• Vaccine Name: Pvs25 mRNA–LNP
• Target Pathogen: Plasmodium spp.
• Target Disease: Malaria
• Type: Protein Subunit Vaccine
• Status: Research
• Host Species for Licensed Use: None
• Antigen: Pvs25, the P. vivax ookinete surface protein expressed on the surface of zygotes/ookinetes and essential for the survival of ookinetes in the mosquito midgut (Kunkeaw et al., 2023)
• Pvs25 gene engineering:
  • Type: Recombinant protein preparation
  • Description: Pvs25A constructs were designed with wildtype signal peptide without the C-terminal GPI anchor, which is essential for parasite cell surface localization. Pvs25A has the wildtype sequence, and Pvs25A I130T contains the I130T substitution predominant in the Asian P. vivax isolates. Pvs25F encodes the full-length sequence of the Pvs25 gene from Sal I with wild-type signal peptide. Pvs25F I130T construct contains the full-length sequence of Pvs25 with the I130T mutation. (Kunkeaw et al., 2023)
  • Detailed Gene Information: Click Here.
• Preparation: Four nucleoside-modified Pvs25 mRNAs were designed based on the sequence of the Pvs25 gene from the reference P. vivax strain Sal I. Two constructs (Pvs25A and Pvs25A I130T) express Pvs25 with wildtype signal peptide without the C-terminal glycosylphosphatidylinositol (GPI) anchor. The other two constructs (Pvs25F and Pvs25F I130T) encode the full-length Pvs25 with its C-terminal GPI anchor. mRNAs were in vitro transcribed using T7 RNA polymerase (Megascript; Ambion) on linearized plasmids encoding mammalian codon-optimized Pvs25. mRNAs were generated to contain 101 nucleotide-long poly(A) tails and modified by replacing uridine-5′-triphosphate with m1Ψ-5′-triphosphate (TriLink BioTechnologies). mRNA capping was performed alongside transcription through the addition of a trinucleotide cap1 analog, CleanCap (TriLink), and mRNA was purified with cellulose-based purification. Purified mRNAs and poly(C) RNA (Sigma) were LNP-encapsulated using a self-assembly process where a mixture of an ionizable cationic lipid, phosphatidylcholine, cholesterol, and polyethylene glycol-lipid in ethanol was rapidly combined with an aqueous solution containing mRNA at acidic pH. (Kunkeaw et al., 2023)
• Immunization Route: Intramuscular injection (i.m.)
• Description: The Pvs25 mRNA-LNP uses nucleoside-modified Pvs25 mRNA, the P.vivax surface protein, as the vaccine antigen encapsulated in liquid nanoparticles (LNP).

Human Response

• Host Strain: Human embryonic kidney (HEK) 293 cells
• Vaccination Protocol: Pvs25 mRNA–LNPs were produced and transfected into human embryonic kidney 293 cells. (Kunkeaw et al., 2023)
• Immune Response: Western blot analysis revealed protein production from each Pvs25 mRNA. The levels of Pvs25 protein production from the two full-length constructs (Pvs25F and Pvs25F I130T) were higher than those from the truncated constructs (Pvs25A and Pvs25A I130T).

Mouse Response

• Host Strain: BALB/c mice
• Vaccination Protocol: Vaccination followed a prime-boost schedule (week 0 and 4) via intramuscular injection using three different doses (3, 10, or 30 µg). Serum from each animal was collected 4 weeks after each immunization to determine the level of anti-Pvs25 antibody by ELISA. (Kunkeaw et al., 2023)
• Immune Response: At 4 weeks after the first (prime) vaccination, the Pvs25-specific IgG induced by mRNA–LNPs was detectable with a geometric mean of reciprocal titer (GMT) value between 630–5300. After the booster dose, the antibody levels rose significantly reaching a GMT between 42,000–169,000. At each dose, the Pvs25F mRNA–LNP outperformed other formulations. The mRNA/mRNA homologous vaccination elicited the strongest memory B cell response and induced the most robust Pvs25-specific CD4+ T cell responses as measured by IFN-γ and IL-2 production of CD4+ T cells whereas this was almost absent in the protein/protein homologous (Pvs25 protein/ISA-51) vaccination group.
• Efficacy: All samples from Pvs25 mRNA–LNP-immunized mice exhibited complete (100%) transmission-blocking activity at 1:2 dilution. The percent reduction in the average oocyst number per mosquito by each serum sample (transmission-reducing activity) of these sera remained nearly complete at 1:10 dilution and were ~80% at 1:50 dilution.