Abstract
BackgroundLigands 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.MethodsNaïve 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.ResultsSLPs 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.ConclusionThese data allow further optimization of vaccination-based immunotherapy of cancer by use of the improved TLR2-ligand Amplivant.
Highlights
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
Dendritic cell (DC) were loaded with titrating doses of either Pam3CSK4 or AV conjugated to SLPOVA CTL or with mixtures of free Synthetic long peptide (SLP) and TLR2-L and subsequently co-cultured with SIINFEKL-specific CD8+ T cells derived from the transgenic OT1 mouse
We show that the enhanced activation of DCs in vitro translates to enhanced endogenous T cell priming in vivo when AV is conjugated to SLP
Summary
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. The receptors of the innate immune system recognizing these structures, pattern-recognition receptors (PRRs), are classified into major receptor families, such as the Toll-like receptor (TLR) family The latter family consists of 10 receptors in human beings and 13 in mice, which are either expressed on the cell surface or in endosomes of many antigen-presenting cells (APCs) such as dendritic cells (DCs) [1]. Triggering of TLRs by their cognate ligand generally results in activation of APCs and concomitant secretion of pro-inflammatory cytokines and upregulation of co-stimulatory molecules on the cell surface. These effects are essential in the induction of isotype class switching of B cells and induction of efficient T cell responses. Vaccinologists have extensively explored the use of several TLR-ligands as adjuvants in vaccines to prevent or treat infections and cancer [2,3,4,5,6,7,8]
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