SPR-based fragment screening with neurotensin receptor 1 generates novel small molecule ligands

Abstract

The neurotensin receptor 1 represents an important drug target involved in various diseases of the central nervous system. So far, the full exploitation of potential therapeutic activities has been compromised by the lack of compounds with favorable physicochemical and pharmacokinetic properties which efficiently penetrate the blood-brain barrier. Recent progress in the generation of stabilized variants of solubilized neurotensin receptor 1 and its subsequent purification and successful structure determination presents a solid starting point to apply the approach of fragment-based screening to extend the chemical space of known neurotensin receptor 1 ligands. In this report, surface plasmon resonance was used as primary method to screen 6369 compounds. Thereby 44 hits were identified and confirmed in competition as well as dose-response experiments. Furthermore, 4 out of 8 selected hits were validated using nuclear magnetic resonance spectroscopy as orthogonal biophysical method. Computational analysis of the compound structures, taking the known crystal structure of the endogenous peptide agonist into consideration, gave insight into the potential fragment-binding location and interactions and inspires chemistry efforts for further exploration of the fragments.

Discovery of fragments that target key interactions in the signal recognition particle (SRP) as potential leads for a new class of antibiotics

“…Fragments from primary FBDD screens typically bind to their targets in the micromolar to millimolar range [ 26 ]. For example, in a study of neurotensin I, the compounds identified had K D values between 18 and 400 μM [ 27 ].…”

Applied Biophysical Methods in Fragment-Based Drug Discovery

“…Of these variables, we have some control over the protein immobilization level; however, the NSB problem is more readily attenuated by keeping the compound concentration low. A survey of published SPR FBS conditions 42,[52][53][54] reveals that many users are opting to screen at fragment concentrations of <300 µM in order to combat the problem of NSB. Although it might attenuate NSB, low-concentration SPR fragment screening limits the fragment binding affinity range that is detectable.…”

“…More detail on the configuration and analysis of SPR competition experiments can be found in the following references. [54][55][56]…”

What is the current value of fluorescence polarization assays in small molecule screening?

“…Thus, improved screening assays are an important component of the discovery of novel compounds. Alternatively, SPR has been used to screen 6,369 compounds as potential GPCR ligands (neurotensin receptor 1) [8].…”