Preparation, characterization and biochemical studies of nanoformulations of chlorambucil to enhance anticancer efficacy

Cancer, as a multifactorial disease, damages healthy cells and organs. The major drawback of anticancer drugs is that they have a severe impact on healthy cells due to their toxic effects. Encapsulation of anticancer drugs into a nanosized formulation enhances the drug delivery system. Thus, nanoformulation (NF) increases the absorption rate and speeds up the circulation time of conventional water-insoluble drugs. Chlorambucil is a well-studied anticancer drug that controls chronic lymphocytic leukaemia and malignant lymphomas. Chlorambucil, a nitrogen mustard alkylating agent, shows hepatotoxicity, secondary malignancies, etc., in normal cells and tissues. In this study, NFs of chlorambucil were prepared through an ultrasonication technique by varying the surfactant concentration. The characterization studies optimized the size, surface charge, heterogeneity, entrapment efficiency, drug loading capacity, in vitro drug release profile, stability, and interaction of the drug and excipient by using dynamic light scattering (DLS), UV-visible spectroscopy, transmission electron microscopy (TEM), Fourier-transform infrared (FTIR) spectroscopy, and nuclear magnetic resonance (NMR) methods. Moreover, MTT assay, cell cycling, and dye staining methods were performed to study the cytotoxicity and apoptotic potential of the NF of chlorambucil in HCT-116 cell lines. The cellular uptake study elucidated the internalization of NF by cancer cells. Microtubule affinity regulating kinase-4 (MARK-4) as a cancer-targeted protein was purified to study the complex formation, conformational changes, dynamics, and stability of proteins upon binding of CH by using multi-spectroscopic and in silico methods. Cancer, as a multifactorial disease, damages healthy cells and organs.