Mechanism of a COVID-19 nanoparticle vaccine candidate that elicits a broadly neutralizing antibody response to SARS-CoV-2 variants

TL;DR: In this paper, a mouse plasma induced by protein nanoparticles that present rationally designed S2GΔHR2 spikes can neutralize the B.1.7, B.2.1, and P.351 variants with comparable titers.

Abstract: Vaccines against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are essential for combating the coronavirus disease 2019 (COVID-19) pandemic. Neutralizing antibody responses to the original Wuhan-Hu-1 strain that were generated during infection and vaccination showed lower effectiveness against variants of concern. Here, we demonstrated that mouse plasma induced by protein nanoparticles that present rationally designed S2GΔHR2 spikes can neutralize the B.1.1.7, B.1.351, and P.1 variants with comparable titers. The mechanism of nanoparticle vaccine-induced immunity was examined in mice for an I3-01v9 60-mer that presents 20 stabilized spikes. Compared with the soluble spike, this nanoparticle showed 6-fold longer retention, 4-fold greater presentation on follicular dendritic cell dendrites, and 5-fold higher germinal center reactions in lymph node follicles. Intact nanoparticles in lymph node tissues were visualized by transmission electron microscopy. In conclusion, spike-presenting protein nanoparticles that induce robust long-lived germinal centers may provide a vaccine solution for emerging SARS-CoV-2 variants. ONE-SENTENCE SUMMARY With prolonged lymph node retention and robust germinal centers, nanoparticles elicit neutralizing antibodies to diverse SARS-CoV-2 variants.

Summary

INTRODUCTION

• The COVID-19 pandemic has led to more than 188 million infection cases and 4 million deaths globally.
• As previously reported (34) (35) (36) (37) (38) , the production of a bNAb response relies on long-lived germinal center (GC) reactions to activate precursor B cells, stimulate affinity maturation, and form long-term immune memory.
• When a different injection route was tested in mouse immunization, E2p and I3-01v9 SApNPs sustained neutralizing titers against the four variants, even at a low dosage of 3.3 μg, whereas a significant reduction of plasma neutralization was observed for the soluble spike.
• The authors study thus demonstrates that a spike-presenting SApNP vaccine derived from the "ancestral" SARS-CoV-2 strain may confer broad protection against emerging variants.

Spike and SApNP vaccine-induced neutralizing responses to SARS-CoV-2 variants.

• The authors previously demonstrated that the rationally designed S2GΔHR2 spike was more immunogenic than the S2P spike (42) , and SApNPs displaying 8-20 spikes outperformed soluble spikes in NAb elicitation (41) (Fig. 1A ).
• The authors first assessed the neutralizing activity of polyclonal plasma induced by various spike and SApNP vaccine formulations from their previous study (41) against the wildtype SARS-CoV-2 strain, Wuhan-Hu-1, as a baseline for comparison (Fig. 1B ).
• Mouse plasma collected at week 5 after two intraperitoneal (i.p.) injections of adjuvanted vaccine antigens (50 μg) was analyzed in pseudoparticle (pp) neutralization assays (43) .
• All three spike-presenting SApNPs elicited superior neutralizing responses than the soluble S2P ECTO spike (41) .
• Overall, the E2p and I3-01v9 SApNP groups exhibited similar or slightly stronger plasma neutralization against the four variants relative to the wildtype strain, Wuhan-Hu-1 (Fig. 1E ).

Adjuvant effect on vaccine-induced neutralizing antibody and T-cell responses.

• Innate immunity plays an important role in regulating adaptive immunity, including humoral and cellular immune responses (47) (48) (49) .
• In most cases, adjuvants combined with AP further boosted plasma neutralizing activity.
• Compared with the non-adjuvanted control, STING and CpG (TLR9) induced 3.7 and 5.5-fold more IFN-γ-producing CD4 +.
• The authors results demonstrate that the I3-01v9 SApNP itself is immunogenic, and adjuvants can further enhance vaccine-induced NAb responses in plasma by up to 35-fold.

Diverse variant-neutralizing mouse antibody lineages identified by single-cell analysis.

