The technology at the heart of this shift is the single-cell or single-nucleus assay for transposase-accessible chromatin sequencing, known as scATAC-seq or snATAC-seq. Rather than measuring which genes are switched on, as single-cell RNA sequencing does, these methods measure where the genome is physically open for business. Chromatin accessibility reveals the cis-regulatory landscape: the enhancers and promoters that transcription factors can reach, and therefore the regulatory programs a cell is competent to activate. snATAC-seq has proven especially robust for human colorectal cancer specimens because it works with frozen and archived tissue, integrates cleanly with transcriptomic data, and captures an upstream, mechanistic readout of cellular identity that gene expression alone cannot provide. In effect, it reads the control panel of the cell rather than just the lights that happen to be on.

Applying this lens has produced a striking reframing of how colorectal cancer should be classified. The traditional consensus molecular subtype scheme, based on bulk transcriptomes, sorts tumors into four groups, but single-cell RNA sequencing showed that this classification mixes in signals from stromal and immune cells, obscuring the tumor cells themselves. A refined framework, the intrinsic consensus molecular subtype system, isolates the epithelial compartment and collapses the picture into two states: iCMS2 and iCMS3. Crucially, large-scale snATAC-seq analyses have now shown that this dichotomy is written into the chromatin. iCMS2 tumors retain open regulatory elements bearing the footprints of intestinal lineage factors such as HNF4A, CDX2, PPARA and ASCL2, controlling genes for differentiation, lipid metabolism and Wnt-driven stem cell renewal. In essence, these tumors co-opt a corrupted version of normal intestinal stem cell circuitry. iCMS3 tumors instead display a dedifferentiated landscape, with accessibility at stress-responsive and mesenchymal enhancers enriched for AP-1, TEAD, SOX and MAFK motifs, reflecting an active disengagement from intestinal identity in favor of a primitive, stress-tolerant configuration.

The two states even map onto anatomy. iCMS2 tumors arise predominantly in the left colon, iCMS3 mainly in the right, hinting that the split may trace back to differences in embryonic endodermal origin. Patient-derived organoids faithfully maintain their tumor-specific regulatory landscapes through extended culture, and deep learning models trained on pan-cancer chromatin data can predict tumor lineage, differentiation state and oncogenic dependencies from accessibility patterns alone. Together these findings suggest that chromatin encodes a cellular identity fingerprint, one that explains why genetically similar tumors can behave so differently in the clinic.

The review then traces how this regulatory architecture is rewired as tumors progress, proposing a four-stage temporal model. The earliest detectable changes occur as normal mucosa becomes adenomatous polyp: enhancers bearing motifs for differentiation factors such as GATA4/6, KLF4/5 and HOX family members progressively close, while distal regulatory elements tied to Wnt and beta-catenin signaling, including elements targeting MYC, open up. Notably, this Wnt-associated chromatin remodeling appears in adenoma-resident stem cells but not in isolated Apc-null stem cells in mouse models, implying that interactions within the mutant crypt environment drive early epigenetic change. That finding challenges the linear genetic model and positions epigenetic plasticity as a potential initiating layer of tumorigenesis itself.

The most extensive remodeling happens at the transition from late adenoma to invasive carcinoma, where tumors commit to one of the two epigenetic trajectories. In iCMS2 tumors, accessibility at intestinal factor loci such as HNF4A dips during early adenoma formation and then rebounds in established cancer, a stage-dependent shift that may let cells exploit altered metabolic programs while staying partially differentiated. iCMS3 tumors take the opposite route, with widespread AP-1-enriched accessibility; organoid studies suggest AP-1 actively suppresses intestinal lineage factors like CDX2 and HNF4A to drive dedifferentiation toward a fetal state, and NF-kappa-B signaling, which alone can dedifferentiate intestinal epithelium in mice, may fuel the inflammatory character of this state.

A particularly viral concept emerging from this literature is oncofetal reprogramming, the reactivation of fetal gene programs in cancer. LGR5-positive cells behave as classic cancer stem cells, yet ablating them does not shrink tumors because LGR5-negative cells replenish the pool. Lineage tracing shows that it is the LGR5-negative cells that preferentially migrate to metastatic sites and then switch back to the LGR5-positive state to proliferate. Multiple studies now indicate that these LGR5-negative cells reactivate fetal enhancer programs, and in mouse tumoroids resembling iCMS2, chemotherapy with FOLFIRI upregulated oncofetal genes, with the oncofetal cells showing reduced drug sensitivity. Spatial transcriptomics places oncofetal cells at the invasive front of early human tumors, driven by TGF-beta signals from specialized fibroblasts. At the mechanistic level, enhancer accessibility differs sharply between adult and fetal intestine even when promoter accessibility and three-dimensional genome architecture look similar, and YAP/TAZ-TEAD motifs mark the oncofetal state while ASCL2 and TCF-LEF motifs mark the stem state. Whether chemotherapy selects for pre-existing oncofetal cells or induces them, and whether YAP/TAZ signaling operates as a shared, subtype-agnostic vulnerability across both iCMS states, remain key open questions.

