BelumosudilKD025SLx-2119AbMoleM9209是口服有效的 Rho 相关卷曲螺旋蛋白激酶 2ROCK2选择性抑制剂。ROCK1 与 ROCK2 是 Rho GTP 酶下游的丝氨酸/苏氨酸激酶调控细胞骨架动力学、细胞迁移、增殖与存活。尽管两种亚型高度同源功能分工却存在差异ROCK2 在免疫细胞与神经、血管组织中优势表达特异性驱动 Th17/Tfh 促炎应答与促纤维化转录程序。Belumosudil 对 ROCK2 的 IC50 约 60-105 nMKi 41 nM对 ROCK1 的 IC50 约 24 μM亚型选择性超过 200 倍为区分 ROCK 亚型功能提供了高分辨率工具。分子层面BelumosudilCAS No.911417-87-3竞争性占据 ROCK2 的 ATP 结合口袋通过两条并行机制重塑细胞表型。在免疫细胞中ROCK2 与磷酸化 STAT3 结合并参与 JAK2/STAT3 复合物形成Belumosudil抑制 ROCK2 后 STAT3 磷酸化及其对 IL-17、IL-21 启动子的占据下降IRF4、RORγt、Bcl6 等转录因子表达下调Th17 与 Tfh 细胞应答受抑同时 ROCK2 与 JAK3 的相互作用增强STAT5 磷酸化与 Foxp3 表达上调调节性 T 细胞功能增强Th17/Treg 平衡向免疫稳态方向偏移。Belumosudil在间质细胞中的ROCK2 抑制活性能导致RhoA-ROCK 驱动的肌动蛋白聚合受阻MRTF 核转位减少胶原、α-SMA、CTGF 等促纤维化基因转录下降。BelumosudilKD025SLx-2119在人外周血 T 细胞体外刺激体系中可使 IL-21、IL-17 分泌分别下降约 90% 与 60%而不影响 IFN-γsiRNA 敲低 ROCK2而非 ROCK1可复现该表型间接证实Belumosudil的生物学效应是通过抑制ROCK2实现的。在大鼠肺微血管内皮细胞PMVEC中7.5 μM的 Belumosudil 能剂量依赖性抑制糖酵解、促进氧化磷酸化、降低细胞内 pH 并抑制细胞迁移有趣的是该代谢重编程不被 ROCK2 敲低复现提示Belumosudil存在 ROCK2 非依赖的代谢调控效应。BelumosudilCAS No.911417-87-3的体内研究涵盖缺血、自身免疫与纤维化等多个模型。在小鼠短暂性大脑中动脉栓塞tMCAO模型中Belumosudil 剂量依赖性缩小梗死体积治疗时间窗至少 3 h疗效维持 4 周以上且不像非选择性 ROCK 抑制剂那样引起低血压远端 MCAO 模型显示其可改善皮层灌注、增强侧支血流。在胶原诱导关节炎CIA小鼠模型中200 mg/kg的Belumosudil 腹腔给药后能抑制Th17 通路和小鼠的关节炎进展。在慢性移植物抗宿主病cGVHD小鼠模型中150 mg/kg的Belumosudil 干预能使肺功能恢复正常、减少肺组织抗体与胶原沉积脾脏 Tfh 减少而 Tfr 增多。在博来霉素Bleomycin大鼠肺纤维化模型中饮水给予 Belumosudil 可降低肺血管通透性。综上BelumosudilKD025SLx-2119AbMoleM9209作为亚型选择性优异的 ROCK2 抑制剂为解析 ROCK2 在免疫失衡、血管张力与器官纤维化中的亚型特异性功能提供了工具。其免疫调节与抗纤维化的双重机制、良好的动物实验效果使其广泛应用于 Th17/Treg 平衡、缺血再灌注与纤维化大/小鼠模型的体内机制研究。* 本文所述均为科研试剂仅供科学研究参考参考文献[1] Zanin-Zhorov A, et al. Selective oral ROCK2 inhibitor down-regulates IL-21 and IL-17 secretion in human T cells via STAT3-dependent mechanism. Proceedings of the National Academy of Sciences of the United States of America, 2014, 111(47): 16814-16819.[2] Flynn R, et al. Targeted Rho-associated kinase 2 inhibition suppresses murine and human chronic GVHD through a Stat3-dependent mechanism. Blood, 2016, 127(17): 2144-2154.[3] Lee J H, et al. Selective ROCK2 inhibition in focal cerebral ischemia. Annals of Clinical and Translational Neurology, 2014, 1(1): 2-14.[4] Lee J Y, et al. KD025 shifts pulmonary endothelial cell bioenergetics and decreases baseline lung permeability. American Journal of Respiratory Cell and Molecular Biology, 2020, 63(4): 519-530.[5] Julian L, Olson M F. Rho-associated coiled-coil containing kinases (ROCK): structure, regulation, and functions. Small GTPases, 2014, 5(2): e29846.