@article{e3bb7280596c474fac0e6fc2541051a6,
title = "Association of SOD3 promoter DNA methylation with its down-regulation in breast carcinomas",
abstract = "Superoxide dismutase 3 (SOD3) is a secreted antioxidant enzyme that regulates reactive oxygen species in the microenvironment. It is also a potential tumour suppressor gene that is significantly downregulated in breast cancer. We have previously shown that its mRNA expression is inversely correlated with relapse free survival in breast cancer patients. This study aimed to investigate the correlation of SOD3 promoter DNA methylation with its expression in different molecular subtypes of breast carcinoma. We found that SOD3 expression was significantly reduced in breast carcinoma samples compared to normal tissues with the lowest levels observed in Luminal B subtype. Pyrosequencing analysis showed significant increase in methylation levels in the SOD3 promoter region (−108 and −19 from the TSS) in tumours vs normal tissues (53.6% vs 25.2%). The highest degree of correlation between methylation and SOD3 expression levels was observed in Luminal B subtype (Spearman{\textquoteright}s R = −0.540, P < 0.00093). In this subtype, the −78 CpG position is the most significantly methylated site. The Spearman{\textquoteright}s coefficient analysis also indicated the most significant correlation of DNA methylation at this site with SOD3 gene expression levels in tumours vs. normal tissues (R = −0.5816, P < 6.9E-12). Moreover, copy number variation analysis of TCGA database revealed that the more aggressive Triple Negative and Her2+ subtypes had higher levels of SOD3 gene deletion. The predominantly down-regulated expression pattern of SOD3 and the various genetic and epigenetic deregulations of its expression suggest that loss of this antioxidant promotes an advantageous tumour-promoting microenvironment in breast cancer.",
keywords = "DNA methylation, SOD3, breast cancer, extracellular superoxide dismutase, pyrosequencing",
author = "Brandon Griess and David Klinkebiel and Alice Kueh and Michelle Desler and Kenneth Cowan and Matthew Fitzgerald and Melissa Teoh-Fitzgerald",
note = "Funding Information: Excess DNA samples collected through the Breast Cancer Collaborative Registry (BCCR) (IRB# 253–13 EP, PI: Dr. Kenneth Cowan) were used for pyrosequencing analysis. BCCR is a web-based biomedical data and biospecimen repository developed by the Fred & Pamela Buffett Cancer Centre. The BCCR provides a critical platform for the Northern Great Plains Personalized Breast Cancer Program (NGPPBCP) funded by The Leona M. and Harry B. Helmsley Charitable Trust. Seventy one locations across the U.S. including seven cancer centres in the Northern Great Plains that are actively enrolling patients on the BCCR. Formalin fixed tissue blocks collected from the cancer centres were sent to the Fred & Pamela Buffett Cancer Centres (FPBCC) for centralized review. Specimens that are deemed adequate for whole exome DNA sequence studies were then sent to the tissue facility at the FPBCC for sectioning. Trained tissue technologist then performed macro-dissection on each specimen to concentrate the number of tumour cells and reduce the contamination of adjacent normal breast tissue. DNA isolation from the FFPE tumour specimens was performed by the FPBCC{\textquoteright}s Molecular Biology/High-Throughput Screening Facility. Normal tissues used are not the adjacent tissues of tumour samples, but are breast tissues isolated from normal patients. DNA was extracted and purified using QIAamp DNA FFPE Tissue (QIAGEN) kits, quantified by Nanodrop 2000, followed by double-stranded DNA assessment using Qubit (3.0) dsDNA HS Assay kit (Invitrogen). Following the extraction of DNA from each patient{\textquoteright}s breast cancer, the quality and quantity of DNA in each sample is determined. RNAs were extracted by using the QIAGEN RNeasy FFPE kit. Funding Information: The Northern Great Plains Personalized Breast Cancer Program (NGPPBCP) was funded by The Leona M. and Harry B. Helmsley Charitable Trust. This work was financially supported by grants from the NIH R0I-CA182086A (Teoh-Fitzgerald). Brandon Griess was supported by the Eppley Institute in Cancer Biology Training Grant (NCI T32CA009476). We thank Oleg Shats (Assistant Director for Cancer Informatics at UNMC), Dr. David Kelly (Director, Molecular Biology Core Facility, Eppley Cancer Institute, UNMC) for clinical data verification. We also acknowledge the contribution by Dr. Peng Xiao (Director, Bioinformatics and Systems Biology Core, UNMC) for analyzing the Agendia microarray data and providing bioinformatics supports. We thank the Epigenomic Core Facility and the director, Dr. David Klinkebiel at UNMC for performing the pyrosequencing analysis. In addition, we thank Agendia Inc. USA for performing the gene array expression profiling. Publisher Copyright: {\textcopyright} 2020 Informa UK Limited, trading as Taylor & Francis Group.",
year = "2020",
month = dec,
doi = "10.1080/15592294.2020.1777666",
language = "English (US)",
volume = "15",
pages = "1325--1335",
journal = "Epigenetics",
issn = "1559-2294",
publisher = "Landes Bioscience",
number = "12",
}