Cancer epigenetic therapy: recent advances, challenges, and emerging opportunities

ABSTRACT

Epigenetic dysregulation is an important nexus in the development and maintenance of human cancers. This review provides an overview of how understanding epigenetic dysregulation in cancers has led to insights for novel cancer therapy development. Over the past two decades, significant strides have been made in drug discovery efforts targeting cancer epigenetic mechanisms, leading to successes in clinical development and approval of cancer epigenetic therapeutics. This article will discuss the current therapeutic rationale guiding the discovery and development of epigenetic therapeutics, key learnings from clinical experiences and new opportunities on the horizon.

1. Introduction

Epigenetics is the study of heritable changes in gene function that are not coded in DNA sequences [1,2]. The human body encompasses trillions of cells, and each cell contains an identical genetic blueprint encoded in DNA. The two-meter-long DNA is highly compacted into a chromatin structure in the cell nucleus. Epigenetic modifications, which involve alterations in gene expression without changes to the underlying DNA sequences, play a critical role in normal development and disease progression. Numerous modifications to DNA and histone proteins are now characterized to form an epigenetic code that regulates chromatin accessibility for transcription, replication, and DNA damage repair [3]. Dysregulation of epigenetic regulatory mechanisms leads to many human diseases, including developmental disorders, neurological diseases, autoimmune diseases, and cancers [4]. In this review, we will discuss epigenetic dysregulation in cancers, opportunities to develop precision therapeutics targeting cancer-specific epigenetic mechanisms and highlight recent progress made in the drug discovery field. Due to space limitation, we will focus our discussion on therapies undergoing active clinical investigation. We will also share our perspectives on a few potential future avenues in developing the next wave of cancer epigenetic therapies.

2. Cancer epigenetic drug discovery

Cancer epigenetics represents a promising frontier in the field of oncology drug discovery, offering new avenues for targeted therapies against cancers. There have been significant efforts to develop novel chemical inhibitors targeting protein families which regulate DNA and histone modifications. Most notable of these targets include DNA cytosine methylation, histone lysine and arginine methylation, and histone lysine acetylation and “reader” protein families that bind to specific DNA and histone modifications. The aim is to deliver anti-tumor activity and to enhance overall cancer treatment efficacy by specifically targeting and modulating these epigenetic regulators. Through a combination of innovative drug screening approaches and advanced drug design strategies (Figure 1), the quest for effective cancer epigenetic drugs holds significant promise in delivering new cancer treatment paradigms.

Figure 1.

Drug discovery targeting different epigenetic target classes. (a) Proteins involved in epigenetic regulation can deposit (writers), detect (readers), and remove (erasers) the various covalent modifications on DNA and histones [5]. Direct methylation of DNA and post-translational modifications on histones are two major epigenetic mechanisms involved in regulating gene expression. The complex interplay between these factors controls genomic accessibility, which in turn regulates expression of genes involved in a variety of cellular processes. Histone acetylation and methylation networks constitute a large component of the human epigenome and have been the focus of small molecule cancer epigenetic drug discovery in the past two decades. Selected examples of inhibitors targeting writers, readers and erasers are illustrated. (b) Chromatin remodeling enzymes are responsible for the rearrangement, displacement, and replacement of nucleosomes on the chromatin. Currently, there are successful examples of small-molecule inhibitors and proteolysis-targeting chimeras (PROTACs) targeting this enzyme class. Created in BioRender. Vatapalli, R. (2024) BioRender.com/t18f526.

2.1. Reverse dysregulated epigenetic and transcriptional programs in cancers

All cancers have highly dysregulated epigenetic landscapes and transcriptional signatures [6,7]. For example, cancer cells generally exhibit focal DNA hypermethylation in gene promoters, accompanied by global DNA hypomethylation. Transcriptomic profiling of human cancers has shown heterogeneous transcriptomic features, including features reminiscent of early progenitor cell development signature, or epithelial–mesenchymal transition (EMT) gene signature or embryonic stem cell-like gene signature [8–10]. Disruption of normal gene expression contributes to tumor initiation, progression, and resistance to therapies. One of the earliest concepts of cancer epigenetic therapy aims to restore normal gene expression. While genetic alterations and mutations are largely irreversible, it was hypothesized that epigenetic changes are more readily reversible with small-molecule therapeutics [11,12].

