Topoisomerase II inhibitor 3 (Compound 6 h) is an acridone analogue and an inhibitor (blocker/antagonist) of Type II DNA topoisomerase (topo II). It has the effect of topo IIα/β inhibitor and the IC50 of topo IIα subtype is 0.17 μM, and topo IIβ isoform has IC50 of 0.23 μM. It can also cause significant DNA damage and cause apoptosis by triggering the loss of mitochondrial membrane potential.

Physicochemical Properties

Molecular Formula	C18H20N4O4
Molecular Weight	356.375803947449
Exact Mass	356.148
CAS #	99140-25-7
PubChem CID	13753489
Appearance	Typically exists as solid at room temperature
LogP	3.4
Hydrogen Bond Donor Count	3
Hydrogen Bond Acceptor Count	7
Rotatable Bond Count	5
Heavy Atom Count	26
Complexity	524
Defined Atom Stereocenter Count	0
InChi Key	VNSZLRBNUNIGRI-UHFFFAOYSA-N
InChi Code	InChI=1S/C18H20N4O4/c1-21(2)9-3-8-19-14-6-7-15(22(25)26)17-16(14)18(24)12-10-11(23)4-5-13(12)20-17/h4-7,10,19,23H,3,8-9H2,1-2H3,(H,20,24)
Chemical Name	1-[3-(dimethylamino)propylamino]-7-hydroxy-4-nitro-10H-acridin-9-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	Topoisomerase II inhibitor 3 (compound 6h) is a powerful inhibitor of topoisomerase II α/β that causes substantial damage to DNA and induces death by causing the loss of mitochondrial membrane potential. With an IC50 value of 0.17 μM for topoisomerase IIα and an IC50 value of topoisomerase IIβ isoform, topoisomerase II inhibitor 3 (Compound 6 h) is a dual inhibitor of topoisomerase IIα/β. 0.23 micrograms. In a dose-dependent way, compound 6 h, or topoisomerase II inhibitor 3, can similarly cause the development of DSBs [1].
Cell Assay	Cell proliferation experiment [1] Cell Types: human breast cancer MDA-MB-231 cells; human lung cancer A549; human acute myeloid leukemia KG1 cells; rat cardiac H9C2 cells Tested Concentrations: 100 μM Incubation Duration: 12 hrs (hours) Experimental Results: In MDA- The most potent anti-proliferative activity was exerted in MB-231 cells (IC50: 0.42 μM), A549 cells (IC50: 1.10 μM), KG1 cells (IC50: 0.15 μM) and H9C2 cells (IC50 >20 μM). Apoptosis analysis [1] Cell Types: MDA-MB-231 Cell Tested Concentrations: 0.5-10 μM Incubation Duration: 24 h Experimental Results: Caused significant loss of mitochondrial membrane potential (MMP) in MDA-MB-231 cells.
References	[1]. Zhi-Ying Li, et al. Structural optimizations and bioevaluation of N-substituted acridone derivatives as strong topoisomerase II inhibitors. Bioorg Chem. 2022 Feb;119:105543.

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	2.8060 mL	14.0300 mL	28.0599 mL
	5 mM	0.5612 mL	2.8060 mL	5.6120 mL
	10 mM	0.2806 mL	1.4030 mL	2.8060 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.