A Multifunctionalized Potyvirus-Derived Nanoparticle That Targets and Internalizes into Cancer Cells.

Plant viral nanoparticles (VNPs) are attractive to nanomedicine researchers because of their safety, ease of production, resistance, and straightforward functionalization. In this paper, we developed and successfully purified a VNP derived from turnip mosaic virus (TuMV), a well-known plant pathogen, that exhibits a high affinity for immunoglobulins G (IgG) thanks to its functionalization with the Z domain of staphylococcal Protein A via gene fusion. We selected cetuximab as a model IgG to demonstrate the versatility of this novel TuMV VNP by developing a fluorescent nanoplatform to mark tumoral cells from the Cal33 line of a tongue squamous cell carcinoma. Using confocal microscopy, we observed that fluorescent VNP-cetuximab bound selectively to Cal33 and was internalized, revealing the potential of this nanotool in cancer research.

Increased miR-21-3p and miR-487b-3p serum levels during anaphylactic reaction in food allergic children.

BACKGROUND: Anaphylaxis is the most severe manifestation of allergic disorders. The poor knowledge of its molecular mechanisms often leads to under-diagnosis. MicroRNAs (miRNA) regulate physiologic and pathologic processes, and they have been postulated as promising diagnostic markers. The main objectives of this study were to characterize the human miRNA profile during anaphylaxis and to assess their capacity as diagnostic markers and determine their participation in the molecular mechanisms of this event. METHODS: The miRNA serum profiles from the acute and baseline phase of 5 oral food-challenged anaphylactic children (<18 years old) were obtained by next-generation sequencing (NGS). From the panel of statistically significant miRNAs obtained, several candidates were selected and analyzed in 19 anaphylactic children by qPCR. We performed system biology analysis (SBA) on their target genes to identify main functions and canonical pathways. A functional in vitro assay was carried out incubating endothelial cells (ECs) in anaphylactic conditions. RESULTS: The NGS identified 389 miRNAs among which 41 were significantly different between acute and baseline samples. The high levels of miR-21-3p (fold change = 2.28, P = .006) and miR-487b-3p (fold change = 1.04, P = .039) observed by NGS in acute serum samples were confirmed in a larger group of 19 patients. The SBA revealed molecular pathways related to the inflammation and immune system regulation. miR-21-3p increased intracellularly and in acute phase serum after EC stimulation. CONCLUSIONS: These findings provide, for the first time, some insights into the anaphylactic miRNA serum profile in children and point to miR-21-3p and miR-487b-3p as candidate biomarkers. Furthermore, the SBA revealed a possible implication of these molecules in the underlying molecular mechanisms. Moreover, ECs increased miR-21-3p intracellularly and released it to the environment in response to anaphylaxis.

The TNF-like weak inducer of the apoptosis/fibroblast growth factor-inducible molecule 14 axis mediates histamine and platelet-activating factor-induced subcutaneous vascular leakage and anaphylactic shock.

BACKGROUND: Anaphylaxis includes mast cell (MC) activation, but less is known about downstream mechanisms (ie, vascular permeability controlled by endothelial cells [ECs]). The TNF-like weak inducer of apoptosis (TWEAK) and its sole receptor, fibroblast growth factor-inducible molecule 14 (Fn14), belong to the TNF superfamily and are involved in proinflammatory responses. OBJECTIVE: We sought to investigate the role of TWEAK/Fn14 axis in anaphylaxis. METHODS: In vivo vascular permeability and mouse models of passive systemic anaphylaxis (PSA) and active systemic anaphylaxis were applied to wild-type (WT), TWEAK- and Fn14-deficient mice (TWEAK-/- and Fn14-/-, respectively). Primary bone marrow-derived mast cells (BMMCs) and ECs from WT and Fn14-/- or TWEAK-/- mice were studied. The TWEAK/Fn14 axis was also investigated in human samples. RESULTS: Mice with PSA and active systemic anaphylaxis had increased Fn14 and TWEAK expression in lung tissues and increased serum soluble TWEAK concentrations. TWEAK and Fn14 deficiencies prevent PSA-related symptoms, resulting in resistance to decreased body temperature, less severe reactions, and maintained physical activity. Numbers of MCs after PSA are similar between genotypes in different tissue regions, such as ear skin and the trachea, tongue, peritoneum, lungs, and bone marrow. Moreover, in vitro studies revealed no differences in degranulation or mediator release between WT and Fn14-/- BMMCs after IgE-FcεRI stimulation. In vivo and in vitro histamine and platelet-activating factor administration increases Fn14 receptor expression in lungs and ECs. Moreover, Fn14 deficiency in ECs maintained in vitro impermeability when stimulated by mediators or activated BMMCs but not by TWEAK-/- BMMCs, indicating that Fn14 is crucial for endothelial barrier function. TWEAK/Fn14 deletion or TWEAK-blocking antibody prevented histamine/platelet-activating factor-induced vascular subcutaneous permeability. Circulating soluble TWEAK levels were increased in patients with anaphylaxis, and plasma from those patients increased Fn14 expression in ECs. CONCLUSION: The TWEAK/Fn14 axis participates in anaphylactic reactions. Inhibition of TWEAK/Fn14 interaction could be efficacious in anaphylaxis therapy.

