microorganisms

Review
Chronic Hepatitis D Virus Infection and Its Treatment:
A Narrative Review
Poonam Mathur * , Arshi Khanam and Shyam Kottilil

Institute of Human Virology, University of Maryland School of Medicine, Baltimore, MD 21201, USA;
akhanam@ihv.umaryland.edu (A.K.); skottilil@ihv.umaryland.edu (S.K.)
* Correspondence: pmathur@ihv.umaryland.edu

Abstract: More than 12 million individuals worldwide are chronically infected with the hepatitis D
virus (HDV). HDV infection is the most severe form of viral hepatitis since it requires hepatitis B
virus co-infection and accelerates progression to cirrhosis and hepatocellular carcinoma. Therefore,
treatment modalities to slow the progression of the disease are essential but not yet available. In addi-
tion, no antiviral treatment to date has been shown to reliably eradicate HDV. Pegylated interferon
(PEG-IFN) is the only universally used treatment to suppress HDV RNA replication and improve
liver inflammation and fibrosis. This treatment can be completed in 12–18 months, but cure rates
remain low, and success does not reliably increase with the addition of a nucleos(t)ide analog. PEG-
IFN therapy is also limited by poor tolerability and multiple adverse effects, including neutropenia,
thrombocytopenia, and neuropsychiatric symptoms. Newer antiviral therapies in development target
unique aspects of HDV viral replication and show promising results in combination with PEG-IFN
for long-term HDV RNA suppression. These newer antiviral therapies include buleviritide (which
blocks HDV entry), lonafarnib (which prevents HDV assembly), and REP-2139 (which prevents HDV
export). In this manuscript, we discuss the characteristics of HDV infection and review the new
antiviral therapies approved for treatment and those under investigation.

Keywords: hepatitis D virus; hepatitis; treatment

Citation: Mathur, P.; Khanam, A.;

1. Introduction
Kottilil, S. Chronic Hepatitis D Virus
Infection and Its Treatment: A The hepatitis D virus (HDV), first discovered in 1977, is estimated to chronically infect
Narrative Review. Microorganisms 12 million people worldwide [1,2]. HDV requires concomitant hepatitis B virus (HBV)
2024, 12, 2177. https://doi.org/ infection to establish productive infection [3]. The estimated prevalence of HDV among
10.3390/microorganisms12112177 those with hepatitis B surface antigen (HBsAg) positivity is 0.8–15% [1,4,5]. HDV causes
the most severe form of viral hepatitis (since it requires HBV co-infection), significantly
Academic Editors: Isabelle Chemin
and Flor Helene Pujol
accelerates the progression of liver cirrhosis, and increases the risk of hepatic decompen-
sation and hepatocellular carcinoma (HCC) compared to HBV mono-infection [1,4,6–9].
Received: 10 October 2024 Furthermore, HDV is an independent risk factor for liver-related and all-cause mortality
Revised: 24 October 2024 in patients with chronic HBV infection [10]. This review provides a summary of HDV
Accepted: 25 October 2024 infection and its treatment.
Published: 29 October 2024
2. Epidemiology
The true global prevalence of HDV is unknown due to the low proportion of HBsAg+
Copyright: © 2024 by the authors.
individuals that are screened for HDV, variations in screening guidance, limited epidemio-
Licensee MDPI, Basel, Switzerland. logic data from several countries, and lack of standardized confirmatory anti-HDV antibody
This article is an open access article and PCR tests and availability of these tests [4,11,12]. There is high HDV endemicity in
distributed under the terms and central and west Africa, central and east Asia, the Amazon Basin, eastern Europe, and
conditions of the Creative Commons the western Pacific region [1,4,12,13]. More specific prevalence estimates among HBsAg
Attribution (CC BY) license (https:// carriers show that Mongolia has the highest rate of HDV RNA prevalence (61%) [14]. Low
creativecommons.org/licenses/by/ HDV endemicity areas include northern Europe, North America, and Japan, likely in part
4.0/). due to hepatitis B vaccination campaigns and low HBV endemicity [1,3,4,13].

Microorganisms 2024, 12, 2177. https://doi.org/10.3390/microorganisms12112177 https://www.mdpi.com/journal/microorganisms

Microorganisms 2024, 12, 2177 2 of 15

Like HBV, HDV is a blood-borne pathogen, so transmission and risk of infection relate
to contact with blood from infected individuals. Since HDV infection requires the presence
of HBV, vaccination against HBV can also prevent HDV [15]. Patients with HBV infection
should be screened at least once for HDV co-infection [1,16]. Screening is particularly
indicated in patients with HBsAg+ and low/undetectable HBV DNA but persistently
high ALT [16]. Additional populations that are at increased risk for HDV infection and
warrant repeated screening include people who inject drugs, sex workers, men who have
sex with men, and those with iatrogenic exposures [1,16]. HIV and hepatitis C virus (HCV)
co-infection also increase the risk of HDV due to similar modes of transmission, so patients
with the former infections should also be screened for HDV infection [1,17]. However,
high HDV prevalence outside of these populations is also possible; in an anti-HDV test
implementation program in Barcelona, 60% of HDV RNA-positive patients had no risk
factors identified [18].

