Meningococcal disease is responsible for considerable morbidity and mortality worldwide. The case-fatality rate is above 10%, and up to 20% of survivors suffer substantial neurological sequelae [1-3]. In India, endemic disease is relatively low, but pyogenic meningitis is associated with up to 3% of acute admissions to pediatric hospitals [4]. This is probably an underestimate given the difficulty verifying Neisseria meningitidis disease involvement. Sinclair, et al. [4] reviewed the reports of meningococcal disease in India and showed that four meningococcal disease epidemics have occurred since 1930; the bulk of the disease in these epidemics and in other cases reported during non-epidemic periods was caused by serogroup A strains [4].

Vaccination offers effective protection against meningococcal illness targeting the key polysaccharide antigens in clinically relevant meningococcal serogroups: A, C, Y, and W-135; however, PS vaccines are poorly immunogenic in those below 2 years of age and elicit a T-cell-independent immune response that provides neither a memory response nor antibody affinity maturation. Repeated PS vaccine dosing may lead to reduced immunogenicity known as hypo-responsiveness [1].

MenACYW-DT, a vaccine composed of polysaccharide antigens A, C, Y and W-135, separately conjugated to diphtheria toxoid (Menactra, Sanofi Pasteur Inc., Swiftwater PA, USA) was licensed in the US in 2005 for those aged 9 month to 5 years. MenACYW-DT confers direct protection as good as polysaccharide vaccines but can also promote an anamnestic response and reduce acquisition of N. meningitidis virulent strains and transmission [3]. Circulating antibody levels were shown to be higher among adolescents 3 years after receiving a conjugate vaccine than among those given polysaccharide vaccines [1,5]. Results indicate that MenACYW-DT produces serogroup-specific protective titers in 86%-100% of 9- to 12-month-old children [6] and nearly universal protection among adolescents [5]. Serious adverse events are uncommon and rarely linked to vaccination [6,7]. A second conjugated quadrivalent meningococcal vaccine, MenACYW-CRM (Menveo; Novartis Vaccines and Diagnostics), was also recently licensed in the US.

We report on an open-label study conducted in India to assess the safety and immunogenicity of a single dose of MenACYW-DT in participants 2 to 55 years of age.

The trial followed the Declaration of Helsinki (Edinburgh revision) and Good Clinical Practice defined by the International Conference on Harmonization. The final protocol was approved prior to the start of the trial by the relevant committees at the three study sites: Institutional Ethics Committee of Maualana Azad Medical College (New Delhi), Kempegowda Institute of Medical Sciences (Bangalore), and Lokmanya Tilak Municipal Medical College and Lokmanya Tilak Municipal General Hospital (Mumbai). Participants or their parents were approached during a routine visit at the investigational site by a site employee (unaffiliated with the investigator or investigation team) and their relatives were solicited to participate. All potential, eligible, participants or their parents were invited to visit the sites where the study was explained in detail. At this point, informed consent was explained, and forms signed prior to participation.

This multicenter, open-label, non-randomized, phase III clinical trial assigned participants to one of three cohort groups: children aged 2-11 years (Child Group), adolescents aged 12-17 years (Adolescent Group), and adults aged 18-55 years (Adult Group). One objective was to describe titers of the Serum bactericidal assay in the presence of baby rabbit complement (SBA-BR) prior to and 30 days after a single dose of MenACYW-DT. Another objective was to describe the safety profile of participants after receiving 1 dose of MenACYW-DT. Participants received in the deltoid a single 0.5 mL dose of MenACYW-DT (Lot U3091AA) containing 4 µg of each of the polysaccharides (A, C, Y, and W-135) conjugated to ~48 µg diphtheria toxoid protein carrier in phospho-buffered saline.

Sera were collected before immunization and ~30-35 days post-vaccination. SBA-BR were performed by Sanofi Pasteur Global Clinical Immunology laboratories (Swiftwater, PA, USA) as per standard methodology [6,10].

Participants were observed for 30 minutes post-vaccination for reactions and to treat immediate adverse events. On Day 1 and the next 7 days, participants graded and recorded solicited adverse reactions. For injection site swelling and erythema for those aged 2–11 years, the grading criteria were Grade 1<2.5cm, Grade 2 ≥2.5cm and <5cm, and Grade 3 ≥5cm. For injection-site swelling and erythema for those aged ≥12 years, criteria were Grade 1≥2.5cm to ≤5cm, Grade 2≥5.1cm to ≤10cm, Grade 3 >10cm. Injection-site pain was recorded as Grade 1, easily tolerated; Grade 2, discomfort interfering with usual activities; and Grade 3, unable to perform usual activities. Fever was graded as Grade 1 (≥38.0ºC and ≤38.4ºC), Grade 2 (≥38.5ºC and ≤38.9ºC), and Grade 3 (≥39.0ºC). Headache, malaise, and myalgia intensity was graded as Grade 1, noticeable discomfort without interference with daily activities; Grade 2, interferes with daily activities; Grade 3, prevents daily activities.

