In this study, we identified new compounds that selectively inhibited hepatitis B virus (HBV) entry, and did not impair bile acid uptake. Our evidence offers a new strategy for developing anti-HBV drugs with fewer side effects.

Anti-hepatitis B virus (HBV) drugs are currently limited to nucleos(t)ide analogs (NAs) and interferons. A challenge of drug development is the identification of small molecules thatC hronic hepatitis B virus (HBV) infection, constituting a public health problem, with an estimated 240 million carriers worldwide (1), elevates the risk of development of liver cirrhosis and hepatocellular carcinoma (2). Antiviral agents against HBV include nucleos(t)ide analogs (NAs) and interferons (IFNs), which can achieve significant reductions in HBV loads (3). Although IFN-␣ and its pegylated form (peg-IFN-␣) modulate host immune responses to HBV infection or directly inhibit HBV replication in hepatocytes, these regimens show low tolerability because of serious adverse effects (3, 4). NAs, including lamivudine (LMV), adefovir, entecavir (ETV), tenofovir, and telbivudine, inhibit reverse transcription to suppress HBV replication, but longterm treatment with some of these NAs often results in selection for a significant number of drug-resistant viruses, which decreases treatment efficacy; i.e., the introduction of two substitutions, L180M and M204V, in the polymerase region leads to resistance to LMV, and an additional mutation of either T184, S202, or M250 with the L180M/M204V mutations confers further ETV resistance (5). More notably, it is difficult for the above-mentioned anti-HBV drugs to completely eliminate HBV from infected cells. Future antiviral strategies include multidrug treatment with an existing drug and a new anti-HBV agent. Consequently, there is a

Hepatitis B virus (HBV) constitutes a significant public health burden, and currently available treatment options are not generally curative, necessitating the development of new therapeutics. Here we have applied random non-standard peptide integrated discovery (RaPID) screening to identify small macrocyclic peptide inhibitors of HBV entry that target the cell-surface receptor for HBV, sodium taurocholate cotransporting polypeptide (NTCP). In addition to their anti-HBV activity, these molecules also inhibit cellular entry by the related hepatitis D virus (HDV), and are active against diverse strains of HBV (including clinically relevant nucleos(t)ide analog-resistant and vaccine escaping strains). Importantly, these macrocyclic peptides, in contrast to other NTCP-binding HBV entry inhibitors, exhibited no inhibition of NTCP-mediated bile acid uptake, making them appealing candidates for therapeutic development.