Nanoparticle vaccines can be designed to induce pDC support of mDCs for increased antigen display

Butkovich, Nina; Tucker, Jo A.; Ramírez, A. Gonzaga; Li, Enya; Meli, Vijaykumar S.; Nelson, Edward L.; Wang, Szu-Wen

doi:10.1039/d2bm01132h

Cited by 7 publications

(8 citation statements)

References 87 publications

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“…Relative to the truncated E2(D381C) mutant used previously, 23,27,29−31 the E2_152 and E2_158 mutants have an additional 20 and 14 amino acids of the wild-type protein sequence, respectively, added to their N-termini. 64,65 Introduction of 60 internal cavity cysteines allows for adjuvant conjugation. 27,66 The CBU1910 protein was recombinantly fused to E2 (Materials and Methods; Table SI-2), and the fusion proteins were expressed in E. coli (Figure SI-1).…”

Section: ■ Results and Discussionmentioning

confidence: 99%

“…The ST peptide was genetically fused to the N-terminus of E2 with a spacer sequence (Figure and Figure SI-3). The E2 mutant D381C possessed 60 internal cavity cysteines, which would enable conjugation of adjuvant. , Because a high-resolution protein structure of CBU1910 has not yet been determined, we used the protein folding prediction tool Alphafold2 to predict the structure of CBU1910 (Figure SI-4). Based on this predicted structure of N-terminal truncated CBU1910 (to enable a soluble antigen), , we decided to fuse SC to the N-terminus of CBU1910 (Figure ; Figure SI-4).…”

Section: Resultsmentioning

confidence: 99%

“…We created two E2 mutants with different N-terminal linker lengths and 60 internal cavity cysteines [E2_152(D381C) and E2_158(D381C)]. Relative to the truncated E2(D381C) mutant used previously, ,,− the E2_152 and E2_158 mutants have an additional 20 and 14 amino acids of the wild-type protein sequence, respectively, added to their N-termini. , Introduction of 60 internal cavity cysteines allows for adjuvant conjugation. , …”

Section: Resultsmentioning

confidence: 99%

“…We note that this result is different than attachment of HA to E2, which had yielded reliable conjugation and stable, monodisperse particles, suggesting that these effects are highly antigen-dependent. Furthermore, the use of the maleimide-tNTA for surface conjugation via cysteine residues limits the use of cysteines internally for conjugation of immune-stimulating adjuvants. ,− , …”

Section: Resultsmentioning

confidence: 99%

“…Previously established E2 mutants E2_152 and E2_158 were used to engineer CBU1910-E2 fusion constructs. 64,65 D381C is an E2 mutation that introduces 60 cysteines to the internal cavity of the nanoparticle, allowing for internal conjugation. 23,27 To introduce the D381C mutation to E2_158 and E2_152 via site directed mutagenesis (SDM) 89,90 the forward primer: 5′-/5Phos/GCCGATCGTTCGTTGCGGTGAAA-TCGTTGC-3′ and reverse primer: 5′-/5Phos/TTTTCGGC-TATACGACCAATACCCAG-3′ were used.…”

Section: ■ Materials and Methodsmentioning

confidence: 99%

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Engineering Protein Nanoparticles Functionalized with an Immunodominant Coxiella burnetii Antigen to Generate a Q Fever Vaccine

Ramirez,

Felgner,

Jain

et al. 2023

Bioconjugate Chem.

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Coxiella burnetii is the causative agent of Q fever, for which there is yet to be an FDA-approved vaccine. This bacterial pathogen has both extra- and intracellular stages in its life cycle, and therefore both a cell-mediated (i.e., T lymphocyte) and humoral (i.e., antibody) immune response are necessary for effective eradication of this pathogen. However, most proposed vaccines elicit strong responses to only one mechanism of adaptive immunity, and some can either cause reactogenicity or lack sufficient immunogenicity. In this work, we aim to apply a nanoparticle-based platform toward producing both antibody and T cell immune responses against C. burnetii. We investigated three approaches for conjugation of the immunodominant outer membrane protein antigen (CBU1910) to the E2 nanoparticle to obtain a consistent antigen orientation: direct genetic fusion, high affinity tris-NTA-Ni conjugation to polyhistidine-tagged CBU1910, and the SpyTag/SpyCatcher (ST/SC) system. Overall, we found that the ST/SC approach yielded nanoparticles loaded with the highest number of antigens while maintaining stability, enabling formulations that could simultaneously co-deliver the protein antigen (CBU1910) and adjuvant (CpG1826) on one nanoparticle (CBU1910-CpG-E2). Using protein microarray analyses, we found that after immunization, antigen-bound nanoparticle formulations elicited significantly higher antigen-specific IgG responses than soluble CBU1910 alone and produced more balanced IgG1/IgG2c ratios. Although T cell recall assays from these protein antigen formulations did not show significant increases in antigen-specific IFN-γ production compared to soluble CBU1910 alone, nanoparticles conjugated with a CD4 peptide epitope from CBU1910 generated elevated T cell responses in mice to both the CBU1910 peptide epitope and whole CBU1910 protein. These investigations highlight the feasibility of conjugating antigens to nanoparticles for tuning and improving both humoral- and cell-mediated adaptive immunity against C. burnetii.

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Section: ■ Results and Discussionmentioning

confidence: 99%