Direct-Acting Antiviral Therapy Restores Immune Tolerance to Patients With Hepatitis C Virus-Induced Cryoglobulinemia Vasculitis.

BACKGROUND & AIMS: Interferon-free direct-acting antiviral (DAA) therapies are effective in patients with hepatitis C virus-induced cryoglobulinemia vasculitis (HCV-CV). We analyzed blood samples from patients with HCV-CV before and after DAA therapy to determine mechanisms of these drugs and their effects on cellular immunity. METHODS: We performed a prospective study of 27 consecutive patients with HCV-CV (median age, 59 y) treated with DAA therapy (21 patients received sofosbuvir plus ribavirin for 24 weeks, 4 patients received sofosbuvir plus daclatasvir for 12 weeks, and 2 patients received sofosbuvir plus simeprevir for 12 weeks) in Paris, France. Blood samples were collected from these patients before and after DAA therapy, and also from 12 healthy donors and 12 individuals with HCV infection without CV. HCV load, cryoglobulins, and cytokines were quantified by flow cytometry, cytokine multiplex assays, and enzyme-linked immunosorbent assay. RESULTS: Twenty-four patients (88.9%) had a complete clinical response of CV to DAA therapy at week 24, defined by improvement of all the affected organs and the absence of relapse. Compared with healthy donors and patients with HCV infection without CV, patients with HCV-CV, before DAA therapy, had a lower percentage of CD4+CD25hiFoxP3+ regulatory T cells (P < .01), but higher proportions of IgM+CD21-/low memory B cells (P < .05), CD4+IFNγ+ cells (P < .01), CD4+IL17A+ cells (P < .01), and CD4+CXCR5+interleukin 21+ follicular T-helper (Tfh) cells (P < .01). In patients with HCV-CV, there was a negative correlation between numbers of IgM+CD21-/low memory B cells and T-regulatory cells (P = .03), and positive correlations with numbers of Tfh cells (P = .03) and serum levels of cryoglobulin (P = .01). DAA therapy increased patients' numbers of T-regulatory cells (1.5% ± 0.18% before therapy vs 2.1% ± 0.18% after therapy), decreased percentages of IgM+CD21-/low memory B cells (35.7% ± 6.1% before therapy vs 14.9% ± 3.8% after therapy), and decreased numbers of Tfh cells (12% ± 1.3% before therapy vs 8% ± 0.9% after therapy). Expression levels of B lymphocyte stimulator receptor 3 and programmed cell death 1 on B cells increased in patients with HCV-CV after DAA-based therapy (mean fluorescence units, 37 ± 2.4 before therapy vs 47 ± 2.6 after therapy, P < .01; and 29 ± 7.3 before therapy vs 48 ± 9.3 after therapy, P < .05, respectively). CONCLUSIONS: In a prospective clinical trial of patients with HCV-CV, DAA-based therapy restored disturbances in peripheral B- and T-cell homeostasis.

Retrovirus-Based Virus-Like Particle Immunogenicity and Its Modulation by Toll-Like Receptor Activation.

