ABSTRACT Chronic kidney disease is a significant healthcare issue affecting >15% of the U.S. population and costing billions in healthcare dollars annually. Transplantation is the best option for most patients with progressive disease, resulting in a significant increase in life expectancy and improved quality of life compared to dialysis. The potential U.S. deceased donor organ supply is estimated to exceed the current number of organs transplanted by a factor of 4- to 5-fold, with a major limitation to the number of acceptable organs for transplant being the ischemic injury sustained between recovery and implantation. A method to cryopreserve or “bank” kidneys prior to transplant would effectively remove the influence of time from the supply chain of organ distribution. This would allow a new paradigm for transplantation that would improve donor/recipient matching, allow for better patient preparation, facilitate tolerance induction protocols, and increase organ utilization while improving graft and patient survival. One promising approach that overcomes the limitations of conventional strategies is vitrification—that is, cooling organs so quickly that they cannot undergo the phase transition from liquid to solid ice. With the help of cryoprotective agents (CPAs), the organ enters a stable glass-like state wherein viable storage is theoretically indefinite. The critical challenge, however, is rewarming without ice formation or cracking: if rewarming is too slow, ice crystals form, and if rewarming is not uniform, thermal stress causes cracking. During our initial R01 funding, we developed a novel approach termed “nanowarming” that achieved both objectives. Iron oxide nanoparticles were perfused throughout the vasculature of the organ along with CPA solutions. The organ was then vitrified by cooling and rewarmed on-demand by placing it in a radiofrequency coil that induces heating in the nanoparticles and, therefore, from within the organ. We found that nanowarming could rewarm vitrified organs, including kidneys, in animal models. We have recently shown, for the first time, that nanowarmed organs function in vitro and in vivo following transplantation. Further, we showed successful vitrification and nanowarming of human-sized (porcine) kidneys. These new data support the feasibility of our approach to cryopreserve and nanowarm whole human organs for transplantation. Nevertheless, many questions remain, including how nanowarmed kidneys function compared to control organs, what, if any, injury occurs during nanowarming, and how to scale up to human-sized organs. In this renewal R01, we propose to: (1) Quantitatively assess cryopreserved and nanowarmed kidney transplant function in a rat model, including long- term preservation, long-term function, modes of injury, and alterations of the host immune response, (2) Engineer and optimize scale-up for nanowarming vitrified human-sized organs, and (3) Vitrify and nanowarm human-sized kidneys while measuring viability, structural integrity, and organ function.

PROJECT NARRATIVE We are developing a novel technology for indefinitely preserving donor kidneys for transplant in an extremely low-temperature, vitrified (glass-like) state and subsequently rewarming them on-demand using nanotechnology. This method for cryopreserving or “banking” kidneys prior to transplant will allow a new paradigm for transplantation that improves donor/recipient matching, allows for better patient preparation, facilitates tolerance induction protocols, and increases organ utilization; all while improving kidney durability and patient survival.