Ha-Na Oh1, Ah-Won Kwak1, Mee-Hyun Lee2, Eunae Kim3, Goo Yoon1, Seung-Sik Cho1, Kangdong Liu4, Jung-Il Chae5, Jung-Hyun Shim6. 1. Department of Pharmacy, College of Pharmacy, Mokpo National University, Jeonnam 58554, Republic of Korea. 2. College of Korean Medicine, Dongshin University, Naju-si, Jeollanam-do 58245, Republic of Korea. 3. College of Pharmacy, Chosun University, Gwangju 61452, Republic of Korea. 4. China-US (Henan) Hormel Cancer Institute, Zhengzhou, Henan, 450008, P.R. China; Basic Medical College, Zhengzhou University, Zhengzhou 450001, Henan, China. 5. Department of Dental Pharmacology, School of Dentistry and Institute of Oral Bioscience, Jeonbuk National University, Jeonju 54896, Republic of Korea. Electronic address: jichae@jbnu.ac.kr. 6. Department of Pharmacy, College of Pharmacy, Mokpo National University, Jeonnam 58554, Republic of Korea; China-US (Henan) Hormel Cancer Institute, Zhengzhou, Henan, 450008, P.R. China; Department of Biomedicine, Health & Life Convergence Sciences, BK21 Four, College of Pharmacy, Mokpo National University, Jeonnam 58554, Republic of Korea. Electronic address: s1004jh@gmail.com.
Abstract
BACKGROUND: Lung cancer has the highest incidence and cancer-related mortality of all cancers worldwide. Its treatment is focused on molecular targeted therapy. c-MET plays an important role in the development and metastasis of various human cancers and has been identified as an attractive potential anti-cancer target. Podophyllotoxin (PPT), an aryltetralin lignan isolated from the rhizomes of Podophyllum species, has several pharmacological activities that include anti-viral and anti-cancer effects. However, the mechanism of the anti-cancer effects of PPT on gefitinib-sensitive (HCC827) or -resistant (MET-amplified HCC827GR) non-small cell lung cancer (NSCLC) cells remains unexplored. PURPOSE: In the present study, we investigated the underlying mechanisms of PPT-induced apoptosis in NSCLC cells and found that the inhibition of c-MET kinase activity contributed to PPT-induced cell death. METHODS: The regulation of c-MET by PPT was examined by pull-down assay, ATP-competitive binding assay, kinase activity assay, molecular docking simulation, and Western blot analysis. The cell growth inhibitory effects of PPT on NSCLC cells were assessed using the MTT assay, soft agar assay, and flow cytometry analysis. RESULTS: PPT could directly interact with c-MET and inhibit kinase activity, which further induced the apoptosis of HCC827GR cells. In contrast, PPT did not significantly affect EGFR kinase activity. PPT significantly inhibited the cell viability of HCC827GR cells, whereas the PPT-treated HCC827 cells showed a cell viability of more than 80%. PPT dose-dependently induced G2/M cell cycle arrest, as shown by the downregulation of cyclin B1 and cdc2, and upregulation of p27 expression in HCC827GR cells. Furthermore, PPT treatment induced Bad expression and downregulation of Mcl-1, survivin, and Bcl-xl expression, subsequently activating multi-caspases. PPT thereby induced caspase-dependent apoptosis in HCC827GR cells. CONCLUSION: These results suggest the potential of PPT as a c-MET inhibitor to overcome tyrosine kinase inhibitor resistance in lung cancer.
BACKGROUND:Lung cancer has the highest incidence and cancer-related mortality of all cancers worldwide. Its treatment is focused on molecular targeted therapy. c-MET plays an important role in the development and metastasis of various humancancers and has been identified as an attractive potential anti-cancer target. Podophyllotoxin (PPT), an aryltetralin lignan isolated from the rhizomes of Podophyllum species, has several pharmacological activities that include anti-viral and anti-cancer effects. However, the mechanism of the anti-cancer effects of PPT on gefitinib-sensitive (HCC827) or -resistant (MET-amplified HCC827GR) non-small cell lung cancer (NSCLC) cells remains unexplored. PURPOSE: In the present study, we investigated the underlying mechanisms of PPT-induced apoptosis in NSCLC cells and found that the inhibition of c-MET kinase activity contributed to PPT-induced cell death. METHODS: The regulation of c-MET by PPT was examined by pull-down assay, ATP-competitive binding assay, kinase activity assay, molecular docking simulation, and Western blot analysis. The cell growth inhibitory effects of PPT on NSCLC cells were assessed using the MTT assay, soft agar assay, and flow cytometry analysis. RESULTS:PPT could directly interact with c-MET and inhibit kinase activity, which further induced the apoptosis of HCC827GR cells. In contrast, PPT did not significantly affect EGFR kinase activity. PPT significantly inhibited the cell viability of HCC827GR cells, whereas the PPT-treated HCC827 cells showed a cell viability of more than 80%. PPT dose-dependently induced G2/M cell cycle arrest, as shown by the downregulation of cyclin B1 and cdc2, and upregulation of p27 expression in HCC827GR cells. Furthermore, PPT treatment induced Bad expression and downregulation of Mcl-1, survivin, and Bcl-xl expression, subsequently activating multi-caspases. PPT thereby induced caspase-dependent apoptosis in HCC827GR cells. CONCLUSION: These results suggest the potential of PPT as a c-MET inhibitor to overcome tyrosine kinase inhibitor resistance in lung cancer.