Current childhood immunization guidelines in the United States call for up to 19 different injections in the first 6 months of an infant’s life, with as many as 5 injections at a single visit, if licensed monovalent vaccines are administered.1 Over the next several years, infant immunizations are likely to become even more complex as additional vaccines reach the marketplace. Some physicians and parents are reluctant to administer multiple injections at a single visit, which may result in deferred vaccination or extra office visits.2–7 If follow-up visits are missed, the introduction of new vaccines may result in an overall decline in vaccine coverage rates and, ultimately, increased disease burden.
Combination vaccines offer a practical solution to the increasingly complex and crowded childhood immunization schedule by improving on-time compliance with recommended vaccination schedules, while reducing the number of injections required during the first 2 years of life.8–13 By reducing the number of needle sticks, combination vaccines decrease fear and pain in infants and toddlers and improve acceptance among providers and parents.14,15 The use of combination vaccines can improve the day-to-day efficiency of the practice because they reduce the cost associated with preparation and administration of each of the individual vaccines and potentially reduce clinic visits.9,16 Finally, the inclusion of new or additional vaccines in existing programs is facilitated by the use of combination vaccines.17
Despite the advantages of combination vaccines regarding compliance, new combination vaccines should be as safe and effective as their component vaccines. Simultaneous exposure to multiple antigens can result in either enhanced or diminished immune responses.18 Regarding immunogenicity, combination vaccines should be noninferior to the separately administered component vaccines. Adverse events (AEs) after administration of combination vaccines should also be reported at similar frequencies compared with the rates reported after concomitant administration of separately injected component vaccines.
Two phase IIa studies in infants and toddlers tested various concentrations of the Haemophilus influenzae type b (Hib) and hepatitis B virus (HepB) components within a liquid hexavalent DTaP-IPV-Hib-HepB combination vaccine that also contained a 5-component acellular pertussis component (DTaP5).19,20 The studies revealed robust antibody responses and good safety profile of the DTaP5-IPV-Hib-HepB vaccine, a liquid, preservative-free, hexavalent combination vaccine developed for active immunization against diphtheria, tetanus, pertussis, poliovirus (types 1, 2 and 3), Hib and HepB.
PARTICIPANTS AND METHODS
This clinical trial was a phase IIb, randomized, open-label, multicenter study conducted at 8 sites in Canada. The study protocol and informed consent forms were approved by the institutional ethics committee at each study site. Parent(s) or legal guardian(s) of all participants provided written consent before initiation of any study-specific procedures. The trial was conducted in compliance with the Declaration of Helsinki.
The overall aim of the study was to assess the safety and immunogenicity of a candidate DTaP5-IPV-Hib-HepB vaccine. A secondary aim was to corroborate that schedules consisting of 3 doses of DTaP5-IPV-Hib-HepB with a heptavalent pneumococcal conjugate vaccine (PCV7) given either concomitantly or 1 month apart provide comparable safety and immunogenicity to those of separately administered licensed vaccines used for routine infant vaccination in Canada. The control vaccines were chosen as representative examples of the standard of care in Canada during the time the study was conducted.
Eligible participants were aged 42–89 days at study entry and had been born after a full-term pregnancy (>37 weeks). Eligible participants should not have: a personal or immediate family history of congenital or acquired immunodeficiency; received immunosuppressive therapy; known or suspected systemic hypersensitivity to any of the vaccine components; chronic illness that could interfere with trial conduct or completion; received blood or blood-derived products; any vaccination preceding the first trial vaccination or planned in the 4 weeks after any trial vaccination; coagulation disorder contraindicating intramuscular vaccination; developmental delay or neurologic disorder; documented seropositivity to HepB surface antigen (HBsAg) in the participant or the participant’s mother; or history of Hib, HepB, diphtheria, tetanus, pertussis or poliovirus disease.
The following vaccines were administered in this study: liquid, preservative-free hexavalent DTaP5-IPV-Hib-HepB vaccine (Sanofi Pasteur Inc., Swiftwater, PA; Merck & Co., Whitehouse Station, NJ); DTaP5-IPV/Hib vaccine (Pentacel; Sanofi Pasteur Inc., Toronto, Ontario, Canada); HepB vaccine (Engerix-B; GlaxoSmithKline, Research Triangle Park, NC); and PCV7 vaccine (Prevnar, Pfizer Inc., New York, NY). Components of all vaccines are shown in Table 1.
