Determinants of drug-target interactions at the single cell level.

Vlad Elgart,Jia-Ren Lin,Joseph Loscalzo,Turkan Haliloglu

doi:10.1371/journal.pcbi.1006601

Vlad Elgart, Jia-Ren Lin + Show 2 more

Open Access

https://doi.org/10.1371/journal.pcbi.1006601

Copy DOI

Abstract

The physiochemical determinants of drug-target interactions in the microenvironment of the cell are complex and generally not defined by simple diffusion and intrinsic chemical reactivity. Non-specific interactions of drugs and macromolecules in cells are rarely considered formally in assessing pharmacodynamics. Here, we demonstrate that non-specific interactions lead to very slow incorporation kinetics of DNA binding drugs. We observe a rate of drug incorporation in cell nuclei three orders of magnitude slower than in vitro due to anomalous drug diffusion within cells. This slow diffusion, however, has an advantageous consequence: it leads to virtually irreversible binding of the drug to specific DNA targets in cells. We show that non-specific interactions drive slow drug diffusion manifesting as slow reaction front propagation. We study the effect of non-specific interactions in different cellular compartments by permeabilization of plasma and nuclear membranes in order to pinpoint differential compartment effects on variability in intracellular drug kinetics. These results provide the basis for a comprehensive model of the determinants of intracellular diffusion of small-molecule drugs, their target-seeking trajectories, and the consequences of these processes on the apparent kinetics of drug-target interactions.

Highlights

Drug efficacy in vivo is notoriously difficult to predict owing, in part, to the complexity of the underlying biochemical processes that govern drug–target interactions in situ
The extent to which these nonspecific interactions influence the availability and action of the drug for its specific target depends upon the relative concentrations of drug, the specific target, and nonspecific targets
Using small-molecule binding to DNA and single cell monitoring, we demonstrate that this effect results in apparently anomalous small molecule-DNA binding kinetics in cells

Summary

Introduction

Drug efficacy in vivo is notoriously difficult to predict owing, in part, to the complexity of the underlying biochemical processes that govern drug–target interactions in situ. Semi-empiric pharmacokinetic/pharmacodynamic (PK/PD) models typically describe accumulation of the drug in tissue(s) and, do not address the question of variability in efficacy for individual cells, which is determined by the drug’s access to and interaction with its target(s) within a cell. Variability in drug efficacy may, be a key factor driving resistance, selection, and toxicity. Our model system is a monolayer cell culture that allows continuous monitoring of drug binding to its target in individual cells. While this model system is, far from ideal, provided that the free drug concentration in a given tissue is fairly uniform, cell culture experiments can meaningfully address the question of heterogeneity of response in a cell population

Methods

Results

Discussion

Conclusion