• Vaccine Name: Marburg Virus Vaccine PHV01
• Target Pathogen: Marburg Virus
• Target Disease: Hemorrhagic fever
• Type: Recombinant vector vaccine
• Status: Research
• Host Species for Licensed Use: None
• Antigen: MARV variant Angola glycoprotein (GP)
• GP from Marburgvirus gene engineering:
  • Type: Recombinant vector construction
  • Description:
  • Detailed Gene Information: Click Here.
• Preparation: The virus was generated via reverse genetics rescue essentially as described previously for a similar construct expressing the MARV variant Musoke GP [12]. A passage level 2 (P2) stock of virus was derived from the rescued virus (considered P0) via amplification on Vero E6 cells at the Public Health Agency of Canada. The P2 stock virus was subsequently amplified and plaque purified in qualified CGMP Vero cells in proprietary serum-free tissue culture media to generate premaster viral seeds at P8. A single P8 clone (Clone 5) was chosen based on plaque morphology, growth kinetics and productivity in Vero cells, and MARV GP transgene sequence fidelity. This clone was amplified to P9 under CGMP conditions to generate the PHV01 MVS. PHV01 MVS release criteria included the well-defined requirements for GMP viral seed manufacturing for purity (including adventitious agents), potency/strength, identity, and sterility. PHV01 MVS was further amplified to P10 under CGMP conditions to generate the PHV01 Working Viral Seed (WVS). The PHV01 Formulated Drug Substance (FDS) at P11 was produced from the PHV01 WVS in a disposable bioreactor using the same CGMP Vero cells and proprietary media described above. PHV01 FDS was harvested from cell culture medium containing virus, clarified, and further purified by nuclease digestion, depth filtration, and tangential flow ultrafiltration/diafiltration in a recombinant human albumin and Tris buffer formulation. Separately, an rVSVΔG-MARV P3 virus was generated from the initial P2 stock virus for research purposes, and PHV01 MVS was passaged three additional times (PHV01 MVS+3) to P12. (Zhu et al., 2022)
• Immunization Route: Intramuscular injection (i.m.)

Guinea pig Response

• Vaccination Protocol: This study was composed of two experiments, termed Experiment #1 and Experiment #2. Both were designed to test the ability of PHV01 to protect female Hartley guinea pigs (Charles River Laboratories) from morbidity and mortality resulting from inoculation with a lethal dose of GPA-MARV/Ang. In Experiment #1, rVSVΔG-MARV research virus P3 and PHV01 MVS were evaluated at three different dose levels: 2 x 10^6 plaque-forming units (PFU; high), 2 × 10^4 PFU (medium), and 2 × 102 PFU (low). Groups of 6 guinea pigs were immunized with one of the two vaccine preparations at one of the three specified doses (prepared with sterile, nontoxic, nonpyrogenic 0.9% saline as diluent), while a control group of 6 animals received an equivalent volume of 0.9% saline. In Experiment #2, the PHV01 FDS was evaluated at two different doses, high and medium (as described above), and PHV01 MVS was evaluated again at the high dose for comparative purposes. Groups of 6 guinea pigs were immunized with one of the two vaccine preparations at the specified doses (as described above), and a control group of 6 animals receiving 0.9% saline was also included. All vaccines were administered intramuscularly (IM) in a total volume of 300 µL, with 150 µL delivered to each of the rear quadriceps muscles. Twenty-eight (28) days postvaccination (DPV), animals were inoculated with 1000 times the median lethal dose (LD50) of GPA-MARV/Ang via intraperitoneal (IP) injection. Animals were monitored for disease and survival up to 29 days postinfection (DPI), equivalent to 57 DPV in Experiment #1 and up to 28 DPI (56 DPV) in Experiment #2. EDTA blood and plasma (Experiment #1) or serum (Experiment #2) samples were obtained at 0 DPV (prior to vaccination), 2 DPV, 27 DPV, 5 DPI (33 DPV), and 29 or 28 DPI (57 or 56 DPV). (Zhu et al., 2022)
• Immune Response: Vaccinemia:
  At 2 days postvaccination (DPV), the majority of vaccinated animals showed robust levels of RNA that loosely correlated with the dose level of vaccine administered. As expected, unvaccinated control animals exhibited no detectable levels of MARV GP RNA during the vaccination phase. With the exception of two guinea pigs, the three remaining vaccinated, non-surviving guinea pigs showed no detectable levels of MARV GP RNA postvaccination. By 27 DPV, MARV GP RNA was no longer detectable in any animal.
  
