Accepted Article
Received Date : 06-Feb-2014 Accepted Date : 11-May-2014 Article type
: Reviews
Title: Making the Case for the Development of a Vaccination against Hepatitis E Virus
Authors: Samir Haffar, MD1,*; Fateh Bazerbachi, MD2,*; John R. Lake, MD 2, †
1
Department of gastroenterology, Al-Mouassat University Hospital, Damascus, Syria; e-mail:
[email protected] 2
Department of gastroenterology, University of Minnesota, Minneapolis, MN, United States;
e-mail: [email protected] *
These authors contributed equally to this work.
† Corresponding author
Corresponding Author: John R. Lake, MD Professor of Medicine and Surgery 406 Harvard St SE MMC 36, VCR V366 Minneapolis, MN 55455 Email: [email protected] The authors declare no conflict of interest, and deny any financial support to this review
Abstract Hepatitis E virus (HEV) infection is a global problem that affects 20 million individuals, and cause acute hepatitis in 3.5 million, with approximately 70,000 deaths worldwide. This article has been accepted for publication and undergone full peer review but has not been through the copyediting, typesetting, pagination and proofreading process, which may lead to differences between this version and the Version of Record. Please cite this article as doi: 10.1111/liv.12590 This article is protected by copyright. All rights reserved.
Accepted Article
While the acute disease is generally self-limited, however, it may progress to fatal fulminant liver failure in certain individuals. Contaminated water supplies disseminate this virus through the fecal-oral route, and swine is thought to be its zoonotic reservoir. Attempts have been made to develop effective HEV vaccines, and two candidates have undergone successful clinical trials. In this review, we discuss HEV epidemiology, genotypes, microbiological structure, as well as the most recent advances in vaccination developments. Keywords: Hepatitis E virus (HEV); HEV Vaccine; Vaccination; Viral hepatitis; Primary prevention
Introduction: Hepatitis E virus (HEV) is found primarily in developing regions of the world, mainly, India, Africa, Asia, and the Middle East. It is a waterborne, enteric virus, and is acquired from contaminated water supplies (1). Although most cases of HEV infection cause a mild clinical illness, some infections lead to acute liver failure, with a mortality rate between 4-30%. The highest mortality occur in pregnant women. In non-endemic areas of the world, the prevalence of antibodies to HEV can be as high as 20%, although clinically significant disease is very uncommon. Although HEV is thought to have a zoonotic reservoir, primarily in swine, nonetheless, imported cases from endemic areas, through immigrants and refugees, are observed.
Why does the world need a vaccine for hepatitis E?
HEV is a major cause of acute hepatitis globally, but it has been neglected over the years due to the lack of a readily available, validated assays for HEV RNA. It is now estimated
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that as high as one third of the population in some developing countries has been infected with HEV (2).
In endemic areas, the overall mortality rate of acute HEV infection is 1–3%, and death occurs in up to 30% in pregnant women (3, 4). Several epidemics of HEV infections have been reported in Asia and Africa, with the most recent in Uganda between 2007 and 2010 (5).
In patients with chronic liver disease, HEV infection have a mortality rate as high as 70% (2). Moreover, HEV-associated chronic liver disease has been described recently among organ transplant recipients, and HIV- infected and other immunocompromised individuals (6).
In 2005, Rein et al. estimated the burden of HEV genotypes 1 and 2 in 9 out of 21 world health regions, which represents 71% of the world’s population, to be 20 million cases per year, with 70,000 deaths and 3,000 stillbirths (7). In contrast to hepatitis A, an estimation of the incidence of hepatitis E disease in developed countries has not yet been performed.
The Virus
HEV was identified as a distinct viral agent in 1983 and was subsequently cloned and sequenced in 1991 (8, 9). It is a spherical, non-enveloped RNA virus, with a diameter of 2734 nanometers, and is the only member of the Hepeviridae family.
The HEV genome is comprised of a single-stranded positive-sense RNA of approximately 7.2 kilobasepairs (figure 1). It consists of short 5′ and 3′ untranslated regions (UTRs), and three partially overlapping open reading frames (ORF), called ORF1, ORF2 and ORF3, that
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encode structural and non-structural proteins(10). ORF1 encodes a polypeptide with nonenzymatic activities required for viral replication. ORF2 encodes the capsid protein which is involved in particle assembly into virus-like or subviral particles and includes neutralizing epitopes. This protein is also crucial for binding to host cells and eliciting neutralizing and protective antibodies during preclinical studies (11). ORF3 overlaps the capsid gene, and appears to be necessary for cellular egress.
Assembly of virions begins with the production of capsid monomers (with or without an Nterminal region), which self-assemble into dimers and subsequently decamers (12, 13). Decamers without the capsid N-terminal assemble into small virus like particles (VLP) that are the source of HEV vaccines and serologic reagents. Decamers of full-length capsid monomers form the full-size virions (figure 2).
HEV has four major genotypes. These genotypes fall into two major groups (table 1). Genotypes 1 and 2 are human viruses, causing epidemic acute hepatitis in developing countries, and are transmitted orally through contact with infected water sources.
