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Original Articles Indian Pediatrics 2002; 39:427-436 |
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Safety and Immunogenicity of Haemophilus Influenzae type B Vaccine Given in Combination with DTwP at 6, 10 and 14 Weeks of Age |
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Manuscript received:
August 22, 2001, Initial review completed: September 25, 2001,
CHILDHOOD immunization programs have been responsible for reducing mortality and morbidity due to many diseases. A cornerstone of these programs is the combined diphtheria-tetanus-pertussis vaccine that allows administration of three antigens in one injection. This has greatly facilitated the inclusion of these three agents in vaccination schedules, with consequent improvements in compliance and coverage. As new pediatric vaccines have been introduced to protect against other diseases, efforts have been made to improve their acceptance by including them with the DTwP combination. To achieve this, various DTwP combinations have been developed to include vaccines against Hepatitis B, Poliovirus and Haemo-philus influenzae type b (Hib), which can be administered in one injection. During the development of such combinations great care has to be taken to ensure that the co-administration does not interfere with the safety, tolerability, immuno-genicity and eventually clinical efficacy of the individual components. To this end vaccine manufacturers may make slight alterations to their formulations to ensure that inclusion of a new antigen is acceptable. However, medical practitioners are often faced with the prospect of giving two vaccines from different manufacturers as separate injections simply because the combined product is not available, or when the DTwP vaccine procured for the national EPI program is from a different manufacturer. Such is the case in India where Hib immunization is recommended but not mandatory(1). At the time of the present study two monocomponent Hib conjugate vaccines were available in India, differing in the protein carrier used; HbOC (Hib TITER, Wyeth Lederle) in which oligosaccharides of the Hib capsular polysaccharide (PRP) are conjugated to a non-toxic mutant diphtheria toxin (CRM 197), and PRP-T (Act-HibTM, Aventis Pasteur) in which PRP is conjugated to tetanus toxoid. Act-Hib is also supplied in the form of a DTwP-combination vaccine (Tetr Act-HibTM). However, supply logistics may require that the reference Act-Hib vaccine be administered with a locally produced DTwP vaccine. It is likely that vaccines containing whole cell pertussis vaccine will increasingly be those produced by local manufacturers as developed countries move to using preparations containing acellular pertussis vaccine. The present study was performed to determine whether extemporaneous mixing of Act-Hib with a DTwP vaccine manufactured locally (BE, Hyderabad, India) interfered with the clinical protection afforded by the Hib vaccine. The combined DTwP-Hib (TetrAct-HibTM) vaccine was used as comparator. Subjects and Methods The study was performed in the Well Baby and Immunization Clinic of the Christian Medical College Hospital, Vellore, Tamil Nadu, India in accordance with the current Declaration of Helsinki, GCP and ICH regulatory guidelines. Approval was obtained from the Institutional Research Committee and written informed consent was obtained from the parent or guardian of all enrolled subjects. A test license was obtained from the Drugs Controller General of India. This was an open, comparative, randomized study. Three groups of subjects received three primary doses of the respective vaccines according to the national EPI schedule at 6, 10 and 14 weeks of age. The primary objective was to demonstrate that the immunogenicity of the Act-Hib vaccine when combined with the locally produced DTwP vaccine (BE) was non-inferior to the reference DTwP-Hib combination vaccine, TetrAct-Hib (Aventis Pasteur, France). For non-inferiority the largest clinically acceptable difference between the two vaccines was not to exceed 10% in terms of anti-PRP seroprotection rate (defined as the percentage of subjects with an anti-PRP response ³1 µg/ml). With a type I error of 5% and type II error of 10% (i.e., power = 90%), 130 evaluable subjects per group were necessary, but in anticipation of a drop-out rate (including missing blood samples) of 19%, 160 subjects were to be enrolled for each group. A third group of 60 subjects was to receive the BE-DTwP and Act-Hib vaccines concomitantly, but at separate injection sites, to observe differences between these vaccines when given as a single combined injection or in association. Secondary objectives were to verify that the antibody responses to diphtheria, tetanus and pertussis were not impaired when the antigens were presented in combination with the Hib vaccine by