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eCF309

ATP-competitive inhibitor of MTOR

Structure

Information

  • MTOR
  • ATP-competitive inhibitor

In Vitro Validations

Uniprot ID: P42345
Target Class: Protein kinase
Target SubClass: Atypical
Potency: IC50
Potency Value: 15 nM
Potency Assay: MTOR kinase activity relative to DMSO was determined by measuring 33P incorporation on its substrate (poly[Glu,Tyr]4:1) by Reaction Biology Corp., USA. eCF309 was tested at 10 different doses with 3-fold serial dilution starting from 10 μM.
PDB ID for probe-target interaction (3D structure): --
Structure-activity relationship: Analysis of the kinase inhibition profile of eCF309 and several analogues indicated that the presence of oxygen atoms in the alkyl group at the position N1 of the pyrazolopyrimidine ring results in increased mTOR selectivity.
Target aliases:
Serine/threonine-protein kinase mTOR, RAPT1, RAFT1 ...

In Cell Validations

In Vivo Data

No in Vivo Validations

Off-Target Selectivity Assesments

Potency end-point : IC50 DDR1/2 2,110 nM, DNA-PK 320 nM, PI3Kα 981 nM, PI3Kβ >10,000 nM, PI3Kγ 1,340 nM, PI3Kδ 1,840 nM
Probe Selectivity in Vitro:
Kinome profiling of eCF309 against 375 wild type and mutant kinases was performed at 10 μM. mTOR was the only protein inhibited >99%.The activity of five kinases was inhibited between 65-90%: DDR1, DNA-PK, PI3Kγ and PI3Kα(E545K). S-score (35) = 0.01
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SERP ratings and comments

SERP Ratings

In Cell Rating

SERP Comments:

eCF309 appears sufficiently potent in cell-free systems (15 nM). Strong evidence is provided with regards to kinase selectivity in biochemical assays. Solid kinase profiling was performed in the original manuscript. However, users should note eCF309 was not tested versus non-kinase targets. Users should consider and potentially evaluate the effects of eCF309 on other kinases (particularly DDR1, DNA-PK, PI3K) in their experimental system in addition to examining mTOR-related effects. Users should note the cell-based readouts (phosphorylation, cell proliferation/viability, cell cycle analysis) are indirect measures of target engagement. Additional studies to confirm direct and specific target engagement in cells at the concentrations commonly tested would enhance confidence in its purported cellular target and mechanism of action. This compound may be useful in model organisms, but no data is available at this point to offer a recommendation in this regard.

(last updated: 21 Dec 2016 )

SERP Ratings

In Cell Rating

SERP Comments:

eCF309 is a potent inhibitor of mTOR signaling. The compound is potent in a target-based assay and in cellular systems. eCF309 has been profiled against 375 wild-type and mutant kinases and has an S(35) of 0.01 when tested at 10 micromolar. Other targets that are inhibited (but more weakly) are PI3K gamma (85%), PI3Ka-E545K mutant (65%), DNA-PK (90%), and DDR1 (77%). The compound contains a diethyl acetal, which is not commonly seen in optimized, medicinal-chemistry molecules. The authors have done a good job addressing potential issues around this functional group. For example, it appears stable in the cellular context, and the aldehyde analogue is inactive. This compound is a useful addition to the tool kit for studying mTOR signaling.

(last updated: 20 Jan 2017 )

SERP Ratings

In Cell Rating

SERP Comments:

eCF309 has been thoroughly profiled against other kinases and moderate activity was detected for only a few of them. Potential in vitro affinities against targets that are phylogenetically more distantly related has not been addressed. Among them, I would perhaps suggest testing against the Adenosine Receptor A1. Other than that, it seems safe to use eCF309 as a selective chemical probe of mTOR signaling, if it is used at concentrations below 1 uM (preferably, below 100 nM).

(last updated: 17 Mar 2017 )