*Important notice: bioRxiv publishes preliminary scientific reports that are not peer-reviewed and, therefore, should not be regarded as conclusive, guide clinical practice/health-related behavior, or treated as established information.
In a recent study posted to the bioRxiv* preprint server, researchers assessed the effectiveness of a multivalent exosome-based coronavirus disease 2019 (COVID-19) vaccine.
Study: Exosome based multivalent vaccine: achieving potent immunization, broaden reactivity and T cell response with nanograms of proteins without any adjuvant. Image Credit: Ground Picture/Shutterstock
Background
Despite the evident usefulness of COVID-19 messenger ribonucleic acid (mRNA)-based vaccines, studies have demonstrated that these vaccines did not provide long-term protection, perhaps because of a lack of cross-reactivity against new variants of concern (VOCs). This necessitated periodic booster injections to ensure long-term protection against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2).
Thus, there is an urgent need to create novel COVID-19 vaccination methods. An option could involve the SARS-CoV-2 spike antigen (StealthTM X-Spike, STX-S) combined with a more conserved SARS-CoV-2 antigen, like the SARS-CoV-2 nucleocapsid (STX-N), in a "mix and match" strategy. This strategy would extend the immune response at the humoral and T cell levels, resulting in enhanced protection against the virus's present and future forms.
About the study
In the present study, researchers employed exosomes to develop a "cocktail" protein-based vaccination involving two distinct viral proteins delivered via the exosome membrane.
By lentiviral transduction, STX cells were produced, and the surface expression of SARS-CoV-2 proteins was assessed using flow cytometry. Utilizing Capricor's labscale purification method, STX exosomes were isolated from the modified 293F cell culture supernatant. Transmission electron microscopy (TEM) imaging was used to assess STX exosomes. Furthermore, enzyme-linked immunosorbent assay (ELISA) was used to determine the concentrations of the SARS-CoV-2 nucleocapsid (Ncap) antigen in STX-N exosomes and SARS-CoV-2 spike antigen in STX-S exosomes.
To confirm the capacity of STX-N and STX-S exosomes to generate an immunological response, the team injected the spike and nucleocapsid antigens along with 10ng formulations of STX-N and STX-S exosomes into mice. To demonstrate the effectiveness of exosome delivery, either Ncap or spike recombinant protein was conjugated with adjuvant and delivered.
The immunological reaction of a multivalent vaccine elicited by combining STX-N and STX-S exosomes was then evaluated. Mice received three different dosages of STX-S+N via two intramuscular (i.m.) injections. After three weeks, a second i.m. injection, the boost injection, was administered. In the study, phosphate-buffered saline (PBS) was employed as a negative control. Evaluation of immunization with the STX-S+N vaccine was performed by quantifying antibodies generated against spike and Ncap. Additionally, antigen-specific T cell responses were evaluated by ELISpot to assess the T cell response towards STX-S+N.
Results
In comparison to parental 293F cells, the expression of SARS-CoV-2 Ncap and the spike was over 95% more than that in STX cells. The estimated average diameter of the purified STX-N and STX-S exosomes was 140.4 nm and 144.6 nm, respectively. The expected polydispersity index (PDI) was less than 0.2.
Nanoparticles with a visible lipid bilayer and a characteristic exosome size and shape were detected. These nanoparticles were spherical, smooth, and contained a lipid bilayer. Significantly, the presence of spike protrusions on the STX-S nanoparticle surface indicated the detection of the spike protein on the exosomes. Nucleocapsid had a distinctive lipid bilayer thicker than naive exosomes, indicating the accumulation of particles within the exosome membrane.
Spike protein was found in STX-S cells and exosomes, with exosome samples containing a higher concentration of spike protein. Nucleocapsid levels were high in exosomes and cells generated from STX-N. Using a bead-based CD81 test, SARS-CoV-2 protein Ncap and spike were identified on the exosome membrane, with over 75% expression observed in conjunction with the exosome-specific marker CD81.
The production of antibodies increased after the initial injection and rose consistently after the booster dose. A single dose of STX-S+N increased immunoglobulin (Ig)-G levels against a spike by up to 30 times, with no remarkable difference among the three doses. After full vaccination, doses 1 and 2 increased antibodies against Spike by a factor of 1,500, but dose 3 increased antibodies by a factor of 280. In contrast, a dose-response relationship was reported for Ncap. A 1.5-fold rise was noted for dose 1, an approximately four-fold increase for dose 2, and an almost seven-fold increase for dose 3. Mice treated with STX-S+N exhibited a 24- to 43-fold rise in IgG against Ncap following the completion of the vaccination cycle, with no significant differences between doses.
STX-S+N vaccination produced antigen-specific and multifunctional T-cell responses. Although baseline interferon (IFN)-Ɣ response was similar between cohorts, examination of IFN-Ɣ-secreting cells towards ex vivo stimulation with Spike or Ncap revealed a significant increase in STX-S+N vaccine-immunized spleens, indicating a Th1-biased CD8+ T cell response. Despite the STX-S+N vaccination, an average seven-fold rise in IFN-Ɣ response was found following spike stimulation. A dose-response trend was observed following Ncap stimulation, with a seven-fold rise in mice receiving the lowest dose of Ncap and a three-fold increase in animals receiving either doses 2 or 3.
Overall, the study findings showed that a single dose of dual-antigen vaccination was effective against various SARS-CoV-2 VOCs, possessed a larger immunological capacity, and may be used to augment the protection established by previously licensed vaccines. The StealthXTM vaccine technology employed in the study elicited a robust, wider immune response without the need for adjuvants. The researchers believe this study could facilitate the study of the next generation of vaccines along with the capacity for multiplexing and rapid turnaround.
*Important notice: bioRxiv publishes preliminary scientific reports that are not peer-reviewed and, therefore, should not be regarded as conclusive, guide clinical practice/health-related behavior, or treated as established information.
Journal reference:
- Preliminary scientific report.
Mafalda Cacciottolo, Justin B Nice, Yujia Li, Michael J LeClaire, Ryan Twaddle, Ciana L. Mora, Stephanie Y. Adachi, Esther R. Chin, Meredith Young, Jenna Angeles, Kristi Elliott, Minghao Sun. (2023). Exosome based multivalent vaccine: achieving potent immunization, broaden reactivity and T cell response with nanograms of proteins without any adjuvant. bioRxiv. doi: https://doi.org/10.1101/2023.01.10.523356 https://www.biorxiv.org/content/10.1101/2023.01.10.523356v1
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