Site-specific antibody-toxin conjugates for cancer therapy

描述（由申请人提供）：抗体提供了一种利用细菌蛋白毒素的极端效力的方法-能够在皮摩尔浓度下杀死健康和转化的细胞-尽管目前没有方法将这些毒素与高亲和力的二价单克隆抗体联系起来。位点特异性附着在这种应用中是至关重要的，因为这两种蛋白质都含有不能被干扰的区域，以维持肿瘤靶向性和细胞毒性。原则上，这可以通过生产单克隆抗体毒素作为重组融合蛋白来实现。然而，以目前的技术，这是不可能的，因为生产单克隆抗体所需的真核细胞被蛋白质毒素的表达杀死。我们已经描述了由细菌酶分类酶A （SrtA）催化的两个反应，这些反应允许单抗和毒素通过有效的、经过验证的方法分别产生，然后在体外特异性地结合。我们相信这将提供一种将蛋白毒素（如白喉毒素和假单胞菌外毒素a）与独立于其靶抗原的单克隆抗体结合的通用机制，为产生针对多种肿瘤抗原的抗体-毒素偶联物提供了一种方法，而单克隆抗体已经可用。

DESCRIPTION (provided by applicant): Antibodies offer a means to harness the extreme potency of bacterial protein toxins-capable of killing both healthy and transformed cells at picomolar concentrations-though no methods are currently available to link these toxins to high-affinity, bivalent mAbs. Site-specific attachment is critical in this application, as both proteins contain regions that cannot be perturbed in order to maintain tumor targeting and cytotoxicity. In principle, this could be accomplished by producing a mAb-toxin as a recombinant fusion protein. However, this is not possible with current technology, as the eukaryotic cells required to produce mAbs are killed by expression of protein toxins. We have characterized two reactions catalyzed by the bacterial enzyme sortase A (SrtA) that allow the mAb and the toxin to be produced separately by effective, proven methods, then site- specifically joined in vitro. We believe that this will provide a general mechanism to conjugate protein toxins such as diphtheria toxin and pseudomonas exotoxin A to mAbs independent of their target antigen, providing a means to generate antibody-toxin conjugates against a variety of tumor antigens for which mAbs are already available.