WARNING: This product is for research use only, not for human or veterinary use.
MedKoo CAT#: 526754
Description: D-4476, also known as Casein Kinase I Inhibitor, is a potent, selective and cell-permeant inhibitor of casein kinase 1 (CK1; IC50 = 200 nM from S. pombe,1 300 nM for CK1δ2). D4476 suppresses the site-specific phosphorylation and nuclear exclusion of FOXO1a. D4476 specifically inhibits the phosphorylation of endogenous forkhead box transcription factor O1a (FOXO1a) on Ser322 and Ser325 within its MPD, without affecting the phosphorylation of other sites.
MedKoo Cat#: 526754
Chemical Formula: C23H18N4O3
Exact Mass: 398.1379
Molecular Weight: 398.422
Elemental Analysis: C, 69.34; H, 4.55; N, 14.06; O, 12.05
Synonym: D-4476; D 4476; D4476; Casein Kinase I Inhibitor.
IUPAC/Chemical Name: 4-[4-(2,3-dihydro-1,4-benzodioxin-6-yl)-5-(2-pyridinyl)1H-imidazol-2-yl]-benzamide
InChi Key: DPDZHVCKYBCJHW-UHFFFAOYSA-N
InChi Code: InChI=1S/C23H18N4O3/c24-22(28)14-4-6-15(7-5-14)23-26-20(21(27-23)17-3-1-2-10-25-17)16-8-9-18-19(13-16)30-12-11-29-18/h1-10,13H,11-12H2,(H2,24,28)(H,26,27)
SMILES Code: O=C(N)C1=CC=C(C2=NC(C3=CC=C(OCCO4)C4=C3)=C(C5=NC=CC=C5)N2)C=C1
Appearance: Solid powder
Purity: >98% (or refer to the Certificate of Analysis)
Shipping Condition: Shipped under ambient temperature as non-hazardous chemical. This product is stable enough for a few weeks during ordinary shipping and time spent in Customs.
Storage Condition: Dry, dark and at 0 - 4 C for short term (days to weeks) or -20 C for long term (months to years).
Solubility: Soluble in DMSO
Shelf Life: >2 years if stored properly
Drug Formulation: This drug may be formulated in DMSO
Stock Solution Storage: 0 - 4 C for short term (days to weeks), or -20 C for long term (months).
HS Tariff Code: 2934.99.9001
|Biological target:||D4476 is an inhibitor of casein kinase 1(CK1) with an IC50 value of 0.3 μM.|
|In vitro activity:||To investigate the role of CK1α in AML (acute myeloid leukemia) cells, CK1α was inhibited using D4476. D4476 significantly reduced the cell viability of three AML cell lines HL-60, HEL and THP-1 in a time- and dose-dependent manner (Fig. 2A), while only a dose-dependent manner was shown after treatment with D4476 lower than 10 µM in patient blast cells. Whether D4476 affects the colony formation ability of AML cells was further investigated, and it was found that D4476 at 40 µM dramatically impaired the colony formation, as indicated with colony number and total cell number, in both HL-60 and HEL cells (Fig. 2B). Notably, there were many small cell clusters, but did not reach the standard of a colony in the D4476 treatment group in these two cell lines (Fig. 2B). Furthermore, D4476 dramatically induced apoptosis in a dose-dependent manner in three AML cell lines tested as well as patient blast cells (Fig. 3A). D4476 treatment promoted the cleavage of PARP, a classical marker of apoptosis, and decreased the expression of survivin, a negative marker protein of apoptosis, in HL-60, THP-1 and HEL cells as well as in patient blast cells (Fig. 3A). Reference: Oncol Rep. 2020 Nov;44(5):1895-1904. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7550986/|
|In vivo activity:||In order determine the therapeutic effect of D4476 in vivo in a clinically relevant model of GBM (glioblastoma multiforme), a glioma stem cell (GSC)-driven xenograft tumour mouse model was used. The tumour size in treatment with D4476 was smaller than those in the control group. Therefore, D4476-treated mice showed increased survival rate. Significantly, D4476 treatment combined with radiotherapy obviously inhibited the growth of tumour (Figure 6C). The survival time of mice bearing GBM was significantly increased in the combination group (Figure 6D). Compared with the control group, the median survival time of mice in the D4476 treatment group was extended by 8 days; radiotherapy treatment extended survival by 6 days, and D4476 combined with radiotherapy resulted in a 22-day extension (Table S3). Reference: J Cell Mol Med. 2021 Jul 3. https://onlinelibrary.wiley.com/doi/10.1111/jcmm.16767|
|Solvent||Max Conc. mg/mL||Max Conc. mM|
|DMSO:PBS (pH 7.2) (1:10)||0.1||0.25|
The following data is based on the product molecular weight 398.422 Batch specific molecular weights may vary from batch to batch due to the degree of hydration, which will affect the solvent volumes required to prepare stock solutions.
