Exploration of the Key Proteins in the Normal-Adenoma-Carcinoma Sequence of Colorectal Cancer Evolution Using In-Depth Quantitative Proteomics

Purpose: In most cases, the carcinogenesis of colorectal Cancer (CRC) follows the normal-adenoma-carcinoma (N-A-C) sequence. In this study, we aimed to identify the key proteins in the N-A-C sequence.

Methods: Differentially expressed proteins (DEPs) in normal, adenoma, and carcinoma tissues were identified using the Tandem Mass Tag- (TMT-) based quantitative proteomics approach. The landscape of proteomic variation in the N-A-C sequence was explored using gene set enrichment analysis (GSEA) and Proteomaps. Key proteins in the N-A-C sequence were identified, verified, and validated based on our proteomic data, external proteomic data, and external transcriptomic data in the ProteomeXchange, CPTAC, GEO, and TCGA databases. The prognostic value of the key proteins in our database was evaluated by univariate and multivariate COX regression analysis. The effects of the key proteins on adenoma organoids and colorectal Cancer cells were explored in functional studies.

Results: Based on our proteomic profiles, we identified 1,294 DEPs between the carcinoma (CG) and normal (NG) groups, 919 DEPs between the adenoma group (AG) and NG, and 1,030 DEPs between the CG and AG. Ribosome- and spliceosome-related pathways were mainly enriched in the N-A process. Extracellular matrix- and epithelial-mesenchymal transition- (EMT-) related pathways were mainly enriched in the A-C process. RRP12 and SERPINH1 were identified, verified, and validated as candidate key proteins in the N-A and A-C processes, respectively. Furthermore, RRP12 and SERPINH1 knockdown impeded the viability and proliferation of adenoma organoids. SERPINH1 was validated as a risk factor for disease-free survival (DFS) based on the TCGA and our database, whereas RRP12 did not show prognostic value. SERPINH1 knockdown was accompanied by EMT-related protein variation, increased Apoptosis, and reduced proliferation, invasion, and migration of CRC cells in vitro.

Conclusions: RRP12 and SERPINH1 may play an important role in the N-A and A-C processes, respectively. Furthermore, SERPINH1 showed favorable prognostic value for DFS in CRC patients. We speculate that SERPINH1 might promote not only the A-C process but also the development of CRC.