Vaccine 35 (2017) 2999–3006

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Vaccine
journal homepage: www.elsevier.com/locate/vaccine

Effect of prophylactic or therapeutic administration of paracetamol on

immune response to DTwP-HepB-Hib combination vaccine in Indian
infants
Arijit Sil a,⇑, Mandyam D. Ravi d, Badri N. Patnaik a, Mandeep S. Dhingra c, Martin Dupuy b,
Dulari J. Gandhi e, Sangappa M. Dhaded f, Anand P. Dubey g, Ritabrata Kundu h, Sanjay K. Lalwani i,
Jugesh Chhatwal j, Leni G. Mathew k, Madhu Gupta l, Shiv D. Sharma m, Sandeep B. Bavdekar n,
Soumya P. Rout a, Midde V. Jayanth a, Naveena A. D’Cor a, Somnath A. Mangarule a, Suresh Ravinuthala a,
Jagadeesh Reddy E a
a
Shantha Biotechnics Private Limited – A Sanofi Company, Hyderabad, India
b
Sanofi Pasteur, Marcy-l’Étoile, France
c
Sanofi Pasteur, Swiftwater, USA
d
Dept. of Pediatrics, JSS Medical College, Mysore, India
e
Dept. of Pediatrics, SBKS MI & RC, Sumandeep Vidyapeeth, Vadodara, India
f
Dept. of Pediatrics, KLE University’s, Jawaharlal Nehru Medical College, Belagavi, India
g
Dept. of Pediatrics, Maulana Azad Medical College, Delhi, India
h
Dept. of Pediatrics, Institute of Child Health, Kolkata, India
i
Dept. of Pediatrics, Bharati Vidyapeeth Deemed University Medical College, Pune, India
j
Dept. of Pediatrics, Christian Medical College, Ludhiana, India
k
Dept. of Pediatrics, Christian Medical College, Vellore, India
l
Dept. of Community Medicine, School of Public Health, Post Graduate Institute of Medical Education & Research, Chandigarh, India
m
Dept. of Pediatrics, Sawai Man Singh Medical College, Jaipur, India
n
Dept. of Pediatrics, Topiwala National Medical College and BYL Nair Ch. Hospital, Mumbai, India

a r t i c l e i n f o a b s t r a c t

Article history: Background: Vaccination is considered as the most cost effective method for preventing infectious dis-
Received 12 September 2016 eases. Low grade fever is a known adverse effect of vaccination. In India, it is a common clinical practice
Received in revised form 4 March 2017 to prescribe paracetamol either prophylactically or therapeutically to manage fever. Some studies have
Accepted 6 March 2017
shown that paracetamol interferes with antibody responses following immunization. This manuscript
Available online 24 April 2017
reports the outcome of a post hoc analysis of data from a clinical trial of a pentavalent vaccine in
Indian infants where paracetamol was not used or was used either as prophylaxis or for treatment of
Keywords:
fever.
Paracetamol
Vaccine
Methods: Pre and post vaccine antibody levels against Diphtheria, Tetanus, Pertussis, Hepatitis B,
Prophylactic Haemophilus influenzae type B were assessed in no paracetamol and paracetamol groups. The paracetamol
Immunogenicity group was further divided into prophylactic and treatment groups.
Pentavalent Results: Similar rates of seroprotection/seroresponse for anti-D, anti-T, anti-wP, anti-PT, anti-HBs and
India anti-PRP were observed in all the groups. There was no clear tendency for difference in percentage sero-
protection/seroresponse and geometric mean (GM) titers in any of the groups.
Conclusion: The study found no evidence that paracetamol usage either as prophylactic or for treatment
impact immunological responses to DTwP-HepB-Hib combination vaccine.
Conclusion: [Clinical trial registry of India (study registration number CTRI/2012/08/002872)].
Ó 2017 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY-NC-ND license
(http://creativecommons.org/licenses/by-nc-nd/4.0/).

⇑ Corresponding author at: Shantha Biotechnics Private Limited, 3rd and 4th
Floor, Vasantha Chambers, Fateh Maidan Road, Basheer Bagh, Hyderabad, Telangana
500004, India.
E-mail address: arijit.sil@sanofi.com (A. Sil).
1. Introduction
Vaccination is an efficient method to prevent a number of infec-
tious diseases ranging from Polio, Pertussis, Hepatitis B, to HPV

