S-Space College of Medicine/School of Medicine (의과대학/대학원) Dept. of Medicine (의학과) Theses (Ph.D. / Sc.D._의학과)
The Comparison of HER2 Targetability between Affibody and Antibody in Nude Mice Bearing Human Breast Cancer Xenograft
유방암 이식 마우스에서 아피바디와 항체의 HER2 표적능 비교
- 의과대학 의학과
- Issue Date
- 서울대학교 대학원
- 학위논문 (박사)-- 서울대학교 대학원 : 의학과, 2014. 2. 강건욱.
- Introduction: Human epidermal growth factor receptor type 2 (HER2) is a tyrosine kinase that is overexpressed in many carcinomas and is associated with a poor prognosis. Imaging HER2 expression in malignant tumors can provide important prognostic and predictive information. The analyses of anti-HER2 tracers by using both fluorescence and radionuclide technologies may characterize the macroscopic and microscopic localization of tracers. The object of this study was to investigate whether accurate macroscopic and microscopic distribution of affibody in a xenograft model could be achieved by using fluorescent and radionuclide labels, and the potential of affibody as a diagnostic or therapeutic agent.
Methods: The anti-HER2 affibody ZHER2:477 and the humanized monoclonal antibody trastuzumab were labeled with 64Cu-DOTA using maleimide, amine reaction or a fluorescent tag using isothiocyanate with an amine reaction. An in vitro cellular binding assay for both tracers was performed using HER2-overexpressing cells, and with fluorescent microscopy for the qualitative analysis of the fluorescent label and quantitative assessment of radionuclide activity. The biodistributions of the radiolabeled affibody ZHER2:477 (64 Cu -affibody) and radiolabeled antibody, trastuzumab (64 Cu -antibody) were compared in BALB/C nu/nu mice bearing both HER2 overexpressing KPL-4, NCI-N87 xenografts and MDA-MB-231 xenografts, which express only negligible levels of HER2. In vivo PET imaging and ex vivo autoradiographic, confocal microscopic analysis of affibody and antibody in tumor-bearing mice were performed at 4 h and 24 h after intravenous administration.
Results: Both 64 Cu -affibody and 64 Cu -antibody demonstrated specific binding to HER2 overexpressing cells in vitro. Similarly, both 64 Cu -affibody and 64 Cu -antibody only specifically targeted HER2 overexpressing xenografts in vivo PET imaging (64 Cu -affibody; 4 h 2.84 ± 0.05 % ID/g, 24 h 3.51 ± 0.53 % ID/g, 64 Cu -antibody; 2 h 0.60 ± 0.73 % ID/g, 24 h 2.84 ± 1.67 % ID/g). When the intratumoral distributions of tracers were analyzed by confocal microscopy using fluorescence, affibody showed tumor penetration similar to antibody at 4 h and it disappeared at 24 h. When the wholebody distributions of tracers were analyzed by biodistribution using radioisotope, 64 Cu -antibody localized more in tumor than 64 Cu -affibody both at 4 h and 24 h (64 Cu -affibody; 4 h 2.56 ± 0.13 % ID/g, 24 h 4.11 ± 0.41 % ID/g, 64 Cu -antibody; 4 h 3.05 ± 0.56 % ID/g, 24 h 5.79 ± 1.28 % ID/g), but 64 Cu -affibody showed better tumor to blood ratio than 64Cu -antibody (64 Cu -affibody; 4 h 2.77 ± 0.96, 24 h 1.95 ± 0.26, 64 Cu -antibody; 4 h 0.07 ± 0.01, 24 h 0.43 ± 0.07).
Conclusions: From confocal microscopy using fluorescence, affibody showed tumor penetration similar to antibody at 4 h and it disappeared at 24 h. From biodistribution study using radioisotope, 64 Cu -antibody localized more in tumor than 64 Cu -affibody both at 4 h and 24 h, but 64 Cu -affibody showed better tumor to blood ratio than 64 Cu -antibody. These analyses showed that affibody can be applied as a diagnostic, rather than a therapeutic agent.