Perspective on Emerging Therapies to Achieve Functional Cure of Chronic Hepatitis B

Despite the availability of a safe and effective vaccine for over four decades, the global health burden of hepatitis B virus (HBV) infection remains high. Recent estimates suggest there are ~ 300 million people with chronic infection resulting in 820,000 deaths annually, primarily due to complications of cirrhosis and hepatocellular carcinoma (HCC) [1]. Additionally, there are approximately 1.5 million incident infections annually. Acknowledging this substantial health burden, the World Health Organization (WHO) has set an ambitious goal of reducing HBV incidence by 90% and HBV-related mortality by 65% by 2030 compared to 2015 levels [2].

Of the two approved classes of therapy for chronic hepatitis B (CHB), peginterferon alfa (Peg-IFN) and nucleos(t)ide analogs (NAs), NAs are the mainstay of treatment. NAs effectively suppress viral replication, improve hepatic inflammation, and reduce progression to cirrhosis and HCC [3]. However, only a small percentage of patients (≤ 1%) achieve the desired endpoint of functional cure, defined as the loss of hepatitis B surface antigen (HBsAg) and HBV DNA below the level of quantitation, that permits safe discontinuation of therapy [4, 5•]. Consequently, most patients require long-term therapy. This is because NAs do not directly target the two sources of HBsAg in serum-covalently closed circular DNA (cccDNA), a non-integrated form of the viral DNA located within the hepatocyte nucleus that serves as the template for viral transcription, and integrated HBV DNA [6, 7]. Although NAs have an excellent safety profile [8], long-term use is associated with increased healthcare costs, medication noncompliance, and risk of renal and bone toxicity. Therefore, there is a need for a novel, finite duration therapy that can achieve high rates of functional cure [9•].

This review focuses on novel therapies in development that hold promise for achieving functional cure. Advancements in therapeutics in combination with more widespread use of HBV vaccination will be crucial for reducing the global burden of HBV infection and achieving the WHO’s goal of HBV eradication.

HBV is a small hepatotropic DNA virus with a partially double-stranded circular genome of approximately 3.2 kilobases. The infectious virion or Dane particle consists of an outer lipid envelope made up of large, middle, and small HBsAg and an inner icosahedral viral nucleocapsid, consisting of core protein monomers with the viral DNA and HBV polymerase [10]. The viral life cycle and druggable targets are shown in Fig. 1.

The ideal endpoint of treatment for HBV is a sterilizing cure. This is defined as the elimination of all traces of the virus including cccDNA and integrated HBV DNA [11]. However, achieving a sterilizing cure for HBV is currently not attainable. Therefore, the current focus of HBV treatment is aimed at obtaining a functional cure defined as sustained HBsAg loss for 24 weeks off-treatment, with or without seroconversion to anti-HBs, and HBV DNA below the limit of quantitation [12, 13]. A functional cure is associated with the reduction in clinical outcomes (cirrhosis, decompensated liver disease, and HCC), is durable off-treatment, and is easy to assess [14•, 15]. However, recent studies revealing that HBsAg can originate from both cccDNA and integrated HBV DNA highlight the complexity of achieving a functional cure [16, 17, 18••].

Given the challenges of achieving a functional cure, some have advocated for using a more achievable endpoint, partial cure, defined as HBsAg < 100 IU/mL, negative HBeAg status, and persistent HBV DNA levels below the lower limit of detection (LLOD) after 24 weeks off treatment [12]. Although a partial cure is also associated with the improvement in clinical outcomes, it is not a durable off-treatment endpoint. Additionally, it does not result in an HBsAg loss which is important for removing stigma for many patients. Thus, a partial cure is considered an intermediate endpoint, and a functional cure continues to be the preferred endpoint of novel therapies in the development for CHB [12].

As shown in Table 1, novel therapies in development for CHB can be categorized into two main groups: those that target key steps in the viral lifecycle, referred to as direct acting antiviral agents, and those that target the immune response, referred to as immunomodulators.

Targeting the viral entry can block the infection of uninfected hepatocytes and thereby prevent the replenishment of the cccDNA pool. Bulevirtide, a small synthetic myristoylated lipopeptide mimicking the pre-S1 sequence of the large S protein, is an entry inhibitor which blocks HBV entry into hepatocytes by binding to the sodium taurocholate co-transporting polypeptide (NTCP) receptor [37]. Although there is evidence on the safety and efficacy of bulevirtide among subjects with chronic hepatitis D virus infection, there is only a limited data among patients with HBV mono-infection [19]. Bulevirtide has minimal effect on HBsAg levels, suggesting that its effectiveness in achieving a functional cure over a short dosing period may be limited. Hepalatide is another NTCP receptor blocker currently under evaluation [22].

