Accelerating the clinical development of protein-based vaccines for malaria by efficient purification using a four amino acid C-terminal ‘C-tag’

Jin, Jing; Hjerrild, Kathryn A.; Silk, Sarah E.; Brown, Rebecca; Labbé, Geneviève; Marshall, Joyce; Wright, Katherine E.; Bezemer, Sandra; Clemmensen, Stine; Biswas, Sumi; Li, Yuanyuan; El-Turabi, Aadil; Douglas, Alexander D.; Hermans, Pim; Detmers, Frank; Jongh, Willem A. de; Higgins, Matthew K.; Ashfield, Rebecca; Draper, Simon J.

doi:10.1016/j.ijpara.2016.12.001

Cited by 63 publications

(71 citation statements)

References 57 publications

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“…The ability of RH5.1 protein to bind to recombinant basigin was subsequently analyzed by SPR, which measured an affinity of 1.2 µM, consistent with previously reported measurements of K D = 1–2 µM 21 , 28 , 33 , 35 (Fig. 3a ).…”

Section: Resultssupporting

confidence: 84%

“…We have previously reported the production of preclinical-grade full-length PfRH5 protein vaccines using polyclonal Drosophila S2 stable cell lines. 33 , 35 For cGMP biomanufacture, we designed a final protein variant, termed “RH5.1” (Fig. 1a ).…”

Section: Resultsmentioning

confidence: 99%

“… 33 , 34 Full-length PfRH5 protein was expressed from stable S2 cell lines and secreted into the supernatant from where it was purified using a newly available affinity purification system that makes use of a C-terminal tag known as “C-tag,” composed of the four amino acids (aa) glutamic acid–proline–glutamic acid–alanine (E-P-E-A). 35 This C-tag is selectively captured on a resin coupled to a camelid single-chain antibody specific for this short sequence 36 that has now been developed into a CaptureSelect™ affinity resin by Thermo Fisher Scientific.…”

Section: Introductionmentioning

confidence: 99%

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Production, quality control, stability, and potency of cGMP-produced Plasmodium falciparum RH5.1 protein vaccine expressed in Drosophila S2 cells

et al. 2018

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Plasmodium falciparum reticulocyte-binding protein homolog 5 (PfRH5) is a leading asexual blood-stage vaccine candidate for malaria. In preparation for clinical trials, a full-length PfRH5 protein vaccine called “RH5.1” was produced as a soluble product under cGMP using the ExpreS2 platform (based on a Drosophila melanogaster S2 stable cell line system). Following development of a high-producing monoclonal S2 cell line, a master cell bank was produced prior to the cGMP campaign. Culture supernatants were processed using C-tag affinity chromatography followed by size exclusion chromatography and virus-reduction filtration. The overall process yielded >400 mg highly pure RH5.1 protein. QC testing showed the MCB and the RH5.1 product met all specified acceptance criteria including those for sterility, purity, and identity. The RH5.1 vaccine product was stored at −80 °C and is stable for over 18 months. Characterization of the protein following formulation in the adjuvant system AS01B showed that RH5.1 is stable in the timeframe needed for clinical vaccine administration, and that there was no discernible impact on the liposomal formulation of AS01B following addition of RH5.1. Subsequent immunization of mice confirmed the RH5.1/AS01B vaccine was immunogenic and could induce functional growth inhibitory antibodies against blood-stage P. falciparum in vitro. The RH5.1/AS01B was judged suitable for use in humans and has since progressed to phase I/IIa clinical trial. Our data support the future use of the Drosophila S2 cell and C-tag platform technologies to enable cGMP-compliant biomanufacture of other novel and “difficult-to-express” recombinant protein-based vaccines.

