Zhengrong Cui

Strong antibody responses induced by protein antigens conjugated onto the surface of lecithin-based nanoparticles.

An accumulation of research over the years has demonstrated the utility of nanoparticles as antigen carriers with adjuvant activity. Herein we defined the adjuvanticity of a novel lecithin-based nanoparticle engineered from emulsions. The nanoparticles were spheres of around 200nm. Model protein antigens, bovine serum albumin (BSA) or Bacillus anthracis protective antigen (PA) protein, were covalently conjugated onto the nanoparticles. Mice immunized with the BSA-conjugated nanoparticles developed strong anti-BSA antibody responses comparable to that induced by BSA adjuvanted with incomplete Freunds adjuvant and 6.5-fold stronger than that induced by BSA adsorbed onto aluminum hydroxide. Immunization of mice with the PA-conjugated nanoparticles elicited a quick, strong, and durable anti-PA antibody response that afforded protection of the mice against a lethal dose of anthrax lethal toxin challenge. The potent adjuvanticity of the nanoparticles was likely due to their ability to move the antigens into local draining lymph nodes, to enhance the uptake of the antigens by antigen-presenting cells (APCs), and to activate APCs. This novel nanoparticle system has the potential to serve as a universal protein-based vaccine carrier capable of inducing strong immune responses.

Nanoparticles engineered from lecithin-in-water emulsions as a potential delivery system for docetaxel

Docetaxel is a potent anticancer drug. However, there continues to be a need for alternative docetaxel delivery systems to improve its efficacy. We reported the engineering of a novel spherical nanoparticle formulation ( approximately 270 nm) from lecithin-in-water emulsions. Docetaxel can be incorporated into the nanoparticles, and the resultant docetaxel-nanoparticles were stable when stored as an aqueous suspension. The release of the docetaxel from the nanoparticles was likely caused by a combination of diffusion and Case II transport. The docetaxel-in-nanoparticles were more effective in killing tumor cells in culture than free docetaxel. Moreover, the docetaxel-nanoparticles did not cause any significant red blood cell lysis or platelet aggregation in vitro, nor did they induce detectable acute liver damage when injected intravenously into mice. Finally, compared to free docetaxel, the intravenously injected docetaxel-nanoparticles increased the accumulation of the docetaxel in a model tumor in mice by 4.5-fold. These lecithin-based nanoparticles have the potential to be a novel biocompatible and efficacious delivery system for docetaxel.

Nasal Immunization with Anthrax Protective Antigen Protein Adjuvanted with Polyriboinosinic–Polyribocytidylic Acid Induced Strong Mucosal and Systemic Immunities

PurposeThe current anthrax vaccine adsorbed (AVA) was originally licensed for the prevention of cutaneous anthrax infection. It has many drawbacks, including the requirement for multiple injections and subsequent annual boosters. Thus, an easily administrable and efficacious anthrax vaccine is needed to prevent the most lethal form of anthrax infection, inhalation anthrax. We propose to develop a nasal anthrax vaccine using anthrax protective antigen (PA) protein as the antigen and synthetic double-stranded RNA in the form of polyriboinosinic–polyribocytidylic acid (pI:C) as an adjuvant.MethodsMice were nasally immunized with recombinant PA admixed with pI:C. The resulting PA-specific antibody responses and the lethal toxin neutralization activity were measured. Moreover, the effect of pI:C on dendritic cells (DCs) was evaluated both in vivo and in vitro.ResultsMice nasally immunized with rPA adjuvanted with pI:C developed strong systemic and mucosal anti-PA responses with lethal toxin neutralization activity. These immune responses compared favorably to that induced by nasal immunization with rPA adjuvanted with cholera toxin. Poly(I:C) enhanced the proportion of DCs in local draining lymph nodes and stimulated DC maturation.ConclusionsThis pI:C-adjuvanted rPA vaccine has the potential to be developed into an efficacious nasal anthrax vaccine.

Non-invasive immunization on the skin using DNA vaccine.