At study entry, healthy infants were randomly assigned in a 1:1:1 ratio to 1 of 3 treatment groups: DTaP5-IPV-Hib-HepB administered concomitantly with PCV7 (group A), DTaP5-IPV-Hib-HepB followed by PCV7 1 month later (group B) and DTaP5-IPV/Hib plus HepB plus PCV7 administered concomitantly (group C). Participants in group A and group C received all assigned vaccinations at ages 2, 4 and 6 months. Participants in group B received DTaP5-IPV-Hib-HepB at ages 2, 4 and 6 months and received PCV7 at ages 3, 5 and 7 months.
DTaP5-IPV-Hib-HepB was given as a 0.5 mL intramuscular injection in the mid-lateral aspect of the thigh, and PCV7 (0.5 mL intramuscularly) was given in the contralateral thigh. Participants were monitored by study staff for 30 minutes after each injection for treatment of any immediate reactions. Parents recorded daily temperature, solicited injection site reactions and systemic reactions between day 0 and day 7 after each vaccination (except for PCV7 administered alone).
Sera were obtained at baseline to assess levels of antibodies to pertussis antigens (pertussis toxoid [PT], filamentous hemagglutinin [FHA], pertactin [PRN], and fimbriae types 2 and 3 [FIM]) and 28–42 days post-dose 3 to assess antibody levels of all DTaP5-IPV-Hib-HepB and pneumococcal antigens.
The primary immunogenicity endpoints were: (1) proportion of participants with seroconversion to pertussis antigens (group A) and (2) proportion of participants with seroprotection against polyribosylribitol phosphate (PRP), HBsAg, diphtheria toxoid, tetanus toxoid and poliovirus antigens (group A). Seroconversion to pertussis antigens was defined as a ≥4-fold rise in antibody concentrations (post-dose 3/pre-dose 1). Seroprotection was defined as anti-PRP antibody concentrations ≥1.0 µg/mL, anti-HBsAg antibody concentrations ≥10 mIU/mL, antidiphtheria toxoid antibody concentrations ≥0.1 IU/mL, antitetanus toxoid antibody concentrations ≥0.1 IU/mL and antipoliovirus types 1, 2 and 3 antibody titers ≥8 reciprocal dilution.
The secondary immunogenicity endpoints were: (1) seroconversion rates for pertussis antigens (as described in the primary immunogenicity endpoint) for groups B and C and (2) geometric mean titers (GMTs) for antibodies against all antigens contained in DTaP5-IPV-Hib-HepB for all study groups and against the PCV7 antigens (groups A and C).
Safety endpoints included (1) the occurrence, time to onset, number of days of occurrence, grading (intensity and seriousness) of solicited injection site and systemic reactions occurring up to 7 days after each vaccination (except PCV7 in group B) and across all vaccinations combined; (2) the occurrence, nature, time to onset, duration, intensity and relationship to vaccination of unsolicited AEs occurring within 30 days after each vaccination (except PCV7 in group B); and (3) occurrence, nature, time to onset, duration and relationship to vaccination of any serious AE (SAE) throughout the study for all groups. AEs were defined using preferred terms from the Medical Dictionary for Regulatory Activities version 9. The intensity of solicited injection site and systemic reactions is described in Table 2.
Antibody titers to diphtheria toxoid were measured by the micrometabolic inhibition test. Antibody titers to tetanus toxoid and pertussis antigens (PT, FHA, PRN and FIM) were assessed by enzyme-linked immunosorbent assay. Antibody titers to poliovirus antigens were measured by neutralization of growth in Vero cells. Antibody titers to PRP were assessed by a Farr-type radioimmunoassay. Antibody titers to HBsAg were measured by an enhanced chemiluminescence sandwich enzyme-linked immunosorbent assay. Response to PCV7 was assessed by antibody titers elicited by to pneumococcal polysaccharide-specific antigens to serotypes 4, 6B, 9V, 14, 18C, 19F and 23F as measured by enzyme-linked immunosorbent assay.
