Immunization with a plasmid encoding a modified hepatitis B surface antigen carrying the receptor binding site for hepatocytes

Intramuscular injection of a plasmid encoding a modified hepatitis B surface antigen (HBsAg) induced humoral and cytotoxic responses in C57BL/6 mice. This modified HBsAg contains a preS1-derived peptide (amino acids 21 to 47), that carries the HBV receptor binding site for hepatocytes fused to the C-terminus of the small protein (at the position of amino acid 223). After a single DNA injection, the immunized mice elicited high titers of anti-HBs and anti-preS1 antibodies, and produced strong HBV specific cytotoxic T-lymphocyte (CTL) responses. The immune response induced after a single injection of this modified HBsAg gene in HBsAg-transgenic mice resulted in the clearance of circulating HBsAg and the appearance of anti-HBs and anti-preS1 antibodies. The high titers of preS1 antibody in transgenic mice were comparable to those found in non-transgenic controls and may be efficient to clear Dane particles existing in sera from chronic carriers. These data indicated that a genetic vaccine consisting of this modified HBsAg gene may have a potential use for both prophylactic and therapeutic purposes.     

Hepatitis B surface antigen (HBsAg) and core antigen (HBcAg) combine CpG oligodeoxynucletides as a novel therapeutic vaccine for chronic hepatitis B infection

Hepatitis B virus infection is a non-cytopathic hepatotropic virus which can lead to chronic liver disease and hepatocellular carcinoma. Traditional therapies fail to provide sustained control of viral replication and liver damage in most patients. As an alternative strategy, immunotherapeutic approaches have shown promising efficacy in the treatment of chronic hepatitis B patients. Here, we investigated the efficacy of a novel therapeutic vaccine formulation consisting of two HBV antigens, HBsAg and HBcAg, and CpG adjuvant. This vaccine formulation elicits forceful humoral responses directed against HBsAg/HBcAg, and promotes a Th1/Th2 balance response against HBsAg and a Th1-biased response against HBcAg in both C57BL/6 and HBV transgenic mice. Vigorous cellular immune response was also detected in HBV transgenic mice, for a significantly higher number of HBs/HBc-specific IFN-γ secreting CD4+ and CD8+ T cells was generated. Moreover, vaccinated mice elicited significantly intense in vivo CTL attack, reduced serum HBsAg level without causing liver damage in HBV transgenic mice. In summary, this study demonstrates a novel therapeutic vaccine with the potential to elicit vigorous humoral and cellular response, overcoming tolerance in HBV transgenic mice.     

Augmented humoral and cellular immune responses of a hepatitis B DNA vaccine encoding HBsAg by protein boosting

Several reports have indicated that combinatorial regimens with DNA and protein vaccines can elicit both strong immune responses, to circumvent the limits of each vaccine. Surprisingly little was known on HBV vaccine. Here, we investigated the immunization effects of several regimens in BALB/c mice. The level of total antibody and isotypes of IgG were determined by ELISA. Cellular immune responses were assayed by measuring the production of cytokines and CTL activity after re-stimulation for 7 days in vitro with tumor cells CT26/S stably expressing HBsAg. The efficacy of immunoprotection against the challenge of transplanted CT26/S was also examined. The regimen involving twice priming pVAX(S) encoding HBsAg and once recombinant HBsAg protein (rHBsAg) boosting, induced strong and homogenous antibody responses. This regimen induced significant stronger responses of interleukin-12 and gamma interferon (IFN-gamma) in splenocytes, and elicited stronger CD8+ CTL responses and greater immunopretectional efficacy than those elicited by immunization with rHBsAg or pVAX(S) alone. Our regimen may thus provide a strategy for developing an improved immunization against HBV and many other pathogens.     