Metastasis adds a further twist: organotropic chromatin remodeling. In mouse models of liver colonization, snATAC-seq shows metastatic cells progressively losing access to colon-specific enhancers bearing IRF1, ELF1 and STAT1 motifs while gaining access to liver-specific elements bearing HNF4A, FOXA2 and CTCF motifs, generating hybrid cells that express both epithelial and hepatocyte-like programs. The acquisition of HNF4A-driven landscapes in liver metastases echoes the iCMS2 signature, and the clinical observation that left-sided, iCMS2-enriched tumors preferentially spread to the liver raises the tantalizing possibility that primary tumor epigenotype predicts metastatic destination. Co-accessibility analyses suggest that long-range enhancer-promoter contacts expand around HNF4A and FOXA2, wiring metastatic cells into liver-specific growth and metabolic signals.

The tumor microenvironment is not a bystander in this process. Cancer-associated fibroblasts split into tumor-promoting subtypes with accessibility enriched for RUNX1, AP-1, NF-kappa-B and HIF1A motifs, and tumor-restraining subtypes that retain homeostatic extracellular matrix programs and are progressively depleted during malignant transformation. Normal fibroblasts acquire CAF-like histone marks when exposed to tumor-conditioned media, confirming that this divergence is epigenetic rather than genetic. In the immune compartment, CD8-positive T cells lose accessibility at the TCF7 locus, a pioneer factor for T cell stemness, marking terminal exhaustion that is already partially established in adenomas, suggesting immunosuppression is an enabling event rather than a late consequence. Regulatory T cells split into IL-10-positive and IL-10-negative subtypes with opposite prognostic implications, potentially resolving the paradox that T regulatory infiltration predicts good outcomes in colorectal cancer but poor outcomes elsewhere. Macrophages divide into C1QC-positive homeostatic and SPP1-positive tumor-enriched subtypes with distinct regulatory architectures. The authors propose viewing all of this as an iCMS-defined epigenomic ecosystem, in which malignant, stromal and immune compartments co-evolve through chromatin-level crosstalk.

Genomic alterations and chromatin states also feed back on each other. Recurrent noncoding mutations rewire transcription factor binding, as in the ETS2 enhancer that promotes epithelial-mesenchymal transition, or TERT promoter mutations that create de novo ETS sites and reactivate telomerase. Structural variations hijack enhancers to supercharge MYC and CCND1, and extrachromosomal DNA circles carry amplified oncogenes with their enhancers as mobile regulatory platforms, segregating nonrandomly during cell division. The same mutation can even have opposite accessibility effects in malignant versus stromal cells, as deep learning models trained on single-cell chromatin data have revealed. Translationally, these insights are already bearing fruit: DNA methyltransferase and EZH2 inhibitors can reopen silenced immune gene enhancers, and early-phase trials combining low-dose DNMT inhibitors with anti-PD-1 antibodies show promising responses in microsatellite-stable colorectal cancer, a population that normally ignores immunotherapy. Cell-free ATAC-seq promises liquid biopsies that read a tumor’s global regulatory state rather than single mutations, spatial epigenomic methods are mapping regulatory domains in intact tissue, and AI foundation models trained on single-cell epigenomes may one day simulate the iCMS2-to-iCMS3 transition and nominate the transcription factors whose manipulation could reverse it. The review’s boldest suggestion is that a therapy capable of reversing the epigenetic clock, restoring oncofetal and dedifferentiated cells to mature, lineage-committed states, could constrain tumor plasticity at its source, turning the cancer’s own regulatory logic against it.

