Filovirus-like particles as vaccines and discovery tools

Ebola and Marburg viruses are members of the family Filoviridae, which cause severe hemorrhagic fevers in humans. Filovirus outbreaks have been sporadic, with mortality rates currently ranging from 30 to 90%. Unfortunately, there is no efficacious human therapy or vaccine available to treat disease caused by either Ebola or Marburg virus infection. Expression of the filovirus matrix protein, VP40, is sufficient to drive spontaneous production and release of virus-like particles (VLPs) that resemble the distinctively filamentous infectious virions. The addition of other filovirus proteins, including virion proteins (VP)24, 30 and 35 and glycoprotein, increases the efficiency of VLP production and results in particles containing multiple filovirus antigens. Vaccination with Ebola or Marburg VLPs containing glycoprotein and VP40 completely protects rodents from lethal challenge with the homologous virus. These candidate vaccines are currently being tested for immunogenicity and efficacy in nonhuman primates. Furthermore, the Ebola and Marburg VLPs are being used as a surrogate model to further understand the filovirus life cycle, with the goal of developing rationally designed vaccines and therapeutics. Thus, in addition to their use as a vaccine, VLPs are currently being used as tools to learn lessons about filovirus pathogenesis, immunology, replication and assembly requirements.     

Prospects for immunisation against Marburg and Ebola viruses

For more than 30 years the filoviruses, Marburg virus and Ebola virus, have been associated with periodic outbreaks of hemorrhagic fever that produce severe and often fatal disease. The filoviruses are endemic primarily in resource‐poor regions in Central Africa and are also potential agents of bioterrorism. Although no vaccines or antiviral drugs for Marburg or Ebola are currently available, remarkable progress has been made over the last decade in developing candidate preventive vaccines against filoviruses in nonhuman primate models. Due to the generally remote locations of filovirus outbreaks, a single‐injection vaccine is desirable. Among the prospective vaccines that have shown efficacy in nonhuman primate models of filoviral hemorrhagic fever, two candidates, one based on a replication‐defective adenovirus serotype 5 and the other on a recombinant VSV (rVSV), were shown to provide complete protection to nonhuman primates when administered as a single injection. The rVSV‐based vaccine has also shown utility when administered for postexposure prophylaxis against filovirus infections. A VSV‐based Ebola vaccine was recently used to manage a potential laboratory exposure. Copyright © 2010 John Wiley & Sons, Ltd.     

Progress in filovirus vaccine development: evaluating the potential for clinical use

Marburg and Ebola viruses cause severe hemorrhagic fever in humans and nonhuman primates. Currently, there are no effective treatments and no licensed vaccines; although a number of vaccine platforms have proven successful in animal models. The ideal filovirus vaccine candidate should be able to provide rapid protection following a single immunization, have the potential to work postexposure and be cross-reactive or multivalent against all Marburg virus strains and all relevant Ebola virus species and strains. Currently, there are multiple platforms that have provided prophylactic protection in nonhuman primates, including DNA, recombinant adenovirus serotype 5, recombinant human parainfluenza virus 3 and virus-like particles. In addition, a single platform, recombinant vesicular stomatitis virus, has demonstrated both prophylactic and postexposure protection in nonhuman primates. These results demonstrate that achieving a vaccine that is protective against filoviruses is possible; the challenge now is to prove its safety and efficacy in order to obtain a vaccine that is ready for human use.     

Protection against filovirus infection: virus-like particle vaccines

Significant progress has been made in vaccine development against infection by Ebola and Marburg viruses, members of the Filoviridae, which cause severe hemorrhagic fevers in humans with no effective treatment and a mortality rate of up to 90%. Several vaccine strategies have been shown to effectively protect immunized animals against filovirus infection. Among these candidate vaccine strategies, virus-like particles represent a promising approach and have been shown to protect small laboratory animals as well as nonhuman primates against lethal challenge by Ebola and/or Marburg viruses. This review briefly summarizes filovirus epidemiology and pathogenesis, and focuses on the discussion of recent advances in filovirus vaccine development and the current understanding of protective immune responses against filovirus infection with an emphasis on the progress and challenge of filovirus virus-like particle vaccine development.     

