Journal: Molecules

Article Title: Kinetic and Structural Aspects of Glycosaminoglycan–Monkeypox Virus Protein A29 Interactions Using Surface Plasmon Resonance

doi: 10.3390/molecules27185898

Figure Lengend Snippet: The binding affinity of MPXV L1R–heparin interactions.

Article Snippet: MPXV L1R and MPXV A29 proteins were purchased from Sino Biological Inc. Porcine intestinal heparin, with an average molecular weight of 15 kDa and polydispersity of 1.4, was purchased from Celsus Laboratories (Cincinnati, OH).

Techniques: Binding Assay

Journal: medRxiv

Article Title: JYNNEOS vaccination induced short-lived neutralizing antibody responses to monkeypox virus in naïve individuals

doi: 10.1101/2024.08.28.24312561

Figure Lengend Snippet: Serum specimens from eight JYNNEOS vaccine recipients were analyzed for IgG reactivity to recombinant protein antigens derived from mpox or vaccinia virus by microsphere immunoassay. One donor with who received ACAM2000 prior to JYNNEOS is shown separately in A and B. Means of seven individuals who had no prior smallpox vaccination are shown in C and D. Mean index values (MFI/cutoff) of MPXV E8L (gray squares), MPXV A35R (blue triangles) and MPXV H3L (white diamonds) were plotted for days 0, 8, 26, 56, 118, 231, and 434 post-vaccination (A,C). Mean index values (MFI/cutoff) of VACV L1R (white circles), VACV A33R (orange triangles) and VACV B5R (gray circles) were plotted for 0, 8, 26, 56, 118, 231, and 434 post-vaccination (B, D). The black dashed line at y = 1.0 indicates the cutoff value. The dotted line indicates the second dose of vaccine at day 28 post vaccination.

Article Snippet: Recombinant A33R (VAC-WR-A33R; Cat# NR545), B5R (VAC-WR-B5R; Cat# NR-546) and L1R (VAC-WR-L2R; Cat# NR-21986) were obtained from BEI Resources, Manassas VA. Mpox A35R (Cat # 230-30238), E8L (Cat# 230-30232) and H3L (Cat# 230-30233) were purchased from Ray Biotech, Peachtree Corners, GA.

Techniques: Recombinant, Derivative Assay, Virus

Journal: medRxiv

Article Title: The impact of antigenic distance on Orthopoxvirus Vaccination and Mpox Infection for cross-protective immunity

doi: 10.1101/2024.01.31.24302065

Figure Lengend Snippet: a, Cohort timeline overview indicated by time post vaccination and vaccine regimen received. Participants received Dryvax, JYNNEOS, or both Dryvax and JYNNEOS vaccines and plasma samples were collected at the indicated time points. Participants were then stratified in five main groups: C0, non-vaccinated (C0=24); C1, Dryvax (C1=78); C2, JYNNEOS (C2=62); C3, Dryvax + JYNNEOS (C3=29); C4, Mpox-Infected patients (C4=25). Dryvax was administered as a single dose at least 40 years prior to this study, whereas the JYNNEOS regimen consisted of two vaccination doses and was recently administered to these participants. Created with BioRender. b,c, Plasma reactive IgG to viral antigens was measured across all full vaccinated individuals and non-vaccinated controls (C0-C3) (C0=12, C1=40, C2=59, C3=31). b, Plasma reactivity to VACV proteins B5R, A33R, A27L, and L1R. c, Plasma reactivity to MPXV proteins B6R, A35R, A29L, L1R and E8L. d,e, LOWESS regression analysis of virus-specific IgG levels over time following JYNNEOS vaccination. d, Plasma reactivity to VACV proteins. e, Plasma reactivity to MPXV proteins. Regression lines are shown as purple (JYNNEOS) and green (JYNNEOS + Dryvax); shading represents 95% confidence interval. For the JYNNEOS cohort, baseline controls consisted of non-vaccinated individuals. Baseline controls for the Dryvax + JYNNEOS cohort were individuals who had previously been vaccinated with Dryvax. Significance was assessed by one-way ANOVA corrected for multiple comparisons using Tukey’s method. Baseline: C2=12, C3=40; 7 days post 1 st vaccine dose: C2=31, C3=16; 30-60 days post 1 st vaccine dose C2=31, C3=14; 7 days post 2 nd vaccine dose: C2=30, C3=13; 30-60 days post 2 nd vaccine dose: C2=22, C3=14; 180-240 days post 2nd vaccine dose: C2=10, C3=4. Horizontal bars represent median fold change. Non-Vax, non-vaccinated; TP5:180+, 180-240 days. ****P < 0.0001, ***P < 0.001, **P < 0.01, and *P < 0.05.

