Human Antibody Responses Following Vaccinia Immunization Using Protein Microarrays and Correlation With Cell-Mediated Immunity and Antibody-Dependent Cellular Cytotoxicity Responses

Abstract

Background: There are limited data regarding immunological correlates of protection for the modified vaccinia Ankara (MVA) smallpox vaccine.

Methods: A total of 523 vaccinia-naive subjects were randomized to receive 2 vaccine doses, as lyophilized MVA given subcutaneously, liquid MVA given subcutaneously (liquid-SC group), or liquid MVA given intradermally (liquid-ID group) 28 days apart. For a subset of subjects, antibody-dependent cellular cytotoxicity (ADCC), interferon-γ release enzyme-linked immunospot (ELISPOT), and protein microarray antibody-binding assays were conducted. Protein microarray responses were assessed for correlations with plaque reduction neutralization titer (PRNT), enzyme-linked immunosorbent assay, ADCC, and ELISPOT results.

Results: MVA elicited significant microarray antibody responses to 15 of 224 antigens, mostly virion membrane proteins, at day 28 or 42, particularly WR113/D8L and WR101H3L. In the liquid-SC group, responses to 9 antigens, including WR113/D8L and WR101/H3L, correlated with PRNT results. Three were correlated in the liquid-ID group. No significant correlations were observed with ELISPOT responses. In the liquid-ID group, WR052/F13L, a membrane glycoprotein, correlated with ADCC responses.

Conclusions: MVA elicited antibodies to 15 vaccinia strain antigens representing virion membrane. Antibody responses to 2 proteins strongly increased and significantly correlated with increases in PRNT. Responses to these proteins are potential correlates of protection and may serve as immunogens for future vaccine development.

Clinical trials registration: NCT00914732.

Keywords: ADCC; ELISPOT; MVA vaccine; antibody responses; correlation of protection; membrane proteins; modified vaccinia Ankara; protein microarray; smallpox; vaccinia western reserve.

© The Author(s) 2021. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.

Figures

Figure 1.

Overview of modified vaccinia Ankara (MVA) vaccine study and assays. Abbreviations: ADCC, antibody-dependent cellular cytotoxicity; ELISPOT, enzyme-linked immunospot; IFN, interferon; liquid-ID, liquid MVA given intradermally; liquid-SC, liquid MVA given subcutaneously; lyophilized-SC, lyophilized MVA given subcutaneously; TCID50, median tissue culture infective dose.

Figure 2.

Median adjusted spot-forming cell (SFC) assessed using interferon γ enzyme-linked immunospot (400 000 cells per well), by modified vaccinia Ankara (MVA) vaccine study visit and MVA vaccine group. The first vaccination was on day 0, and the second vaccination on day 28. Error bars represent 95% bootstrap confidence intervals (CIs). Abbreviations: liquid-ID, liquid MVA given intradermally; liquid-SC, liquid MVA given subcutaneously; lyophilized-SC, lyophilized MVA given subcutaneously.

Figure 3.

Antibody-dependent cellular cytotoxicity results by modified vaccinia Ankara (MVA) vaccine study visit and MVA vaccine group. A, Median response in normalized lytic units (NLUs). B, Median fold change in response. The first vaccination was on day 0, and the second vaccination on day 28. Error bars represent 95% bootstrap confidence intervals (CIs). Abbreviations: liquid-ID, liquid MVA given intradermally; liquid-SC, liquid MVA given subcutaneously.

Figure 4.

Among vaccinia antigens that elicited significant responses, 2 virion membrane proteins (WR101/H3L and WR113/D8L) induced the strongest modified vaccinia Ankara (MVA) antibody responses, which were also correlated with day 42 plaque reduction neutralization titer (PRNT) for the group receiving liquid MVA given subcutaneously (liquid-SC). A, Radar plot summarizing fold change of MVA antibody responses against Vaccinia virus Western Reserve (VACV-WR) antigens that elicited statistically significant antibody responses. Each ray represents 1 of 15 VACV-WR antigens that elicited significant antibody responses (antigens that are known viral membrane proteins [black] and those that are not [gray]). Fold rise is represented by circles and numbers 0–6. Lines represent fold changes for each day (day 28 [gray] and day 42 [black]) and MVA vaccine groups (liquid-SC [solid lines] and liquid MVA given intradermally [liquid-ID; dotted lines]). ID and SC labels below antigen names indicate that the change from prevaccination values was significant for the liquid-ID or liquid-SC group, respectively (day 28 [gray] and day 42 [black]). ID+ indicates that the liquid-ID group had significantly higher responses than the liquid-SC group for that day. B, Scatterplots summarizing correlations between WR101/H3L and WR113/D8L day 42 log fold change and Saint Louis University (SLU) PRNT. Spearman correlation coefficients and associated P values are included in each panel. Lines represent locally weighted scatterplot smoothing trend lines.