Subject of Research: Single-cell epigenomic profiling of chromatin accessibility in colorectal cancer

Article Title: Single-cell epigenomics of colorectal cancer

Article References: Kang, D., Lee, J. H., & Jung, I. (2026). Single-cell epigenomics of colorectal cancer. Experimental & Molecular Medicine. https://doi.org/10.1038/s12276-026-01855-4

Image Credits: AI Generated

DOI: 10.1038/s12276-026-01855-4

Keywords: colorectal cancer, single-cell epigenomics, snATAC-seq, chromatin accessibility, iCMS subtypes, oncofetal reprogramming, tumor microenvironment, cancer-associated fibroblasts, T cell exhaustion, enhancer hijacking, extrachromosomal DNA, liquid biopsy

The new study, published in the journal Molecular Cell and led by Johnathan R. Whetstine, director of the Cancer Epigenetics Institute, demonstrates that gene copy-number amplification, long viewed as a random byproduct of genomic chaos, is in fact a biologically regulated process governed by specific chromatin machinery. By identifying the enzymatic gatekeepers that control how MYC amplification begins and progresses, the team has opened a potential therapeutic route that sidesteps the protein entirely and instead constrains the DNA structural events that multiply the gene. According to Whetstine, this strategy of targeting the DNA structural assembly line rather than the finished protein could have relevance for at least 30 percent of all cancers.

The technical heart of the discovery lies in two chromatin-regulating proteins, KDM4C and SETD2, which the researchers found act as gatekeepers controlling access to the MYC gene region. SETD2 is a histone methyltransferase that helps maintain a stable, properly marked chromatin environment, while KDM4C is a histone demethylase that remodels the epigenetic landscape. Under normal conditions, these enzymes help keep DNA packaged and regulated within the cell. But when SETD2 is lost or blocked, KDM4C gains improper access to the MYC locus and alters the surrounding DNA environment. Tumors, the team found, frequently increase KDM4C levels, which further drives the deleterious amplification process.

Using human cell models, genomic analyses, and mouse studies, the researchers showed that KDM4C does more than loosen chromatin. It actively recruits the cell’s DNA-copying machinery to the MYC region, causing the gene to be copied repeatedly and generating extra MYC copies. The effect was not circumstantial. In a striking proof of causality, the team used dCas9 technology to direct KDM4C to the MYC region artificially, and that alone was sufficient to trigger amplification, even in cells with a completely normal set of chromosomes. As Benjamin I. Ferman, a PhD student in the Whetstine lab and first author of the study, explained, the findings demonstrate that MYC amplification is not simply a random byproduct of genomic instability, but a process actively controlled by specific epigenetic programs.

Copying MYC, however, is only half the story. The real danger emerges when cells carrying dangerous levels of MYC fail to eliminate themselves. MYC overactivation normally triggers programmed cell death, a built-in safeguard against runaway proliferation. The researchers found that disabling this fail-safe allowed hyperamplification and tumor formation to proceed. When cells lost their normal ability to undergo apoptosis, abnormal MYC amplifications could reach high levels and continue expanding. This effect was especially pronounced when cells also lost TP53, a crucial tumor suppressor gene that normally helps eliminate damaged cells before they can do harm.

Over time, the combination of disrupted epigenetic control and TP53 loss drove MYC amplification to high levels, including the formation of circular extrachromosomal DNA, a notoriously aggressive form of oncogene amplification that can accelerate tumor evolution and drug resistance. In mouse models, the consequences were decisive: the starting cells were non-transformed and did not cause tumors without MYC amplification, but once the epigenetic program ran its course and cellular defenses failed, tumor formation followed. The findings suggest that MYC amplification develops in stages, with chromatin changes first triggering the formation of extra gene copies, and the failure of normal defense mechanisms then allowing those abnormal cells to survive, spread, and grow more aggressive. Whetstine notes that this staged relationship echoes the Two-Hit Theory of cancer causation, the landmark concept originated at Fox Chase by Alfred Knudson.

The translational implications are significant. Because the newly identified regulatory mechanisms contain enzymatic targets, they could in principle be exploited through small-molecule drug development, opening therapeutic avenues for MYC-driven cancers that have resisted every conventional approach. The Whetstine lab found that blocking KDM4C activity, either genetically or with targeted drugs, reduced MYC amplification across multiple experimental systems, including MYC-amplified cancer cells and animal models. Notably, a clinically used KDM4 inhibitor successfully suppressed MYC amplification in vivo, an early proof of concept that a challenge long considered untouchable may be addressable in the clinic.

Beyond drug development, the work points to a new biomarker paradigm. If specific epigenetic imbalances, such as SETD2 loss combined with elevated KDM4C, reliably predict which tumors are poised to develop aggressive amplification states, clinicians could potentially identify high-risk cancers earlier and intervene before extrachromosomal DNA and treatment resistance take hold. Whetstine emphasized that MYC amplification appears to be far more dynamic and controlled than previously appreciated, raising the possibility that these amplification states could be therapeutically constrained rather than simply endured. Biomarker and therapeutic studies are already under development at the Cancer Epigenetics Institute and Fox Chase.

