Abstract Body: Missense pathogenic variants in ACTA2, encoding α-smooth muscle actin (SMA), predispose to thoracic aortic aneurysms, but pathogenic variants affecting the specific amino acid arginine 179 (R179) cause a more severe childhood onset phenotype termed Smooth Muscle Dysfunction Syndrome (SMDS). SMDS patients have fully penetrant aortic disease, moyamoya-like cerebrovascular disease, and many other clinical issues stemming from defective smooth muscle function throughout the body. We previously confirmed that SMA localizes to the nucleus in wild-type (WT) smooth muscle cells (SMCs), is enriched in the nucleus over β-actin with SMC differentiation, associates with the INO80 chromatin remodeling complex, and selectively binds to the promoters of SMC contractile genes. R179 variants prevent SMA nuclear localization, leading to incomplete differentiation of SMCs. We generated a conditional knock-in Acta2^SMC-R179C/+ mouse model and previously showed that approximately 67% of aortic SMCs have the variant correctly introduced. These mutant cells recapitulate the incompletely differentiated phenotype observed in vitro. Left carotid artery injury (LCAL) was performed on the Acta2^SMC-R179C/+ and littermate wild-type (WT) mice, and 25% of Acta2^SMC-R179C/+ die within 5 days after LCAL due to ischemic strokes, with a more severe phenotype observed in female mice. We sought to correct the incomplete differentiation phenotype by using a repurposed cancer drug targeting the epigenetic regulator EZH2. The drug, tazemetostat, inhibits the function of EZH2, which is the catalytic subunit of the polycomb repressor complex 2 and is responsible for the trimethylation modification of histone 3 lysine 27 (H3K27me3). Importantly, a prior publication showed that SMCs with ACTA2 p.R179 have globally increased H3K27me3 levels, and our own studies confirm this finding in the endogenous heterozygous context. Treatment with tazemetostat increases SMC contractile protein accumulation and decreases migration in Acta2^R179C/+ SMCs. Chromatin immunoprecipitation studies confirm that H3K27me3 levels are decreased on SMC gene promoters after tazemetostat in both Acta2^R179C/+ and WT SMCs. In-progress work will define locations where EZH2 is differentially active in Acta2^R179C/+ and WT SMCs. We will also pursue a treatment trial in our Acta2^SMC-R179C/+ mouse model after LCAL injury. Taken together, our work identifies a promising mechanistic therapy that could benefit ACTA2 p.R179 patients.