ALK5 12 nM (IC50) ALK4 45 nM (IC50) ALK7 7.5 nM (IC50)

A83-01 sodium is a strong inhibitor of type I lymph node receptor ALK7, type I activin/lymph node receptor ALK4, and TGF-β type I receptor ALK5 kinase. It can lessen the amount of transcription that ALK-5 induces. ALK4-TD and ALK7-TD-induced transcription is also blocked by IC50 of Mv1Lu 12 nM in cells. The expression of constitutively active ALK-6, ALK-2, ALK-3, and ALK-1 induction is weakly inhibited by the IC50 in R4-2 cells, which is 45 nM and 7.5 nM, respectively. Effectively blocking the growth inhibitory impact of TGF-β, A 83-01 sodium (0.03-10 μM) totally suppresses this action at 3 μM. In HaCaT cells, TGF-β-induced Smad activation is inhibited by 83-01 sodium (1–10 μM) [1]. While TGF-β1 increases cell motility, adhesion, and invasion in HM-1 cells, 83-01 sodium (1 μM) does not alter cell proliferation [2].

Mice without body weight or neurobehavioral symptoms can have a considerably higher survival rate when administered with 83-01 (50, 150, and 500 μg/mouse) intraperitoneally [2]. In mice bearing M109 cells, 83-01 sodium (0.5 mg/kg, ip) sodium shows strong antitumor activity [3].

The original constructions of constitutively active forms of ALK-1 through -7 in mammalian expression vectors were described previously. The 9xCAGA-luciferase plasmid contains nine repeats of the CAGA Smad binding element driving luciferase expression. The (BRE)2-luciferase plasmid contains two repeats of the BMP responsive elements of the Id1 promoter cloned upstream of a minimal promoter driving luciferase expression. The 3GC2-luciferase plasmid contains three repeats of a GC-rich sequence derived from the proximal BMP responsive element of the Smad6 promoter. [1]

Mv1Lu cells were seeded in duplicate at a density of 2.5 × 104 cells/well in 24-well plates. The following day, cells were pretreated for 1 h with 1 µM small molecule inhibitors and then cultured with TGF-β 1 ng/mL) for 24 h, 48 h, or 72 h. Cells were trypsinized and counted with a Coulter counter. To explore whether small molecule inhibitors reduced the growth-inhibitory effects of TGF-β in concentration-dependent fashion, Mv1Lu cells were seeded as above and pretreated for 1 h with various concentrations of small molecule inhibitors. After pretreatment, cells were cultured with TGF-β 1 ng/mL) for 48 h and counted.[1]

Female B6C3F1 mice used for the in vivo studies are maintained under specific pathogen-free conditions. To evaluate the effect of A 83-01 on the survival of mice bearing peritoneal dissemination, HM-1 cells (1×106) are injected into the abdominal cavity via the left flank of the mouse. Starting the next day, A 83-01 (150 μg/body) or vehicles (PBS with 0.5% DMSO) are injected into the abdominal cavity three times per week. Mice are euthanized before reaching the moribund state.

[1]. The ALK-5 inhibitor A-83-01 inhibits Smad signaling and epithelial-to-mesenchymal transition by transforming growth factor-beta. Cancer Sci. 2005 Nov;96(11):791-800.

[2]. The activated transforming growth factor-beta signaling pathway in peritoneal metastases is a potential therapeutic target in ovarian cancer. Int J Cancer. 2012 Jan 1;130(1):20-8.

[3]. Enhanced antitumor efficacy of folate-linked liposomal doxorubicin with TGF-β type I receptor inhibitor. Cancer Sci. 2010 Oct;101(10):2207-13.

