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

Chem Commun (Camb). 2017 Jun 29;53(53):7234-7237. 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.

MeSH terms

  • Acrylamide / chemistry
  • Acrylamide / pharmacology*
  • Antineoplastic Agents / chemistry
  • Antineoplastic Agents / pharmacology*
  • Colorectal Neoplasms / drug therapy*
  • Colorectal Neoplasms / metabolism
  • Colorectal Neoplasms / pathology
  • Cysteine / chemistry*
  • Endoplasmic Reticulum / drug effects*
  • Endoplasmic Reticulum / metabolism
  • Humans
  • Ligands
  • Nogo Proteins / antagonists & inhibitors*
  • Nogo Proteins / genetics
  • Nogo Proteins / metabolism
  • Nuclear Envelope / drug effects
  • Nuclear Envelope / metabolism
  • Proteomics*


  • Antineoplastic Agents
  • Ligands
  • Nogo Proteins
  • RTN4 protein, human
  • Acrylamide
  • Cysteine