Polycomb complex protein BMI1 confers resistance to tamoxifen in estrogen receptor positive breast cancer

Highlights

• BMI1 promotes tamoxifen (TAM) resistance in ER-positive breast cancers.

• BMI1 upregulates the transcription activity of estrogen receptor (ER).

• BMI1 increases the expression of MUC1 and androgen receptor in breast cancer.

• BMI1 upregulation is associated with decreases in immunosurveillance of breast cancer.

• Breast cancers with high levels of BMI1 are at risk of developing TAM resistance.

Abstract

We report that BMI1 promotes tamoxifen resistance in estrogen receptor (ER)-positive breast cancer (BC). BMI1 overexpression conferred MCF7 and TD47 cells resistance to tamoxifen; BMI1 knockdown sensitized the process. In MCF7-derived tamoxifen resistant cells, BMI1 expression was upregulated and BMI1 knockdown reduced the resistance. BMI1 is an oncogene; its oncogenic activity is attributed to BMI1-stimulated E3 ubiquitin ligase activity, a process that requires BMI1's ring finger (RF) domain. However, a BMI1 mutant without RF conferred tamoxifen resistance. Tamoxifen significantly reduced the growth of xenografts derived from MCF7 cells, but accelerated the growth of tumors produced by BMI1 overexpressing MCF7 cells. BMI1 enhances the pathways that promote resistance to endocrine therapy, including ER, androgen receptor, and MUC1. In patients with ER + BCs (n = 177), BMI1 expression was associated with BC recurrence. In the Curtis dataset consisting of ER + BCs (n = 1506) and ER- BCs (n = 474; cBioPortal), upregulations in BMI1 mRNA expression were correlated with ER + BCs; the upregulation was associated with a set of differentially expressed genes (DEGs). These DEGs were enriched with reductions in immunological processes, indicating a role of BMI1 in downregulation of the immune surveillance.

Introduction

Breast cancer (BC) is the most common female malignancy and the second leading cause of cancer death in women worldwide; approximately 1.7 million women are diagnosed annually and the disease results in 500,000 deaths per year [20]. BC is a heterogeneous disease, consisting of different histologic subtypes that are classified according to the expression status of estrogen receptor (ER), progesterone receptor, and HER2. Approximately 75% of BCs are ER+ [43]. ER signaling is critical for BC progression in part through upregulation of Myc, cyclin D1, and vascular endothelial growth factor [41]. Therapeutically targeting ER is the first option for patients with ER + BC. Endocrine therapy blocks ER signaling through the use of selective estrogen receptor modulators (SERMs: tamoxifen, raloxifene, arzoxifen, and lasofoxifene), estrogen biosynthesis inhibitors (aromatase inhibitors/AIs: anastrozole, letrozole, and exemestane), and selective estrogen receptor downregulators (SERDs: fulvestrant and ICI 164,384) [42,47]. Among these three groups of anti-estrogen drugs, tamoxifen (TAM) was first used in the clinics over 40 years ago, and remains an important drug for adjuvant therapy particularly for premenopausal women [1,43]. Furthermore, recent clinical trials supported TAM treatment for 10 years for patients with early stage ER + BC [14]. TAM in combination with ovarian suppression improved disease free survival [8,21]. Nevertheless, approximately 30% of patients treated with adjuvant TAM develop resistance [42]. Despite the steady advance in our understanding of the factors contributing to TAM resistance, our current knowledge of these mechanisms remains incomplete.

BMI1 is a well-established oncogene that promotes tumorigenesis in part through maintaining the self-renewal of stem cells and cancer stem cells. BMI1 is a polycomb group protein of the polycomb repressive complex 1 (PRC1) [34], and contributes to the E3 ubiquitin ligase activity of PRC1 via binding to the catalytic subunit RING2 or RING1B [9,16,35,54]. The interaction is mediated through BMI1's ring finger (RF) domain [9,16,35,48,54]. The E3 ligase activity is an important attribute of BMI1-derived oncogenic activities. BMI1 suppresses the INK4A-ARF locus [7,40], which encodes two tumor suppressors p16^INK4A and p19/p14^ARF. The process requires BMI1-associated E3 ubiquitin ligase activity. In mice deficient in BMI1, hematopoietic stem cells, neural stem cells, and intestinal cancer tumorigenesis are all impaired; these effects are reversed by co-knockout of the INK4A-ARF locus [7,39,40]. Upregulation of BMI1 transformed lymphocytes [2], and was detected in non-small cell lung cancer [52], prostate cancer [19], colon cancer [32], nasopharyngeal carcinoma [50], brain cancer [29], cervical cancer, and ovarian cancer [3].

BMI1 plays a role in BC tumorigenesis. BMI1 is upregulated in BC. The upregulation is correlated with c-Myc expression [49] and ER + BC [18,53]. BMI1 expression is associated with poor prognosis [4,6]. A BMI1-derived 11-gene signature displayed stem cell features and was associated with reductions in overall survival in patients with early BC [25]. In line with this clinical evidence, BMI1 sustains BC stem cells [11,44,51]. Overexpression of BMI1 immortalizes human mammary epithelial cells [13,17]. Collectively, there is a large body of evidence supporting an important role of BMI1 in BC tumorigenesis and development. Nevertheless, whether BMI1 contributes to resistance to endocrine therapy remains unclear.

We present evidence supporting a direct involvement of BMI1 in the development of TAM resistance. BMI1 confers resistance to TAM-derived cytotoxicity in vitro and in vivo. Intriguingly, the E3 ubiquitin ligase activity can be dispensable. BMI1 facilitates ER-derived transcription activity and upregulates androgen receptor (AR) and MUC1. BMI1 upregulation in primary ER + BC may lead to downregulation of immune surveillance.