Week 13: Poster Session

Submitted by sbrewer on Wed, 04/26/2017 - 08:54

Meeting Location

BCRC (M311S)

Overview

On the last class of the day, we will conduct a poster session.  Your poster should be printed before the beginning of class. Note that we will print your poster for free -- you do not need to pay for it yourself.

I will be available in the BCRC on Thursday, April 27 by ~5pm and will stay as long as anyone needs help getting their poster finished and printed.  (In point of fact, I will be around most of time during the week and will be happy to help, so feel free to drop in and work and get me if you have questions.)

Required Activities

  1. Complete data collection and analysis
  2. Complete your Research Poster prior to the beginning of class

 

    PP: AI Beats Out Supreme Court Experts

    Submitted by eriklee on Fri, 05/05/2017 - 01:07

    Researchers designed a new algorithm that predicts Supreme Court decisions with pinpoint accuracy. The algorithm draws from data from the Supreme Court Database, which contains records dating back to 1791. The algorithm compiles information between 1816 and 2015 and analyzes factors such as judges, votes, term, issues, and court circuits. To test the algorithm, it sequentially goes through the years, starting at 1816, and tries to make a prediction based on the previous year’s data. The algorithm was able to predict the results with 70.2% accuracy. Well qualified experts have 66% accuracy. This can be big as court decision predictions can help the plaintiffs chose which courts to appeal, companies can choose how to handle cases, lawyers can determine how to proceed with the case, and appellants in high profile cases can decide whether to pursue the case. The researchers hope to factor in expert and public statistics and opinions to make the predictions more precise. 

    Deep-Stack Can Beat World Class Poker Players

    Submitted by eriklee on Fri, 05/05/2017 - 01:06

    Already, algorithms, which have surpassed human skill, have been developed for games such as chess, checkers, go, backgammon, and more. A team of researchers seek to develop an algorithm that can beat world class poker players in a game that is termed a symmetric. That game is heads-up no-limit Texas hold’em. This game combines a public state of probability between the hands of each player with the strategy of wit and deception. Previous iterations of algorithms have tried and failed to challenge players at major tournaments. The previous renditions used an abstract set of states to predict the how the entire game will unfold. They do this by condensing the available possible states to a fraction of options. However, the loss of information has set these algorithms to fail. A new algorithm, known as DeepStack, is lauded to have the ability to beat these algorithms and players. It uses a modified version of learning and combines past experience with reason and probabilistic states. In essence, DeepStack is beginning to model a human mind, and its success shows that these techniques can be effective in predicting asymmetric situations outside of games like poker.

    Link to Article: http://science.sciencemag.org/content/356/6337/508.full

    Explaining T cell activation and its pathway

    Submitted by abnguyen on Thu, 05/04/2017 - 23:46

     

    The affinity of T-cell receptors for MHC is usually too low to mediate a functional interaction between the two cells by itself.  In order to increase signal strength and cell to cell interactions, accessory receptors are required.  Unlike T-cell receptors or MHC proteins, accessory receptors do not bind foreign antigens and are invariant.  The accessory receptors that have a direct role in activating T-cells are called co-receptors.  The most understood co-receptors are CD4 and CD8.  Both proteins are single-pass transmembrane protein and recognize MHC, however, unlike T-cell receptors, CD4 and CD8 bind to non variable parts of the protein. The antibodies to CD4 and CD8 are used to help distinguish between the 2 main classes of T cells. (Smith-Garvin et al, 2017)

        Signaling through TCR alone results in a non responsive state (anergy).  Additional binding to other surface receptors are needed to enhance TCR signals.  The most used and robust receptor is the CD28 protein.  A study done on mice with CD28 knocked out showed a plethora of immune defects such as impaired T-cell activation, a lack of T-cell help for B cells and poor memory of T-cell response (Harris NL. and Ronchese F 1999).  CD28 promotes T-cell proliferation, cytokine production, cell survival, and cellular metabolism.  Many of these processes are activated when TCR binding occurs alone but the signal is not sufficient enough to pass a threshold and requires CD28 co-binding. (Alberts et al, 2002).  CD28 is located on T cells and binds with either CD80 (B7-1) or CD86 (B7-2) on APC.  CD80 and CD86 are expression is upregulated in response to inflammatory stimuli thus leading ligand upregulation to be seen as a key link between danger signals and an immune response.

