Liposome induction of CD8+ T cell responses depends on CD169+ macrophages and Batf3-dependent dendritic cells and is enhanced by GM3 inclusion

Cancer vaccines aim to efficiently prime cytotoxic CD8+ T cell responses which can be achieved by vaccine targeting to dendritic cells. CD169+ macrophages have been shown to transfer antigen to dendritic cells and could act as an alternative target for cancer vaccines. Here, we evaluated liposomes containing the CD169/Siglec-1 binding ligand, ganglioside GM3, and the non-binding ligand, ganglioside GM1, for their capacity to target antigens to CD169+ macrophages and to induce immune responses. CD169+ macrophages demonstrated specific uptake of GM3 liposomes in vitro and in vivo that was dependent on a functional CD169 receptor. Robust antigen-specific CD8+ and CD4+ T and B cell responses were observed upon intravenous administration of GM3 liposomes containing the model antigen ovalbumin in the presence of adjuvant. Immunization of B16-OVA tumor bearing mice with all liposomes resulted in delayed tumor growth and improved survival. The absence of CD169+ macrophages, functional CD169 molecules, and cross-presenting Batf3-dependent dendritic cells (cDC1s) significantly impaired CD8+ T cell responses, while B cell responses were less affected. In conclusion, we demonstrate that inclusion of GM3 in liposomes enhance immune responses and that splenic CD169+ macrophages and cDC1s are required for induction of CD8+ T cell immunity after liposomal vaccination

Incorporation of Toll-Like Receptor Ligands and Inflammasome Stimuli in GM3 Liposomes to Induce Dendritic Cell Maturation and T Cell Responses

Cancer vaccination aims to activate immunity towards cancer cells and can be achieved by delivery of cancer antigens together with immune stimulatory adjuvants to antigen presenting cells (APC). APC maturation and antigen processing is a subsequent prerequisite for T cell priming and anti-tumor immunity. In order to specifically target APC, nanoparticles, such as liposomes, can be used for the delivery of antigen and adjuvant. We have previously shown that liposomal inclusion of the ganglioside GM3, an endogenous ligand for CD169, led to robust uptake by CD169-expressing APC and resulted in strong immune responses when supplemented with a soluble adjuvant. To minimize the adverse effects related to a soluble adjuvant, immune stimulatory molecules can be incorporated in liposomes to achieve targeted delivery of both antigen and adjuvant. In this study, we incorporated TLR4 (MPLA) or TLR7/8 (3M-052) ligands in combination with inflammasome stimuli, 1-palmitoyl-2-glutaryl-sn-glycero-3-phosphocholine (PGPC) or muramyl dipeptide (MDP), into GM3 liposomes. Incorporation of TLR and inflammasome ligands did not interfere with the uptake of GM3 liposomes by CD169-expressing cells. GM3 liposomes containing a TLR ligand efficiently matured human and mouse dendritic cells in vitro and in vivo, while inclusion of PGPC or MDP had minor effects on maturation. Immunization with MPLA-containing GM3 liposomes containing an immunogenic synthetic long peptide stimulated CD4 + and CD8 + T cell responses, but additional incorporation of either PGPC or MDP did not translate into stronger immune responses. In conclusion, our study indicates that TLRL-containing GM3 liposomes are effective vectors to induce DC maturation and T cell priming and thus provide guidance for further selection of liposomal components to optimally stimulate anti-cancer immune responses

Mimicking Pathogens to Augment the Potency of Liposomal Cancer Vaccines

Liposomes have emerged as interesting vehicles in cancer vaccination strategies as their composition enables the inclusion of both hydrophilic and hydrophobic antigens and adjuvants. In addition, liposomes can be decorated with targeting moieties to further resemble pathogenic particles that allow for better engagement with the immune system. However, so far liposomal cancer vaccines have not yet reached their full potential in the clinic. In this review, we summarize recent preclinical studies on liposomal cancer vaccines. We describe the basic ingredients for liposomal cancer vaccines, tumor antigens, and adjuvants, and how their combined inclusion together with targeting moieties potentially derived from pathogens can enhance vaccine immunogenicity. We discuss newly identified antigen-presenting cells in humans and mice that pose as promising targets for cancer vaccines. The lessons learned from these preclinical studies can be applied to enhance the efficacy of liposomal cancer vaccination in the clinic