|Concentration / Solvent Volume / Mass||1 mg||5 mg||10 mg|
|1 mM||1.15 mL||5.76 mL||11.51 mL|
|5 mM||0.23 mL||1.15 mL||2.3 mL|
|10 mM||0.12 mL||0.58 mL||1.15 mL|
|50 mM||0.02 mL||0.12 mL||0.23 mL|
|Formulation protocol:||1. Rena G, Bain J, Elliott M, Cohen P. D4476, a cell-permeant inhibitor of CK1, suppresses the site-specific phosphorylation and nuclear exclusion of FOXO1a. EMBO Rep. 2004 Jan;5(1):60-5. doi: 10.1038/sj.embor.7400048. PMID: 14710188; PMCID: PMC1298959. 2. Xu W, Huang Z, Gan Y, Chen R, Huang Y, Xue B, Jiang S, Yu Z, Yu K, Zhang S. Casein kinase 1α inhibits p53 downstream of MDM2‑mediated autophagy and apoptosis in acute myeloid leukemia. Oncol Rep. 2020 Nov;44(5):1895-1904. doi: 10.3892/or.2020.7760. Epub 2020 Sep 9. PMID: 32901886; PMCID: PMC7550986. 3. Liu G, Li H, Zhang W, Yu J, Zhang X, Wu R, Niu M, Liu X, Yu R. Csnk1a1 inhibition modulates the inflammatory secretome and enhances response to radiotherapy in glioma. J Cell Mol Med. 2021 Jul 3. doi: 10.1111/jcmm.16767. Epub ahead of print. PMID: 34216174.|
|In vitro protocol:||1. Rena G, Bain J, Elliott M, Cohen P. D4476, a cell-permeant inhibitor of CK1, suppresses the site-specific phosphorylation and nuclear exclusion of FOXO1a. EMBO Rep. 2004 Jan;5(1):60-5. doi: 10.1038/sj.embor.7400048. PMID: 14710188; PMCID: PMC1298959. 2. Xu W, Huang Z, Gan Y, Chen R, Huang Y, Xue B, Jiang S, Yu Z, Yu K, Zhang S. Casein kinase 1α inhibits p53 downstream of MDM2‑mediated autophagy and apoptosis in acute myeloid leukemia. Oncol Rep. 2020 Nov;44(5):1895-1904. doi: 10.3892/or.2020.7760. Epub 2020 Sep 9. PMID: 32901886; PMCID: PMC7550986.|
|In vivo protocol:||1. Liu G, Li H, Zhang W, Yu J, Zhang X, Wu R, Niu M, Liu X, Yu R. Csnk1a1 inhibition modulates the inflammatory secretome and enhances response to radiotherapy in glioma. J Cell Mol Med. 2021 Jul 3. doi: 10.1111/jcmm.16767. Epub ahead of print. PMID: 34216174.|
1: Chia R, Haddock S, Beilina A, Rudenko IN, Mamais A, Kaganovich A, Li Y, Kumaran R, Nalls MA, Cookson MR. Phosphorylation of LRRK2 by casein kinase 1α regulates trans-Golgi clustering via differential interaction with ARHGEF7. Nat Commun. 2014 Dec 15;5:5827. doi: 10.1038/ncomms6827. PubMed PMID: 25500533; PubMed Central PMCID: PMC4268884.
2: Bhattacharya D, Dwivedi VP, Kumar S, Reddy MC, Van Kaer L, Moodley P, Das G. Simultaneous inhibition of T helper 2 and T regulatory cell differentiation by small molecules enhances Bacillus Calmette-Guerin vaccine efficacy against tuberculosis. J Biol Chem. 2014 Nov 28;289(48):33404-11. doi: 10.1074/jbc.M114.600452. Epub 2014 Oct 14. PubMed PMID: 25315774; PubMed Central PMCID: PMC4246096.
