Ability of 3 Different Meningococcal C Conjugate ...

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Ability of 3 Different Meningococcal C Conjugate Vaccines to Induce Immunologic Memory after a Single Dose in UK Toddlers Peter Richmond,1,2 Ray Borrow,3 David Goldblatt,2 Jamie Findlow,3 Sarah Martin,3 Rhonwen Morris,4 Keith Cartwright,4 and Elizabeth Miller1

1 Immunisation Division, Communicable Disease Surveillance Centre, Public Health Laboratory Service, and 2Immunobiology Unit, Institute of Child Health, London, 3Public Health Laboratory Service, Meningococcal Reference Unit, Withington Hospital, Manchester, and 4Gloucester Vaccine Evaluation Unit, Public Health Laboratory, Gloucestershire Royal Hospital, Gloucester, United Kingdom

The incidence of meningococcal C disease has increased rapidly in the United Kingdom over the last 5 years and is now responsible for an estimated 1530 cases and 150 deaths per year, many in young infants and adolescents [1]. The short duration of protection from meningococcal C polysaccharide vaccines and their lack of effectiveness in young children have led to the development of meningococcal C conjugate (MCC) vaccines that are safe, immunogenic, and prime for memory in infants with a 3-dose schedule [2] and in toddlers with a 2-dose schedule [3]. To achieve rapid disease control with MCC vaccines, a catchup immunization program was planned in the United Kingdom for all children aged 1–18 years. Therefore, it was essential to determine whether a single dose would be sufficient to provide long-term protection in children after the age of 12 months.

Received 22 May 2000; revised 5 September 2000; electronically published 14 November 2000. Parents gave written informed consent for study participation. The study was approved by the Public Health Laboratory Service and local ethics committees. Financial support: Research and Development Division, UK Department of Health, London. Reprints or correspondence: Dr. Elizabeth Miller, Immunisation Division, Communicable Disease Surveillance Centre, Public Health Laboratory Service, 61 Colindale Ave., London NW9 5EQ, United Kingdom (emiller@phls .org.uk). The Journal of Infectious Diseases 2001; 183:160–3 q 2001 by the Infectious Diseases Society of America. All rights reserved. 0022-1899/2001/18301-0023$02.00

We evaluated the ability of 3 different MCC vaccines to induce immunologic memory after a single dose in toddlers. Materials and Methods Study population. Children 12–18 months old who were eligible for routine measles, mumps, and rubella (MMR) immunization were recruited from October 1997 through July 1998 from general practices in Hertfordshire and Gloucestershire. Toddlers were excluded if they were immunocompromised or had a history of meningococcal disease. Subjects were randomized to receive 1 of 3 candidate MCC vaccines by a computerized block randomization procedure. Vaccines and immunization. Two of the 3 MCC vaccines contained short-chain oligosaccharides (10 mg) derived from serogroup C capsular polysaccharide (O-acetylated), coupled to CRM197, a nontoxic mutant diphtheria toxin (Chiron Vaccines and Wyeth Lederle Vaccines and Pediatrics). The third contained serogroup C polysaccharide (10 mg; de-O-acetylated), conjugated to tetanus toxoid (MCC-TT; North American Vaccine [NAVA]). MCC vaccines (0.5mL dose) were given by intramuscular injection into the right anterolateral thigh or deltoid, according to local practice; 0.5 mL of MMR vaccine (MMR-II; Pasteur Me´rieux) was given at the same time by intramuscular injection into the contralateral limb. Six months later, children received a 0.1-mL dose of meningococcal AC polysaccharide (MACP) vaccine (Mengivac [A 1 C]; Pasteur Me´rieux), containing 10 mg each of meningococcal A and C polysaccharides. Reactogenicity was documented by 7-day diaries kept by parents, in which they recorded axillary temperatures, local reactions, and systemic symptoms, and by interviews with study nurses at 24 h,

