- Email: [email protected]
A combined liquid Hib (PRP-OMP), hepatitis B, diphtheria, tetanus and whole-cell pertussis vaccine: uncontrolled preliminary clinical trial of immunogenicity and reactogenicity
PII: SO264-410X(98)00074-7
Vaccine, Vol. 16, No. 20, pp. 20852089, 1998 0 1998 Elsevier Science Ltd. All rights reserved Printed in Great Britain 0264-410X/98 $19+0.00
ELSEVIER
A combined liquid Hib (PRP-OMP), hepatitis B, diphtheria, tetanus and whole-cell pertussis vaccine: uncontrolled preliminary clinical trial of immunogenicity and reactogenicity Terry Nolan*§, Geoff Hog&, Mary-Ann Darcy*, Maryanne Skeljol and John Carlin” We have conducted a preliminary uncontrolled clinical trial of the immunogenic@ and reactogenicity of a new filly liquid pentavalent combination vaccination which incorporates a diphtheria, tetanus and whole-cell pertussis vaccine with Hib (PRPOMP) and hepatitis B vaccines. Forty-jive infants received three doses of the pentavalent vaccination at 2, 4, and 6 months of age, and then a fourth dose at 18 months of age. Subjects were bled prior to each vaccination, and a month after the third and fourth vaccinations. A 7-day diary card was used to record subject temperatures and other systemic and local clinical signs after each vaccination. After the third dose, 98% of subjects had anti-PRP titres above 1 pg ml-’ (95%ci 88%, 100Y0). Following boosting, the geometric mean titre (GMT) rose a mean 27-fold (95%ci 19-fold, 38-fold) to 33 /g ml-‘, and all suqjects’ titres (lower bound of 95%ci 92%) exceeded 1 pg ml-‘. For hepatitis B antibodji there was a GMT of 100 mIU ml-’ after the third dose, and 86% of infants (95%ci 73%, 9.5%) had antibody levels 210 mIU ml-‘. After the fourth dose, there was a mean 77-fold boost (95%ci 48-fold, 130-fold) to a GMT of 860 mIU ml-’ and 95% (95%ci 84%, 99%) of subjects had titres 210 mIU ml- . Diphtheria, tetanus, and pertussis antibody levels were all at acceptable levels after the first three doses and again after the fourth vaccination. The pentavalent vaccine was well tolerated at all administration times, and had a minor reactogenicity profile similar to DTPw alone 17s reported in previous studies. This study has provided preliminary evidence for both the safety and immunogenic@ of the pentavalent vaccine given as a course at 2, 4, 6 and 18 months. 0 1998 Elsevier Science Ltd. All rights reserved Keywords: DTF’w-Hib-hepatitis
B; vaccine: immunogenicity
Global control of diphtheria, tetanus, pertussis, hepatitis B and Hib disease are realizable objectives with modern vaccine development. Hib vaccines are *Clinical Epidemiology and Biostatistics Unit, Melbourne University Department of Paediatrics, at the Royal Children’s Hospital, Melbourne, Australia, tWomen’s and Children’s Health Care Network Department of Microbiology and Infectious Diseases, at the Royal Children’s Hospital, Melbourne, Australia and :tCSL Ltd, Poplar Rd, Parkville, Victoria, Australia. 9Author to whom correspondence should be addressed. Assoc. Prof. Terrv Nolan. Clinical Epidemiology and Biostatistics Unit, koyal dhildren’s Hospital, Parkville 3052, Australia. Tel: +613-93456368; fax: +613-9345-6000; e-mail: [email protected] (Received 8 October 1997; revised version received 12 February 1998; accepted 18 February 1998)
routinely administered in more than 20 countries, and it is possible that substantially greater use of Hib vaccine will occur in the developing world following confirmation that the burden of invasive Hib is sufficient to justify vaccination ‘-j. Universal vaccination of infants with hepatitis B vaccine is now recommended by the World Health Organization, the US Adviso4y Committee on Immunisation Practices, and others . Whole-cell pertussis vaccines have been used successfully for many years, with worldwide coverage estimated to be 80% 3. The Scientific Advisory Group of Experts (SAGE) of the World Health Organization’s Global Program on Vaccination recently recognized the importance of combination vaccines in the future of immunization, stating that ‘new vaccines should be introduced into national programs in a manner which does not jeopardize current immunisa-
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A combined
liquid Hib (PRP-OMP), hepatitis B, and DTP, vaccine: T. Nolan et al.
tion priorities’ 3. Efforts to streamline delivery, reduce overall cost of vaccination, and to increase parent acceptability and infant tolerance of an increasingly complicated immunisation schedule have focused attention on the development of combination vaccines ‘. However, difficulties with vaccine component interactions have frustrated the development of some combinations ‘. We have conducted a pilot evaluation of a new, fully liquid pentavalent combination vaccine which incorporates DTPw with Hib (PRP-OMP) and hepatitis B vaccines.
MATERIAL AND METHODS Study population The study was conducted between May 1995 and December 1996 in 45 healthy infants (28 male, 17 female) aged 6-12 weeks who were recruited through community maternal and child health centres in Melbourne. Infants were ineligible for study: if they had contracted pertussis, pertussis-like syndrome or Haemophiius infiuenzae type b disease; if they had immune deficiency syndromes; if their mothers were known to be hepatitis B carriers; or if they had been given any blood products (e.g. transfusion), high doses of corticosteroids, cytotoxic agents or radiotherapy.
Vaccine The pentavalent vaccine (adsorbed diphtheria, tetanus, pertussis, H. injluenzae type b and hepatitis B vaccines) contained 30 Lf diphtheria toxoid, 6 Lf tetanus toxoid, not more than 20 x 10” inactivated Bordetella pertussis organisms, 5 /lg hepatitis B surface antigen (recombinant), 7.5 pg H. influenzae PRP-OMP and 225 pg aluminium as aluminium phosphate (423.5 /kg) and aluminium hydroxide (379 ;tg) per dose. Each single 0.5 ml dose was contained in a 2 ml-glass vial with a rubber stopper and aluminium cap, and was stored between 2 and 8°C and protected from light.
