How to achieve a functional cure for chronic hepatitis B infection

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KEY POINTS

• Functional cure of HBV is defined as undetectable HBsAg and unquantifiable HBV DNA 24 weeks after the completion of a finite course of therapy.

• Suppression of HBsAg production from both covalently closed circular DNA and integrated HBV DNA, and the restoration of HBV-specific immune response are necessary to achieve functional cure of HBV.

• HBsAg seroclearance occurs in 3%–5% of patients after 10 years of nucleos(t)ide analog therapy and in 8%–14% of patients 3–5 years after a 48-week course of pegylated interferon.

• Some new antivirals in clinical trials, notably siRNA and antisense oligonucleotide, result in 2–3 log₁₀ decrease in HBsAg levels after 24–48 weeks of treatment but sustained HBsAg seroclearance is uncommon.

• Adding pegylated interferon or immune-modulatory therapy after HBV DNA suppression and HBsAg reduction may enhance the likelihood of functional cure.

INTRODUCTION

Chronic hepatitis B treatment aims to prevent liver-related complications including cirrhosis, hepatic decompensation, and HCC. HBsAg seroclearance is associated with improvement in clinical outcomes, and a low risk of relapse after treatment is discontinued. HBsAg seroclearance confers a further decrease in the risk of HCC compared to HBV DNA suppression and removes the stigma of HBV infection. Thus, sustained off-treatment HBsAg seroclearance with unquantifiable HBV DNA has been chosen as the definition of a functional HBV cure.¹ HBsAg seroclearance rarely occurs with current therapies, 3%–5% after 10 years of continuous nucleos(t)ide analogs (NAs) and 8%–14% after 48 weeks pegylated interferon-alfa (peg-IFN), and 3–5 years posttreatment follow-up.² Challenges to eliminating HBV include the long half-life of covalently closed circular DNA which are not targeted by NAs and can be replenished intracellularly, continued production of HBsAg from integrated HBV DNA even after transcription of covalently closed circular DNA (cccDNA) is suppressed, and HBV-specific immune impairment/exhaustion.

STRATEGIES TO INCREASE HBsAg SEROCLEARANCE WITH CURRENT THERAPIES

While HBsAg seroclearance rarely occurs during NA treatment, retrospective studies and 2 randomized controlled trials showed that HBsAg seroclearance rates were higher in patients who had been on NA for >2–4 years with undetectable HBV DNA and who discontinued NA, with rates up to 13%–39% 5–6 years after NA discontinuation compared to 0%–1% in those who continued NA. However, these encouraging results are not observed in all studies and rates are substantially lower in Asians compared to Caucasians.³ De novo combination of peg-IFN and NA, adding peg-IFN or switching to peg-IFN after HBV DNA had been suppressed by NA has been reported to result in higher HBsAg seroclearance rates in some patients but benefits need to be balanced against the adverse effects of IFN.⁴

NEW THERAPIES AIMED AT HBV CURE

Several new classes of antiviral drugs targeting different steps in HBV lifecycle: viral entry, capsid assembly, viral protein production, and secretion are in clinical development (Figure 1A and Supplemental Materials, http://links.lww.com/XCL/A12).

FIGURE 1.

New HBV therapies and mode of action. (A) HBV lifecycle and targets for direct-acting antiviral agents. NTCP receptor blocker inhibits HBV entrance into hepatocytes. siRNAs and ASO interfere with the translation of RNA transcribed from covalently circular DNA: messenger RNA and/or pregenomic RNA, and RNA transcribed from integrated HBV DNA. CAMs interfere with capsid formation resulting in empty or aberrant capsids, decreasing pregenomic RNA packaging and HBV DNA replication. Nucleic acid polymer blocks subviral HBsAg assembly and secretion, leading to HBsAg reduction. Vaccinal antibodies bind and neutralize circulating HBsAg and potentially restore exhausted HBV-specific immune response. (B) Restoration or stimulation of innate and/or adaptive immune system by immunomodulatory agents. Toll-like receptor 7 and 8 agonists activate innate immune response. Checkpoint inhibitors restore exhausted host immune antiviral response by blocking PD-1 or PDL-1. Therapeutic vaccine stimulates CD4 and CD8 T-cell responses. Vaccinal antibodies bind circulating HBsAg, which may enhance antigen delivery to dendritic cells and T-cell response. peg-IFN acts through the JAK-STAT pathway, upregulating the interferon stimulating genes enhancing HBV-specific immune response, and may also increase the degradation of cccDNA. Abbreviations: ASO, antisense oligonucleotide; CAM, capsid assembly modulators; cccDNA, covalently closed circular deoxyribonucleic acid; DC, dendritic cell; ISG, interferon stimulating gene; MDSC, myeloid-derived suppressor cells; NK, natural killer cell; NTCP, Sodium taurocholate cotransporting polypeptide; PD-1, programmed death 1; PDL-1, programmed death ligand 1; rcDNA, relaxed circular DNA; siRNA, short-interfering ribonucleic acid; TCR, T cell receptor; Th, T helper cell; Treg, regulatory T cell.