Genotypes 3 and 4 are swine viruses that are common in domestic and wild pigs in developed countries. As humans are affected as accidental hosts, they are considered zoonoses. They can cause acute sporadic hepatitis and sometimes chronic hepatitis in immunocompromised patients. All four genotypes are believed to represent a single serotype which has facilitated efforts to develop the hepatitis E vaccines (14).
The capsid protein encoded by ORF2 of HEV is a 72 kiloDalton (kDa) protein (15). It is composed of 660-amino acid and has an N-terminal signal sequence (SS), with three putative
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domains (figure 3). Domain 1 (~100 residues) has a high density of arginine residues, and is likely to be involved in RNA encapsidation. Domain 2 (~240 residues) is predicted to form the core of the viral capsid. Domain 3 (~300 residues) is predicted to be exposed on the virus particle. A neutralization epitope (amino acids 458 – 607) is localized to domain 3 and is likely to be involved in the binding of HEV to its cellular receptor (16). Antibodies against ORF2 in humans and infected animals are long lived, cross-reactive among diverse HEV genotypes, and neutralizes HEV in vitro. This is the reason why ORF2 protein has been the antigen used in all vaccine studies, thus far (14).
Clinical studies of HEV vaccine candidates: Full-length or truncated forms of the ORF2 protein expressed in bacterial, insect, yeast, animal and plant cells have emerged as potential HEV vaccine candidates (Table 2). Two of these have already been shown to be efficacious in phase II/III clinical trials (17, 18). An alternative approach is a DNA vaccine administered either alone or with the addition of protein to augment the immune response. The advantage of DNA vaccination is its stability and ease of preparation but at this point, it is still in the preclinical stage (19).
Clinical studies of HEV vaccine candidates: Two candidates for hepatitis E vaccines have been investigated in clinical trials (Table 3). First, a 56 kDa truncated HEV ORF2 protein has been produced from a recombinant baculovirus that forms virus-like particles. This vaccine has undergone safety and efficacy studies in humans. In a randomized, double-blind, placebo controlled, phase II trial in Nepal sponsored by SmithKline Beecham Biologicals (now Glaxo SmithKline: GSK), anti-HEV negative healthy adults from the Nepalese Army were randomized to receive either three doses of 20 μg of 56 kDa vaccine or placebo at 0, 1, and 6 months, and were followed for an
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average of 804 days. The vaccine was well-tolerated and highly immunogenic, with 95.5% of the study subjects acquiring significant antibody titer (95% CI: 85.6–98.6) (17).
This study had several limitations. First, virtually all the study subjects were young men (mean age 25 years). Second, the study focused on clinical disease rates and did not look at the HEV infection rate. Third, anti-HEV antibodies titer had declined significantly by the end of the study, so that nearly 44% of subjects had antibody titres below what is considered protective (19).
Nonetheless, the publication of this first successful clinical trial of an efficacious HEV vaccine against generated enthusiasm and optimism. However, this vaccine has not reached the market because of concerns regarding its ability to generate sufficient revenue.
Second, Chinese company (Xiamen Innovax Biotech) has developed another hepatitis E vaccine, dubbed the HEV 239 vaccine. HEV 239 is a 26 kDa protein encoded by ORF2 of HEV1, expressed in Escherichia coli, and presents as virus-like particles of 23 nm in diameter.
In a large scale, randomized, double-blind, placebo controlled, phase III trial in 11 townships in eastern China, 112,604 healthy men and women, aged 16–65 years, were randomized to receive either three intramuscular injections of 30 μg of HEV 239 at 0, 1, and 6 months or hepatitis B vaccine as a placebo and were followed for 13-months after the third vaccine dose (18). The vaccine was well tolerated and protected against hepatitis E, with an efficacy of 100% after the second and third dose (95% CI: 72.1–100.0). No vaccine-related adverse events were documented. The predominance of HEV genotype 4 in this region demonstrated
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the cross-protective efficacy of the HEV 239 vaccine, which is based on genotype 1. However, the efficacy of this vaccine against genotype 3 is yet unknown(20).
The study endpoint was clinical disease rate rather than HEV infection rate. Moreover, the persistence of the protection conferred by the vaccine was not studied. The study did not assess the safety of the vaccine in pregnant women, people younger than 15 years or individuals older than 65 years, or in chronic liver disease patients.
In a more recent study, Zhang et al. showed that, although naturally occurring immunity against HEV lowered the subsequent infection risk significantly (to 0.52%), the administration of Hecolin® provided greater protection with an ensuing HEV infection risk of 0.3% (21). In December 2011, the China Food & Drug Administration approved the hepatitis E vaccine Hecolin® for use in subjects older than 16 years (22). The vaccine will be sold in China at a cost of 110 Renminbi (~$17.6 US dollars) per dose (23). It is unclear whether it will be developed in other countries.
Who should receive the vaccine?
HEV vaccination would be most used in developing countries where the virus is endemic, especially in high risk groups such as pregnant women in the second and third trimesters, the elderly, and children less than 2 years of age. The vaccine may also have a role in HEV outbreaks. Since vaccine efficacy after two doses is nearly 100%, protection occurs quickly and can be obtained by two doses given within 1 month (19).
