Role of tumor cell membrane-bound serine proteases in tumor-induced target cytolysis.
The tumor-induced marrow and red blood cell cytolysis assays have been used to explore the mechanism of cancer cell destruction of normal cells. Previously, we suggested that tumor-induced cytolysis was caused by tumor cell membrane-bound serine proteases. In this study, we have shown that concentrations of the broad-spectrum serine protease inhibitor diisopropylfluorophosphate that did not inhibit tumor cell DNA and protein synthesis completely abrogated tumor-induced red blood cell cytolysis. In addition, tumor cell membranes isolated by differential and sucrose density gradient centrifugation and characterized by electron microscopy and enzyme marker analysis were cytolytic for rat 59Fe-labeled red blood cells. The specific activity expressed as release index (%) per microgram of protein was 1.620 for the tumor cell membrane preparations as compared to 0.002 for intact Walker 256 tumor cells. Tumor cell membranes solubilized in Triton X-100 had activity in the p-toluenesulfonyl-L-arginine methyl ester assay for trypsin-like enzymes and the N-benzoyl-L-tyrosine ethyl ester assay for chymotrypsin-like enzymes. The enzyme activities demonstrated in these assays could be inhibited by N-alpha-p-tosyl-L-lysine chloromethyl ketone HCl and L-1-tosylamide-alpha-phenyl-ethyl chloromethyl ketone, respectively. Using [3H]diisopropylfluorophosphate affinity labeling of the tumor cell membrane proteins followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, we have identified membrane-bound serine protease(s) that appear to be responsible for tumor-induced marrow and red blood cell cytolysis.
D000818:Animals; D001854:Bone Marrow Cells; D002279:Carcinoma 256, Walker / Q000201:enzymology* / Q000648:ultrastructure; D002462:Cell Membrane / Q000201:enzymology*; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D003602:Cytotoxicity, Immunologic / Q000187:drug effects; D010450:Endopeptidases / Q000378:metabolism*; D006461:Hemolysis; D007531:Isoflurophate / Q000494:pharmacology; D008297:Male; D051381:Rats; D012697:Serine Endopeptidases
D007531:Isoflurophate; D010450:Endopeptidases; D012697:Serine Endopeptidases