ACTIVATION OF CATHEPSIN-B, SECRETED BY A COLORECTAL-CANCER CELL-LINE REQUIRES LOW PH AND IS MEDIATED BY CATHEPSIN-D

The aim of our study was to identify changes in secreted procathepsin B levels in a model of the human colorectal adenoma to carcinoma seque nce and to determine the factors required for its extracellular activa tion. Conversion of the non-tumorigenic adenoma-derived cell line PC/A A to a highly tumorigenic phenotype (designated AA/C1/SB10/M) was asso ciated with an 8-fold increase in the presence of the preform of cathe psin B in 24 hr conditioned serum-free medium (SFM). In addition, matu re enzyme was only detected in the cell lines of this model with incre ased malignant potential. This is in agreement with the findings of a previous study, in which mature cathepsin B was only present in the 24 hr conditioned SFM of cancer-derived cell lines and not in SFM from a denoma-derived cell lines. Having demonstrated a reduction in the pH o f conditioned medium from cell lines with increased malignant potentia l, we used a range of specific proteinase inhibitors to show that an a spartyl proteinase was involved in the initial activation of procathep sin B. Consistent with this finding, we subsequently demonstrated an i ncreased secretion of the aspartyl proteinase cathepsin D in the mediu m of the AA/C1/SB10/M adenocarcinoma cells compared with the non-tumor igenic AA/C1 cell line. Therefore, the presence of mature cathepsin B in the conditioned medium of the more malignant cell lines coincided w ith a reduction in pH and an increase in the amount of cathepsin D sec reted. Data from the human colorectal derived adenoma to carcinoma seq uence indicate that an in vivo mechanism may exist that, dependent on the simultaneous presence of both a tumour-generated acidic extracellu lar environment and an elevated secretion of procathepsin D, could res ult in the activation of latent procathepsin outside the cell. (C) 199 6 Wiley-Liss, Inc.