3: Rachidi N, Taly JF, Durieu E, Leclercq O, Aulner N, Prina E, Pescher P, Notredame C, Meijer L, Späth GF. Pharmacological assessment defines Leishmania donovani casein kinase 1 as a drug target and reveals important functions in parasite viability and intracellular infection. Antimicrob Agents Chemother. 2014;58(3):1501-15. doi: 10.1128/AAC.02022-13. Epub 2013 Dec 23. PubMed PMID: 24366737; PubMed Central PMCID: PMC3957854.
4: Wang L, Lu A, Zhou HX, Sun R, Zhao J, Zhou CJ, Shen JP, Wu SN, Liang CG. Casein kinase 1 alpha regulates chromosome congression and separation during mouse oocyte meiotic maturation and early embryo development. PLoS One. 2013 May 15;8(5):e63173. doi: 10.1371/journal.pone.0063173. Print 2013. PubMed PMID: 23690993; PubMed Central PMCID: PMC3655170.
5: León-Espinosa G, García E, García-Escudero V, Hernández F, Defelipe J, Avila J. Changes in tau phosphorylation in hibernating rodents. J Neurosci Res. 2013 Jul;91(7):954-62. doi: 10.1002/jnr.23220. Epub 2013 Apr 19. PubMed PMID: 23606524.
6: Stavenuiter F, Gale AJ, Heeb MJ. Phosphorylation of protein S by platelet kinases enhances its activated protein C cofactor activity. FASEB J. 2013 Jul;27(7):2918-25. doi: 10.1096/fj.12-225961. Epub 2013 Apr 11. PubMed PMID: 23580615; PubMed Central PMCID: PMC3688756.
7: Foucher AL, Rachidi N, Gharbi S, Blisnick T, Bastin P, Pemberton IK, Späth GF. Apoptotic marker expression in the absence of cell death in staurosporine-treated Leishmania donovani. Antimicrob Agents Chemother. 2013 Mar;57(3):1252-61. doi: 10.1128/AAC.01983-12. Epub 2012 Dec 21. PubMed PMID: 23263009; PubMed Central PMCID: PMC3591890.
8: Zelenak C, Eberhard M, Jilani K, Qadri SM, Macek B, Lang F. Protein kinase CK1α regulates erythrocyte survival. Cell Physiol Biochem. 2012;29(1-2):171-80. doi: 10.1159/000337598. Epub 2012 Mar 1. PubMed PMID: 22415086.
9: Iguchi Y, Katsuno M, Takagi S, Ishigaki S, Niwa J, Hasegawa M, Tanaka F, Sobue G. Oxidative stress induced by glutathione depletion reproduces pathological modifications of TDP-43 linked to TDP-43 proteinopathies. Neurobiol Dis. 2012 Mar;45(3):862-70. doi: 10.1016/j.nbd.2011.12.002. Epub 2011 Dec 13. PubMed PMID: 22198567.
10: Zyss D, Ebrahimi H, Gergely F. Casein kinase I delta controls centrosome positioning during T cell activation. J Cell Biol. 2011 Nov 28;195(5):781-97. doi: 10.1083/jcb.201106025. PubMed PMID: 22123863; PubMed Central PMCID: PMC3257584.
11: Biswas A, Mukherjee S, Das S, Shields D, Chow CW, Maitra U. Opposing action of casein kinase 1 and calcineurin in nucleo-cytoplasmic shuttling of mammalian translation initiation factor eIF6. J Biol Chem. 2011 Jan 28;286(4):3129-38. doi: 10.1074/jbc.M110.188565. Epub 2010 Nov 17. PubMed PMID: 21084295; PubMed Central PMCID: PMC3024805.
12: Huart AS, MacLaine NJ, Meek DW, Hupp TR. CK1alpha plays a central role in mediating MDM2 control of p53 and E2F-1 protein stability. J Biol Chem. 2009 Nov 20;284(47):32384-94. doi: 10.1074/jbc.M109.052647. Epub 2009 Sep 15. PubMed PMID: 19759023; PubMed Central PMCID: PMC2781653.
13: Rena G, Bain J, Elliott M, Cohen P. D4476, a cell-permeant inhibitor of CK1, suppresses the site-specific phosphorylation and nuclear exclusion of FOXO1a. EMBO Rep. 2004 Jan;5(1):60-5. PubMed PMID: 14710188; PubMed Central PMCID: PMC1298959.