Liposomal Nanovaccine Containing α-Galactosylceramide and Ganglioside GM3 Stimulates Robust CD8+ T Cell Responses via CD169+ Macrophages and cDC1

Successful anti-cancer vaccines aim to prime and reinvigorate cytotoxic T cells and should therefore comprise a potent antigen and adjuvant. Antigen targeting to splenic CD169+ macrophages was shown to induce robust CD8+ T cell responses via antigen transfer to cDC1. Interestingly, CD169+ macrophages can also activate type I natural killer T-cells (NKT). NKT activation via ligands such as α-galactosylceramide (αGC) serve as natural adjuvants through dendritic cell activation. Here, we incorporated ganglioside GM3 and αGC in ovalbumin (OVA) protein-containing liposomes to achieve both CD169+ targeting and superior DC activation. The systemic delivery of GM3-αGC-OVA liposomes resulted in specific uptake by splenic CD169+ macrophages, stimulated strong IFNγ production by NKT and NK cells and coincided with the maturation of cDC1 and significant IL-12 production. Strikingly, superior induction of OVA-specific CD8+ T cells was detected after immunization with GM3-αGC-OVA liposomes. CD8+ T cell activation, but not B cell activation, was dependent on CD169+ macrophages and cDC1, while activation of NKT and NK cells were partially mediated by cDC1. In summary, GM3-αGC antigen-containing liposomes are a potent vaccination platform that promotes the interaction between different immune cell populations, resulting in strong adaptive immunity and therefore emerge as a promising anti-cancer vaccination strategy

Liposome induction of CD8+ T cell responses depends on CD169+ macrophages and Batf3-dependent dendritic cells and is enhanced by GM3 inclusion

Cancer vaccines aim to efficiently prime cytotoxic CD8+ T cell responses which can be achieved by vaccine targeting to dendritic cells. CD169+ macrophages have been shown to transfer antigen to dendritic cells and could act as an alternative target for cancer vaccines. Here, we evaluated liposomes containing the CD169/Siglec-1 binding ligand, ganglioside GM3, and the non-binding ligand, ganglioside GM1, for their capacity to target antigens to CD169+ macrophages and to induce immune responses. CD169+ macrophages demonstrated specific uptake of GM3 liposomes in vitro and in vivo that was dependent on a functional CD169 receptor. Robust antigen-specific CD8+ and CD4+ T and B cell responses were observed upon intravenous administration of GM3 liposomes containing the model antigen ovalbumin in the presence of adjuvant. Immunization of B16-OVA tumor bearing mice with all liposomes resulted in delayed tumor growth and improved survival. The absence of CD169+ macrophages, functional CD169 molecules, and cross-presenting Batf3-dependent dendritic cells (cDC1s) significantly impaired CD8+ T cell responses, while B cell responses were less affected. In conclusion, we demonstrate that inclusion of GM3 in liposomes enhance immune responses and that splenic CD169+ macrophages and cDC1s are required for induction of CD8+ T cell immunity after liposomal vaccination

Optimization of Liposomes for Antigen Targeting to Splenic CD169+ Macrophages

Despite promising progress in cancer vaccination, therapeutic effectiveness is often insufficient. Cancer vaccine effectiveness could be enhanced by targeting vaccine antigens to antigen-presenting cells, thereby increasing T-cell activation. CD169-expressing splenic macrophages efficiently capture particulate antigens from the blood and transfer these antigens to dendritic cells for the activation of CD8+ T cells. In this study, we incorporated a physiological ligand for CD169, the ganglioside GM3, into liposomes to enhance liposome uptake by CD169+ macrophages. We assessed how variation in the amount of GM3, surface-attached PEG and liposomal size affected the binding to, and uptake by, CD169+ macrophages in vitro and in vivo. As a proof of concept, we prepared GM3-targeted liposomes containing a long synthetic ovalbumin peptide and tested the capacity of these liposomes to induce CD8+ and CD4+ T-cell responses compared to control liposomes or soluble peptide. The data indicate that the delivery of liposomes to splenic CD169+ macrophages can be optimized by the selection of liposomal constituents and liposomal size. Moreover, optimized GM3-mediated liposomal targeting to CD169+ macrophages induces potent immune responses and therefore presents as an interesting delivery strategy for cancer vaccination

