Recombinant H1N1 virus-like particle vaccine elicits protective immunity in ferrets against the 2009 pandemic H1N1 influenza virus

The pandemic virus of 2009 (2009 H1N1) continues to cause illness worldwide, especially in younger age groups. The widespread H1N1 virus infection further emphasizes the need for vaccine strategies that are effective against emerging pandemic viruses and are not dependent on the limitations of traditional egg-based technology. This report describes a recombinant influenza virus-like particle (VLP) vaccine consisting of hemagglutinin (HA), neuraminidase (NA), and matrix (M1) proteins of influenza A/California/04/2009 (H1N1) virus. Influenza H1N1 VLPs with a diameter of approximately 120 nm were released into the culture medium from Sf9 insect cells infected with recombinant baculovirus coexpressing HA, NA, and M1 proteins. Purified recombinant H1N1 VLPs morphologically resembled influenza virions and exhibited biological characteristics of influenza virus, including HA and NA activities. In the ferret challenge model, 2009 influenza H1N1 VLPs elicited high-titer serum hemagglutination inhibition (HI) antibodies specific for the 2009 H1N1 virus and inhibited replication of the influenza virus in the upper and lower respiratory tract tissues following A/Mexico/4482/09 (H1N1) virus challenge. Moreover, a single 15 μg dose of H1N1 VLPs resulted in complete virus clearance in the ferret lung. These results provide support for the use of recombinant influenza VLP vaccine as an effective strategy against pandemic H1N1 virus.     

Preparation of a peptide vaccine against GnRH by a bioprocess system based on asparaginase

GnRH is a promising target in hormone-dependent cancer immunotherapy. In our previous study, we have designed and purified a peptide vaccine GhM (GnRH₃-hinge-MVP) by use of the bioprocess system based on asparaginase. Active immunization with GhM in the presence of CFA/IFA evoked strong humoral response. In this study, the motif NRLLLTG with high affinity to nanoparticle carrier VLP HBcΔ-SBD was fused to the C terminus of GhM to form a new peptide vaccine GhMNR (GnRH₃-hinge-MVP-NRLLLTG). The fusion protein ansB-C-GhMNR was controlled by vigorous T7lac promotor and expressed effectively as inclusion bodies after induction by lactose and then purified by means of cell disruption, washing and cold ethanol fractionation. After hydrolyzed for 72 h, GhMNR was liberated from the fusion partner ansB-C and purified by CM52 cation exchange chromatography. These results suggested that the bioprocess system is suitable for large-scale expression and purification of the peptide vaccine GhMNR, and even some other proteins or peptides which may be important for industrial or laboratory purposes.     

Comparative immunogenicity evaluations of influenza A virus M2 peptide as recombinant virus like particle or conjugate vaccines in mice and monkeys

Immunization against M2 peptide, also called M2e, from influenza A virus is an innovative vaccine approach for induction of cross-strain protective immunity. Two promising M2 vaccine compositions reported to date are M2 peptide chemically conjugated to carrier proteins or M2 peptide recombinantly expressed on the surface of virus like particles (VLPs) of hepatitis B virus core antigen (HBVc). To conduct a head-to-head comparison of these approaches, we constructed two recombinant HBVc VLPs expressing M2 peptide and prepared two conjugate vaccines with M2 peptide chemically coupled to Neisseria meningitidis outer membrane complex (OMPC) or HBVc VLP, respectively. Here, we showed superior immunogenicity of M2 peptide conjugated to OMPC and M2 peptide expressed on the surface of HBVc antigen based on dose–titration responses in mice. Surprisingly, HBVc expressing M2 peptide was an inferior vaccine in rhesus monkeys, whether as a primary vaccine or as a booster vaccine, when compared with M2-OMPC conjugate vaccine.     

Blocking IL-1α but not IL-1β increases susceptibility to chronic Mycobacterium tuberculosis infection in mice

IL-1α and IL-1β are potent inflammatory cytokines and important mediators of immune responses to intracellular pathogens such as Mycobacterium tuberculosis (Mtb). Here, we investigated the role of IL-1α and IL-1β during chronic Mtb infection and spontaneous reactivation in mice. For long-term neutralization of IL-1α, IL-1β or both, mice were immunized with virus-like particles (VLPs) displaying either of the cytokines, inducing strong and long-lasting neutralizing IgG responses. Blocking of IL-1α but not of IL-1β resulted in increased susceptibility to chronic infection with Mtb. Neutralizing either IL-1α or IL-1β alone did not lead to increased reactivation of latent tuberculosis. The generation of antibodies neutralizing both IL-1α and IL-1β simultaneously, did not influence weight gain during Mtb reactivation and the slight increase in pulmonary bacillary counts were not significant when compared to control-immunized group. Thus, the results suggest that IL-1α is the major mediator of the IL-1RI-dependent and protective innate immune responses to Mtb in mice.     

