Abstract
Recent progress in three-dimensional (3D) cell culture systems has attracted much attention in the fields of basic life science and drug development. Newly established methods include 3D co-culture, spheroid culture, and organoid culture; these methods enable more human tissue-like culture and have largely replaced traditional two-dimensional (2D) monolayer culture. By combining 3D culture methods with high-content imaging analysis, it is possible to obtain diverse and convincing data even during initial screening (which requires rapid and easy operating procedures). Until recently, 3D culture methods were considered expensive, time-consuming, complex, and unstable. However, by exploiting the self-assembling nature of cells and adding several technical improvements, we have developed several phenotypic screenings aimed at discovering anticancer compounds.
This is a preview of subscription content, access via your institution
Access options
Subscribe to this journal
Receive 12 print issues and online access
$259.00 per year
only $21.58 per issue
Buy this article
- Purchase on SpringerLink
- Instant access to full article PDF
Prices may be subject to local taxes which are calculated during checkout
Similar content being viewed by others
References
Swinney DC, Anthony J. How were new medicines discovered? Nat Rev Drug Discov. 2011;10:507–19.
Hapke RY, Haake SM. Hypoxia-induced epithelial to mesenchymal transition in cancer. Cancer Lett. 2020;487:10–20.
Nakano I. Stem cell signature in glioblastoma: therapeutic development for a moving target. J Neurosurg. 2015;122:324–30.
Shimokawa M, Ohta Y, Nishikori S, Matano M, Takano A, Fujii M, et al. Visualization and targeting of LGR5+ human colon cancer stem cells. Nature. 2017;545:187–204.
Phi LTH, Sari IN, Yang YG, Lee SH, Jun N, Kim KS, et al. Cancer stem cells (CSCs) in drug resistance and their therapeutic implications in cancer treatment. Stem Cells Int. 2018;28:5416923.
Choi SH, Kim YH, Hebisch M, Sliwinski C, Lee S, D’Avanzo C, et al. A three-dimensional human neural cell culture model of Alzheimer’s disease. Nature. 2014;515:274–8.
Kim YH, Choi SH, D’Avanzo C, Hebisch M, Sliwinski C, Bylykbashi E, et al. A 3D human neural cell culture system for modeling Alzheimer’s disease. Nat Protoc. 2015;10:985–1006.
Kwak SS, Washicosky KJ, Brand E, von Maydell D, Aronson J, Kim S, et al. Amyloid-β42/40 ratio drives tau pathology in 3D human neural cell culture models of Alzheimer’s disease. Nat Commun. 2020;11:1377.
Miller DJ. Sydney Ringer; physiological saline, calcium and the contraction of the heart. J Physiol. 2004;555:585–87.
Gähwiler BH. Nerve cells in culture: the extraordinary discovery by Ross Granville Harrison. Brain Res Bull. 1999;50:343–4.
Sanford KK, Earle W, Likely GD. The growth in vitro of single isolated tissue cells. J Natl Cancer Inst. 1948;9:229–46.
Scherer WF, Syverton JT, Gey GO. Studies on the propagation in vitro of poliomyelitis viruses. IV. Viral multiplication in a stable strain of human malignant epithelial cells (strain HeLa) derived from an epidermoid carcinoma of the cervix. J Exp Med. 1953;97:695–710.
Dvorak HF. Tumors: wounds that do not heal. Similarities between tumor stroma generation and wound healing. N Engl J Med. 1986;315:1650–9.
Tuxhorn JA, Ayala GE, Smith MJ, Smith VC, Dang TD, Rowley DR. Reactive stroma in human prostate cancer: induction of myofibroblast phenotype and extracellular matrix remodeling. Clin Cancer Res. 2002;8:2912–23.
Kawada M, Yoshimoto Y, Minamiguchi K, Kumagai H, Someno T, Masuda T, et al. A microplate assay for selective measurement of growth of epithelial tumor cells in direct coculture with stromal cells. Anticancer Res. 2004;24:1561–8.
Kawada M, Atsumi S, Wada SI, Sakamoto S. Novel approaches for identification of anti-tumor drugs and new bioactive compounds. J Antibiot. 2018;71:39–44.
Simian M, Bissell M. Organoids: a historical perspective of thinking in three dimensions. J Cell Biol. 2017;216:31–40.
Fatehullah A, Tan SH, Barker N. Organoids as an in vitro model of human development and disease. Nat Cell Biol. 2016;18:246–54.
Sato T, Vries RG, Snippert HJ, Wetering M, Barker N, Stange DE. Single Lgr5 stem cells build crypt–villus structures in vitro without a mesenchymal niche. Nature. 2009;459:262–66.
Chen X, Wei B, Han X, Zheng Z, Huang J, Liu J, et al. LGR5 is required for the maintenance of spheroid-derived colon cancer stem cells. Int J Mol Med. 2014;3:35–42. 4
Hirsch D, Barker N, McNeil N, Hu Y, Camps J, McKinnon K, et al. LGR5 positivity defines stem-like cells in colorectal cancer. Carcinogenesis. 2014;35:849–58.