Study design The study was an open evaluation, and was approved by the Royal Children’s Hospital Ethics in Human Research Committee. A research assistant obtained written informed consent from the parents of the infants prior to the first blood sample and vaccination. A study doctor visited the infants’ homes and gave a single dose of pentavalent vaccine intramuscularly into the antero-lateral aspect of the thigh at 2, 4 and 6 months of age, and into the deltoid at 18 months. Oral paracetamol (15 mg kg-‘) was routinely given prior to vaccination. Further doses were given only if required at 4 h intervals. Oral sabin vaccine (SmithKline Beecham Biologicals) was administered 7-10 days after the pentavalent vaccine at 2, 4 and 6 months. Local and general reactions were recorded by parents via a structured diary filled in 2-3 h after vaccination, on the evening of vaccination, and thereafter daily until the seventh day after each dose. Temperatures were measured per axilla by Terumo digital thermometers (model C202). The research assistants measured axillary temperature at a home visit 24 h after vaccination. A 5 ml venous blood sample was taken immedia-
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Vaccine 1998 Volume 16 Number 20
tely prior to each vaccination, third and fourth doses.
and 4-6 weeks after the
Laboratory analyses All antibody assays were performed by CSL Ltd except hepatitis B and H. injhenzae type b assays before and after the fourth dose which were performed by Merck Research Laboratories (USA). Antibody to hepatitis B surface antigen (anti-HBs) before and after the first three doses was assayed by enzyme immunoassay (EIA) against a human serum standard calibrated against the WHO first international reference preparation (IU 1 ‘); before and after the fourth dose it was assayed by radioimmunoassay (RIA) against a human serum standard calibrated against the WHO standard (WHO lot 26.1.77)(mIU mll’); H. injhenzae type b capsular polysaccharide polyribosyl-ribitol phosphate (PRP) by RIA against a human serum standard calibrated against the US standard for human anti-Hib PRP serum (,rlg ml-‘) before and after the first three doses, and calibrated against the Centre for Biologics Evaluation antisera and Research (lot# 1983@70000 ng ab mll’) for human anti-Hib PRP serum before and after the fourth dose; tetanus antitoxin by RIA utilizing purified tetanus prototoxin as the capture phase against a human serum standard calibrated against the WHO third international standard for tetanus immunoglobulin (IU ml ‘); essential virulence antigens of Bordetefla pertussis - pertussigen (Ptx), filamentous haemagglutonin (Fha), agglutinogen Type 2 (Agg 2) agglutinogen Type 3 (Agg 3) and pertactin (69kD) by enzyme immunoassay (EIA) utilising monoclonal antibody captured antigen as the capture phase calibrated against the US reference pertussis antisera (human) lots 3 and 4 (u mll’). Assays for diphtheria antitoxin were by cell culture assay in Vero cells at the LmilOOOO level. The human diphtheria standard was calibrated against the WHO first international standard for diphtheria antitoxin. Statistical
analysis
Antibody responses were summarized both as a proportion of subjects reaching recognized protective levels of antibody titre, with standard exact binomial confidence intervals, and as geometric mean titres with 95% confidence intervals, obtained in the standard way by applying normal-theory methods to log-transformed data. Geometric mean fold changes (ratios) comparing pre- and post-vaccination titres were also calculated, with confidence intervals. Antibody titre thresholds used for calculating protected proportions were: antitetanus and anti-diphtheria, 0.01 IU rn;-‘; anti-HBs, 10 mIU ml-‘; and anti-PRP, t big ml-‘. The frequency of adverse events was tabulated with 95% confidence intervals obtained by the binomial method. Fever was defined as axillary temperature 238°C. and excessive fussiness as occurring in babies who were crying and could not be comforted at times or at all ‘.
RESULTS A high proportion of infants had pre-existing titres to tetanus, but immunisation produced a mean 12-fold increase (95%ci 7.4-fold, 20-fold) in titre (Table 1). The first three doses of vaccine produced a 100% serocon-
A combined liquid Hib (PRP-OMP), hepatitis 6, and DTP, vaccine: T. Nolan et al. version to protective level or greater for diphtheria. The pre-fourth dose antibody levels were substantially boosted following vaccination for both antigens. Postvaccination geometric mean titres (GMTs) were very high, and all subjects had antibody levels well in excess of protective thresholds. Anti-HBs antibodies were detected in 12 of the 45 subjects prior to immunisation. This presumably reflected maternal exposure to vaccine (11 mothers had completed a full course of hepatitis B vaccination; another mother had received two doses). The anti-HBs GMT rose sharply after the second vaccination (Table I). Following the I;hird dose, there was a greater response with a mean 14-fold increase in titre. After the three doses, 86% of mfants (95%ci 73%, 95%) had antibody levels at or beyond the protective level of 10 mIU ml-‘. At 18 months of age, 95% of subjects (95%ci 84%, 99%) had titres above 10 mIU ml-’ following the fourth dose. Following the first dose of vaccine, there was a mean fourfold increase in anti-PRP titre, rising smoothly to a mean 16-fold increase after three doses. At this time, 98% of subjects had titres above the protective level of 1 pg m,lK’ (95%ci 88%, 100%). At 18 months, the Hib GMT had fallen so that 57% of evaluatable subjects had titres above 1 ,ug ml-’ (95%ci 41%, 72%). Following boosting, the titres of all subjects exceeded the I. pg ml-’ protective threshold (lower bound of 95%ci 92%). Pertussis antibody titres are shown in Table 2. PostTable 1
Antibody
titres to tetanus, diphtheria,
fourth dose boosting resulted in fold changes in titre of between 14 (Ptx) and 24 (Fha). These fold changes correspond to on average between 2.5 and 10 times higher GMTs than those observed immediately following the third dose. Of the 42 subjects available for evaluation following the third dose, 88.1% (95%ci 74%, 96%) had antibody titres above protective thresholds for all four antigens for which such thresholds are available, namely tetanus, diphtheria, Hib and hepatitis B. The other two subjects (who had inadequate blood for diphtheria and tetanus antibody estimation) had anti-HBs titres below 10 mIU ml-‘. Following the fourth dose, the proportion with antibody titres above protective thresholds for all four antigens was 95.3% (95%ci 84%, 99%). No serious adverse event was recorded after any vaccination with the pentavalent vaccine. There were no seizures, hypotonic hyporesponsive episodes, encephalopathic or anaphylactic episodes. Parents or research assistants recorded no other unusual or unexpected event. Minor systemic and local clinical signs are detailed in Table 3. Episodes of temperature 238°C were most common after the second (33.3%, 95%ci 20%, 49%) and fourth doses (27.3%, 95%ci 15%, 43%). Excessive fussiness occurred in 15.6% of infants after each of the second and third doses. There was a high level of reported tenderness (84.1%) after the 18 month dose, consistent with higher levels of visible local inflammation greater than 25 mm (29.5%).
