Mapping Proteolytic Processing in the Secretome of Gastric Cancer-Associated Myofibroblasts Reveals Activation of MMP-1, MMP-2, and MMP‑3
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Abstract
Cancer
progression involves changes in extracellular proteolysis,
but the contribution of stromal cell secretomes to the cancer degradome
remains uncertain. We have now defined the secretome of a specific
stromal cell type, the myofibroblast, in gastric cancer and its modification
by proteolysis. SILAC labeling and COFRADIC isolation of methionine
containing peptides allowed us to quantify differences in gastric
cancer-derived myofibroblasts compared with myofibroblasts from adjacent
tissue, revealing increased abundance of several proteases in cancer
myofibroblasts including matrix metalloproteinases (MMP)-1 and -3.
Moreover, N-terminal COFRADIC analysis identified cancer-restricted
proteolytic cleavages, including liberation of the active forms of
MMP-1, -2, and -3 from their inactive precursors. In vivo imaging
confirmed increased MMP activity when gastric cancer cells were xenografted
in mice together with gastric cancer myofibroblasts. Western blot
and enzyme activity assays confirmed increased MMP-1, -2, and -3 activity
in cancer myofibroblasts, and cancer cell migration assays indicated
stimulation by MMP-1, -2, and -3 in cancer-associated myofibroblast
media. Thus, cancer-derived myofibroblasts differ from their normal
counterparts by increased production and activation of MMP-1, -2,
and -3, and this may contribute to the remodelling of the cancer cell
microenvironment