A chikungunya virus-like particle vaccine reduces chikungunya disease in cynomolgus macaques and protection is mediated by antibody transferred from vaccinated humans

Abstract

Chikungunya virus (CHIKV) causes periodic outbreaks and is endemic in more than 110 countries. VIMKUNYA, a CHIKV virus-like particle (CHIKV VLP) vaccine, was recently approved by regulators in the United States, European Union, and United Kingdom. Efficacy of VIMKUNYA in endemic settings is difficult to evaluate due to outbreak unpredictability. We used cynomolgus macaques, which model human CHIKV viremia and disease, to assess CHIKV VLP vaccine efficacy. Doses as low as 1.25 mg of CHIKV VLP with aluminum hydroxide adjuvant and passively transferred IgG from vaccinated humans significantly reduced viremia, disease, and joint pathology. Despite having IgG doses yielding neutralizing titers below the established predicted protective threshold of ≥100, animals fared better than the CHIKV-infected control animals, suggesting clinical benefits may occur at lower antibody levels. These findings demonstrate immunogenicity and protective efficacy of CHIKV VLP and relevance of neutralizing antibodies in protection, reinforcing its use in humans to protect against chikungunya disease.

Figure 1:. CHIKV plasma viremia kinetics in cynomolgus macaques inoculated with one of 3 different virus doses, Study A.

a) Experimental design. b) CHIKV RNA levels in plasma over time c) Peak CHIKV RNA levels by dose group. d) Area under the curve (AUC) for CHIKV RNA levels. e) Infectious CHIKV levels in plasma. f) Peak infectious CHIKV levels. g) AUC for infectious CHIKV levels. Data are based on log-transformed values above the LOD. Group means are indicated by short lines. Dotted lines show LOD and LOQ. Statistical significance was assessed using Kruskal-Wallis tests.

Figure 2:. CHIKV plasma viremia kinetics in cynomolgus macaques vaccinated with one of 3 doses of CHIKV VLP + aluminum hydroxide (alum), or VLP or aluminum hydroxide only and then challenged with 10⁷ PFU CHIKV LR-OPY1, Study B. a) Experimental design. b-f) CHIKV RNA levels in plasma, and g) (left) peak and (right) AUC of CHIKV RNA.

AUC was calculated on log-transformed RNA levels using only the area above the LOD. The LOD and LOQ are indicated by horizontal dotted lines. P-values are based on Mann-Whitney tests.

Figure 3:. Infectious CHIKV levels in plasma measured in plaque forming units for CHIKV VLP vaccinated or aluminum hydroxide (alum) administered and CHIKV challenged cynomolgus macaques, Study B.

The LOD is indicated by the line and ranged from 0.9 to 2.1 log₁₀ PFU/ml depending on the input volume. Samples that were undetectable were given a value of the LOD for graphing. AUC analysis of CHIKV PFU over time was based on log-transformed levels using the area above the LOD. Short lines show means. The 3 animals from Study A (A04-A06) that received the same dose (10⁷ PFU) of CHIKV LR-OPY1 are shown for comparison.

Figure 4:. Temporal kinetics of CHIKV neutralizing antibody titers after CHIKV VLP vaccination or aluminum hydroxide (alum) treatment and CHIKV challenge in cynomolgus macaques. a) Kinetic changes in mean neutralization antibody titers (NT₈₀).

Each line represents the cohort mean and error bars show standard deviations. Pairwise comparisons of day 56 pre-challenge titers between groups used Tukey’s multiple comparisons and pink shading highlights p-values <0.05. b) NT₈₀ titers on the day of CHIKV challenge, study day 56. Each dot represents an individual; horizonal lines indicate group means, also shown above groups. The accompanying table shows geometric mean titers (GMTs) on the day of challenge. For calculating GMTs, NT₈₀ titers below the LOD of 19 were assigned a value one half the LOD (9.5) and values greater than the assay upper limit of quantification (ULOQ) were assigned a value of the ULOQ. LLOD is the lower limit of detection.

Figure 5:. CHIKV RNA levels and histopathology scores in joint tissues and muscles and tendons after CHIKV VLP vaccination or aluminum hydroxide (alum) treatment and CHIKV challenge in cynomolgus macaques. a) CHIKV RNA levels in ankle joint capsule and cartilage, right quadricep with tendon, stifle joint capsule and cartilage, and wrist joint capsule and cartilage. b) Histopathology scores in joints and all tissues.

For all panels, each dot represents one animal, box plot lines indicate group means, and whiskers indicate the full range. Samples with undetectable CHIKV RNA were assigned a value halfway between the LOD and the LOQ. P values are from Tukey-Kramer and Mann-Whitney tests.

Figure 6:. CHIKV plasma viremia kinetics in cynomolgus macaques administered IgG from CHIKV VLP vaccinated or naive people and then challenged with 10⁷ PFU CHIKV LR-OPY1, Study C. a) Experimental design, b) Neutralizing antibody titers on the day of challenge 1 day after passive transfer of IgG, c) CHIKV RNA levels in plasma, d) AUC analysis of CHIKV RNA over time and, e) Peak CHIKV RNA levels.

The LOD and LOQ are indicated by horizontal dotted lines. AUC was calculated on log-transformed RNA levels using only the area above the LOD of the RT-PCR assay.

Figure 7:. Infectious CHIKV levels in plasma measured in plaque forming units for animals that received antibody from CHIKV VLP vaccinated or naïve people.

The LOD is indicated by the line at 1.7 log₁₀ PFU/ml. Samples that were undetectable were given a value of the LOD.

Figure 8:. CHIKV RNA levels and histopathology scores in joint tissues and muscles and tendons after CHIKV IgG administration and CHIKV challenge in cynomolgus macaques. a) CHIKV RNA levels in ankle joint capsule and cartilage, right quadricep with tendon, stifle joint capsule and cartilage, and wrist joint capsule and cartilage and b) Histopathology scores in joints and all tissues.

Each dot shows the value from one animal, lines in boxes show means, and whiskers show ranges. Samples with undetectable CHIKV RNA were assigned a value of the midpoint between the LOD and LOQ for graphing and statistical analysis. Statistical designations are based on ANOVA with Tukey’s multiple comparisons tests.

Figure 9:

Logistic regression model of binary outcome (viremia/no viremia) as RT-qPCR 10 day of challenge serum neutralizing antibody titer.

Figure 10:. Correlation between serum neutralizing antibody levels and plasma viral RNA kinetics.

NT₈₀ titer shows neutralizing antibody levels on day 1 (the day of CHIKV challenge) correlated with plasma CHIKV RNA AUC (left) and peak viremia (right).

Figure 11:

Multivariate analysis. a). Bubble plot showing neutralizing antibody titers on day 1, (just prior to CHIKV challenge), total joint histology scores, and peak and AUC values of CHIKV RNA levels in plasma. b). Spearman r multiple variable correlation matrix.