• Development and Evaluation of Micro/Nanoparticulate Vaccines for Human Papillomavirus and Influenza

      Vo, Trinh; College of Pharmacy
      Influenza and Human papillomavirus (HPV) are among the most common infectious deseases prevalent globally. The influenza viruses cause approximately 200,000 hospitalizations each year, resulting in a significant financial burden to the healthcare industry. Similarly, HPVs cause virulent infections that have multiple serotypes capable of producing genital warts and cervical cancer. In both cases the infection is spread through viral agents, The influenza virus is transmitted by aerosolized mist of coughs and sneezes with HPV is a sexually transmitted disease. Prevention of influenza and HPV is possible with the help of vaccines, and currently, there are several vaccines for both infectious diseases. However, in the case of influenza, development of a vaccine is challenging as the virus undergoes rapid mutations and hence, the vaccine needs to be modified every season. Along with the constant need for vaccination rates. In the case of HPV, the need for a novel vaccine delivery system is more crucial. HPV is directly linked to cervical cancer and developing a novel strategy to improve vaccination globally is a major priority. Commercial vaccines are not only expensive but in developing countries the financial cost of maintaining cold storage and vaccine administration is significantly challenging. The developing non-parenteral vaccines is essential to improve patient compliance without affecting safety and efficacy of the vaccine. This project has two main sections focusing on development of an oral microparticulate vaccine for influenza and HPV. In the first part of this project, we developed an oral microparticulate vaccine for influenza. The influenza vaccine consists of a M2e protein which is a viral protein common to all influenza viruses encapsulated into a microparticle. Adjuvants such as Alum, MPL. R848, MF59, Flagellin, CpG and P4 were encapsulated in microparticles separately. Microphages werestimulated with the vaccine-adjuvants combinations and subsequently., splenocytes were incubated with activated macrophages to investigate the T-cell activity. We observed significant enhancement of immune markers such as nitric oxide, CD80, CD86, and MHC II with several adjuvants. Furthermore, CD4 and CD8 T cell expression was also significantly enhanced in groups treated with particulate adjuvants. In the second part of this project, we formulated a novel oral and transdermal vaccine delivery system for HPV. Development of potent vaccines against HPV can prevent serious complications such as genital warts and cervical cancer. Therefore a novel non-parenteral vaccine against HPV may significantly reduce incidence of HPV worldwide. In this study, three different microparticualte vaccine formulations have been prepared and characterized: Bovine Albumin Serum (BSA) cross0linked with glutaradehyde; b-cyclodextrin, and cellulose acetate succinate matrices. We systematically studied activation of the immune markers such as nitirc oxide. CD80, CD86, CD40 and MHC-II. In vivo studies demonstrated elevated humoral and cell-mediated immune response in murine models. We observed elevated IgG titers in mice vaccinated with microparticulate HPV16 VLP vaccine, Furthermore, flow cytometry analysis revealed expansion of specific T-cell populations in vivo. This study proves the efficacy of the HPV-16 oral microparticulate vaccine as a promising alternative to conventional vaccines, In the alter part of the HPV vaccine project we investigated the potential of transdermal HPV vaccine as a novel strategy to improve vaccinations globally. In order to combat HPV infection, vaccines must be affordable, self-administered and efficacious. Transdermal vaccines can be self-administered. are almost painless and tap the immune cells present under the skin. Transdermal vaccines have gained immense attention due to their efficacy. In this project, our main aim was to develop a microparticulate HPV16 VLP vaccine for transdermal vaccination. HPV-16 VLP was encapsulated in microparticles and characterized for size, zeta potential, encapsulation efficiency and microparticle yield. Scanning electron microscope images confirmed the particles were irregular with surface indentations. Western blot analysis was performed on HPV16 VLP extracted from microparticles, which confirmed the retention of antigenicity of the epitope following microparticles, which confirmed the retention of antigenicity of the epitope following microparticle fromulation. Following in vitro characterization studies, we performed extensive in vivo studies to investigate the potential of the microparticulate vaccine to initiate a robust and sustained immune response. IgC titers for mice vaccinated with microparticulate vaccine were significantly elevated as compared to antigen solutions adminstered by the transdermal route. Flow cytometry was performed on vaccinated and control groups to investigate the expansion of specific T-cell populations. Vaccinated mice demonstrated significant increase in CD4, CD45R, CD27 and CD62L cell populations. Expansion of these specific T-cell subsets demonstrates the efficacy of the microparticulate vaccine when administered transdermally. Thus, our study proves the efficacy of the microparticulate vaccine over vaccine solutions and the novel approach of administering the vaccine through a minimally invasive, pain-free route that will help to improve acceptability of vaccination against HPV.