Oncolytic adenovirus drives specific immune response generated by a poly-epitope pDNA vaccine encoding melanoma neoantigens into the tumor site

Abstract Background DNA vaccines against cancer held great promises due to the generation of a specific and long-lasting immune response. However, when used as a single therapy, they are not able to drive the generated immune response into the tumor, because of the immunosuppressive microenvironment, thus limiting their use in humans. To enhance DNA vaccine efficacy, we combined a new poly-epitope DNA vaccine encoding melanoma tumor associated antigens and B16F1-specific neoantigens with an oncolytic virus administered intratumorally. Methods Genomic analysis were performed to find specific mutations in B16F1 melanoma cells. The antigen gene sequences were designed according to these mutations prior to the insertion in the plasmid vector. Mice were injected with B16F1 tumor cells (n = 7–9) and therapeutically vaccinated 2, 9 and 16 days after the tumor injection. The virus was administered intratumorally at day 10, 12 and 14. Immune cell infiltration analysis and cytokine production were performed by flow cytometry, PCR and ELISPOT in the tumor site and in the spleen of animals, 17 days after the tumor injection. Results The combination of DNA vaccine and oncolytic virus significantly increased the immune activity into the tumor. In particular, the local intratumoral viral therapy increased the NK infiltration, thus increasing the production of different cytokines, chemokines and enzymes involved in the adaptive immune system recruitment and cytotoxic activity. On the other side, the DNA vaccine generated antigen-specific T cells in the spleen, which migrated into the tumor when recalled by the local viral therapy. The complementarity between these strategies explains the dramatic tumor regression observed only in the combination group compared to all the other control groups. Conclusions This study explores the immunological mechanism of the combination between an oncolytic adenovirus and a DNA vaccine against melanoma. It demonstrates that the use of a rational combination therapy involving DNA vaccination could overcome its poor immunogenicity. In this way, it will be possible to exploit the great potential of DNA vaccination, thus allowing a larger use in the clinic

Oncolytic adenovirus drives specific immune response generated by a poly-epitope pDNA vaccine encoding melanoma neoantigens into the tumor site

Background: DNA vaccines against cancer held great promises due to the generation of a specific and long lasting immune response. However, when used as a single therapy, they are not able to drive the generated immune response into the tumor, because of the immunosuppressive microenvironment, thus limiting their use in humans. To enhance DNA vaccine efficacy, we combined a new poly-epitope DNA vaccine encoding melanoma tumor associated antigens and B16F1-specific neoantigens with an oncolytic virus administered intratumorally. Methods: Genomic analysis were performed to find specific mutations in B16F1 melanoma cells. The antigen gene sequences were designed according to these mutations prior to the insertion in the plasmid vector. Mice were injected with B16F1 tumor cells (n = 7-9) and therapeutically vaccinated 2, 9 and 16 days after the tumor injection. The virus was administered intratumorally at day 10, 12 and 14. Immune cell infiltration analysis and cytokine production were performed by flow cytometry, PCR and ELISPOT in the tumor site and in the spleen of animals, 17 days after the tumor injection. Results: The combination of DNA vaccine and oncolytic virus significantly increased the immune activity into the tumor. In particular, the local intratumoral viral therapy increased the NK infiltration, thus increasing the production of different cytokines, chemokines and enzymes involved in the adaptive immune system recruitment and cytotoxic activity. On the other side, the DNA vaccine generated antigen-specific T cells in the spleen, which migrated into the tumor when recalled by the local viral therapy. The complementarity between these strategies explains the dramatic tumor regression observed only in the combination group compared to all the other control groups. Conclusions: This study explores the immunological mechanism of the combination between an oncolytic adenovirus and a DNA vaccine against melanoma. It demonstrates that the use of a rational combination therapy involving DNA vaccination could overcome its poor immunogenicity. In this way, it will be possible to exploit the great potential of DNA vaccination, thus allowing a larger use in the clinic.Peer reviewe