• The nature of this response was unclear.
• The authors then examined the biological function of these mouse mAbs.
• By comparison, fewer putative somatic variants were identified for other NAbs in the antigen-specific B-cell repertoires regardless of the sorting probe used (fig. S4D-F ), suggesting that these NAbs either were from less prevalent lineages or were generated in response to a previous injection (each mouse received four doses) (41) .
• Single-cell isolation identified a panel of mouse mAbs with different neutralization breadth and potency against the wildtype SARS-CoV-2 strain and four major variants.

Distribution and trafficking of I3-01v9 SApNP in mouse lymph node.

• After validating these vaccines against variants at both the plasma and mAb levels, the authors studied in vivo behaviors of the S2GΔHR2 spike and two large 60-meric SApNPs to understand why SApNPs outperform soluble spikes in bNAb elicitation.
• The axillary, brachial, and popliteal sentinel lymph nodes were isolated for histological analysis.
• Consistent with their previous study (73) , SApNPs accumulated in lymph node follicles, regardless of the number of doses.
• The injection dose was normalized to the total amount of protein (10 μg) per injection into each footpad (40 μg/mouse).
• Similar patterns of antigen retention were observed after the second injection, although the boost appeared to exert a more positive effect on the soluble spike, which could be detected in lymph node follicles at 48 h (Fig. 4D ).

Retention and presentation of I3-01v9 SApNP on follicular dendritic cell dendrites.

• Antigen retention and presentation in lymph node follicles are prerequisites to the stimulation of robust B cell responses and GC reactions (34, 36) .
• This result confirmed the critical role of FDC networks in mediating vaccine retention in lymph node follicles.
• The processed tissue samples were sectioned and stained on copper grids for TEM analysis.
• The TEM images indicated that FDCs can present many SApNPs to neighboring B cells in this "hugging mode", in which their long dendrites brace B cells to maximize interactions between multivalently displayed spikes and B cell receptors.

Robust germinal center reactions induced by spike-presenting SApNPs.

• Long-lived GC reactions induce immune stimulation for B-cell selection and affinity maturation, as well as production of immune memory and bNAb responses (34, 35, 40) .
• Two metrics, the GC/FDC ratio (i.e., whether GC formation is associated with an FDC network, %) and GC size (i.e., occupied area), were used.
• Single-cell immune profiling and antibody isolation (89) may provide further insights into the clonality of vaccineinduced B-cell lineages within lymph nodes.

DISCUSSION

• To end the COVID-19 pandemic, vaccines need to effectively block current and emerging SARS-CoV-2 variants that evade NAb responses by mutating key epitopes on the viral spike (31) .
• Based on these findings, the authors hypothesized that SApNPs presenting stabilized ancestral Wuhan-Hu-1 spikes may provide an effective vaccine against SARS-CoV-2 variants.
• The authors results revealed that a plethora of NAb lineages were generated upon vaccination, with I3-01v9 SApNP being the most effective at eliciting bNAbs.
• Protein vaccines, despite the well-established records of safety and effectiveness, have yet to be deployed to mitigate the COVID-19 pandemic (93) (94) (95) .
• Indeed, the authors found that the I3-01v9 SApNP, their leading vaccine candidate (41), elicited 6-fold longer retention and 4-fold greater accumulation in lymph node follicles than the stabilized S2GΔHR2 spike alone with a prime-boost regimen.

Rational design of next-generation COVID-19 vaccines requires an in-depth

• The I3-01v9 SApNP elicited a potent bNAb response to four variants, overcoming a major challenge facing the current COVID-19 vaccines.
• Mechanistic studies of vaccine trafficking, retention, presentation, and GC reactions provided valuable insights into the spike and SApNPinduced immunity (95, 105, 106) .
• Such knowledge, if can be obtained for other vaccine platforms (e.g., inactivated whole virions, mRNAs, and viral vectors) will facilitate rational selection of the most effective vaccine candidates to mitigate the pandemic and ultimately stop the spread of SARS-CoV-2.

SARS-CoV-2 spike and SApNP vaccine antigens

• The design, expression, and purification of a stabilized SARS-CoV-2 spike, S2GΔHR2, and three SApNPs that present either 8 or 20 S2GΔHR2 spikes were described in their recent study (41) .
• The ExpiFectamine TM CHO/plasmid DNA complexes were prepared for 100-ml transfection in ExpiCHO cells according to the manufacturer's instructions.
• Protein concentration was determined using UV 280 absorbance with theoretical extinction coefficients.