HBs, and PRP n
% > threshold
(IU ml-‘) An;$tanus Post-first Post-second Post-third Ratio (post-third/pre-first) Pre-fourth Post-fourth Ratio (post-fourth/pre-fourth)
41 44 42 42 38 41 43 40
98 100 100 100 _ 100 100 _
Anti-diphtheria (/U ml- ‘) Pre Post-first Post-second Post-third Ratio (post-third/pre-first) Pre-fourth Post-fourth Ratio (post-fourth/pre-fourth)
37 41 39 42 34 42 43 41
59 37 90 100 _ 95 100 _
Anti-HBs (m/U ml-‘) Pre Post-first Post-second Post-third Ratio (post-third/pre-first) Pre-fourth Post-fourth Ratio (post-fourth/pre-fourth)
45 45 44 44 44 42 43 44
27 24 70 86 _ 62 95 _
(15, (13, (55, (73,
Anti-PRP (pg ml-‘) Pre Post-first Post-second Post-third Ratio (post-third/pre-first) Pre-fourth Post-fourth Ratio (post-fourth/pre-fourth)
43 44 43 43 41 42 43 41
37 81 93 98 _ 57 100 _
(23, 53) (67, 92) (81, 99) (88, 100)
GM
(95% Cl)
(87, (92, (92, (92,
100) 100) 100) 100)
(91, 100) (92, 100) _
(42, (22, (76, (92,
75) 53) 97) 100)
(84, 99) (92, 100) _ 42) 40) 83) 95)
(46, 76) (84, 99)
(41, 72) (92, 100) _
Vaccine
(95% Cl)
0.57 0.21 0.89 7.2 12 0.61 21 35
(0.37, 0.88) (0.15, 0.29) (0.58, 1.3) (5.5, 9.5) (7.4, 20) (0.45, 0.83) (16, 27) (27, 47)
0.03 0.02 0.10 0.83 18 0.17 11 67
(0.02, 0.06) (0.01, 0.02) (0.07, 0.14) (0.57, 1.2) (7.8, 44) (0.11, 0.25) (8.6, 15) (50,91)
7.2 5.3 21 100 14 11 860 77 0.54 2.1 4.3 9.2 16 1.2 33 27
(4.1,13) (3.6,7.9) U4,31)
(56, 180) (5.5,35) (5.6, 20) (450, 1600) (48, 130) (0.38, 0.78) (1.5, 2.9) (3.1, 6.1) (6.4, 13) (10, 26) (0.85, 1.7) (23, 47) (19, 38)
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A combined
liquid Mb (PRP-O/UP), hepatitis B, and DTP, vaccine: T. Nolan et al. Table 3 Systemic and local clinical signs experienced during the 24 h period following vaccination
DISCUSSION A combination DTPw-Hib vaccine has been available for some years ‘, and although there have been recent reports of a quadrivalent DTPw-hepatitis B vaccine I”, a vaccine which combines DTPw, Hib and hepatitis B vaccines has not yet been licensed. This pilot study has provided preliminary evidence for both the safety and immunogenicity of a new fully liquid pentavalent vaccine given as a four-dose course at 2, 4, 6 and 18 months. Because of the small sample size, there were wide confidence intervals on both adverse effect estimates and antibody responses, and a larger study is now under way. No serious adverse event was recorded after any vaccination with the pentavalent vaccine. There were no seizures, hypotonic hyporesponsive episodes, encephalopathic or anaphylactic episodes. The pentavalent vaccine was well tolerated by all subjects, both as infants and at 18 months of age. The relatively high level of reported tenderness after the 18 month dose was consistent both with higher levels of visible local
Table 2 Antibodies
Pertussis antibody to:
responses n
GM
(95% Cl)
Ptx (u ml-‘) Pre Post-first Post-second Post-third Ratio (post-third/pre-first) Pre-fourth Post-fourth Ratio (post-fourthipre-fourth)
41 44 42 42 38 41 43 40
Fha (u ml-‘) Pre Post-first Post-second Post-third Ratio (post-third/pre-first) Pre-fourth Post-fourth Ratio (post-fourth/pre-fourth)
41 44 42 42 38 41 43 40
2.9 1.8 5.7 14 5.3 4.3 110 24
(2.0,4.2) (1.4,2.3) (4.1,8.0) (9.7,19) (3.0,9.5) (3.1, 6.1) (88, 130) (17, 33)
Agg2 (u ml-‘) Pre Post-first Post-second Post-third Ratio (post-third/pre-first) Pre-fourth Post-fourth Ratio (post-fourth/pre-fourth)
41 44 42 42 38 41 43 40
6.4 3.3 5.8 26 3.5 12 180 15
(4.6,8.8) (2.7,4.1) (4.0,8.4) (16, 42) (1.9,6.6) (8.9, 17) (110, 300) (11, 22)
Agg3 (u ml ‘) Pre Post-first Post-second Post-third Ratio most-third/ore-first) Pre-fourth Post-fourth Ratio (post-fourthipre-fourth)
41 44 42 42 38 41 43 40
2.7 1.7 8.4 56 19 11 160 16
(2.1,3.6) (1.4,2.0) (4.8,15) (35, 89) (10, 36) (7.6,16) (110, 220) (11,211
Pert (u ml-‘) Pre Post-first Post-second Post-third Ratio (post-third/pre-first) Pre-fourth Post-fourth Ratio (post-fourth/pre-fourth)
41 44 42 42 38 41 43 40
2088
1.4 0.68 1.2 3.9 3.1 2.8 38 14
1 0.59 2 16 15 2.4 41 19
Vaccine 1998 Volume 16 Number 20
(1.1 ,1.9) (0.54,0.87) (0.75,2.0) (2.1,7.4) (1.5,6.8) (2.1, 3.8) (23, 64) (9.4, 22)
(0.71 ,1.5) (0.47,0.75) (1.3,3.1) (9.5,25) (8.2,26) (1.7, 3.3) (27, 64) (14, 25)
at any time
%
n
95% ci
First vaccination Fever 238°C Excessive fussiness Drowsiness z 25 mm redness > 25 mm swelling Tenderness
15.6 6.7 60.0 11 .l 11.1 46.7
45 45 45 45 45 45
(6.5, 29.5) (1.4, 18.3) (44.3, 74.3) (3.7, 24.1) (3.7, 24.1) (31.7, 62.1)
Second vaccination Fever 2 38°C Excessive fussiness Drowsiness > 25 mm redness > 25 mm swelling Tenderness
33.3 15.6 46.7 2.2 6.7 71.1
45 45 45 45 45 45
(20.0, 49.0) (6.5, 29.5) (31.7, 62.1) (0.1, 11.8) (1.4, 18.3) (55.7, 83.6)
Third vaccination Fever ~38°C Excessive fussiness Drowsiness > 25 mm redness z 25 mm swelling Tenderness
13.3 15.6 40.0 4.4 4.4 46.7
45 45 45 45 45 45
(5.1, 26.8) (6.5, 29.5) (25.7, 55.7) (0.5, 15.1) (0.5, 15.1) (31.7, 62.1)
18 month fourth vaccination Fever 238°C Excessive fussiness Drowsiness > 25 mm redness > 25 mm swelling Tenderness
27.3 6.8 25.0 29.5 25.0 84.1
44 44 44 44 44 44
(15.0, 42.8) (1.4, 18.7) (13.2, 40.3) (16.8, 45.2) (13.2, 40.3) (69.9, 93.4)
inflammation, and also with an older child being able to respond more specifically. The adverse effect profile was generally similar to that recorded in our previous studies with the currently licensed Australian DTP vaccine ” which is a component of the pentavalent vaccine. Although an acellular pertussis component may have produced a lower incidence of transient side effects, recent data demonstrate that whole-cell and acellular pertussis vaccines are similar in terms of rates of serious adverse events I’. Additionally, acellular vaccines do not offer any improvement in efficacy over otent European-type formulation whole-cell vaccines L . Gtven that acellular vaccines are more costly, the pedtavalent vaccine was formulated to contain a wholecell pertussis component with the aim of developing an efficacious and cost-effective vaccine which would satisfy many of the goals of the Children’s Vaccine Initiative for a global paediatric vaccine ‘. Following the first three doses, the seroresponse to the pentavalent vaccine was excellent for Hib antibody, and also good for hepatitis B although not as high as commonly seen with monovalent hepatitis B vaccine 14. This is presumably the result of component interference, though the exact mechanism is unknown. Higher levels of hepatitis B seroconversion at 7 months are desirable, however, and are likely for infants who receive monovalent hepatitis B vaccine shortly after birth 15.This is currently under evaluation. The pertussis immunogenicity profile was similar to that previously documented for the currently licensed Australian DTP vaccine ‘I. Both diphtheria and tetanus responses were well within acceptable margins for protection. The dose at 18 months produced a substantial rise in antibody titre for all antigens. As more vaccines are added to childhood immunisa-
A combined
liquid Hib (PRP-OMP), hepatitis El, and DTP, vaccine:
tion schedules, there is a need to keep those schedules simple by combining as many as possible to reduce the number of encounters, injections, and packaging and delivery costs. A reduction from eleven (DTP, 2, 4, 6, 18 months; Hib, 2, 4, 6, 18 months; hep B, 0, 1, 7 months) to four separate injections (pentavalent, 2, 4, 6, 18 months) may be possible with a pentavalent vaccine such as the one evaluated in this study. A full controlled pre-licensure clinical evaluation of the pentavalent vaccine (in approximately 2000 infants) is now nearing completion.