http://dx.doi.org/10.1016/j.vaccine.2017.03.009
0264-410X/Ó 2017 The Authors. Published by Elsevier Ltd.
This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
3000 A. Sil et al. / Vaccine 35 (2017) 2999–3006
associated cancers. The government of India provides vaccines
against seven vaccine preventable diseases which include diphthe-
ria, whooping cough, tetanus, polio, tuberculosis, measles and hep-
atitis B, as a part of National Immunization Program, in the name of
‘‘Mission Indradhanush” to infants [1]. Among these vaccines, the
whole cell pertussis vaccine has been traditionally associated with
post vaccination fever and injection site pain [2,3]. Many clinicians
therefore prefer to prescribe medications to prevent or treat such
reactions. There is some evidence in literature that antipyretics
blunt the antibody response to vaccines [4,5].
In 2013, Shantha Biotechnics Private Ltd. (a Sanofi Company)
conducted a randomized clinical trial in which 1085 infants were
enrolled across India in order to assess the lot to lot consistency
among 3 lots of investigational vaccine Shan5TM (DTwP-HepB-
Hib); as well as immune non-inferiority and safety in comparison
to Pentavac SDTM (comparator DTwP-HepB-Hib vaccine manufac-
tured by Serum Institute of India). The results of this main study
were published earlier [6]. On reviewing the source data from this
study (post hoc analysis), it was observed that paracetamol (Acet-
aminophen) was the most commonly used antipyretic. Different
patterns of paracetamol usage (depending upon individual clinical
judgments and practices of investigators) to prevent or treat febrile
reactions during routine immunization were observed in the study.
Overall four distinct trends regarding the usage of paracetamol
were observed irrespective of the vaccine received: some clinicians
did not prescribe paracetamol at all, some prescribed it as a pro-
phylactic only and some prescribed it for treatment only, whereas
some others prescribed it as prophylactic with one vaccine dose
and for treatment after another vaccine dose for the same subject
during the study period. The reporting rates of fever and pain were
also similar between the vaccine groups showing that any possible
difference in paracetamol usage had hardly any impact [6]. The
reporting rates of fever and pain were also similar to the previously
reported data from other DTwP-HepB-Hib vaccine studies in India
[7,8].
The primary objective of the 2013 study was not to generate the
data pertaining to the effects of paracetamol usage following pen-
tavalent vaccine immunization [6]. Hence the subjects were not
randomized according to the paracetamol usage [No or Yes (Pro-
phylactic or Treatment)] during the study]. The data was collected
on the usage of paracetamol and also the way it has been used
(Prophylactic or Treatment). Subsequently, a post hoc analysis of
the effect of paracetamol on immunogenicity levels following
administration of three doses of DTwP-HepB-Hib combination vac-
cine in Indian infants was performed and the results are presented
here.
2. Material and methods
2.1. Study design
A multi-center (11 study sites), randomized, single blinded, sta-
tistically powered study was conducted across India to assess lot to
lot consistency among 3 lots of investigational vaccine, as well as
immune non-inferiority and safety in comparison to the existing
licensed pentavalent vaccine in India (comparator vaccine) [6].
The study was initiated upon receipt of approval from national reg-
ulatory authority of India [Drug Controller general of India (DCGI)],
and respective institutional ethics committees. The trial was regis-
tered with the Clinical trial registry of India (study registration
number CTRI/2012/08/002872). Once the infant’s parent/guardian
provided written informed consent and the infant was found to
be eligible for the study, he/she was enrolled. Study was conducted
in accordance with the protocol, ICH-GCP, the current rules and
regulations for conducting clinical trials in India including Sched-
ule Y of Drugs and Cosmetics Act, India, Indian GCP, and Ethical
Guidelines for Biomedical Research on Human Participants (ICMR
guidelines) [6].
A total of 1085 infants aged 6–8 weeks were enrolled in the
study. The infants were randomized to receive investigational or
comparator vaccines in a ratio of 6:1. Out of 1085 infants enrolled,
930 received investigational vaccine and 155 received comparator
vaccine. Since the main objective of the post hoc analysis was to
evaluate the effect of paracetamol on antibody titers, the subjects
were segregated according to the usage of paracetamol in addition
to the type of vaccine administered i.e. investigational or compara-
tor. For the post hoc analysis of paracetamol effects, all the subjects
in each vaccine group were categorized primarily into 3 sub-
