Human Polyclonal Antibodies Produced through DNA Vaccination of Transchromosomal Cattle Provide Mice with Post-Exposure Protection against Lethal Zaire and Sudan Ebolaviruses

Abstract

DNA vaccination of transchromosomal bovines (TcBs) with DNA vaccines expressing the codon-optimized (co) glycoprotein (GP) genes of Ebola virus (EBOV) and Sudan virus (SUDV) produce fully human polyclonal antibodies (pAbs) that recognize both viruses and demonstrate robust neutralizing activity. Each TcB was vaccinated by intramuscular electroporation (IM-EP) a total of four times and at each administration received 10 mg of the EBOV-GPco DNA vaccine and 10 mg of the SUDV-GPco DNA vaccine at two sites on the left and right sides, respectively. After two vaccinations, robust antibody responses (titers > 1000) were detected by ELISA against whole irradiated EBOV or SUDV and recombinant EBOV-GP or SUDV-GP (rGP) antigens, with higher titers observed for the rGP antigens. Strong, virus neutralizing antibody responses (titers >1000) were detected after three vaccinations when measured by vesicular stomatitis virus-based pseudovirion neutralization assay (PsVNA). Maximal neutralizing antibody responses were identified by traditional plaque reduction neutralization tests (PRNT) after four vaccinations. Neutralizing activity of human immunoglobulins (IgG) purified from TcB plasma collected after three vaccinations and injected intraperitoneally (IP) into mice at a 100 mg/kg dose was detected in the serum by PsVNA up to 14 days after administration. Passive transfer by IP injection of the purified IgG (100 mg/kg) to groups of BALB/c mice one day after IP challenge with mouse adapted (ma) EBOV resulted in 80% protection while all mice treated with non-specific pAbs succumbed. Similarly, interferon receptor 1 knockout (IFNAR -/-) mice receiving the purified IgG (100 mg/kg) by IP injection one day after IP challenge with wild type SUDV resulted in 89% survival. These results are the first to demonstrate that filovirus GP DNA vaccines administered to TcBs by IM-EP can elicit neutralizing antibodies that provide post-exposure protection. Additionally, these data describe production of fully human IgG in a large animal system, a system which is capable of producing large quantities of a clinical grade therapeutic product.

Development of a micro-neutralization assay for ebolaviruses using a replication-competent vesicular stomatitis hybrid virus and a quantitative PCR readout

“…4) agree with the values of 2.1 -2.8 mg/mL for EBOV and 2.5-11.3 mg/mL for SUDV obtained using a pseudotyped lentivirus luciferase assay with an IC80 endpoint [37]. However, both results are lower than those obtained with a PRNT 80 , where titers of 229-277 and 1108-1834 mg/mL were obtained for EBOV and SUDV, respectively [37]. Despite this difference in titers obtained from different assays, it is not yet clear which titer is clinically relevant.…”

“…We also measured IC80, 80% viral inhibition, titers of SAB132 and SAB139 to compare our results with the 80% neutralization titers obtained from other assays such as pseudotyped luciferase assay (PsVNA 80 ) and a PRNT performed in a BSL-4 laboratory (PRNT 80 ) [37,38]. The mean IC80 titers of SAB132 for VSV.EBOV.…”

“…The SAB132 and SAB139 antibodies and SAB negative control antibody have been described [37,38]. SAB132 was generated following four inoculations of DNA expression plasmids for codon-optimized GP genes from EBOV and SUDV into transchromosomal bovines [37].…”

“…These antibodies (SAB132 and SAB139) were generated in transchromosomal bovines following inoculation with different immunogens. SAB132 was obtained after inoculation with DNA vaccines expressing the GPs from EBOV and SUDV [37]. SAB139 was obtained following inoculation of purified GP viral-like particles from EBOV Makona [38].…”

Human polyclonal immunoglobulin G from transchromosomic bovines inhibits MERS-CoV in vivo

“…All samples were independently tested in duplicate or more (n ≥ 2). previously used with other viruses (32,34,35,41). Purified, diluted Tc hIgGs were added to Vero E6 cells, exposed to MERS-CoV Jordan strain, and assayed for ability to neutralize MERS-CoV.…”