Testing to assess antibodies elicited by diphtheria, tetanus, pertussis and poliovirus antigens was performed by Sanofi Pasteur Inc, Swiftwater, PA. Testing to assess antibodies elicited by PRP, HBsAg and pneumococcal antigens was performed by Merck Research Laboratories (Upper Gwynedd, PA).
A sample size of 159 participants for group A (assuming a 15% dropout rate) was calculated to provide an estimated overall power of 91.1% to detect a difference in immune responses of 13% (for group B) to 15% (for group A) from targeted responses (described below) based on prior studies (data on file, Merck & Co., Inc. and Sanofi Pasteur Inc.). However, the study was not powered for formal between-group comparisons, and there were no predetermined hypotheses to compare results between study groups. Antibody responses were considered to be satisfactory if the lower bound of the 95% confidence interval (CI) was greater than the predetermined lower limit based on target responses defined in the study endpoints.
The predetermined lower bound limits for pertussis antigens for group A were 75% (PT), 67% (FHA), 60% (PRN) and 73% (FIM) and for group B were 77% (PT), 69% (FHA), 62% (PRN) and 75% (FIM). The predetermined lower bound limits for pertussis antigens for group A were determined to be 2% lower than for group B as some minimal interference was expected from the coadministration of pneumococcal vaccine. No lower bounds limits for pertussis antigens were defined for group C as the study was not designed to power for testing the endpoints for group C.
The predetermined lower bound limits for anti-PRP antibody was 70% achieving >1.0 µg/mL; for anti-HBsAg was 80% achieving >10 mIU/mL; for diphtheria and tetanus antibodies were 80% and 85%, respectively, achieving >0.1 IU/mL; for poliovirus types 1, 2 and 3 were 85% achieving with >8 reciprocal dilution.
The safety analysis set comprised all participants who received at least 1 dose of study vaccine and contributed at least 1 postvaccination safety measurement. The intent-to-treat (ITT) analysis set included participants who received at least 1 dose of DTaP5-IPV-Hib-HepB or DTaP5-IPV/Hib vaccine, contributed at least 1 postvaccination blood draw and had a valid serology test result for at least 1 antigen. The per-protocol (PP) analysis set was a subset of participants in the ITT analysis set who had received correct study vaccines and doses and contributed blood samples according to the protocol-specified schedule.
The study was conducted from August 2006 to April 2008 at 8 sites in Canada. A total of 460 participants enrolled, including 157 in group A, 150 in group B and 153 in group C. Of the 460 participants, 459 were included in the safety analysis set, 434 were included in the ITT analysis set and 327 were included in the PP analysis set (Fig. 1). Of the 459 participants in the safety analysis set, 227 (49.5%) were girls and 232 (50.5%) were boys. The mean (standard deviation) age was 63.2 (9.1) days (range: 42–99 days). Most participants were white (78.2% in group A, 81.3% in group B and 80.4% in group C). Demographic characteristics were generally balanced across groups.
Vaccines were well tolerated by the participants, and rates of solicited injection site reactions (Fig. 2A) and solicited systemic reactions (Fig. 2B) were generally similar across the 3 groups. The proportion of participants (95% CI) who experienced at least 1 solicited reaction within 7 days of any dose during the infant series was 96.8% (92.7–99.0) in group A, 98.0% (94.3–99.6) in group B and 96.7% (92.5–98.9) in group C. Tenderness after any dose was experienced by 56.4% (48.2–64.3), 49.3% (41.1–57.6) and 47.7% (39.6–55.9) of participants in groups A, B and C, respectively. Severe tenderness after any dose was experienced by 3.2% (1.0–7.3), 3.3% (1.1–7.6) and 6.5% (3.2–11.7) of participants in groups A, B and C, respectively. Erythema after any dose was experienced by 51.9% (43.8–60.0), 55.3% (47.0–63.4) and 48.4% (40.2–56.6) of participants in groups A, B and C, respectively. Swelling after any dose was experienced by 35.9% (28.4–44.0), 34.0% (26.5–42.2) and 35.9% (28.4–44.1) of participants in groups A, B and C, respectively. Severe erythema or swelling was rare and similar among the 3 groups. Most reactions occurred within the first 3 days after vaccination and resolved within 7 days.