groups, ‘Without paracetamol’, ‘Paracetamol prophylactic’ and
‘Paracetamol treatment’.
Among subjects who received antipyretics/analgesics, all of
them received paracetamol either alone or in combination with
other analgesics like ibuprofen or mefenamic acid. The proportion
of subjects who used paracetamol combination was very low
(0.3%). Since all the subjects who received antipyretic/analgesic
had received paracetamol, the group was simply termed as ‘Parac-
etamol’ group (‘Paracetamol prophylactic’ or ‘Paracetamol
treatment’).
Subjects in the ‘‘Without paracetamol” group include subjects
who did not receive any paracetamol (or paracetamol containing
medication) during the study. The ‘Paracetamol treatment’ group
includes subjects who received at least one dose of paracetamol
(or paracetamol containing medication) for the purpose of treat-
ment during the study. The ‘Paracetamol prophylactic’, group
includes subjects who received at least one dose of paracetamol
(or paracetamol containing medication) for the purpose of prophy-
laxis during the study. Subjects who received paracetamol both as
prophylactic with one vaccine dose and for treatment after another
vaccine dose (‘heterogeneous’ use) were not included in this anal-
ysis. Additionally, the paracetamol subgroups of both vaccine
groups were pooled to evaluate the impact of paracetamol use irre-
spective of the vaccine (investigational or comparator) received i.e.
pooled vaccine analysis for paracetamol use. Subject disposition is
outlined in Figs. 1 and 2.
2.2. Study participants
Healthy infants aged 6–8 weeks were enrolled in the study after
verifying the inclusion and exclusion criteria. Infants born at
37 weeks gestational period and with 2.5 kg birth weight were
included in the study. Receipt of any other vaccine except BCG,
birth dose of OPV and Hepatitis B, history of diseases targeted by
the study vaccine, etc. were considered as exclusion criteria [6].
2.3. Study vaccines
Both the investigational and comparator vaccines are pentava-
lent pediatric combination vaccines for the prevention of Diphthe-
ria, Tetanus, Pertussis, Hepatitis B and invasive disease caused by
Haemophilus influenzae type b. Composition of investigational and
comparator vaccines are detailed in the earlier publication [6].
2.4. Objectives
As a post hoc analysis, the main study data was analyzed with
the objective of understanding the effect of paracetamol usage as
well as mode of usage i.e. prophylactic or treatment on the
immune response to three doses of DTwP-HepB-Hib combination
vaccine administered to Indian infants at 6–8, 10–12 and 14–
16 weeks of age. We based our analysis on the following research
question: Is there any observed difference in immune responses
 A. Sil et al. / Vaccine 35 (2017) 2999–3006
Fig. 1. Subject disposition as per vaccine received and paracetamol use. Note: Number of subjects (N) in each group for all the valences was not same and varied with a
maximum difference of 3.
between the subjects who have not received paracetamol versus
who have received paracetamol either as prophylactic or for treat-
ment? This was analyzed in each vaccine arm (investigational and
comparator) separately and then as a whole irrespective of the vac-
cine received.
2.5. Study procedures
The study procedures have been detailed and described in the
publication outlining the main results [6]. Criteria for seroprotec-
tion for anti-D, anti-T, anti-HBs and anti-PRP were taken as
0.01 IU/mL, 0.01 IU/mL, 10 mIU/mL and 0.15 mcg/mL
respectively at post dose sample [9–12]. Since no correlate of pro-
tection is available for anti-wP, seroresponse was used as an indi-
cator of protection against pertussis. Criteria for seroresponse:
more than or equal to the pre-vaccination titer in initially seropos-
itive (>11 NTU) subjects; and in case of initial seronegative sub-
jects (11 NTU) the response was considered as per assay cut-off
i.e. those subject having pre-vaccination titer 11 NTU was consid-
ered seroresponders only if they are able to attain a titer of
>11 NTU during post dose sample. Subjects who achieved 4-
fold anti-PT titers rise from baseline titer were considered as
seroresponders of the vaccine [13]. The effect of paracetamol on
anti-D, anti-T, anti-PRP, anti-HBs, anti-wP and anti-PT immuno-
genicity titers were analyzed according to the usage and mode of
usage of paracetamol.
2.6. Statistical analysis
An adhoc a posteriori analysis was conducted on the study
regarding presence or absence of potential impact of paracetamol
on immunogenicity. Using concomitant medication data, subjects
were divided according to the use of paracetamol as described
above. Binary endpoints were selected for each antigen (typically
3001
seroprotection/seroresponse), in order to present the percentage
