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 (TGF)-beta signaling facilitates tumor growth and metastasis in advanced cancer. Use of inhibitors of TGF-beta signaling may thus be a novel strategy for the treatment of patients with such cancer. In this study, we synthesized and characterized a small molecule inhibitor, A-83-01, which is structurally similar to previously reported ALK-5 inhibitors developed by Sawyer et al. (2003) and blocks signaling of type I serine/threonine kinase receptors for cytokines of the TGF-beta superfamily (known as activin receptor-like kinases; ALKs). Using a TGF-beta-responsive reporter construct in mammalian cells, we found that A-83-01 inhibited the transcriptional activity induced by TGF-beta type I receptor ALK-5 and that by activin type IB receptor ALK-4 and nodal type I receptor ALK-7, the kinase domains of which are structurally highly related to those of ALK-5. A-83-01 was found to be more potent in the inhibition of ALK5 than a previously described ALK-5 inhibitor, SB-431542, and also to prevent phosphorylation of Smad2/3 and the growth inhibition induced by TGF-beta. In contrast, A-83-01 had little or no effect on bone morphogenetic protein type I receptors, p38 mitogen-activated protein kinase, or extracellular regulated kinase. Consistent with these findings, A-83-01 inhibited the epithelial-to-mesenchymal transition induced by TGF-beta, suggesting that A-83-01 and related molecules may be useful for preventing the progression of advanced cancers.[1]

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Peritoneal dissemination including omental metastasis is the most frequent route of metastasis and an important prognostic factor in advanced ovarian cancer. We analyzed the publicly available microarray dataset (GSE2109) using binary regression and found that the transforming growth factor (TGF)-beta signaling pathway was activated in omental metastases as compared to primary sites of disease. Immunohistochemical analysis of TGF-beta receptor type 2 and phosphorylated SMAD2 indicated that both were upregulated in omental metastases as compared to primary disease sites. Treatment of the mouse ovarian cancer cell line HM-1 with recombinant TGF-β1 promoted invasiveness, cell motility and cell attachment while these were suppressed by treatment with A-83-01, an inhibitor of the TGF-β signaling pathway. Microarray analysis of HM-1 cells treated with TGF-β1 and/or A-83-01 revealed that A-83-01 efficiently inhibited transcriptional changes that are induced by TGF-β1. Using gene set enrichment analysis, we found that genes upregulated by TGF-β1 in HM-1 cells were also significantly upregulated in omental metastases compared to primary sites in the human ovarian cancer dataset, GSE2109 (false discovery rate (FDR) q = 0.086). Therapeutic effects of A-83-01 in a mouse model of peritoneal dissemination were examined. Intraperitoneal injection of A-83-01 (150 μg given three times weekly) significantly improved survival (p = 0.015). In summary, these results show that the activated TGF-β signaling pathway in peritoneal metastases is a potential therapeutic target in ovarian cancer. [2]

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Tumor cell targeting of drug carriers is a promising strategy and uses the attachment of various ligands to enhance the therapeutic potential of chemotherapy agents. Folic acid is a high-affinity ligand for folate receptor, which is a functional tumor-specific receptor. The transforming growth factor (TGF)-β type I receptor (TβR-I) inhibitor A-83-01 was expected to enhance the accumulation of nanocarriers in tumors by changing the microvascular environment. To enhance the therapeutic effect of folate-linked liposomal doxorubicin (F-SL), we co-administrated F-SL with A-83-01. Intraperitoneally injected A-83-01-induced alterations in the cancer-associated neovasculature were examined by magnetic resonance imaging (MRI) and histological analysis. The targeting efficacy of single intravenous injections of F-SL combined with A-83-01 was evaluated by measurement of the biodistribution and the antitumor effect in mice bearing murine lung carcinoma M109. A-83-01 temporarily changed the tumor vasculature around 3 h post injection. A-83-01 induced 1.7-fold higher drug accumulation of F-SL in the tumor than liposome alone at 24 h post injection. Moreover F-SL co-administrated with A-83-01 showed significantly greater antitumor activity than F-SL alone. This study shows that co-administration of TβR-I inhibitor will open a new strategy for the use of FR-targeting nanocarriers for 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.)