Article Snippet: MaxiSorp plates (96 wells; 442404, Thermo Scientific) were coated with 50□μl per well of either recombinant MPox A35R (A3R-M52H3-100 μg, ACROBiosystems), MPox A29L (A2L-M52H3-100 μg, ACROBiosystems), MPox E8L (E8L-M52H3-50 μg, ACROBiosystems), MPox L1R (L1R-M5241-50 μg, ACROBiosystems), MPox H3L (H3L-M52H1-50 μg, ACROBiosystems), MPox A30L (A3L-M5243-50 μg, ACROBiosystems), VACV A33R (40896-V07E-100 μg, Sino Biological), VACV A27L (40897-V07E-100 μg, Sino Biological), VACV L1R (40903-V07H-100 μg, Sino Biological), or VACV OPC005 [vD8L] (CSB-EP322653VAA-100 μg, CusaBio) at a concentration of 2□μg/ml in PBS and were incubated overnight at 4□°C.

Techniques: Vaccines, Clinical Proteomics, Infection, Virus

Journal: medRxiv

Article Title: The impact of antigenic distance on Orthopoxvirus Vaccination and Mpox Infection for cross-protective immunity

doi: 10.1101/2024.01.31.24302065

Figure Lengend Snippet: Participants received the JYNNEOS vaccine and plasma samples were collected longitudinally at multiple points: baseline (prior to the JYNNEOS vaccination), 7-30 days after the first vaccine dose, and 7-240 days after the second dose. The participants were then categorized into two groups based on their previous vaccination history: 1) the JYNNEOS group, consisting of naïve, non-vaccinated individuals who received the JYNNEOS regimen; and 2) the Dryvax + JYNNEOS group, comprising individuals who were previously vaccinated with Dryvax and subsequently received the JYNNEOS regimen. Importantly, for the JYNNEOS cohort, baseline consists of non-vaccinated individuals. Baseline controls for the Dryvax + JYNNEOS cohort were individuals who had previously been vaccinated with Dryvax. a,b,c,d Plasma reactivity to VACV proteins B5R, A33R, A27L, and L1 (top panel) and to MPXV proteins B6R, A35R, A29L, L1R and E8L (bottom panel). JYNNEOS: Baseline=12; Dose 1=62; Dose 2=62; JYNNEOS + Dryvax: Baseline=40; Dose 1= 30; Dose 2= 31. a, Plasma virus-specific IgG responses on naïve, non-vaccinated individuals following JYNNEOS vaccine doses. b, Plasma virus-specific IgG responses on individuals previously vaccinated with Dryvax and boosted with the JYNNEOS vaccine doses. c, Comparison of baseline plasma virus-specific IgG levels with those measured at the final collection time point (180-240 days post the second vaccination dose), in naïve, non-vaccinated individuals who received the JYNNEOS vaccine regimen. d, Comparison of baseline plasma virus-specific IgG levels with those measured at the final collection time point (180-240 days post the second vaccination dose) on individuals previously vaccinated with Dryvax and boosted with the JYNNEOS vaccine doses. Significance was assessed by one-way ANOVA corrected for multiple comparisons using Tukey’s method. JYNNEOS: Baseline=12; 180-240 days=10; JYNNEOS + Dryvax: Baseline=40; 180-240 days=4. Each dot represents a single individual. Horizontal bars represent median fold change. Boxes represent average ± SD. TP5:180+, 180-240 days. ****P < 0.0001, ***P < 0.001, **P < 0.01, and *P < 0.05.

Article Snippet: MaxiSorp plates (96 wells; 442404, Thermo Scientific) were coated with 50□μl per well of either recombinant MPox A35R (A3R-M52H3-100 μg, ACROBiosystems), MPox A29L (A2L-M52H3-100 μg, ACROBiosystems), MPox E8L (E8L-M52H3-50 μg, ACROBiosystems), MPox L1R (L1R-M5241-50 μg, ACROBiosystems), MPox H3L (H3L-M52H1-50 μg, ACROBiosystems), MPox A30L (A3L-M5243-50 μg, ACROBiosystems), VACV A33R (40896-V07E-100 μg, Sino Biological), VACV A27L (40897-V07E-100 μg, Sino Biological), VACV L1R (40903-V07H-100 μg, Sino Biological), or VACV OPC005 [vD8L] (CSB-EP322653VAA-100 μg, CusaBio) at a concentration of 2□μg/ml in PBS and were incubated overnight at 4□°C.

