Shaoqiong Yi

Vaccination with a DNA vaccine based on human PSCA and HSP70 adjuvant enhances the antigen-specific CD8+ T-cell response and inhibits the PSCA+ tumors growth in mice

DNA vaccines have been shown to be an effective approach to induce antigen‐specific cellular and humoral immunity. However, the lower immune intensity in clinical trials limits the application of DNA vaccine. Here we intend to develop a new DNA vaccine based on prostate stem‐cell antigen (PSCA), which has been suggested as a potential target for prostate cancer therapy, and enhance the DNA vaccine potency with heat shock proteins (HSPs) as adjuvant.

Prediction and identification of mouse cytotoxic T lymphocyte epitopes in Ebola virus glycoproteins

BackgroundEbola viruses (EBOVs) cause severe hemorrhagic fever with a high mortality rate. At present, there are no licensed vaccines or efficient therapies to combat EBOV infection. Previous studies have shown that both humoral and cellular immune responses are crucial for controlling Ebola infection. CD8+ T cells play an important role in mediating vaccine-induced protective immunity. The objective of this study was to identify H-2d-specific T cell epitopes in EBOV glycoproteins (GPs).ResultsComputer-assisted algorithms were used to predict H-2d-specific T cell epitopes in two species of EBOV (Sudan and Zaire) GP. The predicted peptides were synthesized and identified in BALB/c mice immunized with replication-deficient adenovirus vectors expressing the EBOV GP. Enzyme-linked immunospot assays and intracellular cytokine staining showed that the peptides RPHTPQFLF (Sudan EBOV), GPCAGDFAF and LYDRLASTV (Zaire EBOV) could stimulate splenoctyes in immunized mice to produce large amounts of interferon-gamma.ConclusionThree peptides within the GPs of two EBOV strains were identified as T cell epitopes. The identification of these epitopes should facilitate the evaluation of vaccines based on the Ebola virus glycoprotein in a BALB/c mouse model.

Monoclonal Antibody Targeting Staphylococcus aureus Surface Protein A (SasA) Protect Against Staphylococcus aureus Sepsis and Peritonitis in Mice

Epidemic methicillin-resistant Staphylococcus aureus (MRSA) imposes an increasing impact on public health. Due to multi-antibiotics resistance in MRSA strains, there is an urgent need to develop novel therapeutics such as effective monoclonal antibodies (mAbs) against MRSA infections. Staphylococcus aureus surface protein A (SasA), a large surface-located protein (~240 kDa), is one of MSCRAMMs (microbial surface components recognizing adhesive matrix molecules) and a potential target for immunotherapeutic approaches against S. aureus infections. In the present study, we analyzed the sequence of SasA with bioinformatics tools and generated a protective monoclonal antibody (2H7) targeting the conserved domain of SasA. 2H7 was shown to recognize wild-type S. aureus and promote opsonophagocytic killing of S. aureus. In both sepsis and peritoneal infection models, prophylactic administration of 2H7 improved the survival of BALB/c mice challenged by S. aureus strain USA300 and ST239 (prevalent MRSA clones in North America and Asian countries, respectively) and enhanced bacterial clearance in kidneys. Additionally, 2H7 prophylaxis prevented the formation of intraperitoneal abscess in a murine model of peritoneal infection and therapeutic administration of 2H7 showed protective efficacy in a murine sepsis model. Our results presented here provide supporting evidences that an anti-SasA mAb might be a potential component in an antibody-based immunotherapeutic treatment of MRSA infections.

Creation of a Six-fingered Artificial Transcription Factor That Represses the Hepatitis B Virus HBx Gene Integrated into a Human Hepatocellular Carcinoma Cell Line

