Biopolym. Cell. 2021; 37(1):62-72.
Bioorganic Chemistry
Evaluation of in vitro activity and SAR study of the novel hetarylamino-3-aryl-1H-indazole derivatives as inhibitors of protein kinase CK2
1Protopopov M. V., 1Vdovin V. S., 1Lukashov S. S., 1Ostrynska O. V., 1, 2Borysenko I. P., 1Borovykov O. V., 1Starosyla S. A., 3Bilokin Y. V., 1Kukharenko O. P., 1Bdzhola V. G., 1Yarmoluk S. M.
  1. Institute of Molecular Biology and Genetics, NAS of Ukraine
    150, Akademika Zabolotnoho Str., Kyiv, Ukraine, 03143
  2. LLC “Scientific and service firm “Otava”
    150, Akademika Zabolotnoho Str., Kyiv, Ukraine, 03143
  3. OTAVA Ltd
    400 Applewood Crescent, Unit 100, Vaughan, Ontario, L4K 0C3 Canada


Aim. To identify novel protein kinase CK2 inhibitors among the 5-hetarylamino-1H-indazoles. Methods. Biochemical testing was carried out with the aid of γ-32P ATP in vitro kinase assay. Molecular docking via the Autodock 4.2.6 program package was executed, rescoring of docking results was performed using DrugScore scoring function. Results. Among the 17 studied 5-amino-3-arylindazole derivatives 11 inhibitors of protein kinase CK2 with IC50 in nanomolar range were identified. The most active compound has IC50 = 2 nM. SAR study and additional molecular modeling of these compounds allowed us to select prospective substituents for construction of novel compounds with improved activity and physicochemical properties. Conclusions. As a result of this work 11 nanomolar protein kinase CK2 inhibitors were developed and the binding modes of these compounds with the ATP-acceptor site were proposed using molecular docking methods. The physicochemical properties and SAR of substituents of studied compounds were analyzed and 6 novel compounds were designed for further development as protein kinase CK2 inhibitors. Summarizing, 5-heterylamino-1H-indazoles are a good basis for further CK2 inhibitors development.
Keywords: protein kinase CK2 inhibitor, in vitro kinase assay, molecular docking, indazole, pyrazolo[3,4-d]pyrimidine


[1] Burnett G, Kennedy EP. The enzymatic phosphorylation of proteins. J Biol Chem. 1954;211(2):969-80.
[2] Qiao Y, Chen T, Yang H, Chen Y, Lin H, Qu W, Feng F, Liu W, Guo Q, Liu Z, Sun H. Small molecule modulators targeting protein kinase CK1 and CK2. Eur J Med Chem. 2019;181:111581.
[3] Poole A, Poore T, Bandhakavi S, McCann RO, Hanna DE, Glover CV. A global view of CK2 function and regulation. Mol Cell Biochem. 2005;274(1-2):163-70.
[4] Niefind K, Pütter M, Guerra B, Issinger OG, Schomburg D. GTP plus water mimic ATP in the active site of protein kinase CK2. Nat Struct Biol. 1999;6(12):1100-3.
[5] Bian Y, Ye M, Wang C, Cheng K, Song C, Dong M, Pan Y, Qin H, Zou H. Global screening of CK2 kinase substrates by an integrated phosphoproteomics workflow. Sci Rep. 2013;3:3460.
[6] Meggio F, Pinna LA. One-thousand-and-one substrates of protein kinase CK2? FASEB J. 2003;17(3):349-68.
[7] Hanks SK, Hunter T. Protein kinases 6. The eukaryotic protein kinase superfamily: kinase (catalytic) domain structure and classification. FASEB J. 1995;9(8):576-96.
[8] Wilson LK, Dhillon N, Thorner J, Martin GS. Casein kinase II catalyzes tyrosine phosphorylation of the yeast nucleolar immunophilin Fpr3. J Biol Chem. 1997;272(20):12961-7.
[9] Faust M, Montenarh M. Subcellular localization of protein kinase CK2. A key to its function? Cell Tissue Res. 2000;301(3):329-40.
[10] Protopopov MV, Vdovin VS, Lukashov SS, Ostrynska OV, Borysenko IP, Borovykov OV, Starosyla SA, Bilokin YV, Kukharenko OP, Bdzhola VG, Yarmoluk SM. The synthesis of 5-hetarylamino-3-aryl-1H-indazoles as inhibitors of protein kinase CK2. Biopolym Cell. 2020; 36(6):466–76.
[11] Morris GM, Huey R, Lindstrom W, Sanner MF, Belew RK, Goodsell DS, Olson AJ. AutoDock4 and AutoDockTools4: Automated docking with selective receptor flexibility. J Comput Chem. 2009;30(16):2785-91.
[12] Ferguson AD, Sheth PR, Basso AD, Paliwal S, Gray K, Fischmann TO, Le HV. Structural basis of CX-4945 binding to human protein kinase CK2. FEBS Lett. 2011;585(1):104-10.
[13] Guerra B, Bischoff N, Bdzhola VG, Yarmoluk SM, Issinger OG, Golub AG, Niefind K. A Note of Caution on the Role of Halogen Bonds for Protein Kinase/Inhibitor Recognition Suggested by High- And Low-Salt CK2α Complex Structures. ACS Chem Biol. 2015;10(7):1654-60.
[14] Pedretti A, Villa L, Vistoli G. VEGA--an open platform to develop chemo-bio-informatics applications, using plug-in architecture and script programming. J Comput Aided Mol Des. 2004;18(3):167-73.
[15] Hastie CJ, McLauchlan HJ, Cohen P. Assay of protein kinases using radiolabeled ATP: a protocol. Nat Protoc. 2006;1(2):968-71.
[16] Lukashov SS, Kukharenko OP, Bdzhola VG, Yarmoluk SM. Synthesis of 5-amino-3-arylindazole derivatives and study of their in vitro activity towards Ser/Thr and Tyr protein kinases. V International Conference «Chemistry of Nitrogen Containing Heterocycles», Kharkiv. 2009:O.21.
[17] Johnson TW, Gallego RA, Edwards MP. Lipophilic Efficiency as an Important Metric in Drug Design. J Med Chem. 2018;61(15):6401-6420.
[18] Shultz MD. Setting expectations in molecular optimizations: Strengths and limitations of commonly used composite parameters. Bioorg Med Chem Lett. 2013;23(21):5980-91.
[19] Roskoski R Jr. Properties of FDA-approved small molecule protein kinase inhibitors. Pharmacol Res. 2019;144:19-50.
[20] Neudert G, Klebe G. DSX: a knowledge-based scoring function for the assessment of protein-ligand complexes. J Chem Inf Model. 2011;51(10):2731-45.