of subjects according to paracetamol use. Similarly, GM post-
Dose 3 antibody titer was computed for each antigen. The effect
of paracetamol on immunogenicity was analyzed both separately
for each vaccine group and also irrespective of the vaccine group.
The analysis was descriptive using full analysis set, based on
observed value with 95% CI. The CI for the single proportion was
calculated using the exact binomial method (Clopper-Pearson
method, quoted by Newcombe) [14]. For immunogenicity results,
assuming that Log10 transformation of the titers follows a normal
distribution, at first, the mean and the 95% CI was calculated on
Log10 (titers) using the usual calculation for normal distribution
(using Student’s t distribution with n 1 degree of freedom), then
antilog transformation was applied to the results of calculations, in
order to get geometric means (GMs) and their 95% CI.
3. Results
Out of 1085 infants enrolled in the study, data from 975 sub-
jects were considered for this post hoc analysis and the data from
the rest of the 110 subjects were not considered due to various rea-
sons such as subject drop-out, non-availability of post dose sample,
and haemolysis of blood sample. Out of the 975 subjects, 407 did
not receive paracetamol (356 from investigational group and 51
from comparator group) and 568 subjects received paracetamol
as a prophylactic and/or for treatment during the study. Out of
these 568 subjects who received paracetamol, 47 subjects received
paracetamol both as a prophylactic with one vaccine dose and for
treatment after another vaccine dose (‘heterogeneous’ use) were
excluded from this analysis. Out of the remaining 521 subjects
who received paracetamol either as a prophylactic or for treatment
during the study, 71 subjects received paracetamol as a prophylac-
tic only (60 from investigational group and 11 from comparator
group) and 450 received paracetamol for treatment only (387 from
3002 A. Sil et al. / Vaccine 35 (2017) 2999–3006
Fig. 2. Subject disposition as per paracetamol use irrespective of vaccine received. Note: Number of subjects (N) in each group for all the valences was not same and varied
with a maximum difference of 3.
investigational group and 63 from comparator group). Overall it
was observed that similar proportion of subjects have received
paracetamol, either as prophylactic or as treatment, irrespective
of vaccine group (investigational or comparator vaccine group)
that they were randomized to (Fig. 1).
Among the subjects who received paracetamol, multiple occur-
rences of paracetamol use, as high as 7 times in a single subject
was observed during the complete study period. Pre and post vac-
cination blood samples for all the subjects were analyzed for anti-
bodies against each antigen. Seroprotection/seroresponse and GM
titers of all the valences for ‘Without paracetamol’, ‘With paraceta-
mol prophylactic’ and ‘With paracetamol treatment’ subgroups
with respect to the vaccine received are shown in (Table 1). The
results for ‘Without paracetamol’, ‘With paracetamol prophylactic’
and ‘With paracetamol treatment’ groups irrespective to the vac-
cine received’ are shown in (Table 2).
3.1. Anti-D immunogenicity
All the subjects (100%) in the study had achieved seroprotection
levels for anti-D irrespective of vaccine received and the paraceta-
mol use (Tables 1 and 2).
In the investigational group, the anti-D GMTs were similar
among the 3 subgroups irrespective of type of paracetamol use.
However, in the comparator group, the anti-D GMT in the sub-
group ‘Paracetamol prophylactic’ (0.81 IU/mL) was observed to be
numerically lower than the ‘without paracetamol’ subgroup
(1.71 IU/mL) without overlapping 95% CI. The anti-D GMTs in the
‘Paracetamol treatment’ subgroup of comparator group were sim-
ilar to both the ‘Without paracetamol’ and ‘Paracetamol prophylac-
tic’ subgroups (Table 1).
In the pooled vaccine analysis, the anti-D GMTs were similar
among the 3 groups irrespective of type of paracetamol use
(Table 2).
3.2. Anti-T immunogenicity
All the subjects (100%) in the study had achieved seroprotection
levels for anti-T irrespective of vaccine received and the paraceta-
mol use (Tables 1 and 2).
In both the investigational and comparator groups, the anti-T
GMTs were similar among the subgroups irrespective of type of
paracetamol use (Table 1). In the pooled analysis, some differences
in anti-T GMT among the groups were observed (Table 2). The GMT
in ‘Without paracetamol’ group (2.11 IU/mL) was observed to be
numerically higher than that of ‘With paracetamol treatment’
group (1.75 IU/mL) without overlapping of 95% CI (Table 2). While
the GMT of ‘With paracetamol prophylactic’ group was similar to
both ‘Without paracetamol’ and ‘With paracetamol treatment’
groups.
 A. Sil et al. / Vaccine 35 (2017) 2999–3006 3003

