Insertion of foreign epitopes in HBcAg: how to make the chimeric particle assemble

Karpenko, Larisa I.; Иванисенко, В. А.; Pika, I. A.; Chikaev, N. A.; Eroshkin, Alexey; Veremeiko, T. A.; Ilyichev, A.

doi:10.1007/s007260070072

Cited by 45 publications

(39 citation statements)

References 16 publications

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“…Second, the localization of the targeted epitope close to the cell membrane was regarded as beneficial for the Fc-mediated cytolytic activity of induced antibodies (26,27). Third, biophysical and biochemical properties had to facilitate VLP assembly (28). Fourth, since we wanted to evaluate these VLPs in mice and rabbits as proof-ofconcept for subsequent clinical testing in humans, the epitope had to be conserved across these species.…”

Section: Resultsmentioning

confidence: 99%

Highly Specific Auto-Antibodies against Claudin-18 Isoform 2 Induced by a Chimeric HBcAg Virus-Like Particle Vaccine Kill Tumor Cells and Inhibit the Growth of Lung Metastases

et al. 2011

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Strategies for antibody-mediated cancer immunotherapy, such as active immunization with virus-like particle (VLP)-based vaccines, are gaining increasing attention. We developed chimeric hepatitis B virus core antigen (HBcAg)-VLPs that display a surface epitope of the highly selective tumor-associated cell lineage marker claudin-18 isoform 2 (CLDN18.2) flanked by a mobility-increasing linker. Auto-antibodies elicited by immunization with these chimeric HBcAg-VLPs in 2 relevant species (mouse and rabbit) bind with high precision to native CLDN18.2 at physiologic densities on the surface of living cells but not to the corresponding epitope of the CLDN18.1 splice variant that differs by merely one amino acid. The induced auto-antibodies are capable of efficiently killing CLDN18.2 expressing cells in vitro by complement-dependent and antibody-dependent cellmediated cytotoxicity. Moreover, they provide partial protective immunity against the challenge of mice with syngeneic tumor cells stably expressing CLDN18.2. Our study provides a first proof-of-concept that immunization combining VLPs as antigen carriers with specific conformational epitopes of a highly selective differentiation antigen may elicit auto-antibodies with high cytocidal and tumoricidal potential. Cancer Res; 71(2); 516-27. Ó2011 AACR.

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Section: Resultsmentioning

confidence: 99%

Highly Specific Auto-Antibodies against Claudin-18 Isoform 2 Induced by a Chimeric HBcAg Virus-Like Particle Vaccine Kill Tumor Cells and Inhibit the Growth of Lung Metastases

et al. 2011

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“…Antigen modules are fused in the region of the viral capsid protein that is exposed on the surface, including the N-and C-termini . Karpenko et al 2000, Loktev et al 1996. Nevertheless, the production of modular VLPs, in which antigen modules are genetically fused into the viral capsid proteins, is highly unpredictable, depending on (i) the interaction between antigen modules and the viral capsid proteins, (ii) glycosylation efficacy, (iii) the expression system, and (iv) the length of the antigen module.…”

Section: Virus-like Particles (Vlps)mentioning

confidence: 99%

“…A hydrophobic stretch has been known to cause formation of insoluble aggregates, as well as incorrect folding of proteins. Although the negative effects of a hydrophobic stretch 8 have been widely recognised (Aleksaitė and Gedvilaitė 2006, Chiti et al 2003, Esler et al 1996, Karpenko et al 2000, Kazaks et al 2004, Otzen et al 2000, approaches to minimise its effects on VLP assembly still rely on simultaneous expression of modular and unmodularised viral capsid proteins, which yields mosaic VLPs (Karpenko et al 2000, Loktev et al 1996.Various approaches have been commonly utilised to improve the solubility of proteins. These approaches can be classified into two classes, i.e.…”

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confidence: 99%

“…Additionally, irregular aggregates were also observed (Aleksaitė and Gedvilaitė 2006). Similarly, the study by Karpenko et al (Karpenko et al 2000) showed that when an antigen module with a high hydrophobicity score was modularised into Hepatitis B core protein (HBcAg), the modular protein could not assemble into VLPs. They suggested that the hydrophobic antigen module was likely to fold in such a way that it blocked the contact region between the subunits of HBcAg dimer.…”

mentioning

confidence: 99%

“…Although the influences of hydrophobic antigen modules are known, approaches to minimise these impacts are still limited to the use of mosaic VLPs (Karpenko et al 2000, Kazaks et al 2004. Mosaic VLPs are VLPs composed of a modular capsid protein and sterically less challenging capsid protein, e.g.…”

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confidence: 99%

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Engineering of virus-like particles for alternative vaccine candidate targeting a hypervariable peptide antigen element