Inhibitors of DNA methyltransferase (DNMT) and histone deacetylase (HDAC) were developed to restore epigenetic imbalance in cancers. These agents led to the initial clinical successes of cancer epigenetic therapy, with FDA approval of DNMT inhibitors azacytidine and decitabine for the treatment of myelodysplastic syndromes (MDS) and leukemia, and HDAC inhibitor vorinostat (also known as suberoylanilide hydroxamic acid (SAHA)) for the treatment of cutaneous T cell lymphoma (CTCL). The first generation of DNMT and HDAC inhibitors were developed prior to the concept of targeted drug discovery, and they are not selective against close family members and carry off-target activities. Azacytidine, a nucleoside analog, was first synthesized by Sorm and colleagues in the 1960s as a cytotoxic chemotherapeutic agent [13]. The discovery and development of DNMT inhibitors have been extensively reviewed [14,15]. Both azacytidine and decitabine are prodrugs. After uptake into the cell via nucleoside transporters, these molecules undergo a series of phosphorylation modifications, are converted to cytidine triphosphate (CTP) and deoxycytidine triphosphate (dCTP) analogs and then get incorporated into DNA and RNA. Azacytidine and decitabine can directly cause DNA damage by forming covalent adducts with DNMTs on DNA. The “trapped” DNMTs are degraded by a proteasome-mediated mechanism leading to genome-wide DNA hypomethylation [14]. Peter Jones and colleagues in 1980 made the seminal observation that azacitidine can alter the differentiated state of cultured cells and inhibit the methylation state of newly synthesized DNA [16]. DNMT inhibitors are typically used at low doses in the clinic to inhibit DNA methylation following a few rounds of cell cycle progression. At high doses, they cause acute cytotoxicity typical of chemo agents and mask the desirable effect of reprogramming the DNA methylation state. There has been continued interest in developing reversible, isoform selective DNMTi to improve therapeutic index [17]. GSK-3685032 is a novel first-in-class reversible DNMT1 selective inhibitor showing great promise in preclinical studies [17]. The highly selective DNMT inhibitors are much better chemical tools to selectively modulate DNA methylation levels in cells and to evaluate therapeutic hypotheses of inhibiting DNMT1. Thus far, no clinical study has been reported with this new generation of reversible and isoform selective DNMT1 inhibitors.

HDACs catalyze the removal of acetyl groups from lysine residues on histones [18]. These enzymes modify chromatin structure and contribute to aberrant gene expression in cancers. Therefore, targeting HDACs to restore normal cellular gene expression were actively investigated as a potential anticancer therapeutic strategy [19]. All approved HDAC inhibitors (HDACi), including vorinostat, belinostat, romidepsin, and panobinostat, have broad activities to elevate global histone acetylation by inhibiting multiple HDACs within the Zn²⁺ containing HDAC subfamily (HDAC1–11) [5,20]. Rich sources of natural products have greatly accelerated the discovery and development of HDAC inhibitors. Trichostatin A and trapoxin were among the first natural products verified to inhibit HDACs [21]. Cyclic depsipeptides, such as FK228 (also known as romidepsin), are a natural product originally isolated from the bacterium Chromobacterium violaceum that also potently inhibits HDACs [22]. HDACi demonstrated potent cytotoxic activity against human cancer cell lines, inducing differentiation and apoptosis of leukemic cell lines. Extensive preclinical and clinical investigations eventually led to multiple FDA approvals, including 1) vorinostat for the treatment of CTCL in 2006; 2) romidepsin for the treatment of CTCL in 2009 and peripheral T-cell lymphoma (PTCL) in 2011; 3) belinostat for the treatment of PTCL in 2014; and 4) panobinostat for the treatment of multiple myeloma (MM) in combination with other drugs in 2015. In 2019, tucidinostat (also known as Chidamide) was approved by China’s National Medical Products Administration for the treatment of breast cancer. This is the only HDAC inhibitor approved to date for a solid cancer treatment [23–25]. For details on the discovery and development of HDACi, please see reviews [26–28].

