; (b) treatment with 100 mg of lamivudine, 0.5 mg entecavir or 245 mg tenofovir disoproxil fumarate once daily ; (c) a baseline viral load of at least 2000 IU/mL before treatment; (d) duration of treatment for at least 6 months; (e) neither previous treatment with NA nor interferon-based treatment. We also included patients with cirrhosis diagnosed histologically or by imaging with compatible clinical features.

 

Patients were excluded if they had incomplete information for the analysis, were lost to follow-up, and had evidence of HCC, viral coinfections (hepatitis C, D or human immunodeficiency viruses) or pregnancy. The main cause of lost patients was transfer of patients to other clinics due to the moving of our hospital to another location in 2010, and the other causes of noncompliance included assurance and monetary problems, lack of family support, and alternative medication.

 

The study protocol was approved by the Central Research Ethics Committee. All patients were followed up at the out-patient clinics and monitored for serum HBV DNA, liver chemistry and other laboratory parameters every 3-6 months to evaluate the responses at several time points during therapy. The frequency of follow-up measurements was determined by the treating physicians according to the severity of liver disease and type of NA. Patient's follow-up was ended at the time of the VBR or the last visit. For a patient experiencing VBR, the therapy was changed appropriately with a drug that does not share cross-resistance (tenofovir, for lamivudine VBR).

 

Virological response (VR) was thought to achieve undetectable HBV DNA by polymerase chain reaction (PCR) assay. Biochemical response was defined as the decline of alanine aminotransferase (ALT) levels within the normal range in ≥2 consecutive determinations during therapy. Serological response for HBeAg was applied only to HBeAg-positive patients, and was defined as HBeAg loss and seroconversion to anti-HBe. PVR, which was defined as a decrease in HBV DNA of more than 1 lg IU/mL but a detectable HBV DNA level by real-time PCR assay after at least 24 weeks of therapy,^[23] was assessed after 24 and 48 weeks of therapy in our study. VBR was considered the increase of serum HBV DNA level more than 1 lg IU/mL compared to the nadir HBV DNA level during continued therapy. Primary non-response was defined as the inability to reduce serum HBV DNA by 1 lg IU/mL from baseline after the first 12 weeks of therapy. BBR was considered as the increase of ALT levels to >1.5 upper limit of normal (ULN) after ALT normalization during continued therapy. Hepatitis flare was defined as an abrupt elevation of serum ALT over five folds of the ULN.^{[24, 25]}

 

Serum HBV DNA levels were determined by using a quantitative real-time PCR assay, the COBAS AmpliPrep-COBAS TaqMan HBV test (CAP-CTM; Roche Molecular Systems, Inc., Branchburg, NJ, USA), which had a lower limit of detection of 20 copy/mL (1 IU/mL is equivalent to 5.82 copy/mL).

 

Genotypic resistance analysis was conducted with a population-based sequencing method (i.e., a direct sequence analysis of the HBV polymerase gene) in two entecavir patients: one with a VBR, and another with a suboptimal response after prolonged treatment.

 

Continuous data were expressed as mean±standard deviation (SD) or median with range, and compared using the independent-samples t test, one-way ANOVA, the Mann-Whitney test and the Kruskal-Wallis test where appropriate. Categorical variables were compared using the Chi-square test or Fisher's exact test.

 

The cumulative probability of VR and VBR events were calculated using the Kaplan-Meier method and comparison between groups was performed using the log-rank test. Cox regression model was applied to identify the factors that were independently associated with the VBR to lamivudine therapy, by fitting variables that demonstrated a P value <0.05 in the univariate model.

 

When retrospective data were not available at some point of the treatment, the analyses were performed on the existing percent of patients with data.

 

A P value <0.05 was considered statistically significant. All the data were analyzed using SPSS version 17.0 (Chicago, IL, USA).

