A couple of articles appearing in the last few months brought my attention to a topic that medicinal chemists don't always think about and need to pay more attention to; the important role of kinetics in drug discovery, especially in its early stages.
Anyone who is involved in drug discovery knows the importance of the dissociation constant that signifies the affinity of a therapeutic ligand for a protein. SAR around changing affinities (usually represented by Kd or IC50 values) drive lead design and optimization. But as some of the recent reviews note, the problem with this number is that it's a ratio of the on and off rates of binding of the ligand to the protein. A fast on rate and a fast off rate will give you the same number as a slow on and a slow off rate. But the two situations are not identical.
The most important point emphasized by these reviews is that slow off rates can sometimes lead to prolonged drug efficacy in ways that are not apparent from just the affinity. And this is quite logical if you consider that a slow off rate means that a ligand has a longer residence time in the protein's active site and is spending more time modulating its action. What this means in practice is that even compounds with relatively low affinities can have quite significant efficacies resulting from slow off rates.
So how do you modulate off rates? One good thing about off rates is that unlike on rates, they don't depend on concentration. The benefit of this is that you could have a compound which has a low concentration at the target site and is rapidly cleared away, but which nonetheless spends a lot of time in the protein and therefore provides good efficacy. The other good thing about off rates is that they are essentially dependent on the interactions between ligand and target. So you should in principle be able to improve them just by optimizing these interactions. Again, this won't result in better affinity if you are also slowing down on rates, but it might give you improved efficacy.
In part these studies remind us that we need to clearly distinguish between terms like affinity, IC50, efficacy and the other vocabulary of lingua pharmaceutica. But they also ask an important question; why aren't pharmaceutical scientists paying more attention to kinetic measurements in the early stages of drug discovery? That this is indeed the case became apparent when I looked at the website BindingDB which lists key biological, thermodynamic and kinetic parameters for ligands bound to popular targets. One prominent pharmaceutical target that I looked at had 277 different ligand structures bound to it along with many cases where affinities, IC50s and even free energies had been measured. But out of those 277 I could find only 5 cases (less than 2%) where on and off rates had been recorded. Clearly this is not a focus in preclinical drug discovery.
But as the recent article note, it should be. There are several cases of drugs - HIV protease inhibitors for instance - where differing efficacies for compounds with similar affinities essentially result from differing off rates and residence times. In fact as illustrated by the blood pressure lowering drug amlodipine, off rates can mean the difference between a best-in-class drug and the second-best contender; amlodipine is a better drug than others partly because of its longer residence time in the pocket of the calcium channel protein which it inhibits.
The neglect of kinetic rate measurements reminds me of an almost equal neglect of thermodynamic measurements by ITC. As described in other articles, careful measurement of enthalpy and entropy (and not just free energy) can be very useful in early stage drug discovery. This shouldn't be surprising at all; after all kinetics and thermodynamics are the twin pillars of protein-ligand binding, and you neglect them at your own peril.
Anyone who is involved in drug discovery knows the importance of the dissociation constant that signifies the affinity of a therapeutic ligand for a protein. SAR around changing affinities (usually represented by Kd or IC50 values) drive lead design and optimization. But as some of the recent reviews note, the problem with this number is that it's a ratio of the on and off rates of binding of the ligand to the protein. A fast on rate and a fast off rate will give you the same number as a slow on and a slow off rate. But the two situations are not identical.
The most important point emphasized by these reviews is that slow off rates can sometimes lead to prolonged drug efficacy in ways that are not apparent from just the affinity. And this is quite logical if you consider that a slow off rate means that a ligand has a longer residence time in the protein's active site and is spending more time modulating its action. What this means in practice is that even compounds with relatively low affinities can have quite significant efficacies resulting from slow off rates.
So how do you modulate off rates? One good thing about off rates is that unlike on rates, they don't depend on concentration. The benefit of this is that you could have a compound which has a low concentration at the target site and is rapidly cleared away, but which nonetheless spends a lot of time in the protein and therefore provides good efficacy. The other good thing about off rates is that they are essentially dependent on the interactions between ligand and target. So you should in principle be able to improve them just by optimizing these interactions. Again, this won't result in better affinity if you are also slowing down on rates, but it might give you improved efficacy.
In part these studies remind us that we need to clearly distinguish between terms like affinity, IC50, efficacy and the other vocabulary of lingua pharmaceutica. But they also ask an important question; why aren't pharmaceutical scientists paying more attention to kinetic measurements in the early stages of drug discovery? That this is indeed the case became apparent when I looked at the website BindingDB which lists key biological, thermodynamic and kinetic parameters for ligands bound to popular targets. One prominent pharmaceutical target that I looked at had 277 different ligand structures bound to it along with many cases where affinities, IC50s and even free energies had been measured. But out of those 277 I could find only 5 cases (less than 2%) where on and off rates had been recorded. Clearly this is not a focus in preclinical drug discovery.
But as the recent article note, it should be. There are several cases of drugs - HIV protease inhibitors for instance - where differing efficacies for compounds with similar affinities essentially result from differing off rates and residence times. In fact as illustrated by the blood pressure lowering drug amlodipine, off rates can mean the difference between a best-in-class drug and the second-best contender; amlodipine is a better drug than others partly because of its longer residence time in the pocket of the calcium channel protein which it inhibits.
The neglect of kinetic rate measurements reminds me of an almost equal neglect of thermodynamic measurements by ITC. As described in other articles, careful measurement of enthalpy and entropy (and not just free energy) can be very useful in early stage drug discovery. This shouldn't be surprising at all; after all kinetics and thermodynamics are the twin pillars of protein-ligand binding, and you neglect them at your own peril.
