Alk5 inhibitor | BI-4659
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Highlights
BI-4659 is a potent inhibitor of Alk5 (TGFßR1) with an IC50 value of 19 nM and shows selectivity against a broad panel of other kinases (see Selectivity section). BI-4659 blocks the phosphorylation of Smad2 and Smad3 in HaCaT cells with an EC50 of 185 nM. Together with the offered negative control BI-4101, BI-4659 is a suitable tool to test biological hypotheses in vitro.1 (BI-4659 = compound 47i, BI-4104 = compound 48 in reference 1).
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Target information
Transforming growth factor β (TGFβ) is a pluripotent cytokine involved in the regulation of various biological processes such as cell proliferation, differentiation, migration, adhesion, apoptosis, and epithelial-to-mesenchymal transition (EMT). Therapeutic approaches to inhibit its signaling by targeting TGFβ receptor I (TGFβRI/Alk5) are discussed for the treatment of diseases such as idiopathic pulmonary fibrosis (IPF) and cancer.2,3,4
Figure 3: Crystal structure of Alk5 complexed with a close analog of BI-4659 (PDB code 2X7O).1
In vitro Activity
|
Probe name / Negative control |
BI-4659 |
BI-4101 |
|
MW [Da] |
440.5 |
418.5 |
|
Inhibition of Alk5 (Kinase Glow assay, IC50) [nM]a |
19 |
>50000 |
|
Inhibition of phosphorylation of Smad2 and Smad3 in HaCaT cells (EC50) [nM] |
185 |
n.d. |
a for detailed assay conditions see reference 1
In vitro DMPK and CMC parameters
| Probe name / negative Control | BI-4659 | BI-4101 | ||||
|
Solubility @ pH 6.8 [µg/ml] |
2 | n.d. | ||||
|
Solubility @ pH 4 [µg/ml] |
84 | n.d. | ||||
| Solubility @ pH 2 [µg/ml] | 71 | n.d. | ||||
| CACO permeability @pH7.4 [*10-6 cm/s] | 8 | n.d. | ||||
| CACO efflux ratio | 6 | n.d. | ||||
| Microsomal stability (human/mouse/rat) [% QH] | 26 | 23 | <22 | n.d. | ||
| Hepatocyte stability (human/mouse/rat) [% QH] | n.d. | n.d. | ||||
| Plasma protein binding (human/mouse/rat) [%] | n.d. | n.d. | ||||
| hERG [inh. % @ 10 µM] | 38.5 | n.d. | ||||
| hERG IC50 [µM] | >10 | n.d. | ||||
| CYP 3A4 (IC50) [µM] | >50 | n.d. | ||||
| CYP 2C8 (IC50) [µM] | >50 | n.d. | ||||
| CYP 2C9 (IC50) [µM] | >50 | n.d. | ||||
| CYP 2C19 (IC50) [µM] | >50 | n.d. | ||||
| CYP 2D6 (IC50) [µM] | >50 | n.d. | ||||
In vivo DMPK parameters
BI-4659 showed very high clearance in rat and therefore is not suited for in vivo studies
| BI-4659 | Rat |
|
Clearance [%QH] |
123 |
|
Mean residence [l/kg]a |
1.7 |
|
tmax [h]b |
3.3 |
|
Cmax [nM]b |
27.7 |
|
F [%]b |
50 |
|
Vss [l/kg]a |
9 |
ai.v. 4.5 mg/kg
bp.o. 45 mg/kg
Negative control
BI-4101 shows no inhibition of Alk5 in the Kinase Glow assay (IC50 > 50 µM) and therefore is a suitable negative control for in vitro experiments.1 (BI-4101 = Compound 48 in reference 1.)
Figure 4: BI-4101 which serves as a negative control
Selectivity
BI-4659 sows good selectivity in kinase panels. The compound shows no inhibition of 218/232 kinases tested at 2 µM. Cross-reactive kinases (%inhibition data at 0.2 µM): ABL1 (67), BLK (85), CSF1R (FMS) (82), FGR (91), FLT3 (66), FYN (74), LCK (100), LYN A (79), LYN (78), MAP4K5 (KHS1) (77), MELK (80), NTRK3 (TRKC) (78), RET (79), SNF1LK2 (96), YES1 (90).
|
BI-4659 |
Selectivity data available |
|
Cerep® |
No |
|
Panlabs® |
No |
|
Invitrogen® |
Yes |
|
DiscoverX® |
Yes |
|
Dundee |
Yes |
Co-crystal structure of the BI probe compound and the target protein
No in-house structure is available for BI-4659, but for related compound (PDB code 2x7o).
reference molecules
SB-505124, SB-525334, GK6604, SD-208, LY-2157299, EW-7197, GW788388 and others. For a review on Alk5 kinase inhibitors in oncology see reference 5.
Summary
BI-4659 is a potent and selective inhibitor of Alk5 (IC50 = 19 nM) which effectively blocks cellular phosphorylation of Smad2/Smad3 in HaCaT cells (EC50 = 185 nM). We also offer BI-4101 as negative control. BI-4659 (5 mg) can be ordered free of charge together with BI-4101.1,6
Supplementary data
References
Design, Synthesis, and Evaluation of Indolinones as Inhibitors of the Transforming Growth Factor β Receptor I (TGFβRI)
Gerald J. Roth, Armin Heckel, Trixi Brandl, Matthias Grauert, Stefan Hoerer, Joerg T. Kley, Gisela Schnapp, Patrick Baum, Detlev Mennerich, Andreas Schnapp, and John E. Park
J. Med. Chem. 2010, 53, 7287-7295.
Inhibition of ALK5 as a new approach to treat liver fibrotic diseases
Anne-Charlotte de Gouville and Stephane Hue
Drug News Perspect 2006, 19, 85-90.
Inhibition of TGF- Signaling for the Treatment of Tumor Metastasis and Fibrotic Diseases
Ota Fuchs
Current Signal Transduction Therapy 2011, 6, 29-43.
Smad-dependent and Smad-independent pathways in TGF-beta family signaling
Rik Derynck and Ying E. Zhang
Nature 2003, 425, 577-584.
TGF-Beta Type I Receptor (Alk5) Kinase Inhibitors in Oncology
Leona E. Ling, Wen-Cherng Lee
Curr. Pharm. Biotechnol. 2011, 12, 2190-2202.
Phenocopy – A Strategy to Qualify Chemical Compounds during Hit-to-Lead and/or Lead Optimization
Patrick Baum , Ramona Schmid , Carina Ittrich, Werner Rust, Katrin Fundel-Clemens, Susanne Siewert, Martin Baur, Lisa Mara, Lore Gruenbaum, Armin Heckel, Roland Eils, Roland E. Kontermann, Gerald J. Roth, Florian Gantner, Andreas Schnapp, John E. Park, Andreas Weith, Karsten Quast, Detlev Mennerich
PLoS One 2010, 5, e14272.
