in vitro PHARMACOLOGY 2011 CATALOG - Cerep

107
Cerep
services
Kinases
Receptors
Cerep’s panel of kinases has increased from 205 biochemical assays as of January,1, 2009 to 271 (including 16 cellular kinase assays)
as of January 1, 2011 allowing to provide a wider coverage of all major families in the human kinome (Manning G. et al. (2002)
Science, 298: 1912-1934).
Kinases sub-families 1 P.
Protein-tyrosine kinases 108
RTK Receptor Tyrosine Kinases 108
CTK Cytoplasmic Tyrosine Kinases 118
Protein-serine/threonine kinases 124
CMGC Cyclin-dependent-(CDK), Mitogen-activated-(MAPK), Glycogen-synthase-(GSK) and CDK-like kinases 124
CaMK Ca 2+ /calmodulin-dependent protein kinases 130
AGC protein kinases A, G and C cyclic nucleotide regulated and phospholipid-regulated kinases and ribosomal S6 kinases 137
CK1 Casein Kinases 1 144
STE homologs of yeast sterile kinases 144
TKL Tyrosine Kinase-Like 149
other kinases 151
atypical kinases 156
1 Cohen P. (2002), Nature Reviews/Drug Discovery, 1: 309-315
Cerep kinase assays
Ion
channels
Transporters
Kinases
Epigenetic &
DNA-related
enzymes
❚ Biochemical kinase assays
Majority of our biochemical kinase assays use activated kinases. They are usually full length kinase or cytoplasmic domain of RTK. Assays
are designed to be as close as possible to ATP and substrate Km.
The technology used to measure substrate phosphorylation is TR-FRET (HTRF ® or LANCE ® ). A few assays are cascade of activation.
NOTE: A majority of our kinase assays have been converted from HTRF ® to LANCEUltra ® technology. Both technologies are TR-FRET
with the difference in europium labelling: europium cryptate for HTRF ® and europium chelate for LANCEUltra ® .
Assays are converted following a standard procedure consisting of Time course experiment, Km determination and pharmacology
characterization of the enzyme by inhibiting its activity with known inhibitors. IC 50 values obtained for known inhibitors are compared
to literature and to those obtained using HTRF ® technology. If all values are in agreement with the previous ones, LANCE ® technology
is validated for the kinase of interest. This conversion allows us to standardize and automate our kinase assays providing shorter
turnaround times for both profiling and screening experiments.
❚ cellular Kinase assays
In order to complement our existing biochemical assay platform, we have implemented cellular kinase assays. These assays allow to
confirm inhibitors activity in a relevant cellular background and profile their selectivity against multiple signaling pathways.
- Cellular kinase phosphorylation assays
Cellular kinase phosphorylation assays are developed using AlphaScreen ® Surefire ® assay kits. The assays are optimized for directly
measuring kinase activation following treatment of cells with activators of signaling pathways.
- Cellular tyrosine kinase receptor activity assays
We have demonstrated that the impedance-based technology can be used to monitor the activity of the main tyrosine kinase receptor
families. These label-free assays allow the identification of inhibitors targeting either the ligand binding domain or the kinase domain.
These assays can be developed in virtually any cancer cell lines or primary cells. As an example, we have developed and validated
the EGFR cellular assay in the following cancer cell lines: A431, HELA and MDA-MB-231.
❚ binding Kinase assays
To expand our existing kinase services, we offer binding kinase assays to identify type II inhibitors on unphosphorylated kinases, to
compare compounds activity on unphosphorylated and phophorylated kinases and to discriminate between ATP and non-ATP competitive
inhibitors. In addition, we are now using ADP readout to develop kinase assays on intractable substrates (for which no antibody exist)
which allows us to profile kinase activity on different natural substrates in order to identify substrate selective inhibitors. These assays are
displayed as part of our custom research.
For further information, please contact us at customresearch@cerep.com.
Other
enzymes
Specialized
cellular
assays
Standard
profiles
Testing
conditions
Ordering
information
Assay list
& index