Chemically Programmable DNA Nanostructures for Multiplexed High-Throughput Screening of Cardiovascular Therapeutics

The development of small-molecule therapeutics for cardiac hypertrophy is limited by the lack of high-throughput screening platforms capable of multiplexed molecular detection. Here, we present the Multifunctional Nucleic-acid-label-free hESC-CMs Molecular Evolution ( MNCME ) platform, which integrates tetrahedral DNA nanostructures with human embryonic stem cell-derived cardiomyocytes (hESC-CMs) for chemically programmable, label-free detection of DNA, RNA, proteins, and small molecules. By dynamically substituting three label-free primers, MNCME enables flexible molecular profiling and rapid target adaptation. Machine learning-assisted analysis of known anti-hypertrophic compounds identified key molecular signatures, optimizing high-throughput screening workflows. Structurally organized into five functional zones, MNCME screened a 600-compound library, identifying nine bioactive molecules, including caffeic acid (CA), previously unrecognized for its cardioprotective potential. Mechanistic studies revealed that CA modulates ATP synthesis, Ca²⁺ homeostasis, and Mucin-1 signaling, with validation in cellular and in vivo models. By integrating DNA nanotechnology, chemical biology, and high-throughput phenotypic screening, MNCME provides a scalable and versatile platform for accelerating the discovery of small-molecule modulators targeting cardiac hypertrophy. This approach has the potential to transform therapeutic development for cardiovascular diseases and other complex pathologies, offering a powerful tool for both fundamental research and clinical translation.