在浓度高达 100 μM 的情况下持续 48 小时，SB-505124 对肾上皮 A498 细胞没有毒性。 505124 的 IC50 值为 129±11 nM，比 ALK5 的灵敏度低约 2.5 倍，即使浓度高达 10 μM，也能抑制密切相关的 ALK4，但不会抑制 ALK2。在所有这三种细胞系中，SB-505124 (1 μM) 以浓度依赖性方式抑制 TGF-β 诱导的 Smad2 磷酸化。尽管这些细胞表现出不同的激活模式，SB-505124（1 或 5 μM）仍能有效抑制 TGF-β 诱导的 JNK/SAP、细胞外信号调节激酶 1/2 和 p38 的激活[1]。在体外，SB-505124 (10 µM) 抑制 Smad2 的磷酸化以及 CTGF 和 α-SMA 的表达。通过使用免疫荧光，SB-505124 能够抑制 CTGF 和 α-SMA。虽然用 MMC 处理的眼睛的细胞生长较差，但在 GFS 期间应用 SB-505124 的眼睛解剖的外植体却很强劲[3]。

---

凝胶的特性用于体外药物释放和粘度研究。使用培养的兔结膜下成纤维细胞通过MTT法检测Pluronic®F-127的细胞毒性。体外药物释放研究表明，12小时内药物释放率为100%。MTT法检测凝胶对培养的兔结膜下细胞没有细胞毒性。[2]

在携带 A549 异种移植物的 C57Bl6 小鼠中，SB-505124（5 mg/kg；腹膜内注射）本身没有显示出影响；然而，当与单剂量卡铂 (60 mg/kg) 联合使用时，SB-505124 在 5 个物种中引起了持久的反应，从而无需维持治疗 [4]。

---

本研究旨在研究在青光眼滤过手术（GFS）中使用Pluronic®F-127递送激活素受体样激酶5（ALK-5）抑制剂SB-505124的热可逆凝胶。此外，在体内兔GFS模型的手术部位应用了含有5mg SB-505124的Pluronic®F-127凝胶（18%w/v）。在体外粘度研究中，凝胶的粘度从低温（10°C）到体温（37°C）发生了变化（从1000 cps到45000 cps）。在体内兔GFS模型中，药物通过注射成功递送，没有观察到严重的术后并发症。成功制备并递送了SB-505124热可逆凝胶系统用于兔GFS模型，这可能为GFS提供一种新的递送系统。[2]

---

在体内兔GFS模型中，SB-505124在手术期间以乳糖片剂的形式递送。通过裂隙灯检查眼睛，并测量眼压（IOP），直至滤过泡衰竭或手术后28天。术后第5天的组织切片经苏木精和伊红染色后进行组织学评估。切片还进行了CTGF和α-SMA免疫染色。[3]

临床上，非常需要小分子抑制剂来控制转化生长因子（TGF-β）的致病作用和/或调节TGF-β在正常反应中的作用。预计抑制TGF-β信号传导会增强皮肤伤口的再上皮化，减少瘢痕纤维化。为治疗开发的TGF-β信号通路的选择性小分子抑制剂也将成为实验解剖这一复杂通路的有力工具，特别是它与其他信号通路的串扰。在这项研究中，我们表征了2-（5-苯并[1,3]二氧杂环戊醇-5-基-2-叔丁基-3H-咪唑-4-基）-6-甲基吡啶盐酸盐（SB-505124），这是一类与p38咪唑抑制剂相关的新型小分子抑制剂，可以抑制TGF-βI型受体丝氨酸/苏氨酸激酶，即激活素受体样激酶（ALK）5。我们证明，该化合物选择性地和浓度依赖性地抑制下游细胞质信号转导子Smad2和Smad3以及TGF-β诱导的丝裂原活化蛋白激酶途径成分的ALK4、ALK5和ALK7依赖性激活，但不改变ALK1、ALK2、ALK3或ALK6诱导的Smad信号传导。SB-505124还阻断了TGF-β作用的更复杂的终点，其消除TGF-β1治疗引起的细胞死亡的能力证明了这一点。SB-505124的效力是之前描述的相关ALK5抑制剂SB-431542[1]的三到五倍。

