Size | Price | Stock | Qty |
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1mg |
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5mg |
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Other Sizes |
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Targets |
KRAS G12C (IC50 <0.01 μM)
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ln Vitro |
The EC50 of divarasib (Compound 17a) in K-Ras G12C-alkylated HCC1171 cells is 2 nM [2].
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ln Vivo |
Divarasib (10-100 mg/kg/day; PO for 7 days) lowers the amount of free KRAS G12C [1].
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Enzyme Assay |
Preparation of GDC-6036-Bound KRAS G12C [1]
GDC-6036-bound KRAS G12C protein was prepared through alkylation reaction. Briefly, 5 μM of both unlabeled and isotopically labeled (Arg/Lys 13C15N) KRAS G12C proteins were reacted with 20 μM of GDC-6036 in 20 mM HEPE, pH 8, 150 mM NaCl, 5 mM MgCl2, 1 mM EDTA, 1 mM TCEP, and 5 mM of GDP. After overnight incubation at room temperature, both unlabeled and labeled KRAS G12C were transferred to Amicon 3 kDa centrifugal filters for buffer exchange to 10 mM HEPES, 150 mM NaCl, pH 7.4. A 10-μL aliquot was used to assess the extent of GDC-6036 alkylation using a high-resolution accurate-mass mass spectrometry approach on a Q Exactive Plus mass spectrometer as described previously. [1] Spike-In Experiments[1] Free and GDC-6036-bound KRAS G12C proteins were spiked into aliquots of 20 μg total protein (sufficient for 5 replicate runs) from blank tumor tissue lysates with a twofold serial dilution. For the xenograft sample analysis, the blank tissue lysate was prepared from the A549 mouse xenograft model bearing a non-G12C mutant form of KRAS (KRAS G12S). For the human tumor sample analysis, the blank tissue lysate was prepared from a resected human tumor sample that does not harbor the KRAS G12C mutation. Quality control (QC) samples were prepared by mixing free and GDC-6036-bound KRAS G12C standard materials in blank tissue lysates at a ratio of 4:1, 1:1, and 1:4 with the total KRAS G12C protein concentration constant at 5 fmol/μg. Isotopically labeled (Arg/Lys 13C15N) free and GDC-6036-bound KRAS G12C were spiked into each sample at a fixed amount of 125 fmol. |
Cell Assay |
Core Needle Biopsy Collection and Preservation [1]
Tumor biopsies were collected using 18-gauge biopsy needles at 2, 8, or 24 h after the last dose of GDC-6036. Samples were either snap-frozen in cryovial tubes or embedded in optimal cutting temperature (OCT) compound and stored at −80 °C until further analysis.[1] Human NSCLC Tumor Samples[1] Three fresh frozen human NSCLC tumor samples were procured from BioIVT, including two KRAS G12C-positive samples and one KRAS G12C-negative sample. All samples were stored at −80 °C until further analysis.[1] Sample Preparation from Frozen Tumor Biopsies[1] Frozen tumor biopsies were transferred to 0.5 mL polypropylene tubes pre-filled with 1.4 mm ceramic beads and 200 μL of ice-cold Pierce lysis buffer supplemented with cOmplete EDTA-free protease inhibitor cocktail Tablet and then homogenized using a Bead Ruptor 24 Cryo Cooling Unit. Tissue lysates were transferred to 1.5 mL Protein LoBind tubes and centrifuged at 17,000 g for 5 min at 4 °C. The protein concentration of the supernatant was determined using a Pierce BCA Protein Assay Kit |
Animal Protocol |
Animal/Disease Models: Female CB-17 SCID (inbred) mice (20-21 weeks old; 24.1 g) human NSCLC NCI-H2030.X1.1 cells [1]
Doses: 10, 25 or 100 mg/kg Route of Administration: po (oral gavage) daily (QD) for 7 days (Vehicle: 0.5% methylcellulose) Experimental Results: diminished ratio of free KRAS G12C to internal standard. Dose-dependent target engagement was observed at all time points (2, 8, and 24 hrs (hrs (hours)) after the last dose), with >90% KRAS G12C engagement observed at the highest dose evaluated, 100 mg/kg. Female C.B-17 SCID (Inbred) mice that were 20–21 weeks old and weighed an average of 24.1 g were obtained from the Charles River Lab. Human NSCLC NCI-H2030.X1.1 cells were cultured in vitro, harvested during log-phase growth, and resuspended in Hank’s Balanced Salt Solution containing Matrigel (BD Biosciences) at a 1:1 ratio. The cells were then implanted subcutaneously in