Dec 11, 2014
Editor-in-Chief
BMC Infectious Diseases
Re: MS: 1554678346122581
Title: Safety and Immunogenicity of a freeze-dried, Vero cell culture-derived, inactivated
Japanese encephalitis vaccine (KD-287, ENCEVAC(R)) versus a mouse brain-derived
inactivated Japanese encephalitis vaccine in children: a phase III, multicenter, doubleblinded, randomized trial
Dear Professor Philippa Harris,
Thank you very much for your 2nd review of our revised manuscript. Please find our
responses to the expert reviewers’ comments. We appreciate each comment and have made
a series of revisions that we believe have much improved the manuscript. Please do not
hesitate to contact me if you have further questions or concerns.
Sincerely,
Hoan Jong Lee, MD
Professor of Pediatrics
Seoul National University College of Medicine
Seoul National University Children’s Hospital
101 Daehak-ro, Jongno-gu Seoul, 110-769, Republic of Korea
Tel: 82-2-2072-3633
Fax: 82-2-745-4703
E-mail: hoanlee@snu.ac.kr
1
[Reviewer #1]
Reviewer's report:
Although there are some study limitations remaining, the authors answered to this
reviewer's questions appropriately.
- Thank you very much.
[Reviewer #2]
1. Discussion, page 21, lines 18-19.
The comparison of serious adverse events should be done for subjects rather than events.
- Thank you for noting this. We revised the Table 2 and changed the following sentence.
Table 1 Solicited adverse events after vaccinations at each dose in the study subjects (safety
population)
No.(%) of subjects
Group (n)
Total
After 1st Vaccination
After 2nd Vaccination
After 3rd Vaccination
AE
no AE
KD-287 (102)
87 (85.29)
15 (14.71)
JEV-GCC (102)
77 (75.49)
25 (24.51)
KD-287 (102)
65 (63.73)
37 (36.27)
JEV-GCC (102)
54 (52.94)
48 (47.06)
KD-287 (102)
45 (44.12)
57 (55.88)
JEV-GCC (100)
45 (45.00)
55 (55.00)
KD-287 (100)
46 (46.00)
54 (54.00)
JEV-GCC (96)
50 (52.08)
46 (47.92)
p-value*
0.050
0.118
0.900
0.394
AE, adverse event
*The p-value was calculated using the chi-square test.
“Furthermore, severe AEs were rare in both vaccine groups (17/385 [4.4%] in the KD-287
group, 11/390 [2.8%] in the JEV-GCC group, p>0.05).” (Page 21, Lines 18-19)
 “Furthermore, severe AEs were infrequent in both vaccine groups (12/102 [13.8%] in the
KD-287 group, 10/102 [13.0%] in the JEV-GCC group, p>0.05).” (Page 20, Lines 17-18)
2
2. Discussion, page 23, lines 3-4.
It is not possible to say that there were no serious vaccine-related adverse events due to KD287.
- We appreciate the reviewer’s comment. As indicated by the reviewer, there were SAE
associated with the KD-287. Therefore, we changed the following sentences.
“The three-dose regimen of KD-287 showed a favorable safety profile with no serious
vaccine-related adverse events.”
 “In this study, we confirmed that KD-287, the JE-VC prepared with the Beijing-1 strain,
showed a comparable safety profile with and a higher immunogenicity than the JE-MB
prepared with the Nakayama strain.” (Page 21, Lines 14-16)
3. Table 2.
The data for “No. of events” are not useful. It would be more useful to know the number
and proportion of subjects in each group that had mild, moderate, or severe events,
particularly for moderate or severe events following the first dose of vaccine.
- Thank you for this comment. As the reviewers 2 and 3 commented, we revised the Table 2
as following. We decided to omit the incidence of AEs according to the severity for clarifying
the comparison between the two groups and for simplifying new Table 1. Instead, we added
the data regarding to the severity of AEs in new Table 2 (revised Table 3 of the original
manuscript).
Table 1 Solicited adverse events after vaccinations at each dose in the study subjects (n=204,
safety population)
No.(%) of subjects
Group (n)
Total
After 1st Vaccination
After 2nd Vaccination
AE
no AE
KD-287 (102)
87 (85.29)
15 (14.71)
JEV-GCC (102)
77 (75.49)
25 (24.51)
KD-287 (102)
65 (63.73)
37 (36.27)
JEV-GCC (102)
54 (52.94)
48 (47.06)
KD-287 (102)
45 (44.12)
57 (55.88)
3
p-value*
0.050
0.118
0.900
After 3rd Vaccination
JEV-GCC (100)
45 (45.00)
55 (55.00)
KD-287 (100)
46 (46.00)
54 (54.00)
JEV-GCC (96)
50 (52.08)
46 (47.92)
0.394
AE, adverse event
*The p-value was calculated using the chi-square test.
- In addition, we changed the following sentence.
“Most AEs were mild to moderate in both groups (75/87 [86.2%], KD-287; 67/77 [87.0%],
JEV-GCC).” (Page 12, Lines 17-18)
 “Most AEs were mild to moderate in both groups (75/87 [86.2%], KD-287; 67/77 [87.0%],
JEV-GCC, data not shown).” (Page 11, Line 23 – Page 12, Line 1)
4. Table 3.
Suggest presenting the solicited adverse events for the 7 days following each dose during
which those data were actively collected.
- We appreciate the reviewer’s comment. We changed the Table 3 as following.