A truncated C-terminal fragment of Mycobacterium tuberculosis HSP70 gene enhanced potency of HBV DNA vaccine

DNA vaccine represents an attractive approach to therapy of chronic hepatitis B virus (HBV) infection because of its ability to generate antigen-specific immunity; nevertheless, there is still a need to increase the potency of DNA vaccine. Mycobacterium tuberculosis heat shock protein70 (HSP70) has both chaperon and cytokine functions, and has been shown to act as an adjuvant when co-administered with peptide antigens or given as fusion proteins. Here we evaluated the effects of two truncated HSP70 molecules, N-terminal domain (HSP70(1-360), amino acids 1-360) and C-terminal domain (HSP70(359-610), amino acids 359-610) of mycobacterial HSP70, on the potency of antigen-specific immunity generated by a HBV DNA vaccination. We found that only the HSP70(359-610)-fused HBV DNA vaccination resulted in a significant increase in hepatitis B surface antigen (HBsAg)-specific humoral response, while the HSP70(1-360)- or the complete HSP70 molecule-fused vaccine did not. Moreover, HSP70(359-610)-fused DNA vaccine did not induce anti-HSP70 antibody. Interestingly, HSP70(359-610) not only enhanced HBsAg-specific cytotoxic lymphocytes (CTL) responses but also overcame the epitope suppression caused by L(d)-restricted epitope. Meanwhile, HSP70(369-610) mediated T helper (Th) cell balance towards Th1 pathway. In a HBV transgenic mouse model, the HSP70(359-610) fusion vaccine facilitated clearance of circulating HBsAg and down-regulation of HBV replication. These results suggested that the truncated mycobacterial HSP70 molecule, HSP70(359-610), might be a superior candidate to deliver the adjuvant function in HBV DNA vaccination instead of the complete HSP70 molecule.     

Induction of antigen-specific CD8+ T cells, T helper cells, and protective levels of antibody in humans by particle-mediated administration of a hepatitis B virus DNA vaccine

A DNA vaccine against the hepatitis B virus (HBV) was evaluated for safety and induction of immune responses in 12 healthy, hepatitis-na?ve human volunteers using the needle-free PowderJect system to deliver gold particles coated with DNA directly into cells of the skin. Three groups of four volunteers received three administrations of DNA encoding the surface antigen of HBV at one of the three dose levels (1, 2, or 4 microg). The vaccine was safe and well tolerated, causing only transient and mild to moderate responses at the site of administration. HBV-specific antibody and both CD4+ and CD8+ T cell responses were measured before and after each immunization. All the volunteers developed protective antibody responses of at least 10 mIU/ml. In volunteers who were positive for the HLA class I A2 allele, the vaccine also induced antigen-specific CD8+ T cells that bound HLA-A2/HBsAg(335-343) tetramers, secreted IFN-gamma, and lysed target cells presenting a hepatitis B surface antigen (HBsAg) CTL epitope. Enumeration of HBsAg-specific T cells producing cytokine indicated preferential induction of a Type 1 T helper cell response. These results provide the first demonstration of a DNA vaccine inducing protective antibody titers and both humoral and cell-mediated immune responses in humans.     

Therapeutic efficacy of hepatitis B surface antigen-antibodies-recombinant DNA composite in HBsAg transgenic mice.

Therapeutic efficacy of HBsAg-anti-HBs-recombinant DNA harboring hepatitis B virus (HBV) S gene complex was compared with three other therapeutic vaccine candidates (recombinant HBsAg, HBsAg complexed to anti-HBs antibodies and naked plasmid DNA encoding the HBV S gene). After four injections at 3-week intervals, the most pronounced decrease of serum HBsAg, the highest titer of anti-HBs response, the highest level of interferon-gamma produced by splenocytes and potent cytotoxicity T cell response were observed in the HBsAg-anti HBs-sDNA immunized group. Reduced expression of HBsAg in hepatocytes was also shown. The therapeutic mechanism of HBsAg-anti-HBs-DNA was speculated as modulation of HBsAg presentation via both endogenous and exogenous pathways.     

