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Long-term generation of antiPCSK9 antibody using a nanoliposome-based vaccine delivery system

  • Amir Abbas Momtazi-Borojeni
    Affiliations
    Nanotechnology Research Center, Bu-Ali Research Institute, Mashhad University of Medical Sciences, Mashhad, Iran

    Department of Medical Biotechnology, Student Research Committee, Faculty of Medicine, Mashhad University of Medical Sciences, Mashhad, Iran
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  • Mahmoud Reza Jaafari
    Correspondence
    Corresponding author. Department of Pharmaceutical Nanotechnology, School of Pharmacy, Mashhad University of Medical Sciences, Mashhad 91775-1365, Iran
    Affiliations
    Nanotechnology Research Center, Pharmaceutical Technology Institute, Mashhad University of Medical Sciences, Mashhad, Iran

    Biotechnology Research Center, Pharmaceutical Technology Institute, Mashhad University of Medical Sciences, Mashhad, Iran
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  • Ali Badiee
    Affiliations
    Nanotechnology Research Center, Pharmaceutical Technology Institute, Mashhad University of Medical Sciences, Mashhad, Iran

    Biotechnology Research Center, Pharmaceutical Technology Institute, Mashhad University of Medical Sciences, Mashhad, Iran
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  • Amirhossein Sahebkar
    Correspondence
    Corresponding author. Department of Medical Biotechnology, Mashhad University of Medical Sciences, Mashhad 91779-48564, Iran
    Affiliations
    Biotechnology Research Center, Pharmaceutical Technology Institute, Mashhad University of Medical Sciences, Mashhad, Iran

    Neurogenic Inflammation Research Center, Mashhad University of Medical Sciences, Mashhad, Iran

    School of Pharmacy, Mashhad University of Medical Sciences, Mashhad, Iran
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      Highlights

      • PCSK9 inhibition is a pivotal approach for LDL cholesterol-lowering.
      • PCSK9 inhibition using monoclonal antibodies is short-term and not cost-effective.
      • We tested the efficacy of a nanoliposomal PCSK9-specific active vaccine.
      • The tested nanoliposomal vaccine induced humoral immunity against PCSK9 in BALB/c mice.
      • The tested vaccine was also safe and induced long-term generation of anti-PCSK9 antibodies.

      Abstract

      Background and aims

      Proprotein convertase subtilisin kexin type 9 (PCSK9) is a liver secretory enzyme that controls plasma low-density lipoprotein cholesterol (LDL-C) levels through modulation of LDL receptor (LDLR). Inhibition of PCSK9 using monoclonal antibodies (mAbs) can efficiently lower plasma LDL-C. However, the relatively short half-life of mAbs necessitates frequent passive immunization, which is costly. These limitations can be circumvented by active immunization. Here, we evaluated the long-term antiPCSK9 antibody generation in BALB/c mice vaccinated with a nanoliposomal PCSK9-specific active vaccine.

      Methods

      Negatively charged nanoliposomes were used as a vaccine delivery system and prepared via lipid-film hydration method. We constructed a peptide vaccine termed Immunogenic Fused PCSK9-Tetanus (IFPT) by linking a short PCSK9 peptide (as B cell epitope) to a tetanus peptide (as T cell epitope). The IFPT peptide was conjugated to the surface of nanoliposome carriers using a DSPE-PEG- Maleimide (1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[maleimide(PEG)-2000]) linker. Nanoliposomal IFPT (L-IFPT) construct was formulated with alum vaccine adjuvant (L-IFPTA+). To evaluate induction of antiPCSK9 antibody in vivo, BALB/c mice were subcutaneously inoculated four times in bi-weekly intervals with prepared vaccine formulations, including L-IFPT, L-IFPTA+, IFPTA+, IFPT, and empty liposomes as negative control. The long-term efficacy of antiPCSK9 antibodies was evaluated over 48 weeks after prime inoculation. Specificity of generated antiPCSK9 antibodies was assessed using ELISA method. To evaluate immunogenic safety, production of IL-4 and IFN-γ, and population of CD8+ and CD4+ T cells in splenic cells isolated from the vaccinated mice were analyzed.

      Results

      The L-IFPTA+ vaccine was found to elicit the highest IgG antibody response against PCSK9 peptide in the vaccinated mice, when compared with the other vaccine formulations. Antibody titer analyses over 48 weeks post-prime vaccination revealed that the L-IFPTA+ vaccine was able to stimulate a long-lasting humoral immune response against PCSK9 peptide, and thereby decrease plasma PCSK9. Generated antibodies could specifically target PCSK9 and thereby inhibit PCSK9-LDLR interaction. Analysis of splenic cells showed that the population of anti-inflammatory CD4+ Th2 cells and production and secretion of IL-4 cytokine were increased in mice vaccinated with the L-IFPTA+ vaccine, while population of inflammatory CD4+ Th1 cell and cytotoxic CD8+ T cells as well as production and secretion of IFN-γ were not altered.

      Conclusions

      The results indicate efficient activity of the tested nanoliposomal construct (L-IFPTA+) to induce humoral immune response against PCSK9 in BALB/c mice. L-IFPTA+ vaccine can induce immunogenic-safe and long-term generation of antiPCSK9 antibodies in BALB/c mice.

      Graphical abstract

      Keywords

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