(217d) Engineering Cell-Based Therapies That Recognize Defined Combinations of Environmental Cues

Technologies enabling bioengineers to construct cell-based therapies that sense and respond to specific combinations of environmental cues may transform our ability to design and implement novel therapeutic strategies that overcome existing barriers to treatment.  For example, engineering cell-based immunotherapies that better discriminate between tumors and healthy tissue could improve the safety and efficacy of this promising approach.  Implementing such strategies will require synthetic biology “parts” that sense physiologically relevant and exclusively extracellular cues, such as protein mediators of intracellular communication, and relay this sensing event to engineered circuits comprised of synthetic proteins, RNA, and/or DNA.  To address this need, we developed the first fully orthogonal cell surface biosensor platform, termed a modular extracellular sensor architecture (MESA). MESA receptors use a generic signal induction mechanism wherein receptor dimerization results in the release of an engineered transcription factor from the plasma membrane. Here, we leverage MESA technology to engineer cell-based devices that perform multiparametric evaluation of extracellular cues using engineered receptors coupled to intracellular gene circuits. We first evaluated the potential for multiplexing MESA receptors – expressing multiple receptors in the same cells. Orthogonal MESA receptors release distinct transcription factors to serve as inputs to a downstream gene circuit. Finally, we describe MESA receptors that utilize a novel class of affinity reagent as a ligand binding domain – the camelid antibody analog termed a “nanobody” – an approach that may ultimately facilitate engineering cell-based therapies that recognize diverse environmental cues. Like many synthetic biology technologies, these platforms provide powerful tools for scientific inquiry and investigation of disease processes as well as novel therapeutic modalities.