2-MT 63

Reagent Authentication Certificate:

Chemical Probes Portal 2-MT 63.pdf

Protein target name:

TEK

Mechanism of action:

Inhibitor

Recommended Concentration for use in cells:

10-1,000 nM

Probe Information

Target and Probe Information

Target Details

Target class:

Protein kinase

Subclass:

HUGO ID:

HGNC:11724

Entrez gene ID:

7010

UNIPROT ID:

Q02763

Target alias:

TEK receptor tyrosine kinase, CD202B, GLC3E, TIE-2, TIE2, VMCM, VMCM1

Probe Details

Chemical Probes Portal ID:

CPP1307

Probe alias:

2-(Methylamino)triazine 63

PubChem CID:

16109001

ChEMBL ID:

CHEMBL373882

SMILES string:

CNC1=NC=NC(=N1)C2=C(N=CC=C2)OC3=CC(=C(C=C3)F)C(=O)NC4=C(C=CC(=C4)C(F)(F)F)N(C)CCCN(C)C

InChi Key:

HHKGVQGXRGQBBA-UHFFFAOYSA-N

Orthogonal probe:

BAY-826

Physico-chemical properties

NMR:

1H NMR (300 MHz, two inseparable isomers present in DMSO-d6) isomer 1, ä 9.97 (d, J ) 6.9 Hz, 1H), 8.59 (s, 1H), 8.54 (m, 1H), 8.39 (d, J ) 4 Hz, 1H), 8.27 (m, 2H), 8.14 (m, 1H), 7.62 (m, 1H), 7.49 (m, 4H), 7.36 (m, 1H), 2.99 (m, 2H), 2.79 (d, J ) 3.3 Hz, 3H), 2.68 (s, 3H), 2.21 (m, 2H), 2.03 (s, 6H), 1.57 (m, 2H); isomer 2, ä 9.97 (d, J ) 6.9 Hz, 1H), 8.69 (s, 1H), 8.54 (m, 1H), 8.27 (m, 3H), 8.14 (m, 1H), 7.62 (m, 1H), 7.49 (m, 4H), 7.36 (m, 1H), 2.99 (m, 2H), 2.86 (d, J ) 3.3 Hz, 3H), 2.68 (s, 3H), 2.21 (m, 2H), 2.03 (s, 6H), 1.57 (m, 2H).

Validation in Cells and Model Organisms

In vitro validation

On target activity

Potency:

IC50

30 nM

Potency assay:

Homogenous time-resolved fluorescence assays detecting enzyme activity using peptide substrates.

Structure-activity relationship:

Yes, see J Med Chem paper.

Off target activity

Potency (off target):

IC50

Off target protein and potency:

VEGFR: 7,800 nM, IGFR1: 927 nM

Potency assay (off target):

In HTRF assays (as above) on 28 kinases, the compound was at least 30-fold selective for TIE2. Kinases included: TIE2, VEGFR, PDGFR, cFMS, RET, TRKA, cMET, EGFR, INSR, IGFR-1, EPHB4, SYK, ZAP70, JAK3, CSK, LCK, LYN, FGR, ABL, BTK, ITK, FES, p38alpha, CDK5, AURKA, PKBalpha).

Probe Selectivity in Vitro:

Not done.

In cell validation

On target activity in cells

Potency in cells:

IC50

62 nM

Potency assay (cells):

In EA.hy926 cells, the compound inhibited Ang-1-induces TIE2 autophosphorylation in a plate-based immunassay.

Target engagement assay (cells):

Indirect, inhibition of substrate phosphorylation

In vivo validation

Organism:

Sprague-Dawley male rat

Dose:

1 mg/kg (IV), 10 mg/kg (oral)

Route of delivery:

OralIntravenous

Plasma half life:

1.6 h

Systemic clearance:

1,300 mL/h/kg

Fraction of probe bound to plasma protein:

0.93

Cmax:

520 ng/mL (oral)

Tmax:

2 h (oral)

Organ of Interest

Organ (O):

mouse lung

Target engagement assay:

Indirect, inhibition of Ang-1-induced Tie2 autophosphorylation assayed by western blot

Primary Reference

Reference (doi):

10.1021/jm061107l

Reference (PMID):

17253678

SAB Rating

Rating for use in Cells

3 review(s)

Rating for use in Model Organisms

3 review(s)

SAB Members

SAB Comments

2-MT 63 represents a novel inhibitor of the TIE2 receptor. There are multiple kinase inhibitors that have validated activity versus the TIE2 receptor including the advanced clinical agents, Sitravatinib, Glesatinib and Rebastinib. This agent is a useful tool given its unique polypharmacological profile (Rebastinib, for instance, also inhibits VEGFR2, BCR-ABL, LYN and ABL1 kinases while 2-MT 63 does not.) There is a lack of data from cell culture experiments involving this agent in the original publication. It was also shown to inhibit TIE2 autophosphorylation in vivo. However, this outcome was limited - with activity occurring at a high dose (100 mg/kg) and only transiently (positive at 3 hours but the effect was lost by 8 hours).

Mar 18 2017 - 8:23pm

2-MT 63 is a novel TIE-2 kinase inhibitor that demonstrates unique and improved kinase selectivity compared to other inhibitors known to interact with TIE-2. The developmental focus centered on in vivo assessment, and therefore, substantial work in cells has not yet been demonstrated, though the providers adequately bridge enzymatic inhibition with demonstrated potency in a cellular autophosphorylation assay, which translates to in vivo inhibition of TIE-2 autophosphorylation. In vivo efficacy was observed by oral administration in rats at a high dose (100 mg/kg) for a short duration of 3 h. Nevertheless, 2-MT 63 represents one of the best probes one could use to further study the role of TIE-2 in angiogenesis in a selective manner.

Mar 30 2017 - 12:01pm

2-MT 63 has a good kinase selectivity profile in enzyme assays with >30-fold selectivity (typically much greater) against the tested kinases. Importantly, the profile includes selectivity against other targets of relevance for angiogenesis (KDR, PDGFR, EPHB4). Cellular selectiivity data are not published, but the probe does have a comparable level of cellular activity (IC50=62 nM for cellular inhibition of TIE-2 phosphorylation) to enzyme inhibition (IC50=30 nM). In-vivo activity from an oral dose in mouse, albeit at a high dose of 100 mg/kg, is achieved, with inhibition of ANG-1-stimulated TIE-2 phosphorylation observed at 3 h. A time course of in-vivo activity in mouse is not presented but given the short in-vivo half life in rat (t1/2=1.6 h) a short duration of activity would not be surprising. It should be noted that the pharmacokinetic (PK) profile in mouse has not been reported (only PK data for rat are reported), despite the in-vivo PD model being in mouse.

May 15 2017 - 11:31am

2-MT 63

Probe Information

Target Details

TEK receptor tyrosine kinase, CD202B, GLC3E, TIE-2, TIE2, VMCM, VMCM1

Probe Details

Physico-chemical properties

In vitro validation

In cell validation

In vivo validation

Primary Reference

Supporting Organizations

Interested in supporting the portal?

Search form

2-MT 63

Probe Information

Target Details

TEK receptor tyrosine kinase, CD202B, GLC3E, TIE-2, TIE2, VMCM, VMCM1

Probe Details

Physico-chemical properties

In vitro validation

In cell validation

In vivo validation

Primary Reference

Supporting Organizations

Interested in supporting the portal?