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Spleen tyrosine kinase (SYK) is a cytosolic non-receptor protein tyrosine kinase that mediates key signal transduction pathways following the activation of immune cell receptors. SYK regulates cellular events induced by the B cell receptor and Fc receptors with high intrinsic activity. Furthermore, SYK has been regarded as an attractive target for the treatment of autoimmune diseases and cancers. Here, we report the crystal structures of SYK in complex with seven newly developed inhibitors (G206, G207, O178, O194, O259, O272, and O282) to provide structural insights into which substituents of the inhibitors and binding regions of SYK are essential for lead compound optimization. Our kinase inhibitors exhibited high inhibitory activities against SYK, with half-maximal inhibitory concentrations (IC50) of approximately 0.7–33 nM, but they showed dissimilar inhibitory activities against KDR, RET, JAK2, JAK3, and FLT3. Among the seven SYK inhibitors, O272 and O282 exhibited highly specific inhibitions against SYK, whereas O194 exhibited strong inhibition of both SYK and FLT3. Three inhibitors (G206, G207, and O178) more efficiently inhibited FLT3 while still substantially inhibiting SYK activity. The binding mode analysis suggested that a highly selective SYK inhibitor can be developed by optimizing the functional groups that facilitate direct interactions with Asn499. This article is protected by copyright. All rights reserved.

Spleen tyrosine kinase (SYK) is a cytosolic non-receptor protein tyrosine kinase. Because SYK mediates key receptor signaling pathways involving the B cell receptor and Fc receptors, SYK is an attractive target for autoimmune disease and cancer treatments. To date, representative oral SYK inhibitors, including fostamatinib (R406 or R788), entospletinib (GS-9973), cerdulatinib (PRT062070), and TAK-659, have been assessed in clinical trials. Here, we report the crystal structures of SYK in complex with two newly developed inhibitors possessing 4-aminopyrido[4,3-D]pyrimidine moieties (SKI-G-618 and SKI-O-85). One SYK inhibitor (SKI-G-618) exhibited moderate inhibitory activity against SYK, whereas the other inhibitor (SKI-O-85) exhibited a low inhibitory profile against SYK. Binding mode analysis indicates that a highly potent SYK inhibitor might be developed by modifying and optimizing the functional groups that interact with Leu377, Gly378, and Val385 in the G-loop and the nearby region in SYK. In agreement with our structural analysis, one of our SYK inhibitor (SKI-G-618) shows strong inhibitory activities on the β-hexosaminidase release and phosphorylation of SYK/Vav in RBL-2H3 cells. Taken together, our findings have important implications for the design of high affinity SYK inhibitors.

The clinical utility of approved EGFR small-molecule kinase inhibitors is plagued both by toxicity against wild-type EGFR and by metastatic progression in the central nervous system, a disease sanctuary site. Here, we report the discovery and preclinical efficacy of GNS-1486 and GNS-1481, two novel small-molecule EGFR kinase inhibitors that are selective for T790M-mutant isoforms of EGFR. Both agents were effective in multiple mouse xenograft models of human lung adenocarcinoma (T790M-positive or -negative), exhibiting less activity against wild-type EGFR than existing approved EGFR kinase inhibitors (including osimertinib). In addition, GNS-1486 showed superior potency against intracranial metastasis of EGFR-mutant lung adenocarcinoma. Our results offer a preclinical proof of concept for new EGFR kinase inhibitors with the potential to improve therapeutic index and efficacy against brain metastases in patients. Cancer Res; 77(5); 1200–11. ©2017 AACR.

A series of pyrazolylpyrimidine scaffold based Syk inhibitors were synthesized and evaluated for their biological activities and selectivity. Lead optimization efforts provided compounds with potent Syk inhibition in both enzymatic and TNF-α release assay.

Aberrant activations of Fms-like tyrosine receptor kinase (FLT) 3 are implicated in the pathogenesis of 20% to 30% of patients with acute myeloid leukemia (AML). G-749 is a novel FLT3 inhibitor that showed potent and sustained inhibition of the FLT3 wild type and mutants including FLT3-ITD, FLT3-D835Y, FLT3-ITD/N676D, and FLT3-ITD/F691L in cellular assays. G-749 retained its inhibitory potency in various drug-resistance milieus such as patient plasma, FLT3 ligand surge, and stromal protection. Furthermore, it displayed potent antileukemic activity in bone marrow blasts from AML patients regardless of FLT3 mutation status, including those with little or only minor responses to AC220 or PKC412. Oral administration of G-749 yielded complete tumor regression and increased life span in animal models. Thus, G-749 appears to be a promising next-generation drug candidate for the treatment of relapsed and refractory AML patients with various FLT3-ITD/FLT3-TKD mutants and further shows the ability to overcome drug resistance.

Human Pim1 kinase is a serine/threonine protein kinase that plays important biological roles in cell survival, apoptosis, proliferation, and differentiation. Moreover, Pim1 is up-regulated in various hematopoietic malignancies and solid tumors. Thus, Pim1 is an attractive target for cancer therapeutics, and there has been growing interest in developing small molecule inhibitors for Pim1. Here, we describe the crystal structure of Pim1 in complex with a newly developed pyrido[4,3-d]pyrimidine-derivative inhibitor (SKI-O-068). Our inhibitor exhibits a half maximum inhibitory concentration (IC50) of 123 (+/- 14) nM and has an unusual binding mode in complex with Pim1 kinase. The interactions between SKI-O-068 and the Pim1 active site pocket residue are different from those of other scaffold inhibitor-bound structures. The binding mode analysis suggests that the SKI-O-068 inhibitor can be improved by introducing functional groups that facilitate direct interaction with Lys67, which aid in the design of an optimized inhibitor.