Multi-layered proteomic analysis for colorectal cancer

Matrix metalloproteinase-2 (MMP-2) plays a role in cell migration and invasion that involve degradation of extracellular matrix proteins as well as vascular remodeling. Increased expression of MMP-2 has often been observed in the tumors of colorectal cancer (CRC) patients. In order to unveil the perturbed proteomic signal during MMP-2-induced cancer progression, we analyzed MMP-2 dependent secretome change in HCT116 cancer cells by stable isotopic labeling with amino acids in cell culture (SILAC), together with the plasma proteome profile of CRC patients by label-free quantification according to disease progression from TNM stage I to IV. We also analyzed public database of CRC tissue proteome deposited by Clinical Proteomic Tumor Analysis Consortium (NCI/NIH). As a strategy for integrating the multi-layered proteomes, we first performed unsupervised hierarchical clustering on plasma and tissue proteomes according to TNM stage, and then Fisher’s exact test to find out which clusters were significantly represented by the secretome decreased by more than 1.5-fold upon MMP-2 knock down. The resultant cluster included CD9-ITGB1-ITGA7 protein complex as well as MMP-2. The proteins included in this cluster were mostly related to cell-cell communications and cell motility, and showed remarkable increased expression in tissue and plasma upon disease progression from TNM I to II. MMP-2 induced activation of FAK phospho-signaling in an activity-dependent manner. With the comparative and integrative multi-layered proteomic analysis, we suggest that high invasiveness featured in the disease progression of colorectal cancer with increased secretion of MMP-2 is because MMP-2 activates FAK signaling through CD9-integrin complex and consequently induces positive feedback of MMP-2 upregulation.