Chemoproteomics-enabled covalent ligand screen reveals a cysteine hotspot in reticulon 4 that impairs ER morphology and cancer pathogenicity

• Published in: Chemical communications (Cambridge, England) Vol. 53; no. 53; pp. 7234 - 7237
• Main Authors: Bateman, L A, Nguyen, T B, Roberts, A M, Miyamoto, D K, Ku, W-M, Huffman, T R, Petri, Y, Heslin, M J, Contreras, C M, Skibola, C F, Olzmann, J A, Nomura, D K
• Format: Journal Article
• Language: English
• Published: England 29.06.2017
• Subjects: Acrylamide - chemistry; Acrylamide - pharmacology; Antineoplastic Agents - chemistry; Antineoplastic Agents - pharmacology; Cancer; Colorectal Neoplasms - drug therapy; Colorectal Neoplasms - metabolism; Colorectal Neoplasms - pathology; Covalence; Cysteine - chemistry; Endoplasmic reticulum; Endoplasmic Reticulum - drug effects; Endoplasmic Reticulum - metabolism; Fragmentation; Genetics; Hot spots; Humans; Ligands; Morphology; Nogo Proteins - antagonists & inhibitors; Nogo Proteins - genetics; Nogo Proteins - metabolism; Nuclear Envelope - drug effects; Nuclear Envelope - metabolism; Proteomics
• ISSN: 1359-7345; 1364-548X
• DOI: 10.1039/c7cc01480e

Chemical genetics has arisen as a powerful approach for identifying novel anti-cancer agents. However, a major bottleneck of this approach is identifying the targets of lead compounds that arise from screens. Here, we coupled the synthesis and screening of fragment-based cysteine-reactive covalent ligands with activity-based protein profiling (ABPP) chemoproteomic approaches to identify compounds that impair colorectal cancer pathogenicity and map the druggable hotspots targeted by these hits. Through this coupled approach, we discovered a cysteine-reactive acrylamide DKM 3-30 that significantly impaired colorectal cancer cell pathogenicity through targeting C1101 on reticulon 4 (RTN4). While little is known about the role of RTN4 in colorectal cancer, this protein has been established as a critical mediator of endoplasmic reticulum tubular network formation. We show here that covalent modification of C1101 on RTN4 by DKM 3-30 or genetic knockdown of RTN4 impairs endoplasmic reticulum and nuclear envelope morphology as well as colorectal cancer pathogenicity. We thus put forth RTN4 as a potential novel colorectal cancer therapeutic target and reveal a unique druggable hotspot within RTN4 that can be targeted by covalent ligands to impair colorectal cancer pathogenicity. Our results underscore the utility of coupling the screening of fragment-based covalent ligands with isoTOP-ABPP platforms for mining the proteome for novel druggable nodes that can be targeted for cancer therapy.