Retrovirus-derived virus-like particles (VLPs) are particularly interesting vaccine platforms, as they trigger efficient humoral and cellular immune responses and can be used to display heterologous antigens. In this study, we characterized the intrinsic immunogenicity of VLPs and investigated their possible adjuvantization by incorporation of Toll-like receptor (TLR) ligands. We designed a noncoding single-stranded RNA (ncRNA) that could be encapsidated by VLPs and induce TLR7/8 signaling. We found that VLPs efficiently induce in vitro dendritic cell activation, which can be improved by ncRNA encapsidation (ncRNAVLPs). Transcriptome studies of dendritic cells harvested from the spleens of immunized mice identified antigen presentation and immune activation as the main gene expression signatures induced by VLPs, while TLR signaling and Th1 signatures characterize ncRNAVLPs. In vivo and compared with standard VLPs, ncRNAVLPs promoted Th1 responses and improved CD8+ T cell proliferation in a MyD88-dependent manner. In an HIV vaccine mouse model, HIV-pseudotyped ncRNAVLPs elicited stronger antigen-specific cellular and humoral responses than VLPs. Altogether, our findings provide molecular evidence for a strong vaccine potential of retrovirus-derived VLPs that can be further improved by harnessing TLR-mediated immune activation.IMPORTANCE We previously reported that DNA vaccines encoding antigens displayed in/on retroviral VLPs are more efficient than standard DNA vaccines at inducing cellular and humoral immune responses. We aimed to decipher the mechanisms and investigated the VLPs' immunogenicity independently of DNA vaccination. We show that VLPs have the ability to activate antigen-presenting cells directly, thus confirming their intrinsic immunostimulatory properties and their potential to be used as an antigenic platform. Notably, this immunogenicity can be further improved and/or oriented by the incorporation into VLPs of ncRNA, which provides further TLR-mediated activation and Th1-type CD4+ and CD8+ T cell response orientation. Our results highlight the versatility of retrovirus-derived VLP design and the value of using ncRNA as an intrinsic vaccine adjuvant.

Low-Dose IL-2 Induces Regulatory T Cell-Mediated Control of Experimental Food Allergy.

Regulatory T cells (Tregs) are pivotal for maintenance of immune self-tolerance and also regulate immune responses to exogenous Ags, including allergens. Both decreased Treg number and function have been reported in allergic patients, offering new therapeutic perspectives. We previously demonstrated that Tregs can be selectively expanded and activated by low doses of IL-2 (ld-IL-2) inducing immunoregulation without immunosuppression and established its protective effect in autoimmune diseases. In this study, we evaluated the ability of ld-IL-2 to control allergy in an experimental model of food allergy. Ld-IL-2 induced Treg expansion and activation that elicited protection against clinical manifestations of food allergy in two mouse models with OVA and peanut. This clinical effect was lost in Treg-depleted mice, demonstrating the major contribution of Tregs in ld-IL-2 efficacy. Mechanistic studies further indicated that protection from allergy could be explained by a Treg-dependent local modification of the Th1/Th2 balance and an inhibition of mast cell recruitment and activation. Preventive and therapeutic effects of ld-IL-2 were observed over a 7-mo-period, highlighting its long-term efficacy. This study demonstrated that ld-IL-2 is efficient to prevent and to treat allergic immune responses, and thus represents a promising therapeutic strategy for managing allergic diseases.

Early Transcriptome Signatures from Immunized Mouse Dendritic Cells Predict Late Vaccine-Induced T-Cell Responses.

Systems biology offers promising approaches for identifying response-specific signatures to vaccination and assessing their predictive value. Here, we designed a modelling strategy aiming to predict the quality of late T-cell responses after vaccination from early transcriptome analysis of dendritic cells. Using standardized staining with tetramer, we first quantified antigen-specific T-cell expansion 5 to 10 days after vaccination with one of a set of 41 different vaccine vectors all expressing the same antigen. Hierarchical clustering of the responses defined sets of high and low T cell response inducers. We then compared these responses with the transcriptome of splenic dendritic cells obtained 6 hours after vaccination with the same vectors and produced a random forest model capable of predicting the quality of the later antigen-specific T-cell expansion. The model also successfully predicted vector classification as low or strong T-cell response inducers of a novel set of vaccine vectors, based on the early transcriptome results obtained from spleen dendritic cells, whole spleen and even peripheral blood mononuclear cells. Finally, our model developed with mouse datasets also accurately predicted vaccine efficacy from literature-mined human datasets.

Regulatory T-cell development and function are impaired in mice lacking membrane expression of full length intercellular adhesion molecule-1.

