Recombinant ClearColi™-derived outer membrane vesicles as an effective carrier for development of neoepitope-based vaccine candidate against colon carcinoma

Abstract

Background: Colorectal carcinoma (CRC) is the third most common cancer worldwide, with high clonal heterogeneity due to somatic mutations. Poly neoepitope vaccines can inhibit the tumor's escape from the immune system. However, they have rapid clearance and low immunogenicity. Bacteria-derived recombinant outer membrane vesicles (OMVs) have gained increased attention as ideal cancer vaccine candidates due to their unique adjuvant properties and ability to carry antigens. Herein, the benefits of OMV-based and polyneoepitope-based vaccines were combined to obtain a functional individualized cancer vaccine.

Methods: OMVs and rOMVs displaying CT26 polytopes were isolated from ClearColi™ and recombinant ClearColi™ containing pET-22b (ClyA-CT26 polytope) by the AS (70 %) + UDF method. BALB/c mice were immunized with OMVs (40 µg) and rOMVs (20 and 40 µg) and subcutaneously challenged with CT26 cells. Then, IgG1 and IgG2a antibodies specific for CT26 M90 and CT26 polytope, the stimulated IFN-γ, TNF-α, and IL-10 cytokines and the stimulated CTL responses by measuring granzyme B were evaluated. To investigate whether pooled sera and pooled splenocytes are indicators of individual responses, pooled and individual methods for determining the elicited immunity were compared. Additionally, the ability of OMVs and rOMVs (20 and 40 µg) to prevent tumor growth against the CT26 challenge was investigated.

Results: Immunization with rOMVs displaying CT26 polytopes induced a higher titer of CT26 polytope- and CT26 M90 peptide-specific IgG2a than IgG1 antibodies in a dose-dependent manner, thus directing immunity to Th1. The antibody responses determined by pooled sera can be used as indicators of individual responses. In addition, both OMVs and rOMVs displaying CT26 polytopes could induce tumor-suppressing cytokines (IFN-γ and TNF-α). The ability of rOMVs displaying CT26 polytopes to induce these cytokines was higher than OMVs in a dose-dependent way. The results of the granzyme B assay were also in agreement with the cytokine assay. The survival of mice after the CT26 challenge was 100 % in the OMVs and rOMVs groups, and inhibition of tumor growth was significantly higher by rOMVs (40 µg) compared to OMVs.

Conclusion: The bioengineered OMVs displaying CT26 neoepitopes have the potential for the development of personalized tumor vaccines. Our results can provide new insights for developing rOMV-based vaccines displaying polytopes against diseases containing highly variable antigens.

Keywords: Bacterial outer membrane vesicles (OMVs); Cancer immunotherapy; Mouse model of colon carcinoma; Neoepitope.