Cysteine cathepsins often contribute to cancer progression due to their overexpression in the tumour microenvironment and therefore present attractive targets for non-invasive diagnostic imaging. However, the development of highly selective and versatile small molecule probes for cathepsins has been challenging. Here, we targeted tumour-associated cathepsin B using designed ankyrin repeat proteins (DARPins). The selective DARPin 8h6 inhibited cathepsin B with picomolar affinity (K_i = 35 pM) by binding to a site with low structural conservation in cathepsins, as revealed by the X-ray structure of the complex. DARPin 8h6 blocked cathepsin B activity in tumours ex vivo and was successfully applied in in vivo optical imaging in two mouse breast cancer models, in which cathepsin B was bound to the cell membrane or secreted to the extracellular milieu by tumour and stromal cells. Our approach validates cathepsin B as a promising diagnostic and theranostic target in cancer and other inflammation-associated diseases.

中文翻译：

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高选择性抑制性DARPin对肿瘤相关组织蛋白酶B的无创体内成像

半胱氨酸组织蛋白酶由于其在肿瘤微环境中的过表达而常常促进癌症进展，因此为非侵入性诊断成像提供了有吸引力的靶标。然而，用于组织蛋白酶的高度选择性和通用的小分子探针的开发一直具有挑战性。在这里，我们使用设计的锚蛋白重复蛋白（DARPins）靶向与肿瘤相关的组织蛋白酶B。选择性DARPin 8h6通过与组织蛋白酶中低结构保守性的位点结合，以皮摩尔亲和力（K _i = 35 pM）抑制组织蛋白酶B ，如复合物的X射线结构所示。DARPin 8h6阻断了体内组织蛋白酶B的活性，并成功应用于体内在两个小鼠乳腺癌模型中的光学成像，其中组织蛋白酶B通过肿瘤和基质细胞结合到细胞膜上或分泌到细胞外环境中。我们的方法验证了组织蛋白酶B在癌症和其他炎症相关疾病中是有前途的诊断和治疗目标。