        T cell activation requires multiple steps after the initial bindings of MHC to TCR and CD28 to either CD80 or CD86 as shown in figure 4. (a higher resolution image can be found at http://tinyurl.com/m6fdl99).  The final product of this extensive pathway is the production of IL-2 which leads to an immune response.  To being the process, CD28 and CD45 activates tyrosine kinases Lck and Fyn.  These 2 activations phosphorylate ZAP70, SYK, Vav1 and LAT.  LAT binds to GADS, SLP76, ITK, Vav1 and Tec which leads to the activation of PLC-γ, RLK, CARMA1, BCL10, CDC42 and Rac.  Rac activation is essential for MEKK1, MKKs and JNK activation.  JNKs phosphorylates c-Jun and c-Fos within the nucleus and will allow for IL-2 transcription.  The recruitment of these signal transduction components drive T-cell activation.

        On the left side of the pathway, CD28 binds to PI3K, recruiting it to the membrane.  PI3K activates AKT which promotes T-cell survival.  The PLC-γ protein that was activated earlier uses the PIP2 created from PI3k as a substrate to produce PI3 and DAG.  PI3 releases Ca2+ into the cytoplasm which will send NFAT into the cytoplasm.  While this process is occurring, PKC-ϴ is regulating the phosphorylation of IKK with the aid of CARMA1, BCL10 and MALT1.  IKK phosphorylation will degrade I-κBs, freeing NF-κB allowing it to travel into the nucleus.  

        Once NFAT, NF-κB, c-Jun, and c-Fos are inside of the nucleus, IL-2 will begin to be transcribed.  IL-2 transcription allows the T-cell to enter into the cell cycle, promotes cell survival, cell differentiation, and most importantly, the activation of helper T cells. (Smith-Garvin et al, 2017)

    Smith-Garvin, Jennifer E., Gary A. Koretzky, and Martha S. Jordan. “T Cell Activation.” Annual review of immunology 27 (2009): 591–619. PMC. Web. 4 May 2017.

    Harris NL. and Ronchese F.  1999 "The role of B7 costimulation in T-cell immunity".  Immunology and Cell Biology.  <http://www.nature.com/icb/journal/v77/n4/full/icb199938a.html>.  Accessed 3 May 2017.

    Alberts B, Johnson A, Lewis J, et al. Molecular Biology of the Cell. 4th edition. New York: Garland Science; 2002. T Cells and MHC Proteins.

    http://tinyurl.com/m6fdl99

     

     

    reflective essay PP

    Submitted by jdantonio on Thu, 05/04/2017 - 22:45

    Our first project which we began at the beginning of the semester and have continued through to the end is the weekly journal entry. I think when I was first given the assignment I thought this would largely be busy work, something to check off at the end of each week. However as the semester went on I think I continually got more out of it. The reading of papers to create journals entries and just the act of writing made me a better scientific writer. Seeing many examples of  scientific literature allowed me to compare and contrast styles and improve my own writing by synthesising the best parts for my journal entries. It also helped to get me to do projects in other classes in a more reasonable time frame. In the past I would typically do projects at the last second but spreading them out over several days as journal entries made me go at a slower rate and I feel helped to improve their quality. I think in the future I will spread out my work over a larger period of time which will help me improve my writing quality and be prepared for adverse events. I will also try to write more in general as I definitely feel that I have improved my writing skills simply by the act of writing regularly.  