3. HDV Virology and Immune Response

HDV is a single-stranded RNA virus consisting of a genome of 1672–1697 ribonu-
cleotides [5,19]. Its genome is only slightly larger than that of a plant viroid, making it
one of the smallest viruses known to infect humans [1,20]. HDV is a defective satellite
virus that requires the presence of HBsAg for infection and viral replication; its envelope is
derived from and composed of HBV surface proteins [3,5,19]. The HDV genome has only
one actively transcribed open reading frame that codes for two forms of HDAg, large (L)
and small (S). S-HDAg is necessary to initiate and maintain replication, while L-HDAg
negatively regulates replication and triggers HDV to envelop the HBV surface proteins [5].
HDV uses HBsAg to enter hepatocytes via the sodium/bile acid co-transporter (NTCP) [21];
specifically, HDV binds to the pre-S1 domain of HBsAg so it can dock to NTCP. Once HDV
enters and delivers its genome to the nucleus of the hepatocyte, S-HDAg-encoding mRNA
is transcribed and translated [5]. After, the newly synthesized HDV RNA is transcribed to
HDAg mRNA and uses a host enzyme, ADAR-1, to edit the HDAg mRNA, leading to the
formation of L-HDAg [5]. L-HDAg is then prenylated by the enzyme farnesyltransferase [5].
This post-translational modification of L-HDAg—prenylation—is required for L-HDAg
to interact with HBV envelope proteins and virion assembly [19]. This interaction can
be blocked by prenylation inhibitors, which selectively target the assembly of the HDV
virion [22]. Since HDV has limited protein-coding capacity, it deceives host machinery and
is replicated by cellular RNA polymerases that are driven to replicate the HDV genome
as though it were endogenous DNA, which makes finding therapeutic targets difficult
(Figure 1) [23–25].
After infection, interferon (IFN) is the main innate immune mechanism to contain
viral replication [5]. Unlike HBV, HDV induces an IFN response, which serves as a bridge
to the development of adaptive immunity characterized by anti-HDV antibodies and HDV-
specific T-cells [5]. In patients with active hepatitis, anti-HDV IgM antibody titers are
high but lack neutralizing activity and do little to control viral replication [5]. The role of
virus-specific T-cells is not well-defined for HDV infection, but they are presumed to be the
drivers of viral control, similar to HBV and HCV infection [5].
There are eight distinct HDV genotypes that have different replication efficacies [5].
Genotype (GT) 1 is most common, whereas GT 2–8 are found in distinct geographic
regions [3,5]. GT testing is not routinely undertaken, but there is emerging evidence that
genotype may affect disease severity and treatment response [3,26,27].
 2024, 12,
Microorganisms 2024, 12, 2177
x FOR PEER REVIEW 33of
of 16
15

Figure 1. The hepatitis D virion and replication cycle. BLV = buleviritide. LNF = lonafarnib. NTCP =
Figure 1. The hepatitis D virion and replication cycle. BLV = buleviritide. LNF = lonafarnib. NTCP =
sodium/bile acid co-transporter. S-HDAg = small hepatitis delta antigen. L-HDAg = large hepatitis
sodium/bile acid co-transporter. S-HDAg = small hepatitis delta antigen. L-HDAg = large hepatitis
delta antigen. (Created with BioRender.com).
delta antigen. (Created with BioRender.com).
After infection,
4. Clinical interferon
Presentation (IFN) History
and Natural is the main innate immune
of Chronic mechanism to contain
HDV Infection
viral replication [5]. Unlike HBV, HDV induces an IFN response, which serves as a bridge
HDV can either be acquired as a co-infection with HBV or super-infection in already-
to the development of adaptive immunity characterized by anti-HDV antibodies and
established HBV infection [3]. Co-infection almost always results in spontaneous clearance
HDV-specific T-cells [5]. In patients with active hepatitis, anti-HDV IgM antibody titers
of HDV without progression of the disease, similar to that seen in patients with HBV
are high but lack neutralizing activity and do little to control viral replication [5]. The role
infection in the “immune control” phase [3]. Upon serologic evaluation, these patients
of virus-specific T-cells is not well-defined for HDV infection, but they are presumed to
would have a positive HDV antibody (Ab) but a negative HDV RNA. Super-infection,
be the drivers of viral control, similar to HBV and HCV infection [5].
however, results in disease chronicity and rapidly progressive disease 90% of the time [3].
There
It is not are eight what
well-known distinct HDVthe
causes genotypes thatof
acceleration have
liverdifferent
disease, replication efficacies
but it may be [5].
secondary
Genotype (GT) 1 is most common, whereas GT 2–8 are found in distinct geographic
to the direct cytopathic effects of the virus, especially S-HDAg, as well as the aberrant re-
gions [3,5]. GTcytokine
inflammatory testing release
is not routinely undertaken,
due to innate but there
and adaptive is emerging
immune evidence
responses. that
In multiple
genotype may affect disease severity and treatment response [3,26,27].
cohorts globally, it has been demonstrated that HDV replication is the major characteristic
affecting disease severity. Chronic HDV infection from super-infection can suppress HBV
4.
DNAClinical Presentation
replication and Natural
so that HDV History of
is the dominant Chronic
virus HDV Infection
[3,12,24,25]. However, there are some
casesHDV
where can
HBVeither be acquired
replication as a co-infection
predominates (30% ofwith HBV or
patients) or super-infection
HBV replicationinand already-
HDV
established
replication areHBV infection
equal (15% of[3]. Co-infection
patients) [3]. almost always results in spontaneous clear-
ance People
of HDVwith
without
HBV/HDVprogression of the
infection disease,
usually similar
have to that
chronic seen inand
hepatitis, patients with HDV
persistent HBV
infectionoften
viremia in the “immune liver
accelerates control” phase
fibrosis and[3]. Upon serologic
increases the risk forevaluation,
cirrhosis,these
liverpatients
decom-
would haveand
pensation, a positive HDV antibody
HCC [1,4,6,8,9,28]. (Ab) HDV
Chronic but a infection
negative HDV RNA. Super-infection,
also increases the incidence
however, resultsmorbidity,
of liver-related in disease need
chronicity andtransplantation,
for liver rapidly progressive anddisease 90% ofmortality
liver-related the time [3].
[3].
People
It is not with chronicwhat
well-known infection
causes maythe also present of
acceleration with
liverpersistent
disease, butALT elevation
it may despite
be secondary
suppression
to the direct of HBV DNA
cytopathic withofnucleos(t)ide
effects analog (NA)
the virus, especially therapy
S-HDAg, [29]. as
as well Progression
the aberrant to
cirrhosis can occur as quickly as within 5 years of infection and to HCC
inflammatory cytokine release due to innate and adaptive immune responses. In multiple within 10 years [3].
Moreover,
cohorts the riskit of
globally, hasHCC
beenisdemonstrated
about three-fold thathigher
HDV in patients with
replication is theHBV/HDV infection
major characteristic
affecting disease severity. Chronic HDV infection from super-infection can suppress HBV
Microorganisms 2024, 12, 2177 4 of 15

