title: Proteomic analysis of acquired tamoxifen resistance in MCF-7 cells reveals expression signatures associated with enhanced migration.
abstract: BACKGROUND
  Acquired tamoxifen resistance involves complex signaling events that are not yet fully understood. Successful therapeutic intervention to delay the onset of hormone resistance depends critically on mechanistic elucidation of viable molecular targets associated with hormone resistance. This study was undertaken to investigate the global proteomic alterations in a tamoxifen resistant MCF-7 breast cancer cell line obtained by long term treatment of the wild type MCF-7 cell line with 4-hydroxytamoxifen (4-OH Tam).
  
  
  METHODS
  We cultured MCF-7 cells with 4-OH Tam over a period of 12 months to obtain the resistant cell line. A gel-free, quantitative proteomic method was used to identify and quantify the proteome of the resistant cell line. Nano-flow high-performance liquid chromatography coupled to high resolution Fourier transform mass spectrometry was used to analyze fractionated peptide mixtures that were isobarically labeled from the resistant and control cell lysates. Real time quantitative PCR and Western blots were used to verify selected proteomic changes. Lentiviral vector transduction was used to generate MCF-7 cells stably expressing S100P. Online pathway analysis was performed to assess proteomic signatures in tamoxifen resistance. Survival analysis was done to evaluate clinical relevance of altered proteomic expressions.
  
  
  RESULTS
  Quantitative proteomic analysis revealed a wide breadth of signaling events during transition to acquired tamoxifen resistance. A total of 629 proteins were found significantly changed with 364 up-regulated and 265 down-regulated. Collectively, these changes demonstrated the suppressed state of estrogen receptor (ER) and ER-regulated genes, activated survival signaling and increased migratory capacity of the resistant cell line. The protein S100P was found to play a critical role in conferring tamoxifen resistance and enhanced cell motility.
  
  
  CONCLUSIONS
  Our data demonstrate that the adaptive changes in the proteome of tamoxifen resistant breast cancer cells are characterized by down-regulated ER signaling, activation of alternative survival pathways, and enhanced cell motility through regulation of the actin cytoskeleton dynamics. Evidence also emerged that S100P mediates acquired tamoxifen resistance and migration capacity.
authors.affiliation: Department of Chemistry, Xavier University of Louisiana, 1 Drexel Drive, New Orleans, LA 70125, USA.
authors.forename: Changhua
authors.initials: C
authors.lastname: Zhou
authors.forename: Qiu
authors.initials: Q
authors.lastname: Zhong
authors.forename: Lyndsay V
authors.initials: LV
authors.lastname: Rhodes
authors.forename: Ian
authors.initials: I
authors.lastname: Townley
authors.forename: Melyssa R
authors.initials: MR
authors.lastname: Bratton
authors.forename: Qiang
authors.initials: Q
authors.lastname: Zhang
authors.forename: Elizabeth C
authors.initials: EC
authors.lastname: Martin
authors.forename: Steven
authors.initials: S
authors.lastname: Elliott
authors.forename: Bridgette M
authors.initials: BM
authors.lastname: Collins-Burow
authors.forename: Matthew E
authors.initials: ME
authors.lastname: Burow
authors.forename: Guangdi
authors.initials: G
authors.lastname: Wang
mesh_terms: D008841:Actin Cytoskeleton / Q000378:metabolism; D018931:Antineoplastic Agents, Hormonal / Q000494:pharmacology*; D001943:Breast Neoplasms / Q000188:drug therapy* / Q000235:genetics / Q000378:metabolism* / Q000401:mortality / Q000473:pathology*; D002135:Calcium-Binding Proteins / Q000235:genetics / Q000378:metabolism; D045744:Cell Line, Tumor; D002465:Cell Movement; D015536:Down-Regulation / Q000187:drug effects; D019008:Drug Resistance, Neoplasm; D005260:Female; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D009363:Neoplasm Proteins / Q000235:genetics / Q000378:metabolism; D040901:Proteomics / Q000379:methods; D011960:Receptors, Estrogen / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D013629:Tamoxifen / Q000494:pharmacology*; D053719:Tandem Mass Spectrometry
chemical_list: D018931:Antineoplastic Agents, Hormonal; D002135:Calcium-Binding Proteins; D009363:Neoplasm Proteins; D011960:Receptors, Estrogen; C103003:S100P protein, human; D013629:Tamoxifen