Detection of Protein and Metabolites in Cancer Analyses by MALDI 2000–2025

Cancer metabolomics has become a powerful way of understanding tumor biology, identifying biomarkers and metabolites, and helping precision oncology. Matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS), among many other analytical platforms, has gained popularity over the past two and a half decades due to its unique ability of directly analyzing metabolites in tissue with spatial resolution. This review will study 2000-2025 MALDI-based strategies for cancer metabolite detection, spanning from early proof-of-concept protein profiling to the development of high-resolution MALDI-MS imaging (MALDI-MSI), which is capable of mapping thousands of metabolites at near single-cell resolution. Its applications include the differentiation of tumor versus normal tissue, discovery of stage and subtype specific biomarkers, mapping of metabolic heterogeneity, and the visualization of drug metabolism in situ. Breakthrough technological milestones, such as the advanced matrices, on-tissue derivatization, MALDI-2 post-ionization, and the integration with Orbitrap or Fourier-transform ion cyclotron resonance (FT-ICR) platforms, have significantly improved the overall sensitivity, metabolite coverage, and spatial fidelity. Clinically, MALDI-MS has shown its purpose in breast, prostate, colorectal, lung, and liver cancers by providing metabolic fingerprints that are linked to tumor microenvironments, hypoxia, and therapeutic response. However, challenges such as the inclusion of matrix interface with low-mass metabolites, limited quantitation, ion suppression, and the lack of standardized procedures do not yet allow for the transition from translation to routine diagnostics. Even with these hurdles, the future of MALDI-MS in oncology remains in a good position with major advancements in multimodal imaging, machine learning-based data integration, portable sampling devices, and clinical validation studies that are pushing the field towards precision treatment.