Tai-Shan Cheng

Curcumin-targeting Pericellular Serine Protease Matriptase Role in Suppression of Prostate Cancer Cell Invasion, Tumor Growth and Metastasis

Curcumin has been shown to possess potent chemopreventive and antitumor effects on prostate cancer. However, the molecular mechanism involved in curcumins ability to suppress prostate cancer cell invasion, tumor growth, and metastasis is not yet well understood. In this study, we have shown that curcumin can suppress epidermal growth factor (EGF)- stimulated and heregulin-stimulated PC-3 cell invasion, as well as androgen-induced LNCaP cell invasion. Curcumin treatment significantly resulted in reduced matrix metalloproteinase 9 activity and downregulation of cellular matriptase, a membrane-anchored serine protease with oncogenic roles in tumor formation and invasion. Our data further show that curcumin is able to inhibit the induction effects of androgens and EGF on matriptase activation, as well as to reduce the activated levels of matriptase after its overexpression, thus suggesting that curcumin may interrupt diverse signal pathways to block the protease. Furthermore, the reduction of activated matriptase in cells by curcumin was also partly due to curcumins effect on promoting the shedding of matriptase into an extracellular environment, but not via altering matriptase gene expression. In addition, curcumin significantly suppressed the invasive ability of prostate cancer cells induced by matriptase overexpression. In xenograft model, curcumin not only inhibits prostate cancer tumor growth and metastasis but also downregulates matriptase activity in vivo. Overall, the data indicate that curcumin exhibits a suppressive effect on prostate cancer cell invasion, tumor growth, and metastasis, at least in part via downregulating matriptase function. Cancer Prev Res; 6(5); 495–505. ©2013 AACR.

HAI-2 suppresses the invasive growth and metastasis of prostate cancer through regulation of matriptase

Dysregulation of cell surface proteolysis has been strongly implicated in tumorigenicity and metastasis. In this study, we delineated the role of hepatocyte growth factor activator inhibitor-2 (HAI-2) in prostate cancer (PCa) cell migration, invasion, tumorigenicity and metastasis using a human PCa progression model (103E, N1, and N2 cells) and xenograft models. N1 and N2 cells were established through serial intraprostatic propagation of 103E human PCa cells and isolation of the metastatic cells from nearby lymph nodes. The invasion capability of these cells was revealed to gradually increase throughout the serial isolations (103E<N1<N2). In this series of cells, the expression of HAI-2 but not HAI-1 was significantly decreased throughout the progression and occurred in parallel with increased activation of matriptase. The expression level and activity of matriptase increased whereas the HAI-2 protein level decreased over the course of orthotopic tumor growth in mice, which was consistent with the immunohistochemical profiles of matriptase and HAI-2 in archival PCa specimens. Knockdown of matriptase reduced the PCa cell invasion induced by HAI-2 knockdown. HAI-2 overexpression or matriptase silencing in N2 cells downregulated matriptase activity and significantly decreased tumorigenicity and metastatic capability in orthotopically xenografted mice. These results suggest that during the progression of human PCa, matriptase activity is primarily controlled by HAI-2 expression. The imbalance between HAI-2 and matriptase expression led to matriptase activation, thereby increasing cell migration, invasion, tumorigenicity and metastasis.

Matriptase Is Involved in ErbB-2-Induced Prostate Cancer Cell Invasion

Deregulation of both ErbB-2 signaling and matriptase activity has been associated with human prostate cancer (PCa) progression. In this communication, we investigated the roles of both ErbB-2 signaling in matriptase zymogen activation and matriptase in ErbB-2-induced PCa malignancy. In a human PCa cell progression model, we observed that advanced PCa C-81 LNCaP cells exhibited an aggressive phenotype with increased cell migration and invasion capacity; these cells concurrently showed both enhanced ErbB-2 phosphorylation and increased matriptase zymogen activation compared with parental C-33 LNCaP cells. Moreover, ErbB2 activation, both ligand-dependent (eg, epidermal growth factor treatment) and ligand-independent (eg, overexpression), was able to induce matriptase zymogen activation in this cell line. Inhibition of ErbB-2 activity by either the specific inhibitor, AG825, in epidermal growth factor-treated C-33 LNCaP cells or ErbB-2 knockdown in C-81 LNCaP cells, reduced matriptase activation. These observations were confirmed by similar studies using both DU145 and PC3 cells. Together, these data suggest that ErbB-2 signaling plays an important role in matriptase zymogen activation. ErbB-2-enhanced matriptase activation was suppressed by a phosphatidylinositol 3-kinase inhibitor (ie, LY294002) but not by a MEK inhibitor (ie, PD98059). Suppression of matriptase expression by small hairpin RNA knockdown in ErbB-2-overexpressing LNCaP cells dramatically suppressed cancer cell invasion. In summary, our data indicate that ErbB-2 signaling via the phosphatidylinositol 3-kinase pathway results in up-regulated matriptase zymogen activity, which contributes to PCa cell invasion.

