Hydrogel‐Based Slow Release of a Receptor‐Binding Domain Subunit Vaccine Elicits Neutralizing Antibody Responses Against SARS‐CoV‐2
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Abstract
The development of effective vaccines that can be rapidly manufactured and distributed worldwide is necessary to mitigate the devastating health and economic impacts of pandemics like COVID‐19. The receptor‐binding domain (RBD) of the SARS‐CoV‐2 spike protein, which mediates host cell entry of the virus, is an appealing antigen for subunit vaccines because it is efficient to manufacture, highly stable, and a target for neutralizing antibodies. Unfortunately, RBD is poorly immunogenic. While most subunit vaccines are commonly formulated with adjuvants to enhance their immunogenicity, clinically‐relevant adjuvants Alum, AddaVax, and CpG/Alum are found unable to elicit neutralizing responses following a prime‐boost immunization. Here, it has been shown that sustained delivery of an RBD subunit vaccine comprising CpG/Alum adjuvant in an injectable polymer‐nanoparticle (PNP) hydrogel elicited potent anti‐RBD and anti‐spike antibody titers, providing broader protection against SARS‐CoV‐2 variants of concern compared to bolus administration of the same vaccine and vaccines comprising other clinically‐relevant adjuvant systems. Notably, a SARS‐CoV‐2 spike‐pseudotyped lentivirus neutralization assay revealed that hydrogel‐based vaccines elicited potent neutralizing responses when bolus vaccines did not. Together, these results suggest that slow delivery of RBD subunit vaccines with PNP hydrogels can significantly enhance the immunogenicity of RBD and induce neutralizing humoral immunity.
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SciScore for 10.1101/2021.03.31.437792: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Institutional Review Board Statement not detected. Randomization not detected. Blinding not detected. Power Analysis not detected. Sex as a biological variable 8-10 week-old female mice were used. Cell Line Authentication not detected. Table 2: Resources
Antibodies Sentences Resources Mouse Serum ELISAs: Anti-RBD and Anti-spike trimer antibody titers were measured using an end-point ELISA. Anti-spikesuggested: NoneOne of the following goat-anti-mouse secondary antibodies was used: IgG Fc-HRP (1:10,000, Invitrogen A16084) secondarysuggested: (Thermo Fisher Scientific Cat# A16084, RRID:AB_2534758)For flow cytometry analysis, cells were blocked with anti-CD16/CD38 (clone: 2.4G2) and then stained with fluorochrome … SciScore for 10.1101/2021.03.31.437792: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Institutional Review Board Statement not detected. Randomization not detected. Blinding not detected. Power Analysis not detected. Sex as a biological variable 8-10 week-old female mice were used. Cell Line Authentication not detected. Table 2: Resources
Antibodies Sentences Resources Mouse Serum ELISAs: Anti-RBD and Anti-spike trimer antibody titers were measured using an end-point ELISA. Anti-spikesuggested: NoneOne of the following goat-anti-mouse secondary antibodies was used: IgG Fc-HRP (1:10,000, Invitrogen A16084) secondarysuggested: (Thermo Fisher Scientific Cat# A16084, RRID:AB_2534758)For flow cytometry analysis, cells were blocked with anti-CD16/CD38 (clone: 2.4G2) and then stained with fluorochrome conjugated antibodies: CD19, GL7, CD95, CXCR4, CD86, IgG1, CD4, CXCR5, and PD1. anti-CD16/CD38suggested: NoneCD19suggested: (Diaclone Cat# 873.039.050, RRID:AB_1587088)GL7suggested: NoneCD95suggested: NoneCXCR4suggested: NoneCD86suggested: NoneIgG1suggested: NoneCD4suggested: NoneCXCR5suggested: NonePD1suggested: NoneExperimental Models: Cell Lines Sentences Resources Viral Neutralization Assay: Neutralization assays were conducted as described previously.39 Briefly, SARS-CoV-2 spike-pseudotyped lentivirus was produced in HEK239T cells. HEK239Tsuggested: NoneFor the neutralization assay, ACE2/HeLa cells were plated 1-2 days prior to infection. ACE2/HeLasuggested: NoneExperimental Models: Organisms/Strains Sentences Resources Mice and Vaccination: C57BL/6 mice were purchased from Charles River and housed at Stanford University. C57BL/6suggested: NoneSoftware and Algorithms Sentences Resources For RBD retention, the points were fit with a linear fit in GraphPad Prism and the half-life of release was determined (n=3). GraphPadsuggested: (GraphPad Prism, RRID:SCR_002798)Data were analyzed with FlowJo 10 (FlowJo LLC). FlowJosuggested: (FlowJo, RRID:SCR_008520)P values listed were determined in GraphPad Prism software using a one-way ANOVA with Tukey’s multiple comparison test. GraphPad Prismsuggested: (GraphPad Prism, RRID:SCR_002798)Results from OddPub: We did not detect open data. We also did not detect open code. Researchers are encouraged to share open data when possible (see Nature blog).
Results from LimitationRecognizer: We detected the following sentences addressing limitations in the study:By combining the promising antigen, RBD, and well-studied adjuvants, CpG and Alum, with our hydrogel platform, we have created a vaccine that overcomes many of the current limitations and is suitable for broader distribution. In addition to the choice of antigen and adjuvant, there is a growing need for novel and efficient drug delivery systems like our PNP hydrogel platform.45 The hydrogel is injectable and forms a depot that allows for slow release of the vaccine over several days to weeks, which leads to greater affinity maturation and a more potent, durable humoral response.24 This is critical for eliminating the need for a booster shot, in-turn increasing the feasibility of vaccinating billions of people across the globe. In this work we validated that, unlike other hydrogel or microneedle vaccine platforms, the PNP hydrogel is readily loaded with diverse molecular cargo, allowing for the antigen and adjuvants to be co-presented to immune cells. As discussed in previous work from our lab, the hydrogel likely acts through two main mechanisms: as a local stimulatory niche where infiltrating cells experience high local concentrations of adjuvant and antigen and as a slow release depot that allows for prolonged vaccine exposure.24 By delivering RBD with the clinically de-risked adjuvants, CpG and Alum, in our PNP hydrogel, we were able to achieve titers and neutralization levels that greatly exceeded those of the relevant clinical controls, Alum and MF59, as well as the bolu...
Results from TrialIdentifier: No clinical trial numbers were referenced.
Results from Barzooka: We did not find any issues relating to the usage of bar graphs.
Results from JetFighter: We did not find any issues relating to colormaps.
Results from rtransparent:- Thank you for including a conflict of interest statement. Authors are encouraged to include this statement when submitting to a journal.
- Thank you for including a funding statement. Authors are encouraged to include this statement when submitting to a journal.
- No protocol registration statement was detected.
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