其中一种最具侵袭性的乳腺癌类型，涉及人类表皮生长因子受体2 (HER2) 的过度表达。HER2 在所有乳腺癌中过度表达约 25%，并与增加的增殖率、转移率和不良预后相关。自单克隆抗体特鲁齐单（Herceptin）及其他生物构建物的开发以来，HER2阳性乳腺癌的治疗显著提高。然而，患者常常出现耐药现象，显示对新型治疗的需求。纳米抗体已成为潜在开发HER2靶向成像剂和治疗剂的重要研究领域。纳米抗体具有许多有利特性，包括在热和非生理 pH 条件下具有高稳定性，同时保持与设计目标的低纳摩尔亲和力。具体而言，2Rs15d纳米抗体已用于HER2的靶向，并评估为单光子发射计算机断层扫描（SPECT）和正电子发射断层扫描（PET）的诊断成像剂。虽然带有正电子放射体 68Ga 的2Rs15d构建物目前处于I期临床试验阶段，但在临床前或临床研究中获得的 PET 图像仅在注射后3小时内获得。我们评估了我们自行制备的2Rs15d纳米抗体，与螯合剂除铁螯合酸（DFO）结合并与89Zr进行放射标记，以进行 PET 成像至注射后72小时。[89Zr]Zr-DFO-2Rs15d 在磷酸盐缓冲盐水（PBS）和人血清中表现出高稳定性。细胞结合研究显示高结合和特异性HER2，以及明显的内吞作用。我们的体内 PET 成像证实了HER2阳性肿瘤在注射后72小时内具有高质量的可视化效果，而HER2阴性肿瘤则无法观察到。随后的体内分布研究定量支持了显著的HER2阳性肿瘤摄取与阴性对照组的对比结果。我们的研究填补了对2Rs15d纳米抗体在延长时间点上的成像和结合特性的重要空白。由于许多治疗放射性同位素具有单日或多日的半衰期，这些信息将直接有益于2Rs15d对HER2阳性乳腺癌患者的放射治疗发展潜力。

One of the most aggressive forms of breast cancer involves the overexpression of human epidermal growth factor receptor 2 (HER2). HER2 is overexpressed in ∼25% of all breast cancers and is associated with increased proliferation, increased rates of metastasis, and poor prognosis. Treatment for HER2-positive breast cancer has vastly improved since the development of the monoclonal antibody trastuzumab (Herceptin) as well as other biological constructs. However, patients still commonly develop resistance, illustrating the need for newer therapies. Nanobodies have become an important focus for potential development as HER2-targeting imaging agents and therapeutics. Nanobodies have many favorable characteristics, including high stability in heat and nonphysiological pH, while maintaining their low-nanomolar affinity for their designed targets. Specifically, the 2Rs15d nanobody has been developed for targeting HER2 and has been evaluated as a diagnostic imaging agent for single-photon emission computed tomography (SPECT) and positron emission tomography (PET). While a construct of 2Rs15d with the positron emitter 68Ga is currently in phase I clinical trials, the only PET images acquired in preclinical or clinical research have been within 3 h postinjection. We evaluated our in-house produced 2Rs15d nanobody, conjugated with the chelator deferoxamine (DFO), and radiolabeled with 89Zr for PET imaging up to 72 h postinjection. [89Zr]Zr-DFO-2Rs15d demonstrated high stability in both phosphate-buffered saline (PBS) and human serum. Cell binding studies showed high binding and specificity for HER2, as well as prominent internalization. Our in vivo PET imaging confirmed high-quality visualization of HER2-positive tumors up to 72 h postinjection, whereas HER2-negative tumors were not visualized. Subsequent biodistribution studies quantitatively supported the significant HER2-positive tumor uptake compared to the negative control. Our studies fill an important gap in understanding the imaging and binding properties of the 2Rs15d nanobody at extended time points. As many therapeutic radioisotopes have single or multiday half-lives, this information will directly benefit the potential of the radiotherapy development of 2Rs15d for HER2-positive breast cancer patients.