Journal: Molecular Therapy. Nucleic Acids

Article Title: Antigen-dependent interplay of formulation, systemic innate responses, and antibody responses to multi-component replicon RNA vaccination

doi: 10.1016/j.omtn.2025.102595

Figure Lengend Snippet: RepRNA/LION induces minimal systemic cytokines in NHPs (A) Experimental groups: six pigtail macaques were divided into two groups and immunized with either repRNA/LNP or repRNA/LION formulations ( n = 3 per group). (B) Experimental plan: macaques were intramuscularly injected with multi-component repRNA encoding three virus-like particles (VLPs) of enterovirus D68 (EV-D68) (subclades A1, B1, and C) and two glycoproteins (gF and gG) of respiratory syncytial virus (RSV). Each repRNA (20 μg) was formulated with LNP or LION and mixed to yield a total dose of 100 μg. Immunizations were administered at weeks 0 and 8, with blood samples collected at weeks 0, 1, 3, 6, 8, and 10. (C) Serum IFN-α levels at indicated time points post-immunization, determined by ELISA. (D) Cytokine bead assay results for indicated cytokine levels in sera at baseline (week 0), 16 h post-prime (1°), and 16 h post-boost (2°). Results for repRNA/LNP (left) and repRNA/LION (right) groups are shown as a heatmap. (E–G) Nanostring analysis of PBMCs isolated at day 1 post-prime immunization compared to those isolated before immunization (pre-immune). (E) Circle chart showing the differential regulation of transcripts ( p < 0.05) in response to repRNA/LNP or repRNA/LION. (F) Volcano plots with representative upregulated (red) and downregulated (blue) genes. RNA transcripts from PBMCs isolated before immunization (pre-immune) were used as the baseline for the calculation. (G) Pathway scores calculated from RNA transcripts of PBMCs isolated from macaques receiving repRNA/LNP, relative to those receiving repRNA/LION. Scores are presented as the difference in Z-transformed values between the two groups. Positive values on the x-axis indicate pathways with higher expression in PBMCs from the LNP group compared to the LION group. Error bars indicate mean ± SD. Each dot represents the value of an individual. Statistical significance was determined by unpaired (C) and paired (F) Student’s two-tailed t test. Statistically significant results are denoted as p < 0.05∗, <0.01∗∗, <0.001∗∗∗, and <0.0001∗∗∗∗.

Article Snippet: Briefly, ELISA plates were coated with 2 μg/mL of human RSV A2 F and G proteins (Sino Biological, Cat# 11049-V08B and 40830-V08H, respectively; resources, NR-58648 and NR-59001).

Techniques: Injection, Virus, Enzyme-linked Immunosorbent Assay, Isolation, Transformation Assay, Expressing, Two Tailed Test

Journal: Molecular Therapy. Nucleic Acids

Article Title: Antigen-dependent interplay of formulation, systemic innate responses, and antibody responses to multi-component replicon RNA vaccination

doi: 10.1016/j.omtn.2025.102595

Figure Lengend Snippet: Systemic cytokine levels inversely correlated with anti-EV-D68 Ab titers (A–C) Total IgG titers (Log 10 EC50 values) in macaques immunized with repRNA/LNP (red) or repRNA/LION (green) at week 0 (dashed lines) and week 10 (solid lines), as determined by ELISA. Upper panels: mean optical density (OD) values at 450 nm. Antibody responses to specific antigens are shown: (A) RSV-G, (B) RSV-F, and (C) inactivated EV-D68. (D) Neutralizing antibody (NAb) responses against EV-D68 subclades (A1, B1, and C) in sera from macaques immunized with repRNA/LNP (red) or repRNA/LION (green). Log10 values of the 50% reciprocal NAb titers for the indicated subclades of EV-D68 are shown. (E–G) Correlation analysis results between immune parameters and antibody responses in macaques and C57BL/6 mice: (E) serum IFN-α levels at day 1 post-prime dose versus total IgG levels to RSV-G (black), RSV-F (orange) and inactivated EV-D68 (blue) at week 10 in macaques receiving repRNA/LNP or repRNA/LION. (F) Serum IFN-α levels at day 1 post-prime dose versus neutralizing titers to EV-D68 subclades (A1, B1, and C) in macaques receiving repRNA/LNP or repRNA/LION. (G) Serum IFN-α2 levels at 14 h post-prime dose versus neutralizing titers to EV-D68 subclade B2 in C57BL/6 mice. Pooled data from mice receiving monovalent, multi-component repRNA formulated with both LION and LNP are included. Each dot represents the value of an individual. Lines indicate simple linear regression results. Statistical significance was determined using Spearman correlation analysis, with significant results denoted as p < 0.05∗, <0.01∗∗, <0.001∗∗∗, and <0.0001∗∗∗∗.

Article Snippet: Briefly, ELISA plates were coated with 2 μg/mL of human RSV A2 F and G proteins (Sino Biological, Cat# 11049-V08B and 40830-V08H, respectively; resources, NR-58648 and NR-59001).

Techniques: Enzyme-linked Immunosorbent Assay

Journal: Poultry Science

Article Title: Evaluation of immunoprotective effects of PlpE multi-epitope protein incorporated within the aluminum hydroxide-adjuvanted inactivated vaccine against Pasteurella multocida infection in chickens

doi: 10.1016/j.psj.2025.105426

Figure Lengend Snippet: The antibody response to Pm in chickens after challenge. Chickens were immunized twice (0 d and 15 d). Indirect ELISA was performed to detect total IgG levels in sera collected from different groups at 0, 15, and 30 days post-immunization. Data are expressed as the means ± SD. * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001.

Article Snippet: Briefly, ELISA microtiter plates (2205, Thermo Fisher) were coated with purified recombinant PlpE antigens at concentrations of 2 μg/mL, and then blocked with 100 μL of 5 % skimmed milk (1172GR500, BioFroxx, Germany) for 2 h to prevent nonspecific binding.

Techniques: Indirect ELISA