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In developed countries, vaccination may also be useful in high-risk groups such as food industry workers, immunosuppressed patients and those with chronic liver disease, as well as travelers to endemic areas. In some studies, hepatitis E infection may portend deleterious consequences for patients with chronic liver disease, and mortality in this subpopulation may be as high as 75% (24, 25). Although HEV genotype 1 comprises the majority of these infections, genotype 3 in developed countries has been shown to cause hepatic decompensation portending a worse prognosis. Immunosuppressed patients are also potential candidates (e.g. chemotherapy patients, transplant recipients, HIV infected individuals) because HEV infection in these patients could lead to chronic liver damage(26). Porcine vaccination may also be useful if it is cost effective (27).
Areas for future research:
Further research is needed to determine the titer of antibodies required for protection, the duration of protection afforded by the vaccine, the need for vaccine booster, and whether the vaccine is effective, not only against overt clinical illness, but also against all HEV infections.
Mitigating clinical infection and symptomatic disease, and decreasing the development of liver failure may be advantageous, even in the absence of infection prevention (19). Moreover, studies are needed to determine whether the vaccine would be useful for postexposure prophylaxis, such as in cases of HEV outbreaks. Additionally, despite the serologic cross-reactivity, the efficacy of the vaccine has not been studied in regions of the world with
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HEV genotype 3, which has emerged as the predominant HEV strain in Europe and North America. Additional data is needed regarding the safety and efficacy of this vaccine in individuals at extremes of age (younger than 2 years and older than 65 years), in pregnant women, and patients with chronic liver disease, as well as in immunosuppressed patients. Preliminary evidence showed safety of the Hecolin® vaccine in pregnant women; 37 women who received 1, 2 or 3 doses of the vaccine turned out to be pregnant at time of administration. The vaccine was well tolerated, and the delivered babies were healthy (22). These findings should be confirmed in larger studies. Cost effectiveness (C/E) of the vaccine should also be explored. C/E of an intervention is defined as the additional cost required per additional unit of health benefit produced as compared with the next-most effective alternative; this must be distinguished from economic affordability, as they are not mutually exclusive(28). For vaccines, C/E is dictated by several factors, including severity of disease, vaccine price, durability, and efficiency.
Das et al. (29) constructed a Markov model using decision analysis techniques to evaluate three different strategies regarding the early form of HEV vaccination in a Monte Carlo simulation cohort of 10,000 healthy young individuals living in endemic areas: A- Universal vaccination, B- Screening and vaccination, and C- No vaccination. The cost of illness consisted of the direct cost of treatment, as well as the indirect cost of lost workdays. Vaccine-induced immunity was presumed to last at least 8 years. Outcome parameters consisted of the incremental cost-effectiveness ratio, as well as the discounted cost per patient and quality adjusted life years (QALY) gained in each strategy. The cost-effectiveness (C/E) ratio for plans A, B, and C was $0.33, $0.50, and $0.53, respectively. The authors estimated the cost of vaccine to be $ 9.86 which is almost half of the anticipated price of Hecolin®(30).
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Based on the previous data, it seems that universal vaccination against hepatitis E would be highly cost-effective with a marginal cost-effectiveness ratio below the currently available viral hepatitis vaccines, however, a Markov model analysis based on Hecolin® is needed, when additional data become available (31).
Conclusion Too often, the decision to develop a drug, or in this case a vaccine, is profit driven. Such profits are generally generated in the West and not in developing countries. However, our medical ethics should also motivate us to develop therapies for diseases which wreak havoc in developing countries, but not in the West.
Figures legends
Figure 1. Genome of the hepatitis E virus HEV is an RNA virus with three open reading frames encoding structural and non-structural proteins. NTR: non-translated region. Modified from Kumar et al (32)
Figure 2. Assembly of virions of HEV with or without an N-terminal region. Modified from Xing et al and Hoofnagle et al (12, 13)
Figure 3. Genomic organization of ORF-2 of HEV Modified from Aggarwal et al (15)
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Tables Table 1: Genotype characteristics of HEV infections
Geographis distribution Pattern of spread Species Mode of spread Icteric illness Age distribution Sex distribution Mortality Extrahepatic features Chronic infection
Genotypes 1 and 2 Epidemic Developing countries Epidemic and sporadic Human Fecal–oral, waterborne Common Adolescents and young adults Similar in men and women High in pregnant women Few None
Therapy None known Prevention Vaccine Modified from Acharya et al and Hoofnagle et al (13, 33)
Genotypes 3 and 4 Autochthonous Developing and developed countries Sporadic Swine, human Foodborne uncommon Older aduts Higher in men High in older adults Neurologic complications Common in immunocompromised persons Ribavirin, peginterferon Vaccine
