Development Of Small-molecule Probes For HERG Potassium Channel

The potassium channels encoded by the hERG(human ether-a-go-go related gene)obtained from the human hippocampal nerve are rapidly delayed rectifying type.hERG potassium channels are widely distributed in human cardiac tissue and nerve tissue.The reason of some drugs causing severe myocardial toxicity is that they can inhibit hERG channel and,result in long QT syndrome;the mutation of hERG gene will also lead to inherited long QT syndrome,which both can make patients with arrhythmia,even lead to death.Moreover,it was found that hERG channels are distributed in a large amount of tumor cells,and they are in related with some progresses.In these tumor cells,hERG channels can be used as a marker for tumor progression.At present,the research methods of hERG channel mainly include:patch clamp analysis technique,radioactive ligand competition method,and fluorescence analysis technology.The patch clamp analysis technique is the gold standard of hERG channel,but the technique of patch clamp is very expensive and time-consuming.A large amount of radioactive material is required in radioactive ligand competition method,which may cause radiation.In recent years,fluorescence analysis technology has developed rapidly,which plays an important role in the research of life science.Small molecular fluorescent probe has some advantages,such simple synthesis,small volume,high cell permeability,high sensitivity and specificity,so it has been widely used in research,mainly used to study the mechanism of biological macromolecules,cellular stains and environmental indicator.Small molecule fluorescent probe generally includes three parts:recognition groups,connection chains and fluorophore,recognition groups can be able to combine targets specifically,fluorophore can be able to switch the combining signal into fluorescent signal,link chain connect fluorophore and recognition groups.In this paper,the specific hERG channel probes that have two different switch mechanisms were designed on the basis of the reported inhibitors of the hERG channel,asmidazole and E-4031.Among them,we select astemizole as recognition groups,use of environmentally sensitive fluorophore dansyl chloride and naphthalene diimide to replace methoxy phenyl of astemizole,and then through the aliphatic chain connect identify groups and fluorophore to design the probe M1-M3 that have environment-sensitive fluorescent switch mechanism.When the probes are in freestate,the fluorescence intensity is weak,but the fluorescence intensity is significantly higher than the free state when it combined with the hydrophobic pocket.The probe M4-M7 adopted aggregation-induced emission(AIE)fluorescent switch mechanism,astemizole is selected as recognition group in these probes,but considering the volume of four phenyl vinyl structure is a bit big,so E-4031 is also selected as recognition group because of its small volume.Then the "click" reaction between the azoyl group on the fluorescence group and the acetylene group on the recognition group was carried out by copper(Ⅰ).When the probe is in freestate,the aromatic rings can rotate freely,and the molecules in excited state release energy in a non-radiative manner.However,the free rotation of the aromatic rings is limited after the molecular aggregation,and the excited molecules mostly release energy as fluorescence.The small molecule probes for hERG channel were conducted a series of evaluation(optical properties,the affinity for hERG channel,cell toxicity,specificity and microscopic imaging),the results show that the synthesis of small molecule probe has good optical properties,small cell toxicity,and high affinity for hERG channel,so that they can be applied to the microscopic imaging of hERG channel.In the future studies,we will use small molecule probes to explore the relationship between the up-regulation of hERG channel and the tumor to study the mechanism in the tumor for the study of tumorigenesis and antitumor drugs.At the same time,we will design a series of small molecular probes that have near infrared fluorophore,and would be applied to the imaging of hERG channels at mice.