title: Specifically targeting ERK1 or ERK2 kills melanoma cells.
abstract: BACKGROUND
  Overcoming the notorious apoptotic resistance of melanoma cells remains a therapeutic challenge given dismal survival of patients with metastatic melanoma. However, recent clinical trials using a BRAF inhibitor revealed encouraging results for patients with advanced BRAF mutant bearing melanoma, but drug resistance accompanied by recovery of phospho-ERK (pERK) activity present challenges for this approach. While ERK1 and ERK2 are similar in amino acid composition and are frequently not distinguished in clinical reports, the possibility they regulate distinct biological functions in melanoma is largely unexplored.
  
  
  METHODS
  Rather than indirectly inhibiting pERK by targeting upstream kinases such as BRAF or MEK, we directly (and near completely) reduced ERK1 and ERK2 using short hairpin RNAs (shRNAs) to achieve sustained inhibition of pERK1 and/or pERK2.
  
  
  CONCLUSIONS
  Using A375 melanoma cells containing activating BRAFV600E mutation, silencing ERK1 or ERK2 revealed some differences in their biological roles, but also shared roles by reduced cell proliferation, colony formation in soft agar and induced apoptosis. By contrast, chemical mediated inhibition of mutant BRAF (PLX4032) or MEK (PD0325901) triggered less killing of melanoma cells, although they did inhibit proliferation. Death of melanoma cells by silencing ERK1 and/or ERK2 was caspase dependent and accompanied by increased levels of Bak, Bad and Bim, with reduction in p-Bad and detection of activated Bax levels and loss of mitochondrial membrane permeability. Rare treatment resistant clones accompanied silencing of either ERK1 and/or ERK2. Unexpectedly, directly targeting ERK levels also led to reduction in upstream levels of BRAF, CRAF and pMEK, thereby reinforcing the importance of silencing ERK as regards killing and bypassing drug resistance.
  
  
  CONCLUSIONS
  Selectively knocking down ERK1 and/or ERK2 killed A375 melanoma cells and also increased the ability of PLX4032 to kill A375 cells. Thus, a new therapeutic window is open for future clinical trials in which agents targeting ERK1 and ERK2 should be considered in patients with melanoma.
authors.affiliation: Division of Dermatology, Michigan State University, College of Human Medicine, Grand Rapids, MI 49503, USA.
authors.forename: Jianzhong
authors.initials: J
authors.lastname: Qin
authors.forename: Hong
authors.initials: H
authors.lastname: Xin
authors.forename: Brian J
authors.initials: BJ
authors.lastname: Nickoloff
mesh_terms: D000362:Agar; D001549:Benzamides / Q000494:pharmacology; D020169:Caspases / Q000378:metabolism; D016923:Cell Death / Q000187:drug effects; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D004159:Diphenylamine / Q000031:analogs & derivatives / Q000494:pharmacology; D019008:Drug Resistance, Neoplasm / Q000187:drug effects; D004354:Drug Screening Assays, Antitumor; D055785:Gene Knockdown Techniques; D020868:Gene Silencing / Q000187:drug effects; D006801:Humans; D007211:Indoles / Q000494:pharmacology; D008545:Melanoma / Q000201:enzymology* / Q000473:pathology*; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism*; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism*; D009363:Neoplasm Proteins / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D011480:Protease Inhibitors / Q000494:pharmacology; D034741:RNA, Small Interfering / Q000378:metabolism; D013449:Sulfonamides / Q000494:pharmacology; D014410:Tumor Stem Cell Assay; D000077484:Vemurafenib; D051028:bcl-2-Associated X Protein / Q000378:metabolism
chemical_list: D001549:Benzamides; D007211:Indoles; D009363:Neoplasm Proteins; D011480:Protease Inhibitors; D034741:RNA, Small Interfering; D013449:Sulfonamides; D051028:bcl-2-Associated X Protein; D000077484:Vemurafenib; C506614:mirdametinib; D000362:Agar; D004159:Diphenylamine; D011065:Poly(ADP-ribose) Polymerases; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020169:Caspases