Perspectives towards antiviral drug discovery against Ebola virus

Ebola virus disease (EVD), caused by Ebola viruses, resulted in more than 11 500 deaths according to a recent 2018 WHO report. With mortality rates up to 90%, it is nowadays one of the most deadly infectious diseases. However, no Food and Drug Administration-approved Ebola drugs or vaccines are available yet with the mainstay of therapy being supportive care. The high fatality rate and absence of effective treatment or vaccination make Ebola virus a category-A biothreat pathogen. Fortunately, a series of investigational countermeasures have been developed to control and prevent this global threat. This review summarizes the recent therapeutic advances and ongoing research progress from research and development to clinical trials in the development of small-molecule antiviral drugs, small-interference RNA molecules, phosphorodiamidate morpholino oligomers, full-length monoclonal antibodies, and vaccines. Moreover, difficulties are highlighted in the search for effective countermeasures against EVD with additional focus on the interplay between available in silico prediction methods and their evidenced potential in antiviral drug discovery.     

Complex adenovirus-vectored vaccine protects guinea pigs from three strains of Marburg virus challenges

The Marburg virus (MARV), an African filovirus closely related to the Ebola virus, causes a deadly hemorrhagic fever in humans, with up to 90% mortality. Currently, treatment of disease is only supportive, and no vaccines are available to prevent spread of MARV infections. In order to address this need, we have developed and characterized a novel recombinant vaccine that utilizes a single complex adenovirus-vectored vaccine (cAdVax) to overexpress a MARV glycoprotein (GP) fusion protein derived from the Musoke and Ci67 strains of MARV. Vaccination with the cAdVaxM(fus) vaccine led to efficient production of MARV-specific antibodies in both mice and guinea pigs. Significantly, guinea pigs vaccinated with at least 5 x 10(7) pfu of cAdVaxM(fus) vaccine were 100% protected against lethal challenges by the Musoke, Ci67 and Ravn strains of MARV, making it a vaccine with trivalent protective efficacy. Therefore, the cAdVaxM(fus) vaccine serves as a promising vaccine candidate to prevent and contain multi-strain infections by MARV.     

Determination of Specific Antibody Responses to the Six Species of Ebola and Marburg Viruses by Multiplexed Protein Microarrays

Infectious hemorrhagic fevers caused by the Marburg and Ebola filoviruses result in human mortality rates of up to 90%, and there are no effective vaccines or therapeutics available for clinical use. The highly infectious and lethal nature of these viruses highlights the need for reliable and sensitive diagnostic methods. We assembled a protein microarray displaying nucleoprotein (NP), virion protein 40 (VP40), and glycoprotein (GP) antigens from isolates representing the six species of filoviruses for use as a surveillance and diagnostic platform. Using the microarrays, we examined serum antibody responses of rhesus macaques vaccinated with trivalent (GP, NP, and VP40) virus-like particles (VLP) prior to infection with the Marburg virus (MARV) (i.e., Marburg marburgvirus) or the Zaire virus (ZEBOV) (i.e., Zaire ebolavirus). The microarray-based assay detected a significant increase in antigen-specific IgG resulting from immunization, while a greater level of antibody responses resulted from challenge of the vaccinated animals with ZEBOV or MARV. Further, while antibody cross-reactivities were observed among NPs and VP40s of Ebola viruses, antibody recognition of GPs was very specific. The performance of mucin-like domain fragments of GP (GP mucin) expressed in Escherichia coli was compared to that of GP ectodomains produced in eukaryotic cells. Based on results with ZEBOV and MARV proteins, antibody recognition of GP mucins that were deficient in posttranslational modifications was comparable to that of the eukaryotic cell-expressed GP ectodomains in assay performance. We conclude that the described protein microarray may translate into a sensitive assay for diagnosis and serological surveillance of infections caused by multiple species of filoviruses.     

Why are vaccines against many human viral diseases still unavailable; an historic perspective?

The number of new and improved human viral vaccines licensed in recent years contrasts sharply with what could be termed the golden era (1955-1990) when vaccines against polio-, measles, mumps, rubella, and hepatitis B viruses first became available. Here, we attempt to explain why vaccines, mainly against viruses other than human immunodeficiency virus and hepatitis C virus, are still unavailable. They include human herpesviruses other than varicella-zoster virus, respiratory syncytial and most other respiratory, enteric and arthropod-borne viruses. Improved oral poliovirus vaccines are also urgently required. Their unavailability is attributable to regulatory/economic factors and the properties of individual viruses, but also to an absence of relevant animal models and ethical problems for the conduct of clinical of trials in pediatric and other critical populations. All are portents of likely difficulties for the licensing of effective vaccines against emerging pathogens, such as avian influenza, Ebola, and Zika viruses.     

