Panoply™ Human MMP2 Knockdown Stable Cell Line

The role of matrix metalloproteinase-2 (MMP-2) in tumor cell migration has been extensively studied. However, the characterization and role of MMP-2 in clinical samples of metastatic colorectal cancer (CRC) remain poorly understood. To elucidate the proteomic signaling alterations during MMP-2-induced cancer progression, researchers analyzed plasma proteomes from CRC patients (according to disease progression stage), secretomes from MMP-2 knockdown HCT116 cancer cells, and publicly available CRC tissue proteomic data. A comprehensive multilayer proteomic analysis revealed a protein cluster containing epithelial-mesenchymal transition (EMT)-related proteins (such as CD9 integrin) and MMP-2. Proteins within this cluster are regulated by MMP-2 and show significantly increased expression in both tissues and plasma as the disease progresses from TNM stage I to stage II. Furthermore, their tissue proteomic analysis of CRC patients also revealed a potential association between MMP-2 upregulation and activation of the focal adhesion kinase signaling pathway. These studies indicate that the high invasiveness of metastatic CRC is due to increased secretion of the MMP-2 and CD9-integrin complex mediated by FAK signaling pathway activation.

Here, researchers constructed an MMP2 knockdown HCT116 cell line and confirmed this through quantitative PCR analysis. Compared with control cells, MMP2 knockdown cells exhibited significantly reduced migration ability (Figure 1A and 1B). To comprehensively characterize proteins secreted into conditioned medium, researchers performed SILAC-based quantitative analysis on the secretomes of both control and MMP2 knockdown cell lines (Figure 1C). A total of 3196 human-derived proteins were identified, including 99 bovine proteins derived from fetal bovine serum (FBS) and 13 proteins of unknown origin. Then, researchers performed Gene Ontology Cellular Compartment (GOCC) enrichment analysis on quantifiable proteins from all six secretome datasets to assess the quality of the secretomes. Significantly enriched GOCC entries with an enrichment factor greater than 1 in Fisher's exact test were closely associated with extracellular localization. GOCC entries with an enrichment factor less than 1 were associated with cytoplasmic localization (Figure 1D). When the logarithmically transformed protein ratios were plotted as histograms and volcano plots, the quantitative results showed a typical normal distribution (Figure 1E). A total of 164 differentially expressed proteins (DEPs) were identified. Among them, 142 proteins were downregulated and 22 proteins were upregulated (Figure 1E). Analysis showed that "cellular movement," "cell to cell signaling" and "cell death" were the most significantly enriched annotations, and these activities were predicted to be inhibited by MMP2 knockdown (Figure 1F).

Figure 1. Analysis of differentially secreted proteins in MMP2 knockdown HCT116 cells. (Kwon Y, et al., 2021)