1. Immunology and Inflammation
  2. Microbiology and Infectious Disease

SARS-CoV2 variant-specific replicating RNA vaccines protect from disease following challenge with heterologous variants of concern

  1. David W Hawman  Is a corresponding author
  2. Kimberly Meade-White
  3. Jacob Archer
  4. Shanna S Leventhal
  5. Drew Wilson
  6. Carl Shaia
  7. Samantha Randall
  8. Amit P Khandhar
  9. Kyle Krieger
  10. Tien-Ying Hsiang
  11. Michael Gale
  12. Peter Berglund
  13. Deborah Heydenburg Fuller
  14. Heinz Feldmann  Is a corresponding author
  15. Jesse H Erasmus  Is a corresponding author
  1. Laboratory of Virology, Division of Intramural Research, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Rocky Mountain Laboratories, United States
  2. HDT Bio, United States
  3. Rocky Mountain Veterinary Branch, Division of Intramural Research, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Rocky Mountain Laboratories, United States
  4. Department of Microbiology, University of Washington School of Medicine, United States
  5. Center for Innate Immunity and Immune Disease, Department of Immunology, University of Washington School of Medicine, United States
https://doi.org/10.7554/eLife.75537
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  1. David W Hawman
  2. Kimberly Meade-White
  3. Jacob Archer
  4. Shanna S Leventhal
  5. Drew Wilson
  6. Carl Shaia
  7. Samantha Randall
  8. Amit P Khandhar
  9. Kyle Krieger
  10. Tien-Ying Hsiang
  11. Michael Gale
  12. Peter Berglund
  13. Deborah Heydenburg Fuller
  14. Heinz Feldmann
  15. Jesse H Erasmus
(2022)
SARS-CoV2 variant-specific replicating RNA vaccines protect from disease following challenge with heterologous variants of concern
eLife 11:e75537.
https://doi.org/10.7554/eLife.75537
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7 figures, 1 table and 2 additional files

Figures

Figure 1
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Design of full-length SARS-CoV2 spike immunogens harboring deletions and/or substitutions specific to variants of concern.

(A) Using the full-length, pre-fusion-stabilized (KV995PP) spike (S) of the original A.1 lineage of SARS-CoV2 as a reference, including the signal peptide (sp) and regions corresponding to S1, S2, transmembrane (TM), and cytoplasmic (CD) domains, the various deletions and/or substitutions corresponding to the B.1, B.1.1.7, and B.1.351 lineages were introduced. Additionally, a subset of constructs were prepared on the native version of spike. These six open reading frames were cloned into the replicating RNA (repRNA) backbone downstream of the sub-genomic promoter prior to production of capped mRNA. (B) RNA of each construct was then formulated with lipid inorganic nanoparticle (LION) and serial dilutions added to monolayers of baby hamster kidney cells. Cell surface-bound spike was then measured by sandwich enzyme linked immunosorbent assay (ELISA) of cell lysates harvested at 24 hr post-transfection and (C) half-maximal effective concentrations (EC50) determined for each RNA construct.

Figure 1—source data 1

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https://cdn.elifesciences.org/articles/75537/elife-75537-fig1-data1-v2.xlsx
Figure 2
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Relative binding and neutralizing antibody responses induced by each vaccine candidate.

C57BL/6 mice (n = 8/group) received a 1 µg intramuscular injection on days 0 and 28 of lipid inorganic nanoparticle (LION)/replicating RNA (repRNA) encoding either the native conformation of spike derived from A.1, or B.1 viruses, or the pre-fusion-stabilized conformation of spike derived from B.1, B.1.1.7, or B.1.351 viruses, corresponding to repRNAs 600, 675, 676, 673, and 671, respectively. Mice were bled 14 days after the boost immunization and (A) A.1 spike (S)-, S1 domain-, S2 domain-, or receptor-binding domain (RBD)-binding antibody responses measured by enzyme linked immunosorbent assay (ELISA). Neutralizing antibody responses (B–F) measured by 80% plaque reduction neutralization tests (PRNT80) against A.1, B.1, B.1.1.7, or B.1.351 viruses in samples from mice vaccinated with native A.1 (B), native B.1 (C), pre-fusion B.1 (D), pre-fusion B.1.1.7 (E), or pre-fusion B.1.351 (F) spike-derived vaccines. Data in A are presented as geometric means (±geometric standard deviations) along with each individual sample and differences in S-, S1-, S2-, or RBD-binding titers between the 600 A.1 group and other groups were compared by two-way ANOVA (*p < 0.05). Data in B–F are presented as each individual sample connected by black lines with the geometric mean depicted in red and differences between geometric means were compared by Student’s t test.

Figure 2—source data 1

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https://cdn.elifesciences.org/articles/75537/elife-75537-fig2-data1-v2.xlsx
Figure 3
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Post-boost neutralizing antibody responses in Syrian Golden hamsters.

Sera from animals immunized with lipid inorganic nanoparticle (LION) complexed with replicating RNA (repRNA) vaccine variants A.1, B.1.1.7, or B.1.351 were incubated with live virus of variant A.1 (black), B.1.1.7 (pink), B.1.351 (green), or B.1.617.2 (purple) as indicated. Plaque-reduction neutralizing titers are indicated by individual symbols with geometric mean titers represented by the height of the bars. Indicated statistical comparisons performed using a two-way ANOVA with Tukey’s multiple comparisons test. ns p > 0.05, *p < 0.05, ***p < 0.001, ****p < 0.0001.

