CCB02 is a selective CPAP-tubulin interaction inhibitor that can bind to tubulin and compete for the CPAP binding site of β-tubulin. It has IC50 of 689 nM and has high anti-tumor activity. CCB02 has no inhibitory activity against other proteins like cell cycle and centrosome-related proteins, and has no effect on the phosphorylation status of Aurora A, Plk1, Plk2, CDK2 and CHK1.

Physicochemical Properties

Molecular Formula	C14H9N3O
Molecular Weight	235.240762472153
Exact Mass	235.074
CAS #	2100864-57-9
PubChem CID	129216797
Appearance	Light yellow to yellow solid powder
LogP	2.5
Hydrogen Bond Donor Count	0
Hydrogen Bond Acceptor Count	4
Rotatable Bond Count	1
Heavy Atom Count	18
Complexity	350
Defined Atom Stereocenter Count	0
SMILES	O(C)C1=C(C#N)C2C(C=N1)=CC1C=CC=CC=1N=2
InChi Key	QNJYUHRGCPRPQS-UHFFFAOYSA-N
InChi Code	InChI=1S/C14H9N3O/c1-18-14-11(7-15)13-10(8-16-14)6-9-4-2-3-5-12(9)17-13/h2-6,8H,1H3
Chemical Name	3-methoxybenzo[b][1,6]naphthyridine-4-carbonitrile
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 (e.g. under nitrogen), 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)

Biological Activity

ln Vitro	CCB02 inhibits the CPAP PN2-3-tubulin interaction in the PN2-3 CPAP-GST pull-down test, with an IC50 of 0.441 μM [1]. The phosphorylation status of Aurora A, Plk1, Plk2, CDK2, and CHK1 as well as centrosome-associated kinases are not inhibited by CCB02 [1]. Cancer cells with additional centrosomes are inhibited from proliferating by CCB02 (0.1-15 μM, 72 hours) with an IC50 of 0.86-2.9 μM [1]. CCB02 stimulates the recruitment of PCM proteins to the centrosome, activates the spindle assembly checkpoint, and increases the centrosome's microtubule nucleation activity [1].
ln Vivo	CCB02 (30 mg/kg orally daily for 24 days) demonstrated strong anticancer effects in tumor xenografts of the human lung (H1975T790M cells) subcutaneously implanted in nude mice [1]. In mouse xenografts, CCB02 also results in multipolar mitosis and inhibits MDA-MB-231 cell migration [1].
Cell Assay	Cell Viability Test[1] Cell Types: BT549, MDA-MB-231, Pop10, SCC13, SW1271p53/pRb/CDKN2Adel, KYSE30p53/MYC/CyclinD1, A549G12S, PC-9EGFR-Exon19del, HCC827-GR, HCC1833-GR, H1975T790M Cell Tested Concentrations: 0.1-15 μM Incubation Duration: 72 hrs (hours) Experimental Results: IC50 is 0.86 μM (Pop10), 1.2 μM (HCC827-GR), 1.5 μM (H1975T790Mp53/MYC/CyclinD1), 1.15 μM (HCC1833-GR), 1.61 μM (SW1271p53/pRb/CDKN2Adel), 2.41 μM (SCC13) and 2.94 μM (PC-9EGFR-Exon19del).
Animal Protocol	Animal/Disease Models: Nude mice carrying subcutaneoushuman lung (H1975T790M) tumor xenografts [1] Doses: 30 mg/kg Route of Administration: daily oral administration for 24 days Experimental Results: Tumor volume was Dramatically diminished on day 24.
References	[1]. Inhibition of CPAP-tubulin interaction prevents proliferation of centrosome-amplified cancer cells. EMBO J. 2019 Jan 15;38(2). pii: e99876.

Solubility Data

Solubility (In Vitro)

DMSO : ~25 mg/mL (~106.27 mM)

Solubility (In Vivo)

Solubility in Formulation 1: 2.5 mg/mL (10.63 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 sonication. 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. Solubility in Formulation 2: ≥ 2.5 mg/mL (10.63 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 is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.  (Please use freshly prepared in vivo formulations for optimal results.)

Preparing Stock Solutions		1 mg	5 mg	10 mg
	1 mM	4.2510 mL	21.2549 mL	42.5098 mL
	5 mM	0.8502 mL	4.2510 mL	8.5020 mL
	10 mM	0.4251 mL	2.1255 mL	4.2510 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.