Increased production and secretion of HGF alpha-chain and an antagonistic HGF fragment in a human breast cancer progression model

Invasive human breast carcinomas frequently coexpress increased hepatocyte growth factor (HGF) and its receptor Met, suggesting that establishment of an autocrine HGF loop is important in malignant disease. This study examines the expression patterns of HGF and Met activation during tumorigenesis and metastasis using a MCF10A-based model of Ha-Ras-induced human breast cancer progression. Deregulation of cadherin-based cell-cell adhesions, decreased expression of cytokeratins 8/18 and increased activity of matrix metalloproteinases such as MMP-2 occurs in premalignant and malignant (metastatic) cell lines compared to the parental nonmalignant cell line. Compared to the benign parent cell line, premalignant and malignant cell lines exhibit increased secretion of full length HGF alpha-chain and elevated Met tyrosine phosphorylation in complete medium. Interestingly, the premalignant and malignant cells also secrete a similar to 55 kDa HGF fragment. Epitope mapping of the similar to 55 kDa HGF fragment supports the presence of the N-terminal domain of the HGF alpha-chain with a truncation in the C-terminal domain. The similar to 55 kDa HGF fragment shows mobility in SDS-PAGE faster than HGF alpha-chain, but slightly slower than NK4, a previously established full antagonist of HGF. The separated similar to 55 kDa HGF fragment binds to an immobilized Met-IgG fusion protein, and inhibits both HGF/ Met-IgG binding and HGF-induced Met-tyrosine phosphorylation. These results are the first demonstration of an antagonistic similar to 55 kDa HGF fragment secreted during breast carcinoma pro gression, which may have a negative regulatory effect on HGF signaling in premalignant breast epithelial cells. (C) 2009 UICC