compared to those with HBV mono-infection [12]. Populations with increased risk of liver
disease progression are listed in Table 1.

Table 1. Patient groups infected with hepatitis D virus (HDV) at higher risk for liver disease
progression [6,9,10,23,30–36]. HCV = hepatitis C virus. HBV = hepatitis B virus. NA = nucleos(t)ide
analog.

Patient Group
Chronic HDV infection with persistent viremia
Older age
Male sex
HIV or HCV co-infection
High serum HBV DNA levels
Elevated gamma-glutamyl- or aminotransferase levels despite NA treatment
Chronic liver disease and cirrhosis
Cofactors of chronic liver injury, including alcohol abuse, obesity, and diabetes

Patients with HBV/HDV infection who undergo liver transplantation are younger
than those with HBV mono-infection and more likely to have decompensated liver disease.
Fortunately, outcomes post-transplantation are similar between the two groups. In addition
to clinical endpoints, patient-reported outcomes indicate that those with chronic HDV
infection fare worse in their emotional and physical well-being compared to those with
HBV infection alone.
Patients with HBV infection are monitored every 6–12 months for advancing fibrosis
or HCC, and this is of even more importance for those with chronic HDV infection [28].
Monitoring is routinely carried out by ultrasonography, even if aminotransferase levels
are normal [28]. An abnormal ultrasound of the liver should be followed up by computed
tomography or magnetic resonance imaging of the liver [28]. Liver transplantation may be
required due to the rapid progression of liver disease and/or decompensated cirrhosis [3].
Based on the U.S. national transplant database, patients with HBV/HDV infection receive
liver transplants at younger ages than those with HBV only [3]. For example, in a U.S.-based
retrospective study, patients with HDV undergoing liver transplants had a mean age of 52,
and their HBV mono-infected counterparts had a mean age of 55.

5. Diagnosis and Liver Fibrosis Assessment

Diagnosis of HDV infection is made by screening for anti-HDV antibodies and using
quantitative HDV RNA for confirmation (since anti-HDV IgM and IgG may remain positive
after spontaneous clearance) [16,23,37,38]. Challenges to a uniform testing/diagnostic
strategy are lack of provider knowledge regarding HDV, limited treatment options, and
difficulty in accessing HDV-specific laboratory tests [3,4,12]. In addition, since standardized
and validated real-time HDV RNA PCR assays are neither available worldwide nor FDA-
approved, reflex testing, which would test for HDV IgM and IgG in all HBV-positive
individuals, is not routine [23]. Additional items for consideration when evaluating a
patient with HDV infection are shown in Table 2.
Liver fibrosis assessment is best carried out using transient elastography (TE) or liver
biopsy. TE results in the context of HDV infection used to be difficult to interpret since there
are no clear cut-offs for fibrosis; however, a recent study that compared TE to liver biopsy
proposed numerical cut-offs for non-advanced liver fibrosis and cirrhosis, which may be of
utility [39]. Non-invasive, serologic liver fibrosis tests used for HBV infection, such as the
APRI and FIB-4 tests, do not perform well for HDV [40]. However, two non-invasive tests
have been developed and validated for HDV: the delta fibrosis score and delta-4 fibrosis
score [40,41]. Each score incorporates different variables to calculate the score and can be
used as adjuncts to corroborate liver fibrosis staging derived from TE.
Microorganisms 2024, 12, 2177 5 of 15

Table 2. Evaluation of a patient with chronic hepatitis D virus (HDV) infection. Since no clear
guidelines for surveillance of patients with HDV infection are available, clinicians should follow
guidelines for HBV infection [28]. a No clear cut-offs for transient elastography (TE) have been
established due to the paucity of data regarding TE use in HDV infection.