Irritability was the most frequently reported solicited systemic reaction in all 3 groups, with rates (95% CI) of 84.6% (78.0–89.9) in group A, 82.0% (74.9–87.8) in group B and 80.4% (73.2–86.4) in group C. Fever was experienced more frequently in participants in group A (with a rate [95% CI] of 57.7% [49.5–65.6]) than in group B (40.0% [32.1–48.3]) and in group C (41.2% [33.3–49.4]). Nevertheless, severe fever (temperature >39.5°C) was reported with similar frequency (95% CI) in the 3 groups: 0.6 % (0.0–3.5) in group A, 2.0% (0.4–5.7) in group B and 2.0% (0.4–5.6) in group C. Table 3 presents moderate and severe fever frequency rates by dose. There were no febrile seizures reported in any study arm during the infant series. Most solicited systemic reactions occurred within the first 3 days after vaccination and resolved within 7 days; these events were mild to moderate in severity.
Overall, the proportion (95% CI) of participants reporting at least 1 unsolicited AE (including SAEs) from day 0 to day 30 after each vaccination was similar in all groups; 20.0% (14.5–27.7) in group A, 19.3% (13.3–26.6) in group B and 17.6% (12.0–24.6) in group C. The most frequently reported unsolicited AEs were nasopharyngitis (5.8%), diarrhea (3.2%) and injection site induration (2.6%) in group A; nasal congestion (4.0%), cough (2.7%) and nasopharyngitis (2.7%) in group B; and injection site induration, injection site warmth, nasopharyngitis, nasal congestion, cough and rhinorrhea (all 2.0%) in group C. Overall, the safety of DTaP5-IPV-Hib-HepB with respect to unsolicited AEs was similar when given alone or concomitantly with PCV7.
Twenty-four SAEs were reported during the study, including 8 (5.1%) participants in group A, 10 (6.7%) in group B and 6 (3.9 %) in group C. Two participants discontinued from the study because of an SAE: 1 participant in group A reported hypotonia, which resolved overnight without treatment, and 1 participant in group C had fibrosarcoma not related to vaccination. The only SAE considered by the investigator to be vaccine related was the case of hypotonia. Most reported SAEs were respiratory in nature, occurring during winter months However, these SAEs were not associated with a specific vaccine dose, and none were assessed by the investigators as related to vaccination except the single report of hypotonia. No new or unexpected safety issues were identified when DTaP5-IPV-Hib-HepB was given concomitantly with PCV7. No deaths occurred during the study.
At the post-dose 3 time point, the seroconversion rates (% ≥4-fold rise from baseline) elicited by pertussis antigens were 88.1% (81.3–93.0%) for PT, 70.4% (61.9–77.9%) for FHA, 73.6% (65.2–81.0%) for PRN and 90.0% (83.5–94.6%) for FIM in participants in group A (ITT analysis set; Table 4). When the lower bound values were compared with the prespecified lower bound limits (75% for PT, 67% for FHA, 60% for PRN and 73% for FIM, see Participants and Methods), the observed seroconversion response lower bound was higher and met acceptability criteria. The seroconversion rate for FHA did not meet the acceptability criterion (67%). Similarly, seroconversion rates for PT, PRN and FIM met the acceptability criteria for participants in group B, but the rate for FHA did not. Results from the PP analysis set were in agreement with these findings. The pertussis seroconversion rates in groups A and B were similar to those in the control group (group C) except that FHA was modestly higher in group C.
