分離,培養,與腎臟循環腫瘤細胞特性之研究
楊明昕1、 高建璋1、陳進利1、吳勝堂1、查岱龍1
1三軍總醫院 泌尿外科
Identification, propagation and characterisation of circulating renal cancer stem cells
Ming-Hsin Yang1、Chien-Chang Kao1、Chin-Li Chen1、Sheng-Tang Wu1 、Tai-Lung Cha1
1Division of Urology, Department of Surgery, Tri-Service General Hospital, National Defense Medical Center, Taipei, Taiwan.
Purpose:
Renal cell carcinoma (RCC) is an aggressive tumor with early dissemination and still dismal prognosis. Most cases are inoperable, and biopsies to investigate RCC biology are rarely obtainable. Circulating tumour cells (CTCs) are cancer cells disseminated from the primary tumour or metastatic sites into the blood stream. CTCs could provide a less invasive technique to access tumour material (liquid biopsy) and would also be available at any time and for sequential sampling. On the other hand, cancer stem cells (CSCs) represent a functionally defined sub-population of cancer cells, for which our lab and others have provided conclusive evidence that they are essential for the metastatic behaviour of the tumours and, due to their inherent resistance to current therapies, represent an important source for disease relapse. As such, CTCs are likely to represent a heterogeneous population including a subset of cells, potentially the most aggressive, with self-renewal and high tumour initiating capabilities. i.e. circulating CSCs. However, isolating these cells from patients with RCC has been enormously challenging due to the sparsity of these cells.
Materials and Methods:
To isolate CTCs, we use Isoflux as our platform. Isoflux is used with our cocktail of three antibodies, EPCAM, CD147 and CA9, and thus retain any cell expressing at least one of these antigens including doublets or even larger aggregates of cells with at least one antigen+ cell. We use samples from patients presenting at the TSGH with confirmed diagnosis of RCC. These patients can be enrolled within 3 months. To culture the cells, we use our modified organoid culture medium.
Results:
We have now developed a new platform that allows for the isolation of abundant and viable CTC from patients with RCC. The retrieved CTC displayed functional CSC characteristics, which was sufficient for their subsequent expansion in vitro. The arising models were phenotypically distinct compared to conventional PDX models, highly aggressive with strong metastatic activity, and readily passaged.
Conclusions:
These data demonstrate that liquid biopsies now provide access to viable tissue from local and advanced RCC patients allowing for comprehensive and longitudinal phenotypic and molecular analysis via serial blood sampling. These unique models provide tractable systems for therapy testing, understanding drug resistance mechanisms and may facilitate delivery of personalized medicine for RCC.