Uchida H, Yamazaki K, Fukuma M, Yamada T, Hayashida T, Hasegawa H, et al. Overexpression of leucine-rich repeat-containing G protein-coupled receptor 5 in colorectal cancer. Cancer Sci. 2010;101:1731–37.
Leung C, Tan SH, Barker N. Recent advances in Lgr5+ stem cell research. Trends Cell Biol. 2018;28:380–91.
Morgan RG, Mortensson E, Williams AC. Targeting LGR5 in colorectal cancer: therapeutic gold or too plastic? Br J Cancer. 2018;118:1410–18.
Sato T, Stange DE, Ferrante M, Vries RGJ, van Es JH, van den Brink S, et al. Long-term expansion of epithelial organoids from human colon, adenoma, adenocarcinoma, and Barrett’s epithelium. Gastroenterol 2011;141:1762–72.
Mahe MM, Aihara E, Schumacher MA, Zavros Y, Montrose MH, Helmrath MA. et al. Establishment of gastrointestinal epithelial organoids. Curr Protoc Mouse Biol. 2013;3:217–40.
Sato T, Clevers H. Growing self-organizing mini-guts from a single intestinal stem cell: mechanism and applications. Science. 2013;340:1190–94.
Öhlund D, Handly‑Santana A, Biffi G, Elyada E, Almeida AS, Ponz‑Sarvise M, et al. Distinct populations of inflammatory fibroblasts and myofibroblasts in pancreatic cancer. J Exp Med. 2017;214:57996.
Calon A, Lonardo E, Berenguer-Llergo A, Espinet E, Hernando-Momblona X, Iglesias M, et al. Stromal gene expression defines poor-prognosis subtypes in colorectal cancer. Nat Genet. 2015;47:320–32.
Costanza B, Umelo IA, Bellier J, Castronovo V, Turtoi A. Stromal modulators of TGF-β in cancer. J Clin Med 2017;6:7.
Provenzano PP, Cuevas C, Chang AE, Goel VK, Von Hoff DD, Hingorani SR. Enzymatic targeting of the stroma ablates physical barriers to treatment of pancreatic ductal adenocarcinoma. Cancer Cell. 2012;21:418–29.
Sahai E, Astsaturov I, Cukierman E, DeNardo DG, Egeblad M, Evans RM, et al. A framework for advancing our understanding of cancer-associated fibroblasts. Nat Rev Cancer. 2020;20:174–86.
Yoshida GJ. Regulation of heterogeneous cancer-associated fibroblasts: the molecular pathology of activated signaling pathways. J Exp Clin Cancer Res. 2020;39:112.
Froeling FEM, Feig C, Chelala C, Dobson R, Mein CE, Tuveson DA, et al. Retinoic acid-induced pancreatic stellate cell quiescence reduces paracrine Wnt-β-catenin signaling to slow tumor progression. Gastroenterology. 2011;141:1486–97.
Åkerfelt M, Bayramoglu N, Robinson S, Toriseva M, Schukov HP, Härmä V, et al. Automated tracking of tumor-stroma morphology in microtissues identifies functional targets within the tumor microenvironment for therapeutic intervention. Oncotarget. 2015;6:30035–56.
Seino T, Kawasaki S, Shimokawa M, Tamagawa H, Toshimitsu K, Fujii M, et al. Human pancreatic tumor organoids reveal loss of stem cell niche factor dependence during disease progression. Cell Stem Cell. 2018;22:45467.
Bulin AL, Broekgaarden M, Hasan T. Comprehensive high-throughput image analysis for therapeutic efficacy of architecturally complex heterotypic organoids. Sci Rep. 2017;7:16645.
Hou S, Tiriac H, Sridharan BP, Scampavia L, Madoux F, Seldin J, et al. Advanced development of primary pancreatic organoid tumor models for high-throughput phenotypic drug screening. SLAS Discov. 2018;23:574–84.
Tsai S, McOlash L, Palen K, Johnson B, Duris C, Yang Q, et al. Development of primary human pancreatic cancer organoids, matched stromal and immune cells and 3D tumor microenvironment models. BMC Cancer. 2018;18:335.
Baker LA, Tiriac H, Clevers H, Tuveson DA. Modeling pancreatic cancer with organoids. Trends Cancer. 2016;2:176–90.
Broekgaardena M, Anbila S, Bulina AL, Obaida G, Maia Z, Bagloa Y, et al. Modulation of redox metabolism negates cancer-associated fibroblasts-induced treatment resistance in a heterotypic 3D culture platform of pancreatic cancer. Biomaterials 2019;222:119421.
Driehuis E, Hoeck A, Moore K, Kolders S, Francies HE, Gulersonmez MC, et al. Pancreatic cancer organoids recapitulate disease and allow personalized drug screening. PNAS 2019;116:26580–90.
Davis ME. Glioblastoma: overview of disease and treatment. Clin J Oncol Nurs. 2016;20:S2–8.