4
5
6 7
ACKNOWLEDGEMENTS 8
We would like to acknowledge research assistants Noreen Condon, Margaret Flood, Elizabeth McGrath, Susie Proctor; phlebotomists Patricia Barbante, Dr Paul Carmen, Kathleen Lanigan, Charan Sandhu, and Deborah Saunders; and study doctors Len Hartman, Audrey Penington and Joanne Smart. Suzanna Vidmar assisted with the data analysis. Valuable assistance in study planning was provided by John Varigos, Jillian Bennet, and Paul Fahey. Professor Ian Gust provided helpful comments on the manuscript. Serology was carried out by the CSL Ltd Quality Control Biochemistry Department under the supervision of Lena Miloradovic. Bill ‘Woods, Serge DeBartolo and the Clinical Trials Logistics Group at CSL Ltd provided assistance to field staff and ensured that the vaccine cold chain was maintained. We thank the parents of subjects, the maternal and child health nurses in the participating areas, and staff from the health departments of the municipalities of Monash and Whitehorse. This study was supported by a grant to the Royal Children’s :Hospital Research Foundation from CSL Ltd, and by Merck and Co., Inc.
REFERENCES Mulholland, K., Hilton, S. and Adegbola, R. et al. Randomised trial of Haemophilus influenzae type-b tetanus protein conjugate for prevention of pneumonia and meningitis in Gambian infants. Lancer 1997, 349, 1191-l 197. Steinhoff, MC. Haemophilus influenzae type-b infections are preventable everywhere. /_ancet 1997, 349, 1186-l 187. Report of the Meeting of the Scientific Advisory Group of Experts (SAGE) of the Children’s Vaccine Initiative and the Global Program on Vaccination, Geneva, June 1997, WHO
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13
14
15
T. Nolan et al.
Report No. WHO/GPV/97.05. Hepatitis B virus: a comprehensive strategy for eliminating transmission in the United States through universal childhood immunisation. Recommendations of the Immunization Practices Advisory Committee. Morb. Mortal. Wkly Report 1991, 40RR-13, l-25. The Australian lmmunisation Handbook, 6th edn. National Health and Medical Research Council, Australian Government Publishing Service, Canberra, ACT 2601, 1997. Schleiss, M.R. Ever-changing world of pediatric vaccines: why are we giving more shots? J. Pediatr. 1997, 130, 505-507. Schmitt, H.J. lmmunogenicity and reactogenicity of 2 Hib tetanus conjugate vaccines administered by reconstituting with DTPa or given as separate injections [abstract]. In: Proceedings of the 35th ICAAC (G63), September 1995. American Society for Microbiology, Washington D.C. Pichichero, M.E., Christy, C., Decker, M.D., Steinhoff, MC., Edwards, K.M., Rennels, M.B., Anderson, E.L. and Englund, J.A. Defining the key parameters for comparing reactions among acellular and whole-cell pertussis vaccines. Pediatrics 1995,96,588-592. Paradiso, P.R., Hogerman, D.A., Madore, D.V., Keyserling, H., King, J., Reisinger, KS., Blatter, M.M., Rothstein, E., Bernstein, H.H. and Hackell, J. Safety and immunogenicity of a combined diphtheria, tetanus, pertussis and Haemophilus influenzae type b vaccine in young infants. Pediatrics 1993, 92, 1-6. Aristegui, J., Garrote, E., Gonzalez, A., Arrate, J.-P., Perez, A. and Vandepapeliere, P. Immune response to a combined hepatitis B, diphtheria, tetanus and whole cell pertussis vaccine administered to infants at 2. 4 and 6 months of age. Vaccine 1997, 15, 7-9. Nolan, T.M., Hogg, G., Darcy, M.-A., Varigos, J. and McEwen, J. Primary course immunogenicity and reactogenicity of a new DTP, (diphtheria-tetanus-whole cell pertussis) vaccine. Journal of Paediatrics and Child Health 1997, 33, 413-417. Olin, P., Rasmussen, F., Gustafson, L., Hallander, H.O. and Heijbel, H. Randomised controlled trial of two-component, three component, and five-component acellular pertussis vaccines compared with whole-cell pertussis vaccine. Lancet 1997,350,1569-l 577. Plotkin, S.A. and Cadoz, M. The acellular pertussis vaccine trials: an interpretation. Pediatr infect Dis J 1997, 16, 508-517. Keyserling, H.L., West, D.J., Hesley, T.M., Bosley, C., Wiens, B.L. and Calandra, G.B. Antibody responses of healthy infants to a recombinant hepatitis B vaccine administered at 2, 4, and 12 or 15 months of age. J. Pediatr 1994, 125, 67-69. Aristegui, J., Muniz, J., Perez-Legorburu, A., Imaz, M., Arrate, J.P., Suarez, M.D. and Goiri, M.D. Newborn universal immunisation against hepatitis B: immunogenicity and reactogenicity of simultaneous administration of diphtheria/tetanus/ pertussis (DTP] and oral polio vaccines with hepatitis B vaccine at 0, 2 and 6 months of age. Vaccine 1995, 13, 973-977.