To further investigate the contribution of intercellular adhesion molecule-1 (ICAM-1) to adaptive immune responses, we analysed T-cell development and function in mice lacking full-length ICAM-1 (ICAM-1(tm1Jcgr) ). Compared with wild-type (ICAM-1(WT) ) mice, ICAM-1(tm1Jcgr) mice have impaired thymocyte development. Proportions and numbers of double negative, double positive, mature CD4(+) and CD8(+) thymocytes, as well as of regulatory T (Treg) cells were also significantly decreased. In the periphery, ICAM-1(tm1Jcgr) mice had significantly decreased proportions and numbers of naive and activated/memory CD4(+) and CD8(+) T cells, as well as of Treg cells, in lymph nodes but not in the spleen. In vitro activation of CD4(+) and CD8(+) T cells from ICAM-1(tm1Jcgr) mice with anti-CD3 antibodies and antigen-presenting cells (APCs) resulted in a significantly weaker proliferation, whereas proliferation induced with anti-CD3 and anti-CD28 antibody-coated beads was normal. In vivo immunization of ICAM-1(tm1Jcgr) mice resulted in normal generation of specific effector and memory immune responses that protect against a viral challenge. However, contrary to ICAM-1(WT) mice, immunization-induced specific effectors could not eradicate immunogen-expressing tumours. Treg cells from ICAM-1(tm1Jcgr) mice have abnormal activation and proliferation induced by anti-CD3 antibody and APCs, and have markedly decreased suppressive activity in vitro. In contrast to ICAM-1(WT) mice, they were unable to control experimentally induced colitis in vivo. Hence, our results further highlight the pleiotropic role of ICAM-1 in T-cell-dependent immune responses, with a major role in Treg cell development and suppressive function.

Sustained stimulation and expansion of Tregs by IL2 control autoimmunity without impairing immune responses to infection, vaccination and cancer.

Interleukin 2 (IL2) is the key cytokine supporting survival and function of regulatory T cells (Tregs). We recently reported that low-dose IL2 safely expands/stimulates Tregs and improves autoimmune conditions in humans. Further development of IL2 in autoimmune diseases will require chronic IL2 administration, which could affect beneficial effector immune responses regulated by Tregs. We used recombinant adeno-associated viral vector (rAAV)-mediated gene transfer to continuously release IL2 in mice and assessed its long-term effects on immune responses. A single rAAV-IL2 injection enabled sustained stimulation and expansion of Tregs without inducing Teff activation and prevented diabetes in NOD mice. After several weeks of IL2 production, mice responded normally to a viral challenge and to vaccination, and had pregnancies with offspring that developed normally. They showed no change in the occurrence and growth of chemically-induced tumors. Altogether, chronic low-dose IL2 treatment does not affect beneficial effector immune responses at doses that prevent autoimmune diabetes.

Interleukin-1 regulates follicular T cells during the germinal center reaction.

Introduction: Antibody production and the generation of memory B cells are regulated by T follicular helper (Tfh) and T follicular regulatory (Tfr) cells in germinal centers. However, the precise role of Tfr cells in controlling antibody production is still unclear. We have previously shown that both Tfh and Tfr cells express the IL-1R1 agonist receptor, whereas only Tfr cells express the IL-1R2 decoy and IL-1Ra antagonist receptors. We aimed to investigate the role of IL-1 receptors in the regulation of B cell responses by Tfh and Tfr. Methods: We generated mice with IL-1 receptors inactivated in Tfh or Tfr and measured antibody production and cell activation after immunisation. Results: While IL-1ß levels are increased in the draining lymph node after immunisation, antigen-specific antibody levels and cell phenotypes indicated that IL-1ß can activate both Tfh and Tfr cells through IL-1R1 stimulation. Surprisingly, expression of IL-1R2 and IL-1Ra on Tfr cells does not block IL-1 activation of Tfh cells, but rather prevents IL-1/IL-1R1-mediated early activation of Tfr cells. IL-1Rs also regulate the antibody response to autoantigens and its associated pathophysiology in an experimental lupus model. Discussion: Collectively, our results show that IL-1 inhibitory receptors expressed by Tfr cells prevent their own activation and suppressive function, thus licensing IL-1-mediated activation of Tfh cells after immunisation. Further mechanistic studies should unravel these complex interactions between IL-1ß and follicular helper and regulatory T cells and provide new avenues for therapeutic intervention.