Animal immunization and sample collection

• Similar immunization protocols were reported in their previous vaccine studies (41, 86, 87) .
• The mouse studies were conducted according to Association for the Assessment and Accreditation of Laboratory Animal Care guidelines, and the protocols were approved by the IACUC.
• Vaccines were intradermally administered into mouse footpads using a 29-gauge insulin needle under 3% isoflurane anesthesia with oxygen.
• Plasma was isolated from blood after centrifugation at 14000 rpm for 10 min.
• The axillary, brachial, and popliteal sentinel lymph nodes were collected at the end timepoint for further analysis.

Experimental adjuvants and formulation

• The adjuvants squalene-oil-in-water , aluminum hydroxide (AH), aluminum phosphate (AP), 2'3'-c-di-AM(PS)2 (Rp,Rp) (STING ligand), monophosphoryl lipid A from S. minnesota (MPLA-SM) R595 (TLR4 agonist), imidazoquinoline compound R848 (TLR7/8 agonist), and CpG ODN 1826, Class B (TLR9 agonist) were purchased from InvivoGen.
• PIKA, a TLR3 agonist with enhanced T cell and antibody responses reported for a Phase I rabies vaccine trial (107) , was used as an adjuvant.
• Macrophage inhibitors clodronate liposomes (Liposoma BV, catalog no. CP-005-005) were used to eliminate subcapsular sinus macrophages in lymph nodes to promote more robust B-cell activation.
• Each mouse was intradermally immunized at weeks 0, 3, and 6 with 120-140 μl of antigen/adjuvant mix containing 20 μg of vaccine antigen (I3-01v9 SApNP) and 80-100 μl of adjuvant, which was evenly split and injected into four footpads.
• Splenocytes were spun down at 400 × g for 10 min, washed with PBS, and treated with the ammonium-chloride-potassium (ACK) lysing buffer .

SARS-CoV-2 pseudovirus neutralization assay

• The plates were incubated overnight at 4 °C, and then washed five times with wash buffer containing PBS and 0.05% (v/v) Tween 20.
• Each well was then coated with 150 µl of blocking buffer consisting of PBS and 40 mg/ml blotting-grade blocker (Bio-Rad).
• The plates were incubated with blocking buffer for 1 h at room temperature, and then washed five times with wash buffer.
• The plates were incubated with the secondary antibody for 1 h at room temperature and then washed six times with PBS containing 0.05% Tween 20.
• The resulting plate readouts were measured at a wavelength of 450 nm.

Histology, immunostaining, and imaging

• The axillary, brachial, and popliteal sentinel lymph nodes were isolated for histological analysis.
• For immunofluorescent staining, tissue sections were stained for FDCs using anti-CD21 antibody (Abcam, catalog no. ab75985, 1:1800) followed by anti-rabbit secondary antibody conjugated with Alexa Fluor 555 (Thermo Fisher, catalog no.

Electron microscopy analysis of protein nanoparticles and lymph node tissues

• Electron microscopy (EM) analysis was performed by the Core Microscopy Facility at The Scripps Research Institute.
• For the negative-staining EM analysis of protein nanoparticles, the S2GΔHR2-10GS-I3-01v9-L7P SApNP samples were prepared at a concentration of 0.01 mg/ml.
• Carbon-coated copper grids (400 mesh) were glow-discharged, and 10 µl of each sample was adsorbed for 2 min.
• Excess sample was wicked away and grids were negatively stained with 2% uranyl formate for 2 min.
• The tissue samples were washed in double-distilled H 2 O and dehydrated through a graded series of ethanol followed by acetone, infiltrated with LX-112 (Ladd) epoxy resin, and polymerized at 60°C.

Lymph node disaggregation, cell staining, and flow cytometry

• The mice were sacrificed 2, 5, and 8 weeks after a single-dose immunization and 2 and 5 weeks after the boost immunization.
• Fresh axillary, brachial, and popliteal sentinel lymph nodes were collected and mechanically disaggregated.
• The cell samples were stored in HBSS blocking solution for the flow cytometry study.
• Sample events were acquired by a 5-laser BD Biosciences LSR II analytical flow cytometer with BD FACS Diva 6 software at the Core Facility of The Scripps Research Institute.

