Small Molecule Modulator of the mTORC2 Pathway Discovered\nfrom a DEL Library Designed to Bind to Pleckstrin Homology Domains

Pleckstrin homology\n(PH) domains are structural motifs critical\nfor cellular processes, such as signal transduction and cytoskeletal\norganization. Due to their involvement in various diseases, PH domains\nare promising therapeutic targets, yet their highly charged and hydrophobic\nbinding sites are not ideal for traditional small drugs. In this study,\nwe designed a DNA-encoded library (DEL) mimicking phospholipids to\nidentify novel modulators targeting PH domains with uncharted chemical\nproperties. Screening against several PH domains led to the discovery\nof 2DII, a small molecule that selectively binds to mSin1<sup>PH</sup>. This compound can modulate mTORC2 activity by impairing mTORC2’s\nmembrane interactions, resulting in reduced AKT1 phosphorylation.\nA micromapping via Dexter energy transfer based on 2DII bearing an\niridium catalyst (2DII-Ir), along with a biotin-diazirine small molecule\nwas used for target identification by proteomics, which confirmed\nmSin1 as the primary intracellular target of 2DII, demonstrating its\npotential for selective mTORC2 pathway modulation. These findings\nintroduce a novel strategy for targeting PH domains and provide a\nfoundation for the development of therapeutic interventions that modulate\nPH-domain-dependent signaling pathways.