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1998 Volume 16 Number 20
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Vaccine, Vol. 16, No. 20, pp. 20852089, 1998 0 1998 Elsevier Science Ltd. All rights reserved Printed in Great Britain 0264-410X/98 $19+0.00
ELSEVIER
A combined liquid Hib (PRP-OMP), hepatitis B, diphtheria, tetanus and whole-cell pertussis vaccine: uncontrolled preliminary clinical trial of immunogenicity and reactogenicity Terry Nolan*§, Geoff Hog&, Mary-Ann Darcy*, Maryanne Skeljol and John Carlin” We have conducted a preliminary uncontrolled clinical trial of the immunogenic@ and reactogenicity of a new filly liquid pentavalent combination vaccination which incorporates a diphtheria, tetanus and whole-cell pertussis vaccine with Hib (PRPOMP) and hepatitis B vaccines. Forty-jive infants received three doses of the pentavalent vaccination at 2, 4, and 6 months of age, and then a fourth dose at 18 months of age. Subjects were bled prior to each vaccination, and a month after the third and fourth vaccinations. A 7-day diary card was used to record subject temperatures and other systemic and local clinical signs after each vaccination. After the third dose, 98% of subjects had anti-PRP titres above 1 pg ml-’ (95%ci 88%, 100Y0). Following boosting, the geometric mean titre (GMT) rose a mean 27-fold (95%ci 19-fold, 38-fold) to 33 /g ml-‘, and all suqjects’ titres (lower bound of 95%ci 92%) exceeded 1 pg ml-‘. For hepatitis B antibodji there was a GMT of 100 mIU ml-’ after the third dose, and 86% of infants (95%ci 73%, 9.5%) had antibody levels 210 mIU ml-‘. After the fourth dose, there was a mean 77-fold boost (95%ci 48-fold, 130-fold) to a GMT of 860 mIU ml-’ and 95% (95%ci 84%, 99%) of subjects had titres 210 mIU ml- . Diphtheria, tetanus, and pertussis antibody levels were all at acceptable levels after the first three doses and again after the fourth vaccination. The pentavalent vaccine was well tolerated at all administration times, and had a minor reactogenicity profile similar to DTPw alone 17s reported in previous studies. This study has provided preliminary evidence for both the safety and immunogenic@ of the pentavalent vaccine given as a course at 2, 4, 6 and 18 months. 0 1998 Elsevier Science Ltd. All rights reserved Keywords: DTF’w-Hib-hepatitis
B; vaccine: immunogenicity
Global control of diphtheria, tetanus, pertussis, hepatitis B and Hib disease are realizable objectives with modern vaccine development. Hib vaccines are *Clinical Epidemiology and Biostatistics Unit, Melbourne University Department of Paediatrics, at the Royal Children’s Hospital, Melbourne, Australia, tWomen’s and Children’s Health Care Network Department of Microbiology and Infectious Diseases, at the Royal Children’s Hospital, Melbourne, Australia and :tCSL Ltd, Poplar Rd, Parkville, Victoria, Australia. 9Author to whom correspondence should be addressed. Assoc. Prof. Terrv Nolan. Clinical Epidemiology and Biostatistics Unit, koyal dhildren’s Hospital, Parkville 3052, Australia. Tel: +613-93456368; fax: +613-9345-6000; e-mail: [email protected] (Received 8 October 1997; revised version received 12 February 1998; accepted 18 February 1998)
routinely administered in more than 20 countries, and it is possible that substantially greater use of Hib vaccine will occur in the developing world following confirmation that the burden of invasive Hib is sufficient to justify vaccination ‘-j. Universal vaccination of infants with hepatitis B vaccine is now recommended by the World Health Organization, the US Adviso4y Committee on Immunisation Practices, and others . Whole-cell pertussis vaccines have been used successfully for many years, with worldwide coverage estimated to be 80% 3. The Scientific Advisory Group of Experts (SAGE) of the World Health Organization’s Global Program on Vaccination recently recognized the importance of combination vaccines in the future of immunization, stating that ‘new vaccines should be introduced into national programs in a manner which does not jeopardize current immunisa-
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A combined
liquid Hib (PRP-OMP), hepatitis B, and DTP, vaccine: T. Nolan et al.
tion priorities’ 3. Efforts to streamline delivery, reduce overall cost of vaccination, and to increase parent acceptability and infant tolerance of an increasingly complicated immunisation schedule have focused attention on the development of combination vaccines ‘. However, difficulties with vaccine component interactions have frustrated the development of some combinations ‘. We have conducted a pilot evaluation of a new, fully liquid pentavalent combination vaccine which incorporates DTPw with Hib (PRP-OMP) and hepatitis B vaccines.
MATERIAL AND METHODS Study population The study was conducted between May 1995 and December 1996 in 45 healthy infants (28 male, 17 female) aged 6-12 weeks who were recruited through community maternal and child health centres in Melbourne. Infants were ineligible for study: if they had contracted pertussis, pertussis-like syndrome or Haemophiius infiuenzae type b disease; if they had immune deficiency syndromes; if their mothers were known to be hepatitis B carriers; or if they had been given any blood products (e.g. transfusion), high doses of corticosteroids, cytotoxic agents or radiotherapy.
Vaccine The pentavalent vaccine (adsorbed diphtheria, tetanus, pertussis, H. injluenzae type b and hepatitis B vaccines) contained 30 Lf diphtheria toxoid, 6 Lf tetanus toxoid, not more than 20 x 10” inactivated Bordetella pertussis organisms, 5 /lg hepatitis B surface antigen (recombinant), 7.5 pg H. influenzae PRP-OMP and 225 pg aluminium as aluminium phosphate (423.5 /kg) and aluminium hydroxide (379 ;tg) per dose. Each single 0.5 ml dose was contained in a 2 ml-glass vial with a rubber stopper and aluminium cap, and was stored between 2 and 8°C and protected from light.