Table 2 Incidence of solicited adverse events within 7 days after each vaccination (safety
population)
1st dose, n*(%)
Severity
Erythema
Pain
KD-287
JEV-GCC
KD-287
JEV-GCC
KD-287
JEV-GCC
(n=102)
(n=102)
(n=102)
(n=100)
(n=100)
(n=96)
9 (8.8)
18 (17.6)
7 (6.9)
14 (14.0)
7 (7.0)
14 (14.6)
Moderate
3 (2.9)
2 (2.0)
5 (4.9)
2 (2.0)
7 (7.0)
6 (6.3)
Severe
1 (1.0)
0 (0)
0 (0)
0 (0)
1 (1.0)
0 (0)
14 (13.7)
10 (9.8)
3 (2.9)
5 (5.0)
18 (18.0)
11 (11.5)
2 (2.0)
2 (2.0)
2 (2.0)
1 (1.0)
2 (2.0)
3 (3.1)
0 (0)
0 (0)
0 (0)
0 (0)
0 (0)
1 (1.0)
Mild
3 (2.9)
7 (6.9)
6 (5.9)
7 (7.0)
5 (5.0)
11 (11.5)
Moderate
2 (2.0)
1 (1.0)
2 (2.0)
1 (1.0)
1 (1.0)
2 (2.1)
Severe
1 (1.0)
0 (0)
0 (0)
0 (0)
1 (1.0)
0 (0)
19 (18.6)
10 (9.8)
13 (12.7)
12 (12.0)
13 (13.0)
14 (14.6)
8 (7.8)
4 (3.9)
2 (2.0)
4 (4.0)
1 (1.0)
3 (3.1)
Mild
Severe
Fever
3rd dose, n*(%)
Mild
Moderate
Swelling
2nd dose, n*(%)
Mild
Moderate
4
Severe
Crying
Irritability
4 (3.9)
1 (1.0)
2 (2.0)
3 (3.0)
0 (0)
1 (1.0)
27 (26.5)
20 (19.6)
15 (14.7)
6 (6.0)
14 (14.0)
14 (14.6)
Moderate
5 (4.9)
4 (3.9)
3 (2.9)
6 (6.0)
2 (2.0)
3 (3.1)
Severe
2 (2.0)
0 (0)
0 (0)
1 (1.0)
0 (0)
1 (1.0)
12 (11.8)
11 (10.8)
5 (4.9)
6 (6.0)
13 (13.0)
4 (4.2)
1 (1.0)
4 (3.9)
2 (2.0)
5 (5.0)
0 (0)
6 (6.3)
0 (0)
0 (0)
0 (0)
0 (0)
0 (0)
0 (0)
Mild
8 (7.8)
11 (10.8)
4 (3.9)
3 (3.0)
1 (1.0)
4 (4.2)
Moderate
2 (2.0)
0 (0)
0 (0)
0 (0)
0 (0)
0 (0)
Severe
1 (1.0)
0 (0)
0 (0)
0 (0)
0 (0)
0 (0)
17 (16.7)
11 (10.8)
3 (2.9)
5 (5.0)
3 (3.0)
4 (4.2)
4 (3.9)
1 (1.0)
0 (0)
2 (2.0)
0 (0)
2 (2.1)
0 (0)
1 (1.0)
0 (0)
0 (0)
1 (1.0)
1 (1.0)
10 (9.8)
13 (12.7)
5 (4.9)
8 (8.0)
6 (6.0)
8 (8.3)
Mild
Mild
Moderate
Severe
Vomiting
Diarrhea
Mild
Moderate
Severe
Decreased
Mild
appetite
Moderate
2 (2.0)
0 (0)
0 (0)
1 (1.0)
1 (1.0)
0 (0)
Severe
2 (2.0)
0 (0)
1 (1.0)
0 (0)
0 (0)
1 (1.0)
Mild
9 (8.8)
12 (11.8)
8 (7.8)
7 (7.0)
14 (14.0)
17 (17.7)
Moderate
0 (0)
1 (1.0)
0 (0)
1 (1.0)
0 (0)
0 (0)
Severe
0 (0)
0 (0)
0 (0)
0 (0)
0 (0)
0 (0)
Somnolence
*Number of subjects reporting the specific characteristic.
- Additionally, we changed the following sentences.
“All AEs occurring after the three doses of vaccine in both groups are shown in Table 3.
Overall, the local tolerability and systemic safety profile of KD-287 was promising in the
study, with no serious safety concerns recorded. The most common AEs indicated by
participants after immunization were fever (52/102 [51.0%], KD-287; 41/102 [40.2%], JEVGCC), crying (47/102 [46.1%], KD-287; 40/102 [39.2%], JEV-GCC), and diarrhea (30/102
[29.4%], KD-287; 25/102 [24.5%], JEV-GCC). The frequency of most AEs was similar between
the two groups. Although nasopharyngitis was more prevalent in the KD-287 group and
gastroenteritis was more prevalent in JEV-GCC group (p=0.005 and 0.018, respectively),
these AEs may not have been related to the vaccination.
In addition, we compared AEs between the two groups within 7 days after each vaccination.
There were no significant differences in any of the AEs except for fever, which was
significantly different only after the first vaccination (30/102 [29.4%], KD-287; 15/102
5
[14.7%], JEV-GCC, p=0.007, data not shown). We performed further analysis for the daily
cumulative incidence of fever after each vaccination, and we concluded that more mild fever
events occurred within 2 days after the first vaccination in the KD-287 group than in the JEVGCC group (18/102 [17.6%], KD-287; 7/102 [6.9%], JEV-GCC, p=0.019, Figure 2).” (Page 12,
Line 20 – Page 13, Line 12)
 “All solicited AEs occurring within 7 days after each vaccination in both groups are shown
in Table 2. Overall, the local tolerability and systemic safety profile of KD-287 was promising
in the study, with no serious safety concerns identified. The most common AE indicated by
participants was crying in mild severity (27/102 [26.5%], KD-287; 20/102 [19.6%], JEV-GCC)
after first vaccination. The frequency of most AEs was similar between the two groups and
most local and systemic events were mild or moderate in severity. Only the total incidence of
fever after the first vaccination was significantly different between the two groups (31/102
[30.4%], KD-287; 15/102 [14.7%], JEV-GCC, p=0.012). Among the fever events after the first
vaccination in KD-287 and JEV-GCC groups, 61.3% (19/31) and 66.7% (10/15) were mild
severity, respectively.” (Page 12, Lines 7-16)
“The rates of all solicited AEs were not different between the vaccine groups. However,
when the evaluation was confined to the 7-day period after each injection, the incidence of
fever events after the first injection of KD-287 was apparently higher than that after the first
injection of JEV-GCC. Further analysis revealed that only low-grade fever was more prevalent
after KD-287 injection than after control vaccine injection, especially within two days after
the first vaccination.” (Page 21, Line 21 – Page 22, Line 3)
 “The rates of all solicited AEs within 7 days after each vaccination were also not
significantly different between the vaccine groups, except the higher incidence of mild fever
events after the first injection of KD-287 than that of JEV-GCC.” (Page 20, Lines 18-21)
- We deleted the following sentences.
