B lymphocytes are a key pathologic feature of multiple sclerosis (MS) and are becoming an important therapeutic target for this condition. Currently, there is no approved technique to noninvasively visualize B cells in the central nervous system (CNS) to monitor MS disease progression and response to therapies. Here, we evaluated ⁶⁴Cu-rituximab, a radiolabeled antibody specifically targeting the human B cell marker CD20, for its ability to image B cells in a mouse model of MS using PET. Methods: To model CNS infiltration by B cells, experimental autoimmune encephalomyelitis (EAE) was induced in transgenic mice that express human CD20 on B cells. EAE mice were given subcutaneous injections of myelin oligodendrocyte glycoprotein fragment_1–125 emulsified in complete Freund adjuvant. Control mice received complete Freund adjuvant alone. PET imaging of EAE and control mice was performed 1, 4, and 19 h after ⁶⁴Cu-rituximab administration. Mice were perfused and sacrificed after the final PET scan, and radioactivity in dissected tissues was measured with a γ-counter. CNS tissues from these mice were immunostained to quantify B cells or were further analyzed via digital autoradiography. Results: Lumbar spinal cord PET signal was significantly higher in EAE mice than in controls at all evaluated time points (e.g., 1 h after injection: 5.44 ± 0.37 vs. 3.33 ± 0.20 percentage injected dose [%ID]/g, P < 0.05). ⁶⁴Cu-rituximab PET signal in brain regions ranged between 1.74 ± 0.11 and 2.93 ± 0.15 %ID/g for EAE mice, compared with 1.25 ± 0.08 and 2.24 ± 0.11 %ID/g for controls (P < 0.05 for all regions except striatum and thalamus at 1 h after injection). Similarly, ex vivo biodistribution results revealed notably higher ⁶⁴Cu-rituximab uptake in the brain and spinal cord of huCD20tg EAE, and B220 immunostaining verified that increased ⁶⁴Cu-rituximab uptake in CNS tissues corresponded with elevated B cells. Conclusion: B cells can be detected in the CNS of EAE mice using ⁶⁴Cu-rituximab PET. Results from these studies warrant further investigation of ⁶⁴Cu-rituximab in EAE models and consideration of use in MS patients to evaluate its potential for detecting and monitoring B cells in the progression and treatment of this disease. These results represent an initial step toward generating a platform to evaluate B cell–targeted therapeutics en route to the clinic.

B淋巴细胞是多发性硬化症（MS）的关键病理特征，正在成为这种疾病的重要治疗靶标。当前，尚无批准的技术可无创地观察中枢神经系统（CNS）中的B细胞，以监测MS疾病的进展和对治疗的反应。在这里，我们评估了⁶⁴ Cu-rituximab（一种专门针对人B细胞标记CD20的放射性标记抗体）在使用PET的MS小鼠模型中对B细胞成像的能力。方法：为模拟B细胞对CNS的浸润，在B细胞上表达人CD20的转基因小鼠中诱导了实验性自身免疫性脑脊髓炎（EAE）。EAE小鼠皮下注射髓鞘少突胶质细胞糖蛋白片段_1–125在完全的弗氏佐剂中乳化。对照小鼠仅接受完全的弗氏佐剂。在⁶⁴ Cu-利妥昔单抗给药后1、4和19 h对EAE和对照小鼠进行PET成像。在最后一次PET扫描后对小鼠进行灌注并处死，并使用γ计数器测量解剖组织中的放射性。对这些小鼠的CNS组织进行免疫染色以定量B细胞，或通过数字放射自显影进一步分析。结果：在所有评估的时间点（例如，注射后1 h：5.44±0.37 vs. 3.33±0.20百分比注射剂量[％ID] / g，EAE小鼠的腰脊髓PET信号显着高于对照组），P <0.05 ）。⁶⁴对于EAE小鼠，大脑区域中的Cu-rituximab PET信号范围在1.74±0.11和2.93±0.15％ID / g之间，而对照组的1.25±0.08和2.24±0.11％ID / g（除纹状体和其他区域以外的所有区域P <0.05注射后1小时的丘脑）。同样，离体生物分布结果显示，huCD20tg EAE的大脑和脊髓中⁶⁴ Cu-rituximab的摄取显着增加，B220免疫染色证实CNS组织中⁶⁴ Cu-rituximab摄取的增加与B细胞升高相对应。结论：使用⁶⁴ Cu-rituximab PET可以在EAE小鼠的中枢神经系统中检测到B细胞。这些研究的结果值得进一步调查⁶⁴在EAE模型中使用Cu-rituximab并考虑在MS患者中评估其在疾病进展和治疗中检测和监测B细胞的潜力。这些结果代表了朝着建立一个平台以评估针对B细胞的治疗药物到临床的第一步。