Mertens Nico, Devos Filip, Leoen Jannick, Van Deynse Els, Willems An, Schoonooghe Steve, Burvenich Ingrid, De Koker Stefaan, Vlieghe Dominique, Grooten Johan, Kelly Andrew, Van de Wiele Christophe
Department of Molecular Biomedical Research, Flanders Interuniversity Institute of Biotechnology (VIB), Ghent University, Ghent, Belgium.
Cancer Biother Radiopharm. 2004 Feb;19(1):99-109. doi: 10.1089/108497804773391748.
The synthesis of radioligands can benefit considerably from optimized recombinant protein production, both on the aspect of economy of production and on the level of improving the targeting and pharmacokinetics of the ligand. This paper first describes a general production optimization strategy, and then elaborates on a protein design strategy tailored to targeting applications. Production in Escherichia coli will benefit from economy of goods and time as compared to other organisms. In order to increase the chance of finding a successful production system in this host, we have assembled a large number of expression strategies in a single, uniform expression system (FastScreen). The system allows rapid optimization of direct production of native proteins or via a fusion protein strategy with subsequent recovery of the desired protein. As an example of recombinant radioligand synthesis for improved targeting and clearing, a manifold of intermediate molecular size was synthesized by fusing one Fab and two single-chain variable fragments (scFv) antibody binding fragments into a trifunctional molecule (Tribody). Due to the use of the specific heterodimerization of the Fab chains, trispecific, bispecific, or trivalent antibody derived targeting reagents can easily be obtained. Recombinant production techniques also allow for specific incorporation of amino acids favoring a site specific labeling (labeling tags).
放射性配体的合成在生产经济性以及改善配体的靶向性和药代动力学水平方面，都能从优化的重组蛋白生产中受益匪浅。本文首先描述了一种通用的生产优化策略，然后详细阐述了一种针对靶向应用的蛋白质设计策略。与其他生物体相比，在大肠杆菌中生产将在成本和时间方面受益。为了增加在这种宿主中找到成功生产系统的机会，我们在一个统一的表达系统（快速筛选）中整合了大量表达策略。该系统允许快速优化天然蛋白质的直接生产，或通过融合蛋白策略随后回收所需蛋白质。作为用于改善靶向和清除的重组放射性配体合成的一个例子，通过将一个Fab和两个单链可变片段（scFv）抗体结合片段融合成一个三功能分子（三特异性抗体），合成了多种中等分子大小的物质。由于使用了Fab链的特异性异源二聚化，很容易获得三特异性、双特异性或三价抗体衍生的靶向试剂。重组生产技术还允许特异性掺入有利于位点特异性标记的氨基酸（标记标签）。
