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Aminoacid sequences and localization within the ASP-2 protein of the peptide pools used in this study.
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Targeting antigens to dendritic cells (DCs) by using hybrid monoclonal antibodies (mAbs) directed against DC receptors is known to improve activation and support long-lasting T cell responses. In the present work, we used the mAb αDEC205 fused to the Trypanosoma cruzi amastigote surface protein 2 (ASP-2) to identify a region of this protein recogni...
Citations
... As previously demonstrated with proteins from other pathogens, it was possible to map a new CD4+ T cell epitope present in this protein. 48 Toxoplasma gondii tachyzoites main surface antigen, SAG1, was fused to a variable chain fragment of the αDEC205 mAb (scDEC). Intranasal and subcutaneous immunizations together with Poly (I:C) induced protective responses against chronic infection that were probably mediated by polyfunctional Th1 cells. ...
Dendritic cells are central to the development of immunity, as they are specialized in initiating antigen-specific immune responses. In this review, we briefly present the existing knowledge on dendritic cell biology and how their division in different dendritic cell subsets may impact the development of immune responses. In addition, we explore the use of chimeric monoclonal antibodies that bind to dendritic cell surface receptors, with an emphasis on the C-type lectin family of endocytic receptors, to deliver antigens directly to these cells. Promising preclinical studies have shown that it is possible to modulate the development of immune responses to different pathogens when monoclonal antibodies fused to pathogen-derived antigens are used to deliver the antigen to different subsets of dendritic cells. This approach can be used to improve the efficacy of vaccines against different pathogens.
... To improve the immunogenicity of the DNA vaccines, we combined two different approaches: delivery of the DNA vaccine by electroporation and targeting the encoded antigen to a specialized dendritic cell subset (CD8α + ) using a single-chain antibody to the endocytic receptor DEC205 (scDEC). Previous studies revealed promising results using DNA vaccines that targeted antigens to dendritic cells against a variety of pathogens, including HIV [50,52], influenza [57], Yersinia pestis [58], P. yoelii [59], P. falciparum [47], T.cruzi [60], M. tuberculosis [61], T. gondii [62], dengue virus [55] and cancer [63]. This platform offers attractive advantages including stability, low cost of production, and the absence of the Fc domain [51]. ...
Chikungunya virus (CHIKV) has become a significant public health concern due to the increasing number of outbreaks worldwide and the associated comorbidities. Despite substantial efforts, there is no specific treatment or licensed vaccine against CHIKV to date. The E2 glycoprotein of CHIKV is a promising vaccine candidate as it is a major target of neutralizing antibodies during infection. In this study, we evaluated the immunogenicity of two DNA vaccines (a non-targeted and a dendritic cell-targeted vaccine) encoding a consensus sequence of E2 CHIKV and a recombinant protein (E2* CHIKV). Mice were immunized with different homologous and heterologous DNAprime-E2* protein boost strategies, and the specific humoral and cellular immune responses were accessed. We found that mice immunized with heterologous non-targeted DNA prime-E2* CHIKV protein boost developed high levels of neutralizing antibodies, as well as specific IFN-γ producing cells and poly-functional CD4 + and CD8 + T cells. We also identified 14 potential epitopes along the E2 CHIKV protein. Furthermore, immunization with recombinant E2* CHIKV combined with the adjuvant AS03 presented the highest humoral response with neutralizing capacity. Finally, we show that the heterologous prime-boost strategy with the non-targeted pVAX-E2 DNA vaccine as the prime followed by E2* protein + AS03 boost is a promising combination to elicit a broad humoral and cellular immune response. Together, our data highlights the importance of E2 CHIKV for the development of a CHIKV vaccine.
... Much has been done to identify the unique roles of cDC1 and cDC2, specially on their differential ability to prime different CD4 + T cell responses (12)(13)(14)(15)(16)(17)(18)(19)(20)(21)(22)(23)(24)(25)(26). An efficient strategy to assess the function and biology of cDC in vivo consists of delivering antigens directly to these cells using chimeric monoclonal antibodies (mAb). ...