Among the new antivirals, short-interfering RNAs (siRNAs) and antisense oligonucleotide (ASO) have shown the most promise with 1–3 log₁₀ decrease in HBsAg levels after a few doses (Table 1 and Supplemental Materials, http://links.lww.com/XCL/A12). Studies with different siRNAs have shown a plateau in HBsAg decline around weeks 16–20 despite continued dosing, and HBsAg seroclearance has not been reported (Figure 2 and Supplemental Materials, http://links.lww.com/XCL/A12). Recovery of HBsAg-specific T-cell immune responses was observed in some but not all patients after a marked decline in HBsAg levels, suggesting that the addition of immune-modulatory therapy may help in immune recovery and potentially HBsAg seroclearance.^5,6 This hypothesis has been tested in 3 trials with 1 trial showing addition of peg-IFN to siRNA (VIR-2218) and NA resulted in HBsAg seroclearance in 6%–31% versus 0% at end-of-treatment compared to those who received VIR-2218 and NA. The addition of a vaccinal monoclonal HBsAb engineered to stimulate T cells (VIR-3434) to VIR-2218 and NA showed 90% of participants achieved HBsAg <10 IU/mL at end-of-treatment but none had HBsAg seroclearance (Figure 2 and Supplemental Materials, http://links.lww.com/XCL/A12).⁷

TABLE 1.

Summary of HBsAg and HBV DNA responses in key phase 2 trials of single new antiviral or single immunomodulatory therapy in patients who were NA naïve or virally suppressed on NA

		Duration of treatment (wk)		End-of-treatment (log10 reduction, IU/mL)			Posttreatment (log10 reduction, IU/mL)
	NA status	IP	NA	HBV DNA	HBsAg	FU wk	HBV DNA	HBsAg
CAMs
Vebicorvir (ABI-H0731)	Naive	24	24	5.3	<1	NR	—	—
	VS	24	24	n/a	<1	NR	—	—
JNJ-6379	Mixed (JADE)	24–48	72–96	5.5 to 5.9 (E+) 3.7 to 4.1 (E−)	<1	NR	—	—
	Mixed (REEF-1)	48	48–96	6.7 (E+) 4.8 (E−)	<1	24	6.7 (E+) 4.4 (E−)	<1
GLS-4	Mixed	96	96	6.0	<1	NR	—	—
ALG-000184	Naive	24–48	24–60	4.9–5.2	<1	NR	—	—
Posttranscription inhibitors: siRNA
AB-729	Naive	24	24	NR	1.8 (E+)	24	NR	1.6 (E+)
	VS	48	48	n/a	2.6 (E+)	24	n/a	2.3 (E+) SC: 14.3%
					1.9 (E−)	8	n/a	1.8 (E−)
JNJ-3989	Mixed (REEF-1)	48	48–96	5.9–6.6 (E+) 4.0–4.3 (E−)	1.5–2.6a SC: 3%a	24	6.1–6.8 (E+) 4.0–4.3 (E−)	1.0–1.9a
VIR-2218	VS	20	48–60	n/a	2.0b	40	n/a	1.6
Xalnesiran (RG6346)	VS (PIRANGA)	48	48	n/a	1.5–1.8 SC: 3%–7%	24	n/a	1–1.5 SC: 3%–7%
Posttranscription inhibitors: ASO
Bepirovirsen (highest dose group) (B-CLEAR)	Naïvec	24	48	2.5	3.1 SC: 29%	24	2	1.1 SC: 14%
	VS	24	48	n/a	2.8 SC: 26%	24	n/a	1.5 SC: 12%
Toll-like receptor agonists
Vesatolimod (GS-9620, TLR7)	Mixed	12	24–48	n/a	<1	36–48	n/a	<1
Selgantolimod (GS-9688, TLR8)	Naive	24	0	n/a	<1	24	n/a	<1
	Mixed	24	48	5 (Naïve)	<1	24	5.5–5.7 (Naïve)	<1
Therapeutic vaccines
GS-4774 (yeast-based, HBV S, C, and X antigens)	Naive	20	48	n/a	<1	28	n/a	<1
	VS	20	48	n/a	<1	28	n/a	<1

