MSACL 2026 Abstract

Boronic Acid Mass Tag for Spatially Resolved Glucose Detection by MALDI MSI: Application to Prostate Cancer Spheroids

Justine Gatein (1), Nassim Maarouf-Mesli (1), Dzila Aharoutian (1), Sarah Ghasemi (2), Thomas Gervais (2), William D. Lubell (1) and Pierre Chaurand (1)
(1) Département de Chimie, Université de Montréal, Montréal, Québec, Canada, (2) Département de génie physique, Polytechnique Montréal, Montréal, Québec, Canada

Justine Gatein (Presenter) Université de Montréal

Presenter Bio: I am a PhD candidate in analytical chemistry at the Université de Montréal, where I develop MALDI-based mass spectrometry imaging (MSI) approaches for the spatial characterization of biological tissues. My doctoral research, conducted in Prof. Pierre Chaurand's laboratory, focuses on small molecule analysis and methodological development for tissue imaging applications, to understand metabolic reprogramming in cancer.
I am also completing a PharmD thesis at the Université Paul Sabatier (Toulouse III) under the supervision of Dr. Anthony Lemarié. This work explores the contribution of MSI to the understanding of tumoral mechanisms and the identification of novel therapeutic targets in glioblastoma.
Prior to my doctoral studies, I completed a Master's degree in analytical chemistry for drugs and natural products at the Université de Bordeaux in a work-study program at Sanofi, where I was involved in analytical method comparison and validation, supplier qualification, and pharmacopoeial monograph updates. My background originates in pharmaceutical sciences, at the Université Paul Sabatier (Toulouse III), with clinical training in pharmacokinetics and therapeutic drug monitoring at the CHU de Toulouse.

Relevant Financial Disclosures (within past 24 months, reported on May 01, 2026) No relevant financial relationship(s) to disclose.

INTRODUCTION:
Prostate cancer (PCa) affects 1 in 4 men in North America and remains one of the leading causes of cancer-related death in its aggressive forms. A key driver of this aggressiveness is the metabolic reprogramming that occurs within the tumor microenvironment: cells of the hypoxic core develop intense glycolytic activity and engage the glycerol shunt to generate glycerol, a highly diffusible metabolite redistributed toward the proliferative periphery as a fuel source. Understanding this spatial metabolic heterogeneity is essential, as it underlies both tumor progression and resistance to treatment. However, conventional bulk metabolomic approaches lack the spatial resolution required to capture this intratumoral gradient. Matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI MSI) could address this challenge, offering label-free, spatially resolved metabolite detection directly in tissue. Its main limitation for glucose, a central metabolite in this reprogrammed network, is poor ionization efficiency. On-tissue chemical derivatization (OTCD) overcomes this limitation by applying on the tissue section a charged chemical mass tag that selectively reacts with targeted metabolites to enhance their desorption/ionization. Boronic acids are particularly well-suited for glucose derivatization, as the boron group reacts selectively with cis-diol functions. To study glucose metabolism in a clinically relevant context, three-dimensional tumor spheroids derived from PCa cell lines offer a powerful alternative to animal models, faithfully replicating the hypoxic gradients of aggressive tumors.

METHODS:
A new boronic acid mass tag with a permanent positively charged amine function was synthesized and optimized for glucose derivatization, first in solution and then by dry droplet deposition on mouse brain homogenate. Spray deposition parameters were subsequently optimized for both the boronic acid tag and the 2,5-DHA matrix on tissue sections prior to MSI. For validation, OTCD-MALDI MSI was performed in positive ion mode on kidney, ovary and brain cryosections (12 µm). Glucose signal enhancement was determined by comparison to direct MALDI MSI using NEDC matrix in negative ion mode. The boron isotopic signature, exact mass, and MS/MS fragmentation were used to confirm glucose identity. Jumbo (~500 µm diameter) and small (~380 µm diameter) PCa spheroids were generated from C4-2B human prostate cancer cell lines using microfluidic devices. Frozen spheroids were then embedded in OCT blocks, cryosectioned (12 µm thick) and analyzed by MALDI MSI with the optimized OTCD and matrix deposition methods. All acquisitions were performed on a Shimadzu MALDI-iMScope QT system.

RESULTS:
Glucose was successfully detected in kidney, ovary and brain tissue sections by MALDI MSI at a spatial resolution of 25 µm using NEDC as matrix ([glucose+Cl]-, m/z = 215) and after OTCD with boronic acid ([glucose-boronic acid]⁺, m/z = 401) using 2,5-DHA as matrix with a mass accuracy of ~1 ppm. The identity of the glucose-boronic acid adduct was confirmed by exact mass and MS/MS fragmentation. No analyte delocalization was observed during either the OTCD reaction or matrix spray deposition, ensuring the spatial integrity of the detected signal. For brain, glucose was predominantly detected in gray matter, consistent with its role as the primary site of metabolic activity. For kidney, glucose was mostly detected in the inner medulla, and in the medulla for ovaries. When comparing signal intensities, OTCD proved to be approximately 10-fold more sensitive than direct NEDC-based detection. The OTCD method also allows higher spatial resolution imaging at 10 and 5 µm, which was not achievable with the standard NEDC method. MALDI MSI of glucose in PCa spheroids was successfully performed at 25 µm spatial resolution. Glucose signal appears lowest in the hypoxic core and at the periphery, and highest at the interface between these two zones.

CONCLUSION:
These results demonstrate that OTCD using a boronic acid mass tag provides a sensitive and spatially resolved method for glucose imaging by MALDI MSI. The spatial distribution observed in PCa spheroids, with a glucose signal concentrated at the core-periphery interface, is consistent with the known metabolic organization of hypoxic tumors. In PCa spheroids, this spatial distribution likely reflects the metabolomic activity of proliferating cells in the oxygenated periphery, while hypoxic core relies on alternative energy sources, consistent with the metabolomic reprogramming describe in PCa. These findings open perspectives for further investigation of metabolomic reprogramming in PCa and its implications for tumors progression and therapeutic resistance. Beyond PCa, this approach is broadly applicable to any pathology involving glucose metabolism dysregulation, opening perspectives for the study of tumor heterogeneity and histological distribution.