GMTs for antibodies against vaccine antigens are shown in Table 5. Antibody levels against diphtheria toxoid were higher in group B than in the other 2 groups. Antibody GMTs to pneumococcal antigens were similar in the 2 groups tested: groups A and C. Because PCV7 was given concomitantly at each time point in these 2 groups, these data support the notion that other than antidiphtheria toxoid antibodies, there was no effect on immunogenicity of PCV7 when coadministered with DTaP5-IPV-Hib-HepB. The results from the PP analysis set were similar to the results from the ITT analysis set. Seroprotection rates to PRP, HBsAg, diphtheria toxoid, tetanus toxoid and poliovirus antigens were high across all groups and met the criteria for acceptability (Figure 3). For PRP, post-dose 3 seroprotection rates (antibody levels ≥1.0 µg/mL) were 92.7% (87.0–96.4) in group A, 94.0% (88.6–97.4) in group B and 85.4% (78.4–90.8) in group C. For HBsAg, seroprotection rates (antibody levels ≥10 mIU/mL) for group A and group B were high, with >98% of participants achieving seroprotection at post-dose 3. All of the participants (100%) in group C achieved seroprotection (antibody levels ≥10 mIU/mL). For diphtheria, seroprotection rates (95% CI) at post-dose 3 for participants in groups A and C (who received concomitant PCV7) were 89.8% (83.4–94.3) and 90.8% (84.7–95.0), respectively, whereas 100% (97.2–100) of participants in group B achieved seroprotection (antibody level ≥0.1 IU/mL). All participants achieved antidiphtheria antibody titers at levels ≥0.01 IU/mL.21 For tetanus, seroprotection rates (95% CI; antibody level ≥0.1 IU/mL) at post-dose 3 were 100% (97.5–100) in group A, 100% (97.3–100) in group B and 99.3% (96.2–100) in group C. For poliovirus type 3, seroprotection rates (antibody level ≥8 reciprocal dilution) were 100% in all 3 groups. Seroprotection rates (95% CI) for poliovirus type 1 at post-dose 3 were 98.4% (94.2–99.8) in group A, 100% (96.9–100) in group B and 98.3% (94.2–99.8) in group C. Seroprotection rates (95% CI) for poliovirus type 2 at post-dose 3 were 100% (97.1–100) in group A, 99.2% (95.5–100) in group B and 100% (97.1–100) in group C. Thus, although the prespecified criteria for acceptability of seroprotection rates for PRP, HBsAg, diphtheria, tetanus, poliovirus types 1, 2 and 3 in group A were met, there may be evidence of immune interference for antidiphtheria toxoid response from the concomitant PCV7 administration.
The development of combination vaccines raises the possibility of immune interference among the antigen components. Pneumococcal conjugate vaccines are currently licensed for primary and booster vaccination on the same immunization schedules as HepB and the diphtheria-tetanus-pertussis-polio-Hib vaccines. Therefore, vaccine coadministration is tested to determine whether the combination product has similar immunogenicity and safety as the separately administered components.
The purpose of this randomized, open-label, multicenter, phase IIb clinical study was to evaluate the safety and the immunogenicity profile of an investigational, liquid, hexavalent DTaP-IPV-Hib-HepB vaccine (including a 5-component acellular pertussis component) in infants given at 2, 4 and 6 months of age when administered concomitantly or separately from PCV7. Although this study was not powered for formal noninferiority comparisons, the acceptability of antibody responses was evaluated for participants who received either the hexavalent vaccine concomitantly or separately from PCV7 or the control regimen (pentavalent vaccine plus monovalent HepB with PCV7). The formulation selection of this vaccine was based on safety and immunogenicity profiles described in previous phase I and II studies.19,20,22 In those trials, PRP conjugated to an outer membrane protein complex of Neisseria meningitides (3 µg) had the most favorable immunologic response and safety profile when compared to PRP covalently bound to tetanus toxoid formulations or a higher dosage of PRP conjugated to an outer membrane protein complex of Neisseria meningitides. The primary study criterion for the evaluation of the antibody response to PRP was seroconversion rate above a commonly accepted antibody threshold (≥1.0 µg/mL).23 DTaP5-IPV-Hib-HepB surpassed this criterion with high rates of seroprotection of 92.7% (group A) and 94.0% (group B) after the third dose. Concomitant vaccination with PCV7 did not negatively affect the PRP seroprotection rates. This result compares favorably with the rate of seroprotection of 85.5% obtained in infants receiving the routine Canadian immunization schedule (group C). GMTs achieved by the investigative combination vaccine also exceeded those elicited by the current licensed pentavalent product (group C).
The inclusion of HBsAg in the DTaP5-IPV-Hib-HepB vaccine was intended to provide protection against HepB earlier in life, reduce the number of injections and increase vaccination coverage. Several controlled clinical studies have established that a level of anti-HBsAg antibodies ≥10 mIU/mL is indicative of seroprotection against HepB.24,25 Antibody responses to HepB in combination vaccines have been occasionally associated with lower antibody levels.26,27 In our study, no differences were seen among the groups with high HBsAg seroprotection rates (≥10 mIU/mL) of >98% for group A and group B after the third dose. Coadministration of PCV7 did not interfere with response to the HBsAg component of DTaP5-IPV-Hib-HepB vaccine.