Influenza A Virus Utilizes Low-Affinity, High-Avidity Interactions with the Nuclear Import Machinery To Ensure Infection and Immune Evasion.

The incoming influenza A virus (IAV) genome must pass through two distinct barriers in order to establish infection in the cell: the plasma membrane and the nuclear membrane. A precise understanding of the challenges imposed by the nuclear barrier remains outstanding. Passage across is mediated by host karyopherins (KPNAs), which bind to the viral nucleoprotein (NP) via its N-terminal nuclear localization sequence (NLS). The binding affinity between the two molecules is low, but NP is present in a high copy number, which suggests that binding avidity plays a compensatory role during import. Using nanobody-based technology, we demonstrate that a high binding avidity is required for infection, though the absolute value differs between cell types and correlates with their relative susceptibility to infection. In addition, we demonstrate that increasing the affinity level caused a decrease in avidity requirements for some cell types but blocked infection in others. Finally, we show that genomes that become frustrated by low avidity and remain cytoplasmic trigger the type I interferon response. Based on these results, we conclude that IAV balances affinity and avidity considerations in order to overcome the nuclear barrier across a broad range of cell types. Furthermore, these results provide evidence to support the long-standing hypothesis that IAV's strategy of import and replication in the nucleus facilitates immune evasion.IMPORTANCE We used intracellular nanobodies to block influenza virus infection at the step prior to nuclear import of its ribonucleoproteins. By doing so, we were able to answer an important but outstanding question that could not be addressed with conventional tools: how many of the ∼500 available NLS motifs are needed to establish infection? Furthermore, by controlling the subcellular localization of the incoming viral ribonucleoproteins and measuring the cell's antiviral response, we were able to provide direct evidence for the long-standing hypothesis that influenza virus exploits nuclear localization to delay activation of the innate immune response.

Interaction of Alt a 1 with SLC22A17 in the airway mucosa.

BACKGROUND: Despite all the efforts made up to now, the reasons that facilitate a protein becoming an allergen have not been elucidated yet. Alt a 1 protein is the major fungal allergen responsible for chronic asthma, but little is known about its immunological activity. Our main purpose was to investigate the ligand-dependent interactions of Alt a 1 in the human airway epithelium. METHODS: Alt a 1 with and without its ligand (holo- and apo- forms) was incubated with the pulmonary epithelial monolayer model, Calu-3 cells. Allergen transport and cytokine production were measured. Pull-down and immunofluorescence assays were employed to identify the receptor of Alt a 1 using the epithelial cell model and mouse tissues. Receptor-allergen-ligand interactions were analyzed by computational modeling. RESULTS: The holo-form could activate human monocytes, PBMCs, and polarized airway epithelial (Calu-3) cell lines. The allergen was also transported through the monolayer, without any alteration of the epithelial integrity (TEER). Alt a 1 also induced the production of proinflammatory IL8 and specific epithelial cytokines (IL33 and IL25) by Calu-3 cells. The interaction between epithelial cells and holo-Alt a 1 was found to be mediated by the SLC22A17 receptor, and its recognition of Alt a 1 was explained in structural terms. CONCLUSIONS: Our findings identified the Alt a 1 ligand as a central player in the interaction of the allergen with airway mucosa, shedding light into its potential role in the immunological response, while unveiling its potential as a new target for therapy intervention.

Secreted production of assembled Norovirus virus-like particles from Pichia pastoris.

