biostudies-literatureUnknown9(30)Yu CHAfatinib, used for the first-line treatment of non-small-cell lung carcinoma (NSCLC) patients with distinct epidermal growth factor receptor (EGFR) mutations, inactivates EGFR by mimicking ATP structure and forming a covalent adduct with EGFR. We developed a method to unravel potential targets of afatinib in NSCLC cells through immunoprecipitation of afatinib-labeling proteins with anti-afatinib antiserum and mass spectrometry analysis. Ribonucleotide reductase (RNR) is one of target proteins of afatinib revealed by this method. Treatment of afatinib at 10-100 nM potently inhibited intracellular RNR activity in an <i>in vitro</i> assay using permeabilized PC-9 cells (formerly known as PC-14). PC-9 cells treated with 10 μM afatinib displayed elevated markers of DNA damage. Long-term treatmeOncotarget21512-21529https://www.ebi.ac.uk/biostudies/studies/S-EPMC5940374biostudies-literatureAntibody-assisted target identification reveals afatinib, an EGFR covalent inhibitor, down-regulating ribonucleotide reductase.PMC5940374Ho YYYu CHChang GDLee MSTu HFKhoo KHChou CCHuang WCfalseAntibody-assisted target identification reveals afatinib, an EGFR covalent inhibitor, down-regulating ribonucleotide reductase.Afatinib, used for the first-line treatment of non-small-cell lung carcinoma (NSCLC) patients with distinct epidermal growth factor receptor (EGFR) mutations, inactivates EGFR by mimicking ATP structure and forming a covalent adduct with EGFR. We developed a method to unravel potential targets of afatinib in NSCLC cells through immunoprecipitation of afatinib-labeling proteins with anti-afatinib antiserum and mass spectrometry analysis. Ribonucleotide reductase (RNR) is one of target proteins of afatinib revealed by this method. Treatment of afatinib at 10-100 nM potently inhibited intracellular RNR activity in an <i>in vitro</i> assay using permeabilized PC-9 cells (formerly known as PC-14). PC-9 cells treated with 10 μM afatinib displayed elevated markers of DNA damage. Long-term treatme2018-01-01T00:00:00Z2018 Apr2026-05-04T22:59:36.922Z2019-03-26T23:37:41ZS-EPMC59403742976555610.18632/oncotarget.25177