Monoclonal antibodies to Ebola virus: isolation, characteristics, and study of cross reactivity with Marburg virus

Thirteen hybridoma strains producing monoclonal antibodies (Mabs) to Ebola virus were prepared by fusion of NS-O mouse myeloma cells with splenocytes of BALB/c mice immunized with purified and inactivated Ebola virus (Mayinga strain). Mabs directed against viral proteins were selected and tested by ELISA. Protein specificity of 13 Mabs was determined by immunoblotting with SDS-PAGE proteins of Ebola virus. Of these, 11 hybridoma Mabs reacted with 116 kDa protein (NP) and 2 with Ebola virus VP35. Antigenic cross-reactivity between Ebola and Marburg viruses was examined in ELISA and immunoblotting with polyclonal and monoclonal antibodies. In ELISA, polyclonal antibodies of immune sera to Ebola or Marburg viruses reacted with heterologous filoviruses, and two anti-NP Ebola antibodies (Mabs 7E1 and 6G8) cross-reacted with both viruses. Target proteins for cross-reactivity, Ebola NP (116 kDa) and Marburg NP (96 kDa), and VP35 of both filoviruses were detected by immunoblotting with polyclonal and monoclonal antibodies (6G8) to Ebola virus.     

Development of vaccines for Marburg hemorrhagic fever

Marburg (MARV) and Ebola viruses (EBOV) emerged from the rainforests of Central Africa more than 30 years ago causing outbreaks of severe and, usually, fatal hemorrhagic fever. EBOV has garnered the lion's share of the attention, fueled by the higher frequency of EBOV outbreaks, high mortality rates and importation into the USA, documented in such popular works as the best-selling novel 'The Hot Zone'. However, recent large outbreaks of hundreds of cases of MARV infection in the Democratic Republic of the Congo and Angola with case fatalities approaching 90% dramatically highlight its lethal potential. Although no vaccines or antiviral drugs for MARV are currently available, remarkable progress has been made over the last few years in developing potential countermeasures against MARV in nonhuman primate models. In particular, a vaccine based on attenuated recombinant vesicular stomatitis virus was recently shown to have both preventive and postexposure efficacy.     

Inhibition of Lassa virus and Ebola virus infection in host cells treated with the kinase inhibitors genistein and tyrphostin

Arenaviruses and filoviruses are capable of causing hemorrhagic fever syndrome in humans. Limited therapeutic and/or prophylactic options are available for humans suffering from viral hemorrhagic fever. In this report, we demonstrate that pre-treatment of host cells with the kinase inhibitors genistein and tyrphostin AG1478 leads to inhibition of infection or transduction in cells infected with Ebola virus, Marburg virus, and Lassa virus. In all, the results demonstrate that a kinase inhibitor cocktail consisting of genistein and tyrphostin AG1478 is a broad-spectrum antiviral that may be used as a therapeutic or prophylactic against arenavirus and filovirus hemorrhagic fever.     

Towards a vaccine against Ebola virus

Ebola virus infection causes hemorrhagic fever with high mortality rates in humans and nonhuman primates. Currently, there are no vaccines or therapies approved for human use. Outbreaks of Ebola virus have been infrequent, largely confined to remote locations in Africa and quarantine of sick patients has been effective in controlling epidemics. In the past, this small global market has generated little commercial interest for developing an Ebola virus vaccine. However, heightened awareness of bioterrorism advanced by the events surrounding September 11, 2001, concomitant with knowledge that the former Soviet Union was evaluating Ebola virus as a weapon, has dramatically changed perspectives regarding the need for a vaccine against Ebola virus. This review takes a brief historic look at attempts to develop an efficacious vaccine, provides an overview of current vaccine candidates and highlights strategies that have the greatest potential for commercial development.     

