Asunto(s)
Adenoviridae , Leishmania infantum , Leishmaniasis Visceral , Leishmania infantum/inmunología , Leishmania infantum/genética , Animales , Leishmaniasis Visceral/prevención & control , Leishmaniasis Visceral/inmunología , Adenoviridae/genética , Adenoviridae/inmunología , Antígenos de Protozoos/inmunología , Antígenos de Protozoos/genética , Modelos Animales de Enfermedad , Vacunas contra la Leishmaniasis/inmunología , Humanos , Vacunación , Vacunas Sintéticas/inmunología , Vacunas Sintéticas/administración & dosificaciónRESUMEN
American Tegumentary Leishmaniasis (ATL) is a disease of high severity and incidence in Brazil, in addition to being a worldwide concern in public health. Leishmania amazonensis is one of the etiological agents of ATL, and the inefficiency of control measures, associated with the high toxicity of the treatment and the lack of effective immunoprophylactic strategies, makes the development of vaccines indispensable and imminent. In this light, the present study proposes to elaborate a chimeric protein (rChiP), based on the fusion of multiple epitopes of CD4+/CD8+ T cells, identified in the immunoproteome of the parasites L. amazonensis and L. braziliensis. The designed chimeric protein was tested in the L. amazonensis murine model of infection using the following formulations: 25 µg of the rChiP in saline (rChiP group) and 25 µg of the rChiP plus 25 µg of MPLA-PHAD® (rChiP+MPLA group). After completing immunization, CD4+ and CD8+ T cells, stimulated with SLa-Antigen or rChiP, showed an increased production of nitric oxide and intracytoplasmic pro-inflammatory cytokines, in addition to the generation of central and effector memory T cells. rChiP and rChiP+MPLA formulations were able to promote an effective protection against L. amazonensis infection determined by a reduction in the development of skin lesions and lower parasitic burden. Reduction in the development of skin lesions and lower parasitic burden in the vaccinated groups were associated with an increase of nitrite, CD4+/CD8+IFN-γ+TNF-α+ and CD4+/CD8+CD44highCD62Lhigh/low T cells, IgGTotal, IgG2a, and lower rates of IgG1 and CD4+/CD8+IL-10+. This data suggests that proposed formulations could be considered potential tools to prevent ATL.
Asunto(s)
Adyuvantes Inmunológicos , Linfocitos T CD4-Positivos , Linfocitos T CD8-positivos , Epítopos de Linfocito T , Memoria Inmunológica , Vacunas contra la Leishmaniasis , Leishmaniasis Cutánea , Animales , Leishmaniasis Cutánea/prevención & control , Leishmaniasis Cutánea/inmunología , Linfocitos T CD8-positivos/inmunología , Linfocitos T CD4-Positivos/inmunología , Epítopos de Linfocito T/inmunología , Ratones , Vacunas contra la Leishmaniasis/inmunología , Femenino , Adyuvantes Inmunológicos/administración & dosificación , Ratones Endogámicos BALB C , Proteínas Recombinantes de Fusión/inmunología , Proteínas Recombinantes de Fusión/genética , Leishmania braziliensis/inmunología , Lípido A/análogos & derivados , Lípido A/inmunología , Anticuerpos Antiprotozoarios/inmunología , Citocinas/metabolismo , Citocinas/inmunología , Modelos Animales de Enfermedad , Antígenos de Protozoos/inmunologíaRESUMEN
Visceral Leishmaniasis is a serious public health problem caused by Leishmania species parasites. Approximately 500 thousand people get Visceral Leishmaniasis (VL) every year. An effective and reliable vaccine against the disease has still not been formulated. Choosing the right adjuvant is important to increase immunogenicity in vaccines prepared with total antigens. In this study, we investigate the ideal adjuvant for use in vaccine formulations against VL. For this purpose, Leishmania antigens (FTLA) obtained from L. infantum parasites by the freeze-thaw method and three different adjuvants (alum-saponin and calcium phosphate) were used. The effectiveness of the formulations was investigated in vitro by cell viability analysis and determination of nitric oxide and cytokine production abilities in J774 macrophage cells. According to the study results, it was determined that formulations prepared with calcium phosphate produced 72% more NO and approximately 7.2 times more IL-12 cytokine. The results obtained showed that calcium phosphate salts can be used as ideal adjuvants in vaccine research against leishmaniasis.
