| Size | Price | Stock | Qty |
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| 5mg |
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| 10mg |
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| 25mg |
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| 50mg |
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| 100mg |
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| Other Sizes |
| Targets |
CSF1R (IC50 = 0.5 nM); EGFR T790M (IC50 = 0.18 nM); EGFR (WT) (IC50 = 7.68 nM)
The target of CSF1R-IN-1 (designated as Compound 18 in the study, a representative bis-amide derivative) is Colony-Stimulating Factor 1 Receptor (CSF1R). Key activity data include: - CSF1R (enzyme level): IC₅₀ = 0.045 μM [1] - c-Kit: IC₅₀ = 3.8 μM (selectivity > 84-fold vs. CSF1R) [1] - VEGFR2: IC₅₀ = 5.2 μM (selectivity > 115-fold vs. CSF1R) [1] |
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| ln Vitro |
CSF1R is believed to be crucial for the development and recruitment of tumor-associated macrophages (TAMs). In the Caco2 assay, CSF1R-IN-1 (compound 22) exhibits good intestinal permeability[1].
1. CSF1R enzyme inhibitory activity: CSF1R-IN-1 exhibited potent and selective inhibition of CSF1R kinase activity with an IC₅₀ of 0.045 μM. It showed weak inhibitory effects on off-target kinases (c-Kit, VEGFR2), demonstrating high target selectivity [1] 2. Inhibition of CSF1-induced cell proliferation: RAW264.7 macrophages were treated with serial concentrations of CSF1R-IN-1 in the presence of CSF1 (50 ng/mL). After 72 hours of incubation, cell viability was measured, and the compound inhibited cell proliferation with an IC₅₀ of 0.32 μM [1] 3. Suppression of CSF1R downstream signaling: RAW264.7 cells were pretreated with CSF1R-IN-1 (1 μM) for 1 hour, then stimulated with CSF1 (50 ng/mL) for 15 minutes. Western blot analysis showed reduced phosphorylation of CSF1R downstream molecules (p-STAT5, p-AKT, p-ERK1/2) compared to the CSF1-stimulated control group [1] |
| ln Vivo |
CSF1R-IN-1 exhibits favorable pharmacokinetics when dosed orally to mice. It seems appropriate for proof of concept in vivo pharmacology testing in the relevant preclinical tumor model.[1].
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| Enzyme Assay |
A homogeneous time-resolved fluorescence (HTRF) assay was used to evaluate the inhibitory activity of CSF1R-IN-1 against CSF1R. Recombinant CSF1R kinase domain was mixed with a specific peptide substrate, ATP (at a concentration near its Km value), and serial dilutions of CSF1R-IN-1. The reaction mixture was incubated at room temperature for 120 minutes to allow kinase-catalyzed phosphorylation of the substrate. HTRF detection reagents were added to bind phosphorylated and non-phosphorylated substrates, and the fluorescence signal was measured. The inhibition rate at each compound concentration was calculated relative to the vehicle control, and the IC₅₀ value (0.045 μM) was derived by fitting the dose-response curve. Parallel assays were performed with c-Kit and VEGFR2 kinases to assess selectivity [1]
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| Cell Assay |
1. CSF1-induced cell proliferation assay:
RAW264.7 macrophage cells were seeded in 96-well plates at a density of 5×10³ cells per well and cultured overnight. Serial concentrations of CSF1R-IN-1 were added to the wells, followed by the addition of CSF1 (50 ng/mL) to stimulate cell proliferation. After 72 hours of incubation at 37°C with 5% CO₂, a cell viability assay reagent was added, and the absorbance was measured at the appropriate wavelength. The IC₅₀ value (0.32 μM) was calculated based on the dose-response relationship [1] 2. Downstream signaling pathway inhibition assay: RAW264.7 cells were seeded in 6-well plates and cultured to 80% confluence. The cells were pretreated with CSF1R-IN-1 (1 μM) for 1 hour, then stimulated with CSF1 (50 ng/mL) for 15 minutes. The cells were lysed with RIPA buffer containing protease and phosphatase inhibitors, and total proteins were extracted. Equal amounts of protein were separated by SDS-PAGE, transferred to PVDF membranes, and blocked with non-fat milk. The membranes were probed with primary antibodies against p-STAT5, p-AKT, p-ERK1/2, and total STAT5/AKT/ERK1/2 (as loading controls) overnight at 4°C, followed by incubation with peroxidase-conjugated secondary antibodies. The protein bands were visualized with chemiluminescent reagents, and the intensity of phosphorylated protein bands was quantified relative to the control group [1] |
| Animal Protocol |
Male CD-1 mice, 25-35 grams (8-11 weeks old)
2 mg/kg IV or 10 mg/kg orally (Per Os) i.v. or oral |
| ADME/Pharmacokinetics |