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To test for immunologic memory after a single dose of meningococcal C conjugate (MCC) vaccine in toddlers, 226 children 12–18 months old were randomized to receive 1 of 3 MCC vaccines, with a C polysaccharide booster 6 months later. The protein conjugate was diphtheria mutant toxoid in 2 vaccines (MCC-CRM197) and was tetanus toxoid in the third (MCC-TT). One month after the MCC vaccines, 91%–100% of children had serum bactericidal antibody (SBA) titers >8, and 89%–100% had a >4-fold increase. Geometric mean titer (GMT) increased from !4 to 215 (95% confidence interval [CI], 166–279). MCC-TT induced higher SBA GMTs (P ! .001) and higher proportions with SBA >8 (P p .02) than did the MCC-CRM197 vaccines. By 6 months, GMTs had decreased to 55.1 (95% CI, 40–76), but IgG antibody avidity increased (P ! .001). Induction of immunologic memory was confirmed by a GMT of 1977 (range, 1535–2547) after the polysaccharide booster and a further increase in avidity. This evidence justified the use of a single dose in a catch-up immunization program for children 1–18 years old.

JID 2001;183 (1 January)

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Table 1. Meningococcal serogroup C serum bactericidal geometric mean titers (GMTs) for strain C11 to meningococcal C conjugate vaccine (MCC) at a mean age of 13.6 months and to a booster dose of AC polysaccharide vaccine (MACP; 10-mg dose) at a mean age of 20.5 months, according to vaccine formulation. Timing of blood sample

MCC-CRM197 (Wyeth)

MCC-TT (NAVA)

69

68

75

!4

!4

!4

1 (1) 1 (1) 72 123 (78–195) 66 (92) 59 (82) 61/67 (91) 63 19 (11–32) 36 (57) 26 (41) 63 1318 (875–1986) 62 (98) 61 (97)

2 (3) 2 (3) 70 141 (90–222) 64 (91) 58 (83) 58/65 (89) 65 51 (30–85) 49 (75) 43 (66) 63 979 (686–1400) 63 (100) 63 (100)

3 (4) 1 (1) 72 564 (406–783) 72 (100) 70 (97) 72/72 (100) 66 166 (99–281) 57 (86) 55 (83) 71 5272 (3483–7980) 71 (100) 71 (100)

a

P

NA .782 .695 !.001

.022 .004 .006 !.001

.001 !.001 !.001

.64 .20

NOTE. Data are no. (%) unless otherwise indicated. Data are from subjects with sufficient postvaccination serum samples for analysis. CI, confidence interval; NA, not available; NAVA, North American Vaccine; TT, tetanus toxoid. a P values for differences among vaccines.

7 days, and 4 weeks after vaccination. Significant illnesses and hospitalizations in the 7 months after MCC vaccination were also documented. Serum samples were obtained before and 4–6 weeks after immunization. Serum samples were separated, were stored at 2807C, and were transported frozen to the Public Health Laboratory Service Meningococcal Reference Unit (Manchester, UK) for analysis. Serologic studies. Serum samples were tested by using standardized complement-mediated serum bactericidal antibody (SBA) against O-acetylated C11 strain (phenotype C:16:P1.7a,1), as described elsewhere [2]. The complement source was pooled 3–4week-old baby rabbit serum (Pelfreeze Biologicals). SBA titers were expressed as the reciprocal of the final serum dilution giving >50% killing at 60 min. SBA titers !4 were assigned a value of 2 for computational purposes. Serogroup C–specific IgG was measured by standardized ELISA [4], with Centers for Disease Control and Prevention (CDC) 1992 reference serum and O-acetylated serogroup C polysaccharide (NIBSC). The lower limit of the assay was 0.1 mg/mL; serum samples with undetectable antibody levels were assigned a value of 0.05 mg/mL. If sufficient volumes of serum samples with IgG levels 10.6 mg/mL were available, serogroup C–specific IgG avidity was tested by an elution ELISA, using the chaotrope thiocyanate, as described elsewhere [5], and was modified for meningococcal serogroup C assay [6]. All serologic testing was done blinded to the MCC vaccine given. Statistical evaluation. The sample size was based on 95% confidence intervals (CIs) of 83%–97% per group around the proportion with an SBA titer >8 after MCC vaccination, assuming that the true proportion was 90%. Analysis was by intention to treat. Antibody levels and avidity indices were log transformed for the calculation of geometric mean titers (GMTs), geometric mean concentrations (GMCs), geometric mean avidity indices (GMAIs), and 95% CIs. Antibody levels for each vaccine at each time point were

compared by analysis of variance (ANOVA). If ANOVA revealed significant variation, means were compared among vaccines by Student’s t test. Differences between pre- and postvaccination antibody levels and avidity indices were investigated by paired Student’s t tests, and differences among vaccines in frequency of symptoms and proportions above specified titers were investigated by x2 and Fisher’s exact tests as appropriate.