Transforming growth factor β (TGF-β) signaling pathway promotes tumor growth and metastasis in advanced cancers. Therefore, using TGF-β signaling pathway inhibitors may be a novel strategy for treating patients with such cancers. In this study, we synthesized and characterized a small molecule inhibitor, A-83-01, whose structure is similar to the ALK-5 inhibitor developed by Sawyer et al. (2003), and which can block the signaling of type I serine/threonine kinase receptors of TGF-β superfamily cytokines (also known as activin receptor-like kinases; ALK). Using a TGF-β-responsive reporter gene construct from mammalian cells, we found that A-83-01 can inhibit the transcriptional activity induced by TGF-β type I receptor ALK-5, activin IB receptor ALK-4, and Nodal I receptor ALK-7, among which the kinase domains of activin IB and Nodal I receptors are structurally highly correlated with ALK-5. A-83-01 exhibits stronger inhibition of ALK-5 than the previously reported ALK-5 inhibitor SB-431542, and is able to prevent Smad2/3 phosphorylation and TGF-β-induced growth inhibition. In contrast, A-83-01 has little effect on bone morphogenetic protein type I receptor, p38 mitogen-activated protein kinase, or extracellular signal-regulated kinase. Consistent with these findings, A-83-01 inhibits TGF-β-induced epithelial-mesenchymal transition, suggesting that A-83-01 and its related molecules may help prevent the progression of advanced cancer. [1] Peritoneal dissemination, including omental metastasis, is the most common metastatic pathway in advanced ovarian cancer and an important prognostic factor. We used binary regression analysis on the publicly available microarray dataset (GSE2109) and found that the transforming growth factor (TGF)-β signaling pathway was activated in omental metastases compared to the primary lesion. Immunohistochemical analysis of TGF-β receptor type 2 and phosphorylated SMAD2 showed that both were upregulated in omental metastases compared to the primary lesion. Treatment of the mouse ovarian cancer cell line HM-1 with recombinant TGF-β1 promoted its invasiveness, cell migration, and cell adhesion, while treatment with the TGF-β signaling pathway inhibitor A-83-01 inhibited these effects. Microarray analysis of HM-1 cells treated with TGF-β1 and/or A-83-01 revealed that A-83-01 effectively inhibited TGF-β1-induced transcriptional changes. Gene set enrichment analysis revealed that genes upregulated by TGF-β1 in HM-1 cells were also significantly upregulated in omental metastases from the human ovarian cancer dataset GSE2109 (false discovery rate (FDR) q = 0.086). We also investigated the therapeutic effect of A-83-01 in a mouse peritoneal metastasis model. Intraperitoneal injection of A-83-01 (150 μg three times a week) significantly improved survival (p = 0.015). In summary, these results suggest that the activated TGF-β signaling pathway in peritoneal metastases is a potential therapeutic target for ovarian cancer. [2]

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Drug carriers targeting tumor cells are a promising strategy that utilizes various ligand linkages to enhance the therapeutic potential of chemotherapeutic drugs. Folic acid is a high-affinity ligand for the folate receptor, which is a functional tumor-specific receptor. The transforming growth factor (TGF)-β type I receptor (TβR-I) inhibitor A-83-01 is expected to enhance the accumulation of nanocarriers in tumors by altering the microvascular environment. To enhance the therapeutic effect of folate-linked liposomal doxorubicin (F-SL), we co-administered F-SL with A-83-01. The changes in tumor-associated angiogenesis induced by intraperitoneal injection of A-83-01 were examined by magnetic resonance imaging (MRI) and histological analysis. The targeted efficacy of a single intravenous injection of F-SL combined with A-83-01 was evaluated by measuring the biodistribution and antitumor effect in a tumor-bearing mouse lung cancer M109 model. About 3 hours after injection of A-83-01, tumor vasculature underwent transient changes. 24 hours after injection, A-83-01 resulted in 1.7 times higher accumulation of F-SL in the tumor compared to liposomes alone. Furthermore, the antitumor activity of F-SL combined with A-83-01 was significantly higher than that of F-SL alone. This study suggests that the combined use of TβR-I inhibitors will open up new strategies for the application of FR-targeted nanocarriers in cancer treatment. [3]

C25H19N5NAS

443.118

2828431-89-4

A 83-01;909910-43-6;A 83-01;909910-43-6

139034128

Typically exists as White to light yellow solids at room temperature

0

5

3

32

615

0

QEDFBXIADXHNKE-UHFFFAOYSA-M

InChI=1S/C25H19N5S.Na/c1-17-8-7-13-23(27-17)24-21(19-14-15-26-22-12-6-5-11-20(19)22)16-30(29-24)25(31)28-18-9-3-2-4-10-18;/h2-16H,1H3,(H,28,31);/q;+1/p-1

sodium;[3-(6-methylpyridin-2-yl)-4-quinolin-4-ylpyrazole-1-carbothioyl]-phenylazanide

A 83-01 sodium salt; 2828431-89-4; 3-(6-Methylpyridin-2-yl)-N-phenyl-4-(quinolin-4-yl)-1H-pyrazole-1-carbothioamide, sodium salt; A 83-01 sodium; G16241; sodium N-[3-(6-methylpyridin-2-yl)-4-(quinolin-4-yl)pyrazole-1-carbothioyl]anilinide; sodium;[3-(6-methylpyridin-2-yl)-4-quinolin-4-ylpyrazole-1-carbothioyl]-phenylazanide

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: (1). Please store this product in a sealed and protected environment (e.g. under nitrogen), avoid exposure to moisture and light.  (2). This product is not stable in solution, please use freshly prepared working solution for optimal results.

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

DMSO: 100 mg/mL (224.97 mM)

Solubility in Formulation 1: 5 mg/mL (11.25 mM) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), suspension solution; with heating and sonication.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 50.0 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly.
Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.

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Solubility in Formulation 2: ≥ 2.5 mg/mL (5.62 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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Solubility in Formulation 3: ≥ 2.5 mg/mL (5.62 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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Solubility in Formulation 4: 5% DMSO + 40% PEG300 + 5% Tween80 + 50% Saline

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 (Please use freshly prepared in vivo formulations for optimal results.)