Difference between revisions of "Team:CGU Taiwan/Design"

 
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{{CGU_Taiwan}}
 
 
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<h3>★  ALERT! </h3>
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<p>This page is used by the judges to evaluate your team for the <a href="https://2016.igem.org/Judging/Awards#Special_Prizes"> design special prize</a>. </p>
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<p> Delete this box in order to be evaluated for this medal. See more information at <a href="https://2016.igem.org/Judging/Pages_for_Awards/Instructions"> Instructions for Pages for awards</a>.</p>
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<p>
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By talking about your design work on this page, there is one medal criterion that you can attempt to meet, and one award that you can apply for. If your team is going for a gold medal by building a functional prototype, you should tell us what you did on this page.
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<p>This is a prize for the team that has developed a synthetic biology product to solve a real world problem in the most elegant way. The students will have considered how well the product addresses the problem versus other potential solutions, how the product integrates or disrupts other products and processes, and how its lifecycle can more broadly impact our lives and environments in positive and negative ways.</p>
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If you are working on art and design as your main project, please join the art and design track. If you are integrating art and design into the core of your main project, please apply for the award by completing this page.
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<p>Teams who want to focus on art and design should be in the art and design special track. If you want to have a sub-project in this area, you should compete for this award.</p>
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<div class="top">
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<img style="margin-top:-2%;width:100%;height:140%;position:absolute;" src="https://static.igem.org/mediawiki/2016/0/0b/CGU_Taiwan--top.jpg">
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</div>
 
</div>
 
</div>
  
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<header class="top">
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<ul class="header-subnav">
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<li><a href="https://2016.igem.org/Team:CGU_Taiwan">HOME</a></li>
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<li><a href="https://2016.igem.org/Team:CGU_Taiwan/Achievements">ACHIEVEMENTS</a></li>
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<li><a href="https://2016.igem.org/Team:CGU_Taiwan/Description">PROJECT</a></li>
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<li><a href="https://2016.igem.org/Team:CGU_Taiwan/Software">MODELING</a></li>
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<li><a href="https://2016.igem.org/Team:CGU_Taiwan/Human_Practices">HUMAN PRACTICES</a></li>
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<li><a href="https://2016.igem.org/Team:CGU_Taiwan/Team">PEOPLE</a></li>
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<li class="dropdown"><a href="https://2016.igem.org/Team:CGU_Taiwan/Interlab">INTERLAB</a></li>
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<li class="dropdown"><a href="https://2016.igem.org/Team:CGU_Taiwan/Safety">SAFETY</a></li>
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<li><a href="https://2016.igem.org/Team:CGU_Taiwan/Parts">PARTS</a></li>
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</ul>
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<div class="circle"><img style="position:absolute;z-index:+1;width:90%;height:95%;margin-left:3%;margin-top:2%;" src="https://static.igem.org/mediawiki/2016/8/8a/CGU_Taiwan--logo9.jpg"></div>
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</header>
  
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<div class="left">
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<a href="https://2016.igem.org/Team:CGU_Taiwan/Description"><section class="left-tab" style="top:240px;" id="myP" onmousedown="mouseDown()" onmouseup="mouseUp()"><b>TOP</b></section></a>
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<a href="https://2016.igem.org/Team:CGU_Taiwan/Description"><section class="left-tab" style="top:275px;left:50px;">Overview</section></a>
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<a href="https://2016.igem.org/Team:CGU_Taiwan/Design"><section class="left-tab" style="top:310px;left:50px;">Design</section></a>
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<a href="https://2016.igem.org/Team:CGU_Taiwan/Background"><section class="left-tab" style="top:345px;left:50px;">Background</section></a>
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<a href="https://2016.igem.org/Team:CGU_Taiwan/Results"><section class="left-tab" style="top:380px;left:50px;">Results</section></a>
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<a href="https://2016.igem.org/Team:CGU_Taiwan/Proof"><section class="left-tab" style="top:415px;left:50px;">Proof</section></a>
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<a href="https://2016.igem.org/Team:CGU_Taiwan/Notebook"><section class="left-tab" style="top:450px;left:50px;">Notebook</section></a>
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</div>
  
