侵袭性转移性乳腺癌的治疗对于现代生物医学来说仍然是一个巨大的挑战。生物相容性聚合物纳米粒子已在临床上成功应用，并被视为潜在的解决方案。具体来说，研究人员正在探索开发靶向肿瘤细胞膜上相关受体（如HER2）的化疗纳米药物。然而，尚未有针对人类癌症治疗获批准的靶向纳米药物。正在开发新的策略来改变代理的构架并优化其系统管理。在这里，我们描述了这些方法的组合，即有针对性的聚合物纳米载体的设计和一种其向肿瘤部位的系统投递方法。具体而言，我们使用载有诊断染料Nile Blue和化疗化合物Doxorubicin的PLGA纳米胶囊作为两步靶向递送的概念，通过 barnase/barstar蛋白“细菌超级粘合剂”的肿瘤靶向预处理。第一个预处理组分包括抗HER2支架蛋白DARPin9_29与barstar相融合的Bs-DARPin9_29，第二个组分包括化疗PLGA纳米胶囊结合barnase的PLGA-Bn。该系统的有效性在体内进行了评估。为此，我们开发了一种稳定表达人类HER2癌标志物的免疫活性BALB/c小鼠肿瘤模型，以测试癌症治疗纳米PLGA的两步递送潜力。体外和体外研究证实，肿瘤中HER2受体表达稳定，使其成为评估HER2定向药物的可行工具。我们证明，两步递送比单步递送更有效，无论是成像还是治疗肿瘤：两步递送的成像能力比单步更高，肿瘤生长抑制率为94.9％，而单步策略为68.4％。 barnase/barstar蛋白配对已被证明具有优异的生物相容性，通过评估免疫原性和血液毒性的生物安全性测试成功完成。这使得该蛋白配对成为一种高度通用的肿瘤预靶向工具，能够针对不同分子特征的肿瘤进行个性化治疗的发展。

Therapy for aggressive metastatic breast cancer remains a great challenge for modern biomedicine. Biocompatible polymer nanoparticles have been successfully used in clinic and are seen as a potential solution. Specifically, researchers are exploring the development of chemotherapeutic nanoagents targeting the membrane-associated receptors of cancer cells, such as HER2. However, there are no targeting nanomedications that have been approved for human cancer therapy. Novel strategies are being developed to alter the architecture of agents and optimize their systemic administration. Here, we describe a combination of these approaches, namely, the design of a targeted polymer nanocarrier and a method for its systemic delivery to the tumor site. Namely, PLGA nanocapsules loaded with a diagnostic dye, Nile Blue, and a chemotherapeutic compound, doxorubicin, are used for two-step targeted delivery using the concept of tumor pre-targeting through the barnase/barstar protein "bacterial superglue". The first pre-targeting component consists of an anti-HER2 scaffold protein, DARPin9_29 fused with barstar, Bs-DARPin9_29, and the second component comprises chemotherapeutic PLGA nanocapsules conjugated to barnase, PLGA-Bn. The efficacy of this system was evaluated in vivo. To this aim, we developed an immunocompetent BALB/c mouse tumor model with a stable expression of human HER2 oncomarkers to test the potential of two-step delivery of oncotheranostic nano-PLGA. In vitro and ex vivo studies confirmed HER2 receptor stable expression in the tumor, making it a feasible tool for HER2-targeted drug evaluation. We demonstrated that two-step delivery was more effective than one-step delivery for both imaging and tumor therapy: two-step delivery had higher imaging capabilities than one-step and a tumor growth inhibition of 94.9% in comparison to 68.4% for the one-step strategy. The barnase*barstar protein pair has been proven to possess excellent biocompatibility, as evidenced by the successful completion of biosafety tests assessing immunogenicity and hemotoxicity. This renders the protein pair a highly versatile tool for pre-targeting tumors with various molecular profiles, thereby enabling the development of personalized medicine.