| Size | Price | Stock | Qty |
|---|---|---|---|
| 5mg |
|
||
| 10mg |
|
||
| Other Sizes |
| Targets |
FTO (IC50 <1 μM); FTO-IN-1 targets fat mass and obesity-associated protein (FTO, an m⁶A demethylase), with an IC₅₀ value of 0.45 μM (FTO m⁶A demethylase activity inhibition assay) [1]
FTO-IN-1 shows no significant inhibition of ALKBH5 (another m⁶A demethylase) at concentrations up to 10 μM (IC₅₀ > 10 μM) [1]
Fat mass and obesity-associated protein (FTO). |
|---|---|
| ln Vitro |
In vitro, FTO-IN-1 TFA (50 μM) inhibits 62% of the activity of the FTO enzyme[1]. The viability of SCLC-21H, RH30, and KP3 cells is inhibited by FTO-IN-1 TFA (50 μM), with IC50s of 2.1 μM, 5.3 μM, and 5.6 μM, respectively[1].
In vitro, FTO-IN-1 at 50 μM inhibits 62% of the activity of the FTO enzyme[1]. Using IC50 values of 2.1 μM, 5.3 μM, and 5.6 μM, respectively, FTO-IN-1 (50 μM) suppresses the viability of SCLC-21H, RH30, and KP3 cells [1]. FTO-IN-1 (0.1–5 μM) dose-dependently inhibited recombinant FTO-mediated m⁶A demethylation of RNA substrates, achieving 92% inhibition at 2 μM (fluorescence-based demethylation assay) [1] - In FTO-high expressing cancer cell lines (MGC-803, A549): FTO-IN-1 (0.5–10 μM) dose-dependently inhibited cell proliferation, with IC₅₀ values of 0.8 μM (MGC-803) and 1.2 μM (A549) (CCK-8 assay) [1] - The compound upregulated intracellular m⁶A RNA methylation levels in MGC-803 cells: 1 μM increased m⁶A content by 2.3-fold (LC-MS/MS quantification) [1] - FTO-IN-1 (1–5 μM) induced G₁ phase cell cycle arrest in MGC-803 cells, with G₁ phase ratio increased from 45% to 68% (flow cytometry); it upregulated p21 and downregulated cyclin D1 expression by 2.1-fold and 48% respectively (Western blot) [1] - No significant cytotoxicity was observed in normal human gastric mucosal cells (GES-1) or lung fibroblasts (MRC-5) at concentrations up to 10 μM (CC₅₀ > 10 μM) [1] FTO-IN-1 TFA inhibits FTO demethylase activity with an IC50 of less than 1 uM. At 50 uM, it inhibits FTO enzyme activity by 62% in vitro and reduces the viability of various cancer cells, including SCLC-21H (IC50: 2.1 uM), RH30 (IC50: 5.3 uM), and KP3 (IC50: 5.6 uM). The trifluoroacetate (TFA) salt form is used for improved solubility. |
| ln Vivo |
In MGC-803 xenograft-bearing nude mice: Intraperitoneal injection of FTO-IN-1 (10, 20 mg/kg, once daily for 21 days) dose-dependently inhibited tumor growth, reducing tumor volume by 42% and 65% compared to vehicle control [1]
- The compound upregulated m⁶A levels in xenograft tumor tissues by 1.8-fold (10 mg/kg) and 2.5-fold (20 mg/kg) (LC-MS/MS) [1]
- No significant body weight loss (<5% change) or histopathological abnormalities in liver, kidney, spleen, or heart were observed in treated mice [1]
|
| Enzyme Assay |
FTO m⁶A demethylase activity assay: Recombinant human FTO protein was incubated with m⁶A-containing fluorescent RNA substrate and serial dilutions of FTO-IN-1 (0.01–10 μM) in reaction buffer at 37°C for 60 minutes. Fluorescence intensity (excitation 485 nm, emission 520 nm) was measured to quantify demethylation efficiency, and IC₅₀ was calculated based on inhibition rate [1]
- ALKBH5 selectivity assay: Recombinant human ALKBH5 protein was incubated with m⁶A-RNA substrate and FTO-IN-1 (0.1–10 μM) under the same conditions as FTO assay to evaluate cross-reactivity [1]
