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n e w e ng l a n d j o u r na l
of
m e dic i n e
Original Article
A BS T R AC T
BACKGROUND
From the State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, National Institute of Diagnostics
and Vaccine Development in Infectious
Diseases, Collaborative Innovation Center of Biologic Products, School of Public
Health, Xiamen University (J.Z., S.-J.H.,
T.W., J.W.-K.S., M.-H.N., N.-S.X.), and
Xiamen Innovax Biotech Company (Q.Y.,
M.G., X.-H.L., H.-R.P., Y.-M.L.), Xiamen,
Jiangsu Provincial Center for Disease
Control and Prevention, Nanjing (X.-F.Z.,
Y.-M.H., H.W., J.-X.L., F.-C.Z.), Dongtai
Center for Disease Control and Prevention, Dongtai (Z.-Z.W., H.-M.J., Y.-J.W.,
C.-L.Y.), and Yancheng Center for Disease
Control and Prevention, Yancheng (Q.T.,
R.-J.J.) all in China. Address reprint requests to Mr. J. Zhang at the State Key
Laboratory of Molecular Vaccinology and
Molecular Diagnostics, School of Public
Health, Xiamen University, Xiangan
South Rd., Xiamen 361102, China, or at
zhangj@xmu.edu.cn; to Mr. Zhu at the
Jiangsu Provincial Center for Disease
Control and Prevention, Nanjing 210009,
China, or at jszfc@vip.sina.com; or to
Mr. Xia at the National Institute of Diagnostics and Vaccine Development in Infectious Diseases, School of Public Health,
Xiamen University, Xiangan South Rd.,
Xiamen 361102, China, or at n
sxia@xmu
.edu.cn.
Mr. J. Zhang and Mr. X.-F. Zhang contributed equally to this article.
This article was updated on March 5,
2015, at NEJM.org.
Hepatitis E virus (HEV) is a leading cause of acute hepatitis. The long-term efficacy
of a hepatitis E vaccine needs to be determined.
METHODS
During the 4.5-year study period, 60 cases of hepatitis E were identified; 7 cases
were confirmed in the vaccine group (0.3 cases per 10,000 person-years), and 53
cases in the control group (2.1 cases per 10,000 person-years), representing a vaccine efficacy of 86.8% (95% confidence interval, 71 to 94) in the modified intention-to-treat analysis. Of the participants who were assessed for immunogenicity
and were seronegative at baseline, 87% of those who received three doses of the
hepatitis E vaccine maintained antibodies against HEV for at least 4.5 years; HEV
antibody titers developed in 9% in the control group. The rate of adverse events was
similar in the two groups.
CONCLUSIONS
Immunization with this hepatitis E vaccine induced antibodies against HEV and
provided protection against hepatitis E for up to 4.5 years. (Funded by the Chinese
Ministry of Science and Technology and others; ClinicalTrials.gov number,
NCT01014845.)
914
latter vaccine, with the commercial name of Hecolin, has been in use in China since 2012. However,
the long-term efficacy of this hepatitis E vaccine
has not yet been determined.
Me thods
Participants
915
n e w e ng l a n d j o u r na l
The
of
m e dic i n e
Vaccine group
Control group
Blinded extended follow-up study
01
67
13
Vaccine
group
ation
Safety Analysis
Vaccine group
Control group
19
31
43
55
Placebo group
No. of Participants
Assessed for Adverse
Events and Serious
Adverse Events
56,302
56,302
AE:
SAE:
V
Vacc:
56302 Vacc:
V
56302
Plac:
56302
Plac:Per56302
Intention-to-
EfficacyEfficacy
Analysis
Vaccine group
Control group
Protocol
Treat
PPS
PopulationITTPopulation
Vaccine
48693
56,302 56302 48,693
48663
48,663
Placebo
56,302 56302
Immunogenicity
Immunogenicity Analysis
01
67
13
19
31
43
55
Month
No. of Patients
Seronegative at baseline
Vaccine group
Received doses 1, 2, and 3
Received doses 1 and 3
Received doses 1 and 2
Received dose 1
Control group
Seropositive at baseline
Vaccine group
Received doses 1, 2, and 3
Received doses 1 and 3
Received doses 1 and 2
Received dose 1
Control group
2908
97
105
50
3221
2908
97
105
50
3221
418
8
12
6
451
2523
68
63
23
2741
2222
70
59
25
2391
2168
64
62
23
2401
315
7
7
4
338
2659
92
79
38
2833
2659
92
79
38
2833
618
14
16
8
640
1803
57
46
25
1884
2078
64
54
23
2164
2066
61
55
25
2154
505
15
10
8
523
nejm.org
March 5, 2015
With the use of commercial enzyme-linked immunosorbent assay (ELISA) kits (Beijing Wantai
Biological Pharmacy), with the assay performed
according to the manufacturers instructions,
specimens were tested for IgM and IgG antibodies against HEV, IgM antibodies against hepatitis A virus, IgM antibodies against hepatitis B
virus core antigen, hepatitis B virus surface antigen, and antibodies against hepatitis C virus.
