Chlorogenic acid-ZnO nanocomposite derived from Hydrocotyle javanica Thunb. activates ferroptosis-regulated cell death axis in breast cancer cells.

Breast cancer remains one of the leading causes of cancer-related mortality among the women worldwide. The metal oxide nanoparticle-based formulations offer improved drug efficacy which makes them promising candidates for cancer therapy. This study investigates the anti-breast cancer potential of isolated Chlorogenic acid (CGA) from Hydrocotyle javanica (H. javanica) complexed with Zinc oxide (ZnO) nanoparticles. CGA was isolated using systematic chromatography and characterized by 1H NMR, 13C NMR, and mass spectrometry. The isolated CGA was then coated onto ZnO and encapsulated with chitosan (CS). The resulting CGA-ZnO complex was characterized using X-Ray Diffraction, Fourier Transform Infrared Spectroscopy, Field Emission Scanning Electron Microscopy and other techniques. The CGA-ZnO exhibited a hexagonal wurtzite structure (JCPDS card (No. 36-1451)), spherical morphology, +15 mV zeta potential, decreased polydispersity index (0.32) which differentiate CGA-ZnO complex from ZnO alone. CGA-ZnO caused significant decrease in MCF-7 cell's viability (53.14% at 20 μM) compared to CGA (89.21% at 20 μM) or ZnO (97.23 at 20 μM) alone with minimum cytotoxicity to normal cells, MCF-10A (IC50 70 μM). CGA-ZnO elevated ROS levels and increased the apoptotic population in MCF-7 (18.70%), MDA-MB-231 (39.59%) and MDA-MB-468 (45.4%). CGA-ZnO treatment depleted GSH level, SOD and catalase activity, while increasing TBARS levels . The elevated levels of phosphorylated p53, increased p21, and reduced Cyclin A2/CDK2 induced S-phase arrest in MCF-7. Additionally, CGA-ZnO downregulated the expression of phospho-NRF2, GPx, xCT and elevated ferritin, DMT1, cleaved Caspase 3 to activate ferroptosis-regulated cell death axis. Furthermore, rescue assay confirmed the mechanism by which CGA-ZnO efficiently kills the breast cancer cells.