For three consecutive days post-vaccination, participant/guardians were contacted by telephone to report all collected safety information. Information on solicited or unsolicited adverse events occurring over the first 7 days was collected during a house visit 8-10 days post-vaccination. Participants/guardians were asked to record information on other medical events 30 days post-vaccination. Data were collected and collated by electronic data capture system, and Medical Dictionary for Regulatory Activities (MedRA) preferred nomenclature described all adverse events. These were "serious" if life threatening, required hospitalization or disability, or constituted an important medical condition. Solicited systemic and injection-site adverse events were considered vaccination-related. The opinion of the investigator established other instances of vaccination-related events.

Data analysis: The sample size was sufficiently large to identify common adverse events (270 evaluable participants at 95%CI can detect an adverse event occurring with a frequency of ≥1.1%).

Statistical parameters included geometric mean titer (GMT), the % participants with a ≥4-fold increase in titer, and the % participants with SBA-BR titers ≥8 (1/dil) or ≥128 (l/dil). GMTs and their 95% confidence intervals (CIs) were calculated by normal approximation. Safety-related 95%CIs were calculated by exact binomial distribution for percentages (Clopper-Pearson).

The study enrolled 100 participants per group from June to November 2010. Participants in the Child and Adolescent Groups were evenly distributed between the LTM Medical College & General Hospital, Mumbai, India and Maulana Azad Medical College, New Delhi, India; participants in the Adult Group came from Kempegowda Institute of Medical Sciences, Bangalore, India. There were three voluntary withdrawals: one in the Child Group and two in the Adolescent Group. In the Child Group, the mean age was 7.2 years with 47% boys; in the Adolescent Group, the mean age was 14.2 years with 52% boys; in the Adult Group, the mean age was 34.8 years with 62% men.

All participants received a single MenACYW-DT dose and blood was drawn twice but three participants (1 Child Group; 2 Adolescent Group) were prematurely withdrawn due to family relocation before the second blood samples were drawn. For the second blood sampling, most were taken per-protocol, but two participants (1 Child Group; 1 Adolescent Group) were sampled at Day 29, and 8 participants (5 Child Group; 1 Adolescent Group; 2 Adult Group) were sampled between Days 36 and 52.

Temperature variations of some samples resulted in a freezer temperature excursions (TE) to +8.6°C (all samples were to remain ≤-10°C). The affected sera included both prevaccination (49 Child Group; 34 Adolescent Group) and post-vaccination (48 Child Group and 20 Adolescent Group) samples. TEs are technically protocol violations, but the SBA-BR includes a heat inactivation step of 56°C for 30 min prior to testing (a temperature increase ≥ than any sample experienced in a TE). The most likely consequence was a negligible or slightly negative change to the bactericidal activity in the sera; TE-affected samples yielded similar results compared to per-protocol results. Therefore, we pooled the immunogenicity results of all samples.

Immunogenicity: GMTs and % participants achieving a ≥4-fold increase in GMT for all age groups are presented in Table I. In general, titers rose to 1000-4000 (1/dil) 30 days post-vaccination. Exceptions were observed in serogroup C, where Child Group GMTs rose to approximately 600 (1/dil). Across serogroups, 68% to 97% of participants achieved a 4-fold titer increase.

TABLE I  Pre-vaccination and Post-vaccination Geometric Mean Titers (GMT) Measured by SBA-BR 

Contributors: SY, MVM, DHAN, SS, HSR, RA, SA, and PO designed and conducted the study; VBC developed the statistical analysis plan. All authors supervised the writing of the manuscript and subsequent revisions. All authors approved the final draft.

Funding: Sanofi Pasteur.

Competing interests: RA is an employee of Sanofi Pasteur India Pvt Ltd. PO and VBC are employees of Sanofi Pasteur. Serum bactericidal assays in the presence of baby rabbit complement (SBA-BR) were performed by Sanofi Pasteur Global Clinical Immunology laboratories (Swiftwater, PA, USA).

Acknowledgements: Dr John Bukowski of WordsWorld Consulting, Dayton, OH, USA, for professional writing assistance that was funded by Sanofi Pasteur, Swiftwater, PA, USA. The authors also thank Robert Lersch, PhD of Sanofi Pasteur, who coordinated the writing of the manuscript and provided editorial assistance.

1. Centers for Disease Control and Prevention (CDC). Prevention and Control of Meningococcal Disease. Recommendations of the Advisory Committee on Immunization Practices (ACIP). MMWR Recomm Rep. 2005;54:1-21.

2. Granoff DM, Harrison D, Borrow R. Meningococcal Vaccines. In: Plotkin SA, Orenstein WA, Offit PA, editors. Vaccines. 5th ed. Philadelphia, PA: Saunders Elsevier; 2008. p. 399-434.