 

 

Data from the 150 naive patients with CHB treated with NA were analyzed. The mean age of the patients was 48.1 years (range 18-89). In this series, 89 (59.3 %) patients were men, 119 (79.3%) were HBeAg-negative, and 41 (27.3%) had cirrhosis. The median duration of treatment was 19.5 months (range 6-147). One hundred five patients (70.0%) continued to receive antiviral therapy for the second year, 62 (41.3%) for the third year, and 42 (28.0%) for more than 3 years. The median serum HBV DNA level at baseline was 6.09 lg IU/mL (range 3.3-8.9). The baseline characteristics of the patients are presented in Table 1.

 

The baseline median HBV DNA level was significantly higher in patients treated with entecavir or tenofovir than in those treated with lamivudine (P=0.010 and P=0.000, respectively); 17.4% patients treated with lamivudine, 34.3% with entecavir and 43.5% with tenofovir had a baseline HBV DNA level greater than 7 lg IU/mL (P=0.008). HBeAg-negative was related to NA treatment.

 

After 24 and 48 weeks treatment, the percentage of patients achieving VR was 66.7% and 81.8% for lamivudine, 65.0% and 81.3% for entecavir, and 67.6% and 86.4% for tenofovir (P=0.981 and P=0.788), respectively.

 

For HBeAg-negative patients, the cumulative probabilities of achieving VR at week 48 and 96 were 89%, 96% for lamivudine; 92%, 100% for entecavir, and 96%, 100% for tenofovir, respectively. For HBeAg-positive patients, corresponding cumulative VR probability rates were 58%, 58% for lamivudine, 32%, 71% for entecavir, and 93%, 93% for tenofovir, respectively.

 

In patients with high baseline ALT levels, 93.1% treated with lamivudine achieved a biochemical response in 3 (1-18) months, 96.0% with entecavir in 4 (1-45) months, and 96.9% with tenofovir in 5 (1-12) months. In the patients with a biochemical response, the response was achieved within the first 12 months of treatment in 95.7%, 86.4%, and 100% for lamivudine, entecavir, and tenofovir patients, respectively.

 

In HBeAg-positive patients, 3 treated with lamivudine, 2 with entecavir and 4 with tenofovir had serologic response (P=0.941). None of the patients experienced primary nonresponse.

 

PVR rates at 24 weeks were similar among patients treated with lamivudine (16, 33.3%), entecavir (7, 35.0%) and tenofovir (12, 32.4%) (P=0.981). HBV DNA was detectable in 18.2% patients treated with lamivudine, 18.8% with entecavir and 13.6% with tenofovir, respectively after 48 weeks (P=0.788).

 

The baseline median HBV DNA level in patients with PVR was significantly higher among lamivudine treated (6.3 vs 5.1 lg IU/mL, P=0.003) and tenofovir treated patients (7.6 vs 6.6 lg IU/mL, P=0.017) compared with those with VR at 24 weeks, whereas HBV DNA was slightly higher in entecavir treated patients (7.2 vs 6.3 lg IU/mL, P=0.088). Patients with higher baseline viral load (HBV DNA level ≥7 lg IU/mL) or with baseline HBeAg-positive status were more difficult to achieve VR by 24 and 48 weeks of treatment (Fig. 1).

 

VBR was significantly more common in patients treated with lamivudine than in those treated with other two NAs. Eighteen (30.5%) patients treated with lamivudine developed VBR after a median follow-up of 22 months (11-58); 1 (3.6%) in entecavir group developed VBR at 60 months and none of the patients treated with tenofovir experienced VBR. Patients with VBR were switched to tenofovir as the appropriate rescue therapy. HBeAg did not affect VBR occurrence in patients treated with lamivudine (30.8% in HBeAg-negative and 28.6% in HBeAg-positive, P=0.639). Kaplan-Meier estimates for the time to first detection of VBR for patients treated with lamivudine were 9%, 33%, 41%, 53% and 67% at 1, 2, 3, 4 and 5 years, respectively.

 

Of the 18 patients with VBR in the lamivudine group, 4 experienced BBR between 0-9 months after onset of VBR. Hepatitis flare and decompensation of liver disease did not occur in any patient. BBR was not observed in the patient with VBR in the entecavir group.