Persistent elevation of hepatocyte growth factor activator inhibitors in cholangiopathies affects liver fibrosis and differentiation

Alteration of cell surface proteolysis has been proposed to play a role in liver fibrosis, a grave complication of biliary atresia (BA). In this study we investigated the roles of hepatocyte growth factor activator inhibitor (HAI)‐1 and ‐2 in the progression of BA. The expression levels of HAI‐1 and ‐2 were significantly increased in BA livers compared with those in neonatal hepatitis and correlated with disease progression. In BA livers, HAI‐1 and ‐2 were coexpressed in cells involved in ductular reactions. In other selective cholangiopathies, ductular cells positive for HAI‐1 or HAI‐2 also increased in number. Inflammatory cytokines, growth factors, and bile acids differentially up‐regulated expression of HAI‐1 and ‐2 transcripts in fetal liver cells and this induction could be antagonized by a cyclooxygenase‐2 inhibitor. Conditioned media from cell lines stably overexpressing HAI‐1 or HAI‐2 enhanced the fibrogenic activity of portal fibroblasts and stellate cells, suggesting that both proteins might be involved in liver fibrosis. Because HAI‐1 and ‐2 colocalized in ductular reactions sharing similar features to those observed during normal liver development, we sought to investigate the role of HAI‐1 and ‐2 in cholangiopathies by exploring their functions in fetal liver cells. Knockdown of HAI‐1 or HAI‐2 promoted bidirectional differentiation of hepatoblast‐derived cells. In addition, we showed that the hepatocyte growth factor activator, mitogen‐activated protein kinase kinase 1, and phosphatidylinositol 3‐kinase signaling pathways were involved in hepatic differentiation enhanced by HAI‐2 knockdown. Conclusion: HAI‐1 and ‐2 are overexpressed in the liver in cholangiopathies with ductular reactions and are possibly involved in liver fibrosis and hepatic differentiation; they could be investigated as disease markers and potential therapeutic targets. (Hepatology 2012)

Ketamine Increases Permeability and Alters Epithelial Phenotype of Renal Distal Tubular Cells via a GSK‐3β‐Dependent Mechanism

Ketamine, a dissociative anesthetic, is misused and abused worldwide as an illegal recreational drug. In addition to its neuropathic toxicity, ketamine abuse has numerous effects, including renal failure; however, the underlying mechanism is poorly understood. The process called epithelial phenotypic changes (EPCs) causes the loss of cell–cell adhesion and cell polarity in renal diseases, as well as the acquisition of migratory and invasive properties. Madin–Darby canine kidney cells, an in vitro cell model, were subjected to experimental manipulation to investigate whether ketamine could promote EPCs. Our data showed that ketamine dramatically decreased transepithelial electrical resistance and increased paracellular permeability and junction disruption, which were coupled to decreased levels of apical junctional proteins (ZO‐1, occludin, and E‐cadherin). Consistent with the downregulation of epithelial markers, the mesenchymal markers N‐cadherin, fibronectin, and vimentin were markedly upregulated following ketamine stimulation. Of the E‐cadherin repressor complexes tested, the mRNA levels of Snail, Slug, Twist, and ZEB1 were elevated. Moreover, ketamine significantly enhanced migration and invasion. Ketamine‐mediated changes were at least partly caused by the inhibition of GSK‐3β activity through Ser‐9 phosphorylation by the PI3K/Akt pathway. Inhibiting PI3K/Akt with LY294002 reactivated GSK‐3β and suppressed ketamine‐enhanced permeability, EPCs, and motility. These findings were recapitulated by the inactivation of GSK‐3β using the inhibitor 3F8. Taken together, these results provide evidence that ketamine induces renal distal tubular EPCs through the downregulation of several junction proteins, the upregulation of mesenchymal markers, the activation of Akt, and the inactivation of GSK‐3β. J. Cell. Biochem. 117: 881–893, 2016.