Singh SK, Clarke ID, Terasaki M, Bonn VE, Hawkins C, Squire J, et al. Identification of a cancer stem cell in human brain tumors. Cancer Res. 2003;15:5821–8.
Yanae M, Tsubaki M, Satou T, Itoh T, Imano M, Yamazoe Y, et al. Statin-induced apoptosis via the suppression of ERK1/2 and Akt activation by inhibition of the geranylgeranyl-pyrophosphate biosynthesis in glioblastoma. J Exp Clin Cancer Res 2011;30:74–81.
Afshordel S, Kern B, Clasohm J, König H, Priester M, Weissenberger J, et al. Lovastatin and perillyl alcohol inhibit glioma cell invasion, migration,and proliferation – Impact of Ras-/Rho-prenylationlovastatin. Pharm Res. 2015;91:69–77.
Xiao A, Brenneman B, Floyd D, Comeau L, Spurio K, Olmez I, et al. Statins affect human glioblastoma and other cancers through TGF-β inhibition. Oncotarget 2019;10:1716–28.
Graaf MR, Beiderbeck AB, Egberts ACG, Richel DJ, Guchelaar HJ. The risk of cancer in users of statins. J Clin Oncol. 2004;22:2388–94.
Gaist D, Andersen L, Hallas J, Sørensen HT, Schrøder HD, Friis S. Use of statins and risk of glioma: a nationwide case–control study in Denmark. Br J Cancer. 2013;108:715–20.
Gaist D, Hallas J, Friis S, Hansen S, Sørensen HT. Statin use and survival following glioblastoma multiforme. Cancer Epidemiol. 2014;38:722–27.
Behnan J, Finocchiaro G, Hanna G. The landscape of the mesenchymal signature in brain tumours. Brain. 2019;142:847–66.
Fedele M, Cerchia L, Pegoraro S, Sgarra R, Manfioletti G. Proneural-mesenchymal transition: phenotypic plasticity to pcquire pultitherapy pesistance in glioblastoma. Int J Mol Sci. 2019;20:2746–59.
Nakano I. Proneural–mesenchymal transformation of glioma stem cells: do therapies cause evolution of target in glioblastoma? Future Oncol. 2014;10:1527–30.
Guardia GDA, Correa BR, Araujo PR, Qiao M, Burns S, Penalva LOF, et al. Proneural and mesenchymal glioma stem cells display major differences in splicing and lncRNA profiles. NPJ Genom Med. 2020;5:2.
Pavlyukov MS, Yu H, Bastola S, Minata M, Shender VO, Lee Y, et al. Apoptotic cell-derived extracellular vesicles promote malignancy of glioblastoma via intercellular transfer of splicing factors. Cancer Cell. 2018;34:119–35.
Bastola S, Pavlyukov MS, Yamashita D, Ghosh S, Cho H, Kagaya N, et al. Glioma-initiating cells at tumor edge gain signals from tumor core cells to promote their malignancy. Nat Commun. 2020;11:4660.
Gupta PB, Onder TT, Jiang G, Tao K, Kuperwasser C, Weinberg RA, et al. Identification of selective inhibitors of cancer stem cells. Cell. 2009;138:645–59.
Fujii M, Shimokawa M, Date S, Takano A, Matano M, Nanki K, et al. A colorectal tumor organoid library demonstrates progressive loss of niche factor requirements during tumorigenesis. Cell Stem Cell. 2016;18:827–38.
Wetering M, Francies HE, Francis JM, Bounova G, Iorio F, Pronk A, et al. Prospective derivation of a living organoid biobank of colorectal cancer patients. Cell. 2015;161:933–45.
Nanki K, Toshimitsu K, Takano A, Fujii M, Shimokawa M, Ohta Y, et al. Divergent routes toward Wnt and R-spondin niche independency during human gastric carcinogenesis. Cell. 2018;174:856–69.
Broutier L, Mastrogiovanni G, Verstegen MMA, Francies HE, Gavarró LM, Bradshaw CR, et al. Human primary liver cancer-derived organoid cultures for disease modelling and drug screening. Nat Med. 2017;23:1424–35.
Acknowledgements
This work was supported by Technology Research Association for Next Generation Natural Products Chemistry.
Author information
Authors and Affiliations
Contributions
KS designed and managed the study; HS prepared screening samples; NK constructed assay systems and practiced screening; MK and DT established fluorescent labeled Panc-1 cells; TS established CSC cells; HS, NK, and KS wrote the manuscript.
Corresponding author
Ethics declarations
Conflict of interest
The authors declare no competing interests.
Additional information
Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Rights and permissions
About this article
Cite this article
Suenaga, H., Kagaya, N., Kawada, M. et al. Phenotypic screening system using three-dimensional (3D) culture models for natural product screening. J Antibiot 74, 660–666 (2021). https://doi.org/10.1038/s41429-021-00457-8
Received:
Revised:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1038/s41429-021-00457-8
This article is cited by
-
Revolutionizing the female reproductive system research using microfluidic chip platform
Journal of Nanobiotechnology (2023)