Ankith Sherapura1, Vikas H Malojirao1,2, B S Sharath3,4, Prabhu Thirusangu1,5, Riaz Mahmood3, N Suchetha Kumari6, Shrinath M Baliga7, Shaukath Ara Khanum8, B T Prabhakar9. 1. Molecular Biomedicine Laboratory, Postgraduate Department of Studies and Research in Biotechnology, Sahyadri Science College, Kuvempu University, Shivamogga, Karnataka, 577203, India. 2. Division for DNA repair research, Department of Neurosurgery, Center for neuroregeneration, Houston Methodist, Houston, Fannin Street, TX, USA. 3. Postgraduate Department of Studies and Research in Biotechnology and Bioinformatics, Kuvempu University, Shankaraghatta, Shivamogga, Karnataka, 577203, India. 4. School of System Biomedical Science and Department of Bioinformatics and Life Science, Soongsil University, Seoul, South Korea. 5. Division of Experimental Pathology and Laboratory Medicine, Mayo Clinic, Rochester, MN, USA. 6. Department of Biochemistry, K. S. Hegde Medical Academy, NITTE University, Mangalore, Karnataka, India. 7. Mangalore Institute of Oncology, Mangalore, Karnataka, India. 8. Department of Chemistry, Yuvaraja's College (Autonomous), University of Mysore, Mysore, Karnataka, India. 9. Molecular Biomedicine Laboratory, Postgraduate Department of Studies and Research in Biotechnology, Sahyadri Science College, Kuvempu University, Shivamogga, Karnataka, 577203, India. pbtssc@gmail.com.
Abstract
BACKGROUND: Anomalous activation of intra-cellular signalling cascades confers neoplastic properties on malignant cells. The JAK2/STAT3 proteins play a pivotal role in the pathogenesis of most of the solid malignancies. The over expression of STAT3 in these tumours results in an evasion of apoptosis and thereby pathogenesis. Hence, strategy to target STAT3 to regress tumour development is an emerging new concept. As an approach, anti-neoplastic drug, Azo-hydrozone analogue, BT-1F with potential anti-proliferative effect was evaluated to demonstrate its capacity to counteract STAT3 signal with mechanistic approach. METHODS: Cell based screening for cytotoxicity was performed through MTT, LDH and Trypan blue. The BT-1F induced anti-clonogenic property by clonogenic assay. The apoptotic capacity was examined by crystal violet staining, flow cytometry, Annexin-FITC, DAPI and TUNEL assay. The altered signalling events were studied using immunoblot. The drug-induced anti-tumour effect was evaluated in an in-vivo solid tumour model and molecular interaction was further validated by in-silico studies. RESULTS: The BT-1F exerts chemo-sensitivity specifically against EAC and A549 cells without altering its normal counterpart. The anti-proliferative/anti-clonogenic effect was due to the induction of apoptosis through inhibition of STAT3Tyr705 signal. Eventually downstream signalling proteins p53, Bax, Bad and Bcl-xL were significantly altered. Further in-vivo experimental results validated in-vitro findings. The computational approaches assures the BT-1F efficiency in binding with STAT3. CONCLUSION: Systemic validation of STAT3 target drug, BT-1F in in-vitro, in-silico and in-vivo models has promising strategy for solid cancer treatment.
BACKGROUND: Anomalous activation of intra-cellular signalling cascades confers neoplastic properties on malignant cells. The JAK2/STAT3 proteins play a pivotal role in the pathogenesis of most of the solid malignancies. The over expression of STAT3 in these tumours results in an evasion of apoptosis and thereby pathogenesis. Hence, strategy to target STAT3 to regress tumour development is an emerging new concept. As an approach, anti-neoplastic drug, Azo-hydrozone analogue, BT-1F with potential anti-proliferative effect was evaluated to demonstrate its capacity to counteract STAT3 signal with mechanistic approach. METHODS: Cell based screening for cytotoxicity was performed through MTT, LDH and Trypan blue. The BT-1F induced anti-clonogenic property by clonogenic assay. The apoptotic capacity was examined by crystal violet staining, flow cytometry, Annexin-FITC, DAPI and TUNEL assay. The altered signalling events were studied using immunoblot. The drug-induced anti-tumour effect was evaluated in an in-vivo solid tumour model and molecular interaction was further validated by in-silico studies. RESULTS: The BT-1F exerts chemo-sensitivity specifically against EAC and A549 cells without altering its normal counterpart. The anti-proliferative/anti-clonogenic effect was due to the induction of apoptosis through inhibition of STAT3Tyr705 signal. Eventually downstream signalling proteins p53, Bax, Bad and Bcl-xL were significantly altered. Further in-vivo experimental results validated in-vitro findings. The computational approaches assures the BT-1F efficiency in binding with STAT3. CONCLUSION: Systemic validation of STAT3 target drug, BT-1F in in-vitro, in-silico and in-vivo models has promising strategy for solid cancer treatment.
Authors: Carmen Belli; Dario Trapani; Giulia Viale; Paolo D'Amico; Bruno Achutti Duso; Paolo Della Vigna; Franco Orsi; Giuseppe Curigliano Journal: Cancer Treat Rev Date: 2018-02-22 Impact factor: 12.111