Positive selection screen identifies natural product β-catenin inactivators

Finding compounds that can inhibit genetically validated, but difficult to drug, targets is a pressing problem in drug discovery. Natural products often exhibit unexpected pharmacological mechanisms of action that are not easily predicted. Herein, for proof of concept, we sought natural products that can inactivate ?-catenin (?-cat), which is a historically challenging cancer target. Utilizing a positive selection assay that fuses oncogenic ?-cat to a suicide protein, we identified a natural product that activates novel-type protein kinase Cs (PKCs) and thereby mislocalizes and inactivates ?-cat. This work illustrates the opportunity of combining so-called ?up-assays? and complex natural product mixtures to discover active natural products that define new mechanistic paradigms to combat challenging target proteins. Many genetically validated targets in cancer, including the transcription factor ?-catenin (?-cat), have historically been viewed as undruggable. Cell-based phenotypic screening of chemical compounds can reveal unanticipated biological and pharmacological principles. Natural products are powerful probes because of their superior structural diversity, drug-like properties, and biological activities as compared to unoptimized synthetic compounds. We screened 326,304 natural product mixtures (40,744 extracts and 285,560 fractions derived from them) using mammalian cells expressing an oncogenic version of ?-cat fused to a suicide protein. Multiple fractions degraded the ?-cat fusion protein or drove it into a compartment where both fusion partners were apparently inactive. The active natural product from one of the latter specifically activates novel, but not classical, protein kinase Cs and thereby relocates ?-cat to juxtamembrane vacuolar structures. These findings suggest a path for inactivating oncogenic ?-cat and underscore the power of screening natural product collections with robust phenotypic assays.