The path to next-generation disease-modifying immunomodulatory combination therapies in Alzheimer's disease.

The cautious optimism following recent anti-amyloid therapeutic trials for Alzheimer's disease (AD) provides a glimmer of hope after years of disappointment. Although these encouraging results represent discernible progress, they also highlight the need to enhance further the still modest clinical efficacy of current disease-modifying immunotherapies. Here, we highlight crucial milestones essential for advancing precision medicine in AD. These include reevaluating the choice of therapeutic targets by considering the key role of both central neuroinflammation and peripheral immunity in disease pathogenesis, refining patient stratification by further defining the inflammatory component within the forthcoming ATN(I) (amyloid, tau and neurodegeneration (and inflammation)) classification of AD biomarkers and defining more accurate clinical outcomes and prognostic biomarkers that better reflect disease heterogeneity. Next-generation immunotherapies will need to go beyond the current antibody-only approach by simultaneously targeting pathological proteins together with innate neuroinflammation and/or peripheral-central immune crosstalk. Such innovative immunomodulatory combination therapy approaches should be evaluated in appropriately redesigned clinical therapeutic trials, which must carefully integrate the neuroimmune component.

Particulate antigens administrated by intranasal and intravaginal routes in a prime-boost strategy improve HIV-specific TFH generation, high-quality antibodies and long-lasting mucosal immunity.

Mucosal surfaces serve as the primary entry points for pathogens such as SARS- CoV-2 coronavirus or HIV in the human body. Mucosal vaccination plays a crucial role to successfully induce long-lasting systemic and local immune responses to confer sterilizing immunity. However, antigen formulations and delivery methods must be properly selected since they are decisive for the quality and the magnitude of the elicited immune responses in mucosa. We investigated the significance of using particulate antigen forms for mucosal vaccination by comparing VLP- or protein- based vaccines in a mouse model. Based on a mucosal prime-boost immunization protocol combining (i) HIV- pseudotyped recombinant VLPs (HIV-VLPs) and (ii) plasmid DNA encoding HIV- VLPs (pVLPs), we demonstrated that combination of intranasal primes and intravaginal boosts is optimal to elicit both humoral and cellular memory responses in mucosa. Interestingly, our results show that in contrast to proteins, particulate antigens induce high-quality humoral responses characterized by a high breadth, long-term neutralizing activity and cross-clade reactivity, accompanying with high T follicular helper cell (TFH) response. These results underscore the potential of a VLP-based vaccine in effectively instigating long-lasting, HIV-specific immunity and point out the specific role of particulate antigen form in driving high-quality mucosal immune responses.

Intranasal DNA vaccination induces potent mucosal and systemic immune responses and cross-protective immunity against influenza viruses.

The induction of potent virus-specific immune responses at mucosal surfaces where virus transmission occurs is a major challenge for vaccination strategies. In the case of influenza vaccination, this has been achieved only by intranasal delivery of live-attenuated vaccines that otherwise pose safety problems. Here, we demonstrate that potent mucosal and systemic immune responses, both cellular and humoral, are induced by intranasal immunization using formulated DNA. We show that formulation with the DNA carrier polyethylenimine (PEI) improved by a 1,000-fold the efficiency of gene transfer in the respiratory track following intranasal administration of luciferase-coding DNA. Using PEI formulation, intranasal vaccination with DNA-encoding hemagglutinin (HA) from influenza A H5N1 or (H1N1)2009 viruses induced high levels of HA-specific immunoglobulin A (IgA) antibodies that were detected in bronchoalveolar lavages (BALs) and the serum. No mucosal responses could be detected after parenteral or intranasal immunization with naked-DNA. Furthermore, intranasal DNA vaccination with HA from a given H5N1 virus elicited full protection against the parental strain and partial cross-protection against a distinct highly pathogenic H5N1 strain that could be improved by adding neuraminidase (NA) DNA plasmids. Our observations warrant further investigation of intranasal DNA as an effective vaccination route.

