SD VLPs stored at room heat for over 7 months were highly immunogenic and protected mice from genital challenge with HPV16 PsV as well as mice immunized with Gardasil. In summary, spray drying is a one-step, easily scalable pharmaceutical process that can be used to transform candidate L2 VLP vaccines from liquid treatment for a dry powder that does not require refrigeration. First, we assessed the role of antigen dose and boosting on immunogenicity. Mice immunized with 16L2-MS2 VLPs at doses ranging from 2C25 g with or without alum were highly Rabbit Polyclonal to FMN2 immunogenic at all doses; alum appeared to have an adjuvant effect at the lowest dose. Although boosting enhanced antibody titers, even a single immunization could elicit strong and long-lasting antibody responses. We also developed a method to enhance vaccine stability. Using a spray dry apparatus and a combination of sugars & an amino acid as protein stabilizers, we generated dry powder vaccine formulations of our L2 VLPs. Spray drying of our L2 VLPs did not affect the integrity or immunogenicity of VLPs upon reconstitution. Spray dried VLPs were stable at room temperature and at 37C for over one month and the VLPs were highly immunogenic. Taken together, these enhancements are designed to facilitate implementation of a next-generation VLP-based HPV vaccine which addresses U.S. and global disparities in vaccine affordability and access in rural/remote populations. Keywords: MS2 and PP7 bacteriophages, Virus-like Particles, HPV vaccine, Adjuvants, Formulation 1. Introduction Human papillomavirus (HPV) contamination is a necessary cause of nearly all cases of cervical cancer; it is also a significant cause of other anogenital carcinomas as well as a growing percentage of oropharyngeal cancers [1, 2]. The current HPV vaccines (Gardasil and Cervarix) are comprised of virus-like particles (VLPs) derived from the HPV major capsid protein, L1 [3-5]. Both vaccines are highly immunogenic and elicit high titer and long-lasting neutralizing Cefradine antibody responses. Although these vaccines provide strong protection Cefradine against the oncogenic HPV types included in the vaccines (HPV16 and HPV18), they provide very little cross-protection against the other 13-16 high-risk HPV types associated with ~30% of cervical cancer cases [6-11]. More recently a nonavalent HPV vaccine, called Gardasil-9 (which is also based on L1 VLPs), was approved by the Food and Drug Administration Cefradine [12]. While the nonavalent vaccine is likely to increase the breath of HPV protection (it includes VLPs derived from HPV types that cause about 90% of cervical cancer cases), the cost of production and formulation will likely be high, particularly given the fact that the current HPV vaccines are already very expensive [13]. Thus, Cefradine the nonavalent vaccine may not be affordable in underdeveloped countries where ~85% of cervical cancer cases occur. Another limitation of all current HPV vaccines is usually that they require cold-chain for transportation and storage. This requirement is usually a barrier for implementation in the developing world where refrigerated facilities for transportation and storage are often inadequate [14]. As an alternative to the current type-specific HPV vaccines, we have developed vaccines that target highly conserved, broadly neutralizing epitopes from the HPV minor capsid protein, L2 [9, 15-18]. Immunization with L2-displaying VLPs elicits high-titer and broadly neutralizing antibodies against HPV. For example, an RNA bacteriophage MS2-based vaccine displaying a short peptide representing amino acids 17-31 from HPV16 L2 induces antibodies that strongly protect mice from genital contamination with HPV pseudoviruses representing eleven diverse HPV types [17]. The goal of this study was to develop techniques to enhance the clinical applicability of VLP-based vaccines targeting HPV L2, particularly in resource-poor settings. In these studies we asked whether 1) VLP-based vaccines targeting HPV L2 could elicit high titer antibodies responses after a single immunization, and 2) we could develop highly stable formulations of these VLP-based vaccines that were suitable for low-resource settings. We assessed the impact of antigen dose and boosts on antibody responses to L2 and also assessed the longevity of antibody responses. To create a more thermostable vaccine, we spray.