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| Targets |
As a chemical linker rather than a pharmacologically active agent, N-Boc-PEG4-bromide does not bind to biological targets such as receptors or enzymes. Its functional "targets" are the chemical groups on payload molecules, antibodies, and E3 ligase ligands to which it conjugates. In PROTAC applications, the linker connects an E3 ubiquitin ligase ligand to a target protein ligand, enabling the ubiquitin-proteasome system to selectively degrade target proteins. In ADC applications, the linker attaches cytotoxic drugs to monoclonal antibodies. The bromide terminus reacts with nucleophiles (e.g., thiols, amines) via nucleophilic substitution, while the Boc-protected amine provides a handle for deprotection and subsequent amide bond formation. The PEG4 spacer provides optimal solubility and flexibility for bioconjugation.
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| ln Vitro |
Two distinct ligands, one for the E3 ubiquitin ligase and the other for the target protein, are present in PROTAC and are joined by a linker. PROTAC targets and selectively degrades target proteins by means of the intracellular ubiquitin-proteasome system. An ADC cytotoxin is connected to an antibody by use of an ADC linker to form an ADC.
N-Boc-PEG4-bromide itself does not exhibit pharmacological activity in cell-based assays because it is a synthetic linker molecule. Its in vitro utility is demonstrated through the successful synthesis and characterization of PROTAC or ADC constructs incorporating this linker. Researchers evaluate the linker's performance by assessing conjugation efficiency (via HPLC, LC-MS, or NMR), the stability of resulting conjugates in cell culture media, and the biological activity of final constructs in target cell lines. For PROTACs, target protein degradation is measured by Western blot or ELISA; for ADCs, cytotoxicity is assessed using cell viability assays such as MTT or CellTiter-Glo. The PEG4 spacer provides enhanced solubility compared to shorter PEG variants. The compound is soluble in DMSO and other organic solvents, facilitating its use in bioconjugation workflows. |
| ln Vivo |
No intrinsic in vivo pharmacological activity is attributed to N-Boc-PEG4-bromide; its in vivo relevance is through the ADC or PROTAC constructs synthesized with this linker. For in vivo administration, conjugates containing this linker are formulated in standard vehicles such as DMSO/PEG300/Tween-80/saline or other suitable excipients. Typical preclinical studies involve administering the conjugate to rodent models (e.g., tumor xenografts) via intravenous injection. Efficacy endpoints include tumor volume reduction, survival prolongation, and target modulation in tissues. The linker's stability in circulation and its ability to release payloads at target sites are critical parameters evaluated through pharmacokinetic sampling. The PEG4 spacer contributes to favorable pharmacokinetic properties by reducing immunogenicity and improving solubility.
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| Enzyme Assay |
In vitro enzyme/receptor binding assays are not applicable to N-Boc-PEG4-bromide as it is a chemical linker devoid of biological target affinity. Quality control and characterization are performed using standard analytical methods. Purity is assessed by HPLC (≥95-98%) and structural integrity is confirmed by ¹H NMR, ¹³C NMR, and mass spectrometry. The bromide content and functional group integrity are verified through elemental analysis and spectroscopic methods. For researchers using this linker in bioconjugation, reaction progress (bromide displacement by nucleophiles) is typically monitored by TLC, HPLC, or LC-MS. Boc deprotection efficiency is confirmed by NMR or LC-MS following acid treatment (e.g., TFA in DCM). Solubility testing in various solvents is performed to guide formulation development.
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| Cell Assay |
Cell-based assays are not performed directly on N-Boc-PEG4-bromide because the compound lacks biological activity. However, the biological activity of PROTAC or ADC constructs containing this linker is evaluated in relevant cell lines. For PROTAC evaluation, cells are treated with the construct for 4-24 hours, and target protein degradation is quantified by Western blot or ELISA. Cell viability, proliferation, and apoptosis are assessed using standard assays (MTT, CCK-8, flow cytometry). For ADC evaluation, cancer cell lines are treated with the conjugate, and cytotoxicity is measured over 48-72 hours. The linker's contribution to conjugate stability, solubility, and target engagement is inferred from comparative studies. DMSO stock solutions are prepared and diluted in culture media to achieve desired final concentrations. Control groups include vehicle-treated and unconjugated antibody or payload-treated cells.
