Tyrosine kinase inhibitors. 17. Irreversible inhibitors of the epidermal growth factor receptor: 4-(phenylamino)quinazoline- and 4-(phenylamino)pyrido[3,2-d]pyrimidine-6-acrylamides bearing additional solubilizing functions

4-Anilinoquinazoline- and 4-anilinopyrido [3,2-d] pyrimidine-6-acrylamides substituted with solubilizing 7-alkylamine or 7-alkoxyamine side chains wer e prepared by reaction of the corresponding 6-amines with acrylic acid or a crylic acid anhydrides. In the pyrido[3,2-d]pyrimidine series, the intermed iate 6-amino-7-alkylamines were prepared from 7-bromo-6-fluoropyrido[3,2-d] pyrimidine via Stille coupling with the appropriate stannane under palladiu m(0) catalysis. This proved a versatile method for the introduction of cati onic solubilizing side chains. The compounds were evaluated for their inhib ition of phosphorylation of the isolated EGFR enzyme and for inhibition of EGF-stimulated autophosphorylation of EGFR in A431 cells and of heregulin-s timulated autophosphorylation of erbB2 in MBA-MB 453 cells. Quinazoline ana logues with 7-alkoxyamine solubilizing groups were potent irreversible inhi bitors of the isolated EGFR enzyme, with IC50[app] values from 2 to 4 nM, a nd potently inhibited both EG;FR and erbB2 autophosphorylation in cells. 7- Alkylamino- and 7-alkoxyaminopyrido[3,2-d]pyrimidines were also irreversibl e inhibitors with equal or superior potency against the isolated enzyme but were less effective in the cellular autophosphorylation assays. Both quina zoline- and pyrido[3,2-d]pyrimidine-6-acrylamides bound at the ATP site alk ylating cysteine 773, as shown by electrospray ionization mass spectrometry , and had similar rates of absorptive and secretory transport in Caco-2 cel ls. A comparison of two 7-propoxymorpholide analogues showed that the pyrid o[3,2-d]pyrimidine-6-acrylamide had greater amide instability and higher ac rylamide reactivity, being converted to glutathione adducts in cells more r apidly than the corresponding quinazoline. This difference may contribute t o the observed lower cellular potency of the pyrido[3,2-d]pyrimidine-6-acry lamides. Selected compounds showed high in vivo activity against A431 xenog rafts on oral dosing, with the quinazolines being superior to the pyrido [3 ,2-d]pyrimidines. Overall, the quinazolines proved superior to previous ana logues in terms of aqueous solubility, potency, and in vivo antitumor activ ity, and one example (CI 1033) has been selected for clinical evaluation.