Abstract Tendon and ligament injuries represent an acute healthcare burden in the United States, costing >$30 billion annually. Tendon injuries frequently result in scar-like tissue with inferior physical properties - however no regenerative therapy exists to date. Recently, we have identified and characterized perivascular (CD146+) tendon stem/progenitor cells (TSCs) that play an essential role in tendon healing via FAK and ERK1/2 signaling. In our preliminary study, we screened small molecules from a library of FAK and ERK1/2 agonists and identified Oxotremorine M (Oxo-M) and PPBP maleate (4-PPBP) that stimulated TSCs toward regenerative tendon healing. Oxo-M and 4-PPBP were originally developed for treating neuronal diseases but have never been tested in the musculoskeletal system. In vitro, a combination of Oxo-M and 4-PPBP induced significant increases in the expression of tendon-related genes involved in tendon repair. Oxo-M and 4-PPBP showed no cytotoxicity up to 10X working doses. Western blot and siRNA knockdown (KD) confirmed that FAK and ERK1/2 signaling regulate Oxo -M & 4-PPBP-induced tenogenic differentiation of TSCs. In vivo, direct topical delivery of Oxo-M and 4-PPBP onto full-transected rat patellar tendons (PT) significantly improved tendon healing, as observed histologically as densely reorganized collagen fibrils, and functionally as significantly enhanced tensile strength. This process was guided by a rapid but transient increase in the number endogenous TSCs undergoing tenogenic differentiation. In addition, Oxo-M and 4-PPBP specifically targeted CD146+ TSCs through muscarinic acetylcholine receptors (AChRs) and σ1 receptor (σ1R) pathways, with minimal effect on other types of tendon cells. These findings demonstrate a novel and promising activity of the combination of Oxo-M and 4-PPBP in tendon healing by specifically targeting endogenous TSCs. The overall objectives of this STTR grant are to develop a reliable and effective small molecule-based regenerative therapy for tendon injuries by transiently activating regenerative pathways of endogenous TSCs and repair tendon tears. The overarching goal of the STTR phase I grant is to optimize the combination of Oxo-M and 4-PPBP, the 2 compounds and determine their drugability in combination. The 2 aims of the phase I STTR are to obtain robust proof-of-concept and preliminary safety data to establish technical merit, feasibility, and commercial potential of the innovative technology.