Hypoxia-induced epigenetic regulation of breast cancer progression and the tumour microenvironment.

The events that control breast cancer progression and metastasis are complex and intertwined. Hypoxia plays a key role both in oncogenic transformation and in fueling the metastatic potential of breast cancer cells. Here we review the impact of hypoxia on epigenetic regulation of breast cancer, by interfering with multiple aspects of the tumour microenvironment. The co-dependent relationship between oxygen depletion and metabolic shift to aerobic glycolysis impacts on a range of enzymes and metabolites available in the cell, promoting posttranslational modifications of histones and chromatin, and changing the gene expression landscape to facilitate tumour development. Hormone signalling, particularly through ERα, is also tightly regulated by hypoxic exposure, with HIF-1α expression being a prognostic marker for therapeutic resistance in ER+ breast cancers. This highlights the strong need to understand the hypoxia-endocrine signalling axis and exploit it as a therapeutic target. Furthermore, hypoxia has been shown to enhance metastasis in TNBC cells, as well as promoting resistance to taxanes, radiotherapy and even immunotherapy through microRNA regulation and changes in histone packaging. Finally, several other mediators of the hypoxic response are discussed. We highlight a link between ionic dysregulation and hypoxia signalling, indicating a potential connection between HIF-1α and tumoural Na+ accumulation which would be worth further exploration; we present the role of Ca2+ in mediating hypoxic adaptation via chromatin remodelling, transcription factor recruitment and changes in signalling pathways; and we briefly summarise some of the findings regarding vesicle secretion and paracrine induced epigenetic reprogramming upon hypoxic exposure in breast cancer. By summarising these observations, this article highlights the heterogeneity of breast cancers, presenting a series of pathways with potential for therapeutic applications.

ZMIZ1 enhances ERα-dependent expression of E2F2 in breast cancer.

The estrogen receptor-α (ER) drives 75% of breast cancers. On activation, the ER recruits and assembles a 1-2 MDa transcriptionally active complex. These complexes can modulate tumour growth, and understanding the roles of individual proteins within these complexes can help identify new therapeutic targets. Here, we present the discovery of ER and ZMIZ1 within the same multi-protein assembly by quantitative proteomics, and validated by proximity ligation assay. We characterise ZMIZ1 function by demonstrating a significant decrease in the proliferation of ER-positive cancer cell lines. To establish a role for the ER-ZMIZ1 interaction, we measured the transcriptional changes in the estrogen response post-ZMIZ1 knockdown using an RNA-seq time-course over 24 h. Gene set enrichment analysis of the ZMIZ1-knockdown data identified a specific delay in the response of estradiol-induced cell cycle genes. Integration of ENCODE data with our RNA-seq results identified that ER and ZMIZ1 both bind the promoter of E2F2. We therefore propose that ER and ZMIZ1 interact to enable the efficient estrogenic response at subset of cell cycle genes via a novel ZMIZ1-ER-E2F2 signalling axis. Finally, we show that high ZMIZ1 expression is predictive of worse patient outcome, ER and ZMIZ1 are co-expressed in breast cancer patients in TCGA and METABRIC, and the proteins are co-localised within the nuclei of tumour cell in patient biopsies. In conclusion, we establish that ZMIZ1 is a regulator of the estrogenic cell cycle response and provide evidence of the biological importance of the ER-ZMIZ1 interaction in ER-positive patient tumours, supporting potential clinical relevance.

A single-cell atlas enables mapping of homeostatic cellular shifts in the adult human breast.

Here we use single-cell RNA sequencing to compile a human breast cell atlas assembled from 55 donors that had undergone reduction mammoplasties or risk reduction mastectomies. From more than 800,000 cells we identified 41 cell subclusters across the epithelial, immune and stromal compartments. The contribution of these different clusters varied according to the natural history of the tissue. Age, parity and germline mutations, known to modulate the risk of developing breast cancer, affected the homeostatic cellular state of the breast in different ways. We found that immune cells from BRCA1 or BRCA2 carriers had a distinct gene expression signature indicative of potential immune exhaustion, which was validated by immunohistochemistry. This suggests that immune-escape mechanisms could manifest in non-cancerous tissues very early during tumor initiation. This atlas is a rich resource that can be used to inform novel approaches for early detection and prevention of breast cancer.

Transcriptional changes in the mammary gland during lactation revealed by single cell sequencing of cells from human milk.

Under normal conditions, the most significant expansion and differentiation of the adult mammary gland occurs in response to systemic reproductive hormones during pregnancy and lactation to enable milk synthesis and secretion to sustain the offspring. However, human mammary tissue remodelling that takes place during pregnancy and lactation remains poorly understood due to the challenge of acquiring samples. We report here single-cell transcriptomic analysis of 110,744 viable breast cells isolated from human milk or non-lactating breast tissue, isolated from nine and seven donors, respectively. We found that human milk largely contains epithelial cells belonging to the luminal lineage and a repertoire of immune cells. Further transcriptomic analysis of the milk cells identified two distinct secretory cell types that shared similarities with luminal progenitors, but no populations comparable to hormone-responsive cells. Taken together, our data offers a reference map and a window into the cellular dynamics that occur during human lactation and may provide further insights on the interplay between pregnancy, lactation and breast cancer.