Characterization of a Feline Model of Factor XII Deficiency: The In Vivo Role of FXII in Vascular Injury and Inflammatory Responses

For nearly 6 decades scientists have been identifying and defining the many roles of the blood serine protease Factor XII (FXII) in vitro and in vivo. Each year we gain new information about this elusive protein, however conflicts still exist over the importance of FXII in human disease states. Factor XII participates in coagulation, fibrinolysis, renin angiotensin, and immune system responses making it an integral part of mammalian life. Animal models offer the best way to study the importance of a factor in vivo, however only two readily available FXII deficient models exist, i.e. the gene-manipulated mouse and our cats. This work characterizes the origin of the genetic defect causing FXII deficiency in a colony of domestic cats, and investigates in vivo acute responses to intravascular and intradermal (extravascular) injury. Ultimately we aim to provide a well defined animal model for therapeutic targeting strategies, and gain new insights into the importance of FXII in cellular and extracellular interactions. Chapter 1 provides a comprehensive overview of the biological features of FXII: its importance as a central player in coagulation, fibrinolysis, complement activation, the innate immune system, and vascular repair; its molecular and genetic structure and cellular interactions; and the attributed pathophysiology derived from human and mouse studies. Chapter 2 details the feline FXII gene and the mutation discovered in our colony along with protein studies to confirm our findings. Vascular injury and associated fibrinolytic and inflammatory response studies comprise Chapter 3. We found that FXII procoagulant activity is not necessary to initiate clotting and to form stable thrombi upon an initial venous or arterial injury. However, FXII appears to aid in clot stability following a second intravascular injury 48 hours later on the contralateral veins. Inflammatory mediators are also significantly reduced in FXII deficient animals. Chapter 4 addresses the role of FXII in recruiting inflammatory cells to a site of heme-induced skin injury. Our studies explore a new method of nitric-oxide driven innate immune response, finding significantly fewer inflammatory cell infiltrates in FXII deficient cats. Chapter 5 offers future studies to pursue with this model based on results of these dissertation findings

Search for dark matter produced in association with bottom or top quarks in √s = 13 TeV pp collisions with the ATLAS detector

A search for weakly interacting massive particle dark matter produced in association with bottom or top quarks is presented. Final states containing third-generation quarks and miss- ing transverse momentum are considered. The analysis uses 36.1 fb−1 of proton–proton collision data recorded by the ATLAS experiment at √s = 13 TeV in 2015 and 2016. No significant excess of events above the estimated backgrounds is observed. The results are in- terpreted in the framework of simplified models of spin-0 dark-matter mediators. For colour- neutral spin-0 mediators produced in association with top quarks and decaying into a pair of dark-matter particles, mediator masses below 50 GeV are excluded assuming a dark-matter candidate mass of 1 GeV and unitary couplings. For scalar and pseudoscalar mediators produced in association with bottom quarks, the search sets limits on the production cross- section of 300 times the predicted rate for mediators with masses between 10 and 50 GeV and assuming a dark-matter mass of 1 GeV and unitary coupling. Constraints on colour- charged scalar simplified models are also presented. Assuming a dark-matter particle mass of 35 GeV, mediator particles with mass below 1.1 TeV are excluded for couplings yielding a dark-matter relic density consistent with measurements

31st Annual Meeting and Associated Programs of the Society for Immunotherapy of Cancer (SITC 2016) : part two

Background The immunological escape of tumors represents one of the main ob- stacles to the treatment of malignancies. The blockade of PD-1 or CTLA-4 receptors represented a milestone in the history of immunotherapy. However, immune checkpoint inhibitors seem to be effective in specific cohorts of patients. It has been proposed that their efficacy relies on the presence of an immunological response. Thus, we hypothesized that disruption of the PD-L1/PD-1 axis would synergize with our oncolytic vaccine platform PeptiCRAd. Methods We used murine B16OVA in vivo tumor models and flow cytometry analysis to investigate the immunological background. Results First, we found that high-burden B16OVA tumors were refractory to combination immunotherapy. However, with a more aggressive schedule, tumors with a lower burden were more susceptible to the combination of PeptiCRAd and PD-L1 blockade. The therapy signifi- cantly increased the median survival of mice (Fig. 7). Interestingly, the reduced growth of contralaterally injected B16F10 cells sug- gested the presence of a long lasting immunological memory also against non-targeted antigens. Concerning the functional state of tumor infiltrating lymphocytes (TILs), we found that all the immune therapies would enhance the percentage of activated (PD-1pos TIM- 3neg) T lymphocytes and reduce the amount of exhausted (PD-1pos TIM-3pos) cells compared to placebo. As expected, we found that PeptiCRAd monotherapy could increase the number of antigen spe- cific CD8+ T cells compared to other treatments. However, only the combination with PD-L1 blockade could significantly increase the ra- tio between activated and exhausted pentamer positive cells (p= 0.0058), suggesting that by disrupting the PD-1/PD-L1 axis we could decrease the amount of dysfunctional antigen specific T cells. We ob- served that the anatomical location deeply influenced the state of CD4+ and CD8+ T lymphocytes. In fact, TIM-3 expression was in- creased by 2 fold on TILs compared to splenic and lymphoid T cells. In the CD8+ compartment, the expression of PD-1 on the surface seemed to be restricted to the tumor micro-environment, while CD4 + T cells had a high expression of PD-1 also in lymphoid organs. Interestingly, we found that the levels of PD-1 were significantly higher on CD8+ T cells than on CD4+ T cells into the tumor micro- environment (p < 0.0001). Conclusions In conclusion, we demonstrated that the efficacy of immune check- point inhibitors might be strongly enhanced by their combination with cancer vaccines. PeptiCRAd was able to increase the number of antigen-specific T cells and PD-L1 blockade prevented their exhaus- tion, resulting in long-lasting immunological memory and increased median survival