Phase I study of a SARS-CoV-2 mRNA vaccine PTX-COVID19-B (preprint)

PTX-COVID19-B mRNA vaccine encodes for SARS-CoV-2 Spike protein G614 variant and lacks the proline-proline (986-987 position) mutation present in other COVID-19 vaccines. This Phase 1 observer-blinded, randomized, placebo-controlled, ascending dose study evaluated the safety, tolerability, and immunogenicity of two doses of PTX-COVID19-B vaccine in healthy seronegative adults. Participants received two intramuscular doses, 4 weeks apart, of 16-g, 40-g, or 100-g PTX-COVID19-B. Adverse events were generally mild to moderate, self-resolving, and transient. The most common solicited local and systemic adverse event was pain at the injection site and headache, respectively. After the first immunization, all participants seroconverted, producing high titers of anti-receptor-binding-domain, anti-Spike, and neutralizing antibodies, including neutralizing antibodies against the ancestral viral strain and the Alpha, Beta, and Delta variants of concern, in a dose-dependent way, further increasing over 10-20 times after the second dose. All tested doses of PTX-COVID19-B were safe, well-tolerated, and provided a strong immunogenicity response. The 40-g dose showed fewer adverse reactions than the 100-g dose, supporting further investigation of the 40-g dose. Clinical Trial Registration: ClinicalTrials.gov identifier: NCT04765436 (https://clinicaltrials.gov/ct2/show/NCT04765436)

Antibody and T cell responses to SARS-CoV-2 mRNA vaccines during maintenance therapy for immune-mediated inflammatory diseases (preprint)

Background Limited information is available on the impact of immunosuppressants on COVID-19 vaccination in patients with immune-mediated inflammatory diseases (IMID). This study investigated antibody and T cell responses to SARS-CoV-2 mRNA vaccines in IMID patients undergoing immunomodulatory maintenance therapy. Methods This observational cohort study examined the immunogenicity of SARS-CoV-2 mRNA vaccines in adult patients with inflammatory bowel disease, rheumatoid arthritis, ankylosing spondylitis or psoriatic disease, with or without maintenance immunosuppressive therapies. T cell and antibody responses to SARS-COV-2, including neutralization against SARS-CoV-2 variants were determined pre-vaccination and after 1 and 2 vaccine doses. Findings We prospectively followed 150 subjects, 26 healthy controls, 9 IMID patients on no treatment, 44 on anti-TNF, 16 on anti-TNF with methotrexate/azathioprine (MTX/AZA), 10 on anti-IL-23, 28 on anti-IL-12/23, 9 on anti-IL-17, and 8 on MTX/AZA. Most patients showed increased antibody responses from dose 1 to dose 2, with decreases apparent by 3 months post dose 2, albeit with considerable variability within groups. Overall, T cell responses were not consistently different between groups; however, antibody levels and neutralization efficacy in the anti-TNF treated group was lower than controls and waned substantially by 3 months post-dose 2. Implications These findings support the need for a third dose of mRNA vaccine and for continued monitoring of immunity over time.

Neutralizing antibody responses to SARS-CoV-2 variants in vaccinated Ontario long-term care home residents and workers (preprint)

Prioritizing Ontarios long-term care home (LTCH) residents for vaccination against severe acute respiratory syndrome coronavirus 2 has drastically reduced their disease burden; however, recent LTCH outbreaks of variants of concern (VOCs) have raised questions regarding their immune responses. In 198 residents, mRNA vaccine dose 1 elicited partial spike and receptor binding domain antibody responses, while the second elicited a response at least equivalent to convalescent individuals in most residents. Residents administered mRNA-1273 (Moderna) mounted stronger total and neutralizing antibody responses than those administered BNT162b2 (Pfizer-BioNTech). Two to four weeks after dose 2, residents (n = 119, median age 88) produced 4.8-6.3-fold fewer neutralizing antibodies than staff (n = 78; median age 47) against wild-type (with D614G) pseudotyped lentivirus, and residents administered BNT162b2 produced 3.89-fold fewer neutralizing antibodies than those who received mRNA-1273. These effects were exacerbated upon serum challenge with pseudotyped VOC spike, with up to 7.94-fold reductions in B.1.351 (Beta) neutralization. Cumulatively, weaker vaccine stimulation, age/comorbidities, and the VOC produced an [~]130-fold reduction in apparent neutralization titers in LTCH residents and 37.9% of BNT162b2-vaccinated residents had undetectable neutralizing antibodies to B.1.351. Continued immune response surveillance and additional vaccine doses may be required in this population with known vulnerabilities.