measuring the immuno-genicity of the BE and Aventis Pasteur DTwP vaccines, and to assess the safety of these vaccines. Subjects Intended subjects were healthy infants between 6-8 weeks of age, of either sex, who did not have one of the exclusion criteria, i.e., prior immunization with Hib vaccine, participation in another clinical trial, current illness (especially fever) or any acute or congenital illness or disability, undergoing immunosuppressive therapy, known or suspected allergy to any of the vaccine components. Children were to be withdrawn from further participation in the study if after any dose they displayed seizures, encephalopathy or other serious adverse event. Vaccines Each dose of the BE-DTwP vaccine contains at least 25 Lf diphtheria toxoid, 5 Lf tetanus toxoid and 4 IU inactivated Bordetella pertussis with 1.5 mg aluminium phosphate and 0.01% (w/v) thiomersal in 0.5 mL buffered saline. The DTwP component of TetrAct-Hib contains at least 30 IU diphtheria toxoid, 60 IU tetanus toxoid and 4 IU inactivated Bordetella pertussis with 0.65 mg aluminium phosphate and 0.05 mg thiomersal in 0.5 mL buffered saline. Act-Hib vaccine is presented in lyophilized form, each dose containing 10 µg Hib polysaccharide conjugated to tetanus toxoid with 0.6 mg Tris and 42.5 mg sucrose. When administered alone this was reconstituted extemporaneously in 0.5 ml saline, for TetrAct-Hib it was reconstituted in the liquid DTwP component and the same procedure was applied for combined adminis-tration with the BE-DTwP vaccine. All vaccines were given by intramuscular injec-tion in the antero-lateral thigh (opposite thighs for separate DTwP and Hib was used for infants in the third group, i.e., vaccines given in association). Due to the different presentations it was not possible to mask those administering the vaccines, though the parents of children in groups A and B were masked to the study allocation. Safety and Reactogenicity Subjects were monitored in the clinic for 30 minutes after each injection. Parents were supplied with diary cards on which to record any local or systemic adverse events. The cards solicited parents to record any signs of local or systemic reactions such as redness, swelling, pain, fever or irritability during the 72 hours immediately following vaccination, together with the intensity of any such events. Parents were supplied with a special scale to measure the diameter of redness or swelling, and a digital thermometer to measure axillary temperature in case of fever. Instructions on the use of the diary, scale and thermometer were provided during the first visit and were reviewed during subsequent visits. Any serious adverse event was to be reported immediately to the investigator. One of the study physicians checked the diary cards and discussed any events with the parents at the subsequent visit, in order to assess the relationship with the vaccination. As this was an open study only a descriptive analysis of the reactogenicity data is presented. Serology Blood samples were drawn at the first visit before vaccination, and at the final visit 28 days after the third dose. Sera were prepared immediately and maintained at –20ºC during transit to Aventis Pasteur, Val-de-reuil, France for antibody determinations. Antibodies to PRP were determined by radioimmunoassay (RIA), titers being expressed in µg/ml with a cut-off of 0.15 µg/ml to define seropositivity and ³1.0 µg/ml as a mark of long-term seroprotection(2). Anti-diphtheria and anti-tetanus antibodies were measured by ELISA and expressed in IU/ml. Titration of Bordetella pertussis antibodies was performed using a quantitative agglutination method. All sero-logical analyses were performed in a blinded fashion. Where serum volumes were low, priority was given to determining the anti-PRP response. Results A total of 378 infants were enrolled and randomly allocated to the three groups, 160 receiving Act-Hib combined with BE-DTwP vaccine (Group A), 160 the TetrAct-Hib combination vaccine (Group B) and 58 BE-DTwP associated with Act-Hib which was administered in the opposite thigh (Group C) (Table I). There were 7 subjects withdrawn from the study, 4, 1 and 2 from groups A, B and C, respectively. Four of these withdrawals were at the request of the parents and 3 due to relocation of the family. Immunogenicity Evaluable paired sera were available from 364 subjects (Table I), 1 subject from Group A inadvertently received Tetr Act-Hib for the first dose and a further 6 subjects were not included due to protocol violations ( 3 subjects were not 6 weeks old at time of first vaccination, the second blood sample was not taken according to schedule in 2 subjects, and 1 infant did not receive the third dose on schedule). Table I__Demographics of the Study Population
*ATP = According to protocol, # Subject mistakenly administered TetrAct-Hib kept in Group B for analysis.
