HUMANIZED ANTITUMOR BINDING SITES--FRAMEWORK ENGINEERING

描述：（改编自申请人的摘要）抗体变量（V） 每个区域包含3个互补决定区（CDR）， 抗原结合位点和4个框架区（FR），其形成 支撑结合位点的支架。 这些片段穿插在 在每个V区中依次形成FR 1-CDR-1、FR 2-CDR-2-FR 3-CDR 3-FR 4， 整个结合位点需要VH和VL。 该提案探讨了 FR结构的修饰以改善免疫化学 和最小抗体结合位点的溶液性质，包括 单链Fv（sFv，包含VH-接头-VL）或单结构域V区。 在每个V结构域的折叠构象中，FR形成β桶 折叠片和CDR形成连接内部FR的环， 抗原结合位点。 来自人抗体的Fr序列可以融合 到CDR（一个被称为人源化的过程（其消除了大部分的人源化））。 由鼠抗原位点贡献的抗原性。 这项建议旨在 开发先进的FR设计，用于制备杂交sFv或可变 结构域，具有改善的稳定性/溶解性和降低的sFv 免疫原性 研究人员将制作一个人性化的26-10 抗地高辛sFv和具有抗溶菌酶结合的鼠-鼠杂合sFv 位点接枝到26-10骨架上。 此外，调查人员将 探索仅包含26-10 VH结构域的最小结合位点，作为 单体或单链同二聚体。 最后，他们将分析一个 工程化的人体结构， -gly-gly-ser-gly-gly-环以连接FR。 这种非功能性的“框架” 只有“分子”才能成为快速发展的基石， 表现为生物合成抗体结合位点。 这将允许任何FR 直接分析和优化上部结构， 干扰特定的结合位点;特异性CDR将被 重新插入以应用于肿瘤成像、治疗等。

DESCRIPTION: (Adapted from the Applicant's Abstract) Antibody variable (v) regions each comprise 3 complementary determining regions (CDRs) that form the antigen binding site and 4 framework regions (FRs) that form the scaffold supporting the binding site. These segments are interspersed sequentially in each V region to form FR1-CDR-1 FR2-CDR-2-FR3-CDR3-FR4, with the whole binding site requiring VH and VL. This proposal explores the modification of the FR structure for the improvement of immunochemical and solution properties of minimal antibody binding sites, consisting of single-chain Fv (sFv, comprising VH-linker-VL) or single domain V regions. In the folded conformation of each V domain, FRs form a barrel of beta pleated sheet and the CDRs form loops that connect the inner FRs to form the antigen binding site. Fr sequences from human antibodies can be fused to CDRs (a process termed humanization (which eliminates much of the antigenicity contributed by murine antigenic sites. This proposal seeks to develop advanced FR designs to be used in making hybrid sFv or variable domains, having improved stability/solubility and reduced sFv immunogenicity. The investigators will produce a humanized 26-10 antidigoxin sFv and a murine-murine hybrid sFv with an antilysozyme binding site grafted onto the 26-10 framework. In addition, the investigators will explore minimal binding sites comprising only the 26-10 VH domain, as a monomer or single-chain homodimer. Finally, they will analyze an engineered human framework which retains only simple -gly-gly-ser-gly-gly-loops to connect FRs. This nonfunctional "framework only" molecule could be a cornerstone for rapid development of very well behaved biosynthetic antibody binding sites. It would allow any FR superstructure to be directly analyzed and optimized without the perturbation of a particular binding site; specific CDRs would be reinserted for applications to tumor imaging, therapy, etc.