Project Summary Medicinal chemistry saw a paradigm shift in the 1980’s when natural product isolation and phenotypic screens were largely replaced by combinatorial chemistry and structure-based drug design. This transformation was prompted by key advances in molecular biology, structural biology, and synthetic chemistry. For example, the advent of reliable, high yielding coupling chemistries (e.g., peptide coupling, palladium-mediated cross couplings) and a set of concise rules (i.e., Lipinski’s rules) revolutionized the efficiency of medicinal chemistry. Although this expedited the rate of production and size of small-molecule libraries, it was not without a cost. Modern chemical libraries are overpopulated with flat, nondescript compounds that contain fewer than one stereocenter per member and ca. 20% sp3 content. These properties are favorable for engaging deep binding pockets in proteins; however, they are ill-equipped for the shallower surfaces involved in macromolecular (e.g., protein-protein, protein-oligonucleotide) interactions. On the other hand, natural products and their derivatives have proven capable of binding a wide range of challenging targets, including those that are considered “undruggable” due to their skeletal and stereochemical complexity”, and consequently represent the majority of FDA-approved drugs. Diversity-oriented synthesis (DOS), biology-oriented synthesis, and complexity-to- diversity have emerged as organic chemistry strategies that embrace the power of natural product complexity to establish diverse chemical libraries emphasizing densely functionalized, entropically constrained, and stereochemically defined cores. Concurrently, chemical proteomic platforms, such as activity-based protein profiling, are providing a global approach to map the interactions between small molecules and proteins directly in native biological systems for the first time. Chemical proteomic efforts have radically expanded the number of proteins and ligandable sites that can be targeted by small molecules, including many examples that previously lacked chemical probes. This proposal aims to integrate the principles of DOS, covalent chemistry, and chemical proteomics to create highly innovative small-molecule libraries of stereochemical and skeletal complexity and to use these libraries to discover the first selective and cell-active chemical probes for diverse oncoprotein targets. In Specific Aim 1, we will optimize acrylamide ligands that show stereoselective engagement of key cancer targets, including oncogenic transcription factors and nucleotide exchange factors. Advanced compounds, either as direct functional antagonists or bifunctional degraders, will be used to suppress the growth of cancer cells that have been shown to strongly and selectively depend on these oncoproteins. In Specific Aim 2, we will expand our stereoisomeric library to interrogate new three-dimensional chemical space. Specifically, we will establish a library of e...