Cheng Kui, Gao Meng, Godfroy James I, Brown Peter N, Kastelowitz Noah, Yin Hang
Center of Basic Molecular Science, Department of Chemistry, Tsinghua University, Beijing 100082, China.
Department of Chemistry and Biochemistry and the BioFrontiers Institute, University of Colorado Boulder, Boulder, CO 80309, USA.
Sci Adv. 2015;1(3). doi: 10.1126/sciadv.1400139.
Toll-like receptor (TLR) agonists activate both the innate and the adaptive immune systems. These TLR agonists have been exploited as potent vaccine adjuvants and antitumor agents. We describe the identification and characterization of a small molecule, -methyl-4-nitro-2-(4-(4-(trifluoromethyl)phenyl)-1 -imidazol-1-yl)aniline (CU-T12-9), that directly targets TLR1/2 to initiate downstream signaling. CU-T12-9 specifically induces TLR1/2 activation, which can be blocked by either the anti-hTLR1 or the anti-hTLR2 antibody, but not the anti-hTLR6 antibody. Using a variety of different biophysical assays, we have demonstrated the binding mode of CU-T12-9. By binding to both TLR1 and TLR2, CU-T12-9 facilitates the TLR1/2 heterodimeric complex formation, which in turn activates the downstream signaling. Fluorescence anisotropy assays revealed competitive binding to the TLR1/2 complex between CU-T12-9 and PamCSK with a half-maximal inhibitory concentration (IC) of 54.4 nM. Finally, we showed that CU-T12-9 signals through nuclear factor κB (NF-κB) and invokes an elevation of the downstream effectors tumor necrosis factor-α (TNF-α), interleukin-10 (IL-10), and inducible nitric oxide synthase (iNOS). Thus, our studies not only provide compelling new insights into the regulation of TLR1/2 signaling transduction but also may facilitate future therapeutic developments.
Toll样受体（TLR）激动剂可激活先天性和适应性免疫系统。这些TLR激动剂已被用作有效的疫苗佐剂和抗肿瘤药物。我们描述了一种小分子化合物——α-甲基-4-硝基-2-（4-（4-（三氟甲基）苯基）-1H-咪唑-1-基）苯胺（CU-T12-9）的鉴定和特性，该化合物直接靶向TLR1/2以启动下游信号传导。CU-T12-9特异性诱导TLR1/2激活，这可被抗人TLR1或抗人TLR2抗体阻断，但不能被抗人TLR6抗体阻断。通过使用各种不同的生物物理分析方法，我们证明了CU-T12-9的结合模式。通过与TLR1和TLR2两者结合，CU-T12-9促进了TLR1/2异二聚体复合物的形成，进而激活下游信号传导。荧光偏振分析显示CU-T12-9与PamCSK之间对TLR1/2复合物存在竞争性结合，其半数抑制浓度（IC50）为54.4 nM。最后，我们表明CU-T12-9通过核因子κB（NF-κB）发出信号，并引起下游效应分子肿瘤坏死因子-α（TNF-α）、白细胞介素-10（IL-10）和诱导型一氧化氮合酶（iNOS）的升高。因此，我们的研究不仅为TLR1/2信号转导的调控提供了引人注目的新见解，而且可能促进未来的治疗发展。
