Letter: impact of HBV genotypes and PC/BCP mutations on serum HBsAg levels in Chinese HBeAg negative patients—Authors′ reply

Abstract

EDITORS,In their letter, 1 Chen and colleagues pointed out that the patients in our study cohort were predominantly infected with HBV genotype A and D, whereas genotypes B and C were found less frequently. 2 They concluded that this distribution was most likely due to the European origin of our study population and that our results have to be examined in larger study populations with a higher prevalence of HBV genotypes B and C. We agree with this conclusion and we are excited to learn more on this topic in patients with genotype B and C infection as this group was naturally underrepresented in our study of patients living in Europe. Chen and colleagues presented data of 106 HBeAg-negative patients infected with HBV genotypes B and C "without anti-viral therapy before." The authors observed no differences in the HBsAg levels among these genotypes. As they also found no differences in earlier studies analysing patients with nucleos(t)ide analogues therapy, they concluded that genotypes B and C are not associated with differences in HBsAg levels. However, for meaningful interpretation of the genotype-specific impact on HBsAg levels the analysed patient group has to be described more precisely as HBsAg levels also depend strongly on the disease stage. [3][4][5] In the cited studies, 6,7 patients with presumable active hepatitis were included as they qualified for therapy. Both HBeAg-positive (n = 32 and 61, respectively) and HBeAg-negative patients (n = 16 and 33, respectively) were analysed and the overall patient numbers, especially for HBeAg-negative patients, were small. In their letter, the patient numbers are also limited (62 genotype C and 44 genotype B infected patients) and no demographic data were given. In detail, it is unclear if "without anti-viral therapy before" means that these patients were all inactive carriers with normal transaminases etc.without need for anti-viral treatment as in our study. Therefore, interpretation of these data seems to be difficult at this point.In addition, Chen et al presented data regarding the prevalence of the basal core promotor (BCP) A1762T/G1764A and precore (PC) mutation G1896A and their impact on HBsAg levels. The authors wrote that none of these mutations was significantly associated with HBsAg levels, although as shown in figure 1D of their study, statistically significantly more HBsAg was found in patients with PC mutation (P = 0.006, exact patient numbers are missing). In contrast, in our study the PC mutation was associated with lower HBsAg levels and this was also true after excluding patients with additional preS mutations from the analysis (3.0 vs 3.3 log IU/mL for patients with and without G1896A, respectively; P = 0.005). However, this effect seems to be mild and the clinical impact still needs to be explored.The observation that the BCP mutation has no impact on HBsAg levels is in line with our results.In conclusion, we agree with the authors that the impact of the genotype and frequent mutations on HBsAg levels has to be confirmed in studies with a high...

Formation of semi‐enveloped particles as a unique feature of a hepatitis B virus PreS1 deletion mutant

“…However, few studies about the effects of N‐terminal deletions in the PreS1 domain on the levels of intra/extracellular HBsAg have been reported. To analyse the impact of N‐terminal PreS1 deletions on intra/extracellular amounts of HBsAg, an isolate lacking aa 25‐39 in the PreS1 domain (p1.5×HBV_preS1_∆aa25‐39) from an HBV chronically infected patient was expressed in Hepatoma cell line Huh7.5. Western blot analysis using the S‐domain‐specific monoclonal HBO1 of cellular lysate and supernatant from cells transfected with this deletion mutant showed a shift of the LHBs signal to a lower molecular weight compared to the control cells expressing the intact genome (Figure ).…”

“…Plasmid HBV/A carrying 1.5‐fold autologous promoter‐driven genomes of genotype A2 (p1.5×HBV) was used as a reference control. The plasmid HBV PreS1 deletion mutant (p1.5×HBV_preS1_Δaa25‐39) harbours a 1.5‐fold HBV genome genotype A with 15 aa (25FPDHQLDPAFGANSN39) deletion in the N‐terminus of PreS1 domain . The rescued mutants p1.5×HBV_preS1_(+)aa25‐31, p1.5×HBV_preS1_(+)aa32‐39 and p1.5×HBV_preS1_(+)aa25‐39 were generated by overlapping PCR using the following primer: 5′‐tct gtt ccc aac cct ctg gga ttc ttt ccc gat cat cag ttg gac , 5′‐tgg ggt tga agt ccc aat ctg gat t gt cca act gat gat cgg gaa , 5′‐tct gtt ccc aac cct ctg gga ttc cct gca ttc gga gcc aac tca a , 5′‐tgg ggt tga agt ccc aat ctg gat t gt ttg agt tgg ctc cga atg c , 5′‐tct gtt ccc aac cct ctg gga ttc ttt ccc gat cat cag ttg gac cct gca ttc gg , 5′‐tgg ggt tga agt ccc aat ctg gat t gt ttg agt tgg ctc cga atg cag ggt cca ac .…”

“…33,56 N-terminal myristylated peptides corresponding to this epitope specifically prevent HBV infection in permissive cells. 33,56 The missing 15 aa (aa [25][26][27][28][29][30][31][32][33][34][35][36][37][38][39] in the mutant locate in the HBV receptor binding domain and partially overlap with the essential receptor binding residue aa 20-29. 56 Since the essential HBV receptor binding domain is destroyed, it can be assumed that mutant viral particles (∆aa25-39) do not specifically bind to the receptor/complex on hepatocytes and thus fail to establish an infection.…”

“…Recently, an HBV mutant with 15 aa deletion (aa 25‐39) in the N‐terminus of PreS1 domain was isolated based on the analysis of a large European cohort of HBeAg‐negative patients suffering from chronic HBV infection . In this study, we analysed the effects of the N‐terminal deletion in the PreS1 domain on the HBsAg synthesis and release, subcellular distribution, as well as the viral and subviral morphology.…”

“…14,[21][22][23] Many studies showed that these genetic defects are usually caused by in-frame deletions in the C-terminal PreS1 domain or the middle portion of the PreS2 domain or point mutation(s) in the start codon of the PreS2 domain. [14][15][16]18,22,24 Recently, an HBV mutant with 15 aa deletion (aa [25][26][27][28][29][30][31][32][33][34][35][36][37][38][39] in the N-terminus of PreS1 domain was isolated based on the analysis of a large European cohort of HBeAg-negative patients suffering from chronic HBV infection. 25 In this study, we analysed the effects of the N-terminal deletion in the PreS1 domain on the HBsAg synthesis and release, subcellular distribution, as well as the viral and subviral morphology.…”