Triple-negative breast cancer (TNBC) is a major subtype of the disease characterized by the lack of estrogen receptor (ER), progesterone receptor (PR) expression, and human epidermal growth factor receptor 2 (HER2) activation. TNBCs have a high rate of recurrence to distant metastatic sites, and TNBC is currently the only major subtype of breast cancer that lacks targeted therapies. Largely due to this, unfortunately a significant fraction of patients diagnosed with TNBC die of their disease within 5 years of diagnosis. Thus, new, more effective treatment options are strongly needed.

Cancer is a genetic disease initiated and driven by changes in the DNA sequence (i.e., mutation) of genes with key roles in the regulation of cell growth and death. As these mutations are specific for the cancer cells and the cells are dependent on them for their survival, therapeutic inhibition of such mutated genes has been one of the most successful approaches for cancer treatment. In the past decade, systematic sequencing of putative regulators of tumorigenesis has revealed several previously unknown but frequently mutated genes in breast and other cancer types. However, subsequent large-scale sequencing of breast cancer genomes in the past few years has somewhat disappointed initial expectations and identified relatively few recurrently mutated genes that could be explored for therapy. This is especially the case in TNBCs where, aside from the already known cancer genes, very few new commonly mutated genes were found. These findings led to the sobering notion that we probably already know most if not all genes mutated in breast cancer. Thus, the identification of novel therapeutic targets in TNBCs requires other approaches.

Each cell in our body has nearly identical DNA sequence, yet the cells have very different properties and functions and these are stably maintained throughout life. This stable and heritable cellular identity is maintained by epigenetic programs that do not involve DNA sequence changes but include modification of DNA (DNA methylation) and proteins wrapped around it (e.g., histone methylation). As cancer cells have no function in our body, they frequently lose their cellular identity due to perturbed epigenetic programs. Furthermore, cancer cells can also have many different epigenetic states, which leads to high variability in their behavior. Thus, regulators of these epigenetic programs are emerging therapeutic targets in various human cancer types. There are three types of epigenetic regulators: (1) writers add chemicals to the DNA or chromatin proteins, (2) erasers remove the added chemicals, and (3) readers bind to these chemicals to influence the expression of genes. One group of such erasers is histone demethylases (HDMs) with KDM6A as representative member. In the past few years, inhibitors of HDMs have been developed that perturb their eraser function and halt the growth of some cancer types.

In our prior Idea Award, we found that TNBCs that are classified into the same group based on gene expression profiles have differences in epigenetic patterns and that epigenetic profiles better predict tumor cell behavior than gene expression. Furthermore, even within the same tumor there are cancer cells with different epigenetic states and these have different ability to metastasize and respond to treatment. Based on these data, we hypothesize that there is a high degree of epigenetic heterogeneity both between and within TNBCs and that these influence treatment responses and risk of metastatic progression. We propose two specific aims to test these hypotheses:

In Aim 1, we will define epigenetic heterogeneity in TNBCs. To accomplish this, we will analyze the gene expression, DNA methylation, and chromatin profiles of TNBCs; we will use patient samples and xenografts and cultures derived from these. We will integrate all these data types to obtain a comprehensive view of the epigenetic states of TNBC.

In Aim 2, we will investigate the role of HDMs in defining epigenetic heterogeneity in TNBCs. We will inhibit the activity of HDMs by either decreasing their expression or inhibiting their activity and assess the consequences of these on the epigenetic profiles and functional characteristics of the cells with special emphasis on therapeutic sensitivity and metastatic progression.

The successful completion of the proposed project will define epigenetic between and within TNBCs, which will improve our understanding of subclasses of TNBCs and may also identify treatment combinations that more effectively eradicate TNBCs. The development of molecules that modulate the activity of epigenetic regulators such as HDMs is currently one of the most active areas of pharmaceutical research with several agents already in early phases of clinical testing. Thus, the results of our study could potentially be tested in clinical trials in a few years. Furthermore, better understanding of the subclasses of TNBCs would allow the more accurate assignment of patients to already available or currently tested therapies.