Matching of symmetry at interfaces is a fundamental obstacle in molecular assembly. Virus‐like particles (VLPs) are important vaccine platforms against pathogenic threats, including Covid‐19. ...However, symmetry mismatch can prohibit vaccine nanoassembly. We established an approach for coupling VLPs to diverse antigen symmetries. SpyCatcher003 enabled efficient VLP conjugation and extreme thermal resilience. Many people had pre‐existing antibodies to SpyTag:SpyCatcher but less to the 003 variants. We coupled the computer‐designed VLP not only to monomers (SARS‐CoV‐2) but also to cyclic dimers (Newcastle disease, Lyme disease), trimers (influenza hemagglutinins), and tetramers (influenza neuraminidases). Even an antigen with dihedral symmetry could be displayed. For the global challenge of influenza, SpyTag‐mediated display of trimer and tetramer antigens strongly induced neutralizing antibodies. SpyCatcher003 conjugation enables nanodisplay of diverse symmetries towards generation of potent vaccines.
Nanoassembly can play a key role in generating effective vaccines for emerging (SARS‐CoV‐2) and established (influenza) pandemic threats. SpyTag‐mediated display was used to adapt the nanocage to the challenge of bacterial or viral antigens with diverse cyclic and dihedral symmetries. Establishing display of both trimeric and tetrameric antigens may lead to an influenza vaccine with broader protection.
Nanoscale organization is crucial to stimulating an immune response. Using self-assembling proteins as multimerization platforms provides a safe and immunogenic system to vaccinate against otherwise ...weakly immunogenic antigens. Such multimerization platforms are generally based on icosahedral viruses and have led to vaccines given to millions of people. It is unclear whether synthetic protein nanoassemblies would show similar potency. Here we take the computationally designed porous dodecahedral i301 60-mer and rationally engineer this particle, giving a mutated i301 (mi3) with improved particle uniformity and stability. To simplify the conjugation of this nanoparticle, we employ a SpyCatcher fusion of mi3, such that an antigen of interest linked to the SpyTag peptide can spontaneously couple through isopeptide bond formation (Plug-and-Display). SpyCatcher-mi3 expressed solubly to high yields in Escherichia coli, giving more than 10-fold greater yield than a comparable phage-derived icosahedral nanoparticle, SpyCatcher-AP205. SpyCatcher-mi3 nanoparticles showed high stability to temperature, freeze–thaw, lyophilization, and storage over time. We demonstrate approximately 95% efficiency coupling to different transmission-blocking and blood-stage malaria antigens. Plasmodium falciparum CyRPA was conjugated to SpyCatcher-mi3 nanoparticles and elicited a high avidity antibody response, comparable to phage-derived virus-like particles despite their higher valency and RNA cargo. The simple production, precise derivatization, and exceptional ruggedness of this nanoscaffold should facilitate broad application for nanobiotechnology and vaccine development.
For many infectious diseases there is still no vaccine, even though potential protective antigens have been identified. Suitable platforms and conjugation routes are urgently needed to convert the ...promise of such antigens into broadly protective and scalable vaccines. Here we apply a newly established peptide-peptide ligation approach, SnoopLigase, for specific and irreversible coupling of antigens onto an oligomerization platform. SnoopLigase was engineered from a Streptococcus pneumoniae adhesin and enables isopeptide bond formation between two peptide tags: DogTag and SnoopTagJr. We expressed in bacteria DogTag linked to the self-assembling coiled-coil nanoparticle IMX313. This platform was stable over months at 37 °C when lyophilized, remaining reactive even after boiling. IMX-DogTag was efficiently coupled to two blood-stage malarial proteins (from PfEMP1 or CyRPA), with SnoopTagJr fused at the N- or C-terminus. We also showed SnoopLigase-mediated coupling of a telomerase peptide relevant to cancer immunotherapy. SnoopLigase-mediated nanoassembly enhanced the antibody response to both malaria antigens in a prime-boost model. Including or depleting SnoopLigase from the conjugate had little effect on the antibody response to the malarial antigens. SnoopLigase decoration represents a promising and accessible strategy for modular plug-and-display vaccine assembly, as well as providing opportunities for robust nanoconstruction in synthetic biology.
