Abstract 2378: The interaction of epithelial α5β1 integrin and stromal fibronectin is a candidate seed-and-soil mechanism in prostate cancer and bone metastases

Background: Morbidity and mortality from prostate cancer relates principally to its unique proclivity to disseminate with near-exclusivity within the hematopoietic bone marrow microenvironment. Prostate cancer cells compete for the bone marrow niche with CD45+ hematopoietic stem cells (Shiozawa, 2011). Several lines of evidence have implicated mesenchymal stem cells (MSCs) as key architects of the hematopoietic niche (Nakamura-Ishizu, 2013). The molecular mechanisms that determine the tropism of prostate cancer cells to the MSC-regulated niche are undetermined. We hypothesized that specific tropic interactions between human MSCs and prostate cancer cells contribute toward the efficient colonization of the bone marrow. Methods and Results: Bone-derived prostate cancer cells (PC-3) exhibit a strong migratory response when placed in co-culture with human MSCs. Serum-free conditioned media from MSCs similarly induces adhesion, migration and invasion of PC-3 cells, but not proliferation. Heat-inactivation and trypsin digest suggested the bioactive principle was a protein. Size-fractionation by spin-columns determined the bioactivity to reside in the >100kD fraction. Size-exclusion chromatography (SEC) recovered peak activity in a 440kD fraction containing thrombospondin-1 (TSP1). TSP1 put-back assays did not reproduce bioactivity. Further purification of the TSP-1 containing high-molecular weight fraction of the MSC secretome with heparin-affinity chromatography recovered bioactivity with highly restricted bands on polyacrylamide gel electrophoresis, determined by mass spectroscopy to be fragments of fibronectin (FN). Put-back experiments with FN recovered bioactivity. Monospecific antibodies to FN blocked adhesion. Neutralizing monoclonal antibodies to FN receptors, α5 and β1 integrins, but not α4, αv, α6 or β3 integrins blocked the adhesion and migration of prostate cancer cells cells to the MSC secretome and FN. Conclusions: The affinity of bone-derived human prostate cancer cells to the MSC secretome is mediated by fibronectin acting via the α5 and β1 integrins. Human prostate cancer cells isolated from clinical bone marrow specimens express α5β1 integrin in high-frequency (Putz, 1999) and fibronectin is richly expressed in the bone microenvironment (Van der Welde-Zimmermann 1997). The FN-α5β1 interaction requires translational study as a candidate molecular target for therapy, specific to prostate cancer and hematopoietic marrow interaction, the lethal phe