Note: NR, follow-up data not reported; n/a, not available, for example, HBV DNA response in patients virally suppressed on NA at the start of the trial.

^aMixed HBeAg status.

^bNadir responses reported.

^cRemained without NA during trial.

Abbreviations: CAMs, capsid assembly modulators; E, HBeAg; FU, follow-up; IP, investigation product; NA, nucleos(t)ide analog; SC, HBsAg seroclearance; siRNA, short-interfering ribonucleic acid; TLR, Toll-like receptor; VS, virally suppressed; wk, week.

FIGURE 2.

HBsAg seroclearance rates in key phase 2 trials of new antiviral and/or immunomodulatory therapies. All trials were on the backbone of NA therapy except for 1 stratum of the Bepirovirsen trial and 1 NAP trial. Duration of treatment (Tx) and posttreatment FU was shown in weeks (wk), and HBsAg seroclearance rates at EOT and posttreatment FU (FU) are shown in the figure. For trials with more than 1 treatment arm or more than 1 cohort, the highest HBsAg seroclearance rates are shown. For the Bepirovirsen (B-Clear) trial, HBsAg seroclearance rates in the highest dose group regardless of HBV DNA response are shown. For bepirovirsen plus peg-IFN (B-together), the response shown represents undetectable HBsAg and HBV DNA. Abbreviations: CAM, capsid assembly modulator; EOT, end-of-treatment; FU, follow-up; IFN, pegylated interferon; NAP, nucleic acid polymer; siRNA, short-interfering ribonucleic acid; TLR, toll-like receptor; Tx, treatment; wks, weeks.

By contrast, a phase 2 trial of a naked-ASO, bepirovirsen, showed that among patients in the highest dose group (300 mg weekly for 24 weeks), HBsAg seroclearance with undetectable HBV DNA was observed in 28%–29% patients at end-of-treatment and 9%–10% after 24 weeks posttreatment follow-up, with higher rates in those with low baseline HBsAg levels and in patients who are HBeAg-negative and no HBsAg seroclearance in any patients who are HBeAg-positive (Figure 2 and Supplemental Materials, http://links.lww.com/XCL/A12).⁸ Of note, the N-acetylgalactosamine–conjugated formulation that enhances hepatocyte uptake was less effective suggesting that bepirovirsen, which is largely taken up by macrophages, may have additional immunomodulatory effects.

Core assembly modulators (CAMs) interfere with capsid formation resulting in empty or aberrant capsids thereby decreasing HBV DNA replication. Clinical trials showed that the combination of CAM and NA resulted in a greater reduction in HBV RNA levels and an incremental decrease in HBV DNA levels compared to NA alone, but minimal changes in HBsAg levels (Table 1 and Supplemental Materials, http://links.lww.com/XCL/A12). Two studies using different siRNA and CAM showed that the addition of CAM resulted in similar or smaller decrease in HBsAg levels compared to siRNA and NA alone.⁵

Data on bulevirtide, an entry inhibitor, in HBV monoinfection are limited but data in patients with chronic hepatitis D showed bulevirtide monotherapy had minimal effect on HBV DNA or HBsAg levels.⁶

A small trial of nucleic acid polymer which blocks subviral HBsAg assembly and secretion showed 35% patients remained HBsAg-negative more than 1 year after stopping treatment of tenofovir followed by 48 weeks of additional REP-2139-Mg or REP 2165-Mg and peg-IFN (Figure 2 and Supplemental Materials, http://links.lww.com/XCL/A12).⁹ However, most patients experienced hepatitis flares and the results have not been replicated.

