Institut Galien Paris-Sud, UMR 8612, CNRS, Univ Paris-Sud, Université Paris-Saclay, Faculté de Pharmacie, 5 rue Jean-Baptiste Clément, F-92296 Châtenay-Malabry cedex, France.
Institut Paris-Saclay d'Innovation Thérapeutique (IPSIT), UMS IPSIT Université Paris-Sud US 31 INSERM, UMS 3679 CNRS, Microscopy Facility, 5 rue Jean-Baptiste Clément, 92296 Châtenay-Malabry cedex, France.
Eur J Pharm Biopharm. 2019 Sep;142:195-203. doi: 10.1016/j.ejpb.2019.06.019. Epub 2019 Jun 20.
We recently constructed a multicellular spheroid model of pancreatic tumor based on a triple co-culture of cancer cells, fibroblasts and endothelial cells and characterized by the presence of fibronectin, an important component of the tumor extracellular matrix. By combining cancer cells and stromal components, this model recreates in vitro the three-dimensional (3D) architecture of solid tumors. In this study, we used these hetero-type spheroids as a tool to assess the penetration of doxorubicin (used as a model drug) through the whole tumor mass either in a free form or loaded into polymer nanoparticles (NPs), and we investigated whether microscopy images, acquired by Confocal Laser Scanning Microscopy (CLSM) and Light Sheet Fluorescence Microscopy (LSFM), would be best to provide reliable information on this process. Results clearly demonstrated that CLSM was not suitable to accurately monitor the diffusion of small molecules such as the doxorubicin. Indeed, it only allowed to scan a layer of 100 µm depth and no information on deeper layers could be available because of a progressive loss of the fluorescence signal. On the contrary, a complete 3D tomography of the hetero-type multicellular tumor spheroids (MCTS) was obtained by LSFM and multi-view image fusion which revealed that the fluorescent molecule was able to reach the core of spheroids as large as 1 mm in diameter. However, no doxorubicin-loaded polymer nanoparticles were detected in the spheroids, highlighting the challenge of nanomedicine delivery through biological barriers. Overall, the combination of hetero-type MCTS and LSFM allowed to carry out a highly informative microscopic assessment and represents a suitable approach to precisely follow up the drug penetration in tumors. Accordingly, it could provide useful support in the preclinical investigation and optimization of nanoscale systems for drug delivery to solid tumors.
我们最近构建了一种基于癌细胞、成纤维细胞和内皮细胞三重共培养的胰腺肿瘤多细胞球体模型，其特征是存在纤维连接蛋白，这是肿瘤细胞外基质的重要组成部分。通过将癌细胞与基质成分相结合，该模型在体外再现了实体瘤的三维（3D）结构。在这项研究中，我们使用这些异型球体作为工具来评估阿霉素（用作模型药物）通过整个肿瘤质量的渗透，无论是游离形式还是负载在聚合物纳米颗粒（NPs）中，并研究了共聚焦激光扫描显微镜（CLSM）和光片荧光显微镜（LSFM）获取的显微镜图像是否最适合提供有关此过程的可靠信息。结果清楚地表明，CLSM 不适合准确监测小分子（如阿霉素）的扩散。实际上，它只能扫描 100µm 深度的一层，由于荧光信号的逐渐丢失，无法获得更深层的信息。相反，LSFM 和多视图图像融合获得了异型多细胞肿瘤球体（MCTS）的完整 3D 断层扫描，这表明荧光分子能够到达直径达 1mm 的球体核心。然而，在球体中未检测到载有阿霉素的聚合物纳米颗粒，这突出了纳米医学通过生物屏障进行药物传递的挑战。总体而言，异型 MCTS 和 LSFM 的结合允许进行高度信息丰富的显微镜评估，并且是精确跟踪肿瘤内药物渗透的合适方法。因此，它可以在药物输送到实体瘤的纳米级系统的临床前研究和优化中提供有用的支持。