 

Univariate and multivariate Cox regression analysis identified that PVR was significantly associated with VBR in patients treated with lamivudine; patients with PVR at 6 months were 3.09 (95% CI: 1.09-8.74; P=0.034) times more likely to develop VBR than those with VR. Also a tendency to develop VBR was observed in patients older than 50 years of age (HR: 2.80; 95% CI: 0.99-8.18; P=0.052). The variables and their significances in this analysis are presented in Table 2.

 

When patients treated with lamivudine were stratified according to their age and presence of PVR, 6/9 (66.7%) of patients with both PVR and ≥50 years developed VBR, 9/19 (47.4%) of the patients with either PVR or ≥50 years developed VBR, while only 2/13 (15.4%) had VBR in patients without PVR and <50 years (P=0.043), during their median follow-up period of 15.0 (9-52), 17.5 (6-58) and 18.0 (6-147) months, respectively. Corresponding cumulative probabilities of VBR for the three groups at 48, 96 and 144 weeks were 67%, 67%, 83%; 31%, 54%, 54%; and 10%, 26%, 26%, respectively (Fig. 2).

 

Only one of the seven patients among entecavir partial responders, after achieving VR at 24 months, developed VBR at 60 months. Genetic analysis showed that this 39-year-old man had resistant mutations and rtM204V+L180M+S202G. He was a HBeAg-negative, non-cirrhotic patient with a baseline HBV DNA level of 7.8 lg IU/mL and had a detectable HBV DNA level of 69 100 IU/mL at 24 weeks, and 5200 IU/mL at 48 weeks. Another entecavir partial responder, a 42-year-old male, HBeAg-positive, non-cirrhotic patient with a baseline HBV DNA level of 8.8 lg IU/mL, had a detectable HBV DNA level of 46 000 IU/mL at 24 weeks, and 31 900 IU/mL at 48 weeks; he showed a continuous, but slowly declining viral kinetic with a still detectable HBV DNA level of 299 IU/mL at the end of his 49-month follow-up. He showed a 4.37 lg IU/mL HBV DNA decline at the first year of therapy, and afterwards an additional 0.87 lg, 0.35 lg and 0.80 lg IU/mL at his second, third and forth years of therapy, respectively, and neither achieved VR, nor developed VBR. No genotypic resistance to entecavir was detected in this patient. Both patients were checked about medication compliance. Among other 5 entecavir partial responders of 24 weeks, 4 achieved undetectable HBV DNA between 24-48 weeks, and one achieved undetectable HBV DNA at 61 weeks without evidence of VBR throughout their 49 (8-61) months lasting follow-up. Those 5 entecavir partial responders achieving VR had a lower viral load at 24 weeks (103-1193 IU/mL) than the previous 2 partial responders.

 

None of 12 tenofovir patients with detectable HBV DNA at 24 or 48 weeks had evidence for a VBR throughout their 17.5 (8-48) months follow-up. Seven achieved VR between 24-48 weeks and the other 4 at second year of their follow-up and one patient's follow-up ended at 24 weeks. Tenofovir partial responders had a viral load of 82-5 547 670 IU/mL at 24 weeks.

 

The serum HBV DNA level reduced from baseline to 12, 24, 48 and 72 weeks was greater in the tenofovir group than in the lamivudine group. The HBV DNA levels as lg IU/mL for tenofovir vs lamivudine were as follows: 4.00 vs 3.40, P=0.052; 4.97 vs 4.09, P=0.006; 5.64 vs 3.67, P=0.000; 5.72 vs 3.97, P=0.001, respectively. The corresponding HBV DNA levels reduced for entecavir vs lamivudine were: 3.57 vs 3.40, P=1.000; 4.48 vs 4.09, P=0.731; 4.75 vs 3.67, P=0.014; and 4.99 vs 3.97, P=0.062. The change in HBV DNA levels in the entecavir group was greater and tended to be greater than that in the lamivudine group at 48 and 72 weeks, respectively (Table 3 and Fig. 3).

