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Abstract

Macrophage activation plays a pivotal role in the immune response, with the M1-like and M2-like phenotypes orchestrating pro-inflammatory and anti-inflammatory actions, respectively. The dynamic ability of macrophages to shift between these states underpins their significance in pathophysiological conditions, including cancer, where repolarisation towards an M1-like state could potentiate anti-tumour immunity. This study aimed to explore the repolarisation potential of murine bone marrow derived macrophages (BMDMs) and human monocyte derived macrophages (MdMs), focusing on identifying and testing functional molecules capable of inducing shifts toward an M1-like activation state. Through RNA and miRNA sequencing, shared and species-specific pathways in gene expression were identified, facilitating the selection small molecule inhibitors (SMIs) targeting transcription regulators associated with the M2-like phenotype and miRNAs over-expressed in M1-like macrophages and in silico analysed to target transcription regulators. My results revealed that miR-155-5p, along with a pool of SMIs targeting Stat6, Myc, Stat3, and HDACs, effectively repolarised macrophages towards an M1-like phenotype, albeit with notable differences in the extent of repolarisation and functional outcomes between murine and human models. Functional assays assessing the anti-tumour activity and T-cell activation potential of repolarised macrophages demonstrated that IFN-γ+LPS treatment significantly enhanced these capabilities in both species. However, despite molecular indications of repolarisation with miR-155- 5p and the inhibitor pool, these did not translate into expected functional outcomes, highlighting the complexity of macrophage biology and the nuanced interplay of molecular and functional repolarisation mechanisms. This study contributes significantly to our understanding of macrophage polarisation and repolarisation, emphasising the translational relevance of shared pathways across species and identifying potential therapeutic targets for manipulating macrophage states. The findings underscore the importance of evaluating both molecular changes and functional implications of repolarisation strategies, paving the way for the development of targeted therapies that leverage the immune system’s intrinsic anti-tumour potential.