     

    reflective essay jornal entry

    Submitted by jdantonio on Thu, 05/04/2017 - 21:19

    Our first project which we began at the beginning of the semester and have continued through to the end is the weekly journal entry. I think when I was first given the assignment I thought this would largely be busy work, something to check off at the end of each week. However as the semester went on I think I continually got more from it. The reading of papers to create journals entries and just the act of writing made me a better scientific writer seeing many examples of  scientific literature allowed me to compare and contrast styles and improve my own writing by synthesizing the best parts in my journal entries. It also helped to get me to do projects in other classes in a more reasonable time frame. In the past I would typically do projects at the last second but spreading them out with journal entries made me go at a slower rate and I feel helped to improve their quality. I think in the future I will spread out my work over a larger period of time which will help me improve quality and be prepared for adverse events. I will also try to write more as I definitely feel that I have improved my writing skills simply by the act of writing regularly.  

    cancer gentics project 3 excerpt 6

    Submitted by jdantonio on Thu, 05/04/2017 - 20:33

    Prior to the infusion of our selected and modified T-cell lines we will perform a  myeloablation, the destruction of immune cells in the body by high dose full body radiation treatment and the infusion of hematopoietic stem cells (Wrzesinski et al 2010). This pre treatment of patients has been shown to greatly improve the efficacy of ACT treatments in vivo(Perica et al 2015).  This works by reducing the presence of suppressing regulatory lymphocytes that inhibit cytotoxic T-cell activity(Wrzesinski et al 2010). We would infuse both our adaptive cell lines and our chimeric cell lines at the same time following the myeloablation and would concurrently administer IL-2 by and IV (Kochenderfer et al 2012). After the administration of the treatment we would monitor the patient and be on alert for cytokine toxicity a potential side effect of T-cell adaptive therapies caused by T-cell stimulated release of inflammatory cytokines. This can lead to fever and inflammation in the patient and can in very severe cases cause patient death (Barrett et al 2013). For dosage of the treatment, T-cell numbers are a limiting factor in the treatment. So we would infuse all the T-cells we are able to generate in a reasonable amount of time (maybe a month or two of growth), this amount would vary depending on multiple factors.

     

    Barrett DM, Singh N, Porter DL, Grupp SA, and June CH. 2013. Chimeric Antigen Receptor

    Therapy for Cancer. Annual Review 65: 333-347.

    <http://annualreviews.org/doi/full/10.1146/annurev-med-060512-150254>. Accessed

    2017 May 1.



    Kochenderfer JN, Dudley ME, Feldman SA,Wilson HW, Spaner DE, Maric I, Stetler-Stevenson

    M, Phan GQ, Hughes MS, Sherry MR, Yang JC, Kammula US, Devillier L, Carpenter R,

    Nathan DAN, Morgan RA, Laurencot C,and Rosenberg SA. 2012. B-cell depletion and

    remissions of malignancy along with cytokine-associated toxicity in a clinical trial of

    anti-CD19 chimeric-antigen-receptor-transduced T cells. Blood 119(12):2709-20.

    <http://www.bloodjournal.org/content/bloodjournal/119/12/2709.full.pdf>. Accessed

    2017 May 1.

     

    Perica K, Varela JC, Oelke M, and Schneck J. 2015. Adoptive T Cell Immunotherapy for Cancer.

    Rambam Medical Journal 6(1): e0004. National Center of Biotechnology

    Information[NCBI]. <https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4327320/>.

    Accessed 2017 April 29.

     

    Wrzesinski C, Paulos CM, Kaiser A, Muranski P, Palmer DC, Gattinoni L, Yu Z, Rosenberg SA,

    and Restifo NP. 2010. Increased intensity lymphodepletion enhances tumor treatment

    efficacy of adoptively transferred tumor-specific T cells. Journal of Immunotherapy

    33(1): 1-7. National Center of Biotechnology Information[NCBI].

