Distinct Immunogenicity and Efficacy of Poxvirus-Based Vaccine Candidates against Ebola Virus Expressing GP and VP40 Proteins

Abstract

and species cause a severe disease in humans and nonhuman primates (NHPs) characterized by a high mortality rate. There are no licensed therapies or vaccines against Ebola virus disease (EVD), and the recent 2013 to 2016 outbreak in West Africa highlighted the need for EVD-specific medical countermeasures. Here, we generated and characterized head-to-head the immunogenicity and efficacy of five vaccine candidates against Zaire ebolavirus (EBOV) and Sudan ebolavirus (SUDV) based on the highly attenuated poxvirus vector modified vaccinia virus Ankara (MVA) expressing either the virus glycoprotein (GP) or GP together with the virus protein 40 (VP40) forming virus-like particles (VLPs). In a human monocytic cell line, the different MVA vectors (termed MVA-EBOVs and MVA-SUDVs) triggered robust innate immune responses, with production of beta interferon (IFN-β), proinflammatory cytokines, and chemokines. Additionally, several innate immune cells, such as dendritic cells, neutrophils, and natural killer cells, were differentially recruited in the peritoneal cavity of mice inoculated with MVA-EBOVs. After immunization of mice with a homologous prime/boost protocol (MVA/MVA), total IgG antibodies against GP or VP40 from Zaire and Sudan ebolavirus were differentially induced by these vectors, which were mainly of the IgG1 and IgG3 isotypes. Remarkably, an MVA-EBOV construct coexpressing GP and VP40 protected chimeric mice challenged with EBOV to a greater extent than a vector expressing GP alone. These results support the consideration of MVA-EBOVs and MVA-SUDVs expressing GP and VP40 and producing VLPs as best-in-class potential vaccine candidates against EBOV and SUDV. EBOV and SUDV cause a severe hemorrhagic fever affecting humans and NHPs. Since their discovery in 1976, they have caused several sporadic epidemics, with the recent outbreak in West Africa from 2013 to 2016 being the largest and most severe, with more than 11,000 deaths being reported. Although some vaccines are in advanced clinical phases, less expensive, safer, and more effective licensed vaccines are desirable. We generated and characterized head-to-head the immunogenicity and efficacy of five novel vaccines against EBOV and SUDV based on the poxvirus MVA expressing GP or GP and VP40. The expression of GP and VP40 leads to the formation of VLPs. These MVA-EBOV and MVA-SUDV recombinants triggered robust innate and humoral immune responses in mice. Furthermore, MVA-EBOV recombinants expressing GP and VP40 induced high protection against EBOV in a mouse challenge model. Thus, MVA expressing GP and VP40 and producing VLPs is a promising vaccine candidate against EBOV and SUDV.

Deletion of Vaccinia Virus A40R Gene Improves the Immunogenicity of the HIV-1 Vaccine Candidate MVA-B

“…Moreover, it has been reported that insertion of heterologous genes into the MVA HA locus does not affect the expression of neighboring MVA genes and the recombinant viruses generated are functional, indicating that the HA gene region is a suitable insertion site [89]. In fact, we have used the HA locus to insert in the MVA genome heterologous antigens from Leishmania [90] and Ebola virus [42], and this strategy was effective to trigger antigen-specific T cells, humoral immune responses, and protective efficacy.…”

“…This plasmid was used for the insertion of the MVA A40R gene into the MVA hemagglutinin (HA) locus of the MVA-B ∆A40R recombinant virus to generate the MVA-B ∆A40R-rev revertant virus. To construct the plasmid transfer vector pHA-A40R (7126 bp), the MVA A40R gene (526 bp) was amplified by PCR from the MVA-B genome with oligonucleotides LFA40R-XmaI-F (5 -TCCCCCCGGGATGAACAAACATAAGAC-3 ) (XmaI site underlined) and RFA40R-SacII-R (5 -AGGCCGCGGTTATTTTTTTCTAAAACACTC-3 ) (SacII site underlined), digested with XmaI and SacII restriction enzymes, and then inserted into the XmaI/SacII-digested pHA plasmid (6600 bp), the generation of which has been previously described [42]. Thus, pHA-A40R contained the MVA A40R gene under the control of the VACV sE/L promoter introduced in a multiple-cloning site between the MVA HA-L and HA-R flanking regions, and the selectable marker genes for ampicillin and β-glucuronidase (β-gus).…”

“…The MVA-B ∆A40R deletion mutant contained a deletion of the MVA A40R gene in the MVA-B genome and the MVA-B ∆A40R-rev revertant virus contained an insertion of the MVA A40R gene in the MVA HA locus of MVA-B ∆A40R. MVA-B ∆A40R and MVA-B ∆A40R-rev were generated using MVA-B and MVA-B ∆A40R as parental viruses, respectively, and pGem-RG-∆A40R wm and pHA-A40R as plasmid transfer vectors, respectively, employing an infection/transfection protocol previously described [18,20,23,42,43,50]. In the case of MVA-B ∆A40R, after the infection/transfection in DF-1 cells, we selected plaques expressing both Red2/GFP fluorescent proteins, then plaques expressing only GFP, and lastly we selected viruses from plaques that did not express any marker due to the loss of the fluorescent marker, as previously described [18,21].…”

An MVA Vector Expressing HIV-1 Envelope under the Control of a Potent Vaccinia Virus Promoter as a Promising Strategy in HIV/AIDS Vaccine Design

“…The MVA-LEO160-gp120 recombinant virus was generated using MVA-WT as parental virus, and pLZAW1-LEO160-gp120 as plasmid transfer vector; employing an infection/transfection protocol previously described [34][35][36][37][38][39]. After the infection/transfection in DF-1 cells, we initially selected blue plaques stained with 5-bromo-4-chloro-3-indolyl-beta-d-galactopyranoside (X-Gal, Sigma-Aldrich, St. Louis, MO, USA).…”

Ebola virus – prospects for a novel virus-like-particle-expressing modified vaccinia Ankara-based vaccine

“…Despite its abortive infection in primates, MVA has retained desirable features of its vaccinia parent: the elicitation of durable T cell and antibody (Ab) responses [12], the ability to be stored as a lyophilized product at ambient temperature (www.ClinicalTrials.gov; NCT00914732), and the ability to be used without an adjuvant. Due to its large genome size and the amount of coding capacity lost during adaptation (~20%), MVA can be used as a vector to express multiple vaccine antigens [13], a characteristic that readily supports the construction of recombinant MVAs expressing foreign virus-like particles (VLPs) [14][15][16]. Recently, promise for a safer and more stable vaccine has been demonstrated using MVA to express EBOV-like particles to achieve singledose protection in nonhuman primates [14].…”