Laboratory Tests
• Anti-HDV antibodies
• Quantitative HDV RNA
• Screening for HIV, hepatitis C virus, hepatitis A virus immunity
• Complete blood count
• Complete metabolic panel (glucose, electrolytes, and assessment of renal and liver function,
including measurement of liver enzyme levels)
Liver Fibrosis Evaluation
• Liver biopsy
• Transient elastography (TE) a
Health Maintenance
• Hepatocellular carcinoma screening with ultrasonography, computed tomography, or
magnetic resonance imaging every 6–12 months
• Hepatitis A vaccine

6. Treatment
Antiviral treatment is critical for reducing the morbidity and mortality associated with
persistent HDV viremia [3,6,10]. Persistent suppression of HDV RNA is associated with
improved clinical outcomes, including decreased liver-related complications [12,30]. The end-
point for successful treatment response is disputed, but most studies use undetectable or
≥2 log drop in HDV RNA 24 weeks after stopping treatment [16,42,43]. Treatment should
normalize ALT levels, indicating a resolution of liver inflammation [16]. Loss of HBsAg
is critical for the resolution of HDV infection and improves survival; therefore, tracking
HBsAg levels may be a useful guide for when to stop treatment [44–49]. Accordingly, low
HBsAg levels at baseline are a predictor of treatment response [44]. Unfortunately, most
patients with chronic HBV infection do not lose HBsAg, making HDV RNA suppression dif-
ficult for co-infected patients [28]. A summary of approved and investigational treatments
is found in Table 3.

Table 3. Mechanism of action and adverse effects of major therapies available for chronic hepatitis D
virus (HDV) infection. NTCP = sodium/bile acid co-transporter. HDAg = large hepatitis D antigen.
BID = twice daily. RTV = ritonavir. HBsAg = hepatitis B surface antigen. HBV = hepatitis B virus.

Drug Mode of Delivery Mechanism of Considerations Adverse Effects Treatment Considerations

and Dose Action for Use
Pegylated-IFN Subcutaneous Binds to type I Most used for Increase in transaminases, Sustained virologic
and pegylated- injection: interferon receptors, HDV treatment abnormal hematologic response ranges from 23 to
IFNa-2a 180 mg/week for activating laboratory results, fatigue, 57% and does not increase
12–18 months immunomodulatory flu-like symptoms, and with the addition of
and antiviral psychiatric nucleoside analog [16].
proteins. symptoms [45,50].
Pegylated-IFNl Subcutaneous Binds to type III Investigational Flu-like symptoms, Investigated in
injection: 180 mg or interferon receptors, increase in transaminases, proof-of-concept study
120 mg/week for activating and psychiatric only.
48 weeks immunomodulatory symptoms [51].
and antiviral
proteins.
Microorganisms 2024, 12, 2177 6 of 15

Table 3. Cont.

Drug Mode of Delivery Mechanism of Considerations Adverse Effects Treatment Considerations

and Dose Action for Use
Buleviritide Subcutaneous Blocks HDV docking Investigational, Headache, pruritis, fatigue, Maximal efficacy when
injection: 2 mg, to NTCP and not yet eosinophilia, injection site combined with PEG-IFNa.
5 mg, or 10 mg hepatocyte entry. FDA-approved. reactions, upper Reduces HDV RNA levels
abdominal pain, arthralgia, and normalizes ALT [52,55].
asymptomatic bile salt Efficacy not influenced by
increase, and increase in the presence of
transaminases [52–54]. cirrhosis [56]. Daily
injection may limit
adherence.
Lonafarnib Oral: 25 mg, 50 mg, Inhibits L-HDAg Investigational, Nausea, diarrhea, Maximal efficacy and
75 mg, 100 mg, prenylation. not yet abdominal bloating, tolerability when
200 mg, or 300 mg FDA-approved. weight loss, and ALT combined with RTV and
BID flaring [57,58]. PEG-IFNa [58,59]. RTV is a
CYP450 substrate, causing
drug-drug interactions.
REP-2139 Intravenous: Inhibits export/ Investigational, Pyrexia and chills [60]. Combined with PEG-IFNa
250 mg or 500 mg secretion of HDV not yet for efficacy without
from hepatocytes. FDA-approved. compromising
tolerability [60]. May
remove integrated HBV
DNA from the liver [61].

