Gandotinib

Protein target name:

JAK2

Mechanism of action:

Inhibitor, ATP-CompetitiveMutant Specific

Recommended Concentration for use in cells:

~~up to 1 uM~~

Reviewer suggested Concentration for use in cells:

up to 200 nM

Probe Availability

MedChemExpress

Selleck Chemicals

Probe Information

Target and Probe Information

Target Details

Target class:

Protein kinase

Subclass:

HUGO ID:

HGNC:6192

Entrez gene ID:

3717

UNIPROT ID:

O60674

Target alias:

Janus kinase 2, JTK10

Probe Details

Chemical Probes Portal ID:

CPP1211

Probe alias:

LY2784544

PubChem CID:

46213929

ChEMBL ID:

CHEMBL2107823

SMILES string:

CC1=NNC(NC2=NN3C(C(CN4CCOCC4)=C2)=NC(C)=C3CC5=CC=C(C=C5F)Cl)=C1

InChi Key:

SQSZANZGUXWJEA-UHFFFAOYSA-N

Orthogonal probe:

XL019AZ-960BMS-911543

Validation in Cells and Model Organisms

In vitro validation

On target activity

Potency:

IC50, Ki

2.52 nM JAK2, 19.8 nM JAK1, 48.0 nM JAK3, 0.203 nM JAK2 WT, 0.246 nM JAK2 V617F

Potency assay:

JAK LanthaScreen Kinase assay

Structure-activity relationship:

Off target activity

Potency (off target):

IC50

Off target protein and potency:

FLT3 4 nM

Potency assay (off target):

Cerep

Probe Selectivity in Vitro:

Assessed against a CEREP kinase panel (total of 99 kinases). Of these, 20 kinases were inhibited by LY2784544 with IC50 (three concentration points) <0.3 μM FLT3 0.004, FLT4 0.025, FGFR2, 0.032, TYK2 0.044, TRKB 0.095, FGFR3 0.106, KDR 0.109, ALK 0.138, MUSK 0.147, AURKA 0.168, MAP3K9 0.299.

In cell validation

On target activity in cells

Potency in cells:

IC50

45 nM (T), 942 nM (N), 191 nM (A), 20 nM (S) Mutant, 55 nM (P) Mutant

Potency assay (cells):

Cellomics detection assay (TF-1 JAK2 cell-based assay (T), NK-92 JAK3/JAK1 heterodimer cell-based assay (N)), TEL-JAK cell-based AlphaScreen assays (A), pSTAT5 AlphaScreen assay (S), Cell proliferation assay (P)

In vivo validation

Organism:

Mouse, Rat

Dose:

30, 60, 100 mg/Kg

Route of delivery:

Oral

Primary Reference

Reference (doi):

10.1038/bcj.2013.6

Reference (PMID):

23584399

Endorsed probe

The Chemical Probes Portal endorses this compound as chemical probe for use in cells and in model organisms.

SAB Rating

Rating for use in Cells

3 review(s)

Rating for use in Model Organisms

3 review(s)

SAB Members

SAB Comments

Concentration notes: Data suggests it could be dosed up to 20 uM in cellular assays.

BID dosing is required to maintain reasonable drug levels in vivo models. PK parameters are not discussed; dose selection in alternate models would require additional work.

Aug 4 2021 - 9:02pm

Activation of the JAK-STAT signalling pathway has been associated with cancer progression across multiple tumour types either as a tumour intrinsic driver of cancer cell growth and spread or as a modulator of immunosurveillance (10.3390/cancers12071971). Constitutive activation of JAK-STAT signalling can arise through a number of different mechanisms, including the elevated expression of cytokines and/or activating mutations in receptors upstream of JAK-STAT signalling. Activating mutations of JAK2 are commonly found in a group of hematologic malignancies known as myeloproliferative neoplasms (MPNs). An acquired point mutation (JAK2 1849G-T) results in a missense substitution of phenylalanine for valine at amino acid position 617 (V617F) within the autoinhibitory pseudokinase domain of JAK2. This substitution results in a gain-of-function activation mutant of JAK2 (JAK2V617F) with subsequent phosphorylation of STAT5 (10.1038/bcj.2013.6).

Biochemical screening assays demonstrated the small molecule compound LY2784544 exhibits 8- and 16-fold selectivity for JAK2 (IC50= 0.0025) over JAK1(IC50=0.0198) and JAK3 (IC50=0.048), respectively. Further assessment of activity against a CEREP kinase panel (total of 99 kinases) demonstrated 20 kinases were inhibited by LY2784544 with IC50 of <0.3 M. with most potent activity upon FLT3, FLT4, FGFR2 with IC50: 0.004, 0.025 and 0.032 respectively (10.1038/bcj.2013.6). In Ba/F3 cellular assays LY2784544 effectively inhibited JAK2V617F-driven signalling and cell proliferation in Ba/F3 cells (IC50; 0.02 and 0.05 M, respectively). In comparison, LY2784544 was much less potent at inhibiting
interleukin-3-stimulated wild-type JAK2-mediated signalling and cell proliferation (IC50:1.183 and 1.309 M, respectively). In vivo, LY2784544 effectively inhibited STAT5 phosphorylation in Ba/F3-JAK2V617F-GFP (green fluorescent protein) ascitic tumor cells
(TED50.12.7 mg/kg) and significantly reduced Ba/F3-JAK2V617F-GFP tumor burden in the JAK2V617F-induced MPN model (TED50.13.7 mg/kg, twice daily).

LY2784544 is therefore a suitable chemical probe for exploring the activity of the JAK2V617F mutant kinase. LY2784544 may also represent a useful orthogonal probe for exploring the activity of wild type JAK2, however further validation to rule out an activity against other kinases particularly FLT3 and FLT4 would need to be determined.

Cellular concentration notes: I would recommend this is used as a chemical probe specifically for the JAK2V617F mutant protein at a lower concentration of up to 200 nM to avoid off-target activities

Aug 4 2021 - 9:08pm

The cpd should also be tested in vitro and in vivo against FLT4 and FLT3 driven tumors and Baf3 cells (may be testing of this cpd against a large panel of Baf3 cell (ACD-Baf3) would be more beneficial than the CEREP biochemical panel.

Aug 26 2021 - 9:24am

Gandotinib

Probe Availability

Probe Information

Target Details

Janus kinase 2, JTK10

Probe Details

In vitro validation

In cell validation

In vivo validation

Primary Reference

Supporting Organizations

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Gandotinib

Probe Availability

Probe Information

Target Details

Janus kinase 2, JTK10

Probe Details

In vitro validation

In cell validation

In vivo validation

Primary Reference

Supporting Organizations

Interested in supporting the portal?