Poly(ethylene oxide)‐graft‐methotrexate Macromolecular Drugs Conjugating via Aminopteridine Ring Exhibit Potent Anticancer Activity

Abstract

AbstractWater soluble poly(ethylene oxide)‐graft‐methotrexate (PEO‐g‐MTX) conjugates with a robust amide linkage via the amine or carboxylic acid groups of MTX were designed, prepared and investigated for in vitro anti‐tumor effects. MTX was conjugated to multi‐functional PEO containing multiple pendant carboxylic acid (PEO‐g‐COOH) or amine groups (PEO‐g‐NH2) via the carbodiimide chemistry, which afforded PEO‐g‐MTX conjugates with an amide bond to the aminopteridine ring or carboxylic acid groups of MTX (denoted as PEO‐g‐MTX(COOH) and PEO‐g‐MTX(NH2), respectively). Dynamic light scattering (DLS) revealed that all PEO‐g‐MTX conjugates, with MTX contents varying from 4.8 to 19.6 wt%, existed as unimers in phosphate buffer (PB, pH 7.4, 20 mmol·L−1). Interestingly, MTT assays showed that PEO‐g‐MTX(COOH) exhibited potent anti‐tumor activity in HeLa, A549, KB and NIH3T3 cells with cytotoxicity profiles comparable to that of free MTX. In contrast, PEO‐g‐MTX(NH2) revealed diminishing cytostatic effect with IC50 (half maximal inhibitory concentration) ten to hundred times higher than that of PEO‐g‐MTX(COOH). Moreover, PEO‐g‐MTX(COOH) conjugates allowed facile conjugation with targeting ligands. Notably, folate‐decorated PEO‐g‐MTX(COOH) macromolecular drugs showed apparent targetability to folate receptor‐overexpressing KB cells with an IC50 over 12‐fold lower than non‐targeting PEO‐g‐MTX(COOH) control and about 2‐fold lower than free MTX under otherwise the same conditions.