Hammond Scott A, Lutterbuese Ralf, Roff Shannon, Lutterbuese Petra, Schlereth Bernd, Bruckheimer Elizabeth, Kinch Michael S, Coats Steve, Baeuerle Patrick A, Kufer Peter, Kiener Peter A
Medimmune, Inc., Gaithersburg, Maryland 20878, USA.
Cancer Res. 2007 Apr 15;67(8):3927-35. doi: 10.1158/0008-5472.CAN-06-2760.
The EphA2 receptor tyrosine kinase is frequently overexpressed and functionally altered in malignant cells and thus provides opportunities for selective targeting of tumor cells. We describe here the development of a novel, bispecific single-chain antibody (bscAb) referred to as bscEphA2xCD3. This molecule simultaneously targets EphA2 on tumor cells and the T-cell receptor/CD3 complex on T cells and possesses structural and functional characteristics of the recently developed BiTE technology. An EphA2-specific single-chain antibody was selected for recognition of an epitope that is preferentially exposed on malignant cells based on the concept of epitope exclusion; this was fused to a CD3-specific single-chain antibody to generate bscEphA2xCD3. The resultant bscAb redirected unstimulated human T cells to lyse EphA2-expressing tumor cells both in vitro and in vivo. In separate experiments, efficient tumor cell lysis was achieved in vitro at drug concentrations <or=1 microg/mL, at a low T-cell effector-to-tumor target cell ratio (1:1), and with tumor cells that possess few available binding sites (2,400 per cell) for bscEphA2xCD3. Time-lapsed microscopy revealed potent cytotoxic activity of bscEphA2xCD3-activated T cells against monolayers of malignant cells but not against monolayers of nontransformed EphA2-positive cells except at the edges of the monolayer where the target epitope was exposed. BscEphA2xCD3 was also efficacious in human xenograft mouse models modified to show human T-cell killing of tumors. Together, our results reveal opportunities for redirecting the potent activity of cytotoxic T cells towards tumor cells that express selectively accessible epitopes and establish EphA2-specific bscAb molecules as novel and potent therapeutics with selectivity for tumor cells.
EphA2受体酪氨酸激酶在恶性细胞中经常过度表达且功能改变，因此为肿瘤细胞的选择性靶向提供了机会。我们在此描述了一种新型双特异性单链抗体（bscAb），称为bscEphA2xCD3的研发情况。该分子同时靶向肿瘤细胞上的EphA2和T细胞上的T细胞受体/CD3复合物，并具有最近开发的双特异性T细胞衔接器（BiTE）技术的结构和功能特征。基于表位排斥概念，选择了一种EphA2特异性单链抗体来识别优先暴露于恶性细胞上的表位；将其与CD3特异性单链抗体融合以产生bscEphA2xCD3。所得的bscAb在体外和体内均可使未受刺激的人T细胞重定向，以裂解表达EphA2的肿瘤细胞。在单独的实验中，在药物浓度≤1μg/mL、低T细胞效应细胞与肿瘤靶细胞比例（1:1）以及对bscEphA2xCD3具有很少可用结合位点（每个细胞2400个）的肿瘤细胞的情况下，体外实现了有效的肿瘤细胞裂解。延时显微镜显示，bscEphA2xCD3激活的T细胞对恶性细胞单层具有强大的细胞毒活性，但对未转化的EphA2阳性细胞单层则无此活性，除非在单层边缘靶表位暴露处。bscEphA2xCD3在经改造以显示人T细胞对肿瘤的杀伤作用的人异种移植小鼠模型中也有效。总之，我们的结果揭示了将细胞毒性T细胞的强大活性重定向至表达选择性可及表位的肿瘤细胞的机会，并确立了EphA2特异性bscAb分子作为对肿瘤细胞具有选择性的新型强效治疗剂。
