Kinase inhibitors are common. Some 40% of the fragment-derived clinical compounds in our latest list target kinases. Kinase activators, on the other hand, are rare. It is easier to interfere with something than to enhance it, and of the 630+ posts on Practical Fragments, I believe only two discuss enzyme activators. A new paper (here) by Ping Lan, Iyassu Sebhat, and colleagues at Merck and Metabasis provides a third example.
The researchers were interested in adenosine monophosphate-activated protein kinase (AMPK), which plays a critical role in metabolism. As its name suggests, this kinase has naturally occurring activators, though these are nucleotides and thus not particularly useful as chemical probes. It also has fiendishly complex biology: the active enzyme is a heterotrimer of three distinct proteins, each of which comes in two or three flavors, leading to 12 different isoforms with their own unique tissue distributions – which vary among different animals.
After multiple HTS screens failed to produce anything of value, the researchers turned to fragments. Realizing that AMPK was a tough target, they assembled a library of 25,000 highly diverse fragments tending towards super-sized (all of them were greater than 200 Da, and they had up to 22 non-hydrogen atoms). A biochemical screen yielded just three hits, including compound 4.
Despite the generally larger size of fragments in the library, it is interesting that compound 4 follows the rule of three, with just 16 non-hydrogen atoms. Although only modestly active, the small size gave an impressive ligand efficiency, and the activation was nearly 80% that of the natural ligand AMP.
Rigidification of the linker between the benzimidazole and the acid moiety led to compounds such as 27, with low micromolar potency, while growing from the benzimidazole itself led to high nanomolar compounds such as compound 36. Combining these two modifications and further optimization for pharmacokinetic properties led to MK-3903, which was chosen as a development candidate.
MK-3903 activates 10 of the 12 AMPK isoforms and is fairly selective against a panel of off-targets. As predicted mechanistically, administering the compound to mice increases the phosphorylation of downstream substrates of AMPK. It also causes decreased fatty acid synthesis and increased insulin sensitivity. However, a related molecule causes cardiac hypertrophy in rats and monkeys.
In addition to the fact that MK-3903 is an enzyme activator, there are several other notable features about this story. First, despite the difficulty of the target, the team made rapid progress, moving from the initial screen to useful tool compounds in less than a year. Second, as near as I can tell, this optimization was done in the absence of direct structural information on how the compounds bind. (A publication by a separate team, who was closely monitoring the patent literature, describes the crystal structure and mechanistic analysis of a related molecule.) Third, all of this work stemmed from a single fragment: although more ligandable targets may produce lots of hits, in the end you only need one.