Abstract 3303: Investigation of SPAG5 gene expression as a molecular marker for breast cancer prognosis

Breast cancer is a clinically heterogeneous disease, making it imperative to use biomarkers, such as the estrogen receptor (ER) and human epidermal growth factor 2 (HER2), to stratify patients into specific prognostic groups. Biomarker stratification provides patients with more accurate diagnoses, predictions of clinical outcomes, and offers targeted therapeutic options. SPAG5 (Sperm Associated Antigen 5) is involved in the maintenance of spindle-pole integrity, efficient chromosomal alignment, and cell proliferation. Its role in cell division may be a cancer susceptibility factor. Low expression of SPAG5 has been correlated with good prognosis and absence of metastases in ER+ breast cancer. However, its relationship to other molecular subtypes of breast cancer and its functional role in the disease are relatively unknown. The purpose of this study was to investigate the role of SPAG5 in breast cancer through (i) microarray analysis and (ii) correlating its expression with invasiveness, a factor involved in disease progression. In part (i), SPAG5 expression in breast cancers was evaluated by using five Affymetrix, three Agilent and one Illumina microarray datasets. The associations between SPAG5 expression and various tumor pathologies were analyzed via log2 expression ratios. Pathologies that were investigated included the ER+ subtype, HER2 overexpression, lymph node metastasis, and breast cancer stem cell-like/undifferentiated tumors. In part (ii), SPAG5 expression across five breast cancer epithelial cell lines of varying invasive ability (MDA-MB-231, MDA-MB-157, BT549, MCF-7, and CAMA1) was investigated. The cells were cultured at normal conditions and lysed using the CytoBusterTM protein extraction reagent. The supernatants were collected by microcentrifugation at 16,000 rcf at 4°C for 5 minutes. Western blotting was completed using standard techniques. From the microarray analysis, overexpression of SPAG5 showed strong associations with HER2 overexpression, ER-negative and triple-negative cancers, high grade tumors, metastasis, and poor clinical outcomes in a total of 1595 breast cancers. Via immunoblotting, it was shown that higher levels of SPAG5 accompanied highly invasive cell lines (MDA-MB-231, MDA-MB-157, and BT549) and lower levels occurred in weakly invasive cells (MCF-7 and CAMA1). These results suggest that SPAG5 is a biomarker of malignancy and invasiveness and may be a novel therapeutic target for the prevention of metastasis. Stratificat