Senexin A is a novel, potent, selective and ATP site competitive inhibitor of CDK8 and CDK19 with Kd values of 0.83 μM and 0.31 μM for CDK8 and CDK19, respectively. In addition to killing tumor cells, conventional chemotherapy alters the expression of certain genes in tissues damaged by the treatment, which causes the production of several factors that secrete and support tumor growth. It was discovered that the damage-inducible cell-cycle inhibitor p21 (CDKN1A) mediated this secretory phenotype in part. Senexin A prevents transcription that is triggered by damage after p21. Senexin A reverses the increase in tumor engraftment and serum mitogenic activity in mice treated with chemotherapeutic drugs, as well as damage-induced tumor-promoting paracrine activities of tumor cells and normal fibroblasts. Additionally, sensexin A improves the effectiveness of chemotherapy in combating xenografts made of mixtures of tumor cells and fibroblasts. Analysis of microarray data showed remarkable associations between low survival rates in ovarian and breast cancers and CDK8 expression. Senexin A's inhibition of CDK8 presents a viable strategy for boosting the effectiveness of cancer chemotherapy.

Physicochemical Properties

Molecular Formula	C17H14N4
Molecular Weight	274.32
Exact Mass	274.122
Elemental Analysis	C, 74.43; H, 5.14; N, 20.42
CAS #	1366002-50-7
Related CAS #	1366002-50-7
PubChem CID	56927063
Appearance	White to off-white solid powder
LogP	3.229
Hydrogen Bond Donor Count	1
Hydrogen Bond Acceptor Count	4
Rotatable Bond Count	4
Heavy Atom Count	21
Complexity	368
Defined Atom Stereocenter Count	0
SMILES	N#CC1C=CC2=NC=NC(NCCC3C=CC=CC=3)=C2C=1
InChi Key	XBJCNHGQFJFCOY-UHFFFAOYSA-N
InChi Code	InChI=1S/C17H14N4/c18-11-14-6-7-16-15(10-14)17(21-12-20-16)19-9-8-13-4-2-1-3-5-13/h1-7,10,12H,8-9H2,(H,19,20,21)
Chemical Name	4-(2-phenylethylamino)quinazoline-6-carbonitrile
Synonyms	Senexin A
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

Targets	CDK19 (Kd = 0.31 μM); CDK8 (Kd = 0.83 μM)
ln Vitro	Senexin A suppresses CDK8 kinase activity with an IC50 of 0.28 μM and CDK8 and CDK19 ATP site binding with Kd50 of 0.83 μM and 0.31 μM, respectively. Senexin A suppresses transcription in HCT116 colon cancer cells that is dependent on β-catenin. Senexin A potently suppresses the induction of transcription factor EGR1 in HT1080 cells upon serum starvation, subsequent to the readdition of serum. Senexin A solely prevents p21-induced transcription; it has no effect on p21's other biological effects. Senexin A also reduces the expression of numerous secreted factors that promote tumor growth in wild-type HCT116 cells treated with doxorubicin[1].
ln Vivo	Senexin A five times a day completely eliminates the effect of chemotherapy that promotes tumor growth. When used in C57BL/6 mice, sensexin A does not exhibit any observable toxicity and has no appreciable impact on body weight, organ weights, or blood cell counts. Nevertheless, if Senexin A is given after doxorubicin injection, this side effect of the medication is totally eliminated. Treatment with sentexin A significantly enhances the way doxorubicin responds to A549/MEF tumors[1].
Enzyme Assay	Senexin A suppresses CDK8 kinase activity with an IC50 of 0.28 μM and CDK8 and CDK19 ATP site binding with Kd50 of 0.83 μM and 0.31 μM, respectively. Senexin A has no effect on IPTG-induced p21 induction, p21-induced senescent phenotype, or cell growth. Senexin A has no effect on p21's inhibition of gene expression, nor does it obstruct the p21-mediated inhibition of sizable gene sets associated with the DNA replication and mitosis Gene Ontology (GO) categories. Senexin A solely prevents p21-induced transcription; it has no effect on p21's other biological effects. Senexin A suppresses CDK8 kinase activity with an IC50 of 0.28 μM and CDK8 and CDK19 ATP site binding with Kd50 of 0.83 μM and 0.31 μM, respectively. Senexin A suppresses transcription in HCT116 colon cancer cells that is dependent on β-catenin. It does not share cortistatin's potent antiendothelial cell activity and does not inhibit ROCK.
Cell Assay	Conditioned media for mitogenic assays using conditioned media were obtained by washing and cultured MEF for 48 hours without medication, either untreated or treated for 24 hours with 200 nM doxorubicin alone or in combination with 1 μM Senexin A. A549 cells were plated on 12-well plates with 104 cells per well after the media was added. The trypan blue exclusion assay was used to count the cells 48 hours later. Each experiment involved the counting of cells in a minimum of three optical fields and was carried out in triplicate.
Animal Protocol	Mice: Five mice per group are treated with 20 mg/kg Senexin A or a carrier (80% propylene glycol) with five daily intraperitoneal injections as part of Taconic's Senexin A toxicity study in C57BL/6 mice. On days three and six, mice are weighed; on day six, they are killed. The brain, kidney, spleen, thymus, lung, and liver are assigned organ weights. Total white blood cells, neutrophils, lymphocytes, monocytes, eosinophils, and basophils are counted from terminal blood samples[1].
References	[1]. Cyclin-dependent kinase 8 mediates chemotherapy-induced tumor-promoting paracrine activities. Proc Natl Acad Sci U S A. 2012 Aug 21;109(34):13799-804.

Solubility Data

Solubility (In Vitro)

DMSO: >50 mg/mL Water: < 1mg/mL Ethanol: >50 mg/mL

Solubility (In Vivo)

Solubility in Formulation 1: ≥ 2.5 mg/mL (9.11 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 (9.11 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	3.6454 mL	18.2269 mL	36.4538 mL
	5 mM	0.7291 mL	3.6454 mL	7.2908 mL
	10 mM	0.3645 mL	1.8227 mL	3.6454 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.