PROJECT SUMMARY Immune tolerance is mediated via a number of molecular and cellular mechanisms, including regulatory T cells (Treg). Breakdown of this immune tolerance leads to debilitating autoimmune disease, of which rheumatoid arthritis (RA) is exemplary. Polymorphisms in alleles associated with immune signaling can predispose individuals to the development of RA. Genetic variants at the protein tyrosine phosphatase non-receptor 2 (PTPN2) locus are one such set of mutations that are well characterized to contribute to disease pathogenesis and RA severity in a well-established subset of patients. This altered signaling results in changes to the immunological landscape, and particularly influences the balance between anti-inflammatory Treg and the pro- inflammatory T helper 17 (Th17) cells that are involved in RA development. Diminished PTPN2 function results in Treg instability, in which Treg convert to pathogenic “Th17-like” effector T cells in the inflammatory arthritic microenvironment, leading to self-perpetuating cycle of severe inflammation. However, currently there are no therapies that target Treg instability to rescue Treg function and promote immune homeostasis to help resolve inflammation. Cyclodextrin/all-trans retinoic acid complexes (CAC) are a promising tool to address the need for a clinically translatable therapy to restore and maintain Treg function in PTPN2 haploinsufficient (PTPN2+/-) individuals, as CAC are capable of preventing Treg destabilization in inflammatory conditions. However, the systemic administration of CAC to promote Treg is not a feasible strategy, as this may run the risk of systemic immune suppression that could increase the risk of serious infection and cancer. To address this, it is hypothesized that the local, sustained delivery of CAC at the joint will enhance Treg stability and improve clinical outcomes in the PTPN2 compromised endotype. To achieve prolonged, site specific immune modulation, an injectable biomaterial system for the release of CAC is necessary. The goal of this project is to (i) develop and characterize an injectable biomaterial system for the sustained delivery of CAC and (ii) elucidate the efficacy, mechanisms, and specificity of CAC-mediated Treg stabilization. This will be achieved in the following Specific Aims: Aim 1. A microparticle formulation (CAC-MP) comprising CAC encapsulated by poly (lactic-co- glycolic) acid (PLGA) will be characterized and optimized for in vivo release kinetics. Subsequently, a dosage study will be conducted to examine efficacy of CAC-MP at different doses in the Ptpn2+/- SKG model of RA. Aim 2. The mechanism of action by which CAC-MP mediate improvement in Ptpn2+/- SKG arthritis will be elucidated, and the impact of CAC-MP on systemic immune response will be examined. The immediate results of this project will determine the feasibility of localized immune modulation to rescue Treg instability to improve outcomes in a well-defined subset of RA patients us...