Genome-wide epigenetic and mRNA-expression profiling followed by CRISPR/Cas9-mediated gene-disruptions corroborate the MIR141/MIR200C-ZEB1/ZEB2-FGFR1 axis in acquired EMT-associated EGFR TKI-resistance in NSCLC cells

Johan Vad-Nielsen; Nicklas Heine Staunstrup; Magnus Lindkvist Kjeldsen; Nina Dybdal; Guillaume Flandin; Claudia De Stradis; Tina Fuglsang Daugaard; Trine Vilsbøll-Larsen; Christoffer Trier Maansson; Thomas Koed Doktor; Boe Sandahl Sorensen; Anders Lade Nielsen

doi:10.21037/tlcr-22-507

Genome-wide epigenetic and mRNA-expression profiling followed by CRISPR/Cas9-mediated gene-disruptions corroborate the MIR141/MIR200C-ZEB1/ZEB2-FGFR1 axis in acquired EMT-associated EGFR TKI-resistance in NSCLC cells

Johan Vad-Nielsen, Nicklas Heine Staunstrup, Magnus Lindkvist Kjeldsen, Nina Dybdal, Guillaume Flandin, Claudia De Stradis, Tina Fuglsang Daugaard, Trine Vilsbøll-Larsen, Christoffer Trier Maansson, Thomas Koed Doktor, Boe Sandahl Sorensen, Anders Lade Nielsen^*

^*Kontaktforfatter

Publikation: Bidrag til tidsskrift › Tidsskriftartikel › Forskning › peer review

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Abstract

Background: Epithelial-mesenchymal-transition (EMT) is an epigenetic-based mechanism contributing to the acquired treatment resistance against receptor tyrosine kinase inhibitors (TKIs) in non-small cell lung cancer (NSCLC) cells harboring epidermal growth factor receptor (EGFR)-mutations. Delineating the exact epigenetic and gene-expression alterations in EMT-associated EGFR TKI-resistance (EMT-E-TKI-R) is vital for improved diagnosis and treatment of NSCLC patients. Methods: We characterized genome-wide changes in mRNA-expression, DNA-methylation and the histone-modification H3K36me3 in EGFR-mutated NSCLC HCC827 cells in result of acquired EMT-E-TKI-R. CRISPR/Cas9 was used to functional examine key findings from the omics analyses. Results: Acquired EMT-E-TKI-R was analyzed with three omics approaches. RNA-sequencing identified 2,233 and 1,972 up- and down-regulated genes, respectively, and among these were established EMT-markers. DNA-methylation EPIC array analyses identified 14,163 and 7,999 hyper- and hypo-methylated, respectively, differential methylated positions of which several were present in EMT-markers. Finally, H3K36me3 chromatin immunoprecipitation (ChIP)-sequencing detected 2,873 and 3,836 genes with enrichment and depletion, respectively, and among these were established EMT-markers. Correlation analyses showed that EMT-E-TKI-R mRNA-expression changes correlated better with H3K36me3 changes than with DNA-methylation changes. Moreover, the omics data supported the involvement of the MIR141/MIR200C-ZEB1/ZEB2-FGFR1 signaling axis for acquired EMT-E-TKI-R. CRISPR/Cas9-mediated analyses corroborated the importance of ZEB1 in acquired EMT-E-TKI-R, MIR200C and MIR141 to be in an EMT-E-TKI-R-associated auto-regulatory loop with ZEB1, and FGFR1 to mediate cell survival in EMT-E-TKI-R. Conclusions: The current study describes the synchronous genome-wide changes in mRNA-expression, DNA-methylation, and H3K36me3 in NSCLC EMT-E-TKI-R. The omics approaches revealed potential novel diagnostic markers and treatment targets. Besides, the study consolidates the functional impact of the MIR141/MIR200C-ZEB1/ZEB2-FGFR1-signaling axis in NSCLC EMT-E-TKI-R.

Originalsprog	Engelsk
Tidsskrift	Translational Lung Cancer Research
Vol/bind	12
Udgave nummer	1
Sider (fra-til)	42-65
ISSN	2218-6751
DOI	https://doi.org/10.21037/tlcr-22-507
Status	Udgivet - 31. jan. 2023

Bibliografisk note

Funding Information:
The project was supported by Kræftfonden, Dagmar Marshalls Mindelegat, Fabrikant Einar Willumsens Mindelegat, Marie og Børge Kroghs Fond, P. A. Messerschmidt og Hustrus Fond, Thora og Viggo Grove's Mindelegat, Familien Erichsens Familiefond, Axel Muusfeldts Fond, Direktør Emil C. Hertz og Hustru Inger Hertz' Fond, Købmand Sven Hansen og Hustru Ina Hansens Fond, Peetz legat, Frimodt-Heineke Fonden, Else og Mogens Wedell-Wedellsborgs Fond, and Harboefonden.

