The combi-targeting concept: in vitro and in vivo fragmentation of a stable combi-nitrosourea engineered to interact with the epidermal growth factor receptor while remaining DNA reactive.
OBJECTIVE
JDA58 (NSC 741282), a "combi-molecule" optimized in the context of the "combi-targeting concept," is a nitrosourea moiety tethered to an anilinoquinazoline. Here, we sought to show its binary epidermal growth factor receptor (EGFR)/DNA targeting property and to study its fragmentation in vitro and in vivo.


METHODS
The fragmentation of JDA58 was detected in cells in vitro and in vivo by fluorescence microscopy and tandem mass spectrometry. EGFR phosphorylation and DNA damage were determined by Western blotting and comet assay, respectively. Tumor data were examined for statistical significance using the Student's t test.


RESULTS
JDA58 inhibited EGFR tyrosine kinase (IC(50), 0.2 micromol/L) and blocked EGFR phosphorylation in human DU145 prostate cancer cells. It induced significant levels of DNA damage in DU145 cells in vitro or in vivo and showed potent antiproliferative activity both in vitro and in a DU145 xenograft model. In cell-free medium, JDA58 was hydrolyzed to JDA35, a fluorescent amine that could be observed in tumor cells both in vitro and in vivo. In tumor cells in vitro or in vivo, or in plasma collected from mice, the denitrosated species JDA41 was the predominant metabolite. However, mass spectrometric analysis revealed detectable levels of the hydrolytic product JDA35 in tumor cells both in vitro and in vivo.


CONCLUSIONS
The results in toto suggest that growth inhibition in vitro and in vivo may be sustained by the intact combi-molecule plus JDA35 plus JDA41, three inhibitors of EGFR, and the concomitantly released DNA-damaging species. This leads to a model wherein a single molecule carries a complex multitargeted-multidrug combination.
Qiu
Qiyu
Q
Cancer Drug Research Laboratory, Department of Medicine, McGill University Health Center/Royal Victoria Hospital, Montreal, Quebec, Canada.
Domarkas
Juozas
J
Banerjee
Ranjita
R
Merayo
Nuria
N
Brahimi
Fouad
F
McNamee
James P
JP
Gibbs
Bernard F
BF
Jean-Claude
Bertrand J
BJ
D000818:Animals; D045744:Cell Line, Tumor; D020552:Comet Assay; D004247:DNA / Q000737:chemistry*; D004249:DNA Damage; D004354:Drug Screening Assays, Antitumor; D066246:ErbB Receptors / Q000378:metabolism*; D006801:Humans; D020128:Inhibitory Concentration 50; D008297:Male; D013058:Mass Spectrometry; D051379:Mice; D008856:Microscopy, Fluorescence; D009368:Neoplasm Transplantation; D009607:Nitrosourea Compounds / Q000494:pharmacology*; D010766:Phosphorylation
D009607:Nitrosourea Compounds; D004247:DNA; D066246:ErbB Receptors