Study design The study was an open evaluation, and was approved by the Royal Children’s Hospital Ethics in Human Research Committee. A research assistant obtained written informed consent from the parents of the infants prior to the first blood sample and vaccination. A study doctor visited the infants’ homes and gave a single dose of pentavalent vaccine intramuscularly into the antero-lateral aspect of the thigh at 2, 4 and 6 months of age, and into the deltoid at 18 months. Oral paracetamol (15 mg kg-‘) was routinely given prior to vaccination. Further doses were given only if required at 4 h intervals. Oral sabin vaccine (SmithKline Beecham Biologicals) was administered 7-10 days after the pentavalent vaccine at 2, 4 and 6 months. Local and general reactions were recorded by parents via a structured diary filled in 2-3 h after vaccination, on the evening of vaccination, and thereafter daily until the seventh day after each dose. Temperatures were measured per axilla by Terumo digital thermometers (model C202). The research assistants measured axillary temperature at a home visit 24 h after vaccination. A 5 ml venous blood sample was taken immedia-
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tely prior to each vaccination, third and fourth doses.
and 4-6 weeks after the
Laboratory analyses All antibody assays were performed by CSL Ltd except hepatitis B and H. injhenzae type b assays before and after the fourth dose which were performed by Merck Research Laboratories (USA). Antibody to hepatitis B surface antigen (anti-HBs) before and after the first three doses was assayed by enzyme immunoassay (EIA) against a human serum standard calibrated against the WHO first international reference preparation (IU 1 ‘); before and after the fourth dose it was assayed by radioimmunoassay (RIA) against a human serum standard calibrated against the WHO standard (WHO lot 26.1.77)(mIU mll’); H. injhenzae type b capsular polysaccharide polyribosyl-ribitol phosphate (PRP) by RIA against a human serum standard calibrated against the US standard for human anti-Hib PRP serum (,rlg ml-‘) before and after the first three doses, and calibrated against the Centre for Biologics Evaluation antisera and Research (lot# 1983@70000 ng ab mll’) for human anti-Hib PRP serum before and after the fourth dose; tetanus antitoxin by RIA utilizing purified tetanus prototoxin as the capture phase against a human serum standard calibrated against the WHO third international standard for tetanus immunoglobulin (IU ml ‘); essential virulence antigens of Bordetefla pertussis - pertussigen (Ptx), filamentous haemagglutonin (Fha), agglutinogen Type 2 (Agg 2) agglutinogen Type 3 (Agg 3) and pertactin (69kD) by enzyme immunoassay (EIA) utilising monoclonal antibody captured antigen as the capture phase calibrated against the US reference pertussis antisera (human) lots 3 and 4 (u mll’). Assays for diphtheria antitoxin were by cell culture assay in Vero cells at the LmilOOOO level. The human diphtheria standard was calibrated against the WHO first international standard for diphtheria antitoxin. Statistical
analysis
Antibody responses were summarized both as a proportion of subjects reaching recognized protective levels of antibody titre, with standard exact binomial confidence intervals, and as geometric mean titres with 95% confidence intervals, obtained in the standard way by applying normal-theory methods to log-transformed data. Geometric mean fold changes (ratios) comparing pre- and post-vaccination titres were also calculated, with confidence intervals. Antibody titre thresholds used for calculating protected proportions were: antitetanus and anti-diphtheria, 0.01 IU rn;-‘; anti-HBs, 10 mIU ml-‘; and anti-PRP, t big ml-‘. The frequency of adverse events was tabulated with 95% confidence intervals obtained by the binomial method. Fever was defined as axillary temperature 238°C. and excessive fussiness as occurring in babies who were crying and could not be comforted at times or at all ‘.
RESULTS A high proportion of infants had pre-existing titres to tetanus, but immunisation produced a mean 12-fold increase (95%ci 7.4-fold, 20-fold) in titre (Table 1). The first three doses of vaccine produced a 100% serocon-
A combined liquid Hib (PRP-OMP), hepatitis 6, and DTP, vaccine: T. Nolan et al. version to protective level or greater for diphtheria. The pre-fourth dose antibody levels were substantially boosted following vaccination for both antigens. Postvaccination geometric mean titres (GMTs) were very high, and all subjects had antibody levels well in excess of protective thresholds. Anti-HBs antibodies were detected in 12 of the 45 subjects prior to immunisation. This presumably reflected maternal exposure to vaccine (11 mothers had completed a full course of hepatitis B vaccination; another mother had received two doses). The anti-HBs GMT rose sharply after the second vaccination (Table I). Following the I;hird dose, there was a greater response with a mean 14-fold increase in titre. After the three doses, 86% of mfants (95%ci 73%, 95%) had antibody levels at or beyond the protective level of 10 mIU ml-‘. At 18 months of age, 95% of subjects (95%ci 84%, 99%) had titres above 10 mIU ml-’ following the fourth dose. Following the first dose of vaccine, there was a mean fourfold increase in anti-PRP titre, rising smoothly to a mean 16-fold increase after three doses. At this time, 98% of subjects had titres above the protective level of 1 pg m,lK’ (95%ci 88%, 100%). At 18 months, the Hib GMT had fallen so that 57% of evaluatable subjects had titres above 1 ,ug ml-’ (95%ci 41%, 72%). Following boosting, the titres of all subjects exceeded the I. pg ml-’ protective threshold (lower bound of 95%ci 92%). Pertussis antibody titres are shown in Table 2. PostTable 1
Antibody
titres to tetanus, diphtheria,
fourth dose boosting resulted in fold changes in titre of between 14 (Ptx) and 24 (Fha). These fold changes correspond to on average between 2.5 and 10 times higher GMTs than those observed immediately following the third dose. Of the 42 subjects available for evaluation following the third dose, 88.1% (95%ci 74%, 96%) had antibody titres above protective thresholds for all four antigens for which such thresholds are available, namely tetanus, diphtheria, Hib and hepatitis B. The other two subjects (who had inadequate blood for diphtheria and tetanus antibody estimation) had anti-HBs titres below 10 mIU ml-‘. Following the fourth dose, the proportion with antibody titres above protective thresholds for all four antigens was 95.3% (95%ci 84%, 99%). No serious adverse event was recorded after any vaccination with the pentavalent vaccine. There were no seizures, hypotonic hyporesponsive episodes, encephalopathic or anaphylactic episodes. Parents or research assistants recorded no other unusual or unexpected event. Minor systemic and local clinical signs are detailed in Table 3. Episodes of temperature 238°C were most common after the second (33.3%, 95%ci 20%, 49%) and fourth doses (27.3%, 95%ci 15%, 43%). Excessive fussiness occurred in 15.6% of infants after each of the second and third doses. There was a high level of reported tenderness (84.1%) after the 18 month dose, consistent with higher levels of visible local inflammation greater than 25 mm (29.5%).