Levels of IgG antibodies against HEV were measured in World Health Organization units (WU),
which were calculated by a comparison of the
measured level with a reference level of the as-
Efficacy was estimated with the use of data collected during both the initial study and the extended follow-up study. We defined three sets of
efficacy analyses: a per-protocol analysis, which
included data collected from 1 month after the last
vaccination (month 7) through month 54 from
participants who received three doses of either
the hepatitis E vaccine or the control vaccine; an
intention-to-treat analysis, which included data
collected from month 7 through month 54 from
participants who received at least one dose of
either vaccine; and a modified intention-to-treat
analysis, which included data collected from baseline through month 54 from participants who received at least one dose of either vaccine.
Vaccine efficacy and 95% confidence intervals
were calculated on the basis of the observed difference between the vaccine group and the control group and the accrued person-years. An exact conditional procedure was used to evaluate
vaccine efficacy under the assumption that the
numbers of patients with hepatitis E in the vaccine and control groups were independent Poisson random variables.24 To test the robustness of
the results, we also assessed the vaccine efficacy
using a Cox proportional-hazards model and
compared the cumulative incidence of hepatitis
E between the study groups with a log-rank test.
For the analysis of immunogenicity, we included participants who received at least one dose
of a vaccine and who had available serologic test
results at baseline, month 7, and at least one later
time point. In calculating the geometric mean
concentrations, we arbitrarily set seronegative
samples at 0.04 WU per milliliter.
Long-term safety was assessed according to
917
The
n e w e ng l a n d j o u r na l
100
90
Vaccine group
100.0
Participants (%)
80
Control group
70
99.9
60
50
99.8
40
30
0.0
20
10
0
19
31
43
55
19
31
43
55
Vaccine group
Control group
Cumulative No.
of Participants
with Hepatitis E
Vaccine group
Control group
56,302 56,215
56,302 56,209
0
0
0
6
56,022
55,977
55,774
55,718
55,484
55,436
55,240
55,185
1
22
4
38
5
45
7
53
the rate of serious adverse events in the intentionto-treat population. The rates and 95% confidence intervals (unadjusted for multiple comparisons) were compared between the groups with
the use of a two-sided Fishers exact test.
The data analyses were performed with the
use of SAS software, version 9.1 (SAS Institute).
All reported P values are two-sided, and values of
less than 0.05 were considered to indicate statistical significance. To maintain the study blinding,
an independent external statistician performed
the efficacy analysis.
R e sult s
m e dic i n e
of hepatitis E was significantly lower in the vaccine group than in the control group (P<0.001 in
the modified intention-to-treat analysis) (Fig.2);
53 of the cases were detected in the control group,
and 7 were detected in the vaccine group (Table1). Of the 7 participants in the vaccine group
who had hepatitis E, 3 had received all three
doses of the vaccine, 1 had received two doses,
and 3 had received one dose. The illness in these
participants was moderate or mild; no deaths or
episodes of liver failure were recorded. There
were no notable differences between the two
groups with respect to clinical disease manifestations. (Details of the clinical features of the
cases of hepatitis E that occurred during the
extended study are provided in Tables S2 and S3
in the Supplementary Appendix.) Laboratory test
results among participants who had hepatitis E
were generally similar in the two groups, although
a difference in the avidity of antibodies was seen
during the acute phase. Participants in the vaccine group in whom hepatitis E developed had
antibodies with high avidity, a finding that is
consistent with the anamnestic response in persons with previous immunity; in contrast, the
majority of participants in the control group in
whom hepatitis E developed had antibodies with
low avidity, a finding that is consistent with the
antibody response in persons without previous
immunity (Fig. S2 and Table S4 in the Supplementary Appendix). Of the 29 participants whose
viral isolates were sequenced, 26 were infected
with HEV genotype 4, and 3 were infected with
HEV genotype 1.
