Journal Article

Fluorescence based live cell imaging identifies exon 14 skipped hepatocyte growth factor receptor (MET) degraders

Alvarez, Sophie, Naldrett, Michael J, Natarajan, Amarnath, Kizhake, Smitha, Rana, Sandeep, Tolosa, Ezequiel, Zahid, Muhammad, Ayers-Ringler, Jennifer, Borad, Mitesh, Kosari, Farhad, Anastasiadis, Panos Z, Mallareddy, Jayapal Reddy, Kubica, Sydney P, Boghean, Lidia, Schaefer-Klein, Janet, Yang, Lin, Feathers, Ryan, Lin, Wan-Hsin, Singh, Sarbjit, Mansfield, Aaron S, Roden, Anja, Kizhake, Amritha G

2024

Overview

Despite ongoing efforts to employ structure-based methods to discover targeted protein degraders (TPD), the prevailing strategy continues to be the synthesis of a focused set of heterobifunctional compounds and screen them for target protein degradation. Here we used a fluorescence based live cell imaging screen to identify degraders that target exon 14 skipped hepatocyte growth factor receptor (MET). MET is a known oncogenic driver. MET exon 14 skipping mutations (METex14Δ) are found in lung cancers and result in the loss of a degron that is required for E3-ligase recognition and subsequent ubiquitination, prolonging the half-life and oncogenicity of MET. Since proteolysis targeting chimeras (PROTACs) are heterobifunctional molecules that promote target degradation by the proteosome, we sought to restore degradation of MET lost with METex14Δ using a MET-targeting PROTAC. We generated a library of sixty PROTACs of which 37 used the MET inhibitor capmatinib as the protein of interest targeting ligand. We screened this PROTAC library for targeted degradation of METex14Δ-GFP using live cell imaging. We benchmarked out MET-targeting PROTACs to that of a previously reported MET-targeting PROTAC, SJF8240. Curve fitting live cell imaging data affords determination of time required to degrade 50% of the target protein (DT50), which was used in determining structure activity relationships. A promising candidate, 48-284, identified from the screen, exhibited classic PROTAC characteristics, was > 15-fold more potent than SJF8240, had fewer off targets compared to SJF8240, and degraded MET in multiple cell lines.

• Publisher: Cold Spring Harbor Laboratory Press,Cold Spring Harbor Laboratory
• Subject: c-Met protein / Cancer Biology / Cell lines / Chimeras / Degradation / Growth factors / Hepatocyte growth factor / Protein biosynthesis / Protein folding / Protein structure / Proteins / Proteolysis / Ubiquitination