Chronic hepatitis B virus (HBV) infection is an independent risk factor for the development of hepatocellular carcinoma (HCC). The HBV HBx gene is frequently identified as an integrant in the chromosomal DNA of patients with HCC. HBx encodes the X protein (HBx), a putative viral oncoprotein that affects transcriptional regulation of several cellular genes. Therefore, HBx may be an ideal target to impede the progression of HBV infection–related HCC. In this study, integrated HBx was transcriptionally downregulated using an artificial transcription factor (ATF). Two three-fingered Cys2-His2 zinc finger (ZF) motifs that specifically recognized two 9-bp DNA sequences regulating HBx expression were identified from a phage-display library. The ZF domains were linked into a six-fingered protein that specified an 18-bp DNA target in the Enhancer I region upstream of HBx. This DNA-binding domain was fused with a Krüppel-associated box (KRAB) transcriptional repression domain to produce an ATF designed to downregulate HBx integrated into the Hep3B HCC cell line. The ATF significantly repressed HBx in a luciferase reporter assay. Stably expressing the ATF in Hep3B cells resulted in significant growth arrest, whereas stably expressing the ATF in an HCC cell line lacking integrated HBx (HepG2) had virtually no effect. The targeted downregulation of integrated HBx is a promising novel approach to inhibiting the progression of HBV infection–related HCC.

Antiviral Potential of Exogenous Human Omega Interferon to Inhibit Pandemic 2009 A (H1N1) Influenza Virus

The pandemic 2009 H1N1 influenza virus broke out in North America and spread rapidly throughout the world. The type I interferon (IFN) response represents one of the first lines of defense against influenza virus infections. In this study, the protective potential of human exogenous IFN-ω against pandemic 2009 A (H1N1) influenza virus was assessed both in vitro and in guinea pigs. The viral loads of pandemic 2009 A (H1N1) influenza virus strains A/California/04/2009 and A/Beijing/501/2009 were reduced by up to 5000-fold in Caco-2 cells by the addition of human IFN-ω. With daily intranasal treatment with human IFN-ω the viral load of pandemic 2009 A (H1N1) influenza virus strain A/California/04/2009 decreased by 1000-fold in lung tissues of guinea pigs. These results provide strong support for the application of human IFN-ω pretreatment to human influenza control.

Intramuscular Delivery of Adenovirus Serotype 5 Vector Expressing Humanized Protective Antigen Induces Rapid Protection against Anthrax That May Bypass Intranasally Originated Preexisting Adenovirus Immunity

ABSTRACT Developing an effective anthrax vaccine that can induce a rapid and sustained immune response is a priority for the prevention of bioterrorism-associated anthrax infection. Here, we developed a recombinant replication-deficient adenovirus serotype 5-based vaccine expressing the humanized protective antigen (Ad5-PAopt). A single intramuscular injection of Ad5-PAopt resulted in rapid and robust humoral and cellular immune responses in Fisher 344 rats. Animals intramuscularly inoculated with a single dose of 108 infectious units of Ad5-PAopt achieved 100% protection from challenge with 10 times the 50% lethal dose (LD50) of anthrax lethal toxin 7 days after vaccination. Although preexisting intranasally induced immunity to Ad5 slightly weakened the humoral and cellular immune responses to Ad5-PAopt via intramuscular inoculation, 100% protection was achieved 15 days after vaccination in Fisher 344 rats. The protective efficacy conferred by intramuscular vaccination in the presence of preexisting intranasally induced immunity was significantly better than that of intranasal delivery of Ad5-PAopt and intramuscular injection with recombinant PA and aluminum adjuvant without preexisting immunity. As natural Ad5 infection often occurs via the mucosal route, the work here largely illuminates that intramuscular inoculation with Ad5-PAopt can overcome the negative effects of immunity induced by prior adenovirus infection and represents an efficient approach for protecting against emerging anthrax.

Human Prostate Stem Cell Antigen and HSP70 Fusion Protein Vaccine Inhibits Prostate Stem Cell Antigen-Expressing Tumor Growth in Mice