The analyses showed significant efficacy of the
hepatitis E vaccine through month 54 (Table2).
Efficacy did not decrease during the extended
follow-up study. Vaccine efficacy was 86.8%
(95% confidence interval [CI], 71.0 to 94.0) in
the modified intention-to-treat analysis, 85.1%
(95% CI, 67.1 to 93.3) in the intention-to-treat
analysis, and 93.3% (95% CI, 78.6 to 97.9) in the
per-protocol analysis.
Immunogenicity
Efficacy
Participants who were enrolled in the initial efficacy study were followed for 544.2 months
(Fig.1, and Table S1 in the Supplementary Appendix). A total of 60 cases of hepatitis E were confirmed, including 23 in the initial study23 and 37
in the extended study. The cumulative incidence
918
of
We determined the geometric mean concentrations of IgG antibodies against HEV and the
seropositivity rates before and after vaccination
and plotted them according to the immune status of the participants at baseline (Fig.3). Of the
5567 participants in the vaccine group who were
assessed for immunogenicity, 52% were seroneg-
Vaccine Group
(N=7)
Control Group
(N=53)
P Value
4711.2
549.2
0.07
0.67
Male
37
Female
16
0.37
3
23
13.63.5 x ULN
23.32.6 x ULN
0.32
4929.6
5121.9
0.53
4.3 (1.611.4)
8.5 (7.110.3)
0.09
15.8 (1.2215.2)
57.4 (37.887.3)
0.51
64 (5077)
6 (48)
<0.001
* Plusminus values are means SD. CI denotes confidence interval, and WU World Health Organization units.
Alanine aminotransferase levels were measured in units per liter. The upper limit of the normal range (ULN) is 40 U
per liter in men and 31 U per liter in women.
The test for avidity of IgG antibodies against HEV was conducted on serum samples collected during the acute phase,
before day 60 of the illness (see Fig. S2 and Table S4 in the Supplementary Appendix). Avidity was the residual antibody
level in the presence of 5M urea expressed as a percentage of the residual antibody level in the absence of urea. A residual antibody level greater than 60% indicates high avidity, a level of 40 to 60% indicates moderate avidity, and a level
lower than 40% indicates low avidity. Of the participants who had hepatitis E, 1 of the 7 in the vaccine group and 5 of
the 53 in the placebo group were not tested for avidity because not enough samples were available.
ative at baseline, of whom 99.9% underwent seroconversion after vaccination; 87% of those who
were seronegative at baseline and received three
doses of the hepatitis E vaccine remained seropositive at the end of month 54 (i.e., the beginning
of month 55) (Table S5 in the Supplementary Appendix). The seroconversion rates were similar
among participants who were seronegative at
baseline and received either one, two, or three
doses of the hepatitis E vaccine. From month 7
through month 43, antibody levels were higher in
participants who received three doses of the hepatitis E vaccine than in those who received two
doses; antibody levels were lowest in participants
who received only one dose of the vaccine. Participants who were seropositive at baseline had a
response to a single dose of the vaccine (Fig.3).
Safety
Discussion
The hepatitis E vaccine we tested provided sustained protection against hepatitis E for up to
4.5 years after the first vaccination. The immune
response persisted in participants with and
those without preexisting immunity to HEV. No
concerns about the safety of the vaccine were
identified.
Vaccine efficacy did not appear to decrease
over time. In addition to one breakthrough infection that was observed in a vaccinated participant
during the initial study, six more breakthrough
infections occurred in vaccinated participants
during the extended follow-up study. The immune
responses to the vaccine in participants with
breakthrough infections are unknown, which
makes it impossible to determine the level of anti-
919
The
n e w e ng l a n d j o u r na l
of
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Table 2. Vaccine Efficacy in the Modified Intention-to-Treat, Intention-to-Treat, and Per-Protocol Analyses.