BACKGROUND: Norovirus virus-like particles (NoV VLPs) have recently been explored as potential vaccine platforms due to their ability to produce an effective immune response. Expression of the main structural protein, VP1, leads to formation of self-assembled particles with similar characteristics to the original virus. These NoV VLPs have been expressed in Escherichia coli, yeast and insect cells. Expression in E. coli and insect cells share downstream processing issues due to the presence of inclusion bodies or the need for numerous purification steps. NoV VLPs have also been produced in the yeast P. pastoris; however the protein was only expressed intracellularly. RESULTS: We have successfully expressed and secreted the VP1 protein in the novel P. pastoris strain, Bg11, using the methanol inducible pJ912 expression vector, containing the cDNA of NoV VP1. Expression of the VP1 protein in Bg11 was carried out in a 1.5 L bioreactor resulting in a total yield of NoV VLPs greater than 0.6 g/L. NoV VLPs obtained from the culture supernatant were purified via ion-exchange chromatography, resulting in particles with a purity over 90%. The average size of the particles after purification was 40 nm. Transmission electron microscopy was used to visualize the morphology of the particles and saliva-binding assay confirmed that the NoV VLPs bind to Histo-Blood Group Antigens (HBGA). CONCLUSIONS: In this study we describe the expression and characterization of fully assembled Norovirus virus-like particles obtained from P. pastoris. The particles are similar in size, morphology and binding capacity, as previously described, for the original NoV. Our results detail the successful expression and secretion of VLPs in P. pastoris, improving their candidacy as a vaccine platform.

Production and use of antigen tetramers to study antigen-specific B cells.

B cells generate antibodies that provide protection from infection, but also cause pathology in autoimmune and allergic conditions. Antigen-specific B cells can be detected by binding their surface antibody receptors with native antigens conjugated to fluorescent probes, a technique that has revealed substantial insight into B cell activation and function. This protocol describes the process of generating fluorescent antigen tetramer probes and delineates a process of enriching large samples based on antigen-specificity for high-resolution analyses of the antigen-specific B cell repertoire. Enrichment of tetramer-binding cells allows for detection of antigen-specific B cells as rare as 1 in 100 million cells, providing sufficient resolution to study naive B cells and IgE-expressing cells by flow cytometry. The generation of antigen tetramers involves antigen biotinylation, assessment of biotin:antigen ratio for optimal tetramer loading and polymerization around a streptavidin-fluorophore backbone. We also describe the construction of a control tetramer to exclude B cells binding to the tetramer backbone. We provide a framework to validate whether tetramer probes are detecting true antigen-specific B cells and discuss considerations for experimental design. This protocol can be performed by researchers trained in basic biomedical/immunological research techniques, using instrumentation commonly found in most laboratories. Constructing the antigen and control tetramers takes 9 h, though their specificity should be assessed before experimentation and may take weeks to months depending on the method of validation. Sample enrichment requires ~2 h but is generally time and cost neutral as fewer cells are run through the flow cytometer.

A New Optimized Version of a Colorectal Cancer-Targeted Immunotoxin Based on a Non-Immunogenic Variant of the Ribotoxin α-Sarcin.

Due to its incidence and mortality, cancer remains one of the main risks to human health and lifespans. In order to overcome this worldwide disease, immunotherapy and the therapeutic use of immunotoxins have arisen as promising approaches. However, the immunogenicity of foreign proteins limits the dose of immunotoxins administered, thereby leading to a decrease in its therapeutic benefit. In this study, we designed two different variants of non-immunogenic immunotoxins (IMTXA33αSDI and IMTXA33furαSDI) based on a deimmunized variant of the ribotoxin α-sarcin. The inclusion of a furin cleavage site in IMTXA33furαSDI would allow a more efficient release of the toxic domain to the cytosol. Both immunotoxins were produced and purified in the yeast Pichia pastoris and later functionally characterized (both in vitro and in vivo), and immunogenicity assays were carried out. The results showed that both immunotoxins were functionally active and less immunogenic than the wild-type immunotoxin. In addition, IMTXA33furαSDI showed a more efficient antitumor effect (both in vitro and in vivo) due to the inclusion of the furin linker. These results constituted a step forward in the optimization of immunotoxins with low immunogenicity and enhanced antitumor activity, which can lead to potential better outcomes in cancer treatment.