Peptides-Coated Oncolytic Vaccines for Cancer Personalized Medicine

Publisher Copyright: Copyright © 2022 Feola, Russo, Martins, Lopes, Vandermeulen, Fluhler, De Giorgi, Fusciello, Pesonen, Ylösmäki, Antignani, Chiaro, Hamdan, Feodoroff, Grönholm and Cerullo.Oncolytic Viruses (OVs) work through two main mechanisms of action: the direct lysis of the virus-infected cancer cells and the release of tumor antigens as a result of the viral burst. In this sc.enario, the OVs act as in situ cancer vaccines, since the immunogenicity of the virus is combined with tumor antigens, that direct the specificity of the anti-tumor adaptive immune response. However, this mechanism in some cases fails in eliciting a strong specific T cell response. One way to overcome this problem and enhance the priming efficiency is the production of genetically modified oncolytic viruses encoding one or more tumor antigens. To avoid the long and expensive process related to the engineering of the OVs, we have exploited an approach based on coating OVs (adenovirus and vaccinia virus) with tumor antigens. In this work, oncolytic viruses encoding tumor antigens and tumor antigen decorated adenoviral platform (PeptiCRAd) have been used as cancer vaccines and evaluated both for their prophylactic and therapeutic efficacy. We have first tested the oncolytic vaccines by exploiting the OVA model, moving then to TRP2, a more clinically relevant tumor antigen. Finally, both approaches have been investigated in tumor neo-antigens settings. Interestingly, both genetically modified oncolytic adenovirus and PeptiCRAd elicited T cells-specific anti-tumor responses. However, in vitro cross-representation experiments, showed an advantage of PeptiCRAd as regards the fast presentation of the model epitope SIINFEKL from OVA in an immunogenic rather than tolerogenic fashion. Here two approaches used as cancer oncolytic vaccines have been explored and characterized for their efficacy. Although the generation of specific anti-tumor T cells was elicited in both approaches, PeptiCRAd retains the advantage of being rapidly adaptable by coating the adenovirus with a different set of tumor antigens, which is crucial in personalized cancer vaccines clinical setting.Peer reviewe

DNA electrotransfer after intradermal injection for immunisation

The specific immunologic environment of the skin provides both innate and adaptive immunities and efficiently protects the individual from external aggressions; making the skin an attractive organ for the delivery of DNA vaccines. However, the development of such vaccines requires appropriate delivery technologies. Electrotransfer is one of the most efficient methods of in vivo non-viral gene transfer. The introduction reviews the literature and gives information about the main fields evoked in the thesis: the skin, methods for DNA transfer in the skin, electrotransfer and DNA vaccines. The aim of this work was to optimise immunisation by intradermal DNA electrotransfer. First, we optimised the delivery method itself. The effects of the injection method and the DNA dose on gene expression after intradermal DNA electrotransfer were studied. Pre-treatments were evaluated to enhance DNA diffusion in the skin. Then, DNA immunisation with a weakly immunogenic model antigen, luciferase, was investigated. The immune response after intradermal DNA electrotransfer was compared with immunisation to the luciferase recombinant protein. Finally, the influence of the delivery route was studied. Second, we tried to find adjuvants to modulate and/or enhance the immune response after intradermal DNA electrotransfer. Four potential adjuvants, acting on different targets were studied: hyaluronidase, imiquimod, monophosphoryl lipid A and tape stripping. Finally, we worked on the optimisation of the plasmid vector. We investigated if it was possible to induce immune response when only some specific cells of the skin were targeted. Specific promoters of cutaneous cells were cloned and we investigated if fascin, keratin14 or CD11c promoter plasmids were able to induce immune response. We also followed kinetic of luciferase expression after DNA electrotransfer in the muscle and in the skin of a small eukaryotic expression vector which was devoid of antibiotic resistance markers and optimised for gene therapy.Thèse de doctorat en sciences pharmaceutiques (FARM 3) --UCL, 200

Electrotransfer of DNA vaccine

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Chitosan-based siRNA delivery systems

Recently, chitosan has attracted significant attention in the formulation of small interfering RNA (siRNA). Because of its cationic nature, chitosan can easily complex siRNA, thus readily forming nanoparticles. Moreover, chitosan is biocompatible and biodegradable, which make it a good candidate for siRNA delivery in vivo. However, chitosan requires further development to achieve high efficiency. This review will describe the major barriers that impair the efficiency of the chitosan-based siRNA delivery systems, including the stability of the delivery system in biological fluids and endosomal escape. Several solutions to counteract these barriers have been developed and will be discussed. The parameters to consider for designing powerful delivery systems will be described, particularly the possibilities for grafting targeting ligands. Finally, optimized systems that allow in vivo therapeutic applications for both local and systemic delivery will be reviewed. This review will present recent improvements in chitosan-based siRNA delivery systems that overcome many of these system's previous pitfalls and pave the way to a new generation of siRNA delivery systems