| Size | Price | Stock | Qty |
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| 5mg |
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| Other Sizes |
| Targets |
Activin-like kinase 5 (ALK5, also known as TGF-beta receptor type I, TGFBR1). TGFbetaRI-IN-2 is an ATP-competitive inhibitor that binds to the kinase domain of ALK5. It has a pIC50 of 7.6 (IC50 ∼25 nM) and a pEC50 of 6.63 (EC50 ∼234 nM) in cellular assays. At 1 uM, it also inhibits MAP3K2 (MEKK2) by 31%, with an IC50 value of 2.2 uM, indicating moderate selectivity. It is selective over other ALK receptors and related kinases.
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| ln Vitro |
At 1 μM, TP-008 inhibits MAP3K2 (MEKK2) by 31%, with an IC50 value of 2.2 μM[1].
In vitro, TGFbetaRI-IN-2 potently inhibits ALK5 autophosphorylation and the subsequent phosphorylation of downstream SMAD2/3 proteins. In TGF-beta-responsive cell lines (e.g., Mv1Lu mink lung epithelial cells, human HaCaT keratinocytes), it blocks TGF-beta1-induced SMAD2/3 nuclear translocation and transcriptional activity. It suppresses TGF-beta1-induced epithelial-mesenchymal transition (EMT) markers (e.g., reduced E-cadherin, increased vimentin). It also inhibits TGF-beta1-induced collagen production in human dermal fibroblasts (IC50 ∼100 nM). It has no significant effect on the BMP (bone morphogenetic protein) pathway, which signals through ALK1/2/3/6. |
| ln Vivo |
TP-008 (oral; 50, 150, and 500 mg/kg; 5 days) causes cardiovascular toxicity that manifests as hemorrhage, fibrin deposition in heart valves, and proliferation of valvular interstitial cells [ 1].
In vivo, oral administration of TGFbetaRI-IN-2 at 50, 150, and 500 mg/kg for 5 days induces cardiovascular toxicity, characterized by valvular interstitial cell proliferation, neutrophil infiltration, hemorrhage, and fibrin deposition in the heart valves. This cardiotoxicity is an on-target effect of prolonged ALK5 inhibition, as TGF-beta signaling is essential for maintaining heart valve integrity. No in vivo efficacy studies have been reported. It is predicted to have antifibrotic activity in models of renal, hepatic, or pulmonary fibrosis at lower, non-cardiotoxic doses, but this has not been confirmed. |
| Enzyme Assay |
Non-cellular kinase inhibition assays are performed using recombinant human ALK5 kinase domain (0.5-2 ug) in reaction buffer (50 mM HEPES pH 7.5, 10 mM MgCl2, 1 mM EGTA, 2 mM DTT, 0.01% Tween-20, 10 uM ATP, and 0.2 mg/mL poly(Glu4Tyr) peptide substrate). Varying concentrations of TGFbetaRI-IN-2 (0.1-1000 nM) are added, and the reaction is incubated for 60 minutes at 30degC. The incorporation of phosphate is detected using an anti-phosphotyrosine antibody in an ELISA format (or by radiolabeled 33P-ATP). The IC50 is calculated from the inhibition curve. Selectivity profiling against a panel of >50 kinases (including ALK2, ALK3, ALK4, MAP2K2, MAP3K2) is performed at 1 uM to determine specificity.
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| Cell Assay |
Mv1Lu (mink lung epithelial) cells stably transfected with a SMAD2/3-responsive luciferase reporter gene (CAGA12-Luc) are used. Cells are seeded in 96-well plates (2×104 cells/well) in DMEM with 1% FBS. After 24 hours, cells are pre-treated with TGFbetaRI-IN-2 (0.1-1000 nM) for 1 hour, then stimulated with TGF-beta1 (1-2 ng/mL) for 16-24 hours. Luciferase activity is measured using a luciferase assay reagent. The EC50 is defined as the concentration that inhibits the TGF-beta1-induced luciferase signal by 50%. For SMAD2/3 phosphorylation, cells are treated with compound (0.1-1000 nM) plus 1 ng/mL TGF-beta1 for 1-2 hours, and cell lysates are analyzed by western blot using anti-phospho-SMAD2 (Ser465/467) and anti-phospho-SMAD3 (Ser423/425) antibodies.
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| Animal Protocol |
Animal/Disease Models: Rat[1]
Doses: 50, 150 and 500 mg/kg Route of Administration: Oral; 50, 150 and 500 mg/kg; 5 days Experimental Results: Heart valve disease was induced in both the medium-dose and high-dose animal groups. No in vivo efficacy studies have been reported. A toxicology study in rodents (likely rats or mice) showed that oral administration of TGFbetaRI-IN-2 at doses of 50, 150, and 500 mg/kg for 5 days induces dose-dependent cardiovascular toxicity. Animals are administered the compound (formulated in 0.5% methylcellulose or similar vehicle) by oral gavage once daily. At the end of the 5-day treatment period, animals are euthanized, hearts are harvested, and heart valves (aortic, mitral, tricuspid, pulmonary) are examined by histopathology. Staining with hematoxylin and eosin (H&E) and Masson's trichrome reveals valvular interstitial cell proliferation, neutrophil infiltration, hemorrhage, and fibrin deposition. The NOAEL (no-observed-adverse-effect level) for cardiotoxicity is likely below 50 mg/kg. |
| ADME/Pharmacokinetics |
No detailed PK data have been reported. As an orally active ALK5 inhibitor with a pIC50 of 7.6, TGFbetaRI-IN-2 is expected to be absorbed with moderate oral bioavailability. Based on its physicochemical properties (Molecular weight: 411.82, LogP ∼3-4, H-bond donors: 1, H-bond acceptors: 7), it likely has a moderate terminal half-life (2-6 hours) in rodents. The high oral doses required to observe cardiotoxicity (50-500 mg/kg) suggest that compound exposure may be limited by solubility or metabolism, or that the therapeutic window is narrow. Detailed PK parameters (Cmax, Tmax, AUC, t½, F%) have not been reported.