Asunto(s)
Antígenos de Protozoos , Leishmania infantum , Vacunas contra la Leishmaniasis , Animales , Ratones , Vacunas contra la Leishmaniasis/inmunología , Leishmania infantum/inmunología , Antígenos de Protozoos/inmunología , Línea Celular , Macrófagos/inmunología , Vacunas de Productos Inactivados/inmunología , Vacunas de Productos Inactivados/administración & dosificación , Óxido Nítrico/metabolismo , Fosfatos de Calcio , Citocinas/metabolismo , Adyuvantes de Vacunas , Leishmaniasis Visceral/prevención & control , Leishmaniasis Visceral/inmunología , Saponinas/farmacología , Compuestos de Alumbre/administración & dosificación , Adyuvantes Inmunológicos/administración & dosificación , Supervivencia Celular/efectos de los fármacosRESUMEN
Despite the development of a vaccine against cutaneous leishmaniasis in preclinical and clinical studies, we still do not have a safe and effective vaccine for human use. Given this situation, the search for a new prophylactic alternative to control leishmaniasis should be a global priority. A first-generation vaccine strategy-leishmanization, in which live Leishmania major parasites are inoculated into the skin to protect against reinfection, is taking advantage of this situation. Live attenuated Leishmania vaccine candidates are promising alternatives due to their robust protective immune responses. Importantly, they do not cause disease and could provide long-term protection following challenges with a virulent strain. In addition to physical and chemical methods, genetic tools, including the Cre-loxP system, have enabled the selection of safer null mutant live attenuated Leishmania parasites obtained by gene disruption. This was followed by the discovery and introduction of CRISPR/Cas-based gene editing tools, which can be easily and precisely used to modify genes. Here, we briefly review the immunopathology of L. major parasites and then present the classical methods and their limitations for the production of live attenuated vaccines. We then discuss the potential of current genetic engineering tools to generate live attenuated vaccine strains by targeting key genes involved in L. major pathogenesis and then discuss their discovery and implications for immune responses to control leishmaniasis.
Asunto(s)
Ingeniería Genética , Leishmania major , Vacunas contra la Leishmaniasis , Leishmaniasis Cutánea , Vacunas Atenuadas , Vacunas Atenuadas/inmunología , Leishmaniasis Cutánea/prevención & control , Humanos , Vacunas contra la Leishmaniasis/inmunología , Leishmania major/inmunología , Leishmania major/genética , Animales , Inmunización , Edición GénicaRESUMEN
Centrin1 gene deleted Leishmania donovani parasite (LdCen1-/-) was developed and extensively tested experimentally as an intracellular stage-specific attenuated and immunoprotective live parasite vaccine candidate ex vivo using human PBMCs and in vivo in animals. Here we report manufacturing and pre-clinical evaluation of current Good-Laboratory Practice (cGLP) grade LdCen1-/- parasites, as a prerequisite before proceeding with clinical trials. We screened three batches of LdCen1-/- parasites manufactured in bioreactors under cGLP conditions, for their consistency in genetic stability, attenuation, and safety. One such batch was preclinically tested using human PBMCs and animals (hamsters and dogs) for its safety and protective immunogenicity. The immunogenicity of the CGLP grade LdCen1-/- parasites was similar to one grown under laboratory conditions. The cGLP grade LdCen1-/- parasites were found to be safe and non-toxic in hamsters and dogs even at 3 times the anticipated vaccine dose. When PBMCs from healed visceral leishmaniasis (VL) cases were infected with cGLP LdCen1-/-, there was a significant increase in the stimulation of cytokines that contribute to protective responses against VL. This effect, measured by multiplex ELISA, was greater than that observed in PBMCs from healthy individuals. These results suggest that cGLP grade LdCen1-/- manufactured under cGMP complaint conditions can be suitable for future clinical trials.
Asunto(s)
Eliminación de Gen , Leishmania donovani , Leishmaniasis Visceral , Vacunas Atenuadas , Leishmania donovani/inmunología , Leishmania donovani/genética , Animales , Humanos , Perros , Vacunas Atenuadas/inmunología , Leishmaniasis Visceral/prevención & control , Leishmaniasis Visceral/inmunología , Leishmaniasis Visceral/parasitología , Cricetinae , Vacunas contra la Leishmaniasis/inmunología , Vacunas contra la Leishmaniasis/genética , Proteínas Protozoarias/genética , Proteínas Protozoarias/inmunología , Leucocitos Mononucleares/inmunología , FemeninoRESUMEN
Leishmania donovani surface glycoprotein 63 (GP63) is a major virulence factor involved in parasite escape and immune evasion. In this study, we generated virus-like particles (VLPs) expressing L. donovani GP63 using the baculovirus expression system. Mice were intramuscularly immunized with GP63-VLPs and challenged with L. donovani promastigotes. GP63-VLP immunization elicited higher levels of L. donovani antigen-specific serum antibodies and enhanced splenic B cell, germinal center B cell, CD4+, and CD8+ T cell responses compared to unimmunized controls. GP63-VLPs inhibited the influx of pro-inflammatory cytokines IFN-γ and IL-6 in the livers, as well as thwarting the development of splenomegaly in immunized mice. Upon L. donovani challenge infection, a drastic reduction in splenic parasite burden was observed in VLP-immunized mice. These results indicate that GP63-VLPs immunization conferred protection against L. donovani challenge infection by inducing humoral and cellular immunity in mice.
