Single hepatocytes show persistence and transcriptional inactivity of hepatitis B
Ashwin Balagopal, Tanner Grudda, Ruy M. Ribeiro, Yasmeen S. Saad, Hyon S. Hwang, Jeffrey Quinn, Michael Murphy, Kathleen Ward, Richard K. Sterling, Yang Zhang, Alan S. Perelson, Mark S. Sulkowski, William O. Osburn, Chloe L. Thio
Ashwin Balagopal, Tanner Grudda, Ruy M. Ribeiro, Yasmeen S. Saad, Hyon S. Hwang, Jeffrey Quinn, Michael Murphy, Kathleen Ward, Richard K. Sterling, Yang Zhang, Alan S. Perelson, Mark S. Sulkowski, William O. Osburn, Chloe L. Thio
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Research Article Hepatology Infectious disease

Single hepatocytes show persistence and transcriptional inactivity of hepatitis B

Abstract

There is no cure for the more than 270 million people chronically infected with HBV. Nucleos(t)ide analogs (NUCs), the mainstay of anti-HBV treatment, block HBV reverse transcription. NUCs do not eliminate the intranuclear covalently closed circular DNA (cccDNA), from which viral RNAs, including pregenomic RNA (pgRNA), are transcribed. A key gap in designing a cure is understanding how NUCs affect HBV replication and transcription because serum markers yield an incomplete view of intrahepatic HBV. We applied single-cell laser capture microdissection and droplet digital PCR to paired liver biopsies collected from 5 HBV/HIV-coinfected persons who took NUCs over 2–4 years. From biopsy 1 to 2, proportions of HBV-infected hepatocytes declined with adherence to NUC treatment (P < 0.05); we extrapolated that eradication of HBV will take over 10 decades with NUCs in these participants. In individual hepatocytes, pgRNA levels diminished 28- to 73-fold during NUC treatment, corresponding with decreased tissue HBV core antigen staining (P < 0.01). In 4 out of 5 participants, hepatocytes with cccDNA but undetectable pgRNA (transcriptionally inactive) were present, and these were enriched in 3 participants during NUC treatment. Further work to unravel mechanisms of cccDNA transcriptional inactivation may lead to therapies that can achieve this in all hepatocytes, resulting in a functional cure.

Authors

Ashwin Balagopal, Tanner Grudda, Ruy M. Ribeiro, Yasmeen S. Saad, Hyon S. Hwang, Jeffrey Quinn, Michael Murphy, Kathleen Ward, Richard K. Sterling, Yang Zhang, Alan S. Perelson, Mark S. Sulkowski, William O. Osburn, Chloe L. Thio

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Figure 1

HBV eradication from the liver is slow.

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HBV eradication from the liver is slow. (A) The proportion of infected h...
(A) The proportion of infected hepatocytes for each participant at each biopsy is shown. Points indicate when each biopsy was taken; the line indicates the inferred trajectories of the proportion infected, extrapolated from the proportions at each point; 95% confidence intervals are shown in the light shaded portion around each line. (B) The log10(number of infected hepatocytes) was estimated from the proportion of infected hepatocytes that was calculated at each liver biopsy in the 5 participants. The changes in log10(number of infected hepatocytes) for each participant were extrapolated to estimate when the liver could be considered eradicated of all infection. Extrapolated values are shown by dotted lines. For comparison, 90%, 80%, and 50% annual cure rates (i.e., the number of HBV-infected hepatocytes that are cleared each year) are shown as dashed lines. DAART, dually active antiretroviral therapy.