Publisher Copyright:
© Translational Lung Cancer Research. All rights reserved.

Dokumenter og links

10.21037/tlcr-22-507Licens: CC BY-NC-ND

Open Access VersionForlagets udgivne version, 4,97 MBLicens: CC BY-NC-ND

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Citationsformater

Vad-Nielsen, J., Staunstrup, N. H., Kjeldsen, M. L., Dybdal, N., Flandin, G., De Stradis, C., Daugaard, T. F., Vilsbøll-Larsen, T., Maansson, C. T., Doktor, T. K., Sorensen, B. S., & Nielsen, A. L. (2023). Genome-wide epigenetic and mRNA-expression profiling followed by CRISPR/Cas9-mediated gene-disruptions corroborate the MIR141/MIR200C-ZEB1/ZEB2-FGFR1 axis in acquired EMT-associated EGFR TKI-resistance in NSCLC cells. Translational Lung Cancer Research, 12(1), 42-65. https://doi.org/10.21037/tlcr-22-507

Vad-Nielsen, Johan ; Staunstrup, Nicklas Heine ; Kjeldsen, Magnus Lindkvist et al. / Genome-wide epigenetic and mRNA-expression profiling followed by CRISPR/Cas9-mediated gene-disruptions corroborate the MIR141/MIR200C-ZEB1/ZEB2-FGFR1 axis in acquired EMT-associated EGFR TKI-resistance in NSCLC cells. I: Translational Lung Cancer Research. 2023 ; Bind 12, Nr. 1. s. 42-65.

@article{6a12a19f47c84dce838daf2f31ad4db7,

title = "Genome-wide epigenetic and mRNA-expression profiling followed by CRISPR/Cas9-mediated gene-disruptions corroborate the MIR141/MIR200C-ZEB1/ZEB2-FGFR1 axis in acquired EMT-associated EGFR TKI-resistance in NSCLC cells",

abstract = "Background: Epithelial-mesenchymal-transition (EMT) is an epigenetic-based mechanism contributing to the acquired treatment resistance against receptor tyrosine kinase inhibitors (TKIs) in non-small cell lung cancer (NSCLC) cells harboring epidermal growth factor receptor (EGFR)-mutations. Delineating the exact epigenetic and gene-expression alterations in EMT-associated EGFR TKI-resistance (EMT-E-TKI-R) is vital for improved diagnosis and treatment of NSCLC patients. Methods: We characterized genome-wide changes in mRNA-expression, DNA-methylation and the histone-modification H3K36me3 in EGFR-mutated NSCLC HCC827 cells in result of acquired EMT-E-TKI-R. CRISPR/Cas9 was used to functional examine key findings from the omics analyses. Results: Acquired EMT-E-TKI-R was analyzed with three omics approaches. RNA-sequencing identified 2,233 and 1,972 up- and down-regulated genes, respectively, and among these were established EMT-markers. DNA-methylation EPIC array analyses identified 14,163 and 7,999 hyper- and hypo-methylated, respectively, differential methylated positions of which several were present in EMT-markers. Finally, H3K36me3 chromatin immunoprecipitation (ChIP)-sequencing detected 2,873 and 3,836 genes with enrichment and depletion, respectively, and among these were established EMT-markers. Correlation analyses showed that EMT-E-TKI-R mRNA-expression changes correlated better with H3K36me3 changes than with DNA-methylation changes. Moreover, the omics data supported the involvement of the MIR141/MIR200C-ZEB1/ZEB2-FGFR1 signaling axis for acquired EMT-E-TKI-R. CRISPR/Cas9-mediated analyses corroborated the importance of ZEB1 in acquired EMT-E-TKI-R, MIR200C and MIR141 to be in an EMT-E-TKI-R-associated auto-regulatory loop with ZEB1, and FGFR1 to mediate cell survival in EMT-E-TKI-R. Conclusions: The current study describes the synchronous genome-wide changes in mRNA-expression, DNA-methylation, and H3K36me3 in NSCLC EMT-E-TKI-R. The omics approaches revealed potential novel diagnostic markers and treatment targets. Besides, the study consolidates the functional impact of the MIR141/MIR200C-ZEB1/ZEB2-FGFR1-signaling axis in NSCLC EMT-E-TKI-R.",