细胞实验参考细胞系Human telomerase reverse transcriptase-immortalized retinal pigment epithelial (hTERT RPE-1) cell line方法Cells were seeded in culture plates or glass coverslips and cultured to subconfluence, then treated with serially diluted belumosudil; cilia incidence and cilia length were quantified by acetylated α-tubulin immunofluorescence staining followed by confocal laser scanning microscopy; ROCK2 inhibitory efficacy was verified via Western blotting detection of phosphorylated MYPT1 (Thr696) protein level; cell viability was assessed by trypan blue exclusion to rule out non-specific cytotoxicity浓度0.5, 1, 2.5, 5 μM处理时间48 h参考文献PLoS Biol. 2025 Jul 24;23 (7):e3003275上述方法来自公开文献仅供相同目的实验参考。如实验目的、材料、方法不同请参考其他文献。动物实验参考动物模型Female BALB/c recipient mice, 8–10 weeks old, sclerodermatous chronic graft-versus-host disease model induced by allogeneic bone marrow plus splenocyte transplantation from C57BL/6 donor mice配制The complete solvent composition was not described in original paper剂量30, 100, 300 mg/kg给药处理Oral gavage, once daily, initiated on the day of allogeneic cell transplantation and sustained for 8 consecutive weeks; clinical cGVHD scores were monitored weekly; target organ (skin, lung, liver) histopathology, collagen deposition quantification and immune cell phenotyping were performed at experimental endpoint; serum inflammatory cytokine profiles were detected by multiplex immunoassay参考文献Drugs. 2021 Oct;81 (15):1787-1796上述方法来自公开文献仅供相同目的实验参考。如实验目的、材料、方法不同请参考其他文献。体内实验的工作液建议现用现配当天使用如在配制过程中出现沉淀、析出现象可以通过超声和或加热的方式助溶切勿直接全部溶解 AbMole 抑制剂篇丨 Belumosudil（KD025，SLx-2119）：选择性 ROCK2 抑制剂，在免疫平衡与纤维化机制研究中的应用
BelumosudilKD025SLx-2119AbMoleM9209是口服有效的 Rho 相关卷曲螺旋蛋白激酶 2ROCK2选择性抑制剂。ROCK1 与 ROCK2 是 Rho GTP 酶下游的丝氨酸/苏氨酸激酶调控细胞骨架动力学、细胞迁移、增殖与存活。尽管两种亚型高度同源功能分工却存在差异ROCK2 在免疫细胞与神经、血管组织中优势表达特异性驱动 Th17/Tfh 促炎应答与促纤维化转录程序。Belumosudil 对 ROCK2 的 IC50 约 60-105 nMKi 41 nM对 ROCK1 的 IC50 约 24 μM亚型选择性超过 200 倍为区分 ROCK 亚型功能提供了高分辨率工具。分子层面BelumosudilCAS No.911417-87-3竞争性占据 ROCK2 的 ATP 结合口袋通过两条并行机制重塑细胞表型。在免疫细胞中ROCK2 与磷酸化 STAT3 结合并参与 JAK2/STAT3 复合物形成Belumosudil抑制 ROCK2 后 STAT3 磷酸化及其对 IL-17、IL-21 启动子的占据下降IRF4、RORγt、Bcl6 等转录因子表达下调Th17 与 Tfh 细胞应答受抑同时 ROCK2 与 JAK3 的相互作用增强STAT5 磷酸化与 Foxp3 表达上调调节性 T 细胞功能增强Th17/Treg 平衡向免疫稳态方向偏移。Belumosudil在间质细胞中的ROCK2 抑制活性能导致RhoA-ROCK 驱动的肌动蛋白聚合受阻MRTF 核转位减少胶原、α-SMA、CTGF 等促纤维化基因转录下降。BelumosudilKD025SLx-2119在人外周血 T 细胞体外刺激体系中可使 IL-21、IL-17 分泌分别下降约 90% 与 60%而不影响 IFN-γsiRNA 敲低 ROCK2而非 ROCK1可复现该表型间接证实Belumosudil的生物学效应是通过抑制ROCK2实现的。在大鼠肺微血管内皮细胞PMVEC中7.5 μM的 Belumosudil 能剂量依赖性抑制糖酵解、促进氧化磷酸化、降低细胞内 pH 并抑制细胞迁移有趣的是该代谢重编程不被 ROCK2 敲低复现提示Belumosudil存在 ROCK2 非依赖的代谢调控效应。BelumosudilCAS No.911417-87-3的体内研究涵盖缺血、自身免疫与纤维化等多个模型。在小鼠短暂性大脑中动脉栓塞tMCAO模型中Belumosudil 剂量依赖性缩小梗死体积治疗时间窗至少 3 h疗效维持 4 周以上且不像非选择性 ROCK 抑制剂那样引起低血压远端 MCAO 模型显示其可改善皮层灌注、增强侧支血流。在胶原诱导关节炎CIA小鼠模型中200 mg/kg的Belumosudil 腹腔给药后能抑制Th17 通路和小鼠的关节炎进展。在慢性移植物抗宿主病cGVHD小鼠模型中150 mg/kg的Belumosudil 干预能使肺功能恢复正常、减少肺组织抗体与胶原沉积脾脏 Tfh 减少而 Tfr 增多。在博来霉素Bleomycin大鼠肺纤维化模型中饮水给予 Belumosudil 可降低肺血管通透性。