Xiaoyu Wang

Rational design of thermostable vaccines by engineered peptide-induced virus self-biomineralization under physiological conditions

The development of vaccines against infectious diseases represents one of the most important contributions to medical science. However, vaccine-preventable diseases still cause millions of deaths each year due to the thermal instability and poor efficacy of vaccines. Using the human enterovirus type 71 vaccine strain as a model, we suggest a combined, rational design approach to improve the thermostability and immunogenicity of live vaccines by self-biomineralization. The biomimetic nucleating peptides are rationally integrated onto the capsid of enterovirus type 71 by reverse genetics so that calcium phosphate mineralization can be biologically induced onto vaccine surfaces under physiological conditions, generating a mineral exterior. This engineered self-biomineralized virus was characterized in detail for its unique structural, virological, and chemical properties. Analogous to many exteriors, the mineral coating confers some new properties on enclosed vaccines. The self-biomineralized vaccine can be stored at 26 °C for more than 9 d and at 37 °C for approximately 1 wk. Both in vitro and in vivo experiments demonstrate that this engineered vaccine can be used efficiently after heat treatment or ambient temperature storage, which reduces the dependence on a cold chain. Such a combination of genetic technology and biomineralization provides an economic solution for current vaccination programs, especially in developing countries that lack expensive refrigeration infrastructures.

Rational design of a flavivirus vaccine by abolishing viral RNA 2'-O methylation.

ABSTRACT Viruses that replicate in the cytoplasm cannot access the host nuclear capping machinery. These viruses have evolved viral methyltransferase(s) to methylate N-7 and 2′-O cap of their RNA; alternatively, they “snatch” host mRNA cap to form the 5′ end of viral RNA. The function of 2′-O methylation of viral RNA cap is to mimic cellular mRNA and to evade host innate immune restriction. A cytoplasmic virus defective in 2′-O methylation is replicative, but its viral RNA lacks 2′-O methylation and is recognized and eliminated by the host immune response. Such a mutant virus could be rationally designed as a live attenuated vaccine. Here, we use Japanese encephalitis virus (JEV), an important mosquito-borne flavivirus, to prove this novel vaccine concept. We show that JEV methyltransferase is responsible for both N-7 and 2′-O cap methylations as well as evasion of host innate immune response. Recombinant virus completely defective in 2′-O methylation was stable in cell culture after being passaged for >30 days. The mutant virus was attenuated in mice, elicited robust humoral and cellular immune responses, and retained the engineered mutation in vivo. A single dose of immunization induced full protection against lethal challenge with JEV strains in mice. Mechanistically, the attenuation phenotype was attributed to the enhanced sensitivity of the mutant virus to the antiviral effects of interferon and IFIT proteins. Collectively, the results demonstrate the feasibility of using 2′-O methylation-defective virus as a vaccine approach; this vaccine approach should be applicable to other flaviviruses and nonflaviviruses that encode their own viral 2′-O methyltransferases.

Human IgG Subclasses against Enterovirus Type 71: Neutralization versus Antibody Dependent Enhancement of Infection

The emerging human enterovirus 71 (EV71) represents a growing threat to public health, and no vaccine or specific antiviral is currently available. Human intravenous immunoglobulin (IVIG) is clinical used in treating severe EV71 infections. However, the discovery of antibody dependent enhancement (ADE) of EV71 infection illustrates the complex roles of antibody in controlling EV71 infection. In this study, to identify the distinct role of each IgG subclass on neutralization and enhancement of EV71 infection, different lots of pharmaceutical IVIG preparations manufactured from Chinese donors were used for IgG subclass fractionation by pH gradient elution with the protein A-conjugated affinity column. The neutralization and ADE capacities on EV71 infection of each purified IgG subclass were then assayed, respectively. The neutralizing activity of human IVIG is mainly mediated by IgG1 subclass and to less extent by IgG2 subclass. Interestingly, IgG3 fraction did not have neutralizing activity but enhanced EV71 infection in vitro. These results revealed the different roles of human IgG subclasses on EV71 infection, which is of critical importance for the rational design of immunotherapy and vaccines against severe EV71 diseases.

Biomineralization-Based Virus Shell-Engineering: Towards Neutralization Escape and Tropism Expansion

Biomineralization-based virus shell-engineering (BVSE) is a potential surface modification strategy to afford a biocompatible and biodegradable calcium phosphate (CaPi) shell onto single virus, allowing development of Trojan virus with enhanced infection, expanded tropism and neutralization escape, which open up the multiple applications of virus in biomedicines and materials.

