Aehtesham Hussain1,2, Mohd Saleem Dar1,3,4, Nasima Bano1,3, Md Mehedi Hossain1,3, Rafia Basit1,3, Aadil Qadir Bhat1,3, Mushtaq A Aga5, Sabeena Ali2, Qazi Parvaiz Hassan6,7, Mohd Jamal Dar8,9. 1. Academy of Scientific and Innovative Research, New Delhi, India. 2. Microbial Biotechnology Division, CSIR - Indian Institute of Integrative Medicine, Sanat Nagar, Srinagar, Jammu and Kashmir, 190005, India. 3. Cancer Pharmacology Division, CSIR - Indian Institute of Integrative Medicine, Jammu Tawi, 180001, India. 4. Department of Biochemistry, Purdue University, West Lafayette, IN, 47907-2063, USA. 5. Medicinal Chemistry Division, CSIR - Indian Institute of Integrative Medicine, Jammu Tawi, Jammu and Kashmir, 180001, India. 6. Academy of Scientific and Innovative Research, New Delhi, India. qphassan@iiim.ac.in. 7. Microbial Biotechnology Division, CSIR - Indian Institute of Integrative Medicine, Sanat Nagar, Srinagar, Jammu and Kashmir, 190005, India. qphassan@iiim.ac.in. 8. Academy of Scientific and Innovative Research, New Delhi, India. jamal@iiim.ac.in. 9. Cancer Pharmacology Division, CSIR - Indian Institute of Integrative Medicine, Jammu Tawi, 180001, India. jamal@iiim.ac.in.
Abstract
PURPOSE: Despite the fact that hyper-activation of Wnt/β-catenin signaling pathway has been seen in many cancers, including liver, colorectal and lung carcinoma, no small molecule inhibitors are available that specifically target this pathway. In this study, we analyzed the impact of dinactin (DA), an antibiotic ionophore produced by Streptomyces species, as an effective small molecule targeting Wnt/β-catenin signaling pathway in cancer cells. METHODS: We performed MTT assays to investigate cell viability and proliferation after exposure to small molecules. Protein expression analysis was carried out by western blotting. Top-Flash reporter assays were used to score for β-catenin signaling and cell cycle analysis was carried out by flow cytometry. RESULTS: In the first set of experiments, DA was seen to selectively inhibit the proliferation of HCT-116 and HepG2 cancer cells, unlike HEK-293 cells (a low tumorigenic cell line), in apoptosis-independent manner. Further, DA was seen to block the G1/S progression and decrease the expression of cyclin D1 in cancer cells. Since cyclin D1 is the downstream target gene of Wnt/β-catenin signaling, we examined the impact of DA on TCF-dependent β-catenin activity using Top-Flash reporter assay. Interestingly, DA significantly decreased Top-Flash activity at lower nano-molar concentrations when compared with salinomycin in HCT-116 and HepG2 cells. CONCLUSION: We report the identification of dinactin as a natural product-based small molecule that effectively blocks the Wnt/β-catenin signaling pathway in cancer cells at nano-molar concentration. We anticipate that DA could be developed as a novel drug for anti-cancer therapy and for the management of neuropathic pain.
PURPOSE: Despite the fact that hyper-activation of Wnt/β-catenin signaling pathway has been seen in many cancers, including liver, colorectal and lung carcinoma, no small molecule inhibitors are available that specifically target this pathway. In this study, we analyzed the impact of dinactin (DA), an antibiotic ionophore produced by Streptomyces species, as an effective small molecule targeting Wnt/β-catenin signaling pathway in cancer cells. METHODS: We performed MTT assays to investigate cell viability and proliferation after exposure to small molecules. Protein expression analysis was carried out by western blotting. Top-Flash reporter assays were used to score for β-catenin signaling and cell cycle analysis was carried out by flow cytometry. RESULTS: In the first set of experiments, DA was seen to selectively inhibit the proliferation of HCT-116 and HepG2cancer cells, unlike HEK-293 cells (a low tumorigenic cell line), in apoptosis-independent manner. Further, DA was seen to block the G1/S progression and decrease the expression of cyclin D1 in cancer cells. Since cyclin D1 is the downstream target gene of Wnt/β-catenin signaling, we examined the impact of DA on TCF-dependent β-catenin activity using Top-Flash reporter assay. Interestingly, DA significantly decreased Top-Flash activity at lower nano-molar concentrations when compared with salinomycin in HCT-116 and HepG2 cells. CONCLUSION: We report the identification of dinactin as a natural product-based small molecule that effectively blocks the Wnt/β-catenin signaling pathway in cancer cells at nano-molar concentration. We anticipate that DA could be developed as a novel drug for anti-cancer therapy and for the management of neuropathic pain.