LIM Domain Kinase (LIMK) Inhibitor R10015

- The best-characterized LIMK inhibitor

LIM domain kinases (LIMKs) are serine/threonine and tyrosine kinases which phosphorylate and inactivate the ADF/cofilin family of proteins through serine 3 phosphorylation (Yang et al., 1998). LIMK regulates actin dynamics through phosphorylating cofilin, and this phosphorylation inhibits cofilin-mediated depolymerization of the actin filaments. Both LIMKs (LIMK1 and LIMK2) contain two N-terminal LIM domains, a PDZ domain, and a C-terminal protein kinase domain. LIMKs are regulated by small GTPases of the Rho family, Rho-dependent kinase (ROCK), and the p21-activated kinases PAK1/PAK4. LIMKs are involved in many cellular processes associated with actin dynamics and actin cytoskeletal rearrangement, such as cell migration, chemotaxis, cell development, neuronal differentiation, cell-cell interaction, tissue repair, wound healing, cancer metastasis, and viral infection (Foletta et al., 2004; Manetti, 2012; Yoshioka et al., 2003). LIMK is highly expressed in many metastatic cancer cells, and has been identified as a possible therapeutic target (Endo et al., 2007; Nishimura et al., 2006). In addition, LIMK has been shown to be involved in the infection and pathogenesis of multiple viruses such as HIV, influenza A, pseudorabies virus, EBOV, RVFV, VEEV, and HSV-1 (Liu et al., 2014; Vorster et al., 2011). LIMK inhibitors have also been shown to possess broad-spectrum anti-viral activities.

The LIMK inhibitor R10015 is the best biochemically and biologically characterized LIMK inhibitor (Yi et al., 2017). R10015 has been profiled against a panel of 62 kinases (Table 1). It directly inhibits the kinase activity of LIMK by binding to the ATP-binding pocket of LIMK. The in vitro IC50 value for inhibiting recombinant LIMK1 protein was approximately 38 nM (Fig. 1). R10015 blocks the phosphorylation of cofilin at serine 3 in human T cells (Fig. 2). R10015 also inhibits T cell chemotaxis and chemokine-induced actin polymerization (Fig. 3).

R10015

R10015 highlights:

  • Molecular weight (M.W.) = 525.9
  • LIM Kinase inhibition IC50 = 38±5 nM
  • Dissolves in DMSO (Dimethyl sulfoxide)
  • Best biochemically characterized LIMK inhibitor (profiled against 62 kinases for specificity in vitro)
  • Best biologically characterized LIMK inhibitor (tested for specificity in vivo in T cells)
  • Profiled against multiple viruses

Table 1. Profiling R10015 against 62 kinases (DMSO control, 100% activity)

Kinase % Activity Kinase % Activity
ABL1 89.2 LIMK1 6.4
AKT2 73.1 LRRK2 (G2019S) 6.7
ALK 95.5 MEK1 75.6
AMPK(A1/B1/G1) 76.6 MLCK/MYLK 83.3
Aurora A 60.3 MRCKa/CDC42BPA 96.5
BRAF 91.9 mTOR/FRAP1 100.0
BTK 90.2 NEK1 67.2
CAMK2b 90.8 P38a/MAPK14 98.6
CDK2/cyclin A 82.7 p70S6K/RPS6KB1 10.0
CDK5/p35 85.5 PAK2 98.2
CK1d 85.0 PDGFRb 78.6
CK2a 94.2 PIM2 97.6
c-Kit 81.3 PKA 24.0
c-MET 82.1 PKCa 47.5
c-Src 92.4 PKN1/PRK1 58.6
DAPK1 87.8 PLK2 103.2
EGFR 93.2 RET 46.5
EPHA3 91.1 ROCK1 42.3
ERK2/MAPK1 87.2 ROCK2 29.5
FAK/PTK2 81.5 RSK1 47.2
FGFR1 86.5 SGK1 61.0
FLT3 32.2 SLK/STK2 108.0
GSK3b 67.8 STK16 48.6
IGF1R 90.0 STK33 56.5
IKKa/CHUK 90.5 TAK1 82.4
IKKb/IKBKB 85.1 TAOK1 54.0
IKKe/IKBKE 84.1 TBK1 82.0
JAK3 61.1 TESK1 90.2
JNK1 95.2 TLK1 98.5
JNK3 85.7 WEE1 97.2
KDR/VEGFR2 87.2 ZIPK/DAPK3 98.5
  • The % data were the average of two determinations.
  • R10015 used was 1 µM.
  • The ATP concentration used was 10 µM.
  • Staurosporine was used as the control for assay validation.

References

Endo, M., Ohashi, K., and Mizuno, K. (2007). LIM kinase and slingshot are critical for neurite extension. J Biol Chem 282, 13692-13702.
Foletta, V.C., Moussi, N., Sarmiere, P.D., Bamburg, J.R., and Bernard, O. (2004). LIM kinase 1, a key regulator of actin dynamics, is widely expressed in embryonic and adult tissues. Exp Cell Res 294, 392-405.
Liu, G., Xiang, Y., Guo, C., Pei, Y., Wang, Y., and Kitazato, K. (2014). Cofilin-1 is involved in regulation of actin reorganization during influenza A virus assembly and budding. Biochem Biophys Res Commun 453, 821-825.
Manetti, F. (2012). Recent Findings Confirm LIM Domain Kinases as Emerging Target Candidates for Cancer Therapy. Curr Cancer Drug Targets 12, 543-560.
Nishimura, Y., Yoshioka, K., Bernard, O., Bereczky, B., and Itoh, K. (2006). A role of LIM kinase 1/cofilin pathway in regulating endocytic trafficking of EGF receptor in human breast cancer cells. Histochem Cell Biol 126, 627-638.
Vorster, P.J., Guo, J., Yoder, A., Wang, W., Zheng, Y., Xu, X., Yu, D., Spear, M., and Wu, Y. (2011). LIM kinase 1 modulates cortical actin and CXCR4 cycling and is activated by HIV-1 to initiate viral infection. J Biol Chem 286, 12554-12564.
Yang, N., Higuchi, O., Ohashi, K., Nagata, K., Wada, A., Kangawa, K., Nishida, E., and Mizuno, K. (1998). Cofilin phosphorylation by LIM-kinase 1 and its role in Rac-mediated actin reorganization. Nature 393, 809-812.
Yi, F., Guo, J., Dabbagh, D., Spear, M., He, S., Kehn-Hall, K., Fontenot, J., Yin, Y., Bibian, M., Park, C.M., et al. (2017). Discovery of Novel Small Molecule Inhibitors of LIM Domain Kinase for Inhibiting HIV-1. J Virol. 91:e02418-16.
Yoshioka, K., Foletta, V., Bernard, O., and Itoh, K. (2003). A role for LIM kinase in cancer invasion. Proc Natl Acad Sci U S A 100, 7247-7252.

 

R10015 may be obtained with a Limited Use License. R10015 is intended for research use only and is not for diagnostic or therapeutic purposes or uses in humans.