Lung cancer growth inhibition and autophagy activation by tetrazole via ERK1/2 up-regulation and mTOR/p70S6K signaling down-regulation

Liangjian Zheng; Jun Zhang; Jia Fan; Yuxin He; Tingting Zhan; Liwen Rong; Mengzhen Yuan; Hongyan Zhang

doi:10.18388/abp.2020_5776

Liangjian Zheng 1Department of Oncology, The Third People’s Hospital of Chengdu, Chengdu, Sichuan, 610031, China; 2Department of Oncology, The Affiliated Hospital of Southwest Jiaotong University, Chengdu, Sichuan, 610031, China
Jun Zhang 1Department of Oncology, The Third People’s Hospital of Chengdu, Chengdu, Sichuan, 610031, China; 2Department of Oncology, The Affiliated Hospital of Southwest Jiaotong University, Chengdu, Sichuan, 610031, China
Jia Fan 1Department of Oncology, The Third People’s Hospital of Chengdu, Chengdu, Sichuan, 610031, China; 2Department of Oncology, The Affiliated Hospital of Southwest Jiaotong University, Chengdu, Sichuan, 610031, China
Yuxin He 1Department of Oncology, The Third People’s Hospital of Chengdu, Chengdu, Sichuan, 610031, China; 2Department of Oncology, The Affiliated Hospital of Southwest Jiaotong University, Chengdu, Sichuan, 610031, China
Tingting Zhan 1Department of Oncology, The Third People’s Hospital of Chengdu, Chengdu, Sichuan, 610031, China; 2Department of Oncology, The Affiliated Hospital of Southwest Jiaotong University, Chengdu, Sichuan, 610031, China
Liwen Rong 1Department of Oncology, The Third People’s Hospital of Chengdu, Chengdu, Sichuan, 610031, China; 2Department of Oncology, The Affiliated Hospital of Southwest Jiaotong University, Chengdu, Sichuan, 610031, China
Mengzhen Yuan 1Department of Oncology, The Third People’s Hospital of Chengdu, Chengdu, Sichuan, 610031, China; 2Department of Oncology, The Affiliated Hospital of Southwest Jiaotong University, Chengdu, Sichuan, 610031, China
Hongyan Zhang Department of Laboratory Medicine, Eastern Hospital, Sichuan Academy of Medical Sciences and Sichuan Provincial People’s Hospital, Chengdu, Sichuan, 610101, China

DOI: https://doi.org/10.18388/abp.2020_5776

Abstract

Lung cancer, a most common clinically diagnosed malignancy grows rapidly and undergoes metastasis/diffusion to distant organs at a fast rate. In the present study gravacridondiol tetrazole (tetrazole) was synthesized and investigated for lung cancer growth inhibition potential in vitro. MTT assay and flow cytometry using propidium iodide were used to determine viability changes and DNA content distribution. Protein expression and apoptotic changes were detected by western blotting and Annexin-V/PI assays. Treatment with 12 μM tetrazole suppressed viabilities to 23% and 20% in A549 and NCI-H1819 cells, respectively. In tetrazole exposed cells, G1-phase cell count increased significantly compared to the control. Tetrazole-treatment of A549 and NCI-H1819 cells caused a prominent raise in LC3‑II and p-ERK1/2 expression at 72 h. The SQSTM1/p62 level, p-mTOR and p-p70S6K expression was lowered significantly in A549 and NCI-H1819 cells on exposure to tetrazole. Exposure to U1026 alleviated tetrazole mediated LC3II/I ratio increase in A549 and NCI-H1819 cells significantly (P<0.02) compared to tetrazole treated cells. Treatment with tetrazole and 3‑MA in combination led a significant (P<0.02) elevation in A549 and NCI-H1819 cell apoptotic count relative to tetrazole (12 μM) alone treated cells. Moreover, tetrazole and 3‑MA combination increased cleavage of caspase‑3 to a greater extent compared to tetrazole. In summary, tetrazole manifested anti-proliferative effect on lung cancer cells via autophagy over-activation and arrest of cell cycle. It deactivated ERK1/2 signalling and promoted mTOR signaling in A549 and NCI-H1819 cells to regulate cancer proliferation. Thus, tetrazole needs to be studied further as an anti-proliferative agent for treatment of lung cancer.