Abstract

Fragment based screening has been a successful hit discovery approach for reversible inhibitors in providing better chemical space coverage and higher probability of binding due to lower molecular weight complexity. One of the challenges of fragment based screening is the requirement of sensitive biophysical detection methods due to the weak binding affinity of fragment hits. In addition, in the absence of crystallography, rationalization of which functional groups within the fragment are driving target binding is often unknown. The screening of covalent fragments looks to address these limitations, given covalent binders are easy to detect by mass spectrometry and the dominant interaction is unambiguous.

Domainex has compiled a cysteine targeted covalent fragment screening library and established a rapid mass spectrometry based screening platform. This platform has been used to identify covalent fragment hits against a protein of interest, which to our knowledge, has not been prosecuted by a covalent fragment screen.

Due to a low signal to noise ratio seen in the complex mixtures of the wild type, a truncated version of the target protein was selected for this study. Through an optimization process, conditions were selected to allow identification of binders from pools of 5 fragments using a Waters G2-XS QToF.

10 strong binders were identified from our acrylamide covalent fragment library and these fall into three unique clusters of chemotypes, which present potential new start points for further exploration.