The Turkish Journal of Gastroenterology
2002, Volume 13, No 1, Page(s) 6-10
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|The effect of recombinant hepatitis B vaccine therapy in chronic hepatitis B infection
|Cengiz PATA1, Aziz YAZAR1, Kamuran KONCA1, Gökhan BÝLGÝÇ1, Gülçin ESKANDARI2, Candan ÖZTÜRK3
|Mersin Universty Medical Faculty, Depatments of Internal Medicine1, Biochemistry2 and Microbiology3, Mersin
|Keywords: Chronic hepatitis B, healthy hepatitis B virus carrier, recombinant hepatitis B vaccine..
Background/aims: Immune-modulator and antiviral treatments
for carriers of hepatitis B virus are known to have poor
efficacy with a high cost and frequent side effects, which has led
to investigation of new treatment modalities. The aim of this
study was to determine the efficacy of recombinant hepatitis B
vaccine in the treatment of patients with chronic hepatitis B
(HBV DNA>5 pg/ml, ALT>60) infection diagnosed histopathologically
and asymptomatic carriers (HBV DNA<5 pg/ml, ALT
<40, Anti Hbe positive) of the virus. Methods: The vaccine (Gen
Hevac B Pasteur) was administered at baseline and at months
one and six to patients with chronic hepatitis and asymptomatic
carriers. Ten cases with chronic hepatitis B infection
were assigned to a control group to whom no treatment was
given. Biochemical and microbiological investigations were
performed at baseline and at months three, six and twelve in all
cases. Seroconversion of Hbe Ag, loss of HBV DNA and normalization
of ALT were considered as a positive response.
Results: Patients with chronic hepatitis B who were given the
vaccine were found to have significantly low levels of HBV DNA
at 12 months (63.2±20pg/ml) compared to baseline values
(174.4±36.9pg/ml), while controls were found to have high levels
of HBV DNA at 12 months (223.1±33pg/ml) compared to
baseline values (165.2±33.2pg/ml) (p<0.05). At 12 months,
HBV DNA had become negative in seven of 19 patients given
the vaccine (36.8%) Four patients with chronic hepatitis
(36.35%) were observed to have HBeAg seroconversion and one
patient (5.2%) HBsAg seroconversion at the end of 12 months
and there were four (21.05%) patients who responded positively
to vaccine therapy in this group. Asymptomatic carriers and
controls did not have seroconversion of HBs Ag. Also, HBV
DNA did not become negative in controls. Conclusion: It may
be concluded that recombinant hepatitis B vaccine is effective
in the treatment of chronic hepatitis B.
Materials And Methods
Hepatitis B, a non-cytopathic virus, infects the
liver cells and cellular and humoral immune
responses determine the disease outcome (1). Five
percent of cases develop chronic hepatitis following
infection and in time face an increased risk of
cirrhosis and hepatocellular carcinoma (HCC) (2). Efficacy of the most satisfactory combinations of
immuno-modulating and antiviral treatments has
been around 30-45% in chronic hepatitis (3-6).
Furthermore, these treatment regimes are costly
and cause frequent and serious side-effects, which
has lead to increasing efforts to find new treatment modalities (7). In addition, the treatment of
asymptomatic hepatitis B carriers is of concern
due to periods of flares and the potency of cirrhosis
and HCC in this group of patients (8,9).
Recombinant hepatitis B vaccines have long been
used for protection in the serum of and three
doses have been shown to produce Anti HBs in the
serum of approximately 95% of people who have
not encountered the virus (10). Two kinds of
recombinant vaccine are used for active immunization
against hepatitis B; one of them contains
the PreS1 and PreS2 antigenic domains while the
other kind contains S and PreS polypeptide. No
important differences between the effectiveness of
these two types of vaccine have been detected (11).