Various immune-modulatory approaches including stimulating innate immune response with toll-like receptors 7 and 8; activating T-cell responses with therapeutic vaccines comprising HBV surface as well as core, polymerase, and X antigens, with or without adjuvants; and the removal of immune blockade using checkpoint inhibitors have been studied but few of these therapies have resulted in >1 log₁₀ HBsAg decline and even fewer in HBsAg seroclearance (Table 1, Figures 1B and 2, and Supplemental Materials, http://links.lww.com/XCL/A12).^5–7 Recent studies have explored the role of neutralizing antibodies and augmentation of B-cell responses, with promising results in 1 study where a vaccinal HBsAb was combined with an siRNA (discussed above). Other studies have combined peg-IFN with siRNA or ASO, together with NA or other immune-modulatory therapies with HBsAg seroclearance rates ≥10% at follow-up week 24 (Table 2, Figure 2, and Supplemental Materials, http://links.lww.com/XCL/A12) though long-term durability remains unclear.

TABLE 2.

Summary of HBV DNA and HBsAg responses in key phase 2 trials of a combination of 2 new antivirals or 1 new antiviral with immunomodulatory therapy on the backbone of NA

Combination				End-of-treatment responses					Follow-up responses
Direct antiviral	Immune-modulatory	HBeAg	NA status	Duration of treatment (wk)	HBV DNA	HBsAg	HBsAg seroclearance	wk	HBV DNA	HBsAg	HBsAg seroclearance
Vebicorvir (CAM)	peg-IFN	Pos	Naive	24	6.2	<1	0%	24	6.5	<1	0%
Vebicorvir (CAM) + AB-729 (siRNA)	—	Neg	VS	48	n/a	1.9	0%	8	n/a	1.9	0%
JNJ-6379 (CAM) + JNJ-3989 (siRNA)	—	Mixed	Mixed	48	6.7 (E+) 3.6 (E−)	1.4–2.0 (E+) 1.1–1.3 (E−)	0%	24	6.9 (E+) 3.7 (E−)	1.4–2.0 (E+) 1.1–1.3 (E−)	0%
		Neg	VS	48	n/a	1.9	0%	24	n/a	1.8	0%
JNJ-6379 (CAM) + JNJ-3989 (siRNA)	peg-IFN	Mixed	VS	24	n/a	2.2	2.1%	NR	—	—	—
		Pos (IT)	Naïve	48a	6.2	3.6	12.2%	24	6.5	2.9	16.3%
RO7049389 (CAM)	peg-IFN	Mixed	Naive	48	7.0	1.6 (E+) 0.9 (E−)	0%	24	n/a	1.6 (E+) 0.9 (E−)	0%
AB-729 (siRNA)	peg-IFN	Neg	VS	12–24	n/a	2.0–3.4	9%	12	n/a	1.2	0%
VIR-2218 (siRNA)	peg-IFN 12-48 wk	Mixed	VS	20–48	n/a	2.0–2.9	6%–31%	24	n/a	1.1–1.2	0%–17%
VIR-2218 (siRNA)	VIR-3434 (vaccinal monoclonal Ab)	Mixed	VS	20	n/a	2.7–3.1	15%	48	n/a	0.8–0.9	0%
VIR-2218 (siRNA)	VIR-3434 (vaccinal monoclonal Ab) + peg-IFN	Mixed	VS	24	n/a	n/a	14.3%	12	n/a	n/a	9.5%
Xalnesiran (RG6346) (siRNA)	Ruzotolimod (TLR7)	Mixed	VS	48	n/a	2.0	20%	24	n/a	1.5	13%
	peg-IFN	Mixed	VS	48	n/a	2.0	30%	24	n/a	1.8	23%
Bepirovirsen (ASO)	peg-IFN	Mixed	VS	48	n/a	NR	17%–22%b	24	n/a	NR	9%–15%b
—	Nivolumab (anti-PD-1) + VTP-300 (Therapeutic vaccine)	Neg	VS	1 dose Nivo and 2 dose VTP-300	n/a	n/a	n/a	32	n/a	1.0	11%
		Mixed	VS	4–12	n/a	n/a	n/a	4–12	n/a	0.2–1.7	2.5%
—	Nivolumab (anti-PD-1) + GS-4774 (therapeutic vaccine)	Neg	VS	12	n/a	<1	0%	12	n/a	<1	0%
—	BRII-179 (therapeutic vaccine) + IFN	Mixed	VS	24	n/a	<1	0%	NR	—	—	—
	BRII-179 (therapeutic vaccine) + IFN	Neg	VS and peg-IFN lead-in 24–48 wk	24	n/a	NR	32.6%	12	n/a	NR	31.8%
REP2139 or REP2165	peg-IFN	Neg	Naïve, TDF added	48	7	3.6	60%	48	n/a	3.6	35%

