Green tea (-)-epigalocatechin-3-gallate inhibits KIT activity and causes caspase-dependent cell death in gastrointestinal stromal tumor including imatinib-resistant cells.

Imatinib, a selective tyrosine kinase inhibitor, has been used as a standard first-line therapy for gastrointestinal stromal tumor (GIST) patients. Unfortunately, most patients responding to imatinib will eventually exhibit the resistance, the cause of which is not fully understood. The serious clinical problems of imatinib-resistance demand alternative treatment strategy. (-)-Epigallocatechin-3-gallate (EGCG), a main component of green tea catechin, has been demonstrated potential anti-tumor effects on various types of cancer cells. Here, we report for the first time that EGCG has shown anti-tumor effects on gastrointestinal stromal tumor cell line GIST-T1 by suppressing cell proliferation and eventually inducing cell death via caspase-dependent pathways. GIST-T1 and imatinib resistant GIST-T1 (GIST-T1 IR) cells were used to assess the effects of EGCG. In both cell types, KIT activity was completely inhibited after 4 h treatment with 60 muM EGCG. EGCG specifically inhibited activated KIT, which was demonstrated by using Ba/F3 cells transfected with human wild-type KIT construct. At a dose of 30 muM EGCG, the KIT activity remains but at more than 40 muM EGCG, the KIT activity was abolished in these transfected-Ba/F3 cells. Our results suggest that EGCG has a promising potential as a natural KIT inhibitor and therefore it could be used as a novel therapeutic or preventive reagent for GISTs including the imatinib-resistant cases.

Cas-L was overexpressed in imatinib-resistant gastrointestinal stromal tumor cells.

Gastrointestinal stromal tumors (GISTs) are the most common mesenchymal tumors in the gastrointestinal tract. Most GISTs patients respond to imatinib, yet will eventually exhibit resistance, and the mechanisms of imatinib resistance have not yet been fully elucidated. To clarify the mechanisms of secondary imatinib-resistant gastrointestinal stromal tumors, we generated resistant cells from the imatinib-sensitive GIST-T1 cells by exposing them to increasing concentrations of imatinib for 6 m. GIST-T1 IR (imatinib-resistant) cells showing an IC50 of imatinib 5-7 microM were generated. In GIST-T1 IR cells, KIT and its downstream signaling molecules remained phosphorylated with the presence of 1 microM imatinib, and no new mutations were found in KIT, PDGFRA, PKCtheta and JAK2. DNA micro-array analysis showed the overexpression of Cas-L in the resistant cells with 513 fold higher than that in the parental cells. Cas-L overexpression and SRC hyper-activation were also observed in the resistant cells at protein level and they were markedly decreased in KIT siRNA transfected GIST-T1 IR cells. Interestingly, GIST-T1 IR cells transfected with Cas-L siRNA turned out to become again sensitive to imatinib. Imatinib or PP1, a SRC inhibitor, alone was not enough to suppress the activation of KIT and its downstream signaling molecules, but the combination of them showed strong inhibitory effects on those in the resistant cells. We report for the first time that the mechanism of imatinib-resistant GISTs, at least in one cell line, involves KIT/Cas-L/SRC signaling. Cas-L depletion sensitized the resistant GIST-T1 IR cells to imatinib.