| Size | Price | Stock | Qty |
|---|---|---|---|
| 1mg |
|
||
| 5mg |
|
||
| Other Sizes |
| Targets |
WRN (Werner syndrome RecQ helicase). VVD‑214 covalently binds to cysteine 727 of WRN and inhibits both its ATP hydrolysis and helicase activities.
|
|---|---|
| ln Vitro |
In cell‑free biochemical assays, VVD‑214 binds to WRN protein and inhibits its helicase activity. In cellular assays, VVD‑214 treatment causes widespread double‑stranded DNA breaks, nuclear swelling, and cell death in MSI‑high cancer cells (e.g., HCT116, LoVo, DLD‑1 MSI cells) but not in microsatellite stable (MSS) cells or normal cells. This demonstrates its synthetic lethal mechanism of action.
|
| ln Vivo |
In vivo, VVD‑214 exhibits robust tumor regression in multiple MSI‑high colorectal cancer cell line‑derived xenografts (CDX) and patient‑derived xenograft (PDX) models, including models derived from patients progressing on immune checkpoint therapies. It is a synthetic lethal allosteric inhibitor that selectively targets MSI‑H tumors. Dose‑dependent tumor growth inhibition and regression have been observed.
|
| Enzyme Assay |
A general cell‑free protocol for assessing WRN helicase inhibition: A fluorescence‑based DNA unwinding assay is used. Recombinant WRN helicase protein (10‑50 nM) is incubated with a forked DNA substrate (50 nM) containing a fluorophore and quencher (e.g., a 5′-fluorescein-labeled strand annealed to a complementary strand with a 3′‑black hole quencher) in helicase buffer (25 mM Tris‑HCl, pH 7.5, 2 mM MgCl2, 2 mM ATP, 50 mM KCl, 1 mM DTT, 0.1 mg/mL BSA) with varying concentrations of VVD‑214 (0.1 nM to 10 uM). The reaction is initiated by adding ATP and incubated at 37degC for 30‑60 minutes. The reaction is stopped with 20 mM EDTA and 0.5% SDS. The increase in fluorescence (due to strand separation) is measured (excitation 485 nm, emission 528 nm). The IC₅0 for helicase inhibition is calculated. For binding studies, a cellular thermal shift assay (CETSA) can be used to assess target engagement.
|
| Cell Assay |
A general cellular protocol for VVD‑214: MSI‑high colorectal cancer cells (e.g., HCT116, LoVo, or RKO) and MSS cells (e.g., HT‑29 or SW480) are seeded in 96‑well plates at 5,000‑10,000 cells/well. Cells are treated with serial dilutions of VVD‑214 (0.01 nM to 10 uM) for 72‑120 hours. Cell viability is measured using CellTiter‑Glo assay. The synthetic lethal effect is calculated as the ratio of IC₅0 in MSS cells to IC₅0 in MSI cells. For DNA damage assessment, cells are treated for 24‑48 hours, fixed, and stained with an anti‑gammaH2AX (Ser139) antibody, and the number of gammaH2AX foci per nucleus is quantified by immunofluorescence microscopy. Nuclear morphology is assessed by DAPI staining (nuclear swelling, micronuclei formation). Apoptosis is assessed by Annexin V/PI staining and flow cytometry.
|
| Animal Protocol |
A general animal protocol for VVD‑214: Female NOD/SCID mice are subcutaneously implanted with 5×10⁶ HCT116 MSI‑high colorectal cancer cells or other MSI‑H PDX models. When tumors reach ~150‑200 mm3, mice are randomized into treatment groups (n=8‑10 per group). VVD‑214 is formulated in a suitable vehicle (e.g., 10% DMSO, 40% PEG300, 5% Tween‑80, 45% saline) and administered via intraperitoneal (IP) injection or oral gavage at doses of 10, 30, and 100 mg/kg once daily for 21‑28 days. Tumor volume is measured twice weekly. At the end of the study, mice are euthanized, and tumors are harvested for IHC (gammaH2AX, Ki‑67, cleaved caspase‑3), Western blot (gammaH2AX, cleaved PARP), and TUNEL staining. For efficacy in resistant models, VVD‑214 is also evaluated in MSI‑H PDX models derived from patients progressing on immune checkpoint inhibitors.