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| Animal Protocol |
In vivo animal studies are conducted with ADC or PROTAC constructs incorporating N-Boc-PEG4-bromide, not with the linker alone. Typical protocols utilize immunocompromised mice bearing tumor xenografts. The conjugate is administered via intravenous, intraperitoneal, or subcutaneous injection at doses determined by the payload's maximum tolerated dose. For in vivo formulation, the conjugate is prepared in standard vehicles such as DMSO/PEG300/Tween-80/saline or other suitable excipients. Efficacy is assessed by caliper measurement of tumor volumes every 2-3 days, body weight monitoring, and survival analysis. Pharmacodynamic endpoints include target protein degradation (PROTACs) or tumor growth inhibition (ADCs). Plasma samples are collected for pharmacokinetic analysis of conjugate stability and payload release. The linker's stability in biological matrices is a key parameter evaluated in these studies.
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| ADME/Pharmacokinetics |
As a chemical linker, N-Boc-PEG4-bromide does not have a conventional pharmacokinetic profile. However, the pharmacokinetic properties of bioconjugates incorporating this linker are evaluated in preclinical studies. Following intravenous administration in rodents, key parameters such as half-life, clearance, volume of distribution, and AUC are determined from plasma concentration-time data. The PEG4 spacer contributes to enhanced aqueous solubility, reduced protein binding, and prolonged circulation time of the conjugate. Linker stability in plasma is assessed by measuring free payload release over time using LC-MS/MS. The Boc-protected amine is stable at physiological pH (7.4) and is only deprotected under acidic conditions. Formulation studies indicate good solubility in standard in vivo vehicles. The compound itself is not administered systemically; therefore, its absorption, distribution, metabolism, and excretion are not independently characterized.
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| Toxicity/Toxicokinetics |
Toxicological data for N-Boc-PEG4-bromide are limited because it is a research-grade reagent not intended for human use. As a bromoalkyl compound, it may cause skin and eye irritation upon contact. The reactive bromide group can act as an alkylating agent, necessitating standard laboratory safety precautions: use of personal protective equipment (gloves, safety goggles, lab coat) and handling in a well-ventilated fume hood. The compound should be stored at -20°C for long-term stability (up to 3 years) and can be shipped at ambient temperature. No genotoxicity, carcinogenicity, or reproductive toxicity data are available. Researchers should consult the safety data sheet (SDS) before handling. The compound is not classified as a hazardous drug but should be treated with care due to the presence of the alkyl bromide moiety.
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| References | |
| Additional Infomation |
Additional information for N-Boc-PEG4-bromide: The compound has a CAS number of 1392499-32-9. Its molecular formula is C₁₅H₃₀BrNO₆ and molecular weight is 400.31 g/mol. Purity is typically ≥95%. The compound appears as a liquid. The Boc-protected amine can be deprotected under mild acidic conditions to form a free amine. The bromide group enables efficient nucleophilic substitution reactions. The hydrophilic PEG spacer increases solubility in aqueous media. This product is a non-cleavable linker for bioconjugation. It is for research use only and is not approved for clinical or diagnostic applications. No clinical trials or regulatory approvals exist for this compound as it is a synthetic building block. Related compounds include shorter and longer PEG variants with similar functional groups.
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| Molecular Formula |
C15H30BRNO6
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|---|---|
| Molecular Weight |
400.305804729462
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| Exact Mass |
399.125
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| CAS # |
1392499-32-9
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| PubChem CID |
88853548
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| Appearance |
Light yellow to brown liquid
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| Density |
1.223±0.06 g/cm3
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| Boiling Point |
459.3±40.0 °C
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| LogP |
1.1
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| Hydrogen Bond Donor Count |
1
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| Hydrogen Bond Acceptor Count |
6
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| Rotatable Bond Count |
16
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| Heavy Atom Count |
23
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| Complexity |
286
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| Defined Atom Stereocenter Count |
0
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| SMILES |
BrCCOCCOCCOCCOCCNC(=O)OC(C)(C)C
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| InChi Key |
KYOVSWOWVSWAII-UHFFFAOYSA-N
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| InChi Code |
InChI=1S/C15H30BrNO6/c1-15(2,3)23-14(18)17-5-7-20-9-11-22-13-12-21-10-8-19-6-4-16/h4-13H2,1-3H3,(H,17,18)
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| Chemical Name |
tert-butyl N-[2-[2-[2-[2-(2-bromoethoxy)ethoxy]ethoxy]ethoxy]ethyl]carbamate
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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 | 2.4981 mL | 12.4903 mL | 24.9806 mL | |
| 5 mM | 0.4996 mL | 2.4981 mL | 4.9961 mL | |
| 10 mM | 0.2498 mL | 1.2490 mL | 2.4981 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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