Friday, June 3, 2011

Automated Profiling Applications using Simple, Robust Instrumentation

Lead compounds, identified during hit and hit to lead screening, are normally profiled to determine selectivity for the drug target. Compounds demonstrating inhibitory effects to other non-target enzymes may then be sent back for modification or be removed from further testing. The procedure typically involves performing the test assay with a dilution series of the compound in order to ascertain the IC50 or EC50 potency. Therefore, a separate set of instrumentation is required from that used for primary screening. Liquid handling must be able to serially titrate compounds, as well as dispense multiple assay reagents to high density plates.

Here we demonstrate the ability to perform automated compound profiling applications for two separate drug target families, known for selectivity issues.

Kinases are one of the most diverse and highly studied gene families. It is becoming increasingly important for compounds to be profiled against other kinases due to the non-specificity of ATP competitive inhibitors. In this application, we used a luminescent assay chemistry which is able to detect small changes in ATP consumption from kinases having large ranges of ATPKm app values. A single set of reagents is used for all kinase/substrate combinations.


Figure 1 – Compound titration data for the PI3 Kinase specific inhibitor Wortmannin.
Figure 1 – Compound titration data for the PI3 Kinase specific inhibitor Wortmannin.
Histone deacetylases (HDACs) are an emerging drug target family. Histone hyperacetylation correlates with an open, decondensed chromatin structure and gene activation, while hypoacetylation correlates with chromatin condensation and transcriptional repression. Due to their importance in gene regulation, HDACs are becoming an increasingly accepted target for cancer therapy. Here we incorporated a green-emitting fluorescent assay which is capable of detecting the activity and inhibition of multiple HDAC enzymes using the same set of reagents.


Figure 2 – Compound titration data for various known HDAC specific inhibitors.
Figure 2 – Compound titration data for various known HDAC specific inhibitors.
The examples above demonstrate how automated compound profiling applications can be easily accomplished using BioTek instrumentation.



By, BioTek Instruments, Brad Larson, Principal Scientist

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