| Size | Price | Stock | Qty |
|---|---|---|---|
| 1mg |
|
||
| Other Sizes |
| Targets |
Topoisomerase I. Glycyl-Exatecan-d5 hydrochloride is a deuterated analog of Glycyl-Exatecan, which is a derivative of exatecan (DX-8951). Exatecan is a camptothecin analog that inhibits DNA topoisomerase I. Topoisomerase I is an essential nuclear enzyme that relieves torsional stress in DNA during replication, transcription, and recombination by creating single-strand breaks. Exatecan stabilizes the topoisomerase I-DNA cleavage complex, preventing religation of DNA strands, leading to accumulation of DNA breaks, replication fork arrest, and ultimately cell death (apoptosis). The deuterium labeling (d5) does not alter the pharmacological activity of the parent compound but provides a stable isotope tag for quantification. Glycyl-Exatecan-d5 is not intended to be used as a therapeutic agent but as an analytical standard to quantify exatecan or its derivatives in biological samples.
|
|---|---|
| ln Vitro |
Not applicable (internal standard). Glycyl-Exatecan-d5 hydrochloride is not used in biological activity assays to measure enzyme inhibition or cell proliferation because its purpose is to serve as an internal standard for mass spectrometry. The parent compound, exatecan (DX-8951), is a potent topoisomerase I inhibitor with IC50 values in the nanomolar range. In vitro, exatecan inhibits topoisomerase I-mediated DNA relaxation and induces DNA damage in cancer cell lines, leading to cell cycle arrest (G2/M phase) and apoptosis. Exatecan is more potent than irinotecan (CPT-11) and has shown efficacy against various cancer cell lines. Glycyl-Exatecan, the non-deuterated form, is a water-soluble derivative that maintains topoisomerase I inhibition. However, the deuterated version is not typically used in functional assays because the isotopic substitution does not significantly alter biological activity. For research purposes, the non-labeled analog should be used for activity assays, and the labeled analog is reserved for analytical quantification. No specific EC50 or IC50 data are available for the d5-labeled compound.
|
| ln Vivo |
Not applicable. The deuterated compound is intended to be used as an internal standard in bioanalytical assays. In vivo activity (antitumor efficacy) of Glycyl-Exatecan-d5 has not been studied because the purpose is analytical. However, the parent compound exatecan has demonstrated in vivo antitumor activity in mouse xenograft models of various cancers (breast, lung, colon, ovarian, pancreatic) at doses of 5-20 mg/kg, given intravenously or intraperitoneally. Exatecan mesylate (also known as DX-8951f) has been investigated in clinical trials for the treatment of solid tumors. The deuterated version would be expected to have similar in vivo activity if administered, but this is not the intended use. The hydrochloride salt form enhances water solubility for formulation. No in vivo data are available for this specific isotopic variant.
|
| Enzyme Assay |
For non-cellular enzyme assays using Glycyl-Exatecan-d5, the compound is not used to measure enzyme activity; it is used as an internal standard in LC-MS/MS analysis of biological samples. Therefore, no receptor/enzyme binding experimental protocols are applicable. However, for the non-deuterated parent compound (exatecan), the inhibition of topoisomerase I can be measured using a DNA relaxation assay. Briefly, supercoiled plasmid DNA (pBR322) is incubated with recombinant human topoisomerase I in reaction buffer (50 mM Tris-HCl pH 7.5, 100 mM KCl, 10 mM MgCl2, 0.5 mM DTT, 30 ug/mL BSA) in the presence of varying concentrations of exatecan (0.01-10 uM) at 37degC for 30 min. The reaction is terminated by adding SDS and proteinase K, followed by phenol-chloroform extraction. The DNA products are separated by agarose gel electrophoresis, and the relaxed (and nicked) forms are visualized by ethidium bromide staining. The IC50 is the concentration that inhibits 50% of DNA relaxation. For a fluorescence-based topoisomerase I inhibition assay, a fluorogenic DNA substrate can be used. The assay is performed in a 96-well plate; the reaction is initiated by adding topoisomerase I, and fluorescence (e.g., from intercalating dyes) is measured over time. The IC50 is determined. For the deuterated compound, if used in a competition experiment to displace a labeled ligand, standard topoisomerase I binding assays (e.g., DNA cleavage assay using radiolabeled oligonucleotide) can be performed, but again, not typical.
