Novel Dual PET and Fluorescent Labelling Reagents for Multiscale Cell Tracking

作为医疗保健的第四支柱，基于细胞的免疫疗法提供了一种新的、快速发展的技术，具有改善人类疾病的巨大潜力。例如，嵌合抗原受体(CAR)技术正越来越多地被用于治疗癌症和其他类型的疾病。1 Maher(生物医学/临床联合主管)开发了一种名为T1E28z的CAR，它针对在几个实体肿瘤中上调的9个ErbB同源和异源二聚体中的8个。2使用T1E28z工程的T细胞的免疫治疗目前正在对局部晚期/复发的头颈部鳞状细胞癌患者进行I期临床试验评估(Clinicaltrials.gov NCT01818323)。3细胞治疗的医学研究和临床应用中的一个基本挑战是了解注入的细胞的体内行为。成像研究可以动态跟踪被管理细胞的迁移、增殖和最终命运。它将提供成功靶向的确凿证据，并允许量化细胞向靶点和非靶点的迁移程度。因此，在这种实验性治疗的临床前和临床开发的最早阶段纳入细胞跟踪研究是至关重要的，以提供对其安全性、作用机制和有效性的早期洞察。4采用正电子发射断层扫描(PET)、核磁共振(MRI)或荧光等单一方式的成像方法已被探索以动态跟踪注入的治疗细胞的持久性、迁移和增殖。4然而，我们对体内治疗细胞的命运的了解仍然有限，因为没有一种单一的成像方式能满足高分辨率、高灵敏度和深层组织渗透的需要。因此，跨细胞到宏观尺度研究受控细胞的体内行为的能力仍然是细胞跟踪的基本目标。配备双报告的细胞标记试剂可以通过组合使用两种成像模式的强度来克服其中一些限制。5例如，PET是高度敏感和非侵入性的。它可以产生放射性标记细胞的实时图像，并定量测量其全身分布，而不受探测深度的限制。通过选择适当的放射性核素，可以在临床相关的时间范围内获得连续的图像。同时，通过体外高分辨率荧光成像可以检测标记细胞在靶组织和非靶组织中的分布。这一跨学科合作项目旨在开发一种新型的双PET和荧光标记试剂，以跟踪从全身到微观尺度的治疗细胞。商业上可用的碘-124将被使用，因为它在临床使用的PET放射性同位素中具有最长的半衰期。原则上，碘-124可以为直接用PET标记的细胞提供尽可能长的跟踪时间。此外，通过标记试剂的分解代谢产生的任何游离碘-124的甲状腺和胃摄取都可以很容易地被碘化钾预处理所阻断。因此，它将提供低背景以可视化标记的细胞。然而，使用基于碘-124的有机生物标记试剂进行细胞标记的主要挑战是如何克服脱碘酶介导的脱碘反应。由于脱碘酶主要存在于细胞内，我们预计通过在细胞表面偶联基于碘-124的双重标记剂可以最大限度地减少脱碘。

Emerging as the fourth pillar of healthcare, cell-based immunotherapies offer a novel and rapidly developing technology that has great potential to ameliorate human disease. Exemplifying this, chimeric antigen receptor (CAR) technology is increasingly being harnessed for the treatment of cancer and other disease types.1 Maher (biomedical/clinical co-supervisor) developed a CAR named T1E28z that targets 8 of 9 ErbB homo- and heterodimers upregulated in several solid tumours.2 Immunotherapy using T1E28z-engineered T-cells is currently undergoing Phase I clinical trial evaluation in patients with locally advanced/ recurrent head and neck squamous cell carcinoma (clinicaltrials.gov NCT01818323).3 One fundamental challenge in both medical research and clinical applications of cell therapy is to understand the in vivo behaviours of the infused cells. Imaging studies can dynamically track the migration, proliferation, and final fate of the administered cells. It will provide definitive evidence of successful targeting and allow quantification of the degree of cell migration to the target and non-target sites. Thus, it is vital to incorporate cell tracking studies at the earliest stage of preclinical and clinical development of such experimental treatments to provide early insight into their safety, mechanism of action, and efficacy. 4 Imaging methods employed single modality such as positron emission tomography (PET), MRI, or fluorescence etc. have been explored to dynamically track the persistence, migration, and proliferation of the infused therapeutic cells.4 However, our understanding of the fate of the therapeutic cells in vivo remains limited because no single imaging modality meets the need for high resolution, high sensitivity, and deep tissue penetration. The ability to study the in vivo behaviour of the administered cells across cellular to macroscopic scales therefore remains a fundamental goal for cell tracking. Cell labelling reagents equipped with dual reporters can overcome some of these restraints by allowing combinational use of the strengths from both imaging modalities.5 For example, PET is highly sensitive and non-invasive. It can produce real-time images of the radiolabelled cells and quantitatively measure their whole-body distribution without the limitation of detection depth. By selecting an appropriate radionuclide, sequential images can be acquired over a clinically relevant timeframe. Meanwhile, the distribution of the labelled cells in the target and non-target tissues can be examined by ex vivo high-resolution fluorescence imaging.This interdisciplinary collaborative project aims to develop a novel dual PET and fluorescent labelling reagent to track the therapeutic cells from the whole body to the microscopic scale. The commercially available iodine-124 will be employed as it has the longest half-life among the clinically used PET radioisotopes. In principle, iodine-124 can provide the longest possible tracking time for directly labelled cells with PET. In addition, the thyroid and stomach uptake of any free iodine-124 generated through the catabolism of the labeling reagent can be readily blocked by the potassium iodide pre-treatment. Thus, it will provide a low background to visualise the labelled cells. However, the major challenge to use iodine-124 based organic bioconjugation reagents for cell labelling is how to overcome the deiodinases mediated deiodination. As the deiodinases mainly present intracellularly, we envisage that the deiodination would be minimised by coupling the iodine-124 based dual labelling reagent on the cell surface.