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Section: Resultssupporting

confidence: 84%

Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Production, quality control, stability, and potency of cGMP-produced Plasmodium falciparum RH5.1 protein vaccine expressed in Drosophila S2 cells

et al. 2018

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“…The concentrated supernatant was then loaded onto CaptureSelect™ C-tag affinity column (Thermo Fisher Scientific) equilibrated in Tris-buffered saline and bound proteins were eluted with 20 mM Tris–HCl, 2 M MgCl 2 , pH 7.4. Fractions containing Psf48/45 were then pooled, concentrated and subjected to size-exclusion chromatography using a Superdex 200 16/60 PG column (GE Healthcare) 33 . 10C and 6C truncation variants were cloned, expressed and purified as described above.…”

Section: Methodsmentioning

confidence: 99%

Structural basis for recognition of the malaria vaccine candidate Pfs48/45 by a transmission blocking antibody

et al. 2018

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The quest to develop an effective malaria vaccine remains a major priority in the fight against global infectious disease. An approach with great potential is a transmission-blocking vaccine which induces antibodies that prevent establishment of a productive infection in mosquitos that feed on infected humans, thereby stopping the transmission cycle. One of the most promising targets for such a vaccine is the gamete surface protein, Pfs48/45. Here we establish a system for production of full-length Pfs48/45 and use this to raise a panel of monoclonal antibodies. We map the binding regions of these antibodies on Pfs48/45 and correlate the location of their epitopes with their transmission-blocking activity. Finally, we present the structure of the C-terminal domain of Pfs48/45 bound to the most potent transmission-blocking antibody, and provide key molecular information for future structure-guided immunogen design.

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“…Arginine is strong protein binders which stabilize protein-protein interaction from aggregation, especially during the process of folding (39). These four amino acids have been used for the induction of protein binding ability of peptides in many vaccine preparation against broad range of diseases including cancer, pneumonia, and malaria (40)(41)(42)(43). The introduction of cysteine zipper in the candidate vaccine leads to formation of inter-promoter disulfide rings that enabled stable coiledcoil trimers, generating satisfactory immunogenicity against hRSV (44).…”

Section: Discussionmentioning

confidence: 99%

An immunoinformatic approach to design a novel vaccine against the human respiratory syncytial virus (hRSV) by targeting M2-1 protein

Momtaz¹,

Foysal²

2019

Af J Clin Exp Micro

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Background: Human respiratory syncytial virus (hRSV) is the leading cause of upper and lower respiratory infection in infants, adults and immunocompromised persons. The matrix protein, M2-1 of hRSV is a cofactor of viral RNA polymerase that plays a crucial role during replication. This programmed study was designed to scrutinize potential immunogens from the M2-1 protein characterized from four different continents. Methods: Sequence data obtained from NCBI databases were analysed by using a series of web and software based bioinformatics tools to find out the best epitope against hRSV. Results: The phylogenetic data revealed a homogenized clustering of M2-1 protein for the African, European, and Asian clades while proteins from North American collections found to have a significant evolutionary detachment compared to three other clusters. Using various web-based bioinformatics tools, the study identified four common B-cell epitopes present in all the M2-1 proteins from four different clusters with higher antigenicity and conservancy. Among the 17 M2-1 protein investigated for T-cell epitopes, "VLQNLDVGL" peptide from A2 super-type, and "QSACVAMSK" and "CLNGRRCHY" from A3 super-type showed the highest antigenicity at >0.80 conservancy cutoff value. After evaluation of all antigenic properties, only "CLNGRRCHY" peptide qualified as a potential vaccine candidate against hRSV. Molecular docking revealed strong and stable binding of the epitope to major histocompatibility complexes (MHC) molecules in terms of hydrogen bonding. Conclusion: The designed epitope could be used as a possible vaccine candidate against hRSV.

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Accelerating the clinical development of protein-based vaccines for malaria by efficient purification using a four amino acid C-terminal ‘C-tag’

Abstract: Graphical abstract

Cited by 63 publications

References 57 publications

Production, quality control, stability, and potency of cGMP-produced Plasmodium falciparum RH5.1 protein vaccine expressed in Drosophila S2 cells

Production, quality control, stability, and potency of cGMP-produced Plasmodium falciparum RH5.1 protein vaccine expressed in Drosophila S2 cells

Structural basis for recognition of the malaria vaccine candidate Pfs48/45 by a transmission blocking antibody

An immunoinformatic approach to design a novel vaccine against the human respiratory syncytial virus (hRSV) by targeting M2-1 protein

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