Acid-sensitive polyethylene glycol conjugates of doxorubicin: Preparation,in vitro efficacy and intracellular distribution

Coupling anticancer drugs to synthetic polymers is a promising approach of enhancing the antitumor efficacy and reducing the side-effects of these age nts. Doxorubicin maleimide derivatives containing an amide or acid-sensitiv e hydrazone linker were therefore coupled to cl-methoxy-poly(ethylene glyco l)-thiopropionic acid amide (MW 20000 Da), alpha,w-bis-thiopropionic acid a mide poly(ethylene glycol) (MW 20000 Da) or alpha-tert-butoxy-poly(ethylene glycol)-thiopropionic acid amide (MW 70000 Da) and the resulting polyethyl ene glycol (PEG) conjugates isolated through size-exclusion chromatography. The polymer drug derivatives were designed as to release doxorubicin insid e the tumor cell by acid-cleavage of the hydrazone bond after uptake of the conjugate by endocytosis. The acid-sensitive PEG conjugates containing the carboxylic hydrazone bonds exhibited in vitro activity against human BXF T 24 bladder carcinoma and LXFL 529L lung cancer cells with IC70 values in th e range 0.02-1.5 mu m (cell culture assay: propidium iodide fluorescence or colony forming assay). In contrast, PEG doxorubicin conjugates containing an amide bond between the drug and the polymer showed no in vitro activity. Fluorescence microscopy studies in LXFL 529 lung cancer cells revealed tha t free doxorubicin accumulates in the cell nucleus whereas doxorubicin of t he acid-sensitive PEG doxorubicin conjugates is primarily localized in the cytoplasm. Nevertheless, the acid-sensitive PEG doxorubicin conjugates reta in their ability to bind to calf thymus DNA as shown by fluorescence and vi sible spectroscopy studies. Results regarding the effect of an acid-sensiti ve PEG conjugate of molecular weight 20000 in the chorioallantoic membrane (CAM) assay indicate that this conjugate is significantly less embryotoxic than free doxorubicin although antiangiogenic effects were not observed. (C ) 1999 Elsevier Science Ltd. All rights reserved.