Distinctive Immune\nSignatures Driven by Structural\nAlterations in Desmuramylpeptide NOD2 Agonists

Herein we report\non the design, synthesis and biological evaluation\nof a series of nucleotide-binding oligomerization-domain-containing\nprotein 2 (NOD2) desmuramylpeptide agonists. The structural prerequisites\nthat shape both physicochemical and immunomodulatory profiles of desmuramylpeptide\nNOD2 agonists have been delineated. Within this context, we identified <b>3</b>, a butyrylated desmuramylpeptide, as a potent <i>in\nvitro</i> NOD2 agonist (EC<sub>50</sub> = 4.6 nM), exhibiting\nan almost 17-fold enhancement in potency compared to its unsubstituted\ncounterpart <b>1</b> (EC<sub>50</sub> = 77.0 nM). The novel\nset of desmuramylpeptides demonstrate unique <i>in vitro</i> immunomodulatory activities. They elicited cytokine production in\nperipheral blood mononuclear cells (PBMCs), both alone and in conjunction\nwith lipopolysaccharide (LPS). The spermine-decorated <b>32</b> also stimulated the LPS-induced cytotoxic activity (2.95-fold) of\nPBMCs against K562 cancer cells. Notably, the cholesterol-conjugate <b>26</b> displayed anti-inflammatory actions, highlighted by its\ncapacity to convert the inflammatory monocyte subset into an anti-inflammatory\nphenotype. Finally, the eicosapentaenoylated derivative <b>23</b> augmented antigen presentation by mouse bone marrow-derived dendritic\ncells (BMDCs), thus highlighting its potential as a vaccine adjuvant.