“Although nasopharyngitis was more prevalent in the KD-287 group and gastroenteritis was
more prevalent in JEV-GCC group (p=0.005 and 0.018, respectively), these AEs may not have
been related to the vaccination.” (Page 13, Lines 3-5 of the original manuscript)
“Among the unsolicited AEs, nasopharyngitis was more prevalent in the KD-287 group, and
6
gastroenteritis was more prevalent in the JEV-GCC group. All unsolicited AEs, including the
observed nasopharyngitis and gastroenteritis, may not have been related to the injected
vaccine.” (Page 22, Lines 8-11 of the original manuscript)
5. Table 4.
As above, the data for “No. of events” are not useful. Suggest only presenting and
comparing the number and proportion of subjects with serious adverse events.
- Thank you for the comment. During the revision process by the comment of reviewers 2
and 3, we identified the erroneous gathering and interpretation of SAE incidence in both
groups. Thus, we revised Table 4 and changed the p-value in the table and main text
correctly as following.
Table 3 Incidence of serious adverse events requiring hospitalization (safety population)
Adverse Events
No. of Subjects, n (%)
p-value*
KD-287 (n=102)
JEV-GCC (n=102)
17 (16.67)
23 (22.55)
0.373
Febrile seizure
1 (0.98)
-
1.000
Otitis Media
1 (0.98)
1 (0.98)
1.000
Pharyngitis
2 (1.96)
1 (0.98)
1.000
Laryngitis
1 (0.98)
-
1.000
Bronchitis
4 (3.92)
4 (3.92)
1.000
Pneumonia
5 (4.90)
13 (12.75)
0.127
-
1 (0.98)
1.000
Gastroenteritis
1 (0.98)
3 (2.94)
0.369
Others†
2 (1.96)
-
0.498
Total
Influenza
*The p-value was calculated using the chi-square or Fisher’s exact test.
†Others included a case of vaccine overdose and a case of accessory skin tag.
“In total, 58 SAEs were reported: 26 occurred in 23 children in the KD-287 group and 32
occurred in 30 children in the JEV-GCC group (p=0.378, Table 4). These events were regarded
as most likely not related to the vaccine itself except for one event of fever and one event of
febrile seizure in different children, both in the KD-287 group.” (Page 13, Lines 13-16)
7
 “In total, 52 SAEs were reported: 23 occurred in 17 children in the KD-287 group and 29
occurred in 23 children in the JEV-GCC group (p=0.373, Table 3). These events were regarded
as most likely not related to the vaccine itself except for one event of febrile seizure in the
KD-287 group.” (Page 12, Lines 17-20)
“Only one fever event and one febrile seizure event in the KD-287 group may have been
associated with vaccination.” (Page 22, Lines 11-13)
 “Only one febrile seizure event in the KD-287 group may have been associated with
vaccination.” (Page 21, Lines 1-2)
6. Title and abstract.
Despite the well-established correlate of protection and the examples of other safety and
immunogenicity studies use the term “phase 3”, I still believe this is a phase 2 study. No
response needed.
- Thank you for the comment.
7. Background, page 6, line 1.
Suggest adding reference for Kaltenbock et al. Vaccine 2010;28:834.
- Thank you very much for noting this. We added this article as reference number 11.
8. Methods, page 6, lines 22-23.
The ITT population consisted of participants with at least one blood sample collected postvaccination. It appears that two subjects in the safety population received one dose of JEVGCC (and therefore had a pre-vaccination blood sample collected) but were excluded from
the ITT analysis. I accept the authors explanation regarding the exclusion criteria for the ITT
group but still believe all subjects who were randomized to a study group should be included
in the ITT analysis whether or not they received a dose of vaccine or had a post-vaccination
specimen.
- We appreciated and agree with the reviewer’s comment. We revised the following
sentences and Fig.1.
8
“The safety population comprised all participants receiving at least one vaccination. The
intention-to-treat (ITT) population consisted of all participants that had at least one blood
sample among the safety population.” (Page 6, Line 22 – Page 7, Line 1)
 “The intention-to-treat (ITT) population comprised all participants who were randomized
to a study group, and the safety population comprised all participants receiving at least one
vaccination.” ” (Page 6, Line 22 – Page 7, Line 1)
“Primary immunogenicity assessments were based on the PP population (n=188), excluding
participants with major protocol violations. All secondary immunogenicity data were based
on the ITT population (n=202).” (Page 10, Lines 19-21)
 “All immunogenicity assessments were based on the PP population (n=188), excluding
participants with major protocol violations.” (Page 10, Lines 21-22)
“All of the randomized participants except one, who voluntarily withdrew before the first
vaccination, were included in the safety population (n=204, 99.5%).” (Page 11, Lines 21-22)
 “All of the randomized participants (n=205, 100%; ITT population) except one, who
voluntarily withdrew before the first vaccination, were included in the safety population
(n=204, 99.5%).” (Page 11, Lines 17-19)
- We deleted the following sentence.
“The first two doses were given two weeks apart in 102 (99.0%) children in the KD-287
group and in 100 (98.0%) children in the JEV-GCC group (ITT population).” (Page 11, Lines 1516 of the original manuscript)
- We added the following sentence.
