Inhibitory effect of 4‐methylesculetin on hyaluronan synthesis slows the development of human pancreatic cancer in vitro and in nude mice

Abstract

We report the inhibitory effect of 4-methylesculetin (ME), a 4-methylumbelliferone derivative, on hyaluronan (HA) synthesis by pancreatic cancer cells, and its resulting anticancer action. First, HA in cell culture was analyzed using competitive inhibition with hyaluronic acid-binding protein (HABP) to study HA synthesis by the human pancreatic cancer cell line KP1-NK, and cell-surface HA was visualized using a particle-exclusion assay to study the synthesis of extracellular matrix HA. We also analyzed the inhibitory effect of ME on cell adhesion and invasion, which play a role in the invasion, growth and metastasis of human pancreatic cancer. Furthermore, we examined HA in human pancreatic cancer cells transplanted into the hypodermis of nude mice to study the inhibitory effect of ME on HA synthesis. Moreover, pancreatic cancer cells were also transplanted into the abdomen of nude mice to study whether ME would have the potential to prolong the survival of patients with end-stage pancreatic cancer. ME at 10 lM did not inhibit the growth of human pancreatic cancer cells, but inhibited HA synthesis in cell culture by 40%, adhesion by 44% and invasion by 40%. ME inhibited the proliferation of subcutaneous tumors and HA synthesis (by 50%) of pancreatic cancer transplanted into the hypodermis of nude mice. ME also prolonged the survival time of nude mice bearing abdominally transplanted pancreatic cancer cells. ME inhibited pancreatic cancer growth and metastasis by inhibition of HA synthesis. These results suggest that ME may prolong the survival time of patients with end-stage pancreatic cancer. ' 2007 Wiley-Liss, Inc.Key words: hyaluronan; pancreatic cancer; 4-methylumbelliferone; 4-methylesculetin; KP1-NL Pancreatic cancer remains one of the most malignant neoplasms because the disease is diagnosed frequently at an advanced stage, and only 3% of patients survive 5 years. [1][2][3][4] One of the pathological characteristics of this carcinoma is its early invasion of surrounding tissues and metastasis. Pancreatic cancer has various modes of spread, such as hepatic metastasis, peritoneal dissemination, retroperitoneal recurrence, lymph node metastasis and local and distant metastasis from an early stage. 5 Recently, the standard drug for treatment of progressive pancreatic cancer has been gemcitabine, 6 but this does not improve survival time because pancreatic cancer is highly resistant to chemotherapy. 1 Therefore, more effective therapy for progressive pancreatic cancer is needed.Hyaluronan (HA) is a nonsulfated linear glycosaminoglycan composed of repeated b-1,4-GlcUA-b-1-3-GlcNAc disaccharide units. 7 It is present in many tissues as a major component of the extracellular matrix and is crucial for various physiological processes. 8,9 Previous studies have shown that the amounts and composition of glycosaminoglycan undergo marked changes in various neoplastic tissues. 10,11 Increased levels of HA are often associated with certain human tumors such as colon cancer, breast cancer, glioma, lung cancer and Wilms' t...

Accumulation of Extracellular Hyaluronan by Hyaluronan Synthase 3 Promotes Tumor Growth and Modulates the Pancreatic Cancer Microenvironment

“…In HAS3-overexpressing tumors, PEGPH20 removes most of the hyaluronan and has a strong inhibitory effect on tumor growth, whereas in HAS2-overexpressing tumors PEGPH20 removes the extracellular but not the intracellular hyaluronan and causes a weaker inhibitory effect on tumor growth. Our results are in line with previous work showing that depletion of hyaluronan or suppression of its synthesis leads to inhibition of tumor growth in multiple tumor models [9, 36–38, 41, 42]. …”

“…Suppression of hyaluronan synthesis by downregulation of HASs has been previously shown to inhibit the growth of implanted breast, prostate, squamous cell carcinoma, and osteosarcoma tumors [15, 36–38]. Similarly, hyaluronan synthesis inhibitor 4-methylumbelliferone or its derivatives suppress metastasis of several tumor types [38–41]. …”

The glycocalyx and its significance in human medicine

“…Hyaluronan synthesis inhibitor 4-methylumbelliferone (4-MU), a dietary supplement or its derivatives suppress metastasis of several tumour types [118][119][120][121]. Yates et al [122] demonstrated that 4-MU inhibits HA synthesis in part by downregulating HA receptors and the phosphatidylinositol 3-kinase/ CD44 complex.…”

“…It acts to promote GCX reconstitution and control GCX degrading enzymes and has antiinflammatory, anti-apoptotic and antisenescence effects on ECs. Frati Munari [103] noted that treatment with sulodexide improved chronic venous disease and promoted healing of chronic Pentosan polysulphate Reduces MMP activity [104] Wheat germ agglutinin Lectin that binds to HS and HA [60] Rhamnan sulphate Enhances HS synthesis [105][106][107] Hawthorn extract WSS 1442 Enhances HS (indirect evidence) [108,110] Cationic copolymers Bind to the GCX [111] MicroRNAs Restore PGs and their biosynthetic enzymes [129] Degrade the glycocalyx PEGPH20 Depletes HA [112][113][114][115] 4-MU/4-ME Suppresses HA synthesis [92,[116][117][118][119][120][121][122][123] A6 monoclonal antibody Blocks CD44 [124] Naphthalene methanol-D-xyloside Blocks HS assembly [125] Peracetylated 6-fluoro-GalNAc Inhibits sulphated GAG synthesis [126] siRNAs Block GAG synthetic enzymes Unpublished data PEGPH20, pegylated human recombinant hyaluronidase; 4-MU, 4-methylumbelliferone; 4-ME, 4-methlesculetin; GAG, glycosaminoglycan; MMP, matrix metalloproteinase; HS, heparan sulphate; HA, hyaluronic acid; GCX, glycocalyx; PG, proteoglycan.…”

Hyaluronan stimulates pancreatic cancer cell motility

“…Because we found that treatment with 4-MU robustly attenuates the migratory ability of PDAC cells, inhibiting HA production could be an ideal therapeutic strategy to prevent invasion and/or metastasis of PDAC. In fact, inhibition of HA synthesis by treatment with 4-MU, or its derivative 4-methylesculetin, has demonstrated antitumor activity in a variety of cancers, including PDAC [32][33][34][35][36][37]. Of particular importance, 4-MU (or hymecromone) is a dietary supplement for improving liver health and can be administered orally without any toxicities [38].…”