Multiplex Molecular Imaging of Drugs and Metabolites in Dosed Tissues by DESI Mass Spectrometry

Library Number:
PSTR134959884
Author(s):
Bindesh Shrestha, Philippa Hart , Mark Towers, Emmanuelle Claude
Source:
Waters
Content Type:
Posters
Content Subtype:
Other Symposium
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Introduction

Liquid chromatography (LC) based assays are gold standards for determining the concentrations of drugs and metabolites in plasma during the drug discovery process. However, in many instances, drug candidates have different effects on various target tissues. Often, there is an asymmetrical relationship between the concentrations of drugs and metabolites in the plasma versus the targeted tissues. Whole-body autoradiography has been utilized to determine the distribution of radiolabeled drugs and their metabolite candidates in preclinical animal models. Information if a drug candidate reached the desired target(s) of preclinical animal models is an essential step in the pharmaceutical discovery. Mass spectrometry imaging (MSI) offers a possibility of mapping several drug molecules without isotope labeling.  Here we showcase the utility of DESI MSI to assess simultaneous detection of drug and metabolites.

Method

One cohort of mice were orally administered with terfenadine, olanzapine, moxifloxacin and erlotinib at the pharmacologically relevant levels of 25, 10, 10 and 25 mg/kg, respectively. A second cohort was dosed with vehicle to act as a control. Liver sections were taken from each cohort at 2 and 6 hours post administration from the euthanized animals. DESI MSI analysis was performed using a DESI source integrated on a quadruple time of flight (Xevo G2-XS Q-Tof, Waters Corporations). High Definition Imaging (HDI) version 1.4 software was used to define DESI imaging experiments, as well as, process and visualize the collected ion images.

Results

Limits of detection (LOD) of drugs were established by using standard drug dilution series spotted on glass slide. The LODs were from range 5pg to 500ng, dependent on compound and background ions. For example, with the same settings and backgrounds, the LOD for olanzapine and erlotinib analyzed were 5 ng and 50pg, respectively. Ion images showing the localization of the drugs and endogenous metabolite ions were generated from a single DESI imaging experiment. Additionally, within the same experiment, we could detect some phase I metabolites from olanzapine, terfenadine and erlotinib. Spatial correlation between detected drugs and phase I metabolites were established by using spatial correlation analysis based on Pearson product-moment correlation coefficient and hierarchical clustering analysis. Molecular identification of metabolites was aided using high mass accuracy, database search, and elemental composition. Regions of interest (ROIs) were drawn on the DESI imaging datasets to compare the relative normalized intensities between sections. In order to obtain the relative quantitative information, a stable isotope labelled compounds could also be added as internal standards in DESI electrospray solvent. Currently, we routinely add an in-line internal standard, such as Le-Enk (200 pmol/μL), for lock mass correction. The concentration of stable isotope labeled compound depends on the ionization efficiency of the molecule, typically between 50 to 200 pmol/μL.

Conclusions

The presented work shows the capability of DESI to simultaneously detect and image various drug molecules and their metabolites at pharmacologically relevant levels. The relative quantitative imaging of those drugs and metabolites are possible using stable isotope labeling. Further work needs to be conducted on targeted experiments with increased sensitivity (e.g. MRM) and quantitative capabilities to further establish DESI-MSI as a molecular imaging tool in drug discovery process.


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