“In addition, immunogenicity data for the ITT population (data not shown) were comparable
to those for the PP population (Tables 4 and S1).” (Page 14, Lines 2-4)
9

9. Results, page 11, lines 17-18.
What were the eligibility criteria violations (N=3) and the protocol violations before the third
vaccination (N=9)?
- Thank you for the comment. The eligibility criteria violations were other vaccine (MMR)
administration within 4 weeks preceding the first dose of the study vaccine (n=1) and
underlying chronic diseases (n=2; one congenital hypothyroidism and one bronchopulmonary dysplasia). The protocol violations before the third vaccination were use of
prohibited drugs (n=1), visit window deviation (n=6), over dose (n=1), and violation of 3rd
vaccination criteria (occurrence of SAE [febrile convulsion]; n=1) (Figure 1).
We change the following sentences.
“Seventeen (8.3%) children were dropped because of eligibility criteria violations (n=3),
follow-up loss (n=2), withdrawal of consent (n=3), and protocol violation before the third
vaccination (n=9).” (Page 11, Lines 16-18)
”Seventeen (8.3%) children were dropped because of eligibility criteria violations (n=3;
other vaccination 2 weeks before enrollment [n=1] and underlying chronic diseases [n=2]),
follow-up loss (n=2), withdrawal of consent (n=3), and protocol violation before the third
vaccination (prohibited drug use [n=1], visit window deviation [n=6], over dose [n=1], and
violation of third vaccination criteria [neurologic SAE; n=1]).” (Page 11, Lines 11-16)
10
10. Results, page 13, line 9.
Delete data not shown; they are presented in this sentence.
- Thank you for the comment. We deleted the sentence.
11. Results, page 13-15.
Consider presenting all of the homologous immunogenicity results first and then the
heterologous results.
- Thank you for the comment. We revised the following sentences and Tables 5 and S1 as
suggested.
“In the homologous analysis (matching vaccine and target strains), the SCR after the booster
injection (primary endpoint) with KD-287 (100%) was higher than the SCR with JEV-GCC
(98.9%), but the opposite was observed in the heterologous analysis (the SCR with KD-287
was lower than that with JEV-GCC) (p>0.05 in both analyses). Only one subject in each
vaccine group was not seroconverted to JEV when tested with the virus of the Nakayama
strain. Because the lower limit of the 95% CI for the seroconversion difference was higher
than -10% in the homologous analysis (-1.00%), we concluded that KD-287 achieved the
non-inferiority criterion. The lower limit of the 95% CI for the seroconversion difference in
the heterologous analysis was also higher than -10% (-3.17%). Furthermore, the SCRs against
the homologous strain after the second vaccination and before the third vaccination were
100% and 100%, respectively, in the KD-287 group, and 97.9% and 94.7%, respectively, in the
JEV-GCC group (p>0.05 in both). Against the heterologous strain, the SCRs in the KD-287
group were significantly higher than those in the JEV-GCC group, both after the second
vaccination (94.62% and 54.74%, respectively, p<0.01) and before the third vaccination
(93.55% and 34.74%, respectively, p<0.01).
The GMTs were also higher in the KD-287 group than they were in the JEV-GCC group after
the second vaccination and before and after the third vaccination against the homologous
strain (GMT ratio: 5.59, 20.13, and 13.79, respectively, all p<0.01; Table S1) and the
heterologous strain (GMT ratio: 4.05, 5.15, and 4.19, respectively, all p<0.01 in all; Table S1).
The reverse cumulative curve of neutralizing antibodies against the homologous strain also
revealed that immune responses of the KD-287 group were increased relative to those of
the JEV-GCC group after the second vaccination and before and after the third vaccination,
11
respectively (Figure 3). Because the lower limits of the 95% CI for the GMT ratio were higher
than 0.5 in both the homologous (10.8) and the heterologous (3.2) analyses (Table S1), we
concluded that KD-287 was also not inferior to JEV-GCC with regard to the GMT of the
neutralizing antibody.” (Page 12, Line 1 – Page 13, Line 3)
 “The immunogenicity assessed by SCRs and GMTs at 4 weeks after the third injection was
excellent in both vaccines. Only one subject in JEV-GCC group was not seroconverted to JEV.
The GMTs were higher in the KD-287 group than they were in the JEV-GCC group after the
third vaccination (GMT ratio: 13.79, p<0.001). Because the lower limit of the 95% CI for the
seroconversion difference (-1.00%) and for the GMT ratio (10.8) was higher than -10% and
0.5, respectively, we concluded that KD-287 achieved the non-inferiority criterion.
Furthermore, the SCRs after the second vaccination and before the third vaccination were
excellent in both groups. The GMTs were also higher in the KD-287 group than they were in
the JEV-GCC group after the second vaccination and before and after the third vaccination
(GMT ratio: 5.59 and 20.13, respectively, p<0.001 in both). The reverse cumulative curve of
neutralizing antibodies also revealed that immune responses of the KD-287 group were
increased relative to those of the JEV-GCC group after the second vaccination and before
and after the third vaccination, respectively (Figure S1).