HBs, and PRP n
% > threshold
(IU ml-‘) An;$tanus Post-first Post-second Post-third Ratio (post-third/pre-first) Pre-fourth Post-fourth Ratio (post-fourth/pre-fourth)
41 44 42 42 38 41 43 40
98 100 100 100 _ 100 100 _
Anti-diphtheria (/U ml- ‘) Pre Post-first Post-second Post-third Ratio (post-third/pre-first) Pre-fourth Post-fourth Ratio (post-fourth/pre-fourth)
37 41 39 42 34 42 43 41
59 37 90 100 _ 95 100 _
Anti-HBs (m/U ml-‘) Pre Post-first Post-second Post-third Ratio (post-third/pre-first) Pre-fourth Post-fourth Ratio (post-fourth/pre-fourth)
45 45 44 44 44 42 43 44
27 24 70 86 _ 62 95 _
(15, (13, (55, (73,
Anti-PRP (pg ml-‘) Pre Post-first Post-second Post-third Ratio (post-third/pre-first) Pre-fourth Post-fourth Ratio (post-fourth/pre-fourth)
43 44 43 43 41 42 43 41
37 81 93 98 _ 57 100 _
(23, 53) (67, 92) (81, 99) (88, 100)
GM
(95% Cl)
(87, (92, (92, (92,
100) 100) 100) 100)
(91, 100) (92, 100) _
(42, (22, (76, (92,
75) 53) 97) 100)
(84, 99) (92, 100) _ 42) 40) 83) 95)
(46, 76) (84, 99)
(41, 72) (92, 100) _
Vaccine
(95% Cl)
0.57 0.21 0.89 7.2 12 0.61 21 35
(0.37, 0.88) (0.15, 0.29) (0.58, 1.3) (5.5, 9.5) (7.4, 20) (0.45, 0.83) (16, 27) (27, 47)
0.03 0.02 0.10 0.83 18 0.17 11 67
(0.02, 0.06) (0.01, 0.02) (0.07, 0.14) (0.57, 1.2) (7.8, 44) (0.11, 0.25) (8.6, 15) (50,91)
7.2 5.3 21 100 14 11 860 77 0.54 2.1 4.3 9.2 16 1.2 33 27
(4.1,13) (3.6,7.9) U4,31)
(56, 180) (5.5,35) (5.6, 20) (450, 1600) (48, 130) (0.38, 0.78) (1.5, 2.9) (3.1, 6.1) (6.4, 13) (10, 26) (0.85, 1.7) (23, 47) (19, 38)
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liquid Mb (PRP-O/UP), hepatitis B, and DTP, vaccine: T. Nolan et al. Table 3 Systemic and local clinical signs experienced during the 24 h period following vaccination
DISCUSSION A combination DTPw-Hib vaccine has been available for some years ‘, and although there have been recent reports of a quadrivalent DTPw-hepatitis B vaccine I”, a vaccine which combines DTPw, Hib and hepatitis B vaccines has not yet been licensed. This pilot study has provided preliminary evidence for both the safety and immunogenicity of a new fully liquid pentavalent vaccine given as a four-dose course at 2, 4, 6 and 18 months. Because of the small sample size, there were wide confidence intervals on both adverse effect estimates and antibody responses, and a larger study is now under way. No serious adverse event was recorded after any vaccination with the pentavalent vaccine. There were no seizures, hypotonic hyporesponsive episodes, encephalopathic or anaphylactic episodes. The pentavalent vaccine was well tolerated by all subjects, both as infants and at 18 months of age. The relatively high level of reported tenderness after the 18 month dose was consistent both with higher levels of visible local
Table 2 Antibodies
Pertussis antibody to:
responses n
GM
(95% Cl)
Ptx (u ml-‘) Pre Post-first Post-second Post-third Ratio (post-third/pre-first) Pre-fourth Post-fourth Ratio (post-fourthipre-fourth)
41 44 42 42 38 41 43 40
Fha (u ml-‘) Pre Post-first Post-second Post-third Ratio (post-third/pre-first) Pre-fourth Post-fourth Ratio (post-fourth/pre-fourth)
41 44 42 42 38 41 43 40
2.9 1.8 5.7 14 5.3 4.3 110 24
(2.0,4.2) (1.4,2.3) (4.1,8.0) (9.7,19) (3.0,9.5) (3.1, 6.1) (88, 130) (17, 33)
Agg2 (u ml-‘) Pre Post-first Post-second Post-third Ratio (post-third/pre-first) Pre-fourth Post-fourth Ratio (post-fourth/pre-fourth)
41 44 42 42 38 41 43 40
6.4 3.3 5.8 26 3.5 12 180 15
(4.6,8.8) (2.7,4.1) (4.0,8.4) (16, 42) (1.9,6.6) (8.9, 17) (110, 300) (11, 22)
Agg3 (u ml ‘) Pre Post-first Post-second Post-third Ratio most-third/ore-first) Pre-fourth Post-fourth Ratio (post-fourthipre-fourth)
41 44 42 42 38 41 43 40
2.7 1.7 8.4 56 19 11 160 16
(2.1,3.6) (1.4,2.0) (4.8,15) (35, 89) (10, 36) (7.6,16) (110, 220) (11,211
Pert (u ml-‘) Pre Post-first Post-second Post-third Ratio (post-third/pre-first) Pre-fourth Post-fourth Ratio (post-fourth/pre-fourth)
41 44 42 42 38 41 43 40
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1.4 0.68 1.2 3.9 3.1 2.8 38 14
1 0.59 2 16 15 2.4 41 19
Vaccine 1998 Volume 16 Number 20
(1.1 ,1.9) (0.54,0.87) (0.75,2.0) (2.1,7.4) (1.5,6.8) (2.1, 3.8) (23, 64) (9.4, 22)
(0.71 ,1.5) (0.47,0.75) (1.3,3.1) (9.5,25) (8.2,26) (1.7, 3.3) (27, 64) (14, 25)
at any time
%
n
95% ci
First vaccination Fever 238°C Excessive fussiness Drowsiness z 25 mm redness > 25 mm swelling Tenderness
15.6 6.7 60.0 11 .l 11.1 46.7
45 45 45 45 45 45
(6.5, 29.5) (1.4, 18.3) (44.3, 74.3) (3.7, 24.1) (3.7, 24.1) (31.7, 62.1)
Second vaccination Fever 2 38°C Excessive fussiness Drowsiness > 25 mm redness > 25 mm swelling Tenderness
33.3 15.6 46.7 2.2 6.7 71.1
45 45 45 45 45 45
(20.0, 49.0) (6.5, 29.5) (31.7, 62.1) (0.1, 11.8) (1.4, 18.3) (55.7, 83.6)
Third vaccination Fever ~38°C Excessive fussiness Drowsiness > 25 mm redness z 25 mm swelling Tenderness
13.3 15.6 40.0 4.4 4.4 46.7
45 45 45 45 45 45
(5.1, 26.8) (6.5, 29.5) (25.7, 55.7) (0.5, 15.1) (0.5, 15.1) (31.7, 62.1)
18 month fourth vaccination Fever 238°C Excessive fussiness Drowsiness > 25 mm redness > 25 mm swelling Tenderness
27.3 6.8 25.0 29.5 25.0 84.1
44 44 44 44 44 44
(15.0, 42.8) (1.4, 18.7) (13.2, 40.3) (16.8, 45.2) (13.2, 40.3) (69.9, 93.4)
inflammation, and also with an older child being able to respond more specifically. The adverse effect profile was generally similar to that recorded in our previous studies with the currently licensed Australian DTP vaccine ” which is a component of the pentavalent vaccine. Although an acellular pertussis component may have produced a lower incidence of transient side effects, recent data demonstrate that whole-cell and acellular pertussis vaccines are similar in terms of rates of serious adverse events I’. Additionally, acellular vaccines do not offer any improvement in efficacy over otent European-type formulation whole-cell vaccines L . Gtven that acellular vaccines are more costly, the pedtavalent vaccine was formulated to contain a wholecell pertussis component with the aim of developing an efficacious and cost-effective vaccine which would satisfy many of the goals of the Children’s Vaccine Initiative for a global paediatric vaccine ‘. Following the first three doses, the seroresponse to the pentavalent vaccine was excellent for Hib antibody, and also good for hepatitis B although not as high as commonly seen with monovalent hepatitis B vaccine 14. This is presumably the result of component interference, though the exact mechanism is unknown. Higher levels of hepatitis B seroconversion at 7 months are desirable, however, and are likely for infants who receive monovalent hepatitis B vaccine shortly after birth 15.This is currently under evaluation. The pertussis immunogenicity profile was similar to that previously documented for the currently licensed Australian DTP vaccine ‘I. Both diphtheria and tetanus responses were well within acceptable margins for protection. The dose at 18 months produced a substantial rise in antibody titre for all antigens. As more vaccines are added to childhood immunisa-
A combined
liquid Hib (PRP-OMP), hepatitis El, and DTP, vaccine:
tion schedules, there is a need to keep those schedules simple by combining as many as possible to reduce the number of encounters, injections, and packaging and delivery costs. A reduction from eleven (DTP, 2, 4, 6, 18 months; Hib, 2, 4, 6, 18 months; hep B, 0, 1, 7 months) to four separate injections (pentavalent, 2, 4, 6, 18 months) may be possible with a pentavalent vaccine such as the one evaluated in this study. A full controlled pre-licensure clinical evaluation of the pentavalent vaccine (in approximately 2000 infants) is now nearing completion.
