SAR by NMR is the Nuclear Magnetic Resonance (NMR) based method in which small organic molecules that bind to the proximal site are identified, optimized and finally linked together to produce high-affinity ligands. It is called “SAR by NMR” because the structure-activity relationship (SAR) is obtained by the Nuclear Magnetic Resonance (NMR). It is based on the fragments approaches to drug design.  With this technique, compounds with nanomolar affinity for a target protein can be rapidly discovered by tethering two ligands with micromolar affinities. The method reduces the chemical synthesis and the time required for the discovery of high-affinity ligands and is particularly useful in target-directed drugs research.

1. SAR BY NMR
SAKEEL AHMED (PhD Scholar)
Department of Pharmacology (NIPER, Mohali)

2. SAR BY NMR
 SAR by NMR is the Nuclear Magnetic Resonance (NMR) based method in which
small organic molecules that bind to the proximal site are identified, optimized and
finally linked together to produce high-affinity ligands.
 It is called “SAR by NMR” because structure activity relationship (SAR) is obtained by
the Nuclear Magnetic Resonance (NMR).
 It is based on the fragments approaches to drug design.
 With this technique, compounds with nanomolar affinity for a target protein can be
rapidly discovered by tethering two ligands with the micromolar affinities.
 The method reduce the chemical synthesis and the time required for the discovery
of high affinity ligands and is particularly useful in the target- directed drugs
research.
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3. Figure. The SAR by NMR concept. Multiple, low molecular weight
compounds (“fragments”) are identified that can simultaneously bind to
proximal sites on the protein surface. Based on structural information,
these fragments can be linked to produce a ligand whose binding affinity
is theoretically the sum of the individual pieces.
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4.  Library of low molecular weight compounds is screen to identify molecules that bind to the target protein.
 Binding is confirmed by the 15N and 1H Amide chemical shift in 2D HSQC Spectra upon addition of ligand.
 These spectra can be rapidly obtained, making it possible to screen a large number of compounds.
 Once a lead molecule is identified, analogs are screened to optimize binding to this site.
 Next, a ligand is sought that interacts with a nearby site. Binding to a second Pocket is determined by observing changes
in a different set of amide chemical shifts in either the original screen or a screen conducted in the presence of the first
fragment.
 From an analysis of the chemical shift changes, the approximate location of the second ligand is identified. Optimization
of the second ligand for binding to this site is then carried out by screening structurally related compounds
 When two "lead" fragments have been selected, their location and orientation in the ternary complex are determined
experimentally either by NMR spectroscopy or by x-ray crystallography.
 Finally, on the basis of this structural information, compounds are synthesized in which the two fragments are linked
together with the goal of producing a high affinity ligand.
SCREENING BY SAR BY NMR

5. Fig. An outline of the SAR by NMR method.

6. An HTS of a relatively large (~106) compound collection typically serves as the entry point to lead
identification and optimization.
However, this approach is failing to produce high-quality clinical candidates at a rate
commensurate with the resources that have been dedicated to these efforts.
There are several very specific reasons for these trends:
1. First, the quality of the leads that have resulted from these large-scale screens tend to lack the
appropriate physicochemical properties for clinical success. For example, typical leads tend to
be large and lipophilic––significantly reducing the chances that oral bioavailability can be
achieved.
2. An HTS fails to produce any chemical matter that is suitable for further discovery efforts. 50%
of all screens produce quality leads that are worthy of further evaluation.
CONVENTIONAL HTS vs FRAGMENT BASED SCREENING

7. 1. One of the greatest advantages of fragment-based screening as compared to conventional
HTS is the substantially greater coverage of chemical diversity space (i.e., the universe of
compounds that can potentially exist) that can be achieved.
2. Even small fragment libraries represent a substantially larger fraction of diversity space
than conventional compound repositories.
3. Another factor contributing to this increased diversity is that the ultimate goal is to link or
tether the individual fragments found at the multiple subsites. This means that the
effective number of ligands that is virtually assessed in a fragment screen is a power
function of the number of subsites evaluated and the number of different ways to link the
fragments together.
Continue……

8. Case Study: Matrix Metalloproteinase Inhibitors
 Matrix metalloproteinases are a family of zinc-dependent endopeptidases that are
implicated in a variety of diseases, including arthritis and tumor metastasis.
 Fragment screen against MMP-3 and discovered that acetohydroxamate (a zinc-chelating
moiety with a KD value of 11 mM for the protein) could bind to the protein
simultaneously with a number of biaryl compounds (with KD values in the 20–100 μM
range).
 The three-dimensional structure of a ternary complex clearly revealed that these two
fragments could be linked. In fact, one of the first linked compounds exhibited an IC50
value of 57 nM and bound to the protein as designed.
 ABT-518 which exhibited excellent oral anti-tumor efficacy in animal trials and was
approved for Phase 1 clinical trials.
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9. Figure: SAR by NMR on matrix metalloproteinases