... The aDEC205 mAb has been widely used to target antigens to cDC1 via the DEC205 receptor. Antigen targeting to cDC1 promotes Th1 and Th1-like Tfh CD4 + T cell responses after immunization (8,11,(13)(14)(15)(17)(18)(19)(20)(21)(22)(23)(24)(25)(26)(27)(28)(29). The aDCIR2 (33D1) mAb has been used to target antigens to cDC2. ...
Conventional dendritic cells (cDC) are a group of antigen-presenting cells specialized in priming T cell responses. In mice, splenic cDC are divided into conventional type 1 DC (cDC1) and conventional type 2 (cDC2). cDC1 are specialized to prime the Th1 CD4⁺ T cell response, while cDC2 are mainly associated with the induction of follicular helper T cell responses to support germinal center formation. However, the mechanisms that control the functions of cDC1 and cDC2 are not fully understood, especially the signaling pathways that can modulate their ability to promote different CD4⁺ T cell responses. Here, we targeted a model antigen for cDC1 and cDC2, through DEC205 and DCIR2 receptors, respectively, to study the role of the STAT3 signaling pathway in the ability of these cells to prime CD4⁺ T cells. Our results show that, in the absence of the STAT3 signaling pathway, antigen targeting to cDC2 induced similar frequencies of Tfh cells between STAT3-deficient mice compared to fully competent mice. On the other hand, Th1 and Th1-like Tfh cell responses were significantly reduced in STAT3-deficient mice after antigen targeting to cDC1 via the DEC205 receptor. In summary, our results indicate that STAT3 signaling does not control the ability of cDC2 to promote Tfh cell responses after antigen targeting via the DCIR2 receptor, but modulates the function of cDC1 to promote Th1 and Th1-like Tfh T cell responses after antigen targeting via the DEC205 receptor.
... Therefore, antigens have been targeted in vivo to the DEC-205 receptor by conjugation with a specific anti-DEC-205 mAb to stimulate antigen presentation by DCs. Moreover, potent protective responses against different infectious agents and cancer have been achieved when used together with a maturation stimulus (33)(34)(35)(36)(37)(38)(39)(40)(41)(42)(43). Thus, targeting tumor antigens to DCs through DEC-205 is a promising alternative for the treatment of malignant tumors. ...
HPV E5 is an oncoprotein mainly expressed in premalignant lesions, which makes it an important target for a vaccine to prevent or cure cervical cancer (CC). In this study, we evaluated whether E5 targeted to DEC-205, present in dendritic cells (DCs), could induce a therapeutic protection against HPV16-induced tumor cells in a mouse model. The HPV-16 E5 (16E5) protein was cross-linked to a monoclonal antibody (mAb) specific to mouse DEC-205 (anti-DEC-205:16E5) or to an isotype control mAb (isotype:16E5). Rotavirus VP6 was cross-linked to the mouse anti-DEC-205 mAb (anti-DEC-205:VP6) as a non-specific antigen control. BALB/c mice were inoculated subcutaneously (s.c.) with the 16E5-expressing BMK-16/myc tumor cells, and 7 and 14 days later the mice were immunized s.c. with the conjugates, free 16E5 or PBS in the presence of adjuvant. Tumor growth was monitored to evaluate protection. A strong protective immune response against the tumor cells was induced when the mice were inoculated with the anti-DEC-205:16E5 conjugate, since 70% of the mice controlled the tumor growth and survived, whereas the remaining 30% developed tumors and died by day 72. In contrast, 100% of the mice in the control groups died by day 30. The anti-DEC-205:16E5 conjugate was found to induce 16E5-specific memory T cells, with a Th1/Th17 profile. Both CD4⁺ and CD8⁺ T cells contributed to the observed protection. Finally, treating mice that had developed tumors with an anti-PD-1 mAb, delayed the tumor growth for more than 20 days. These results show that targeting 16E5 to DEC-205, alone or combined with an immune checkpoint blockade, could be a promising protocol for the treatment of the early stages of HPV-associated cancer.