Techniques: Clinical Proteomics, Virus, Comparison

Journal: medRxiv

Article Title: The impact of antigenic distance on Orthopoxvirus Vaccination and Mpox Infection for cross-protective immunity

doi: 10.1101/2024.01.31.24302065

Figure Lengend Snippet: Blood samples were collected from individuals that received first-generation and/or third generation VACV-based vaccines along with MPXV infected patients. Analysis of immunogenicity were performed using neutralization assays with authentic Vaccinia virus (VACV), Cowpox virus (CWPXV) and Mpox virus (MPXV) virus. a, Plasma neutralization titers against VACV, CWPXV and MPXV across full vaccinated individuals and non-vaccinated controls (C0=21, C1=40, C2=55, C3=28). b, Schematic figure illustrating antigenic similarity from VACV across Orthopoxviruses used within this study. Colors indicate different viruses, VACV (pink), CWPXV (purple), MPXV (blue) and arrows indicate antigenic distance. Full genomic similarity is indicated outside the parentheses, while the percentage of genomic similarity across the eight immunogenic proteins, which are the main targets for neutralization, is specified within the parentheses. Created with BioRender. c, Plasma neutralization capacity against VACV, CWPXV, and MPXV in MPXV-infected patients (C4=25). Each dot represents a single patient. Squares represent patients that were previously vaccinated with Dryvax vaccine. d,e,f,g, Plasma neutralization capacity against VACV, CWPXV and MPXV over time following JYNNEOS vaccination. d, Neutralization titers over time in naïve, non-vaccinated individuals after the JYNNEOS regimen. e, LOWESS regression comparisons of plasma neutralization capacity against each orthopoxvirus over time in naïve, individuals, following the JYNNEOS regimen. f, Neutralization titers over time after the JYNNEOS regimen, in individuals previously vaccinated with Dryvax. e , LOWESS regression comparisons of plasma neutralization capacity against each orthopoxvirus over time following the JYNNEOS regimen, in individuals previously vaccinated with Dryvax. Regression lines are shown as pink (VACV), purple (CWPXV) and light blue (MPXV); shading represents 95% confidence interval. Significance was assessed using mixed-effect analysis corrected for multiple comparisons using by one-way ANOVA corrected for multiple comparisons using Tukey’s method. Baseline: C2=22, C3=40; 7 days post 1 st vaccine dose: C2=31, C3=16; 30-60 days post 1 st vaccine dose C2=31 C3=14; 7 days post 2 nd vaccine dose: C2=30, C3=13; 30-60 days post 2 nd vaccine dose: C2=21, C3=14; 180-240 days post 2nd vaccine dose: C2=8, C3=4. Lines indicate neutralization dynamics across vaccine regimen. Horizontal bars represent average ± SD. TP5:180+, 180-240 days. ****P < 0.0001, ***P < 0.001, **P < 0.01, and *P < 0.05.

Article Snippet: MaxiSorp plates (96 wells; 442404, Thermo Scientific) were coated with 50□μl per well of either recombinant MPox A35R (A3R-M52H3-100 μg, ACROBiosystems), MPox A29L (A2L-M52H3-100 μg, ACROBiosystems), MPox E8L (E8L-M52H3-50 μg, ACROBiosystems), MPox L1R (L1R-M5241-50 μg, ACROBiosystems), MPox H3L (H3L-M52H1-50 μg, ACROBiosystems), MPox A30L (A3L-M5243-50 μg, ACROBiosystems), VACV A33R (40896-V07E-100 μg, Sino Biological), VACV A27L (40897-V07E-100 μg, Sino Biological), VACV L1R (40903-V07H-100 μg, Sino Biological), or VACV OPC005 [vD8L] (CSB-EP322653VAA-100 μg, CusaBio) at a concentration of 2□μg/ml in PBS and were incubated overnight at 4□°C.

