29 June 2020

Quality control of the iNEXT poised fragment library

Two weeks ago we described fragment libraries built for crystallographic screening, and this week we continue the library theme. A primary challenge after validating a fragment hit is what to do next. You can look for similar compounds, either in-house or from vendors, but ultimately you’ll need to do chemistry. And this might not be trivial: you don’t want to embark on a multistep synthesis if you can avoid it. One solution is to build “poised fragments,” which contain at least one functional group amenable to easy chemistry. Characterization of such a library has just been published (open access) in J. Biomol. NMR. by Harald Schwalbe (Goethe University) and a large multinational group of collaborators.

The researchers are part of the European iNEXT (Infrastructure for NMR, EM, and X-rays for Translational research) consortium. A set of 11,677 commercial fragments was computationally analyzed, and 782 were purchased from several vendors. To efficiently cover chemical space, the researchers took a “minimum fragments and maximum diversity” approach: similarity analysis revealed 391 clusters, most with just 1-3 members. The fragments are mostly rule of three compliant, though a bit on the large side, with 80% of fragments in the 200-250 Da range and an average molecular weight of 220 Da.

Fragments were characterized by NMR and LC-MS. NMR experiments were done in both d6-DMSO (at 50 mM) and phosphate buffer (at 1 mM). The DMSO solutions were spiked with 10% D2O; this mixture remains liquid at 4 °C, thus avoiding freeze-thaw problems. Fragment concentrations were established by NMR using either external or internal standards.

Some 30% of fragments did not pass quality control. This probably will not come as a shock to long-time readers, though it is a bit worse than some previous studies. Just like unhappy families, fragments failed QC for a variety of reasons, including impurities, degradation, solubility, and even inconsistency with the expected structures. There were also a couple cases where two fragments were mixed together, suggesting operator error while assembling the library. NMR spectra for various types of QC failures are provided in the paper.

The researchers are honest about deficiencies in the library, noting that it contains PAINful molecules such as catechols and hydroquinones, though one wonders why these were not removed in the first place. Laudably, they provide SMILES for all 782 compounds along with an extensive set of physicochemical calculations (see here – opens as an Excel spreadsheet). Weirdly though, the researchers do not specify which molecules failed QC or which vendors they came from. At first I thought these had been weeded out of the final set, but five examples shown as failures appear in the spreadsheet. The library is sold commercially by Enamine as the DSI-poised Library, and since this library contains only 768 compounds perhaps the most egregious bad actors have been removed. Enamine was rated highly in a reader poll, so presumably the compounds have all passed QC.

So how does the fragment library perform? That – unfortunately – is not addressed in this paper, though the DSI-poised library was among those screened against the SARS-CoV-2 main protease (MPro). Have you used it? And if so, has the “poised” nature of the library allowed you to efficiently grow fragment hits? Hopefully these questions will be answered in the literature, if not in the comments.

1 comment:

Anonymous said...

It performed very well for us. We got 80 hits in a crystallization screen for our target. All fragments look interesting and chemistry is feasible. Still, I must say we spend a lot of time on the optimization/SAR phase...Buying close analogues could perhaps speed up things, but previously this strategy was not succesful for us, and it is expensive. So we design and synthesize instead.