A fentanyl vaccine constructed upon opsonizing antibodies specific for the Galα1-3Gal epitope.
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ABSTRACT:
SUBMITTER: Wang J
PROVIDER: S-EPMC7398699 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Ricin toxin (RT) is the second most lethal toxin known; it has been designated by the CDC as a select agent. RT is made by the castor bean plant; an estimated 50,000 tons of RT are produced annually as a by-product of castor oil. RT has two subunits, a ribotoxic A chain (RTA) and galactose-binding B chain (RTB). RT binds to all mammalian cells and once internalized, a single RTA catalytically inactivates all of the ribosomes in a cell. Administered as an aerosol, RT causes rapid lung damage and fibrosis followed by death. There are no Food and Drug Administration-approved vaccines and treatments are only effective in the first few hours after exposure. We have developed a recombinant RTA vaccine that has two mutations V76M/Y80A (RiVax). The protein is expressed in Escherichia coli and is nontoxic and immunogenic in mice, rabbits, and humans. When vaccinated mice are challenged with injected, aerosolized, or orally administered (gavaged) RT, they are completely protected. We have now developed a thermostable, aluminum-adjuvant-containing formulation of RiVax and tested it in rhesus macaques. After three injections, the animals developed antibodies that completely protected them from a lethal dose of aerosolized RT. These antibodies neutralized RT and competed to varying degrees with a panel of neutralizing and nonneutralizing mouse monoclonal antibodies known to recognize specific epitopes on native RTA. The resulting antibody competition profile could represent an immunologic signature of protection. Importantly, the same signature was observed using sera from RiVax-immunized humans.
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Project description:Human cytomegalovirus (CMV) is a highly prevalent pathogen with ~60%-90% seropositivity in adults. CMV can contribute to organ rejection in transplant recipients and is a major cause of birth defects in newborns. Currently, there are no approved vaccines against CMV. The epitope of a CMV neutralizing monoclonal antibody against a conserved region of the envelope protein gH provided the basis for a new CMV vaccine design. We exploited the influenza A virus as a vaccine platform due to the highly immunogenic head domain of its hemagglutinin envelope protein. Influenza A variants were engineered by reverse genetics to express the epitope of an anti-CMV gH neutralizing antibody that recognizes native gH into the hemagglutinin antigenic Sa site. We determined that the recombinant influenza variants expressing 7, 10, or 13 residues of the anti-gH neutralizing antibody epitope were recognized and neutralized by the anti-gH antibody 10C10. Mice vaccinated with the influenza/CMV chimeric viruses induced CMV-specific antibodies that recognized the native gH protein and inhibited virus infection. In fact, the influenza variants expressing 7-13 gH residues neutralized a CMV infection at ~60% following two immunizations with variants expressing the 13 residue gH peptide produced the highest levels of neutralization. Collectively, our study demonstrates that a variant influenza virus inserted with a gH peptide can generate a humoral response that limits a CMV infection.
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Project description:BackgroundHuman Cytomegalovirus (HCMV) is a ubiquitous herpesvirus affecting approximately 90% of the world population. HCMV causes disease in immunologically naive and immunosuppressed patients. The prevention, diagnosis and therapy of HCMV infection are thus crucial to public health. The availability of effective prophylactic and therapeutic treatments remain a significant challenge and no vaccine is currently available. Here, we sought to define an epitope-based vaccine against HCMV, eliciting B and T cell responses, from experimentally defined HCMV-specific epitopes.ResultsWe selected 398 and 790 experimentally validated HCMV-specific B and T cell epitopes, respectively, from available epitope resources and apply a knowledge-based approach in combination with immunoinformatic predictions to ensemble a universal vaccine against HCMV. The T cell component consists of 6 CD8 and 6 CD4 T cell epitopes that are conserved among HCMV strains. All CD8 T cell epitopes were reported to induce cytotoxic activity, are derived from early expressed genes and are predicted to provide population protection coverage over 97%. The CD4 T cell epitopes are derived from HCMV structural proteins and provide a population protection coverage over 92%. The B cell component consists of just 3 B cell epitopes from the ectodomain of glycoproteins L and H that are highly flexible and exposed to the solvent.ConclusionsWe have defined a multiantigenic epitope vaccine ensemble against the HCMV that should elicit T and B cell responses in the entire population. Importantly, although we arrived to this epitope ensemble with the help of computational predictions, the actual epitopes are not predicted but are known to be immunogenic.