Analysis
Vaccine Group
Person-Yr
at Risk*
Cases of
Hepatitis E
Control Group
Rate
Person-Yr
at Risk*
no./10,000
person-yr
(95% CI)
number
Cases of
Hepatitis E
Vaccine Efficacy
Rate
no./10,000
person-yr
(95% CI)
number
P Value
% (95% CI)
Modified intention-to-treat
018 mo
87,354
0.1 (0.00.6)
87,323
22
2.5 (1.63.8)
95.5 (66.399.4)
<0.001
030 mo
143,249
0.3 (0.10.7)
143,173
38
2.7 (1.93.6)
89.5 (70.596.2)
<0.001
042 mo
198,873
0.3 (0.10.6)
198,751
45
2.3 (1.73.0)
88.9 (72.095.6)
<0.001
054 mo
254,380
0.3 (0.10.6)
254,198
53
2.1 (1.62.7)
86.8 (71.094.0)
<0.001
56,105
0.2 (0.01.0)
56,081
16
2.8 (1.64.6)
93.8 (52.999.2)
<0.001
Intention-to-treat
718 mo
730 mo
112,000
0.4 (0.10.9)
111,931
32
2.9 (2.04.0)
87.5 (64.795.6)
<0.001
742 mo
167,624
0.3 (0.10.7)
167,509
39
2.3 (1.73.2)
87.2 (67.595.0)
<0.001
754 mo
223,131
0.3 (0.10.6)
222,956
47
2.1 (1.52.8)
85.1 (67.193.3)
<0.001
1954 mo
167,026
0.4 (0.10.8)
166,875
31
1.9 (1.32.6)
80.1 (53.791.9)
<0.001
718 mo
48,595
0.0 (0.00.0)
48,555
15
730 mo
97,031
0.1 (0.00.6)
96,946
30
3.1 (2.14.4)
96.7 (75.699.6)
<0.001
742 mo
145,262
0.1 (0.00.4)
145,122
37
2.6 (1.83.5)
97.3 (80.399.6)
<0.001
754 mo
193,409
0.2 (0.00.5)
193,197
45
2.3 (1.73.1)
93.3 (78.697.9)
<0.001
1954 mo
144,814
0.2 (0.00.6)
144,642
30
2.1 (1.43.0)
90.0 (67.397.0)
<0.001
Per-protocol
* The number of person-years at risk was defined as the cumulative number of follow-up years of the at-risk participants at the indicated time
point.
the Supplementary Appendix). These findings suggest that, in some situations, the virus can evade
the immunity induced by vaccination in the same
manner that it can evade naturally acquired immunity, as reported previously.9
The sustained vaccine efficacy observed in
this study was associated with persistent immunity. The level of vaccine-induced antibodies decreased quickly in the first 2 years after vaccination and subsequently decreased more slowly. Most
of the vaccinated participants had detectable antibodies up to 4.5 years after the first vaccination.
The antibody levels induced by two doses of
the hepatitis E vaccine were slightly lower than
the levels induced by three doses. Previous reports have suggested that the first two doses of
the hepatitis E vaccine prevent illness and infection in the 5 months after vaccination, until the
third dose is given.23,25 These findings raised interest in the use of a two-dose schedule in future
100.00
14.96
10.00
4.09
1.47
1.00
0.56
0.39
0.27
0.10
0.04
0.01
13
19
31
43
55
0
0
2904
103
418
30
2512
151
2165
158
1996
165
274
29
2908
3221
2908
3221
418
451
2523
2741
2222
2391
2168
2401
315
338
Total No.
Vaccine group
Control group
100.00
24.74
10.85
10.00
7.44
4.41
3.77
2.27
1.00
0.10
0.04
0.01
13
19
31
43
55
Total No.
Vaccine group
Control group
2659
2833
2658
2760
618
629
1801
1834
2074
2066
2059
1970
504
466
2659
2833
2659
2833
618
640
1803
1884
2078
2164
2066
2154
505
523
921
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