MMP activity appears to play an important role in bone matrix turnover when prostate cancer cells are present in bone. Bone matrix turnover and metastatic tumor growth appear to be involved in a mutually supportive cycle that is disrupted by MMP inhibition.

Emerging evidence indicates that tumor-associated proteolytic remodeling of bone matrix may underlie the capacity of tumor cells to colonize and survive in the bone microenvironment. Of particular importance, urokinase-type plasminogen activator (uPA) has been shown to correlate with human prostate cancer (PC) metastasis. The importance of this protease may be related to its ability to initiate a proteolytic cascade, leading to the activation of multiple proteases and growth factors. Previously, we showed that maspin, a serine protease inhibitor, specifically inhibits PC-associated uPA and PC cell invasion and motility in vitro. In this article, we showed that maspin-expressing transfectant cells derived from PC cell line DU145 were inhibited in in vitro extracellular matrix and collagen degradation assays. To test the effect of tumor-associated maspin on PC-induced bone matrix remodeling and tumor growth, we injected the maspin-transfected DU145 cells into human fetal bone fragments, which were previously implanted in immunodeficient mice. These studies showed that maspin expression decreased tumor growth, reduced osteolysis, and decreased angiogenesis. Furthermore, the maspin-expressing tumors contained significant fibrosis and collagen staining, and exhibited a more glandular organization. These data represent evidence that maspin inhibits PC-induced bone matrix remodeling and induces PC glandular redifferentiation. These results support our current working hypothesis that maspin exerts its tumor suppressive role, at least in part, by blocking the pericellular uPA system and suggest that maspin may offer an opportunity to improve therapeutic intervention of bone metastasis.A ndrogen deprivation therapy has been the mainstay of treatment of metastatic prostate cancer (PC) for Ͼ50 years. Usually, this therapy produces tremendous tumor shrinkage and significant clinical improvement. However, the duration of response is limited and disease always recurs. Cytotoxic chemotherapy is commonly added, although this approach offers little chance for meaningful, long-term survival. Thus, new approaches are needed. The skeleton is the major target organ of metastasis in patients with PC, and bone metastasis is associated with poor survival. Can bone-targeted therapy improve survival? In a recent clinical trial (1), a therapeutic, bone-seeking radioisotope was added to standard chemotherapy in patients showing an initial chemotherapeutic response. Survival was prolonged in the patients receiving the bone-seeking radioisotope compared with those receiving chemotherapy alone, suggesting that targeting skeletal metastases is a promising approach in PC treatment.Clinicians have traditionally classified bone metastases as either osteolytic or osteoblastic (2). However, tumor deposits in bone usually contain both bone formation and bone degradation (3-8). A ''vicious cycle'' is created whereby metastatic tumor stimulates bone turnover and bone turnover promotes local tumor growth. To date, various molecules have been implicated ...

We previously characterized a purine-specific Na(+)-nucleoside cotransport system in bile canalicular membrane. The function of this transport system may be related to conserving nucleosides and preventing cholestasis. We report here the isolation of a cDNA encoding a Na(+)-dependent nucleoside transporter from rat liver using an expression cloning strategy. The substrate specificities and kinetic characteristics of the cloned cotransporter are consistent with the properties of the Na(+)-dependent, purine-selective nucleoside transporter in bile canalicular membranes. The nucleotide sequence predicts a protein of 659 amino acids (72 kDa) with 14 putative membrane-spanning domains. Northern blot analysis showed that the transcripts are present in liver and several other tissues. Data base searches indicate significant sequence similarity to the pyrimidine-selective nucleoside transporter (cNT1) of rat jejunum. Although these two subtypes of Na(+)-nucleoside cotransporter have different substrate specificities and tissue localizations, they are members of a single gene family.

The secretion of bile by the liver is primarily determined by the ability of the hepatocyte to transport bile acids into the bile canaliculus. A carrier-mediated process for the transport of taurocholate, the major bile acid in humans and rats, was previously demonstrated in canalicular membrane vesicles from rat liver. This process is driven by an outside-positive membrane potential that is, however, insufficient to explain the large bile acid concentration gradient between the hepatocyte and bile. In this study, we describe an ATP-dependent transport system for taurocholate in inside-out canalicular membrane vesicles from rat liver. Taurocholate is the major bile acid in humans and rats. Its uptake from plasma by hepatocytes utilizes a Na' cotransport system that requires Na',K+-ATPase for generation of a Na' gradient (1, 2). Two pathways for intracellular bile acid transport have been demonstrated. One involves microtubule-dependent vesicular transport from the Golgi apparatus to the bile canaliculus (3, 4). The other is the major physiologic pathway and involves direct transport to the canaliculus, possibly by cytosolic binding proteins (5). The secretion of bile acids into the canaliculus is a major determinant ofbile flow. Although the contents ofthe bile canaliculus have never been measured due to its inaccessibility, studies ofductal bile suggest that there is a large bile acid concentration gradient between the hepatocyte and canalicular contents (6, 7). Previous studies using vesicles selectively derived from the canalicular domain of hepatocyte plasma membrane revealed carrier-mediated transport of taurocholate and other bile acids that is driven by an outside-positive membrane potential (8, 9). However, the membrane potential is insufficient to explain the concentration gradients of bile acids that are postulated to occur across the canalicular membrane (6).We have described (10, 11) two distinct ATP-dependent transport systems in the canalicular membrane. One system utilizes P-glycoprotein, the product of the multidrugresistance gene, and transports mainly hydrophobic cations, such as daunomycin (10). The carrier protein for the second system has not been purified but its substrates are nonbile acid organic anions, such as bilirubin diglucuronide, oxidized glutathione (GSSG), and glutathione S-conjugates (11-13).The latter transport system is defective in TR-mutant rats, which have a phenotype that resembles the defect in patients with Dubin-Johnson syndrome (11,14). While studying these transport processes, an ATP-dependent transport of bile acids in canalicular membrane vesicles (CMVs) was detected and has been now characterized. (GSBSP) and dinitrophenyl glutathione (GSDNP) were synthesized nonenzymatically (16,17). Rabbit polyclonal antibody against the extracellular domain of -glutamyltransferase (y-GT) was kindly provided by Masayasu Inoue (Kumamoto University Medical School, Kumamoto, Japan). Bilirubin diglucuronide, which was purified by HPLC (18), was kindly provided by N. Roy-Cho...