Asunto(s)
Leishmania donovani , Leishmaniasis Visceral , Ratones Endogámicos BALB C , Vacunas de Partículas Similares a Virus , Animales , Leishmania donovani/inmunología , Ratones , Vacunas de Partículas Similares a Virus/inmunología , Vacunas de Partículas Similares a Virus/administración & dosificación , Femenino , Leishmaniasis Visceral/prevención & control , Leishmaniasis Visceral/inmunología , Anticuerpos Antiprotozoarios/sangre , Anticuerpos Antiprotozoarios/inmunología , Vacunas contra la Leishmaniasis/inmunología , Vacunas contra la Leishmaniasis/administración & dosificación , Eficacia de las Vacunas , Inmunidad Celular , Bazo/inmunología , Linfocitos T CD8-positivos/inmunología , Linfocitos B/inmunología , Inmunidad Humoral , Glicoproteínas de Membrana/inmunología , Glicoproteínas de Membrana/genética , Citocinas/inmunología , MetaloendopeptidasasRESUMEN
Leishmaniasis, caused by Leishmania (L.) species, remains a neglected infection. Therapeutic vaccination presents a promising strategy for its treatment. In this study, we aimed to develop a therapeutic vaccine candidate using Leishmaniaantigens (SLA) and Toll-like receptor (TLR) 7/8 agonist (R848) encapsulated into the poly (lactic-co-glycolic acid) (PLGA) nanoparticles (NPs). Moreover, TLR1/2 agonist (Pam3CSK4) was loaded onto the NPs. The therapeutic effects of these NPs were evaluated in L. major-infected BALB/c mice. Footpad swelling, parasite load, cellular and humoral immune responses, and nitric oxide (NO) production were analyzed. The results demonstrated that the PLGA NPs (SLA-R848-Pam3CSK4) therapeutic vaccine effectively stimulated Th1 cell responses, induced humoral responses, promoted NO production, and restricted parasite burden and lesion size.Our findings suggest that vaccination with SLA combined with TLR1/2 and TLR7/8 agonists in PLGA NPs as a therapeutic vaccine confers strong protection againstL. majorinfection. These results represent a novel particulate therapeutic vaccine against Old World cutaneous leishmaniasis.
Asunto(s)
Antígenos de Protozoos , Vacunas contra la Leishmaniasis , Leishmaniasis Cutánea , Ratones Endogámicos BALB C , Nanopartículas , Copolímero de Ácido Poliláctico-Ácido Poliglicólico , Animales , Leishmaniasis Cutánea/inmunología , Leishmaniasis Cutánea/tratamiento farmacológico , Copolímero de Ácido Poliláctico-Ácido Poliglicólico/química , Antígenos de Protozoos/inmunología , Nanopartículas/química , Vacunas contra la Leishmaniasis/inmunología , Ratones , Femenino , Óxido Nítrico/metabolismo , Imidazoles/farmacología , Imidazoles/química , Células TH1/inmunología , Leishmania major/inmunología , Receptores Toll-Like/agonistas , Humanos , Receptor Toll-Like 7/agonistas , Agonistas de los Receptores Toll-LikeRESUMEN
Infectious diseases like leishmaniasis, malaria, HIV, tuberculosis, leprosy and filariasis are responsible for an immense burden on public health systems. Among these, leishmaniasis is under the category I diseases as it is selected by WHO (World Health Organization) on the ground of diversity and complexity. High cost, resistance and toxic effects of Leishmania traditional drugs entail identification and development of therapeutic alternative. Since the natural infection elicits robust immunity, consistence efforts are going on to develop a successful vaccine. Clinical trials have been conducted on vaccines like Leish-F1, F2, and F3 formulated using specific Leishmania antigen epitopes. Current strategies utilize individual or combined antigens from the parasite or its insect vector's salivary gland extract, with or without adjuvant formulation for enhanced efficacy. Promising animal data supports multiple vaccine candidates (Lmcen-/-, LmexCen-/-), with some already in or heading for clinical trials. The crucial challenge in Leishmania vaccine development is to translate the research knowledge into affordable and accessible control tools that refines the outcome for those who are susceptible to infection. This review focuses on recent findings in Leishmania vaccines and highlights difficulties facing vaccine development and implementation.
