title: Design and synthesis of novel tyrosine kinase inhibitors using a pharmacophore model of the ATP-binding site of the EGF-R.
abstract: One of the most promising targets for the rational design of anti-cancer drugs is the family of the EGF-receptor protein tyrosine kinases. Despite the high sequence homology within the ATP-binding region of protein tyrosine and/or serine threonine kinases, ATP-competitive compounds have the potential to be selective inhibitors of protein kinases. Dianilino-phthalimides CGP 52 411 and CGP 53,353 have been identified as potent and ATP-competitive inhibitors of the EGF-R tyrosine kinase with no or only minor activity against a panel of tyrosine and serine/threonine kinases. Using a calculated 3-D computer model of the catalytic domain of the EGF-R-tyrosine kinase together with CGP 52 411 as example of an ATP-competitive inhibitor, a pharmacophore model for ATP-competitive inhibitors in the active site of the EGF-R PTK was developed. With the help of this model, 4-phenylamino-7H-pyrrolo[2,3-d]pyrimidines were then identified as new potent EGF-R PTK inhibitors. In an interactive process, the class of the 4-phenylamino-pyrrolo-pyrimidines was optimized and structure-activity-relationship of a series of derivatives thereof are discussed. In vitro, the most active compounds (CGP 59 326, CGP 60 261, CGP 62 706) inhibited the EGF-R tyrosine kinase with IC50 value between 6-30 nM. High selectivity towards a panel of non-receptor tyrosine kinases (c-SRC, v-Abl) and serine/threonine kinases (PKC alpha, PKA) was observed. Kinetic analysis revealed competitive type kinetics relative to ATP. In cells, EGF-stimulated cellular tyrosine phosphorylation was inhibited by these compounds at IC50 values between 0.1-0.3 microM, whereas the ligand-induced receptor autophosphorylation of the PDGR-R was not effected by concentrations up to 100 microM. Furthermore, CGP 59 326, CGP 60 261, CGP 62 706 were able to selectively inhibit c-fos mRNA expression in EGF-dependent cell lines with (IC50) approx. 0.1-1 microM) but not in EGF-independent cell systems (IC50 > 100 microM). Proliferation of the EGF-dependent MK cell line was inhibited with similar IC50 values. In addition, CGP 59 326 and CGP 62 706 showed good in vivo efficacy at low doses after oral or subcutaneous administration in nude mice tumor models using xenografts of the EGF-dependent A431 cell lines. The ED50 values were between 1.5-2 mg/kg. Phenylamino-pyrrolo-pyrimidines therefore represent a new series of tyrosine kinase inhibitors which preferentially inhibit the EGF-mediated signal transduction pathway and have the characteristics for further evaluation as anticancer agents.
authors.affiliation: CIBA Pharmaceuticals Division, Cancer and Bone Metabolism Research Department, CIBA Limited, Basel, Switzerland.
authors.forename: P
authors.initials: P
authors.lastname: Traxler
authors.forename: P
authors.initials: P
authors.lastname: Furet
authors.forename: H
authors.initials: H
authors.lastname: Mett
authors.forename: E
authors.initials: E
authors.lastname: Buchdunger
authors.forename: T
authors.initials: T
authors.lastname: Meyer
authors.forename: N
authors.initials: N
authors.lastname: Lydon
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism*; D000818:Animals; D001665:Binding Sites / Q000187:drug effects; D015195:Drug Design; D004791:Enzyme Inhibitors / Q000138:chemical synthesis* / Q000494:pharmacology*; D066246:ErbB Receptors / Q000737:chemistry* / Q000187:drug effects*; D006801:Humans; D008958:Models, Molecular; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors*
chemical_list: D004791:Enzyme Inhibitors; D000255:Adenosine Triphosphate; D066246:ErbB Receptors; D011505:Protein-Tyrosine Kinases