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Evaluation of candidate vaccine approaches for MERS-CoV.

Wang L, Shi W, Joyce MG, Modjarrad K, Zhang Y, Leung K, Lees CR, Zhou T, Yassine HM, Kanekiyo M, Yang ZY, Chen X, Becker MM, Freeman M, Vogel L, Johnson JC, Olinger G, Todd JP, Bagci U, Solomon J, Mollura DJ, Hensley L, Jahrling P, Denison MR, Rao SS, Subbarao K, Kwong PD, Mascola JR, Kong WP, Graham BS - Nat Commun (2015)

Bottom Line: Efforts focused solely on the receptor-binding domain (RBD) of the viral Spike (S) glycoprotein may not optimize neutralizing antibody (NAb) responses.Here we show that immunogens based on full-length S DNA and S1 subunit protein elicit robust serum-neutralizing activity against several MERS-CoV strains in mice and non-human primates.Multiple neutralization mechanisms were demonstrated by solving the atomic structure of a NAb-RBD complex, through sequencing of neutralization escape viruses and by constructing MERS-CoV S variants for serological assays.

View Article: PubMed Central - PubMed

Affiliation: Vaccine Research Center, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland 20892, USA.

ABSTRACT
The emergence of Middle East respiratory syndrome coronavirus (MERS-CoV) as a cause of severe respiratory disease highlights the need for effective approaches to CoV vaccine development. Efforts focused solely on the receptor-binding domain (RBD) of the viral Spike (S) glycoprotein may not optimize neutralizing antibody (NAb) responses. Here we show that immunogens based on full-length S DNA and S1 subunit protein elicit robust serum-neutralizing activity against several MERS-CoV strains in mice and non-human primates. Serological analysis and isolation of murine monoclonal antibodies revealed that immunization elicits NAbs to RBD and, non-RBD portions of S1 and S2 subunit. Multiple neutralization mechanisms were demonstrated by solving the atomic structure of a NAb-RBD complex, through sequencing of neutralization escape viruses and by constructing MERS-CoV S variants for serological assays. Immunization of rhesus macaques confers protection against MERS-CoV-induced radiographic pneumonia, as assessed using computerized tomography, supporting this strategy as a promising approach for MERS-CoV vaccine development.

No MeSH data available.


