RESUMEN
A series of new 6-substituted-4-(3-bromophenylamino)quinazoline derivatives that may function as irreversible inhibitors of epidermal growth factor receptor (EGFR) and human epidermal growth factor receptor (HER-2) tyrosine kinases have been prepared. These inhibitors have, at the C-6 position, butynamide, crotonamide, and methacrylamide Michael acceptors bearing water-solublilizing substituents. These compounds were prepared by acylation of 6-amino-4-(3-bromophenylamino)quinazoline with unsaturated acid chlorides or mixed anhydrides. We show that attaching a basic functional group onto the Michael acceptor results in greater reactivity, due to intramolecular catalysis of the Michael addition and/or an inductive effect of the protonated basic group. This, along with improved water solubility, results in compounds with enhanced biological properties. We present molecular modeling and experimental evidence that these inhibitors interact covalently with the target enzymes. One compound, 16a, was shown to have excellent oral activity in a human epidermoid carcinoma (A431) xenograft model in nude mice.
Asunto(s)
Antineoplásicos/síntesis química , Inhibidores Enzimáticos/síntesis química , Receptores ErbB/antagonistas & inhibidores , Quinazolinas/síntesis química , Receptor ErbB-2/antagonistas & inhibidores , Administración Oral , Animales , Antineoplásicos/química , Antineoplásicos/farmacología , Western Blotting , División Celular/efectos de los fármacos , Ensayos de Selección de Medicamentos Antitumorales , Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/farmacología , Femenino , Fluorometría , Glutatión/química , Humanos , Espectroscopía de Resonancia Magnética , Ratones , Ratones Desnudos , Modelos Moleculares , Fosforilación , Pruebas de Precipitina , Quinazolinas/química , Quinazolinas/farmacología , Trasplante Heterólogo , Células Tumorales CultivadasRESUMEN
The substrate specificity for phospholipase D from Streptomyces chromofuscus (PLD(Sc)) has been determined utilizing an assay based on the quantitation of inorganic phosphate. 1,2-Di-n-hexanoyl phosphatidylcholine (C6PC), phosphatidylethanolamine (C6PE), phosphatidylserine (C6PS), phosphatidylglycerol (C6PG), and an unnatural phospholipid bearing a neohexyl headgroup (C6PDB) were examined as substrates. The assay relies on the quenching of the PLD(Sc)-catalyzed hydrolysis of the phospholipid substrates with EDTA followed by the hydrolysis of the phosphatidic acid product with alkaline phosphatase. The inorganic phosphate thus released is quantitated through the formation of a complex with ammonium molybdate, which has an absorbance maximum at 700 nm. To minimize the time involved and the reagents consumed, the assay is conducted in 96-well plates. The results of this study indicate that the catalytic efficiency for PLD(Sc) on the substrates is C6PC >> C6PS approximately C6PE > C6PG >> C6PDB.