| Size | Price | Stock | Qty |
|---|---|---|---|
| 100mg |
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| 500mg |
| Targets |
The primary target is cereblon (CRBN), a substrate receptor of the CRL4 E3 ubiquitin ligase complex. The thalidomide moiety binds to CRBN, inducing a conformational change that promotes ubiquitination of neo-substrates. The C6 alkyl chain (six methylene groups) provides a flexible hydrophobic spacer between the CRBN ligand and the terminal amine, which is used to conjugate to target protein ligands. The resulting PROTAC recruits CRBN to the target protein, leading to ubiquitination and proteasomal degradation. This conjugate alone does not induce degradation; it is a synthetic intermediate.
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| ln Vitro |
One ligand is for an E3 ubiquitin ligase, and the other is for the target protein; these two ligands are joined by a linker to form PROTACs. PROTACs target and selectively degrade target proteins by taking advantage of the intracellular ubiquitin-proteasome system.
In vitro, Thalidomide-NH-C6-NH2 TFA is used as a building block for PROTAC synthesis. The thalidomide-CRBN binding affinity is retained (KD typically in the low micromolar to nanomolar range). The C6 alkyl linker offers longer reach compared to C4 or shorter linkers, which may be optimal for certain protein-protein interactions in the ternary complex. The NH linker (secondary amine) is stable; the terminal primary amine is available for conjugation to carboxylate-containing warheads via amide coupling. The compound is soluble in DMSO (≥50 mM). Due to the TFA salt, it may have improved solubility in polar organic solvents. It is not directly active in cells without conjugation to a target ligand. |
| ln Vivo |
In vivo, active PROTACs synthesized from Thalidomide-NH-C6-NH2 TFA have been studied in animal models. The C6 alkyl linker provides a balance between flexibility and hydrophobicity, which can influence cell permeability, metabolic stability, and pharmacokinetics of the final PROTAC. PROTACs with this linker have shown efficacy in xenograft models by degrading targets such as BET proteins, BRD4, and others. This linker-ligand conjugate is not administered alone; it must be conjugated to a target protein ligand to form a functional degrader.
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| Enzyme Assay |
For CRBN binding evaluation, use fluorescence polarization (FP) or TR-FRET. Label the terminal NH2 with a fluorescent tag (e.g., FITC, TAMRA) to create a tracer. Incubate recombinant CRBN-DDB1 complex (20-50 nM) with fluorescent tracer (5-10 nM) and increasing concentrations of unlabeled Thalidomide-NH-C6-NH2 TFA (0.1 nM to 10 uM) in assay buffer (50 mM Tris pH 8.0, 150 mM NaCl, 1 mM DTT, 0.01% Triton X-100) for 60 min at room temperature. Measure FP or TR-FRET. Calculate IC₅0 and Ki. For direct binding, use surface plasmon resonance (SPR) with immobilized CRBN-DDB1; flow compound at concentrations from 1 nM to 10 uM. For orthogonal validation, use cellular thermal shift assay (CETSA) in cells expressing CRBN.
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| Cell Assay |
For cellular degradation studies, first synthesize a PROTAC by conjugating the terminal NH2 of this linker-ligand to a target protein ligand bearing a carboxylate group using standard amide coupling (EDC/NHS, HATU, or TBTU in DMF or DMSO). Purify by preparative HPLC. Culture target-expressing cells (e.g., HEK-293, HeLa, MV-4-11, or other cancer cell lines) in RPMI or DMEM with 10% FBS at 37degC with 5% CO2. Treat cells with PROTAC (1 nM to 10 uM) for 4-24 h. Harvest lysates, perform Western blot for target protein (e.g., BRD4, BET, BTK, etc.). Quantify degradation efficiency (DC₅0, Dmax). Confirm CRBN dependence by co-treatment with lenalidomide (1-10 uM) or by using CRBN-knockout cells. Assess ubiquitination: immunoprecipitate target protein, blot for ubiquitin. Evaluate apoptosis via caspase-3/7 assay, and cell viability via CellTiter-Glo after 48-72 h treatment.
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| Animal Protocol |
In vivo studies with PROTACs derived from this linker: Use female BALB/c nude or NSG mice (6-8 weeks, 18-25 g) bearing subcutaneous xenografts of cancer cell lines expressing the target protein. Administer PROTAC (10-100 mg/kg) via intraperitoneal (i.p.) or intravenous (i.v.) injection, daily or every other day. Formulate in 10% DMSO + 40% PEG300 + 5% Tween-80 + 45% saline or in 20% SBE-beta-CD in saline. Monitor tumor volume by calipers and body weight every 2-3 days. At endpoint (2-4 weeks), collect tumors and major organs. Measure target protein levels in tumor lysates by Western blot or IHC. Assess degradation pharmacokinetics by collecting tumors at 2, 6, 12, 24 h post-dose. For plasma PK, collect blood at multiple time points (0.25, 0.5, 1, 2, 4, 6, 8, 12, 24 h) and quantify PROTAC by LC-MS/MS. Include vehicle control and CRBN competitive control (e.g., pomalidomide) to confirm on-target degradation. Perform histopathology on liver, kidney, and spleen.