In the heterologous analysis, the SCRs after the third injection were also excellent in both
vaccines, so that only one subject in KD-287 group was not seroconverted (Table S1). The
SCRs in the KD-287 group were higher than those in the JEV-GCC group, both after the
second vaccination (94.62% and 54.74%, respectively) and before the third vaccination
(93.55% and 34.74%, respectively). Against the heterologous strain, the GMTs were also
higher in the KD-287 group than they were in the JEV-GCC group after the second
vaccination and before and after the third vaccination (GMT ratio: 4.05, 5.15, and 4.19,
respectively, p<0.001 in all). The lower limits of the 95% CI both for the seroconversion
difference (-3.2) and for the GMT ratio (3.2) in the heterologous analysis also fulfilled the
non-inferiority criteria. In addition, immunogenicity data for the ITT population (data not
shown) were comparable to those for the PP population (Tables 4 and S1).” (Page 13, Line 4
– Page 14, Line 4)
12
Table 4 Seroconversion rates and geometric mean titers after vaccination compared to
before vaccination (per-protocol population, homologous response)
Time point
Value
KD-287 (n=93)
JEV-GCC (n=95)
Seropositive
n (%)
0 (0.0)
1 (1.0)
at baseline
95% CI
93 (100.0)
93 (97.9)
After 2nd dose
SCR
Before 3rd dose
After 3rd dose
Before Vaccination
After 2nd dose
GMT
rd
Before 3 dose
After 3rd dose
n (%)
*Difference/Ratio
95% CI
n (%)
2.1
(-0.78, 4.99)
93 (100.0)
90 (94.7)
95% CI
5.3
(0.77, 9.75)
n (%)
93 (100.0)
94 (98.9)
1.1
95% CI
(100.0, 100.0)
(96.9, 100.0)
(-1.00, 3.10)
log10n
5
5
1.0
95% CI
(5, 6)
log10n
601
107
5.6†
95% CI
(509, 709)
(84, 137)
(4.2, 7.5)
log10n
917
45
20.1†
95% CI
(757, 1110)
(36, 56)
(15.2, 26.7)
log10n
13347
967
13.8†
95% CI
(11455, 15551)
(798, 1172)
(10.8, 17.6)
(1.0, 1.1)
SCR, seroconversion rate; GMT, geometric mean titer; CI, confidence interval
*Difference is KD-287 minus JEV-GCC for SCR and ratio is KD-287 divided by JEV-GCC for GMT.
†The p-value was calculated using the t-test for GMT; p<0.001 in all.
13
Table S1 Seroconversion rates and geometric mean titers after vaccination compared to
before vaccination (per-protocol population, heterologous response)
Time point
Value
KD-287 (n=93)
JEV-GCC (n=95)
Seropositive
n (%)
0 (0.0)
1 (1.0)
at baseline
95% CI
88 (94.6)
52 (54.7)
After 2nd dose
SCR
Before 3rd dose
After 3rd dose
Before Vaccination
After 2nd dose
GMT
n (%)
95% CI
n (%)
After 3rd dose
39.9
(28.88, 50.90)
87 (93.6)
33 (34.7)
95% CI
58.8
(48.01, 69.61)
n (%)
92 (98.9)
95 (100.0)
-1.08
95% CI
(96.8, 100.0)
(100.0, 100.0)
(-3.17, 1. 20)
log10n
5
5
1.0
95% CI
(5, 6)
log10n
45
11
4.1†
95% CI
(37, 53)
(9, 13)
(3.2, 5.2)
log10
47
9
5.2†
95% CI
(39, 57)
(8, 11)
(4.0, 6.7)
log10n
705
168
4.2†
95% CI
(591, 843)
(137, 207)
(3.2, 5.5)
n
Before 3rd dose
*Difference/Ratio
(1.0, 1.1)
SCR, seroconversion rate; GMT, geometric mean titer; CI, confidence interval
*Difference is KD-287 minus JEV-GCC for SCR and ratio is KD-287 divided by JEV-GCC for GMT.
†The p-value was calculated using the t-test for GMT; p<0.001 in all.
14
12. Discussion, pages 16-18.
The information regarding the other available JE vaccines could be reduced as it does not
substantially contribute to the interpretation of this study’s findings.
- We appreciate the reviewer’s comment. We deleted the following sentence.
“A live-attenuated virus vaccine prepared with the SA14-14-2 strain in primary hamster
kidney cells was licensed in China in 1988, and is now also available in Nepal, India, Sri Lanka,
and South Korea. A JE-CV (IMOJEV®) was developed by Sanofi-Pasteur by replacing the
cDNA-encoding pr M and E proteins of the yellow fever virus vector, YFV17D, with proteins
of the SA14-14-2 strain. This JE-CV was licensed in Australia and Thailand in 2010. However,
potential safety hazards with live vaccines and genetically modified strains are a constant
source of controversy in the scientific community and among public-health officials,
particularly with regard to the potential threat of mutagenic reversion into pathogenic
strains [19].” (Page 16, Line 17 – Page 17, Line 2 of the original manuscript)
“IC51 is available in 38 countries in North America, Europe, Asia, and Oceania.” (Page 17,
Lines 8-9 of the original manuscript)
“In 1965, a placebo-controlled (tetanus toxoid) trial of an inactivated, unpurified JE vaccine
(Nakayama strain) was conducted in Taiwan, and in 1984, a placebo-controlled (tetanus
toxoid) trial of an inactivated, purified JE vaccine (Nakayama strain and bivalent
Nakayama/Beijing-1 strains) was conducted in Thailand [16]. The bivalent and monovalent
vaccines showed the same (91%) protective efficacies.” (Page 17, Line 21 – Page 18, Line 2 of
the original manuscript)
13. Table 4.
Although there is no significant difference between the two groups, the proportion of
subjects with serious adverse events is remarkably high in both groups. Please confirm that
23% of KD-287 and 29% of JEV-GCC subjects were hospitalized during the study period? If so,
do the authors have an explanation?
- Thank you for noting this. During the revision process, we identified the erroneous
gathering and interpretation of SAE incidence in both groups. However, the proportion of
15
subject with SAEs is still high as 16.67% of KD-287 and 22.55% of JEV-GCC groups.
Because the study subjects were primarily patients that frequently visited the each site,
there could be a selection bias for medical attention and risk factors. Moreover, children
with a lower respiratory tract infection, acute febrile illness, or dehydration are used to be
hospitalized in the easily accessible and relatively inexpensive Korean health care system.
Thus, we added the following sentences.