Note: HBsAg and HBV DNA responses were reported as log10 IU/mL reduction from baseline or percentage with HBsAg seroclearance.

^aJNJ3989 +/− JNJ-6379 lead in 36 weeks followed by peg-IFN 12 weeks.

^bHBsAg below detection AND HBV DNA below quantification.

Abbreviations: Ab, antibody; CAMs, capsid assembly modulators; E, HBeAg; IP, investigation product; IT, immune-tolerant; n/a, not available; NA, nucleos(t)ide analog; Neg, negative; NR, follow-up data not reported; PD-1, programmed death 1; peg-IFN, pegylated interferon alpha-2a; Pos, positive; SC, HBsAg seroclearance; siRNA, short-interfering ribonucleic acid; TDF, tenofovir disoproxil fumarate; TLR, toll-like receptor; VS, virally suppressed; wk, weeks.

PATH TO HBV CURE

Combination therapy will be necessary to fully suppress HBV DNA replication and HBsAg production followed by stimulation/restoration of HBV-specific immune response (Figure 3 and Supplemental Materials, http://links.lww.com/XCL/A12). NAs will remain the backbone for suppressing HBV DNA replication, complemented by CAM, siRNA/ASO, and possibly an entry inhibitor. siRNA/ASO will be pivotal in suppressing HBsAg production and could be designed to target transcripts from both covalently closed circular DNA and integrated HBV DNA. peg-IFN and some immune-modulatory therapies may facilitate HBsAg seroclearance after levels are lowered by siRNA/ASO, though data from 1 trial of sequential therapy with bepirovirsen (ASO) followed by peg-IFN showed negligible incremental HBsAg seroclearance and did not prevent HBsAg relapse.¹⁰ For many patients, additional immune modulation will be needed. Two major challenges in developing an HBV cure involve individual patient immune responsiveness and safety concerns. NAs have an excellent safety profile and any attempt to improve HBV therapy must not compromise safety. Thus, combination therapies that include peg-IFN with its well-known adverse effects or immune checkpoint inhibitors with potential for systemic immune-mediated complications can only be justified if they lead to significantly higher HBsAg seroclearance rates compared to simpler and safer therapies and must be accompanied by careful patient selection and monitoring. Contrary to direct-acting antivirals, immune-modulatory therapies may need to be tailored to individual HBV-specific immune impairment to enhance the likelihood of HBV cure. As guidelines expand treatment indications including patients in the immune-tolerant phase, new therapies must evaluate their efficacy in these patients as well as children, patients with cirrhosis, and those with HIV or HDV coinfection, who are currently excluded from clinical trials.

FIGURE 3.

Strategies toward combination of novel therapies toward HBV cure. Multiple steps are required to achieve HBV functional cure including inhibition of viral replication, reduction of viral antigen production, and boosting/restoring HBV-specific immune response. The major concern for combining different regimens is possible adverse events. Abbreviations: ASO, antisense oligonucleotide; CAM, core assembly modulator; IFN, interferon; NA, nucleos(t)ide analog; NAP, nucleic acid polymer; siRNA, short-interfering ribonucleic acid.