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A tractable covalent linker strategy for the production of immunogenic antigen-TLR7/8L bioconjugates.

ABSTRACT: Despite the ease of production and improved safety profiles of recombinant vaccines, the inherently low immunogenicity of unadjuvanted proteins remains an impediment to their widespread adoption. The covalent tethering of TLR agonists to antigenic proteins offers a unique approach to co-deliver both constituents to the same cell-enhancing vaccine efficacy while minimizing reactogenicity. However, the paucity of simple and effective linker chemistries continues to hamper progress. Here, we present a modular, PEG-based linker system compatible with even extremely lipophilic and challenging TLR7/8 agonists. To advance the field and address previous obstacles, we offer the most straightforward and antigen-preserving linker system to date. These antigen-adjuvant conjugates enhance antigen-specific immune responses in mice, demonstrating the power of our approach within the context of modern vaccinology.

SUBMITTER: Massena CJ

PROVIDER: S-EPMC9118693 | biostudies-literature | 2021 May

REPOSITORIES: biostudies-literature

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A tractable covalent linker strategy for the production of immunogenic antigen-TLR7/8L bioconjugates.

Massena C J CJ Lathrop S K SK Davison C J CJ Schoener R R Bazin H G HG Evans J T JT Burkhart D J DJ

Chemical communications (Cambridge, England) 20210501 38

Despite the ease of production and improved safety profiles of recombinant vaccines, the inherently low immunogenicity of unadjuvanted proteins remains an impediment to their widespread adoption. The covalent tethering of TLR agonists to antigenic proteins offers a unique approach to co-deliver both constituents to the same cell-enhancing vaccine efficacy while minimizing reactogenicity. However, the paucity of simple and effective linker chemistries continues to hamper progress. Here, we presen ...[more]

PMID: 33977971

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Project description:Prostate cancer is the second leading cause of cancer-related death in men in the USA; death occurs when patients progress to metastatic castration-resistant prostate cancer (CRPC). Although immunotherapy with the Food and Drug Administration-approved vaccine sipuleucel-T, which targets prostatic acid phosphatase (PAP), extends survival for 2-4 months, the identification of new immunogenic tumor-associated antigens (TAAs) continues to be an unmet need. We evaluated the differential expression profile of castration-resistant prostate epithelial cells that give rise to CRPC from mice following an androgen deprivation/repletion cycle. The expression levels of a set of androgen-responsive genes were further evaluated in prostate, brain, colon, liver, lung, skin, kidney, and salivary gland from murine and human databases. The expression of a novel prostate-restricted TAA was then validated by immunostaining of mouse tissues and analyzed in primary tumors across all human cancer types in The Cancer Genome Atlas. Finally, the immunogenicity of this TAA was evaluated in vitro and in vivo using autologous coculture assays with cells from healthy donors as well as by measuring antigen-specific antibodies in sera from patients with prostate cancer (PCa) from a neoadjuvant clinical trial. We identified a set of androgen-responsive genes that could serve as potential TAAs for PCa. In particular, we found transglutaminase 4 (Tgm4) to be highly expressed in prostate tumors that originate from luminal epithelial cells and only expressed at low levels in most extraprostatic tissues evaluated. Furthermore, elevated levels of TGM4 expression in primary PCa tumors correlated with unfavorable prognosis in patients. In vitro and in vivo assays confirmed the immunogenicity of TGM4. We found that activated proinflammatory effector memory CD8 and CD4 T cells were expanded by monocyte-derived dendritic cell (moDCs) pulsed with TGM4 to a greater extent than moDCs pulsed with either PAP or prostate-specific antigen (PSA), and T cells primed with TGM4-pulsed moDCs produce functional cytokines following a prime/boost regiment or in vitro stimulation. An IgG antibody response to TGM4 was detected in 30% of vaccinated patients, while fewer than 8% of vaccinated patients developed antibody responses to PSA or prostate-specific membrane antigen (PSMA). These results suggest that TGM4 is an immunogenic, prostate-restricted antigen with the potential for further development as an immunotherapy target.

Project description:Current influenza vaccines have relatively low effectiveness, especially against antigenically drifted strains, the effectiveness is even lower in the elderly and immunosuppressed individuals. We have previously shown in a randomized clinical trial that the topical application of a toll-like receptor 7 agonist, imiquimod, just before intradermal influenza vaccine could expedite and augment antibody response, including to antigenically-drifted strains. However, the mechanism of this vaccine and imiquimod combination approach is poorly understood. Here, we demonstrated that imiquimod alone directly activated purified mouse peritoneal B cells. When combined with inactivated H1N1/415742Md influenza virus particle (VP) as vaccine, co-stimulation of mouse peritoneal B cells in vitro induced stronger activation, proliferation, and production of virus-antigen specific IgM and IgG. Intraperitoneal injection of a combination of VP and imiquimod (VCI) was associated with an increased number of activated B cells with enhanced expression of CD86 in the mesenteric draining lymph nodes (mesLN) and the spleen at 18 h after injection. Three days after immunization with VCI, mouse spleen showed significantly more IgM and IgG secreting cells upon in vitro re-stimulation with inactivated virus, mouse sera were detected with viral neutralizing antibody. Transfer of these spleen B cells to naïve mice improved survival after lethal dose of H1N1/415742Md challenge. More importantly, the functional response of VCI-induced B cell activation was demonstrated by early challenge with a lethal dose of H1N1/415742Md influenza virus at 3 days after immunization. The spleen and mediastinal lymph nodes (mdLN) in mice immunized with VCI had germinal center formation, and significantly higher number of plasmablasts, plasma cells, and virus-antigen specific IgM and IgG secreting cells at only 3-4 days post virus challenge, compared with those of mice that have received imiquimod, inactivated virus alone or PBS. Serum virus-specific IgG2a, IgG2b, and IgG1 and bronchoalveolar lavage fluid (BALF) virus-specific IgA at 3 or 4 days post challenge were significantly higher in mice immunized with VCI, which had significantly reduced lung viral load and 100% survival. These findings suggested that imiquimod accelerates the vaccine-induced antibody production via inducing rapid differentiation of naïve B cells into antigen-specific antibody producing cells.

Dataset Information

A tractable covalent linker strategy for the production of immunogenic antigen-TLR7/8L bioconjugates.

Publications

A tractable covalent linker strategy for the production of immunogenic antigen-TLR7/8L bioconjugates.

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