Synergistic Protective Activity of Tumor-Specific Epitopes Engineered in Bacterial Outer Membrane Vesicles

Abstract

IntroductionBacterial outer membrane vesicles (OMVs) are naturally produced by all Gram-negative bacteria and, thanks to their plasticity and unique adjuvanticity, are emerging as an attractive vaccine platform. To test the applicability of OMVs in cancer immunotherapy, we decorated them with either one or two protective epitopes present in the B16F10EGFRvIII cell line and tested the protective activity of OMV immunization in C57BL/6 mice challenged with B16F10EGFRvIII.Materials and methodsThe 14 amino acid B cell epitope of human epidermal growth factor receptor variant III (EGFRvIII) and the mutation-derived CD4+ T cell neo-epitope of kif18b gene (B16-M30) were used to decorate OMVs either alone or in combination. C57BL/6 were immunized with the OMVs and then challenged with B16F10EGFRvIII cells. Immunogenicity and protective activity was followed by measuring anti-EGFRvIII antibodies, M30-specific T cells, tumor-infiltrating cell population, and tumor growth.ResultsImmunization with engineered EGFRvIII-OMVs induced a strong inhibition of tumor growth after B16F10EGFRvIII challenge. Furthermore, mice immunized with engineered OMVs carrying both EGFRvIII and M30 epitopes were completely protected from tumor challenge. Immunization was accompanied by induction of high anti-EGFRvIII antibody titers, M30-specific T cells, and infiltration of CD4+ and CD8+ T cells at the tumor site.ConclusionOMVs can be decorated with tumor antigens and can elicit antigen-specific, protective antitumor responses in immunocompetent mice. The synergistic protective activity of multiple epitopes simultaneously administered with OMVs makes the OMV platform particularly attractive for cancer immunotherapy.

Vaccination With a FAT1-Derived B Cell Epitope Combined With Tumor-Specific B and T Cell Epitopes Elicits Additive Protection in Cancer Mouse Models

“…A vaccine based on EGFRvIII peptide was tested in glioblastoma patients, with promising results even though EGFRvIII-negative tumor cells ultimately escaped vaccine-induced protection ( 27 ). We previously demonstrated that OMVs decorated with EGFRvIII peptide elicited specific antibodies which could inhibit the growth of a B16F10 cell line derivative expressing EGFRvIII in syngeneic C57bl6 mice ( 24 ). Since EGFRvIII-B16F10 cells, like their progenitor B16F10, express mD8-FAT1 on their surface (Figure 4A ), we tested whether the combination of mD8-FAT1-OMVs and EGFRvIII-OMVs could further enhance the anti-tumor activity of EGFRvIII-OMVs immunization in mice challenged with EGFRvIII-B16F10.…”

“…The construction of pET-Nm-fHbp-vIII plasmid expressing the Neisseria meningitidis FHbp fused to three repeated copies of EGFRvIII peptide was previously described ( 24 ).…”

“…The recent data demonstrating that vaccines constituted by mutation-derived CD4+/CD8+ T cell neoepitopes induce anti-tumor immune responses in both preclinical and clinical settings ( 20 , 21 ), had prompted us to test whether the unique properties of OMVs could be exploited in cancer immunotherapy. We already showed that immunization with OMVs engineered with the EGFRvIII tumor-specific B cell epitope ( 22 ) and with M30, a mutation-derived CD4+ T cell epitope expressed in B16F10 murine melanoma cells ( 23 ), fully prevented tumor growth in C57bl6 mice challenged with B16F10 cells expressing EGRFvIII ( 24 ). We also showed that protection was associated to the elicitation of both anti-EGFRvIII antibodies and M30-specific T cells.…”

Outer Membrane Vesicles: Current Status and Future Direction of These Novel Vaccine Adjuvants

“…Furthermore, a recent experiment decked OMVs with tumor antigens that elicited protective anti-tumor responses in immune-competent mice. The engineered OMVs offered synergistic protective activity, resulting in OMV platform that was particularly attractive for cancer immunotherapy (Grandi et al, 2017 ).…”

Outer membrane vesicles: moving within the intricate labyrinth of assays that can predict risks of reactogenicity in humans

“…Bacterial strains can be engineered not only to modulate LPS endotoxicity but also to increase the blebbing, 4,[13][14][15] overexpress key target antigens, 16 and simultaneously express multiple variants or heterologous antigens (either proteins or polysaccharides). 17,18 This enabled to produce and test, to date mainly at preclinical level, several engineered OMVs carrying either bacterial, 19 viral, 20 parasitic, 21 and even cancer 22 antigens. Finally, the development of ad hoc industrial production, purification, and characterization processes has allowed to obtain well-defined and stable vaccine products in high yield and at low costs.…”