Both DNMT and HDAC inhibitors have broad mechanisms of action with pleiotropic toxicity in normal tissue, which has limited their clinical utility. Overall, these agents have a narrow therapeutic index. FDA approval is one measure of success; however, the overall clinical benefit of these agents has been fairly modest. Patients treated with pan-HDAC inhibitors have experienced severe adverse events such as fatigue or asthenia that lack a robust mitigation strategy. Ongoing efforts to develop more targeted and isoform-selective DNMT and HDAC inhibitors may overcome some of the challenges associated with the early generation of DNMT and HDAC inhibitors [29]. Beyond drug discovery efforts, better patient selection strategies are needed to maximize therapeutic benefit by identifying and enriching for responder populations. A comprehensive review of the ongoing clinical investigation on HDAC and DNMT inhibitors can be found here [30–32].

2.2. Precision targeted epigenetic therapeutics

Targeted cancer therapy has transformed cancer treatment where oncogenic driver proteins can be identified and selectively targeted by small molecule drugs [33,34]. Precision targeted cancer drugs are designed to specifically target molecular and genetic alterations in cancers. This approach helps improve tumor treatment efficacy and minimize normal tissue toxicities, potentially leading to fewer side effects. Key characteristics of precision targeted medicine include 1) identification of specific tumor biomarkers for patient selection; 2) personalized treatment matching patient with specific biomarkers with corresponding targeted therapies; and 3) therapeutic agents are highly selective against the protein drug targets of interest.

Large-scale cancer genome sequencing efforts have uncovered numerous genetic mutations of epigenetic regulators, adding new evidence that altered epigenetic regulatory mechanisms are a major hallmark of cancer [35]. Cancer genome sequencing projects have uncovered many alterations of epigenetic regulators in cancers, which fueled multiple waves of targeted drug discovery efforts to systematically target emerging enzyme classes important for epigenetic regulation, including protein methyltransferases, demethylases, acetyltransferases, DNA/RNA helicases, and numerous histone code reader family proteins [36].

Oncogenic driver mutations of epigenetic regulators are frequently observed in hematological cancers and rare pediatric cancers, including 1) hotspot gain-of-function (GoF) mutation of enhancer of zeste homolog 2 (EZH2) in lymphoma [37]; 2) chromosomal translocation of mixed lineage leukemia protein-1 (MLL1) in leukemia [38]; 3) chromosomal translocation and hotspot gain-of-function mutation of nuclear receptor binding SET domain protein 2 (NSD2) in multiple myeloma and acute lymphoblastic leukemia [39,40]; 4) chromosomal translocation of bromodomain-containing protein 3 (BRD3) and BRD4 in NUT midline carcinoma [41]; 5) chromosomal translocation of SS18 in synovial sarcoma [42]; and 6) the well-studied gain-of-function mutation in isocitrate dehydrogenase 1 (IDH1) and IDH2 in leukemia and glioma [43–45] and many more. Histones themselves are also mutated and directly linked to oncogenesis [46,47]. Missense mutations in histone H3 always occur at or near well-characterized regulatory residues that impact its post-translational modifications. For example, H3K27M mutation is found in nearly 80% of pediatric diffuse intrinsic pontine glioma (DIPG) and these mutations diminish global H3K27 trimethylation by locally trapping and inactivating the polycomb repressive complex 2 (PRC2) [48]. In most cases of DIPG, H3K27M, or H3G34V mutations are the only genetic alterations identified in the patient. H3K36M mutation is found in nearly 90% of chondroblastoma and this alteration suppresses global H3K36 methylation [49]. All histone mutations occurring in cancers have been comprehensively reviewed [50,51]. The identification of epigenetic driver oncogenes provides a strong rationale for precision-focused cancer epigenetic drug discovery. In this next section, we will discuss recent advancements in targeting epigenetic regulators as oncogenic drivers, as well as novel strategies to target loss-of-function alterations of epigenetic regulators.

