Tracking Drugs and Lipids: Quantitative Mass Spectrometry Imaging of Liposomal Doxorubicin Delivery and Bilayer Fate in Three-Dimensional Tumor Models

Targeted therapy to the tumor would greatly advance precision
medicine.
Many drug delivery vehicles have emerged, but liposomes are cited
as the most successful to date. Recent efforts to develop liposomal
drug delivery systems focus on drug distribution in tissues and ignore
liposomal fate. In this study, we developed a novel method to elucidate
both drug and liposomal bilayer distribution in a three-dimensional
cell culture model using quantitative matrix-assisted laser desorption/ionization
mass spectrometry imaging (MALDI qMSI) alongside fluorescence microscopy.
Imaging liposomal distribution in a cell culture model is challenging,
as lipids forming the bilayer are endogenous to the model system.
To resolve this issue, we functionalized the bilayer by chemically
cross-linking a fluorescent tag to the alkyne-containing lipid hexynoyl
phosphoethanolamine (HPE). We synthesized liposomes incorporating
the tagged HPE lipid and encapsulated within them doxorubicin, yielding
a theranostic liposome capable of both drug delivery and monitoring
liposomal uptake. We employed an “in-tissue” MALDI qMSI
approach to generate a calibration curve with R2 = 0.9687, allowing for quantification of doxorubicin within
spheroid sections at multiple time points. After 72 h of treatment
with the theranostic liposomes, full doxorubicin penetration was observed.
The metabolites doxorubicinone and 7-deoxydoxorubicinone were also
detected after 48 h. Modification of the bilayer allowed for fluorescence
microscopy tracking of liposomes, while MALDI MSI simultaneously permitted
the imaging of drugs and metabolites. While we demonstrated the utility
of our method with doxorubicin, this system could be applied to examine
the uptake, release, and metabolism of many other liposome-encapsulated
drugs.