Geometric mean concentrations (GMC) of anti-PRP antibodies one month after the third vaccination are shown in Table II. All three groups achieved high mean antibody levels from 16.9 to 17.5 µg/ml. In group A the combined administration of BE-DTwP and Act-Hib induced long-term seroprotective anti-PRP concentrations (³1.0 µg/ml) in 96.7% of the subjects, and seropositivity (³ 0.15 µg/ml) in all but one child (99.3%). All subjects in Group B, given Act-Hib as part of the TetrAct-Hib combination, achieved the seroprotective anti-PRP concentration (100% seropositivity), giving a difference of 3.3% (90% CI, 0.28%; 7.52%) between Groups A and B. As the upper limit of the 90% CI was lower than 10% this showed non-inferiority of the BE-DTwP/Act-Hib combination with respect to TetrAct-Hib. In Group C all infants given Act-Hib as a separate injection were seropositive with only one not achieving the seroprotective concentration (98%) indicating that the combination with BE-DTwP did not interfere with the response to Act-Hib. Table II-Anti-PRP Responses in the Three Study Groups Pre-Vaccinaltion and One Month After the Third Dose
Immune responses to the three components of the DTwP vaccines are shown in Table III. Protection against diphtheria and tetanus is attained at titers between 0.01 and 0.1 IU/ml(2). A large proportion of the subjects already displayed antidiphtheria titers, probably due to maternal antibodies. After vaccination all subjects given TetrAct-Hib achieved the higher concentration against diphtheria, but while all subjects given BE-DTwP achieved 0.01 IU/ml, only 93.9% in Group A and 96.4% in Group C achieved 0.1 IU/ml. This was reflected in the GMTs, TetrAct-Hib eliciting a higher anti-diphtheria titer than BE-DTwP as a combination or a separate injection. Table III-Immune Response to Components of the DTwP Vaccines in the Three Study Groups
Almost all the children in each group were already seroprotected ( ³0.1 IU/ml) against tetanus before vaccination (96.8-98.2%). Although seroprotection was maintained in all subjects post-vaccination, when some decline in the maternal antibody levels would have been anticipated, only TetrAct-Hib elicited a significant increase in GMT for tetanus, from 1.36 to 3.27 IU/ml. There is no currently accepted serologic correlate for pertussis, but the table shows a marked increase in titers in all three groups such that all subjects had a titer ³1 : 80 dilution. The response was similar in all three groups. Reactogenicity Reactogenicity data are reported for all children who received vaccinations. The most clinically important anticipated systemic reaction to DTwP vaccination is fever(3). As shown in Table IV, more episodes of fever occurred with BE-DTwP (40.4-66.0% of doses in Group A and 46.4-74.1% of doses in Group C) than with TetrAct-Hib (23.8 – 31.1% of doses in Group B). In all three groups the majority of cases were mild (axillary temperature from 37.5% to 38.4ºC) or moderate (38.5º to 39.4ºC), but there were some reports of severe fever (³39.5ºC), 6, 3 and 1 in groups A, B and C, respectively. The other solicited systemic symptom was irritability, defined to include unusual or inconsolable crying, which was also reported more frequently with BE-DTwP than TetrAct-Hib (63.9-80.0%, 55.6-64.5% and 55.4-74.1% of doses in Groups A, B and C, respectively) and with proportionally more severe cases (Table IV). Three other systemic symptoms were also reported as being possibly related to the vaccination (defined as a temporal association between vaccination and illness, but the etiology not investigated). They included one case of mild diarrhea after the first dose in Group A, and two cases of mild upper respiratory tract infection, one each in groups A and B, after the second vaccination. All systemic symptoms resolved without sequelae. Table IV-Percentage of Subjects in Each Study Group for Which the Solicited Systemic Symptoms, Fever and Irritability, were Reported After Each Vaccine Dose, and the Severity According to the Parents/Guardians
The majority of infants were reported to suffer at least one local reaction to each dose (Table V). There was a trend to lower rates with TetrAct-Hib than with both of the BE-DTwP groups (the site of BE-DTwP injection being considered in Group C), and in each group a trend to fewer reactions to subsequent doses. The most frequently reported local symptom was pain at the injection site (Table V). Most cases were mild or moderate but 13.8-15.4%, 5.0-8.7% and 15.5-19.6% of doses in Groups A, B and C, respectively, were reported to cause severe pain (infant cried when limb was moved). The reporting rates were similar for BE-DTwP when administered combined or associated with Act-Hib showing that the combination did not further aggravate the reactions to this vaccine. Importantly however, the separately administered Act-Hib produced almost as many local reactions as the DTwP. Giving the two vaccines in combination therefore effectively halved the number of such reactions. Discussion The incorporation of additional antigens into DTwP combinations has been readily accepted as a means to include new vaccines into pediatric vaccination schedules while promoting compliance by limiting the number of additional injections required. Such combi-nations also simplify logistic and adminis-trative functions and cold-chain problems by keeping the necessary material to a minimum. Their use however is conditional upon the combination not affecting the tolerability or immunogenicity of the individual vaccine components. For this reason vaccine manufacturers have to extensively test any new such combination in comparison with the separately administered components. Table V-Percentage of Subjects in Each Group for Which atleast One of the Solicited Local Symptoms, Pain, Redness and Swelling, were Reported After Each Vaccine Dose.
