Design and mathematical analysis of activating transcriptional amplifiers that enable modular temporal control in synthetic juxtacrine circuits

The ability to control mammalian cells such that they self-organize or enact therapeutic effects as desired has incredible implications. Not only would it further our understanding of native processes such as development and the immune response, but it would also have powerful applications in medical fields such as regenerative medicine and immunotherapy. This control is typically obtained by synthetic circuits that use synthetic receptors, but control remains incomplete. The synthetic juxtacrine receptors (SJRs) are widely used as they are fully modular and enable spatial control, but they have limited gene expression amplification and temporal control. As these are integral facets to cell control, I therefore designed transcription factor based amplifiers that amplify gene expression and enable unidirectional temporal control by prolonging duration of target gene expression. Using a validated in silico framework for SJR signaling, I combined these amplifiers with SJRs and show that these SJR amplifier circuits can direct spatiotemporal patterning and improve the quality of self-organization. I then show that these circuits can improve chimeric antigen receptor (CAR) T cell tumor killing against various heterogenous antigen expression tumors. These amplifiers are flexible tools that improve control over SJR based circuits with both basic and therapeutic applications.