Table 1
Immune response to Diphtheria, Tetanus, Hib, Hepatitis B and Pertussis antigens based on the nature of the vaccine administered.

Antigen Statistics Investigational Comparator

Without With paracetamol With paracetamol Without With paracetamol With paracetamol
paracetamol prophylactic treatment paracetamol prophylactic treatment
N = 356 N = 60 N = 387 N = 51 N = 11 N = 63
Anti-D Antibody-Serology- % 100.0 100.0 100.0 100.0 100.0 100.0
IU/mL Seroprotection (99.0; 100.0) (94.0; 100.0) (99.1; 100.0) (99.0; 100.0) (71.5; 100.0) (94.3; 100.0)
(95% CI)
GM 1.65 1.76 1.51 1.71 0.81 1.47
(95% CI) (1.53; 1.79) (1.50;2.05) (1.39; 1.65) (1.41; 2.08)* (0.51;1.30)* (1.22; 1.76)
Anti-T Antibody-Serology- % 100.0 100.0 100.0 100.0 100.0 100.0
IU/mL Seroprotection (99.2; 100.0) (94.0; 100.0) (99.1; 100.0) (93.0; 100.0) (71.5; 100.0) (94.3; 100.0)
(95% CI)
GM 2.06 2.04 1.72 2.44 1.84 1.95
(95% CI) (1.87; 2.28) (1.59;2.63) (1.57; 1.89) (1.87; 3.17) (1.21;2.79) (1.56; 2.45)
Anti-PRP Antibody- % 99.8 100.0 99.7 100.0 100.0 100.0
Serology-mcg/mL Seroprotection (98.9; 100.0) (94.0; 100.0) (98.6; 100.0) (93.0; 100.0) (71.5; 100.0) (94.3; 100.0)
(95% CI)
GM 8.56 11.2 9.04 10.4 3.58 10.4
(95% CI) (7.68; 9.53) (8.91; 14.1) (8.20; 9.96) (7.56; 14.4) (1.20; 10.7) (8.12; 13.3)
Anti-HBs Antibody- % 97.4 96.7 98.2 100.0 100.0 100.0
Serology-mIU/mL Seroprotection (95.2; 98.8) (88.5; 99.6) (96.3; 99.3) (93.0; 100.0) (71.5; 100.0) (94.2; 100.0)
(95% CI)
GM 926 508 824 2246 1360 1645
(95% CI) (780; 1098) (326; 793) (707; 961) (1515; 3331) (410;4514) (1117; 2424)
Anti-wP Antibody- % 69.6 73.3 71.1 68.6 81.8 68.3
Serology-Novotech units Seroresponse (64.5; 74.3) (60.3; 83.9) (66.3; 75.5) (54.1; 80.9) (48.2; 97.7) (55.3; 79.4)
(95% CI)
GM 17.6 18.1 17.5 17.2 20.6 17.1
(95% CI) (15.9; 19.5) (13.9; 23.7) (16.0; 19.2) (13.4; 22.1) (11.1; 38.4) (13.5; 21.6)
Anti-PT antibody-Serology- % 4-fold rise 63.5 71.7 66.7 64.7 54.5 61.9
IU/mL (95% CI) (58.2; 68.5) (58.6; 82.5) (61.7; 71.3) (50.1; 77.6) (23.4; 83.3) (48.8; 73.9)
GM 16.0 18.7 25.1 16.5 10.5 23.0
(95% CI) (12.9; 19.9)* (10.4; 33.8) (20.5; 30.7)* (10.1; 27.1) (2.71; 41.0) (14.4; 36.8)

Note: Number of subjects (N) in each group for all the valences was not same and varied with a maximum difference of 3.
*
Non overlapping 95% C.

Table 2
Immune response to Diphtheria, Tetanus, Hib, Hepatitis B and Pertussis antigens regardless of the nature of the vaccine administered.

Antigen Statistics Without paracetamol With paracetamol prophylactic With paracetamol treatment
N = 407 N = 71 N = 450
Anti-D Antibody-Serology-IU/mL % Seroprotection 100.0 100.0 100.0
(95% CI) (99.1; 100.0) (94.9; 100.0) (99.2; 100.0)
GM 1.66 1.56 1.50
(95% CI) (1.54; 1.79) (1.33;1.83) (1.39; 1.63)
Anti-T Antibody-Serology-IU/mL % Seroprotection 100.0 100.0 100.0
(95% CI) (99.1; 100.0) (94.9; 100.0) (99.2; 100.0)
GM 2.11 2.01 1.75
(95% CI) (1.92; 2.31)a (1.61;2.50) (1.61; 1.91)a
Anti-PRP Antibody-Serology-mcg/mL % Seroprotection 99.3 100.0 99.8
(95% CI) (97.9; 99.8) (94.9; 100.0) (98.8; 100.0)
GM 8.77 9.38 9.22
(95% CI) (7.92; 9.72) (7.23; 12.2) (8.42; 10.1)
Anti-HBs Antibody-Serology-mIU/mL % Seroprotection 97.8 97.2 98.4
(95% CI) (95.8; 99.0) (90.2; 99.7) (96.8; 99.4)
GM 1035 592 907
(95% CI) (883; 1215) (391; 897) (785; 1047)
Anti-wP Antibody-Serology-Novotech units % Seroresponse 69.5 74.6 70.7
(95% CI) (64.7; 73.9) (62.9; 84.2) (66.2; 74.8)
GM 17.5 18.5 17.5
(95% CI) (15.9; 19.3) (14.5; 23.6) (16.1; 19.0)
Anti-PT antibody-Serology-IU/mL % 4-fold rise 63.6 69.0 66.0
(95% CI) (58.8; 68.3) (56.9; 79.5) (61.4; 70.4)
GM 16.1 17.1 24.8
(95% CI) (13.2; 19.6)a (10.1; 29.1) (20.6; 29.8)a

