title: Epigenetic modulation of MAGE-A3 antigen expression in multiple myeloma following treatment with the demethylation agent 5-azacitidine and the histone deacetlyase inhibitor MGCD0103.
abstract: OBJECTIVE
  Immunotherapy targeting MAGE-A3 in multiple myeloma (MM) could eradicate highly aggressive and proliferative clonal cell populations responsible for relapse. However, expression of many cancer-testis antigens, including MAGE-A3, can be heterogeneous, leading to the potential for tumor escape despite MAGE-A3-induced immunity. We hypothesized that a combination of the hypomethylating agent 5-azacitidine (5AC) and the histone deacetylase inhibitor (HDACi) MGCD0103 (MGC) could induce MAGE-A3 expression in MAGE-A3-negative MM, resulting in recognition and killing of MM cells by MAGE-A3-specific cytotoxic T lymphocytes (CTL).
  
  
  METHODS
  Gene expression analyses of MAGE-A3 expression in primary MM patient samples at diagnosis and relapse were completed to identify populations that would benefit from MAGE-A3 immunotherapy. MM cell lines were treated with 5AC and MGC. Real-time polymerase chain reaction (PCR) and Western blotting were performed to assess MAGE-A3 RNA and protein levels, respectively. Chromium-release assays and interferon (IFN) secretion assays were employed to ascertain MAGE-A3 CTL specificity against treated targets.
  
  
  RESULTS
  Gene expression analysis revealed that MAGE-A3 is expressed in MM patients at diagnosis (25%) and at relapse (49%). We observed de novo expression of MAGE-A3 RNA and protein in MAGE-A3-negative cell lines treated with 5AC. MGC treatment alone did not induce expression but sequential 5AC/MGC treatment led to enhanced expression and augmented recognition by MAGE-A3-specific CTL, as assessed by (51)Cr-release assays (P = 0.047) and enzyme-linked immunosorbent assay (ELISA) for IFN-γ secretion (P = 0.004).
  
  
  CONCLUSIONS
  MAGE-A3 is an attractive target for immunotherapy of MM and epigenetic modulation by 5AC, and MGC can induce MAGE-A3 expression and facilitate killing by MAGE-A3-specific CTL.
authors.affiliation: Myeloma Institute for Research and Therapy, University of Arkansas for Medical Sciences, Little Rock, Arkansas 72205, USA.
authors.forename: Amberly
authors.initials: A
authors.lastname: Moreno-Bost
authors.forename: Susann
authors.initials: S
authors.lastname: Szmania
authors.forename: Katie
authors.initials: K
authors.lastname: Stone
authors.forename: Tarun
authors.initials: T
authors.lastname: Garg
authors.forename: Antje
authors.initials: A
authors.lastname: Hoerring
authors.forename: Jackie
authors.initials: J
authors.lastname: Szymonifka
authors.forename: John
authors.initials: J
authors.lastname: Shaughnessy
authors.forename: Bart
authors.initials: B
authors.lastname: Barlogie
authors.forename: H Grant
authors.initials: HG
authors.lastname: Prentice
authors.forename: Frits
authors.initials: F
authors.lastname: van Rhee
mesh_terms: D000951:Antigens, Neoplasm / Q000235:genetics* / Q000276:immunology; D000971:Antineoplastic Combined Chemotherapy Protocols; D001374:Azacitidine / Q000627:therapeutic use*; D001549:Benzamides / Q000627:therapeutic use*; D045744:Cell Line, Tumor; D019175:DNA Methylation / Q000187:drug effects; D044127:Epigenesis, Genetic / Q000187:drug effects*; D005260:Female; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D056572:Histone Deacetylase Inhibitors / Q000627:therapeutic use*; D006801:Humans; D008297:Male; D009101:Multiple Myeloma / Q000235:genetics / Q000276:immunology / Q000628:therapy*; D009363:Neoplasm Proteins / Q000235:genetics* / Q000276:immunology; D011743:Pyrimidines / Q000627:therapeutic use*; D055502:Secondary Prevention; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology / Q000637:transplantation*
chemical_list: D000951:Antigens, Neoplasm; D001549:Benzamides; D056572:Histone Deacetylase Inhibitors; C072013:MAGEA3 protein, human; D009363:Neoplasm Proteins; D011743:Pyrimidines; C523184:mocetinostat; D001374:Azacitidine