Preclinical development of virus‐like particle (VLP)‐based immunotherapy against HDM allergy
Authors: Piyachat Patniboon, Chanatip Thanasarnthungcharoen, Supapich Chanasit, Laurence Fievez, Theerayuth Kaewamatawong, Fabrice Bureau, Alain Jacquet
Journal: Allergy (2024)

The unique disease-modifying treatment for house dust mite (HDM)-induced allergic rhinitis or allergic asthma consists in allergen-specific immunotherapy (AIT) through sublingual or subcutaneous administrations of allergen extracts. The mechanisms by which AIT dampens Th2-biased HDM allergic response leading to immune tolerance include the generation of allergen-specific T/B regulatory or Th1 cells and IgG1/IgG4 blocking antibodies.1 AP205 bacteriophage-derived virus-like particles (VLPs) displaying major HDM allergens Der p 1 (VLP-ProDer p 1, VLP-PD1N) or Der p 2 (VLP-D2) are highly hypoallergenic, promote Th1-biased immune responses and elicit high levels of blocking specific IgG.2, 3 Such chimeric VLPs represent promising candidates for the design of shorter, safer, and more effective AIT. Using a pre-established adjuvant-free mouse model of HDM-induced airway inflammation,4 we assessed the therapeutic capacities of a bivalent vaccine VLP-PD1N/VLP-D2 (Figure 1A). BALB/c mice were intranasally sensitized and challenged with HDM extracts. Sensitized animals were intramuscularly vaccinated with the bivalent vaccine (VLP-PD1N/VLP-D2, 5 μg/5 μg in 20 μL Phosphate-buffered saline-PBS). This dose was selected according to our previous immunogenicity studies.2, 3 As controls, animals were vaccinated with undecorated SpyCatcher-VLP (SC-VLP, 10 μg) or PBS (allergic group). As expected, the typical markers of HDM-induced airway inflammation were evidenced in allergic mice: large influx of inflammatory cells in bronchoalveolar fluids (BALF), accumulation of pulmonary eosinophils (Figure 1B), peribronchiolar/ perivascular inflammation and induction of mucus-producing cells in lung tissues (Figure 1C), high IL-5 levels in BALF (Figure 1D). Remarkably, all these key parameters of the HDM-specific inflammatory responses were significantly and similarly dampened by specific and non-specific VLP treatments (Figure 1B–D). Moreover, large levels of IFNγ were detected in BALFs from VLP-treated mice (Figure 1D). We observed a significant high positive and inverse correlation between the number of eosinophils and the levels of IL-5 and IFNγ respectively (Figure S1). In supernatants from restimulated splenocytes, IL-5 levels measured in the allergic group were equally blunted by specific and control VLP-AIT which, in turn, triggered large increase in IFNγ (Figure 2A). Treatment with VLP-PD1N/D2 elicited higher production of IFNγ than AIT with SC-VLP. Der p 1-/Der p 2-specific IgE antibody levels measured in allergic mice at post-AIT time point were significantly dampened at the same levels by vaccinations with specific or non-specific VLPs. The robust downregulation of the specific IgE levels persisted after intranasal challenges (Figure 2B). Contrary to SC-VLP, treatments with VLP-PD1N/D2 elicited the development of high Der p 1-/Der p 2-specific IgG1/IgG2a titers (Figure 2C). Our results highlighted that specific and non-specific VLP-based AIT alleviate pulmonary inflammatory response to HDM (lung eosinophilia/mucus-producing cells) but also HDM-specific Th2 cytokine as well as Der p 1-/Der p 2-specific IgE serum levels. In our present animal model, the effective downregulation of the HDM-induced airway inflammation is independent on the potent specific IgG antibody responses, mediated only by the specific bivalent vaccine. Although the induction of specific blocking IgG1/IgG4 antibodies represents one of the main hallmarks of successful AIT, its correlation with clinical outcomes remains debatable.5 However, the persistence for years of the inhibitory activity of IgG antibodies following discontinuation of AIT is involved with long-term immune tolerance.6 Consequently, specific and non-specific VLPs remain to be tested in more prolonged mouse models of HDM-induced airway inflammation to confirm the efficacy of VLP-AIT in the absence of HDM-specific IgG responses. We hypothesize that the anti-allergic effects shared by VLP-PD1N/D2 and SC-VLP are dependent on TLR3/TLR7-specific innate immune signalling mediated by the prokaryotic RNA cargo encapsulated into AP205 VLPs.3 Of note, several preclinical/clinical studies evidenced the effectiveness of non-specific treatments including TLR9 (CpG motifs alone or encapsulated in VLPs (QbG10)) or TLR7 (Resiquimod) ligand7 as well as non-specific mRNA formulated with LNPs.4 VLP AIT could trigger the induction of innate immune brakes in the lung which are at least mediated by interstitial macrophages expressing TLR7 at high level.8 These regulatory macrophages could restore lung homeostasis. Alternatively, non-specific VLPs could induce trained immunity through the reprograming of innate immune cells as monocytes/ Type 2 conventional dendritic cells (cDC2) into tolerogenic dendritic cells by mechanisms depending on metabolic and epigenetic rewiring.9 Upcoming studies are required to elucidate extensively the mechanism of immune modulation mediated by AP205-based VLPs in the context of HDM allergy. The contribution of prokaryotic RNA cargo encapsulated into AP205 VLPs in successful AIT must be particularly investigated through RNAse pretreatment and/or the use of TLR7-deficient mice. Important and still unresolved points must be addressed for the clinical translation of the VLP approach including finding the optimal dose, route of administration and confirming the dispensability of additional adjuvants. Further large-scale controlled studies will be necessary to evidence the safety, the effectiveness of VLP-based AIT and the duration of these protective effects. Particularly, the significance of VLP-specific B cell response for the positive clinical outcomes remains to be investigated. The interaction between VLPs and antigen-presenting and regulatory T cells also deserves our full attention. For human trials, VLPs must be produced under GMP (Good Manufacturing Procedures) conditions and consistent amounts of RNA cargo in the VLP preparations must be secured. A.J., L.F. and F.B. designed the study; P.P. prepared the different VLPs; P.P. S.C., C.T. and T.K. performed the experiments on the design of the animal model and immunotherapy. A.J. provided supervision and analyzed the data. A.J. drafted the manuscript. All authors contributed to and approved the final version of the manuscript. This work was partly funded by Thailand Science Research and Innovation (TSRI) Fund Chulalongkorn University (HEA663000026), by a 90th Anniversary of Chulalongkorn University Ratchadaphiseksomphot Fund. We warmly thank Prof. Adam F. Sander (University of Copenhagen, Denmark) for providing the genetic construction for SC-VLP production. The other authors declare they have no conflict of interest. The data that support the findings of this study are available from the corresponding author upon reasonable request. Appendix S1. Please note: The publisher is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article.