Abstract A273: TAS-5567, a highly potent and selective inhibitor of SYK kinase, demonstrated antitumor activity in diffuse large B-cell lymphoma (DLBCL) cellsin vitro and in vivo.
Authors: Irie Hiroki , Kawagishi Aki , Ishihara Keiko , Tsuji Shingo , Nakatsuru Yoko , Kuze Jiro , Fujino Naoko , Ogino Yoshio , Komatani Hideya , Utsugi Teruhiro 
Published: August 2014

Abstract
Background: Chronic activation of the B-cell receptor (BCR) pathway is critical for proliferation and survival of malignant B cells, and is implicated in the pathogenesis of B-cell lymphomas. Spleen tyrosine kinase (SYK) is a central regulator of the BCR pathway. Although there are several reports of SYK inhibitory compounds in development, there appears to still be a need for a highly selective SYK inhibitor. Here we report on a novel SYK inhibitor, TAS-5567, which displays high potency with excellent kinase selectivity. In this study, we characterized the in vitro cytotoxic effect and the mechanism of action of TAS-5567 in DLBCL cells, and evaluated in vivo antitumor activity in DLBCL xenograft models.
Materials and Methods: In vitro enzyme inhibition of SYK was determined by using purified SYK and the peptide substrate. Selectivity for kinases of TAS-5567 was determined in a panel of 196 kinases. To evaluate the target engagement of TAS-5567, the cellular phosphorylation of BLNK(Y84), a substrate of SYK kinase, was examined by Flow cytometry. For a growth inhibition assay, DLBCL cells were treated with TAS-5567 for 3 days, and the number of living cells was determined by measuring cellular ATP. In order to demonstrate TAS-5567’s mechanism of action in DLBCL cells, a cell cycle analysis was conducted by Flow cytometry. For evaluation of pharmacokinetics (PK), pharmacodynamics (PD), and antitumor effect of TAS-5567 in vivo, we orally dosed TAS-5567 in human DLBCL xenograft models.
Results: TAS-5567 was a potent inhibitor of SYK with an IC50 value of 0.37nM, and demonstrated little activities against the other 196 kinases. In SU-DHL10 DLBCL cells, TAS-5567 potently inhibited the phosphorylation of BLNK (IC50 = 10 nM), and inhibited cell proliferation in a concentration-dependent manner (IC50 = 68 nM). Cell cycle arrest at G1 phase and apoptosis were observed in the DLBCL cells by continuous exposure to TAS-5567 at 300 nM. Also, TAS-5567 exhibited broad antiproliferation activity across various DLBCL cell lines in vitro. In vivo, TAS-5567 demonstrated favorable oral PK properties in mice, and exhibited a sustained and robust inhibition of the pharmacodynamics marker pBLNK in tumor tissues of SU-DHL10 DLBCL xenograft model. Consistent with the results of the pharmacodynamics study, TAS-5567 displayed significant antitumor effect in the SU-DHL10 xenograft models at PD effective doses with minimum body weight changes.
Conclusion: TAS-5567 is an orally available, highly potent, and selective inhibitor of SYK. TAS-5567 demonstrated substantial antitumor activity in DLBCL cells both in vitro and in vivo, with an accompanying sustained and robust inhibition of SYK in the cells. These data suggest that TAS-5567 may be a promising SYK inhibitor for the treatment of DLBCL.
Citation Information: Mol Cancer Ther 2013;12(11 Suppl):A273.
Citation Format: Hiroki Irie, Aki Kawagishi, Keiko Ishihara, Shingo Tsuji, Yoko Nakatsuru, Jiro Kuze, Naoko Fujino, Yoshio Ogino, Hideya Komatani, Teruhiro Utsugi. TAS-5567, a highly potent and selective inhibitor of SYK kinase, demonstrated antitumor activity in diffuse large B-cell lymphoma (DLBCL) cellsin vitro and in vivo. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr A273.