Results In total, 226 children (128 boys and 98 girls) received an MCC vaccine dose at a mean age of 13.6 months (range, 12–18 months), 75 each in the Chiron and Wyeth groups and 76 in the NAVA group. A booster dose of 10 mg of C polysaccharide was given to 217 children (mean age, 20.5 months; range, 17–28 months). Nine children withdrew before the polysaccharide booster, 4 from the Chiron, 2 from the Wyeth, and 3 from the NAVA groups. Local erythema or swelling >3 cm within 7 days at the MCC injection site occurred in 2.3% and 1.8% of children, respectively. Elevated temperatures >387C were reported in 5% of subjects within 3 days of vaccination. There were no significant differences in reactogenicity among the 3 MCC vaccines. One child had a febrile convulsion 9 days after vaccination that was attributed to the MMR vaccine. No serious illnesses or sequelae related to vaccination were reported in the 7 months after immunization. One child developed urticaria 6 h after MACP vaccination that resolved uneventfully within 6 days without treatment. SBA responses. Before vaccination, 94% of children had a


Before MCC, n GMT (95% CI) Titer >8 Titer >32 After MCC, n GMT (95% CI) Titer >8 Titer >32 >4-Fold increase Before MACP, n GMT (95% CI) Titer >8 Titer >32 After MACP, n GMT (95% CI) Titer >8 Titer >32

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Richmond et al.


Table 2. Meningococcal serogroup C–specific IgG geometric mean antibody concentrations (GMCs; mg/mL) and geometric mean avidity indices (GMAIs; with 95% confidence intervals [CIs]) before and after meningococcal C conjugate (MCC) vaccines at a mean age of 13.6 months and before and after a booster of AC polysaccharide vaccine (MACP; 10-mg dose) at a mean age of 20.5 months, according to vaccine formulation. Timing of blood sample

NOTE.

n

MCC-CRM197 (Wyeth)

n

MCC-TT (NAVA)

n

0.81 (0.68–0.97) NA

64 NA

0.84 (0.71–1.01) NA

63 NA

0.88 (0.75–1.05) NA

65 NA

8.03 (6.3–10.2) 69.8 (63.9–76.3)

64 47

11.1 (8.4–14.6) 72.5 (66.4–79.2)

57 45

13.30 (11.0–16.0) 52.9 (49.8–56.2)

62 47

0.94 (0.73–1.20) 91.9 (84.6–99.9)

61 42

1.70 (1.3–2.2) 93.9 (85.5–105.2)

61 41

1.84 (1.46–2.33) 87.0 (81.6–94.0)

68 43

10.82 (8.2–14.2) 127.6 (119.7–136.1)

61 48

9.00 (6.9–11.8) 123.5 (115.7–131.9)

58 46

14.20 (11.4–17.7) 132.3 (126.3–138.7)

70 41

NA, not available; NAVA, North American Vaccine; TT, tetanus toxoid.

titer !4. After a single dose, 91%–100% of children achieved titers >8 (table 1), and 89%–100% had a >4-fold increase in titer. There was no difference in SBA response between MCCCRM197 vaccines (P p .68), but GMTs were higher after MCCTT vaccination (P ! .001 for both). The proportions of children with SBA titers >8 or >32, or with >4-fold increase in SBA, were also higher after MCC-TT vaccination. Antibody levels declined by 6 months after vaccination (P ! .001 ), with an overall GMT across the 3 vaccine groups of 55.1 (95% CI, 40–76), although the majority (57%–86%) of children had titers >8. SBA levels remained higher in children after MCC-TT vaccination than after MCC-CRM197 vaccinations (P ! .001) and were higher in the Wyeth group than in the Chiron group (P p .01). GMTs in each group after the MACP booster were 7–11fold higher than after MCC vaccination (P ! .001 ). In all, 99% of children (including 19 of 21 with a titer !32 after MCC vaccination) had titers >32. There were no significant differences among the groups (P p .2). GMT after boosting was higher in toddlers primed with MCC-TT than in toddlers primed with MCC-CRM197 vaccines (P ! .001). IgG responses. IgG GMCs after MCC were similar among vaccines (P p .08). After MACP administration, IgG levels were similar to those after MCC vaccination (table 2). GMAIs were similar after both MCC-CRM197 vaccinations (P p .54) but were lower after MCC-TT vaccination (P ! .001 ; table 2). There was an increase in GMAI at 6 months, compared with that 1 month after MCC vaccination (P ! .001 in all groups; table 2). GMAIs were similar among vaccines at 6 months (P p .415). GMAI increased in all groups from before and after MACP administration (P ! .001 ); there were no differences in GMAI among vaccine groups after administration of MACP vaccine (P p .294).

