Identifying the key players in osteopontin/alpha(v)beta(3)-mediated migration and invasion of metastatic prostate cancer cells

Abstract

Advanced stages of prostatic carcinoma have a high incidence of metastases to the bones. Our research examines the mechanisms that can facilitate local invasion of prostate cancer (PC) cells inside the extracellular matrix (ECM) of bone. PC3 is a stable cancer cell line derived from bony metastasis of prostate cancer. Osteopontin (OPN), which is an autocrine motility factor secreted by both osteoclasts and osteoblasts, is an important component of the ECM in bone. PC3 cells degrade ECM by secretion of matrix metalloproteinases (MMP-s), primarily through secretion of active MMP-9. Experimental approaches that employed both an over expression and knockdown of endogenous OPN in PC3 cells demonstrated that OPN regulates migration of PC3 cells by increasing active MMP-9 secretion. OPN-mediated upregulation of MMP-9 expression and secretion occurs in response to activation of the hyaluronan receptor CD44. Bisphosphonate-mediated inhibition of Rho kinase, which is an upstream activator of CD44, attenuated the stimulatory effects of OPN on both migration and secretion of MMP-9. In addition to a standard form, multiple CD44 isoforms exist in several cell systems. We have identified CD44 isoforms, which exist in PC3 cells. At the cellular level, migration is a consequence of increased MMP activity in conjunction with changes in the actin cytoskeleton. PC3 cells exhibited punctate evaginations (similar to invadopodia), enriched in actin, which could degrade the underlying gelatin matrix. MMP-9 activity was essential for the invasiveness of invadopodia, but had no role in their formation. The WAVE protein family member WASP was observed to have a role in actin polymerization in PC3 cells. Active MMP-9 was demonstrated to be associated with WASP, thereby pointing out to a mechanistic possibility of WASP's association to MMP-9, with or without an intervening adaptor protein. OPN increased the incidence of invadopodia formation as well as WASP co-localization with invadopodia. The contiguous activation from OPN, CD44, and MMP-9 to WASP and dynamic actin changes provides deeper insight into OPN-mediated PC3 cell invasion and paves the way for therapeutically targeting this pathway in future.