Difference between revisions of "Team:Evry/Design"

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<h1>In vitro and in vivo proof of concept</h1>
  
<p class="text-justify"> Next, we cloned the full fusion protein OVA1-DEC205-HAtag inside our yeast with AGA1P. It was correctly displayed on yeast membrane (figure 4). HA-tag was fused at the end C-terminal of the protein to ensure complete translation upon detection. In both experiment (figure 2), the yeast was transformed to express AGA2P-OVA1-DEC205-HAtag and labelled with antibody anti-HA conjugated to the fluorochrome emitting at 650 nm.</p>
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After creating software prediction of tumor antigen and a functional surface displaying system, we intend to deliver the predicted antigen to the immune system. As a proof of concept, we tested in vitro on dendritic cells and macrophages extracted from mice spleen the yeast displaying system. </p>
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<p class="text-justify">Then, we performed an in vivo assay on melanoma bearing mice. The melanoma chosen was the very aggressive cell line B16-OVA. Our yeast chassis was injected after the large tumoral development of melanoma in order to assess immune induction in a true clinical context with patients presenting advance cancer. Now it's time to turn to the results.</p>
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<p class="text-justify"> We cloned the full fusion protein OVA1-DEC205-HAtag inside our yeast with AGA1P. It was correctly displayed on yeast membrane (figure 4). HA-tag was fused at the end C-terminal of the protein to ensure complete translation upon detection. In both experiment (figure 2), the yeast was transformed to express AGA2P-OVA1-DEC205-HAtag and labelled with antibody anti-HA conjugated to the fluorochrome emitting at 650 nm.</p>
  
 
<img border="0" class='img-responsive' src="https://static.igem.org/mediawiki/2015/8/80/Microscopieresults.jpg" alt="" />
 
<img border="0" class='img-responsive' src="https://static.igem.org/mediawiki/2015/8/80/Microscopieresults.jpg" alt="" />
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<img border="0" class='img-responsive' src="https://static.igem.org/mediawiki/2015/thumb/9/94/TTM_CD8%2B_OVA1.jpg/800px-TTM_CD8%2B_OVA1.jpg" alt="" />
 
<img border="0" class='img-responsive' src="https://static.igem.org/mediawiki/2015/thumb/9/94/TTM_CD8%2B_OVA1.jpg/800px-TTM_CD8%2B_OVA1.jpg" alt="" />
 
<p class="text-justify"><strong> Figure 10: Tetramer assay for CD8+ OVA1 specific T cells extracted from blood on melanoma mice C57BLC/6.</strong> Mice were sacrifice 18 days after tumor challenge with B16-OVA melanoma cell line. Mice received 3 injections of 2.10^7 yeasts.</p>
 
<p class="text-justify"><strong> Figure 10: Tetramer assay for CD8+ OVA1 specific T cells extracted from blood on melanoma mice C57BLC/6.</strong> Mice were sacrifice 18 days after tumor challenge with B16-OVA melanoma cell line. Mice received 3 injections of 2.10^7 yeasts.</p>
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<h3>Conclusion</h3>
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<p class="text-justify">We have designed a new chassis that was tested in vivo to induce a strong immune response against tumor antigen. In combination with our software predicting the best tumor antigen from the patient sequencing data. This chassis establishes a new paradigm. Instead of adapting the patient to the therapies available, we can adapt the therapy to the patient. Our chassis is scalable because it targets immune cells in vivo, making a personalized medicine a reality. In addition of tumor antigen delivery system, we have developed a recombinant encapsulated yeast to break the tolerogenic tumor environment. Finally, a yeast bio-sensor could detect and target hypoxic resistant cancer cells and prevent further metastasis.</p>
  
 
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<p class="text-justify"> 32. Bookman MA, Swerdlow R, Matis LA: Adoptive chemoimmunotherapy of murine leukemia with helper T lymphocyte clones. J Immunol 1987, 139:3166-3170.</p>
 
<p class="text-justify"> 32. Bookman MA, Swerdlow R, Matis LA: Adoptive chemoimmunotherapy of murine leukemia with helper T lymphocyte clones. J Immunol 1987, 139:3166-3170.</p>
 
  <p class="text-justify">33. Johnsen G & Elsayed S, Antigenic and allergenic determinants of ovalbumin-111. MHC Ia-binding peptide (OA 323-339) interacts with human and rabbit specific antibodies, 1990, Mol Immunol. 27821-7.</p>
 
  <p class="text-justify">33. Johnsen G & Elsayed S, Antigenic and allergenic determinants of ovalbumin-111. MHC Ia-binding peptide (OA 323-339) interacts with human and rabbit specific antibodies, 1990, Mol Immunol. 27821-7.</p>
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Revision as of 03:50, 19 September 2015

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