keywords = "epigenetics, epithelial-mesenchymal-transition (EMT), Non-small cell lung cancer (NSCLC), tyrosine kinase inhibitor-resistance (TKI-resistance)",

author = "Johan Vad-Nielsen and Staunstrup, {Nicklas Heine} and Kjeldsen, {Magnus Lindkvist} and Nina Dybdal and Guillaume Flandin and {De Stradis}, Claudia and Daugaard, {Tina Fuglsang} and Trine Vilsb{\o}ll-Larsen and Maansson, {Christoffer Trier} and Doktor, {Thomas Koed} and Sorensen, {Boe Sandahl} and Nielsen, {Anders Lade}",

year = "2023",

month = jan,

day = "31",

doi = "10.21037/tlcr-22-507",

language = "English",

volume = "12",

pages = "42--65",

journal = "Translational Lung Cancer Research",

issn = "2218-6751",

publisher = "Pioneer Bioscience Publishing Company",

number = "1",

}

Vad-Nielsen, J, Staunstrup, NH, Kjeldsen, ML, Dybdal, N, Flandin, G, De Stradis, C, Daugaard, TF, Vilsbøll-Larsen, T, Maansson, CT, Doktor, TK, Sorensen, BS & Nielsen, AL 2023, 'Genome-wide epigenetic and mRNA-expression profiling followed by CRISPR/Cas9-mediated gene-disruptions corroborate the MIR141/MIR200C-ZEB1/ZEB2-FGFR1 axis in acquired EMT-associated EGFR TKI-resistance in NSCLC cells', Translational Lung Cancer Research, bind 12, nr. 1, s. 42-65. https://doi.org/10.21037/tlcr-22-507

Genome-wide epigenetic and mRNA-expression profiling followed by CRISPR/Cas9-mediated gene-disruptions corroborate the MIR141/MIR200C-ZEB1/ZEB2-FGFR1 axis in acquired EMT-associated EGFR TKI-resistance in NSCLC cells. / Vad-Nielsen, Johan; Staunstrup, Nicklas Heine; Kjeldsen, Magnus Lindkvist et al.

I: Translational Lung Cancer Research, Bind 12, Nr. 1, 31.01.2023, s. 42-65.

Publikation: Bidrag til tidsskrift › Tidsskriftartikel › Forskning › peer review

TY - JOUR

T1 - Genome-wide epigenetic and mRNA-expression profiling followed by CRISPR/Cas9-mediated gene-disruptions corroborate the MIR141/MIR200C-ZEB1/ZEB2-FGFR1 axis in acquired EMT-associated EGFR TKI-resistance in NSCLC cells

AU - Vad-Nielsen, Johan

AU - Staunstrup, Nicklas Heine

AU - Kjeldsen, Magnus Lindkvist

AU - Dybdal, Nina

AU - Flandin, Guillaume

AU - De Stradis, Claudia

AU - Daugaard, Tina Fuglsang

AU - Vilsbøll-Larsen, Trine

AU - Maansson, Christoffer Trier

AU - Doktor, Thomas Koed

AU - Sorensen, Boe Sandahl

AU - Nielsen, Anders Lade

PY - 2023/1/31

Y1 - 2023/1/31

N2 - Background: Epithelial-mesenchymal-transition (EMT) is an epigenetic-based mechanism contributing to the acquired treatment resistance against receptor tyrosine kinase inhibitors (TKIs) in non-small cell lung cancer (NSCLC) cells harboring epidermal growth factor receptor (EGFR)-mutations. Delineating the exact epigenetic and gene-expression alterations in EMT-associated EGFR TKI-resistance (EMT-E-TKI-R) is vital for improved diagnosis and treatment of NSCLC patients. Methods: We characterized genome-wide changes in mRNA-expression, DNA-methylation and the histone-modification H3K36me3 in EGFR-mutated NSCLC HCC827 cells in result of acquired EMT-E-TKI-R. CRISPR/Cas9 was used to functional examine key findings from the omics analyses. Results: Acquired EMT-E-TKI-R was analyzed with three omics approaches. RNA-sequencing identified 2,233 and 1,972 up- and down-regulated genes, respectively, and among these were established EMT-markers. DNA-methylation EPIC array analyses identified 14,163 and 7,999 hyper- and hypo-methylated, respectively, differential methylated positions of which several were present in EMT-markers. Finally, H3K36me3 chromatin immunoprecipitation (ChIP)-sequencing detected 2,873 and 3,836 genes with enrichment and depletion, respectively, and among these were established EMT-markers. Correlation analyses showed that EMT-E-TKI-R mRNA-expression changes correlated better with H3K36me3 changes than with DNA-methylation changes. Moreover, the omics data supported the involvement of the MIR141/MIR200C-ZEB1/ZEB2-FGFR1 signaling axis for acquired EMT-E-TKI-R. CRISPR/Cas9-mediated analyses corroborated the importance of ZEB1 in acquired EMT-E-TKI-R, MIR200C and MIR141 to be in an EMT-E-TKI-R-associated auto-regulatory loop with ZEB1, and FGFR1 to mediate cell survival in EMT-E-TKI-R. Conclusions: The current study describes the synchronous genome-wide changes in mRNA-expression, DNA-methylation, and H3K36me3 in NSCLC EMT-E-TKI-R. The omics approaches revealed potential novel diagnostic markers and treatment targets. Besides, the study consolidates the functional impact of the MIR141/MIR200C-ZEB1/ZEB2-FGFR1-signaling axis in NSCLC EMT-E-TKI-R.

