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Diversity of Human Pancreatic Cancer Cell Proteinases: Role of Cell Membrane Metalloproteinases in Collagenolysis and Cytolysis¹

Department of Medicine and Research, Veterans Administration Medical Center, Northport, New York 11768, and School of Medicine, Health Science Center, State University of New York at Stony Brook, Stony Brook, New York 11794

In this study we have examined the tissue-destructive proteinases of human pancreatic ductal cancer cel lines derived initially from xenogenic transplants. Cancer cell organelles were isolated following nitrogen cavitation using sucrose density gradient centrifugation. Serine, cysteine, and metalloproteinases were assayed using radiolabeled protein and synthetic substrates. Tumor-induced RBC lysis was quantitated by measuring the release of isotope from ⁵⁹Fe-labeled RBCs co-cultivated with tumor cells or subcellular fractions. Enzyme inhibitors with specificity toward different classes of proteinases were used in the above assays to categorize the enzymes responsible for substrate degradation. Results indicated that intact pancreatic cancer cells (RWP-1 and RWP-2 cell lines), cell homogenate, and cytosol contain proteinases which were able to degrade [³H]collagen (type I) and [³H] gelatin and lyse normal RBCs. Cancer cell membrane fractions were enriched in collagenolytic, gelatinolytic, and cytolytic activities which could be abrogated by EDTA but not by inhibitors of serine or cysteine proteinases, which indicates that metalloproteinases are the active enzymes in these assays. Although plasminogen activator and cysteine proteinases were also enriched in the tumor cell membranes, these activities were not required for collagen degradation or cytolysis. We conclude that human cancer cell membrane proteinases are advantageously situated to facilitate damage to surrounding normal tissues.

¹ Supported by Merit Review Funds from the Veterans Administration.

² To whom requests for reprints should be addressed.

Received 5/24/85. Revised 8/ 8/85. Accepted 9/ 6/85.