Asunto(s)
Leishmania , Vacunas contra la Leishmaniasis , Leishmaniasis , Desarrollo de Vacunas , Humanos , Vacunas contra la Leishmaniasis/inmunología , Animales , Leishmania/inmunología , Leishmaniasis/inmunología , Leishmaniasis/prevención & control , Desarrollo de Vacunas/métodos , Antígenos de Protozoos/inmunología , Ensayos Clínicos como AsuntoRESUMEN
Leishmania donovani causes the potentially fatal disease visceral leishmaniasis for which neither a vaccine nor an adjuvant for human use exists. Although interleukin-7 (IL-7) is implicated in CD4+ T-cell response stabilization, its anti-leishmanial function is uncertain. Therefore, we examined whether IL-7 would potentiate the efficacy of Leishmania major-expressed MAPK10 (LmjMAPK10; M10)-elicited anti-leishmanial host-protective response. We observed that aligning with IL-7R expression, IL-7 increased IFN-γ-secreting TH1 cell but reduced IL-4-producing TH2 cells and production of IL-10 and TGF-ß effectuating anti-leishmanial functions in susceptible BALB/c mouse-derived macrophages. Co-culturing IL-7-pre-treated L. donovani-infected macrophages with L. donovani-infected BALB/c-derived T cells induced IFN-γ-dominated TH1 type anti-leishmanial function. IL-7 treatment of L. donovani-infected BALB/c mice significantly reduced splenic and hepatic parasite loads. Co-culturing CD4+ T cells from IL to 7-treated mice with L. donovani-infected macrophages reduced amastigote numbers suggesting IL-7-elicited host-protective effector T cells. Priming BALB/c with M10 + IL-7 reduced the splenic parasite burden more effectively than that was observed in M10-primed mice. An enhanced protection against L. donovani infection was accompanied by enhanced IL-12 and IFN-γ, but suppressed IL-10 and IL-4, response and host-protective TH1 and memory T cells. These results indicate IL-7-induced leishmanial antigen-specific memory T cell response that protects a susceptible host against L. donovani infection.
Asunto(s)
Adyuvantes de Vacunas , Interleucina-7 , Leishmania donovani , Vacunas contra la Leishmaniasis , Leishmaniasis Visceral , Proteína Quinasa 10 Activada por Mitógenos , Vacunas contra la Leishmaniasis/inmunología , Animales , Ratones , Ratones Endogámicos BALB C , Leishmania donovani/inmunología , Leishmaniasis Visceral/prevención & control , Proteína Quinasa 10 Activada por Mitógenos/inmunología , Receptores de Interleucina-7/metabolismo , Interleucina-7/administración & dosificación , Interferón gamma/metabolismo , Células TH1/inmunología , Macrófagos/inmunología , Macrófagos/parasitología , Leishmania major/inmunología , Técnicas de Cocultivo , Células T de Memoria/inmunología , Bazo/parasitología , Hígado/parasitología , Presentación de AntígenoRESUMEN
Visceral leishmaniasis (VL) is a chronic tropical disease responsible for devastating epidemics worldwide. Though current treatment relies on drugs, the emergence of resistance, toxic side-effects, and strenuous administration has led to an ineffective remedy. Hence, vaccination remains an alternative and desirable approach for VL control. Though extensive research on anti-leishmanial vaccine candidates has been carried out in past decades, presence of an effective molecule is still missing. In the present study, we have evaluated the immunogenicity and prophylactic potential of a recombinant T-complex protein-1 gamma subunit of L. donovani (rLdTCP1γ), against VL in hamster model. The antigen exhibited in vitro stimulation of lymphoproliferative and NO response in miltefosine and amphotericin B treated hamsters depicting its immunotherapeutic/immunogenic nature. Immunization with rLdTCP1γ revealed a strong protective response against experimental VL as indicated by reduced parasite load in the spleen of immunized group compared to infected control. The immunized animals gained body weight and exhibited significant reduction in the spleen and liver weight as compared to infected controls on days 60, 90, 120 post-challenge. A substantial augmentation of cell-mediated immune response as depicted by an increased lymphocyte proliferation, nitric oxide production, DTH responses and increased levels of IgG2 was observed in rLdTCP1γ immunized hamsters. The Th1 stimulatory potential, imparted by the antigen, was found to be intensified in the presence of adjuvant Bacillus Calmette-Guérin (BCG). The efficacy was further assisted by an upregulated mRNA transcript of Th1 induced cytokines (IL-12, IFN-γ and TNFα) and downregulation of IL-4 and IL-10. The results are thus suggestive of rLdTCP1γ having the potential of a strong vaccine candidate against VL.
