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Abstract
Introduction: The monolayer cell culture model is a popular model for screening anti-tumor activity of plant extracts. However, almost the extracts selected for screening in this model have failed in subsequent animal models. Therefore, there is only about 5 % of candidates from the original thousands of drugs that are screened which ultimately reach clinical trial. This study aimed to compare the differences in anti-tumor activity of 34 plant extracts against breast cancer cells in 2 models of monolayer cell culture (2D) and in three-dimensional (3D) cell culture.
 Methods: Four breast cancer cell lines (MCF-7, CD44+CD24- MCF-7, VN9, and CD44+CD24- VN9) were used to generate the 2D and 3D models (the 3D model was developed by culturing breast cancer cells in matrigel). The extracts were got from the plant extract library that prepared in the previous study. The anti-tumor activity was evaluated via half inhibitory concentrations( IC50 values).
 Results: Of the 34 extracts, E12, E7, E5 and E6 of them had an effect on MCF-7, CD44+CD24- MCF-7, VN9 and CD44+CD24- VN9 cells, respectively. The results indicated 10 potentially strong candidates for future drug development targeting hypoxic areas in breast cancer.
 Conclusion: The 3D culture model exhibited higher resistance to extracts than the 2D culture model. The CD44+CD24- cell population of both VN9 and MCF-7 cell lines showed higher drug resistance than the original cell lines (VN9 and MCF-7).
Highlights
The monolayer cell culture model is a popular model for screening anti-tumor activity of plant extracts
The results indicated 10 potentially strong candidates for future drug development targeting hypoxic areas in breast cancer
The half inhibitory concentration (IC50) results of doxorubicin and tirapazamine showed that both 2D and 3D models were successfully established for anti-tumor activity evaluation (Table 2)
Summary
The monolayer cell culture model is a popular model for screening anti-tumor activity of plant extracts. This study aimed to compare the differences in anti-tumor activity of 34 plant extracts against breast cancer cells in 2 models of monolayer cell culture (2D) and in three-dimensional (3D) cell culture. Screening done on a cancer model in 3dimensional (3D) culture may be better for studying drug effects since the 3D culture model is more similar to the in vivo animal models (and possibly clinical trials); the 3D model more closely reflects characteristics of in vivo tumors, such as differentiation, tumor microenvironment, and distribution of hypoxia in certain populations [5,6,7]. The advantage is that gravity is used to precipitate the cells together and thereby stimulate the cells to stick together into 3D spheres 10 This method has a disadvantage of using very gentle manipulations and is difficult to develop if screened at high throughput automation. Matrigel promotes the differentiation of different cell lines (e.g. prostate, salivary gland, mammary epithelium, pancreas, Schwann cells, intestinal cells, and bone cells), of primary cell lines (e.g. sertoli cells, blood cells, cartilage cells, epithelial cells, endometrial cells, and fallopian epithelial cells), and even tissue explants
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