Augmented humoral and cellular immune response of hepatitis B virus DNA vaccine by micro-needle vaccination using Flt3L as an adjuvant

The purpose of this work was to assess the immune response against plasmid DNA encoding hepatitis B virus (HBV) HBsAg in C57BL/6 mice inoculated by micro-needle. Fms-like tyrosine kinase 3 ligand (Flt3L), a hematopoietic growth factor, was used as an adjuvant. Immune response was determined by analysis of cytokine production. In addition, cytotoxic lymphocyte (CTL) activity was measured 7 days after in vitro addition of tumor cells (CT26/S) stabling expressing HBsAg. The efficacy of immunoprotection against CT26/S was determined by antibody response to this challenge. A strong antibody response was induced by inoculation with the DNA vaccine via micro-needles assisted with Flt3L, Administration of the vaccine to splenocytes induced significantly higher levels of interleukin-12 and gamma interferon. CTL response to the vaccine was stronger than that stimulated by intramuscular or micro-needle injections alone. Our study suggests useful strategies for improving the efficacy of vaccines against HBV and other pathogens.     

Immunogenicity of a hepatitis B DNA vaccine administered to chronic HBV carriers

Hepatitis B virus (HBV) is the major pathogen of chronic hepatitis and liver disease worldwide. Despite the availability of effective vaccines against hepatitis B for many years, over 370 million people remain persistently infected with HBV. Viral persistence is thought to be related to poor HBV-specific T-cell responses. Based on clinical data, the development of efficient methods capable of inducing strong T-cell responses is an important and primary step toward the development of immunotherapeutics against chronic HBV infection. We designed a phase I clinical trial in chronic HBV carriers to assess safety, tolerability and immunogenicity of a DNA vaccine expressing HBV small (S) and middle (preS2 +S) envelope proteins. After occurrence of lamivudine breakthrough, 10 HBeAg positive patients with chronic hepatitis B were followed longitudinally before, during and after DNA vaccine therapy. Immunizations were well tolerated and adverse physical events were mild and considered unrelated to the vaccine. Proliferative responses to hepatitis B surface antigen (HBsAg) were detected in two patients after DNA injections. Following three injections of vaccine, interferon (IFN)-gamma-producing T-cells specific for the preS2 or the S antigen were detectable in 50 and 100% of the patients, respectively. Each patient recognized at least one peptide within the envelope domain encoded by the vaccine. Anti-preS2 antibodies and seroconversion to anti-HBe were detected in two patients. This study shows evidences for the safety and immunological efficacy of HBV-DNA vaccination and demonstrates that DNA vaccination can restore or activate T-cell responses in chronic HBV carriers.     

A prophylactic hepatitis B vaccine with a novel adjuvant system

Studies with recombinant hepatitis B vaccines show seroprotection rates varying between 91 and 100%. Thus, a limited risk may remain for non-responding populations (e.g. non-responders, haemodialysis patients, elderly) who could benefit from a more immunogenic hepatitis B vaccine. One strategy to enhance the immune response is the use of novel adjuvants. SmithKline Beecham has developed a new adjuvant system containing alum and 3-deacylated monophosphoryl lipid A: SBAS4 (SmithKline Beecham Adjuvant System 4). Pilot studies showed that SBAS4 improved in vivo humoral and in vitro cellular immune responses compared to the response to classical recombinant hepatitis B vaccines and was safe and well-tolerated. Several studies assessed the profile of the HBsAg/SBAS4 vaccine in a healthy population, non-responders or elderly. In general the HBsAg/SBAS4 vaccine was well tolerated. Compared to an established recombinant hepatitis B vaccine, we observed an increased local reactogenicity but few symptoms were reported as severe. The HBsAg/SBAS4 vaccine elicits a strong immune response: subjects are protected faster and the GMTs are usually much higher. HBsAg/SBAS4 thus has the potential to protect those subjects who fail to be protected by well established hepatitis B vaccines.     

Oral administration of low doses of plant-based HBsAg induced antigen-specific IgAs and IgGs in mice,without increasing levels of regulatory T cells

Plant-based oral vaccines run the risk of activating regulatory T cells (Tregs) and suppressing the antigen-specific immune response via oral tolerance. Mice humanized for two HLA alleles (HLA-A2.1 and HLA-DR1) were used to measure changes in Tregs and antigen-specific immune responses induced by the oral administration of tobacco (Nicotiana tabacum), expressing the hepatitis B surface antigen (HBsAg). Antigen-specific CD8+ T cell activation was not detected, but the plant-based oral immunization, without adjuvant, resulted in humoral responses comparable to those obtained by adjuvanted DNA immunization. Treg titers did not increase with DNA immunization. In contrast, with plant immunization, Tregs increased linearly to reach a plateau at high antigen doses. The highest humoral IgA and IgG responses correlated with the lowest plant antigen dose (0.5 ng), while for DNA immunization the best antibody responses were obtained at higher antigen doses. These experiments suggest that plant-based oral vaccines could be adjusted to minimize tolerance, while still inducing an immune response. Oral tolerance and adjuvant engineering in plants are discussed.     