Abstract Previous studies have shown promising results when using an agonistic anti-4-1BB antibody treatment against established tumors. While this is promising, this type of treatment can induce ...severe side effects. Therefore, we decided to incorporate the membrane form of 4-1BB ligand (4-1BBL) in a replicative deficient adenovirus vaccine expressing the invariant chain (Ii) adjuvant fused to a tumor associated antigen (TAA). The Ii adjuvant increases and prolongs TAA specific CD8+ T cells as previously shown and local expression of 4-1BBL was chosen to avoid the toxicity associated with systemic antibody administration. Furthermore, adenovirus encoded 4-1BBL expression has previously been successfully used to enhance responses toward Plasmodium falciparum and Influenza A antigens. We showed that the incorporation of 4-1BBL in the adenovirus vector led to surface expression of 4-1BBL on antigen presenting cells, but it did not enhance T cell responses in mice towards the Ii linked antigen. In tumor-bearing mice, our vaccine was found to decrease the frequency of TAA specific CD8+ T cells, but this difference did not alter the therapeutic efficacy. In order to reconcile our findings with the previous reports of increased anti-cancer efficacy using systemically delivered 4-1BB agonists, we incorporated a secreted version of 4-1BBL (Fc-4-1BBL) in our vaccine and co-expressed it with the Ii linked to TAA. In tumor bearing mice, this vaccine initially delayed tumor growth and slightly increased survival compared to the vaccine expressing the membrane form of 4-1BBL. Accordingly, secreted 4-1BBL co-encoded with the Ii linked antigen may offer a simplification compared to administration of drug and vaccine separately.
Adhesion G protein‐coupled receptors (aGPCRs) constitute the second largest subclass of the GPCR superfamily. Although canonical GPCRs are explored pharmacologically as drug targets, no clinically ...approved drugs target the aGPCR family so far. The aGPCR GPR56/ADGRG1 stands out as an especially promising target, given its direct link to the monogenetic disease bilateral frontoparietal polymicrogyria and implications in cancers. Key to understanding GPCR pharmacology has been mapping out intracellular signalling activity. Detection of GPCR signalling in the Gαs/Gαi/Gαq G protein pathways is feasible with second messenger detection systems. However, in the case of Gα12/13‐coupled receptors, like GPR56, signalling detection is more challenging due to the lack of direct second messenger generation. To overcome this challenge, we engineered a Gαq chimera to translate Gα12/13 signalling. We show the ability of the chimeric GαΔ6q12myr and GαΔ6q13myr to translate basal Gα12/13 signalling of GPR56 to a Gαq readout in transcription factor luciferase reporter systems and show that the established peptide ligands (P7 and P19) function to enhance this signal. We further demonstrate the ability to directly influence the generation of second messengers in inositol‐3‐phosphate assays. In the future, these chimeric G proteins could facilitate basic functional studies, drug screenings and deorphanization of other aGPCRs.
Breast cancer liver metastases (BCLM) is a common cause of breast cancer-related death. The prognostic and predictive value of receptor expression and St Gallen classification is challenged by ...receptor status discordance in distant metastases. The aim of this study was to determine the rate of receptor conversion from breast cancer to BCLM and the impact on survival.
Patients registered with BCLM in two Swedish national cancer registers were recruited retrospectively. Data on receptor expression in primary breast cancer and BCLM were collected, as well as information about predictive factors for survival. The rate of receptor and subtype conversion was analyzed. A Cox regression model was used to investigate predictive factors for survival.
A cohort of 132 patients with BCLM was identified. Estrogen receptor (ER), progesterone receptor (PgR) and HER2 converted in 17, 33 and 10%, respectively. PgR was lost in BCLM while 8/10 HER2 conversions went from negative to positive. The BC subtype was re-classified in 21% of the BCLM. Median survival after BCLM was 13 months and HER2 amplification was associated with improved survival (HR 0.28 CI 0.085-0.90). The highest predictive value (Harrell´s C-index) was obtained when including both BC and BCLM status.
Receptor and subtype conversions are common in BCLM, and a liver biopsy is warranted to tailor BCLM treatment. HER2 amplification is associated with improved survival in a BCLM cohort.
Introduction: Despite their appeal as vaccine vectors, adenoviral vectors are yet unable to induce protective immune responses against some weakly immunogenic antigens. Additionally, the maximum ...doses of adenovirus-based vaccines are limited by vector-induced toxicity, causing vector elimination and diminished immune responses against the target antigen. In order to increase immune responses to the transgene, while maintaining a moderate vector dose, new technologies for improved transgene presentation have been developed for adenoviral vaccine vectors.