|
| Cell Assay |
For cell-based assays, the non-deuterated parent compound exatecan (DX-8951) or Glycyl-Exatecan is used to evaluate antiproliferative activity. Cancer cells (e.g., HCT116, MCF-7, A549, PC-3, BxPC-3) are seeded in 96-well plates at 5 × 10^3 cells/well in appropriate medium (RPMI-1640 or DMEM) with 10% FBS. After 24 h, cells are treated with various concentrations (0.1-1000 nM) of exatecan or Glycyl-Exatecan for 48-72 hours. Cell viability is measured by MTT, CCK-8, or CellTiter-Glo assays. The IC50 is calculated. Apoptosis is assessed by Annexin V-FITC/PI staining and flow cytometry. Cell cycle analysis is performed by propidium iodide staining and flow cytometry. For DNA damage, gamma-H2AX staining (Ser139) is visualized by immunofluorescence or Western blot. For the deuterated version (Glycyl-Exatecan-d5), cell-based assays are not typically conducted because it is used as an analytical standard. The hydrochloride salt (HCl) of the deuterated compound is used for preparing calibration standards in biological matrices. For researchers who want to use the d5 compound for metabolic stability assays, hepatocytes or microsomes are incubated with the compound, and the disappearance of the parent or appearance of metabolites is monitored by LC-MS/MS, using the deuterated compound itself as the substrate and the non-deuterated compound as the internal standard, or vice versa. A typical metabolism assay: Human liver microsomes (0.5 mg/mL) are incubated with Glycyl-Exatecan-d5 (1-10 uM) in 100 mM phosphate buffer (pH 7.4) containing NADPH (1 mM) at 37degC for 0-60 min. Reactions are terminated by adding acetonitrile containing internal standard, and samples are analyzed by LC-MS/MS. Percent remaining is calculated. This is a pharmacokinetic/metabolism assay, not an activity assay.
|
| Animal Protocol |
Not applicable. Glycyl-Exatecan-d5 hydrochloride is used as an internal standard for bioanalysis and is not administered to animals for efficacy studies. Therefore, in vivo animal experiment protocols for this specific compound are not available. However, for the non-deuterated exatecan or Glycyl-Exatecan, standard xenograft protocols are used. For example, female BALB/c nude mice (5-6 weeks old) are injected subcutaneously with 5 × 10^6 HCT116 colon cancer cells (or other cancer cells). When tumors reach 100-150 mm3, mice are randomized (n=8-10 per group). Exatecan mesylate (DX-8951f) is formulated in saline or 5% glucose and administered intravenously (i.v.) at doses of 5-20 mg/kg on days 1, 5, 9 (q4d × 3). Control groups receive vehicle. Tumor volumes are measured every 2-3 days. Efficacy is evaluated by tumor growth inhibition (TGI%). The d5-labeled compound is not used in such studies due to its cost and labeling. For PK studies using the labeled compound as a tracer, the d5 compound can be co-administered with the non-deuterated compound to study absorption, distribution, metabolism, and excretion (ADME) in rats or mice. A typical PK study: Male Sprague-Dawley rats (n=3-4 per time point) receive an intravenous bolus (1-2 mg/kg) or oral gavage (5-10 mg/kg) of a mixture of Glycyl-Exatecan (non-labeled) and Glycyl-Exatecan-d5 (as an internal standard for LC-MS/MS calibration). Blood samples are collected at pre-dose, 5, 15, 30, 60, 120, 240, 360, 480, 720, and 1440 min. Plasma is separated, and drug concentration is quantified by LC-MS/MS, using the d5 compound as the internal standard for the non-labeled compound (or vice versa). PK parameters (AUC, Cmax, Tmax, t1/2, CL, Vd, bioavailability) are calculated using non-compartmental analysis. This is the primary in vivo use of deuterated compounds. However, the Glycyl-Exatecan-d5 itself is not the pharmacologically active agent in these studies. All animal procedures must be approved by IACUC.