    cancer gentics project 3 excerpt 5

    Submitted by jdantonio on Thu, 05/04/2017 - 18:59

    We will augment the endogenous TIL lines by the creation of additional cell lines that have genetically engineered chimeric antigen receptors (CAR). These chimeric receptor will be designed to bind to one of the remaining 10 most prevalent neoantigens that we identified as previously described. We will create ten cell lines, each being given one of chimeric receptors, creating ten cell lines which each bind to a different neoantigen. We will introduce a gene which encodes for a genetically engineered T-cell receptor by a viral vector in vitro. The CTR will be  created by the fusion of the Vh and VL gene sequence segments of a monoclonal antibody that binds to our neoantigen, a hinge-Ch2-Ch3 IgG1 , a transmembrane CD4 domain, and zeta chain of the CD3 cytoplasmic domain (Park et al 2007). This creates a chimeric TCR that recognises one of our discovered neoantigen and can activate T-cell activation (Park et al 2007). The monoclonal antibodies that bind to our neoantigens will be created by using a generalized method of immunization of rodent models with our neoantigens, the rodents will create antibodies against the neoantigen and they will be harvested and screened in a manner similar to the adoptive cells for bind specificity to the neoantigen (Gerdes et al 1983). These antibody are then used to make the gene fragment that is placed in the chimeric vector. We would perform this procedure for the 10 most common neoantigens for our tumor that were not found to have endogenous TIL’s during the adoptive cell therapy. The chimeric gene would be delivered to the cells in vitro by an IVT (in vitro transcription) RNA vector which is one of the standard delivery systems for CARs (Zhao et al 2010). Once the viral vector has been incorporated into the T-Cells we would then modify them further by using CRISPR to knockout the PDL-1 and CTLA-4 receptors on the cells(Albain et al 2015).

     

    Ablain J, Durand EM, Yang S,  Zhou Y,  Zon LI. 2015. A CRISPR/Cas9 Vector System for

    Tissue-Specific Gene Disruption in Zebrafish. Developmental Cell 32(6): 756–764.

    Sciencedirect.<http://www.sciencedirect.com/science/article/pii/S1534580715000751>.

    Accessed 2017 May 1.

    Gerdes J, Schwab U, Lemke H, Stein H. 1983. Production of a mouse monoclonal antibody

    reactive with a human nuclear antigen associated with cell proliferation. International

    Journal of Cancer 31(1): 13-20. Wiley Online Library. <http://onlinelibrary.wiley.com/doi/10.1002/ijc.2910310104/full>. Accessed 2017 May 1.

    Park JR, DiGiusto DL, Slovak M, Wright C, Naranjo A, Wagner J, Meechoovet HB, Bautista C, Chang WC, Ostberg JR, Jensen MC. 2007. Adoptive Transfer of Chimeric Antigen Receptor Re-directed Cytolytic T Lymphocyte Clones in Patients with Neuroblastoma. Molecular Therapy 15(4):  825–833. Science Direct. <http://www.sciencedirect.com/science/article/pii/S152500161631351X>. Accessed 2017 May 1.

     

    Zhao Y, Moon E, Carpenito C, Paulos CM, Liu X, Brennan AL, Chew A, Carroll RG, Scholler J,

    Levine BL, Albelda SM, June CH. 2010. Multiple injections of electroporated autologous

    T cells expressing a chimeric antigen receptor mediate regression of human disseminated

    tumor. Cancer Research

    70(22):9053-906.<https://www.ncbi.nlm.nih.gov/pubmed/20926399>. Accessed 2017

    May 1.

    cancer gentics project 3 excerpt 4

    Submitted by jdantonio on Thu, 05/04/2017 - 17:47

    For the selection of our T-cells that possess endogenous neoantigen receptors we first harvest T-cells by taking blood samples from our patient. The Cells would then be separated from the plasma and the neoantigen cytotoxic T-cells would be selected out by using autologous DC cells that have been loaded with the protein epitome of the neoantigens which we have identified (Yee et al 2002). We will be following the standard procedure for adoptive T-cell generation as described in Yee et al 1999(Yee et al 1999). In short the T-cells are separated and cloned, creating identical clone lines, then screened by testing them for their ability to bind to antigen loaded dendritic cells which are placed in a set of loaded wells for testing. The population of T-cells that are found to bind to the neoantigen are then put through a chromium lystic test. where in the TILs are put on a plate with harvested tumor cells that present the target neoantigen and have been loaded with chromium, the lysing of these cells causes the release of chromium and can be used to test whether the antigen binding T-cells are cytotoxic (Yee et al 2002).  The antigen binding cytotoxic  T-cells will then be raised in culture containing IL-2 and anti CD3-antibodies which activated the growth and proliferation of T-cells, this will be done in 14 day cycles.(Yee et al 2002) We will perform theses screening tests to find a population of cells that can bind to the neoantigens we experimentally derived, we would hope to find cell lines cable of binding to about 10 of the 20 most common of our neoantigens.We will also give preference to those T-cells capable of binding neoantigens caused by driver mutations, mutations that are critical to cancer progression(Stratton et al 2009). With this we will have 10,or more hopefully, T-cell lines that can bind to our tumor and initiate cell death. The use of multiple adoptive cell lines will help to prevent the cancer from become resistant to our treatment, as one single mutation, or even a few, would not be enough for the tumor to evade treatment. Finally we will further the effectiveness by knocking out the T-cell suppression CTLA-4 receptor and the T-cell death initiating PDL-1 receptor by CRISPR knockout as described in detail later in the project (Albain et al 2015).

