Breast cancer is the most common cancer among women worldwide. Although the five-, ten- and fifteen-year survival rates are good for breast cancer patients diagnosed with early-stage disease, some cancers recur many years after completion of primary therapy. Tumor heterogeneity and clonal evolution may lead to distant metastasis and therapy resistance, which are the main causes of breast cancer-associated deaths. In the clinic today, imaging techniques like mammography and tissue biopsies are used to diagnose breast cancer. Even though these methods are important in primary diagnosis, they have limitations when it comes to longitudinal monitoring of residual disease after treatment, disease progression, therapy responses, and disease recurrence. Over the last few years, there has been an increasing interest in the diagnostic, prognostic, and predictive potential of circulating cancer-derived material acquired through liquid biopsies in breast cancer. Thanks to the development of sensitive devices and platforms, a variety of tumor-derived material, including circulating cancer cells (CTCs), circulating DNA (ctDNA), and biomolecules encapsulated in extracellular vesicles, can now be extracted and analyzed from body fluids. Here we will review the most recent studies on breast cancer, demonstrating the clinical potential and utility of CTCs and ctDNA. We will also review literature illustrating the potential of circulating exosomal RNA and proteins as future biomarkers in breast cancer. Finally, we will discuss some of the advantages and limitations of liquid biopsies and the future perspectives of this field in breast cancer management.

The long noncoding RNA (lncRNA) NEAT1 (nuclear enriched abundant transcript 1) is the architectural component of nuclear paraspeckles, and it has recently gained considerable attention as it is abnormally expressed in pathological conditions such as cancer and neurodegenerative diseases. NEAT1 and paraspeckle formation are increased in cells upon exposure to a variety of environmental stressors and believed to play an important role in cell survival. The present study was undertaken to further investigate the role of NEAT1 in cellular stress response pathways. We show that NEAT1 is a novel target gene of heat shock transcription factor 1 (HSF1) and is up-regulated when the heat shock response pathway is activated by sulforaphane (SFN) or elevated temperature. HSF1 binds specifically to a newly identified conserved heat shock element in the NEAT1 promoter. In line with this, SFN induced the formation of NEAT1-containing paraspeckles via an HSF1-dependent mechanism. HSF1 plays a key role in the cellular response to proteotoxic stress by promoting the expression of a series of genes, including those encoding molecular chaperones. We have found that the expression of HSP70, HSP90, and HSP27 is amplified and sustained during heat shock in NEAT1-depleted cells compared with control cells, indicating that NEAT1 feeds back via an unknown mechanism to regulate HSF1 activity. This interrelationship is potentially significant in human diseases such as cancer and neurodegenerative disorders.

The long non-coding RNA NEAT1 locus is transcribed into two overlapping isoforms, NEAT1_1 and NEAT1_2, of which the latter is essential for the assembly of nuclear paraspeckles. NEAT1 is abnormally expressed in a wide variety of human cancers. Emerging evidence suggests that the two isoforms have distinct functions in gene expression regulation, and recently it was shown that NEAT1_2, but not NEAT1_1, expression predicts poor clinical outcome in cancer. Here, we report that NEAT1_2 expression correlates with HER2-positive breast cancers and high-grade disease. We provide evidence that NEAT1_1 and NEAT1_2 have distinct expression pattern among different intrinsic breast cancer subtypes. Finally, we show that NEAT1_2 expression and paraspeckle formation increase upon lactation in humans, confirming what has previously been demonstrated in mice.

Epithelial-mesenchymal transition (EMT) is a cellular plasticity program critical for embryonic development and tissue regeneration, and aberrant EMT is associated with disease including cancer. The high degree of plasticity in the mammary epithelium is reflected in extensive heterogeneity among breast cancers. Here, we have analyzed RNA-sequencing data from three different mammary epithelial cell line-derived EMT models and identified a robust mammary EMT gene expression signature that separates breast cancers into distinct subgroups. Most strikingly, the basal-like breast cancers form two subgroups displaying partial-EMT and post-EMT gene expression patterns. We present evidence that key EMT-associated transcription factors play distinct roles at different stages of EMT in mammary epithelial cells.

Paraspeckles are stress-induced nuclear RNA-protein condensates that assemble on the long non-coding RNA NEAT1. Their increased formation under certain cellular circumstances has gained growing interest due to their association with serious human diseases such as neurodegenerative disorders and cancer. The biological functions of paraspeckles still appear obscure, but increasing evidence suggests that they contribute to regulation of gene expression by recruiting specific proteins and RNA molecules. Here, we have characterized and compared two stress-enriched interactomes of the essential paraspeckle protein NONO in both wild type and paraspeckle-deficient NEAT1 knockout cells. We identified Hsp70 as part of stress-enriched NONO complexes in wild type, but not in NEAT1-depleted cells. We show that proteotoxic stress-induced paraspeckle formation and NEAT1 expression are strictly dependent on Hsp70 chaperone activity. Our data demonstrate that both NONO and Hsp70 transiently translocate to the nucleolus during heat shock and that paraspeckle formation during recovery follows Hsp70-dependent relocation of NONO from the nucleolus to the nucleoplasm. Taken together, we demonstrate an important role of Hsp70 in paraspeckle assembly and identify a possible link between the nuclear protein quality control system and paraspeckles.