Anggraeni¹

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A promising alternative to replace the current egg-or cell culture-based technology for vaccine production from live viruses is virus-like particle (VLP) technology based on a microbial platform. VLPs are macromolecular assemblies of viral capsid proteins, which have been shown to tolerate insertion of antigen modules via genetic recombinant technology, yielding modular VLPs. Many studies on modular VLPs presume that when a peptide antigen element is taken out from the intact proteins and then modularised on VLPs, it is unable to fold into its native structure. However, until now, presentation of a peptide antigen element on a VLP and the impact of the display strategy to present the antigen element on the quality of the resulting antibodies (i.e. the ability of the antibodies to recognise the intact protein) are not fully understood. This thesis aims to understand the underlying fundamentals regarding modularisation of peptide antigen elements on VLPs for induction of high-quality antibodies. A hypervariable receptor-binding domain, Helix 190 (H190), from the hemagglutinin protein of influenza A virus was used as a model for modularisation on VLPs from murine polyomavirus (MuPyV) VP1 protein. Four major findings are presented. Firstly, two display strategies, i.e. arraying of H190 in tandem repeats and the use of helix promoter elements, were shown to display H190 in its immunogenic form equally. However, modularisation using tandem repeat display induced antibodies of a higher quality than modularisation using helix promoter elements.Secondly, the quality of antibodies induced by the tandem repeat display bearing two copies of H190 was optimum, thus no significant improvement was observed following the use of adjuvant or increasing the copy number of H190. Additionally, the increase in the copy number of H190 was shown to reduce the assembly capability and solubility of modular VP1 in an environment that was optimised for wild-type VP1. Thirdly, this thesis shows the novel finding in the use of flanking ionic elements to stabilise VLP precursors, termed as capsomeres, bearing two copies of H190 containing a hydrophobic stretch, which caused aggregation. Fourthly, the first steps towards obtaining the atomic crystal structure of presented H190 on a modular protein were performed, i.e. a mild and satisfactory laboratory process was developed to achieve high-purity modular VP1 capsomeres, unattainable using previously established expression and purification process of wild-type MuPyV VP1. This thesis shows a step forward towards understanding the presentation of a peptide antigen element on a VLP that enables induction of highquality antibodies, and towards VLP engineering to manipulate the aggregation and solubility of modular VP1. VLP technology based on a microbial platform presented here is iii a potentially safe and effective alternative vaccine candidate that targets a hypervariable peptide antigen element. The speed of the microbial platform allows a rapid response to the hypervariability of the pe...

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Melanoma vaccine candidates from chimeric hepatitis B core virus‐like particles carrying a tumor‐associated MAGE‐3 epitope

Kazāks

Balmaks

Voronkova

et al. 2008

Biotechnology Journal

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Vaccination of melanoma patients with tumor-specific antigens recognized by cytotoxic T lymphocytes (CTLs) may produce significant tumor regressions. Here, we suggest a novel type of tumor vaccines, with well-studied CTL epitopes presented on highly immunogenic virus-like particle (VLP) carriers. Cancer-germline gene MAGE-3 encodes for an antigenic nonapeptide (MAGE-3(168-176) peptide) that is recognized by CTLs on human leukocyte antigen (HLA)-A1 and HLA-B35 molecules. A set of recombinant genes encoding hepatitis B virus core protein carrying MAGE-3 epitope was constructed and expressed in Escherichia coli cells. Variants that led to formation of chimeric VLPs in vivo were purified and analyzed for their DNA binding properties in vitro. VLPs exhibiting the most pronounced nucleic acid binding affinity were selected and loaded either with single-stranded DNA oligodeoxynucleotides rich in nonmethylated CG motifs, or with longer double-stranded DNA fragments. Packaged DNA was protected, at least partially, against the action of bacterial DNase. Such highly purified chimeric VLPs with entrapped immunomodulatory sequences could possibly be used as antitumor vaccines.

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Insertion of foreign epitopes in HBcAg: how to make the chimeric particle assemble

Cited by 45 publications

References 16 publications

Highly Specific Auto-Antibodies against Claudin-18 Isoform 2 Induced by a Chimeric HBcAg Virus-Like Particle Vaccine Kill Tumor Cells and Inhibit the Growth of Lung Metastases

Highly Specific Auto-Antibodies against Claudin-18 Isoform 2 Induced by a Chimeric HBcAg Virus-Like Particle Vaccine Kill Tumor Cells and Inhibit the Growth of Lung Metastases

Engineering of virus-like particles for alternative vaccine candidate targeting a hypervariable peptide antigen element

Melanoma vaccine candidates from chimeric hepatitis B core virus‐like particles carrying a tumor‐associated MAGE‐3 epitope

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