Table 2: Pre-clinical evaluation of hepatitis E vaccine candidates (14, 19) Proteins expressed in E. coli
Proteins expressed in insect cells Baculovirus-mediated
Spodopteralitura larvae Proteins expressed in other milieus Yeast Pichia pastoris Transgenic tomato plants DNA vaccines Naked
DNA plus protein VLPs: Virus-Like-Particles
ORF-2 protein TrpE-C2
Amino acids 221 – 660
Source Burma
pE2 HEV 239
394 – 607 368 – 606
China China
56-kDa protein 53-kDa protein 62-kDa 50 kDa (VLPs) 62-kDa
112 – 607 112 – 577 112 – 660 112 – 534 112 – 660
Pakistan Burma Burma Burma India
HBV/HEV pE2
551 – 607 394 – 607
China China
1 – 660 1 – 660 1 – 660
Burma India Burma India
pJHEV pcDNA-ORF2 pcHEVORF2 +26 kDa
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Remarks
Human clinical trials Human clinical trials
Oral route
Stability Ease of preparation
Accepted Article
Table 3: Phase II/III trials of HEV vaccine Shrestha et al
Zhu et al
NEJM – March 2007
Lancet – August 2010
Phase II trial
Phase III trial
Randomized, double-blind,
Randomized, double-blind,
placebo controlled
placebo controlled
Company
GlaxoSmithKline Biologicals
Xiamen Innovax Biotech
Country
Nepal
Jiangsu province, China
Baculovirus expressed 56 kDa
E. coli expressed HEV 239
1794 healthy subjects
112 604 healthy subjects
HEV vaccine versus placebo
HEV versus HBV vaccine
Mostly males ( 99.6%)
males and females, 16 – 65 years
Phase study Type of study
Recombinant protein ORF2 Number of vaccinees Randomization Population
Young (mean age 25 years)
HEV genotypes
genotype 1 prevalent
Genotypes 1 and 4 prevalent with predominance of genotype 4
20 μg
30 μg
Route of administration
Intramuscularly
Intramuscularly
Intervals between doses
0, 1, 6 months
0, 1, 6 months
Prevention of clinically overt
Prevention of clinically overt
HEV disease
HEV disease
2 years post-vaccination
13 month post-vaccination
87.5%
95.5%
After 2nd dose
85.7%
100%
After 3rd dose
95,5%
100%
Increased injection-site pain
No side effect
Not further developed
Hecolin® (Innovax)
Doses
Primary end-point
Follow-up period Efficacy: After 1st dose
Side effects Commercialization
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References 1. KRAWCZYNSKI K. Hepatitis E. Hepatology 1993; 17(5): 932-41. 2. Hepatitis E vaccine: why wait? Lancet 2010; 376(9744): 845. 3. KHUROO M S, KAMILI S. Aetiology, clinical course and outcome of sporadic acute viral hepatitis in pregnancy. Journal of viral hepatitis 2003; 10(1): 61-9. 4. KHUROO M S. Discovery of hepatitis E: the epidemic non-A, non-B hepatitis 30 years down the memory lane. Virus research 2011; 161(1): 3-14. 5. TESHALE E H, HOWARD C M, GRYTDAL S P, et al. Hepatitis E epidemic, Uganda. Emerging infectious diseases 2010; 16(1): 126-9. 6. MIYAMURA T. Hepatitis E virus infection in developed countries. Virus research 2011; 161(1): 40-6. 7. REIN D B, STEVENS G A, THEAKER J, WITTENBORN J S, WIERSMA S T. The global burden of hepatitis E virus genotypes 1 and 2 in 2005. Hepatology 2012; 55(4): 988-97. 8. BALAYAN M S, ANDJAPARIDZE A G, SAVINSKAYA S S, et al. Evidence for a virus in non-A, nonB hepatitis transmitted via the fecal-oral route. Intervirology 1983; 20(1): 23-31. 9. REYES G R, YARBOUGH P O, TAM A W, et al. Hepatitis E virus (HEV): the novel agent responsible for enterically transmitted non-A, non-B hepatitis. Gastroenterologia Japonica 1991; 26 Suppl 3: 142-7. 10. KRAWCZYNSKI K. Hepatitis E vaccine--ready for prime time? The New England journal of medicine 2007; 356(9): 949-51. 11. ZHANG J, GU Y, GE S X, et al. Analysis of hepatitis E virus neutralization sites using monoclonal antibodies directed against a virus capsid protein. Vaccine 2005; 23(22): 2881-92. 12. XING L, LI T C, MAYAZAKI N, et al. Structure of hepatitis E virion-sized particle reveals an RNA-dependent viral assembly pathway. The Journal of biological chemistry 2010; 285(43): 3317583. 13. HOOFNAGLE J H, NELSON K E, PURCELL R H. Hepatitis E. The New England journal of medicine 2012; 367(13): 1237-44. 14. KAMILI S. Toward the development of a hepatitis E vaccine. Virus research 2011; 161(1): 93100. 15. AGGARWAL R, JAMEEL S. Hepatitis E vaccine. Hepatology international 2008; 2(3): 308-15. 16. HOLLA R P, AHMAD I, AHMAD Z, JAMEEL S. Molecular virology of hepatitis E virus. Semin Liver Dis 2013; 33(1): 3-14. 17. SHRESTHA M P, SCOTT R M, JOSHI D M, et al. Safety and efficacy of a recombinant hepatitis E vaccine. The New England journal of medicine 2007; 356(9): 895-903. 18. ZHU F C, ZHANG J, ZHANG X F, et al. Efficacy and safety of a recombinant hepatitis E vaccine in healthy adults: a large-scale, randomised, double-blind placebo-controlled, phase 3 trial. Lancet 2010; 376(9744): 895-902. 19. THOMAS D. BOYER M P M, ARUN J. SANYAL. Zakim and Boyer's Hepatology: A Textbook of Liver Disease. 6th ed: Saunders; 2011. 20. RIVEIRO-BARCIELA M, RODRIGUEZ-FRIAS F, BUTI M. Hepatitis E virus: new faces of an old infection. Annals of hepatology 2013; 12(6): 861-70. 21. ZHANG J, ZHANG X F, ZHOU C, et al. Protection against hepatitis E virus infection by naturally acquired and vaccine induced immunity. Clin Microbiol Infect 2013. 22. WU T, LI S W, ZHANG J, NG M H, XIA N S, ZHAO Q. Hepatitis E vaccine development: a 14 year odyssey. Human vaccines & immunotherapeutics 2012; 8(6): 823-7. 23. RIEDMANN E M. Chinese biotech partnership brings first hepatitis E vaccine to the market. Human vaccines & immunotherapeutics 2012; 8(12): 1743-4. 24. RAMACHANDRAN J, EAPEN C E, KANG G, et al. Hepatitis E superinfection produces severe decompensation in patients with chronic liver disease. Journal of gastroenterology and hepatology 2004; 19(2): 134-8.
