Department of Radiology, Washington University School of Medicine, St. Louis, Missouri.
Department of Biomedical Engineering, Washington University in St. Louis, St. Louis, Missouri.
J Nucl Med. 2018 Feb;59(2):216-222. doi: 10.2967/jnumed.117.196063. Epub 2017 Oct 12.
Multiple myeloma (MM) is a plasma B-cell hematologic cancer that causes significant skeletal morbidity. Despite improvements in survival, heterogeneity in response remains a major challenge in MM. Cluster of differentiation 38 (CD38) is a type II transmembrane glycoprotein overexpressed in myeloma cells and is implicated in MM cell signaling. Daratumumab is a U.S. Food and Drug Administration-approved high-affinity monoclonal antibody targeting CD38 that is clinically benefiting refractory MM patients. Here, we evaluated [Zr]Zr-desferrioxamine (DFO)-daratumumab PET/CT imaging in MM tumor models. Daratumumab was conjugated to DFO--benzyl-isothiocyanate (DFO-Bz-NCS) for radiolabeling with Zr. Chelator conjugation was confirmed by electrospray ionization-mass spectrometry, and radiolabeling was monitored by instant thin-layer chromatography. Daratumumab was conjugated to Cyanine5 (Cy5) dye for cell microscopy. In vitro and in vivo evaluation of [Zr]Zr-DFO-daratumumab was performed using CD38 human myeloma MM1.S- (MM1.S) cells. Cellular studies determined the affinity, immunoreactivity, and specificity of [Zr]Zr-DFO-daratumumab. A 5TGM1- (5TGM1)/KaLwRij MM mouse model served as control for imaging background noise. [Zr]Zr-DFO-daratumumab PET/CT small-animal imaging was performed in severe combined immunodeficient mice bearing solid and disseminated MM tumors. Tissue biodistribution (7 d after tracer administration, 1.11 MBq/animal, = 4-6/group) was performed in wild-type and MM1.S tumor-bearing mice. A specific activity of 55.5 MBq/nmol (0.37 MBq/μg) was reproducibly obtained with [Zr]Zr-daratumumab-DFO. Flow cytometry confirmed CD38 expression (>99%) on the surface of MM1.S cells. Confocal microscopy with daratumumab-Cy5 demonstrated specific cell binding. Dissociation constant, 3.3 nM (±0.58), and receptor density, 10.1 fmol/mg (±0.64), was obtained with a saturation binding assay. [Zr]Zr-DFO-daratumumab/PET demonstrated specificity and sensitivity for detecting CD38 myeloma tumors of variable sizes (8.5-128 mm) with standardized uptake values ranging from 2.1 to 9.3. Discrete medullar lesions, confirmed by bioluminescence images, were efficiently imaged with [Zr]Zr-DFO-daratumumab/PET. Biodistribution at 7 d after administration of [Zr]Zr-DFO-daratumumab showed prominent tumor uptake (27.7 ± 7.6 percentage injected dose per gram). In vivo blocking was achieved with a 200-fold excess of unlabeled daratumumab. [Zr]Zr-DFO- and Cy5-daratumumab demonstrated superb binding to CD38 human MM cells and significantly low binding to CD38 cells. Daratumumab bioconjugates are being evaluated for image-guided delivery of therapeutic radionuclides.
多发性骨髓瘤（MM）是一种浆细胞血液系统恶性肿瘤，会导致严重的骨骼发病率。尽管生存有所改善，但反应的异质性仍然是 MM 的主要挑战。CD38 是一种在骨髓瘤细胞中过度表达的 II 型跨膜糖蛋白，与 MM 细胞信号有关。Daratumumab 是一种美国食品和药物管理局批准的靶向 CD38 的高亲和力单克隆抗体，可使难治性 MM 患者受益。在这里，我们评估了[Zr]Zr-去铁胺（DFO）-daratumumab PET/CT 成像在 MM 肿瘤模型中的应用。Daratumumab 与 DFO-苯异硫氰酸酯（DFO-Bz-NCS）缀合用于 Zr 的放射性标记。电喷雾电离质谱法证实了螯合剂的缀合，通过即时薄层层析监测放射性标记。Daratumumab 与 Cy5 染料缀合用于细胞显微镜。使用 CD38 人类骨髓瘤 MM1.S-（MM1.S）细胞进行 [Zr]Zr-DFO-daratumumab 的体外和体内评估。细胞研究确定了 [Zr]Zr-DFO-daratumumab 的亲和力、免疫反应性和特异性。5TGM1-（5TGM1）/KaLwRij MM 小鼠模型作为成像背景噪声的对照。在携带实体和弥散性 MM 肿瘤的严重联合免疫缺陷小鼠中进行 [Zr]Zr-DFO-daratumumab 的 PET/CT 小动物成像。在野生型和 MM1.S 肿瘤荷瘤小鼠中进行了组织生物分布（示踪剂给药后 7 天，1.11 MBq/动物，每组 4-6 只）。用 [Zr]Zr-daratumumab-DFO 可重复性地获得 55.5 MBq/nmol（0.37 MBq/μg）的比活度。流式细胞术证实 MM1.S 细胞表面表达 CD38（>99%）。用 daratumumab-Cy5 进行共焦显微镜显示出特异性细胞结合。通过饱和结合测定获得 3.3 nM（±0.58）的解离常数和 10.1 fmol/mg（±0.64）的受体密度。[Zr]Zr-DFO-daratumumab/PET 特异性和灵敏度均用于检测大小（8.5-128 mm）不同的 CD38 骨髓瘤肿瘤，标准摄取值范围为 2.1-9.3。通过生物发光图像证实离散的骨髓病变可有效成像。[Zr]Zr-DFO-daratumumab 给药 7 天后的生物分布显示肿瘤摄取明显（27.7±7.6%注入剂量/克）。用未标记的 daratumumab 200 倍过量进行体内阻断。[Zr]Zr-DFO 和 Cy5-daratumumab 与 CD38 人类 MM 细胞具有极好的结合能力，与 CD38 细胞的结合能力明显较低。Daratumumab 生物缀合物正在被评估用于放射性核素的图像引导递送。
