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Role of Glycosidases in Human Ovarian Carcinoma Cell Mediated Degradation of Subendothelial Extracellular Matrix¹

Department of Experimental Therapeutics [M. J. N., M. S., R. J. B.] and Gynecologic Research Department [R. M., K. M.], Roswell Park Memorial Institute, Buffalo, New York 14263, and Department of Gynecology and Obstetrics, Buffalo General Hospital, Buffalo, New York 14203 [K. C.]

Penetration of the extracellular matrix (ECM) by tumor cells, an event which occurs at various stages of the metastatic process, involves tumor cell glycosidase mediated hydrolysis of proteoglycans (PG). Recently, we observed that human ovarian carcinoma cell lines (HOCC) derived from primary tumors, peritoneal effusions, and distant metastases possess a varying ability to degrade radiolabeled PG of the ECM, while normal cells (human mesothelial cells or ovarian fibroblasts) fail to do so. To determine whether a quantitative relationship exists between glycosidase activity and degradation of ECM, both intracellular and extracellular glycosidase activities were measured for HOCC and normal cell lines. No relationship was found between intracellular glycosidase activities and the ability of cells to degrade ECM. However, a correlation was observed between extracellular or secretory glycosidase activities and HOCC mediated ECM degradation. In particular, a 5–8-fold increase, as compared to normal cells, was observed for HOCC extracellular ß-N-acetylglucosaminidase (EC 3.2.2.30) activity. The accumulation or secretion of this enzyme from HOCC into culture medium was found to be time dependent and not related to intracellular levels. Purified hexosaminidase derived from invasive HOCC was able to hydrolyze [³H]-glucosamine radiolabeled ECM (up to 30% radiolabel) and resulted in the cumulative release of free [³H]-N-acetylglucosamine. This enzyme mediated hydrolysis could be completely prevented with 2-acetamido-2-deoxy-l,5-D-gluconolactone, a competitive inhibitor (K_i 10^-6 M). Finally, HOCC mediated degradation of radiolabeled ECM was discerned to be dependent upon active hexosaminidase action, since tumor cell mediated degradation of ECM could be inhibited by up to 60% in the presence of this synthetic competitive inhibitor. In summary, these studies indicate a strong association between HOCC solubilization of glycoconjugates present in the ECM and extracellular levels of hexosaminidase.

¹ This research was partially supported by the National Cancer Institute (Grants CA-42898 and CA-13038) and the NIH (Grant GM-31425).

² To whom requests for reprints should be addresed, at Department of Experimental Therapeutics, Roswell Park Memorial Institute, 666 Elm Street, Buffalo, NY 14263.

Received 2/13/87. Revised 6/ 8/87. Accepted 6/ 9/87.