title: Acquired resistance to erlotinib in A-431 epidermoid cancer cells requires down-regulation of MMAC1/PTEN and up-regulation of phosphorylated Akt.
abstract: Erlotinib (Tarceva), an epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor, has clinical activity in advanced lung cancer, but disease that initially responds to erlotinib eventually progresses. The mechanism of this acquired resistance is unclear. We established two erlotinib-resistant pools of A-431 cells, a well-characterized epidermoid cancer cell line that constitutively overexpresses EGFR and is sensitive to erlotinib, by continuous exposure to erlotinib over a 6-month period. The extent of EGFR gene amplification or mutation of the EGFR tyrosine kinase domain was not altered in the resistant cells. Intracellular erlotinib concentrations, determined by liquid chromatography-tandem mass spectrometry, were almost the same in all three cell lines. Immunoprecipitation with EGFR antibody followed by detection with phosphotyrosine antibody revealed that erlotinib effectively reduced EGFR phosphorylation in both parental cells and resistant cells. Erlotinib induced mutated in multiple advanced cancers 1/phosphatase and tensin homologue (MMAC1/PTEN) and suppressed phosphorylated Akt (Ser(473)) but not in the erlotinib-resistant cells. Overexpression of MMAC1/PTEN by transfection with Ad.MMAC1/PTEN or by pharmacologic suppression of Akt activity restored erlotinib sensitivity in both resistant pools. Further, transfection of parental A-431 cells with constitutively active Akt was sufficient to cause resistance to erlotinib. We propose that acquired erlotinib resistance associated with MMAC1/PTEN down-regulation and Akt activation could be overcome by inhibitors of signaling through the phosphatidylinositol 3-kinase pathway.
authors.affiliation: Breast Cancer Translational Research Laboratory, The University of Texas M. D. Anderson Cancer Center, Houston, Texas 77030-4009, USA.
authors.forename: Fumiyuki
authors.initials: F
authors.lastname: Yamasaki
authors.forename: Mary J
authors.initials: MJ
authors.lastname: Johansen
authors.forename: Dongwei
authors.initials: D
authors.lastname: Zhang
authors.forename: Savitri
authors.initials: S
authors.lastname: Krishnamurthy
authors.forename: Edward
authors.initials: E
authors.lastname: Felix
authors.forename: Chandra
authors.initials: C
authors.lastname: Bartholomeusz
authors.forename: Richard J
authors.initials: RJ
authors.lastname: Aguilar
authors.forename: Kaoru
authors.initials: K
authors.lastname: Kurisu
authors.forename: Gordon B
authors.initials: GB
authors.lastname: Mills
authors.forename: Gabriel N
authors.initials: GN
authors.lastname: Hortobagyi
authors.forename: Naoto T
authors.initials: NT
authors.lastname: Ueno
mesh_terms: D000970:Antineoplastic Agents / Q000494:pharmacology*; D001483:Base Sequence; D015153:Blotting, Western; D002294:Carcinoma, Squamous Cell / Q000378:metabolism*; D045744:Cell Line, Tumor; D002853:Chromatography, Liquid; D015536:Down-Regulation; D019008:Drug Resistance, Neoplasm / Q000502:physiology*; D066246:ErbB Receptors / Q000235:genetics / Q000378:metabolism; D000069347:Erlotinib Hydrochloride; D005434:Flow Cytometry; D018628:Gene Dosage; D006801:Humans; D047468:Immunoprecipitation; D017404:In Situ Hybridization, Fluorescence; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D009154:Mutation; D051059:PTEN Phosphohydrolase / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism*; D011799:Quinazolines / Q000494:pharmacology*; D014162:Transfection; D015854:Up-Regulation
chemical_list: D000970:Antineoplastic Agents; D047428:Protein Kinase Inhibitors; D011799:Quinazolines; D000069347:Erlotinib Hydrochloride; D066246:ErbB Receptors; D051057:Proto-Oncogene Proteins c-akt; D051059:PTEN Phosphohydrolase; C494929:PTEN protein, human