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| Toxicity/Toxicokinetics |
In vivo, TGFbetaRI-IN-2 is cardiotoxic at high doses (50-500 mg/kg oral, daily for 5 days). The cardiac lesions include valvular interstitial cell proliferation (reminiscent of myxomatous valve disease), neutrophil infiltration, hemorrhage, and fibrin deposition in the heart valves. This is an on-target class effect of ALK5 inhibitors, as TGF-beta signaling is critical for maintaining the quiescence of valvular interstitial cells and for endothelial integrity. No other significant toxicities (e.g., hepatotoxicity, nephrotoxicity, gastrointestinal toxicity) have been reported at these high doses. At lower doses, the compound is expected to be well-tolerated with no significant adverse effects. No genotoxicity or hERG data have been reported.
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| References | |
| Additional Infomation |
TGFbetaRI-IN-2 (TP-008) is a research-grade ALK5 inhibitor for studying the role of TGF-beta signaling in cellular processes such as EMT, fibrosis, and cancer metastasis. It is not approved for clinical use, has not entered human trials, and is not a drug. It serves as a tool compound for target validation, but its clinical development is likely limited by on-target cardiotoxicity observed in vivo. The cardiotoxicity of ALK5 inhibitors (also known as TGFBR1 inhibitors) has been well documented (e.g., galunisertib, LY364947) and represents a major challenge for their therapeutic application in chronic diseases. This compound is available from chemical suppliers for research use only.
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| Molecular Formula |
C₂₀H₁₅CLFN₅O₂
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|---|---|
| Molecular Weight |
411.82
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| Exact Mass |
411.089
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| CAS # |
1976038-41-1
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| PubChem CID |
145925651
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| Appearance |
White to off-white solid powder
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| LogP |
2
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| Hydrogen Bond Donor Count |
1
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| Hydrogen Bond Acceptor Count |
5
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| Rotatable Bond Count |
4
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| Heavy Atom Count |
29
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| Complexity |
640
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| Defined Atom Stereocenter Count |
0
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| SMILES |
CC1=CN=C(C=C1N2C3=C(C=NC=C3)N(C2=O)CC(=O)N)C4=C(C=CC(=C4)Cl)F
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| InChi Key |
LVEUPFUJRKZPEN-UHFFFAOYSA-N
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| InChi Code |
InChI=1S/C20H15ClFN5O2/c1-11-8-25-15(13-6-12(21)2-3-14(13)22)7-17(11)27-16-4-5-24-9-18(16)26(20(27)29)10-19(23)28/h2-9H,10H2,1H3,(H2,23,28)
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| Chemical Name |
2-[1-[2-(5-chloro-2-fluorophenyl)-5-methylpyridin-4-yl]-2-oxoimidazo[4,5-c]pyridin-3-yl]acetamide
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| Synonyms |
TGFβRIIN2; TGFβRI IN 2
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| HS Tariff Code |
2934.99.9001
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| Storage |
Powder -20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month |
| Shipping Condition |
Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)
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| Solubility (In Vitro) |
May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples
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| Solubility (In Vivo) |
Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.
Injection Formulations
Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)(e.g. IP/IV/IM/SC) *Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution. Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline) Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil) Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals). View More
Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)] Oral Formulations
Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium) Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals). View More
Oral Formulation 3: Dissolved in PEG400 (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 2.4282 mL | 12.1412 mL | 24.2825 mL | |
| 5 mM | 0.4856 mL | 2.4282 mL | 4.8565 mL | |
| 10 mM | 0.2428 mL | 1.2141 mL | 2.4282 mL |
*Note: Please select an appropriate solvent for the preparation of stock solution based on your experiment needs. For most products, DMSO can be used for preparing stock solutions (e.g. 5 mM, 10 mM, or 20 mM concentration); some products with high aqueous solubility may be dissolved in water directly. Solubility information is available at the above Solubility Data section. Once the stock solution is prepared, aliquot it to routine usage volumes and store at -20°C or -80°C. Avoid repeated freeze and thaw cycles.
Calculation results
Working concentration： mg/mL；
Method for preparing DMSO stock solution： mg drug pre-dissolved in μL DMSO (stock solution concentration mg/mL). Please contact us first if the concentration exceeds the DMSO solubility of the batch of drug.
Method for preparing in vivo formulation:：Take μL DMSO stock solution, next add μL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL ddH2O，mix and clarify.
(1) Please be sure that the solution is clear before the addition of next solvent. Dissolution methods like vortex, ultrasound or warming and heat may be used to aid dissolving.
(2) Be sure to add the solvent(s) in order.
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