SCT60103 Group 4 Assignment
- 1. THE POTENTIAL OF USING THREE- DIMENSIONAL (3D) CELL CULTURES FOR ANTICANCER DRUG SCREENING Group 4 Dave Aranka Thomas, Elaine Lee, Satish Paul, Sheryn Yeo, Teo Yi Heng
- 2. BASIC CONCEPTS & PRINCIPLES 2 What are 3D cell cultures? In vitro cultures where immortalized cell lines, stem cells, or explants are placed within hydrogel matrices Mimic in vivo cell environments Dimensionality of 3D cultures leads to varying cellular responses Spatial and physical aspects in 3D cultures affect signal transduction of cells, influencing gene expression and cellular behaviour Provide a more accurate prediction of drug toxicity and efficacy than traditional in vitro tumour cells cultured in monolayer (2D) (Antoni et al. 2015, Edmonsond 2014)
- 3. 3 Type of culture 2D 3D In vivo imitation Do not mimic the natural structure of the tissue or tumour mass In vivo tissues and organs are in 3D form Cells interactions Deprived of cell-cell and cell-extracellular environment interactions Proper cell-cell and cell-extracellular environment interactions Characteristics of cells Changed morphology, loss of diverse phenotype and polarity Preserved morphology, diverse phenotype and polarity Access to essential compounds Unlimited access to oxygen, nutrients, metabolites and signaling molecules (in contrast to in vivo) Variable access to oxygen, nutrients, metabolites and signaling molecules (same as in vivo) Cost of maintaining a culture Low cost, commercially available tests Expensive, time-consuming, fewer commercially available tests Table 1: Comparison between 2D and 3D cell cultures (Kapałczyńska et al. 2018)
- 4. DISCUSSION THE 3D MODEL: MULTICELLULAR TUMOUR SPHEROIDS (MCTS) 4 MCTS Most well-characterized organotypic model of cancerScaffold-free spherical self-assembled aggregates of cancer cells Display an intermediate complexity between 2D in vitro cell cultures and in vivo solid tumors Suitable 3D model for drug evaluation (Lazzari, Couvreur & Mura 2017, Nath & Devi 2016)
- 5. ● Spheroids in MCTS systems are constructed with different zones of cells a) proliferating cells on the outside b) quiescent viable cells in the middle c) necrotic cells at the inner core ● Creates a gradient of oxygen and nutrients from the outside of tumour spheroids to the core 5 Figure 2. The structure of MCTS with different regions of cells: necrotic zone (inner core), quiescent viable cell zone (middle), proliferating zone (outermost) Figure 3. Simulation of the basic structure of MCTS (Sant & Johnston 2017)
- 6. APPLICATION IN ANTICANCER DRUG SCREENING • Use 3D cell culture technology and patient-derived tumour cells • Yield better predictive value for preclinical screening • Reduce side effects of chemotherapy Preclinical anticancer drug screening • Target identification is the rate-limiting step in anticancer drug discovery • Develop novel mechanisms to accelerate target identification and validation Accelerate target identification • 3D tumour spheroids can design microenvironment similar to cancer cells in vivo • Able to stimulate proliferation, enhance cell motility, or induce cell dormancy • e.g. target dormant cancer cells to enhance cytostatic drug efficacy Promote desired cell behaviour 6 (Fang & Eglen 2017, Langhans 2018, Lv et al. 2017, Sant & Johnston 2017, Wenzel et al. 2014)
- 7. EVALUATION OF DRUG SENSITIVITY 7 MCTS enables study of the dynamic relationship between cancer cells and the microenvironment, including cell- cell and cell-matrix interactions Cancer-ECM interactions help to determine their therapeutic resistance which directly affects treatment efficacy Examples: Pancreatic tumour – enhanced expression of cell-adhesion molecules increases chemotherapeutic resistance A549 human lung cancer cells – MCTS establishes contact with their microenvironment, decreasing its sensitivity to chemotherapeutic drugs Increased adhesions may activate downstream signaling pathways leading to changes in gene expression, influencing sensitivity of cancer cells to drugs Many ECM-targeted chemotherapeutic drugs have been developed to inhibit specific integrin interactions, or to either synthesise or degrade the ECM. (Holle, Young & Spatz 2016, LaBarbera, Reid & Yoo 2012, Millard et al. 2017)
- 8. Monitor Changes in Viability & Apoptosis of Tumour Spheroids High-throughput screening (HTS) and Kinetic Apoptosis Assay To monitor change in viability of spheroids in real-time To study the effectiveness of drug on tumour spheroids To characterize cytotoxicity and apoptosis of cancer cells – chemotherapeutic drugs target extrinsic pathway of apoptosis to induce cell death 8 (Brunelle & Zhang 2010, Hesley 2016, Kessel et al. 2018)
- 9. CHALLENGES & CURRENT DEVELOPMENTS • Traditional methods are time- consuming and labour intensive • Cultures on non-adherent surfaces produce cells of varying size • Advanced methods need high level of expertise •Solution: Use of bioreactor yields uniform-sized cells and consistent life span of cell culture Forming and maintaining spheroids of uniform size • Interaction of multiple cell types with each other might exhibit malignancy •Solution: Formation of tumour histoid from stroma cells can be used to control malignancy Making tissue-like spheroids with multiple cell types 9 (Mehta et al. 2012)
