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?:abstract
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Background To address the dire need for a safe and effective vaccine to protect individuals from and reduce transmission of SARS-CoV-2, we developed a COVID-19 vaccine that elicits not only robust humoral responses but also activates T cells. Our bivalent vaccine expresses both an optimized viral spike (S) protein (S-Fusion) and the viral nucleocapsid (N) protein with an Enhanced T-cell Stimulation Domain (N-ETSD) that directs N to the endo/lysosomal subcellular compartment to enhance MHC class II responses. The vaccine antigens are delivered by the second-generation adenovirus serotype 5 [E1-, E2b-, E3-] platform (hAd5) that has been safely administered and found to be effective in generating tumor-specific T cells even in the presence of pre-existing adenovirus immunity. Here, we report our findings on the safety and efficacy of our hAd5 S-Fusion + N-ETSD subcutaneous (SC) prime and thermally-stable oral boost vaccine in generating SARS-CoV-2-neutralizing antibodies, eliciting N- and S-specific T-cell responses, and providing complete protection with the clearing of virus after challenge in Non-Human Primates (NHP). A key objective of the study was to explore the efficacy of a novel thermally-stable oral hAd5 S-Fusion + N-ETSD to serve as a booster dose following an SC prime. Methods Group 1 NHP received the hAd5 S-Fusion + N-ETSD vaccine on Days 0 and 14 by SC injection (1011 VP), and on Day 28 by a single oral boost (1010 VP); Group 2 received vaccination on the same schedule, but with an SC prime and two oral boosts. Group 3 placebo NHP were dosed with vehicle-only SC-oral-oral. Blood for the isolation of sera and PBMCs was collected throughout the study. ELISA was used for determination of anti-S IgG levels, cPass™ for presence of neutralizing antibodies, and ELISpot for interferon-γ(IFN-γ) and interleukin-4 (IL-4) secretion by T cells. On Study Day 56, all NHP were challenged with intratracheal/intranasal 1 × 106 TCID50/mL SARS-CoV-2. Bronchoalveolar lavage (BAL) samples were collected on Day 42 pre-challenge and at several time points post-challenge; nasal swabs were collected daily post-challenge. NHP were euthanized on Study Day 70 and tissue collected for histopathological analyses. Viral load and active viral replication were determined in BAL and nasal swab specimens by RT qPCR of genomic and subgenomic RNA, respectively. Safety was determined by cage-side observations such as weight as well as hematology and clinical chemistry analyses of blood. Results The hAd5 S-Fusion + N-ETSD vaccine, both SC and oral, elicited no apparent toxicity seen in clinical chemistry, hematology, or cage-side observations. Neutralizing antibodies were induced in 9 of 10 vaccinated NHP and anti-S IgG positive titers in 10 out of 10. Th1 dominant T-cell responses were elicited by both S and N antigens, with responses being greater for N. Viral replication was inhibited from Day 1 post-SARS-CoV-2 challenge with complete protection in all (10/10) primates within 7 days of challenge from both nasal passages and lung. Replicating SARS-CoV-2 dropped immediately and was undetectable as soon 3 days post-challenge. There was a rapid decline in lymphocytes from the periphery on Day 1 post-challenge and a rebound within 3 days following challenge that was significantly higher by Day 14 post-challenge in Group 1 as compared to Group 3 placebo NHP. Conclusions In the rhesus macaque NHP model, the bivalent hAd5 S-Fusion + N-ETSD subcutaneous and oral vaccine provided complete protection of nasal passages and lung against SARS-CoV-2 challenge by eliciting neutralizing antibodies plus Th1 dominant N- and S-specific T-cell responses. Inhibition of viral replication within the first 24 hours post-challenge, in vaccinated NHP compared to placebo NHP, suggests the presence of SARS-CoV-2-specific cytotoxic T cells that rapidly cleared infected cells. The rapidity of clearance implies that shedding of live viruses may be attenuated as a result of vaccination and thus the vaccine has the potential to prevent transmission of virus by infected individuals. Clinical trials of hAd5 S-Fusion + N-ETSD are ongoing. The hAd5 S-Fusion + N-ETSD subcutaneous prime/boost vaccine has completed Phase 1 clinical trials and Phase 2/3 trials are actively recruiting. The thermally-stable oral vaccine will enter Phase 1 trials as a prime and boost, as well as explored to provide a boost to subcutaneous vaccination.
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