Skin has evolved to protect not only by acting as a physical barrier, but also by its role in our powerful immune system. As a frontline of the hosts defense against pathogens, skin is well equipped for immune surveillance. For example, compared to many other tissues, the epidermis of the skin contains a high population of Langerhans cells, which are very potent immature dendritic cells. Thus, targeting antigens to the skin epidermis should be able to efficiently induce strong immune responses. However, the forbidden barrier posed by the stratum corneum layer of the epidermis prevents effective entrance of antigens into the epidermis. Nevertheless, non-invasive immunization onto the skin has proven in the last several years to be a viable immunization modality. DNA vaccine is a vaccine made of bacterial plasmid DNA encoding an antigen of interest. Upon uptake of the plasmid, host express and process the encoding antigen, and then mount immune responses against it. DNA vaccine is advantageous over many other types of vaccines. The feasibility of non-invasive immunization onto the skin with DNA vaccine has been confirmed. Although the potency of the immune response has proven to be weak, many skin stratum corneum disrupting chemical and physical approaches and DNA vaccine carriers/adjuvants that significantly enhance the resulting immune response have been reported. In addition, research on elucidating the mechanism of immune induction from non-invasively, topically applied DNA vaccine has also been carried out. With further improvement and optimization, non-invasive immunization onto the skin with DNA vaccine should be able to elicit reliable and efficacious immune response to a variety of antigens.

A Thermo-Sensitive Polymeric Gel Containing a Gadolinium (Gd) Compound Encapsulated into Liposomes Significantly Extended the Retention of the Gd in Tumors

Gadolinium neutron capture therapy (Gd-NCT) is a promising approach to fight cancer. One key factor for the success of Gd-NCT is to deliver and maintain a sufficient amount of Gd inside tumors. A large amount of Gd can be readily introduced into tumors by direct intratumor injection. However, an innovative approach is needed to maintain the Gd in the tumors. We encapsulated a Gd compound into a liposome formulation and then dispersed the liposomes into a thermo-sensitive polymeric gel. In murine tumor models, we showed that this liposome-in-thermo-sensitive gel system significantly extended the retention of the Gd compound in tumors. This similar concept may be applied to prolong the retention of other cytotoxic chemicals in tumors, and thus, improve their anti-tumor efficacy.

Localized irradiation of tumors prior to synthetic dsRNA therapy enhanced the resultant anti-tumor activity

BACKGROUND AND PURPOSE Despite the potent tumoricidal activity of the synthetic dsRNA in culture, its in vivo anti-tumor activity has proven to be limited. We sought to devise and validate a new strategy to improve the in vivo anti-tumor activity by integrating localized irradiation into dsRNA therapy. MATERIALS AND METHODS Using a mouse lung cancer model and a mouse melanoma model in immuno-competent mice or athymic nude mice, we evaluated the combined anti-tumor activity using a synthetic dsRNA, polyinosine-cytosine (poly(I:C)). RESULTS Localized irradiation of tumors prior to the poly(I:C) therapy significantly delayed the tumor growth as compared to monotherapies using the radiation or poly(I:C) alone. The poly(I:C) enhanced the tumor response to radiation with a dose modification factor as large as 20. The combined effect was synergistic only in immuno-competent mice with highly immunogenic tumors. The anti-tumor activity of the combination therapy was significantly impaired when the type I interferons in the mice were neutralized. CONCLUSIONS This combination modality may represent a promising approach to exploit synthetic dsRNA in cancer therapy and to enhance tumor response to radiation. T cell-mediated immunity was likely responsible for the combined synergistic effect. Type I interferons contributed significantly to the combined anti-tumor activity.

