FUNCTIONAL-HETEROGENEITY OF CD44 MOLECULES IN OVARIAN-CANCER CELL-LINES

We have previously shown that CD44 partly mediates ovarian cancer cell attachment to peritoneal mesothelium through recognition of mesotheli al-associated hyaluronate. CD44 is a major receptor for hyaluronate an d exists as a standard 90-180-kDa form (CD44H), as well as several hig her molecular mass variant forms produced by alternative splicing. To determine whether functional differences exist between CD44H and its v ariants we have investigated the relationship between CD44 isoform exp ression and mesothelial adhesion in 12 ovarian cancer cell lines. Eigh t lines were CD44 positive (range, 83-94%) and demonstrated significan t binding to mesothelium and hyaluronate, whereas two lines showed red uced CD44 levels (3-13%) and demonstrated decreased binding. Interesti ngly, two other lines (OVC-3 and SW626) expressed CD44 in the majority of cells (>93%) and yet bound weakly to mesothelium. Mean linear fluo rescence intensity of CD44 expressed by OVC-3 and SW626 cells was appr oximately one-half that of strongly binding cell lines, suggesting tha t the ability to adhere may be partly related to CD44 surface density. However, immunoprecipitation and immunoblot analyses revealed that st andard CD44H represented only 23-31% of total CD44 in weakly binding c ells, with the majority of species being comprised of CD44 variants. I ndirect immunofluorescence of OVC-3 and SW626 cells confirmed the pres ence of CD44 variants containing exons v3, v6, and v9. In contrast, CD 44H represented the majority (75-86%) of total CD44 expressed by stron gly binding cell lines such as CAOV-3 and UPN36T. Transfection of CD44 H cDNA into weakly binding OVC-3 cells restored significant mesothelia l binding which was partly blocked by anti-CD44 antibody. These data s uggest that the expression of CD44 is necessary but not sufficient for mediating attachment of ovarian cancer cells to mesothelium. Although CD44 variants may constitute the major CD44 species in certain ovaria n cancer cell lines, it appears that these CD44 species are not always capable of mediating significant binding to mesothelium or hyaluronat e. Rather, an adequate level of CD44H is the critical determinant of b inding in this system. The role of CD44 variants in the process of ova rian cancer metastasis will require further investigation.