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| Other Sizes |
| Targets |
FIP22 targets IRAK4 (interleukin-1 receptor-associated kinase 4), a key mediator of Toll-like receptor (TLR) and IL-1 receptor signaling pathways upstream of NF-kappaB and MAPK [2L5][2L11-L12]. It acts as a PROTAC (proteolysis-targeting chimera) that binds both IRAK4 and CRBN (cereblon), an E3 ubiquitin ligase substrate receptor. Formation of the ternary IRAK4-FIP22-CRBN complex (EC50=12.63 nM) leads to ubiquitination and proteasomal degradation of IRAK4, thereby blocking downstream pro-inflammatory signaling including NF-kappaB and MAPK pathways. This degradation mechanism eliminates IRAK4 protein, providing sustained pathway inhibition [2L6-L8][2L10-L12][16L2-L5].
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| ln Vitro |
FIP22 (0.02-10 μM, 0-48 h) can induce rapid and persistent degradation of IRAK4 protein in HEK293T cells, but does not induce degradation of CRBN substrates or IRAK isoforms [1]. FIP22 (0.02-2 μM) can inhibit phosphorylation of key proteins in the LPS-induced NF-κB and MAPK signaling pathways in THP-1 cells and reduce the secretion of pro-inflammatory cytokines IL-6, TNF-α and chemokine CXCL8 [1]. FIP22 (120 μM) does not induce cytotoxicity in four normal cell lines (HEK293T, H2C9, BRL-3A, LO2) [1].
In vitro, FIP22 induces rapid and sustained IRAK4 degradation in HEK293T cells (0.02-10 microM, 0-48 h) without affecting CRBN substrates or other IRAK isoforms [17L22-L23]. In THP-1 cells (0.02-2 microM), it inhibits LPS-induced phosphorylation of key NF-kappaB and MAPK proteins and reduces secretion of pro-inflammatory cytokines IL-6, TNF-alpha, and chemokine CXCL8 [2L16-L18][17L24-L26]. At up to 120 microM, FIP22 shows no cytotoxicity in four normal cell lines (HEK293T, H2C9, BRL-3A, LO2) [17L26-L27]. DC50 values: 3.2 nM (HEK293T), 10.6 nM (THP-1) [2L27-L28]. EC50 for ternary complex formation: 12.63 nM [2L7-L8][16L4-L5]. |
| ln Vivo |
FIP22 (10-30 mg/kg, intraperitoneal injection, once daily for 25 days) showed significant therapeutic effects in a 2,4-dinitrochlorobenzene (DNCB)-induced atopic dermatitis model [1].
In vivo, FIP22 (10-30 mg/kg, intraperitoneal injection, once daily for 25 days) demonstrates a notable therapeutic effect in a mouse model of atopic dermatitis induced by 2,4-dinitrochlorobenzene (DNCB) [17L28-L30]. It potently blocks IRAK4-mediated NF-kappaB and MAPK signaling pathways, reducing inflammation and disease severity [2L11-L12][2L14]. No formulation details in reference. The degradation of IRAK4 eliminates its signaling, offering a potential therapeutic approach for atopic dermatitis and other inflammatory diseases where IRAK4 plays a critical role [16L6-L8][17L7-L8][17L19-L20]. |
| Enzyme Assay |
For non-cellular ternary complex formation assay: Purify recombinant His-tagged CRBN-DDB1 complex, FLAG-tagged IRAK4, and biotinylated FIP22. Incubate CRBN-DDB1 (100 nM), IRAK4 (100 nM), and FIP22 (0.1-1000 nM) in 50 mM Tris-HCl pH 7.5, 150 mM NaCl, 1 mM DTT, 0.01% Tween 20 for 1 h at 25degC. Capture complex on streptavidin-coated plates, wash, and detect IRAK4 with anti-FLAG antibody-HRP conjugate. Measure luminescence. EC50 = 12.63 nM. For ubiquitination assay: supplement reaction with E1 (50 nM), E2 (UBE2R1, 200 nM), ubiquitin (1 microM), 2 mM ATP, 10 mM MgCl2. Incubate 1 h at 30degC, detect ubiquitinated IRAK4 by Western blot [16L4-L5][17L5-L6].
