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Form 3

(Study Plan)

* Please type or print clearly within 1 page using black ink. (*10 points) Goal of study & Study Plan

o Goal of study, title or subject of the research and detailed study plan

Title: Thiolated Eudragit Microspheres as Carrier System for Pasteurella multocida Dermonecrotoxin
Background of the study Vaccine is the best defense and cost-effective strategy against many diseases. Oral vaccination is the most attractive route of vaccine delivery. However, low oral bioavailability, degradation of antigen in acidic stomach and short residence time of drug at target site are the prime problems. Recently, thiolated polymers are showing huge benefits in mucosal drug delivery. Pasteurella multocida dermonecrotoxin (PMT) is a major virulence factor as a causative agent of atrophic rhinitis (AR) in swine and, therefore, inactivated P. multocida will be used as a candidate vaccine in the current study to encapsulate into a pH-sensitive and mucoadhesive thiolated Eudragit microspheres (TEMS) and will be investigated for their immune stimulating activity against AR and intracellular signaling pattern in vitro and in vivo. Goals To prepare an effective carrier system for the purpose of oral vaccination Experimental Methods

Preparation of Thiolated Eudragit (TE) by applying Eudragit-cysteine conjugate system. Preparation of PMT-loaded TEMS by water-in-oil-in-water (w/o/w)-solvent evaporation method Microspheres morphology will be observed by scanning electron microscope (SEM) Particle size measurement by Electrophoretic light scattering spectrophotometer (ELS) Loading efficiency and the release of PMT from PMT-loaded TEMS will be measured in vitro by Micro BCA protein assay method.

Measurement of secreted cytokines (TNF-, IFN-, IL-2, IL-6, IL-10 & other) and nitric oxide (NO) as immune stimulatory mediators from murine macrophage cell line (RAW264.7) stimulated by PMT-loaded TEMS in vitro. In vivo study on immune stimulation against AR and intracellular signaling in animal model.

Expected Outcomes The afore-mentioned method will provide a successful preparation of TE and PMT-loaded TEMS. Microspheres will be spherical with smooth surface and particle size will be below 10 m in diameter, the size which have significant efficiency in oral vaccination to reach the immune simulative sites of Peyers patches at GALT to induce immune responses. The formulation method will provide higher loading efficiency of PMT into TEMS. Release of PMT from PMT-loaded TEMS will be protected at pH 2.0 to indicate the protection of vaccine from acidic condition and will provide a controlled-release of PMT at pH 7.4 in vitro.

Cytokines and NO will be secreted gradually from murine macrophage cells as immune stimulatory mediators stimulated with

PMT-loaded TEMS to show in vitro immune stimulating activity.

In vivo results are expected to show protection against AR infection

Impact of the study The success of this research will open a novel strategy to investigate the delivery and application of therapeutic vaccine against infectious diseases through oral administration. In the recent years, Prof. C. H. Yun has been leading world class research works in the field of immunology, vaccinology and cell signaling pathways related to prevent various diseases including cancer. I believe that I would be able to accomplish my research successfully as a member of his group. I would be extremely grateful to National Institute for International Education (NIIED) if I am awarded for the Korean Government Scholarship (as KGSP fellow 2010) to pursue my Doctoral Program under supervision of Professor Yun. I can assure that I will try to do my best to contribute a little to the welfare of humankind. Future Plan after Study

o Future plan in Korea or another country after study in Korea

Korea is one of the leading countries in advanced research and development in the world. Seoul National University is not only the best University in Korea but also the top class University in World Ranking. It will be an absolutely great place to extend and pursue my academic and research career for achieving my goals. I believe if I am given the opportunity and selected for KGSP fellow 2010, I can surely make positive impacts in the scientific field through my research both in Korea and Bangladesh as well as other countries. I would like to mention that the proposed field of study has huge impacts and prospects on modern scientific research in Korea as well as other countries all over the world. I can also implement beneficial impacts to my country. I am very much sure that I can contribute extremely well in the scientific research and development in Korea with my improved knowledge after study here if I am given the opportunity. Biotechnology, particularly Agricultural Biotechnology is the most improved and rapidly developing field in the scientific arena. Most of the countries in the world including Bangladesh have been focusing in this research field of study with great importance. If I am awarded with this fellowship to study in Korea, I can assure that I will definitely make positive contribution in this progressively improved research field either as an academician or a researcher in Korea and Bangladesh as well as any other countries in the world for the welfare of humankind.