Abstract Body: Familial dilated cardiomyopathy (DCM) is a genetic heart disorder characterized by enlargement of one or both ventricles, rendering the heart unable to pump blood efficiently. Genetic segregation identified a mutation in α-Tropomyosin (Tm) at residue 230 (D230N-Tm) linked to severe, early onset DCM. To meaningfully link genotype to phenotype our group sought to investigate the effects of D230N-Tm on the biophysical structure of the Tm filament. It was shown that D230N-Tm caused a compaction of the cardiac Troponin T (cTnT)–Tm overlap region and a concomitant decrease in flexibility. Given the altered biophysical properties, we screened for molecules that could target these structural changes utilizing a fully atomistic cardiac thin filament model. Computational modeling and biophysical studies identified small molecule 4-Hydroxy Duloxetine β-D-Glucuronide (Z06) which was shown to alleviate compaction of the overlap and restore flexibility of the Tm filament in vitro. We hypothesized that restoring the flexibility of the overlap region in D230N-Tm would improve regulatory and cardiac function in our D230N-Tm transgenic mice. To investigate the effect of Z06 on D230N-Tm myofibrils we performed an NADH-coupled ATPase assay. ATPase activity was measured at saturating calcium (100uM CaCl₂) conditions in the presence (250uM) and absence (0uM) of Z06. In the absence of Z06, D230N-Tm exhibited a decrease in ATPase activity (0.065±0.001 umol/min) compared to NTg littermates (0.094±0.005 umol/min). Addition of 250uM Z06, restored D230N-Tm ATPase rates (0.089±0.003 umol/min) to levels comparable to NTg mice. To then assess the ability of Z06 in improving systolic function we administered Z06 to 2-month-old D230N-Tm mice and their NTg littermates. At baseline, D230N-Tm mice exhibited a decrease in percent fractional shortening (%FS) of 22.50±0.563 compared to their NTg littermates (29.90±0.653. After four weeks of treatment we observed an improvement in %FS in D230N-Tm mice to 24.341±0.812 with no significant changes to systolic function in NTg littermates. Future work will investigate the efficacy of Z06 as a late-stage treatment and assess its general applicability in other thin filament mutations, D219N-Tm (HCM) and R173W-cTnT (DCM).