Detection of all known filovirus species by reverse transcription-polymerase chain reaction using a primer set specific for the viral nucleoprotein gene

The filoviruses, Marburg virus (MARV) and Ebola virus (EBOV), are causative agents of severe hemorrhagic fever with high mortality rates in humans and non-human primates. Sporadic outbreaks of filovirus infection have occurred in Central Africa and parts of Asia. Identification of the natural reservoir animals that are unknown yet and epidemiological investigations are current challenges to forestall outbreaks of filovirus diseases. The filovirus species identified currently include one in the MARV group and five in the EBOV group, with large genetic variations found among the species. Therefore, it has been difficult to develop a single sensitive assay to detect all filovirus species, which would advance laboratory diagnosis greatly in endemic areas. In this study, a highly sensitive universal RT-PCR assay targeting the nucleoprotein (NP) gene of filoviruses was developed. The genomic RNAs of all known MARV and EBOV species were detected by using an NP-specific primer set. In addition, this RT-PCR procedure was verified further for its application to detect viral RNAs in tissue samples of animals infected experimentally and blood specimens of infected patients. This assay will be a useful method for diagnostics and epidemiological studies of filovirus infections.     

Immunotherapy for Infectious Diseases: Past,Present, and Future

Passive immunotherapy for established infections, as opposed to active immunization to prevent disease, remains a tiny niche in the world of antimicrobial therapies. Many of the passive immunotherapies currently available are directed against bacterial toxins, such as botulism, or are intended for agents of bioterrorism such as anthrax, which fortunately has remained rare. The emergence of Ebola virus and multi-drug resistant pathogens, however, may breathe new life into the immunotherapy field as researchers seek non-antibiotic interventions for infectious diseases.     

Retracted: Ebola virus: an introduction and its pathology

The Ebola viruses are causative agent of a severe Ebola virus disease (EVD) or Ebola hemorrhagic fever (EHF) in human and other primates. Transmission of EVD occurs through the contact of body fluids from infected persons or animals, making it one of the most epidemic diseases worldwide. Underestimating the Ebola virus has cost loss of precious human lives in recent years. Ebola virus outbreak in year 2014 created a history, affecting a larger population in a wide geographical region of African sub‐continent. EVD outbreaks have a case fatality rate of up to 70%. Ebola viruses are endemic in regions of Africa. Ebola viruses mainly target the hepatocytes, endothelial, and macrophage‐rich lymphoid tissues and are characterized by immune suppression and a systemic inflammatory response that causes impairment of the vascular, coagulation, and immune systems. This impairment leads to multifocal necrosis and multi organ failure, and thus, in some ways, resembling septic shock. Currently, neither a specific treatment nor a vaccine licensed for use in humans is available. This review is focused on general characteristic of Ebola viruses, its pathogenesis, immunological response of host, and recent approaches for vaccine development against EVD. Copyright © 2015 John Wiley & Sons, Ltd.     

Progress in recombinant DNA-derived vaccines for Lassa virus and filoviruses

Developing vaccines for highly pathogenic viruses such as those causing Lassa, Ebola, and Marburg hemorrhagic fevers is a daunting task due to both scientific and logistical constraints. Scientific hurdles to overcome include poorly defined relationships between pathogenicity and protective immune responses, genetic diversity of viruses, and safety in a target population that includes a large number of individuals with compromised immune systems. Logistical obstacles include the requirement for biosafety level-4 containment to study the authentic viruses, the poor public health infrastructure of the endemic disease areas, and the cost of developing these vaccines for use in non-lucrative markets. Recombinant DNA-based vaccine approaches offer promise of overcoming some of these issues. In this review, we consider the status of various recombinant DNA candidate vaccines against Lassa virus and filoviruses which have been tested in animals.     

埃博拉疫苗研究进展

埃博拉病毒病是由埃博拉病毒引起的死亡率极高的烈性传染病。埃博拉疫情的大爆发促进了埃博拉疫苗的研制进程,多种在临床前阶段取得良好效果的埃博拉疫苗进入临床试验阶段,且已有两种疫苗在本国获批注册。本文对目前埃博拉疫苗的研究进展进行综述,旨在为疫苗研究者提供参考。     

Conservation of the 3' terminal nucleotide sequences of Ebola and Marburg virus

The 3' RNA base sequences of several Marburg (MBG) and Ebola (EBO) virus isolates have been determined. A comparison of these 3' terminal noncoding sequences with those of other negative strand RNA viruses suggests a unique phylogenic niche for Marburg and Ebola viruses. The translation initiation site and 35 N-terminal amino acids of the 3' proximal coding gene of a Zaire strain of Ebola virus was predicted. In addition, putative leader RNA sequences preceding the first gene are discussed in terms of possible regulatory functions.