4
5
6 7
ACKNOWLEDGEMENTS 8
We would like to acknowledge research assistants Noreen Condon, Margaret Flood, Elizabeth McGrath, Susie Proctor; phlebotomists Patricia Barbante, Dr Paul Carmen, Kathleen Lanigan, Charan Sandhu, and Deborah Saunders; and study doctors Len Hartman, Audrey Penington and Joanne Smart. Suzanna Vidmar assisted with the data analysis. Valuable assistance in study planning was provided by John Varigos, Jillian Bennet, and Paul Fahey. Professor Ian Gust provided helpful comments on the manuscript. Serology was carried out by the CSL Ltd Quality Control Biochemistry Department under the supervision of Lena Miloradovic. Bill ‘Woods, Serge DeBartolo and the Clinical Trials Logistics Group at CSL Ltd provided assistance to field staff and ensured that the vaccine cold chain was maintained. We thank the parents of subjects, the maternal and child health nurses in the participating areas, and staff from the health departments of the municipalities of Monash and Whitehorse. This study was supported by a grant to the Royal Children’s :Hospital Research Foundation from CSL Ltd, and by Merck and Co., Inc.
REFERENCES Mulholland, K., Hilton, S. and Adegbola, R. et al. Randomised trial of Haemophilus influenzae type-b tetanus protein conjugate for prevention of pneumonia and meningitis in Gambian infants. Lancer 1997, 349, 1191-l 197. Steinhoff, MC. Haemophilus influenzae type-b infections are preventable everywhere. /_ancet 1997, 349, 1186-l 187. Report of the Meeting of the Scientific Advisory Group of Experts (SAGE) of the Children’s Vaccine Initiative and the Global Program on Vaccination, Geneva, June 1997, WHO
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Report No. WHO/GPV/97.05. Hepatitis B virus: a comprehensive strategy for eliminating transmission in the United States through universal childhood immunisation. Recommendations of the Immunization Practices Advisory Committee. Morb. Mortal. Wkly Report 1991, 40RR-13, l-25. The Australian lmmunisation Handbook, 6th edn. National Health and Medical Research Council, Australian Government Publishing Service, Canberra, ACT 2601, 1997. Schleiss, M.R. Ever-changing world of pediatric vaccines: why are we giving more shots? J. Pediatr. 1997, 130, 505-507. Schmitt, H.J. lmmunogenicity and reactogenicity of 2 Hib tetanus conjugate vaccines administered by reconstituting with DTPa or given as separate injections [abstract]. In: Proceedings of the 35th ICAAC (G63), September 1995. American Society for Microbiology, Washington D.C. Pichichero, M.E., Christy, C., Decker, M.D., Steinhoff, MC., Edwards, K.M., Rennels, M.B., Anderson, E.L. and Englund, J.A. Defining the key parameters for comparing reactions among acellular and whole-cell pertussis vaccines. Pediatrics 1995,96,588-592. Paradiso, P.R., Hogerman, D.A., Madore, D.V., Keyserling, H., King, J., Reisinger, KS., Blatter, M.M., Rothstein, E., Bernstein, H.H. and Hackell, J. Safety and immunogenicity of a combined diphtheria, tetanus, pertussis and Haemophilus influenzae type b vaccine in young infants. Pediatrics 1993, 92, 1-6. Aristegui, J., Garrote, E., Gonzalez, A., Arrate, J.-P., Perez, A. and Vandepapeliere, P. Immune response to a combined hepatitis B, diphtheria, tetanus and whole cell pertussis vaccine administered to infants at 2. 4 and 6 months of age. Vaccine 1997, 15, 7-9. Nolan, T.M., Hogg, G., Darcy, M.-A., Varigos, J. and McEwen, J. Primary course immunogenicity and reactogenicity of a new DTP, (diphtheria-tetanus-whole cell pertussis) vaccine. Journal of Paediatrics and Child Health 1997, 33, 413-417. Olin, P., Rasmussen, F., Gustafson, L., Hallander, H.O. and Heijbel, H. Randomised controlled trial of two-component, three component, and five-component acellular pertussis vaccines compared with whole-cell pertussis vaccine. Lancet 1997,350,1569-l 577. Plotkin, S.A. and Cadoz, M. The acellular pertussis vaccine trials: an interpretation. Pediatr infect Dis J 1997, 16, 508-517. Keyserling, H.L., West, D.J., Hesley, T.M., Bosley, C., Wiens, B.L. and Calandra, G.B. Antibody responses of healthy infants to a recombinant hepatitis B vaccine administered at 2, 4, and 12 or 15 months of age. J. Pediatr 1994, 125, 67-69. Aristegui, J., Muniz, J., Perez-Legorburu, A., Imaz, M., Arrate, J.P., Suarez, M.D. and Goiri, M.D. Newborn universal immunisation against hepatitis B: immunogenicity and reactogenicity of simultaneous administration of diphtheria/tetanus/ pertussis (DTP] and oral polio vaccines with hepatitis B vaccine at 0, 2 and 6 months of age. Vaccine 1995, 13, 973-977.
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