Countering epigenetic control of cell identity gone awry

Every cell in the body carries the same DNA, yet a neuron, a blood stem cell and a muscle cell each perform vastly different functions. What sets them apart is not the genetic code itself, but how it is used. Through epigenetic regulation, cells activate or deactivate genes, guiding development, establishing and preserving cell identity, and helping tissues function as they should.

When epigenetic control breaks down, the consequences can be profound. In pediatric cancers and other diseases, cells can become trapped in immature developmental states, unable to reach their fated cell identities. At St. Jude, researchers are studying how epigenetic programs shape these identities in health and disease, revealing vulnerabilities that may lead to new therapies.

“Many pediatric cancers can be thought of as disorders of developmental pathways,” said Adam Durbin, MD, PhD, Department of Oncology. “As cells differentiate into their fated lineages, they turn different sets of genes off and on; childhood cancer cells may stop this process early, keeping cells in a more proliferative and less differentiated state.”

Windows of epigenetic opportunity

Durbin studies the epigenetics of pediatric solid and brain tumors, such as neuroblastoma, rhabdomyosarcoma and medulloblastoma, which have very few therapeutically targetable genetic cancer mutations. Rather than focusing directly only on those mutations, his lab asks how they reshape the epigenetic landscape of a developing cell and whether that altered state creates new vulnerabilities.

“Surprisingly, we’ve found that how you target epigenetic and transcriptional regulators in these different settings makes an immense difference to the resulting outcomes,” Durbin explained. “Altering the genes that tumors need to sustain the transformed state can promote strikingly different outcomes.”

With this in mind, Durbin investigated specific transcription factors in rhabdomyosarcoma. In work published in Cell Reports, his lab identified that transcription factors, such as SIX1, that were only present at certain points in development, were required to sustain cancer cell growth. Without this factor, rhabdomyosarcoma cells differentiated into skeletal muscle. In parallel, they studied mechanisms of targeting epigenetic regulators that control these transcription factors. In a work published in Nature Communications, they demonstrated that targeting the same epigenetic regulators using different methods can cause strikingly different outcomes, ranging from no effect to cell death.  

That window of susceptibility to transformation is mirrored in most pediatric brain tumors, including ependymoma, medulloblastoma and pediatric high-grade glioma. Stephen Mack, PhD, Department of Developmental Neurobiology, studies ependymoma, which is the third most common childhood brain cancer. “We’ve found that there are specific sites in the genome that are epigenetically open during a critical time in development in cells that become ependymoma,” said Mack. “The primary cancer-causing protein binds and activates those sites at a specific time during development.”

Together, these findings suggest that in some pediatric cancers, the mutation alone is not enough. The cell’s preexisting identity and the epigenetic state that defines it can determine whether an oncogenic mutation takes hold.

Fusion oncoproteins hijack the epigenetics of cell identity

Ependymoma and other cancers can be driven by an epigenetically active fusion oncoprotein that is present during that critical window of vulnerability. Fusion oncoproteins occur when two genes fuse, creating a new hybrid protein comprised of two portions, one from each of the original genes, gaining new functions — including altering cell identity.

Understanding how these hybrids distort the epigenetics of cell identity is of great interest in ependymoma, as 70% of cases are driven by the ZFTA–RELA fusion oncoprotein. Mack’s lab showed in Nature that if epigenetic controls make the right genes accessible during development, the ZFTA portion of the fusion will bind to them. The RELA region will then inappropriately activate those genes, resulting in a cancer-promoting cell state.

“We established that this fusion promotes oncogene expression,” Mack said, “but we also found that these fusion events must be paired with a specific cell type early in development that’s primed for transformation. In these cells, the default state is proliferation, a time when the brain is growing at its fastest rate.”

The transformed cells also express markers associated with early developmental states, reinforcing the idea that the fusion takes over these early cells, pushing them epigenetically off their fated trajectory and instead causing them to become malignant cells.

Turning aberrant epigenetics into a therapeutic opportunity

Fusions also present a therapeutic opportunity, but targeting them can be difficult because targeting the genes that comprise a fusion also means targeting their nonfused versions in healthy cells — a challenge for drug development. Instead, understanding how these fusions promote cancer by altering chromatin, the complex of DNA and proteins that control gene expression, may reveal alternative therapeutic options.  

