Quantitative proteomic profiling of paired cancerous and normal colon epithelial cells isolated freshly from colorectal cancer patients

Abstract

Purpose: The heterogeneous structure in tumor tissues from colorectal cancer (CRC) patients excludes an informative comparison between tumors and adjacent normal tissues. Here, we develop and apply a strategy to compare paired cancerous (CEC) versus normal (NEC) epithelial cells enriched from patients and discover potential biomarkers and therapeutic targets for CRC.

Experimental design: CEC and NEC cells are respectively isolated from five different tumor and normal locations in the resected colon tissue from each patient (N = 12 patients) using an optimized epithelial cell adhesion molecule (EpCAM)-based enrichment approach. An ion current-based quantitative method is employed to perform comparative proteomic analysis for each patient.

Results: A total of 458 altered proteins that are common among >75% of patients are observed and selected for further investigation. Besides known findings such as deregulation of mitochondrial function, tricarboxylic acid cycle, and RNA post-transcriptional modification, functional analysis further revealed RAN signaling pathway, small nucleolar ribonucleoproteins (snoRNPs), and infection by RNA viruses are altered in CEC cells. A selection of the altered proteins of interest is validated by immunohistochemistry analyses.

Conclusion and clinical relevance: The informative comparison between matched CEC and NEC enhances our understanding of molecular mechanisms of CRC development and provides biomarker candidates and new pathways for therapeutic intervention.

Keywords: Biomarker discovery; Colorectal cancer; Ion current-based analysis; Tumor cell enrichment.