Ciliobrevin D

Ciliobrevin D is an inhibitor of dynein ATPase activity, preventing the cycling activity of the motor protein^[1]. Ciliobrevin D has been used to investigate dynein's role in axonal transport and in the targeting of membrane proteins and organelles‌^[2][3][5]. Ciliobrevin D can also interfere with Hedgehog signaling and primary cilia formation^[4].

In vitro, chicken dorsal root ganglion (DRG) neurons were treated with Ciliobrevin D (20μM) for 45min prevented both retrograde and anterograde movement of mitochondria. The percentage of motile lysosomes was reduced. DRG cultures were incubated in 20μM Ciliobrevin D for 60 minutes also reversibly inhibits axon extension^[2]. T-ALL cells MOLT3 were pre-treated with Ciliobrevin D (20µM) for 24h and then with vehicle or pan-HDAC inhibitor Trichostatin A (TSA) (0.5µM) for 16h. TSA treatment reduced Notch3 surface levels and increased accumulation of Notch3 protein in the lysosomal compartment. Surface Notch3 levels were rescued by inhibition of dynein with Ciliobrevin D^[3]. Human retinal pigment epithelial (hTERT RPE-1) cells were treated with either 50μM Ciliobrevin D, 10nM MI-181, 100nM MI-181, 50μM Ciliobrevin D + 10nM MI-181, or 50μM Ciliobrevin D + 100nM MI-181 for 24h after serum withdrawal. Ciliobrevin D treatment alone led to a marked decrease in the average length of cilia and the percentage of ciliated cells. The cilia length defects and decrease in percent ciliation observed in Ciliobrevin D treated cells were rescued to near control levels when cells were cotreated with MI-181^[4].

In vivo, 5.7μL of 90μM Ciliobrevin D was stereotaxically injected into cerebral lateral ventricle of C57BL/6 mice 45min before adeno-associated virus (AAV)-9 variant, AAV.CAP-B10, administration. Ciliobrevin D restrained AAV.CAP-B10 transduction in the hippocampus^[5]. Ciliobrevin D was used at 15μM per testis (assuming a testis volume of ~1.6ml at 1.6g) on adult male Sprague-Dawley rats. The needle was inserted from the apical to the basal end of the testis vertically. Ciliobrevin D treatment induced defects in spermatogenesis by leading to significant disorganization of the F-actin network and compromising the blood-testis barrier integrity^[6].

References:
[1] See S K, Hoogendoorn S, Chung A H, et al. Cytoplasmic Dynein Antagonists with Improved Potency and Isoform Selectivity. ACS Chem Biol. 2016 Jan 15;11(1):53-60.
[2] Sainath R, Gallo G. The dynein inhibitor Ciliobrevin D inhibits the bidirectional transport of organelles along sensory axons and impairs NGF-mediated regulation of growth cones and axon branches.Dev Neurobiol. 2015 Jul;75(7):757-77.
[3] Pinazza M, Ghisi M, Minuzzo S, et al. Histone deacetylase 6 controls Notch3 trafficking and degradation in T-cell acute lymphoblastic leukemia cells. Oncogene. 2018 Jul;37(28):3839-3851.
[4] Gholkar A A, Gimeno T V, Edgemon J E, et al. MI-181 Modulates Cilia Length and Restores Cilia Length in Cells with Defective Shortened Cilia. ACS Chem Biol. 2024 Aug 16;19(8):1733-1742.
[5] Wang J C, Ding S H, He Z H,et al. Dynein-regulated brain transduction of AAV.CAP-B10 via cerebral lateral ventricle enhances hippocampal function after traumatic brain injury through Ngf gene delivery in mice. Exp Neurol. 2025 May 13:391:115285.
[6] Wen Q, Tang E I, Lui W Y, et al. Dynein 1 supports spermatid transport and spermiation during spermatogenesis in the rat testis. Am J Physiol Endocrinol Metab. 2018 Nov 1;315(5):E924-E948.

Ciliobrevin D是一种动力蛋白ATP酶活性抑制剂，可阻止马达蛋白的动力循环活动‌^[1]。Ciliobrevin D已被用于研究动力蛋白在轴突运输以及膜蛋白和细胞器靶向中的作用^[2][3][5]。Ciliobrevin D还可干扰Hedgehog信号通路和原发性纤毛的形成^[4]。

体外实验中，鸡背根神经节（DRG）神经元经20μM Ciliobrevin D处理45min后，线粒体的顺行和逆行运动均被抑制，运动性溶酶体的百分比也降低。DRG培养物在20μM Ciliobrevin D中孵育60min，可以可逆性地抑制轴突延伸^[2]。T-ALL细胞MOLT3先用20μM Ciliobrevin D预处理24h，再用载体或泛HDAC抑制剂三氟司他（TSA）（0.5μM）处理16h。TSA处理降低了Notch3的表面水平，并增加了Notch3蛋白在溶酶体中的积累。通过Ciliobrevin D抑制动力蛋白可恢复Notch3的表面水平^[3]。人视网膜色素上皮（hTERT RPE-1）细胞在血清撤除后，分别用50μM Ciliobrevin D、10nM MI-181、100nM MI-181、50μM Ciliobrevin D+10nM MI-181或50μM Ciliobrevin D+100nM MI-181处理24h。单独使用Ciliobrevin D处理会导致纤毛平均长度明显缩短，纤毛细胞百分比降低。Ciliobrevin D处理细胞中观察到的纤毛长度缺陷和纤毛百分比下降，当细胞与MI-181联合处理时，可恢复至接近对照水平^[4]。

体内实验中，将5.7μL的90μM Ciliobrevin D立体定向注射到C57BL/6小鼠脑侧脑室，于腺相关病毒（AAV）-9变体AAV.CAP-B10给药前45min进行。Ciliobrevin D限制了AAV.CAP-B10在海马区的转导^[5]。在成年雄性Sprague-Dawley大鼠上，按每睾丸15μM Ciliobrevin D（假设睾丸体积约为1.6ml，重1.6g）进行处理，将针从睾丸的顶端垂直插入到底端。Ciliobrevin D处理导致精子发生缺陷，表现为F-肌动蛋白网络显著紊乱，破坏了血-睾屏障的完整性^[6]。