Bradley martyn scannerz series#

(25,26) Nevertheless, we limited our screen to commercially available compounds to enable the broadest future use of the library. (44) Not many fragments containing these electrophiles were available for purchase from commercial vendors, likely due to the longstanding bias against covalent modifiers. Although chloroacetamides are more reactive, (51) they still show selectivity in chemical-proteomic screens. Acrylamides are represented in many rationally designed covalent drugs, such as ibrutinib and osimertinib. We constructed our electrophile-fragment library by focusing on two mild electrophilic “warheads”: acrylamides and chloroacetamides.

This is because covalent binders are easy to detect by mass spectrometry, because the dominant interaction is unambiguous (namely, the covalent bond), which simplifies the design of follow-up series, and because the primary hits are already potent. Screening covalent fragments addresses these limitations.

Bradley martyn scannerz series#

In particular, it requires a large compound series with typically ambiguous structure–activity relationships, because no method to date can reliably rationalize which are the dominant interactions of the original fragment. (32,33) The major limitation in fragment-based screening is the weak binding affinity of fragment hits, which not only necessitates very sensitive biophysical detection methods, coupled with elaborate validation cascades, to eliminate attendant artifacts but also makes progressing hits to potency difficult and expensive. (30,31) In comparison to traditional HTS, fragment-based screening offers better coverage of chemical space and higher probability of binding due to lower molecular complexity. (28)įragment-based screening, which focuses on very low molecular weight compounds, is a successful hit discovery approach for reversible inhibitors (29,30) that has led to several drugs and chemical probes. (25−27) A major risk in screening large covalent libraries is that hits will be dominated by overly reactive compounds rather than by specific recognition.

Empirical high-throughput screening (HTS) for covalent binders is typically avoided, (24) owing to concerns about promiscuous activity. (17−23) While successful, in silico docking still has its limitations: it is limited to targets for which a crystal structure (or a high-quality model) is available, it cannot efficiently address protein flexibility, and it cannot predict the intrinsic reactivity of electrophiles and thus may result in highly reactive compounds. (4,12−16) More recently, large-scale covalent virtual screens have emerged as a method for the discovery of covalent binders. Historically, the most widespread approach to designing such inhibitors has relied on the incorporation of an electrophile into an already optimized reversible recognition element, (4,9−11) most notably in kinase inhibitors. This study highlights the potential of electrophile-fragment screening as a practical and efficient tool for covalent-ligand discovery.ĭiscovering new covalent inhibitors, however, remains challenging. No inhibitors were previously known for either. Combining our approach with high-throughput crystallography allowed rapid progression to potent and selective probes for two enzymes, the deubiquitinase OTUB2 and the pyrophosphatase NUDT7. In contrast, we found hits for most targets. Highly reactive and promiscuous fragments were rare and could be easily eliminated. We characterized this library by a new high-throughput thiol-reactivity assay and screened them against 10 cysteine-containing proteins. We hypothesized that mild electrophiles might overcome the selectivity challenge and constructed a library of 993 mildly electrophilic fragments. In principle, fragments that can irreversibly bind their target can overcome the low affinity that limits reversible fragment screening, but such electrophilic fragments were considered nonselective and were rarely screened. Covalent probes can display unmatched potency, selectivity, and duration of action however, their discovery is challenging.