Project Summary My laboratory combines mechanistic cell biology with synthetic biology, focusing on understanding cellular behaviors like recognition and communication. We aim to advance the applications of engineered multicellular systems, particularly in engineered T cells targeting solid tumors. Our research has a foundation on two main questions. 1) How can we engineer highly specific cellular recognition? We seek to understand the limits and synergies of strategies for enhancing cellular specificity. Through a comparative analysis of synthetic circuits, we will explore mechanisms such multi-step signaling, and molecular titration to optimize T cell discrimination of tumors from bystander tissue based on antigen density. We also aim to uncover general principles for engineering gene expression systems and its relationship with genome organization. This may lead to general rules to engineer robust circuit behavior that could help in therapeutics. 2) how does multicellular organization affect cellular recognition? In addressing the complexity and heterogeneity of tumors, we aim for a quantitative understanding of how tissue architecture impacts T cell activity. Using an engineered spheroid platform, our research will focus into how variation in antigen density in solid tumors influence the antigen density sensing T cell activity. We anticipate extending this research to include factors like tumor inhibitory signals, inter T cell communication and the role of chemokine secretion, on T cell trafficking and tumor infiltration. The fundamental idea in this project is to control the composition and spatial organization of a spheroid and to study how these properties affect the activity of engineered T cells. In summary, our group will apply principles of molecular recognition and novel methods in cell and tissue engineering to understand and control cellular behavior. By systematically deconstructing the problem of how T cells recognize tumors from bystander tissue based on antigen density and studying the influence of tumor organization to the immune response, we aim to layout fundamental rules to engineering recognition at the cellular level and improve therapeutic cells.