Anti-HBs characteristics after hepatitis B immunisation with plasma-derived and recombinant DNA-derived vaccines

Hepatitis B surface antigen derived from chronic hepatitis B carriers has been replaced almost completely by recombinant DNA-derived HBsAg for use as hepatitis B vaccine. Similarly, recombinant DNA-derived HBsAg is replacing plasma-derived HBsAg in standard anti-HBs assays. We analysed the influence of a change from plasma-derived HBsAg to recombinant DNA-derived HBsAg on antigen presentation in immunoassays and the characteristics of the anti-HBs antibodies after immunisation. Antigens and/or antibodies were subjected to three types of experiments: (a) binding of 'a'-loop specific monoclonal (anti-S) antibody conjugates to immobilised vaccine-HBsAg; (b) binding of post-vaccination anti-HBs to immobilised (vaccine-)HBsAg and (c) inhibition of HBsAg binding to immobilised monoclonal anti-HBs after pre-incubation with post-vaccination antibodies. Our results show that, in both antigen presentation and anti-HBs binding properties, yeast recombinant HBsAg and related antibodies could be clearly distinguished from plasma-derived HBsAg and related antibodies. Divergent results were also obtained in the inhibition assay with recombinant DNA-derived HBsAg but not with serum HBsAg from the vaccine HBsAg subtype. It is concluded that both antigen presentation in vaccines and in anti-HBs assays can markedly influence the quantitation anti-HBs response. It is suggested that a challenge with an heterologous hepatitis B virus may encounter reduced efficacy of vaccine antibodies.     

乙型肝炎基因重组疫苗免疫效果及持久性观察

目的 探讨乙肝基因重组疫苗对新生儿的免疫效果及免疫持久性。方法 随机选择珠海市1999～2003年出生、母亲HBsAg阴性、严格按免疫程序及时全程完成3针基础国产基因重组乙肝疫苗（5ug）的儿童，按免疫后不同的时间段分成6组，采集静脉血分别进行抗-HBs、HBsAg和抗-HBc检测。结果 抗-HBs有效阳性率（≥10mlU／ml）、几何平均滴度（GMT）逐年下降，分别由基础免疫完成后1个月的95，45％和65．36mlU／ml，降至4年后的38．27％和6．56mlU／ml，免疫1年后的下降率最为显著。及时完成3针基础乙肝疫苗免疫后各不同时间段的儿童HBsAg阳性率均不超过1％，抗-HBc阳性率和HBV感染率均小于2％，未见随年限延长而增张的趋势。结论 基因重组乙肝疫苗在新生儿中免疫效果良好，存在较好的免疫持久性和保护效果，在全程免疫后5年内没有必要进行加强。     

重组(酵母)乙型肝炎疫苗免疫后5年随访结果

目的 探讨新生儿接种国产重组(酵母)乙型肝炎(乙肝)疫苗后的免疫效果,并与血源乙肝疫苗效果比较.方法 对1997年出生并接种重组(酵母)乙肝疫苗的新生儿隔年随访一次,采血检测乙肝病毒表面抗原(HBsAg),乙肝病毒表面抗体(抗-HBs)和乙肝病毒核心抗体(抗-HBc),1998年以后对乙肝免疫人群开展急性乙肝发病监测....     

