Cell Culture

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Figure 2: qPCR gene expression analysis. The quantitative gene expression results indicate NSC differentiation in 3D spheroid can express more neuronal marker gene (TUBB3) than NSCs maintained and differentiated in 2D adherent culture (n=10; *confidence interval of 2D and 3D, p=0.043<0.05). Source: Corning Life Sciences

complex mouse models that seldom reflect the pathophysiology of human tumour progression. In a climate where the clinical trial success rate for oncology drugs is only 3.4%12, cancer researchers are adopting 3D culture techniques to study human cancer biology and develop novel, effica- cious treatments for cancer patients. While cancer cell organoids are typically derived

from primary cells rather than HiPSCs, solid tumour models grown in 3D systems encourage cell-cell and cell-matrix interactions that closely imitate the natural environment. Compared to 2D cell culture systems, these models:

l Better mimic solid tumour gene and protein expression, metabolic activity, cell stress response, structure, signal transduction and cellular trans- port proteins13. l Exhibit patient variability and resistance to drug uptake and metabolism, and drug sensitivity14. l Reproduce most parameters of the tumour micro-environment, including oxygen and nutrient gradients, as well as the development of dormant tumour regions13.

Two clear examples of these advantages can be found in research related to breast cancer. Pickl et

Drug Discovery World Spring 2019

al15 demonstrated that breast tumours cultured in 3D express higher sensitivity to trastuzumab than those cultured in 2D, as the cells of the 3D cultures display increased activation and dependence on HER2 and HER3 signalling. Trastuzumab blocked HER2 and HER3 activation and proliferation of 3D cultures but not 2D-cultured cells. Wenzel et al16 used imaging technologies to dis-

tinguish the inner core of T47D breast cancer cells cultured in 3D from those in the outer core of the culture. Inner core cells had less access to oxygen and nutrients and showed reduced metabolic activ- ity compared with outer core cells. By screening small molecule libraries against these 3D cell cul- tures, these authors14 identified nine novel com- pounds that selectively killed the inner core cancer cells without affecting the more active proliferating outer core cancer cells. The field is now moving toward more predictive

3D cancer models that utilise patient-derived tumours, as well as high throughput drug screening using these 3D models. Numerous methods have been reported that consistently produce 3D tumour organoids and that are compatible with HTS automation instrumentation, increasing the potential for large-scale drug screening using patient-derived tumour models17.

11Takasato, Er et al. Kidney organoids from human iPS cells contain multiple lineages and model human nephrogenesis. Nature (2015); 526(7574):564-8. 12Wong CH, Siah, KW, Lo, AW. Estimation of clinical trial success rates and related parameters. Biostatistics. 2018;20(2):273-286. 13 Senkowski, W, Jarvius, M, Rubin, J et al. Large-scale gene expression profiling platform for identification of context- dependent drug responses in multicellular tumor spheroids. Cell Chem. Biol. 23(11), 1428-1438. 14 Stock, K, Estrada, MF, Vidic, S et al. Capturing tumor complexity in vitro: Comparative analysis of 2D and 3D tumor models for drug discovery. Sci Rep. 2016; 6: 28951. 15 Pickl, M and Ries, CH. Comparison of 3D and 2D tumor models reveals enhanced HER2 activation in 3D associated with an increased response to trastuzumab. Oncogene 28: 461-468, 2009. 16Wenzel, C et al. 3D high- content screening for the identification of compounds that target cells in dormant tumor spheroid regions. Experimental cell research 323, 131-143. 17 Madoux, F, Tanner, A, Vessels, M. A 1536-Well 3D Viability Assay to Assess the Cytotoxic Effect of Drugs on Spheroids. SLAS Discov. 2017, 22 (5), 516-524. 18 Jorfi, Mehdi et al. Three Dimensional Models of the Human Brain Development and Diseases. Advanced healthcare materials 7.1 (2018): 1700723. 19 Monzel, AS et al. Derivation of human midbrain-specific organoids from neuroepithelial stem cells. Stem Cell Rep. 8, 1144-1154 (2017).

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