Note: Number of subjects (N) in each group for all the valences was not same and varied with a maximum difference of 3.
a
Non overlapping 95% CI.
3004 A. Sil et al. / Vaccine 35 (2017) 2999–3006
3.3. Anti-PRP immunogenicity
Similar rates of seroprotection (at least 99.3%) and GMT for anti-
PRP were observed in all the groups irrespective of vaccine
received and the paracetamol use (Tables 1 and 2).
3.4. Anti-HBs immunogenicity
Similar rates of seroprotection (at least 96.7%) for anti-HBs were
observed in all the groups irrespective of vaccine received and the
paracetamol use (Tables 1 and 2).
In the investigational group, the seroprotection rate was
numerically lowest in ‘‘With paracetamol prophylactic’ subgroup
(96.7%). In the comparator group, there was no numerical differ-
ence in the seroprotection rates among the subgroups.
GM antibody titers for anti-HBs were numerically lowest in the
‘With paracetamol prophylactic’ subgroup of the investigational
and comparator groups (Table 1). It was also lowest in ‘With parac-
etamol prophylactic’ group in the pooled analysis irrespective of
the vaccine received (Table 2) However, the above differences were
not considered to have any significance due to the overlapping of
95% CI.
3.5. anti-wP immunogenicity
Similar rates of seroresponse for anti-wP were observed in all
the groups irrespective of vaccine received and the paracetamol
use. At least 68.3% of subjects achieved seroresponse in all the
groups (Tables 1 and 2).
The seroresponse and the GM antibody titers for anti-wP were
numerically highest in the ‘With paracetamol prophylactic’ sub-
group of the investigational and comparator groups (Table 1). They
were also highest in ‘With paracetamol prophylactic’ group in the
pooled vaccine analysis (Table 2).
However, the differences were not considered to have any sig-
nificance due to the overlapping of 95% CI.
3.6. Anti-PT immunogenicity
Similar rates of seroresponse (4-fold rise) for anti-PT were
observed in all the groups (Tables 1 and 2). At least 61.9% of sub-
jects achieved seroresponse among all the groups.
Numerically, the proportion of subjects achieving seroresponse
was observed to be highest in ‘With paracetamol prophylactic’ sub-
group of the investigational group and also among the pooled vac-
cine analysis groups. However, in the comparator group, the lowest
seroresponse rate was observed in ‘With paracetamol prophylactic’
subgroup. But these differences were insignificant.
Some differences in GM antibody titers for anti-PT among the
groups were observed.
In the investigational group, the anti-PT GMT in the subgroup
‘Without Paracetamol’ (16.0 IU/mL) was observed to be lower than
the ‘With paracetamol treatment’ subgroup (25.1 IU/mL) without
overlapping 95% CI. While the anti-PT GMT in the ‘With Paraceta-
mol prophylactic’ subgroup was similar to both the ‘Without
paracetamol’ and ‘With Paracetamol treatment’ subgroups. In the
comparator group, the anti-PT GMTs were similar among the 3
subgroups irrespective of type of paracetamol use (Table 1).
In the pooled analysis also, some differences in anti-PT GMT
among the groups were observed. The GMT in ‘Without paraceta-
mol’ group (16.1 IU/mL) was observed to be numerically lower
than that of ‘With paracetamol treatment’ group (24.8 IU/mL)
without overlapping of 95% CI. While the GMT of ‘With paraceta-
mol prophylactic’ group was similar to both ‘Without paracetamol’
and ‘With paracetamol treatment’ groups (Table 2).
4. Discussion
Fever and pain are common adverse reactions following immu-
nization. Many clinicians therefore prefer to advice medications to
prevent or treat such reactions. There is literature evidence to sug-
gest that changing injection technique may also decrease pain
response [15]. In this study, it was observed that paracetamol
(Acetaminophen) was the most commonly used antipyretic/anal-
gesic. This could be due to its easy availability as an over-the-
counter (OTC) medication in India, low cost, high therapeutic index
and availability in multiple types of formulations. As per one pub-
lished study, administration of paracetamol as a prophylactic with
each vaccine dose decreased the immunogenicity titers when com-
pared with the subjects who did not receive paracetamol. However
the article also states that the antibody titers do not decrease
below the protective level [5]. In another study, it was also
observed that the administration of paracetamol as a prophylactic
decreased the immunogenicity titers when compared with the
subjects who received paracetamol as a treatment [4]. One more
study observed that the lower immune response with prophylactic
paracetamol after vaccination is of transient nature [16].
In our post hoc analysis, similar immune responses for anti-PRP,
anti-HBs and anti-wP were observed in all groups, irrespective of
paracetamol usage and the vaccine received.
