One of the unsolved problems of cancer is metastatic dissemination, the spread of the cancer into distant organs. Shedding cells to the blood stream is a common phenomenon of most cancers and these malignant cells are released into circulation very early (circulating tumor cells or CTCs). Recent data indicate, however, that only those CTCs can initiate metastasis that show cancer stem cell features. Moreover, these cells are effective only if they form small clusters in circulation, called spheroids.
Spheroid or tumorsphere formation is a typical feature of cancer cells in vitro as well. We have been studying cancer spheroids in vitro and found that they form hypoxic centers that carry cancer stem cells. We also investigated tumorsphere formation in vivo in cancer patients, using flow cytometry. We detected and characterized circulating cancer spheroids in the blood of metastatic patients. Our preliminary data suggest that tumorspheres in blood can be a reliable indicator of metastatic predisposition and a specific marker of metastasis. The goal of this project was to work out the chemical/physical procedures to microfabricate controlled pore-size devices to capture circulating cancer spheroids.
We used parylene-based technology with micromachining methods, a process well established in the Micro/Nano Fabrication Facility at Tufts University. We applied a thin film of Parylene-C, a biocompatible polymer that is commonly used in medical devices and has become popular as structural material in the microfabrication field. Parylene-C was deposited from vapor phase using a coating process and it is pinhole free. The material has a high toughness and can be used as a substrate for electrical contacts as well. Pores can be dry etched through the Parylene membrane using oxygen plasmas. As the average size of cellular elements in blood is normally less than 15 um, while circulating spheroids were shown to be between 40-60 um, we selected a pore size at 20 um to capture spheroids specifically.
We used an in vitro tissue culture model and show that cancer cell spheroids can be isolated and characterized using the chemically microfabricated parylene filters. We also used control mesh filters and our preliminary results demonstrate that cancer spheroids can be isolated from patient blood as well. These devices represent the first steps to develop a new tool in the early diagnosis of metastatic cancer.
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