EP9158H: An Immunoinformatics-Designed mRNA Vaccine Encoding Multi-Epitope Antigens and Dual TLR Agonists for Tuberculosis Prevention
Zhang M. Ali S.L. Tian Y. Abduldayeva A. Zhou S. An Y. Li Y. Ni R. Zhang L. Liu Y. Sun W. Gong W.
December 2025Multidisciplinary Digital Publishing Institute (MDPI)
Bioengineering
2025#12Issue 12
Background: Tuberculosis (TB) remains a pressing global health crisis. The inadequate efficacy of the BCG vaccine against adult pulmonary TB underscores the urgent need for novel, effective vaccines. This study aimed to design a novel mRNA vaccine candidate against TB using a rational immunoinformatics approach. Methods: From 13 antigens, >12,000 epitopes were filtered to select 60 optimal peptides (36 CTL, 16 HTL, 8 B-cell), assembled into 25 scaffolds with 49 TLR2/4 agonist configurations. EP9158H underwent structural modeling, 100 ns molecular dynamics, docking, immune simulation, RNAfold, and conservation analysis across 76 strains. Results: EP9158H, encoding 15 CTL, 9 HTL, and 8 B-cell epitopes flanked by TLR2 agonist ESAT-6 and TLR4 agonist HBHA, emerged as the optimal candidate. All 32 constituent epitopes showed >81% conservation, with 81.25% exhibiting perfect identity across MTBC lineages. The scaffold demonstrated high solubility (0.531), broad population coverage (73.76% MHC-I, 88.91% MHC-II), optimal TLR2/4 docking scores (−1359.7 and −1348.3), and robust structural stability (ProSA Z-score −6.18; RMSD 22–27 Å). Immune simulation predicted strong Th1-biased T-cell responses and high levels of antibody titers. RNAfold analysis revealed stable mRNA secondary structures (MFE −1127.5 kcal/mol) supporting efficient translation. Conclusions: EP9158H integrates broad epitope coverage, dual TLR agonism, and validated stability. Compared to single-antigen vaccines, it offers superior strain coverage, enhanced innate activation, and mRNA advantages for CTL induction, warranting experimental validation.
immune simulation , immunoinformatics , molecular dynamics , mRNA vaccine , TLR agonist , tuberculosis
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Senior Department of Tuberculosis, Chinese PLA General Hospital, Beijing, 100091, China
Graduate School, Hebei North University, Zhangjiakou, 075000, China
Department of Biochemistry, Abdul Wali Khan University, Mardan, 23200, Pakistan
Department of Research Institute of Preventive Medicine Named Academician E. Dalenov, Astana Medical University, Astana, 010000, Kazakhstan
Department of Geriatrics, The Eighth Medical Center of PLA General Hospital, Beijing, 100091, China
Senior Department of Tuberculosis
Graduate School
Department of Biochemistry
Department of Research Institute of Preventive Medicine Named Academician E. Dalenov
Department of Geriatrics
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