“We’ve found that one of the unique features of many fusion oncoproteins is their tendency to form aberrant biomolecular condensates on chromatin,” said Richard Kriwacki, PhD, Department of Structural Biology. “Condensates are liquid-like structures that bring certain proteins and other biomolecules together within the three-dimensional space of the cell nucleus. In the case of ZFTA–RELA fusions in ependymoma, the condensates they make bring together chromatin-modifying enzymes and certain regions of the genome in an abnormal way.”

These fusion-caused condensates do not exist in normal cells, representing an opportunity. If scientists can inhibit a part or parts of these aberrant, liquid-like (or fluid) assemblies, they may find a way to affect only ependymoma cells selectively. “Maybe some of those interacting proteins could be targets,” said Mack, who worked with Kriwacki to characterize these structures, which were published in Nature Cell Biology.

Scientists are further down the pathway to identifying such therapies in leukemia caused by NUP98 fusions, which account for about 8% of pediatric acute myeloid leukemia cases. Kriwacki co-authored a study with Charles Mullighan, MBBS (Hons), MSc, MD, St. Jude Comprehensive Cancer Center senior deputy director and Department of Pathology member, and colleagues in Cancer Discovery examining which proteins interacted with NUP98 fusions and DNA in condensates. The researchers identified the acetyltransferases MOZ/KAT6A and HBO1/KAT7 as critical to its function, then tested them as therapeutic targets, finding that inhibiting them greatly reduced leukemia cell growth.  

“Our results with the Mullighan laboratory provide a proof of concept that targeting these condensates may reduce the aberrant gene expression fueled by fusions and possibly reverse disease,” Kriwacki explained.

While studying epigenetics can lead to novel therapies, adding epigenetic modulators may be especially powerful when used to reshape cell identity to improve the effects of other treatments. In NUP98-rearranged leukemia, Kriwacki, Mullighan and collaborators found that combining acetyltransferase inhibition with inhibition of menin, another molecule that helps the fusion activate oncogenes, had a synergistic effect, even in patient samples from difficult-to-treat relapsed disease.

Understanding normal cell identity development

To understand how cell identity goes awry, scientists must also understand how it is established and maintained under normal conditions. Blood stem cells are an informative model because they have many epigenetic controls to balance self-renewal and differentiation without becoming cancerous.

“Hematopoietic stem cells express many genes to be able to respond to different cues, making them ‘noisy’ to study,” said Shannon McKinney-Freeman, PhD, Department of Hematology. “Instead, examining the epigenetic patterns controlling that expression has been a better indicator of their cell identity and what kind of cells they will make.”

Using that approach, McKinney-Freeman’s lab has uncovered how blood stem cells maintain a balance of self-renewal and progeny creation during blood cell formation at the epigenetic level. Published in Blood Advances, the researchers uncovered how regulatory networks of transcription factors help restore the entire blood cell system after transplantation. They further investigated this in preclinical models that revealed the importance of cellular signaling and transport proteins to the process, also published in Blood Advances.  

McKinney-Freeman’s research also shows that disrupted identity programs are not limited to cancer. In studies of sickle cell disease, she and her colleagues found that blood stem cells from affected individuals have a distinct functional identity. Compared to stem cells from unaffected individuals, they behave differently and are less functional, suggesting that chronic disease-associated signals may reshape their epigenetic state.

“One of the impacts of the trademark sickling is an increase in red blood cell death,” said Terri Cain, St. Jude Children’s Research Hospital Graduate School of Biomedical Sciences, who is working on her doctorate in the McKinney-Freeman lab. “That triggers the release of inflammatory cytokines into the bloodstream, and studies in other contexts have shown these molecules have a large influence over the epigenetic landscape, so we’ve begun to look at how that impacts blood stem cells during sickle cell disease, and how that could open new therapeutic opportunities.”

Together, those studies provide an important reference point: Scientists must first understand how healthy identity is built and maintained before learning how it can be warped in disease, but if they can do so, it may lead to new therapies.

Centering cell identity

From blood stem cells maintaining normal function to fusion proteins trapping cells in malignant developmental states, these studies point to a shared theme: Epigenetic regulation helps determine what a cell is, what it can become and how it responds to therapy.

“An emerging theme across fields is that cells can become epigenetically trapped in an abnormal state,” Mack said. “If we can find ways to alter the epigenome to push cells towards their normal trajectory, that could become a generalizable approach to identify new therapies that don’t focus on killing diseased cells.”

“Cell identities are incredibly complex, and every layer of epigenetic control we understand reveals even more intricacies,” McKinney-Freeman said. “But as we grow our understanding, we are beginning to enter the realm of enhancing or changing cell identity in novel ways to benefit patients facing these catastrophic diseases.”