综上BelumosudilKD025SLx-2119AbMoleM9209作为亚型选择性优异的 ROCK2 抑制剂为解析 ROCK2 在免疫失衡、血管张力与器官纤维化中的亚型特异性功能提供了工具。其免疫调节与抗纤维化的双重机制、良好的动物实验效果使其广泛应用于 Th17/Treg 平衡、缺血再灌注与纤维化大/小鼠模型的体内机制研究。* 本文所述均为科研试剂仅供科学研究参考参考文献[1] Zanin-Zhorov A, et al. Selective oral ROCK2 inhibitor down-regulates IL-21 and IL-17 secretion in human T cells via STAT3-dependent mechanism. Proceedings of the National Academy of Sciences of the United States of America, 2014, 111(47): 16814-16819.[2] Flynn R, et al. Targeted Rho-associated kinase 2 inhibition suppresses murine and human chronic GVHD through a Stat3-dependent mechanism. Blood, 2016, 127(17): 2144-2154.[3] Lee J H, et al. Selective ROCK2 inhibition in focal cerebral ischemia. Annals of Clinical and Translational Neurology, 2014, 1(1): 2-14.[4] Lee J Y, et al. KD025 shifts pulmonary endothelial cell bioenergetics and decreases baseline lung permeability. American Journal of Respiratory Cell and Molecular Biology, 2020, 63(4): 519-530.[5] Julian L, Olson M F. Rho-associated coiled-coil containing kinases (ROCK): structure, regulation, and functions. Small GTPases, 2014, 5(2): e29846.细胞实验参考细胞系Human telomerase reverse transcriptase-immortalized retinal pigment epithelial (hTERT RPE-1) cell line方法Cells were seeded in culture plates or glass coverslips and cultured to subconfluence, then treated with serially diluted belumosudil; cilia incidence and cilia length were quantified by acetylated α-tubulin immunofluorescence staining followed by confocal laser scanning microscopy; ROCK2 inhibitory efficacy was verified via Western blotting detection of phosphorylated MYPT1 (Thr696) protein level; cell viability was assessed by trypan blue exclusion to rule out non-specific cytotoxicity浓度0.5, 1, 2.5, 5 μM处理时间48 h参考文献PLoS Biol. 2025 Jul 24;23 (7):e3003275上述方法来自公开文献仅供相同目的实验参考。如实验目的、材料、方法不同请参考其他文献。动物实验参考动物模型Female BALB/c recipient mice, 8–10 weeks old, sclerodermatous chronic graft-versus-host disease model induced by allogeneic bone marrow plus splenocyte transplantation from C57BL/6 donor mice配制The complete solvent composition was not described in original paper剂量30, 100, 300 mg/kg给药处理Oral gavage, once daily, initiated on the day of allogeneic cell transplantation and sustained for 8 consecutive weeks; clinical cGVHD scores were monitored weekly; target organ (skin, lung, liver) histopathology, collagen deposition quantification and immune cell phenotyping were performed at experimental endpoint; serum inflammatory cytokine profiles were detected by multiplex immunoassay参考文献Drugs. 2021 Oct;81 (15):1787-1796上述方法来自公开文献仅供相同目的实验参考。如实验目的、材料、方法不同请参考其他文献。体内实验的工作液建议现用现配当天使用如在配制过程中出现沉淀、析出现象可以通过超声和或加热的方式助溶切勿直接全部溶解