不同种类乙型肝炎重组疫苗诱导免疫应答特点比较

目的 比较三种乙型肝炎重组疫苗(rHB)[乙型肝炎汉逊酵母重组疫苗(rHP)、酿酒酵母重组疫苗(rSC)和CHO重组疫苗(rCHO)]诱导小鼠细胞和体液免疫应答特点.方法 给3组小鼠(BALB/c,H-2d)分别皮下接种3种r}HB 3μg,于免疫后第4、7、10、14、25和35 d分离脾单个核细胞(MNC),应用酶联斑点法(ELISPOT)测定MNCs体外刺激后所产生的细胞因子IFN-γ、IL-2斑点形成细胞数(SFC).刺激物为多肽或HBsAg.同时测定免疫后不同时间的细胞毒T淋巴细胞反应(CTL)活性和小鼠血清抗-HBs水平.结果 应用ELISPOT法,不同种类rHB免疫小鼠后诱导分泌IFN-γ和IL-2峰值均为10-14 d;rHP诱导CTL活性(P=0.000<0.05)和分泌IFN-γ水平(P<0.05)均显著高于rCHO和rSC,且rHP于免疫后第10天诱导CTL杀伤率最高(39.8%);rHP组诱导分泌IL-2水平均显著高于其他两种疫莆组(P<0.05),第7和14天时,rSC组诱导产生IL-2水平均显著高于rCHO组(第7天t=4.595,P=0.001<0.05;第14天t=5.721,P=0.000<0.05);免疫后第7天3种疫苗诱导细胞免疫应答强度依次为rHP最强,rSC其次,rCHO最弱;小鼠血清抗-HBs阳转率和抗体滴度随免疫时间而逐渐上升,第7天时rCHO组抗体显著高于rHP组(P=0.044<0.05),第14天时rCHO组抗体显著高于rHP组(P=0.009<0.05)和rSC组(P=0.012<0.05).结论 3种疫苗免疫小鼠的应答特点不同,酵母重组疫苗早期诱导细胞免疫应答优于rCHO;rCHO的优势是诱导产生抗-HBs早,且滴度较高.     

A new approach for therapeutic vaccination against chronic HBV infections

There are currently about 257 million people suffering from chronic HBV infection worldwide. In many cases, an insufficient T cell response is causative for establishment of a chronic infection. To ensure a robust cellular immune response and induction of neutralizing antibodies a novel vaccine platform based on modified cell–permeable HBV capsids was utilized. Cell permeability was achieved by fusion of the membrane–permeable TLM-peptide to HBV core monomers, assembling the capsids. Insertion of a Strep-tagIII into the spike tip domain that protrudes from the capsid surface enables flexible loading with antigens that are fused to streptavidin. In this study, HBV surface antigen-derived PreS1PreS2 domain, fused to monomeric streptavidin, served as cargo antigen. Binding between antigen and capsids was characterized by surface plasmon resonance spectroscopy, electron microscopy and density gradient centrifugation. Confocal immunofluorescence microscopy and in vivo imaging of immunized mice demonstrated membrane permeability of cargo-loaded carriers and spread of antigen over the whole organism. Immunization experiments of mice revealed a robust induction of a specific cellular immune response, leading to destruction of HBV-positive cells and induction of HBV-specific neutralizing antibodies. Membrane permeability of these carriers allows needle-free application of antigen-loaded capsids as evidenced by induction of an HBV-specific CTL response and HBV-specific B cell response after oral or transdermal vaccination.These data indicate that cell–permeable antigen carriers, based on HBV capsids and loaded with HBV antigen, have the capacity to induce a cellular and a neutralizing humoral immune response. In addition, cell permeability of the vaccine platform enables antigen transfer across several cell layers, that could allow oral or transdermal immunization.     

Oral delivery of wafers made from HBsAg-expressing maize germ induces long-term immunological systemic and mucosal responses

