BackgroundOver 14 million people die each year from infectious diseases despite extensive vaccine use [1]. The needle and syringe—first invented in 1853—is still the primary delivery device, injecting liquid vaccine into muscle. Vaccines could be far more effective if they were precisely delivered into the narrow layer just beneath the skin surface that contains a much higher density of potent antigen-presenting cells (APCs) essential to generate a protective immune response. We hypothesized that successful vaccination could be achieved this way with far lower antigen doses than required by the needle and syringe.Methodology/Principal FindingsTo meet this objective, using a probability-based theoretical analysis for targeting skin APCs, we designed the Nanopatch™, which contains an array of densely packed projections (21025/cm2) invisible to the human eye (110 µm in length, tapering to tips with a sharpness of <1000 nm), that are dry-coated with vaccine and applied to the skin for two minutes. Here we show that the Nanopatches deliver a seasonal influenza vaccine (Fluvax® 2008) to directly contact thousands of APCs, in excellent agreement with theoretical prediction. By physically targeting vaccine directly to these cells we induced protective levels of functional antibody responses in mice and also protection against an influenza virus challenge that are comparable to the vaccine delivered intramuscularly with the needle and syringe—but with less than 1/100th of the delivered antigen.Conclusions/SignificanceOur results represent a marked improvement—an order of magnitude greater than reported by others—for injected doses administered by other delivery methods, without reliance on an added adjuvant, and with only a single vaccination. This study provides a proven mathematical/engineering delivery device template for extension into human studies—and we speculate that successful translation of these findings into humans could uniquely assist with problems of vaccine shortages and distribution—together with alleviating fear of the needle and the need for trained practitioners to administer vaccine, e.g., during an influenza pandemic.
We have identified a major T-cell epitope, amino acids [48][49][50][51][52][53][54] in the E7 open reading frame protein of human papillomavirus (HPV) type 16. Lymph node cells from mice immunized with synthetic peptides containing DRAHYNI proliferated and produced interleukin when challenged in vitro with peptide or whole HPV-16 E7 fusion protein. The T epitope was recognized in association with all five major histocompatibility complex class II I-A and I-E alleles tested. Synthetic peptides consisting of DRAHYNI linked to major B-cell epitopes on the E7 molecule formed immunogens capable of eliciting strong antibody responses to HPV-16 E7. The T epitope could provide help for the production of antibody to several B epitopes simultaneously, including a B epitope of HPV-18 E7 protein. Mice immunized with a peptide containing DRAHYNI and B epitope and, at a later date, infected with recombinant vaccinia E7 virus, displayed secondary antibody responses to E7. Because E7 has a role in cell transformation and is the most abundant viral protein in HPV-associated neoplastic cervical epithelial cells, the data have implications for vaccine strategies.Circumstantial evidence implicates host immune mechanisms in the control of human papillomavirus (HPV)-associated tumors of the anogenital epithelium (1), and there is an increased risk of squamous cell carcinoma of the cervix and vulva but not of control organs, such as breast and rectum, in immunosuppressed allograft recipients (2). E7 is the most abundant viral protein in HPV-16-containing CaSki and SiHa squamous carcinoma cell lines and in HPV-18-containing HeLa and C4-1 lines (3). DNA-transfection experiments implicate the E7 open reading frame protein in in vitro transformation of mouse fibroblasts (4), rat epithelial cells (5), and primary human keratinocytes (6). Cooperation with an active ras oncogene leads to full transformation (5), and there is a requirement for continued expression of the E7 gene to maintain the transformed phenotype (7). The E7 protein may be immunogenic after infection with HPV, as anti-E7 antibodies have been described in the serum of "20% of patients with HPV-16-associated cervical lesions (8, 9).These observations suggest that the E7 protein merits consideration as a candidate antigen for a potential vaccine against HPV infection. We have recently described the major immunodominant B epitopes in HPV-16 E7 (10) and HPV-18 E7 (11) defined by monoclonal antibodies. In this study we have defined, using synthetic peptides, a major T helper (Th) epitope in HPV-16 E7.
MATERIALS AND METHODSSynthetic Peptides. Peptides were synthesized by using derivatized N-tert-butoxycarbonyl (t-Boc) amino acids on benzhydryl resin (12) or using 9-fluorenylmethoxycarbonyl (Fmoc) chemistry on an Applied Biosystems 431A peptide synthesizer. The amino acid composition, toxicity, and mitogenicity of all peptides were checked. Peptides 8Q and GF15 were amino acid sequenced.HPV-16 E7 Protein. HPV-16 E7 protein was produced as MS2 fusion protein from a heat-ind...
Influenza vaccination using the NP appeared to be safe, and acceptable in this first time in humans study, and induced similar immune responses to vaccination by IM injection.
The 'Nanopatch' (NP) comprises arrays of densely packed projections with a defined geometry and distribution designed to physically target vaccines directly to thousands of epidermal and dermal antigen presenting cells (APCs). These miniaturized arrays are two orders of magnitude smaller than standard needles-which deliver most vaccines-and are also much smaller than current microneedle arrays. The NP is dry-coated with antigen, adjuvant, and/or DNA payloads. After the NP was pressed onto mouse skin, a protein payload co-localized with 91.4 + or - 4.1 APC mm(-2) (or 2925 in total) representing 52% of the delivery sites within the NP contact area, agreeing well with a probability-based model used to guide the device design; it then substantially increases as the antigen diffuses in the skin to many more cells. APC co-localizing with protein payloads rapidly disappears from the application area, suggesting APC migration. The NP also delivers DNA payloads leading to cutaneous expression of encoded proteins within 24 h. The efficiency of NP immunization is demonstrated using an inactivated whole chikungunya virus vaccine and a DNA-delivered attenuated West Nile virus vaccine. The NP thus offers a needle-free, versatile, highly effective vaccine delivery system that is potentially inexpensive and simple to use.
Background The Vaxxas high-density microarray patch (HD-MAP) consists of a high density of microprojections coated with vaccine for delivery into the skin. Microarray patches (MAPs) offer the possibility of improved vaccine thermostability as well as the potential to be safer, more acceptable, easier to use, and more cost-effective for the administration of vaccines than injection by needle and syringe (N&S). Here, we report a phase I trial using the Vaxxas HD-MAP to deliver a monovalent influenza vaccine that was to the best of our knowledge the first clinical trial to evaluate the safety, tolerability, and immunogenicity of lower doses of influenza vaccine delivered by MAPs. Methods and findings HD-MAPs were coated with a monovalent, split inactivated influenza virus vaccine containing A/Singapore/GP1908/2015 H1N1 haemagglutinin (HA). Between February 2018 and March 2018, 60 healthy adults (age 18-35 years) in Melbourne, Australia were enrolled into part A of the study and vaccinated with either: HD-MAPs delivering 15 μg of A/Singapore/ GP1908/2015 H1N1 HA antigen (A-Sing) to the volar forearm (FA); uncoated HD-MAPs; intramuscular (IM) injection of commercially available quadrivalent influenza vaccine (QIV) containing A/Singapore/GP1908/2015 H1N1 HA (15 μg/dose); or IM injection of H1N1 HA antigen (15 μg/dose). After 22 days' follow-up and assessment of the safety data, a further 150 healthy adults were enrolled and randomly assigned to 1 of 9 treatment groups.
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