Synthesis and studies on three-compartment flavone-containing combi-molecules designed to target EGFR, DNA, and MEK.

In order to induce a tandem targeting of EGFR, DNA, and MEK, we built complex combi-molecules containing an EGFR targeting quinazoline and an aminoethyltriazene moiety linking the entire molecule to PD98059. Two complex molecules were synthesized: one with a short aminoethyl spacer, AL232, and the other AL414 with a longer aminoethylaminoethyl spacer. AL414 was a more potent inhibitor of EGFR tyrosine kinase than AL232. Both combi-molecules blocked EGFR phosphorylation in whole cells and downregulated extracellular signaling-regulated kinases (ERK1,2). However, only AL414 was capable of inducing DNA damage. Thus, it was taken in vivo for metabolic analysis. The results showed that 3 h after injection, AL414 was hydrolyzed to an EGFR inhibitor FD105, which was further acetylated to FD105Ac, a more potent inhibitor of EGFR. The detected flavone derivative was PD98059 linked to the hydroxyalkyl moiety resulting from the decomposition of the alkyldiazonium species. Independent synthesis of the latter metabolite and further in vitro analysis showed that it was deprived of antiproliferative activity. The results in toto suggest that while AL414 is a three-compartment combi-molecule, only the EGFR and DNA targeting species can be released and the cleavage to the intact MEK inhibitor PD98059 was mitigated by the stability of the carbamate.