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Project description:The Immune Epitope Database (IEDB) is a free public resource which catalogs experiments characterizing immune epitopes. To accommodate data from next generation repertoire sequencing experiments, we recently updated how we capture and query epitope specific antibodies and T cell receptors. Specifically, we are now storing partial receptor sequences sufficient to determine CDRs and VDJ gene usage which are commonly identified by repertoire sequencing. For previously captured full length receptor sequencing data, we have calculated the corresponding CDR sequences and gene usage information using IMGT numbering and VDJ gene nomenclature format. To integrate information from receptors defined at different levels of resolution, we grouped receptors based on their host species, receptor type and CDR3 sequence. As of August 2018, we have cataloged sequence information for more than 22,510 receptors in 18,292 receptor groups, shown to bind to more than 2,241 distinct epitopes. These data are accessible as full exports and through a new dedicated query interface. The later combines the new ability to search by receptor characteristics with previously existing capability to search by epitope characteristics such as the infectious agent the epitope is derived from, or the kind of immune response involved in its recognition. We expect that this comprehensive capture of epitope specific immune receptor information will provide new insights into receptor-epitope interactions, and facilitate the development of novel tools that help in the analysis of receptor repertoire data.
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Project description:Elicitation of antibodies against targets that are immunorecessive, cryptic, or transient in their native context has been a challenge for vaccine design. Here we demonstrate the elicitation of structure-specific antibodies against the HIV-1 gp41 epitope of the broadly neutralizing antibody 2F5. This conformationally flexible region of gp41 assumes mostly helical conformations but adopts a kinked, extended structure when bound by antibody 2F5. Computational techniques were employed to transplant the 2F5 epitope into select acceptor scaffolds. The resultant "2F5-epitope scaffolds" possessed nanomolar affinity for antibody 2F5 and a range of epitope flexibilities and antigenic specificities. Crystallographic characterization of the epitope scaffold with highest affinity and antigenic discrimination confirmed good to near perfect attainment of the target conformation for the gp41 molecular graft in free and 2F5-bound states, respectively. Animals immunized with 2F5-epitope scaffolds showed levels of graft-specific immune responses that correlated with graft flexibility (p < 0.04), while antibody responses against the graft-as dissected residue-by-residue with alanine substitutions-resembled more closely those of 2F5 than sera elicited with flexible or cyclized peptides, a resemblance heightened by heterologous prime-boost. Lastly, crystal structures of a gp41 peptide in complex with monoclonal antibodies elicited by the 2F5-epitope scaffolds revealed that the elicited antibodies induce gp41 to assume its 2F5-recognized shape. Epitope scaffolds thus provide a means to elicit antibodies that recognize a predetermined target shape and sequence, even if that shape is transient in nature, and a means by which to dissect factors influencing such elicitation.
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Project description:Pituitary hormones have long been thought solely to regulate single targets. Challenging this paradigm, we discovered that both anterior and posterior pituitary hormones, including FSH, had other functions in physiology. We have shown that FSH regulates skeletal integrity, and, more recently, find that FSH inhibition reduces body fat and induces thermogenic adipose tissue. A polyclonal antibody raised against a short, receptor-binding epitope of FSHβ was found not only to rescue bone loss postovariectomy, but also to display marked antiobesity and probeiging actions. Questioning whether a single agent could be used to treat two medical conditions of public health importance--osteoporosis and obesity--we developed two further monoclonal antibodies, Hf2 and Mf4, against computationally defined receptor-binding epitopes of FSHβ. Hf2 has already been shown to reduce body weight and fat mass and cause beiging in mice on a high-fat diet. Here, we show that Hf2, which binds mouse Fsh in immunoprecipitation assays, also increases cortical thickness and trabecular bone volume, and microstructural parameters, in sham-operated and ovariectomized mice, noted on microcomputed tomography. This effect was largely recapitulated with Mf4, which inhibited bone resorption by osteoclasts and stimulated new bone formation by osteoblasts. These effects were exerted in the absence of alterations in serum estrogen in wild-type mice. We also reconfirm the existence of Fshrs in bone by documenting the specific binding of fluorescently labeled FSH, FSH-CH, in vivo. Our study provides the framework for the future development of an FSH-based therapeutic that could potentially target both bone and fat.
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Project description:BackgroundTumor angiogenesis is critical for tumor growth, infiltration and metastasis. Vascular endothelial growth factor (VEGF) is a potent angiogenic factor and targeting it is important in reducing angiogenesis. Bevacizumab (Avastin), a monoclonal antibody that reacts directly against VEGF, has been demonstrated to be an effective treatment for various cancers such as rectal cancer, colon carcinoma, and non-small cell lung cancer, etc.ResultsIn the current study, we used the phage display technique to generate mimotopes that complemented the screening Avastin antibody (Ab). The candidate mimotopes of VEGF were isolated from a 12-mer peptide library. The phage displaying peptide DHTLYTPYHTHP (designated as 12P) exhibited high affinity to Avastin. The chemically synthesized 12P was conjugated to keyhole limpet hemocyanin (KLH) by glutaraldehyde (GA) to form vaccine KLH-12 peptide (KLH-12P). This epitope vaccine significantly induced humoral immunity in mice. The blood serum from KLH-12P-immunized mice associated with VEGF and blocked its binding to VEGFR, thus inhibiting vascular endothelial cell proliferation and migration.ConclusionsOur data indicate that the isolated mimotope 12P reported here could potentially elicit specific antibodies against VEGF and result in the induction of anti-angiogenesis responses.