A Thermostable Oral SARS-CoV-2 Vaccine Induces Mucosal and Protective Immunity.

An ideal protective vaccine against SARS-CoV-2 should not only be effective in preventing disease, but also in preventing virus transmission. It should also be well accepted by the population and have a simple logistic chain. To fulfill these criteria, we developed a thermostable, orally administered vaccine that can induce a robust mucosal neutralizing immune response. We used our platform based on retrovirus-derived enveloped virus-like particles (eVLPs) harnessed with variable surface proteins (VSPs) from the intestinal parasite Giardia lamblia, affording them resistance to degradation and the triggering of robust mucosal cellular and antibody immune responses after oral administration. We made eVLPs expressing various forms of the SARS-CoV-2 Spike protein (S), with or without membrane protein (M) expression. We found that prime-boost administration of VSP-decorated eVLPs expressing a pre-fusion stabilized form of S and M triggers robust mucosal responses against SARS-CoV-2 in mice and hamsters, which translate into complete protection from a viral challenge. Moreover, they dramatically boosted the IgA mucosal response of intramuscularly injected vaccines. We conclude that our thermostable orally administered eVLP vaccine could be a valuable addition to the current arsenal against SARS-CoV-2, in a stand-alone prime-boost vaccination strategy or as a boost for existing vaccines.

High diversity of the immune repertoire in humanized NOD.SCID.gamma c-/- mice.

The diversity of the human immune repertoire and how it relates to a functional immune response has not yet been studied in detail in humanized NOD.SCID.gammac(-/-) immunodeficient mice. Here, we used a multiplex PCR on genomic DNA to quantify the combinatorial diversity of all possible V-J rearrangements at the TCR-beta chain and heavy chain Ig locus. We first show that the combinatorial diversity of the TCR-beta chain generated in the thymus was well preserved in the periphery, suggesting that human T cells were not vastly activated in mice, in agreement with phenotypic studies. We then show that the combinatorial diversity in NOD.SCID.gammac(-/-) mice reached 100% of human reference samples for both the TCR and the heavy chain of Ig. To document the functionality of this repertoire, we show that a detectable but weak HLA-restricted cellular immune response could be elicited in reconstituted mice after immunization with an adenoviral vector expressing HCV envelope glycoproteins. Altogether, our results suggest that humanized mice express a diversified repertoire and are able to mount antigen-specific immune responses.

Identification of a dominant epitope in the hemagglutinin of an Asian highly pathogenic avian influenza H5N1 clade 1 virus by selection of escape mutants.

H5N1 avian influenza virus has caused widespread infection in poultry and wild birds, and has the potential to emerge as a pandemic threat to humans. The hemagglutinin (HA) is a glycoprotein on the surface of the virus envelope. Understanding its antigenic structure is essential for designing novel vaccines that can inhibit virus infection. The aim of this study was to map the amino acid substitutions that resulted in resistance to neutralization by monoclonal antibodies (MAbs) of the highly pathogenic A/crested eagle/Belgium/01/2004 (H5N1), a clade 1 virus. Two hybridomas specific to H5N1 clade 1 viruses were selected by enzyme-linked immunosorbent assay, virus neutralization test, and immunofluorescence assay. Escape mutant populations resisting neutralization by those MAbs (8C5 and 5A1) were then selected, and sequencing of these mutants allowed the prediction of the HA protein structure by molecular homology. We could detect an amino acid change in our escape mutants at position K189E corresponding to antigenic site 2 of H5 HA1 and site B of H3 HA1. Interestingly, 336 out of 350 available HA sequences from H5N1 clade 1 and clade 2.3 viruses had Lys (K) at position 189 in the HA1, whereas HA sequences analyzed from dade 2.1 and 2.2 viruses had Arg (R). This residue also interacts with the receptor-binding site, and it is thus important for the evolution of H5N1 viruses. An additional substitution K29E in HA2 subunit was also observed and identified with the use of NetChop software as a loss of a proteasomal cleavage site, which seems to be an advantage for H5N1 viruses.

