Design and synthesis of NLR and TLR based ligand-antigen conjugates

The mammalian immune system protects, amongst others, against invading pathogens and consists of an innate and adaptive component. The innate system is the first line of defense in which pattern recognition receptors, like TLR2, NOD1 and NOD2 receptors, detect pathogen associated molecular patterns (PAMPS) that are specific for pathogens. PAMPs exhibit a broad structural variety and the exact molecular structures of ligands that bind to the corresponding PRRs are mostly unknown. The research presented in this Thesis is directed to the design, synthesis and immunological evaluation of new NOD1, NOD2 and TLR2 ligands as well as conjugates in which these ligands are covalently bound to an antigenic OVA-derived peptide. The designed NLR and TLR ligand-antigen conjugates contribute to the insight of the processes of the mammalian immune system at molecular level. Further elucidation of pattern recognition receptor acti vation and antigen presentation with these conjugates may eventually result in synthetic vaccine modalities

Design and synthesis of NLR and TLR based ligand-antigen conjugates

The mammalian immune system protects, amongst others, against invading pathogens and consists of an innate and adaptive component. The innate system is the first line of defense in which pattern recognition receptors, like TLR2, NOD1 and NOD2 receptors, detect pathogen associated molecular patterns (PAMPS) that are specific for pathogens. PAMPs exhibit a broad structural variety and the exact molecular structures of ligands that bind to the corresponding PRRs are mostly unknown. The research presented in this Thesis is directed to the design, synthesis and immunological evaluation of new NOD1, NOD2 and TLR2 ligands as well as conjugates in which these ligands are covalently bound to an antigenic OVA-derived peptide. The designed NLR and TLR ligand-antigen conjugates contribute to the insight of the processes of the mammalian immune system at molecular level. Further elucidation of pattern recognition receptor acti vation and antigen presentation with these conjugates may eventually result in synthetic vaccine modalities.UBL - phd migration 201

Design, automated synthesis and immunological evaluation of NOD2-ligand-antigen conjugates

Tumorimmunolog

Lipophilic Muramyl Dipeptide-Antigen Conjugates as Immunostimulating Agents

Tumorimmunolog

Lipophilic muramyl dipeptide-antigen conjugates as immunostimulating agents

Muramyl dipeptide (MDP) is the smallest peptidoglycan fragment capable of triggering the innate immune system through interaction with the intracellular NOD2 receptor. To develop synthetic vaccine modalities composed of an antigenic entity (typically a small peptide) and a molecular adjuvant with well-defined activity, we previously assembled covalent MDP-antigen conjugates. Although these were found to be capable of stimulating the NOD2 receptor and were processed by dendritic cells (DCs) leading to effective antigen presentation, DC maturation-required for an apt immune response-could not be achieved with these conjugates. To improve the efficacy of these vaccine modalities, we equipped the MDP moiety with lipophilic tails, well-known modifications to enhance the immune-stimulatory activity of MDPs. Herein we report the design and synthesis of a lipophilic MDP-antigen conjugate and show that it is a promising vaccine modality capable of stimulating the NOD2 receptor, maturing DCs, and delivering antigen cargo into the MHC-I cross-presentation pathway.Bio-organic Synthesi

Dual Synthetic Peptide Conjugate Vaccine Simultaneously Triggers TLR2 and NOD2 and Activates Human Dendritic Cells

Tumorimmunolog

Novel TLR2-binding adjuvant induces enhanced T cell responses and tumor eradication

Ligands for the Toll-like receptor (TLR) family can induce activation of cells of the innate immune system and are widely studied for their potential to enhance adaptive immunity. Conjugation of TLR2-ligand Pam3CSK4 to synthetic long peptides (SLPs) was shown to strongly enhance the induction of antitumor immunity. To further improve cancer vaccination, we have previously shown that the novel TLR2-L Amplivant (AV), a modified Pam3CSK4, potentiates the maturation effects on murine DCs. In the current study, we further assessed the immunological properties of AV.\nNaïve mice were vaccinated with a conjugate of either Pam3CSK4 or AV and an SLP to assess specific T cell priming efficiency in vivo. The potency of AV and Pam3CSK4, either as free compounds or conjugated to different SLPs, to mature murine DCs was compared by stimulating murine dendritic cells overnight followed by ELISA and flow cytometry analysis. Murine tumor experiments were carried out by vaccinating mice carrying established HPV16 E6 and E7-expressing tumors and subsequently analyzing myeloid and lymphoid cells infiltrating the tumor microenvironment. Furthermore, tumor outgrowth after vaccination was monitored to enable comparison of the efficiency to induce antitumor immunity by Pam3CSK-SLP and AV-SLP conjugates. To enhance therapeutic efficacy, AV-SLP conjugate vaccination was combined with ablative therapies to assess whether synergism between such therapies would occur.\nSLPs conjugated to AV induce stronger DC maturation, in vivo T cell priming and antitumor immunity compared to conjugates with Pam3CSK4. Interestingly, AV-SLP conjugates modulate the macrophage populations in the tumor microenvironment, correlating with a therapeutic effect in an aggressive murine tumor model. The potency of AV-SLP conjugates in cancer vaccination operates optimally in combination with chemotherapy or photodynamic therapy.\nThese data allow further optimization of vaccination-based immunotherapy of cancer by use of the improved TLR2-ligand Amplivant.\nBACKGROUND\nMETHODS\nRESULTS\nCONCLUSIONBio-organic Synthesi

N-Tetradecylcarbamyl Lipopeptides as Novel Agonists for Toll-like Receptor 2

Tumorimmunolog