On the different experimental manifestations of two‐state ‘induced‐fit’ binding of drugs to their cellular targets

Abstract

'Induced-fit' binding of drugs to a target may lead to high affinity, selectivity and a long residence time, and this mechanism has been proposed to apply to many drugs with high clinical efficacy. It is a multistep process that initially involves the binding of a drug to its target to form a loose RL complex and a subsequent isomerization/conformational change to yield a tighter binding R'L state. Equations with the same mathematical form may also describe the binding of bivalent antibodies and related synthetic drugs. Based on a selected range of 'microscopic' rate constants and variables such as the ligand concentration and incubation time, we have simulated the experimental manifestations that may go along with induced-fit binding. Overall, they validate different experimental procedures that have been used over the years to identify such binding mechanisms. However, they also reveal that each of these manifestations only becomes perceptible at particular combinations of rate constants. The simulations also show that the durable nature of R'L and the propensity of R'L to be formed repeatedly before the ligand dissociates will increase the residence time. This review may help pharmacologists and medicinal chemists obtain preliminary indications for identifying an induced-fit mechanism. Abbreviations[L], concentration of ligand in bulk of solution; RL and R'L, loose and tight binding ligand-receptor complexes; △G o , difference in Gibbs free energy between R and R'L; k 1 , k 2 … k 1A … k 1M …, microscopic rate constants; k obs , k on , k off , macroscopic rate constants for association and dissociation; K D , 'thermodynamic' equilibrium dissociation constant based on △G o ; K D *, [L] at which this observed binding is half-maximal at equilibrium; app K D *, [L] at which the observed binding is half-maximal under earlier non-equilibrium conditions; p, -LogThese Tables list key protein targets and ligands in this article which are hyperlinked to corresponding entries in http://www.guidetopharmacology. org, the common portal for data from the IUPHAR/BPS Guide to PHARMACOLOGY (Pawson et al., 2014) and are permanently archived in the Concise Guide to PHARMACOLOGY 2015/16 (Alexander et al., 2015.

Distinct in vivo target occupancy by bivalent‐ and induced‐fit‐like binding drugs

“…The model (Figure A) and equations (4 and 5 in Figure B) that are dedicated to genuine induced‐fit binding also apply to bivalent drugs or ligands (Vauquelin et al, , ). In both scenarios, the equations allow long residence to be achieved, albeit by different means.…”

“…Such plots are linear with k obs = k off + k on . [L] for one‐step binding drugs but hyperbolic for induced‐fit‐like binders such as Drug B (denoted as ‘F’ in Figure A of Vauquelin et al, ). This precludes the utilization of k on as a simple second‐order rate constant.…”

“…To this end, their (k 2 .k 4 )/(k 1 .k 3 ) ratio is arbitrarily set to 4.10 −9 M, and k 1 is also kept constant at 1.10 6 M −1 ·min −1 , as in Vauquelin et al . (). The binding properties are then only controlled by k 2 and k 3 (because k 4 = 4.10 −3 .k 3 /k 2 ·min −1 ).…”

“…Equation 5: K D * is the macroscopic/pseudo affinity constant; it equals [L] when the occupancy (i.e. [RL] + [R′L]) is half maximal at equilibrium (Vauquelin et al, ). Equation 6 is an alternative formulation of Equation 5; it emphasizes the equivalence of the (k 3 + k 4 )/k 2 and k 4 /(k 1 .…”

Effects of target binding kinetics on in vivo drug efficacy: koff, kon and rebinding

“…With respect to the PD models, multi-step 'induced fit'-type binding mechanisms are now rather considered to play a key role for achieving high affinity and clinical efficacy of many drugs (Copeland, 2016). Yet despite of this increased complexity, radioligand washout experiments ( Figure 1B and Vauquelin, 2012) most often yield mono-exponential dissociation curves so that the overall dissociation process can still be satisfactorily described by a 'macroscopic' k off (Neubig et al, 2003;Tummino and Copeland, 2008;Vauquelin et al, 2016). Regardless of the binding mechanism, it is of note that such in vitro experiments still essentially focus on obtaining 'genuine' k off values and, to this end, an excess of unlabelled competing ligand has been routinely added to the washout medium to prevent 'rebinding' of the radioligand ( Figure 1B).…”