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Figure 4.1 A schematic overview of the development of FBS (and HTS) hits with respect
to typical potencies achieved at different stages. The primary fragment hits typically have
potencies in the range 0.1-1mM. Selected fragment hits are optimized with respect to affin-
ity without substantially increasing the molecular weight. The optimized fragments are then
slightly expanded, usually by testing in-house or commercially available analogous com-
pounds or by applying one-step chemistries if the fragment library is designed to facilitate
this. Alternatively, fragments are linked or merged. After a couple of iterations, compounds
with low micromolar or better potencies and with high ligand efficiencies have normally been
identified. These compounds are good starting points for a full-scale chemistry program. Mean-
while, the HTS assay has been developed and formatted. Confirmed HTS hits refer to hits with
high purity and stability and good dose-response curves. Further, they should be confirmed
in an orthogonal assay, preferably a biophysical binding assay, to ascertain that they are active
via a 'sound' mechanism. [17, 18] Finally, hit series should be identified and prioritized and com-
mercially available analogues purchased. From this stage, further development of compounds
originating from the fragment screen and the confirmed HTS hits does not differ significantly
and could be fed into the same medicinal chemistry optimization program.
Figure 4.2 The workflow in a fragment-based screening project.
function of the target protein in the desired way. The biochemical assay does not have
to be employed during the first iteration, but could wait until the affinity of the developed
fragments has increased. This could very well be a low-throughput assay; in many instances
it is possible to use NMR as an enzymatic activity assay. [ 19 ] As discussed in Figure 4.1,
analogues to fragment hits are either synthesized by one-step chemistries or acquired from
in-house compound collections or commercial sources. In the first iteration, the fragment
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