24^th International Conference on
Vaccine Design, Production & Safety

Chiungyi Huang works in National Health Research Institutes of Taiwan for more than 10 years. The objective of the research group focuses on design and development of emulsion adjuvants based on degradable polymers

ABSTRACT: Emulsion-based adjuvants were demonstrated as an effective tool to increase the vaccine efficacy. This study provides the size matter and compositions of emulsions for modulating the immune responses. We use a mouse model with ovalbumin (OVA) as the antigen to deepen our understanding of the properties of the emulsion at micro-/nanoscale interacting with immune cells. First of all, we explored mechanistic reasoning toward embracing the interface between the physicochemical and biological signatures for core oil selection in antigen uptake. Our results showed that treatment of dendritic cells (DCs) and splenocytes with a squalene-based emulsion (referred as SqE) induced reactive oxidative species (ROS) production and resulted in an increase in apoptotic and necrotic cells in a concentration- and time-dependent manner. Furthermore, DCs co-cultured with cellular debris of SqE-pretreated splenocytes resulted in a higher level of ovalbumin (OVA) antigen uptake by DCs than those co-cultured with untreated splenocytes. Interestingly, the potency was rather attenuated when splenocytes pretreated with a typical ROS inhibitor, N-acetyl-cysteine. Notably, SqE possesses a high impact on eliciting ROS-mediated antigen uptake compared with a squalane-based emulsion (SqA). Next, we formulated various emulsions using different types of core oils and fatty acids, and planned to control the size of the emulsions range from macro- to nanoscale by passing through the extruder membrane. The results showed that emulsions at nanoscale could enhance activation of DCs in vitro, compared with those at submicron scale. These results highlight the importance of unsaturated squalene core oil in the adjuvant activity of emulsions and offer insight into the design and development of vaccine against infectious diseases or cancer immunotherapy technology. This work was supported by grant 107A1-IVPP22-014 from the National Health Research Institutes of Taiwan and grant MOST 106-2314-B-400-016-MY3 from the Ministry of Science and Technology of Taiwan.

Graduated in 2009 from the INSA (National Institute of Applied Sciences), I wanted to be a pharmaceutical researcher. Therefore, I did a PhD thesis in Chemistry dedicated to oncology, and more specifically to the targeting of tumors, to improve diagnosis and radiotherapy treatment of radioresistant cancers (glioblastoma, melanoma and chondrosarcoma). After completing my PhD in 2013, I joined the CEA, the French Atomic Energy and Alternative Energies Commission, among the Technological Research Division. I used to work on just as transversal themes, i.e., the identification of new biomarkers or new therapeutic targets of prostate cancer, the delivery of novel therapeutic agents to breast cancer, as well as the development of a vaccine against HIV. Today, I work in an R&D platform at Sanofi Pasteur: I design new conjugation chemistries for the development of conjugate vaccines.

Pathogenic bacteria exhibit specific polysaccharides on their surface that have been identified as major virulence factors. , While capsular polysaccharides are considered T-cell independent antigens, coupling them to a carrier protein can activate a T-cell dependent response. , Unlike plain polysaccharide vaccines, glycoconjugates are able to recruit T cells that help B cell activation, resulting in immunogenic responses in the young children under the age of two and induction of a sustained immune memory.Since the success of Hib conjugate vaccine in reducing significantly the burden of Haemophilus Influenzae type b-related disease, two other major conjugate vaccines have been marketed, i.e. against Streptococcus Pneumoniae and Neisseria Meningitidis infections, and many others are currently under development.4,6 In this respect, our Development Platform is willing to expand its conjugation activities by implementing a chemistry laboratory for the design and development of novel glycoconjugate vaccines.A laboratory covering a surface area of 90m2 has been established and adapted to chemical purposes. Conventional chemical laboratory facilities have been installed (e.g. fume hoods, lab benches, weigh station) and process equipments suitable for biomolecule conjugation have been purchased, such as pH titrator or purification systems to perform gel filtration chromatography and tangential flow filtration. With the ultimate objective to characterize synthesized glycoconjugates and to monitor each step of the conjugation reaction, we have also integrated a microplate reader to perform absorbance and fluorescence measurements. Additionally, a wide panel of analytical equipments, such as an HPLC, a GPC/SEC system coupled to quadruple detection (UV, RI, SLS, Viscosity), or a DLS apparatus will be available. This chemistry laboratory will increase the capacity of our Development Platform to develop new conjugate vaccines.