6.1. Pegylated Interferons

The first treatment for HDV infection was pegylated-interferon-alpha (PEG-IFNa) for
48 weeks. Interferon is thought to interfere with HDV replication by inhibiting cell-to-cell
spread during cell division and reducing the risk of liver decompensation, even if there
is no virologic response. PEG-IFNa is a type I interferon administered subcutaneously
(SC) (usually weekly) [6,50]. It has a sustained viral response rate (SVR) of 23–57% when
treatment is administered for 48 weeks [16]. ALT normalization usually follows a virologic
response, as well as improvement in liver histology [16,62]. Some patients lose HBsAg
with 12–18 months’ treatment of PEG-IFNa, but sustained HBsAg loss usually requires
prolonged treatment [49,62]. Studies of ≥2 years of treatment with PEG-IFNa have found
higher rates of sustained HDV RNA (58%) and HBsAg clearance (33%), and improvements
in liver histology [45,63]. Relapse of HDV viremia has been seen in about 50% of patients
even after prolonged PEG-IFNa treatment, occurring up to 9 years after the completion
of therapy [47,50,63,64]. Thus, the optimal treatment duration of PEG-IFNa is yet to be
established [62,65]. Unfortunately, PEG-IFNa is contraindicated in advanced liver disease
or major extrahepatic comorbidities [23]. It is also associated with neuropsychiatric and
hematologic side effects, as well as paradoxical increases in aminotransferases, requiring
frequent monitoring [23,45,50].
Success with PEG-IFNa is not enhanced by the addition of HBV-targeting NAs, as seen
in randomized, placebo-controlled trials using adefovir or tenofovir disoproxil fumarate
(TDF) in addition to PEG-IFNa or PEG-IFNa-2a [45,50,55,66]. In addition to the lack of
efficacy enhancement, Anastasiou et al. noted that with the addition of an NA there were
paradoxical increases in HBV DNA towards the end of treatment that were positively
associated with HBsAg loss and HDV RNA suppression [66]. Moreover, biochemical
markers of cell death (M30 and ALT) were higher during the HBV DNA peaks, suggesting
that PEG-IFNa mediated death of hepatocytes may parallel increases in HBV DNA. In a
systematic review and meta-analysis of randomized controlled trials using IFN and NAs,
IFN and IFN+ NAs had high rates of viral relapse 24 weeks after the end of treatment,
confirming that there is no apparent benefit of adding NAs to IFN [67]. Despite high rates
of relapse, the AASLD and EASL recommend IFN+ NA treatment in HDV patients if they
have cirrhosis or serum HBV DNA levels are elevated since there are significant benefits if,
by chance, HDV suppression is achieved [16,23].
Microorganisms 2024, 12, 2177 7 of 15

Due to the intolerable side effects associated with PEG-IFNa, a type III IFN, PEG-
IFNl, was investigated in an open-label, proof-of-concept study, LIMT-1 [51]. Like IFNa,
IFNl has a capacity for broad induction of antiviral activity with a similar downstream
signaling pathway, but its receptors are mainly in gastrointestinal and respiratory tracts’
epithelial cells and not in hematopoietic cells. Due to the limited receptor distribution,
the investigators of LIMT-1 assessed if the safety and tolerability of IFNl were better than
IFNa. Thirty-three patients received IFNl 180 mg or 120 mg SC weekly for 48 weeks.
The IFN 180 mcg group demonstrated higher efficacy: HDV RNA negativity at 24-week
follow-up was 5/14 (36%) for participants in the 180 mg and only 3/19 (16%) in the 120 mg
group. There were no significant changes in HBsAg levels throughout the study for either
treatment group, and there were no significant associations between the HBsAg levels, HDV
RNA, or ALT. Common side effects were flu-like symptoms and elevated transaminases.
Only one patient had a neuropsychiatric side effect. Though PEG-IFNl appeared more
tolerable than PEG-IFNa in this trial, no further investigations are planned due to four
patients experiencing hepatobiliary events that resulted in hepatic decompensation in a
separate phase III trial [68].

6.2. Novel HDV-Specific Antivirals

HDV-specific antiviral therapies inhibit the HDV life cycle via three mechanisms [12]:
1. Blocking HDV particles from entering hepatocytes (buleviritide, BLV).
2. Preventing assembly of mature infectious HDV particles (lonafarnib, LNF).
3. Preventing export of HDV particles (REP-2139).

6.2.1. Buleviritide (BLV)

BLV is a myristoylated, synthetic lipopeptide corresponding to the preS1 sequence
of HBsAg. Essentially, BLV is a mimic of the functional lipopeptide pre-S1 of HBsAg that
HDV uses to dock to the NTCP co-transporter. Therefore, by acting as a pre-S1 lipopeptide
decoy, BLV blocks HDV from entering hepatocytes. Overall, BLV is well-tolerated without
drug-related serious AEs or treatment discontinuations [52,53,55]. The most common
AEs have been headache, pruritus, fatigue, eosinophilia, injection site reactions, upper
abdominal pain, and arthralgia. Asymptomatic dose-dependent increases in bile acid levels
also occur; bile acid level increases are expected since the NTCP functions as a bile acid
transporter [53–56,69,70].
To assess efficacy, the first-in-human, randomized phase 1b/2a study of BLV included
24 patients 1:1:1 to receive BLV 2 mg, PEG-IFNa-2a, or a combination for 24 weeks. BLV was
well-tolerated without serious adverse events (SAEs), and there were significant reductions
in HDV RNA after 24 weeks, with a higher mean of on-therapy decline when BLV was
combined with PEG-IFNa [52]. Viral kinetic modeling confirmed the strong, synergistic
effect of BLV and PEG-IFNa-2a on both HBV and HDV RNA levels. Also, ALT normalized
with BLV monotherapy in 6 of 8 patients. However, the primary endpoint, ≤0.5 log decrease
in HBsAg, was not reached in any patient.
MYR202 was a multicenter, parallel-group, randomized, open-label phase II study that
assessed the efficacy of BLV at different doses for 24 weeks. Participants were randomized
1:1:1:1 for 2, 5, and 10 mg of BLV with TDF 245 mg once daily or TDF 245 mg once daily
alone [53]. The study enrolled 120 patients, and 59 patients had cirrhosis. The primary
endpoint was HDV RNA below the level of detection or ≥2 log IU/mL decline in HDV
RNA at week 24. The results for participants with virologic responses were: BDV 2 mg +
TDF, 15/28 (54%); BDV 5 mg + TDF, 16/32 (50%); BDV 10 mg + TDF, 23/30 (77%); and
TDF 1/28 (4%) (p < 0.0001 for all BDV/TDF compared to TDF alone). Interestingly, efficacy
was not dose-dependent. Liver biopsies from 22 participants at baseline and at the end
of treatment were analyzed, and all showed decreased HDV RNA; however, 89% (49/55)
of participants developed relapse after treatment discontinuation, with aminotransferase
flares in 22% of patients, suggesting BLV efficacy with an NA is not sufficient for HDV
suppression, unlike what was seen when BLV was combined with PEG-IFNa-2a. Similar to
Microorganisms 2024, 12, 2177 8 of 15