the right lateral thorax of C.B-17 SCID (Inbred) mice. Each mouse was injected with 10 × 106 cells in a volume of 100 μL. Tumors were monitored until they reached a mean tumor volume of 238–524 mm3. The mice were distributed into 10 groups on the basis of tumor volumes, with n = 4 mice/group. The mean tumor volume across all 10 groups was 363 mm3 at the initiation of dosing with GDC-6036. Mice were given vehicle (100 μL 0.5% methylcellulose), 10, 25, or 100 mg/kg GDC-6036. The vehicle and GDC-6036 were administered by oral gavage (PO) every day (QD) for 7 days in a volume of 100 μL. Tumor volumes were measured at group out (day 0) and following 1 week of dosing (day 7), and the percent change in tumor volume was plotted in GraphPad Prism software. Core needle biopsies were collected on day 7. |
References | |
Additional Infomation |
Divarasib is an orally available inhibitor of the oncogenic KRAS substitution mutation, G12C, with potential antineoplastic activity. Upon oral administration, divarasib selectively targets the KRAS G12C mutant and inhibits KRAS G12C mutant-dependent signaling. KRAS, a member of the RAS family of oncogenes, serves an important role in cell signaling, division and differentiation. Mutations of KRAS may induce constitutive signal transduction leading to tumor cell growth, proliferation, invasion, and metastasis.
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Molecular Formula |
C29H32CLF4N7O2
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Molecular Weight |
622.06
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Exact Mass |
621.22
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Elemental Analysis |
C, 55.99; H, 5.19; Cl, 5.70; F, 12.22; N, 15.76; O, 5.14
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CAS # |
2417987-45-0
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Related CAS # |
2762240-36-6 (adipate); 2417987-45-0 (active atropisomer);2417917-17-8 (atropisomers)
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PubChem CID |
146624881
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Appearance |
White to light yellow solid
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LogP |
5.4
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Hydrogen Bond Donor Count |
1
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Hydrogen Bond Acceptor Count |
12
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Rotatable Bond Count |
6
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Heavy Atom Count |
43
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Complexity |
997
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Defined Atom Stereocenter Count |
2
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SMILES |
C[C@H]1CN(CCN1C2=NC(=NC3=C(C(=C(C=C32)Cl)C4=C(C(=CC(=N4)N)C)C(F)(F)F)F)OC[C@@H]5CCCN5C)C(=O)C=C
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InChi Key |
ZRBPIAWWRPFDPY-IRXDYDNUSA-N
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InChi Code |
InChI=1S/C29H32ClF4N7O2/c1-5-21(42)40-9-10-41(16(3)13-40)27-18-12-19(30)22(26-23(29(32,33)34)15(2)11-20(35)36-26)24(31)25(18)37-28(38-27)43-14-17-7-6-8-39(17)4/h5,11-12,16-17H,1,6-10,13-14H2,2-4H3,(H2,35,36)/t16-,17-/m0/s1
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Chemical Name |
1-((S)-4-(7-((R)-6-amino-4-methyl-3-(trifluoromethyl)pyridin-2-yl)-6-chloro-8-fluoro-2-(((S)-1-methylpyrrolidin-2-yl)methoxy)quinazolin-4-yl)-3-methylpiperazin-1-yl)prop-2-en-1-one
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Synonyms |
GDC-6036; RG6330; GDC 6036; RG-6330; GDC6036; RG 6330
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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) |
DMSO : ~100 mg/mL (~160.76 mM)
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Solubility (In Vivo) |
Solubility in Formulation 1: ≥ 2.5 mg/mL (4.02 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 (4.02 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. View More
Solubility in Formulation 3: ≥ 2.5 mg/mL (4.02 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution. |
Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
1 mM | 1.6076 mL | 8.0378 mL | 16.0756 mL | |
5 mM | 0.3215 mL | 1.6076 mL | 3.2151 mL | |
10 mM | 0.1608 mL | 0.8038 mL | 1.6076 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.