서강대학교

목차

1. Introduction 1
1.1. Respiratory Syncytial Virus (RSV) 1
1.2. Vaccine associated Enhanced Respiratory Disease (ERD) 2
1.3. Lipid Nanoparticle (LNP) 3
1.4. Cellular uptake mechanism of LNP 6
1.5. The Limitation of LNP 11
1.6. Cell-penetrating Peptides (CPPs) and Recent Researches 11
1.7. Melittin 13
2. Materials and Methods 16
2.1. Materials 16
2.2. Synthesis of lipid-C-MEL 17
2.3. Preparation of LNP and LNP-MEL 18
2.4. Characterization of LNP and LNP-MEL 19
2.5. Encapsulation Efficiency of LNP and LNP-MEL 20
2.6. Cell culture 20
2.7. RSV (RSV A2) preparation 21
2.8. Cell cytotoxicity analysis 21
2.9. Hemolysis assay 22
2.10. mRNA transfection efficiency analysis 22
2.11. Cellular uptake analysis 23
2.12. Endosomal escape efficiency analysis 23
2.13. Animal studies 24
2.14. Bioimaging studies 24
2.15. Histopathological analysis 25
2.16. Serum collection 25
2.17. ELISA 25
2.18. Lung supernatant collection 26
2.19. Bronchoalveolar Lavage (BAL) fluid collection 27
2.20. Immunization and RSV challenge studies 27
2.21. Flow cytometry 28
3. Results and Discussion 29
3.1 Synthesis of lipid-C-MEL 29
3.2 Toxicity analysis of lipid-C-MEL 31
3.3 Optimization and characterization of LNP-MEL 33
3.4 In vitro endosomal escape and mRNA delivery of LNP-MEL 39
3.5 In Vivo mRNA delivery efficiency and toxicity of LNP-MEL 44
3.6 In vivo immunogenicity of RSV mRNA LNP-MEL vaccine 47
4. Conclusion 51
5. References 53