  
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<br><br><br>
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<div class="mid">
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<div style="font-size:60px;color:#FF8800;text-decoration:none;margin-left:22%;margin-top:20%;">
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Design
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</div>
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<br><br><br>
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<b style="color:#0D014D;font-size:28px;margin-left:22%;">Project Design</b>
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<br><br>
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<div style="color:black;font-size:18px;margin-left:22%;margin-right:5%;text-align:justify;text-decoration:none;">
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Leijuvant is a whole new kind of adjuvant that uses Leishmania as an effective T cell stimulator. Leishmania is a parasite that specifically lives within macrophage, a professional antigen presenting cell (APC). As a potential vaccine adjuvant, Leishmania possess many advantages, including APC recruitment, pattern recognition receptor (PRR) activation, inflammasome activation, activation of MHC-presenting pathway and most important of all, T cell activation<sup>[1],[2],[3]</sup>. We adapted a combinational approach by loading the Leishmania DT mutants both endogenously and exogenously with different chemicals. After the chemicals are illuminated by specific wavelength of light, double-photo inactivation will kill Leishmania thoroughly. Thus, transgenic mutant of Leishmania that can be inactivated by light exposure acts as a safe carrier to deliver specific antigens to APCs<sup>[4]</sup>.
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</div>
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<br><br>
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<img src="https://static.igem.org/mediawiki/2016/2/25/CGU_Taiwan--design1.jpg" style="float:left;width:40%;height:160%;margin-left:22%;margin-right:7%;">
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<div style="color:black;font-size:18px;margin-left:22%;margin-right:5%;text-align:justify;text-decoration:none;">
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Photo-inactivated Leishmania has been proven to activate CD8 and CD4 T cells<sup>[5]</sup>. However, further activation of B cells has not been confirmed. As a vaccine adjuvant, persistent antibody production against specific antigen is an essential parameter in evaluating vaccine efficacy. B cell needs the stimulation from both the intact antigen and antigen-specific CD4 T cell simultaneously in order to differentiate into plasma cell in producing antibodies.Therefore, we hypothesized that genetically engineered Leishmania that express specific antigen can serve as an adjuvant that deliver antigens into APCs.
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<br><br>
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The photo-inactivated Leishmania will be engulfed by APCs and activate T cells via MHC molecules. With the company of intact vaccine antigen, they can stimulate B cell form both ways and induce antibody production.
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</div>
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<br><br>
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<b style="color:#0D014D;font-size:28px;margin-left:22%;">Experimental design</b>
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<br><br>
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<b style="color:#0D014D;font-size:20px;margin-left:22%;">Our experiments can be divided into four main parts:</b>
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<br>
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<div style="color:black;font-size:18px;margin-left:24%;margin-right:5%;text-align:justify;text-decoration:none;">
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<ol>
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<li>Ecoli-Leishmania shuttle vector-the Leishmania antigen expression system</li>
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<li>In vivo test for validation of our concept of LEIJUVANT</li>
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<li>In vitro test for proving our software prediction system, McHug, and our concept</li>
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<li>As for the social experiments, we designed questionnaires to study the public's general knowledge level on vaccines</li></ol>
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<br><br>
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</div>
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<img src="https://static.igem.org/mediawiki/2016/d/d8/CGU_Taiwan--design2.jpg" style="width:50%;height:200%;margin-left:35%;">
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<br><br>
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<div style="color:black;font-size:18px;margin-left:22%;margin-right:5%;text-align:justify;text-decoration:none;">
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In order to use Leishmania to deliver antigen into APCs to further activate immune response, we designed an E. coli-Leishmania shuttle vector to carry the antigen sequence into Leishmania through electroporation. Successfully tranfected Leishmania was isolated by drug selection and then subjected to double-photo inactivation. After insuring the death of Leishmania, we tested its effects on antibody production and T cell response through in vivo experiments. We co-injected Leijuvant with OVA protein subcutaneously into C57BL/6 mice. We also performed in vitro experiments to identify antigen peptide sequences binding to MHC molecules by mass spectrometry. It will validate the accuracy of our McHug prediction system.
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</div>
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<br><br>
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<img src="https://static.igem.org/mediawiki/2016/f/fa/CGU_Taiwan--design3.jpg" style="width:50%;height:180%;margin-left:35%;">
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<br><br>
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<div style="color:black;font-size:18px;margin-left:22%;margin-right:5%;text-align:justify;text-decoration:none;">
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To launch a medical product, it must go through multiple processes including R&D, scale-up production, risk-benefit assessment, and public perception survey. Thus, we visited professionals in different divisions of vaccine industries to learn more details about vaccine research and development as well as their opinions on our project. We also carried out a survey to acquire public perception toward our product and their knowledge on vaccines.
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<br><br><br><br>
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<div style="font-size:12px;">
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References:<br>
 +
1. Lima-Junior, D.S., et al., Inflammasome-derived IL-1[beta] production induces nitric oxide-mediated resistance to Leishmania. Nat Med, 2013. 19(7): p. 909-915.<br>
 +
2. Srivastava, S., et al., Leishmania expressed lipophosphoglycan interacts with Toll-like receptor (TLR)-2 to decrease TLR-9 expression and reduce anti-leishmanial responses. Clin Exp Immunol, 2013. 172(3): p. 403-9.<br>
 +
3. Lang, T., et al., Distribution of MHC class I and of MHC class II molecules in macrophages infected with Leishmania amazonensis. J Cell Sci, 1994. 107 ( Pt 1): p. 69-82.<br>
 +
4. Dutta, S., K. Waki, and K.P. Chang, Combinational sensitization of Leishmania with uroporphyrin and aluminum phthalocyanine synergistically enhances their photodynamic inactivation in vitro and in vivo. Photochem Photobiol, 2012. 88(3): p. 620-5.<br>
 +
5. Dutta, S., et al., Intracellular targeting specificity of novel phthalocyanines assessed in a host-parasite model for developing potential photodynamic medicine. PLoS One, 2011. 6(6): p. e20786.<br>
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</div>
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</div>
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<br><br>
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</div>
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<div class="end2" style="margin-top:230%;">
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<br>
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<ul class="list3" class="margin-left:1%;margin-top:2%;">
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<li><a href="#anchor1"><b style="color:white;font-size:28px;margin-left:7%;">TOP</a></li>
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</ul>
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<br>
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<div class="sitemap" style="margin-left:5%;">
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Latest revision as of 06:43, 2 December 2016