The research was supported by the National Institutes of Health and developmental funding from Fox Chase Cancer Center. In addition to the Fox Chase team, collaborators from Massachusetts General Hospital, Harvard Medical School, Stanford University, the Broad Institute, and the Icahn School of Medicine at Mount Sinai contributed to the study. Taken together, the findings reframe one of cancer biology’s oldest puzzles: the amplification of a master growth gene is not a stochastic accident but a regulated epigenetic program with identifiable switches, and those switches, unlike the MYC protein itself, may finally be druggable.

Subject of Research: Epigenetic regulation of MYC gene amplification in aggressive cancers

Article Title: Cracking the 'undruggable' code: Fox Chase Cancer Center researchers unveil epigenetic blueprint fueling aggressive MYC amplification driving cancer

Article References: Cracking the 'undruggable' code: Fox Chase Cancer Center researchers unveil epigenetic blueprint fueling aggressive MYC amplification driving cancer. (n.d.). Original publication

Image Credits: AI Generated

DOI: Not provided

Keywords: MYC, gene amplification, epigenetics, KDM4C, SETD2, extrachromosomal DNA, TP53, chromatin, undruggable targets, Fox Chase Cancer Center, Molecular Cell, cancer therapy

Extrachromosomal DNA first attracted attention decades ago when researchers noticed strange, ring-shaped DNA fragments floating outside the chromosomes of tumor cells. Unlike chromosomal DNA, ecDNA is structurally independent, frequently circular and acentromeric, meaning it lacks the centromere structures that normally ensure chromosomes are distributed evenly during cell division. This absence has profound consequences. During mitosis, ecDNA molecules are inherited through noncanonical behaviors that allow unequal segregation between daughter cells, meaning one daughter cell can receive a windfall of oncogene copies while the other receives few or none. The review emphasizes that this non-Mendelian inheritance pattern also permits coordinated co-inheritance, in which multiple ecDNA species carrying different oncogenes and regulatory elements travel together through cell generations, allowing advantageous combinations of genes and regulatory states to persist and propagate within a tumor population.

The classical view of ecDNA centered on copy-number biology: more copies of an oncogene such as MYC or EGFR meant more messenger RNA and more oncogenic protein, fueling aggressive growth. The new synthesis argues that this framing misses the most consequential part of the story. ecDNA molecules carry not just genes but enhancers, the short DNA sequences that act as volume controls for gene expression. Because ecDNA is circular, enhancers that were once distant from their target genes on linear chromosomes can be brought into immediate physical proximity, rewiring enhancer-oncogene communication in ways that linear DNA cannot easily achieve. The review describes how ecDNA can hijack pre-existing enhancers, activate noncanonical regulatory elements that would normally remain silent, and thereby transform a raw genome rearrangement event into genuine epigenetic innovation.

At the physical level, ecDNA does not float passively in the nucleus. Advanced imaging and genomic mapping studies summarized in the review show that ecDNA molecules cluster together into transcriptionally active nuclear hubs, dense assemblies where multiple circular molecules and their target genes on chromosomes converge. These hubs behave like condensate-associated structures, membrane-less compartments that concentrate transcription machinery, transcription factors and co-activators at extraordinarily high local concentrations. The result is a hyperactive transcriptional factory that can drive oncogene expression to levels unattainable through ordinary chromosomal regulation. In this sense, ecDNA-positive tumor cells are not merely carrying extra genetic cargo; they are constructing an entirely new layer of nuclear architecture dedicated to amplifying and fine-tuning oncogenic signaling.

This architectural rewiring has direct consequences for chromatin itself. The review details how ecDNA reshapes chromatin accessibility, the degree to which DNA is physically open and available to transcription factors. ecDNA tends to maintain an unusually open chromatin state, rich in active histone marks, which keeps its passenger enhancers and promoters in a perpetually transcription-ready configuration. Because ecDNA hubs can also contact chromosomal targets, this openness can extend to the ordinary genome, altering which chromosomal genes are switched on or off. Through these mechanisms, ecDNA generates transcriptional plasticity, the capacity of a tumor cell to shift its gene-expression program rapidly in response to changing conditions, whether that condition is a drug, an immune attack or a change in nutrient supply.