There was no observed difference among the groups in anti-D
seroprotection rates as all the subjects in the study achieved sero-
protection levels. In case of anti-D GMT, the subjects in the inves-
tigational group had similar GMT levels irrespective of
paracetamol use. In the comparator group, subjects who used
paracetamol as treatment had similar GMT levels as that of sub-
jects who did not use paracetamol, while subjects who used parac-
etamol as prophylactic had markedly lower GMT levels. However,
no such difference was observed with prophylactic paracetamol
use when analyzed irrespective of the vaccine received (in pooled
vaccine analysis). Moreover the observed difference may not have
clinical relevance as all the study subjects achieved seroprotective
levels irrespective of paracetamol usage and the vaccine received.
No difference was observed among the groups in anti-T sero-
protection rates as all the subjects in the study achieved seropro-
tection levels. There was also no observed difference in anti-T
GM levels with paracetamol use when analyzed separately for each
vaccine. But in case of pooled vaccine analysis, the subjects who
received paracetamol for treatment had much lower anti-T GM
titers than those who did not receive paracetamol, while there
was no difference when paracetamol was used as prophylactic.
However, the observed difference may not be clinically relevant
as all the study subjects achieved seroprotective levels irrespective
of paracetamol usage.
Similar seroprotection rates for anti-HBs were observed in all
the groups. With respect to anti-HBs GMT, the subjects who
received paracetamol as prophylactic had much lower titers than
those who did not receive paracetamol. Use of prophylactic parac-
etamol was associated with lower GMT even when evaluated sep-
arately for investigational and comparator vaccines. The difference
was not clinically relevant since not only the 95% CI of GMT over-
lapped for all the groups but the seroprotection rates amongst the
various groups were also comparable. This effect of paracetamol
usage on anti-HBs is in concordance with an earlier study which
found similar results in Dutch adults. However, even in the men-
tioned study the anti-HBs level in the prophylactic paracetamol
group (4257 mIU/mL) was well above the seroprotective cut-off
level (10 mIU/mL) [5]. In the present analysis also, the anti-HBs
level in the prophylactic paracetamol group (592 mIU/mL) was
well above the seroprotective cut-off level. Even if the Hepatitis
B titer was in the range of 500 mIU/mL (With paracetamol prophy-
lactic group), it was far above the protective level i.e. 10 mIU/mL.
 A. Sil et al. / Vaccine 35 (2017) 2999–3006
Published literature indicates minimal clinical risk to duration and
boostability of immune response even with a relatively lower GMT
i.e. the titers remain above the protective level after 12 months
after the 3rd dose (time point for booster dosing for Hepatitis B)
even with a relatively lower GMT [8].
With respect to anti-pertussis immunogenicity, similar immune
responses for anti-wP were observed in all groups. Similar rates of
seroresponse for anti-PT were also observed in all groups but there
were some differences in GMT. In the investigational group and in
the pooled vaccine analysis, the anti-PT GMT was highest in sub-
jects who had taken paracetamol as treatment and lowest in sub-
jects who had not taken paracetamol. However, this difference
observed in the anti-PT immune response may be clinically
insignificant as no difference was observed in anti-wP immune
responses among the groups. It is possible that in children whom
the GMTs of PT antibodies (the most important mediator of clinical
protection for pertussis) were higher, a fever resulted, which had to
be treated with paracetamol.
In this analysis, among subjects who received antipyretics/anal-
gesics all of them received paracetamol alone or in combination
with other analgesics like ibuprofen and mefenamic acid. Since
all the subjects who received antipyretic/analgesic had received
paracetamol, the group was simply termed as ‘With Paracetamol’
group. Making a sub set analysis of the paracetamol alone vs parac-
etamol combination was deemed unnecessary for this analysis as
only 0.3% of subjects received paracetamol combined with another
antipyretic medication.
Health authorities of various countries have provided guideli-
nes on paracetamol use at the time of vaccination: The US Advisory
Committee on Immunization Practice recommends that ‘‘it is rea-
sonable to consider administering antipyretics (such as Acetami-
nophen) at age-appropriate doses at the time of vaccination and
every 4–6 h for 48–72 h to children at higher risk for seizures than
the general population” [17]. Public Health Agency of Canada rec-
ommends that paracetamol can only be used with whole-cell per-
tussis vaccines rather than with acellular pertussis vaccines,