     

    Ablain J, Durand EM, Yang S,  Zhou Y,  Zon LI. 2015. A CRISPR/Cas9 Vector System for

    Tissue-Specific Gene Disruption in Zebrafish. Developmental Cell 32(6): 756–764.

    Sciencedirect.<http://www.sciencedirect.com/science/article/pii/S1534580715000751>.

    Accessed 2017 May 1.

     

    Stratton MR,Campbell PJ, Futreal PA. 2009. The cancer genome. Nature 458: 719-724.

    <http://www.nature.com/nature/journal/v458/n7239/full/nature07943.html>. Accessed

    2017 may 1.

     

    Yee C, Thompson JA, Byrd D, Riddell SR, Roche P, Celis E, and Greenberg PD. 2002.Adoptive

    T cell therapy using antigen-specific CD8+ T cell clones for the treatment of patients

    with metastatic melanoma: In vivo persistence, migration, and antitumor effect of

    transferred T cells. Proceedings of the National Academy of Sciences 99(25):

    16168–16173. <http://www.pnas.org/content/99/25/16168.full.pdf> Accessed 2017 April

    30



    Yee C,  Savage PA,  Lee PP, Davis MM, and Greenberg PD. 1999.Isolation of High Avidity

    Melanoma-Reactive CTL from Heterogeneous Populations Using Peptide-MHC

    Tetramers. The Journal of Immunology 162 (4) 2227-2234. <http://

    www.jimmunol.org/content/162/4/2227?ijkey=5e73920a74aaa8e458988ce6c5acc1d1f2f9

    ef72&keytype2=tf_ipsecsha>. Accessed 2017 April 30.

    cancer gentics project 3 excerpt 3

    Submitted by jdantonio on Thu, 05/04/2017 - 16:00

    T-cells play a key role as a part of the immune system. They identify pathogenic entities by recognition of non-self antigens on the damaged/infected/foreign cells (Neumann et al 2002). T-cells are able to destroy cells that possess an antigen that they are cable of binding to by three main pathways.(neumann et al. 2002). The first method of lysis for cytotoxic T-cells in the release of cytotoxic granules such as perforin, which break down the target cell's membrane causing death(Neumann et al. 2002). The second is by the activation of the RAS receptor pathway, T-cells can have the ligand for this receptor (CD95L) which activates the apoptosis pathway in the target cell (Cox and Der 2003). The third method that cytotoxic T-cells use to kill their target cells is the release of cytokines such as TNF-α, which can induce apoptotic pathways within the cell (Neumann et al 2002). Our T-cell will use a combination of these pathways to effectively kill tumor cells that present neoantigens that the T-cells bind to.

     

    Cox AD and CJ Der. 2002. The dark side of Ras: regulation of apoptosis. Oncogene 22:

    8999–9006.<http://www.nature.com/onc/journal/v22/n56/full/1207111a.html>. Accessed

     

    Neumann H, Medana IM, Bauer J, Lassmann H. 2003. Cytotoxic T lymphocytes in autoimmune

    and degenerative CNS diseases. Trends in Neuroscience 25(6): 313-319.

    <http://www.cell.com/trends/neurosciences/fulltext/S0166-2236(02)02154-9>. Accessed

    2017 May 3.

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