DC production, T cell culture, activation, and flow cytometry analysis

• Mouse bone marrow (BM) was cultured in RPMI 1640 medium containing 10% fetal bovine serum (FBS) and recombinant mouse Fms-like tyrosine kinase 3 ligand (Flt3L, 50 ng/ml) and stem cell factor (SCF, 10 ng/ml) for 9 days as previously described (109) .
• To induce DC activation, immature DCs were incubated with lipopolysaccharide (LPS, 100 ng/ml) plus R848 (Resiquimod, 100 ng/ml) overnight, which activated TLR4 or TLR7/8 signaling, respectively.
• CD11c + DCs were sorted using magnetic beads (Miltenyi-Biotech, CA).
• Recombinant mouse Flt3 ligand (Flt3L) and mouse SCF were purchased from Shenandoah Biotech (Warwick, PA).
• Cells were stained with appropriate concentrations of mAbs.

Bulk and single-cell sorting of SARS-CoV-2 antigen-specific mouse B cells.

• Spleens or lymph nodes were harvested from mice 15 days after the last immunization, and the cell suspension was prepared.
• Dead cells were excluded by staining with the Fixable Aqua Dead Cell Stain kit (Thermo Fisher, catalog no. L34957).
• In the SEC profiles, the probe peak was well separated from the peak of biotin ligase (fig. S3A ).
• Cells and biotinylated proteins were incubated for 5 min at 4 °C, followed by the addition of 2.5 µl of anti-mouse IgG fluorescently labeled with FITC (Jackson ImmunoResearch catalog no.
• For bulk sorting, positive cells were sorted into an Eppendorf microtube with 20 μl of lysis buffer.

Antibody cloning from Env-specific single B cells and antibody production.

• The antibody cloning of SARS-CoV2-2 antigen-sorted single B cells was conducted as follows.
• A second PCR reaction was then performed using 5 μl of the first PCR as the template and respective mouse primers (110) according to the same recipe as the first PCR.
• The PCR products were run on 1% Agarose gel and those with correct heavy and light chain bands were then used for Gibson ligation (New England Biolabs), cloning into human IgG expression vectors, and transformation into competent cells.
• Mouse mAbs were expressed by the transient transfection of ExpiCHO cells (Thermo Fisher) with equal amounts of paired heavy and κ-light chain plasmids.
• Antibody proteins were purified from the culture supernatant after 12-14 days using Protein A bead columns (Thermo Fisher).

NGS and bioinformatics analysis of mouse B cells.

• Previously, a 5′-rapid amplification of cDNA ends (RACE)-PCR protocol was developed for the deep sequencing analysis of mouse B-cell repertoires (69) .
• Briefly, 5´-RACE cDNA was obtained from bulk-sorted B cells of each mouse with the SMART-Seq v4 Ultra Low Input RNA Kit for Sequencing .
• The 5′-RACE primer contained a PGM/S5 P1 adaptor, and the reverse primer contained a PGM/S5 A adaptor.
• A total of 25 cycles of PCR was performed and the expected PCR products (500-600 bp) were gel purified .
• NGS was performed on the Ion S5 GeneStudio system.

Statistical analysis

• Data were collected from 4-7 mice per group.
• All of the statistical analyses were performed and graphs were generated using GraphPad Prism 9.1.2 software.
• In the analysis of vaccine-induced plasma neutralization, different vaccine groups were compared using one-way analysis of variance , whereas for a given vaccine group ID 50 titers of the same plasma sample against different variants were compared using repeated measures one-way .
• In both cases, they were followed by Dunnett's multiple comparison post hoc test.
• For the vaccine accumulation and GC study, different vaccine groups were compared using one-way ANOVA, followed by Tukey's multiple comparison post hoc test.

