Pacifichem 2021

Pacifichem, the last significant conference of 2021, has just ended. Traditionally held every five years, these meetings usually bring thousands of visitors from Pacific Rim countries to Honolulu. They are planned years in advance: symposia proposals were due in early 2018. Pacifichem 2015 saw the first symposium dedicated to FBLD, and that was so popular that a few of us planned one for 2020. The conference organizers decided to postpone the 2020 meeting in the hope that we could all meet in person. But SARS-CoV-2 had other plans, and instead of meeting in Hawaii we met on Zoom.

 

Time zones were challenging. Four-hour sessions started in the morning or evening Hawaiian time, which translated to 02:00 in Shanghai or 23:00 in Boston, respectively. In contrast to other virtual meetings almost all the presentations were live and not recorded, which meant that you only had one chance to see a talk.

 

Despite these challenges and universal Zoom-fatigue, the event came off quite well. With more than two dozen presentations I won’t attempt to cover everything but will instead just touch on a few themes.

 

Methods

Quite a few talks focused on methods, with NMR being especially well-represented. The symposium started with Will Pomerantz (University of Minnesota) discussing Protein-Observed Fluorine (PrOF) NMR, in which fluorinated amino acids are introduced into proteins. We’ve written about this previously, including Will’s longstanding interest in assessing shapely fragments. After reading about Mads Clausen’s fluorinated Fsp³-rich library, Will established a collaboration and has found 8 hits from 79 fragments screened against BET bromodomain proteins. He has also been able to optimize potent leads selective for either the BD1 or BD2 domains of BRD4.

 

Scott Prosser (University of Toronto) is also using NMR to study fluorine-labeled proteins, in this case GPCRs. And Michael Overduin (University of Alberta), one of the symposium organizers, is also studying membrane-bound proteins using NMR techniques.

 

On the extreme side of spectrometers, Chojiro Kojima described the 950 MHz NMR at Osaka University. This enables a ¹H-¹³C HSQC experiment on protein as dilute as 0.2 micromolar, which could be valuable for insoluble or hard-to-purify proteins. The facility is open to international collaborators. Chojiro also described isotopically labeling proteins with transglutaminase and ¹H{¹⁹F} STD NMR, which works even when the fluorine peak itself is broadened to invisibility.

 

But you don’t need a big magnet to do good science. Brian Stockman’s group at Adelphi University is composed entirely of undergraduates who use substrate-detected NMR to follow enzymatic reactions to find inhibitors of neglected parasitic infections.

 

All techniques can give false positives, and NMR can be very effective at weeding these out. As we described two years ago Steven LaPlante (NMX) has been developing methods to rapidly identify aggregators and has been assembling something of a taxonomy; more later.

 

But the conference was not all NMR all the time. Rebecca Whitehouse (Monash University) described a 91-compound “MicroFrag” library consisting of fragments containing 5-8 atoms, “somewhere in the land between solvents and fragments.” NMR and crystallographic screens of the difficult antimicrobial target DsbA gave very high hit rates, and both techniques successfully identified the large but shallow substrate-binding groove. In contrast, screening actual solvents or using the well-established FTMap computational approach did not clearly identify this groove.

 

The push in crystallography is towards increased speed, and Debanu Das described the high-throughput platform at Acclero BioStructures, which is capable of screening 1000 fragments per week, similar to XChem. But if even that is too slow for you, Marius Schmidt (University of Wisconsin-Madison) described mix-and-inject experiments using the European X-ray free-electron laser (XFEL). Much of the focus with this technique has been on high-speed enzymology, but since 100 datasets can be collected in 10 hours it can be used for high-throughput crystallographic screening too. An upgrade next year will increase this to 1000 datasets in 3 hours, though the resulting petabytes of data will no doubt create headaches for IT departments.

 

It’s not enough to find hits, you need to figure out what to do with them, and symposium organizer Martin Scanlon (Monash University) discussed a computational approach (GRADe, similar to Fragment Network) as well as the experimental (REFiL) approach we’ve discussed previously. Across 9 projects the techniques were successful at improving affinity, in some cases from unmeasurable levels.

 

Covalent Fragments

Several talks focused on covalent FBLD. Alexander Statsyuk (University of Houston) proposed five rules for covalent fragments: 1) ease of synthesis; 2) non-promiscuous electrophiles; 3) intrinsic reactivity should be the same within the library; 4) a given library should use the same electrophile; and 5) the electrophile should be on the end of the molecule with a minimal linker connecting it to the variable fragment. Some of these make sense, but it would have been fun to debate others over Mai Tais.

 

Dan Nomura (UC Berkeley) described using covalent fragments in chemoproteomic experiments, where he has identified more than 100,000 potentially ligandable hotspots in more than 16,000 human proteins. Among other applications, this allows him to make bifunctional molecules to bring two proteins together. A clear application is PROTACs, where the electrophilic fragment targets an E3 ligase, but he also described targeting the deubiquitinase OTUB1 to stabilize proteins.

 

Earlier this year we celebrated the approval of the KRAS^G12C inhibitor sotorasib. This target had long resisted drug discovery efforts; Ratmir Derda (University of Alberta) evocatively mentioned “waves and waves of medicinal chemists washing off its shore for 30 years.” Success was finally enabled using disulfide Tethering, and David Turner (Frederick National Laboratory) is now using this approach to interrogate nearly every surface-exposed residue by systematically mutating them to cysteines and screening against more than 1000 disulfide-containing fragments. He is well over half-way through the 85 mutants, and the resulting dataset should be valuable not just for drug discovery but for understanding molecular interactions.

 

Success Stories

With more than 50 drugs in the clinic derived from fragments, there were several success stories. Masakazu Atobe (Asahi Kasei) presented the discovery of the potent PKCζ inhibitors we wrote about here. And Chaohong Sun (AbbVie) described inhibitors of TNFα (see here), emphasizing the importance of robust biophysics and early committed chemistry.

 

Finally, Emiliano Tamanini (Astex) presented a nice fragment-to-lead effort to find a selective inhibitor of HDAC2. Despite some successes, histone deacetylase inhibitors tend to be non-specific and come with side effects. Emiliano described a fragment screen that identified a new metal chelator and used fragment merging to develop a molecule capable of crossing the blood-brain-barrier.

 

These last two stories in particular are examples of pursuing difficult targets, another theme throughout the conference. When asked about the challenges of targeting cancer-resistance-causing glucuronosyltransferases, Katherine Borden (University of Montreal) responded, “if you don’t try, where will you be?”

 

Bright words for these darkling days.