the BLV + PEG-IFNa-2a combination, there were no significant changes in serum HBsAg
levels with BLV + NA.
A subsequent phase II study, MYR203, was a multicenter, open-label, randomized
study that allocated 90 participants into six treatment groups: PEG-IFNa 180 µg weekly,
BLV 2 mg daily + PEG-IFNa 180 µg weekly, BLV 5 mg daily + PEG-IFNa 180 µg weekly,
BLV 10 mg + PEG-IFNa 180 µg weekly, and BLV 10 mg + TDF, or BLV 2 mg daily for
48 weeks [55]. Twenty-four weeks after the end of treatment, the highest rates of HDV
RNA below the level of detection were in participants who were treated with BLV 2 mg
daily + PEG-IFNa 180 µg weekly (53%). Also, there were more BLV 2 mg dose participants
with ≥1 log HBsAg log decrease (40% patients) than in the other groups, including those
treated with 5 mg and 10 mg of BLV, confirming a lack of dose-dependent efficacy for BLV,
as was suggested by MYR202. At follow-up, HDV RNA clearance persisted in patients
who had decreased HBsAg.
BLV was also investigated in a randomized, phase III study, MYR301 [54]. There
were 150 participants randomly assigned 1:1:1 to receive BLV 2 mg or 10 mg daily for
144 weeks or no treatment for 48 weeks, followed by BLV 10 mg daily for 96 weeks (the
control group). The primary endpoint, which was a combined response at week 48 of
undetectable HDV RNA or level that decreased by ≤2 log IU/mL and normalization of
ALT, occurred in 45% of participants in the BLV 2 mg group, 48% in the BLV 10 mg group,
and 2% in the control group (p < 0.001 for comparison with control group). ALT normalized
in 12% of participants in the control group, 51% in the BLV 2 mg group, and 56% in the BLV
10 mg group. As seen in phase II studies, there was no efficacy advantage of BLV 10 mg
over 2 mg for HDV RNA suppression (20% vs. 12%, respectively, p = 0.41). HBsAg level
loss or level decrease by ≥1 log IU/mL did not occur in either group at week 48. In the
ongoing study of MYR301 to week 96, out of 150 participants who did not have a virologic
response at week 24, 43% achieved a virologic response at week 96. Biochemical response
also occurred, but this was often independent of virologic response [71]. Also, post-study
analyses of liver biopsy samples from patients in the MYR202, MYR203, and MYR301
trials show that BLV effectively decreased the number of HDV-positive hepatocytes, which
results in a concomitant decrease in transcriptional levels of inflammatory chemokines and
interferon-stimulated genes, compared to placebo, reinforcing the notion that long-term
use of BLV for more than 24 weeks may increase HDV SVR rates [72].
MYR204 was a phase 2b, open-label trial comparing the efficacy of (1) PEG-IFNa-2a
(180 µg weekly) for 48 weeks, (2) BLV 10 mg for 96 weeks, and (3) BLV (2 mg or 10 mg
daily) in combination with PEG-IFNa-2a (180 µg weekly) for 48 weeks, followed by the
same dose of BLV alone for another 48 weeks [73]. The primary endpoint was HDV RNA
levels 24 weeks after the end of treatment, and the primary comparison was between the
BLV 10 mg alone and BLV 10 mg + PEG-IFNa-2a groups. The combination of BLV 10 mg +
PEG-IFNa-2a was superior in regard to undetectable HDV RNA at week 24 (34 percentage
point difference; 95% confidence interval 15 to 50; p < 0.001).
Last, BLV has been investigated in prospective studies of patients with HDV infection
and compensated cirrhosis [56,74]. In a cohort of 18 Caucasian patients who received BLV
2 mg daily for 48 weeks, there were no participants who developed hepatic decompensation
or HCC. The regimen was well-tolerated, with no discontinuations for AEs or symptoms
of increased bile acids. Virologic response (HDV RNA below the level of detection or
decline of ≥2 log IU/mL at week 48) was met in 78% (14/18) of participants. Most
participants (15/18, 83%) also had ALT normalization. In another study, two patients with
HDV-related compensated cirrhosis were treated with BLV 10 mg daily for up to 3 years.
Virologic and biochemical responses were maintained, and in one patient, liver function
tests improved, and esophageal varices disappeared. Though HBsAg levels declined, they
did not disappear completely. These data suggest BLV can be used to safely treat HDV
infection in patients with cirrhosis who have contraindications to IFN treatment.
Overall, BLV seems to be well-tolerated and without dose-dependent efficacy on HDV
RNA decline. HBsAg levels are unaffected by BLV monotherapy, but the addition of PEG-
Microorganisms 2024, 12, 2177 9 of 15