2.2.1. Direct targeting of oncogenic drivers

There have been extensive efforts to develop novel therapeutics that target oncogenic driver mutations of epigenetic regulators. A number of these efforts have led to active clinical investigations (Table 1). The first success with this strategy is the FDA approval of EZH2 inhibitor tazemetostat for the treatment of follicular lymphoma (FL) in 2020 [52]. EZH2 is a member of the Polycomb-group (PcG) proteins which maintain the transcriptional repressive state by methylating histone H3 lysine 27 [53]. These proteins play a key role in regulating embryonic stem cell differentiation, renewal, cell fate decision, and the normal cellular differentiation program. Mutation of EZH2 and the PRC2 complex subunits are observed in multiple cancers, and they can be oncogenic or tumor suppressive depending on the cancer type. Oncogenic EZH2 hotspot, missense point mutations (including Y641, Y677, Y687) are concentrated in FL and diffuse large B cell lymphoma (DLBLC) of the GCB subtype [37,54–57]. Elegant biochemical investigation has demonstrated that these hotspot mutations result in neomorphic biochemical and cellular properties. While the EZH2 wild type protein is a very efficient enzyme in converting H3K27me0 to H3K27me1/2, most of the mutated EZH2 proteins showed elevated enzymatic activity that resulted in an elevated level of H3K27me3 [37,55]. Cancer cells containing EZH2 hotspot mutation display genome-wide hyper-H3K27 trimethylation. EZH2 inhibitors and EED inhibitors demonstrated profound anti-tumor activity in preclinical models, which promoted their clinical development [58–60]. Tazemetostat had an overall response rate (ORR) of 69% in EZH2 mutant FL patients, which led to its approval in FL [52]. Interestingly, tazemetostat is also efficacious in a subset of EZH2 wild type FL patients, with an ORR of 34% [61].

Table 1.

Inhibitors targeting epigenetic oncogenic drivers in clinical development (active trials only).

Target	Biomarkers	Inhibitors	Indication	Clinical trial number
EZH2	EZH2 GOF mutation	Tazemetostat, GSK2816126, PF-06821497, FTX-6058, SHR2554)	B-cell Non-Hodgkin lymphoma, follicular lymphoma, solid tumors	NCT03456726 NCT05228158 NCT03603951 NCT03213665 NCT05467943 NCT04104776 NCT05551936 NCT04224493
		Tazemetostat, SHR2554, PF-06821497, Valmetostat, HH-2853	PTCL, MPNST, Breast cancer, TNBC, mCRPC, R/R B-cell NHL, R/R ATL	NCT05559008 NCT04355858 NCT03805399 NCT06122389 NCT04917042 NCT03460977 NCT04703192 NCT04842877 NCT04102150 NCT05633969 NCT06244485 NCT03930953 NCT04390737
EED	EZH2 GOF mutation	MAK-683, APG-5918, ORIC-944	DLBCL, solid tumors	NCT02900651 NCT05415098. NCT05413421
Menin	MLL rearrangements/ NPM1m/FLT3m	BMF-219, SNDX-5613, KO-539, DSP-5336 JNJ-75276617	Leukemia, R/R Leukemias, MRD-positive AML	NCT05153330 NCT04065399 NCT04067336 NCT04811560 NCT04988555 NCT05326516 NCT05453903 NCT05521087 NCT05406817 NCT05761171 NCT04065399 NCT05360160 NCT06284486 NCT06226571 NCT05886049 NCT06222580 NCT05735184 NCT06001788 NCT04988555
	KRAS mutation	BMF-219	CRC, NSCLC, PDAC	NCT05631574
	Hox upregulation	SNDX-5613	Leukemias	NCT06229912
		SNDX-5613	AML, Pediatric AML, CRC, Solid tumors	NCT06313437 NCT06177067 NCT05731947
NSD2	NSD2 translocation	KTX-1001	Multiple myeloma	NCT05651932
SMARCA2/4		FHD-286	Metastatic uveal melanoma, AML, MDS	NCT04879017 NCT04891757
KAT6A		PF-07248144	metastatic ER+HER2- breast cancer, CRPC, NSCLC	NCT04606446