That plasticity feeds directly into some of the most clinically feared phenomena in oncology. The review positions ecDNA as a key driver of intratumoral heterogeneity, the coexistence of genetically and epigenetically distinct cell populations within a single tumor. Because ecDNA segregates unequally during mitosis, daughter cells diverge quickly in oncogene dosage and regulatory state, generating a diverse portfolio of cellular phenotypes from a single founding lineage. This diversity provides abundant raw material for clonal evolution: when chemotherapy eliminates cells with one regulatory configuration, subpopulations with different ecDNA compositions and enhancer landscapes survive and repopulate the tumor. The same logic applies to metastatic adaptation, where disseminated cells must reprogram their transcriptional identity to colonize new tissues, and to therapeutic resistance, where ecDNA-mediated enhancer rewiring can rapidly upregulate drug targets or bypass inhibited pathways.

Yet the very features that make ecDNA so powerful for the tumor may also constitute its Achilles’ heel. The review highlights that the high transcriptional burden carried by ecDNA creates a form of genomic stress known as transcription-replication conflict. When the transcription machinery and the DNA replication machinery collide on the same stretch of DNA, the resulting collisions can stall replication forks and generate DNA damage. ecDNA-positive cells, with their extraordinarily high transcriptional output, may live permanently near the threshold of tolerable transcription-replication interference. This dependence suggests selective vulnerabilities that could be exploited therapeutically: drugs that further stress transcription or replication, or that disrupt the specialized regulatory architecture of ecDNA hubs, might preferentially harm ecDNA-rich tumors while sparing normal cells.

The authors also point to the dependence of ecDNA-positive tumors on the integrity of their nuclear hubs and condensates as a second class of vulnerability. If transcriptional output in these tumors relies on the physical coalescence of ecDNA molecules and their associated regulatory factors, then interventions that dissolve or destabilize those assemblies, for example by targeting condensate-forming proteins or the chromatin regulators that maintain open chromatin on ecDNA, could collapse the tumor’s oncogenic transcription program. Similarly, because ecDNA inheritance depends on noncanonical mitotic behaviors rather than centromeres, the machinery that distributes ecDNA during cell division represents another potential point of attack. None of these strategies has yet produced approved therapies, but the review frames them as a conceptual frontier for precision oncology, in which treatment decisions could one day be guided by mapping a tumor’s ecDNA complement and its regulatory architecture.

Perhaps the most significant contribution of the review is conceptual. By shifting the focus from copy-number biology to regulatory biology, the authors reframe ecDNA not as a passive amplifier of oncogenes but as an active platform for epigenetic reprogramming, a mobile, self-perpetuating module that converts structural genome chaos into functional transcriptional diversity. This reframing has implications for how scientists model tumor evolution, how pathologists assess tumor aggressiveness and how drug developers search for targets in the least stable, most adaptable corners of the cancer genome. As long-read sequencing, spatial genomics and chromatin mapping technologies mature, the ability to read a tumor’s ecDNA landscape may become as routine as today’s copy-number profiling, opening a window onto the epigenetic strategies that cancers use to survive everything medicine throws at them.

The review, published open access in Molecular Cancer, was supported by the Shandong Provincial Traditional Chinese Medicine Science and Technology Project and the Health Commission of Jiangxi Province. Its authors span institutions across China, including Nanchang University, Hunan University, Naval Medical University and Shandong First Medical University, reflecting the rapidly growing international effort to understand the nonchromosomal genome of cancer. What emerges from their synthesis is a picture of tumors that are not just collections of mutated genes but ecosystems of mobile regulatory elements, and of ecDNA as the engine that keeps those ecosystems adaptable, heterogeneous and, for now, one step ahead of treatment.

Subject of Research: The role of extrachromosomal DNA as a regulatory platform for epigenetic reprogramming in cancer

Article Title: Extrachromosomal DNA as a platform for epigenetic reprogramming in cancer

Article References: Liu, B.-X., Zhou, L., Lai, S., Han, X., Dai, X.-J., Liu, R., Wang, Y., Chen, D., Wei, J., Hu, H., Zu, W., Jiang, L., Chang, J., & Zha, L. (2026). Extrachromosomal DNA as a platform for epigenetic reprogramming in cancer. Molecular Cancer. https://doi.org/10.1186/s12943-026-02789-1

Image Credits: AI Generated

DOI: 10.1186/s12943-026-02789-1

Keywords: extrachromosomal DNA, ecDNA, epigenetic reprogramming, chromatin accessibility, enhancer rewiring, transcriptional plasticity, tumor evolution, intratumoral heterogeneity, therapeutic resistance, nuclear hubs, precision oncology, Molecular Cancer