10 July 2017

Reagents as covalent fragments

Covalent drugs are a thing these days: as long as you can get selectivity, there’s nothing like a covalent bond to juice up affinity for a target. Screening for covalent fragments is thus a reasonable approach, and multiple researchers have assembled libraries of fragments containing either irreversible or reversible covalent “warheads”. The latest example, by Marion Lanier, Mark Hixon, and collaborators at Takeda, appears in J. Med. Chem.

Boronic acids can form reversible covalent bonds with the side chains of serines or threonines in proteins, with a predilection for the highly reactive active-site residues found in hydrolytic enzymes. Indeed, three approved drugs – bortezomib, tavaborole, and ixazomib – contain boronic acids.

When medicinal chemists think of boronic acids, they probably think of them as reagents for the Suzuki coupling, a useful method for forming carbon-carbon bonds. Because the reaction is so popular, some 6000 boronic acids are commercially available, many of them fragment-sized. Thus, the researchers assembled a set of 650 into a boronic acid library (BAL).

To determine whether this BAL would be useful, the researchers screened it against autotaxin, a phospholipase with anti-cancer and anti-inflammatory potential. Fragments were tested at 100 µM in a functional assay, with hits retested in 11-point dose-response curves. This yielded a whopping 51 molecules with IC50 values better than 10 µM, some as good as 200 nM.

The researchers also screened autotaxin against a set of 1750 non-boronic acid containing fragments, this time at 500 µM. Not surprisingly, hits tended to be significantly weaker despite the similar sizes of the fragments. The BAL fragments had average ligand efficiencies of 0.61 kcal mol-1 per heavy atom, while the conventional fragments averaged a lower but still respectable 0.41. Some of the BAL fragments were also crystallized bound to autotaxin, revealing that they do in fact form bonds with the catalytic threonine. 

This is a nice paper, though I do wish that the researchers had tried to calculate the inherent reactivities of the boronic acids to determine how these differences affected their affinities for the protein, as has been done for other warheads such as aryl acrylamides. Also, it would be interesting to see how a docking program such as DOCKovalent performs against this target with the same set of fragments. Hopefully we’ll see these questions addressed in the future. In the meantime, expect to see commercial vendors start offering libraries of boronic acid fragments.


Peter Kenny said...

Hi Dan, it’s not straightforward to calculate (or even define) ‘inherent reactivity’ in a meaningful way for warheads that form covalent bonds with targets because non-covalent interactions between target and ligand can potentially influence both on-rate and affinity. I believe that it can be useful to label warheads as ‘reversible’ or ‘irreversible’ but it’s important to bear in mind that optimization of fragment hit affinity is likely to lead to a decrease in off-rate.

Dan Erlanson said...

Hi Pete,

Actually, it's quite common to assess the inherent reactivity of warheads using a common nucleophile. In the case of thiol-reactive warheads this is usually glutathione or some other cysteine derivative. The Amgen paper I mentioned above does this for aryl acrylamides, and the researchers found a linear Hammett correlation. They also got good agreement between experimental results and density functional theory calculations.

Another paper from Pfizer looks at a more diverse set of irreversible warheads.

The reason this is important is that if you have the same fragment with different warheads, and the warheads differ significantly in reactivity, you may think you are optimizing molecular complementarity when in fact you are just tweaking reactivity. Alexander Statsyuk has a nice discussion of this, covered here.

All of this also holds true for reversible covalent inhibitors, as discussed here.

You are absolutely correct that "non-covalent interactions between target and ligand can potentially influence both on-rate and affinity", and these interactions can in fact be improved during lead optimization.