Full text

1
Mechanism of a COVID-19 nanoparticle vaccine candidate that elicits a broadly 1
neutralizing antibody response to SARS-CoV-2 variants 2
3
Yi-Nan Zhang
1
, Jennifer Paynter
1
, Cindy Sou
1
, Tatiana Fourfouris
1
, Ying Wang
3,4
, Ciril 4
Abraham
3
, Timothy Ngo
1
, Yi Zhang
3,4
, Linling He
1
, and Jiang Zhu
1, 2,
* 5
6
1
Department of Integrative Structural and Computational Biology,
2
Department of Immunology 7
and Microbiology, The Scripps Research Institute, La Jolla, California 92037, USA
8
3
Fels Institute for Cancer Research and Molecular Biology, and
4
Department of Microbiology 9
and Immunology, Temple University, Philadelphia, Pennsylvania 19140, USA. 10
11
*Corresponding author 12
JZ: Phone +1 (858) 784-8157; Email: jiang@scripps.edu
13
14
KEYWORDS 15
Ancestral strain; broadly neutralizing antibody (bNAb); coronavirus disease 2019 (COVID-19); 16
severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2); self-assembling protein 17
nanoparticle (SApNP); vaccine; variant of concern (VOC). 18
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2
ABSTRACT (150 words) 19
Vaccines that induce potent neutralizing antibody (NAb) responses against emerging variants of 20
severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are essential for combating the 21
coronavirus disease 2019 (COVID-19) pandemic. We demonstrated that mouse plasma induced 22
by self-assembling protein nanoparticles (SApNPs) that present 20 rationally designed 23
S2GΔHR2 spikes of the ancestral Wuhan-Hu-1 strain can neutralize the B.1.1.7, B.1.351, P.1, 24
and B.1.617 variants with the same potency. The adjuvant effect on vaccine-induced immunity 25
was investigated by testing 16 formulations for the multilayered I3-01v9 SApNP. Using single-26
cell sorting, monoclonal antibodies (mAbs) with diverse neutralization breadth and potency were 27
isolated from mice immunized with the receptor binding domain (RBD), S2GΔHR2 spike, and 28
SApNP vaccines. The mechanism of vaccine-induced immunity was examined in mice. 29
Compared with the soluble spike, the I3-01v9 SApNP showed 6-fold longer retention, 4-fold 30
greater presentation on follicular dendritic cell dendrites, and 5-fold stronger germinal center 31
reactions in lymph node follicles. 32
33
34
ONE-SENTENCE SUMMARY (125 characters) 35
With a well-defined mechanism, spike nanoparticle vaccines can effectively counter SARS-36
CoV-2 variants. 37
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3
INTRODUCTION 38
The COVID-19 pandemic has led to more than 188 million infection cases and 4 million deaths 39
globally. Antibody responses to SARS-CoV-2 spike antigens can be sustained for several months 40
in most COVID-19 patients after infection (1-4). However, recently identified variants of 41
concern (VOCs) exhibit higher transmissibility and resistance to prior immunity as SARS-CoV-2 42
continues to adapt to the human host (5, 6). One such variant, B.1.1.7 (WHO classification: 43
Alpha), emerged from southeast England in October 2020 and accounted for two-thirds of new 44
infections in London in December 2020, with a higher transmission rate (43-90%) and risk of 45
mortality (32-104%) than previously circulating strains (7, 8). Other variants, such as B.1.351 46
(Beta) and P.1 (Gamma), also became prevalent in three provinces in South Africa and Manaus, 47
Brazil, respectively (6, 9, 10). The B.1.617.2 (Delta) variant, which was initially identified in 48
India, is becoming a dominant strain in many countries (11, 12) and responsible for the majority 49
of new COVID-19 cases. This variant was found to be ~60% more transmissible than the highly 50
infectious B.1.1.7 variant (12). The rise of SARS-CoV-2 VOCs and their rapid spread worldwide 51
result in more infection cases, hospitalizations, and potentially more deaths, further straining 52
healthcare resources (10). 53
To date, eight COVID-19 vaccines have been approved for emergency use in humans, 54
with more than 90 candidates assessed in various phases of clinical trials (13). With the 55
exception of inactivated whole-virion vaccines, diverse platforms have been used to deliver the 56
recombinant SARS-CoV-2 spike, such as mRNA-encapsulating liposomes (e.g., BNT162b2 and 57
mRNA-1273), adenovirus vectors (e.g., ChAdOx1 nCoV-19 [AZD1222], CTII-nCoV, Sputnik 58
V, and Ad26.COV2.S), and micelle-attached spikes (e.g., NVX-CoV2373). These vaccines 59
demonstrated 65-96% efficacy in Phase 3 trials, with lower morbidity and mortality associated 60
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4
