Fragments vs MTH1: a chemical probe

As mentioned last week, CHI’s FBDD Meeting was chock-full of success stories. Some of these have recently been published, including work in J. Med. Chem. by Jenny Viklund (Sprint Biosciences) and collaborators at Bayer, the University of Oxford, and the Structural Genomics Consortium.

The researchers were interested in the protein MutT Homologue 1 (MTH1), which helps clear the cell of oxidized nucleotide triphosphates. The enzyme is upregulated in several cancers, and previous research involving non-selective MTH1 inhibitors had implicated it in cancer cell survival. But other research suggested that the effects on cancer cells were due to off-target effects. Clearly what was needed was a high-quality chemical probe.

The researchers started with a thermal shift assay of just 723 fragments screened at 1 mM, of which 166 increased the melting temperature by at least 1°C – a remarkably high hit rate suggesting good ligandability. Of the 49 fragments tested in full dose response thermal shift assays, 48 showed dose dependence. Compound 1 was not the most potent or ligand efficient, but it was synthetically tractable and different from other reported MTH1 inhibitors. Isothermal titration calorimetry revealed a dissociation constant of 49.5 µM, and the compound was also active in an enzymatic assay.

A crystal structure of compound 1 bound to MTH1 guided the selection of similar molecules from an in-house collection, such as compound 3. The structure also revealed a small pocket near the 2-position of the azaindole ring, and compound 5 – also available from the in-house collection – gave a nice pop in potency. Synthesis of a few analogs quickly led to compound 7, with mid-nanomolar activity. Crystallography revealed that the molecule bound mostly as expected. But because an asparagine side chain shifted to accommodate it, standard rigid-protein computational techniques would likely not have predicted its binding.

Further optimization for both potency and DMPK properties ultimately led to BAY-707, which is orally bioavailable in mice. In the interest of space I won’t go into details, but the paper is worth reading for a lovely, well-written account of lead optimization. Astute readers will recognize that all these molecules contain a 7-azaindole core, which is the same moiety that led to three clinical kinase inhibitors. The researchers tested representative molecules against a large panel of kinases as well as other ATPases and determined that the series is quite selective.

With probe in hand, the researchers set off to test whether inhibiting MTH1 would be useful for treating cancer. Unfortunately, as reported in another paper, the results actually “devalidate” the target. Despite potently inhibiting enzymatic activity in cells, BAY-707 showed no growth inhibition on several cancer cell lines, nor did it show activity in mouse xenograft models. While certainly disappointing, the results with this selective inhibitor at least provide a better understanding of biology.

This is also an example of just how quickly FBLD can yield results: at the CHI meeting Jenny said that it took 3.5 FTEs just 14 months from the start of synthesis to discover BAY-707, and the paper says this required only 35 compounds. A nice counterexample the next time someone says fragment approaches take too long.