“Although there was remarkably high incidence of SAEs in both groups, it was possibly due
to the high medical attention of parents of study subjects and specific situation in Korean
healthcare system convenient to be hospitalized.” (Page 21, Lines 2-5)
14. Table 4.
Delete first footnote. The authors should not speculate on what adverse events were related
to vaccination.
- We appreciate the reviewer’s comment. We deleted the footnote.
16
[Reviewer #3]
Reviewer's report:
[Major]
1. It is not appropriate to test the difference in baseline characteristics in a randomised trial.
These tests should be removed from the paper.
- Thank you for noting this. We deleted the following sentences and Table 1.
“Demographic characteristics, including sex, age, medical history, prior medication, and
concomitant medication and vaccination, were compared between treatment groups using
Student’s t-test or the Wilcoxon rank sum test for continuous variables and the chi-square or
Fisher’s exact test for categorical variables.” (Page 11, Lines 2-5 of the original manuscript)
“The demographics and baseline characteristics of the study subjects were similar in the two
groups in the safety population (Table 1). There were no significant differences in the
number of males and females between the two groups (p=0.159). The median ages of the
children who received the test and the control vaccine were 12.0 (range 12.0–22.0) and 12.0
(range 12.0–18.0) months, respectively (p=0.233). The rates of concomitant medications
were high because the subjects might possibly contract a common cold or bacterial
respiratory infection such as otitis media and sinusitis during the relatively long monitoring
period of 14 weeks. Cold preparations and antibiotics were prescribed to 72 (71%) and 66
(65%) subjects in the KD-287 group, respectively and 70 (69%) and 68 (67%) subjects in the
JEV-GCC group, respectively during the study (data not shown).” (Page 11, Line 23 – Page 12,
Line 9 of the original manuscript)
2. It is not clear what analyses where done looking at ‘daily cumulative incidence’ of fever.
There is a reference to mild fever – this sounded as if mild fever was looked at separately
from all fever. This would not be appropriate but hopefully this just a problem of wording. As
an analysis of a difference in when fever occurred will not have been planned it may be
better to display the data graphically rather than perform formal analysis if it is considered
important to report when fever was most likely to occur. What test was done is not clear. It
would not be appropriate to statistically test at each day. This would be an extreme case of
the separate testing of the data after each dose. These are the same people so the results
17
are not independent of one another and correctly an analysis taking this into account should
have been performed. As most analyses have been done on the global measure of any
occurrence I have not suggested a more correct mixed model analysis be performed.
However if separate statistical tests on cumulative data from consecutive days was
performed this is too extreme and the analysis should be removed. If this is in fact reporting
some other analysis then what was done needs to be made clear in the methods if it
remains in the paper. The discussion also needs adjustment in light of this.
- We appreciated and agree with the reviewer’s comment. We deleted the following
sentences regarding the analysis for the ‘daily cumulative incidence of fever’ and Figure 2.
“We performed further analysis for the daily cumulative incidence of fever after each
vaccination, and we concluded that more mild fever events occurred within 2 days after the
first vaccination in the KD-287 group than in the JEV-GCC group (18/102 [17.6%], KD-287;
7/102 [6.9%], JEV-GCC, p=0.019, Figure 2).” (Page 13, Lines 9-12 of the original manuscript)
“Further analysis revealed that only low-grade fever was more prevalent after KD-287
injection than after control vaccine injection, especially within two days after the first
vaccination.” (Page 22, Lines 1-3 of the original manuscript)
- Additionally, we changed the following sentence.
“The safety profile of KD-287 was good, and its local and systemic tolerability profiles were
comparable to those of JEV-GCC, except for a higher incidence of mild fever within two days
after the first vaccination.” (Page 15, Lines 20-22)
 “The safety profile of KD-287 was good, and its local and systemic tolerability profiles
were comparable to those of JEV-GCC, except for a higher incidence of fever after the first
vaccination.” (Page 15, Lines 8-10)
18
[Minor]
1. The breakdown of the study dropouts given in the text is one short – it adds to 16, not 17
- Thank you very much for noting this. We had a mistake for counting the number of subjects
who dropped out. We change the following sentences.
“Seventeen (8.3%) children were dropped because of eligibility criteria violations (n=3),
follow-up loss (n=2), withdrawal of consent (n=3), and protocol violation before the third
vaccination (n=9).” (Page 11, Lines 16-18)
”Seventeen (8.3%) children were dropped because of eligibility criteria violations (n=3;
other vaccination 2 weeks before enrollment [n=1] and underlying chronic diseases [n=2]),
follow-up loss (n=2), withdrawal of consent (n=3), and protocol violation before the third
vaccination (prohibited drug use [n=1], visit window deviation [n=6], over dose [n=1], and
violation of third vaccination criteria [neurologic SAE; n=1]).” (Page 11, Lines 13-18)
2. Table 2 would be better displayed with the no AE group as one of the categories rather
with the % with reaction out to the side and the percentages being the percentage of the
whole group, not those with a reaction. It needs to be made clear in the table that the p
values are for the test of no reaction compared to a reaction rather than the severity of
reaction.
- Thank you for the comment. We revised the Table 2 as following.
Table 1 Solicited adverse events after vaccinations at each dose in the study subjects (safety
population)
No.(%) of subjects
Group (n)
Total
After 1st Vaccination
After 2nd Vaccination
After 3rd Vaccination
AE
no AE
KD-287 (102)
87 (85.29)
15 (14.71)
JEV-GCC (102)
77 (75.49)
25 (24.51)
KD-287 (102)
65 (63.73)
37 (36.27)
JEV-GCC (102)
54 (52.94)
48 (47.06)
KD-287 (102)
45 (44.12)
57 (55.88)
JEV-GCC (100)
45 (45.00)
55 (55.00)
KD-287 (100)
46 (46.00)
54 (54.00)
JEV-GCC (96)
50 (52.08)
46 (47.92)
19
p-value*
0.050
0.118
0.900
0.394
- In addition, we changed the following sentences.