In the absence of standard serologic correlates of protection against pertussis,28 seroconversion rates to pertussis antigens were defined as described in the Endpoints section of Participants and Methods. For PT, PRN and FIM, the lower bounds of the 95% CIs for the observed responses were greater than the predefined lower bound limits and met the acceptability criterion. The seroconversion rate for FHA did not meet the acceptability criteria. The biologic explanation for the less than optimal response to FHA antigen is not clear and should be explored in future studies. Nevertheless, the clinical importance of this finding is unknown, given that anti-FHA antibody responses have not been correlated with efficacy against disease and responses to PT, PRN or FIM have been sufficient to confer protection.29,30
In the present study, the PCV7 vaccine was highly immunogenic in early infancy among participants receiving DTaP5-IPV-Hib-HepB and DTaP5-IPV/Hib plus HepB; GMT values for all 7 serotypes after 3 doses were similar to those reported in previous studies in which PCV7 was administrated concomitantly with other recommended pediatric vaccines.31–34 Our results were also consistent with the level of pneumococcal antibodies after vaccination in the pivotal study performed at Northern California Kaiser Permanente,35 although direct comparisons cannot be made between our study and other studies because of different populations, concomitant vaccines and serology laboratories. Antibodies to the pneumococcal antigens were not measured in participants who received PCV7 1 month after administration of DTaP5-IPV-Hib-HepB. Therefore, potential differences in response to pneumococcal antigens when PCV7 was coadministered versus administered 1 month later could not be determined. Mean GMTs for antibodies against diphtheria toxoid were lower when PCV7 was administrated concomitantly with DTaP5-IPV-Hib-HepB or DTaP5-IPV/Hib plus HepB than when PCV7 was given 1 month apart. This finding could be related to an immune interference between the diphtheria protein CRM197, used as a carrier protein in PCV7, and similar antigens contained in the simultaneously administered combination vaccines.36,37
Although the prespecified criteria for acceptability of seroprotection rates to PRP, HBsAg, diphtheria toxoid, tetanus toxoid and poliovirus types 1, 2 and 3 in group A were met, these results suggested that for the diphtheria toxoid antigen there was evidence of immune interference from concomitant administration of PCV. However, considering that a series of 4 vaccine doses will be administered overall, one could anticipate a booster response in diphtheria antibodies on a subsequent dose. The hexavalent vaccine has demonstrated a good safety and tolerability profile as measured by rates of solicited local and systemic reactions and unsolicited AEs and SAEs; rates were generally comparable among the 3 groups, including group C, in which routinely given, licensed, Canadian infant vaccine was administered. No new or unexpected safety issues were identified when DTaP5-IPV-Hib-HepB was given concomitantly with PCV7.
In conclusion, DTaP5-IPV-Hib-HepB administered as primary vaccination was generally safe, well tolerated and immunogenic in infants vaccinated at 2, 4 and 6 months of age. The immunogenicity and safety profiles were similar to those observed with the currently administered vaccine in Canada except for GMTs to PT and tetanus toxoid, which were higher with DTaP5-IPV-Hib-HepB. Rates of seroconversion to FHA did not meet the acceptability criterion but there is likely no clinical impact of this modestly low antibody response given the robust response to the other pertussis antigens. GMTs to FIM, PRP and diphtheria toxoid were higher when the hexavalent vaccine was administered alone. The higher diphtheria toxoid GMT when PCV7 was administered nonconcomitantly likely reflects immune interference of the CRM197 carrier protein of PCV7 to the response to diphtheria toxoid. The acceptability of immunogenicity and safety profiles of the vaccine when coadministered with PCV7 support further development of this combination vaccine.
The authors thank the study investigators and their staff at the participating trial centers for their careful attention to detail and to the children and their families for participating in the study. The authors thank the serology groups of Sanofi Pasteur Inc., Swiftwater, PA and Merck, Research Laboratories, Upper Gwynedd, PA for serology testing. The authors thank Robert Lersch of Sanofi Pasteur Inc. who coordinated the writing of the manuscript; the authors also thank Julia R. Gage of Gage Medical Writing, LLC for providing the first draft of the introduction, methods and results sections and for editing and revising the manuscript per the authors’ instructions.
