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DNA Polymerase Alpha Subunit B Is a Binding Protein for Erlotinib Resistance in Non-Small Cell Lung Cancer.

ABSTRACT: Erlotinib inhibits epithelial growth factor receptor (EGFR) kinase activity and is used to treat non-small cell lung cancer (NSCLC). Despite its high efficacy, recurrence can occur in patients who become resistant to the drug. To address the underlying mechanism of Erlotinib resistance, we investigated additional mechanisms related to mode-of-drug-action, by multiple protein-binding interactions, besides EGFR by using drug affinity responsive target stability (DARTS) and liquid chromatography-mass spectrometry (LC-MS/MS) methods with non-labeled Erlotinib. DNA polymerase alpha subunit B (POLA2) was identified as a new Erlotinib binding protein that was validated by the DARTS platform, complemented with cellular thermal shift assays. Genetic knock-down of POLA2 promoted the anti-proliferative effect of the drug in the Erlotinib-resistant cell line H1299 with high POLA2 expression, whereas the overexpression of POLA2 restored anti-proliferative effects in the Erlotinib-sensitive cell line HCC827 with low POLA2 expression. Importantly, POLA2 expression levels in four NSCLC cell lines were positively correlated with anti-proliferative Erlotinib efficacy (Pearson correlation coefficient, R = 0.9886). These results suggest that POLA2 is a novel complementary target protein of Erlotinib, and could clinically provide validity as a surrogate marker for drug resistance in patients with NSCLC.

SUBMITTER: Kim TY

PROVIDER: S-EPMC7564424 | biostudies-literature | 2020 Sep

REPOSITORIES: biostudies-literature

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DNA Polymerase Alpha Subunit B Is a Binding Protein for Erlotinib Resistance in Non-Small Cell Lung Cancer.

Kim Tae Young TY Ji Eun Sun ES Lee Ju Yeon JY Kim Jin Young JY Yoo Jong Shin JS Szasz A Marcell AM Dome Balazs B Marko-Varga Gyorgy G Kwon Ho Jeong HJ

Cancers 20200913 9

Erlotinib inhibits epithelial growth factor receptor (EGFR) kinase activity and is used to treat non-small cell lung cancer (NSCLC). Despite its high efficacy, recurrence can occur in patients who become resistant to the drug. To address the underlying mechanism of Erlotinib resistance, we investigated additional mechanisms related to mode-of-drug-action, by multiple protein-binding interactions, besides EGFR by using drug affinity responsive target stability (DARTS) and liquid chromatography-ma ...[more]

PMID: 32933200

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Project description:Currently, resistance to tyrosine kinase inhibitors, such as erlotinib, has become a major obstacle for improving the clinical outcome of patients with metastatic and advanced‑stage non-small cell lung cancer (NSCLC). While cell behavior can be modulated by long non-coding RNAs (lncRNAs), the roles of lncRNAs within extracellular vesicles (exosomes) are largely unknown. To this end, in this study, the involvement and regulatory functions of potential lncRNAs wrapped by exosomes during the development of chemoresistance in human NSCLC were investigated. Erlotinib-resistant cell lines were established by grafting HCC827 and HCC4006 cells into mice and which were treated with erlotinib. After one treatment course, xenografted NSCLC cells were isolated and transplanted into nude mice again followed by erlotinib treatment. This process was repeated until 4th generation xenografts were isolated and confirmed to be erlotinib-resistant NSCLC cells. lncRNA microarray assays followed by RT‑qPCR were then performed which identified that lncRNA RP11‑838N2.4 was upregulated in erlotinib-resistant cells when compared to normal NSCLC cells. Furthermore, bioinformatics analysis and chromatin immunoprecipitation revealed that forkhead box protein O1 (FOXO1) could bind to the promoter region of lncRNA RP11‑838N2.4, resulting in its silencing through the recruitment of histone deacetylase. Functional experiments demonstrated that the knockdown of lncRNA RP11‑838N2.4 potently promoted erlotinib-induced cytotoxicity. Furthermore, extracellular lncRNA RP11‑838N2.4 could be incorporated into exosomes and transmitted to sensitive cells, thus disseminating erlotinib resistance. Treatment-sensitive cells with exosomes containing lncRNA RP11‑838N2.4 induced erlotinib resistance, while the knockdown of lncRNA RP11‑838N2.4 abrogated this effect. In addition, the serum expression levels of exosomal lncRNA RP11‑838N2.4 were upregulated in patients exhibiting resistance to erlotinib treatment. On the whole, exosomal lncRNA RP11‑838N2.4 may serve as a therapeutic target for patients with NSCLC.

Dataset Information

DNA Polymerase Alpha Subunit B Is a Binding Protein for Erlotinib Resistance in Non-Small Cell Lung Cancer.

Publications

DNA Polymerase Alpha Subunit B Is a Binding Protein for Erlotinib Resistance in Non-Small Cell Lung Cancer.

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