Related in: MedlinePlus

MERS-CoV Spike immunogens elicit potent, long-lived neutralization in NHPs and protect from severe lung infiltrates.Selected immunogens from mouse studies were evaluated in NHPs. (a) Schematic representation of full-length MERS-CoV Spike protein cDNA and recombinant S1 protein. The DNA construct consists of full-length Spike and transmembrane domain. The protein construct contains a truncated Spike with the S1 subunit. RBD, receptor-binding domain; SP, signal peptide; TM, transmembrane domain; 3CHis, Human rhinovirus 3C protease cleavage site, followed by 6 × histidine tag. (b) Immunogenicity of three vaccine regimens. Six NHPs, per group, were immunized intramuscularly using plasmid DNA with electroporation at weeks 0, 4 and 8; plasmid DNA with electroporation at weeks 0 and 4; and protein plus AlPO4 at week 8 or protein plus AlPO4 at weeks 0 and 8. Two weeks after immunization and at weeks 12 and 18, neutralizing antibody titres were measured against pseudotyped MERS-CoV England1. Different symbols indicate sera from six NHPs per group collected at indicated time points. IC90 neutralization titres (GMT with 95% confidence interval) from sera were determined. Nonparametric two-tailed t-test (Mann–Whitney) was used for statistical analysis. (c) Spike immunogens protect against pulmonary disease in NHPs. Six unimmunized NHPs and 12 NHPs were immunized with one of two selected candidate vaccine immunogens ((1) full-length S DNA prime/S1 subunit protein boost; (2) S1 subunit protein prime/S1 subunit protein boost) and challenged with MERS-CoV 19 weeks after last vaccine boost. Intratracheal inoculation of 5 × 106 p.f.u. of the JordanN3 strain (GenBank ID: KC776174.1) was performed on each animal. The per cent abnormal lung volume in all NHPs peaked on day 3 post challenge; however, the lung infiltrates were significantly more extensive and prolonged in the unvaccinated compared with vaccinated NHPs. A nonparametric two-tailed t-test (Mann–Whitney) was used. *P value <0.05; ** P value<0.01. (d) Abnormal lung segmental images from selected animals on day 6 post challenge. The images correspond to NHP lung volume data points circled in black in Fig. 5c. The CT images and abnormal lung segments for all 18 animals are shown in Supplementary Fig. 14 a–c.
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f5: MERS-CoV Spike immunogens elicit potent, long-lived neutralization in NHPs and protect from severe lung infiltrates.Selected immunogens from mouse studies were evaluated in NHPs. (a) Schematic representation of full-length MERS-CoV Spike protein cDNA and recombinant S1 protein. The DNA construct consists of full-length Spike and transmembrane domain. The protein construct contains a truncated Spike with the S1 subunit. RBD, receptor-binding domain; SP, signal peptide; TM, transmembrane domain; 3CHis, Human rhinovirus 3C protease cleavage site, followed by 6 × histidine tag. (b) Immunogenicity of three vaccine regimens. Six NHPs, per group, were immunized intramuscularly using plasmid DNA with electroporation at weeks 0, 4 and 8; plasmid DNA with electroporation at weeks 0 and 4; and protein plus AlPO4 at week 8 or protein plus AlPO4 at weeks 0 and 8. Two weeks after immunization and at weeks 12 and 18, neutralizing antibody titres were measured against pseudotyped MERS-CoV England1. Different symbols indicate sera from six NHPs per group collected at indicated time points. IC90 neutralization titres (GMT with 95% confidence interval) from sera were determined. Nonparametric two-tailed t-test (Mann–Whitney) was used for statistical analysis. (c) Spike immunogens protect against pulmonary disease in NHPs. Six unimmunized NHPs and 12 NHPs were immunized with one of two selected candidate vaccine immunogens ((1) full-length S DNA prime/S1 subunit protein boost; (2) S1 subunit protein prime/S1 subunit protein boost) and challenged with MERS-CoV 19 weeks after last vaccine boost. Intratracheal inoculation of 5 × 106 p.f.u. of the JordanN3 strain (GenBank ID: KC776174.1) was performed on each animal. The per cent abnormal lung volume in all NHPs peaked on day 3 post challenge; however, the lung infiltrates were significantly more extensive and prolonged in the unvaccinated compared with vaccinated NHPs. A nonparametric two-tailed t-test (Mann–Whitney) was used. *P value <0.05; ** P value<0.01. (d) Abnormal lung segmental images from selected animals on day 6 post challenge. The images correspond to NHP lung volume data points circled in black in Fig. 5c. The CT images and abnormal lung segments for all 18 animals are shown in Supplementary Fig. 14 a–c.

Mentions: Of the eight vaccine regimens tested in mice, the three most immunogenic were taken forward for evaluation in NHPs (Fig. 5a). Comparable to the mice, NHPs primed with either S1 protein (plus aluminium phosphate (AlPO4) adjuvant) or S DNA (followed by electroporation) and boosted with S1 protein (plus AlPO4 adjuvant) generated the highest NAb titres, as measured by the pseudotyped virus neutralization assay, compared with the S DNA-only group (Fig. 5b). Both groups initially had low antibody titres after priming that increased 10- to 100-fold after boosting. IC90 neutralization titers after the final boost were ∼1 log10 higher in mice than in NHPs, which could be because of the different animal models, vaccine doses or adjuvants used in the two studies. Antibody titres remained high, however, at more than 2.5 log10 at 10-week post boost and persisted at higher levels in the DNA–protein group. A microneutralization assay with the MERS-CoV JordanN3 strain was compared with the pseudotyped neutralization assay and demonstrated similar results. Sera from the two groups immunized with S DNA bound all epitopes recognized by the four previously characterized murine antibodies (Supplementary Fig. 13). Sera from NHPs immunized with S1 protein alone, however, blocked mAbs targeted to the RBD (D12 and F11) and non-RBD S1 subunit (G2) but not the S2 subunit (G4).


