1 / 25

OPA methods in clinical vaccine trials: experience with killing and flow cytometric OPA methods

OPA methods in clinical vaccine trials: experience with killing and flow cytometric OPA methods. Nina Ekstrom Vaccine Immunology Laboratory National Public Health Institute (KTL) Helsinki, Finland. OPA in clinical vaccine trials. Killing type OPA (Romero-Steiner et al.1997)

kirti
Download Presentation

OPA methods in clinical vaccine trials: experience with killing and flow cytometric OPA methods

An Image/Link below is provided (as is) to download presentation Download Policy: Content on the Website is provided to you AS IS for your information and personal use and may not be sold / licensed / shared on other websites without getting consent from its author. Content is provided to you AS IS for your information and personal use only. Download presentation by click this link. While downloading, if for some reason you are not able to download a presentation, the publisher may have deleted the file from their server. During download, if you can't get a presentation, the file might be deleted by the publisher.

E N D

Presentation Transcript

  1. OPA methods in clinical vaccine trials: experience with killing and flow cytometric OPA methods Nina Ekstrom Vaccine Immunology Laboratory National Public Health Institute (KTL) Helsinki, Finland

  2. OPA in clinical vaccine trials • Killing type OPA (Romero-Steiner et al.1997) • Various phase 2 studies with different Pnc-conjugate vaccines in Finnish infants (Anttila et al. 1999) • 7-valent PncCRM and PncOMPC in Finnish infants in The Finnish Otitis Media Vaccine Trial (FinOM) • 11-valent PncDT in Filipino infants (Puumalainen et al. 2003) • 23-valent PS in HIV+ Ugandan adults (French et al. 2004) • 11-valent PncDT in Finnish and Israeli infants (Wuorimaa et al. 2005) • 11-valent Pn-PD in Finnish infants (Nurkka et al. 2005) • Flow cytometric OPA (Martinez et al. 1999) • 11-valent PncDT in Filipino infants (Lucero et al. 2004) • 9-valent PncCRM in South-African infants (Mahdi et al. 2005)

  3. The Finnish Otitis Media Vaccine Trial(FinOM) • Randomized, double-blind, phase 3 cohort study designed to evaluate in parallel two 7-valent Pnc conjugate vaccines (N= 2497 Finnish infants) • Study vaccines ( 2, 4, 6 and 12 months of age) • PncCRM (Wyeth Vaccines) N=835 • PncOMPC (Merck & Co. Inc.) N= 831 • Hepatitis B (Merck & Co. Inc.) N=831

  4. Aims of the FinOM Vaccine Trial • Efficacy of two PCVs against serotype-specific pneumococcal acute otitis media (AOM) compared with control vaccine • Immunogenicity • Quality of antibodies (avidity) • Functionality of antibodies (OPA)  choice of OPA method • Serological correlates of protection

  5. Comparison of OPA methods • Killing type OPA (Romero-Steiner et al.1997) • Radio-OPA (Vidarsson et al.1994) • Flow cytometric OPA-1 (Jansen et al.1998) • Flow cytometric OPA-2 (Martinez et al.1999) • Sera from infants (n=10-16) immunized at 2, 4, 6 with heptavalent PncCRM and at 15 mo with PncCRM or 23-valent PncPS • Pnc serotypes 6B and 19F (reference strains from CDC) • IgG concentrations were determined by EIA

  6. Differences in OPA protocols Vakevainen et al. CDLI 2001

  7. Relationship between killing, radio and flow-1 OPAs 6B 19F Vakevainen et al. CDLI 2001

  8. Relationship between OPA and EIA 6B 19F Vakevainen et al. CDLI 2001

  9. Flow-2 OPA vs. other OPA methods 6B 19F Vakevainen et al. CDLI 2001

  10. Summary of comparisons • Different OPAs gave comparable results • levels of OPA were different • serotype-specific differences (19F > 6B) • OPA correlated with IgG concentration • Killing, radio > flow-1 • Highest correlation between killing and radio OPAs • Differences in sensitivity (emphasized for 19F) • Killing > Radio > Flow-2 > Flow-1

  11. The Killing OPA was chosen to be used in the FinOM Trial • Most sensitive • Measured the killing of bacteria • Standardised, ”the golden standard” • Reproducibility between laboratories had been evaluated in the multilaboratory study (Romero-Steiner et al. 2003) • Laborious • Slow (max. 90 analyses/week) • Long-term repeteability ? • Price