Liposomal Nanovaccine Containing α-Galactosylceramide and Ganglioside GM3 Stimulates Robust CD8+ T Cell Responses via CD169+ Macrophages and cDC1

10.3390/vaccines9010056Vaccines91Jan-1

Liposomal Nanovaccine Containing α-Galactosylceramide and Ganglioside GM3 Stimulates Robust CD8+ T Cell Responses via CD169+ Macrophages and cDC1

Successful anti-cancer vaccines aim to prime and reinvigorate cytotoxic T cells and should therefore comprise a potent antigen and adjuvant. Antigen targeting to splenic CD169+ macrophages was shown to induce robust CD8+ T cell responses via antigen transfer to cDC1. Interestingly, CD169+ macrophages can also activate type I natural killer T-cells (NKT). NKT activation via ligands such as α-galactosylceramide (αGC) serve as natural adjuvants through dendritic cell activation. Here, we incorporated ganglioside GM3 and αGC in ovalbumin (OVA) protein-containing liposomes to achieve both CD169+ targeting and superior DC activation. The systemic delivery of GM3-αGC-OVA liposomes resulted in specific uptake by splenic CD169+ macrophages, stimulated strong IFNγ production by NKT and NK cells and coincided with the maturation of cDC1 and significant IL-12 production. Strikingly, superior induction of OVA-specific CD8+ T cells was detected after immunization with GM3-αGC-OVA liposomes. CD8+ T cell activation, but not B cell activation, was dependent on CD169+ macrophages and cDC1, while activation of NKT and NK cells were partially mediated by cDC1. In summary, GM3-αGC antigen-containing liposomes are a potent vaccination platform that promotes the interaction between different immune cell populations, resulting in strong adaptive immunity and therefore emerge as a promising anti-cancer vaccination strategy

Optimization of Liposomes for Antigen Targeting to Splenic CD169+ Macrophages

Despite promising progress in cancer vaccination, therapeutic effectiveness is often insufficient. Cancer vaccine effectiveness could be enhanced by targeting vaccine antigens to antigen-presenting cells, thereby increasing T-cell activation. CD169-expressing splenic macrophages efficiently capture particulate antigens from the blood and transfer these antigens to dendritic cells for the activation of CD8+ T cells. In this study, we incorporated a physiological ligand for CD169, the ganglioside GM3, into liposomes to enhance liposome uptake by CD169+ macrophages. We assessed how variation in the amount of GM3, surface-attached PEG and liposomal size affected the binding to, and uptake by, CD169+ macrophages in vitro and in vivo. As a proof of concept, we prepared GM3-targeted liposomes containing a long synthetic ovalbumin peptide and tested the capacity of these liposomes to induce CD8+ and CD4+ T-cell responses compared to control liposomes or soluble peptide. The data indicate that the delivery of liposomes to splenic CD169+ macrophages can be optimized by the selection of liposomal constituents and liposomal size. Moreover, optimized GM3-mediated liposomal targeting to CD169+ macrophages induces potent immune responses and therefore presents as an interesting delivery strategy for cancer vaccination

Optimization of Liposomes for Antigen Targeting to Splenic CD169+ Macrophages

Despite promising progress in cancer vaccination, therapeutic effectiveness is often insufficient. Cancer vaccine effectiveness could be enhanced by targeting vaccine antigens to antigen-presenting cells, thereby increasing T-cell activation. CD169-expressing splenic macrophages efficiently capture particulate antigens from the blood and transfer these antigens to dendritic cells for the activation of CD8+ T cells. In this study, we incorporated a physiological ligand for CD169, the ganglioside GM3, into liposomes to enhance liposome uptake by CD169+ macrophages. We assessed how variation in the amount of GM3, surface-attached PEG and liposomal size affected the binding to, and uptake by, CD169+ macrophages in vitro and in vivo. As a proof of concept, we prepared GM3-targeted liposomes containing a long synthetic ovalbumin peptide and tested the capacity of these liposomes to induce CD8+ and CD4+ T-cell responses compared to control liposomes or soluble peptide. The data indicate that the delivery of liposomes to splenic CD169+ macrophages can be optimized by the selection of liposomal constituents and liposomal size. Moreover, optimized GM3-mediated liposomal targeting to CD169+ macrophages induces potent immune responses and therefore presents as an interesting delivery strategy for cancer vaccination