IFNa seems to enhance initial HDV RNA decline and decrease HBsAg; this effect may be
due to PEG-IFN enhancing the kinetics of viral decline [75]. A systematic review and meta-
analysis of randomized controlled trials confirms IFN + BLV’s superiority to other HDV
treatment regimens (including BLV alone) in HDV suppression at least 24 weeks after the
end of treatment [67]. In addition, BLV seems to have a high barrier to virologic resistance,
based on post-study analysis of non-responders and those with virologic breakthroughs
after BLV monotherapy in the MYR202 and MYR 301 studies [76].
As monotherapy, the 2 mg dose of BLV has been approved for HDV treatment in
Europe as of July 2023 [23]. In the United States, the FDA granted BLV a breakthrough
therapy and orphan drug designation in October 2022, but it has not been fully FDA-
approved [77]. After European approval, several real-world studies support BLV’s safety
and efficacy. A large, multicenter, real-world cohort of 114 patients in Germany treated
with BLV 2 mg (without IFN) confirmed the efficacy of the drug, with virologic response
in 76% (87/114) of cases and declines in ALT levels [78]. Other real-world studies show
BLV’s safety and efficacy in treating HDV in patients with HIV co-infection and advanced
liver disease, with normalization of ALT between 2 and 6 months of treatment [79–81]. In
addition, economic models suggest that BLV can prevent significantly more liver events
than PEG-IFNa in patients with chronic HDV infection and contribute to cost savings
through reductions in liver complications [82]. BLV is a promising HDV treatment, but a
remaining challenge with BLV treatment is the lack of a decline in HBsAg and identifying
predictors of non-response to treatment; it has been suggested that low body mass index
and high alpha-fetoprotein prior to treatment are possible predictors of delayed treatment
response [81].

6.2.2. Lonafarnib (LNF)

LNF inhibits farnesyl transferase, the enzyme necessary for the prenylation modifi-
cation of L-HDAg, thus preventing the assembly of HDV virions. LNF is an oral capsule
first investigated in a proof-of-concept, double-blind, placebo-controlled study for antiviral
efficacy against HDV [57]. The study randomized 14 participants to receive LNF 100 mg
twice daily (BID) or LNF 200 mg BID for 28 days with 6 months’ follow-up. The primary
efficacy endpoint was a decrease in HDV RNA. There was a mean log HDV RNA decline
of −0.73 log IU/mL in the LNF 100 mg group and a decline of −1.54 log IU/mL in the
LNF 200 mg group. LNF serum concentrations correlated with HDV RNA changes, but
there were no changes in serum HBsAg levels. AEs associated with LNF were mainly mild,
consisting of nausea, diarrhea, abdominal bloating, and weight loss, which were more
profound in the 200 mg group.
LOWR HDV-1 was a single-center, phase II proof-of-concept study exploring optimal
LNF dosing regimens [58]. Fifteen participants received LNF 200 mg or 300 mg BID
for 12 weeks, LNF 100 mg three times daily for 5 weeks, LNF 100 mg BID + PEG-IFNa-
2a 180 mg weekly for 8 weeks, or LNF 100 mg BID + ritonavir (RTV) 100 mg daily for
8 weeks (RTV is an oral protease inhibitor that can increase half-lives of other drugs
due to CYP450 inhibition). The best antiviral responses were seen with the LNF+ PEG-
IFNa combinations. LNF monotherapy caused gastrointestinal AEs, but RTV improved
tolerability. The addition of RTV to LNF also resulted in substantial suppression of HDV
RNA compared to the LNF monotherapy regimens. In two participants who received the
12-week regimens, there was a transient, post-treatment ALT elevation that was associated
with suppression of HDV RNA and subsequent ALT normalization (like that seen with
PEG-IFNa + NA). This was not seen in patients with LNF < 12 weeks and was not due to
HBV reactivation. The LOWR HDV-1 study showed that (1) adding RTV to LNF gave better
antiviral responses and tolerability, (2) PEG-IFNa-2a in combination with LNF was more
effective than PEG-IFNa-2a monotherapy, and (3) transient ALT flaring may be beneficial
and therapeutic.
The LOWR HDV-2 study expanded on the findings of the LOWR HDV-1 study to
investigate varying doses and durations of LNF + RTV [59]. In this single-center, open-label,
Microorganisms 2024, 12, 2177 10 of 15

non-randomized, phase II study, 55 participants were enrolled into three treatment groups
to receive high-dose LNF (LNF ≥ 75 mg BID + RTV for 12 weeks), all-oral low-dose LNF
(LNF 25 or 50 mg BID + RTV for 24 weeks), or combination low-dose LNF with PEG-IFNa
(LNF 25 or 50 mg BID + RTV + PEG-IFNa for 24 weeks). The primary endpoint was ≤2 log
decline or HDV RNA below the lower limit of quantification at the end of treatment. The
primary endpoint was reached in 46% (6/13) and 89% (8/9) of participants with the LNF
50 mg BID + RTV and LNF (25 or 50 mg BID) + RTV+ PEG-IFNa regimens, respectively.
Similar to the LOWR HDV-1 study, multiple patients had post-treatment ALT increases, but
these were well-tolerated and followed by HDV RNA suppression and ALT normalization.
More gastrointestinal AEs were in the high-dose LNF group.
The LNF studies confirmed that the addition of PEG-IFNa enhanced antiviral efficacy.
The LOWR HDV-2 study also found that adding RTV to PEG-IFNa had the most efficacy
and tolerability for participants. A follow-up pharmacokinetic modeling study confirmed
that LNF combined with RTV was the most efficient to achieve antiviral efficacy without
increasing AEs [83]. Though LNF + RTV + PEG-IFNa is well-tolerated and effective, RTV’s
potential to induce drug-drug interactions via CYP450 inhibition may present issues with
concomitant medications. After completion of phase 3 trials with LNF, it was granted
orphan drug designation in the U.S. and Europe. A remaining challenge in LNF’s efficacy is
that it seems limited to suppression of HDV RNA with no to little changes in HBsAg levels.

