Fragments vs Trypanosoma parasites

Last month we highlighted how fragments could be used to discover inhibitors of protein-protein interactions (PPIs). Today we continue the theme of fragments vs PPIs, in this case the interaction between PEX14 and PEX5, proteins which are important for glucose metabolism in disease-causing protists such as Trypanosoma.

The research, published recently in Science, was done by a large multinational team led by Grzegorz Popowicz, Michael Sattler (both at Helmholtz Zentrum München), and Ralf Erdmann (Ruhr University Bochum). They started by solving the NMR structure of the N-terminal domain of PEX14 from T. brucei, the organism that causes sleeping sickness. Previous work had shown that PEX5 binds to this domain, with two aromatic side chains of PEX5 binding in adjacent hydrophobic pockets. With this information in hand, the team performed a virtual screen of several million (non-fragment-sized) molecules. Eight of the best-scoring hits were tested, and four showed binding in an NMR assay, with compound 1 having the highest affinity.

Next, the researchers screened a library of 1500 fragments (each at 1 mM in pools of 5) using ¹H, ¹⁵N HMQC NMR. This led to 12 hits with affinities better than 2 mM. Strikingly, all of these fragments contained fused bicylic aromatic ring systems, three of which were substituted naphthyls. Appending these onto compound 1 led to compound 4, with low micromolar affinity. Introducing an amine to interact with a glutamic acid residue in PEX14 led to compound 5, with high nanomolar affinity. This compound also showed activity against several species of pathogenic Trypanosoma. Further tweaking led to a molecule with activity in a mouse model of infection.

This example of fragment-assisted drug discovery (FADD) is reminiscent of other cases (described here, here, and here) in which fragments were used to replace elements of a previously identified molecule. While it is possible that traditional medicinal chemistry could have achieved the same result, fragments probably helped winnow down the number of molecules to be synthesized. It is also nice to see this technology applied to understudied diseases.