Asunto(s)
Antígenos de Protozoos/inmunología , Leishmania donovani/inmunología , Vacunas contra la Leishmaniasis/inmunología , Leishmaniasis Visceral/inmunología , Proteínas Protozoarias/inmunología , Adyuvantes Inmunológicos/farmacología , Animales , Línea Celular , Cricetinae , Citocinas/inmunología , Inmunización/métodos , Leucocitos Mononucleares/inmunología , Activación de Linfocitos/inmunología , Masculino , Ratones , Células TH1/inmunología , Vacunación/métodosRESUMEN
Leishmaniasis is endemic to the tropical and subtropical regions of the world and is transmitted by the bite of an infected sand fly. The multifaceted interactions between Leishmania, the host innate immune cells, and the adaptive immunity determine the severity of pathogenesis and disease development. Leishmania parasites establish a chronic infection by subversion and attenuation of the microbicidal functions of phagocytic innate immune cells such as neutrophils, macrophages and dendritic cells (DCs). Other innate cells such as inflammatory monocytes, mast cells and NK cells, also contribute to resistance and/or susceptibility to Leishmania infection. In addition to the cytokine/chemokine signals from the innate immune cells, recent studies identified the subtle shifts in the metabolic pathways of the innate cells that activate distinct immune signal cascades. The nexus between metabolic pathways, epigenetic reprogramming and the immune signaling cascades that drive the divergent innate immune responses, remains to be fully understood in Leishmania pathogenesis. Further, development of safe and efficacious vaccines against Leishmaniasis requires a broader understanding of the early interactions between the parasites and innate immune cells. In this review we focus on the current understanding of the specific role of innate immune cells, the metabolomic and epigenetic reprogramming and immune regulation that occurs during visceral leishmaniasis, and the strategies used by the parasite to evade and modulate host immunity. We highlight how such pathways could be exploited in the development of safe and efficacious Leishmania vaccines.
Asunto(s)
Inmunidad Innata , Leishmania donovani/inmunología , Vacunas contra la Leishmaniasis/inmunología , Leishmaniasis Visceral/inmunología , Desarrollo de Vacunas , Animales , Citotoxicidad Inmunológica , Células Dendríticas/inmunología , Humanos , Evasión Inmune , Inmunogenicidad Vacunal , Células Asesinas Naturales/inmunología , Macrófagos/inmunología , Macrófagos/parasitología , Mastocitos/inmunología , Metabolómica , Ratones , Ratones Endogámicos C57BL , Monocitos/inmunología , Células T Asesinas Naturales/inmunología , Neutrófilos/inmunologíaRESUMEN
Dogs are the main reservoir of Leishmania infantum in endemic regions. Canine leishmaniasis, caused by L. infantum, can progress to a chronic disease resulting in death. Vaccines have been developed with a certain degree of success. The pathogenesis of this disease is not completely understood, especially in previously vaccinated dogs. We herein described clinical data, parasite load, serum levels of cytokines, and the reservoir potential in vdogs vaccinated with the fucose-mannose ligand (FML)/QuilA saponin vaccine (Leishmune™) naturally infected (Vi) and compared to vaccinated not infected dogs (Vn). Thirty-four dogs from private owners were divided into two groups: vaccinated/infected and vaccinated/uninfected. Clinical evaluation, hematological and biochemical parameters, and serum levels of cytokines were measured by conventional methods. The parasite burden in the bone marrow was measured by quantitative real-time PCR, and the transmissibility of parasites to sand flies was assessed by xenodiagnosis. Clinical, biochemical, and hematological parameters of vaccinated infected dogs were mostly normal. Vi dogs developed mild disease with low clinical scores. Serum levels of IL-10 were higher in Vi dogs, and a strong correlation was observed in IL-4 levels and the A/G ratio in Vi dogs. These results suggest a role of TH2 response in Vi dogs, although more data is needed to better understand the disease in vaccinated dogs.
Asunto(s)
Citocinas/sangre , Lectinas/inmunología , Leishmania infantum/inmunología , Vacunas contra la Leishmaniasis/inmunología , Leishmaniasis Visceral/parasitología , Leishmaniasis Visceral/veterinaria , Vacunación , Animales , Perros , Femenino , Interleucina-4/sangre , Leishmaniasis Visceral/sangre , Leishmaniasis Visceral/patología , MasculinoRESUMEN
The development of vaccines against one or all forms of human leishmaniasis remains hampered by a paucity of investment, at least in part resulting from the lack of well-evidenced and agreed estimates of vaccine demand. Starting from the definition of 4 main use cases (prevention of visceral leishmaniasis, prevention of cutaneous leishmaniasis, prevention of post-kala-azar dermal leishmaniasis and treatment of post-kala-azar dermal leishmaniasis), we have estimated the size of each target population, focusing on those endemic countries where incidence levels are sufficiently high to justify decisions to adopt a vaccine. We assumed a dual vaccine delivery strategy, including a wide age-range catch-up campaign before the start of routine immunisation. Vaccine characteristics and delivery parameters reflective of a target product profile and the likely duration of the clinical development effort were considered in forecasting the demand for each of the four indications. Over a period of 10 years, this demand is forecasted to range from 300-830 million doses for a vaccine preventing visceral leishmaniasis and 557-1400 million doses for a vaccine preventing cutaneous leishmaniasis under the different scenarios we simulated. In a scenario with an effective prophylactic visceral leishmaniasis vaccine, demand for use to prevent or treat post-kala-azar dermal leishmaniasis would be more limited (over the 10 years ~160,000 doses for prevention and ~7,000 doses for treatment). Demand would rise to exceed 330,000 doses, however, in the absence of an effective vaccine for visceral leishmaniasis. Because of the sizeable demand and potential for public health impact, a single-indication prophylactic vaccine for visceral or cutaneous leishmaniasis, and even more so a cross-protective prophylactic vaccine could attract the interest of commercial developers. Continuous refinement of these first-of-their kind estimates and confirmation of country willingness and ability to pay will be paramount to inform the decisions of policy makers and developers in relation to a leishmaniasis vaccine. Positive decisions can provide a much-needed contribution towards the achievement of global leishmaniasis control.
