Abstract 289: Aberrant DNA methylation of DLX4 and SIM1 is a predictive marker for disease progression of uterine cervical low-grade squamous intraepithelial lesion

Background: Cervical cancer is the second-most common cancer in women worldwide. In Japan, it was the 11th leading cause of death from cancer for women in 2008 but there has been a unique increase of cervical cancer incidence among women in the 20-39-year age group in spite of the decrease of it among women over age 40. High-risk (hr) HPV infection initiates an early event through which an accumulation of genetic and epigenetic aberrations follows. Aberrant DNA methylation (abMet) is shown to provide disease biomarkers with great potential applicable to clinical specimens. The authors undertook a prospective cohort study, as the first trial in the world up to our knowledge, to examine the relationship between the abMet of DLX4 and SIM1 genes and progression of LSIL. Methods: A total of 113 patients were selected from the Comparative Study on Conventional Pap smear and Liquid-based Cytology (ThinPrep) for Uterine Cervix (CCLBC), in which 11,039 samples were enrolled between October 2007 and March 2010 to compare the cytological features of conventional Pap smears and liquid-based cytology specimens using the ThinPrep method. They were classified into 4 groups according to their cervical cytology, hrHPV infection and follow up. Cytology samples were examined for abMet of DLX4 and SIM1 genes and their protein expressions. CaSki cells were treated with 5-Aza-2´-deoxycytidine (5-aza-dC). Results: 40 samples in group 1 were negative for intraepithelial lesion or malignancy. 21 LSILs in group 2 showed a continuance of LSIL for longer than 365 days, and 12 LSILs in group 3 showed an up-grading to high-grade (H) SIL+ within 365 days after the diagnosis of LSIL. 40 in group 4 were squamous cell carcinoma. All but group 1 were infected with hrHPV. Significant difference existed in frequency of abMet between groups 2 and 3 (p=0.044), between groups 3 and 4 (p=0.020) for DLX4, and between groups 1 and 3 (p=0.0003), as well as between groups 2 and 3 (p=0.005) for SIM1 gene. DLX4 protein expression was significantly reduced in the DLX4 abMet positive tissues, as compared to the negative tissues (p=0.008), and 5-aza-dC treatment extracted DLX4 protein expression of CaSki cells in a dose-dependent manner (p