 

The HBV DNA levels as lg IU/mL reduced for tenofovir vs entecavir were 4.00 vs 3.57, P=0.292; 4.97 vs 4.48, P=0.458; 5.64 vs 4.75, P=0.066; and 5.72 vs 4.99, P=0.378 (Table 3 and Fig. 3).

 

 

The present study assessed the PVR rates and treatment outcomes of partial responders of lamivudine, entecavir and tenofovir in our routine clinical practice. The clinical relevance of PVR is whether these patients are at risk for developing resistance, and whether adaptation of treatment can achieve sustained viral suppression.

 

Because primary nonresponse is a rare situation, we concentrated the PVR and VR rates in patients treated with monotherapy of lamivudine, entecavir or tenofovir. The rates of HBV DNA detectability for HBeAg-negative patients were 29% for lamivudine, 7.2% for entecavir, and 15% for tenofovir at 24 weeks, but 27%-28% for lamivudine, 6.5%-11% for entecavir, 7% for tenofovir at 48 weeks (or 52 weeks in some studies).^{[18, 26-29]} The rates for HBeAg-positive patients were 68% for lamivudine, 23.3% for entecavir, and 51% for tenofovir at 24 weeks, but 60% for lamivudine, 14.8%-33% for entecavir, and 15%-24% for tenofovir at 48 weeks.^{[18, 28, 30-32]}

 

In our study, the PVR rates of the HBeAg-negative patients at 24 weeks for lamivudine, entecavir and tenofovir were very similar. Whereas the PVR rates in HBeAg-negative patients were comparable to HBV DNA detectability rates of previous studies. The rates of HBeAg-positive patients seemed to be overestimated because of the relatively low number of the patients. There were marked differences in the PVR rates between the patients with baseline HBeAg-positive and negative status, and with a higher and lower baseline viral load for each group of three NAs, and both for 24 and 48 weeks.

 

Multivariate regression analyses identified PVR at 24 weeks as a significant, independent risk factor to develop VBR for lamivudine patients in this study and therefore, these patients required adaptation of antiviral therapy. Since age is a risk factor for developing VBR, patients older than 50 years are identified as a population warranting for closer follow-up during the lamivudine treatment. Therefore, early modification to drugs with different resistance profiles, such as tenofovir, should be offered when a suboptimal response at 24 weeks is detected, considering the unacceptably high VBR probability. Initiating therapy with lamivudine in patients aged <50 years and it was continued according to the PCR negativity at 6 months of therapy, which may result in a satisfactory long-term outcome and also may be a cost benefit.

 

In the present study, the relationship between residual viremia at 24 weeks and poor subsequent therapeutic outcome for patients treated with lamivudine was consistent with that reported elsewhere.^{[11, 18-22]} This finding supports the recommendation of the current EASL guideline to use a more potent drug in lamivudine patients with PVR at 24 weeks.^[23] Since the risk of developing lamivudine-resistance directly correlates with a slow response, it seems that still ongoing viral replication at 24 weeks facilitates the emergence of lamivudine mutated strains. The development of resistance during the treatment of CHB with NAs is associated with adverse treatment outcome.^{[13, 33]} The modification of treatment regimen according to the on-treatment response at this time point would achieve more favorable long-term outcomes while minimizing the treatment failure among patients receiving lamivudine therapy. The high levels of residual viremia are considered to be associated with a higher risk. In a study,^[11] patients with HBV DNA levels of more than 10³ copies/mL at 6 months of therapy had a chance of 63.2% for the development of YMDD variants within 29 months, in contrast to a chance of 13% for those patients with HBV DNA levels of less than 10³ copies/mL. Also, elderly patients seemed to have a tendency for higher VBR rates for lamivudine treatment in our study. There were no clear-cut data on age as a pretreatment predictor of VBR. In another study,^[34] lamivudine resistance was no more prevalent in patients of more than 60 years old. A similar study investigated the determinants of emergence of lamivudine resistant mutations and found that age >50 years is associated with the emergence of YMDD mutations only in the univariate model.^[35]

 

