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1.
Mol Ther Nucleic Acids ; 19: 1098-1109, 2020 Jan 16.
Artigo em Inglês | MEDLINE | ID: mdl-32059336

RESUMO

In vitro transcribed mRNAs hold the promises of many medical applications in disease prevention and treatment, such as replacement or supplement of missing or inadequately expressed endogenous proteins and as preventive vaccines against infectious diseases, therapeutic vaccines, or other protein-based biopharmaceutics for cancer therapy. A safe and efficient delivery system for mRNA is crucial to the success of mRNA therapeutic applications. In this study, we report that InstantFECT, a liposome-based transfection reagent, can pack pseudouridine-incorporated mRNA into nanocomplexes that are highly efficient in mediating in vivo transfection in multiple organs after local delivery. High levels of expression of EGFP and luciferase reporters after intratumoral and intramuscular injections were observed, which lasted for up to 96 hrs. Immunogenicity of antigens encoded by mRNA delivered with nanocomplex was investigated by subcutaneous delivery of modified mRNAs encoding Staphylococcus aureus adenosine synthase A (AdsA) and a model tumor-associated antigen ovalbumin (OVA). Strong T cell responses were provoked by both mRNAs delivered. Therapeutic and protective treatment with the OVA mRNA-liposome nanocomplex significantly inhibited B16-OVA tumor progression and increased mouse survival. There was no sign of obvious toxicity related to the treatment both in tissue culture and in mice. An intravenous injection of the same dosage of the modified mRNA-lipid nanocomplex showed minimal transfection in major organs, indicating an excellent safety feature as the gene transfer occurred only at the injection sites, whereas intravenous (i.v.) injection with the same amount of mRNA complexed with a commercial transfection reagent Trans-IT showed luciferase expression in the spleen. In summary, InstantFECT cationic liposomes provide a safe and efficient in vivo locoregional delivery of mRNA and could be a useful tool for basic research and for the development of mRNA-based therapies.

2.
Nanoscale ; 11(45): 21782-21789, 2019 Nov 21.
Artigo em Inglês | MEDLINE | ID: mdl-31709434

RESUMO

For efficient cancer vaccines, the antitumor function largely relies on cytotoxic T cells, whose activation can be effectively induced via antigen-encoding mRNA, making mRNA-based cancer vaccines an attractive approach for personalized cancer therapy. While the liposome-based delivery system enables the systemic delivery and transfection of mRNA, incorporating an adjuvant that is non-lipid like remains challenging, although the co-delivery of mRNA (antigen) and effective adjuvant is key to the activation of the cytotoxic T cells. This is because the presence of an adjuvant is important for dendritic cell maturation-another necessity for cytotoxic T cell activation. In the present work, we designed a poly (lactic-co-glycolic acid) (PLGA)-core/lipid-shell hybrid nanoparticle carrier for the co-delivery of mRNA and gardiquimod (adjuvant that cannot be incorporated into the lipid shell). We demonstrated in the present work that the co-delivery of mRNA and gardiquimod led to the effective antigen expression and DC maturation in vitro. The intravenous administration of the hybrid nanovaccine resulted in the enrichment of mRNA expression in the spleen and a strong immune response in vivo. The simultaneous delivery of the antigen and adjuvant both spatially and temporally via the core/shell nanoparticle carrier is found to be beneficial for tumor growth inhibition.

3.
mSphere ; 4(5)2019 09 04.
Artigo em Inglês | MEDLINE | ID: mdl-31484738

RESUMO

The demand for a prophylactic vaccine against methicillin-resistant Staphylococcus aureus (MRSA) has motivated numerous dedicated research groups to design and develop such a vaccine. In this study, we have developed a multivalent vaccine, Sta-V5, composed of five conserved antigens involved in three important virulence mechanisms. This prototype vaccine conferred up to 100% protection against multiple epidemiologically relevant S. aureus isolates in five different murine disease models. The vaccine not only elicits functional antibodies that mediate opsonophagocytic killing of S. aureus but also mounts robust antigen-specific T-cell responses. In addition, our data implied that γδ T cells contribute to the protection induced by Sta-V5 in a murine skin infection model.IMPORTANCE Staphylococcus aureus infections, especially MRSA infections, are becoming a major global health issue and are resulting in mortality rates that are increasing every year. However, an effective vaccine is lacking due to the complexity of the infection process of S. aureus In this study, we found that the addition of two novel protein components to three well-studied vaccine candidates significantly improved the efficacy of the combined vaccine. Furthermore, the five-component vaccine not only elicits a robust antibody response but also induces cytokine secretion by T cells, making it a promising vaccine candidate to fill the void.