Discussion In November 1999, the United Kingdom became the first country to introduce MCC vaccines [1]. The routine schedule consists of 3 doses at ages 2, 3, and 4 months with no booster and was based on the evidence of induction of immunologic memory with this regimen [2]. The high morbidity from meningococcal C disease in older age groups indicated the need for catch-up immunization. With the marked age dependency of antibody response to C polysaccharide, it was argued that, if toddlers 12–14 months old were primed for memory after a single dose, this would allow for extrapolation to older age groups. The putative correlate of protection for meningococcal C disease is an SBA titer >4 with a human complement source [7]. On the basis of a reported 10-fold difference in IgG titer when rabbit was substituted for human complement [8], we used a cutoff >32, in addition to a cutoff >8, as used in earlier MCC studies [2, 9, 10]. The proportion of children achieving the higher level was 82%–97% after MCC vaccination, declining to 41%–83% 6 months later. Large decreases in SBA level were also seen 6 months after the infant course [2, 10]. This suggests that, as with Haemophilus influenzae type b (Hib) conjugate vaccines [11], reliance on immunologic memory, rather than maintenance of SBA levels above a protective threshold, will be necessary if MCC vaccines are to protect long term. Evidence of immunologic memory was based on the booster response to MACP vaccine and maturation of antibody avidity. The higher SBA response to 10 mg of C polysaccharide in our study children, compared with that in naive UK children !2 years old receiving a full 50-mg dose (GMT, 3.2; 95% CI, 1.98–5.7; 7% SBA, >8) [9], is a clear indication of priming for memory. Furthermore, the increase in IgG avidity 6 months after MCC vaccination suggests the selective stimulation of high avidity memory B cells. The functional importance of avidity maturation is re-


Before MCC GMC GMAI After MCC GMC GMAI Before MACP GMC GMAI After MACP GMC GMAI

MCC-CRM197 (Chiron)


Meningococcal Vaccine in Toddlers

Acknowledgments We thank the study nurses in Hertfordshire and Gloucestershire who conducted the study; Pauline Kaye, Jo Southern, Joan Vurdien, and Teresa Gibbs (Public Health Laboratory Service [PHLS] Communicable Disease Surveillance Centre, London) for study administration and data management; and Nick Andrews (PHLS Statistics Unit, London) for statistical advice. We acknowledge the assistance of Moya Burrage and Carol Thornton (Centre for Applied Microbiology and Research, Salisbury, UK) in sample processing and assay standardization, Janet Suker and Ian Feavers (National Institute for Biological Standards and Research, Hertfordshire, UK) for provision of meningococcal C polysaccharide, and Maggie Vickers (Institute of Child Health, London) for performing the avidity assays. We thank David Salisbury (Department of Health, London) for support of the study.