The predictive values of lamivudine for other baseline and on-treatment factors have been also studied in our and previous studies. Baseline viral load, HBeAg status, presence of cirrhosis, gender, and high ALT level were not used to predict the VBR in our cohort, although some of them showed a significant role in different series. The HBV DNA level before treatment was significantly effective in univariate analysis of four lamivudine-treatment series from Japan, China, Australia and Bangladesh although the on-treatment HBV DNA level was considered as the most significant predictor for resistance or VBR in these studies.^{[22, 36-38]} The last series also reported failure to convert HBeAg to negative at 12 months as another on-treatment predictor.^[38] The pretreatment predictors including gender, high baseline ALT, and HBeAg positivity which were reported as either significant in univariate analysis or in multivariate analysis were totally insignificant in the mentioned series, indicating the inconsistency of these factors which may be related to the characteristics of the patients. The predictive value of the on-treatment predictor HBV DNA response in many studies emphasizes the importance of monitoring patients on lamivudine therapy.

 

In choosing which antiviral agent is used as the first line therapy, baseline and on-treatment predictors of outcome should be applied clinically in decision-making. Our analysis showed that the outcomes of treatment with lamivudine differed significantly according to these predictors. Several medical societies^{[23, 39]} around the world recommend the withdrawal of lamivudine, and favor the use of highly potent drugs with a high genetic barrier to resistance as the first choice. Despite this, lamivudine is still a widely prescribed first line drug in many high endemic areas of the world, with low-income.^[17] Identifying the predictors of therapeutic outcomes and comparing the clinical and cost utility of different therapeutic strategies become critical when guiding to prioritize the use of one drug among pharmacological alternatives. It is an important issue to translate clinical outcomes into economic value for many low-income countries; therefore, taking into account these predictors as well as the availability and cost effectiveness of antiviral agents is needed.

 

In our study, the change of the level of HBV DNA was characterized by a rapid and profound decrease of 3.40-4.00 lg IU/mL with all three NAs at 12 weeks, and a relatively slow decrease afterwards (Fig. 3). Compared with lamivudine, tenofovir showed a more potent HBV DNA suppression at 12-72 weeks, and also showed a greater but not significant suppression at 12-72 weeks than entecavir. The change of HBV DNA levels in the entecavir group was similar to that in the lamivudine group at 12 and 24 weeks, but entecavir resulted in a greater reduction of the HBV DNA levels at 48 and 72 weeks than did lamivudine (Table 3).

 

In our study, the PVR rate at 24 weeks was observed with entecavir and tenofovir with a similar proportion as in the lamivudine group. The high PVR rates similar to those of lamivudine do not reflect the low potency or facilitate the selection of resistant strains for the two drugs, rather reflect the longer time for viral clearance in our entecavir and tenofovir patients with higher baseline HBV DNA levels. The tenofovir and entecavir group consisted of more patients than lamivudine patients with a HBV DNA level of ≥7 lg IU/mL and these patients had more residual viremia at 24 and 48 weeks. The high PVR rates for these two NAs in our study can be explained by physician's favor to use drugs known as a higher genetic barrier and potency for patients with a higher baseline viral load. Similar trends with a baseline HBV DNA level and VR have been observed in two studies.^{[40, 41]} In one study, treatment-naive CHB patients received entecavir for 3 years. All patients with baseline HBV DNA <8 lg copies/mL achieved VR, whereas 75% of the patients with baseline HBV DNA ≥8 lg copies/mL did not; but it was possible that complete viral suppression might have been attained in more patients with a longer follow-up.^[40] In another study, time course for VR with tenofovir therapy was longer among patients with baseline HBV DNA ≥9 lg copies/mL, but at 96 weeks, the percentage of patients with VR was similar, and by the end of 240 weeks, persistent viremia or tenofovir resistance was not observed in any patient.^[41]

 