- 10. CHALLENGES & CURRENT DEVELOPMENTS • Testing methods of drug efficacy in 2D cell cultures are incompatible for testing 3D cell cultures • Difficult to integrate previous research results to current 3D cell culture methods •Solution: Development of new specialized testing method for 3D cell culture (i.e. bioluminescence) Compatibility with current methods • MCTS model mimic only the avascular region of in vivo tumour tissues, leaving out the vasculature, immune system components and fluid dynamics Solution: Microfluidics incorporation Easy and controlled replenishing of cell cultures medium Impart mechanical stimulus Absence of Vasculature 10 (Lazzari, Couvreur & Mura 2017, Mehta et al. 2012)
- 11. CONCLUSION Anticancer drug screening is crucial in cancer research 3D cell culture systems serve as platforms for drug screening and used as reliable models for in vitro testing, given that MCTS have greater structure and cellular zone components similarity to in vivo tumours New culture methods are continuously developed, and older methods are constantly evolving to address the current challenges. The fluidity of this field is what allows ongoing advances to be made (Duval et al. 2017) 11
- 12. REFERENCES Antoni, D, Burckel, H, Josset, E, Noel, G 2015, ‘Three Dimensional Cell Culture: A Breakthrough in Vivo’, International Journal of Molecular Science, vol.16, no. 3, pp. 5517–5527. Brunelle, J, Zhang, B 2010, ‘Apoptosis Assays for Quantifying the Bioactivity of Anticancer Drug Products’, Drug Resistance Updates, vol. 13, no. 6, pp. 172-179. Duval, K, Grover, H, Han, LH, Mou, Y, Pegoraro, A, Fredberg, J & Chen, Z 2017, ‘Modeling Physiological Events in 2D vs. 3D Cell Culture’, Physiology (Bethesda), vol. 32, no. 4, pp. 266-277. Edmondson, R, Broglie, JJ, Adcock, AF & Yang L 2014, ‘Three-dimensional Cell Culture Systems and their Applications in Drug Discovery and Cell-based Biosensors’, Assay Drug Dev Technologies, vol. 12, no. 4, pp.207–218. Fang, Y & Eglen, RM 2017, ‘Three-Dimensional Cell Cultures in Drug Discovery and Development’, Slas Discovery, vol. 22, no. 5, pp. 456-472. Hesley, J 2016, ‘Screening for Cancer Therapeutics Using Spheroids’, Genetic Engineering and Biotechnology News, vol. 36, no. 11, p. 130. Holle, A, Young, J & Spatz, J 2016, ‘In Vitro Cancer Cell-ECM Interactions Inform In Vivo Cancer Treatment’, Advanced Drug Delivery Reviews, vol. 97, pp. 270-279. Kapałczyńska, M, Kolenda, T, Przybyła, W, Zajączkowska, M, Teresiak, A, Filas, V, Ibbs, M, Bliźniak, R, Łuczewski, Ł & Lamperska, K 2018, ‘2D and 3D Cell Cultures - A Comparison of Different Types of Cancer Cell Cultures’ Archives of Medical Science, vol. 14 no. 4, pp. 910-919. Kessel, S, Cribbes, S, Bonasu, S, Qiu, J & Chan, L 2018, ‘Real-Time Apoptosis and Viability High Throughput Screening of 3D Multicellular Tumor Spheroids Using the Celigo Image Cytometer’, SLAS Discovery, vol. 23, no. 2, pp. 202-210. 12
- 13. REFERENCES LaBarbera, D, Reid, B, Yoo, BH 2012, ‘The multicellular tumor spheroid model for high-throughput cancer drug discovery’, ResearchGate, vol. 7, no. 9, p. 819. Langhans, SA 2018, ‘Three-Dimensional in Vitro Cell Culture Models in Drug Discovery and Drug Repositioning’, Frontiers in Pharmocology, vol. 9, no. 6. Lazzari, G, Couvreur, P, Mura, S 2017, ‘Multicellular Tumour Spheroids: A Relevant 3D Model for the In Vitro Preclinical Investigation of Polymer Nanomedicines’, Polymer Chemistry, no. 34. Lv, D, Hu, Z, Lu, L, Lu, H, Xu, X 2017, ‘Three-dimensional cell culture: A powerful tool in tumor research and drug discovery’, Oncology Letters, vol. 14, no. 6, pp. 6999-7010. Mehta, G, Hsiao, A, Ingram, M, Luker, G, Takayama, S 2012, ‘Opportunities and challenges for use of tumor spheroids as models to test drug delivery and efficacy’, Journal of Controlled Release, vol. 164, no. 2, pp. 192-204. Millard, M, Yakavets, I, Zorin, V, Kulmukhamedova, A, Marchal, S, Bezdetnaya, L 2017, ‘Drug delivery to solid tumors: the predictive value of the multicellular tumor spheroid model for nanomedicine screening’, International journal of nanomedicine, vol. 12, pp. 7993-8007. Nath, S, Devi, G 2016, ‘Three-Dimensional Culture Systems in Cancer Research: Focus on Tumor Spheroid Model’, Pharmacology & therapeutics, vol. 163, pp. 94-108. Sant, S & Johnston, PA 2017, ‘ The Production of 3D Tumour Spheroids for Cancer Drug Discovery’ , Drug Discovery Today: Technologies, vol. 23, pp. 27-36. Wenzel, C, Riefke, B, Gründemann, S, Krebs, A, Christian, S, Prinz, F, Osterland, M, Golfier, S, Räse, S, Ansari, N, Esner, M, Bickle, M, Pampaloni, F, Mattheyer, C, Stelzer, E, Parcyzk, K, Prechtl, S, Steigemann, P 2014, ‘3D high-content screening for the identification’, Experimental Cell Research, vol. 323. 13