Corrigendum: Suitability of potyviral recombinant virus-like particles bearing a complete food allergen for immunotherapy vaccines.

[This corrects the article DOI: 10.3389/fimmu.2022.986823.].

Novel mediator in anaphylaxis: decreased levels of miR-375-3p in serum and within extracellular vesicles of patients.

Introduction: Anaphylaxis is among the most severe manifestations of allergic disorders, but its molecular basis remains largely unknown and reliable diagnostic markers are not currently available. MicroRNAs (miRNAs) regulate several pathophysiological processes and have been proposed as non-invasive biomarkers. Therefore, this study aims to evaluate their involvement in anaphylactic reaction and their value as biomarkers. Methods: Acute (anaphylaxis) and baseline (control) serum samples from 67 patients with anaphylaxis were studied. Among them, 35 were adults with drug-induced anaphylaxis, 13 adults with food-induced anaphylaxis and 19 children with food-induced anaphylaxis. The circulating serum miRNAs profile was characterized by next-generation sequencing (NGS). For this purpose, acute and baseline samples from 5 adults with drug-induced anaphylaxis were used. RNA was extracted, retrotranscribed, sequenced and the readings obtained were mapped to the human database miRBase_20. In addition, a system biology analysis (SBA) was performed with its target genes and revealed pathways related to anaphylactic mediators signaling. Moreover, functional and molecular endothelial permeability assays were conducted with miR-375-3p-transfected cells in response to cAMP. Results: A total of 334 miRNAs were identified, of which 21 were significant differentially expressed between both phases. Extracellular vesicles (EVs) were characterized by Western blot, electron microscopy and NanoSight. A decrease of miR-375-3p levels was determined by qPCR in both serum and EVs of patients with anaphylaxis (****p<.0001). Precisely, the decrease of miR-375-3p correlated with the increase of two inflammatory cytokines: monocyte chemoattractant protein-1 (MCP-1) and granulocyte macrophage colony-stimulating factor (GM-CSF). On the other hand, functional and molecular data obtained showed that miR-375-3p partially blocked the endothelial barrier maintenance and stabilization by disassembly of cell-cell junctions exhibiting low Rac1-Cdc42 levels. Discussion: These findings demonstrate a differential serum profile of circulating miRNAs in patients with anaphylaxis and exhibit the miR-375-3p modulation in serum and EVs during drug- and food-mediated anaphylactic reactions. Furthermore, the in silico and in vitro studies show a negative role for miR-375-3p/Rac1-Cdc42 in the endothelial barrier stability.

Lipid Ligands and Allergenic LTPs: Redefining the Paradigm of the Protein-Centered Vision in Allergy.

Lipid Transfer Proteins (LTPs) have been described as one of the most prevalent and cross-reactive allergen families in the general population. They are widely distributed among the plant kingdom, as well as in different plant organs ranging from pollen to fruits. Thus, they can initiate allergic reactions with very different outcomes, such as asthma and food allergy. Several mouse models have been developed to unravel the mechanisms that lead LTPs to promote such strong sensitization patterns. Interestingly, the union of certain ligands can strengthen the allergenic capacity of LTPs, suggesting that not only is the protein relevant in the sensitization process, but also the ligands that LTPs carry in their cavity. In fact, different LTPs with pro-allergenic capacity have been shown to transport similar ligands, thus positioning lipids in a central role during the first stages of the allergic response. Here, we offer the latest advances in the use of experimental animals to study the topic, remarking differences among them and providing future researchers a tool to choose the most suitable model to achieve their goals. Also, recent results derived from metabolomic studies in humans are included, highlighting how allergic diseases alter the lipidic metabolism toward a pathogenic state in the individual. Altogether, this review offers a comprehensive body of work that sums up the background evidence supporting the role of lipids as modulators of allergic diseases. Studying the role of lipids during allergic sensitization might broaden our understanding of the molecular events leading to tolerance breakdown in the epithelium, thus helping us to understand how allergy is initiated and established in the individuals.

Identification and molecular characterization of a novel non-specific lipid transfer protein (TdLTP2) from durum wheat.