如前所述或通过使用修饰的四唑盐WST-1测量细胞活力。将大约 2000 个细胞接种在 96 孔培养皿中的 100 μL 0.2% FBS 无酚红培养基中过夜。将细胞用 50 μL SB-505124（以达到指定的最终浓度）处理 30 分钟，然后使用或不使用 TGF-β1 和 TNF-α 处理至最终体积 200 μL。通过将每个孔与 10 μL WST-1 在 37°C 下孵育 3 小时，在指定的时间点测量细胞生长。代谢活跃的细胞将 WST-1 裂解为水溶性甲臜，可使用酶联免疫吸附测定板读数器直接定量。每个实验至少进行两次，并且每个细胞系的处理一式三份进行。

Animal/Disease Models: C57Bl6 mice with A549 xenografts [4]
Doses: 5 mg/kg
Route of Administration: intraperitoneal (ip) injection; Daily
Experimental Results: No effect alone, but a single dose of carboplatin (60 mg/kg) gave The drug produced durable responses in 5 animals, requiring no maintenance treatment.

---

In an in vivo rabbit GFS model, SB-505124 was delivered in a lactose tablet during surgery. Eyes were examined by slit-lamp and intraocular pressure (IOP) was measured until the time of bleb failure or up to 28 days after surgery. Tissue sections on day 5 after surgery were histologically evaluated after staining with hematoxylin and eosin. The sections were also immunostained for CTGF and α-SMA.[3]

[1]. SB-505124 is a selective inhibitor of transforming growth factor-beta type I receptors ALK4, ALK5, and ALK7. Mol Pharmacol. 2004 Mar;65(3):744-52.

[2]. Thermoreversible gel for delivery of receptor-like kinase 5 inhibitor SB-505124 for glaucoma filtration surgery. Pharm Dev Technol. 2013 Jul-Aug;18(4):957-62.

[3]. Suppression of transforming growth factor-β effects in rabbit subconjunctival fibroblasts by receptor-like kinase 5 inhibitor. Mol Vis. 2010 Sep 16;16:1880-92.

[4]. Inhibition of activin signaling in lung adenocarcinoma increases the therapeutic index of platinum chemotherapy. Sci Transl Med. 2018 Jul 25;10(451). pii: eaat3504.

Purpose: Transforming growth factor-β (TGF-β) activity has been implicated in subconjunctival scarring in eyes following glaucoma filtration surgery (GFS). The purpose of this study is to determine whether an inhibitor for activin receptor-like kinase (ALK) 5 (also known as TGF-β receptor type I) could suppress TGF-β activity and thereby promote filtering bleb survival after GFS in a rabbit model. Results: The docking study indicated hydrogen bond interactions between SB-505124 and amino acids His-283 and Ser-280 of ALK-5. Suppression of pSmad2, CTGF, and α-SMA by SB-505124 was observed in cultured fibroblasts. Filtering blebs in the GFS with SB-505124 group were maintained for more than 10 days, and the period of bleb survival was significantly longer than that in controls. IOP levels after surgery seemed to be related to bleb survival. Histologically, subconjunctival cell infiltration and scarring at the surgical site in the GFS with SB-505124 and mitomycin C (MMC) groups were much subsided compared to controls. Suppression of CTGF and α-SMA by SB-505124 was also observed by immunofluorescence. Cell outgrowth from explants dissected from eyes to which SB-505124 was applied during GFS was robust while outgrowth was poor from those treated with MMC. Conclusions: The ALK-5 inhibitor SB-505124 was efficacious both in vitro and in vivo in suppressing the TGF-β action. The inhibitor may provide a novel therapy for preventing ocular inflammation and scarring.[3]

---

Resistance to platinum chemotherapy is a long-standing problem in the management of lung adenocarcinoma. Using a whole-genome synthetic lethal RNA interference screen, we identified activin signaling as a critical mediator of innate platinum resistance. The transforming growth factor-β (TGFβ) superfamily ligands activin A and growth differentiation factor 11 (GDF11) mediated resistance via their cognate receptors through TGFβ-activated kinase 1 (TAK1), rather than through the SMAD family of transcription factors. Inhibition of activin receptor signaling or blockade of activin A and GDF11 by the endogenous protein follistatin overcame this resistance. Consistent with the role of activin signaling in acute renal injury, both therapeutic interventions attenuated acute cisplatin-induced nephrotoxicity, its major dose-limiting side effect. This cancer-specific enhancement of platinum-induced cell death has the potential to dramatically improve the safety and efficacy of chemotherapy in lung cancer patients.[4]