Recombinant retrovirus-like particle forming DNA vaccines in prime-boost immunization and their use for hepatitis C virus vaccine development.

BACKGROUND: The expression of Moloney murine leukemia virus (Mo-MLV) gag proteins is sufficient to generate retrovirus-like particles (retroVLPs) that can be used as antigen-display platforms by pseudotyping with heterologous envelope proteins or by insertion of epitopes in structural constituents. To circumvent the in vitro production of such retroVLPs, we used DNA plasmids generating recombinant retroVLPs (plasmo-retroVLPs) as immunogens. We previously demonstrated that plasmo-retroVLPs induce significantly better antigen-specific T cell responses and antiviral immune protection than plasmids bearing a single mutation preventing retroVLPs assembly. In the present study, we investigated the possibility of using such plasmo-retroVLPs in prime-boost immunization strategies for hepatitis C virus (HCV) vaccine development. METHODS: To define the best immunization regimen with plasmo-retroVLPs and serotype 5 recombinant adenovirus vectors (rAd5), we used standardized methodologies measuring immune responses to the GP(33-41) 'gold standard' antigen. The protective efficacy of these immunization schedules was also evaluated in mice after tumor challenge. We then applied the optimal prime-boost immunization strategy using vectors expressing HCV-E1/E2 envelope glycoproteins. RESULTS: Using vectors expressing the model antigen, we demonstrated that rAd5(GP33-41)/plasmo-retroVLP(GP33-41) regimen induced significantly higher cellular immune responses than plasmo-retroVLP(GP33-41)/rAd5(GP33-41). Consequently, HCV-specific plasmo-retroVLPs (plasmo-retroVLP(E1E2)) were used as boost in mice primed with rAd5(E1E2) and we observed that plasmo-retroVLP(E1E2) significantly increased E1/E2-specific interferon-gamma cellular responses and E2-specific antibody generation. By contrast, plasmids unable to form E1/E2-pseudotyped retroVLPs had no boosting effect, revealing the importance of presenting E1/E2 in a particulate form. CONCLUSIONS: Altogether, combining plasmo-retroVLPs that represent a new class of genetic vaccines in a heterologous prime-boost vaccination strategy appears to be a promising strategy for HCV vaccine development.

Efficient oral vaccination by bioengineering virus-like particles with protozoan surface proteins.

Intestinal and free-living protozoa, such as Giardia lamblia, express a dense coat of variant-specific surface proteins (VSPs) on trophozoites that protects the parasite inside the host's intestine. Here we show that VSPs not only are resistant to proteolytic digestion and extreme pH and temperatures but also stimulate host innate immune responses in a TLR-4 dependent manner. We show that these properties can be exploited to both protect and adjuvant vaccine antigens for oral administration. Chimeric Virus-like Particles (VLPs) decorated with VSPs and expressing model surface antigens, such as influenza virus hemagglutinin (HA) and neuraminidase (NA), are protected from degradation and activate antigen presenting cells in vitro. Orally administered VSP-pseudotyped VLPs, but not plain VLPs, generate robust immune responses that protect mice from influenza infection and HA-expressing tumors. This versatile vaccine platform has the attributes to meet the ultimate challenge of generating safe, stable and efficient oral vaccines.

Replication-competent vectors and empty virus-like particles: new retroviral vector designs for cancer gene therapy or vaccines.

Replication-defective vectors based on murine oncoretroviruses were the first gene transfer vectors to be used in successful gene therapies. Despite this achievement, they have two major drawbacks: insufficient efficacy for in vivo gene transfer and insertional mutagenesis. Attempts to overcome these problems have led to two retroviral vector designs of principally opposite character: replication-competent vectors transducing largely intact genomes and genome-free vectors. Replication-competent retroviral vectors have achieved dramatically improved efficacy for in vivo cancer gene therapy and genome-free retroviral vectors expressing different kinds of antigens have proven excellent as immunogens. Current developments aim to improve the safety of the replication-competent vectors and to augment the production efficiency of the genome-free vectors by expression from heterologous viral or non-viral vectors. Together with the continuous advances of classical defective retroviral vectors for ex vivo gene therapy, these developments illustrate that, due to their tremendous design versatility, retroviral vectors remain important vectors for gene therapy applications.

