A Lyme disease mRNA vaccine targeting Borrelia burgdorferi OspA induces strong immune responses and prevents transmission in mice

Abstract

Lyme borreliosis (LB), caused by Borrelia burgdorferi sensu lato, is one of the most common tick-borne diseases in the northern hemisphere. Given its increasing global incidence, LB remains a major public health concern and the development of an effective vaccine is recognized as a key component of the overall disease prevention strategy. Here, we present results obtained with newly developed lipid nanoparticle-encapsulated mRNA vaccine candidates encoding the outer surface protein A (OspA) of B. burgdorferi sensu stricto (Bbss) serotype 1 (mRNA-OspA) with or without a secretion signal (SS) or a transmembrane domain. We evaluated the immunogenicity and protective efficacy of the mRNA-OspA vaccine candidates in a tick-fed mouse challenge model compared with an adjuvanted OspA protein subunit vaccine and the licensed canine vaccine Recombitek Lyme. At the doses tested, the mRNA-OspA vaccines induced significantly higher OspA-specific immunoglobulin G titers than the protein-based vaccines, as well as functional antibodies measured by serum bactericidal assay against Bbss strain B31. Complete protection against transmission was observed in the group immunized with the mRNA-OspA without SS. Overall, these data demonstrate that an mRNA-OspA vaccine can be effective against LB infection and could be used in the future for the prevention of Lyme disease.

Keywords: Borrelia burgdorferi; Lyme borreliosis; MT: Oligonucleotides: Therapies and Applications; OspA; immunogenicity; mRNA vaccines; outer surface protein A; tick-borne pathogens; ticks; vector-borne disease control.

Graphical abstract

Figure 1

In vitro expression of mRNA-OspA in HEK293T (A) Western blots of mRNA-OspA ST1 native, mRNA-HA-SS-OspA ST1 native, mRNA-HA-SS-OspA ST1 Gly(−), mRNA-TMB-OspA ST1 native, and mRNA-TMB-OspA ST1 Gly(−) in intracellular fraction, cell supernatant and crude extracts (transmembrane). Cell’s supernatants, crude extracts and intracellular compartments were collected after 48 or 72 h. A negative control (buffer) and a positive control (recombinant OspA) were also used. Blotted proteins (duplicates) were detected with polyclonal rabbit antibodies that recognize OspA. Antigenicity of secreted OspA ST1 antigen delivered by mRNA in HEK293 cells was also assessed. Transfected cell supernatants were used to perform sandwich ELISAs with functional monoclonal antibodies (B) LA-2, (C) 857-2, and (D) 221-7. (E) Depict localization of mAbs target epitopes on OspA ST1.

Figure 2

Schematic representation of the study procedures and timeline One hundred C3H/HeNRj mice were allocated into 10 groups and intramuscularly immunized with each vaccine preparation on days 0 and 21. Two weeks later (35 days p.i.), mice were infested with Bbss-infected Ixodes ricinus nymphs. HA-SS, HA secretion signal; Gly(-), deglycosylated; TMB , HA transmembrane domain.

Figure 3

Humoral immune responses to protein- and mRNA-based vaccines (A) Protein-based vaccines (B) mRNA-based vaccine candidates. OspA-specific IgG ELISA titers were measured following IM vaccination of C3H/HeNRj mice (n = 10 mice per group) on day 0 and day 21 with mRNA vaccines coding for OspA ST1 with or without secretion signal (HA-SS) or transmembrane domain (TMB). Bars represent geometric mean titers (GMTs) ± 95% confidence interval (10 or 9 mice in each group on days 0, 21, 28, 45, and 70, and 5 or 4 mice in each group on day 92). Horizontal dotted lines represent the limit of detection (LOD) of the assay. (C) Humoral immune responses to mRNA-OspA vaccine candidates compared with protein-based vaccines during plateau phase reached between day 35 and day 45. Individual mice are shown. OspA-specific IgG ELISA titers measured on day 45 p.i. Bars represent geometric mean titers ± 95% confidence interval. ∗p < 0.05; ∗∗p < 0.01; ∗∗∗p < 0.001; ∗∗∗∗p < 0.0001.

Figure 4

Antibody titers of different IgG subclasses measured on day 45 post-immunization. Titers are presented in individual value with the geometric mean indicated for all groups (n = 6 mice in each group). (A) IgG1 and (B) IgG2a. ∗p < 0.05; ∗∗p < 0.01; ∗∗∗p < 0.001.

Figure 5

Functional antibody responses determined by serum bactericidal assay (A) Viable numbers of B. burgdorferi B31 (log bacterial count) after 48 h of incubation with different serum pools collected on day 45. The x axis represents the serial 2-fold dilutions (from 1:8 to 1:16,384) of each serum pool. A mean of 2 × 10⁸ spirochetes were counted in the negative control wells. The red dashed horizontal line represents the bactericidal activity for which the sera dilution is able to kill 50% (1 × 10⁸) of spirochetes present in the well. Spirochetes survivability was counted by DFM. (B) Survival of B. burgdorferi ss strain B31 GCB in the presence of guinea pig complement and sera collected from mice immunized with protein- or mRNA-based vaccines coding for OspA ST1 with or without secretion signal (HA-SS) or transmembrane domain (TMB). The viability of spirochetes (in green) was assessed after 48 h of incubation, using fluorescent microscope (Leica Microsystems). Scale bar, 200 μm. Gly(-), deglycosylated.