R406

R406 is an orally available spleen tyrosine kinase (Syk) inhibitor with an IC₅₀ value of 41nM and competitive inhibitor for ATP binding with a K_i of 30nM^[1]. R406 also inhibits the isolated enzymes Lyn with IC₅₀ value of 63nM and Lck with IC₅₀ value of 37nM. Despite similar IC₅₀ values on isolated kinases, R406 shows selectivity in cell-based assays^[2].

In vitro, human diploid fibroblasts were treated with 1, 2, 5, 10 and 20μM of R406 for 24 hours in the growth media. R406 induced apoptotic cell death by inhibiting FAK and p38 activity as well as increasing ROS^[3]. Two patient-derived glioma stem cell (GSC) lines, GSC-1 (Syk-positive) and GSC-2 (Syk-negative), were treated with 0.75–0.89μM R406 for 72h. R406 significantly inhibited neurosphere formation and triggered apoptosis in GSCs. R406 also induced a metabolic shift from glycolysis to oxidative phosphorylation (OXPHOS) and subsequently production of excess ROS in GSCs^[4]. PBMC or purified T cells from chronic lymphocytic leukemia (CLL) patients were pretreated with R406 (0.1 or 1µM) for 30min and then activation markers, cell proliferation and chemotaxis assay were evaluated. R406 impaired the expression of activation markers and cytokine-secretion in response to TCR/CD3 stimulation on T cells. R406 also impaired T cell proliferation in response to TCR/CD3 and IL-15 stimulation and T cell migration in response to CCL21, CCL19 and CXCL12^[5].

In vivo, streptozotocin (STZ) induced diabetic mellitus rat models were given with 5 and 10mg/kg/day R406, respectively via gavage administration for 12 consecutive weeks. R406 could ameliorate STZ induced pericyte loss, acellular capillary formation and retinal vascular leakage; exerted protective effect on blood–retinal barrier; attenuated retinal cell apoptosis; and enhanced retinal cell proliferation^[6]. Mouse chronic migraine (CM) models were pretreated with R406 (5mg/kg) via intraperitoneal administration. R406 significantly ameliorated pain hypersensitivity in CM mice and decreased the protein levels of both CGRP and c-fos in the vicinity of trigeminal nucleus caudalis. R406 inhibited microglia activation and NLRP3 inflammasome at trigeminal nucleus caudalis sites, accompanied by a down-regulation of mature IL-1β expression^[7].

References:
[1] Sylvia Braselmann, Boyle L D, Inoue T, et al. R406, an orally available spleen tyrosine kinase inhibitor blocks fc receptor signaling and reduces immune complex-mediated inflammation. J Pharmacol Exp Ther. 2006 Dec;319(3):998-1008.
[2] Hoon-Suk Cha, Taylor V, Zhao H R, et al. A novel spleen tyrosine kinase inhibitor blocks c-Jun N-terminal kinase-mediated gene expression in synoviocytes. J Pharmacol Exp Ther. 2006 May;317(2):571-8.
[3] Cho H J, Yang E J, Park J T, et al. Identification of SYK inhibitor, R406 as a novel senolytic agent. Aging (Albany NY). 2020 May 7;12(9):8221-8240.
[4] Sun S X, Xue D D, Chen Z J, et al. R406 elicits anti-Warburg effect via Syk-dependent and -independent mechanisms to trigger apoptosis in glioma stem cells. Cell Death Dis. 2019 May 1;10(5):358.
[5] Colado A, Almejún M B, Podaza E, et al. The kinase inhibitors R406 and GS-9973 impair T cell functions and macrophage-mediated anti-tumor activity of rituximab in chronic lymphocytic leukemia patients. Cancer Immunol Immunother. 2017 Apr;66(4):461-473.
[6] Su X, Sun Z H, Ren Q, et al. The effect of spleen tyrosine kinase inhibitor R406 on diabetic retinopathy in experimental diabetic rats. Int Ophthalmol. 2020 Sep;40(9):2371-2383.
[7] Fan Z Z, Su D D, Li Z C, et al. Metformin attenuates central sensitization by regulating neuroinflammation through the TREM2-SYK signaling pathway in a mouse model of chronic migraine. J Neuroinflammation. 2024 Dec 3;21(1):318.

R406是一种口服可用的脾酪氨酸激酶（Syk）抑制剂，IC₅₀值为41nM，并以K_i=30nM竞争性地抑制ATP结合^[1]。R406还能抑制分离酶Lyn，IC₅₀值为63nM，以及Lck，IC₅₀值为37nM。尽管在分离激酶上的IC₅₀值相近，R406在细胞水平实验中表现出选择性^[2]。

体外实验中，将人二倍体成纤维细胞在生长培养基中用1、2、5、10和20μM的R406处理24小时。R406通过抑制FAK和p38活性并增加ROS，诱导细胞凋亡^[3]。用0.75–0.89μM的R406处理两株患者来源的胶质瘤干细胞（GSC），Syk阳性GSC-1和Syk阴性GSC-2，持续72小时。R406显著抑制神经球形成并触发GSCs凋亡。R406还诱导代谢从糖酵解向氧化磷酸化（OXPHOS）转变，随后产生过量ROS^[4]。将慢性淋巴细胞白血病（CLL）患者的外周血单个核细胞（PBMC）或纯化T细胞用R406（0.1或1μM）预处理30分钟，随后评估激活标志物、细胞增殖和趋化实验。R406抑制T细胞在TCR/CD3刺激下的激活标志物表达和细胞因子分泌。R406还抑制T细胞在TCR/CD3和IL-15刺激下的增殖，以及T细胞对CCL21、CCL19和CXCL12的迁移^[5]。

体内实验中，链脲佐菌素（STZ）诱导的糖尿病大鼠模型分别经口给予5和10mg/kg/day的R406，连续12周。R406可减轻STZ诱导的周细胞缺失、无细胞毛细血管形成和视网膜血管渗漏；对血-视网膜屏障发挥保护作用；减轻视网膜细胞凋亡；并增强视网膜细胞增殖^[6]。在慢性偏头痛（CM）小鼠模型中，经腹腔注射R406（5mg/kg）预处理。R406显著减轻CM小鼠的痛觉过敏，并降低三叉神经尾核区域CGRP和c-fos蛋白水平。R406抑制三叉神经尾核部位的小胶质细胞激活和NLRP3炎症小体，伴随成熟IL-1β表达下调^[7]。