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3D-cultivation of NSCLC cell lines induce gene expression alterations of key cancer-associated pathways and mimic in-vivo conditions.

ABSTRACT: This work evaluated gene expression differences between a hanging-drop 3D NSCLC model and 2D cell cultures and their in-vivo relevance by comparison to patient-derived data from The Cancer Genome Atlas. Gene expression of 2D and 3D cultures for Colo699 and A549 were assessed using Affymetrix HuGene 1.0 ST gene chips. Biostatistical analyses tested for reproducibility, comparability and significant differences in gene expression profiles between cell lines, experiments and culture methods. The analyses revealed a high interassay correlation within specific culture systems proving a high validity. 979 genes were altered in A549 and 1106 in Colo699 cells due to 3D cultivation. The overlap of changed genes between the cell lines was small (149), but the involved pathways in the reactome and GO- analyses showed a high overlap with DNA methylation, cell cycle, SIRT1, PKN1 pathway, DNA repair and oxidative stress as well known cancer-associated representatives. Additional specific GSEA-analyses revealed changes in immunologic and endothelial cell proliferation pathways, whereas hypoxic, EMT and angiogenic pathways were downregulated. Gene enrichment analyses showed 3D-induced gene up-regulations in the cell lines 38 to be represented in in-vivo samples of NSCLC patients using data of The Cancer Genome Atlas. Thus, our 3D NSCLC model might provide a tool for early drug development and investigation of microenvironment-associated mechanisms. However, this work also highlights the need for further individualization and model adaption to address remaining challenges.

SUBMITTER: Gamerith G

PROVIDER: S-EPMC5762538 | biostudies-literature | 2017 Dec

REPOSITORIES: biostudies-literature

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3D-cultivation of NSCLC cell lines induce gene expression alterations of key cancer-associated pathways and mimic <i>in-vivo</i> conditions.

Gamerith Gabriele G Rainer Johannes J Huber Julia M JM Hackl Hubert H Trajanoski Zlatko Z Koeck Stefan S Lorenz Edith E Kern Johann J Kofler Reinhard R Kelm Jens M JM Zwierzina Heinz H Amann Arno A

Oncotarget 20171106 68

This work evaluated gene expression differences between a hanging-drop 3D NSCLC model and 2D cell cultures and their <i>in-vivo</i> relevance by comparison to patient-derived data from The Cancer Genome Atlas. Gene expression of 2D and 3D cultures for Colo699 and A549 were assessed using Affymetrix HuGene 1.0 ST gene chips. Biostatistical analyses tested for reproducibility, comparability and significant differences in gene expression profiles between cell lines, experiments and culture methods. ...[more]

PMID: 29348853

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Project description:Background: The main focus of the work was the evaluation of gene expression differences between our established NSCLC 3D cell culture model and the 2D cell culture in regard to the use of our model for drug screening applications. Methods: The non-small cell lung cancer (NSCLC) cell lines Colo699 and A549 were cultivated as monolayer (2D) on cell culture plates for five days or as microtissues (3D) in a hanging-drop system for five and ten days, respectively. Cells and microtissues were harvested and Affymetrix chip analyses were performed with the prior isolated RNA. This was repeated in three independent experiments. Subsequent biostatistical data analyses tested for reproducibility, comparability and significant differences in gene expression profiles between cell lines, experiments and culture methods. Results: The analyses revealed a high interassay correlation within the distinct culture systems, thus proving a high validity of our data. The comparison of 3D versus 2D cell cultures revealed significant differences in RNA expression (979 genes for A549; 1106 genes for Colo699), but the overlap of changes in RNA profiles between the cell lines at the individual gene level was small (149 genes), potentially reflecting overall heterogeneity and their origin, i.e. primary vs pleural effusion. Nevertheless, these RNA expression changes affected most relevant cancer-associated pathways as DNA methylation, cell cycle, rRNA expression and meiosis pathways. Furthermore, the expression differences between 2D and 3D were more evident after longer cultivation time, which supports the hypothesis of cultivation related mechanisms and the usage of long-time cultivation systems. Conclusion: In summary, our data support the need of innovative 3D drug testing systems to close the gap between in-vitro drug screening and in-vivo data. Thus, our 3D NSCLC model might provide a model to address the challenge of microenviroment associated resistance mechanisms, as well as cell-cell interaction related effects.

Dataset Information

3D-cultivation of NSCLC cell lines induce gene expression alterations of key cancer-associated pathways and mimic in-vivo conditions.

Publications

3D-cultivation of NSCLC cell lines induce gene expression alterations of key cancer-associated pathways and mimic <i>in-vivo</i> conditions.

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