Evaluation of candidate vaccine approaches for MERS-CoV.

Wang L, Shi W, Joyce MG, Modjarrad K, Zhang Y, Leung K, Lees CR, Zhou T, Yassine HM, Kanekiyo M, Yang ZY, Chen X, Becker MM, Freeman M, Vogel L, Johnson JC, Olinger G, Todd JP, Bagci U, Solomon J, Mollura DJ, Hensley L, Jahrling P, Denison MR, Rao SS, Subbarao K, Kwong PD, Mascola JR, Kong WP, Graham BS - Nat Commun (2015)

MERS-CoV Spike immunogens elicit potent, long-lived neutralization in NHPs and protect from severe lung infiltrates.Selected immunogens from mouse studies were evaluated in NHPs. (a) Schematic representation of full-length MERS-CoV Spike protein cDNA and recombinant S1 protein. The DNA construct consists of full-length Spike and transmembrane domain. The protein construct contains a truncated Spike with the S1 subunit. RBD, receptor-binding domain; SP, signal peptide; TM, transmembrane domain; 3CHis, Human rhinovirus 3C protease cleavage site, followed by 6 × histidine tag. (b) Immunogenicity of three vaccine regimens. Six NHPs, per group, were immunized intramuscularly using plasmid DNA with electroporation at weeks 0, 4 and 8; plasmid DNA with electroporation at weeks 0 and 4; and protein plus AlPO4 at week 8 or protein plus AlPO4 at weeks 0 and 8. Two weeks after immunization and at weeks 12 and 18, neutralizing antibody titres were measured against pseudotyped MERS-CoV England1. Different symbols indicate sera from six NHPs per group collected at indicated time points. IC90 neutralization titres (GMT with 95% confidence interval) from sera were determined. Nonparametric two-tailed t-test (Mann–Whitney) was used for statistical analysis. (c) Spike immunogens protect against pulmonary disease in NHPs. Six unimmunized NHPs and 12 NHPs were immunized with one of two selected candidate vaccine immunogens ((1) full-length S DNA prime/S1 subunit protein boost; (2) S1 subunit protein prime/S1 subunit protein boost) and challenged with MERS-CoV 19 weeks after last vaccine boost. Intratracheal inoculation of 5 × 106 p.f.u. of the JordanN3 strain (GenBank ID: KC776174.1) was performed on each animal. The per cent abnormal lung volume in all NHPs peaked on day 3 post challenge; however, the lung infiltrates were significantly more extensive and prolonged in the unvaccinated compared with vaccinated NHPs. A nonparametric two-tailed t-test (Mann–Whitney) was used. *P value <0.05; ** P value<0.01. (d) Abnormal lung segmental images from selected animals on day 6 post challenge. The images correspond to NHP lung volume data points circled in black in Fig. 5c. The CT images and abnormal lung segments for all 18 animals are shown in Supplementary Fig. 14 a–c.
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Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4525294&req=5

