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Fragment Hit Validation
Validation of Hits from a TINS screen is a critical step in the drug discovery process.
We offer three different approaches:
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One powerful method of validating hits is to perform a competition binding assay using TINS. Here known ligands (inhibitors) are used to validate the binding of fragments. When performing a competition study in TINS each hit is reassayed as a singleton, thereby providing confirmation of binding, and subsequently in the presence of saturating amounts of a competitor, providing binding site information. |
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If it is possible to incorporate stable, NMR visible isotopes into the target, for example by bacterial expression, than an entire process of hit validation and potentially structural biology becomes available. The use of isotopic labelling allows efficient acquisition of NMR spectra of the protein that are extremely sensitive to ligand binding (Protein Observed NMR - HSQC). This orthogonal technique can be used to validate and quantify binding as well as to define a low resolution binding site on the target.
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Surface Plasmon Resonance (SPR) is a powerful, industry accepted ligand discovery technology that is orthogonal to NMR. In particular, TINS and SPR are highly complementary since both require target immobilization, while TINS provides extraordinary sensitivity to weak interactions and SPR is highly quantitative. We have recently shown that TINS screening followed by SPR characterization of fragment hits (Kobayashi et al, J. Biomol. Screening, 2010, 15, p.978) forms a powerful work flow for a fragment-based discovery program. ZoBio has a unique, highly integrated platform of TINS and and a Biacore T200 instrument that allows us to further leverage our unparalleled expertise in protein immobilization for the benefit of our customers.
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