Biomedical Engineering Reference
In-Depth Information
templates containing three adjacent DTS-coding regions. Three consecutive DNA-
programmed acylations and one Wittig macrocyclization reaction allowed for the
construction of a (64
4 plus a built-
in phenyl sulfonamide derivative, which was known to bind carbonic anhydrase
with nanomolar affinity) (Figure 11.11). Following panning experiments against
carbonic anhydrase and PCR amplification of the library before and after selection,
enzymatic digestion of the
NlaIII
domain (used as a coding sequence for sulfonamide
positive control) revealed remarkable enrichment of the expected phenyl sulfonamide
macrocycle, thus confirming the successful application of DTS principles to DNA-
encoded chemical library construction [60].
More recently, using a strategy very similar to the one described in Figure 11.14,
the same laboratory reported the construction of a larger macrocyclic fumaramide
library containing 13,824 [(12
+
1)-membered macrocycle library (4
×
4
×
8] structures, starting from eight
independent scaffolds and 36 different building blocks [61].
The library was used for in vitro selection against 36 different target proteins.
Resynthesis of the selected macrocyclic structures resulted in the identification of
various kinase inhibitors and activators. The two most potent compounds isolated
specifically inhibited Src kinase with IC
50
values of 680 and 960 nM, respectively
(see Table 11.1) [77]. Starting from these compounds, Georghiou et al. developed and
characterized in molecular detail a second generation of macrocycles with potencies
as high as IC
50
≤
×
12
×
12)
×
4 nM [78].
In analogy with DNA-routed libraries (see Section 11.2.2.2), the DNA appendage
serves for both encoding and programming the library assembly. Therefore, templates
surviving selection process can be used as an evolutionary input for further rounds
of chemical translation and selection. However, for DNA-templated libraries, such a
evolutionary process has not been experimentally demonstrated yet.
As at low concentrations DTS reactions depend exclusively on the hybridiza-
tion of reagent and template, each DNA pair behaves as an independent molecu-
lar reactor [70,79]. Only DNA reactants linked to complementary oligonucleotides
yield intramolecular products, whereas reactions between mismatching strands are
substantially negligible. Following this principle, Liu's group applied DTS for the
discovery of new chemical reactions [79,80]. In a first proof-of-concept experiment,
two pools of DNA-linked substrates were mixed at nanomolar concentrations and
evaluated simultaneously for bond-forming reactions. The novel reaction discov-
ery method led to the identification of a new Pd(II)-mediated carbon-carbon bond
reaction between a terminal alkyne and a monosubstituted alkene, as depicted in
Figure 11.15a [79]. Notably, the newly discovered DNA-templated reaction also
operated in a non-DNA-templated format setup (Figure 11.15b) [79].
11.2.2.3.1 DPC Platform
On the groundwork by Liu's group on DNA-templated
synthesis (DTS) and its implications in DNA-encoded chemical library construction,
Ensemble Therapeutics developed DPC (DNA-programmed chemistry platform), an
integrated system for the synthesis and selection of DNA-encoded libraries based on
macrocyclic scaffolds (also known as
Ensemblins
) [81].