Recently, vaccine therapy has been proposed for
the treatment of some chronic hepatitis B viral
Materials And Methods
|Materials And Methods
This study included 19 patients diagnosed with
chronic hepatitis (Group I), 18 asymptomatic carriers
(Group II) and 10 patients diagnosed with
chronic hepatitis (controls) who were referred to
Internal Disease Outpatient Clinics of Mersin
University Hospital, Turkey, between 1999 and
2000. The patients with chronic hepatitis were
positive for HBs Ag and HBV DNA and their ALT
values were 1.5 times higher than the normal values.
The asmyptomatic carriers of hepatitis B
virus were negative for HBV DNA, positive for
Anti HBe, positive for HBs Ag and had normal
level of ALT (<40). The controls were positive for
HBs Ag and HBV DNA and their ALT levels were
1.5 times higher than the normal.
All patients and carriers underwent physical
examination, biochemical examination and investigations
of hepatitis markers (HBs Ag, Anti HBs,
HBe, Hbc IgM, Anti HCV) and HBV DNA. Biopsy
of the liver was performed under ultrasonography
using Tru-cut biopsy needle and the material
obtained was evaluated by the same pathologist
according to Knodell’s histologic activity index
(14). Patients with Anti HCV antibody and Anti
HIV antibody, with alcohol dependency, previously
treated for chronic hepatitis and those found to
have cirrhosis on biopsy were excluded from the study. The vaccine Gen Hevac B Pasteur (contains
S and PreS polypeptide) 0.5 ml was administered
by intramuscular injection into the deltoid region
of patients and carriers in both study groups at
baseline and at months one and six. Control
patients were not given any treatment.
Biochemical analyses and investigations of
hepatitis markers and HBV DNA were performed
at baseline and three, six and 12 months. On the
basis that antibody synthesis starts two weeks
after vaccination (15), biochemical assessments
were made 15 days after the third vaccination. A
positive response to treatment was defined at the
end of one year as loss of serum HBV DNA (<5
pg/ml), seroconversion from HBe Ag (if present) to
Anti HBe and normalization of ALT level (<40)
Measurement of hepatitis markers were made in
accordance with random access Elisa system
using Elisa kits (Abbott USA) and AXYM device.
Cut-off points for HBs Ag, HBe Ag, Anti HBe and
Anti HCV were considered 2, 10, 1, <1 and 1
HBV Hybridization assay
HBV DNA solution hybridization was carried out
in the sera of patients using appropriate kits and
with probes of HBV pointed by I 125
(Diagen/USA). A set of oligonucleotide probes bind
single-stranded HBV DNA to a solid phase, which
is detected by a second set of oligonucleotide
probes. The assay protocol was according to the
manufacturer's instructions. Briefly, 10 µl of test
samples or HBV DNA-positive standards (0.5 to
4,400 MEq/ml) was added to 10 µl of lysis reagent
on a microwell plate coated with oligonucleotide
probes and incubated for 30 min at 63°C to release
HBV DNA from the viral particles. Ten microliters
of the second set of probes in denaturing
buffer was then added to the wells and incubated
for 30 min at 63°C. Ten microliters of neutralizing
reagent was then added, and the hybridization
process continued for 16 hours. After washing,
amplifiers were added and incubated for 30 min at
53°C. After washing, 30 µl of dioxetane substrate
was added. The plate was incubated for 25 min at
37°C in the Chiron luminometer, and light emission
was measured for each well (16).
Man Whitney U, One Way Anova and Chi-square
tests were used for statistical analysis. P<0.05
was considered significant.
Materials And Methods
Age and sex were comparable in study groups I
(12 female, seven male; age: 36±3.4 years) and II
(10 female, eight male; age: 41±2.5 years) and in
the control group (six female, four male; age:
39.2±8.2 years). At baseline, there was no difference
between ALT levels in Group I
(63.52±4.18U/L) and the control group
(64.18±5.25U/L) but when these two groups were
compared with the Group II (32.77±1.97U/L), the
difference was significant (p<0.05). At baseline
and 12 months there was significant decrease in
ALT levels of Group I (p<0.05) while in the control
group, ALT level was 64.18±5.35U/L at baseline
and 72.9±7.35U/L at 12 months (p<0.05) (Table 1).