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25. KUMAR ACHARYA S, KUMAR SHARMA P, SINGH R, et al. Hepatitis E virus (HEV) infection in patients with cirrhosis is associated with rapid decompensation and death. J Hepatol 2007; 46(3): 387-94. 26. DALTON H R, HAZELDINE S, BANKS M, IJAZ S, BENDALL R. Locally acquired hepatitis E in chronic liver disease. Lancet 2007; 369(9569): 1260. 27. KAMAR N, BENDALL R, LEGRAND-ABRAVANEL F, et al. Hepatitis E. Lancet 2012; 379(9835): 2477-88. 28. KIM J J. The role of cost-effectiveness in U.S. vaccination policy. The New England journal of medicine 2011; 365(19): 1760-1. 29. DAS A, SINHA M. A cost-effectiveness analysis of a candidate vaccine against hepatitis E virus using the institute of medicine model. The American journal of gastroenterology 2001; 96(9, Supplement 1): S263. 30. RIEDMANN E M. Human Vaccines & Immunotherapeutics: News. Human vaccines & immunotherapeutics 2013; 9(10). 31. WIERZBA T F, PANZNER U. Report on the International Symposium on Hepatitis E, Seoul, South Korea, 2010. Emerging infectious diseases 2012; 18(5). 32. KUMAR S, SUBHADRA S, SINGH B, PANDA B K. Hepatitis E virus: the current scenario. International journal of infectious diseases : IJID : official publication of the International Society for Infectious Diseases 2013; 17(4): e228-33. 33. ACHARYA S K, PANDA S K. Hepatitis E: water, water everywhere - now a global disease. J Hepatol 2011; 54(1): 9-11.
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Accepted Article This article is protected by copyright. All rights reserved.
Received Date : 06-Feb-2014 Accepted Date : 11-May-2014 Article type
: Reviews
Title: Making the Case for the Development of a Vaccination against Hepatitis E Virus
Authors: Samir Haffar, MD1,*; Fateh Bazerbachi, MD2,*; John R. Lake, MD 2, †
1
Department of gastroenterology, Al-Mouassat University Hospital, Damascus, Syria; e-mail:
[email protected] 2
Department of gastroenterology, University of Minnesota, Minneapolis, MN, United States;
e-mail: [email protected] *
These authors contributed equally to this work.
† Corresponding author
Corresponding Author: John R. Lake, MD Professor of Medicine and Surgery 406 Harvard St SE MMC 36, VCR V366 Minneapolis, MN 55455 Email: [email protected] The authors declare no conflict of interest, and deny any financial support to this review
Abstract Hepatitis E virus (HEV) infection is a global problem that affects 20 million individuals, and cause acute hepatitis in 3.5 million, with approximately 70,000 deaths worldwide. This article has been accepted for publication and undergone full peer review but has not been through the copyediting, typesetting, pagination and proofreading process, which may lead to differences between this version and the Version of Record. Please cite this article as doi: 10.1111/liv.12590 This article is protected by copyright. All rights reserved.
Accepted Article
While the acute disease is generally self-limited, however, it may progress to fatal fulminant liver failure in certain individuals. Contaminated water supplies disseminate this virus through the fecal-oral route, and swine is thought to be its zoonotic reservoir. Attempts have been made to develop effective HEV vaccines, and two candidates have undergone successful clinical trials. In this review, we discuss HEV epidemiology, genotypes, microbiological structure, as well as the most recent advances in vaccination developments. Keywords: Hepatitis E virus (HEV); HEV Vaccine; Vaccination; Viral hepatitis; Primary prevention
Introduction: Hepatitis E virus (HEV) is found primarily in developing regions of the world, mainly, India, Africa, Asia, and the Middle East. It is a waterborne, enteric virus, and is acquired from contaminated water supplies (1). Although most cases of HEV infection cause a mild clinical illness, some infections lead to acute liver failure, with a mortality rate between 4-30%. The highest mortality occur in pregnant women. In non-endemic areas of the world, the prevalence of antibodies to HEV can be as high as 20%, although clinically significant disease is very uncommon. Although HEV is thought to have a zoonotic reservoir, primarily in swine, nonetheless, imported cases from endemic areas, through immigrants and refugees, are observed.