Monoclonal and polyclonal antibodies that specifically bind to the N-terminal region of pre-S1 and block HBV attachment to hepatocytes are under development [38]. VIR-3434 is a monoclonal antibody designed to block the HBV entry into hepatocytes and reduce the level of virions and subviral particles. It may also function as a T cell vaccine against HBV. In a proof-of-principle study, VIR-3434 demonstrated a rapid HBsAg decline of ≥ 1 log IU/mL in virally suppressed subjects [21]. Studies in combination with pegIFN with or without a small interfering RNA (siRNA) and NA are ongoing [39]. Additionally, GC1102, a recombinant hepatitis B immunoglobulin, showed HBsAg loss in 2 (22%) out of 9 participants with low pretreatment HBsAg levels [22].

Targeting HBV mRNA using siRNA or antisense oligonucleotides (ASO) are promising approaches to lower viral antigen production and indirectly inhibit HBV replication. siRNAs are short, double-stranded RNAs that lead to gene silencing in a sequence-specific manner by utilizing the endogenous RNA interference pathway. A single siRNA can target multiple HBV mRNAs due to shared terminal 3′ sequences in all HBV mRNAs. siRNAs require a carrier vehicle for their uptake as they do not readily cross the vascular endothelium. ASOs are single-stranded DNA or RNA sequences that bind to complementary sequences of mRNAs and degrade the mRNAs via a ribonuclease H–dependent mechanism. ASOs have the advantage of targeting pre-mRNA, enhancing their specificity and reducing off-target effects without the need for carrier vehicles [40].

In chronic HBV infection, patients often have impaired innate and adaptive immune responses. High levels of viral antigens, such as HBsAg, can lead to the exhaustion of HBV-specific T cells, further compromising the immune response [74]. To achieve long-term control of HBV infection, it is crucial to restore immune function.

In chronic HBV infection, the innate immune system has difficulty sensing and effectively responding to the HBV. Additionally, HBV may actively evade the innate immune response leading some to consider it a stealth virus [74]. A number of therapeutic approaches are being developed to restore the innate immune response. These include Toll-like receptor (TLRs) agonists, retinoid acid–inducible gene-I (RIG-I), and stimulator of interferon genes (STING), which aim to induce the production of interferons and proinflammatory cytokines that can help clear HBV [75].

TLRs play a critical role in the early innate immune response by sensing highly conserved bacterial or viral products such as LPS, bacterial DNA, and viral double-stranded RNA. The stimulation of TLRs results in a variety of cellular responses that lead to the production of IFNs, pro-inflammatory cytokines, and effector cytokines that eliminate pathogens. The TLR-7 agonists vesatolimod (GS-9620), RO7020531 and JNJ-4964, and TLR-8 agonist selgantolimod (GS-9688) [34, 76] have shown promise in preclinical trials by activating intrahepatic dendritic cells and triggering the production of interferons, as well as activate natural killer (NK) cells and T cells [77, 78]. However, clinical trials with vesatolimod and selgantolimod did not show substantial benefit in reducing HBsAg levels [34, 76]. In a phase 2 study, selgantolimod led to HBsAg loss in 5% of virally suppressed subjects and a modest reduction in HBsAg levels [34].

Inarigivir (SB9200), an oral agonist of RIG-I and NOD2 that induces interferon-mediated antiviral immune responses in virus-infected cells, is no longer being developed due to hepatotoxicity and death of a research subject [79], 80.

Further research is needed to define the optimal innate immune targets, and to establish the role of agonists of innate immunity in combination with antiviral therapy. Moreover, understanding the timing and duration of immune modulation in relation to viral inhibition will be important for achieving a functional cure.

Restoring an adequate adaptive immune response will likely be necessary for controlling and potentially curing HBV infection.

Checkpoint inhibition, using antibodies targeting the programmed death receptor (PD-1) or programmed death ligand (PD-L1), is one non-specific approach to restore T cell function. PD-1 and its ligands downregulate T cell activation. PD-1 has been shown to be overexpressed by HBV-specific T cells. Thus, agents that block PD-1 and its ligand checkpoint inhibitors represent a therapeutic strategy to restore function of HBV-specific T cells and enhancing antibody production [81]. Proof-of-concept has been demonstrated in cancer treatment. There is, however, a concern for autoimmunity, and an uncontrolled immune response may lead to flares of hepatitis. In a pilot study, the use of PD-1 inhibitor nivolumab with or without GS-4774, a therapeutic vaccine in virally suppressed subjects, resulted in only modest declines in HBsAg levels. HBsAg seroclearance was achieved in one patient in the nivolumab-only group [82]. In another phase 2 study using the PD-L1 antibody envafolimab (ASC22), 3 out of 7 subjects (43%) with baseline HBsAg levels below 100 IU/mL achieved HBsAg loss [35]. Oral, liver-specific checkpoint inhibitors are in development. The optimal duration and sequence of checkpoint inhibition in a therapeutic regimen remain to be determined.

