Exploring natural products as Bcl-2 inhibitors for acute myeloid leukemia therapy using In vitro, STD-NMR spectroscopy, and In silico approaches

Acute myeloid leukemia (AML) is the predominant form of acute leukemia, affecting elderly individuals, typically diagnosed at an average age of 68 years. AML cells rely on the Bcl-2 protein for their survival. Overexpression of Bcl-2 protein in various cancer types renders it as a potential candidate for targeted therapies. The present study aimed to identify natural compounds as Bcl-2 inhibitors using in vitro , biophysical, and integrated computational approaches. The MTT assay was performed for cell proliferation, followed by apoptosis and gene expression analysis. STD-NMR spectroscopy, molecular docking and molecular dynamics simulations were performed for protein-ligand interactions. In the in vitro anti-proliferative assay, three natural compounds, gossypol ( 1 ), camptothecin ( 2 ), and jaceidin ( 3 ), were found active against the HL-60 cell line with IC₅₀ concentrations of 1.634 ± 0.072, 0.137 ± 0.029, and 13.492 ± 2.292 μM, respectively. These compounds triggered apoptosis and decreased cellular viability in a dose-dependent manner. The gene expression analysis of Bax , Bcl-2 , and Caspase 3 in HL-60 cells revealed that these compounds induce apoptosis by regulating essential apoptotic genes. Among the three identified potential hits, only gossypol ( 1 ) was buffer soluble and subjected to STD-NMR experiment to evaluate its protein-ligand interactions. Furthermore, molecular docking, binding free energies and MD simulation analyses demonstrated stable interactions of these compounds with the Bcl-2 protein. The three natural products showed potent to significant activity, effectively inducing apoptosis in the HL-60 cell line. Hence, this study identifies three potential lead candidates for drug discovery against Bcl-2-related cancers after further mechanistic and pre-clinical studies.