Genes & Tissue CulturePresentation

(Group 4)

By:Asiah Salleh

Chua Qin LingNg Chu Xin

Lim Su Shen

Development of cancer therapeutics is often carried out in 2D cultures prior to testing on animal model. In comparison to 2D cultures,

discuss the potential of using 3D in vitro models for drug efficiency testing

General Introduction of 3D & 2D culture

Video Introduction

Comparison between 2D and 3D culture

2D/3D Culture in drug efficacy testing ➔ Studies have found that cells cultured in 3D models are more resistant to

anticancer drug than in 2D cultures - geno- or phenotypical change induced by 3D spheroid formation - difference in gene expression - different cell stage

➔ 2D culture is non-predictive for in vivo test- Drug candidates fail in clinical trials due to adverse events, low efficacy etc.

➔ 3D culture emerged as a more reliable model to test for cancer cell viability in response to drug treatment- Photodynamic therapy

(Zang et. al 2012)

(Edmondson et. al 2014)

(Chen et. al 2015)

Differences in efficacy of PDT between 2D monolayer cell cultures and 3D spheres

● Perform Photodynamic Therapy (PDT) to compare conventional 2D culture and 3D culture

● LIVE/DEAD staining to check for cell viability

● After 10 minutes, about 50% of the cells in 2D culture are viable, but most of the cells are still viable in 3D spheroid

● After 1 hour, all cells in 2D culture were killed, but many cells in the sphere are still viable

(Chen etl al 2015)

Advantages of 3D cell culture systems for drug discovery.

★ Matrices contain ECM components lead to better cell-cell contact, communication and signalling pathway activation.

★ Restored cell functional and morphological differentiation.

★ Culture condition can be modified to include factors/proteins found in particular tumour microenvironment.

★ The gene and protein expression levels of cells as well as the cellular behaviours are similar to the in vivo levels.

★ Provide in vitro models for including different types of cells to build multicellular systems.

★ Bridges the gap between in vitro and in vivo drug screening,

(Edmondson et. al, 2014)

Figure : Advantages of microenvironment for drug discovery programmes. (Lovitt et. al, 2014)

Disadvantages of 3D cell culture systems for drug delivery❖ variability in biologically derived matrices leads to non-

reproducible experimental results

❖ high cost needed for large scale studies and high throughput assay

❖ High variability result ( difference ECM components in matrices)

❖ Low throughput in certain technique, due to the necessity of manual media changes (Katt et. al 2016)

(LabAutopedia, 2016)

Figure : Hanging Drop Plate Assembly.

Application of 3D cell culture · study basic mechanisms of organ development· to develop artificial organs for replacement or support of natural organs· to optimize bioproduction, or to obtain realistic pharmacotoxicological test systems.

Drug discovery:Undergo high throughput screening to generate hits.Useful in cancer drug research

organotypic tissue or slice cultures for drug testing:artificial liver (hepatocyte spheroids)hormone-producing tissuesbrain cell cultures heart cellsextracellular matrix and skin (by using fibroblasts and/or keratinocytes)

Artificial skin

Embryoid BodiesEmbryoid bodies are derived from embryonic stem cell linesretained capacity of lineage commitment (i.e., of generating cells of the hematopoietic,

endothelial, muscle, and neuronal lineages)

ReferencesChen, YC, Lou, X, Zhang, Z, Ingram, P & Yoon, E 2015, ‘High Throughput Cancer Cell Sphere Formation for Characterizing the Efficacy of Photodynamic Therapy in 3D cultures’, Scientific Reports, viewed 29 April 2016,<http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4495468/>

Edmondson, R, Broglie, JJ, Adcock, AF &Yang LJ 2014, ‘Three-Dimensional Cell Culture Systems and Their Applications in Drug Discovery and Cell-Based Biosensors’, Assay and Drug Development Technologies, vol 12, no. 4, pp. 207-218.

Katt, ME, Placone, AL, Wong AD, Xu, ZS, Searson, PC 2016, ' In Vitro Tumour Models: Advantages, Disadvantages, Variables and Selecting the Right Platform', Frontiers in Bioengineering and Biotechnology, vol. 4, no. 12.

Kunz-Schughart, L 2004, ‘The Use of 3-D Cultures for High-Throughput Screening: The Multicellular Spheroid Model’, Journal of Biomolecular Screening, vol 9, no.4, pp.273-285.

LabAutopedia 2016, Automated Cell Dispensing and Image-based Spheroid Formation Tracking, viewed 26 April 2016, <http://www.labautopedia.org/mw/Automated_Cell_Dispensing_and_Image-Based_Spheroid_Formation_Tracking>

Lovitt, CJ, Shelper, TB & Avery, VM 2014, ‘Advanced Cell Culture Techniques for Cancer Drug Discovery’, Discovery Biology, vol 3, no. 2, pp. 345-367.

Mueller-Klieser, W 1997, ‘Three-dimensional cell cultures: from molecular mechanisms to clinical applications’, American Journal of Physiology - Cell Physiology, vol 273, no.4, pp.C1109-C1123.

Zhang,R, Li, D, Tang, IC, Wang, J & Yang, ST 2012, ‘Cell-Based Assays in High-Throughput Screening for Drug Discovery’, International Journal of Biotechnology for Wellness Industries, vol. 1, pp. 31-51.

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