Figure 3—source data 1

Source data for Figure 3.

https://cdn.elifesciences.org/articles/75537/elife-75537-fig3-data1-v2.xlsx
Figure 4
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Replicating RNA (repRNA) vaccination significantly reduces viral shedding.

Mock or repRNA-vaccinated hamsters were challenged with 1000 tissue-culture infectious dose 50 assay (TCID50) of the indicated SARS-CoV2 strains via the IN route. At day 2 or 4 post-infection (PI), oral swabs were collected. SARS-CoV2 in the swabs was quantified by qRT-PCR specific for the sub-genomic (SgE) RNA (A, C, E) or infectious virus by TCID50 assay (B, D, F). N = 5 (mock-vaccinated, B.1.351 challenge) or 6 (all other groups). A two-way ANOVA with Dunnett’s multiple comparison test against mock-vaccinated hamsters was performed. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. Comparisons without indicated p-values were considered non-significant (p > 0.05).

Figure 4—source data 1

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https://cdn.elifesciences.org/articles/75537/elife-75537-fig4-data1-v2.xlsx
Figure 5
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Replicating RNA (repRNA) vaccination significantly reduces viral burden in the lungs.

Mock or repRNA-vaccinated hamsters were challenged with 1000 tissue-culture infectious dose 50 assay (TCID50) of the indicated SARS-CoV2 strains via the IN route. At day 4 post-infection (PI), hamsters were euthanized, and lung tissue collected. SARS-CoV2 burden in the lung was quantified by qRT-PCR specific for the sub-genomic (SgE) RNA (A, C, E). Infectious virus in the lungs was quantified by TCID50 assay (B, D, F). Indicated statistical comparisons performed using a one-way ANOVA with Dunnett’s multiple comparison test against mock-vaccinated hamsters. *p < 0.05, ****p < 0.0001. Comparisons without indicated p-values were non-significant (p > 0.05).

Figure 5—source data 1

Source data for Figure 5.

https://cdn.elifesciences.org/articles/75537/elife-75537-fig5-data1-v2.xlsx
Figure 6
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Replicating RNA (repRNA) vaccination reduces lung pathology and SARS-CoV2 antigen burden in lung tissue.

Mock or repRNA-vaccinated hamsters were challenged with 1000 tissue-culture infectious dose 50 assay (TCID50) of the indicated SARS-CoV2 strains via the IN route. At day 4 post-infection (PI), hamsters were euthanized and lungs formalin-fixed and paraffin-embedded. Sections were stained with H&E (top row of each challenge) or for SARS-CoV2 viral antigen (bottom row of each challenge). Representative images of each group are shown.

Figure 7
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Replicating RNA (repRNA) vaccination significantly protects against lung pathology in hamsters.

Mock or repRNA-vaccinated hamsters were challenged with 1000 tissue-culture infectious dose 50 assay (TCID50) of the indicated SARS-CoV2 strains via the IN route. At day 4 post-infection (PI), hamsters were euthanized, lung weighed and lungs formalin-fixed and paraffin-embedded. Lung weights as percentage of body weight are reported (A–C). Lung sections were stained with H&E (D–F) or for SARS-CoV2 viral antigen (G–I). Sections were scored by a pathologist blind to study groups and assigned a score for percent area affected by SARS-CoV2 lesions and cumulative score presented (A–C) or presence of viral antigen (D–F). Indicated statistical comparisons performed using a one-way ANOVA with Dunnett’s multiple comparison test against mock-vaccinated hamsters. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. Comparisons without indicated p-values were considered non-significant (p > 0.05).

Figure 7—source data 1

Source data for Figure 7.

https://cdn.elifesciences.org/articles/75537/elife-75537-fig7-data1-v2.xlsx

Tables

Table 1
Fraction of hamster samples with any detectable infectious virus.
ChallengeRNAOral swabsLung tissue
D2D4D4
A.1Mock6/66/66/6
A.1 (604)6/62/61/6
B.1.351 (671)6/66/61/6
B.1.17 (673)3/60/60/6
B.1.351Mock5/55/55/5
A.1 (604)6/62/60/6
B.1.351 (671)2/62/60/6
B.1.17 (673)3/62/60/6
B.1.1.7Mock6/66/66/6
A.1 (604)6/62/60/6
B.1.351 (671)2/63/60/6
B.1.17 (673)1/63/60/6

Additional files

Supplementary file 1

Complete histological and immunohistochemistry (IHC) findings.

Formalin-fixed lung tissue was stained with H&E or viral antigen detected via IHC. Sections were scored by a certified pathologist blinded to study groups. Representative images are shown in Figure 6, scores in Figure 7, and complete summary of findings are provided here.

https://cdn.elifesciences.org/articles/75537/elife-75537-supp1-v2.xlsx
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  1. David W Hawman
  2. Kimberly Meade-White
  3. Jacob Archer
  4. Shanna S Leventhal
  5. Drew Wilson
  6. Carl Shaia
  7. Samantha Randall
  8. Amit P Khandhar
  9. Kyle Krieger
  10. Tien-Ying Hsiang
  11. Michael Gale
  12. Peter Berglund
  13. Deborah Heydenburg Fuller
  14. Heinz Feldmann
  15. Jesse H Erasmus
(2022)
SARS-CoV2 variant-specific replicating RNA vaccines protect from disease following challenge with heterologous variants of concern
eLife 11:e75537.
https://doi.org/10.7554/eLife.75537