Strategies to restore T cell responsiveness include creation of genetically engineered T cells. T cell receptors cloned from HBV-reactive cells can be introduced into T cells. Alternatively, T cells can be modified to express chimeric antigen receptors that specifically target HBV antigens on the hepatocyte surface [83]. These genetically modified T cells have been shown to effectively eliminate HBV-infected cells in vitro and have shown safety in animal models [84, 85]. The safety and efficacy of using genetically engineered T cells were shown in a small study in which adoptive transfer of autologous T cells expressing HBV-specific T cell receptors was achieved in participants with HBV-related HCC [86]. This may open the possibility of using this approach in patients with CHB.

Therapeutic vaccination aims to induce both B and T cell immune responses, as the resolution of acute HBV infection is associated with lifelong immunity mediated by these components of the immune system. Previous attempts to develop a therapeutic vaccine using a variety of antigens, protein-based, protein-antibody complex–based, and DNA-based vaccines, have been unsuccessful [87]. Novel vaccination strategies are exploring use of viral recombinant vectors, dendritic cell vaccines, and mRNA vaccines. Viral vector-based vaccines, in particular, offer the advantage of intracellular antigen expression, triggering a robust cytotoxic T lymphocyte response [88].

TG1050, an adenovirus 5–based vaccine expressing HBV antigens, and lentiviral vectors encoding HBV antigens, have demonstrated the induction of specific T cell responses but only modest declines in HBsAg levels [89]. Studies involving GS-4774, a yeast-based vaccine expressing HBsAg, HBcAg, and HBX, have shown minimal reduction in serum HBsAg levels, despite robust cytokine production by HBV-specific CD8 T cells [90].

Innovative strategies are being developed to enhance the immunogenicity of peptide-based vaccines. These include the utilization of novel nasal formulations such as NASVAC and derivatives of Sci-B-Vac, like BRII-179 [36, 91]. Clinical trials evaluating the safety and efficacy of these approaches have yielded promising results. The mucoadhesive carboxy-vinyl polymer (CVP-NASVAC) nasal therapeutic vaccine, containing HBsAg and HBcAg, has demonstrated significant reductions in HBsAg levels in participants receiving both NA therapy and HBV carriers. Notably, 9.5% of subjects (6 out of 71) achieved a functional cure [91]. Another study investigating BRII-179, which encompasses all three HBV surface envelope proteins, has shown moderate antibody responses; however, it did not lead to a noticeable reduction in HBsAg levels in virally suppressed subjects under NA therapy [36].

Prime-boost vaccination, involving sequential administration of vaccines with different antigens and delivery systems, is one of the more promising approaches to overcome HBV-specific tolerance and have been shown to generate high levels of memory T cells in preclinical studies. Additionally, combining viral antigenemia knockdown using siRNAs with therapeutic vaccination has resulted in the development of polyfunctional HBV-specific CD8 + T cells and elimination of HBV [92].

An advantage of vaccination is its theoretical safety, as it specifically stimulates HBV-specific immunity without non-specific activation of innate responses or autoimmunity. However, important considerations include determining the optimal vaccine composition, use of T or B cell adjuvants, the vaccine regimen, and the mode of administration need to be addressed in future studies.

Success with combination therapy for other infectious diseases such as those caused by human immunodeficiency virus and hepatitis C virus implies that a similar approach will be needed to achieve a functional cure of chronic HBV infection. Although over 40 compounds with multiple mechanisms of action are currently in development, Table 1, combination approaches are largely empirical due to the lack of knowledge of the mechanisms required to achieve viral control and HBsAg loss. The most promising approaches focus on agents that inhibit viral replication, reduce antigenic burden, and partially enhance/restore immune control against HBV, Fig. 2 [24, 25•, 33, 35, 45]. However, determining the optimal combinations, timing and sequence of use, and treatment duration remains a challenge. Personalized treatment approaches may be necessary based on factors such as HBeAg status, viral load, HBsAg levels, and presence of cirrhosis.

Combination therapy with currently approved agents peg-IFN and NAs can achieve functional cure in ~ 10% of patients. However, many patients decline or are ineligible to receive peg-IFN due to its poor tolerability. Functional cure rates with NAs alone range from 1 to 3%. This highlights the need for newer therapy that can achieve higher rates of a functional cure to reduce the morbidity and mortality associated with chronic HBV infection. Given that the bulk of patients with chronic HBV infection reside in low- and middle-income countries, the ideal regimen would be one that is of finite duration, orally administered, easy to manufacture, and scalable. To realize this goal, continued research, investment, and commitment from both the industry and the scientific community will be crucial.