References 1. Chief medical officer, chief nursing officer, chief pharmaceutical officer. Introduction of immunization against group C meningococcal infection. London: Department of Health, 1999 (PL/CMO/99/2, PL/CNO/99/4, PL/ CPHO/99/1). 2. Richmond PC, Borrow R, Miller E, et al. Meningococcal serogroup C conjugate vaccine in immunogenic in infancy and primes for memory. J Infect Dis 1999; 179:1569–72. 3. Macdonald NE, Halperin SA, Law BJ, Forrest B, Danzig LE, Granoff DM. Induction of immunologic memory by conjugated vs. plain meningococcal C polysaccharide vaccine in toddlers. JAMA 1998; 280:1685–9. 4. Gheesling LL, Carlone GM, Pais L, et al. Multicenter comparison of Neisseria meningitidis serogroup C anti-capsular polysaccharide antibody levels measured by a standardized enzyme-linked immunosorbent assay. J Clin Microbiol 1994; 32:1475–82. 5. Goldblatt D, Pinto Vas RPJ, Miller E. Antibody avidity as a surrogate marker of successful priming by Haemophilus influenzae type b conjugate vaccines following infant immunization. J Infect Dis 1998; 177:1112–5. 6. Richmond P, Goldblatt D, Fusco PC, et al. Safety and immunogenicity of a new Neisseria meningitidis serogroup C–tetanus toxoid conjugate vaccine in healthy adults. Vaccine 1999; 18:641–6. 7. Goldschneider I, Gotschlich EC, Artenstein MS. Human immunity to the meningococcus. I. The role of humoral antibodies. J Exp Med 1969; 129: 1307–26. 8. Griffiss JM, Goroff DK. IgA blocks IgM and IgG-initiated immune lysis by separate molecular mechanisms. J Immunol 1983; 130:2882–5. 9. Borrow R, Richmond P, Kaczmarski EB, et al. Meningococcal serogroup C–specific IgG antibody responses and serum bactericidal titers in children following vaccination with a meningococcal A/C polysaccharide vaccine. FEMS Immunol Med Microbiol 2000; 28:79–85. 10. Fairley CK, Begg N, Borrow R, Fox AJ, Jones DM, Cartwright K. Conjugate meningococcal serogroup A and C vaccine: reactogenicity and immunogenicity in United Kingdom infants. J Infect Dis 1996; 174:1360–3. 11. Eskola J, Ward J, Dagan R, Goldblatt D, Zepp F, Seigrist CA. Combination vaccination of Haemophilus influenzae type b conjugate and diphtheriatetanus-pertussis containing acellular pertussis vaccines. Lancet 1999; 354: 2063–8. 12. Granoff DM, Maslanka SE, Carlone GM, et al. A modified enzyme-linked immunosorbent assay for the measurement of antibody responses to meningococcal C polysaccharide that correlates with bactericidal responses. Clin Diagn Lab Immunol 1998; 5:479–85. 13. Liebermann JM, Chiu SS, Wong VK, et al. Safety and immunogenicity of a serogroups A/C Neisseria meningitidis oligosaccharide-protein conjugate vaccine in young children: a randomized controlled trial. JAMA 1996; 275:1499–503. 14. Antilla M, Eskola J, Ahman H, Kahty H. Avidity of IgG for Streptococcus pneumoniae type 6B and 23F polysaccharides in infants primed with pneumococcal conjugates and boosted with polysaccharide or conjugate vaccines. J Infect Dis 1998;177:1614–21. 15. Frasch CE. Specificity of antibodies to O-acetyl-positive and O-acetyl-negative group C meningococcal polysaccharides in sera from vaccinees and carriers. Infect Immun 1991; 59:4349–56.


flected in the higher bactericidal activity of the antibodies that are produced after MACP vaccination, compared with that produced 1 month after MCC vaccination, despite similar IgG levels. In contrast, polysaccharide vaccine in young children induces low avidity IgG antibodies that lack bactericidal activity [12] and are not protective [13]. The increase in IgG antibody avidity after MCC vaccine supports the use of this measure as a surrogate marker for immunologic priming for memory, as with Hib and pneumococcal conjugate vaccines [5, 14]. The superior immunogenicity of the MCC-TT vaccine versus the MCC-CRM197 vaccines may reflect better T cell priming by the TT carrier. Differences in the size of the serogroup C polysaccharide or oligosaccharide chains linked to the particular carrier proteins may also result in differences in immunogenicity and priming among the vaccines. The acetylation status of the meningococcal C polysaccharide can also affect immunogenicity and the specificity of antibodies produced [15]. The de-Oacetylated MCC-TT vaccine may induce antibodies that are not measured with an O-acetylated–specific ELISA but may still have functional activity that results in higher SBA levels. This highlights the importance of functional antibody assays in vaccine evaluation. The results of this study formed the foundation for the UK decision to use a single dose of MCC vaccine in the catch-up campaign for children >12 months old. A postlicensure surveillance program has been implemented and will allow for the conclusions based on the immunologic criteria to be validated against measures of clinical effectiveness and will provide confidence for other countries that are contemplating a single-dose catch-up immunization program for MCC vaccines.

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