Techniques: Vaccines, Infection, Immunopeptidomics, Neutralization, Virus, Clinical Proteomics

Journal: medRxiv

Article Title: The impact of antigenic distance on Orthopoxvirus Vaccination and Mpox Infection for cross-protective immunity

doi: 10.1101/2024.01.31.24302065

Figure Lengend Snippet: Participants received the JYNNEOS vaccine and plasma samples were collected longitudinally at multiple points: baseline (prior to the JYNNEOS vaccination), 7-30 days after the first vaccine dose, and 7-240 days after the second dose. The participants were then categorized into two groups based on their previous vaccination history: 1) the JYNNEOS group, consisting of naïve, non-vaccinated individuals who received the JYNNEOS regimen; and 2) the Dryvax + JYNNEOS group, comprising individuals who were previously vaccinated with Dryvax and subsequently received the JYNNEOS regimen. Importantly, for the JYNNEOS cohort, baseline consists of non-vaccinated individuals. Baseline controls for the Dryvax + JYNNEOS cohort were individuals who had previously been vaccinated with Dryvax. Analysis of immunogenicity were performed using neutralization assays with authentic Vaccinia virus (VACV), Cowpox virus (CWPXV) and Mpox virus (MPXV) virus. a,b, Plasma neutralization titers against VACV, CWPXV and MPXV following JYNNEOS vaccination doses. a, Neutralization capacity in naïve, non-vaccinated individuals following JYNNEOS vaccine doses, Baseline, n=22; 1 Dose, n=62; 2 Doses=60. b, Neutralization capacity on individuals previously vaccinated with Dryvax and boosted with the JYNNEOS vaccine doses, Baseline, n=40; 1 Dose, n=30; 2 Doses=31. c, Linear regression analysis assessing the relationship between the neutralization titers against VACV and the IgG levels to VACV as determined by ELISA antigens. Regression lines are shown as blue purple (vB5R), red (vA33R), (vA27L) or green (vL1R), or Pearson’s correlation coefficients and linear regression significance are listed accordingly; shading represents 95% confidence interval. d, Linear regression analysis assessing the relationship between the neutralization titers against VACV and neutralization titers against MPXV (top) and CWPXV (bottom). Pearson’s correlation coefficients and linear regression significance are listed accordingly; shading represents 95% confidence interval. e, Plasma neutralization capacity against VACV, CWPXV and MPXV over time following JYNNEOS vaccination. f, Comparison of baseline neutralization titers with those measured at the final collection time point (180-240 days post the second vaccination dose), in both naïve, non-vaccinated individuals and those previously vaccinated with Dryvax who received the JYNNEOS vaccine regimen. JYNNEOS: Baseline=12; 180-240 days=8; Dryvax: Baseline=40; 180-240days=4. g, Neutralization capacity against VACV, CWPXV, and MPXV in participants that received the 2 nd JYNNEOS vaccine dose at the regular interval, 4-5 weeks (C2, n=12; C3, n=7) or an extended interval, 7-8 weeks (C2, n=34; C3, n=12). Significance was assessed by a non-parametric Mann-Whitney t test. Each dot represents a single individual, values chosen for this analysis were closest to peak of antibody response (7-30 days post second dose). Horizontal bars represent average ± SD. Boxes represent average ± SD. TP5:180+, 180-240 days. ****P < 0.0001, ***P < 0.001, **P < 0.01, and *P < 0.05.

Article Snippet: MaxiSorp plates (96 wells; 442404, Thermo Scientific) were coated with 50□μl per well of either recombinant MPox A35R (A3R-M52H3-100 μg, ACROBiosystems), MPox A29L (A2L-M52H3-100 μg, ACROBiosystems), MPox E8L (E8L-M52H3-50 μg, ACROBiosystems), MPox L1R (L1R-M5241-50 μg, ACROBiosystems), MPox H3L (H3L-M52H1-50 μg, ACROBiosystems), MPox A30L (A3L-M5243-50 μg, ACROBiosystems), VACV A33R (40896-V07E-100 μg, Sino Biological), VACV A27L (40897-V07E-100 μg, Sino Biological), VACV L1R (40903-V07H-100 μg, Sino Biological), or VACV OPC005 [vD8L] (CSB-EP322653VAA-100 μg, CusaBio) at a concentration of 2□μg/ml in PBS and were incubated overnight at 4□°C.

