A TYRPHOSTIN-DERIVED INHIBITOR OF PROTEIN-TYROSINE KINASES - ISOLATION AND CHARACTERIZATION

We recently reported that tyrphostin 23 (3,4-dihydroxybenzylidene malo nonitrile) is unstable in solution and that some of the degradation pr oducts are better inhibitors of the tyrosine kinase activity of Src an d the EG;F-receptor kinase than the parent compound itself (Ramdas et al., Cancer Res. 54, 867-868, 1994), In this study, the tyrphostin as- derived compound designated PS, which is a more stable and potent prot ein tyrosine kinase inhibitor, was isolated, P-3 was purified from oxi dized tyrphostin 23 by solvent extraction, silica-gel hash chromatogra phy, and reverse-phase high-pressure liquid chromatography, The physic al characteristics of the isolated compound were determined and its ch emical structure elucidated by H-1 and C-13 NMR spectroscopy, The prop osed structure of this new inhibitor is that of a tyrphostin 23 dimer joined at the benzylidene carbon, P-3 was evaluated in vitro as an inh ibitor of four different protein tyrosine kinases (Src, Csk, EGF-recep tor, and FGF-re ceptor) and two protein serine kinases (PK-A and PKC), This compound exhibited the most inhibitory activity against Src with a K-i value of 6 mu M and was less inhibitory toward the other protei n kinases with K-i values ranging from 35 to 300 E mu M P-3 did not in hibit other nucleotide-utilizing enzymes such as lactate dehydrogenase and hexokinase, The growth and colony formation of HT-29 colon adenoc arcinoma cells that contain activated Src was inhibited by P-3 with an IC50 value of approximately 10 mu M. (C) 1995 Academic Press, Inc.