Integrating Theory with Experimental Studies on the Directed Assembly of Phenforminium 5-Fluorouracilate toward Enhanced In Vitro/Vivo Properties and Synergistic Antitumor Efficacy

The codrug efficacy increase approach is developed to improve the biopharmaceutical properties of the conventional antitumor drug 5-fluorouracil (Fu) through molecular salification by tapping the potential of biguanide agents in property optimization and further antitumor synergism. In this route, the biguanide phenformin (Phf) is noncovalently bonded with Fu through a proton transference procedure based on its acidic–alkalinity differences, thereby improving the physicochemical and pharmacokinetic behaviors of Fu and ultimately achieving enhanced efficacy by leveraging the synergized antitumor effect of Phf. Oriented by this, a novel codrug molecular salt, phenforminium 5-fluorouracilate (Fu-Phf), is assembled and systematically characterized for its precise structure employing single-crystal/powder X-ray diffraction and other analytical methods. The results revealed the presence of robust charge-assisted hydrogen bonds in the asymmetric unit, with its structural characteristics being internally hydrophilic and externally hydrophobic, which affects its in vitro physicochemical performances. Thereinto, the dissolubility and permeability of Fu are proven to be simultaneously improved after forming the molecular salt, which aligns positively with a series of theoretical predictions, including molecular electrostatic potential (MEP), reduced density gradient (RDG), frontier molecular orbitals (FMOs), and topological analysis based on the theory of atoms in molecules (AIMs). All of these result in improved in vivo pharmacokinetic performances of the molecular salt, leading to higher bioavailability over Fu alone. Encouragingly, the strengthened tumor inhibition properties of the Fu-Phf codrug are theoretically simulated through the molecular docking method and experimentally observed against selected cancer cell strains, showing significantly reduced IC50. Furthermore, the calculated CI values of less than 1.0 highlight the achievement of the synergistic effects of the two components in the codrug molecular salt. Consequently, this work introduces a fresh salification case of Fu, presenting promising potential for future development; simultaneously, it also provides insights into exploring the possibilities of biguanide medicine as antitumor reinforcement through dual-drug synergy and optimization strategies.