In vivo immuno-PET imaging of thyroid tumor nodules using Galectin-3 as tumor marker - Revised version

尽管甲状腺结节在成年人群中的患病率很高，但使用放射性碘的传统甲状腺造影术无法区分良性和恶性增殖。活检标本的免疫组织化学和分子分析提供了强有力的证据galectin3作为甲状腺癌的标志物，我们已经获得了初步的实验证据可视化galectin3表达的小鼠模型甲状腺肿瘤的免疫成像。在本项目中，我们的目标是进一步开发用于半乳糖凝集素3的免疫PET成像的放射性免疫缀合物。为了利用与全尺寸抗体相比更好的组织渗透和更快的血浆清除，我们计划开发来自现有半乳糖凝集素3特异性mAb的重组Fab片段，并用锆89放射性标记它们。将从杂交瘤细胞系中克隆V基因，并在E.大肠杆菌，如果需要，随后通过蛋白质工程优化。所得的对半乳糖凝集素3具有高亲和力的人嵌合Fab片段将偶联至金属特异性螯合剂化合物，均通过偶联至Lys侧链随机偶联和以利用新化学的位点定向方式偶联，并用PET同位素放射性标记。放射性免疫偶联物将在携带不同甲状腺皮下和原位异种移植肿瘤的小鼠中进行体外表征和体内成像。这种方法可能允许，在未来，良性和恶性甲状腺结节之间的术前区分，导致更好的患者选择和治疗决策，最终减少不必要的甲状腺手术治疗的数量。

Despite a high prevalence of thyroid nodules in the adult population, conventional thyroid scintigraphy using radioiodine does not allow the distinction between benign and malignant proliferation. Immunohistochemical and molecular analysis of biopsy specimens has provided strong evidence for galectin3 as thyroid cancer marker, and we have obtained preliminary experimental evidence about visualization of galectin3 expression in mouse models of thyroid tumour by immunoimaging. In the present project, we aim to further develop radio-immunoconjugates for immunoPET imaging of galectin3. To take advantage of the better tissue penetration and faster plasma clearance compared with full-size antibodies, we plan to develop recombinant Fab fragments derived from existing galectin3 specific mAbs and radiolabel them with zirconium89. The V genes will be cloned from hybridoma cell lines and the Fab fragments functionally expressed in E. coli, if necessary followed by optimization via protein engineering. Resulting human chimeric Fab fragments with high affinity toward galectin3 will be coupled to metal specific chelator compounds, both randomly via conjugation to Lys side chains and in a site directed fashion utilizing novel chemistries, and radiolabelled with the PET isotopes. The radio-immunoconjugates will be subjected to in vitro characterization and in vivo imaging in mice that carry different thyroid subcutaneous and orthotopic xenograft tumors. This approach may allow, in the future, the preoperative discrimination between benign and malignant thyroid nodules, leading to better patient selection and therapy decision, eventually reducing the number of unnecessary thyroid surgical treatments.