Nasal immunization with the mixture of PA63, LF, and a PGA conjugate induced strong antibody responses against all three antigens

A new generation anthrax vaccine is expected to target not only the anthrax protective antigen (PA) protein, but also other virulent factors of Bacillus anthracis. It is also expected to be amenable for rapid mass immunization of a large number of people. This study aimed to address these needs by designing a prototypic triantigen nasal anthrax vaccine candidate that contained a truncated PA (rPA63), the anthrax lethal factor (LF), and the capsular poly-gamma-D-glutamic acid (gammaDPGA) as the antigens and a synthetic double-stranded RNA (dsRNA), polyriboinosinic-polyribocytodylic acid (poly(I:C)) as the adjuvant. This study identified the optimal dose of nasal poly(I:C) in mice, demonstrated that nasal immunization of mice with the LF was capable of inducing functional anti-LF antibodies (Abs), and showed that nasal immunization of mice with the prototypic triantigen vaccine candidate induced strong immune responses against all three antigens. The immune responses protected macrophages against an anthrax lethal toxin challenge in vitro and enabled the immunized mice to survive a lethal dose of anthrax lethal toxin challenge in vivo. The anti-PGA Abs were shown to have complement-mediated bacteriolytic activity. After further optimization, this triantigen nasal vaccine candidate is expected to become one of the newer generation anthrax vaccines.

Learning from Viruses: The Necrotic Bodies of Tumor Cells with Intracellular Synthetic dsRNA Induced Strong Anti-tumor Immune Responses

PurposeCoaxing dead tumor cells to induce specific immune responses is an attractive tumor therapy. However, there continues to be a need for adjuvants that can promote the cross-presentation of the dead tumor cells to induce specific anti-tumor response. Viral dsRNA has multiple mechanisms to promote the cross-presentation of viral antigens in virus-infected cells. We propose to learn from viruses by generating dead tumor cells having synthetic dsRNA delivered inside them to allow the dsRNA to promote the cross-presentation of dead tumor cells.Materials and MethodsUsing synthetic dsRNA, poly(I:C), and the TC-1 cervical cancer model, we evaluated the extent to which the poly(I:C) can promote the necrotic bodies of TC-1 cells to induce specific anti-tumor immune response. The poly(I:C) was either simply mixed with the dead TC-1 cells or pre-loaded inside them.ResultsImmunization of tumor-bearing mice with the necrotic bodies of tumor cells admixed with poly(I:C) significantly inhibited the tumor growth. More importantly, immunization with the necrotic bodies having poly(I:C) pre-loaded inside led to a significantly stronger anti-tumor response than when the necrotic bodies were simply admixed with the poly(I:C), apparently through a CD8+ CTL response-mediated mechanism.ConclusionsThese findings are expected to be clinically relevant for devising improved whole cell-based tumor vaccines.

Evaluation of the immune response induced by a nasal anthrax vaccine based on the protective antigen protein in anaesthetized and non‐anaesthetized mice

To better protect against inhalational anthrax infection, a nasal anthrax vaccine based on the protective antigen (PA) protein of Bacillus anthracis could be an attractive alternative to the current Anthrax‐Vaccine‐Adsorbed (AVA), which was licensed for cutaneous anthrax prevention. Previously, we have demonstrated that an anti‐PA immune response comparable with that in mice subcutaneously immunized with PA protein adjuvanted with aluminium hydroxide was induced in both the systemic compartment and the mucosal secretions of the nose and lung of anaesthetized mice when they were nasally immunized with PA protein incorporated into previously reported LPD (Liposome—Protamine—DNA) particles. In this study, we evaluated the anti‐PA immune response induced by the nasal PA/LPD particles in non‐anaesthetized mice and compared it with that in anaesthetized mice. Our data showed that the anti‐PA antibody response and the anthrax lethal toxin‐neutralization activity induced by the nasal PA/LPD in non‐anaesthetized mice was relatively weaker than that in anaesthetized mice. However, the splenocytes isolated from the nasally immunized mice, anaesthetized and non‐anaesthetized, proliferated comparably after in‐vitro re‐stimulation. By evaluating the uptake of fluorescence‐labelled LPD particles by phagocytes in the nasal and broncho‐alveolar lavages of mice after the nasal administration, we concluded that the relatively weaker anti‐PA immune response in the non‐anaesthetized mice might be partially attributed to the reduced retention of the PA/LPD particles in the nasal cavity of the non‐anaesthetized mice. Data collected in this study are expected to be useful for future anthrax nasal vaccine studies when mice are used as a model.