Anticancer agent 98 (compound 12k) is a tubulin polymerization inhibitor (Kd=16.9 μM). Anticancer agent 98 exerts antiproliferation effects on tumor cells and displays antiangiogenic effects in vitro. Anticancer agent 98 displays good human and mouse liver microsomal stability with t1/2>300 minutes.

Physicochemical Properties

Molecular Formula	C17H19N5O2
Molecular Weight	325.37
Exact Mass	325.153
CAS #	2857070-72-3
PubChem CID	166090152
Appearance	Typically exists as solid at room temperature
LogP	2.2
Hydrogen Bond Donor Count	2
Hydrogen Bond Acceptor Count	6
Rotatable Bond Count	3
Heavy Atom Count	24
Complexity	476
Defined Atom Stereocenter Count	0
InChi Key	BRTSQLXTBUCXNF-UHFFFAOYSA-N
InChi Code	InChI=1S/C17H19N5O2/c1-18-17-20-12-5-3-4-11(12)16(21-17)22-9-15(23)19-13-8-10(24-2)6-7-14(13)22/h6-8H,3-5,9H2,1-2H3,(H,19,23)(H,18,20,21)
Chemical Name	7-methoxy-4-[2-(methylamino)-6,7-dihydro-5H-cyclopenta[d]pyrimidin-4-yl]-1,3-dihydroquinoxalin-2-one
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)

Biological Activity

ln Vitro	Anticancer agent 98 has an IC50 range of 0.6-3 nM and suppresses the proliferation of cancer in melanoma, breast cancer, and pancreatic cancer[1]. In PC-3 cells, the anticancer drug 98 (300 nM, 1 μM, or 3 μM; 2 hours) increases β-tubulin adducts in a dose-dependent manner [1]. By using SPR spectroscopy, anticancer drug 98 (3.125, 6.25, 12.5, 25 and 50 μM) has a high binding affinity of 16.9 μM for tubulin [1]. Tubulin polymerization is significantly inhibited by anticancer agent 98 (10 μM, 50 μM; 0-60 minutes) for a duration of 60 minutes [1]. On COS-7 cells in vitro, anticancer drug 98 (100 nM; 4 h) exhibits anti-proliferative and anti-angiogenic properties [1]. Molecular stability in vitro[1] t1/2 (min) CLint (μL/min/mg) >300 <2.31 >300 <2.31 human microsomes mouse microsomes
ln Vivo	In male NSG mice with PC3/TxR xenograft tumors, anticancer agent 98 (2.5 mg/kg; IV; twice weekly for 2 weeks) was well tolerated without causing appreciable weight loss. Prostate cancer tumor growth was also markedly reduced by anticancer agent 98 in comparison to the Paclitaxel (HY-B0015; 10 mg/kg once weekly) and control groups [1]. Route Dose (mg/kg) Cmax (ng/mL) tmax (min) AUC (ng·min/mL) t1/2 (min) F (%) iv 4 1247 5.0 173,476 238 / po 10 78.3 10.0 8161 358 2.02
Animal Protocol	Animal/Disease Models: PC3/TxR xenograft model in NSG mouse[1] Doses: 2.5 mg/kg Route of Administration: IV; twice weekly for 2 weeks Experimental Results: Inhibited the tumor growth in volume by approximately 85.6%. And inhibited angiogenesis by 44% related to control group. Overcame taxane resistance at a low, safe, but potent dose in vivo.
References	[1]. Design, Synthesis, and Biological Evaluation of Pyrimidine Dihydroquinoxalinone Derivatives as Tubulin Colchicine Site-Binding Agents That Displayed Potent Anticancer Activity Both In Vitro and In Vivo. ACS Pharmacology & Translational Science, 2023.

Solubility Data

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

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 (e.g. IP/IV/IM/SC) Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline) *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). 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)] *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. Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin → 500 μL Saline) Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO → 100 μLPEG300 → 200 μL castor oil → 650 μL Saline) Injection Formulation 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (i.e. 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline) Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH → 900 μL Corn oil) Injection Formulation 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL 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). Oral Formulation 3: Dissolved in PEG400 Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose Oral Formulation 6: Mixing with food powders Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently.  (Please use freshly prepared in vivo formulations for optimal results.)

Preparing Stock Solutions		1 mg	5 mg	10 mg
	1 mM	3.0734 mL	15.3671 mL	30.7342 mL
	5 mM	0.6147 mL	3.0734 mL	6.1468 mL
	10 mM	0.3073 mL	1.5367 mL	3.0734 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.