Design, synthesis, and evaluation of indolinones as inhibitors of the transforming growth factor ß receptor I (TGFßRI).

Inhibition of transforming growth factor ß (TGFß) type I receptor (Alk5) offers a novel approach for the treatment of fibrotic diseases and cancer. Indolinones substituted in position 6 were identified as a new chemotype inhibiting TGFßRI concomitant with a low cross-reactivity among the human kinome. A subset of compounds showed additional inhibition of platelet-derived growth factor receptor alpha (PDGFRα), contributing to an interesting pharmacological profile. In contrast, p38 kinase, which is often inhibited by TGFßRI inhibitors, was not targeted by derivatives based on the indolinone chemotype. Guided by an X-ray structure of lead compound 5 (BIBF0775) soaked into the kinase domain of TGFßRI, optimization furnished potent and selective inhibitors of TGFßRI. Potent inhibition translated well into good inhibition of TGFßRI-mediated phosphorylation of Smad2/3, demonstrating efficacy in a cellular setting. Optimized compounds were extensively profiled on a 232-kinase panel and showed low cross-reactivities within the human kinome.

Characterization and comparative evaluation of a novel PAI-1 inhibitor.

Plasminogen activator inhibitor-1 (PAI-1), the primary physiological inhibitor of both tissue-type plasminogen activator and urokinase-type plasminogen activator in plasma, is a well established risk factor in thrombotic diseases. Reduction of active PAI-1 levels may lead to a decreased tendency of thrombosis. Compounds that can suppress pharmacologically active PAI-1 levels are therefore considered as putative drugs. In the present study, we describe the PAI-1 neutralizing properties and mechanism of a newly selected compound (i.e. fendosal, HP129) in comparison to four previously reported compounds (i. e. AR-H029953XX, XR1853, XR5118 and the peptide TVASS) using different assays. The inhibitory effect of these compounds on active PAl-1 was analyzed by a plasmin-coupled chromogenic assay (Coaset t-PA), direct chromogenic assays (t-PA, u-PA) and quantification of complex formation by ELISA, SDS-PAGE and surface plasmon resonance. Comparative evaluation of the obtained IC50 values reveals large differences [i.e. IC50 of 15 microM (HP129) vs. >1000 microM (XR5118) determined at 37 degrees C using SDS-PAGE] between the compounds studied. Importantly, the relative potency of the various compounds is also dependent on the method used (10 to 170-fold differences in IC50 values). Characterization of the PAI-1 forms (i.e. active, non-reactive and substrate) generated upon inactivation reveals that the newly described compound HP129 induces a unique pathway (i.e. active to non-reactive conversion via a substrate-behaving intermediate) of inactivation compared to the other compounds. Taken together, these data strongly suggest that the various compounds act through different mechanisms. In addition, the results stress the necessity for a careful selection of the method used for the evaluation of PAI-1 inhibitors, preferably requiring a panel of screening methods.