Different immunogenicity but similar antitumor efficacy of two DNA vaccines coding for an antigen secreted in different membrane vesicle‐associated forms

Abstract

The induction of an active immune response to control or eliminate tumours is still an unfulfilled challenge. We focused on plasmid DNA vaccines using an innovative approach whereby the antigen is expressed in association with extracellular vesicles (EVs) to facilitate antigen cross-presentation and improve induced immunity. Our two groups had independently shown previously that DNA vaccines encoding EV-associated antigens are more efficient at inducing cytotoxic T-cell responses than vaccines encoding the non-EV-associated antigen. Here, we compared our two approaches to associate the ovalbumin (OVA) antigen to EVs: (a) by fusion to the lipid-binding domain C1C2 of MFGE8(=lactadherin), which is exposed on the surface of secreted membrane vesicles; and (b) by fusion to retroviral Gag capsid protein, which is incorporated inside membrane-enclosed virus-like particles. Plasmids encoding either form of modified OVA were used as DNA-based vaccines (i.e. injected into mice to allow in vivo expression of the antigen associated to EVs). We show that both DNA vaccines induced, with similar efficiency, OVA-specific CD8+ T cells and total IgG antibodies. By contrast, each vaccine preferentially stimulated different isotypes of immunoglobulins, and the OVA-C1C2-encoding vaccine favoured antigen-specific CD4+ T lymphocyte induction as compared to the Gag-OVA vaccine. Nevertheless, both OVA-C1C2 and Gag-OVA vaccines efficiently prevented in vivo outgrowth of OVA-expressing tumours and reduced tumour progression when administered to tumour-bearing mice, although with variable efficacies depending on the tumour models. DNA vaccines encoding EV-associated antigens are thus promising immunotherapy tools in cancer but also potentially other diseases.

Proteomic comparison defines novel markers to characterize heterogeneous populations of extracellular vesicle subtypes

“…Such isolation procedures coisolate mixed EV populations, which we will call "small EVs" (sEVs), for lack of better term, in the rest of this article. Because EVs of different intracellular origins probably have different functional properties (9,10), the mixed nature of EV preparations has made the growing literature increasingly confusing, with contradictory proposed functions and clinical uses of vesicles being regularly published. The lack of specific purification and characterization tools prevents a clear understanding of the specific versus shared functional properties of the different EVs.…”

Therapeutic and diagnostic applications of extracellular vesicles

“…To make use of tumor-antigen bearing EVs without the negative characteristics of tumor-derived EVs, a DNA vaccine (delivered via an adenoviral vector or EP) was developed that encodes a fusion protein comprising (the extracellular part of) a known tumor-antigen and an EV-associated protein (C1C2 domain of lactadherin or the gag protein). Expression of this fusion construct shuttles the associated antigens to the surface or lumen of secreted EVs, respectively [157,210]. Nevertheless, this technology is limited to wellcharacterized antigens and would likely benefit from an additional immune modulator.…”

Exosome Isolation: Is There an Optimal Method with Regard to Diagnosis or Treatment?

“…Therefore, when EVs will be used for delivery of specific small RNA molecules generated in the donor cells, it is crucial to distinguish whether obtained EVs are microvesicles or exosomes. While for particular applications such as RNA delivery or drug delivery exosomes may be preferable, for other applications such as vaccination and surface antigen display microvesicles might be more relevant [40]. For diagnostic application, especially in proteomics analyses, free protein contaminants may lead to false-positive results.…”