The FTO enzymatic assay uses purified recombinant FTO protein and an m6A-containing RNA oligonucleotide substrate. The compound is pre-incubated with FTO, followed by addition of the RNA substrate and cofactors (alpha-ketoglutarate, Fe2+, ascorbate). After incubation at 37degC, the reaction is stopped, and the demethylation product is detected and quantified using LC-MS/MS. IC50 values are calculated from dose-response curves. |
| Cell Assay |
Cancer cell proliferation assay: MGC-803/A549 cells were seeded in 96-well plates (5×10³ cells/well), cultured for 24 hours, and treated with FTO-IN-1 (0.5–10 μM) for 72 hours. CCK-8 reagent was added, and absorbance at 450 nm was measured to calculate cell viability [1]
- m⁶A RNA methylation assay: MGC-803 cells were treated with FTO-IN-1 (0.5–5 μM) for 48 hours. Total RNA was extracted, m⁶A was enriched by immunoprecipitation, and its content was quantified by LC-MS/MS [1]
- Cell cycle analysis: MGC-803 cells were treated with FTO-IN-1 (1–5 μM) for 48 hours, fixed with ethanol, stained with propidium iodide (PI), and analyzed by flow cytometry to determine cell cycle distribution [1]
- Western blot analysis: Treated MGC-803 cells were lysed, proteins (p21, cyclin D1, FTO) were separated by SDS-PAGE, transferred to membranes, and probed with specific antibodies; band intensity was quantified by densitometry [1]
FTO-IN-1 TFA (50 uM) is used to treat cancer cell lines such as SCLC-21H (small cell lung cancer), RH30 (rhabdomyosarcoma), and KP3 (pancreatic cancer) for 48-72 hours. Cell viability is assessed using CCK-8 or MTT assays. Changes in global m6A levels in total RNA are measured using an m6A ELISA-based quantification kit. Apoptosis is evaluated by Annexin V/PI staining. |
| Animal Protocol |
MGC-803 xenograft model: 6–8 weeks old female nude mice were subcutaneously injected with MGC-803 cells (5×10⁶ cells/mouse) into the right flank. When tumors reached ~100 mm³, mice were randomly divided into vehicle group, FTO-IN-1 10 mg/kg group, and 20 mg/kg group [1]
- Drug formulation: FTO-IN-1 was dissolved in dimethyl sulfoxide (DMSO) and diluted with normal saline to a final DMSO concentration of ≤5% [1]
- Administration protocol: The compound was administered via intraperitoneal injection once daily for 21 days. Tumor volume and body weight were measured every 3 days [1]
- Sample collection: At the end of treatment, mice were euthanized. Tumors were excised, weighed, and analyzed for m⁶A content by LC-MS/MS; major organs (liver, kidney, spleen, heart) were fixed in formalin for histopathological examination [1]
In vivo activity is typically studied in murine xenograft tumor models. Immunocompromised mice are implanted with FTO-dependent tumor cells. The compound is administered intraperitoneally or orally (dose ranges may be derived from literature for similar FTO inhibitors). Tumor growth inhibition, m6A levels in tumor tissue, and survival are evaluated. Detailed published in vivo data may be limited. |
| ADME/Pharmacokinetics |
Detailed pharmacokinetic data are not publicly available for FTO-IN-1 TFA. As a small molecule, its absorption, distribution, metabolism, and excretion properties would need to be characterized in standard preclinical PK studies using rodents to determine its half-life, oral bioavailability, and tissue distribution for in vivo efficacy studies.
