06 May 2012

Fragments versus Ras

The protein Ras is one of those cancer targets that’s been around forever and has rebuffed countless attacks by many researchers using multiple strategies. The most obvious ligand pocket is the one where GTP and GDP bind. Unfortunately, these molecules bind with picomolar affinity, and they are present at very high concentrations in cells. To try to find an alternative small molecule binding site, Guowei Fang and colleagues at Genentech took a fragment-based approach, and have reported their results in a recent issue of Proc. Nat. Acad. Sci. USA (as well as at a recent meeting).

The researchers used 1D NMR screening (STD) to screen 3300 fragments in pools against GDP-bound KRas; 240 hits were retested as single compounds and further validated by 2D NMR (HSQC). This resulted in 25 confirmed hits. Surprisingly, all of them appeared to bind to one region of the protein some distance from the GDP-binding site. Subsequent crystallography confirmed that these fragments bind to a small pocket about 250 Å3 in size. However, there could be more here than meets the eye, as this is a fairly flexible region of the protein, and the pocket changes shape in response to different fragments.

 At least one of the fragments, DCAI, not only binds to Ras, it also inhibits the association of Ras with the protein SOS, thereby blocking nucleotide exchange and Ras activation. Interestingly, this blocking activity was distinct from binding activity; the fragment BZIM has comparable affinity as judged by NMR, yet does not inhibit the interaction with SOS.

Somewhat surprisingly given its low affinity, DCAI is also active in cell-based assays. And although the molecule is still a long, long way from a drug, the results are encouraging. Perhaps a fragment-based approach will finally succeed against this target. Or perhaps Ras will yet again reveal its intransigence.

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