Carey et al1 recently proposed that HBV pregenomic RNA (pgRNA) is a sensitive indicator of covalently closed circular DNA (cccDNA) transcription status in patients receiving nucleos(t)ide analogue therapy (NA) for hepatitis B. pgRNA is transcribed from the cccDNA minichromosome and therefore reflects a product of cccDNA transcriptional activity and its prevalence within HBV-infected cells. They observed that some patients reached pgRNA negativity 12 months of NA, suggesting a surprisingly rapid decline in transcriptional activity/prevalence of cccDNA. Moreover, studies by Bazinet et al2 and Liu et al3 found that pgRNA and total HBV RNA (totRNA) declined from pre-treatment levels as early as 6 months and 3 months of NA, respectively. In both these studies, pgRNA/totRNA decline lagged behind the HBV DNA (virus) decline, in agreement with the findings in1. Interestingly, Liu et al3 observed a biphasic decline in both totRNA and virus in some rapid responder patients, which is surprising since NA could only be expected to cause similar rates of totRNA and virus decline if an antiviral activity in addition to reverse-transcriptase inhibition was present (Figure).
Figure. Established and potential antiviral effects of nucleos(t)ide analogue (NA) in HBV infection.
Previously described inhibition of reverse transcription by the HBV reverse transcriptase (RT) provides a mechanism for inhibiting secretion of HBV DNA (red) and recycling of HBV DNA (purple) consistent with reduction of HBV DNA but not HBV pregenomic RNA (pgRNA) in the blood. Potential mechanisms consistent with reduction of HBV pgRNA in the blood include transcriptional inactivation / degradation of covalently closed circular DNA (cccDNA) (1), inhibition of RT-binding to pgRNA (2) potentially required for the encapsidation of pgRNA (3). MVB, multivesicular body.
Biphasic virus decline during NA typically consists of a rapid-first phase followed by a slower-second phase. Mathematical modeling suggests this is a result of NA blocking virus production with the first phase driven by its t1/2 in blood. The second phase is driven by the rate of HBV-infected cell loss/death4. Assuming similar serum t1/2 for pgRNA/totRNA and virus, the first phase observed by Liu et al3 might suggest that NAs immediately inhibit pgRNA/totRNA production by novel mechanisms whereas the second phase could reflect inhibition of pgRNA/totRNA and possibly cccDNA loss and/or HBV-infected cell loss/death (Figure).
While persistence of pgRNA is an indicator of ongoing cccDNA transcription in patients with undetectable serum viral levels and quantifiable pgRNA is associated with virus rebound and alanine-aminotransferase flare when NA is discontinued1, undetectable pgRNA does not necessarily indicate cccDNA clearance. Future studies with very early and frequent pgRNA/totRNA and viral sampling will be required to characterize pgRNA/totRNA kinetics and perform theoretical modeling in order to elucidate pgRNA/totRNA dynamics during infection and NA and whether NA promote cccDNA clearance.
ACKNOWLEDGMENTS
The research was supported in part by the U.S. National Institute of Health (NIH) grants R01-AI144112 and R01-AI146917.
References
- 1.Carey I, Gersch J, Wang B, et al. Pregenomic HBV RNA and Hepatitis B Core-Related Antigen Predict Outcomes in Hepatitis B e Antigen-Negative Chronic Hepatitis B Patients Suppressed on Nucleos(T)ide Analogue Therapy. Hepatology 2020; 72(1): 42–57. [DOI] [PubMed] [Google Scholar]
- 2.Bazient M, Anderson M, Pantea V, et al. Analysis of HBsAg levels, HBsAg isoforms, HBsAg immune complexes, HBV pregenomic RNA and HBcrAg dynamics during and after NAP-based combination therapy in the REP 301-LTF and REP 401 studies. J Hepatology 2020; (73): S142. [Google Scholar]
- 3.Liu S, Liu Z, Li W, et al. Factors associated with the biphasic kinetics of serum HBV RNA in patients with HBeAg-positive chronic hepatitis B treated with nucleos(t)ide analogues. Aliment Pharmacol Ther 2020; 52(4): 692–700. [DOI] [PubMed] [Google Scholar]
- 4.Dahari H, Shudo E, Ribeiro RM, Perelson AS. Modeling complex decay profiles of hepatitis B virus during antiviral therapy. Hepatology 2009; 49(1): 32–8. [DOI] [PMC free article] [PubMed] [Google Scholar]