Epigenetic therapy with hydralazine and magnesium valproate added to chemoradiation in cervical carcinoma.

As a number of epigenetic alterations have been described in cervical cancer which may participate in the development and progression of this tumor, and the primary treatment for locally advanced disease is concurrent chemoradiation, we wanted to evaluate the efficacy and safety of hydralazine and valproate added to radiation and cisplatin. In addition, the transcriptome changes induced by the epigenetic drugs were also evaluated. Keywords: Cervical Cancer, epigenetic therapy, valproate, hydralazine, transcriptional response, epigenetic drugs Previously untreated patients with histological diagnosis of carcinoma of the cervix and FIGO staged IIIB were entered into this phase II study. Patients received a single oral dose of 500 mg of sulphamethazine early in the morning, and urine was collected within the ensuing 6 h for phenotyping of acetylator status as reported. Afterward, patients began treatment (day –7) with a daily dose of a slow-release formulation of hydralazine tablets containing either 182 mg for rapid, or 83 mg for slow acetylators. Magnesium valproate tablets of 700 mg were also administered as a slow-release formulation at a dose of 30 mg/kg t.i.d. Both were administered until intracavitary therapy was completed. Punch biopsies were taken from primary cervical tumors at diagnosis and at day 8 of treatment with hydralazine and magnesium valproate prior to the first dose of cisplatin and external radiation. Part of the biopsy was sent to the Institution’s Pathology Department for routine hematoxilin and eosin evaluation. The remaining biopsy specimen was immediately frozen at –70°C for biological analyses. RNA isolation was done in TRIzol according to manufacturer’s instructions. One μg of total RNA was used as input material. All reagents were provided in the CodeLink expression assay kit (Amersham, Piscataway, NJ USA). First-strand cDNA was generated using Superscript II reverse transcriptase and a T7 primer. Subsequently, second-strand cDNA was produced using E. coli DNA polymerase I and RNase H. The resultant double-stranded cDNA was purified on a QIAquick column (Qiagen, Valencia, CA) and cRNA was generated via an in vitro transcription reaction using T7 RNA polymerase and biotin-11-UTP (Perkin-Elmer, Boston, MA). cRNA was purified on an RNeasy column (Qiagen), quantified by UV spectrophotometry, and 10ug were then fragmented by heating at 94°C for 20 minutes in the presence of magnesium. The fragmented cRNA was hybridized overnight at 37°C in hybridization buffer to a CodeLink Whole Human Genome Bioarray in an Innova 4080 shaking incubator (New Brunswick, Edison, NJ) at 300 rpm.