Chemistry Reference
In-Depth Information
benefits of the TINS fragment library designed and tested as collaborative effort between
ZoBio (www.zobio.com) and Pyxis Discovery (www.pyxis-discovery.com) of Delft, The
Netherlands.
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It is now a well-accepted principle that the 'rule of three'
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forms an approximate limit
guiding the chemical nature of compounds that should be considered as a fragment for
inclusion in a collection for ligand screening. At the other end of the spectrum, recent
work from the Shoichet
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laboratory suggests that including very simple fragments of less
than approximately 150 Da could cause difficulties downstream during the lead evolution
process. Clearly, a number of
in silico
filters must also be employed to remove undesir-
able compounds such as known toxicophores or reactive groups. In our efforts, we also
placed great emphasis on water solubility of the compounds. In one of the first publications
concerning fragment library design, only about 50% of the selected fragments possessed
sufficient solubility (1mM) to be screened.
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In more recent publications, better results
for the water solubility of fragment libraries have been reported.
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The prediction of
water solubility, however, remains a challenge because one has to take into consideration
both the crystal and solution states of the compound. Moreover, in our own analysis, we
have not been able to find a simple correlation between the number of hydrogen bond
donors/acceptors and water solubility. Since computational methods for better prediction
of water solubility are still under development, one must determine experimentally the
solubility of a given fragment. However, by applying cut-off values based on experience,
for properties that can be better predicted, such as Clog
P
and the number of hydrogen bond
donors and acceptors, which have a profound influence on water solubility, the fraction of
water-soluble fragments can be increased considerably. In our own efforts, about 90% of
compounds that were selected were soluble as singletons at 500 μM in phosphate-buffered
saline and 5% DMSO. Evotec has recently mentioned an in-house QSAR model to predict
solubility which is claimed to be useful, but no data are currently available.
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While ori-
ginally our emphasis on water solubility stemmed from practical aspects of making mixes
of compounds at 500 μM each in aqueous buffer, this effort has been well served when
screening membrane proteins, since we feel that it is one of the important reasons that we
have so far experienced a very low false positive rate.
Our library, which is intended to serve as a source of chemical diversity, is composed of
compounds selected from four themes: (1) diversity using the scaffold-based classification
approach (SCA),
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(2) amino acid derivatives, (3) scaffolds found in natural products and
(4) shape diversity. All compounds were selected from a carefully prepared database rep-
resenting 70 000 compounds that would make desirable starting points for drug discovery,
including rule of three compliance, and were commercially available from reliable suppli-
ers. One of our explicit intentions in forming the library upon these design principles is to
evaluate the performance of the various classes of compounds against different targets, both
soluble and membrane bound. Although it remains too early to draw sensible conclusions
from the roughly 10 targets that have been screened to date, in many cases there are up to
twofold differences in hit rates between the different themes for a given target.
6.2.2
Immobilization and Reference Protein
The strength of TINS lies in the fact that it is a referential system. That is, the signal acquired
in the presence of the target protein is compared with the signal acquired in the presence
of a reference sample consisting of a known protein immobilized at approximately the