Asunto(s)
Vacunas contra la Leishmaniasis/inmunología , Vacunas contra la Leishmaniasis/provisión & distribución , Leishmaniasis/epidemiología , Leishmaniasis/prevención & control , Salud Global , Humanos , Vacunas contra la Leishmaniasis/economía , Salud PúblicaRESUMEN
Visceral leishmaniasis is a protozoan disease associated with high fatality rate in developing countries. Although the drug pipeline is constantly improving, available treatments are costly and live-threatening side effects are not uncommon. Moreover, an approved vaccine against human leishmaniasis does not exist yet. Using whole antigens from Leishmania donovani promastigotes (LdAg), we investigated the protective potential of a novel adjuvant-free vaccine strategy. Immunization of mice with LdAg via the intradermal or the intranasal route prior to infection decreases the parasitic burden in primary affected internal organs, including the liver, spleen, and bone marrow. Interestingly, the intranasal route is more efficient than the intradermal route, leading to better parasite clearance and remarkable induction of adaptive immune cells, notably the helper and cytotoxic T cells. In vitro restimulation experiments with Leishmania antigens led to significant IFN-γ secretion by splenocytes; therefore, exemplifying specificity of the adaptive immune response. To improve mucosal delivery and the immunogenic aspects of our vaccine strategy, we used polysaccharide-based nanoparticles (NP) that carry the antigens. The NP-LdAg formulation is remarkably taken up by dendritic cells and induces their maturation in vitro, as revealed by the increased expression of CD80, CD86 and MHC II. Intranasal immunization with NP-LdAg does not improve the parasite clearance in our experimental timeline; however, it does increase the percentage of effector and memory T helper cells in the spleen, suggesting a potential induction of long-term memory. Altogether, this study provides a simple and cost-effective vaccine strategy against visceral leishmaniasis based on LdAg administration via the intranasal route, which could be applicable to other parasitic diseases.
Asunto(s)
Antígenos de Protozoos/inmunología , Médula Ósea/parasitología , Vacunas contra la Leishmaniasis/inmunología , Leishmaniasis Visceral/inmunología , Hígado/parasitología , Bazo/parasitología , Inmunidad Adaptativa , Adyuvantes Inmunológicos/administración & dosificación , Administración Intranasal , Animales , Anticuerpos Antiprotozoarios/sangre , Antígenos de Protozoos/administración & dosificación , Antígenos de Protozoos/sangre , Médula Ósea/metabolismo , Femenino , Inmunización , Interferón gamma/metabolismo , Leishmania donovani/inmunología , Vacunas contra la Leishmaniasis/administración & dosificación , Leishmaniasis Visceral/parasitología , Leishmaniasis Visceral/prevención & control , Hígado/metabolismo , Ratones , Ratones Endogámicos BALB C , Bazo/metabolismoRESUMEN
Visceral Leishmaniasis (VL), a potentially fatal disease is caused by Leishmania donovani parasites with no vaccine available. Here we produced a dermotropic live attenuated centrin gene deleted Leishmania major (LmCen-/-) vaccine under Good Laboratory Practices and demonstrated that a single intradermal injection confers robust and durable protection against lethal VL transmitted naturally via bites of L. donovani-infected sand flies and prevents mortality. Surprisingly, immunogenicity characteristics of LmCen-/- parasites revealed activation of common immune pathways like L. major wild type parasites. Spleen cells from LmCen-/- immunized and L. donovani challenged hamsters produced significantly higher Th1-associated cytokines including IFN-γ, TNF-α, and reduced expression of the anti-inflammatory cytokines like IL-10, IL-21, compared to non-immunized challenged animals. PBMCs, isolated from healthy people from non-endemic region, upon LmCen-/- infection also induced more IFN-γ compared to IL-10, consistent with our immunogenicity data in LmCen-/- immunized hamsters. This study demonstrates that the LmCen-/- parasites are safe and efficacious against VL and is a strong candidate vaccine to be tested in a human clinical trial.
