Fragments vs soluble epoxide hydrolase – hundreds of them!

Soluble epoxide hydrolase (sEH) is a potential target for cardiovascular and immune disorders. The enzyme catalyzes the hydrolysis of long chain, lipophilic epoxyeicosatrienoic acids. These bind in a largely hydrophobic “L-shaped” pocket, with the catalytic machinery at the point of the L. Although it is relatively easy to find potent inhibitors of this enzyme, these tend to be greasy, insoluble, and non-druglike. In a recent paper in Bioorg. Med. Chem. Yasushi Amano and colleagues at Astellas describe a fragment-based approach to find better leads.

The researchers started with a high-concentration (up to 2 mM) enzymatic inhibition assay of 4200 fragments, resulting in 307 hits with IC₅₀ values between 700 nM and 1.7 mM. All of these were taken into co-crystallography trials, yielding crystals for about half of them. The other fragments were soaked into apo-crystals of sEH (that is, crystals without bound ligand) to get as many structures as possible. All together 126 crystal structures of fragments bound to sEH were solved, which is all the more impressive considering that there are only three authors on the paper!

Most of the fragments (83) bound at the catalytic site (example shown in green), while 29 bound to one of the lipophilic branches of the L and 9 bound to the other (cyan and magenta). Five fragments bound at two different sites within the enzyme. The researchers discuss ten of the fragments in some detail.

Many of the fragments that bind at the catalytic site contain amides or ureas – moieties in known inhibitors – but some of them (such as compound 3 above) are more unusual. Also, despite the generally lipophilic nature of the binding pocket, many of the fragments – even those that bind in the hydrophobic branches of the L – make hydrogen bonds to the protein or to bound water molecules. This suggests that it should be possible to find potent inhibitors that are less hydrophobic than previously reported molecules. Fragment growing, linking, and merging approaches could all work, and indeed the researchers hint that results of these studies will be reported in future papers.

More importantly, this paper provides a great set of crystallographically validated fragments binding to distinct sites on a well-characterized protein. Helpfully, the researchers have deposited the coordinates of the 10 co-crystal structures discussed in the protein data bank. Moreover, all of the fragments are commercially available. This seems like an ideal model system for validating computational docking and scoring approaches as well as for better understanding the energetics of protein-ligand interactions. If I were an academician working in these areas, I’d jump in feet first!