title: Autophagosome-mediated EGFR down-regulation induced by the CK2 inhibitor enhances the efficacy of EGFR-TKI on EGFR-mutant lung cancer cells with resistance by T790M.
abstract: Protein kinase CK2 has diverse functions promoting and maintaining cancer phenotypes. We investigated the effect of CK2 inhibition in lung cancer cells with T790M-mediated resistance to the EGFR-TK inhibitor. Resistant sublines of PC-9 to gefitinib (PC-9/GR) and erlotinib (PC-9/ER) were established by previous study, and T790M secondary mutation was found in both resistant sublines. A decrease of EGFR by siRNA treatment effectively controlled the growth of resistant cells, thus suggesting that they still have EGFR-dependency. CX-4945, a potent and selective CK2 inhibitor, induced autophagy in PC-9/GR and PC-9/ER, and which was supported by the induction of autophagic vacuoles and microtubule-associated protein 1 light chain 3 (LC3) expression, and the increase of punctate fluorescent signals in resistant cells pre-transfected with green fluorescent protein (GFP)-tagged LC3. However, the withdrawal of CX-4945 led to the recovery of cancer cells with autophagy. We found that the induction of autophagy by CX-4945 in both resistant cells was CK2 dependent by using small interfering RNA against CK2. The treatment with CX-4945 alone induced a minimal growth inhibition in resistant cells. However, combined treatment of CX-4945 and EGFR-TKI effectively inhibited cancer-cell proliferation and induced apoptosis. CX-4945 increased the translocation of EGFR from the cell surface into the autophagosome, subsequently leading to the decrease of EGFR while inhibition of autophagy by 3MA or Atg7-targeted siRNA pretreatment reduced the decrease of EGFR by CX-4945. Accordingly, apoptosis by a combination of CX-4945 and EGFR-TKI was suppressed by 3MA or Atg7-targeted siRNA pretreatment, thus suggesting that autophagosome-mediated EGFR down-regulation would have an important role regarding apoptotic cell death by EGFR-TKI. Combined treatment of the CK2 inhibitor and EGFR-TKI may be a promising strategy for overcoming T790M-mediated resistance.
authors.affiliation: Department of Pulmonary and Critical Care Medicine, Asan Medical Center, College of Medicine, University of Ulsan, Seoul, Korea; College of Pharmacy, Chung-Ang University, Seoul, Korea.
authors.forename: Kwang Sup
authors.initials: KS
authors.lastname: So
authors.affiliation: Department of Internal Medicine, Korea Cancer Center Hosptial, Seoul, Korea.
authors.forename: Cheol Hyeon
authors.initials: CH
authors.lastname: Kim
authors.affiliation: Department of Pulmonary and Critical Care Medicine, Asan Medical Center, College of Medicine, University of Ulsan, Seoul, Korea; Asan Institute for Life Sciences, Asan Medical Center, College of Medicine, University of Ulsan, Seoul, Korea.
authors.forename: Jin Kyung
authors.initials: JK
authors.lastname: Rho
authors.affiliation: Department of Pulmonary and Critical Care Medicine, Asan Medical Center, College of Medicine, University of Ulsan, Seoul, Korea.
authors.forename: Sun Ye
authors.initials: SY
authors.lastname: Kim
authors.affiliation: Department of Pulmonary and Critical Care Medicine, Asan Medical Center, College of Medicine, University of Ulsan, Seoul, Korea.
authors.forename: Yun Jung
authors.initials: YJ
authors.lastname: Choi
authors.affiliation: Department of Pathology, Asan Medical Center, College of Medicine, University of Ulsan, Seoul, Korea.
authors.forename: Joon Seon
authors.initials: JS
authors.lastname: Song
authors.affiliation: Department of Pulmonary and Critical Care Medicine, Asan Medical Center, College of Medicine, University of Ulsan, Seoul, Korea.
authors.forename: Woo Sung
authors.initials: WS
authors.lastname: Kim
authors.affiliation: Department of Pulmonary and Critical Care Medicine, Asan Medical Center, College of Medicine, University of Ulsan, Seoul, Korea; Department of Oncology, Asan Medical Center, College of Medicine, University of Ulsan, Seoul, Korea.
authors.forename: Chang Min
authors.initials: CM
authors.lastname: Choi
authors.affiliation: College of Pharmacy, Chung-Ang University, Seoul, Korea.
authors.forename: Young Jin
authors.initials: YJ
authors.lastname: Chun
authors.affiliation: Department of Oncology, Asan Medical Center, College of Medicine, University of Ulsan, Seoul, Korea.
authors.forename: Jae Cheol
authors.initials: JC
authors.lastname: Lee
mesh_terms: D017209:Apoptosis / Q000187:drug effects; D001343:Autophagy / Q000187:drug effects; D047390:Casein Kinase II / Q000037:antagonists & inhibitors*; D045744:Cell Line, Tumor; D019008:Drug Resistance, Neoplasm; D066246:ErbB Receptors / Q000235:genetics*; D000069347:Erlotinib Hydrochloride; D000077156:Gefitinib; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects*; D006801:Humans; D008175:Lung Neoplasms / Q000235:genetics* / Q000378:metabolism*; D009154:Mutation; D009287:Naphthyridines / Q000494:pharmacology; D010588:Phagosomes / Q000378:metabolism*; D010619:Phenazines; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*; D011799:Quinazolines / Q000494:pharmacology
chemical_list: D009287:Naphthyridines; D010619:Phenazines; D047428:Protein Kinase Inhibitors; D011799:Quinazolines; C555142:silmitasertib; D000069347:Erlotinib Hydrochloride; D066246:ErbB Receptors; D047390:Casein Kinase II; D000077156:Gefitinib