“Most AEs were mild to moderate in both groups (75/87 [86.2%], KD-287; 67/77 [87.0%],
JEV-GCC).” (Page 12, Lines 17-18)
 “Most AEs were mild to moderate in both groups (75/87 [86.2%], KD-287; 67/77 [87.0%],
JEV-GCC, data not shown).” (Page 11, Line 23 – Page 12, Line 1)
“Furthermore, severe AEs were rare in both vaccine groups (17/385 [4.4%] in the KD-287
group, 11/390 [2.8%] in the JEV-GCC group, p>0.05).” (Page 21, Lines 18-19)
 “Furthermore, severe AEs were infrequent in both vaccine groups (12/102 [13.8%] in the
KD-287 group, 10/102 [13.0%] in the JEV-GCC group, p>0.05).” (Page 20, Lines 17-18)
3. Should the heading for table 3 not be after any dose rather than each – it is not separated
out by dose
- Thank you for noting this. We revised Table 3 as following.
Table 2 Incidence of solicited adverse events within 7 days after each vaccination (safety
population)
1st dose, n*(%)
Severity
Erythema
Pain
KD-287
JEV-GCC
KD-287
JEV-GCC
KD-287
JEV-GCC
(n=102)
(n=102)
(n=102)
(n=100)
(n=100)
(n=96)
9 (8.8)
18 (17.6)
7 (6.9)
14 (14.0)
7 (7.0)
14 (14.6)
Moderate
3 (2.9)
2 (2.0)
5 (4.9)
2 (2.0)
7 (7.0)
6 (6.3)
Severe
1 (1.0)
0 (0)
0 (0)
0 (0)
1 (1.0)
0 (0)
14 (13.7)
10 (9.8)
3 (2.9)
5 (5.0)
18 (18.0)
11 (11.5)
2 (2.0)
2 (2.0)
2 (2.0)
1 (1.0)
2 (2.0)
3 (3.1)
0 (0)
0 (0)
0 (0)
0 (0)
0 (0)
1 (1.0)
Mild
3 (2.9)
7 (6.9)
6 (5.9)
7 (7.0)
5 (5.0)
11 (11.5)
Moderate
2 (2.0)
1 (1.0)
2 (2.0)
1 (1.0)
1 (1.0)
2 (2.1)
Severe
1 (1.0)
0 (0)
0 (0)
0 (0)
1 (1.0)
0 (0)
19 (18.6)
10 (9.8)
13 (12.7)
12 (12.0)
13 (13.0)
14 (14.6)
Moderate
8 (7.8)
4 (3.9)
2 (2.0)
4 (4.0)
1 (1.0)
3 (3.1)
Severe
4 (3.9)
1 (1.0)
2 (2.0)
3 (3.0)
0 (0)
1 (1.0)
Mild
Severe
Fever
3rd dose, n*(%)
Mild
Moderate
Swelling
2nd dose, n*(%)
Mild
20
Crying
Irritability
Mild
27 (26.5)
20 (19.6)
15 (14.7)
6 (6.0)
14 (14.0)
14 (14.6)
Moderate
5 (4.9)
4 (3.9)
3 (2.9)
6 (6.0)
2 (2.0)
3 (3.1)
Severe
2 (2.0)
0 (0)
0 (0)
1 (1.0)
0 (0)
1 (1.0)
12 (11.8)
11 (10.8)
5 (4.9)
6 (6.0)
13 (13.0)
4 (4.2)
1 (1.0)
4 (3.9)
2 (2.0)
5 (5.0)
0 (0)
6 (6.3)
0 (0)
0 (0)
0 (0)
0 (0)
0 (0)
0 (0)
Mild
8 (7.8)
11 (10.8)
4 (3.9)
3 (3.0)
1 (1.0)
4 (4.2)
Moderate
2 (2.0)
0 (0)
0 (0)
0 (0)
0 (0)
0 (0)
Severe
1 (1.0)
0 (0)
0 (0)
0 (0)
0 (0)
0 (0)
17 (16.7)
11 (10.8)
3 (2.9)
5 (5.0)
3 (3.0)
4 (4.2)
4 (3.9)
1 (1.0)
0 (0)
2 (2.0)
0 (0)
2 (2.1)
0 (0)
1 (1.0)
0 (0)
0 (0)
1 (1.0)
1 (1.0)
10 (9.8)
13 (12.7)
5 (4.9)
8 (8.0)
6 (6.0)
8 (8.3)
Mild
Moderate
Severe
Vomiting
Diarrhea
Mild
Moderate
Severe
Decreased
Mild
appetite
Moderate
2 (2.0)
0 (0)
0 (0)
1 (1.0)
1 (1.0)
0 (0)
Severe
2 (2.0)
0 (0)
1 (1.0)
0 (0)
0 (0)
1 (1.0)
Mild
9 (8.8)
12 (11.8)
8 (7.8)
7 (7.0)
14 (14.0)
17 (17.7)
Moderate
0 (0)
1 (1.0)
0 (0)
1 (1.0)
0 (0)
0 (0)
Severe
0 (0)
0 (0)
0 (0)
0 (0)
0 (0)
0 (0)
Somnolence
*Number of subjects reporting the specific characteristic.
4. The test for difference in children experiencing SAE’s in the 2 groups has a different p
value in the text from that in table 4
- Thank you for noting this. During the revision process, we identified the erroneous
gathering and interpretation of SAE incidence in both groups. Thus, we revised Table 4 and
changed the p-value in the table and main text correctly as following.
