title: Upregulation of CXCR4 is functionally crucial for maintenance of stemness in drug-resistant non-small cell lung cancer cells.
abstract: The hypothesis of cancer stem cells has been proposed to explain the therapeutic failure in a variety of cancers including lung cancers. Previously, we demonstrated acquisition of epithelial-mesenchymal transition, a feature highly reminiscent of cancer stem-like cells, in gefitinib-resistant A549 cells (A549/GR). Here, we show that A549/GR cells contain a high proportion of CXCR4+ cells that are responsible for having high potential of self-renewal activity in vitro and tumorigenicity in vivo. A549/GR cells exhibited strong sphere-forming activity and high CXCR4 expression and SDF-1α secretion compared with parent cells. Pharmacological inhibition (AMD3100) and/or siRNA transfection targeting CXCR4 significantly suppressed sphere-forming activity in A549 and A549/GR cells, and in various non-small cell lung cancer (NSCLC) cell lines. A549/GR cells showed enhanced Akt, mTOR and STAT3 (Y705) phosphorylation. Pharmacological inhibition of phosphatidyl inositol 3-kinase or transfection with wild-type PTEN suppressed phosphorylation of Akt, mTOR and STAT3 (Y705), sphere formation, and CXCR4 expression in A549/GR cells, whereas mutant PTEN enhanced these events. Inhibition of STAT3 by WP1066 or siSTAT3 significantly suppressed the sphere formation, but not CXCR4 expression, indicating that STAT3 is a downstream effector of CXCR4-mediated signaling. FACS-sorted CXCR4+ A549/GR cells formed many large spheres, had self-renewal capacity, demonstrated radiation resistance in vitro and exhibited stronger tumorigenic potential in vivo than CXCR4- cells. Lentiviral-transduction of CXCR4 enhanced sphere formation and tumorigenicity in H460 and A549 cells, whereas introduction of siCXCR4 suppressed these activities in A549/GR cells. Our data indicate that CXCR4+ NSCLC cells are strong candidates for tumorigenic stem-like cancer cells that maintain stemness through a CXCR4-medated STAT3 pathway and provide a potential therapeutic target for eliminating these malignant cells in NSCLC.
authors.affiliation: Divisions of Radiation Cancer Research, Korea Institute of Radiological and Medical Sciences, Seoul, Korea.
authors.forename: M-J
authors.initials: MJ
authors.lastname: Jung
authors.forename: J-K
authors.initials: JK
authors.lastname: Rho
authors.forename: Y-M
authors.initials: YM
authors.lastname: Kim
authors.forename: J E
authors.initials: JE
authors.lastname: Jung
authors.forename: Y B
authors.initials: YB
authors.lastname: Jin
authors.forename: Y-G
authors.initials: YG
authors.lastname: Ko
authors.forename: J-S
authors.initials: JS
authors.lastname: Lee
authors.forename: S-J
authors.initials: SJ
authors.lastname: Lee
authors.forename: J C
authors.initials: JC
authors.lastname: Lee
authors.forename: M-J
authors.initials: MJ
authors.lastname: Park
mesh_terms: D000818:Animals; D001596:Benzylamines; D002289:Carcinoma, Non-Small-Cell Lung / Q000188:drug therapy / Q000378:metabolism* / Q000473:pathology*; D045744:Cell Line, Tumor; D054377:Chemokine CXCL12 / Q000378:metabolism; D000080027:Cyclams; D019008:Drug Resistance, Neoplasm; D005260:Female; D006571:Heterocyclic Compounds / Q000494:pharmacology; D006801:Humans; D008175:Lung Neoplasms / Q000188:drug therapy / Q000378:metabolism / Q000473:pathology; D051379:Mice; D008819:Mice, Nude; D016513:Mice, SCID; D014411:Neoplastic Stem Cells / Q000187:drug effects / Q000502:physiology*; D051059:PTEN Phosphohydrolase / Q000235:genetics / Q000378:metabolism; D058539:Phosphatidylinositol 3-Kinase / Q000378:metabolism; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010766:Phosphorylation; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D011725:Pyridines / Q000494:pharmacology; D034622:RNA Interference; D034741:RNA, Small Interfering; D019718:Receptors, CXCR4 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D050796:STAT3 Transcription Factor / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D058570:TOR Serine-Threonine Kinases / Q000378:metabolism; D020032:Tyrphostins / Q000494:pharmacology; D015854:Up-Regulation
chemical_list: D001596:Benzylamines; C527276:CXCR4 protein, human; D054377:Chemokine CXCL12; D000080027:Cyclams; D006571:Heterocyclic Compounds; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011725:Pyridines; D034741:RNA, Small Interfering; D019718:Receptors, CXCR4; D050796:STAT3 Transcription Factor; C494086:STAT3 protein, human; D020032:Tyrphostins; C519885:WP1066; C546842:MTOR protein, human; D058539:Phosphatidylinositol 3-Kinase; D051057:Proto-Oncogene Proteins c-akt; D058570:TOR Serine-Threonine Kinases; D051059:PTEN Phosphohydrolase; C494929:PTEN protein, human; C088327:plerixafor