Completed my degree in engineering at Ecole Superieure d'Ingenieurs de Luminy in the year 2009. Was the purification scientist at Sanofi Pasteur from 2009- 2010. From the year 2010 - 2011 was biomanufacturing engineer at Merck Millipore. Was the process development scientist in Merck millipore between 2011- 2015. Now Responsable scientist at Sanofi Pastuer

Purification is a crucial step during vaccine production process development. In the current race to clinic context, the purification process has to be developed in shorter timelines and traditional screening approaches are being challenged. As a central technology of purification, chromatography is a perfect target to increase screening throughput. Our current work is focused on the implementation of an automated chromatography resin screening platform for viral particles or subunit antigens. Robots running multi-conditions simultaneously are combined with high throughput analytical methods. Our global approach also includes data management tools to accelerate data analysis and ensure data integrity.By multiplying tested conditions, this automated methodology speeds up screening studies resulting in shorter development times before clinics. In later development stages, the approach is valuable again: optimization plans can be enlarged with additional conditions, thus enhancing process understanding and ultimately process robustness.

Rinat Nuriev is a scientific fellow in the Laboratory of Bioconjugates of I. Mechnikov Research Institute of Vaccines and Sera. The team of the lab works in the fields of Immunochemistry and Vaccine Design; specifically we are investigating immunochemical structures of the pneumococcal conjugate vaccines and new approaches for the detection of antibiotics. Our scientific work also aims the detailed study of the protein-polysaccharide conjugation process in order to find some new and useful ways to prevent pneumococcal invasion

Ongoing improvement of the conjugate vaccines against S. pneumoniae remains relevant due to the changes in the circulating pneumococcal serotypes. Despite of the achievements in conjugate design, the development of highly effective vaccines requires individual conjugation approaches for certain pneumococcal capsular polysaccharide. The carbodiimide condensation method using adipic acid dihydrazide (ADH) as a spacer is one of the common approaches for synthesis of such conjugates. However, this technique can promote the formation of intramolecular crosslinking, thereby decreasing the efficiency of conjugate. Alternative approaches, such as azide-alkyne cycloaddition, allow avoiding intramolecular crosslinking owing to bioortogonal site-specific interactions. We modified capsular polysaccharides of S.pneumoniae types 9N (Ps9N) and 3 (Ps3) with ADH and propargylamine (PA) in different molar ratios between polysaccharide and a spacer. Immunochemical activity of the modified Ps9N changed insignificantly in all cases, while Ps3 derivatives were considerably different from its native form. Further immunorecognition examinations of Ps-antigens were conducted after conjugation of Ps-ADH and Ps-PA derivatives with tetanus toxoid. The studies showed that Ps9N modification with 100-fold molar excess of ADH was sufficient for effective coupling with carrier, while Ps3 should be treated with 1000-fold ADH excess. We optimized the procedure of azide-alkyne cycloaddition and showed that 1mM Cu (I) as a catalyst promotes effective conjugate formation, whereas reaction time (1-24 h) doesn't affect the yield. Animal studies revealed high immunogenicity in mice and promotion of faster S.pneumoniae elimination from mouse lungs after intranasal infection. In this work we propose some new considerations regarding the synthesis of pneumococcal polysaccharide conjugates, which can help in further vaccine development. We also emphasize that the safety of native polysaccharide structure should be immunochemically monitored for prognosis of conjugates bioefficacy.