“In total, 58 SAEs were reported: 26 occurred in 23 children in the KD-287 group and 32
occurred in 30 children in the JEV-GCC group (p=0.378, Table 4). These events were regarded
as most likely not related to the vaccine itself except for one event of fever and one event of
febrile seizure in different children, both in the KD-287 group.” (Page 13, Lines 13-16)
 “In total, 52 SAEs were reported: 23 occurred in 17 children in the KD-287 group and 29
occurred in 23 children in the JEV-GCC group (p=0.373, Table 3). These events were regarded
as most likely not related to the vaccine itself except for one event of febrile seizure in the
KD-287 group.” (Page 12, Lines 17-20)
21
“Only one fever event and one febrile seizure event in the KD-287 group may have been
associated with vaccination.” (Page 22, Lines 11-13)
 “Only one febrile seizure event in the KD-287 group may have been associated with
vaccination.” (Page 21, Lines 1-2)
Table 3 Incidence of serious adverse events requiring hospitalization (safety population)
Adverse Events
No. of Subjects, n (%)
p-value*
KD-287 (n=102)
JEV-GCC (n=102)
17 (16.67)
23 (22.55)
0.373
Febrile seizure
1 (0.98)
-
1.000
Otitis Media
1 (0.98)
1 (0.98)
1.000
Pharyngitis
2 (1.96)
1 (0.98)
1.000
Laryngitis
1 (0.98)
-
1.000
Bronchitis
4 (3.92)
4 (3.92)
1.000
Pneumonia
5 (4.90)
13 (12.75)
0.127
-
1 (0.98)
1.000
Gastroenteritis
1 (0.98)
3 (2.94)
0.369
Others†
2 (1.96)
-
0.498
Total
Influenza
*The p-value was calculated using the chi-square or Fisher’s exact test.
†Others included a case of vaccine overdose and a case of accessory skin tag.
5. The post booster responses to both the homologous and heterologous challenges were
almost identical in both groups. It is not appropriate to state one is higher than the other
and that they reverse and then at the end say there was no statistical difference. This should
be reworded (it also needed several reads to work out what the p values were referring to
and actual p values should always be quoted – not >.05
- Thank you for the comments. We changed the following sentences.
“In the homologous analysis (matching vaccine and target strains), the SCR after the booster
injection (primary endpoint) with KD-287 (100%) was higher than the SCR with JEV-GCC
(98.9%), but the opposite was observed in the heterologous analysis (the SCR with KD-287
was lower than that with JEV-GCC) (p>0.05 in both analyses).” (Page 14, Lines 1-4)
22
 “The immunogenicity assessed by SCRs and GMTs at 4 weeks after the third injection was
excellent in both vaccines.” (Page 13, Lines 4-5)
“Furthermore, the SCRs against the homologous strain after the second vaccination and
before the third vaccination were 100% and 100%, respectively, in the KD-287 group, and
97.9% and 94.7%, respectively, in the JEV-GCC group (p>0.05 in both).” (Page 14, Lines 9-12)
 “Furthermore, the SCRs after the second vaccination and before the third vaccination
were excellent in both groups.” (Page 13, Lines 10-11)
“Against the heterologous strain, the SCRs in the KD-287 group were significantly higher than
those in the JEV-GCC group, both after the second vaccination (94.62% and 54.74%,
respectively, p<0.01) and before the third vaccination (93.55% and 34.74%, respectively,
p<0.01).
The GMTs were also higher in the KD-287 group than they were in the JEV-GCC group after
the second vaccination and before and after the third vaccination against the homologous
strain (GMT ratio: 5.59, 20.13, and 13.79, respectively, all p<0.01; Table S1) and the
heterologous strain (GMT ratio: 4.05, 5.15, and 4.19, respectively, all p<0.01 in all; Table S1).”
(Page 14, Lines 12-19)
 “In the heterologous analysis, the SCRs after the third injection were excellent in both
vaccines, so that only one subject in KD-287 group was not seroconverted (Table S1). The
SCRs in the KD-287 group were higher than those in the JEV-GCC group, both after the
second vaccination (94.62% and 54.74%, respectively) and before the third vaccination
(93.55% and 34.74%, respectively). Against the heterologous strain, the GMTs were higher in
the KD-287 group than they were in the JEV-GCC group after the second vaccination and
before and after the third vaccination (GMT ratio: 4.05, 5.15, and 4.19, respectively, p<0.001
in all).” (Page 13, Line 17 – Page 14, Line 1)
6. Table S1 has the incorrect method of analysis in the footnote
- Thank you for noting this. We revised the footnote by correct method of analysis as follows.
“*The p-value was calculated using the chi-square or Fisher’s exact test.” (Page 33, Line 3)
 “†The p-value was calculated using the t-test for GMT; p<0.001 in all.” (Pages 33, Line 6)
23
7. 3nd line of the discussion – the study was for non inferiority, not equivalence
- Thank you for the comment. We changed the following phrase.
“whether its safety and immunogenicity are equivalent to” (Page 15, Line 14)
 “whether its immunogenicity is not inferior and its safety is comparable to” (Page 15,
Lines 3-4)
24
[Editorial points]
* Please name the specific ethics committees which approved your study
- Thank you for the comment. We revised the following sentence.
“The study protocol was approved by independent review boards and the Korean Food and
Drug Administration (KFDA).” (Page 6, Lines 20-21)
 “The study protocol was approved by the Institutional Review Board of Seoul National
University Hospital and the Korean Food and Drug Administration, as well as independent
review board at each study site.” (Page 6, Lines 20-22)
* Please include the details of the funding from Boryung in your competing interest section.
In addition we note that the 'Monitoring, data management, and statistical analyses' was
performed by them. Could you clarify which author this was? Their link to Boryung should
also be fully declared (along with any other pharmaceutical company links).
- Persons who have performed monitoring, data management, and statistical analyses are
not included as author in this manuscript. We added the following sentences regarding the
competing interests and the details of the funding from Boryung.
“None of the authors were employee of or reported payments/grants from the Boryung
Pharmaceutical Co., Ltd., which provided all compensation for study involvement and
supported the expenses for medicine, laboratory tests, and the clinical research coordinator.
The funding body had no role in the analysis, interpretation, or writing of the manuscript
from the data.” (Page 21, Line 22 – Page 22, Line 3)
25