title: In vivo targeting of HER2-positive tumor using 2-helix affibody molecules.
abstract: Molecular imaging of human epidermal growth factor receptor type 2 (HER2) expression has drawn significant attention because of the unique role of the HER2 gene in diagnosis, therapy and prognosis of human breast cancer. In our previous research, a novel cyclic 2-helix small protein, MUT-DS, was discovered as an anti-HER2 Affibody analog with high affinity through rational protein design and engineering. MUT-DS was then evaluated for positron emission tomography (PET) of HER2-positive tumor by labeling with two radionuclides, 68Ga and 18F, with relatively short half-life (t1/2<2 h). In order to fully study the in vivo behavior of 2-helix small protein and demonstrate that it could be a robust platform for labeling with a variety of radionuclides for different applications, in this study, MUT-DS was further radiolabeled with 64Cu or 111In and evaluated for in vivo targeting of HER2-positive tumor in mice. Design 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) conjugated MUT-DS (DOTA-MUT-DS) was chemically synthesized using solid phase peptide synthesizer and I2 oxidation. DOTA-MUT-DS was then radiolabeled with 64Cu or 111In to prepare the HER2 imaging probe (64Cu/111In-DOTA-MUT-DS). Both biodistribution and microPET imaging of the probe were evaluated in nude mice bearing subcutaneous HER2-positive SKOV3 tumors. DOTA-MUT-DS could be successfully synthesized and radiolabeled with 64Cu or 111In. Biodistribution study showed that tumor uptake value of 64Cu or 111In-labeled DOTA-MUT-DS was 4.66±0.38 or 2.17±0.15%ID/g, respectively, in nude mice bearing SKOV3 xenografts (n=3) at 1 h post-injection (p.i.). Tumor-to-blood and tumor-to-muscle ratios for 64Cu-DOTA-MUT-DS were attained to be 3.05 and 3.48 at 1 h p.i., respectively, while for 111In-DOTA-MUT-DS, they were 2.04 and 3.19, respectively. Co-injection of the cold Affibody molecule ZHER2:342 with 64Cu-DOTA-MUT-DS specifically reduced the SKOV3 tumor uptake of the probe by 48%. 111In-DOTA-MUT-DS displayed lower liver uptake at all the time points investigated and higher tumor to blood ratios at 4 and 20 h p.i., when compared with 64Cu-DOTA-MUT-DS. This study demonstrates that the 2-helix protein based probes, 64Cu/111In DOTA-MUT-DS, are promising molecular probes for imaging HER2-positive tumor. Two-helix small protein scaffold holds great promise as a novel and robust platform for imaging and therapy applications.
authors: Molecular Imaging Program at Stanford (MIPS), Department of Radiology and Bio-X Program, Stanford University, California, Stanford, CA 94305-5344, USA.
authors: Gang
authors: G
authors: Ren
authors: Jack M
authors: JM
authors: Webster
authors: Zhe
authors: Z
authors: Liu
authors: Rong
authors: R
authors: Zhang
authors: Zheng
authors: Z
authors: Miao
authors: Hongguang
authors: H
authors: Liu
authors: Sanjiv S
authors: SS
authors: Gambhir
authors: Faisal A
authors: FA
authors: Syud
authors: Zhen
authors: Z
authors: Cheng
mesh_terms: D000818:Animals; D001665:Binding Sites; D001943:Breast Neoplasms / Q000000981:diagnostic imaging* / Q000378:metabolism; D003301:Copper Radioisotopes; D005260:Female; D006573:Heterocyclic Compounds, 1-Ring / Q000378:metabolism; D007205:Indium Radioisotopes; D051379:Mice; D008819:Mice, Nude; D018716:Molecular Mimicry; D009368:Neoplasm Transplantation; D010051:Ovarian Neoplasms / Q000000981:diagnostic imaging* / Q000378:metabolism; D049268:Positron-Emission Tomography; D011485:Protein Binding; D011506:Proteins / Q000138:chemical synthesis / Q000737:chemistry / Q000378:metabolism; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000737:chemistry / Q000378:metabolism*; D015899:Tomography, Emission-Computed, Single-Photon; D014183:Transplantation, Heterologous
chemical_list: D003301:Copper Radioisotopes; D006573:Heterocyclic Compounds, 1-Ring; D007205:Indium Radioisotopes; D011506:Proteins; D018719:Erb-b2 Receptor Tyrosine Kinases; C071349:1,4,7,10-tetraazacyclododecane- 1,4,7,10-tetraacetic acid; C508053:ERBB2 protein, human