A dual inhibitor of platelet-derived growth factor beta-receptor and Src kinase activity potently interferes with motogenic and mitogenic responses to PDGF in vascular smooth muscle cells. A novel candidate for prevention of vascular remodeling.
PP1 has previously been described as an inhibitor of the Src-family kinases p56(Lck) and FynT. We have therefore decided to use PP1 to determine the functional role of Src in platelet-derived growth factor (PDGF)-induced proliferation and migration of human coronary artery smooth muscle cells (HCASMCs). A synthetic protocol for PP1/AGL1872 has been developed, and the inhibitory activity of PP1/AGL1872 against Src was examined. PP1/AGL1872 potently inhibited recombinant p60(c-src) in vitro and Src-dependent tyrosine phosphorylation in p60(c-srcF572)-transformed NIH3T3 cells. PP1/AGL1872 also potently inhibited PDGF-stimulated migration of HCASMCs, as determined in the modified Boyden chamber, as well as PDGF-stimulated proliferation of HCASMCs. Surprisingly, in addition to inhibition of Src kinase, PP1/AGL1872 was found to inhibit PDGF receptor kinase in cell-free assays and in various types of intact cells, including HCASMCs. PP1/AGL1872 did not inhibit phosphorylation of the vascular endothelial growth factor receptor KDR (VEGF receptor-2; kinase-insert domain containing receptor) in cell-free assays as well as in intact human coronary artery endothelial cells. In line with the insensitivity of KDR, PP1/AGL1872 had only a weak effect on vascular endothelial growth factor-stimulated migration of human coronary artery endothelial cells. On treatment of cells expressing different receptor tyrosine kinases, the activities of the epidermal growth factor receptor, fibroblast growth factor receptor-1, and insulin-like growth factor-1 receptor were resistant to PP1/AGL1872, whereas PDGF alpha-receptor was susceptible, albeit to a lesser extent than PDGF beta-receptor. These data suggest that the previously described tyrosine kinase inhibitor PP1/AGL1872 is not selective for the Src family of tyrosine kinases. It is also a potent inhibitor of the PDGF beta-receptor kinase but is not a ubiquitous tyrosine kinase inhibitor. PP1/AGL1872 inhibits migration and proliferation of HCASMCs probably by interference with 2 distinct tyrosine phosphorylation events, creating a novel and potent inhibitory principle with possible relevance for the treatment of pathological HCASMC activity, such as vascular remodeling and restenosis.
Waltenberger
J
J
Department of Internal Medicine, Ulm University Medical Center, Ulm, Germany. johannes.waltenberger@medizin.uni-ulm.de
Uecker
A
A
Kroll
J
J
Frank
H
H
Mayr
U
U
Bjorge
J D
JD
Fujita
D
D
Gazit
A
A
Hombach
V
V
Levitzki
A
A
Böhmer
F D
FD
D016475:3T3 Cells; D000818:Animals; D000077214:Becaplermin; D001808:Blood Vessels / Q000187:drug effects; D002465:Cell Movement / Q000187:drug effects; D016228:Endothelial Growth Factors / Q000494:pharmacology; D004730:Endothelium, Vascular / Q000166:cytology / Q000502:physiology; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D006801:Humans; D008222:Lymphokines / Q000494:pharmacology; D051379:Mice; D008938:Mitosis / Q000187:drug effects*; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000187:drug effects*; D010982:Platelet-Derived Growth Factor / Q000037:antagonists & inhibitors / Q000494:pharmacology*; D020574:Proto-Oncogene Proteins c-sis; D016392:Proto-Oncogene Proteins pp60(c-src) / Q000037:antagonists & inhibitors; D011720:Pyrazoles / Q000494:pharmacology; D011743:Pyrimidines / Q000494:pharmacology; D020794:Receptor Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors; D020797:Receptor, Platelet-Derived Growth Factor beta; D017479:Receptors, Platelet-Derived Growth Factor / Q000037:antagonists & inhibitors* / Q000378:metabolism; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors; D019061:src-Family Kinases / Q000037:antagonists & inhibitors*
C108123:4-amino-5-(4-methylphenyl)-7-(tert-butyl)pyrazolo(3,4-d)pyrimidine; D016228:Endothelial Growth Factors; D004791:Enzyme Inhibitors; D008222:Lymphokines; D010982:Platelet-Derived Growth Factor; D020574:Proto-Oncogene Proteins c-sis; D011720:Pyrazoles; D011743:Pyrimidines; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors; D000077214:Becaplermin; D020794:Receptor Protein-Tyrosine Kinases; D020797:Receptor, Platelet-Derived Growth Factor beta; D017479:Receptors, Platelet-Derived Growth Factor; D016392:Proto-Oncogene Proteins pp60(c-src); D019061:src-Family Kinases