Split-CAT, a discovery engine for molecular glues: a novel modality for cancer therapy

Targeted protein degradation is a novel therapy modality for cancer and other diseases, decreasing the level of undruggable proteins rather than inhibiting their activity. In this approach molecular glue molecules augment the ubiquitylation dependent degradation of disease-causing proteins such as transcription factors. However, the discovery of molecular glues is highly difficult as they must have two functional ends that simultaneously bind a ubiquitin ligase and its target. We developed a novel discovery engine for molecular glues that harnesses positive selection in E. coli. The system was structurally designed by a split chloramphenicol acetyltransferase (split-CAT) reporter that genetically links ubiquitylation to antibiotic resistance and growth. We demonstrated the system’s responses and specificity to the known thalidomide-like glues and their targets. The absence of ubiquitylation-dependent degradation cascades and deubiquitylation in E. coli significantly simplify the screen and the results interpretation. We will present the employment of the discovery engine to identify novel ubiquitylation cascades and molecular glues involved in various cancers.