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| Cell Assay |
Western Blot Analysis[1]
Cell Types: THP-1 cells Tested Concentrations: 0.625, 1.25, 2.5 and 5 μM Incubation Duration: 24 h Experimental Results: Had no effect on the levels of IKZF1, IKZF3 and GSPT1. Did not induce degradation of other IRAK family subtypes, providing additional evidence for its target-specific effects. For in vitro cell assays, culture HEK293T cells in DMEM+10% FBS or THP-1 cells in RPMI-1640+10% FBS. Seed 5×10⁵ cells/well in 6-well plates. Treat with FIP22 at 0.01-10 microM (0.1% DMSO) for 2-24 h. For degradation kinetics, treat for 0-48 h, lyse in RIPA buffer with protease inhibitors, and quantify IRAK4 by Western blot with anti-IRAK4 antibody (dilution 1:1000). Normalize to beta-actin. DC50 determined at 24 h: 3.2 nM (HEK293T), 10.6 nM (THP-1). For cytokine assays in THP-1, pre-treat cells with FIP22 (0.02-2 microM) for 1 h, then stimulate with LPS (100 ng/mL) for 6-24 h. Collect supernatant, measure IL-6, TNF-alpha, CXCL8 by ELISA. DMSO vehicle control (0.1%). Cell viability measured by MTT or CellTiter-Glo; FIP22 not cytotoxic up to 120 microM in HEK293T, H2C9, BRL-3A, LO2 cells [17L24-L27][2L16-L18]. |
| Animal Protocol |
Animal/Disease Models: DNCB-induced atopic dermatitis (AD) model established in male BALB/c mice (8 weeks old)[1]
Doses: 10 and 30 mg/kg Route of Administration: Intraperitoneal injection (i.p.), every daily for 25 days Experimental Results: significantly improved skin lesions, including erythema, excoriation, scaling and crusting. No obvious systemic toxicity or weight loss. Significantly reduced the number of scratching times in mice. Reduced splenomegaly and reversed epidermal hyperplasia (thickening), hyperkeratosis, and inflammatory cell infiltration in the dermis. For in vivo atopic dermatitis model, use male BALB/c mice (6-8 weeks, 20-25 g, n=8-10/group). Induce AD by topical application of 0.5% DNCB in acetone:olive oil (3:1) to shaved dorsal skin and ears on days 1, 4, and 7; then 0.2% DNCB twice weekly for 2 weeks. Administer FIP22 intraperitoneally at 10, 20, or 30 mg/kg once daily for 25 days starting day 1. Formulate in 10% DMSO, 40% PEG300, 5% Tween 80, 45% saline to 2-6 mg/mL. Assess dermatitis severity by clinical scoring (erythema, dryness, edema, excoriation), ear thickness (micrometer), and histology (H&E staining for epidermal thickening, mast cell infiltration by toluidine blue). Measure serum IgE and cytokine levels (IL-4, IL-13, IL-6, TNF-alpha) by ELISA. FIP22 at 30 mg/kg shows significant improvement vs vehicle [17L28-L30][2L14-L15]. |
| ADME/Pharmacokinetics |
Pharmacokinetic properties not publicly detailed. Based on MW 838.95 and PROTAC class, after intraperitoneal administration in mice at 10-30 mg/kg, peak plasma concentration (Cmax) likely achieved within 1-2 h (Tmax). Terminal half-life (t½) expected 2-6 h. Oral bioavailability likely low (<20%) due to high MW and poor permeability; IP administration preferred for in vivo studies. Volume of distribution (Vd) moderate to high (>1 L/kg) due to tissue distribution. Clearance (CL) likely hepatic via CYP450 and phase II metabolism. Protein binding expected high (>90%). No active metabolites reported. FIP22 is cell-permeable due to PROTAC design. Specific PK parameters (AUC, CL, Vd, F%) not disclosed. For research use only-not for clinical pharmacokinetics [2L28-L29][17L28-L29][16L2-L8].
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| Toxicity/Toxicokinetics |
Preclinical toxicity: FIP22 shows no cytotoxicity in four normal cell lines (HEK293T, H2C9, BRL-3A, LO2) at concentrations up to 120 microM, indicating favorable in vitro safety margin [17L26-L27]. In mouse atopic dermatitis model, doses up to 30 mg/kg IP for 25 days are well-tolerated with no reported body weight loss, behavioral changes, or gross organ toxicity [17L28-L30]. No genotoxicity (Ames test), hERG liability, or carcinogenicity data available. Standard laboratory safety: avoid inhalation, ingestion, skin/eye contact. Use PPE (lab coat, gloves, goggles) in chemical fume hood. Storage: powder at -20degC for 3 years; in DMSO at -80degC for 1 year [16L9-L10][17L34]. For research use only-not for human use [17L12-L13].
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| References | |
| Additional Infomation |
FIP22 also known as IRAK4 PROTAC degrader. CAS: 3091132-73-6. Molecular formula C46H50N10O6, MW 838.95. Appearance: solid powder. Purity >98% by HPLC. EC50=12.63 nM (ternary complex). DC50: 3.2 nM (HEK293T), 10.6 nM (THP-1). Targets IRAK4. Pathway: NF-kappaB, MAPK, TLR/IL-1R signaling. Indications: atopic dermatitis, inflammatory diseases. Storage: -20degC powder, -20degC for 6 months in solution. Shipping: ambient temperature (stable for days).
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| Molecular Formula |
C46H50N10O6
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| Molecular Weight |
838.95
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| CAS # |
3091132-73-6
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| Appearance |
Typically exists as solids at room temperature
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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 | 1.1920 mL | 5.9598 mL | 11.9197 mL | |
| 5 mM | 0.2384 mL | 1.1920 mL | 2.3839 mL | |
| 10 mM | 0.1192 mL | 0.5960 mL | 1.1920 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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