Non-specific lipid transfer proteins (nsLTPs) are small, cysteine-rich proteins, a part of the pathogenesis-related protein family, and numerous of them act as positive regulators during plant disease resistance, growth, and reproduction. These proteins are involved also in the intracellular transfer of lipids, as well as in plant immune responses. Besides their differences in sequences, they show similar features in their structure. However, they show distinct lipid-binding specificities signifying their various biological roles that dictate further structural study. This study reports the identification, in silico characterization and purification of a novel member of the nsLTP2 protein family from durum wheat, TdLTP2. It was generated and purified using the combination of gel filtration chromatography and reverse-phase high-performance liquid chromatography (RP-HPLC). Its identity was detected by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and mass spectrometry (MALDI-TOF). TdLTP2 had been expressed in different stress to detect its localization; therefore, fluor-immunolocalization studies accomplished this data. In this approach, to assess the allergenicity of TdLTP2, thirty patients with baker's asthma were enrolled and ELISA to detect the presence of specific IgE antibodies tested their sera. Moreover, the lipid-binding properties of TdLTP2 were examined in vitro and validated using a molecular docking study. In summary, our results demonstrate a new addition of member in plant nsLTPs family, TdLTP2, which can develop a better understanding about its biological functions and shed light on future applications.

Plant non-specific lipid transfer proteins: An overview.

Plant non-specific lipid transfer proteins (nsLTPs) are usually defined as small, basic proteins, with a wide distribution in all orders of higher plants. Structurally, nsLTPs contain a conserved motif of eight cysteines, linked by four disulphide bonds, and a hydrophobic cavity in which the ligand is housed. This structure confers stability and enhances the ability to bind and transport a variety of hydrophobic molecules. Their highly conserved structural resemblance but low sequence identity reflects the wide variety of ligands they can carry, as well as the broad biological functions to which they are linked to, such as membrane stabilization, cell wall organization and signal transduction. In addition, they have also been described as essential in resistance to biotic and abiotic stresses, plant growth and development, seed development, and germination. Hence, there is growing interest in this family of proteins for their critical roles in plant development and for the many unresolved questions that need to be clarified, regarding their subcellular localization, transfer capacity, expression profile, biological function, and evolution.

Antitumor applications of polyphenol-conjugated turnip mosaic virus-derived nanoparticles.

Background: Filamentous plant virus-derived nanoparticles are biodegradable and noninfectious to humans. Their structure is also amenable to chemical modifications. They constitute an appealing material for biomedical applications including imaging and drug delivery. We had previously used turnip mosaic virus-derived nanoparticles (TuMV-NPs) to increase antibody-sensing in vivo, to prevent biofilm formation and to build biological nanoscaffolds. Materials & methods: We analyzed TuMV-NP biodistribution and tumor homing using in vivo imaging. We studied in vitro the interaction with human cancer cell lines and the antiproliferative effect of epigallocatechin gallate-functionalized TuMV-NPs. Results & conclusion: TuMV-NPs are efficiently internalized by human cells and show good tumor homing. The antiproliferative effect of epigallocatechin gallate-TuMV-NPs suggests that they could offer a potential anticancer therapy.

Cancer is the second leading cause of death worldwide, just behind cardiovascular disease. It accounts for nearly 10 million deaths annually, and new strategies to improve early detection and drug delivery are urgently needed. Nanoparticles are small structures within the nanometer range (1 billionth of a meter) that can be used to deliver either an imaging probe (tracer) to allow the detection of a tumor or drugs to kill tumor cells. There are many types of nanoparticles; those based on plant viruses are especially appealing for biomedical purposes because they are biodegradable and noninfectious to humans. Also, their physicochemical properties, such as symmetry, uniformity and loading capacity, make them excellent nanocarriers. We report here for the first time the ability of nanoparticles derived from the turnip mosaic virus (TuMV), a well-known virus naturally infecting cruciferous plants (e.g., broccoli, turnip, radish, cabbage) but not humans, to deliver a fluorescent imaging probe that allows tumor detection in vivo. Moreover, TuMV nanoparticles were used to deliver a natural chemotherapeutic agent of plant origin to different types of tumor cells (lung, colorectal, breast, and head and neck), showing increased antiproliferative capacity compared to the nonvehiculized drug.

NLRP3 priming due to skin damage precedes LTP allergic sensitization in a mouse model.