Intranasal delivery of Norwalk virus-like particles formulated in an in situ gelling, dry powder vaccine

The development of a vaccine to prevent norovirus infections has been focused on immunization at a mucosal surface, but has been limited by the low immunogenicity of self-assembling Norwalk virus-like particles (NV VLPs) delivered enterically or at nasal surfaces. Nasal immunization, which offers the advantage of ease of immunization, faces obstacles imposed by the normal process of mucociliary clearance, which limits residence time of applied antigens. Herein, we describe the use of a dry powder formulation (GelVac) of an inert in situ gelling polysaccharide (GelSite) extracted from Aloe vera for nasal delivery of NV VLP antigen. Powder formulations, with or without NV VLP antigen, were similar in structure in dry form or when rehydrated in simulated nasal fluids. Immunogenicity of the dry powder VLP formulation was compared to equivalent antigen/adjuvant liquid formulations in animals. For the GelVac powder, we observed superior NV-specific serum and mucosal (aerodigestive and reproductive tracts) antibody responses relative to liquid formulations. Incorporation of the TLR7 agonist gardiquimod in dry powder formulations did not enhance antibody responses, although its inclusion in liquid formulations did enhance VLP immunogenicity irrespective of the presence or absence of GelSite. We interpret these data as showing that GelSite-based dry powder formulations (1) stabilize the immunogenic structural properties of VLPs and (2) induce systemic and mucosal antibody titers which are equal or greater than those achieved by VLPs plus adjuvant in a liquid formulation. We conclude that in situ gelation of the GelVac dry powder formulation at nasal mucosal surfaces delays mucociliary clearance and thereby prolongs VLP antigen exposure to immune effector sites.     

The L1 major capsid protein of HPV16 differentially modulates APC trafficking according to the vaccination or natural infection context

Human papillomavirus (HPV) infection, particularly type 16, is causally associated with cancer of the uterine cervix. The progression of cervical lesions suggests that viral antigens are not adequately presented to the immune system. The aim of this study was to determine whether HPV16 viral particles can influence the trafficking of human DC/Langerhans cells (LC), either by direct interactions with DC or following incubation with human normal keratinocytes that are in close contact with LC in the squamous epithelium. We first demonstrated that HPV16 L1 major capsid protein, when self-assembled into virus-like particles (VLP), is able to induce in DC an over-expression of CXC receptor 4 (CXCR4) via the activation of the NF-κB signaling pathway and to enhance DC motility in the presence of CXCL12, suggesting an ability to migrate towards lymph nodes. We also showed that conditioned media of HPV16 VLP-treated keratinocytes induce a lower LC migration than those from untreated keratinocytes and that prostaglandin E2 (PGE(2)), detected in HPV16 VLP-treated keratinocyte supernatants, may reduce LC recruitment into the squamous epithelium. Taken together, our data demonstrate that HPV16 VLP may differentially regulate the immune protective response according to their target cells.     

HPV-vaccination against cervical carcinoma: will it really work?

Prophylactic HPV vaccination provides an opportunity to profoundly affect cervical cancer incidence worldwide. The quadrivalent HPV VLP 6, 11, 16, 18 vaccine (Gardasil) and the bivalent HPV VLP 16, 18 vaccine (Cervarix) are effective for prevention of HPV infection and cervical precancerous lesions. The quadrivalent vaccine is also effective for prevention of vulvar and vaginal lesions and genital warts. With the introduction of the vaccines general issues have to be raised such as optimal age for vaccination, duration of protection after vaccination, impact on cervical cancer screening, vaccination of males and feasibility of application to developing countries. The prospects of a vaccine which will protect against the most common viral sexually transmitted infection and thereby, protect against the complications of HPV infection such as cervical cancer is extremely appealing. The success of HPV vaccination as a major public health prevention opportunity, however, will entirely depend on efficient infrastructures to deliver the vaccines and on the acceptance by individuals, parents and health care providers.     

血管迷走性晕厥儿童心室晚电位变化

目的探讨血管迷走性晕厥（VVS）儿童心室晚电位（VLP）的变化。方法直立倾斜试验（HUTT）阳性的VVS儿童45例（研究组），用广东中山SR-1000A心电综合自动分析仪通过体表叠加方法检测VLP。匹配健康儿童45例（对照组）。结果与对照组比较，研究组高频低幅时限（LAS40）明显延长（P〈0．05），心率、总QRS时间（TQRS）、均方根值（RMS40）差异无统计学意义（P〉0．05）。结论VVS儿童心肌电活动异常，存在心脏事件发生的危险性。