In Group I, anti HBe was positive in eight cases
(42%) at baseline, in ten cases (52%) at six months
and in 12 cases (63%) (p<0.05) at 12 months.
Apart from positive Anti HBe cases at baseline,
four cases (36.35%) were found to have seroconversion
from HBe Ag to anti HBe. Anti HBs was
negative in all cases at baseline and there was no
change at three and six months, but Anti HBs
antibodies (>100U/L) were detected in one case (5.2%) at 12 months. In five of the controls (50%),
HBe Ag was positive and in the remaining Anti
HBe was positive at baseline, but no change was
detected in serological tests at three, six and 12
months (Table 1).
There was no significant difference in baseline
HBV DNA levels between the Group I
(174.47±36.93pg/ml) and control patients
(165.27±33.25pg/ml) (p>0.05). However, the HBV
DNA level at 12 months was significantly low
(63.21±20.5319pg/ml) in Group I, while it was significantly
high (223.10±33.8pg/ml) in the control
group (p<0.05). HBV DNA was positive in all cases
of Group 1 at baseline and negative in three cases
(15%) at six months (Table 1). It was observed that
HBV DNA disappeared in seven of 19 cases (three
HBe Ag positive at baseline with no HBe Ag seroconversion
at 12 months, two HBe Ag positive at
baseline with HBe Ag seroconversion at 12 months
and two Anti HBe positive) in Group I (36.8%) at
12 months, and that the ALT level of all these
seven cases was below 40IU. Thus there were four
patients (21.05%) who responded positively to vaccine therapy in Group I. There was no significant
difference in mean ages, HAI and HBV DNA levels
of cases who responded positively and those
who did not. In the control group, HBV DNA did
not become negative in any cases.
Comparison of the four cases with and seven cases
without HBe Ag seroconversion showed no significant
difference in age, HAI scores and ALT levels
at baseline (p>0.05), although HBV DNA levels
were higher in cases with seroconversion (p<0.05).
Also, there was no significant difference in the
mean age of cases responding to treatment, they
were younger than those not responding to treatment.
At 12 months, there was no significant difference
in ALT levels between these two groups,
but HBV DNA levels were significantly lower in
cases with seroconversion of HBe Ag than those
without (p>0.05) (Table 2). While low titres of
HBV DNA were determined in one of the four
cases with seroconversion of HBe Ag, HBV DNA
titres of another was measured as 9 pg/ml. It was
thought that this virus may develop mutations in
the core gene.
Table 1. Results of ALT, HBV DNA, seroconversion of HBe Ag and HBs Ag in study and control groups.
Table 2. Comparison of ALT and HBV DNA levels of chronic hepatitis patients with an without seroconversion of
Materials And Methods
In this study, HBV DNA became negative in
36.8% of patients and seroconversion from HBe
Ag to AntiHBe occurred in 36.3% of patients with
chronic hepatitis who received vaccine therapy;
thus a positive response was determined in
21.05% of these patients while no seroconversion
occurred in the other groups. Pol et al. reported
that HBV DNA became negative in 55.6% of 46
patients six months after treatment with rHB vaccine
given three times at monthly intervals (15).
Couillin et al., in their series of 17 cases who were
administered six doses of vaccine, noted that HBV
DNA became negative in 57.1% of their cases,
Anti HBe occurred in 29.4%, but HBs Ag seroconversion
did not occur in any case at the end of 12
months follow up (17). Similarly, several other
studies have reported that HBV became negative
in almost half of patients, while HBs Ag seroconversion
was not observed (18,19). Akbar et al
reported that of 32 HBV transgenicmice, whose
HBV DNA, HBe Ag and HBs Ag were positive and
which were given 10µg of vaccine every month for
12 months, HBe Ag antibodies disappeared in 30
HBV transgenicmice (93.7%), Anti HBs antibodies developed in five HBV transgenicmice and HBV
DNA levels significantly decreased in all rats (20).
The results of all these studies show that the vaccine
is an effective treatment modality. Although
the findings of the present study are concordant
with previous reports, the small number of cases
limit the conclusions which may be drawn.
