Peritoneal Implantation Human Ovarian Cancer Xenograft in Nude Mice: A Novel Role Inhibition of CD44 Limits Intra-Abdominal Spread of a In Vivo

toneal mesothelium predominantly express the 90-kDa CD44H mol ecule, whereas ovarian cancer lines that bind poorly to mesothelium express either absent or relatively low levels of CD44H (2, 3). Furthermore, transfection of weakly binding ovarian cancer cells with CD44H cDNA is capable of restoring their ability to bind to mesothe hum in vitro (3). Finally, treatment of mesothelial monolayers with hyaluronidase abolishes the CD44 component of ovarian cancer cell binding (2). Taken together, these date suggest a possible role for the CD44H molecule in the process of ovarian cancer metastasis. The in vitro binding assay that we developed to quantitate ovarian cancer cell attachment to mesothelium is performed by allowing peritoneal mesothelial cells obtained from ascitic fluid to grow to confluence in microtiter wells under the stimulatory effects of EGF3 and hydrocortisone (2†" 4). Because it is not known whether mesothe hal cells grown in this fashion are truly representative of the pento neal mesotheial surface, we developed an animal model of ovarian cancer cell metastasis in order to investigate the role of CD44 in vivo and to obtain preclinical evidence in support of strategies designed to inhibit implantation. We now present the results of in vivo expen ments that demonstrate an important role for the CD44H molecule in the implantationof human ovarian cancer cells within the murine peritoneal cavity. Source of Reagents and Antibodies. Munne monoclonalantibodiesused in the characterization of ovarian cancer cell lines and for i.p. treatment are University, Chicago, IL). As expected, both anti-Dl44 and anti-DF3 are incapable of inhibiting ovarian cancer cell binding to mesothelium in vitro over a concentration range of 1†" 50 @Wml (data not shown; Refs. 2 and 3). Anti CD44 antibody clone 515 has been previously shown to neutralize CD44-mediated binding of cells to mesothelium and to hyaluronic acid-coated wells in vitro (2, 3). This antibody does not induce tumor cell clumping of the 36M2 humanovariancancercell line (describedbelow) in suspensioncultureunder conditions that prevent plastic adherence. The use of these antibodies in indirect immunofluorescence analysis by flow cytometry has been described previously (3). All antibodies were affinity-purified using Affi-Gel protein A