|
| ADME/Pharmacokinetics |
General PK protocol for VVD‑214: Male Sprague‑Dawley rats are administered VVD‑214 via oral gavage (PO, 10 mg/kg) and intravenous (IV, 2 mg/kg). Blood samples are collected at 0.083, 0.25, 0.5, 1, 2, 4, 8, 12, and 24 hours. Plasma concentrations are quantified by LC‑MS/MS. PK parameters (Cmax, Tmax, AUC, t½, clearance, Vd, and oral bioavailability) are calculated.
|
| Toxicity/Toxicokinetics |
General toxicity protocol for VVD‑214: A 14‑day repeat‑dose toxicity study is performed in ICR mice. VVD‑214 is administered via oral gavage at doses of 10, 30, and 100 mg/kg/day for 14 days. Parameters include clinical signs, body weight, food consumption, hematology (complete blood count, platelet count), serum chemistry (ALT, AST, BUN, creatinine), and histopathology of major organs (bone marrow, gastrointestinal tract, liver, kidneys). Because MSI tumors are the primary target, MSS tissues may be relatively spared. However, due to the role of WRN in maintaining genome stability, long‑term toxicity studies are necessary to assess potential effects on normal proliferating tissues.
|
| References | |
| Additional Infomation |
VVD‑214 (RO7589831) has the molecular formula C20H21F2N3O4S and a molecular weight of 437.46 g/mol. The compound was discovered using chemoproteomics and is a covalent allosteric inhibitor of WRN helicase. It covalently binds to cysteine 727 of WRN. WRN inhibition is synthetically lethal in MSI‑H tumors (approximately 15% of colorectal cancers, 20‑30% of endometrial cancers, and 15‑20% of gastric cancers) due to their dependence on WRN for replication fork stability. VVD‑214 is a research compound for the study of WRN‑targeted therapies and for developing treatments for MSI‑H cancers that are resistant to immune checkpoint inhibitors.
|
| Molecular Formula |
C20H21F2N3O4S
|
|---|---|
| Molecular Weight |
437.46
|
| Exact Mass |
437.122
|
| CAS # |
3026500-20-6
|
| PubChem CID |
170717998
|
| Appearance |
Solid powder
|
| Density |
1.355±0.06 g/cm3(Temp: 20 °C; Press: 760 Torr)(predicted)
|
| Boiling Point |
627.5±55.0 °C(predicted)
|
| LogP |
2.5
|
| Hydrogen Bond Donor Count |
1
|
| Hydrogen Bond Acceptor Count |
8
|
| Rotatable Bond Count |
8
|
| Heavy Atom Count |
30
|
| Complexity |
731
|
| Defined Atom Stereocenter Count |
1
|
| SMILES |
O(C1C=CC=CC=1)C1=NC(=NC=C1C(=O)N[C@@H](C1CC1)/C=C/S(=O)(=O)C)C(F)(F)C
|
| InChi Key |
UWPFUBNCDABUEE-SIFUEBAJSA-N
|
| InChi Code |
InChI=1S/C20H21F2N3O4S/c1-20(21,22)19-23-12-15(18(25-19)29-14-6-4-3-5-7-14)17(26)24-16(13-8-9-13)10-11-30(2,27)28/h3-7,10-13,16H,8-9H2,1-2H3,(H,24,26)/b11-10+/t16-/m1/s1
|
| Chemical Name |
N-[(E,1S)-1-cyclopropyl-3-methylsulfonylprop-2-enyl]-2-(1,1-difluoroethyl)-4-phenoxypyrimidine-5-carboxamide
|
| Synonyms |
RO7589831; VVD-133214
|
| 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 |
| 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 : 100 mg/mL (228.59 mM; with sonication)
|
|---|---|
| Solubility (In Vivo) |
Solubility in Formulation 1: ≥ 2.5 mg/mL (5.71 mM)(Saturation unknown) in 10% DMSO 40% PEG300 5% Tween-80 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, add 100 μL of 25.0 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix well; then add 50 μL Tween-80 to the above system and mix well; then add 450 μL saline to make it 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 (5.71 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, add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL corn oil and mix well. (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 2.2859 mL | 11.4296 mL | 22.8592 mL | |
| 5 mM | 0.4572 mL | 2.2859 mL | 4.5718 mL | |
| 10 mM | 0.2286 mL | 1.1430 mL | 2.2859 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.
Link: https://clinicaltrials.gov/ct2/show/NCT06004245
Conditions:Advanced Solid Tumors|Colorectal CancerProducts are for research use only; We do not sell to patients
Copyright 2020 InvivoChem LLC | All Rights Reserved