3. Pelton SI, Gilmet GP. Expanding prevention of invasive meningococcal disease. Expert Rev Vaccines. 2009; 8:717-27.

4. Sinclair D, Preziosi MP, Jacob John T, Greenwood B. The epidemiology of meningococcal disease in India. Trop Med Int Health. 2010;15:1421-35.

5. Keyserling H, Papa T, Koranyi K, Ryall R, Bassily E, Bybel MJ, et al. Safety, immunogenicity, and immune memory of a novel meningococcal (groups A, C, Y, and W-135) polysaccharide diphtheria toxoid conjugate vaccine (MCV-4) in healthy adolescents. Arch Pediatr Adolesc Med. 2005;159:907-13.

6. Pina LM, Bassily E, Machmer A, Hou V, Reinhardt A. Safety and immunogenicity of a quadrivalent meningococcal polysaccharide diphtheria toxoid conjugate vaccine in infants and toddlers: three multicenter phase III studies. Pediatr Infect Dis J. 2012;31:1173-83.

7. Menactra® [prescribing information]. Swiftwater, Pa, USA; Sanofi Pasteur Inc.; 2011. Available from: http://www.fda.gov/downloads/biologicsbloodvaccines/vaccines/approvedproducts/ucm131170.pdf. Accessed March 25, 2013.

8. MENVEO® [prescribing information]. Cambridge, MA, USA: Novartis Vaccines and Diagnostics, Inc. Available from: https://　www.　novartisvaccinesdirect.　com/　PDF/Menveo_Full_Promotional_PI.pdf. Accessed March 25, 2014.

9. US National Institutes of Health (NIH). A Study of Meningococcal Vaccine, Menactra® in Healthy Subjects in India.　Available from: http://clinicaltrials.gov/ct2/show/NCT01086969?term=mta51&rank=1. Accessed March 25, 2014.

10. Maslanka SE, Gheesling LL, Libutti DE, Donaldson KB, Harakeh HS, Dykes JK, et al. Standardization and a multilaboratory comparison of Neisseria meningitidis serogroup A and C serum bactericidal assays. The Multilaboratory Study Group. Clin Diagn Lab Immunol. 1997;4:156-67.

11. Keyserling HL, Pollard AJ, DeTora LM, Gilmet GP. Experience with MCV-4, a meningococcal, diphtheria toxoid conjugate vaccine against serogroups A, C, Y and W-135. Expert Rev Vaccines. 2006;5:445-59.

12. Harrison LH. Prospects for vaccine prevention of meningococcal infection. Clin Microbiol Rev. 2006;19:142-64.

13. Maiden MC, Ibarz-Pavon AB, Urwin R, Gray SJ, Andrews NJ, Clarke SC, et al. Impact of meningococcal serogroup C conjugate vaccines on carriage and herd immunity. J Infect Dis. 2008;197:737-43.

14. Kristiansen PA, Diomande F, Ouedraogo R, Sanou I, Sangare L, Ouedraogo AS, et al. Carriage of Neisseria lactamica in 1- to 29-year-old people in Burkina Faso: epidemiology and molecular characterization. J Clin Microbiol. 2012;50:4020-7.

15. Frasch CE, Preziosi MP, LaForce FM. Development of a group A meningococcal conjugate vaccine, MenAfriVac (TM). Hum Vaccin Immunother. 2012;8: 715-24.

16. Khalil M, Al-Mazrou Y, Findlow H, Chadha H, Bosch Castells V, Johnson DR, et al. Safety and immunogenicity of a meningococcal quadrivalent conjugate vaccine in five- to eight-year-old Saudi Arabian children previously vaccinated with two doses of a meningococcal quadrivalent polysaccharide vaccine. Clin Vaccine Immunol. 2012;19:1561-6.

17. Al-Mazrou Y, Khalil M, Findlow H, Chadha H, Bosch Castells V, Johnson DR, et al. Immunogenicity and safety of a meningococcal quadrivalent conjugate vaccine in Saudi Arabian adolescents previously vaccinated with one dose of bivalent and quadrivalent meningococcal polysaccharide vaccines: a phase III, controlled, randomized, and modified blind-observer study. Clin Vaccine Immunol. 2012;19:999-1004.

18. World Health Organization (WHO). Meningococcal vaccines: WHO position paper, November 2011. Wkly Epidemiol Rec. 2011;86:521-39.

19. Lujan-Zilbermann J, Warshaw MG, Williams PL, Spector SA, Decker MD, Abzug MJ, et al. Immunogenicity and safety of 1 vs 2 doses of quadrivalent meningococcal conjugate vaccine in youth infected with Human immunodeficiency virus. J Pediatr. 2012;161:676-81.

20. Siberry GK, Warshaw MG, Williams PL, Spector SA, Decker MD, Jean-Philippe P, et al. Safety and immunogenicity　of quadrivalent meningococcal conjugate vaccine in 2-to 10-year-old human immunodeficiency virus-infected children. Pediatr Infect Dis J. 2012; 31:47-52.