Applications of Microscale Thermophoresis to Small Molecule Drug Discovery and Optimization in the Molecular Targets Program

Authors

• RS Shah

Abstract

Microscale Thermophoresis (MST) is an innovative biophysical tool that can be used to measure the apparent binding affinity of two biomolecules. It works by measuring the diffusion of a fluorescently tagged macromolecule in response to a temperature gradient and can discern changes in the apparent size and shape of said macromolecule through changes in its diffusion rate. This makes MST useful for measuring the interactions of various substances to a target fluorescent protein, and it can be utilized to determine the affinity of potential drug compounds to target proteins of interest. This poster aims to demonstrate the various ways that MST has been utilized by the Molecular Targets Program (MTP) at the NCI to screen and characterize small molecules as potential cancer therapeutics. the MTP works by testing the affinity of target cancer-related proteins or biomolecules against diverse libraries of natural product small molecules in a high throughput screening (HTS) format. The MTP has demonstrated the ability of MST to be used to verify and further characterize HTS hits. Using MST, we have been able to verify the legitimacy of these hits, as well as filter out nonspecific binder, self-aggregators, and assay artifacts from our primary hits. Additionally, we have used MST to classify hit small molecules by their affinity to target proteins and used the exact change in diffusion rate to understand the mode of binding of some of these compounds. This approach has allowed us to find drug candidates for various proteins, such as PKA, Calmodulin, and RBD.

Scientific Focus Area: Molecular Biology and Biochemistry

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