In the face of supply difficulties and parental pressures to minimize discomfort, physicians may be tempted to use extemporaneous vaccine mixing to prepare combinations of vaccines from different manufacturers. Such combinations will not have been tested with such use in mind and so it is important to ensure that the safety and efficacy of such combinations is tested in advance. This issue was recently raised in the USA where the FDA found it necessary to instruct medical practitioners not to coadminister Hib vaccines from the same syringe as DTwP vaccines containing acellular pertussis(4). In India there were only two Hib conjugate vaccines available at the time of this study, one of which, Act-Hib, is also supplied in combination with the DTwP vaccine from the same manufacturer as TetrAct-Hib. Although not part of the national infant vaccination program, the high immunogenicity and exceptional safety of Act-Hib(5) makes Hib vaccination a wise precautionary measure. This is especially true when one considers the fact that Hib is the commonest cause of bacterial meningitis in infants and young children in India(6). Several studies have documented the safety and immunogenicity of DTwP-Hib combinations(7,8). However, DTwP vaccines available through the EPI program in India are often those produced by local manufacturers. Therefore, it would be natural for practitioners to expect to be able to coadminister Hib vaccine with the locally produced DTwP vaccine. Previous studies in India have documented the immunogenicity of the Hib vaccine administered along with locally available DTwP vaccines but were not designed to allow regulatory approval of the combination and did not measure the immune response to all the antigens in the combination(9-10). Our present data, based on mixing Act-Hib with BE-DTwP, confirm that combination of Hib vaccine with DTwP produced by at least one local manufacturer is safe and immunogenic. However, the combination of Hib vaccine with DTwP produced by other local manufacturers may need to be documented before they are approved to routine use. The advantage of lower rates of local reactions due to only one injection being given with no increase in general reactogenicity would increase the acceptability of this "non-official" combina-tion and hence compliance with the Hib vaccination. The high rates of seroprotective responses attained with TetrAct-Hib are in accordance with other published data for this vaccine from studies performed in the Gambia(11), Belgium(12), USA(13) and Thailand(14). While not a specific goal of the study, and hence not analyzed statistically, it does appear evident from the data that the Aventis Pasteur DTwP vaccine component elicits higher antibody levels to diphtheria and tetanus. Indeed, TetrAct-Hib was the only one of the two vaccine to increase the anti-tetanus titers over the pre-vaccination level. However, both vaccines elicited an immune response that may be considered protective. We conclude that on preference the reference vaccine, TetrAct-Hib should be administered wherever possible, but if the supply situation dictates otherwise, it is safe to reconstitute and administer Act-Hib as a combination with BE-DTwP vaccine. Combination with other DTwP vaccines is possible but needs to be confirmed. Acknowledgement The authors gratefully acknowledge the assistance of Ms. M. Rajarshi for local monitoring support, the research medical officers Drs. Sneha Varkki, Ranjeetha Ambett, Beneta Sam and Jeyanthi Jacob, field worker Mr. Andrew, and the study nurses Ms. Sridevi, Ms. Thilagavathy and Ms. Sasirekha, for their assistance with patient enrolement and follow up. The authors would also like to thank Dr. M.K. Lalitha for assistance with prepara-tion of sera and storage of vaccines and sera, and Keith Veitch for his advice and help in reviewing the manuscript. Contributors: TC was the principal investigator and was involved in conceiving and designing the study, supervising all aspects of the study, analysis of data and manuscript preparation; he will act as the guarantor for the paper. NT was a co-investigator and provided day-to-day supervision of patient enrolment, follow up of all reported serious adverse events and participated in data analysis and manuscript preparation. PR was a co-investigator and participated in designing the study and provided supervision for patient enrolment and follow up. AD and ID were involved in designing the study, providing logistical support, supervising shipment of vaccines and sera, serological testing of sera, data management and manuscript preparation. Funding: Financial support and vaccines for the study were provided by Aventis Pasteur International, Lyon, France. BE DTwP was supplied by Biological E, Hyderabad, India. Competing interests: AD and ID are employees of Aventis Pasteur, International, Lyon. None of the other authors have a commercial interest in either of the vaccine companies.
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