Colour-coded surgery in Neuroblastoma: developing a dual PET/Near-Infrared Fluorescence Imaging probe to visualise tumour from diagnosis to resection

神经母细胞瘤是儿童中第二大常见的实体肿瘤，在英国每年约有100名新儿童受其影响。神经母细胞瘤的主要手术挑战之一是如何切除整个肿瘤。这尤其困难，因为成神经细胞瘤的特征性弥漫性生长在腹腔和/或胸腔的后部，而且肿瘤严格附着在主要血管上，增加了严重出血的风险。因此，有必要开发一种肿瘤特异性检测策略，以帮助外科医生提供活的肿瘤细胞与应该保存的正常解剖结构的简单可视化。我们假设，开发一种从诊断到随访追踪活性肿瘤细胞的单一方法，包括手术期间的实时可视化，将是治疗这种侵袭性儿科癌症的重大进步。在过去的十年里，两种成像模式已经展示了改变我们治疗癌症方式的潜力。首先，正电子发射断层扫描（PET）已成为监测癌症对治疗反应的有力工具。其次，近红外荧光（NIRF）成像已被证明是一种非常有前途的图像引导肿瘤组织切除技术，因为它有助于在手术过程中实时、高分辨率地描绘肿瘤边缘。我们的目标是选择性地标记神经母细胞瘤细胞，以便在治疗的不同阶段和术中使用荧光（NIRF）通过全身扫描（PET）对其进行可视化。简而言之，在项目的前18个月，我们将与市场上不同的荧光分子结合一种常用的抗神经母细胞瘤特异性抗体（抗gd2单克隆抗体- mab）。这将使我们了解荧光是否可以选择性地附着在体外的神经母细胞瘤细胞上（目的1）。然后，我们将在成熟的神经母细胞瘤动物模型中进行一系列实验，以证实从体内不同器官观察神经母细胞瘤的可能性（目标2）。作为该目标的一部分，我们将基于荧光成像（NIRF）对小鼠肿瘤进行手术解剖。下一步（目标3）将创建第二个标签，通过PET扫描显示肿瘤。如果上述目标成功，我们将能够注册知识产权，并与专门从事单克隆抗体制备和工程的公司联系，按照良好生产规范（GMP）方法开发双重标记的NIRF/PET单克隆抗体。在项目的第二个18个月，我们将进行一项临床试验，以测试双重NIRF/PET探针在人体中的安全性和有效性（目标4）。学术合作伙伴和合作者提供的专业知识和设施非常适合开发这个项目（参见支持案例）。在项目结束时，我们期望有一种新的分子可以用于治疗患有神经母细胞瘤的儿童。这将在不同治疗期间有效监测活性肿瘤细胞，更容易手术切除，更客观地评估肿瘤残留，以及更好的术后长期随访方面带来巨大的优势。新型的NIRF/PET分子也将允许个体化手术治疗神经母细胞瘤。由于外科医生能更好地观察肿瘤，患者将接受范围更小、更有针对性的手术，并发症的风险也更低。这可能是手术治疗这种恶性疾病进展中的一个里程碑，可以提高生存率并减少残留疾病的机会。

Neuroblastoma is the second most common solid tumour in children, and it affects around 100 new children a year in the UK. One of the main surgical challenges in the treatment of neuroblastoma is to be able to remove the entire tumour. This is particularly difficult because of the characteristic diffuse growth of neuroblastoma in the posterior part of the abdominal and/or thoracic cavities and the fact the tumour is strictly adherent to major blood vessels increasing the risk of severe bleeding. Therefore, there is the need to develop a tumour-specific detection strategy that could assist surgeons in providing easy visualisation of viable tumour cells versus normal anatomical structures that should be preserved. We hypothesise that developing a single way to track active tumour cells from diagnosis to follow-up, including their real-time visualisation during surgery, will be a major advance in the treatment of this aggressive paediatric cancer. Over the past decade, two imaging modalities have demonstrated the potential of transforming the way we treat cancer. First, positron emission tomography (PET) has become a powerful tool in monitoring the cancer response to treatments. Second, near-infrared fluorescence (NIRF) imaging has proven to be a very promising technique for the image-guided resection of tumour tissue, as it facilitates the real-time, high-resolution delineation of tumour margins during surgery. We aim to selectively label neuroblastoma cells so they can be visualised with a whole-body scan (PET) during the different phases of treatment and intraoperatively with the use of fluorescence (NIRF). Briefly, in the first 18 months of the project, we will conjugate a commonly used specific antibody against neuroblastoma (anti-GD2 monoclonal antibody-mAb) with different fluorescent molecules available on the market. This will allow us to understand if fluorescence can be selectively attached to neuroblastoma cells in vitro (Objective 1). Then we will run a series of experiments in well-established animal models of neuroblastoma to confirm the possibility of visualising neuroblastoma from different organs in vivo (objective 2). As part of this objective, we will carry on surgical dissections of the tumour in mice based on fluorescent imaging (NIRF). The next step (objective 3) will be to create a second labelling to visualise the tumour with a PET scan. If the above objectives are successful, we will be able to register the intellectual property and to link with companies specialised in the preparation and engineering of monoclonal antibodies to develop the dual labelled NIRF/PET monoclonal antibodies following Good Manufacturing Practice (GMP) methods. In the second 18 months of the project, we will run a clinical trial to test the safety and efficacy of the dual NIRF/PET probe in humans (objective 4).The expertise and facilities provided by the academic partner and collaborators are excellent to develop this project (see Case for Support). At the end of the project, we expect to have a novel molecule that can be used in the cure of children with neuroblastoma. This will bring huge advantages in term of effective monitoring of active tumour cells during different treatments, easier surgical resection with a more objective assessment of tumour residuals, and better long term post-surgical follow-up. The novel NIRF/PET molecule will also allow personalised surgery to treat neuroblastoma. As a consequence of better visualisation of the tumour by the surgeon, patients will receive less extensive and more targeted surgery with less risk for complications. This can be a milestone in the progress of surgery for this malignant disease and can lead to increase survival and reduce the chance of residual disease.