Areas covered: This review provides an overview of different genetic-fusion adjuvants that aim to improve antigen presentation in the context of adenoviral vector-based vaccines. The influence on both T cell and B cell responses are discussed, with a main focus on two technologies: MHC class II-associated invariant chain and virus-like-vaccines.
Expert opinion: Different strategies have been tested to improve adenovirus-based vaccinations with varying degrees of success. The reviewed genetic adjuvants were designed to increase antigen processing and MHC presentation, or promote humoral immune responses with an improved conformational antigen display. While none of the introduced technologies is universally applicable, this review shall give an overview to identify potential improvements for future vaccination approaches.
Expression of human endogenous retrovirus type W (HERV-W) has been linked to cancer, making HERV-W antigens potential targets for therapeutic cancer vaccines. In a previous study, we effectively ...treated established tumours in mice by using adenoviral-vectored vaccines targeting the murine endogenous retrovirus envelope and group-specific antigen (Gag) of melanoma-associated retrovirus (MelARV) in combination with anti-PD-1. To break the immunological tolerance to MelARV, we mutated the immunosuppressive domain (ISD) of the MelARV envelope. However, reports on the immunogenicity of the HERV-W envelope, Syncytin-1, and its ISD are conflicting. To identify the most effective HERV-W cancer vaccine candidate, we evaluated the immunogenicity of vaccines encoding either the wild-type or mutated HERV-W envelope ISD in vitro and in vivo. Here, we show that the wild-type HERV-W vaccine generated higher activation of murine antigen-presenting cells and higher specific T-cell responses than the ISD-mutated counterpart. We also found that the wild-type HERV-W vaccine was sufficient to increase the probability of survival in mice subjected to HERV-W envelope-expressing tumours compared to a control vaccine. These findings provide the foundation for developing a therapeutic cancer vaccine targeting HERV-W-positive cancers in humans.
Human endogenous retrovirus type W (HERV-W) is expressed in various cancers. We previously developed an adenovirus-vectored cancer vaccine targeting HERV-W by encoding an assembled HERV-W ...group-specific antigen sequence and the HERV-W envelope sequence Syncytin-1. Syncytin-1 is constitutively fusogenic and forms large multinucleated cell fusions when overexpressed. Consequently, immunising humans with a vaccine encoding Syncytin-1 can lead to the formation of extensive syncytia, which is undesirable and poses a potential safety issue. Here, we show experiments in cell lines that restoring an evolutionary lost cleavage site of the fusion inhibitory R-peptide of Syncytin-1 inhibit cell fusion. Interestingly, this modification of the HERV-W vaccine’s fusogenicity increased the expression of the vaccine antigens in vitro. It also enhanced Syncytin-1-specific antibody responses and CD8+-mediated T-cell responses compared to the wildtype vaccine in vaccinated mice, with a notable enhancement in responses to subdominant T-cell epitopes but equal responses to dominant epitopes and similar rates of survival following a tumour challenge. The impairment of cell–cell fusion and the enhanced immunogenicity profile of this HERV-W vaccine strengthens the prospects of obtaining a meaningful immune response against HERV-W in patients with HERV-W-overexpressing cancers.
Virus-Like-Vaccines against HIV Andersson, Anne-Marie C; Schwerdtfeger, Melanie; Holst, Peter J
Vaccines (Basel),
02/2018, Letnik:
6, Številka:
1
Journal Article
Recenzirano
Odprti dostop
Protection against chronic infections has necessitated the development of ever-more potent vaccination tools. HIV seems to be the most challenging foe, with a remarkable, poorly immunogenic and ...fragile surface glycoprotein and the ability to overpower the cell immune system. Virus-like-particle (VLP) vaccines have emerged as potent inducers of antibody and helper T cell responses, while replication-deficient viral vectors have yielded potent cytotoxic T cell responses. Here, we review the emerging concept of merging these two technologies into virus-like-vaccines (VLVs) for the targeting of HIV. Such vaccines are immunologically perceived as viruses, as they infect cells and produce VLPs in situ, but they only resemble viruses, as the replication defective vectors and VLPs cannot propagate an infection. The inherent safety of such a platform, despite robust particle production, is a distinct advantage over live-attenuated vaccines that must balance safety and immunogenicity. Previous studies have delivered VLVs encoded in modified Vaccinia Ankara vectors and we have developed the concept into a single-reading adenovirus-based technology capable of eliciting robust CD8⁺ and CD4⁺ T cells responses and trimer binding antibody responses. Such vaccines offer the potential to display the naturally produced immunogen directly and induce an integrated humoral and cellular immune response.
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