Dorsomorphin (Compound C) 2HCl|1219168-18-9|安捷凯

Background

IC50: Dorsomorphin inhibited BMP4-induced phosphorylation of BMP-responsive SMADs in a dose-dependent manner (half maximal inhibitory concentration (IC50) =0.47 mM).
Bone morphogenetic protein (BMP) signals coordinate developmental patterning and have essential physiological roles in mature organisms. The first known small-molecule inhibitor of BMP signaling, dorsomorphin, were identified in a screen for compounds that perturb dorsoventral axis formation in zebrafish.
In vitro: Previoius researchers found that dorsomorphin selectively inhibits the BMP type I receptors ALK2/ALK3/ALK6 leading to block BMP-mediated SMAD1/5/8 phosphorylation, osteogenic differentiation as well as target gene transcription. Using dorsomorphin, they examined the role of BMP signaling in iron homeostasis.dorsomorphin inhibited the systemic iron regulator hepcidin BMP-, hemojuvelin- and interleukin 6–stimulated expression, indicating that BMP receptors regulate hepcidin induction by all of these stimuli [1].
In vivo: The systemic challenge with iron rapidly induced SMAD1/5/8 phosphorylation and hepcidin expression in the liver, while dorsomorphin treatment could blocke SMAD1/5/8 phosphorylation, normalize hepcidin expression and increase serum iron levels. These suggest an crucial physiological role for hepatic BMP signaling in iron-hepcidin homeostasis [1].
Clinical trial: Dorsomorphin is still in preclinical development stage and no clinicl trial is ongoing currently.

IC50:Dorsomorphin 以剂量依赖性方式抑制 BMP4 诱导的 BMP 反应性 SMAD 磷酸化（半数最大抑制浓度 (IC50) =0.47 mM）。
骨形态发生蛋白 (BMP) 信号协调发育模式和在成熟生物体中具有重要的生理作用。第一个已知的 BMP 信号转导小分子抑制剂 dorsomorphin 在筛选干扰斑马鱼背腹轴形成的化合物时被鉴定出来。
体外:Previioius 研究人员发现，dorsomorphin 选择性抑制 BMP I 型受体 ALK2/ALK3/ALK6，从而阻断 BMP 介导的 SMAD1/5/8 磷酸化、成骨分化以及靶基因转录。他们使用 dorsomorphin 检查了 BMP 信号在铁稳态中的作用。dorsomorphin 抑制全身铁调节剂铁调素 BMP-、血幼素和白细胞介素 6 刺激的表达，表明 BMP 受体通过所有这些刺激调节铁调素诱导 [1]。
在体内:铁的全身性刺激会迅速诱导肝脏中的 SMAD1/5/8 磷酸化和 hepcidin 表达，而 dorsomorphin 治疗可以阻断 SMAD1/5/8 磷酸化，使 hepcidin 表达正常化并增加血清铁水平。这些表明肝脏 BMP 信号转导在铁-铁调素稳态中的重要生理作用 [1]。
临床试验:Dorsomorphin尚处于临床前开发阶段，目前尚无临床试验。

Reference:
[1] Yu PB, Hong CC, Sachidanandan C, Babitt JL, Deng DY, Hoyng SA, Lin HY, Bloch KD, Peterson RT.Dorsomorphin inhibits BMP signals required for embryogenesis and iron metabolism. Nat Chem Biol. 2008;4(1):33-41.

Protocol

Kinase experiment [1]:

AMPK partial purification and in vitro kinase assay

Liver AMPK is partially purified from male SD rats to the blue-Sepharose step. The 100-μl reaction mixture contains 100 μM AMP, 100 μM ATP (0.5 μCi 33P-ATP per reaction), and 50 μM SAMS in a buffer (40 mM HEPES, pH 7.0, 80 mM NaCl, 0.8 mM EDTA, 5 mM MgCl2, 0.025% BSA, and 0.8 mM DTT). The reaction is initiated with addition of the enzyme. After 30-minute incubation at 30°C, the reaction is stopped by addition of 80 μl 1% H3PO4. Aliquots (100 μl) are transferred to 96-well MultiScreen plates. The plate is washed three times with 1% H3PO4 followed by detection in a Top-count. The in vitro AMPK inhibition data obtained with compound C — (6-[4-(2-Piperidin-1-yl-ethoxy)-phenyl)]-3-pyridin-4-yl-pyyrazolo[1,5-a] pyrimidine — are fit to the following equation for competitive inhibition by nonlinear regression using a least-squares Marquardt algorithm in a computer program written by N. Thornberry of Merck Research Laboratories: Vi/Vo = (Km + S)/[S + Km × (1 + I/Ki)], where Vi is the inhibited velocity, Vo is the initial velocity, S is the substrate (ATP) concentration, Km is the Michaelis constant for ATP, I is the inhibitor (compound C) concentration, and Ki is the dissociation constant for compound C.

Cell experiment [2]:

Cell lines

mouse pulmonary artery smooth muscle cells (PASMCs), Hep3B cells

Preparation method

The solubility of this compound in DMSO is limited. General tips for obtaining a higher concentration: Please warm the tube at 37℃ for 10 minutes and/or shake it in the ultrasonic bath for a while. Stock solution can be stored below -20℃ for several months.

Reacting condition

0.1-20 μM for 30 min

Applications

Dorsomorphin (4 μM) inhibited osteogenic differentiation in C2C12 cells and suppressed BMP-mediated SMAD activation by blocking BMP type I receptor function. Moreover, treatment with dorsomorphin (1 μM) blocked BMP- and HJV-mediated hepcidin expression in cultured hepatoma-derived cells.

Animal experiment [2]:

Animal models

Zebrafish embryos model; Wild-type (WT) C57BL/6 adult mice model

Dosage form

10 mM dorsomorphin for 30 h; or 10 mg/ kg, intraperitoneal injection

Applications

Dorsomorphin induced dorsalization in zebrafish embryos model. Moreover, Dorsomorphin inhibited bone mineralization in vivo. Additionally, Dorsomorphin (10 mg/ kg) induced hyperferremia in mice.

Other notes

Please test the solubility of all compounds indoor, and the actual solubility may slightly differ with the theoretical value. This is caused by an experimental system error and it is normal.

参考文献:

1. Zhou, G., Myers, R., Li, Y., Chen, Y., Shen, X., Fenyk-Melody, J., Wu, M., Ventre, J., Doebber, T., Fujii, N., Musi, N., Hirshman, M. F., Goodyear, L. J. and Moller, D. E. (2001) Role of AMP-activated protein kinase in mechanism of metformin action. J Clin Invest. 108, 1167-1174

2. Yu, P. B., Hong, C. C., Sachidanandan, C., Babitt, J. L., Deng, D. Y., Hoyng, S. A., Lin, H. Y., Bloch, K. D. and Peterson, R. T. (2008) Dorsomorphin inhibits BMP signals required for embryogenesis and iron metabolism. Nat Chem Biol. 4, 33-41