T cell immunity ameliorates COVID-19 disease severity and provides post-exposure prophylaxis after peptide-vaccination, in Syrian hamsters

Abstract

Background: The emergence of novel SARS-CoV-2 variants that resist neutralizing antibodies drew the attention to cellular immunity and calls for the development of alternative vaccination strategies to combat the pandemic. Here, we have assessed the kinetics of T cell responses and protective efficacy against severe COVID-19 in pre- and post-exposure settings, elicited by PolyPEPI-SCoV-2, a peptide based T cell vaccine.

Methods: 75 Syrian hamsters were immunized subcutaneously with PolyPEPI-SCoV-2 on D0 and D14. On D42, hamsters were intranasally challenged with 10² TCID₅₀ of the virus. To analyze immunogenicity by IFN-γ ELISPOT and antibody secretion, lymphoid tissues were collected both before (D0, D14, D28, D42) and after challenge (D44, D46, D49). To measure vaccine efficacy, lung tissue, throat swabs and nasal turbinate samples were assessed for viral load and histopathological changes. Further, body weight was monitored on D0, D28, D42 and every day after challenge.

Results: The vaccine induced robust activation of T cells against all SARS-CoV-2 structural proteins that were rapidly boosted after virus challenge compared to control animals (~4-fold, p<0.05). A single dose of PolyPEPI-SCoV-2 administered one day after challenge also resulted in elevated T cell response (p<0.01). The vaccination did not induce virus-specific antibodies and viral load reduction. Still, peptide vaccination significantly reduced body weight loss (p<0.001), relative lung weight (p<0.05) and lung lesions (p<0.05), in both settings.

Conclusion: Our study provides first proof of concept data on the contribution of T cell immunity on disease course and provide rationale for the use of T cell-based peptide vaccines against both novel SARS-CoV-2 variants and supports post-exposure prophylaxis as alternative vaccination strategy against COVID-19.

Keywords: SARS-CoV-2; T cells; adaptive immunity; post-exposure prophylaxis; therapeutic; vaccine.

Figure 1

Study outline and overview of experimental groups. 9-10 week old Syrian hamsters (n=75) were divided into 4 cohorts. Hamsters were immunized with PolyPEPI-SCoV-2 subcutaneously on Day 0 (D0) and D14 (prophylactic setting, P) or as a PEP treatment (T) on D43, one day after challenge with SARS-CoV-2. As controls, hamsters were injected with adjuvant only (A) or were left untreated (C). Where indicated, (A, C) controls were combined as Controls (C):( A+C). Hamsters were challenged with 10² TCID50 SARS-CoV-2 on D42 and 5 animals were sacrificed in each group for immune response (T cell response and antibody, IR) and efficacy evaluation (body weight, virology and pathology, (E) at study days (D) indicated on the figure.

Figure 2

PolyPEPI-SCoV-2 induces strong and rapid adaptive T cell immunity pre- and post-infection. (A) PolyPEPI-SCoV-2 induces adaptive T cell immunity in the spleen. Immunogenicity is illustrated on D14, D28, D46 and D49 as mean ± SD spot forming unit (SFU) per 10⁶ spleen cells of 5 animals against the 30-mer peptide pool (one-tailed Mann-Whitney test, ns, p>0.05, (*) p<0.05, (**) p<0.01. NA, not-analyzed (response not investigated). (B, C) T cell response kinetics was evaluated on D14, D28, D42 (0 d.p.i.), D44 (2 d.p.i.), D46 (4 d.p.i.) and D49 (7 d.p.i.) against (B) the 9-mer and (C) the 30-mer peptide pools (S, N, M and E peptides pooled together). Data are shown as average SFU per 10⁶ spleen cells of 5 animals or SFU per 10⁶ pooled lymph node cells (pool of 5 animals per group). Response to 9-mer represents mainly CD8+ T cell while response to 30-mer depicts combined CD4+ plus CD8+ T cell activation.

