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Regulation of epidermal growth factor receptor signaling and erlotinib sensitivity in head and neck cancer cells by miR-7

ABSTRACT: Elevated expression and activity of the epidermal growth factor receptor (EGFR)/protein kinase B (Akt) signaling pathway is associated with development, progression and treatment resistance of head and neck cancer (HNC). Several studies have demonstrated that microRNA-7 (miR-7) regulates EGFR expression and Akt activity in a range of cancer cell types via its specific interaction with the EGFR mRNA 3′ untranslated region (3′-UTR). In the present study, we found that miR-7 regulated EGFR expression and Akt activity in HNC cell lines, and that this was associated with reduced growth in vitro and in vivo of cells (HN5) that were sensitive to the EGFR tyrosine kinase inhibitor (TKI) erlotinib (Tarceva). miR-7 acted synergistically with erlotinib to inhibit growth of erlotinib-resistant FaDu cells, an effect associated with increased inhibition of Akt activity. Microarray analysis of HN5 and FaDu cell lines transfected with miR-7 identified a common set of downregulated miR-7 target genes, providing insight into the tumor suppressor function of miR-7. Furthermore, we identified several target miR-7 mRNAs with a putative role in the sensitization of FaDu cells to erlotinib. Together, these data support the coordinate regulation of Akt signaling by miR-7 in HNC cells and suggest the therapeutic potential of miR-7 alone or in combination with EGFR TKIs in this disease.

ORGANISM(S): Homo sapiens

PROVIDER: GSE40130 | GEO | 2012/08/16

SECONDARY ACCESSION(S): PRJNA172887

REPOSITORIES: GEO

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Project description:Erlotinib is a tyrosine kinase inhibitor (TKI) that is approved as a second-line monotherapy in patients with advanced non-small cell lung cancer (NSCLC). In these patients, erlotinib prolongs survival but its benefit remains modest since many tumors express wild-type EGF receptor (wtEGFR) lacking a TKI-sensitizing mutation, develop a second-site EGFR mutation, e.g., EGFR-L858R/T790M, or activate an alternate receptor tyrosine kinase, e.g., through MET amplification. To test potential drug combinations that could improve the efficacy of erlotinib, we combined erlotinib with quinacrine, which inhibits the FACT (facilitates chromatin transcription) complex that is required for nuclear factor-M-NM-:B (NF-M-NM-:B) transcriptional activity. In A549 (wtEGFR), H1975 (EGFR-L858R/T790M) and H1993 (MET amplification) NSCLC cells, the combination of erlotinib and quinacrine was highly synergistic, as quantified by Chou-Talalay combination indices. The combination inhibited colony formation, induced cell cycle arrest and apoptosis, and slowed xenograft tumor growth. Quinacrine decreased the level of active FACT subunit SSRP1 and suppressed NF-M-NM-:B-dependent luciferase activity. Knockdown of SSRP1 decreased cell growth and sensitized cells to erlotinib. Transcriptomic profiling showed that quinacrine, alone or in combination with erlotinib, significantly affected cell cycle-related genes that contain binding sites for transcription factors that regulate SSRP1 target genes. As potential biomarkers of drug synergy, we identified genes that were more strongly suppressed by the combination than by either single treatment, and whose increased expression predicted poorer survival in lung adenocarcinoma patients. Combination of quinacrine and erlotinib is synergistic in NSCLC through suppression of FACT, resulting in inhibition of cell cycle progression. RNA was isolated from cells treated with 1 M-NM-<M erlotinib, 3 M-NM-<M or 5 M-NM-<M quinacrine or a combination of both for 6, 12, 24, or 48 h using the RNeasy Mini Kit (QIAGEN) according to the manufacturerM-bM-^@M-^Ys instructions. Microarray analysis was performed using the Affymetrix Human Gene 2.1 ST Array at the Gene Expression & Genotyping Core Facility at Case Comprehensive Cancer Center. Raw CEL files were pre-processed using the Affymetrix Expression Console Software 1.30 with Robust Multi-array Average (RMA) normalization (background correction, quantile normalization and log2 transformation). Probes were annotated using the HuGene 2.1 st hg19 probeset annotation files downloaded from the Affymetrix website.

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Regulation of epidermal growth factor receptor signaling and erlotinib sensitivity in head and neck cancer cells by miR-7

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