Fluorescence activation mechanism and imaging of drug permeation with new sensors for smoking-cessation ligands

eLife

Jonathan H Wang¹  Henry A Lester¹  Aaron L Nichols¹  Amol V Shivange¹  Bruce N Cohen¹  Charlene H Kim¹  Laura Luebbert²  Zack Blumenfeld³  Anand K Muthusamy⁴  Annet EM Blom⁴  Dennis A Dougherty⁴  Chengcheng Fan⁴  Hailey J Knox⁴  Douglas C Rees⁵  Loren L Looger⁶  Philip M Borden⁶  Jonathan S Marvin⁶  Timothy Gallagher⁷  Hugo Rego Campello⁷ 

[1] Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, United States;Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, United States;Institute of Biology, Leiden University, Leiden, Netherlands;Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, United States;Keck School of Medicine, University of Southern California, Los Angeles, United States;Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, United States;Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, United States;Howard Hughes Medical Institute, California Institute of Technology, Pasadena, United States;Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, United States;School of Chemistry, University of Bristol, Bristol, United Kingdom;

关键词: pharmacokinetics;    nicotine;    biosensors;    nicotinic agonists;    iDrugSnFRs;    inside-out pharmacology;    Mouse;   

DOI  :  10.7554/eLife.74648

来源: eLife Sciences Publications, Ltd

【 摘 要 】

Nicotinic partial agonists provide an accepted aid for smoking cessation and thus contribute to decreasing tobacco-related disease. Improved drugs constitute a continued area of study. However, there remains no reductionist method to examine the cellular and subcellular pharmacokinetic properties of these compounds in living cells. Here, we developed new intensity-based drug-sensing fluorescent reporters (iDrugSnFRs) for the nicotinic partial agonists dianicline, cytisine, and two cytisine derivatives – 10-fluorocytisine and 9-bromo-10-ethylcytisine. We report the first atomic-scale structures of liganded periplasmic binding protein-based biosensors, accelerating development of iDrugSnFRs and also explaining the activation mechanism. The nicotinic iDrugSnFRs detect their drug partners in solution, as well as at the plasma membrane (PM) and in the endoplasmic reticulum (ER) of cell lines and mouse hippocampal neurons. At the PM, the speed of solution changes limits the growth and decay rates of the fluorescence response in almost all cases. In contrast, we found that rates of membrane crossing differ among these nicotinic drugs by >30-fold. The new nicotinic iDrugSnFRs provide insight into the real-time pharmacokinetic properties of nicotinic agonists and provide a methodology whereby iDrugSnFRs can inform both pharmaceutical neuroscience and addiction neuroscience.

【 授权许可】

CC BY