Immunohistochemical Validation of Topoisomerase 2 alpha and BubR1 as Novel Breast Cancer Risk Genes in Benign Breast Tissue
A Nasir, DT Chen, AC Culhane, M Gruidl, SM McCarthy, C Venkataramu, J Quackenbush, TJ Yeatman. Moffitt Cancer Center, Tampa, FL; Dana-Farber Cancer Institute, Harvard Medical School, Boston, MA
Background: In order to predict the risk of malignancy in histologically normal breast, we have developed a novel 32-gene malignancy-risk gene signature that was validated by real-time PCR. Here we report further validation of two of our top malignancy-risk genes, topoisomerase 2 alpha (TOP2A) and BubR1 at protein level.
Design: Using our novel malignancy-risk gene signature, we stratified 18 histologically normal breast (HNB) tissues as high-risk normals (HRNs, n=9) or low-risk normals (LRNs, n=9). All HRNs and LRNs were from peri-menopausal female patients (mean ages: 52 and 51 years respectively). All archival sections from mastectomies of these patients were reviewed to select blocks with maximum numbers of histologically-normal terminal duct lobule units (TDLUs). Control set: Six invasive ductal carcinomas (IDC). 5 thick serial sections were stained for TOP2A and BubR1 proteins, using standard immunohistochemical (IHC) protocols. TOP2A staining index (% of stained cells) and BubR1 H-scores based on the product of positively stained cells (%) and immunointensity (0/1+/2+/3+) were all specimen types (TDLUs present in the HRN and LRN samples and the selected IDCs).
Results: TOP2A immunstain was nuclear; BubR1 was cytoplasmic. The immunoreactivity in the tumor cells and in the benign acinar and ductal epithelium of all of the TDLUs was scored. There was significantly higher immunoreactivity for TOP2A (t. test, P < 0.01) and BUBR1 (t. test, P < 0.001) in histologically normal breast tissues, which had been predicted to be at high risk of malignancy using our 32 gene malignancy-risk signature.
Table: TOP2A and BubR1 in IDCs, high-risk and low-risk normal breast tissues.
|Archival specimen type||Average # of TDLUs evaluated/specimen (Range)||Mean TOP2A index (%) (Range)||Mean BubR1 expression score (H-score) (Range)|
|IDC (N=6)||Not applicable||26.7 (15-35)||149.0 (80-200)|
|High-risk normal breast tissue||33 (19-39)||11.4 (2-30)||64.4 (33-113)|
|Low-risk normal breast tissue||24 (17-35)||1.8 (1-3)||16.6 (10-22)|
Conclusions: Topoisomerase 2 alpha and BubR1 are novel breast cancer risk genes that can be used to determine risk of malignancy in benign breast tissue. We are further investigating the clinical utility of these markers on larger cohorts of benign breast tissues.
Monday, March 9, 2009 2:00 PM
Platform Session: Section B, Monday Afternoon