엔도좀 탈출능이 향상된 종양 특이적 세포질 침투항체의 개발
Generation of tumor-specific cytosol-penetrating antibody with improved endosomal escape activity
- 주제(키워드) Cytosol-penetrating antibody , Endosomal escape
- 발행기관 아주대학교
- 지도교수 김용성
- 발행년도 2018
- 학위수여년월 2018. 2
- 학위명 석사
- 학과 및 전공 일반대학원 분자과학기술학과
- 실제URI http://www.dcollection.net/handler/ajou/000000026913
- 본문언어 영어
- 저작권 아주대학교 논문은 저작권에 의해 보호받습니다.
초록/요약
In general, Intact form of IgG antibodies cannot reach the cytosol by escaping from the endosome after receptor-mediated endocytosis because they have no endosomal escape activity. Our group recently reported the cytosol-penetrating antibody TMab4 cytotransmab. Also, we generated TMab4-3 which is an engineered version of TMab4 to improve the endosomal escape efficiency and its expression yield. However, TMab4-3 is not yet suitable for targeted therapy because TMab4-3 has a binding activity against HSPG expressed on the surface of normal cells. In addition, it is necessary to improve the endosomal escape efficiency to increase the therapeutic efficacy of cytotransmab. Here, I generated a tumor-specific cytotransmab with improved endosomal escape efficiency. First, I have created a new cytotransmab called as CT-41 with the elimination of HSPG binding activity. Second, a cyclic peptide for targeting EpCAM which is well known for over-expressed on surface of tumor cell was fused with N-terminus of CT-41 to confer tumor tissue specificity. Third, an additional endosomal escape motif for cytotransmab have been introduced into the EpCAM specific cytotransmab called as CT-ep41. Lastly, charged residue mutations are inserted near the hydrophobic endosomal escape motif to increase its developability and solubility. As a result, CT12-ep61 as a final clone is showed a specificity to EpCAM-positive tumor cells and increased endosome escape efficiency about 34% compared to CT-ep41 and its expression level is higher about 8-fold compared to CT-ep41. These results suggest that tumor-tissue specific cytotransmab can be applied as a tool for targeted cancer therapy.
more목차
1. Introduction 1
2. Material and methods 3
3. Results 9
3.1 Generation of EpCAM-specific Cytotransmab for targeting tumor tissue 9
3.2 Development of endosomal escape motif on VH-CDR3 of Cytotransmab 17
3.3 Generation of Cytotransmab with endosomal escape motif on both of VH-CDR3 and VL-CDR3 for improving endosomal escape efficiency 24
3.4 Improvement of biophysical properties of Cytotransmab with endosomal escape motif on both of VH-CDR3 and VL-CDR3 29
4. Discussion 39
5. References 43
국문 요약 46