4.
BMC Infect Dis ; 18(1): 195, 2018 04 25.
Artigo em Inglês | MEDLINE | ID: mdl-29699491

RESUMO

BACKGROUND: Staphylococcus aureus (S. aureus) causes a wide range of infectious diseases in human and animals. The emergence of antibiotic-resistant strains demands novel strategies for prophylactic vaccine development. In this study, live attenuated S. enterica subsp. enterica serotype Typhimurium (S. Typhimurium) vaccine against S. aureus infection was developed, in which Salmonella Pathogenesis Island-1 Type 3 Secretion System (SPI-1 T3SS) was employed to deliver SaEsxA and SaEsxB, two of ESAT-6-like (Early Secreted Antigenic Target-6) virulence factors of S. aureus. METHODS: Antigens SaEsxA and SaEsxB were fused with the N-terminal secretion and translocation domain of SPI-1 effector SipA. And cytosolic delivery of Staphylococcal antigens into macrophages was examined by western blot. BALB/c mice were orally immunized with S. Typhimurium-SaEsxA and S. Typhimurium-SaEsxB vaccines. Antigen-specific humoral and Th1/Th17 immune responses were examined by ELISA and ELISPOT assays 7-9 days after the 2nd booster. For ELISPOT assays, the statistical significance was determined by Student's t test. The vaccine efficacy was evaluated by lethal challenge with two S. aureus clinical isolates Newman strain and USA 300 strain. Statistical significance was determined by Log rank (Mantel-Cox) analysis. And a P value of < 0.05 was considered statistically significant. RESULTS: Oral administration of S. Typhimurium-SaEsxA and S. Typhimurium-SaEsxB vaccines induced antigen-specific humoral and Th1/Th17 immune responses, which increased the survival rate for vaccinated mice when challenged with S. aureus strains. CONCLUSIONS: The newly developed S. Typhimurium-based vaccines delivering SaEsxA and SaEsxB by SPI-1 T3SS could confer protection against S. aureus infection. This study provides evidence that translocation of foreign antigens via Salmonella SPI-1 T3SS into the cytosol of antigen presenting cells (APCs) could induce potent immune responses against pathogens.


Assuntos
Células RAW 264.7/efeitos dos fármacos , Salmonella typhimurium/imunologia , Infecções Estafilocócicas/prevenção & controle , Sistemas de Secreção Tipo III/imunologia , Vacinas Atenuadas/administração & dosagem , Animais , Imunização , Camundongos , Camundongos Endogâmicos BALB C , Fatores de Virulência/imunologia
5.
J Infect Dis ; 216(2): 245-253, 2017 07 15.
Artigo em Inglês | MEDLINE | ID: mdl-28633319

RESUMO

Staphylococcusaureus is a severe pathogen found in the community and in hospitals. Most notably, methicillin-resistant S. aureus (MRSA) is resistant to almost all antibiotics, which is a growing public health concern. The emergence of drug-resistant strains has prompted the search for alternative treatments such as immunotherapeutic approaches. Previous research showed that S. aureus exploit the immunomodulatory attributes of adenosine to escape host immunity. In this study, we investigated adenosine synthase A (AdsA), an S. aureus cell wall-anchored enzyme as possible targets for immunotherapy. Mice vaccinated with aluminum hydroxide-formulated recombinant AdsA (rAdsA) induced high-titer anti-AdsA antibodies, thereby providing consistent protection in 3 mouse infection models when challenged with 2 S. aureus strains. The importance of anti-AdsA antibody in protection was demonstrated by passive transfer experiments. Moreover, AdsA-specific antisera promote killing S. aureus by immune cells. Altogether, our data demonstrate that the AdsA is a promising target for vaccines and therapeutics development to alleviate severe S. aureus diseases.