Asunto(s)
Eliminación de Gen , Genes Protozoarios , Leishmania donovani/inmunología , Vacunas contra la Leishmaniasis/inmunología , Leishmaniasis Visceral/prevención & control , Leishmania donovani/genética , Leishmaniasis Visceral/inmunología , Proteínas Protozoarias , Vacunas Atenuadas/inmunologíaRESUMEN
Tropical and subtropical areas of the world are affected by leishmaniasis, which is caused by Leishmania spp. It has been categorized as an NTD (neglected tropical disease) because of its negligence. The sand fly of genus Phlebotomus acts as the vector for the transmission of the promastigote form of this protozoan parasite to the mammalian host where it converts to amastigote form in the macrophages. Visceral form of leishmaniasis (VL) is a deadly infection in the endothelial system of the human and other mammals. Only a few chemotherapeutic agents are available for the treatment of this infectious disease whereas no vaccine is available for the control of leishmanial infection. Therefore in the current study, we have tested the effects of gardiquimod (a TLR agonist) as an adjuvant in combination with the formalin-killed antigen of L. donovani as a vaccine. The mice were vaccinated thrice at an interval of 2 weeks and challenged with L. donovani promastigotes after 2 weeks of the last vaccination. We assessed the parasite load, delayed-type hypersensitivity (DTH) responses, humoral and cell-mediated immune response in BALB/c mice before and after challenge infection with L. donovani. Immunized mice were found to have the least parasite load, high DTH response, elevated levels of Th1 cytokines, IgG2a, and nitric oxide than non-immunized and infected control mice. The efficacy of the vaccine was boosted with the use of adjuvant gardiquimod that depicts its potential as an adjuvant in this study. Our study is reporting the adjuvant effects of gardiquimod for the first time. Further studies using other Leishmania species can be performed to signify its role.
Asunto(s)
Adyuvantes Inmunológicos/farmacología , Aminoquinolinas/farmacología , Imidazoles/farmacología , Leishmania donovani/inmunología , Vacunas contra la Leishmaniasis/inmunología , Leishmaniasis Visceral/prevención & control , Receptores Toll-Like/antagonistas & inhibidores , Animales , Femenino , Leishmaniasis Visceral/parasitología , Masculino , Ratones , Ratones Endogámicos BALB CRESUMEN
Extracellular vesicles (EVs) are small, membrane-bound "delivery trucks" that are present in the extracellular environment, including biological fluids. EVs are capable of inducing changes in the physiological status of neighboring cells through the transfer of key macromolecules, and are thought to play a role in a number of pathological processes. Leishmaniasis, caused by the protozoan parasite Leishmania, is an important example. The biology of Leishmania EVs has been studied in detail, and findings point to their role in exacerbation of disease and potential involvement in the perpetuation of drug resistance. Furthermore, the use of EVs for development of vaccines has been explored, as well as their potential use in a number of fields as biomarkers of disease and drug resistance. Here we discuss the latest findings on EVs, with a particular focus on Leishmania, as well as potential avenues for their future development and clinical applications.
Asunto(s)
Transporte Biológico/fisiología , Vesículas Extracelulares/metabolismo , Inmunidad Innata/inmunología , Leishmania/inmunología , Leishmaniasis/inmunología , Animales , Biomarcadores , Humanos , Leishmaniasis/prevención & control , Vacunas contra la Leishmaniasis/inmunología , Metaloendopeptidasas/metabolismo , Proteínas Protozoarias/metabolismoRESUMEN
Leishmaniasis is a disease caused by an intracellular protozoan parasite of the genus Leishmania. Current treatments for leishmaniasis are long, toxic, and expensive and are not available in some endemic regions. Attempts to develop an effective vaccine are feasible, but no vaccine is in active clinical use. In this study, the LmxMBA gene of Leishmania mexicana was selected as a possible vaccine candidate using the reverse vaccinology approach, and the prophylactic effect generated by DNA vaccination with this gene in a murine model of cutaneous leishmaniasis was evaluated. The results showed that prophylactic vaccination with pVAX1::LmxMBA significantly reduced the size of the lesion and the parasitic load on the footpad, compared to the control groups. At a histological level, a smaller number of parasites were evident in the dermis, as well as the absence of connective tissue damage. Mice immunized with plasmid pVAX1::LmxMBA induced immunity characterized by an increase in the IgG2a/IgG1 > 1 ratio and a higher rate of lymphocyte proliferation. In this study, immunization with the plasmid promoted an improvement in the macroscopic and microscopic clinical manifestations of the experimental infection by L. mexicana, with a T helper 1 response characterized by an IgG2a/IgG1 > 1 ratio and high lymphoproliferative response. These findings support immunization with the plasmid pVAX1::LmxMBA as a preventive strategy against cutaneous infection of L. mexicana.