... Murine splenic cDC1s express the DEC205 (CD205) receptor, while cDC2s express DCIR2 (DC immunoreceptor 2), both members of the C-type lectin family of endocytic receptors [22,23]. Anti-DEC205 (αDEC) and anti-DCIR2 (αDCIR2) chimeric mAbs were effectively used to target proteins to either cDC1s or cDC2s, respectively, in an attempt to improve cellular and humoral immune responses [5,14,[24][25][26][27][28][29][30][31]. In some cases, partial protection against different pathogens was observed in vaccine approaches [28,29,31]. ...
... Likewise, antigen targeting to cDC1s and cDC2s via DEC205 and DCIR2 receptors, respectively, was also used as a strategy to study the biology and function of both cDCs subsets [5,6,15]. Antigen targeting to cDC1s via DEC205 receptor is able to induce cross-presentation to CD8 + T cells, cytotoxicity, and a T H 1-biased immune response in the presence of DC maturation stimuli [11,14,24,26,29,30,32]. For this particular model, if the chimeric mAb is delivered in the absence of a maturation stimulus, the outcome is T cell tolerance [11,33]. ...
... Previous studies indicated that immunization with αDEC could induce high antibody titers after the administration of a booster dose [14,24,[27][28][29][30][31]. In this sense, we set out to systematically compare the induction of T H and humoral immune responses using antigen targeting to cDC1s and cDC2s after the administration of one or two doses of the chimeric mAbs. ...
Conventional dendritic cells (cDCs) are specialized in antigen presentation. In the mouse spleen, cDCs are classified in cDC1s and cDC2s, and express DEC205 and DCIR2 endocytic receptors, respectively. Monoclonal antibodies (mAbs) αDEC205 (αDEC) and αDCIR2 have been fused to different antigens to deliver them to cDC1s or cDC2s. We immunized mice with αDEC and αDCIR2 fused to an antigen using Poly (I:C) as adjuvant. The initial immune response was analyzed from days 3 to 6 after the immunization. We also studied the influence of a booster dose. Our results showed that antigen targeting to cDC1s promoted a pro‐inflammatory TH1 cell response. Antigen targeting to cDC2s induced TFH cells, germinal centers and plasma cell differentiation. After boost, antigen targeting to cDC1s improved the TH1 cell response and induced TH1‐like TFH cells that led to an increase in specific antibody titers and IgG class switch. Additionally, a population of regulatory T cells was also observed. Antigen targeting to cDC2s did not improve the specific antibody response after boost. Our results add new information on the immune response induced after the administration of a booster dose with αDEC and αDCIR2 fusion mAbs. These results may be useful for vaccine design using recombinant mAbs.
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... Plasmids encoding the light and heavy chain of the mouse αDEC205 antibody were kindly provided by Dr. Michel C. Nussenzweig (The Rockefeller University, New York, USA). The plasmid encoding the heavy chain of the mouse DEC205 fused to eight HIV-1 epitopes was previously described and contains the following epitopes: p6 (32)(33)(34)(35)(36)(37)(38)(39)(40)(41)(42)(43)(44)(45)(46), p17 (73-89), pol (785-799), gp160 (188-201), rev (11)(12)(13)(14)(15)(16)(17)(18)(19)(20)(21)(22)(23)(24)(25)(26)(27), vpr (65-82), vif (144-158), and nef (180-194) (39). ...
... Antigen targeting to DCs through DEC205 receptor is used as a vaccination strategy to induce strong antigen-specific immune responses against several pathogens (26,34,55) and tumors In the HIV vaccine scenario, antigen targeting to cDC1 through DEC205 was performed using the full-length gag (p24) protein (21,51,(59)(60)(61)(62). The success in different pre-clinical studies using mice and non-human primates (16) quickly pushed forward the translation of this strategy to humans. ...
