DENV (DENV) is the most common human arboviral infection, and the most important public health threat from mosquito-borne viral pathogens causing an estimated 100 million cases of dengue fever and half a million cases of dengue hemorrhagic fever (DHF) per year. Of those infected with DHF, 90% are children who ultimately succumb and make up the 5% of people who die of the infection. As Dengue is endemic in all continents except Europe, over half of the world?s population is at risk of infection. Over the past fifty years, incidents of dengue infection have increased by over 30-fold. Clinical trials of DENV vaccines have shown promising results, but prototype vaccines remain inadequate against all four DENV serotypes. Vaccination is the most important strategy for prevention of flavivirus infections, and monoclonal antibodies show great promise even as a post-exposure treatment strategy. One critical challenge to the development of effective vaccines is our incomplete ability to examine and understand the protective humoral immunity against DENV. This challenge is attributed to the limitation of the current technologies to provide a comprehensive profile of the protective neutralizing antibodies in dengue infection. In this application, we propose using single-cell antibody nanowells (SCAN), an innovative single-cell technology, to quickly and efficiently screen individual B- cells for antigen specific products. Next, these antigen specific products will be isolated and reproduced via cloning and recombinant DNA technologies. This extremely efficient process uses much smaller samples than conventional methodologies, and, most importantly, it will allow for the identification of rare cross-neutralizing epitopes. We propose using SCAN technology to identify and isolate DENV-specific antibodies that will be evaluated for viral neutralizing properties. Antibodies with neutralizing properties will then be characterized to determine mechanism of neutralization. Results are expected to generate a repertoire of DENV-specific antibodies. These antibodies will be screened for neutralizing capacity and mapped for antigenic epitopes. On a more fundamental level, these results will establish this approach and direct it forward towards the creation of immuno therapeutics and vaccine development.
With 3.9 billion people at risk of infection, dengue is a pressing world health concern; however, vaccine clinical trials have not been able to provide complete protection from dengue, and neutralizing antibodies against dengue virus have not been successfully translated into human patients. The goals of this application are to utilize an innovative single-cell technology to provide a virus-specific humoral immune response in dengue patients, identify and evaluate neutralizing antibody properties, and determine the mechanism of neutralization. The results from this study will establish a foundation for effective vaccine targets against dengue infection.
| Voigt, Alexandria; Semenova, Touyana; Yamamoto, Janet et al. (2018) Therapeutic Antibody Discovery in Infectious Diseases Using Single-Cell Analysis. Adv Exp Med Biol 1068:89-102 |
