Molecular interactions are relevant to both prediction-driven and hypothesis-driven molecular design. Design hypotheses are often framed in terms of molecular interactions and a predictive model for affinity that fails to capture the physics of molecular interactions will choke when used outside narrowly-defined congeneric series. Although we think of affinity in terms of contacts between protein and ligand, it is important to remember that the contribution of a particular contact to affinity is not strictly an experimental observable.
In FBDD we think of ligands in terms of their component fragments and are particularly interested in the extent to which the properties of fragments determine the properties of structurally elaborated compounds. Comparing the affinties of ligands with the fragments from which they might have been derived is one way in which this question can be addressed and one will occasionally encounter the term 'deconstruction' in the FBDD field. Just as you need to be careful how you link fragments when assembling a ligand, you also need to be careful how you decompose a ligand into component fragments. In this blog post I'm going to use a well-known deconstruction study to highlight some of the things that you need to think about when deconstructing.
I like to think of molecular interactions in terms of generic atom types such as 'neutral hydrogen bond acceptor' or cationic hydrogen bond donor'. This is a pharmacophoric view of molecular recognition which is also relevant to bioisosterism and scaffold-hopping. Those who take a more physical view of molecular recognition would say that pharmacophoric atom-typing is just cheminformatics and somewhat uncouth. However, you can capture a lot of physics with atom-typing and it's not like placing atomic charges on nuclei is such great physics anyway...
When deconstructing a ligand molecule you want to minimise changes to the way in which a binding site might see atoms in the ligand. For example breaking the carbon-nitrogen bond of an amide and adding hydrogens is not a great idea because you’ll turn hydrogen bond donor into a cation (at physiological pH) and a strong hydrogen bond acceptor into a weaker one. Deconstructions that add or remove hydrogen atoms from nitrogen or oxygen atoms are usually not a good idea. In the featured ligand deconstruction study, fragments 2 and 3 were derived structure from 1. The acyl sulfonamide group of 1 would be expected to be predominantly deprotonated under normal physiological conditions (a pKa of 5.4 has been reported for sulfacetamide). In contrast, fragment 2 would be expected to be predominantly neutral at normal physiological pH (benzenesulfonamide pKa is 10.1). This means that the deconstruction of the acylsulfonamide transforms an anionic nitrogen into a neutral one that is bonded to a donor hydrogen. This makes it difficult to draw conclusions from the observation that the fragment does not bind to the target. Is the interaction between the relevant part of the parent ligand very weak or has the deconstruction changed the pharmacophoric character of interacting atoms?
The deconstruction of 1 to 3 effectively creates a cationic center (a pKa of 5.1 has been reported for dimethylaniline) and having two nitrogen atoms in the piperazine ring does introduce complications. Two pKa values are observed for piperazine and in a recent study these were found to be 9.7 and 5.5 at 298K. This tells us that protonation of one of the nitrogen atoms makes it more difficult to protonate the other one (which makes sense). These measured pKa values also tell us that piperazine will exist predominantly as a monocation at normal physiological pH and the corresponding values for 1,4-dimethylpiperazine are 8.4 and 3.8. If you take a look at the source that I used for the benzenesulfonamide pKa, you'll see that attaching a phenyl ring to a carbon that is bonded to a basic nitrogen will make that nitrogen less basic by about one log unit. Bringing this all together for compound 1 suggests that protonation of piperazine will occur preferentially at the left hand nitrogen (see figure above) and the that the relevant pKa will be about 7.4. Deconstruction to fragment 3 is expected to shift the preferred site of protonation to the other nitrogen.
So what is the protonation state of piperazine when compound 1 binds to its target? The closeness of the likely pKa to normal physiological pH makes it difficult to say and if the relevant proton/lone-pair is directed away from the protein surface then cationic and neutral forms may have similar affinity. At this point, I should mention that I couldn't find the value(s) of the pH at which the NMR experiments were performed (if this information is indeed there, I'll invoke the 'reading PDF on my computer defense') and the information really needs to be communicated in a study such as this one.
I'll mention a couple of other deconstructions to illustrate the point that changing an element may sometimes result in less perturbation of the relevant substructures. Fragment 4 gets round the problem of deconstruction shifting the preferred site of protonation. The nitrogen atom in the parent molecule that is mutated into carbon will be a weak hydrogen bond acceptor because it is linked directly to an aromatic ring. It can be argued that mutating a weak hydrogen bond acceptor into a hydrophobic atom represents a smaller perturbation than mutating it into a cationic center. However, piperidine is more basic than piperazine so there will be less neutral form (which may or may not be relevant). Deconstruction to fragment 5 preserves (actually is likely to strengthen) the hydrogen bond acceptor character of the less basic piperazine nitrogen but is likely to decrease the amount of cationic form because morpholine is less basic than piperazine.
Hopefully this will have got you thinking in a bit more depth about ligand deconstruction and I'll finish off with a cartoon of how we might use deconstruction in lead optimisation. First we check that we can can actually measure affinity for a fragment that is obtained by deconstructing the lead compound. Then we assemble SAR (could be a good way to explore bioisosteric replacements) before incorporating the best fragments into the lead structure. Essentially, the fragment assay allows us to assemble SAR in a more accessible region of chemical space.
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Milletti, Storchi, Goracci, Bendels, Wagner, Kansy, Cruciani, Extending pKa prediction accuracy: High-throughput pKa measurements to understand pKa modulation of new chemical series. Eur. J. Med. Chem. 2010, 45, 4270-4279. DOI
Fickling, Fischer, Mann, Packer & Vaughan, Hammett Substituent Constants for Electron-withdrawing Substituents : Dissociation of Phenols, Anilinium Ions and Dimethylanilinium Ions. JACS 1959,81, 4226-4230. DOI
Khalili, Henni, East, pKa Values of Some Piperazines at (298, 303, 313, and 323) K. J. Chem. Eng. Data 2009, 54, 2914-2917. DOI