Assuntos
Anticorpos Antibacterianos/farmacologia , Proteínas de Bactérias/imunologia , Imunização Passiva , Ligases/imunologia , Infecções Cutâneas Estafilocócicas/terapia , Adenosina/biossíntese , Animais , Antibacterianos/uso terapêutico , Modelos Animais de Doenças , Feminino , Imunoterapia , Camundongos , Camundongos Endogâmicos BALB C , Coelhos , Staphylococcus aureus/enzimologia
6.
Oncol Lett ; 13(1): 177-183, 2017 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-28123538

RESUMO

The antitumor properties of bacteria have been demonstrated over the past decades. However, the efficacy is limited and unclear. Furthermore, systemic infection remains a serious concern in bacteria treatment. In this study, the effect of YB1, a rationally designed 'obligate' anaerobic Salmonella typhimurium strain, on liver tumor growth and metastasis in a nude mouse orthotopic liver tumor model was investigated. The orthotopic liver tumor model was established in nude mice using the hepatocellular carcinoma cell line MHCC-97L. Two weeks after orthotopic liver tumor implantation, YB1, SL7207 and saline were respectively administered through the tail vein of the mice. Longitudinal monitoring of tumor growth and metastasis was performed using Xenogen IVIS, and direct measurements of tumor volume were taken 3 weeks after treatment. In vitro, MHCC-97L and PLC cells were incubated with YB1 or SL7207 under anaerobic conditions. YB1 was observed to invade tumor cells and induce tumor cell apoptosis and death. The results revealed that all mice in the YB1 group were alive 3 weeks after YB1 injection while all mice in the SL7207 group died within 11 days of the SL7207 injection. The body weight decreased by ~9% on day 1 after YB1 injection and but subsequently recovered. Liver tumor growth and metastases were significantly inhibited following YB1 treatment. By contrast to the control group, a large number of Gr1-positive cells were detected on days 1 to 21 following YB1 treatment. Furthermore, YB1 also effectively invaded tumor cells and induced tumor cell apoptosis and death. In conclusion, YB1 suppressed liver tumor growth and metastasis in a nude mice liver tumor model. The potential mechanism may be through enhancing innate immune response and inducing tumor cell apoptosis and cell death.

7.
J Hematol Oncol ; 8: 99, 2015 Aug 19.
Artigo em Inglês | MEDLINE | ID: mdl-26286454

RESUMO

BACKGROUND: Neuroblastoma currently has poor prognosis, therefore we proposed a new strategy by targeting neuroblastoma with genetically engineered anaerobic Salmonella (Sal-YB1). METHODS: Nude and nonobese diabetic-severe combined immunodeficiency (NOD-SCID) orthotopic mouse models were used, and Sal-YB1 was administered via tail vein. The therapeutic effectiveness, bio-safety, and mechanisms were studied. RESULTS: No mice died of therapy-related complications. Tumor size reduction was 70 and 30% in nude and NOD-SCID mice, respectively. No Salmonella was detected in the urine; 75% mice had positive stool culture if diaminopimelic acid was added, but all turned negative subsequently. Tumor tissues had more Sal-YB1 infiltration, necrosis, and shrinkage in Sal-YB1-treated mice. Significantly higher expression of TLR4, TNF-stimulated gene 6 protein (TSG6), and cleaved caspase 1, 3, 8, and 9 was found in the tumor masses of the Sal-YB1-treated group with a decrease of interleukin 1 receptor-associated kinase (IRAK) and nuclear factor of kappa light polypeptide gene enhancer in B-cells inhibitor alpha (IκBα). There was a high release of TNFα both in human macrophages and mouse tumor tissues with Sal-YB1 treatment. The antitumor effect of the supernatant derived from macrophages treated with Sal-YB1 could be reversed with TNFα and pan-caspase inhibitors. CONCLUSIONS: This new approach in targeting neuroblastoma by bio-engineered Salmonella with the assistance of macrophages indirectly may have a clinical therapeutic impact in the future.


Assuntos
Neoplasias Encefálicas/terapia , Neuroblastoma/terapia , Salmonella typhimurium/metabolismo , Animais , Modelos Animais de Doenças , Humanos , Camundongos , Camundongos Endogâmicos NOD , Camundongos Nus , Camundongos SCID
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