because of the higher incidence of fever associated with whole-
cell pertussis vaccines than acellular pertussis vaccines [18]. Two
studies conducted in India observed that there is an overall bene-
ficial effect of prophylactic paracetamol in reducing the adverse
effects following vaccination. Use of prophylactic paracetamol
was observed to reduce the non-compliant parents and the immu-
nization drop out cases [19,20]. However, there are no specific rec-
ommendation/guidance from the Indian Pediatric association or
national health authorities regarding the use of antipyretics during
vaccination.
The impact of paracetamol usage as well as mode of usage dur-
ing vaccination was evaluated in this post hoc analysis of the
immunological data from a recent clinical trial [6]. However, the
dose of paracetamol administered was unknown, since analysis
of paracetamol use was not planned as part of the study protocol.
The paracetamol group was not categorized based on dose or num-
ber of occurrences of paracetamol use during the study. The ad’hoc
groups constructed were based on post-medication data and hence
randomization was not possible. Therefore the possibility of bias
cannot be negated completely. In the original study, the subjects
were randomized into investigational and comparator vaccine
groups but they were not randomized according to paracetamol
use. These are some of the possible limitations of the present anal-
ysis. However since the data was collected as part of the original
protocol and similar proportion of subjects received paracetamol
(either as prophylactic or as treatment) in both the investigational
and comparator vaccine groups, the possibility of bias is minimal.
This study provides additional information on effect of parac-
etamol with vaccine administration and the results indicate that
there appears to be no negative or positive impact of paracetamol
3005
administration, irrespective of its use as a prophylactic or for treat-
ment, on vaccine immunogenicity.
5. Conclusion
In this post hoc analysis study on the possible impact of parac-
etamol use on immune responses to DTwP-HepB-Hib combination
vaccine in Indian infants, no clear trend was observed. The differ-
ences observed in GMT values between the various study groups
are minimal and do not have any clinical relevance.
Contributions
AS, MDR and MSD conceived and designed the study and were
involved in the analysis and interpretation of data and results
along with MD, BNP and MVJ. All the other authors from the sites
(MDR, DJG, SMD, APD, RK, SKL, JC, LGM, MG, SDS and SBB) were
principal investigators and were involved in the daily conduct of
the study at their respective sites, study procedures, collection of
data and the patient medical care. All the authors had access to
the data and were involved in either writing or review of the
manuscript and interpretation of data and results.AS, MDR, BNP,
MSD, SPR, MVJ, NA, SM, MD, SR and JE were involved in the final-
ization of the manuscript. All the authors agree to the interpreta-
tion of the data and the representation of the results as
presented in the manuscript. All authors fulfill ICMJE criteria in
the manuscript.AS will stand as the guarantor for the paper.
Conflict of interests
The site investigators have no financial interest in the vaccine or
the manufacturer but received research grants from Shantha
Biotechnics Private Limited –a sanofi Company; for conducting
the study at their respective sites. AS, BNP, MSD, SPR, MVJ, NA,
SM, SR and JE were all employees of the manufacturer of the inves-
tigational vaccine (Shantha Biotechnics Private Limited – a Sanofi
Company) and MD is an employee of Sanofi Pasteur, France
involved during the planning, analysis and interpretation of the
study.
Financial disclosure
The study was fully funded by Shantha Biotechnics Private Lim-
ited –a Sanofi Company.
Acknowledgements
The authors would like to thank the parents of all study subjects
for consenting to participate in this research study. Contributions
of all the clinicians and para-medical staff of the Departments of
Pediatrics JSS Medical college Mysore, SBKS MI & RC, Sumandeep
Vidyapeeth, Vadodara, KLE University’s J N Medical College, Bela-
gavi, Maulana Azad Medical College Delhi, Institute of Child Health
Kolkata, Bharati Vidyapeeth Deemed University Medical College
Pune, Christian Medical College, Ludhiana, Christian Medical Col-
lege, Vellore, SMS Medical College Jaipur, Topiwala National Med-
ical College and BYL Nair Ch. Hospital, Mumbai, and Department of
Community Medicine, School of Public Health, PGI Chandigarh are
thankfully acknowledged. We also acknowledge the staff of Cog-
nizant India Ltd. (For data management), Quest Diagnostic India
Ltd. and Focus diagnostics USA (For Laboratory analysis).
3006 A. Sil et al. / Vaccine 35 (2017) 2999–3006

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