Studies on the role of two proteins in breast cancer - histone deacetylase 11 in estrogen receptor positive breast cancer and the redox protein memo in metastasis and tumorigenesis

Abstract

Breast cancer is the most diagnosed cancer type in women worldwide. It is a heterogeneous disease that can progress and metastasize into distant organs. After lung cancer, breast cancer is the second leading cause of cancer death among women in developed countries. However, death rates are declining since 1990, mainly due to earlier detection and improved therapies. The development of therapies specifically targeting known oncogenes have improved patient survival. For example the estrogen receptor (ER), a hormone receptor expressed in 70% of breast cancer and the receptor tyrosine kinase ErbB2, which is expressed in 20% of breast cancer, are treated by endocrine therapies and targeted antibodies or kinase inhibitors, respectively. Despite many successes, not all patients respond to these treatments or they develop resistance leading to cancer recurrence. The treatment of metastases remains the biggest challenge. Therefore, new therapies are required and this demands a better understanding of disease development and progression. With the two studies presented here, we aimed to gain more insight in the role of histone deacetylase 11 (HDAC11) in breast cancer, and in the function of Memo (Mediator of ErbB2-driven cell motility), in tumorigenesis and metastasis. Results from several preclinical studies suggest that the inhibition of HDACs is a promising approach to target cancer. However, the role of individual HDACs in breast cancer is largely unknown. In our study, we aimed to find HDACs with an oncogenic role in breast cancer. By screening available online databases, we discovered HDAC11 to be overexpressed in breast cancer compared to normal breast. Furthermore, we found a correlation between HDAC11 expression and ER levels in breast cancer, and could show that HDAC11 expression is induced by estrogen-signaling. Moreover, a knockdown (KD) approach revealed that HDAC11 promotes proliferation in the ER-positive breast cancer cell lines T47D and MCF7, but does not significantly alter proliferation of the ER-negative breast cancer cell lines MDA-MB-231 and SUM159. KD of HDAC11 in the ER-positive murine breast cancer cell line J110 decreased tumor growth in vivo. In addition, RNA sequencing combined with gene set enrichment analyses (GSEA) revealed that HDAC11 controls part of the estrogen responsive transcriptional program, especially genes that are normally downregulated upon estrogen induced signaling. This suggests that HDAC11 is an estrogen induced repressor of gene transcription. Furthermore, Kaplan-Meier analysis revealed that HDAC11 has prognostic value for patients who received endocrine therapy, as the probability of recurrence-free survival (RFS) and distant metastasis-free survival (DMFS) decreased significantly for patients having tumors with high levels of HDAC11. In summary, we show that HDAC11 has oncogenic potential in ER-positive breast cancer and therefore might be a new target for therapy. Memo was initially found through its interaction with ErbB2. Several studies have demonstrated that Memo is required for breast cancer cell migration in response to ligands activating receptor tyrosine kinases (RTKs). In this study we aimed to investigate if Memo is involved in breast cancer metastasis and tumorigenesis. Recently, we discovered that Memo is a copper-dependent redox enzyme, which promotes spontaneous metastasis from a xenograft breast cancer model. By tissue microarray analysis of primary human breast tumors we found Memo to be overexpressed in 40% of breast cancer, while expressed at low levels in the normal human breast. Moreover, high cytoplasmic Memo localization has prognostic value for early distant metastasis and death. However, primary tumor growth was not affected by Memo, neither in a xenograft model using the triple-negative human breast cancer cell line MDA-MB-231, nor in a constitutively active ErbB2 (NeuNT)-driven murine spontaneous breast cancer model. Immunohistochemistry of normal murine mammary glands revealed that Memo is expressed throughout all stages of development, specifically in the luminal cell compartment. To study the role of Memo in tumorigenesis, we generated a spontaneous NeuNT-driven murine breast cancer cell model, in which Memo excision and NeuNT expression should be achieved at the same time. Mice from all strains developed tumors with the same frequency. However, they were all positive for Memo expression, suggesting that either Memo-depleted mammary epithelial cells were outcompeted by Memo expressing cells and/or only Memo-positive cells can give rise to tumors, or that Cre-recombinase mediated excision of Memo was not functional. In summary, this study revealed that Memo is a copper-dependent redox enzyme that promotes breast cancer metastasis.