# Mass Spectrometry Imaging Reveals the Distribution of a GABRG2 Targeting Antisense Oligonucleotide and Its Functional Effect in Rat Brain

## TL;DR

This study uses mass spectrometry imaging to track an antisense oligonucleotide in rat brain and kidney, showing its effects on neurotransmitters, lipids, and proteins.

## Contribution

A novel multiomics mass spectrometry imaging approach is developed to map ASO distribution and its molecular effects in brain tissue.

## Key findings

- An optimized MALDI-MSI protocol successfully visualized phosphorothioate-modified ASOs in rat brain and kidney.
- ASO treatment altered levels of multiple neurotransmitters, lipids, and synaptic-related proteins.
- The spatial omics approach revealed comprehensive molecular changes linked to ASO targeting of GABRG2.

## Abstract

In recent years, the development of antisense oligonucleotides
(ASOs) has gained wide interest as therapeutic agents for their potential
in treating neurodegenerative diseases. ASOs are chemically modified
oligonucleotides that are designed to bind complementary regions of
RNA or DNA and, thereby, modulate the expression of the corresponding
protein. Here, we present a multiomics approach to investigate the
spatial distribution and biological effect of an ASO designed to target
the mRNA that translates for γ-aminobutyric acid A receptor
γ2 subunit (GABRG2), which is abundantly expressed within the
brain. In this study, a rat model was used to develop a multiomics
mass spectrometry (imaging) approach to map ASO distribution in brain
and kidney, followed by in-depth analysis of the lipidome, proteome,
and metabolome. The ASOs’ phosphorothioate-modified backbone
was visualized using an optimized matrix-assisted laser desorption/ionization
mass spectrometry imaging (MALDI-MSI) protocol, which included the
introduction of an organic washing step prior to MALDI-MSI acquisition
and an optimized acquisition method. On consecutive tissue sections,
reactive matrix FMP10 was applied to enable the visualization of neurotransmitters,
which revealed significant alterations for multiple neurotransmitters.
Lastly, on the same slide, the ASOs’ effect on the lipidome
and proteome of the brain was further analyzed. Proteins corresponding
to synaptic activity and plasticity were mainly affected by the ASO.
This spatial omics approach provides insight into the comprehensive
molecular landscape of ASO-mediated interventions and their promise
as treatments for neurological disorders.