High RASSF1A promoter methylation levels are predictive of poor prognosis in fine-needle aspirate washings of breast cancer lesions

Abstract

Previously, we reported that the accuracy of cytological diagnosis of breast lesions could be augmented through the quantitative assessment of DNA methylation of fine-needle aspirate (FNA) washings. Herein, we aimed at the evaluation of the prognostic value of quantitative promoter methylation at three gene loci (APC, CCND2, and RASSF1A) in a large series of FNA washings from breast lesions. Methylation levels of three gene promoters were assessed by quantitative methylation-specific PCR in bisulfite-modified DNA from 211 FNA washings, comprising 178 carcinomas and 33 benign lesions, both histopathologically confirmed. Receiver operator characteristic (ROC) curve analysis was used to determine the diagnostic performance of the gene panel in distinguishing cancer from non-cancerous lesions. Relevant clinicopathologic data and time to progression and/or death from breast cancer were correlated with methylation findings. Log-rank test and Cox-regression model identified independent predictors of prognosis. APC, CCND2, and RASSF1A methylation levels differed significantly between malignant and benign lesions. ROC curve analysis confirmed the diagnostic performance of the gene panel. In univariate analysis, stage was significantly associated with overall, disease-specific and disease-free survival, whereas tumor grade was associated with disease-specific and disease-free survival. Remarkably, RASSF1A methylation was significantly and independently associated with worse disease-free survival in the final multivariate analysis. We confirmed that quantitative gene promoter methylation augments the diagnostic performance of cytopathology. Importantly, and in addition to standard clinicopathologic parameters, RASSF1A high-methylation levels are independent predictors of worse outcome in breast cancer. Thus, epigenetic biomarkers provide valuable tools for breast cancer patient management.

A RASSF1A Polymorphism Restricts p53/p73 Activation and Associates with Poor Survival and Accelerated Age of Onset of Soft Tissue Sarcoma

“…Taken together, these observations support the idea that RASSF1 c.397G>T results in the loss or impairment of a major tumor suppressor pathway in cells (4,7,10,11,24). In survival studies of STS patients, the minor allele associated with poorer survival with a significant RR of tumor-related death (50.5 vs. 100.9 months, P ¼ 0.016; RR ¼ 4.82, P ¼ 0.034, Table 1) in males.…”

“…Interestingly, we and others have previously shown that inactivation of RASSF1A signaling by methylation associates with poorer survival in a number of tumor types including STS (7)(8)(9)(10)(11)(12). In this article, we go on to provide evidence in a cohort of STS patients that this association is sex specific, whereby hypermethylation of the RASSF1A promoter in tumors only associates with poorer survival in males.…”

“…Although LOH has been observed, RASSF1A is more frequently epigenetically inactivated by hypermethylation of CpG islands within the promoter and the first exon, limiting expression in a wide variety of human cancers (3)(4)(5)(6). High promoter methylation levels (implying low RASSF1A mRNA and protein levels) have been shown to associate with poor prognosis and are thus considered an independent prognostic indicator of overall survival in lung, renal, and breast cancers as well as in uveal melanoma, hepatoblastoma, and sarcoma (7)(8)(9)(10)(11)(12). Interestingly, lower levels of RASSF1A transcript have also been correlated with the decreased radiochemosensitivity of hepatoblastoma and colorectal cell lines (7,13).…”

RASSF tumor suppressor gene family: Biological functions and regulation

“…Epigenetic loss of RASSF1A has been observed in urine [138][139][140], nipple aspirates [141,142], and sputum [143] to mention a few. Thus, there is potential in utilizing epigenetic loss of RASSF1A as a diagnostic/prognostic tool for treating cancer and potentially other diseases as well.…”

Breast: Ductal carcinoma

“…The most important isoform for breast cancer seems to be isoform A. The frequency of RASSF1A methylation in breast tumors is high (65%) and methylation in tumor samples and fine-needle aspirate washings was shown to be an independent predictor of poor prognosis (Sharma et al, 2009;Buhmeida et al, 2011;Martins et al, 2011). RASSF1A methylation is an attractive biomarker for early cancer detection and its methylation analysis is applicable to a range of body fluids including serum and nipple fluid (Suijkerbuijk et al, 2008).…”