Necroptotic suppression of lung cancer cell proliferation and migration: A comprehensive in vitro and ın silico study to determine new molecular targets for pexidartinib

Abstract

In this study, the cytotoxic effects of pexidartinib (PLX), a tyrosine kinase inhibitor approved for tenosynovial giant cell tumor through inhibition of colony-stimulating factor 1 receptor (CSF1R), against A549 lung adenocarcinoma cells and Beas-2B healthy bronchial cells were investigated by in detailed in-vitro and in-silico studies. Through MTT assays, PLX demonstrated significant inhibition of A549 cell viability with IC50 values of 2.15 and 1.3 mu M at 24 and 48 h, respectively, while having minimal effects on Beas-2B cells, with IC50 values of 36.2 and 9.3 mu M. The high selectivity index indicates PLX's preferential action against cancerous cells. The mechanism of cell death induced by PLX was further explored using Annexin V/PI staining and flow cytometry, revealing that PLX primarily induces necrosis in A549 cells, with an increase in necrotic cell populations and reduced efficacy at higher concentrations. Western blot analysis showed an upregulation of necroptosis markers (RIP3 and pMLKL) in A549 cells, while apoptotic markers like Caspase-3 remained unchanged. In addition, wound healing assays demonstrated that PLX significantly inhibits A549 cell migration in a dose-dependent manner. Molecular docking studies identified key amino acids involved in PLX binding interactions with target proteins. RIPK1 showed the strongest binding affinity. MD simulations revealed that the PLX-VEGFR2 complex was the most stable. As conclusion, PLX, although approved for tenosynovial giant cell tumors, shows promising potential for lung adenocarcinoma treatment. It selectively inhibits cancer cell viability, induces necroptosis, and reduces cell migration. Its stronger binding to RIPK1 and VEGFR2 more than CSF1R.