title: Up-regulation of HLA class-I antigen expression and antigen-specific CTL response in cervical cancer cells by the demethylating agent hydralazine and the histone deacetylase inhibitor valproic acid.
abstract: BACKGROUND
  DNA hypermethylation and histone deacetylation are epigenetic events that contribute to the absence or downregulated expression of different components of the tumor recognition complex. These events affect the processing and presentation of antigenic peptides to CTLs by HLA class-I molecules. In this work evaluated the effect of the DNA hypomethylating agent hydralazine and the histone deacetylase inhibitor valproic acid, on the expression of HLA class-I molecules and on the antigen-specific immune recognition of cervical cancer cells.
  
  
  METHODS
  Cell lines C33A (HPV-), CaSki (HPV-16+) and MS751 (HPV-18+) were treated with hydralazine and valproic acid to assess the expression of HLA class-I molecules by flow cytometry and RT-PCR. Promoter methylation of HLA class-I -A, -B and C, was also evaluated by Methylation-Specific PCR. Primary cervical tumors of four HLA-A*0201 allele patients were typed for HPV and their CTL's stimulated in vitro with the T2 cell line previously loaded with 50 microM of the HPV peptides. Cytotoxicity of stimulated CTL's was assayed against Caski and MS751 cells pre-treated with hydralazine and valproic acid.
  
  
  RESULTS
  Valproic acid and hydralazine/valproic acid up-regulated the constitutive HLA class-I expression as evaluated by flow cytometry and RT-PCR despite constitutive promoter demethylation at these loci. Hydralazine and valproic acid in combination but no IFN-gamma hyperacetylated histone H4 as evaluated by ChiP assay. The antigenic immune recognition of CaSki and MS751 cells by CTLs specific to HPV-16/18 E6 and E7-derived epitopes, was increased by VA and H/VA and the combination of H/VA/IFN-gamma.
  
  
  CONCLUSIONS
  These results support the potential use of hydralazine and valproic acid as an adjuvant for immune intervention in cervical cancer patients whenever clinical protocols based on tumor antigen recognition is desirable, like in those cases where the application of E6 and E7 based therapeutic vaccines is used.
authors.affiliation: Laboratorio de Inmunobiología, Unidad de Investigación en Diferenciación Celular y Cáncer. FES-Zaragoza, UNAM, México. mogl@servidor.unam.mx
authors.forename: María de Lourdes
authors.initials: Mde L
authors.lastname: Mora-García
authors.forename: Alfonso
authors.initials: A
authors.lastname: Duenas-González
authors.forename: Jorge
authors.initials: J
authors.lastname: Hernández-Montes
authors.forename: Erick
authors.initials: E
authors.lastname: De la Cruz-Hernández
authors.forename: Enrique
authors.initials: E
authors.lastname: Pérez-Cárdenas
authors.forename: Benny
authors.initials: B
authors.lastname: Weiss-Steider
authors.forename: Edelmiro
authors.initials: E
authors.lastname: Santiago-Osorio
authors.forename: Vianney Francisco
authors.initials: VF
authors.lastname: Ortíz-Navarrete
authors.forename: Víctor Hugo
authors.initials: VH
authors.lastname: Rosales
authors.forename: David
authors.initials: D
authors.lastname: Cantú
authors.forename: Marcela
authors.initials: M
authors.lastname: Lizano-Soberón
authors.forename: Martha Patricia
authors.initials: MP
authors.lastname: Rojo-Aguilar
authors.forename: Alberto
authors.initials: A
authors.lastname: Monroy-García