Hydrated Silica Exterior Produced by Biomimetic Silicification Confers Viral Vaccine Heat-Resistance

Heat-lability is a key roadblock that strangles the widespread applications of many biological products. In nature, archaeal and extremophilic organisms utilize amorphous silica as a protective biomineral and exhibit considerable thermal tolerance. Here we present a bioinspired approach to generate thermostable virus by introducing an artificial hydrated silica exterior on individual virion. Similar to thermophiles, silicified viruses can survive longer at high temperature than their wild-type relatives. Virus inactivation assays showed that silica hydration exterior of the modified virus effectively prolonged infectivity of viruses by ∼ 10-fold at room temperature, achieving a similar result as that obtained by storing native ones at 4 °C. Mechanistic studies indicate that amorphous silica nanoclusters stabilize the inner virion structure by forming a layer that restricts molecular mobility, acting as physiochemical nanoanchors. Notably, we further evaluate the potential application of this biomimetic strategy in stabilizing clinically approved vaccine, and the silicified polio vaccine that can retain 90% potency after the storage at room temperature for 35 days was generated by this biosilicification approach and validated with in vivo experiments. This approach not only biomimetically connects inorganic material and living virus but also provides an innovative resolution to improve the thermal stability of biological agents using nanomaterials.

Virus capture and destruction by label-free graphene oxide for detection and disinfection applications.

Graphene oxide (GO) can efficiently capture viruses, destroy their surface proteins, and extract viral RNA in an aqueous environment by using the superficial bioreduction of GO. It follows from these phenomena that GO is an excellent nanomaterial for the high-throughput detection and disinfection of viruses, demonstrating its great potential for the prevention of environmental infections.

Biomineralization: From Material Tactics to Biological Strategy

Biomineralization is an important tactic by which biological organisms produce hierarchically structured minerals with marvellous functions. Biomineralization studies typically focus on the mediation function of organic matrices on inorganic minerals, which helps scientists to design and synthesize bioinspired functional materials. However, the presence of inorganic minerals may also alter the native behaviours of organic matrices and even biological organisms. This progress report discusses the latest achievements relating to biomineralization mechanisms, the manufacturing of biomimetic materials and relevant applications in biological and biomedical fields. In particular, biomineralized vaccines and algae with improved thermostability and photosynthesis, respectively, demonstrate that biomineralization is a strategy for organism evolution via the rational design of organism-material complexes. The successful modification of biological systems using materials is based on the regulatory effect of inorganic materials on organic organisms, which is another aspect of biomineralization control. Unlike previous studies, this study integrates materials and biological science to achieve a more comprehensive view of the mechanisms and applications of biomineralization.

Vaccine Engineering with Dual-Functional Mineral Shell: A Promising Strategy to Overcome Preexisting Immunity.

Dual-functional biomineral-vaccine core-shell nanohybrids are obtained using recombinant adenovirus serotype 5 (rAd5) as templates, which efficiently masks the neutralizing epitope of vaccines and preserve their original immunogenicity. The versatile vaccine hybrid can evade the preexisting anti-Ad5 immunity, leading to boosted multifunctional antigen-specific cytokine-secreting T cell responses and presenting promising applications of vaccine-material hybrid for the rational design of vaccines.

Chemotherapeutic Effects of Bioassay-Guided Extracts of the American Cockroach, Periplaneta americana

The organic extract of Periplaneta americana L. (Dictyoptera; Blattidae) has been traditionally used in southwestern China as an alternative medicine against disorders such as hepatitis, trauma, gastric ulcers, burns, and heart disease. The present study describes bioassay-guided purification and chemotherapeutic evaluation of the 60% ethanolic fraction of P americana organic extracts (PAE60). The most effective cytotoxic fraction was determined by way of repeated in vitro screenings against 12 distinct cultured human carcinoma cell lines: Eca 109, BGC823, HO8910, LS174T, CNE, HeLa, K562, PC-3, A549, BEL 7404, HL-60, and KB, followed by in vivo antitumor assays of the lead fraction (PAE60). The complexity of enriched active fraction was qualitatively evaluated using thin layer chromatography. Reconstituted PAE60 was effective at inhibiting HL-60, KB, CNE, and BGC823 cell growth with IC50 values <20 µg mL−1. PAE60 reduced tumor growth in S180-bearing immunocompetent mice by 72.62% after 10 days following oral doses of 500 mg kg d−1 compared with 78.75% inhibition following 40 mg kg d−1 of cyclophosphamide (CTX). Thymus and spleen indices of S180-bearing mice treated with PAE60 were significantly greater (P < .05) than CTX treatment groups, suggesting potential immunomodulation of antitumor host defenses by PAE60. Antiviral activity was also investigated and PAE60 inhibited herpes simplex type-2 replication (IC50 = 4.11 ± 0.64 µg mL−1) with a selectivity index (CC50 to IC50 ratio) of 64.84 in Vero cells but was less effective on type-1 virus (IC50 of 25.6 ± 3.16 µg mL−1). These results support future clinical trials on P. americana as an alternative or complementary medicinal agent.

Antioxidant and neuroprotective effects of synthesized sintenin derivatives

Three series of sintenin derivatives (compounds 1–14) were designed and prepared and their antioxidative and neuroprotective effects were evaluated. The in vitro models of scavenging 1,1-diphenyl-2-picrylhydrazyl (DPPH) radicals, chelating ferrous ions, inhibiting the rat brain homogenates lipid peroxidation, and protecting neurons damaged by hydrogen peroxide were employed for bioassays. It was found that sintenin derivatives 4 and 13 showed remarkable antioxidative and neuroprotective activities.