Why does the world need a vaccine for hepatitis E?
HEV is a major cause of acute hepatitis globally, but it has been neglected over the years due to the lack of a readily available, validated assays for HEV RNA. It is now estimated
This article is protected by copyright. All rights reserved.
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that as high as one third of the population in some developing countries has been infected with HEV (2).
In endemic areas, the overall mortality rate of acute HEV infection is 1–3%, and death occurs in up to 30% in pregnant women (3, 4). Several epidemics of HEV infections have been reported in Asia and Africa, with the most recent in Uganda between 2007 and 2010 (5).
In patients with chronic liver disease, HEV infection have a mortality rate as high as 70% (2). Moreover, HEV-associated chronic liver disease has been described recently among organ transplant recipients, and HIV- infected and other immunocompromised individuals (6).
In 2005, Rein et al. estimated the burden of HEV genotypes 1 and 2 in 9 out of 21 world health regions, which represents 71% of the world’s population, to be 20 million cases per year, with 70,000 deaths and 3,000 stillbirths (7). In contrast to hepatitis A, an estimation of the incidence of hepatitis E disease in developed countries has not yet been performed.
The Virus
HEV was identified as a distinct viral agent in 1983 and was subsequently cloned and sequenced in 1991 (8, 9). It is a spherical, non-enveloped RNA virus, with a diameter of 2734 nanometers, and is the only member of the Hepeviridae family.
The HEV genome is comprised of a single-stranded positive-sense RNA of approximately 7.2 kilobasepairs (figure 1). It consists of short 5′ and 3′ untranslated regions (UTRs), and three partially overlapping open reading frames (ORF), called ORF1, ORF2 and ORF3, that
This article is protected by copyright. All rights reserved.
Accepted Article
encode structural and non-structural proteins(10). ORF1 encodes a polypeptide with nonenzymatic activities required for viral replication. ORF2 encodes the capsid protein which is involved in particle assembly into virus-like or subviral particles and includes neutralizing epitopes. This protein is also crucial for binding to host cells and eliciting neutralizing and protective antibodies during preclinical studies (11). ORF3 overlaps the capsid gene, and appears to be necessary for cellular egress.
Assembly of virions begins with the production of capsid monomers (with or without an Nterminal region), which self-assemble into dimers and subsequently decamers (12, 13). Decamers without the capsid N-terminal assemble into small virus like particles (VLP) that are the source of HEV vaccines and serologic reagents. Decamers of full-length capsid monomers form the full-size virions (figure 2).
HEV has four major genotypes. These genotypes fall into two major groups (table 1). Genotypes 1 and 2 are human viruses, causing epidemic acute hepatitis in developing countries, and are transmitted orally through contact with infected water sources.
Genotypes 3 and 4 are swine viruses that are common in domestic and wild pigs in developed countries. As humans are affected as accidental hosts, they are considered zoonoses. They can cause acute sporadic hepatitis and sometimes chronic hepatitis in immunocompromised patients. All four genotypes are believed to represent a single serotype which has facilitated efforts to develop the hepatitis E vaccines (14).
The capsid protein encoded by ORF2 of HEV is a 72 kiloDalton (kDa) protein (15). It is composed of 660-amino acid and has an N-terminal signal sequence (SS), with three putative
This article is protected by copyright. All rights reserved.
Accepted Article
domains (figure 3). Domain 1 (~100 residues) has a high density of arginine residues, and is likely to be involved in RNA encapsidation. Domain 2 (~240 residues) is predicted to form the core of the viral capsid. Domain 3 (~300 residues) is predicted to be exposed on the virus particle. A neutralization epitope (amino acids 458 – 607) is localized to domain 3 and is likely to be involved in the binding of HEV to its cellular receptor (16). Antibodies against ORF2 in humans and infected animals are long lived, cross-reactive among diverse HEV genotypes, and neutralizes HEV in vitro. This is the reason why ORF2 protein has been the antigen used in all vaccine studies, thus far (14).
Clinical studies of HEV vaccine candidates: Full-length or truncated forms of the ORF2 protein expressed in bacterial, insect, yeast, animal and plant cells have emerged as potential HEV vaccine candidates (Table 2). Two of these have already been shown to be efficacious in phase II/III clinical trials (17, 18). An alternative approach is a DNA vaccine administered either alone or with the addition of protein to augment the immune response. The advantage of DNA vaccination is its stability and ease of preparation but at this point, it is still in the preclinical stage (19).
Clinical studies of HEV vaccine candidates: Two candidates for hepatitis E vaccines have been investigated in clinical trials (Table 3). First, a 56 kDa truncated HEV ORF2 protein has been produced from a recombinant baculovirus that forms virus-like particles. This vaccine has undergone safety and efficacy studies in humans. In a randomized, double-blind, placebo controlled, phase II trial in Nepal sponsored by SmithKline Beecham Biologicals (now Glaxo SmithKline: GSK), anti-HEV negative healthy adults from the Nepalese Army were randomized to receive either three doses of 20 μg of 56 kDa vaccine or placebo at 0, 1, and 6 months, and were followed for an
This article is protected by copyright. All rights reserved.
