Cell-Based Therapy in Minipig Model of Radiation-Induced Xerostomia

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This project will develop a fully functional, implantable human salivary gland for patients suffering from xerostomia/dry mouth after radiotherapy for head and neck cancer. Despite the regulatory requirement for a close-to-human animal model for clinical translation of tissue engineered organs, large animal models do not exist to test stem-cell approaches to restore salivary function including fluid secretion and protein production needed for digestion and oral health. Rodent models, while useful to test product designs, lack key attributes needed to deliver an implantable biological device to the clinic. To address these shortcomings, we assembled an interdisciplinary team that includes the Farach-Carson/Harrington team in Houston, the Passineau laboratory in Pittsburgh to develop an approach using a radiated pig model amenable to genetic manipulation, and the Lombaert laboratory in Ann Arbor to assess implant integration. Our recent demonstration that a human-in-pig model under immunosuppression is well tolerated presents an unprecedented opportunity to develop xenograft model for the salivary human stem/progenitor cell (hS/PC) transplantation studies and avoid the need to develop a pig-in-pig autograft. This proposal builds on our exciting preliminary results to develop and characterize a novel radiated immunosuppressed mini-pig model for testing the ability of transplanted hS/PCs to restore salivary secretory function. We hypothesize that this model will recapitulate the environment of the radiated human salivary bed, and provide the type of information needed to evaluate the function of human S/P cells one day to be transplanted into patients suffering from xerostomia. Specific Aims are: 1) Establish the radiated immunosuppressed minipig as a suitable host animal to evaluate the long term stability, biocompatibility and fate of matrix-modified hyaluronate (HA) hydrogel materials containing encapsulated hS/PCs; a) Develop and standardize optimized strategies to place the salivary neotissue prototype in the host parotid bed; b) Evaluate stability of implanted acellular hydrogel constructs at various times after placement in the radiated parotid and make formulational changes as needed to ensure long term biointegration; c) Use a quantitative scoring system to evaluate the fate over time of encapsulated hS/PC-loaded constructs after transplantation into the radiated salivary bed including viability, growth without overgrowth, host integration including vasculature and nerve, and differentiation into salivary structures expressing acinar, ductal and myoepithelial markers. 2) Evaluate the ability of the transplanted tissue to restore salivary function: a) Demonstrate functional protein delivery from the salivary neotissues into the irradiated minipig by combining restorative gene therapy with implanted salivary neotissues; b) Demonstrate restoration of fluid flow by evaluating aquaporin 5 expression in neotissues and fluid production using direct saliva collection techniques. The evaluation of salivary functional restoration will be aided by use of a genetic modification to create a secreted Met-Luc S/P cell that when implanted would provide a noninvasive means of following graft survival over time simply by sampling Luc in host blood or saliva.