The peptide HsTX1[R14A] is a potent and selective blocker of the voltage-gated potassium channel KV1.3, which is a highly promising target for the treatment of autoimmune diseases and other conditions.1,2 However, there is limited information on its pharmacokinetics and biodistribution in mice, both of which are important given that many preclinical efficacy studies are conducted in mice.
A bioanalytical method for the quantitative determination of HsTX1[R14A] in C57BL/6J mouse plasma was developed using liquid chromatography-tandem mass spectrometry (LC-MS/MS). The validated method was linear over the concentration range 6.7-66.7 nM for HsTX1[R14A]; and the lower limit of quantitation was 6.7 nM. The average precision and accuracy were <5.4% and <13.5%, respectively. The entire analysis time was only 5 min per sample. The currently developed LC-MS/MS assay will be applied to assess the pharmacokinetics of HsTX1[R14A] in mice following intravenous administration.
In addition, a radiotracer was developed to enable positron emission tomography (PET) studies of peptide distribution in mice over a period of days. HsTX1[R14A] was synthesised1 with a DOTA tag coupled to the N-terminus via a mini PEG, aminoethyloxyethyloxyacetyl (AeeA) linker and radiolabelled with copper-64. The radiochemical purity and in vitro stability of [64Cu]Cu-DOTA-HsTX1[R14A] will be determined by radio-HPLC, prior to administration to mice and PET will be employed to monitor biodistribution.
These analytical methods (LC-MS/MS and PET) can now be applied to evaluate the plasma pharmacokinetics and biodistribution of HsTX1[R14A] in mice in both healthy and diseased states.