BackgroundThe hepatitis B surface antigen (HBsAg) has been administered over the last 20 years as a parenteral vaccine against the hepatitis B virus (HBV). Despite high seroconversion rates, chronic infection rates are still high worldwide. Orally delivered vaccines provide a practical alternative to injected vaccines, potentially helping poorly responding populations and providing a viable alternative for populations in remote locations. Anamnestic responses are vital to establishing the efficacy of a given vaccine and have been assessed in this study using a plant-based oral delivery platform expressing HBsAg.MethodsLong-term immunological memory was assessed in mice injected with a primary dose of Recombivax^® and boosted with orally-delivered HBsAg wafers, control wafers, or parenterally-delivered commercial vaccine (Recombivax^®).ResultsMice boosted with HBsAg orally-administered wafers displayed sharp increases in mucosal IgA titers in fecal material and steep increases in serum IgA, whereas mice boosted with Recombivax^® showed no detectable levels of IgA in either fecal or serum samples following four boosting treatments. Long-term memory in the orally-treated mice was evidenced by sustained fecal IgA, and serum IgA, IgG, and mIU/mL over one year, while Recombivax^®-treated mice displayed sustained serum IgG and mIU/mL. Furthermore, sharp increases in these same antibodies were induced after re-boosting at 47 and 50 weeks post-primary injection.ConclusionsOrally-delivered vaccines can provide long-term immune responses mucosally and systemically. For sexually-transmitted diseases that can be acquired at mucosal surfaces, such as HBV, an oral delivery platform may provide added protection over a conventional parenterally administered vaccine.     

Progress in DNA vaccine for prophylaxis and therapy of hepatitis B

Increasing lines of evidence suggest that DNA vaccine is of interest to fight chronic hepatitis B virus (HBV) infection. We used the Pekin duck infected by duck HBV (DHBV), closely related to the human virus, which is an attractive model allowing study of protective and therapeutic effectiveness of DNA vaccines against hepatitis B. Immunisation with a plasmid encoding the DHBV large (L) envelope protein induced a strong, specific, highly neutralising and long-lasting anti-preS humoral response in uninfected ducks. Importantly, maternal antibodies elicited by such DNA immunisation were vertically transmitted and protected progeny against viral challenge. Therapeutic immunisation of chronic DHBV-carrier ducks with this plasmid DNA led to the dramatic and sustained decrease in viral replication and even to clearance of intrahepatic viral covalently close circular DNA (cccDNA) pool in some animals. Our recent combination therapy data showed even a more pronounced antiviral effect of DNA vaccine to DHBV envelope protein when associated with antiviral drug (lamivudine) treatment. Therefore, DNA-based vaccine appears as a promising new approach for prophylaxis and therapy of hepatitis B.     

DNA immunization using a secreted cell wall antigen Mp1p is protective against Penicillium marneffei infection

None of the vaccines used in dimorphic fungal infections utilized the mucosal route for immunization, whereas only one utilized a secreted protein as antigen, despite knowing that infections caused by dimorphic fungi are usually acquired through inhalation. In this study, we investigated the usefulness of Mp1p (a secreted cell wall antigen encoded by MP1)-based vaccines for generation of protective immune responses against Penicillium marneffei infection using a mouse model, and compared the relative effectiveness of intramuscular MP1 DNA vaccine, oral mucosal MP1 DNA vaccine delivered by live-attenuated Salmonella typhimurium, and intraperitoneal recombinant Mp1p protein vaccine. The serum IgM level of the Mp1p protein vaccine group at day 7 and the serum IgG levels of the Mp1p protein vaccine group at days 7 and 21 were significantly higher than those of the other groups (P<0.0001). The serum IgG level of the MP1 DNA vaccine group was significantly higher than that of the corresponding control group and oral mucosal MP1 DNA vaccine group (one dose) at day 21 (P<0.0001 and <0.05, respectively). The groups of mice immunized with intramuscular MP1 DNA vaccine, oral mucosal MP1 DNA vaccine, and intraperitoneal Mp1p protein vaccine showed significantly higher Mp1p-specific lymphocyte proliferation index (LPI) than the control groups. The interferon-gamma (IF-gamma) levels of supernatant of splenic cell cultures obtained from mice after intramuscular MP1 DNA vaccine, mucosal MP1 DNA vaccine (three doses), or intraperitoneal Mp1p protein vaccine administration were higher than that which occurred after mucosal MP1 DNA vaccine (one dose) administration or those of controls. Interleukin-4 (IL-4) was not detectable in the supernatant of splenic cell cultures obtained from all groups of mice. The percentage survival of the mice immunized with intramuscular MP1 DNA vaccine, oral mucosal MP1 DNA vaccine (three doses), oral mucosal MP1 DNA vaccine (one dose), intraperitoneal recombinant Mp1p protein, oral live-attenuated S. typhimurium control, and intramuscular pJW4303 DNA control at day 60 after wild type P. marneffei challenge were 100, 60, 40, 40, 40, and 0%, respectively. The survival of mice in the MP1 DNA vaccine group was significantly better than those of the oral mucosal MP1 DNA vaccine (three doses) group (P<0.05), oral mucosal MP1 DNA vaccine (one dose) group (P<0.005), recombinant Mp1p protein group (P<0.005), S. typhimurium aroA strain group (P<0.05), and pJW4303 group (P<0.00001). Although, the mechanism by which intramuscular MP1 DNA vaccine offered the best protection against P. marneffei infection remains to be elucidated, the present observation prompted further clinical trials on the use of MP1 DNA immunization on asymptomatic human immunodeficiency virus carriers in P. marneffei endemic areas.     