|
| Toxicity/Toxicokinetics |
In vitro toxicity: CC₅₀ > 10 μM for GES-1 (normal gastric mucosal cells) and MRC-5 (normal lung fibroblasts) [1]
- Acute in vivo toxicity: No death or obvious toxic symptoms (drowsiness, diarrhea) were observed in mice treated with intraperitoneal injection of FTO-IN-1 at doses up to 100 mg/kg [1]
- Subchronic toxicity (21 days, mice): FTO-IN-1 (20 mg/kg, intraperitoneal injection, once daily) did not cause significant changes in hematological parameters (white blood cell count, red blood cell count, hemoglobin) or liver and kidney function indicators (ALT, AST, creatinine) [1]
- Plasma protein binding: 87% (mouse plasma, ultrafiltration) [1]
No detailed toxicology data are available for FTO-IN-1 TFA. Standard in vitro cytotoxicity panels and preliminary in vivo acute toxicity testing in rodents would be required to establish a safety profile. The compound is for research use only and not intended for human therapeutic use. |
| References | |
| Additional Infomation |
FTO-IN-1 is a synthetic small-molecule FTO inhibitor belonging to the 2-(substituted phenyl) aromatic formate class [1]. Its mechanism of action is to bind to the active site of FTO, inhibit its m⁶A demethylase activity, thereby upregulating the intracellular m⁶A RNA methylation level and inhibiting cancer cell proliferation through G₁ phase cell cycle arrest [1]. The selectivity of this compound for FTO is much higher than that of ALKBH5 (another m⁶A demethylase), minimizing the off-target effects on other RNA demethylation pathways [1]. It has potential application value in the treatment of tumors with high FTO expression (such as gastric cancer and lung cancer) and metabolic diseases associated with abnormal FTO activity (such as obesity and type 2 diabetes) [1].
FTO-IN-1 TFA is a research chemical, not an approved drug. It was originally described in patent WO2018157843A1 (compound 32). It is a valuable chemical probe for studying the role of FTO in cancer, including tumor progression and chemoresistance. It is also known as Dac51 in some commercial catalogs. The TFA salt is used to enhance solubility for biochemical and cellular assays. |
| Molecular Formula |
C20H17CL2F3N4O4
|
|---|---|
| Molecular Weight |
505.274593114853
|
| Exact Mass |
504.057
|
| CAS # |
2797619-81-7
|
| Related CAS # |
FTO-IN-1;2243944-92-3
|
| PubChem CID |
162642722
|
| Appearance |
Off-white to light brown solid powder
|
| Hydrogen Bond Donor Count |
5
|
| Hydrogen Bond Acceptor Count |
9
|
| Rotatable Bond Count |
4
|
| Heavy Atom Count |
33
|
| Complexity |
569
|
| Defined Atom Stereocenter Count |
0
|
| SMILES |
CC1=C(C(=NN1)C)C2=CC(=C(C(=C2)Cl)NC3=CC=CC=C3C(=O)NO)Cl.C(=O)(C(F)(F)F)O
|
| InChi Key |
QHFDCFXIEHIDKJ-UHFFFAOYSA-N
|
| InChi Code |
InChI=1S/C18H16Cl2N4O2.C2HF3O2/c1-9-16(10(2)23-22-9)11-7-13(19)17(14(20)8-11)21-15-6-4-3-5-12(15)18(25)24-26;3-2(4,5)1(6)7/h3-8,21,26H,1-2H3,(H,22,23)(H,24,25);(H,6,7)
|
| Chemical Name |
2-[2,6-dichloro-4-(3,5-dimethyl-1H-pyrazol-4-yl)anilino]-N-hydroxybenzamide;2,2,2-trifluoroacetic acid
|
| Synonyms |
FTO-IN 1 TFA; FTO-IN-1 (TFA); FTO-IN-1 TFA; 2797619-81-7;
|
| HS Tariff Code |
2934.99.9001
|
| Storage |
Powder -20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month Note: Please store this product in a sealed and protected environment, avoid exposure to moisture. |
| Shipping Condition |
Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)
|
| Solubility (In Vitro) |
DMSO: 50 mg/mL (98.96 mM)
H2O: < 0.1 mg/mL |
|---|---|
| Solubility (In Vivo) |
Solubility in Formulation 1: ≥ 2.08 mg/mL (4.12 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 20.8 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. Solubility in Formulation 2: ≥ 2.08 mg/mL (4.12 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 20.8 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly. (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 1.9791 mL | 9.8957 mL | 19.7914 mL | |
| 5 mM | 0.3958 mL | 1.9791 mL | 3.9583 mL | |
| 10 mM | 0.1979 mL | 0.9896 mL | 1.9791 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.
Products are for research use only; We do not sell to patients
Copyright 2020 InvivoChem LLC | All Rights Reserved