Allergic sensitization is initiated by protein and epithelia interaction, although the molecular mechanisms leading this encounter toward an allergic phenotype remain unknown. Here, we apply the two-hit hypothesis of inflammatory diseases to the study of food allergy sensitization. First, we studied the effects of long-term depilation in mice by analyzing samples at different time points. Several weeks of depilation were needed until clear immunological changes were evidenced, starting with upregulation of NLRP3 protein levels, which was followed by overexpression of Il1b and Il18 transcripts. Secondly, we assessed the effects of allergen addition (in this case, Pru p 3 in complex with its natural lipid ligand) over depilated skin. Systemic sensitization was evaluated by intraperitoneal provocation with Pru p 3 and measure of body temperature. Anaphylaxis was achieved, but only in mice sensitized with Prup3_complex and not treated with the NLRP3 inhibitor MCC950, thus demonstrating the importance of both hits (depilation + allergen addition) in the consecution of the allergic phenotype. In addition, allergen encounter (but not depilation) promoted skin remodeling, as well as CD45+ infiltration not only in the sensitized area (the skin), but across several mucosal tissues (skin, lungs, and gut), furtherly validating the systemization of the response. Finally, a low-scale study with human ILC2s is reported, where we demonstrate that Prup3_complex can induce their phenotype switch (↑CD86, ↑S1P1) when cultured in vitro, although more data is needed to understand the implications of these changes in food allergy development.

Alt a 1 Promotes Allergic Asthma In Vivo Through TLR4-Alveolar Macrophages.

The mold Alternaria alternata is one of the main sources of asthma exacerbation, being its major allergen, Alt a 1, indispensable for its development. The main objective of this work was to answer two main questions: 1) can Alt a 1 by itself (without any other context) induce an asthmatic profile in vivo?; and 2) Which molecular mechanisms take place during this phenomenon? To answer both questions, we have developed a mouse model of allergic asthma using only Alt a 1 for mice sensitization. We also made use of in-vitro cellular models and computational studies to support some aspects of our hypothesis. Our results showed that Alt a 1 can induce an asthmatic phenotype, promoting tissue remodeling and infiltration of CD45+ cells, especially eosinophils and macrophages (Siglec F+ and F4/80+). Also, we have found that Alt a 1 sensitization is mediated by the TLR4-macrophage axis.

Suitability of potyviral recombinant virus-like particles bearing a complete food allergen for immunotherapy vaccines.

Virus-like particles (VLPs) have been gaining attention as potential platforms for delivery of cargos in nanomedicine. Although animal viruses are largely selected due to their immunostimulatory capacities, VLPs from plant viruses constitute a promising alternative to be considered. VLPs derived from Turnip mosaic virus (TuMV) have proven to present a tridimensional structure suited to display molecules of interest on their surface, making them interesting tools to be studied in theragnostic strategies. Here, we study their potential in the treatment of food allergy by genetically coupling TuMV-derived VLPs to Pru p 3, one of the most dominant allergens in Mediterranean climates. VLPs-Pru p 3 were generated by cloning a synthetic gene encoding the TuMV coat protein and Pru p 3, separated by a linker, into a transient high-expression vector, followed by agroinfiltration in Nicotiana benthamiana plants. The generated fusion protein self-assembled in planta to form the VLPs, which were purified by exclusion chromatography. Their elongated morphology was confirmed by electron microscopy and their size (~400 nm), and monodispersity was confirmed by dynamic light scattering. Initial in vitro characterization confirmed that they were able to induce proliferation of human immune cells. This proliferative capability was enhanced when coupled with the natural lipid ligand of Pru p 3. The resultant formulation, called VLP-Complex, was also able to be transported by intestinal epithelial cells, without affecting the monolayer integrity. In light of all these results, VLP-Complex was furtherly tested in a mouse model of food allergy. Sublingual administration of VLP-Complex could effectively reduce some serological markers associated with allergic responses in mice, such as anti-Pru p 3 sIgE and sIgG2a. Noteworthy, no associated macroscopic, nephritic, or hepatic toxicity was detected, as assessed by weight, blood urea nitrogen (BUN) and galectin-3 analyses, respectively. Our results highlight the standardized production of allergen-coated TuMV-VLPs in N. benthamiana plants. The resulting formula exerts notable immunomodulatory properties without the need for potentially hazardous adjuvants. Accordingly, no detectable toxicity associated to their administration was detected. As a result, we propose them as good candidates to be furtherly studied in the treatment of immune-based pathologies.