Prostate stem cell antigen (PSCA) has been considered a potentially worthwhile target for prostate cancer therapy with its overexpression in both androgen-dependent and androgen-independent prostate cancers. However, PSCA is an autoantigen that can evoke immunological tolerance and hardly incite effective immunologic response. In this study, we sought to construct the fusion protein vaccines based on PSCA and heat shock protein 70 (HSP70) and to evaluate their immune responses and therapeutic efficacy. A series of recombinant proteins were prepared, and then, the male C57BL/6 mice were immunized subcutaneously by inoculation with RM-PSCA/Luc cells. The PSCA-specific cellular immune responses were monitored with ELISPOT and intracellular cytokines staining assay, and ELISA assay was used to detect humoral immune responses. The tumor growth was observed by in vivo bioluminescence imaging. The results showed that the mice vaccinated with PSCA-HSP could induce the PSCA-specific cellular and humoral immune responses. Tumor progression could be quantitatively monitored by in vivo bioluminescence imaging. Animal experiments showed that PSCA-HSP could inhibit the growth of PSCA-expressing tumors and prolong the survival time of vaccinated mice. This study supported and confirmed the potential of HSP70 as a chaperone for protein vaccines, and PSCA-HSP could be of potential value for prostate cancer treatment.

Monitoring luciferase-labeled human prostate stem cell antigen-expressing tumor growth in a mouse model

The aim of this study was to establish a tumor model in mice with the expression of luciferase (Luc) and human prostate stem cell antigen (PSCA), in order to evaluate the activities of anticancer drugs or vaccines for prostate cancer. RM-1 cells were stably transfected with pcDNA-Luc and pcDNA-PSCA plasmids. The Luc-expressing cells were examined using a luminometer and the PSCA-expressing cells were examined using a reverse transcription-polymerase chain reaction (RT-PCR) and flow cytometric analysis. Male C57BL/6 mice were inoculated subcutaneously with the RM-PSCA/Luc cells, prior to the tumor growth and survival time of the mice being measured, respectively. In vivo bioluminescence imaging was used to detect Luc expression and immunohistochemical analysis was used to detect PSCA expression. Inoculation of the tumor cells into the C57BL/6 mice closely mimicked the tumor growth of prostate cancer. All of the inoculated mice exhibited a detectable tumor within two weeks. Tumor progression was able to be quantitatively monitored following the inoculation of 1×106 RM-PSCA/Luc cells. There was an excellent correlation (R2=0.9849) between the photon counts and tumor volume. The expression of PSCA in tumor tissues was confirmed using immunohistochemical analysis. The Luc and PSCA co-expression tumor model was successfully established in mice, which is likely to accelerate the understanding of the pathogenesis of prostate cancer and facilitate the development of novel antitumor drugs or vaccines for the disease.

Immunity induced by Staphylococcus aureus surface protein A was protective against lethal challenge of Staphylococcus aureus in BALB/c mice

Staphylococcus aureus is the most common cause of hospital‐acquired bacteremia. Due to emergence of antibiotic‐resistant strains, these infections present a serious public health threat. In this study, to develop a broadly protective vaccine, we tested whether immune responses induced by several proteins associated with S. aureus toxicity could protect mice from lethal challenge with human clinical S. aureus isolate USA300. We found that the surface protein A (SasA) of S. aureus could protect mice from lethal challenge of the bacteria.

A conformational change of C fragment of tetanus neurotoxin reduces its ganglioside-binding activity but does not destroy its immunogenicity.

ABSTRACT The C fragment of tetanus neurotoxin (TeNT-Hc) with different conformations was observed due to the four cysteine residues within it which could form different intramolecular disulfide bonds. In this study, we prepared and compared three types of monomeric TeNT-Hc with different conformational components: free sulfhydryls (50 kDa), bound sulfhydryls (44 kDa), and a mixture of the two conformational proteins (half 50 kDa and half 44 kDa). TeNT-Hc with bound sulfhydryls reduced its binding activity to ganglioside GT1b and neuronal PC-12 cells compared to what was seen for TeNT-Hc with free sulfhydryls. However, there was no significant difference among their immunogenicities in mice, including induction of antitetanus toxoid IgG titers, antibody types, and protective capacities against tetanus neurotoxin challenge. Our results showed that the conformational changes of TeNT-Hc resulting from disulfide bond formation reduced its ganglioside-binding activity but did not destroy its immunogenicity, and the protein still retained continuous B cell and T cell epitopes; that is, the presence of the ganglioside-binding site within TeNT-Hc may be not essential for the induction of a fully protective antitetanus response. TeNT-Hc with bound sulfhydryls may be developed into an ideal human vaccine with a lower potential for side effects.