Novel Dual PET and Fluorescent Labelling Reagents for Multiscale Cell Tracking

作为医疗保健的第四大支柱，基于细胞的免疫疗法提供了一种新颖且快速发展的技术，具有改善人类疾病的巨大潜力。举例来说，嵌合抗原受体（CAR）技术越来越多地用于治疗癌症和其他疾病类型。1 Maher（生物医学/临床共同主管）开发了一种名为T1 E28 z的CAR，其靶向在几种实体瘤中上调的9种ErbB同源和异源二聚体中的8种。cells目前正在局部晚期/复发性头颈部鳞状细胞癌患者中进行I期临床试验评价（clinicaltrials.gov NCT 01818323）。3细胞疗法的医学研究和临床应用中的一个基本挑战是了解输注细胞的体内行为。成像研究可以动态地跟踪所施用细胞的迁移、增殖和最终命运。它将提供成功靶向的明确证据，并允许量化细胞迁移到靶位点和非靶位点的程度。因此，在这些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至关重要，以早期了解其安全性，作用机制和疗效。4采用单一模式的成像方法，如正电子发射断层扫描（PET）、MRI或荧光等，已被探索用于动态跟踪输注治疗细胞的持续性、迁移和增殖。4然而，我们对治疗细胞在体内的命运的理解仍然有限，因为没有单一的成像模式满足高分辨率、高灵敏度和深组织穿透的需求。因此，在细胞到宏观尺度上研究所施用细胞的体内行为的能力仍然是细胞追踪的基本目标。配备双报告基因的细胞标记试剂可以通过结合使用两种成像模式的优势来克服其中一些限制。5例如，PET具有高度灵敏性和非侵入性。它可以产生放射性标记细胞的实时图像，并定量测量它们的全身分布，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采集连续图像。该跨学科合作项目旨在开发一种新型的双重PET和荧光标记试剂，用于从全身到微观尺度追踪治疗细胞。将使用市售碘-124，因为其在临床使用的PET放射性同位素中半衰期最长。原则上，碘-124可以为PET直接标记的细胞提供最长的跟踪时间。此外，甲状腺和胃对通过标记试剂的催化剂产生的任何游离碘-124的摄取可以容易地被碘化钾预处理阻断。因此，它将提供低背景以使标记的细胞可视化。然而，使用基于碘-124的有机生物缀合试剂用于细胞标记的主要挑战是如何克服脱碘酶介导的脱碘。由于脱碘酶主要存在于细胞内，我们设想通过在细胞表面上偶联基于碘-124的双标记试剂来使脱碘最小化。

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.