Detection of RASSF1A and RAR? Hypermethylation in Serum DNA from Breast Cancer Patients

Abstract

ABBREVIATIONS RASSF1ARAS association domain family protein 1A RARβ retionic acid receptor β MSP methylation specific PCR ERα estrogen receptor α PRB progesterone receptor B ACKNOWLEDGEMENTSWe are grateful to Department of Biotechnology, India, for providing the financial support to conduct this study. S.M. is grateful to University Grant Commission for award of Junior Research Fellowship. The authors thank Dr. Cecile Rochette Egly for the kind gift of RARβ monoclonal antibody. Research PaperDetection of RASSF1A and RARβ Hypermethylation in Serum DNA from Breast Cancer Patients ABSTRACTBreast cancer is fast emerging as the leading cancer amongst females, especially in young females in metropolitan cities in India. The epigenetic alterations involved in the onset and progression of breast cancer may serve as biomarkers for early detection and prognosis of the disease. Furthermore, using body fluids such as serum offers a noninvasive method to procure multiple samples for such analyses. In this study, we examined methylation status of two normally unmethylated but biologically significant cancer genes, RAS association domain family protein 1A (RASSF1A) and retionic acid receptor β (RARβ) by methylation specific PCR (MSP) in invasive ductal carcinomas of the breast and paired serum DNA. RASSF1A was found to be methylated in 17 of 20 (85%) breast tumors; while sera from 15 of 20 (75%) of the patients showed concordant methylated RASSF1A, with a sensitivity of 88%. RARβ was methylated in 2/20 (10%) breast tumors. A gene unmethylated in the tumor DNA was always found to be unmethylated in the matched serum DNA for both RASSF1A and RARβ genes; hence specificity was 100%. Immunohistochemical analysis of RARβ protein in 15 breast carcinoma patients harboring unmethylated RARβ in tumors and serum DNA showed the expression of RARβ protein in tumors and paired normal breast tissues, confirming the MSP findings, suggesting that RARβ promoter is functional in these cases. This study underscores the potential utility of DNA methylation based screening of serum, a readily accessible body fluid, as a surrogate marker for early detection of breast cancer.

The heparan sulfate sulfotransferase 3-OST3A (HS3ST3A) is a novel tumor regulator and a prognostic marker in breast cancer

“…Our demonstration of the HS-sulfotransferase 3-OST3A gene methylation and histone modification is in line with previous findings indicating that epigenetic dysregulation is an important feature of breast cancer pathobiology and that it depends upon the tumor signature. 32 Our results validate the potential value of epigenetic therapies targeting 3-OST3A together with other silenced critical genes such as those coding for the Ras interacting protein RASSF1A 33 , the estrogen receptor α (ERα) 34 and the retinoic acid receptor RARα 35 in TN breast cancers. Altogether, our findings support the attractive idea that the HS-modifying 3-OST3A sulfotransferase enzyme participates ino the matrix-cell dialog through epigenetic mechanisms.…”

Genome-wide identification of OTP gene as a novel methylation marker of breast cancer

“…Recently, several groups, using a panel of multiple genes which are well known to be tumorassociated, have demonstrated the prevalence of methylation positive genes in sera or plasma from breast cancer patients (20)(21)(22)(23)(24)(25)(26)(27). Nevertheless, current multiple gene panel assays still lack the necessary sensitivity for application in the clinical setting.…”

Clinicopathological significance of RASSF1A reduced expression and hypermethylation in hepatocellular carcinoma

“…It has been suggested that RASSF1A methylation is one of the most common aberrations so far identified in human cancers and that the loss of the functional protein may promote the development of many human tumors. Hypermethylation of the RASSF1A has been detected frequently in the tissues of different cancers [10][11][12][13][14][15][16][17], while in the body fluid, methylation of RASSF1A promoter has also been documented in 50% sputum and 84% serum from lung cancer [30], 35% urine from bladder cancer [15], 56-60% serum from breast caner [31][32][33], and 5% serum from undifferentiated nasopharyngeal carcinoma [34], which indicated the value of RASSF1A methylation in DNA for the early-stage diagnosis of tumors. Detection of methylated DNA has then been suggested as a potential biomarker for early detection of cancer.…”