Accepted Article
average of 804 days. The vaccine was well-tolerated and highly immunogenic, with 95.5% of the study subjects acquiring significant antibody titer (95% CI: 85.6–98.6) (17).
This study had several limitations. First, virtually all the study subjects were young men (mean age 25 years). Second, the study focused on clinical disease rates and did not look at the HEV infection rate. Third, anti-HEV antibodies titer had declined significantly by the end of the study, so that nearly 44% of subjects had antibody titres below what is considered protective (19).
Nonetheless, the publication of this first successful clinical trial of an efficacious HEV vaccine against generated enthusiasm and optimism. However, this vaccine has not reached the market because of concerns regarding its ability to generate sufficient revenue.
Second, Chinese company (Xiamen Innovax Biotech) has developed another hepatitis E vaccine, dubbed the HEV 239 vaccine. HEV 239 is a 26 kDa protein encoded by ORF2 of HEV1, expressed in Escherichia coli, and presents as virus-like particles of 23 nm in diameter.
In a large scale, randomized, double-blind, placebo controlled, phase III trial in 11 townships in eastern China, 112,604 healthy men and women, aged 16–65 years, were randomized to receive either three intramuscular injections of 30 μg of HEV 239 at 0, 1, and 6 months or hepatitis B vaccine as a placebo and were followed for 13-months after the third vaccine dose (18). The vaccine was well tolerated and protected against hepatitis E, with an efficacy of 100% after the second and third dose (95% CI: 72.1–100.0). No vaccine-related adverse events were documented. The predominance of HEV genotype 4 in this region demonstrated
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the cross-protective efficacy of the HEV 239 vaccine, which is based on genotype 1. However, the efficacy of this vaccine against genotype 3 is yet unknown(20).
The study endpoint was clinical disease rate rather than HEV infection rate. Moreover, the persistence of the protection conferred by the vaccine was not studied. The study did not assess the safety of the vaccine in pregnant women, people younger than 15 years or individuals older than 65 years, or in chronic liver disease patients.
In a more recent study, Zhang et al. showed that, although naturally occurring immunity against HEV lowered the subsequent infection risk significantly (to 0.52%), the administration of Hecolin® provided greater protection with an ensuing HEV infection risk of 0.3% (21). In December 2011, the China Food & Drug Administration approved the hepatitis E vaccine Hecolin® for use in subjects older than 16 years (22). The vaccine will be sold in China at a cost of 110 Renminbi (~$17.6 US dollars) per dose (23). It is unclear whether it will be developed in other countries.
Who should receive the vaccine?
HEV vaccination would be most used in developing countries where the virus is endemic, especially in high risk groups such as pregnant women in the second and third trimesters, the elderly, and children less than 2 years of age. The vaccine may also have a role in HEV outbreaks. Since vaccine efficacy after two doses is nearly 100%, protection occurs quickly and can be obtained by two doses given within 1 month (19).
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In developed countries, vaccination may also be useful in high-risk groups such as food industry workers, immunosuppressed patients and those with chronic liver disease, as well as travelers to endemic areas. In some studies, hepatitis E infection may portend deleterious consequences for patients with chronic liver disease, and mortality in this subpopulation may be as high as 75% (24, 25). Although HEV genotype 1 comprises the majority of these infections, genotype 3 in developed countries has been shown to cause hepatic decompensation portending a worse prognosis. Immunosuppressed patients are also potential candidates (e.g. chemotherapy patients, transplant recipients, HIV infected individuals) because HEV infection in these patients could lead to chronic liver damage(26). Porcine vaccination may also be useful if it is cost effective (27).
Areas for future research:
Further research is needed to determine the titer of antibodies required for protection, the duration of protection afforded by the vaccine, the need for vaccine booster, and whether the vaccine is effective, not only against overt clinical illness, but also against all HEV infections.
Mitigating clinical infection and symptomatic disease, and decreasing the development of liver failure may be advantageous, even in the absence of infection prevention (19). Moreover, studies are needed to determine whether the vaccine would be useful for postexposure prophylaxis, such as in cases of HEV outbreaks. Additionally, despite the serologic cross-reactivity, the efficacy of the vaccine has not been studied in regions of the world with
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HEV genotype 3, which has emerged as the predominant HEV strain in Europe and North America. Additional data is needed regarding the safety and efficacy of this vaccine in individuals at extremes of age (younger than 2 years and older than 65 years), in pregnant women, and patients with chronic liver disease, as well as in immunosuppressed patients. Preliminary evidence showed safety of the Hecolin® vaccine in pregnant women; 37 women who received 1, 2 or 3 doses of the vaccine turned out to be pregnant at time of administration. The vaccine was well tolerated, and the delivered babies were healthy (22). These findings should be confirmed in larger studies. Cost effectiveness (C/E) of the vaccine should also be explored. C/E of an intervention is defined as the additional cost required per additional unit of health benefit produced as compared with the next-most effective alternative; this must be distinguished from economic affordability, as they are not mutually exclusive(28). For vaccines, C/E is dictated by several factors, including severity of disease, vaccine price, durability, and efficiency.
