Abstract Body: While cell models have been used to study cardiac diseases, there is currently no standardization or fully mature in vitro system. Obtaining a mature cardiomyocyte (CM) phenotype is necessary to create a physiologically relevant environment and accurately model and study cardiac diseases. Increased success in maturation of human induced pluripotent stem cell derived CMs (hiPS-CMs) has been achieved in 3D in vitro models in the form of engineered heart tissues (EHTs). Digital light processing (DLP) offers a high throughput and efficient method to fabricate hydrogels that recapitulate properties of native tissues. In this study, we use DLP-printed EHTs to seed and mature differentiated hiPS-CMs and test their potential for modeling pathological cardiac hypertrophy. To further encourage maturity of CMs, we utilized a low glucose maturation medium supplemented with fatty acids. DLP-printed EHTs displayed high reproducibility and increased CM maturity. To determine degree of maturity of our system, we performed RT-qPCR, western blot analysis, and immunofluorescence (IF) microscopy. We found increased expression of fatty-acid transport and beta oxidation genes and improved sarcomere alignment compared to 2D hiPS-CMs. To model pathological cardiac hypertrophy, we used two different approaches: 1. acute adrenergic agonism with isoproterenol and phenylephrine; 2. chronic culturing of EHTs in pathologically stiff conditions. To assess pathology, we conducted RT-qPCR, western blot, ELISA, IF microscopy, and functional analyses. We found that agonist-treated EHTs displayed increased pathology-associated gene expression compared to standard 2D hiPS-CMs. Both agonist-treated and stiff EHTs had increased secreted B-Type Natriuretic Peptide (BNP) and phosphorylation of proteins involved in pathological remodeling, including ERK and P38. Additionally, we observed increased nuclear area, increased calcium handling, and increased contractile force in the treatment groups. In conclusion, we have established a high throughput, reproducible, mature EHT platform that can be used to model pathological cardiac hypertrophy. This platform can be easily adopted by other laboratories to study a wide array of cardiac diseases.