Figure 3

PolyPEPI-SCoV-2 does not induce antibody secretion and virus neutralization (A, B) Antibody response measured by (A) pre-S and (B) N protein IgG ELISAs from hamster sera. Results are shown as average of secreted antibodies (ELU/mL) ± SD of 5 animals per group (or 10 in the Controls group) measured in duplicates from D14 to D49 (lower panel) and separately illustrated on D49 (upper panel). (C) SARS-CoV-2 neutralization titers were measured by virus neutralization assay. Results are shown as average of virus neutralizing (VN) titer ± SD of 5 animals per group (or 10 in the Controls group) measured in triplicates from D14 to D49 (lower panel) or separately illustrated on D49 (upper panel). (C): Controls (combined challenge and adjuvant controls), P: prophylactic, T: post-exposure prophylactic. ns, non-significant result, p>0.05 by one-tailed Mann-Whitney test.

Figure 4

PolyPEPI-SCoV-2 does not alter viral clearance from the respiratory tract. (A) Viral load in the lung tissue, nasal turbinates and throat, determined on D49 by PCR. Results are shown as average of log10 VP/mL or log10 CP/mL ± SD of 5 animals per group (or 10 in the Controls group). (B) Replication-competent virus isolated from the lung, nasal turbinates and throat, measured by TCID50 assay on D49. Results are shown as average of log10 TCID50/gram tissue ± SD of 5 animals per group (or 10 in the Controls group). Dotted lines indicate the limit of detection (LOD). CP, crossing-point value, TCID50/g: 50% tissue culture infective dose per gram tissue, VP, virus particles, C, Controls (combined challenge and adjuvant controls), P: prophylactic, T: post-exposure prophylactic. ns, non-significant result, p>0.05 by one-tailed Mann-Whitney test.

Figure 5

PolyPEPI-SCoV-2 decreases body weight loss of SARS-CoV-2 infected hamsters. Post challenge daily relative body weight changes are represented for each individual animal (left panel) or on D49 (right panel) compared to D42. Results are shown as average of body weight loss % ± SD of 5 animals (10 Controls) per group (right panel). Solid lines represent average relative weight loss trendlines for the cohorts (left panel). C, Controls (combined challenge and adjuvant controls), P: prophylactic, T: post-exposure prophylactic. *p<0.5, **p<0.01, ***p<0.001 by one-tailed Mann-Whitney test.

Figure 6

PolyPEPI-SCoV-2 decreases lung pathology of SARS-CoV-2 infected hamsters. (A) Lung scores (see subpanel B) were evaluated based on hematoxylin-eosin (H&E) stained tissue sections, examined by light microscopy. Representative results of one animal from each group on D49 are shown as follows: overview of the H&E stained lung (upper left panels), bronchitis (upper right panels), alveolitis (lower left panels), vasculitis (lower right panels). (B–D) Lung pathology was evaluated as lung score (B), lung lesions (C) and relative lung weight (D) on D49. Data are presented as sum of the score of the disease parameters (detailed in Materials and Methods and in Supplementary Figure 5 ). (C) Gross pathology was evaluated on the whole lung lobes via visual observation by an independent pathologist and results provided as the % of lung area affected (lung lesion). (D) Relative lung weight illustrated as percentages of body weight [(lung weight/body weight)*100]. Results are shown as average ± SD of 5 animals (10 Controls) per group. C, challenge and adjuvant controls, P, prophylactic, T, post-exposure prophylactic. ns, p>0.05, (*) p<0.05 calculated using one-tailed Mann-Whitney test.

Figure 7

Impact of SARS-CoV-2 variants of concern (VOC) on PolyPEPI-SCoV-2 peptides. Mutations described for major VOCs are shown and compared to PolyPEPI-SCoV-2 peptides (S2, S5, S9, E, M, N1, N2, N3 and N4) (25).