Preclinical evaluation of a SARS-CoV-2 mRNA vaccine PTX-COVID19-B (preprint)

Safe and effective vaccines are needed to end the COVID-19 pandemic caused by SARS-CoV-2. Here we report the preclinical development of a lipid nanoparticle (LNP) formulated SARS-CoV-2 mRNA vaccine, PTX-COVID19-B. PTX-COVID19-B was chosen among three candidates after the initial mouse vaccination results showed that it elicited the strongest neutralizing antibody response against SARS-CoV-2. Further tests in mice and hamsters indicated that PTX-COVID19-B induced robust humoral and cellular immune responses and completely protected the vaccinated animals from SARS-CoV-2 infection in the lung. Studies in hamsters also showed that PTX-COVID19-B protected the upper respiratory tract from SARS-CoV-2 infection. Mouse immune sera elicited by PTX-COVID19-B vaccination were able to neutralize SARS-CoV-2 variants of concern (VOCs), including the B.1.1.7, B.1.351 and P.1 lineages. No adverse effects were induced by PTX-COVID19-B in both mice and hamsters. These preclinical results indicate that PTX-COVID19-B is safe and effective. Based on these results, PTX-COVID19-B was authorized by Health Canada to enter clinical trials in December 2020 with a phase 1 clinical trial ongoing (ClinicalTrials.gov number: NCT04765436).

A SARS-CoV-2 - host proximity interactome (preprint)

Viral replication is dependent on interactions between viral polypeptides and host proteins. Identifying virus-host protein interactions can thus uncover unique opportunities for interfering with the virus life cycle via novel drug compounds or drug repurposing. Importantly, many viral-host protein interactions take place at intracellular membranes and poorly soluble organelles, which are difficult to profile using classical biochemical purification approaches. Applying proximity-dependent biotinylation (BioID) with the fast-acting miniTurbo enzyme to 27 SARS-CoV-2 proteins in a lung adenocarcinoma cell line (A549), we detected 7810 proximity interactions (7382 of which are new for SARS-CoV-2) with 2242 host proteins (results available at covid19interactome.org). These results complement and dramatically expand upon recent affinity purification-based studies identifying stable host-virus protein complexes, and offer an unparalleled view of membrane-associated processes critical for viral production. Host cell organellar markers were also subjected to BioID in parallel, allowing us to propose modes of action for several viral proteins in the context of host proteome remodelling. In summary, our dataset identifies numerous high confidence proximity partners for SARS-CoV-2 viral proteins, and describes potential mechanisms for their effects on specific host cell functions.

Nanoluciferase complementation-based biosensor reveals the importance of N- linked glycosylation of SARS-CoV-2 Spike for viral entry (preprint)

The ongoing COVID-19 pandemic has highlighted the` immediate need for the development of antiviral therapeutics targeting different stages of the SARS-CoV-2 lifecycle. We developed a bioluminescence-based biosensor to interrogate the interaction between the SARS-CoV-2 viral spike protein and its host entry receptor, angiotensin-converting enzyme 2 (ACE2). The biosensor assay is based on a Nanoluciferase complementation reporter, composed of two subunits, Large BiT and Small BiT, fused to the spike receptor-binding domain (RBD) of the SARS-CoV-2 S protein and ACE2 ectodomain, respectively. Using this biosensor, we uncovered a critical role for glycosylation of asparagine residues within the RBD in mediating successful binding to the cellular ACE2 receptor and subsequent virus infection. Our findings support RBD glycosylation as a therapeutic and vaccine target to blunt SARSCoV- 2 infections.

A simple protein-based SARS-CoV-2 surrogate neutralization assay (preprint)

With the COVID-19 pandemic surpassing 12M confirmed cases and 550K deaths worldwide, defining the key components of the immune response to SARS-CoV-2 infection is critical. Of particular importance is the identification of immune correlates of infection that would support public health decision-making on treatment approaches, vaccination strategies, and convalescent plasma therapy. While ELISA-based assays to detect and quantitate antibodies to SARS-CoV-2 in patient samples have been developed, the detection of neutralizing antibodies typically requires more demanding cell-based viral assays. Here, we present and validate a safe and efficient protein-based assay for the detection of serum and plasma antibodies that block the interaction of the SARS-CoV-2 spike (S) protein receptor binding domain (RBD) with its receptor, angiotensin converting-enzyme 2 (ACE2). This test is performed on the same platform and in parallel with an enzyme-linked immunosorbent assay (ELISA) for the detection of antibodies against the RBD and serves as a surrogate neutralization assay.