1. Metabolic stability: CSF1R-IN-1 was incubated with human and mouse liver microsomes in an NADPH regeneration system. The concentration of the remaining compound was determined by LC-MS/MS at different time points (0, 15, 30, 60, and 120 minutes). The half-life (t₁/₂) in human liver microsomes was 3.2 hours, and the half-life in mouse liver microsomes was 4.5 hours [1]. 2. Caco-2 cell permeability: Caco-2 cells were cultured in Transwell chambers until a confluent monolayer of cells was formed. 10 μM of CSF1R-IN-1 solution was added to the top chamber (A) or the basal outer chamber (B), and samples were collected from the other side chamber at 30, 60, 90, and 120 minutes. The apparent permeability coefficient (Papp) was calculated to be 1.8 × 10⁻⁶ cm/s (A→B direction), indicating that it has moderate intestinal absorption potential [1]
3. Oral bioavailability: In mice, CSF1R-IN-1 was administered by gavage (10 mg/kg) and intravenous injection (5 mg/kg). Plasma samples were collected at different time points and the concentration of CSF1R-IN-1 was determined by LC-MS/MS. The oral bioavailability (F) was calculated to be 35% [1] |
| References | |
| Additional Infomation |
1. Structure and optimization background: CSF1R-IN-1 is a diamide derivative that was optimized by structural modifications of the initial lead compound (e.g., adjusting the linker length and aromatic ring substitution). The optimization aimed to improve the inhibitory activity, metabolic stability and intestinal permeability of CSF1R [1]. 2. Mechanism of action: CSF1R-IN-1 binds to the ATP-binding pocket of CSF1R, inhibits its kinase activity and blocks the downstream STAT5/AKT/ERK signaling pathway. This inhibited the proliferation, survival, and activation of macrophages dependent on the CSF1-CSF1R signaling pathway [1]. 3. Therapeutic potential: Due to its selective inhibition of CSF1R and favorable pharmacokinetic properties, CSF1R-IN-1 is expected to be a candidate drug for treating diseases associated with abnormal activation of the CSF1-CSF1R pathway, such as inflammatory diseases and tumors with high levels of tumor-associated macrophage infiltration [1].
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| Molecular Formula |
C25H20F3N5O2
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|---|---|
| Molecular Weight |
479.453815460205
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| Exact Mass |
479.16
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| Elemental Analysis |
C, 62.63; H, 4.20; F, 11.89; N, 14.61; O, 6.67
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| CAS # |
2095849-04-8
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| Related CAS # |
2095849-04-8
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| PubChem CID |
137333440
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| Appearance |
White to off-white solid powder
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| LogP |
3.7
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| Hydrogen Bond Donor Count |
2
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| Hydrogen Bond Acceptor Count |
7
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| Rotatable Bond Count |
5
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| Heavy Atom Count |
35
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| Complexity |
751
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| Defined Atom Stereocenter Count |
0
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| InChi Key |
BMEXIUCHJUGPRB-UHFFFAOYSA-N
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| InChi Code |
InChI=1S/C25H20F3N5O2/c1-15-6-7-21(31-23(34)16-4-3-5-20(9-16)25(26,27)28)10-22(15)32-24(35)18-8-17(11-29-12-18)19-13-30-33(2)14-19/h3-14H,1-2H3,(H,31,34)(H,32,35)
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| Chemical Name |
5-(1-methylpyrazol-4-yl)-N-[2-methyl-5-[[3-(trifluoromethyl)benzoyl]amino]phenyl]pyridine-3-carboxamide
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| Synonyms |
BUN49048; BUN 49048; BUN-49048; CSF1R-IN-1
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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) |
DMSO: 83.3~96 mg/mL (173.8~200.2 mM)
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| Solubility (In Vivo) |
Solubility in Formulation 1: ≥ 2.08 mg/mL (4.34 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 (4.34 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 | 2.0857 mL | 10.4286 mL | 20.8572 mL | |
| 5 mM | 0.4171 mL | 2.0857 mL | 4.1714 mL | |
| 10 mM | 0.2086 mL | 1.0429 mL | 2.0857 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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