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Cultivation and characterization of cancer stem cells in vitro
Accumulating reports indicate that cancer cells with stem cell-like characteristics (CSCs) exist in various types of cancer. It has been reported that these cells are responsible for malignant traits such as metastasis and chemoresistance.
Establishment of in vitro culture system of CSCs
In order to understand biological nature of CSCs, we started biological and biochemical characterization of CSCs. We isolated CSCs from human colon cancer specimens, and established conditions that allow stable in vitro propagation of colon CSCs in a spheroid form (Ohata et al., Cancer Res. 2012; Ishiguro et al., Cancer Sci. 2017, Review).
We found that inhibition of Rho kinase greatly facilitated the establishment of spheroids from primary colon cancer. Under such conditions, the spheroid cells expressed CSC markers, showed the ability to differentiate, and induced tumors in mice. The spheroids were composed of cells that express various levels of CD44, and CD44high cells exhibited characteristics similar to cancer stem cells. As expected from the predicted hierarchy, CD44high cells differentiated into CD44-/low cells. Unexpectedly, a fraction of CD44-/low cells generated CD44high cells, and we hypothesize the model in which the transition from CD44-/low to CD44high state may enhance tumorigenicity by maintaining a CD44high fraction in colon cancer (Ohata et al., Cancer Res. 2012).
In addition to colon cancer, we also found that inhibition of Rho kinase also promoted the establishment of spheroids from primary ovarian cancer. Biochemical and biological evaluation of the established spheroids revealed that reciprocal regulatory relationship between ALDH1A1 and SOX2 are involved in malignant progression of ovarian cancer as well as CSC proliferation (Ishiguro et al., Cancer Res. 2016).
These CSC spheroids prove to be instrumental for pre-clinical examination of anti-tumor compounds as well. The colon cancer spheroids and the tumors generated after transplantation of the spheroids in immuno-deficient mice were used for investigating efficacy of the newly-developed anti-cancer compounds that target the Wnt signaling (Masuda et al., Nature Comm. 2016).
The Cancer 3D Culture Workshop
Due to rapid progress of basic and translational research of cancer in recent years, a growing number of therapeutic agents, including small compounds, nucleic acid drugs, and antibody drugs, has been identified as potent candidates for next-generation anti-cancer medicines. Before they are used at bedsides, detailed pre-clinical examination of these medicines using cancer cell lines and/or animal models needs to be performed to predict their clinical efficacy.
However, current pre-clinical research tends to suffer from critical drawbacks; examination of therapeutic efficacy using cancer cell lines in many cases does not accurately predict their effectiveness at bedside, and testing in animal models tends to be financially demanding and to take a long time. In order to circumvent such problems, establishment of novel systems will be necessary for better assessment of anti-cancer agents.
Recently, novel three-dimensional (3D) cultivation techniques of cancer cells, such as spheroid culture or organoid culture, have been established. Because cancer cells can be cultivated without losing their original traits under such culture conditions, it is highly likely that they are suitable for the experimental platforms to evaluate therapeutic efficacy.
With this in mind, here we started The Cancer 3D Culture Workshop, in which basic and translational researcher can discuss various issues related to further development of the 3D cultures and their application for cancer research in future.