TGF-ß and VEGF cooperatively control the immunotolerant tumor environment and the efficacy of cancer immunotherapies.

Tregs imprint an early immunotolerant tumor environment that prevents effective antitumor immune responses. Using transcriptomics of tumor tissues, we identified early upregulation of VEGF and TGF-ß pathways compatible with tolerance imprinting. Silencing of VEGF or TGF-ß in tumor cells induced early and pleiotropic modulation of immune-related transcriptome signatures in tumor tissues. These were surprisingly similar for both silenced tumors and related to common downstream effects on Tregs. Silencing of VEGF or TGF-ß resulted in dramatically delayed tumor growth, associated with decreased Tregs and myeloid-derived suppressor cells and increased effector T cell activation in tumor infiltrates. Strikingly, co-silencing of TGF-ß and VEGF led to a substantial spontaneous tumor eradication rate and the combination of their respective inhibitory drugs was synergistic. VEGF and/or TGF-ß silencing also restored tumor sensitivity to tumor-specific cell therapies and markedly improved the efficacy of anti-PD-1/anti-CTLA-4 treatment. Thus, TGF-ß and VEGF cooperatively control the tolerant environment of tumors and are targets for improved cancer immunotherapies.

Intra-cheek immunization as a novel vaccination route for therapeutic vaccines of head and neck squamous cell carcinomas using plasmo virus-like particles.

Despite current therapy, head and neck squamous cell carcinomas (HNSCCs) arising from various mucosal sites of the upper aero-digestive tract frequently relapse in a loco-regional manner and have a poor prognosis. Our objective was to validate an innovative mucosal route of vaccination using plasmo virus-like particles (pVLPs) in a pre-clinical orthotopic model of HNSCCs. For this purpose, we used pVLP-E7, that are plasmid DNA encoding retroviral virus-like particles carrying a truncated E7 oncoprotein from HPV-16 as antigen model, to vaccinate mice bearing pre-established TC-1 tumors implanted into the buccal mucosa. pVLP-E7 were combined with clinical grade TLR agonists (Imiquimod and CpG-ODN). In this pre-clinical orthotopic model, whose tumor microenvironment resembles to those of human HNSCCs, different mucosal vaccination routes were tested for their ability to elicit efficient immune and antitumoral responses. Results showed that mucosal intra-cheek (IC) vaccinations using pVLP-E7, comparatively to intradermic vaccinations (ID), gave rise to higher mobilization of mucosal (CD49a(+)) CD8(+) specific effector T cells in both tumor draining lymph nodes (TdLNs) and tumor microenvironment resulting in better antitumor effects and in a long-term protection against tumor rechallenge. In vivo CD8(+) depletion demonstrated that antitumoral effects were fully dependent upon the presence of CD8(+) T cells. Validation of IC mucosal vaccinations with pVLPs combined with adjuvants using a pre-clinical orthotopic model of HNSCC provides valuable pre-clinical data to rapidly envision the use of such therapeutic vaccines in patients with HNSCCs, inasmuch as vaccinal components and adjuvants can be easily obtained as clinical grade reagents.

Enveloped virus-like particle platforms: vaccines of the future?

The techniques to produce effective vaccines have evolved, and the early vaccines (live, inactivated, subunit...) are no longer considered as the most appropriate for new vaccine development. We question here what will be the future vaccines, and argue that virus-like particle (VLP)-based vaccines are promising candidates. In addition to being effective vaccines against analogous viruses from which they are derived, VLPs can also be used to present foreign epitopes to the immune system. The achievement of this strategy can be illustrated by the recent development of malaria candidate vaccine. We point out recent VLP-based vaccine developments and discuss future perspectives.