Name
Novel antibody-dependent intracellular neutralization assay to assess rotavirus vaccine efficacy
Presenter
Sarah Woodyear, Cornell University
Co-Author(s)
Sarah Woodyear (1), Stanley Cryz (2), Sarah Caddy (1)
1 - Baker Institute of Animal Health, Cornell University, Ithaca, NY, USA
2 - PATH, Seattle, WA, USA
Abstract Category
Vaccines
Abstract
The development of next-generation rotavirus vaccines faces challenges due to the lack of clearly defined correlates of protection. Currently, there is no reliable post-vaccination test to accurately assess vaccine effectiveness. For other viruses, neutralization assays are generally considered more informative than ELISAs as they assess the functional activity of antibodies. However, for rotavirus, neutralization assays have shown limited correlation with protection, and therefore previous clinical trials for rotavirus vaccines have evaluated efficacy using clinical endpoints in conjunction with IgA ELISAs.
We have developed a novel rotavirus neutralization assay aimed at predicting vaccine efficacy. This assay focuses on intracellular neutralization mediated by VP6-specific antibodies, which target viral proteins that are only exposed after the virus enters the cell. We use an electroporation-based method to introduce serially diluted serum antibodies into MA104 cells, which are subsequently infected with rotavirus. Viral load is quantified by fluorescent staining of infected cells and imaging using a high-throughput Cytation 7 instrument.
First, we showed that intracellular neutralization correlates with protection from rotavirus infection in mice, whereas standard neutralization assays do not. Next, we optimized the assay to enable testing of serum samples from 154 infants vaccinated with Rotarix as part of the phase III PATH trial. Samples from 40 infants vaccinated with the non-replicating vaccine (P2-VP8) were also tested. Initial results show significant differences between samples collected pre-vaccination and 85 days post-vaccination. Unblinding of cases (infants succumbed to rotavirus-induced gastroenteritis) and controls will determine the assay's value as a correlate of protection for rotavirus vaccination.
We have developed a novel rotavirus neutralization assay aimed at predicting vaccine efficacy. This assay focuses on intracellular neutralization mediated by VP6-specific antibodies, which target viral proteins that are only exposed after the virus enters the cell. We use an electroporation-based method to introduce serially diluted serum antibodies into MA104 cells, which are subsequently infected with rotavirus. Viral load is quantified by fluorescent staining of infected cells and imaging using a high-throughput Cytation 7 instrument.
First, we showed that intracellular neutralization correlates with protection from rotavirus infection in mice, whereas standard neutralization assays do not. Next, we optimized the assay to enable testing of serum samples from 154 infants vaccinated with Rotarix as part of the phase III PATH trial. Samples from 40 infants vaccinated with the non-replicating vaccine (P2-VP8) were also tested. Initial results show significant differences between samples collected pre-vaccination and 85 days post-vaccination. Unblinding of cases (infants succumbed to rotavirus-induced gastroenteritis) and controls will determine the assay's value as a correlate of protection for rotavirus vaccination.