Abstract 5307: Critical platforms of PROTAC validation

Abstract

Abstract In recent years, proteolysis targeting chimeric (PROTAC) technology has become an effective endogenous protein degradation tool in drug discovery. As a class of heterobifunctional small molecules, PROTACs recruit an E3 ubiquitin ligase to a given substrate protein, resulting in its targeted degradation through the endogenous ubiquitin-proteasome system (UPS). PROTAC consists of three chemical elements: a ligand that binds to a target protein, a ligand that binds to an E3 ubiquitin ligase, and a linker that connects the two ligands. Compared with traditional drug development, PROTAC technology possesses unique advantages including overcoming drug resistance and targeting the “undruggable”. Here we generated a kind of PROTAC target, like KRAS, BRD4, ESR1, associated cell lines to accelerate the discovery of new protein degradation drugs and the target protein degradation therapeutics by an optimized insertion system. In the beginning, we develop a system of different tag targeted cell lines, which can be easy to introduce HiBiT, Flag, GFP and also Halo Tag, et.al to target protein by CRISPaint or CRISPR Cas9 mediated HDR system. We also developed an arbitrary point mutation system based on the optimized PE3 mutation system, which can randomly modify any point mutation to simulate in vivo disease models in vitro. During this process, we have also optimized a series of validation system, like Targeted Protein Degradation (TPD), High-throughput flow cytometry (HTFC) and In-Cell Western (ICW) to shorten the detection time and improve detection stability. It is worth mentioning that we also develop a HiBiT and GFP dual-signal system, which can easily validate the target protein degradation and cell viability. We hope that our detection system can speed up the development of new drugs and contribute to human anti-tumor.Keywords: PROTAC, UPS, drug development, therapeutics Citation Format: Ruifeng Wang. Critical platforms of PROTAC validation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5307.