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From 2013.igem.org

Recombinant expression of FluM1 from Influenza & Melan-A from Melanoma provides protein epitope for antigen recognition

The BactoVax vaccine platform has been engineered to act as a general vaccine platform, suitable for vaccination against bacteria, viruses, and tumors. Our current system has been engineered to express both viral antigen FluM1 and cancer surface antigen Melan-A to illustrate the modular nature of the platform itself. Instead of having to engineer a separate vaccine for every single different strain of a particular disease or group of disease separately, BactoVax provides a chassis that can be easily modified simply by addition of the nucleotide sequence for antigen of interest to trigger a specialized immune response.

MHC Class I Pathway Presentation Depends Upon Pathogenic Characteristics

The immune system has two separate pathways within antigen-presenting immune cells (APCs) that trigger immune responses. The Major Histocompatibility Complex (MHC) Class I Pathway activated CD8, or Cytotoxic, T lymphocytes. The MHC Class I protein complex is present on all human cells and is the complex that changes during tumor development. Activated of CD8 T cells with an antigen present on an MHC Class I Protein Complex allows them to selectively target tumor cells expressing that same antigen on their surface.

However, when a dendritic cell or macrophage engulfs a bacterium or virus, the foreign material is degraded within a phagolysosome and then presented directly onto MHC Class II protein complexes. MHC Class II complexes only activate B cells, which produce antibodies. However, these antibodies are not sufficient to launch an anti-tumor response because they cannot recognize the antigens present on the MHC Class I proteins on tumor cells to distinguish them from healthy cells. This is because antibodies are not familiar with the MHC Class I protein complex; they were activated by MHC Class II protein complexes.

Listeria monogenocytes, a foodborne pathogen, has been explored as a cancer vaccine delivery vector because it contains a protein, Listeriolysin O, that allows it, along with its toxin proteins, to escape the phagolysosome of dendritic cells and macrophages and live within the cytoplasm of immune cells. Proteins and antigens present in the cytoplasm instead of the phagolysosome can be presented on MHC Class I pathway over the MHC Class II pathway. However, because of the inherent pathogenicity of Listeria monogenocytes from other toxins, such as internalin, it is not the most suitable cancer vaccine chassis for translational applications.

Expression of Lysteriolysin in E.coli Nissile can confer difference in MHC presentation

We plan to introduce the listeriolysin O (LLO) gene into lab safe strains of E. coli to provide a safer chassis for vaccine delivery with the same effectiveness as current Listeria vectors. Phagolysosome lysis, induced by LLO after the E. coli membrane is degraded inside a dendritic cell, is necessary for our cancer antigens can be released from the E. coli chassis into the macrophage cytoplasm, which results in presentation on MHC Class I complexes and subsequent cytotoxic T cell activation, needed for destroy tumor cells. B cells produce antibodies, but they will not be able to recruit cytotoxic T cells to tumor cells without the previous existence of T cells that recognize tumor antigens. MHC Class I activation is the only method to activate cytotoxic T cells to recognize antigens on the surface of tumor cells. We are, however, looking into safer alternatives to LLO, such as ubiquitin, to replace LLO in future iterations of the vaccine system as well as determine methods to activate both MHC pathways to provide a joint immune response.