TOP
Overview
Design
Background
Results
Proof
Notebook



Design



Project Design

Leijuvant is a whole new kind of adjuvant that uses Leishmania as an effective T cell stimulator. Leishmania is a parasite that specifically lives within macrophage, a professional antigen presenting cell (APC). As a potential vaccine adjuvant, Leishmania possess many advantages, including APC recruitment, pattern recognition receptor (PRR) activation, inflammasome activation, activation of MHC-presenting pathway and most important of all, T cell activation[1],[2],[3]. We adapted a combinational approach by loading the Leishmania DT mutants both endogenously and exogenously with different chemicals. After the chemicals are illuminated by specific wavelength of light, double-photo inactivation will kill Leishmania thoroughly. Thus, transgenic mutant of Leishmania that can be inactivated by light exposure acts as a safe carrier to deliver specific antigens to APCs[4].


Photo-inactivated Leishmania has been proven to activate CD8 and CD4 T cells[5]. However, further activation of B cells has not been confirmed. As a vaccine adjuvant, persistent antibody production against specific antigen is an essential parameter in evaluating vaccine efficacy. B cell needs the stimulation from both the intact antigen and antigen-specific CD4 T cell simultaneously in order to differentiate into plasma cell in producing antibodies.Therefore, we hypothesized that genetically engineered Leishmania that express specific antigen can serve as an adjuvant that deliver antigens into APCs.

The photo-inactivated Leishmania will be engulfed by APCs and activate T cells via MHC molecules. With the company of intact vaccine antigen, they can stimulate B cell form both ways and induce antibody production.


Experimental design

Our experiments can be divided into four main parts:
  1. Ecoli-Leishmania shuttle vector-the Leishmania antigen expression system
  2. In vivo test for validation of our concept of LEIJUVANT
  3. In vitro test for proving our software prediction system, McHug, and our concept
  4. As for the social experiments, we designed questionnaires to study the public's general knowledge level on vaccines




In order to use Leishmania to deliver antigen into APCs to further activate immune response, we designed an E. coli-Leishmania shuttle vector to carry the antigen sequence into Leishmania through electroporation. Successfully tranfected Leishmania was isolated by drug selection and then subjected to double-photo inactivation. After insuring the death of Leishmania, we tested its effects on antibody production and T cell response through in vivo experiments. We co-injected Leijuvant with OVA protein subcutaneously into C57BL/6 mice. We also performed in vitro experiments to identify antigen peptide sequences binding to MHC molecules by mass spectrometry. It will validate the accuracy of our McHug prediction system.




To launch a medical product, it must go through multiple processes including R&D, scale-up production, risk-benefit assessment, and public perception survey. Thus, we visited professionals in different divisions of vaccine industries to learn more details about vaccine research and development as well as their opinions on our project. We also carried out a survey to acquire public perception toward our product and their knowledge on vaccines.



References:
1. Lima-Junior, D.S., et al., Inflammasome-derived IL-1[beta] production induces nitric oxide-mediated resistance to Leishmania. Nat Med, 2013. 19(7): p. 909-915.
2. Srivastava, S., et al., Leishmania expressed lipophosphoglycan interacts with Toll-like receptor (TLR)-2 to decrease TLR-9 expression and reduce anti-leishmanial responses. Clin Exp Immunol, 2013. 172(3): p. 403-9.
3. Lang, T., et al., Distribution of MHC class I and of MHC class II molecules in macrophages infected with Leishmania amazonensis. J Cell Sci, 1994. 107 ( Pt 1): p. 69-82.
4. Dutta, S., K. Waki, and K.P. Chang, Combinational sensitization of Leishmania with uroporphyrin and aluminum phthalocyanine synergistically enhances their photodynamic inactivation in vitro and in vivo. Photochem Photobiol, 2012. 88(3): p. 620-5.
5. Dutta, S., et al., Intracellular targeting specificity of novel phthalocyanines assessed in a host-parasite model for developing potential photodynamic medicine. PLoS One, 2011. 6(6): p. e20786.





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