Bone sialoprotein, matrix metalloproteinase 2, and αvβ3 integrin in osteotropic cancer cell invasion

Background: Bone sialoprotein (BSP) interacts separately with both matrix metalloproteinase 2 (MMP-2) and integrin alpha(v)beta(3) and is overexpressed in many metastatic tumors. Its role in tumor biology, however, remains unclear. We investigated whether BSP enhances cancer cell invasiveness by forming a trimolecular complex with MMP-2 and cell-surface integrin alpha(v)beta(3). Methods: Invasiveness of breast, prostate, lung, and thyroid tumor cell lines was measured with a modified Boyden chamber assay. Binding and co-localization of BSP, MMP-2, and integrin alpha(v)beta(3) were investigated with immunoprecipitation and in situ hybridization. All statistical tests were two-sided. Results.. Treatment with BSP increased invasiveness of many breast, prostate, lung, and thyroid cancer cells through Matrigel in a dose-dependent manner. BSP at 50 nM increased the invasiveness of SW-579 thyroid cancer cells (95.2 units, 95% confidence interval [CI] = 90.4 to 100 units) by approximately 10-fold compared with that of untreated control SW-579 cells (9.1 units, 95% CI = 5.7 to 12.5 units) (P<.001). Addition of an inactive mutated BSP, in which BSP's integrin-binding RGD tripeptide was altered, or addition of integrin alpha(v)beta(3)-blocking antibodies resulted in invasiveness equivalent to that of untreated cells. Inhibiting cellular MMP-2 activity with chemical inhibitors or a specific antibody also blocked BSP-enhanced invasiveness. Osteopontin and dentin matrix protein 1, proteins related to BSP that also bind integrin alpha(v)beta(3) and form complexes with other MMPs (but not MMP-2), did not enhance invasiveness. Immunoprecipitation showed that a complex containing BSP, integrin alpha(v)beta(3), and MMP-2 formed in vitro. Addition of BSP increased the amount of MMP-2 bound by cells in an integrin-dependent fashion. Co-expression of BSP, integrin alpha(v)beta(3), and MMP-2 in papillary thyroid carcinoma cells was shown by in situ hybridization. Conclusion: Cancer cells appear to become more invasive when BSP forms a cell-surface trimolecular complex by linking MMP-2 to integrin alpha(v)beta(3). [J Natl Cancer Inst 2004;96:956-65]