f5: MERS-CoV Spike immunogens elicit potent, long-lived neutralization in NHPs and protect from severe lung infiltrates.Selected immunogens from mouse studies were evaluated in NHPs. (a) Schematic representation of full-length MERS-CoV Spike protein cDNA and recombinant S1 protein. The DNA construct consists of full-length Spike and transmembrane domain. The protein construct contains a truncated Spike with the S1 subunit. RBD, receptor-binding domain; SP, signal peptide; TM, transmembrane domain; 3CHis, Human rhinovirus 3C protease cleavage site, followed by 6 × histidine tag. (b) Immunogenicity of three vaccine regimens. Six NHPs, per group, were immunized intramuscularly using plasmid DNA with electroporation at weeks 0, 4 and 8; plasmid DNA with electroporation at weeks 0 and 4; and protein plus AlPO4 at week 8 or protein plus AlPO4 at weeks 0 and 8. Two weeks after immunization and at weeks 12 and 18, neutralizing antibody titres were measured against pseudotyped MERS-CoV England1. Different symbols indicate sera from six NHPs per group collected at indicated time points. IC90 neutralization titres (GMT with 95% confidence interval) from sera were determined. Nonparametric two-tailed t-test (Mann–Whitney) was used for statistical analysis. (c) Spike immunogens protect against pulmonary disease in NHPs. Six unimmunized NHPs and 12 NHPs were immunized with one of two selected candidate vaccine immunogens ((1) full-length S DNA prime/S1 subunit protein boost; (2) S1 subunit protein prime/S1 subunit protein boost) and challenged with MERS-CoV 19 weeks after last vaccine boost. Intratracheal inoculation of 5 × 106 p.f.u. of the JordanN3 strain (GenBank ID: KC776174.1) was performed on each animal. The per cent abnormal lung volume in all NHPs peaked on day 3 post challenge; however, the lung infiltrates were significantly more extensive and prolonged in the unvaccinated compared with vaccinated NHPs. A nonparametric two-tailed t-test (Mann–Whitney) was used. *P value <0.05; ** P value<0.01. (d) Abnormal lung segmental images from selected animals on day 6 post challenge. The images correspond to NHP lung volume data points circled in black in Fig. 5c. The CT images and abnormal lung segments for all 18 animals are shown in Supplementary Fig. 14 a–c.
Mentions: Of the eight vaccine regimens tested in mice, the three most immunogenic were taken forward for evaluation in NHPs (Fig. 5a). Comparable to the mice, NHPs primed with either S1 protein (plus aluminium phosphate (AlPO4) adjuvant) or S DNA (followed by electroporation) and boosted with S1 protein (plus AlPO4 adjuvant) generated the highest NAb titres, as measured by the pseudotyped virus neutralization assay, compared with the S DNA-only group (Fig. 5b). Both groups initially had low antibody titres after priming that increased 10- to 100-fold after boosting. IC90 neutralization titers after the final boost were ∼1 log10 higher in mice than in NHPs, which could be because of the different animal models, vaccine doses or adjuvants used in the two studies. Antibody titres remained high, however, at more than 2.5 log10 at 10-week post boost and persisted at higher levels in the DNA–protein group. A microneutralization assay with the MERS-CoV JordanN3 strain was compared with the pseudotyped neutralization assay and demonstrated similar results. Sera from the two groups immunized with S DNA bound all epitopes recognized by the four previously characterized murine antibodies (Supplementary Fig. 13). Sera from NHPs immunized with S1 protein alone, however, blocked mAbs targeted to the RBD (D12 and F11) and non-RBD S1 subunit (G2) but not the S2 subunit (G4).

Bottom Line: Efforts focused solely on the receptor-binding domain (RBD) of the viral Spike (S) glycoprotein may not optimize neutralizing antibody (NAb) responses.Here we show that immunogens based on full-length S DNA and S1 subunit protein elicit robust serum-neutralizing activity against several MERS-CoV strains in mice and non-human primates.Multiple neutralization mechanisms were demonstrated by solving the atomic structure of a NAb-RBD complex, through sequencing of neutralization escape viruses and by constructing MERS-CoV S variants for serological assays.

View Article: PubMed Central - PubMed

Affiliation: Vaccine Research Center, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland 20892, USA.

ABSTRACT
The emergence of Middle East respiratory syndrome coronavirus (MERS-CoV) as a cause of severe respiratory disease highlights the need for effective approaches to CoV vaccine development. Efforts focused solely on the receptor-binding domain (RBD) of the viral Spike (S) glycoprotein may not optimize neutralizing antibody (NAb) responses. Here we show that immunogens based on full-length S DNA and S1 subunit protein elicit robust serum-neutralizing activity against several MERS-CoV strains in mice and non-human primates. Serological analysis and isolation of murine monoclonal antibodies revealed that immunization elicits NAbs to RBD and, non-RBD portions of S1 and S2 subunit. Multiple neutralization mechanisms were demonstrated by solving the atomic structure of a NAb-RBD complex, through sequencing of neutralization escape viruses and by constructing MERS-CoV S variants for serological assays. Immunization of rhesus macaques confers protection against MERS-CoV-induced radiographic pneumonia, as assessed using computerized tomography, supporting this strategy as a promising approach for MERS-CoV vaccine development.

No MeSH data available.


Related in: MedlinePlus