Dynamic combinatorial chemistry
(DCC) sounds incredibly cool. The
idea is that libraries spontaneously form and reform. Add a protein and Le Châtelier's principle favors the formation of the best binders. In other words, not only
does cream rise to the top, more cream is actually created.
The applications of DCC for
fragment linking are obvious, and indeed early reports date back nearly twenty
years to the dawn of practical FBDD. The latest results are described in a new
paper in Angew. Chem. Int. Ed. by
Anna Hirsch and collaborators mostly at the University of Groningen.
The researchers were interested
in the aspartic protease endothiapepsin, which is a model protein for more
disease-relevant targets. This is a dream protein: it is easy to make in large
amounts, crystallizes readily, and is stable for weeks at room temperature.
Readers will recall that this protein has also been the subject of multiple
screening methods. Previous efforts using DCC had generated low micromolar inhibitors
such as 1 and 2. These acylhydrazones form reversibly from hydrazides and
aldehydes. Crystallography had also previously revealed that compound 1 binds
in the so-called S1 and S2 subsites of endothiapesin while compound 2 binds in
the S1 and S2’ subsites. In the current paper, the researchers enlisted DCC to try
to combine the best of the binding elements.
To do this, the researchers chose
isophthalaldehyde, which contains two aldehyde moieties, and nine hydrazides,
which could give a total of 78 different bis-acylhydrazones. They incubated 50 µM
of isophthalaldehyde with either four or five of the hydrazides (each at 100 µM),
with or without 50 µM protein, and in the presence of 10 mM aniline to
accelerate the exchange. Reactions were allowed to incubate at room temperature
at pH 4.6 for 20 hours, after which the protein was denatured and the samples
were analyzed by HPLC to see whether some products were enriched in the
presence of protein.
Biologists may want to consider
whether their favorite proteins would remain folded and functional under these
conditions, and chemists may also balk at molecules containing an acylhydrazone
moiety – let alone two. Leaving aside these concerns, though, what were the
results?
As one would hope, some molecules
were enriched over others when protein was present, though only by a modest two
or three-fold. Two of the enriched molecules – both homodimers – were
resynthesized and tested. Compound 13 was quite potent, and crystallography
revealed that it binds in a similar fashion to compound 1, though electron
density is missing for part of the molecule. Compound 16, on the other hand, is
only marginally more potent than the starting molecules. Unfortunately the
researchers do not discuss the activities of molecules that had not been
enriched at all.
The paper ends by stating rather
hopefully that DCC “holds great promise for accelerating drug development for
this challenging class of proteases, and it could afford useful new lead
compounds. This approach could be also extended to a large number of other
protein targets.”
I’m not so sure.
I’m not so sure.
This is an interesting study; the work was carefully done and thoroughly
documented—but I’m less sanguine about whether DCC will actually ever be practical for lead generation. Indeed,
the very fact that the experiments were done well yet are incapable of
distinguishing a strong binder from a weaker one argues that the technique is
inherently limited. I would love to see DCC work, but it seems to me that, even
after two decades of effort, DCC has not been able to move beyond proof of
concept studies. Does anyone have a good counterexample?
2 comments:
Hi Dan,
I skimmed the paper, it seems they have not done any crystallography for the new compounds, it was all modeled in.
Maybe I misread when you say "...compound 13 was quite potent and crystallography revealed that it binds in a similar fashion to compound 1, though electron density is missing for part of the molecule...".
As regarding examples...nope, not many I have seen..maybe this could be of interest? Oldie but ..goldie:
http://onlinelibrary.wiley.com/doi/10.1002/cbic.200900537/abstract
Thanks FBDD evangelist. The co-crystal structure of compound 13 is deposited as 5HCT in the pdb. One of the indole moieties is visible, along with the central phenyl ring, but you are correct that modeling is used to place the other indole moiety.
The ChemBioChem paper is interesting, though the best compound is only ~5-fold more potent than the starting "fragment". I confess I've actually dabbled in DCC a bit myself and have found other approaches more effective, even if less elegant.
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