Reduced IC50 Values
We compared the IC50 values of 12 samples before and after the secondary recovery ultrasonic process. The IC50 values of all the test fragments left-shifted to lower concentrations; 9 out of the 12 samples showed more than two digits in percent reduction after fragment library secondary recovery, and four showed over 80% reduction.
In particular, the IC50 value of one sample shifted from 56 nM to 0.56 nM—a two logs reduction in IC50 value. This indicates that the potencies of these fragment samples were much stronger than originally thought.
The Hendrix SM100 supports a wide range of common labware: 96-, 384-, and 1,536-well plates; flat bottom, U-bottom, V-bottom labware; 2-D barcoded tubes; and scintillation vials. Because the Hendrix SM100 has 16x24 arrays of piezoelectric acoustic elements, it enables rapid fluid processing and reduces the hours-long solubilization process to minutes.
Recovering a rack of 96-fragment samples took five to ten minutes depending on sample volume and the amount of precipitates; given the small amount of additional time required, the quality improvements observed in both primary and secondary screening were significant.
Fragment libraries are screened at much higher concentrations than small molecule libraries due to their low affinity to biological targets. This makes fragment libraries more susceptible to precipitation and consequently affects the accuracy of screening results.
The data presented here shows that the Hendrix SM100 ultrasonic fluid processor can be used to recover the precipitated fragment samples back into solution, and thus increase both the sample concentration and the accuracy of screening results. After putting the fragment library through secondary recovery using the Hendrix SM100 system, we observed higher percent inhibition in primary screening and lower IC50 values in secondary screening.
By adding secondary recovery into the drug discovery process, we observed improvements in the quantity and the quality of primary screening leads as well as in the quality of secondary screening results.