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
Y1 - 2023/1
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)
UR - http://www.scopus.com/inward/record.url?scp=85147778438&partnerID=8YFLogxK
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 -