The Kunitz Domain I of Hepatocyte Growth Factor Activator Inhibitor-2 Inhibits Matriptase Activity and Invasive Ability of Human Prostate Cancer Cells

Dysregulation of pericellular proteolysis is often required for tumor invasion and cancer progression. It has been shown that down-regulation of hepatocyte growth factor activator inhibitor-2 (HAI-2) results in activation of matriptase (a membrane-anchored serine protease), human prostate cancer cell motility and tumor growth. In this study, we further characterized if HAI-2 was a cognate inhibitor for matriptase and identified which Kunitz domain of HAI-2 was required for inhibiting matriptase and human prostate cancer cell motility. Our results show that HAI-2 overexpression suppressed matriptase-induced prostate cancer cell motility. We demonstrate that HAI-2 interacts with matriptase on cell surface and inhibits matriptase proteolytic activity. Moreover, cellular HAI-2 harnesses its Kunitz domain 1 (KD1) to inhibit matriptase activation and prostate cancer cell motility although recombinant KD1 and KD2 of HAI-2 both show an inhibitory activity and interaction with matriptase protease domain. The results together indicate that HAI-2 is a cognate inhibitor of matriptase, and KD1 of HAI-2 plays a major role in the inhibition of cellular matritptase activation as well as human prostate cancer invasion.

Abstract 2000: Curcumin prevents human prostate cancer cell invasion by inhibiting of matriptase activity

Curcumin has been shown with potent chemopreventive and antitumor effects on prostate cancer. However, the molecular mechanism how curcumin suppresses prostate cancer cell migration and invasion is not well understood. In this study we showed that curcumin could significantly suppress prostate cancer PC3 cell migration and invasion. Moreover, our data also showed that curcumin was able to suppress EGF- or heregulin-stimulated PC-3 cell invasion, as well as androgen-induced LNCaP cell invasion. Curcumin treatment resulted in not only significant reductions on MMP-9 and MMP-2 activities, but also on activated level of matriptase which is a membrane-bound serine protease and has been proposed to play important roles in tumor invasion and malignancy. Matriptase is synthesized as a full length zymogen, which is rapidly stepped into the maturation process, mature form through a cleavage within the SEA domain to from a latent matriptase. Our data further showed that curcumin can inhibit the latent matriptase formation in the maturation process to affect matriptase activity. Furthermore, the reduction of activated matriptase by curcumin was also partly due to curcumin promoting the shedding of activated matriptase into extracellular environment, but not due to curcumin effect on matriptase gene expression. In addition, we established the stable pools of CWR22Rv1 cells with matriptase overexpression and found that expression of matriptase in CWR22Rv1 cells enhanced their invasive ability. With curcumin treatment, the invasive ability induced by matriptase overexpression was significantly suppressed. In summary, the data indicated that curcumin exhibits a suppressive effect on prostate cancer cell invasion, at least in part by down-regulating matriptase activity. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2000. doi:1538-7445.AM2012-2000

Abstract 1413: Hepatocyte growth factor activator inhibitor-2 inhibits prostate cancer cell migration and invasion

Dysregulation of proteolysis on cell surface has been strongly implicated in cancer cell migration and invasion. In this study, we delineated the role of hepatocyte growth factor activator inhibitor-2 (HAI-2) in prostate cancer cell migration and invasion within a human prostate cancer progression model, established by a serial of intraprostatic injections of prostate cancer cells and isolation of the invasive cells from nearby lymph nodes. The invasion capability of these cells was increased after the serial isolations. Interestingly, the expression of HAI-2 but not HAI-1 was dramatically decreased following the progression and concurrent with an increase of activated matriptase. Overexpression of HAI-2 reduced prostate cancer cell migration and invasion, at least in part due to its inhibitory role in matriptase. Following the orthotopic tumor growth in mice, the level of matriptase was increased while HAI-2 protein level was decreased. These results indicated that during the progression of human prostate cancer, HAI-2 expression was reduced, leading to constitutive matriptase activation, increased cell migration and invasion. Thus, HAI-2 exerted an inhibitory role in prostate cancer progression partly through inhibiting matripase activity. Imbalance between HAI-2 and matriptase may result in prostate tumor progression. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1413. doi:10.1158/1538-7445.AM2011-1413

Abstract A54: Downregulation of hepatocyte growth factor activator inhibitor-2 is involved in the progression of prostate cancer cells

Dysregulation of pericellular proteolysis has been strongly involved in tumorigenesis. In this study, we delineated the role of hepatocyte growth factor activator inhibitor-2 (HAI-2) in prostate cancer cell invasion and tumorigenicity within a human prostate cancer progression model (103E, N1, and N2 cells). This model was newly established by a serial of intraprostatic propagation of a human prostate cancer 103E cell line and isolation of the metastatic cells from nearby lymph nodes. The invasion capability of these cells was gradually increased after the serial isolations (103E Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the Second AACR International Conference on Frontiers in Basic Cancer Research; 2011 Sep 14-18; San Francisco, CA. Philadelphia (PA): AACR; Cancer Res 2011;71(18 Suppl):Abstract nr A54.