Synthetic Chronogenetic Gene Circuits for Circadian Cell Therapies

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Abstract The goal of this project will be to combine principles of synthetic biology, tissue engineering, and circadian biology to generate living stem cell-based implants that can deliver biologic drugs in a prescribed circadian manner for the treatment of a wide range of disease processes that exhibit diurnal rhythmicity. To engineer this regulatory system, the core clock gene circuitry of murine induced pluripotent stem cells (iPSCs) will be engineered to synthesize anti-cytokine biologic drugs. These cells will be used to form tissue-engineered constructs for in vitro and in vivo testing. The first aim of this project will focus on the creation of synthetic “chronogenetic” gene circuits with prescribed frequency, phase, and amplitude of biologic drug delivery. More specifically, cells will be designed to synthesize inhibitors of either interleukin 1 or tumor necrosis factor alpha. These cells will be engineered into stable cartilage constructs in injectable hydrogel carriers that will be tested as a therapeutic approach for the treatment of experimental rheumatoid arthritis (RA) in two different mouse models of this disease. RA is an inflammatory disease that affects approximately 1% of the general population. Several anti-cytokine biologics are currently in clinical use; however, the continuous administration of these therapeutics at high level can lead to significant adverse effects. We posit that drug delivery that aligns with the circadian rhythmicity of peak inflammatory cytokine release will have significantly improved effectiveness. The efficacy of these approaches will be assessed using clinical, histologic, molecular, and pain/behavior testing. The creation of such synthetic chronogenetic systems cells provides the possibility for long-term “chronotherapy”, or timed drug delivery for the treatment of many chronic inflammatory diseases beyond RA.