Lymph node-targeting and antigen-presenting proteinticles for in vivo cancer immunotherapy

학위논문 (석사)-- 서울대학교 융합과학기술대학원 : 분자의학 및 바이오제약학과, 2015. 8. 이동수. ; Cancer immunotherapy is a new class of cancer treatment which uses the bodys own immune system to defeat cancers. To elicit immune responses that attack cancer cells, efficient antigen delivery to lymph nodes (LNs) can be an appropriate strategy, because LNs possess a high population of immune-related cells such as antigen presenting cells (APCs), T cells and B cells. However, realization of this strategy remains challenging. Biocompatible protein-based nanoparticles (named proteinticles) that self-assemble inside cells with precise 3D nanostructure, have a unique advantage over synthetic nanomaterials for LN-targeting immunotherapy. Herein, human ferritin heavy chain (HFt) and red fluorescence protein (RFP) were selected as a promising antigen carrier and a model antigen, respectively. Genetically recombined HFt-RFP vaccine particle with a size of 26 nm showed fast diffusion and long retention in the targeted lymph node. Moreover, HFt-RFP induced strong cytotoxic CD8+ T cell responses, resulting in significant inhibition of tumor growth in RFP-expressing tumor-bearing mice. Here, we show that LN-targeting strategy using antigen-presenting proteinticles holds promises for effective cancer immunotherapy. ; ABSTRACT ……………………………………………………… i Table of Contents ……………………………………………… ii List of Tables …………………………………………………… iv List of Figures ……………………………………………………v 1. Introduction ……………………………………………………1 2. Materials and Methods 2.1 Biosynthesis of HFt-RFP ……………………………… 3 2.2 Characterization of HFt-RFP ……………………………4 2.3 Preparation of Cy5.5 …………………………………… 4 2.4 Lymph node accumulation …………………………… 5 2.5 Intracellular cytokine staining ………………………… 5 2.6 Evaluation of tumor inhibition effect ………………… 6 2.7 Immunofluorescence dual-staining and lymphocytes population analysis …………………………………… 7 2.8 Statistical analysis ………………………………………8 3. Results 3.1 Development and characterization of HFt-RFP …… 9 3.2 Comparison of HFt-RFP with RFP, HFt in time- dependent in vivo and ex vivo ...