  Viremia:
  MARV GP-specific RNA in the blood of animals was quantified via RT-qPCR. At 5 DPI, all control animals exhibited very high levels of MARV RNA in the blood, with an average of approximately 9–10 Log10 GEQ/mL, indicating abundant MARV replication concomitant with severe infection. Conversely, the majority of vaccinated animals exhibited no viral RNA in the blood at 5 DPI, suggesting that vaccine-elicited immunity prevented MARV replication. Indeed, all vaccinated groups of animals showed significantly lower mean MARV RNA levels compared with the control animals.
  
  Humoral Response:
  The geometric mean endpoint titers for all vaccinated groups were significantly higher than that of the unvaccinated animals. Of the vaccinated, non-surviving guinea pigs, most had low or undetectable IgG endpoint titers, which exhibited moderate IgG levels. Pooling the data from the PHV01 MVS and FDS groups showed no significant difference in mean IgG titers between the medium and high dose levels, suggesting that both doses of vaccine elicited similarly robust immune responses. Moreover, logistic regression analysis comparing IgG endpoint titers and survival for all animals immunized with PHV01 MVS or FDS revealed a 90% probability of surviving infection with an IgG endpoint titer of 1600 (~Log10 3.2), which was achieved in almost all vaccinated animals from the medium- and high-dose groups. At 28 or 29 DPI (56 or 57 DPV), IgG endpoint titers increased to similar levels for all surviving animals, signifying an enhancement in the immune response to MARV following infection.
  
  Neutralizing Antibody Response:
  Pooling the data from the clonal PHV01-vaccinated animals revealed that the high-dose groups had statistically significant higher geometric mean PRNT50 endpoint titers compared to both the control animals and the animals vaccinated with the medium dose. Together, these data suggest that immunization with PHV01 elicits a potent and dose-dependent neutralizing antibody response. (Zhu et al., 2022)
• Side Effects: All vaccinated, surviving animals gained weight following GPA-MARV/Ang infection, and the vast majority exhibited no clinical signs of disease. Indeed, only 3 of the 32 vaccinated, surviving animals showed any clinical signs, which were all extremely mild (e.g., minor weight loss, ruffling of fur, and/or moderately reduced activity) and resolved completely within a few days and before 14 DPI. (Zhu et al., 2022)
• Challenge Protocol: 1000 LD50 dose of GPA-MARV/Ang (Zhu et al., 2022)
• Efficacy: In general, vaccination with rVSVΔG-MARV P3 and PHV01 prevented severe disease and death due to MARV infection in nearly all animals. Animals vaccinated with the high dose of rVSVΔG-MARV P3, PHV01 MVS, or PHV01 FDS showed 100% protection from MARV, as did animals vaccinated with the medium dose of PHV01 MVS. The medium and low dose of rVSV-MARV P3, as well as the medium dose of PHV01 FDS, resulted in 83% survival, with one of six guinea pigs in each group (animals 9, 16, and 60) succumbing to MARV infection. The low dose of PHV01 MVS resulted in 67% survival, with two of six guinea pigs (animals 31 and 32) succumbing to infection. In total, of the 54 animals that were vaccinated with any vaccine, 49 survived GPA-MARV/Ang challenge, giving an overall vaccine efficacy of ~91%. To specifically assess the level of protection conferred by the clonal PHV01, we pooled the data from the PHV01 MVS and FDS medium- and high-dose groups. All animals (n = 18) that received the high dose of these vaccines survived, while only 1 of 12 animals that received the medium dose succumbed. Of the 30 animals vaccinated with either the high or medium dose of PHV01 MVS or FDS, 29 survived, giving an overall PHV01 vaccine efficacy of 97%. (Zhu et al., 2022)
• Description: PHV01 provided high survival rates against homologous MARV/Ang challenge, protecting 100% of the animals at the highest dose level tested and offering significant protection even at very low dose levels. (Zhu et al., 2022)