Dell'Isola, Mario (2025) Chemical Biology approaches to disclose TREM2 functions. [Tesi di dottorato]

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Abstract

Currently, TREM2 is recognized as one of the most promising therapeutic targets for neurodegenerative diseases. Nevertheless, substantial gaps persist in elucidating its mechanism of action, largely due to the limited identification and characterization of its putative agonists, hence its classification as an “orphan receptor.” The main goal of this PhD project is to develop innovative tools to advance the mechanistic understanding of TREM2 through the identification of novel endogenous ligands. Building upon prior evidence from the synthetic sulfolipid Sulfavant A (SULF A), the first synthetic reported TREM2 ligand, a search for structurally related endogenous molecules had been conducted in the brains of young mice, where TREM2 is predominantly expressed by microglial cells (brain resident macrophage-like cells). Comprehensive mass spectrometry analyses, supported by MZmine-assisted deconvolution of raw LC-MS/MS data, enabled the identification of three sulfated lipid classes, including sulfatides, seminolipids, and SGDG. Functional assays, including TREM2-reporter and phagocytosis tests, pointed out the seminolipids as candidate TREM2 ligand, eliciting cellular responses comparable to SULF-A. This project investigated also the development of biochemical tools to elucidate the TREM2 signalling. In this context, the inhibitory peptide IA9 was examined as a biochemical probe and experimental findings revealed that its biological activity is strongly influenced by the nature of the associated counterion, which modulates its structural conformation and functional properties. Taken together, this work represents a pioneering effort to unravel the molecular mechanisms governing TREM2 activation and signalling. It establishes a conceptual and methodological framework for future studies aimed at exploring TREM2-mediated pathways and therapeutic strategies. Seminolipids emerge as valuable molecular tools for dissecting TREM2 function and its role in neuroinflammatory contexts, both in vitro and in vivo. Conversely, the IA9-K peptide will require further refinement to define the optimal conditions for its application as a selective TREM2 inhibitor in functional studies.

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