6.2.3. REP-2139
REP-2139, a nucleic acid polymer, prevents the export of HDV particles. REP-2139
was investigated in the open-label, non-randomized phase II trial called REP 301 [60].
This trial enrolled 12 patients with HBV/HDV co-infection in Moldova. Participants
received 63 weeks of treatment with a 1-year follow-up. The treatment regimen was
REP-2139 500 mg intravenous (IV) weekly for 15 weeks, REP-2139 250 mg IV + 250 mg
PEG-IFNa-2a weekly for 15 weeks, then PEG-IFNa-2a 180 mg monotherapy weekly for
33 weeks. The most common AEs during REP-2139 monotherapy were pyrexia and chills.
However, the safety and tolerability of the combination regimen were similar to PEG-IFNa-
2a monotherapy. A majority (11/12, 92%) of participants had RNA suppression during
treatment, 9 (75%) at the end of treatment, and 7 (58%) at follow-up. Long-term follow-up of
REP 301 assessed safety, tolerability, and HDV functional cure at 3.5 years of follow-up [61].
No abnormal liver or kidney abnormalities were seen, HDV RNA suppression persisted in
7 of 11 participants, and HBsAg seroconversion was seen in 5 participants. More clinical
investigations of this molecule are needed, but it shows promise if it can suppress HDV
RNA and lead to HBsAg loss.

7. Conclusions and Future Directions

BLV, LNF, and REP-2139 studies support their safety and efficacy, especially when
combined with PEG-IFNa, suggesting that magnifying the endogenous immune response
is vital for enhancing viral kinetics and HDV RNA clearance. However, the need for
PEG-IFNa is also a downside due to the adverse effects of this therapy. Moreover, studies
with these antiviral agents have not elucidated a fully reliable virologic endpoint to therapy
since they all used different endpoints, making it difficult to compare their efficacy [12]. To
date, BLV is the only approved HDV-specific antiviral. A challenge remains in finding an
antiviral therapy that suppresses HBsAg, since this seems crucial to persistent suppression
of HDV RNA. For this reason, there may be room for development of other antivirals,
including REP-2139.
In addition to the treatment challenges facing HDV, several challenges remain to
limit HDV infections in the United States (U.S.) and worldwide (Table 4). First, there is a
paucity of data on HDV’s prevalence or genotype distribution, and there are no established
cohorts of patients. Second, clinicians do not routinely screen for HDV since there is a
lack of knowledge regarding who should be screened and which diagnostics are available.
In addition, some clinicians do not screen for HDV since no optimal HDV treatment
Microorganisms 2024, 12, 2177 11 of 15

is available. Third, the currently universally-used treatment—PEG-IFNa—has limited

efficacy. Last, if an antiviral drug is approved, it is unclear if patients would be amenable
to treatments due to drug-drug interactions, the need for daily injections, long durations
of treatment with adverse events, and low potential for cure. Ultimately, these concerns
will need to be addressed to understand the scope of the impact that these developing
therapies have on curing and reducing the significant morbidity and mortality associated
with HDV infection.

Table 4. Future directions for HDV research. HDV = hepatitis D virus.

Area of Interest Research Questions about HDV

How can global HDV prevalence estimates and associations be more accurately defined?
Epidemiology
Why do certain population groups report higher prevalences of HDV?
What is the relevance of genotypes on the natural history of HDV infection?
Genotypes
Do genotypes affect treatment outcomes?
How can the sensitivity and specificity of HDV RNA assays be improved?
RNA measurement and fibrosis
Can numerical cut-offs obtained from transient elastography stage liver fibrosis in
detection
HDV-infected individuals accurately?
What are the immunological mechanisms associated with disease progression and severity?
What are the immune correlates of HDV clearance or functional cure?
Immunological parameters
Are there immunological markers associated with disease severity and functional cure that
can be monitored during HDV treatment?
Can there be a common antiviral that can efficiently inhibit both HBV and HDV infection
and effectively eliminate the chances of viral rebound?
What endpoint should be taken into consideration to stop antiviral treatment without the
Antiviral treatment risk of rebound viremia?
What are the factors contributing to treatment non-response?
Should therapies in development also target the host immune response to control
viral replication?

Author Contributions: Conceptualization P.M. and S.K.; methodology P.M.; writing—original draft
preparation P.M.; writing—reviewing and editing P.M., A.K., and S.K. All authors have read and
agreed to the published version of the manuscript.
Funding: The research received no external funding.
Conflicts of Interest: The authors declare no conflicts of interest.

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