|
| ADME/Pharmacokinetics |
The deuterated form (Glycyl-Exatecan-d5) is expected to have identical pharmacokinetic properties to the non-deuterated Glycyl-Exatecan, as deuterium substitution has a minimal isotope effect on PK parameters in most cases. Glycyl-Exatecan is a water-soluble derivative of exatecan. Exatecan (DX-8951) has a plasma half-life in humans of approximately 8-15 hours. In rodents, after intravenous administration, exatecan has a t1/2 of 1-3 hours, a high clearance (CL ~20-40 mL/min/kg), and a large volume of distribution (Vd > 5 L/kg), indicating extensive tissue distribution. It is not highly bound to plasma proteins (40-60%). Exatecan is primarily excreted unchanged in urine and bile. The hydrochloride salt (HCl) provides aqueous solubility. For the deuterated compound, LC-MS/MS quantification is used. For PK sample processing: Plasma samples (50 uL) are mixed with internal standard (e.g., another stable isotope-labeled compound), and proteins are precipitated with acetonitrile (3:1). After centrifugation, the supernatant is injected into LC-MS/MS system. Separation is achieved on a C18 reversed-phase column with a mobile phase of 0.1% formic acid in water and acetonitrile. Detection is in positive ion electrospray ionization (ESI) mode with multiple reaction monitoring (MRM) transitions specific for the d5 compound (e.g., m/z [M+H]+ for d5 compared to non-labeled). The linear range is typically 0.1-1000 ng/mL. No formal PK studies are published for this specific labeled compound.
|
| Toxicity/Toxicokinetics |
No specific toxicity data are available for Glycyl-Exatecan-d5 hydrochloride. The toxicity profile of the parent compound exatecan (DX-8951) is well characterized. As a topoisomerase I inhibitor, exatecan causes myelosuppression (neutropenia, thrombocytopenia), gastrointestinal toxicity (diarrhea, nausea, vomiting), and alopecia, similar to other camptothecins, but with reduced late-onset diarrhea compared to irinotecan. In preclinical studies, the maximum tolerated dose (MTD) in rodents was approximately 10-20 mg/kg (i.v.). At higher doses, severe myelosuppression and intestinal damage were observed. The deuterated compound is expected to have the same toxicity profile because the isotopic substitution does not alter pharmacological activity. However, the d5 compound is used as an analytical standard, not as a therapeutic agent; thus, it is handled only in small quantities (milligrams) in research labs, minimizing exposure risk. The hydrochloride salt is not expected to add additional toxicity. No genotoxicity, carcinogenicity, or reproductive toxicity studies have been performed on the d5 compound. Glycyl-Exatecan-d5 is for research use only and is not approved for human or veterinary use. Standard precautions for handling cytotoxic compounds (gloves, lab coat, fume hood) should be followed, as topoisomerase inhibitors are potential carcinogens and teratogens.
|
| Additional Infomation |
Exatecan (DX-8951) is a synthetic camptothecin analog that was developed for its improved water solubility and reduced toxicity compared to other camptothecins. Exatecan mesylate (DX-8951f) has been evaluated in clinical trials for solid tumors, including gastric, pancreatic, breast, lung, and ovarian cancers, but has not received FDA approval. Glycyl-Exatecan is an amino acid prodrug derivative designed to improve solubility. Deuterium labeling (d5) is a common technique in pharmaceutical analysis to generate internal standards for quantitative mass spectrometry. The stable isotope label (deuterium) does not change the chemical structure substantially but provides a mass shift of 5 Da, allowing differentiation from the unlabeled analyte. Glycyl-Exatecan-d5 is used in LC-MS/MS methods to quantify exatecan and its analogs in biological matrices (plasma, tissue homogenates, cell lysates) for pharmacokinetic and bioequivalence studies. The product is not a drug; it is a research-grade analytical standard. It is not intended for therapeutic use. The hydrochloride salt (HCl) is used to improve solubility and stability. This compound is for research use only.
|
| Molecular Formula |
C26H21D5CLFN4O5
|
|---|---|
| Molecular Weight |
533.99
|
| Appearance |
Typically exists as solid at room temperature
|
| 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 :~110 mg/mL (~206.00 mM)
|
|---|---|
| Solubility (In Vivo) |
Solubility in Formulation 1: ≥ 2.75 mg/mL (5.15 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 27.5 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: 2.75 mg/mL (5.15 mM) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution; with ultrasonication. For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 27.5 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. View More
Solubility in Formulation 3: 2.75 mg/mL (5.15 mM) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), suspension solution; with ultrasonication. |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 1.8727 mL | 9.3635 mL | 18.7269 mL | |
| 5 mM | 0.3745 mL | 1.8727 mL | 3.7454 mL | |
| 10 mM | 0.1873 mL | 0.9363 mL | 1.8727 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