Techniques: Clinical Proteomics, Immunopeptidomics, Neutralization, Virus, Enzyme-linked Immunosorbent Assay, Comparison, MANN-WHITNEY

Journal: medRxiv

Article Title: The impact of antigenic distance on Orthopoxvirus Vaccination and Mpox Infection for cross-protective immunity

doi: 10.1101/2024.01.31.24302065

Figure Lengend Snippet: a, Comparative analysis of conserved amino acid sequences among MPXV, VACV, and CWPXV within immunogenic proteins B6R, A35R, A29L, M1R, and E8L. Amino acids highlighted in white indicate point mutations unique to VACV antigens. Amino acids highlighted in red represent point mutations exclusive to MPXV. b, Heat map analysis depicting the percentage of sequence identity between MPXV and CWPXV compared to VACV’s primary neutralization targets proteins. c,d, Structural models of Mpox antigens, based on Vaccinia antigen structures. Neutralizing antibody binding sites are highlighted in red. MPXV specific residues are highlighted in blue, and MPXV/CPXV specific residues are highlighted in purple. Unique mutations specific to MPXV are indicated by the red dots.

Article Snippet: MaxiSorp plates (96 wells; 442404, Thermo Scientific) were coated with 50□μl per well of either recombinant MPox A35R (A3R-M52H3-100 μg, ACROBiosystems), MPox A29L (A2L-M52H3-100 μg, ACROBiosystems), MPox E8L (E8L-M52H3-50 μg, ACROBiosystems), MPox L1R (L1R-M5241-50 μg, ACROBiosystems), MPox H3L (H3L-M52H1-50 μg, ACROBiosystems), MPox A30L (A3L-M5243-50 μg, ACROBiosystems), VACV A33R (40896-V07E-100 μg, Sino Biological), VACV A27L (40897-V07E-100 μg, Sino Biological), VACV L1R (40903-V07H-100 μg, Sino Biological), or VACV OPC005 [vD8L] (CSB-EP322653VAA-100 μg, CusaBio) at a concentration of 2□μg/ml in PBS and were incubated overnight at 4□°C.

Techniques: Sequencing, Neutralization, Binding Assay

Journal: medRxiv

Article Title: The impact of antigenic distance on Orthopoxvirus Vaccination and Mpox Infection for cross-protective immunity

doi: 10.1101/2024.01.31.24302065

Figure Lengend Snippet: Orthopoxviruses-reactive CD4+ and CD8+ T cells after in vitro PBMC stimulation with orthopoxviruses peptide pools (OPX) or Mpox specific peptide pool (MPX) . a, Gating strategy and representative dot plots showing the percentage of double-positive cells expressing OX40+CD138+ (AIM+) CD4+ T cells (top) and CD69+CD138+ (AIM+) CD8+ T cells (bottom). b, Analysis of AIM+ CD4+ T cells and AIM+ CD8+ T cells in individuals recently fully vaccinated with JYNNEOS compared to those vaccinated with Dryvax long ago. Significance was assessed by one-way ANOVA corrected for multiple comparisons using Tukey’s method. JYNNEOS: CD4=26; CD8=24. Dryvax: CD4=10; CD8=10. Each dot represents a single individual. Horizontal bars represent average ± SD. Stim, non-stimulated cells; PHA, phytohemagglutinin was used as positive control. ****P < 0.0001, ***P < 0.001, **P < 0.01, and *P < 0.05.

Article Snippet: MaxiSorp plates (96 wells; 442404, Thermo Scientific) were coated with 50□μl per well of either recombinant MPox A35R (A3R-M52H3-100 μg, ACROBiosystems), MPox A29L (A2L-M52H3-100 μg, ACROBiosystems), MPox E8L (E8L-M52H3-50 μg, ACROBiosystems), MPox L1R (L1R-M5241-50 μg, ACROBiosystems), MPox H3L (H3L-M52H1-50 μg, ACROBiosystems), MPox A30L (A3L-M5243-50 μg, ACROBiosystems), VACV A33R (40896-V07E-100 μg, Sino Biological), VACV A27L (40897-V07E-100 μg, Sino Biological), VACV L1R (40903-V07H-100 μg, Sino Biological), or VACV OPC005 [vD8L] (CSB-EP322653VAA-100 μg, CusaBio) at a concentration of 2□μg/ml in PBS and were incubated overnight at 4□°C.

Techniques: In Vitro, Expressing, Positive Control