  12. The FinOM Trial - OPA analyses • N = 166 infants in Kangasala cohort • PncCRM N = 56 • PncOMPC N = 52 • Control (Hepatitis B) N = 58 • Immunizations at 2, 4, 6 and 12 months of age • OPA was performed for 7, 12, 13 and 24 mo samples • OPA for serotypes 6B, 19F and 23F

  13. The FinOM Trial - Killing OPA • Killing-OPA as described by Romero-Steiner et al. 1997 • Differentiated HL-60 cells as effector cells • S. Pneumoniae reference strains from CDC • Baby rabbit complement • Colonies counted manually • IgG concentrations were determined by EIA

  14. IgG concetrations & OPA titers in The FinOM Trial

  15. IgG concentration (ng/ml) required to kill 50% of Pnc (EIA:OPA ratio) * < 50% infants had a detectable OPA titer

  16. Correlation between EIA & OPA 7 mo 13 mo 6B 19F 23F

  17. OPA as a correlate of protection • Our results conform that Ab concentration is the primary correlate of protection, but that OPA is needed as a secondary correlate of protection: • Despite equal Ab concentrations the functionality of Ab’s may differ between serotypes • EIA:OPA ratio seems to correlate better with protection against AOM than Ab concentration at population level

  18. Experience with the flow cytometric OPA method • Correlates with killing OPA • Rapid and less laborious • Unaffected by antibiotics insera of e.g HIV+ persons • Multiplexing possible • Long-term repeteability ? • Flow cytometer needed • Price 19F: Alexa Fluor Dye 647 23F: 5,6-carboxyfluorescein, succinimidyl ester

  19. Experience with the flow cytometric OPA method – Soweto Trial Quantitative and Qualitative Antibody Response to Pneumococcal Conjugate Vaccine Among African Human Immunodeficiency Virus-Infected and Uninfected Children Madhi SA †, Kuwanda L*, Cutland C †, Holm A ‡, Kayhty H ‡, and Klugman KP § *National Institute of Communicable Diseases/University of the Witwatersrand/Medical Research Council: Respiratory and Meningeal Pathogens Reasearch Unit, and the †Paediatric Infectious Diseases Research Unit, Wits Health Consortium, University of the Witwatersrand, Johannesburg, South Africa; the ‡National Public Health Institute, Helsinki, Finland; and the Departments of § International Health, Rollins School of Public Health and Infectious Diseases, School of Medicine, Emory University, Atlanta, GA PIDJ 2005;24:410-16

  20. Soweto Trial • Nested study of a larger phase 3 trial that evaluated the efficacy of a 9-valent PncCRM in 39 836 children • Study vaccine or placebo were given at 6, 10 and 14 weeks of age • Blood sample was taken a month after the 3rd vaccine dose • EIA and OPA analyses were performed at KTL, Finland • OPA analyses were performed using the flow cytometric OPA assay (Martinez et al. 1999) • HL-60 cells as effector cells • S.pneumoniae strains from CDC • Baby rabbit complement

  21. Anti-Pnc in HIV + and HIV – children Madhi et al. PIDJ 2005

  22. IgG concentration (ng/ml) required for 50% uptake Madhi et al. PIDJ 2005

  23. Conclusion • Despite equal Ab concentrations PCV induced Ab’s of HIV+ and HIV- children had different functional activity • Abs produced by HIV+children were dysfunctional • The results suggest that although Ab concentration is the primary correlate of protection OPA is needed as a secondary correlate of protection

  24. Summary – OPA methods at KTL

  25. Acknowledgments Vaccine Immunology Laboratory & Clinical Unit, Department of Vaccines, KTL: Merja Vakevainen Hannele Lehtonen Maijastiina Karpala Kaisa Jousimies Merja Ryynanen Nina Nikkanen Jukka Jokinen Helena Käyhty The FinOM Study Group CDC, Atlanta: Sandra Romero-Steiner Joseph Martinez George M. Carlone Eijkman-Winkler Institute for Microbiology, Utrecht University Hospital, The Netherlands: Wouter Jansen Andre Verheul Harm Snippe National University Hospital, Reykjavik, Iceland: Eirikur Saeland Ingileif Jonsdottir Paediatric Infectious Diseases Research Unit, , University of the Witwatersrand, Johannesburg, South Africa The group of Shabir Mahdi

More Related