Cellular immune responses are implicated in resistance to HIV and have been considered for the development of an effective vaccine. Despite their safety profile, subunit vaccines need to be delivered combined with an adjuvant. In the last years, in vivo antigen targeting to dendritic cells (DCs) using chimeric monoclonal antibodies (mAb) against the DC endocytic receptor DEC205/CD205 was shown to support long-term T cell immunity. Here, we evaluated the ability of different adjuvants to modulate specific cellular immune response when eight CD4⁺ HIV-derived epitopes (HIVBr8) were targeted to DEC205⁺ DCs in vivo. Immunization with two doses of αDECHIVBr8 mAb along with poly(I:C) induced Th1 cytokine production and higher frequency of HIV-specific polyfunctional and long-lived T cells than MPL or CpG ODN-assisted immunization. Although each adjuvant elicited responses against the 8 epitopes present in the vaccine, the magnitude of the T cell response was higher in the presence of poly(I:C). Moreover, poly(I:C) up regulated the expression of costimulatory molecules in both cDC1 and cDC2 DCs subsets. In summary, the use of poly(I:C) in a vaccine formulation that targets multiple epitopes to the DEC205 receptor improved the potency and the quality of HIV-specific responses when compared to other vaccine-adjuvant formulations. This study highlights the importance of the rational selection of antigen/adjuvant combination to potentiate the desired immune responses.
... Antigens derived from different pathogens have been targeted to DEC205 + CD8α + DCs using chimeric anti-DEC205 monoclonal antibodies (mAbs) genetically fused to them, administered in the presence of a DC maturation stimulus (26)(27)(28)(29)(30)(31)(32)(33)(34). ...
... CD4 + T cell epitopes derived from different proteins have been more easily mapped in samples derived from animals submitted to antigen targeting to DCs. CD4 + T cells epitopes were detected in the Plasmodium yoelii circunsporozoite protein (27), in the HIV p24 gag (51), in the Leishmania major LACK antigen (64), in the Yersinia pestis LcrV antigen (65) and in the Trypanosoma cruzi ASP-2 protein (31). ...
Dengue fever has become a global threat, causing millions of infections every year. An effective vaccine against all four serotypes of dengue virus (DENV) has not been developed yet. Among the different vaccination strategies available today, DNA vaccines are safe and practical, but currently induce relatively weak immune responses in humans. In order to improve immunogenicity, antigens may be targeted to dendritic cells (DCs), the main antigen presenting cells and orchestrators of the adaptive immune response, inducing T and B cell activation. It was previously shown that a DNA vaccine encoding a fusion protein comprised of an antigen and a single-chain Fv antibody (scFv) specific for the DC endocytic receptor DEC205 induced strong immune responses to the targeted antigen. In this work, we evaluate this strategy to improve the immunogenicity of dengue virus (DENV) proteins. Plasmids encoding the scFv αDEC205, or an isotype control (scFv ISO), fused to the DENV2 envelope protein domain III (EDIII) were generated, and EDIII specific immune responses were evaluated in immunized mice. BALB/c mice were intramuscularly (i.m.) immunized three times with plasmid DNAs encoding either scDEC-EDIII or scISO-EDIII followed by electroporation. Analyses of the antibody responses indicated that EDIII fusion with scFv targeting the DEC205 receptor significantly enhanced serum anti-EDIII IgG titers that inhibited DENV2 infection. Similarly, mice immunized with the scDEC-EDIII plasmid developed a robust CD4⁺ T cell response to the targeted antigen, allowing the identification of two linear epitopes recognized by the BALB/c haplotype. Taken together, these results indicate that targeting DENV2 EDIII protein to DCs using a DNA vaccine encoding the scFv αDEC205 improves both antibody and CD4⁺ T cell responses. This strategy opens perspectives for the use of DNA vaccines that encode antigens targeted to DCs as a strategy to increase immunogenicity.