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| ADME/Pharmacokinetics |
Thalidomide-NH-C6-NH2 TFA has a molecular formula: free base C20H24N4O4 (MW 384.43). The TFA salt (trifluoroacetate, CF3CO2H) adds 114.02, so the salt MW is ~498.45. The compound contains a thalidomide core, an NH linker (secondary amine), and a C6 alkyl chain (hexamethylene) with terminal primary amine. Solubility: soluble in DMSO (≥50 mM); slightly soluble in water. For PROTAC synthesis, dissolve in anhydrous DMSO. Storage: powder at -20degC, desiccated, protected from light. Solutions in DMSO can be stored at -80degC for 6 months. The TFA salt is hygroscopic; keep container tightly sealed. The compound is stable for at least 2 years as powder under inert atmosphere.
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| Toxicity/Toxicokinetics |
Thalidomide-based compounds are potent teratogens. Handle with extreme caution: use double gloves, lab coat, face shield, and work in a certified fume hood. Avoid inhalation, ingestion, skin contact. Do not handle if pregnant or potentially pregnant. For research use only; not for human therapeutic use. The TFA salt is corrosive in large quantities but at milligram scale it is manageable. Consult the safety data sheet before handling. Dispose of waste as hazardous pharmaceutical waste. The compound is not acutely toxic at typical research scales but reproductive toxicity is a serious concern based on the thalidomide core.
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| References | |
| Additional Infomation |
Thalidomide-NH-C6-NH2 TFA is a thalidomide-based CRBN ligand-linker conjugate used for PROTAC synthesis. The C6 alkyl linker (hexamethylene) provides a longer hydrophobic spacer compared to C4 or PEG linkers. This length may be optimal for certain target proteins where a longer reach between the CRBN and target binding sites is required for ternary complex formation. The NH linker (instead of O-amido) provides a different vector geometry. This compound is a valuable building block for constructing PROTAC libraries to degrade a variety of targets including BRD4, BET, BTK, CDK, and others. It is for research use only and is not approved for clinical use. Thalidomide is known for teratogenicity; the conjugate is a chemical tool for targeted protein degradation.
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| Molecular Formula |
C21H25F3N4O6
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|---|---|
| Molecular Weight |
486.441615819931
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| Exact Mass |
486.172
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| CAS # |
2093386-51-5
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| PubChem CID |
146047127
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| Appearance |
Light yellow to yellow solid powder
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| Hydrogen Bond Donor Count |
4
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| Hydrogen Bond Acceptor Count |
11
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| Rotatable Bond Count |
8
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| Heavy Atom Count |
34
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| Complexity |
692
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| Defined Atom Stereocenter Count |
0
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| SMILES |
FC(C(=O)O)(F)F.O=C1C2C(=CC=CC=2C(N1C1C(NC(CC1)=O)=O)=O)NCCCCCCN
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| InChi Key |
VUTCFPKPHCMKEA-UHFFFAOYSA-N
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| InChi Code |
InChI=1S/C19H24N4O4.C2HF3O2/c20-10-3-1-2-4-11-21-13-7-5-6-12-16(13)19(27)23(18(12)26)14-8-9-15(24)22-17(14)25;3-2(4,5)1(6)7/h5-7,14,21H,1-4,8-11,20H2,(H,22,24,25);(H,6,7)
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| Chemical Name |
4-(6-aminohexylamino)-2-(2,6-dioxopiperidin-3-yl)isoindole-1,3-dione;2,2,2-trifluoroacetic acid
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| Synonyms |
ThalidomideNHC6NH2 TFA; Thalidomide NH C6 NH2 TFA
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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 Note: Please store this product in a sealed and protected environment (e.g. under nitrogen), 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)
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| Solubility (In Vitro) |
DMSO : ~100 mg/mL (~205.58 mM)
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| Solubility (In Vivo) |
Solubility in Formulation 1: ≥ 2.5 mg/mL (5.14 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. Solubility in Formulation 2: 2.5 mg/mL (5.14 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 ultrasonication. 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 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. (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 2.0558 mL | 10.2788 mL | 20.5575 mL | |
| 5 mM | 0.4112 mL | 2.0558 mL | 4.1115 mL | |
| 10 mM | 0.2056 mL | 1.0279 mL | 2.0558 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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