Novel Carbazole-Piperazine Hybrid Small Molecule Induces Apoptosis by Targeting BCL-2 and Inhibits Tumor Progression in Lung Adenocarcinoma in Vitro and Xenograft Mice Model.

Mongre Mongre,Lee Lee,Prakash Prakash,Jung Jung,Kumari Kumari,Mishra Mishra,Hong Hong

doi:10.3390/cancers11091245

Abstract

Lung cancer is a type of deadly cancer and a leading cause of cancer associated death worldwide. BCL-2 protein is considered as an imperative target for the treatment of cancer due to their significant involvement in cell survival and death. A carbazole-piperazine hybrid molecule ECPU-0001 was designed and synthesized as a potent BCL-2 targeting agent with effective anticancer cancer activity. Interaction of ECPU-001 has been assessed by docking, molecular dynamics (MD) simulation, and thermal shift assay. Further, in vitro and in vivo anticancer activity was executed by cytotoxicity assay, FACS, colony formation and migration assay, western blotting, immunocyto/histochemistry and xenograft nude mice model. Molecular docking and MD simulation study confirmed that ECPU-0001 nicely interacts with the active site of BCL-2 by displaying a Ki value of 5.72 µM and binding energy (ΔG) of –8.35 kcal/mol. Thermal shift assay also validated strong interaction of this compound with BCL-2. ECPU-0001 effectively exerted a cytotoxic effect against lung adenocarnoma cells A459 with an IC50 value of 1.779 µM. Molecular mechanism of action have also been investigated and found that ECPU-0001 induced apoptosis in A459 cell by targeting BCL-2 to induce intrinsic pathway of apoptosis. Administration of ECPU-0001 significantly inhibited progression of tumor in a xenograft model without exerting severe toxicity and remarkably reduced tumor volume as well as tumor burden in treated animals. Our investigation bestowed ECPU-0001 as an effective tumoricidal agent which exhibited impressive anticancer activity in vitro as well as in vivo by targeting BCL-2 associated intrinsic pathway of apoptosis. Thus, ECPU-0001 may provide a valuable input for therapy of lung adenosarcoma in future, however, further extensive investigation of this compound will be needed.

Highlights

Lung cancer is considered as one of the most prevalent malignant diseases and foremost cause of cancer death worldwide [1,2]
We adopted the recently solved crystal structure of BCL-2 complex with inhibitor S55746 which is suggested that active site of BCL-2 is canonically formed by four α-helices (α1–α4) and the inhibitor is transversely occupied in the active site, stabilized with hydrophobic residues of α2 and α4 [37]
A molecular dynamics (MD) simulation study has been executed to investigate the conformational stability of ECPU-0001 in active site of BCL-2, in water at 300 K

Summary

Introduction

Lung cancer is considered as one of the most prevalent malignant diseases and foremost cause of cancer death worldwide [1,2]. WHO has classified lung cancer in two categories, small cell lung. Cancers 2019, 11, 1245 cancer (SCLC) and non-small cell lung cancer (NSCLC) [3,4]. It is estimated that more than 1.3 million new cases of lung cancer each year are appearing. Current treatment strategies of lung cancer include chemotherapy, radiotherapy, immunotherapy and surgical removal of uncontrolled malignant cells [7]. Widespread efforts have been made to fight with malignancies and due course several anticancer drug molecules have been developed [8]. Currently running anticancer drugs in market is suffering from numerous drawback including normal cell toxicity, resistance and intolerance. The discovery of powerful anticancer agents is a compulsory requirement to selectively kill cancerous cell without exerting severe side effect

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