... Although significant information has been obtained by studying activation markers and cytokines secreted by CD4 + and CD8 + T cells during T. cruzi infection [3], knowledge about the fine specificity of these cells is restricted to a few parasite epitopes. Most of these are peptides from proteins belonging to the trans-sialidase family, like TS, ASP-1, ASP-2 [19][20][21]. ...
Human CD8+ and CD4+ T cell lines and clones are valuable tools to explore the role of these cells in the context of diseases, especially in cases in which the main underlying actor is the immune response, like Chagas disease. These cell lines and clones provide a good experimental system to address the phenotypic and functional features of specific T cell subpopulations and furthermore settle the framework necessary for analyzing their antigen/peptide specificity.This chapter details a culture method for the establishment of T. cruzi-specific memory T cell lines from mononuclear cells isolated from Chagas disease patients' peripheral blood. The presented protocol comprises (1) enrichment of memory CD4+ T cells, (2) stimulation with parasite lysate, (3) evaluation of specificity, and (4) expansion and maintenance of specific T cell lines.
... Chagas Evaluación de la evolución y regulación del proceso infeccioso. (Clemente et al., 2016) Identificación de nuevos epítopos de células T. (Rampazo et al., 2015) Dengue Determinación de las regiones de unión de anticuerpos monoclonales de dengue a proteína de envoltura del virus. (Tang et al., 2015) Evaluación de anticuerpos monoclonales generados para la detección de proteínas de envoltura del virus. ...
Los anticuerpos son glicoproteínas, componentes moleculares y humorales del sistema
inmune, que tienen la capacidad de reconocer y bloquear virus, bacterias, parásitos u hongos. Los anticuerpos monoclonales, producidos por los linfocitos B, son las herramientas más novedosas en el campo clínico y biotecnológico, de gran utilidad en el
diagnóstico y tratamiento de enfermedades infecciosas, inmunológicas y neoplásicas, así como en el estudio de la interacción antígeno–anticuerpo, y la marcación, detección y cuantificación de moléculas y células. Con la incorporación de las técnicas de biología molecular e ingeniería genética y proteica se ha ampliado el horizonte de la generación de anticuerpos monoclonales y sus usos, y se han desarrollado técnicas como la hibridación, la quimerización, la humanización y la producción de anticuerpos monoclonales totalmente humanos. Es una de las áreas de mayor crecimiento en la industria biotecnológica y farmacéutica. El capítulo presenta los aspectos relevantes de los anticuerpos monoclonales, como herramienta para el diagnóstico y la investigación.
... These results are in agreement with those obtained by others showing that in combination with TLR stimulation, DEC205 targeting induced a robust CD4+ T cell immune response (47)(48)(49) and only low level of CD8+ T cell response (25,40). One exception is OVA protein for which both potent CD4+ and CD8+ responses were obtained (39,50,51). ...
Toxoplasmosis is a major public health problem and the development of a human vaccine is of high priority. Efficient vaccination against Toxoplasma gondii requires both a mucosal and systemic Th1 immune response. Moreover, dendritic cells play a critical role in orchestrating the innate immune functions and driving specific adaptive immunity to T. gondii. In this study, we explore an original vaccination strategy that combines administration via mucosal and systemic routes of fusion proteins able to target the major T. gondii surface antigen SAG1 to DCs using an antibody fragment single-chain fragment variable (scFv) directed against DEC205 endocytic receptor. Our results show that SAG1 targeting to DCs by scFv via intranasal and subcutaneous administration improved protection against chronic T. gondii infection. A marked reduction in brain parasite burden is observed when compared with the intranasal or the subcutaneous route alone. DC targeting improved both local and systemic humoral and cellular immune responses and potentiated more specifically the Th1 response profile by more efficient production of IFN-γ, interleukin-2, IgG2a, and nasal IgA. This study provides evidence of the potential of DC targeting for the development of new vaccines against a range of Apicomplexa parasites.
