In this study we tested the hypothesis that EZH2 expression may be clinically useful in predicting prognosis in a challenging group of patients with invasive carcinomas smaller than 2 cm (T1) with negative lymph nodes (N0) or 1–3 positive nodes (N1) treated by standard of care between 1996 and 2002. There are few studies in the literature which focus on biomarker development in early stage breast cancer with limited clinical significance. The identification of patients with early stage biologically aggressive tumors has the potential to assist in patient management.
Based on a cohort of 480 early stage invasive carcinomas of the breast rigorously selected and histopathologically analyzed we made several novel observations. Consistent with its oncogenic function, EZH2 overexpression is associated with high histological grade (Nottingham grade 3) and tumor size, negative estrogen and progesterone receptors and with HER-2/neu overexpression. We discovered that EZH2 expression is higher in early stage breast carcinomas from women with first degree family history of breast and/or ovarian cancer, suggestive of inherited breast cancer susceptibility [25
]. Our finding that EZH2 overexpression is associated with familial breast cancers in early stage disease is especially relevant given previous studies supporting a mechanistic connection between EZH2 and hereditary breast cancer genes BRCA1 and TP53. Our laboratory has shown that EZH2 protein is upregulated in benign appearing lobules of prophylactic mastectomies from BRCA1 mutation carriers [23
]. In estrogen receptor negative invasive breast carcinomas, EZH2 overexpression is able to regulate BRCA1 expression and intracellular localization [10
]. The relationship between EZH2 and BRCA1 is likely complex and reciprocal as EZH2 overexpression was found in breast cancers arising in women with BRCA1 mutations [24
]. Pietersen and colleagues have reported that invasive breast carcinomas with high EZH2 protein harbor TP53 mutations and suggested that during tumorigenesis EZH2 overexpression coincides with TP53 activation [27
]. Our results highlight the relevance of these basic studies to human breast cancer and suggest the hypothesis that EZH2 overexpression may be associated with other molecular determinants of hereditary breast cancer in addition to BRCA1 and TP53 gene mutations, which warrant investigation.
Even though most women with early stage breast cancer in the sporadic or familial settings do not progress, distant metastases may occur long after the first 5 years following primary breast cancer treatment [1
]. A major challenge in the management of women with early stage breast cancer is the identification of those who will develop metastasis from those that will not. We tested the hypothesis that EZH2 overexpression in early stage primary breast carcinoma may predict long-term metastasis up to 15 years following diagnosis of the primary tumor. Indeed, EZH2 expression levels in primary breast carcinoma were associated with metastasis-specific survival in women with first degree family history at 5, 10 and up to 15 years after primary breast carcinoma diagnosis. The 15- year metastasis free survival was 83% and 94% for women with tumors exhibiting high and low EZH2 expression respectively (log rank test p=0.05). The best multivariable model predictive of metastasis-specific survival included high EZH2 expression, high histological grade, higher stage, and absence of anti-hormonal treatment, reflecting negative estrogen and progesterone receptor status. High EZH2 expression was a strong independent predictor of metastasis providing 15 year survival information above other independent prognostic features. These data are clinically relevant because EZH2 may identify women with early stage breast cancer and first degree family history at higher risk to develop distant metastasis. Furthermore our data support the potential clinical utility of incorporating EZH2 into clinical nomograms to help determine the risk of cancer progression in this specific group of patients.
A limitation of our analysis is that tumor tissue samples were not available for 30% of patients identified. However, given that these samples would have not been selected on EZH2 expression it is unlikely to have biased our results. Our study focused on very small tumors so the material resource was very limited. Despite this, we successfully utilized 90% of the available tissues in our analysis. Our findings were focused on the very small group of women in our cohort (n=112 out of 480) who had first degree family history of breast and/or ovarian cancer. Therefore, some of our results included large confidence intervals. Nonetheless, we were able to demonstrate a significant association of EZH2 with distant metastasis even after controlling for important covariates, which strengthens our results.
Since our initial reports of EZH2 overexpression in breast cancer our findings have been supported by other investigations. EZH2 overexpression has been found in breast and other solid tumors including bladder [28
], lung [32
], melanoma [17
], and hepatocellular carcinoma [33
]. In these malignancies, EZH2 has been implicated in neoplastic transformation, progression, invasion and metastasis. Taken together, these data suggest that EZH2 may be involved in a global, rather than a tissue type specific mechanism of tumor progression.
In conclusion, our results indicate that in early stage breast cancer, EZH2 overexpression is associated with features of aggressive disease. We discovered a novel association between EZH2 overexpression and first degree family history of breast and/or ovarian cancer, which paves the way to mechanistic and functional studies. Our finding that EZH2 overexpression is an indicator of metastasis and metastasis-related deaths in women with early stage breast cancer and first degree family history may have important clinical implications. Specifically, EZH2 detection at the time of primary tumor diagnosis may aid clinicians in guiding management decisions and lays the foundation for the development of targeted treatments.