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Project description:Active immunization is an emerging potential modality to combat fatal overdose amid the opioid epidemic. In this study, we described the design, synthesis, formulation, and animal testing of an efficacious vaccine against fentanyl. The vaccine formulation is composed of a novel fentanyl hapten conjugated to tetanus toxoid (TT) and adjuvanted with liposomes containing monophosphoryl lipid A adsorbed on aluminum hydroxide. The linker and hapten N-phenyl-N-(1-(4-(3-(tritylthio)propanamido)phenethyl)piperidin-4-yl)propionamide were conjugated sequentially to TT using amine-N-hydroxysuccinimide-ester and thiol-maleimide reaction chemistries, respectively. Conjugation was facile, efficient, and reproducible with a protein recovery of >98% and a hapten density of 30-35 per carrier protein molecule. In mice, immunization induced high and robust antibody endpoint titers in the order of >106 against the hapten. The antisera bound fentanyl, carfentanil, cyclopropyl fentanyl, para-fluorofentanyl, and furanyl fentanyl in vitro with antibody-drug dissociation constants in the range of 0.36-4.66 nM. No cross-reactivity to naloxone, naltrexone, methadone, or buprenorphine was observed. In vivo, immunization shifted the antinociceptive dose-response curve of fentanyl to higher doses. Collectively, these preclinical results showcased the desired traits of a potential vaccine against fentanyl and demonstrated the feasibility of immunization to combat fentanyl-induced effects.
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Project description:Anti-Gal is a natural antibody, which constitutes as much as 1% of circulating IgG in humans and displays a distinct specificity for the structure Gal alpha 1----3Gal. This glycosidic structure has been found on various tissues of many nonprimate mammals. A comparative study of the occurrence of anti-Gal versus the expression of the Gal alpha 1----3Gal epitope was performed in primates, and a distinct evolutionary pattern was observed. Whereas anti-Gal was found to be present in Old World monkeys and apes in titers comparable to those in humans, its corresponding antigenic epitope is abundantly expressed on erythrocytes of New World monkeys. Immunostaining with anti-Gal of glycolipids from New World monkey erythrocytes indicated that the molecules to which anti-Gal binds are similar to those found in rabbit and bovine erythrocytes. These findings indicate that there is an evolutionary reciprocity between New World and Old World primates in the production of the Gal alpha 1----3Gal structure and the antibody that recognizes it. The expression of the Gal alpha 1----3Gal epitope was evolutionarily conserved in New World monkeys, but it was suppressed in ancestral lineages of Old World primates. The suppression of this epitope was accompanied by the production of anti-Gal. The observed in vivo binding of anti-Gal to human normal senescent and some pathologic erythrocytes implies that the Gal alpha 1----3Gal epitope is present in man in a cryptic form.
| S-EPMC304431 | biostudies-literature
Project description:Previous studies demonstrated that polyclonal antibodies against foot-and-mouth disease virus (FMDV) generated by vaccination can mediate immune functions not only through virus neutralization but also through promoting virus uptake by macrophages and dendritic cells that are otherwise resistant to FMDV infection. This causes abortive infections resulting in activation, enhanced antigen presentation but also cell death. Here we report the use of RAW264.7 cells representing a murine macrophage cells line to characterize opsonizing functions of a collection of monoclonal antibodies (mAbs) against FMDV O and A serotypes. We demonstrate that all neutralizing immunoglobulin G isotype mAbs are able to opsonize FMDV resulting in increased cell death of RAW264.7 cells. In contrast, neutralizing IgM antibodies did not possess this activity. Opsonization was observed with broader reactivity within the serotype when compared to neutralization. Importantly, the anti-O serotype D9 mAb reacting with the continuous epitope within the G-H loop of VP1 that contains the RGD binding site of FMDV, opsonized several FMDV serotypes despite its restricted neutralizing activity within the O serotype. Furthermore, by generating RAW264.7 cells expressing bovine CD32, an easy-to-use cell-based assay system to test for bovine antibody-dependent enhanced infection of FMDV was generated and tested with a collection of sera. The data indicate that opsonizing titers correlated better with vaccine dose when compared to neutralizing titers. On the other hand, neutralization and opsonization titers were similar predictive of protection. We conclude that low avidity interactions are sufficient to mediate Fcγ receptor-mediated immune functions that could contribute to protective immune responses against FMDV.
| S-EPMC9597200 | biostudies-literature