Discovery of Nanomolar Desmuramylpeptide Agonists of the Innate Immune Receptor Nucleotide-Binding Oligomerization Domain-Containing Protein 2 (NOD2) Possessing Immunostimulatory Properties

Abstract

Muramyl dipeptide (MDP), a fragment of bacterial peptidoglycan, has long been known as the smallest fragment possessing adjuvant activity, on the basis of its agonistic action on the nucleotide-binding oligomerization domain-containing protein 2 (NOD2). There is a pressing need for novel adjuvants, and NOD2 agonists provide an untapped source of potential candidates. Here, we report the design, synthesis, and characterization of a series of novel acyl tripeptides. A pivotal structural element for molecular recognition by NOD2 has been identified, culminating in the discovery of compound 9, the most potent desmuramylpeptide NOD2 agonist to date. Compound 9 augmented pro-inflammatory cytokine release from human peripheral blood mononuclear cells in synergy with lipopolysaccharide. Furthermore, it was able to induce ovalbumin-specific IgG titers in a mouse model of adjuvancy. These findings provide deeper insights into the structural requirements of desmuramylpeptides for NOD2-activation and highlight the potential use of NOD2 agonists as adjuvants for vaccines.

Harnessing the untapped potential of nucleotide‐binding oligomerization domain ligands for cancer immunotherapy

“…The finding that the presence of the N-acetylmuramyl moiety is not necessary for the immunomodulatory properties of MDP led to the design and synthesis of a new class of MDP derivatives, termed desmuramylpeptides. [195][196][197][198][199][200][201][202] Desmuramylpeptides are devoid of the N-acetylmuramyl moiety and have therefore more lipophilic character than MDP. This class contains compounds that are able to enhance host defense against microbial infections as well as exhibit strong adjuvant activity and, even, remarkable antitumor potency.…”

Translation of peptidoglycan metabolites into immunotherapeutics

“…The MDP derivative 11 in which MurNAc was replaced with an indole‐2‐ylcarboxamido group showed similar activity as murabutide in both NF‐κB reporter and cytokine secretion assays . The related trans ‐feruloyl analog 12 exhibited low nanomolar activation (EC 50 = 46 n m ), providing functional MDP analogs with equivalent or even more potent activity without the hydrolysable reducing sugar …”

“…B30‐MDP also increased immunisation in mice using X‐irradiated L5178Y‐ML25 lymphoma cells and prevented subsequent metastases better than vaccination with the irradiated cells alone . More recently, liposomal‐encapsulated desmuramyl compound 12 showed an increase in serum levels of antigen‐specific IgG upon OVA peptide vaccination; yet, its effects were less pronounced than MDP . Unfortunately, these preclinical observations have not directly translated to the clinic.…”

“…79 The related transferuloyl analog 12 exhibited low nanomolar activation (EC 50 = 46 nM), providing functional MDP analogs with equivalent or even more potent activity without the hydrolysable reducing sugar. 80…”

“…96 More recently, liposomal-encapsulated desmuramyl compound 12 showed an increase in serum levels of antigenspecific IgG upon OVA peptide vaccination; yet, its effects were less pronounced than MDP. 80 Unfortunately, these preclinical observations have not directly translated to the clinic. The muramyl tripeptide-phosphatidylethanolamine conjugate MTP-PE 15 had been explored in Phase I clinical trials as a vaccine adjuvant for both influenza and human immunodeficiency virus type 1 (HIV-1), but its addition to the vaccines demonstrated significantly increased adverse side effects with little to no improvement of immunogenicity.…”