C20H22CLN3O2

371.860583782196

371.14

C, 64.60; H, 5.96; Cl, 9.53; N, 11.30; O, 8.60

356559-13-2

SB-505124;694433-59-5

16079009

Light yellow to green yellow solid powder

5.275

60.03

2

4

3

26

466

0

BTUOOXPZOVNPMF-UHFFFAOYSA-N

InChI=1S/C20H21N3O2.ClH/c1-12-6-5-7-14(21-12)18-17(22-19(23-18)20(2,3)4)13-8-9-15-16(10-13)25-11-24-15;/h5-10H,11H2,1-4H3,(H,22,23);1H

2-[4-(1,3-benzodioxol-5-yl)-2-tert-butyl-1H-imidazol-5-yl]-6-methylpyridine;hydrochloride

SB-505124 hydrochloride; SB-505124 (hydrochloride); SB-505124 HCl; CHEMBL1824446; 356559-13-2 (HCl); 2-(5-benzo[1,3]dioxol-5-yl-2-tert-butyl-3h-imidazol-4-yl)-6-methylpyridine hydrochloride; SB505124 hydrochloride;

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

注意： 请将本产品存放在密封且受保护的环境中，避免吸湿/受潮。

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

MEthanol : ~125 mg/mL (~336.15 mM)
DMSO : ~50 mg/mL (~134.46 mM)

配方 1 中的溶解度: ≥ 2.5 mg/mL (6.72 mM) (饱和度未知) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (这些助溶剂从左到右依次添加，逐一添加), 澄清溶液。
例如，若需制备1 mL的工作液，可将100 μL 25.0 mg/mL澄清DMSO储备液加入到400 μL PEG300中，混匀；然后向上述溶液中加入50 μL Tween-80，混匀；加入450 μL生理盐水定容至1 mL。
*生理盐水的制备：将 0.9 g 氯化钠溶解在 100 mL ddH₂O中，得到澄清溶液。

---

配方 2 中的溶解度: ≥ 2.5 mg/mL (6.72 mM) (饱和度未知) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (这些助溶剂从左到右依次添加，逐一添加), 澄清溶液。
例如，若需制备1 mL的工作液，可将 100 μL 25.0 mg/mL澄清DMSO储备液加入900 μL 20% SBE-β-CD生理盐水溶液中，混匀。
*20% SBE-β-CD 生理盐水溶液的制备（4°C，1 周）：将 2 g SBE-β-CD 溶解于 10 mL 生理盐水中，得到澄清溶液。

---

请根据您的实验动物和给药方式选择适当的溶解配方/方案：
1、请先配制澄清的储备液(如：用DMSO配置50 或 100 mg/mL母液(储备液));
2、取适量母液，按从左到右的顺序依次添加助溶剂，澄清后再加入下一助溶剂。以 下列配方为例说明 (注意此配方只用于说明，并不一定代表此产品 的实际溶解配方):
10% DMSO → 40% PEG300 → 5% Tween-80 → 45% ddH2O (或 saline);
假设最终工作液的体积为 1 mL, 浓度为5 mg/mL: 取 100 μL 50 mg/mL 的澄清 DMSO 储备液加到 400 μL PEG300 中，混合均匀/澄清；向上述体系中加入50 μL Tween-80，混合均匀/澄清；然后继续加入450 μL ddH2O (或 saline)定容至 1 mL；
3、溶剂前显示的百分比是指该溶剂在最终溶液/工作液中的体积所占比例;
4、 如产品在配制过程中出现沉淀/析出，可通过加热(≤50℃)或超声的方式助溶;
5、为保证最佳实验结果，工作液请现配现用！
6、如不确定怎么将母液配置成体内动物实验的工作液，请查看说明书或联系我们;
7、 以上所有助溶剂都可在 Invivochem.cn网站购买。