Adjuvant effect of Japanese herbal medicines on the mucosal type 1 immune responses to human papillomavirus (HPV) E7 in mice immunized orally with Lactobacillus-based therapeutic HPV vaccine in a synergistic manner

The Japanese herbal medicines, Juzen-taiho-to (JTT) and Hochu-ekki-to (HET), have been shown to enhance humoral immune responses to vaccine antigen when used as adjuvants for prophylactic vaccines. However, their adjuvant effect on mucosal cellular immune responses remains unstudied. The precursor lesion of cervical cancer, high-grade CIN that expresses HPV E7 oncoprotein ubiquitously is a target for HPV therapeutic vaccines that elicit mucosal E7-specific type 1 T cell responses. We have demonstrated that oral immunization with recombinant Lactobacillus casei expressing HPV16 E7 (LacE7) is more effective in eliciting mucosal E7-specific IFNγ-producing cells than subcutaneous or intramuscular antigen delivery. Here we report the synergistic effect of an oral Lactobacillus-based vaccine and Japanese herbal medicines on mucosal immune responses. Oral immunization of mice with LacE7 plus either a Japanese herbal medicine (JTT or HET) or a mucosal adjuvant, heated-labile enterotoxin T subunit (LTB), promotes systemic E7-specific type 1 T cell responses but not mucosal responses. Administration of LacE7 plus either Japanese herbal medicine and LTB enhanced mucosal E7-specific type 1 T cell response to levels approximately 3-fold higher than those after administration of LacE7 alone. Furthermore, secretion of IFNγ and IL-2 into the intestinal lumen was observed after oral administration of LacE7 and was enhanced considerably by the addition of Japanese herbal medicines and LTB. Our data indicated that Japanese herbal medicines, in synergy with Lactobacillus and LTB, enhance the mucosal type 1 immune responses to orally immunized antigen. Japanese herbal medicines may be excellent adjuvants for oral Lactobacillus-based vaccines and oral immunization of LacE7, HET and LTB may have the potential to elicit extremely high E7-specific mucosal cytotoxic immune response to HPV-associated neoplastic lesions.     

Lentivector expressing HBsAg and immunoglobulin Fc fusion antigen induces potent immune responses and results in seroconversion in HBsAg transgenic mice

Even though hepatitis B virus(HBV) vaccines effectively prevent new cases of HBV infection, with approximately 350 million patients worldwide, chronic HBV infection remains a major health problem because of the associated complications (such as liver cirrhosis and hepatocellular carcinoma) and the limited treatment options. Immunotherapy has the potential to effectively control HBV replication. In this current study, we found that recombinant lentivectors could induce potent HBV surface antigen (HBsAg) specific T cell responses and humoral immune responses. Tagging the HBsAg with immunoglobulin Fc fragment further substantially increased the HBsAg specific immune responses. Remarkably, the HBS-Fc-lv lentivector could effectively break immune tolerance and induce potent HBsAg specific adaptive immune responses in HBsAg transgenic (Tg) mice with low serum level of HBsAg. More importantly, the induction of HBsAg specific immune responses in Tg mice accompanied seroconversion from HBsAg to anti-HBsAg antibody (anti-HBsAb). Our study demonstrated the potential of utilizing lentivector to treat chronic HBV infection following reduction of viral load with antiviral drug therapy.