Category: Assay Development and Screening
Evaluation of selected cancer drugs on tumor cell growth in 2D monolayer versus 3D tumor spheroids of lung and breast cancer cell lines
T. West, J. Huang, J. Chan, N. Pagratis, and Y. Li
High Throughput Biology & Screening, Gilead Sciences, Foster City, CA 94404
Cancer drug candidates identified from 2D monolayer cultures are often not effective on hypoxic and slow growing tumors in animal models or clinical trials. The differences between 2D culture and in vivo conditions could be one of the reasons for high attrition rate in cancer drug discovery. 3D tumor spheroid culture has shown increasing value in cancer research because it approximates conditions found in human native tumors by allowing cell-cell and cell-ECM interactions, as well as chemical gradients and biological zones in the tumor microenvironment. Fayad et al., (2011) demonstrated that cancer drug candidates screened out from 2D and 3D cultures are different. In addition, recent reports showed that drugs against molecules in KRAS pathways are more efficacious growth inhibitors in 3D tumor spheroids than in 2D monolayer cultures (Wellspring AACR, 2017; Nichols et al. 2017). In this study, we selected a panel of cancer drugs including several chemotherapy agents, inhibitors of RAS pathway MEK1/2 (Trametinib and Selumetinib) and BRAF (Vemuratenib), Akt/mTOR (Temsirolimus and Sirolimus), EGFR (Erlotinib, Afatinib, and Lapatinib), BET (GILD-1), TBK1 (GILD-2), and PI3Kbeta (GILD-3). We evaluated the effect of these inhibitors at various doses (14.3 uM to 2.0 nM in 1:3 dilutions) on tumor growth in 2D versus 3D cultures of SKBR-3 and MDA-MB-231 breast, and A549 lung cancer cell lines, in 384-well format. We demonstrated that in SKBR-3 cell line, Lapatinib, Afatinib, and a PI3Kbeta inhibitor (GILD-3), inhibited 3D tumor spheroid growth more potently than 2D. In MDA-MB-231 cell line, Doxorubicin, Selumetinib, and Trametinib preferentially inhibited 3D spheroid growth over 2D. Interestingly, the BET inhibitor (GILD-1) not only showed more potent IC50 in 3D culture, but also inhibited tumor growth to ~60% at 530 nM, whereas in 2D culture, the same concentration achieved only ~20% tumor growth inhibition. This compound would have not been selected for follow-up if 2D culture was applied for screening. In a NSCLC A549 cell line, we found that Trametinib, Selumetinib, Erlotinib, Afatinib, and BET inhibitor (GILD-1) showed preferential inhibition of 3D spheroid growth over 2D. Cell growth was measured by Cell Titer-Glo for both 2D and 3D. Additionally, growth inhibition for 3D spheroids of MDA-MB-231-GFP and A549-NucLight Red cell lines was measured by Cellomics imaging. Our results indicate that in cancer drug discovery and screening, 3D culture can identify compounds that otherwise would be missed in 2D culture, thus expanding the relevant drug candidates for follow up.
Yvonne Li– Senior Scientist, Gilead Sciences, San Mateo, CA