Assessment of DNA ‘fingerprinting’ as a method for validating the identity of cancer cell lines maintained in long-term culture

AbstractWhile chemical and genetic viability screens in cancer cell lines have identified many promising cancer vulnerabilities, simple univariate readouts of cell proliferation fail to capture the complex cellular responses to perturbations. Complementarily, gene expression profiling offers an information-rich measure of cell state that can provide a more detailed account of cellular responses to perturbations. Relatively little is known, however, about the relationship between transcriptional responses to per-turbations and the long-term cell viability effects of those perturbations. To address this question, we integrated thousands of post-perturbational transcriptional profiles from the Connectivity Map with large-scale screens of cancer cell lines’ viability response to genetic and chemical perturbations. This analysis revealed a generalized transcriptional signature associated with reduced viability across perturbations, which was consistent across post-perturbation time-points, perturbation types, and viability datasets. At a more granular level, we lay out the landscape of treatment-specific expression-viability relationships across a broad panel of drugs and genetic reagents, and we demonstrate that these post-perturbational expression signatures can be used to infer long-term viability. Together, these results help unmask the transcriptional changes that are associated with perturbation-induced viability loss in cancer cell lines.

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Dual Targeting of FGFR3 and ERBB3 Enhances The Efficacy of FGFR Inhibitors in FGFR3 Fusion-Driven Bladder Cancer

10.21203/rs.3.rs-1026540/v1 ◽

2021 ◽

Author(s):

Andrew J Weickhardt ◽

David K Lau ◽

Margeaux Hodgson-Garms ◽

Austen Lavis ◽

Laura J Jenkins ◽

...

Keyword(s):

Bladder Cancer ◽

Cell Lines ◽

Cancer Cell ◽

Combination Treatment ◽

Acquired Resistance ◽

Cancer Cell Lines ◽

Bladder Cancer Cell ◽

Long Term Treatment ◽

Inhibitor Treatment

Abstract Background Mutations and fusions in Fibroblast Growth Factor Receptor 3 (FGFR3) occur in 10-20% of metastatic urothelial carcinomas and confers sensitivity to FGFR inhibitors. However, responses to these agents are often short-lived due to the development of acquired resistance. The objective of this study was to identify mechanisms of resistance to FGFR inhibitors in two previously uncharacterised bladder cancer cell lines harbouring FGFR3 fusions and assess rational combination therapies to enhance sensitivity to these agents. Methods Acquired resistance to FGFR inhibitors was generated in two FGFR3 fusion harbouring cell lines, SW780 (FGFR3-BAIAP2L1 fusion) and RT4 (FGFR3-TACC3 fusion), by long-term exposure to the FGFR inhibitor BGJ398. Changes in levels of receptor tyrosine kinases were assessed by phospho-RTK arrays and immunoblotting. Changes in cell viability and proliferation were assessed by the Cell-Titre Glo assay and by propidium iodide staining and FACS analysis. Results Long term treatment of FGFR3-fusion harbouring SW780 and RT4 bladder cancer cell lines with the BGJ398 resulted in the establishment of resistant clones. These clones were cross-resistant to the clinically approved FGFR inhibitor erdafitinib and the covalently binding irreversible FGFR inhibitor TAS-120, but remained sensitive to the MEK inhibitor trametinib, indicating resistance is mediated by alternate activation of MAPK signalling. The FGFR inhibitor-resistant SW780 and RT4 lines displayed increased expression of pERBB3, and strikingly, combination treatment with an FGFR inhibitor and the ATP-competitive pan-ERBB inhibitor AZD8931 overcame this resistance. Notably, rapid induction of pERBB3 and reactivation of pERK also occurred in parental FGFR3 fusion-driven lines within 24 hours of FGFR inhibitor treatment, and combination treatment with an FGFR inhibitor and AZD8931 delayed the reactivation of pERBB3 and pERK and synergistically inhibited cell proliferation. Conclusions We demonstrate that increased expression of pERBB3 is a key mechanism of adaptive resistance to FGFR inhibitors in FGFR3-fusion driven bladder cancers, and that this also occurs rapidly following FGFR inhibitor treatment. Our findings demonstrate that resistance can be overcome by combination treatment with a pan-ERBB inhibitor and suggest that upfront combination treatment with FGFR and pan-ERBB inhibitors warrants further investigation for FGFR3-fusion harbouring bladder cancers.

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Resistance development after long-term sorafenib exposure in hepatocellular cancer cell lines and risk of rebound growth and epithelial to mesenchymal transition.

Journal of Clinical Oncology ◽

10.1200/jco.2012.30.4_suppl.216 ◽

2012 ◽

Vol 30 (4_suppl) ◽

pp. 216-216 ◽

Cited By ~ 22

Author(s):

Chris Verslype ◽

Hannah van Malenstein ◽

Jeroen Dekervel ◽

Petra Windmolders ◽

Louis Libbrecht ◽

...

Keyword(s):

Gene Expression ◽

Side Effects ◽

Cell Lines ◽

Cancer Cell ◽

Hepg2 Cells ◽

Epithelial To Mesenchymal Transition ◽

Cancer Cell Lines ◽

Mesenchymal Transition ◽

Resistant Cells

216 Background: Sorafenib, a multi tyrosine kinase inhibitor, is the first line treatment in patients with advanced hepatocellular carcinoma (HCC). It leads to a survival benefit but treatment with sorafenib is hampered by two phenomena: patients can develop important side-effects and eventually all patients show progression. We aimed to determine the effects of long-term exposure to sorafenib and its withdrawal in vitro. Methods: We developed sorafenib resistant liver cancer cell lines (HepG2, WRL-68 and Huh-7) by slowly increasing sorafenib concentrations. XTT- and BrdU-assay were used to study the effect of sorafenib withdrawal on proliferation and metabolism. Morphological changes were examined with immuocytochemistry, gene expression changes with RT-PCR and the invasive potential with matrigel invasion chambers. Microarray was performed on resistant HepG2 cells. Results: HepG2 cells (6µM), WRL-68 cells (6µM) and Huh-7 cells (5µM) became resistant to sorafenib. Resistance was confirmed with a shift of the IC50 to ±16µM and ongoing phosphorylation of ERK during sorafenib exposure. All three resistant cell types showed significant increased proliferation and metabolic activity after withdrawal of sorafenib. The HepG2 resistant cells have undergone an epithelial to mesenchymal transition (EMT) with loss of E-cadherin and high expression of vimentin. The cells that displayed EMT became spindle shaped and were highly invasive. Furthermore, gene expression profiling confirmed EMT changes in a large set of EMT-related genes. Considering drug metabolism, the HepG2 sorafenib resistant cells showed a downregulation of UDP glucuronosyltransferases (UGT) and cytochromes P450, such as CYP3A4. There was a strong downregulation of different ABC-transporters, although breast cancer resistance protein (ABCG2) was upregulated. Conclusions: Long-term treatment with sorafenib can lead to the development of resistant cells, even with an aggressive phenotype because the cells can undergo EMT. Furthermore we demonstrated that abrogation of treatment leads to rebound growth, suggesting the importance of aggressive management of side-effects.

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