AB - Background: Epithelial-mesenchymal-transition (EMT) is an epigenetic-based mechanism contributing to the acquired treatment resistance against receptor tyrosine kinase inhibitors (TKIs) in non-small cell lung cancer (NSCLC) cells harboring epidermal growth factor receptor (EGFR)-mutations. Delineating the exact epigenetic and gene-expression alterations in EMT-associated EGFR TKI-resistance (EMT-E-TKI-R) is vital for improved diagnosis and treatment of NSCLC patients. Methods: We characterized genome-wide changes in mRNA-expression, DNA-methylation and the histone-modification H3K36me3 in EGFR-mutated NSCLC HCC827 cells in result of acquired EMT-E-TKI-R. CRISPR/Cas9 was used to functional examine key findings from the omics analyses. Results: Acquired EMT-E-TKI-R was analyzed with three omics approaches. RNA-sequencing identified 2,233 and 1,972 up- and down-regulated genes, respectively, and among these were established EMT-markers. DNA-methylation EPIC array analyses identified 14,163 and 7,999 hyper- and hypo-methylated, respectively, differential methylated positions of which several were present in EMT-markers. Finally, H3K36me3 chromatin immunoprecipitation (ChIP)-sequencing detected 2,873 and 3,836 genes with enrichment and depletion, respectively, and among these were established EMT-markers. Correlation analyses showed that EMT-E-TKI-R mRNA-expression changes correlated better with H3K36me3 changes than with DNA-methylation changes. Moreover, the omics data supported the involvement of the MIR141/MIR200C-ZEB1/ZEB2-FGFR1 signaling axis for acquired EMT-E-TKI-R. CRISPR/Cas9-mediated analyses corroborated the importance of ZEB1 in acquired EMT-E-TKI-R, MIR200C and MIR141 to be in an EMT-E-TKI-R-associated auto-regulatory loop with ZEB1, and FGFR1 to mediate cell survival in EMT-E-TKI-R. Conclusions: The current study describes the synchronous genome-wide changes in mRNA-expression, DNA-methylation, and H3K36me3 in NSCLC EMT-E-TKI-R. The omics approaches revealed potential novel diagnostic markers and treatment targets. Besides, the study consolidates the functional impact of the MIR141/MIR200C-ZEB1/ZEB2-FGFR1-signaling axis in NSCLC EMT-E-TKI-R.

KW - epigenetics

KW - epithelial-mesenchymal-transition (EMT)

KW - Non-small cell lung cancer (NSCLC)

KW - tyrosine kinase inhibitor-resistance (TKI-resistance)

U2 - 10.21037/tlcr-22-507

DO - 10.21037/tlcr-22-507

M3 - Journal article

C2 - 36762066

AN - SCOPUS:85147778438

SN - 2218-6751

VL - 12

SP - 42

EP - 65

JO - Translational Lung Cancer Research

JF - Translational Lung Cancer Research

IS - 1

ER -

Vad-Nielsen J, Staunstrup NH, Kjeldsen ML, Dybdal N, Flandin G, De Stradis C et al. Genome-wide epigenetic and mRNA-expression profiling followed by CRISPR/Cas9-mediated gene-disruptions corroborate the MIR141/MIR200C-ZEB1/ZEB2-FGFR1 axis in acquired EMT-associated EGFR TKI-resistance in NSCLC cells. Translational Lung Cancer Research. 2023 jan. 31;12(1):42-65. doi: 10.21037/tlcr-22-507