Purpose: Gonorrhea is one of the most common sexually transmitted disease. It is caused by Gram negative diplococcus bacteria, Nisseria gonorrhoeae. According to CDC, 468,514 cases of gonorrhea were reported in the United States in the year 2016. The patients suffering from gonorrhea infection experience symptoms like: urethritis and cervicitis, pelvic inflammation, pus discharge, abdominal pain, and dyspareunia. The current treatment choices for gonorrhea involve the use of antimicrobials. But the recent progress of drug resistance in the gonorrhea bacteria has become a great hazard to public health and current antimicrobial treatments for the infection. Hence, novel preventative and therapeutic methods for gonorrhea infections are needed. Some studies have been carried out in the past for developing a vaccine for gonorrhea, which showed some success, but they were not able to generate an adaptive immune response. Thus, there has been an increased focus to identify specific antigens and generation of a protective immune response. A preventative vaccine for gonorrhea can be of great importance in the healthcare because of wide occurrence of the infection. The skin provides a favorable site for the vaccination because of ease of access and presence of immune cells for an efficient immune response against a range of antigens. Langerhans cells (specialized dendritic cells) in the skin are phagocytic cells that signal T-cells. Upon activation, T cells and macrophages drain into nearby lymph nodes causing an increased immune response. The objective was to encapsulate the antigen (formalin fixed gonorrhea bacteria) into microparticles, which would be tested for their antigenicity and immunogenicity using microneedles-based delivery through transdermal route.
Methods: Microparticles loaded with antigens were prepared using spray drying method. The particulate vaccine formulation contains a biodegradable, biocompatible, non-antigenic and sustained releasing polymer components containing cross-linked albumin matrix and formalin-fixed inactivated whole-cell gonococci (vaccine antigen, 10% w/v) and adjuvants. The microparticles were characterized for percent yield, size, charge and poly dispersity index (PDI). The sugar-based microneedles containing the gonorrhea vaccine microparticles were prepared using the mold casting method. Scanning Electron Microscopy of the microneedles was carried out to characterize size and shape of microneedles. In-vivo efficacy of this vaccine was checked in 6-8 weeks old swiss webster (CFW) mice. Mice were administered one prime dose at day 0 followed by two booster doses at week 2 and 4. There were five groups (n=6) in this study; naïve group which received nothing and served as control, second group received the blank microneedles without any vaccine (Blank MN or microneedle control group), third group received gonorrhea vaccine suspension subcutaneously (GnH Susp SubQ), forth group was administered gonorrhea vaccine suspension loaded into the microneedles (GnH Susp MN) via transdermal delivery and fifth group was administered gonorrhea vaccine microparticles loaded in the microneedles (GnH MP MN). Blood samples were collected from these animals bi-weekly and the serum antibody titers were measured using ELISA. Animals were sacrificed at week 10, lymph organs were collected and levels of the immune cells such as the CD4+ and CD8+ T cells in the spleens were counted using BD Accuri™ C6 Plus (BD Accuri Cytometers, Ann Arbor, MI).
Results: The percent yield for vaccine particles was 89 % w/w. Vaccine particles were 4.5 um and PDI was 0.447 with a charge of -25 mV. The group which received the GnH vaccine microparticles in microneedles (GnH MP MN) showed significantly higher antibody titers than the other two vaccine groups at week 6 and 8 (*p<0.001; #p<0.05). The group that received the vaccine, showed significantly higher CD4+ and CD8+ T cells than compared to the controls – naïve and blank microneedles (p<0.05).
Conclusion: The gonorrhea vaccine microparticles were prepared with a novel approach using formalin fixed dead gonorrhea bacteria as the antigen. Since the bacteria is formalin fixed, all the surface proteins, antigenic domains are conserved in their native form which are presented by antigen presenting to the immune cells of the body. Generation of antibodies post vaccination was seen in mice using microneedle-based transdermal route of administration. The vaccination produced both CD4+ and CD8+ T cell based immune response, which is an important factor for the success of a vaccine. The microparticulate based vaccine delivery system is a promising strategy for the delivery of the gonorrhea vaccine. This vaccine can be of substantial importance in battling the antibiotic resistances being established in the bacteria. However, a complete effort on many facades will be needed to bring gonorrhea vaccine into clinical use. Many challenges are inherent in the development process of the gonorrhea vaccine including the time and money that would be needed for the clinical trials but the development of vaccine to prevent this infection will be of significant benefit in public health care.
Rikhav Gala– Research Scientist, University of New Mexico, Atlanta, Georgia
Cherilyn D'Souza– Mercer University, Atlanta, Georgia
Carmen Popescu– Global Pharma Program Coordinator, Roquette America Inc., Geneva, Illinois
Susu Zughaier– Assistant Professor, Qatar University, Doha
Martin D'Souza– Professor, Mercer University, Atlanta, Georgia