Blockade of four-transmembrane L6 family member 5 (TM4SF5)-mediated tumorigenicity in hepatocytes by a synthetic chalcone derivative

Abstract

We previously reported that the four-transmembrane L6 family member 5 (TM4SF5) was highly expressed in hepatocarcinoma, induced morphological elongation and epithelialmesenchymal transition, and caused abnormal cell growth in multilayers in vitro and tumor formation in vivo. In this study, we identified a synthetic compound, 4 -(p-toluenesulfonylamido)-4-hydroxychalcone (TSAHC) that antagonized both the TM4SF5-mediated multilayer growth and TM4SF5-enhanced migration/invasion. TSAHC treatment induced multilayer-growing cells to grow in monolayers, recovering contact inhibition without accompanying apoptosis, and inhibited chemotactic migration and invasion. E pithelial monolayer integrity is maintained by integrin-mediated cell adhesion between the cell and the extracellular matrix (ECM) and by E-cadherin-mediated contact between adjacent cells. 1 Epithelial-mesenchymal transition (EMT) through loss of cellcell contacts disrupts monolayer integrity and alters cell-ECM interactions. 2 Tumor cells disseminated from primary tumors via loss of cell adhesion and contact can migrate to and invade distal tissues. 3 We recently reported that TM4SF5-mediated EMT results in a loss of contact inhibition and multilayer growth. 4 Therefore, altered cell adhesion and contact may lead to both a loss of contact inhibition and a dissemination of metastatic cells from the primary tumor. 5 Integrin-mediated cell adhesion reorganizes actin filaments 6 through activation of diverse intracellular signaling molecules, including focal adhesion kinase (FAK), Rho guanosine triphosphatases (RhoA, Rac1, and CDC42), and others, 7 which are critical for cellular morphology and migration. 8 TM4SF proteins (i.e., tetraspanins or tetraspans) are a group of membrane proteins with four transmembrane

Monoclonal Antibody Targeting of the Cell Surface Molecule TM4SF5 Inhibits the Growth of Hepatocellular Carcinoma

“…As IgG2a has extremely low affinity for Fc Receptors on phagocytic cells in human (34), further consideration of the isotype may be required for the future application of this antibody in humans. Considering that TM4SF5 is involved in EMT and metastasis (13)(14)(15)(16), and that the treatment of TM4SF5-expressing cells with the anti-TM4SF5 monoclonal antibody reduced motility and enhanced expression of E-cadherin, the anti-TM4SF5 monoclonal antibody may have antimetastatic activity and therefore may be able to contribute to preventing metastasis in patients with primary HCC. Therefore, this topic warrants further investigation in the future.…”

“…Lee and colleagues showed that TM4SF5 is involved in HCC development, specifically by inducing morphological elongation, epithelial-mesenchymal transition (EMT), uncontrolled cell proliferation, and angiogenesis (13,14). A synthetic inhibitor targeting TM4SF5, 4 0 -(ptoluenesulfonyl-amido)-4-hydroxychalcone (TSAHC), has been shown to inhibit HCC growth and metastasis in vitro and in vivo (15). Therefore, TM4SF5 seems to play an important role in HCC formation, and it is a rational molecular target for the clinical development of HCC therapeutics (16).…”

Tetraspan TM4SF5-dependent direct activation of FAK and metastatic potential of hepatocarcinoma cells

“…Additionally, control compounds lacking the functional group (49-toluenesulfonylamido group) of TSAHC (i.e. 49-amino-4-hydroxychalcone and 49,4-dihydroxychalcone) did not exert any effects on FAK phosphorylation (Lee et al, 2009b).…”

“…Suppression of TM4SF5 or treatment with an anti-TM4SF5 reagent (TSAHC) or a FAK inhibitor attenuated TM4SF5/FAK interactionmediated FAK phosphorylation and activation. TSAHC was originally screened as an a-glycosidase inhibitor (Seo et al, 2005) and appears to affect the structural integrity or Nglycosylation (at N138/N155) of the extracellular loop 2 (EC2) within TM4SF5, which appears to be important for multilayer growth and migration (Lee et al, 2009b). TSAHC or FAK inhibitor treatment attenuated the interaction between TM4SF5 and FAK, indicating that both the structural aspects of the extracellular region of TM4SF5 and the activity of intracellular FAK are able to modulate the interaction between TM4SF5 and FAK.…”

“…In some cases, cells were transiently transfected with shRNA against control scrambled sequence or TM4SF5 [shTM4SF5 (Lee et al, 2006) or GFP-shTM4SF5 (Origene, Rockville, MD)] for 48 h; treated with DMSO or pharmacological inhibitors, including TSAHC (20 mM) against TM4SF5 (Lee et al, 2009b) or PF271 (1.0 mM) against FAK (Pasapera et al, 2010) for 24 h; or pretreated with normal mouse IgG or neutralizing anti-integrin b1, a5, or av antibody (Invitrogen, clone P4C10, P1D6, LM609, respectively, 20 mg/ml) for 30 min in the middle of rocking prior to replating. For replating, cells were trypsinized and washed with serum-free media containing 1% BSA twice before rocking (60 rpm) at 37˚C for 1 h and then either kept in suspension or replated on PL (Sigma, St. Louis, MO) or FN (10 mg/ml)-precoated culture dishes for the indicated times.…”