LATERALLY ASSOCIATED PROTEINS MODULATE A6 INTEGRIN CLEAVAGE, A PERMISSIVE PROCESS UTILIZED DURING CANCER METASTASIS

Abstract

Expression of A6 integrin, a laminin receptor, on tumor cell surfaces is associated with reduced patient survival and increased metastasis in a variety of tumors. In prostate cancer, tumor extra capsular escape occurs in part via laminin coated nerves and vascular dissemination, resulting in clinically significant bone metastases. Our group previously identified a novel form of A6 integrin, called A6p, generated by urokinase (uPA) dependent cleavage of the laminin binding domain from the tumor cell surface. Although functional consequences of cleavage have been characterized, little is known about how this process is regulated.Regulation of uPA mediated cleavage was identified by a laterally interacting protein expressed on the cellular surface. A direct interaction between the urokinase receptor (uPAR) and A6 integrin was characterized. This direct interaction was responsible for the extracellular cleavage of A6. Transient knockout of A3 integrin, a known interacting partner of uPAR, increased uPAR association with A6 integrin and enhanced production of A6p. Analysis of tissue obtained from human prostate tumors confirmed uPAR and A6 integrin expression in invasive disease. Taken together the results demonstrate a novel and dynamic role for uPAR regulation of integrin dependent adhesion through lateral interaction.Using the known conformation sensitivity of integrin function to I determined if engagement of the extracellular domain by antibodies inhibited integrin cleavage and the extravasation step of metastasis. Both endogenous and inducible levels of A6p were inhibited by engaging the extracellular domain of A6 with monoclonal antibody J8H. J8H inhibited tumor cell invasion through Matrigel. A SCID mouse model of extravasation and bone metastasis produced detectable, progressive osteolytic lesions within three weeks of intracardiac injections. Injection of tumor cells, pre-treated with J8H, delayed the appearance of metastases. Validation of the A6 cleavage effect on extravasation was confirmed through a genetic approach using tumor cells transfected with uncleavable A6 integrin. Uncleavable A6 integrin significantly delayed the onset and progression of osseous metastases out to 6 weeks post injection. The results suggest that A6 integrin cleavage permits extravasation of human prostate cancer cells from circulation to bone and can be manipulated to prevent metastasis.