1. National Center for Immunization and Respiratory Diseases. . General recommendations on immunization: recommendations of the Advisory Committee on Immunization Practices (ACIP). MMWR Recomm Rep. 2011;60:1–64
2. Szilagyi PG, Rodewald LE, Humiston SG, et al. Immunization practices of pediatricians and family physicians in the United States. Pediatrics. 1994;94(4, pt 1):517–523
3. Askew GL, Finelli L, Lutz J, et al. Beliefs and practices regarding childhood vaccination among urban pediatric providers in New Jersey. Pediatrics. 1995;96(5, pt 1):889–892
4. Woodin KA, Rodewald LE, Humiston SG, et al. Physician and parent opinions. Are children becoming pincushions from immunizations? Arch Pediatr Adolesc Med. 1995;149:845–849
5. Zimmerman RK, Bradford BJ, Janosky JE, et al. Barriers to measles and pertussis immunization: the knowledge and attitudes of Pennsylvania primary care physicians. Am J Prev Med. 1997;13:89–97
6. Zimmerman RK, Schlesselman JJ, Mieczkowski TA, et al. Physician concerns about vaccine adverse effects and potential litigation. Arch Pediatr Adolesc Med. 1998;152:12–19
7. Meyerhoff AS, Jacobs RJ. Do too many shots due lead to missed vaccination opportunities? Does it matter? Prev Med. 2005;41:540–544
8. Koslap-Petraco MB, Judelsohn RG. Societal impact of combination vaccines: experiences of physicians, nurses, and parents. J Pediatr Health Care. 2008;22:300–309
9. Koslap-Petraco MB, Parsons T. Communicating the benefits of combination vaccines to parents and health care providers. J Pediatr Health Care. 2003;17:53–57
10. Decker MD, Edwards KM. Haemophilus influenzae type b vaccines: history, choice and comparisons. Pediatr Infect Dis J. 1998;17(suppl 9):S113–S116
11. Happe LE, Lunacsek OE, Marshall GS, et al. Combination vaccine use and vaccination quality in a managed care population. Am J Manag Care. 2007;13:506–512
12. Kalies H, Grote V, Verstraeten T, et al. The use of combination vaccines has improved timeliness of vaccination in children. Pediatr Infect Dis J. 2006;25:507–512
13. Marshall GS, Happe LE, Lunacsek OE, et al. Use of combination vaccines is associated with improved coverage rates. Pediatr Infect Dis J. 2007;26:496–500
14. Reis EC, Roth EK, Syphan JL, et al. Effective pain reduction for multiple immunization injections in young infants. Arch Pediatr Adolesc Med. 2003;157:1115–1120
15. Dodd D. Benefits of combination vaccines: effective vaccination on a simplified schedule. Am J Manag Care. 2003;9(suppl 1):S6–S12
16. Mullany L. Considerations for implementing a new combination vaccine into managed care. Am J Manag Care. 2003;9(suppl 1):S23–S29
17. Advisory Committee on Immunization Practices (ACIP), the American Academy of Pediatrics (AAP), and the American Academy of Family Physicians (AAFP).. Combination vaccines for childhood immunization: recommendations of the Advisory Committee on Immunization Practices (ACIP), the American Academy of Pediatrics (AAP), and the American Academy of Family Physicians (AAFP). Pediatrics. 1999;103:1064–1077
18. Insel RA. Potential alterations in immunogenicity by combining or simultaneously administering vaccine components. Ann N Y Acad Sci. 1995;754:35–47
19. Diaz-Mitoma F, Halperin SA, Tapiero B, et al. Safety and immunogenicity of three different formulations of a liquid hexavalent diphtheria-tetanus-acellular pertussis-inactivated poliovirus-Haemophilus influenzae b conjugate-hepatitis B vaccine at 2, 4, 6 and 12-14 months of age. Vaccine. 2011;29:1324–1331
20. Halperin SA, Tapiero B, Diaz-Mitoma F, et al. Safety and immunogenicity of a hexavalent diphtheria-tetanus-acellular pertussis-inactivated poliovirus-Haemophilus influenzae b conjugate-hepatitis B vaccine at 2, 3, 4, and 12-14 months of age. Vaccine. 2009;27:2540–2547
