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

尽管成年人群中甲状腺结节的发病率很高，但使用放射性碘的常规甲状腺核素扫描不能区分良性和恶性增殖。免疫组织化学和分子分析为Galectin3作为甲状腺癌标记物提供了强有力的证据，我们通过免疫成像技术对Galectin3在小鼠甲状腺肿瘤模型中的表达进行了初步的实验研究。在本项目中，我们的目标是进一步开发用于Galectin 3免疫PET成像的放射免疫偶联物。为了利用与全尺寸抗体相比更好的组织穿透性和更快的血浆清除速度，我们计划开发来自现有Galectin3特异性单抗的重组Fab片段，并用锆89对它们进行放射性标记。V基因将从杂交瘤细胞系中克隆出来，Fab片段将在大肠杆菌中进行功能性表达，如有必要，将通过蛋白质工程进行优化。产生的对Galectin3具有高亲和力的人嵌合Fab片段将通过与Lys侧链的偶联和利用新的化学方法以定点定向的方式随机偶联到金属特异性螯合剂化合物上，并用PET同位素进行放射性标记。这些放射免疫结合物将在携带不同甲状腺皮下和原位异种移植瘤的小鼠身上进行体外鉴定和体内成像。这种方法可能在将来允许术前区分甲状腺结节的良恶性，导致更好的患者选择和治疗决定，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.