Despite advances in this field, the immunological
pathophysiology implicated in the marked differences
in the prognosis of chronic viral hepatitis B
patients has not been fully understood (21). A
virus causing infection in the liver should be recognized
by CD4 TH cells, an important molecule
of the immune system, and dendritic cells should
sufficiently process the antigen for the recognition
of the virus (22). TH1 cells, which are a subtype of
CD4 TH, release interleukin-2 (IL-2) and INFgamma
(23), which have an inductive role in the
synthesis of B lymphocytes with characteristics of
Ig G1 and Ig G2. Anti HBs developing after
hepatitis B virus infection have been shown to
possess the nature of Ig G1 and Ig G2 (24). When
cytokines were measured in healthy hepatitis B
virus carriers and patients with chronic hepatitis
following vaccination, INF-gamma levels were
observed to increase significantly in those
responding to treatment (25,26). Several investigators
suggested that the functions of dendritic
cells were impaired in carriers unable to develop
sufficient antibody to the virus and in turn developing
chronic hepatitis (27,28). With vaccination,
above all recombinant vaccine, it is though that be
administration of antigen in different quantities,
different compositions of the virus and processing
of the virus by antigen presenting cells may cause
activation of the immune system. This hypothesis
is supported by the findings of Akbar et al that
MHC Class II CD 86 receptors on dendritic cells
increased following vaccination (29).
In conclusion, hepatitis B vaccine is inexpensive
and has few side-effects. Although its efficacy in
hepatitis B infection has been demonstrated by a
limited number of studies, it appears to be an
effective treatment and may be an alternative to
other treatment modalities. Further studies, in
which the vaccine can be administered to patients
not responding to other treatments for longer
periods and with vaccines containing different
antigens are needed.
Materials And Methods
1. Szmunnes W, Stevens CE, Harley EJ, et al. Hepatitis B
Vaccine: Demonstration of efficacy in a controlled clinical
trial in a high risk population in the United States. N Eng J Med 1980;303: 833-41.
2. Alexander GJM, Brahm J, Fagan EA. Loss of HbsAg with
interferon therapy in chronic hepatitis virus infection. Lancet 1987; 2: 66-8.,
3. Saracco G, Mazzella G, Rosina F. A controlled trial of
human lymphoblastoid interferon in chronic hepatitis B in
Italy. Hepatology 1989;10: 336-41.
4. Jules L, Dienstang MD, Eugene R et al. Lamivudine as initial
treatment for chronic hepatitis B in the United States.
N Eng J Med 1999;341:1256-63.
5. Liaw YF, Leung NW, Chang TT et al. Effects of extended
lamivudine therapy in Asian patients chronic hepatitis B.
Asia hepatitis lamivudine study group. Gastroenterology
6. Wong DKH, Cheung AM, Q’Rourke K, et al. Effect of alpha
interferon treatment in patients with HBe antigen positive
chronic hepatitis B. Ann Intern Med 1993; 119: 312-23.
7. Peters M, Vierling J, Grenswin ME, et al. Immunology of
liver. Hepatology 1991;13: 977-94.
8. Guan R, Yap I, Wong L, et al. Evidence of viral replication
in HBs Ag positive patients with hepatocelluler carcinoma
: measurement of serum hepatitis B virus deoxyribonucleic
acid. Ann Acad Med Singapore 1989; 88: 557-61.
9. LingaoAL, Domingo EO, Nishioka K. Hepatitis B virus
profile of hepatocelluler carcinoma in the Philippines.
Cancer 1981; 48: 1590-5.
10. Böcher WO, Herzog-Hauff S, Herr W, et al. Regulation of
neutralizing antihepatitis B surface antibody response in
vitro in HBs vaccine recipients and patients with acute or
chronic hepatitis B virus infection. Clin Exp Immunol
11. Ey›gun CP, Y›lmaz S, Gul C, et al. A cooperative trial of
two surface subunit recombinant hepatitis B vaccines vs a
surface and PreS subunit vaccine for immunization of
healthy adults. J Viral Hepat 1998; 5: 265-9.