In our study, the majority of tenofovir and entecavir treated patients who did not accomplish PCR negativity at 24 or 48 weeks finally achieved VR; therefore, slow responder seems to fit better instead of partial responder for these patients. In one study including the entecavir patients from six phase �� and �� clinical trials, the cumulative probability of VBR in NA-naive patients was only 0.8% at 5 years, even though a substantial number of patients achieved VR beyond the first year of treatment.^[42] Another study concluded that adaptation was not necessary in a majority of NA-naive patients with detectable HBV DNA at 48 weeks, because long-term use of entecavir led to VR in 81% after a median follow-up of 19 months; particularly those with HBV DNA <1000 IU/mL at 48 weeks had a higher cumulated probability of VR compared to those with HBV DNA ≥1000 IU/mL (95% vs 57%).^[27] Among the 7 patients without a VR, one experienced VBR without genotypic resistance, and the other 6 had a declined viral load at the end of the study.^[27]

 

Among our two entecavir partial responders, one could not achieve VR, and the other experienced VBR after VR. A high residual level of HBV replication was noted at both 24 and 48 weeks compared with the other entecavir partial responders who achieved and maintained VR throughout the follow-up. In two different case reports,^{[43, 44]} genotypic evolution of entecavir-resistant mutants were described in two NA-naive patients, i.e. one patient had a level of 3.30 lg IU/mL at 24 weeks remaining as high as 4.21 lg IU/mL at 120 weeks, and the other had a level of 4 lg copies/mL between 24-124 weeks of therapy. Another study^[30] reported one patient who developed genotypic resistance among 19 naive CHB patients with detectable HBV DNA at 1 year under entecavir therapy. These data may suggest that even with a high genetic barrier NA, patients with a prolonged high level of residual viremia may face the risk of developing resistance. There is limited evidence in terms of the time point and the level of viremia which is associated with a significant risk that would justify a therapeutic decision to adapt the therapeutic regimen.

 

One entecavir partial responder showed a continuous but slowly declining viral kinetics within 49 months. No genotypic resistance was detected in this patient. In a relevant study,^[45] patients whose serum HBV DNA levels remained high after long-term entecavir treatment. They were defined as slow-responders with HBV DNA levels ≥3.0 lg copies/mL after 2 years. 41.7% of HBeAg-positive and none of the HBeAg-negative patients were defined as slow responders, and no entecavir-resistant mutants were detected in these patients. Pretreatment low baseline HBV DNA levels, low quantitative HBsAg (qHBsAg) levels and HBeAg-negative status were predictive of HBV DNA-negative status at year 2, while multivariate analysis revealed the low qHBsAg level as a significant predictive factor (P=0.03). Higher baseline viral load may not be the only explanation of PVR to highly potent NAs in naive patients. Exact mechanisms and predictors need further study.^[45]

 

Our study has some limitations. Our cohort consisted of a heterogeneous population who were observed retrospectively and not randomized, so the results differed from the registration trials of a controlled environment. Nevertheless, the results might be representative of routine clinical practice, and also the observational design made it possible to compare three NA groups within one study. Although, we did not perform a genotype analysis, many published studies have reported that almost all patients chronically infected with HBV in Turkey have genotype D, constituting 100% of HBV infected patients.^[46-50]

 

In conclusion, the rise in rates of viral suppression leading to a VR over time with minimal risk of VBR, in accordance with the low rates of genotypic resistance in registration trials,^{[26, 28, 41, 51]} suggests that a satisfactory response can be attained without a modification during prolonged treatment of the majority of patients with PVR to entecavir or tenofovir. Since evidence is not enough to drawn a conclusion on the relationship between PVR and subsequent VBR in lamivudine treatment, the rationale for the decision to modify the treatment regimen for low potent drugs should not be directly translated to a high genetic barrier to resistance drugs. Serum qHBsAg levels, altered pharmacodynamic or pharmacokinetic characteristics of drugs like cell entry, hepatocyte phosphorylation to active forms, and bioavailability may be associated with flat HBV DNA kinetics causing a "partial" or "slow" response pattern to more potent NAs such as entecavir and tenofovir, so HBV DNA level at some point of the therapy may not be an independent predictor for subsequent failure. Furthermore, these factors may influence not only on the response to the initially started antiviral drug, but also on that of the salvage regimen. Hence treatment modification may not provide the expected benefit. Further investigation is needed to solve these issues and to propose better antiviral algorithms.

 

 

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