Biocompatibility and cytotoxicity in vitro of surface-functionalized drug-loaded spinel ferrite nanoparticles

In this study, poly(isobutylene-alt-maleic anhydride) (PMA)-coated spinel ferrite (MFe₂O₄, where M = Fe, Co, Ni, or Zn) nanoparticles (NPs) were developed as carriers of the anticancer drugs doxorubicin (DOX) and methotrexate (MTX). Physical characterizations confirmed the formation of pure cubic structures (14–22 nm) with magnetic properties. Drug-loaded NPs exhibited tumor specificity with significantly higher (p < 0.005) drug release in an acidic environment (pH 5.5). The nanoparticles were highlycolloidal (zeta potential = −35 to −26 mV) in deionized water, phosphate buffer saline (PBS), and sodium borate buffer (SBB).They showed elevated and dose-dependent cytotoxicity in vitro compared to free drug controls. The IC50 values ranged from0.81 to 3.97 μg/mL for HepG2 and HT144 cells, whereas IC₅₀ values for normal lymphocytes were 10 to 35 times higher(18.35–43.04 µg/mL). Cobalt ferrite (CFO) and zinc ferrite (ZFO) NPs were highly genotoxic (p < 0.05) in cancer cell lines. Thenanoparticles caused cytotoxicity via oxidative stress, causing DNA damage and activation of p53-mediated cell cycle arrest (significantly elevated expression, p < 0.005, majorly G1 and G2/M arrest) and apoptosis. Cytotoxicity testing in 3D spheroids showedsignificant (p < 0.05) reduction in spheroid diameter and up to 74 ± 8.9% of cell death after two weeks. In addition, they also inhibited multidrug resistance (MDR) pump activity in both cell lines suggesting effectivity in MDR cancers. Among the tested MFe₂O₄NPs, CFO nanocarriers were the most favorable for targeted cancer therapy due to excellent magnetic, colloidal, cytotoxic, andbiocompatible aspects. However, detailed mechanistic, in vivo cytotoxicity, and magnetic-field-assisted studies are required to fullyexploit these nanocarriers in therapeutic applications.