12. Burke DS. Vaccine therapy for HIV, a historic view of the
treatment of infectious diseases by active specific immunization
with microbe-derived antigens. Vaccine 1993; 11:
13. Stanberry LR, Bernstein DI, Burke RL, et al. Vaccination
with herpes simplex virus glyco-proteins; protection
against initial and reccurent genital herpes. J Infect Dis
1987; 155: 914-20.
14. Knodell RG, Ishak KG, Black WC. Formulation and application
of a numerical scoring system for assessing histological
activity in asymptomatic chronic active hepatitis.
Hepatology 1981; 1:431-5.
15. Pol S, Coullin I, Michel ML, et al. Immunotherapy of
chronic hepatitis B by anti HBV vaccine. Acta
Gastroenterol Belg 1998; 61: 28-33.
16. Kapke GF, Watson G, Sheffler S, et al. Comparison of the
Chiron Quantiplex branched DNA (bDNA) assay and the
Abbott Genostics solution hybridization assay for quantification
of hepatitis B viral DNA. J. Viral Hepatitis 1997; 4:67-75
17. Coullin I, Pol S, Mancini M, et al. Specific Vaccine therapy
in Chronic Hepatitis B: Induction of T Cell Proliferative
Responses Specific for envelope Antigens. J Infect Dis
1999; 180: 15-26.
18. Pol S. Driss F, Michel M, et al. Specific vaccine therapy in
chronic hepatitis B infection. Lancet 1994; 216: 342-8.
19. Wen Y-M, Wu X-H, Hu D-C, et al. Hepatitis B vaccine and
anti HBs complex as approach for vaccine therapy. Lancet
20. Akbar SMF, Kazunori K, Kanimota K, et al. Placebo controlled
trial of vaccination with hepatitis B virus surface
antigen in Hepatitis B virus transgenic mice. J Hepatol
1997; 26: 131-7.
21. Löhr HF, Krung S, Herr W, et al. Quantitative and functional
analysis of core specific T helper cell and CTL activities
in acute and chronic hepatitis B. Liver 1998;18: 413-
22. Löhr HF, Weber W, Schlaak J, et al. Proliferative response
CD4 T cells and hepatitis B virus clearance in chronic
hepatitis with or without hepatitis B e-minus hepatitis B
virus mutants. Hepatology 1995; 22: 61-8.
23. Imboden JB: T- Lymphocytes and Natural Killer Cells. In
Medical Immunology. Stites DP, Abba I, Tristram
T,Parslow G ( Eds). Prentice-Hall International Inc.1997, p
24. Mattilla PS, Pelkonen J, Makela O. Proportions of Ig G
subclasses in antibodies formed during vaccination with
hepatitis B surface antigen. Scand J Immunol 1986; 24:
25. Böcher W, Herzog-Hauff S, Schlaak J, et al. Kinetics of
hepatitis B surface antigen-specific immuner response in
acute and chronic hepatitis B or after HBs vaccination:
Stimulation of the in vitro antibody response by interferon
gamma. Hepatology 1999; 29: 238-44.
26. Rahman F, Dahmen A, Herzog-Hauf S, et al. Cellular and
humoral immune responses induced by intradermal or
intramuscular vaccination with the major hepatitis B surface
antigen. Hepatology 2000; 31: 521-7.
27. Egea E, Iglesias A, Salazar M, et al. The cellular basis for
lack of antibody responses to hepatitis B vaccine in
humans. J Exp Med 1991; 173: 531-8.
28. Kurose K, Akbar F, Yamamoto K, et al. Production of antibody
to hepatitis B surface antigen (anti-HBs) by murine
hepatitis B virus carriers: neonatal tolerence versus antigen
presentation by dendritic cells. Immunology 1997; 92:
29. Akbar SMF, Abe M, M Toshikazu, et al. Mechanism of activation
of vaccine therapy in murine hepatitis B virus carriers:
vaccine-induced activation of antigen presenting
dendritic cells. J Hepatol 1999; 30: 755-64.
Materials And Methods
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