Das et al. (29) constructed a Markov model using decision analysis techniques to evaluate three different strategies regarding the early form of HEV vaccination in a Monte Carlo simulation cohort of 10,000 healthy young individuals living in endemic areas: A- Universal vaccination, B- Screening and vaccination, and C- No vaccination. The cost of illness consisted of the direct cost of treatment, as well as the indirect cost of lost workdays. Vaccine-induced immunity was presumed to last at least 8 years. Outcome parameters consisted of the incremental cost-effectiveness ratio, as well as the discounted cost per patient and quality adjusted life years (QALY) gained in each strategy. The cost-effectiveness (C/E) ratio for plans A, B, and C was $0.33, $0.50, and $0.53, respectively. The authors estimated the cost of vaccine to be $ 9.86 which is almost half of the anticipated price of Hecolin®(30).
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Based on the previous data, it seems that universal vaccination against hepatitis E would be highly cost-effective with a marginal cost-effectiveness ratio below the currently available viral hepatitis vaccines, however, a Markov model analysis based on Hecolin® is needed, when additional data become available (31).
Conclusion Too often, the decision to develop a drug, or in this case a vaccine, is profit driven. Such profits are generally generated in the West and not in developing countries. However, our medical ethics should also motivate us to develop therapies for diseases which wreak havoc in developing countries, but not in the West.
Figures legends
Figure 1. Genome of the hepatitis E virus HEV is an RNA virus with three open reading frames encoding structural and non-structural proteins. NTR: non-translated region. Modified from Kumar et al (32)
Figure 2. Assembly of virions of HEV with or without an N-terminal region. Modified from Xing et al and Hoofnagle et al (12, 13)
Figure 3. Genomic organization of ORF-2 of HEV Modified from Aggarwal et al (15)
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Tables Table 1: Genotype characteristics of HEV infections
Geographis distribution Pattern of spread Species Mode of spread Icteric illness Age distribution Sex distribution Mortality Extrahepatic features Chronic infection
Genotypes 1 and 2 Epidemic Developing countries Epidemic and sporadic Human Fecal–oral, waterborne Common Adolescents and young adults Similar in men and women High in pregnant women Few None
Therapy None known Prevention Vaccine Modified from Acharya et al and Hoofnagle et al (13, 33)
Genotypes 3 and 4 Autochthonous Developing and developed countries Sporadic Swine, human Foodborne uncommon Older aduts Higher in men High in older adults Neurologic complications Common in immunocompromised persons Ribavirin, peginterferon Vaccine
Table 2: Pre-clinical evaluation of hepatitis E vaccine candidates (14, 19) Proteins expressed in E. coli
Proteins expressed in insect cells Baculovirus-mediated
Spodopteralitura larvae Proteins expressed in other milieus Yeast Pichia pastoris Transgenic tomato plants DNA vaccines Naked
DNA plus protein VLPs: Virus-Like-Particles
ORF-2 protein TrpE-C2
Amino acids 221 – 660
Source Burma
pE2 HEV 239
394 – 607 368 – 606
China China
56-kDa protein 53-kDa protein 62-kDa 50 kDa (VLPs) 62-kDa
112 – 607 112 – 577 112 – 660 112 – 534 112 – 660
Pakistan Burma Burma Burma India
HBV/HEV pE2
551 – 607 394 – 607
China China
1 – 660 1 – 660 1 – 660
Burma India Burma India
pJHEV pcDNA-ORF2 pcHEVORF2 +26 kDa
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Remarks
Human clinical trials Human clinical trials
Oral route
Stability Ease of preparation
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Table 3: Phase II/III trials of HEV vaccine Shrestha et al
Zhu et al
NEJM – March 2007
Lancet – August 2010
Phase II trial
Phase III trial
Randomized, double-blind,
Randomized, double-blind,
placebo controlled
placebo controlled
Company
GlaxoSmithKline Biologicals
Xiamen Innovax Biotech
Country
Nepal
Jiangsu province, China
Baculovirus expressed 56 kDa
E. coli expressed HEV 239
1794 healthy subjects
112 604 healthy subjects
HEV vaccine versus placebo
HEV versus HBV vaccine
Mostly males ( 99.6%)
males and females, 16 – 65 years
Phase study Type of study
Recombinant protein ORF2 Number of vaccinees Randomization Population
Young (mean age 25 years)
HEV genotypes
genotype 1 prevalent
Genotypes 1 and 4 prevalent with predominance of genotype 4
20 μg
30 μg
Route of administration
Intramuscularly
Intramuscularly
Intervals between doses
0, 1, 6 months
0, 1, 6 months
Prevention of clinically overt
Prevention of clinically overt
HEV disease
HEV disease
2 years post-vaccination
13 month post-vaccination
87.5%
95.5%
After 2nd dose
85.7%
100%
After 3rd dose
95,5%
100%
Increased injection-site pain
No side effect
Not further developed
Hecolin® (Innovax)
Doses
Primary end-point
Follow-up period Efficacy: After 1st dose
Side effects Commercialization
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