21. Efstratiou A, Maple PAC Manual for the Laboratory Diagnosis of Diphtheria. 1994 Copenhagen, Denmark Expanded Programme on Immunization, World Health Organization Regional Office for Europe (ICP/EPI/038C)
22. Halperin SA, Langley JM, Hesley TM, et al. Safety and immunogenicity of two formulations of a hexavalent diphtheria-tetanus-acellular pertussis-inactivated poliovirus-Haemophilus influenzae conjugate-hepatitis B vaccine in 15 to 18-month-old children. Hum Vaccin. 2005;1:245–250
23. Eskola J, Ward J, Dagan R, et al. Combined vaccination of Haemophilus influenzae type b conjugate and diphtheria-tetanus-pertussis containing acellular pertussis. Lancet. 1999;354:2063–2068
24. Szmuness W, Stevens CE, Zang EA, et al. A controlled clinical trial of the efficacy of the hepatitis B vaccine (Heptavax B): a final report. Hepatology. 1981;1:377–385
25. Coutinho RA, Lelie N, Albrecht-Van Lent P, et al. Efficacy of a heat inactivated hepatitis B vaccine in male homosexuals: outcome of a placebo controlled double blind trial. Br Med J (Clin Res Ed). 1983;286:1305–1308
26. Ortega-Barrìa E, Kanra G, Leroux G, et al. DTPw-HBV/Hib 2.5 study group. The immunogenicity and reactogenicity of DTPw-HBV/Hib 2.5 combination vaccine: results from four phase III multicenter trials across three continents. Vaccine. 2007;25:8432–8440
27. Greenberg DP, Wong VK, Partridge S, et al. Immunogenicity of a Haemophilus influenzae type b-tetanus toxoid conjugate vaccine when mixed with a diphtheria-tetanus-acellular pertussis-hepatitis B combination vaccine. Pediatr Infect Dis J. 2000;19:1135–1140
28. Poland GA. Acellular pertussis vaccines: new vaccines for an old disease. Lancet. 1996;347:209–210
29. Cherry JD, Gornbein J, Heininger U, et al. A search for serologic correlates of immunity to Bordetella pertussis cough illnesses. Vaccine. 1998;16:1901–1906
30. Storsaeter J, Hallander HO, Gustafsson L, et al. Levels of anti-pertussis antibodies related to protection after household exposure to Bordetella pertussis. Vaccine. 1998;16:1907–1916
31. Olivier C, Belohradsky BH, Stojanov S, et al. Immunogenicity, reactogenicity, and safety of a seven-valent pneumococcal conjugate vaccine (PCV7) concurrently administered with a fully liquid DTPa-IPV-HBV-Hib combination vaccine in healthy infants. Vaccine. 2008;26:3142–3152
32. Tichmann-Schumann I, Soemantri P, Behre U, et al. Immunogenicity and reactogenicity of four doses of diphtheria-tetanus-three-component acellular pertussis-hepatitis B-inactivated polio virus-Haemophilus influenzae type b vaccine coadministered with 7-valent pneumococcal conjugate Vaccine. Pediatr Infect Dis J. 2005;24:70–77
33. Schmitt HJ, Faber J, Lorenz I, et al. The safety, reactogenicity and immunogenicity of a 7-valent pneumococcal conjugate vaccine (7VPnC) concurrently administered with a combination DTaP-IPV-Hib vaccine. Vaccine. 2003;21:3653–3662
34. Knuf M, Habermehl P, Cimino C, et al. Immunogenicity, reactogenicity and safety of a 7-valent pneumococcal conjugate vaccine (PCV7) concurrently administered with a DTPa-HBV-IPV/Hib combination vaccine in healthy infants. Vaccine. 2006;24:4727–4736
35. Black S, Shinefield H, Fireman B, et al. Efficacy, safety and immunogenicity of heptavalent pneumococcal conjugate vaccine in children. Northern California Kaiser Permanente Vaccine Study Center Group. Pediatr Infect Dis J. 2000;19:187–195
36. Dagan R, Poolman J, Siegrist CA. Glycoconjugate vaccines and immune interference: A review. Vaccine. 2010;28:5513–5523
37. Knuf M, Kowalzik F, Kieninger D. Comparative effects of carrier proteins on vaccine-induced immune response. Vaccine. 2011;29:4881–4890