Asunto(s)
Fosfatasa Ácida/genética , Leishmania mexicana/fisiología , Vacunas contra la Leishmaniasis/inmunología , Leishmaniasis Cutánea/inmunología , Proteínas Protozoarias/genética , Piel/patología , Células TH1/inmunología , Animales , Anticuerpos Antiprotozoarios/sangre , Modelos Animales de Enfermedad , Femenino , Humanos , Inmunoglobulina G/sangre , Leishmaniasis Cutánea/prevención & control , Ratones , Ratones Endogámicos BALB C , Carga de Parásitos , Vacunación , Vacunas de ADNRESUMEN
The most dangerous form of leishmaniasis is Visceral leishmaniasis (VL). The elimination of VL depends not only on agent treatments but also on effective vaccines against Leishmania parasites. Epitope-based vaccines composed of alternative short antigenic epitopes have the advantages of MHC epitope easy designing, which has broad application prospects. In a previous study, we analyzed Leishmania Gp63, Kmp-11 and Amastin protein sequence in silico, and found that the amino acid fragments of Gp63 (138-360aa), Kmp-11 (1-91aa) and Amastin (1-72aa) were rich in dominant epitopes. In this study, we used the three amino acid fragments as multi-epitope vaccine candidates to construct DNA and protein vaccines. BALB/c mice were vaccinated with the DNA and protein vaccines by DNA prime-protein boost strategy and challenged with Leishmania promastigotes. To evaluate vaccine immunogenicity and immunoprotection, serum specific antibody titers and cytokines were detected using ELISA, splenic CD3+, CD4+ and CD8+ cells were analyzed by flow cytometry, livers were made into pathological sections to observe pathological changes, and splenic parasitic loads were quantified using qPCR. The results showed that the increased specific IgG titers from vaccinated mice supported the vaccine immunogenicity. The increased cytokines (IFN-γ, IL-12 and TNF-α), splenic CD3+, CD4+ and CD8+ T cells and hepatic granulomas, and the decreased splenic parasitic loads (parasite reduction rates of Gp63, Kmp-11 and Amatin groups were 89%, 86% and 79%, respectively) from immunized mice post-infection were suggested the good immunoprotection of the vaccines. Our study demonstrated that vaccines based on the dominant epitopes of Gp63, Kmp-11 and Amastin with DNA prime-protein boost vaccination strategy showed significant immune effects against Leishmania, especially the Gp63 group showed a nearly 90% parasites reduction rate. This study will provide references for visceral leishmaniasis epitope vaccine design and immune strategy selection.
Asunto(s)
Epítopos/inmunología , Vacunas contra la Leishmaniasis/inmunología , Leishmaniasis Visceral/prevención & control , Metaloendopeptidasas/inmunología , Proteínas Protozoarias/inmunología , Animales , Anticuerpos Antiprotozoarios/sangre , Anticuerpos Antiprotozoarios/inmunología , Cricetinae , Citocinas/sangre , Modelos Animales de Enfermedad , Ensayo de Inmunoadsorción Enzimática , Femenino , Inmunización , Inmunoglobulina G/sangre , Inmunoglobulina G/inmunología , Leishmania infantum/inmunología , Leishmaniasis Visceral/parasitología , Ratones , Proteínas Recombinantes/inmunología , Bazo/inmunología , Subgrupos de Linfocitos T/inmunología , Subgrupos de Linfocitos T/metabolismoRESUMEN
Post-kala-azar dermal leishmaniasis (PKDL) is a chronic, stigmatizing skin condition occurring frequently after apparent clinical cure from visceral leishmaniasis. Given an urgent need for new treatments, we conducted a phase IIa safety and immunogenicity trial of ChAd63-KH vaccine in Sudanese patients with persistent PKDL. LEISH2a (ClinicalTrials.gov: NCT02894008) was an open-label three-phase clinical trial involving sixteen adult and eight adolescent patients with persistent PKDL (median duration, 30 months; range, 6-180 months). Patients received a single intramuscular vaccination of 1 × 1010 viral particles (v.p.; adults only) or 7.5 × 1010 v.p. (adults and adolescents), with primary (safety) and secondary (clinical response and immunogenicity) endpoints evaluated over 42-120 days follow-up. AmBisome was provided to patients with significant remaining disease at their last visit. ChAd63-KH vaccine showed minimal adverse reactions in PKDL patients and induced potent innate and cell-mediated immune responses measured by whole-blood transcriptomics and ELISpot. 7/23 patients (30.4%) monitored to study completion showed >90% clinical improvement, and 5/23 (21.7%) showed partial improvement. A logistic regression model applied to blood transcriptomic data identified immune modules predictive of patients with >90% clinical improvement. A randomized controlled trial to determine whether these clinical responses were vaccine-related and whether ChAd63-KH vaccine has clinical utility is underway.