with COVID-19 disease (14-19). However, a notable loss of vaccine efficacy against new SARS-61
CoV-2 variants was reported, likely caused by spike mutations in the receptor-binding domain 62
(RBD; e.g., K417N, E484K, and N501Y), N-terminal domain (NTD; e.g., L18F, D80A, D215G, 63
and
Δ
242-244), and other regions that are critical to spike stability and function (e.g., D614G and 64
P681R) (6, 11, 20-25). Among circulating VOCs, the B.1.351 lineage appeared to be most 65
resistant to neutralization by convalescent plasma (9.4-fold) and vaccine sera (10.3- to 12.4-fold) 66
(26), whereas a lesser degree of reduction was observed for an early variant, B.1.1.7 (27-29). 67
Based on these findings, it was suggested that vaccines would need to be updated periodically to 68
maintain protection against rapidly evolving SARS-CoV-2 (30-32). However, in a recent study, 69
convalescent sera from B.1.351 or P.1-infected individuals showed a more visible reduction of 70
B.1.617.2 neutralization than convalescent sera from individuals infected with early pandemic 71
strains (33). Together, these issues raise the concern that herd immunity may be difficult to 72
achieve, highlighting the necessity of developing vaccines that can elicit a broadly neutralizing 73
antibody (bNAb) response to current and emerging variants (25, 31). As previously reported (34-74
38), the production of a bNAb response relies on long-lived germinal center (GC) reactions to 75
activate precursor B cells, stimulate affinity maturation, and form long-term immune memory. In 76
particular, antigen retention and presentation within lymph node follicles are key to the induction 77
of long-lived GC reactions (34, 36, 39) and should be considered in the development of bNAb-78
producing vaccines (40). 79
We previously investigated the cause of SARS-CoV-2 spike metastability and rationally 80
designed the S2GΔHR2 spike, which was displayed on three self-assembling protein 81
nanoparticle (SApNP) platforms, including ferritin (FR) 24-mer and multilayered E2p and I3-82
01v9 60-mers, as COVID-19 vaccine candidates (41). In the present study, we investigated the 83
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5
vaccine-induced NAb response to SARS-CoV-2 VOCs and mechanism by which SApNP 84
vaccines (e.g., I3-01v9) generate such a response. We first examined the neutralizing activity of 85
mouse plasma from our previous study (41) against four representative SARS-CoV-2 variants, 86
B.1.1.7, B.1.351, P.1, and B.1.617
Rec
, which was derived from an early analysis of the B.1.617 87
lineage (11) and shares key spike mutations with VOC B.1.617.2. Mouse plasma induced by the 88
S2GΔHR2 spike-presenting I3-01v9 SApNP potently neutralized all four variants with 89
comparable titers to the wildtype strain, Wuhan-Hu-1. When a different injection route was 90
tested in mouse immunization, E2p and I3-01v9 SApNPs sustained neutralizing titers against the 91
four variants, even at a low dosage of 3.3 μg, whereas a significant reduction of plasma 92
neutralization was observed for the soluble spike. Next, we examined the adjuvant effect on 93
vaccine-induced humoral and T-cell responses for the I3-01v9 SApNP. While detectable plasma 94
neutralization was observed for the non-adjuvanted I3-01v9 group, conventional adjuvants, such 95
as aluminum hydroxide (AH) and phosphate (AP), boosted the titers by 8.6- to 11.3-fold (or 9.6 96
to 12.3 times). Adjuvants that target the stimulator of interferon genes (STING) and Toll-like 97
receptor 9 (TLR9) pathways enhanced neutralization by 21- to 35-fold, alone or combined with 98
AP, in addition to a Th1-biased cellular response. We then performed antigen-specific single-cell 99
sorting and isolated 20 monoclonal antibodies (mAbs) from RBD, spike, and I3-01v9 SApNP-100
immunized mice. These mAbs were derived from diverse B cell lineages, of which some 101
neutralized the wildtype Wuhan-Hu-1 strain and four variants with equivalent potency. Lastly, 102
we investigated how SApNPs behave in lymph nodes and induce GCs by characterizing vaccine 103
delivery and immunological responses at the intraorgan, intracellular, and intercellular levels in 104
mice. The I3-01v9 SApNP showed 6-fold longer retention, 4-fold greater presentation on 105
follicular dendritic cell (DC) dendrites, and 5-fold higher GC reactions than the soluble spike. 106
.CC-BY-NC 4.0 International licenseavailable under a
(which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made
The copyright holder for this preprintthis version posted September 9, 2021. ; https://doi.org/10.1101/2021.03.26.437274doi: bioRxiv preprint