Translocations of the MLL1 gene typify a unique group of acute leukemia that are often associated with poor prognosis. The MLL1 gene encodes a DNA binding protein that methylates histone H3K4 to positively regulate gene expression, including the Hox gene loci [62]. Leukemogenic MLL1 translocations fuse the N-terminus of MLL1 with >80 different fusion partners, and the majority of these fusions can aberrantly recruit DOT1 Like Histone Lysine Methyltransferase (DOT1L) to potentiate transcription activity [63]. DOT1L is a histone H3 lysine 79 methyltransferase [64]. DOT1L inhibitors were developed to inhibit leukemogenic transcription programs driven by MLL1-fusions [65]. DOT1L inhibitors demonstrated profound anti-tumor effects in models carrying MLL1 translocations [66,67] and in solid tumor models [66,68,69], which led to the progression of EPZ-5676 to a clinical trial [70]. Encouraging anti-tumor efficacy was observed early in the trial, though the overall anti-tumor efficacy was diminished when the trial was expanded to a larger patient population. Ultimately, DOT1L remains a target of interest but warrants further investigation.

Multiple alternate strategies were also pursued to inhibit the MLL1 fusion leukemogenic transcription, including inhibitors of Menin, WD repeat-containing protein 5 (WDR5) and MLL1 [71–74]. The most advanced of these efforts is the development of Menin inhibitors to block protein–protein interaction between MLL1 and Menin [75]. The N-terminal of the MLL1 protein contains a domain that interacts with Menin, a protein that serves as a link between MLL1 and the chromatin-binding protein lens epithelium-derived growth factor (LEDGF). The association of MLL1/Menin/LEDGF is required for the survival of leukemic cells [62,63]. Menin inhibitors potently suppress Hox and Meis1 expression and inhibit leukemic cell survival [75,76]. Multiple Menin inhibitors are now in clinical development with very promising anti-tumor efficacy observed (Table 1). While the initial proof-of-concept was demonstrated in MLL1-re-arranged leukemia, it was also found that leukemia with mutations in the nucleophosmin (NPM1) gene are also susceptible to Menin inhibition [75]. Disruption of Menin-MLL1 interaction induced growth arrest, myeloid cell differentiation and downregulation of Hox gene transcription [77]. Clinical studies with ziftomenib (KO-539) and revumenib (SNDX-5613), for which phase I clinical trial data were reported, demonstrated very encouraging efficacy in both MLL1r and NPM1mut leukemia [78]. This is now a highly active area of clinical development, and Menin inhibitors hold the promise to transform the clinical practice of MLL1r and NPM1mut leukemia.

The H3K36 histone methyltransferase NSD2 (also known as WHSC1 or MMSET) is a key oncogenic driver in multiple myeloma (MM) that carry t(4;14) translocation [39,79]. Global genomic profiling in the cancer cell line encyclopedia (CCLE) efforts also revealed hotspot, gain-of-function NSD2 mutations in pediatric acute lymphoblastic leukemia as well as in various solid tumors [40]. Cancers carrying NSD2 oncogenic mutations showed global hypermethylation of H3K36me2, with concomitant global hypomethylation of H3K27me3. Depletion of NSD2 by shRNA showed profound anti-tumor effects [79], validating NSD2 as an attractive drug target. A catalytic inhibitor selective against NSD2 (KTX-1001) has been developed and progressed to Phase 1 clinical trial (NCT05651932).

The success of EZH2 inhibitors in lymphoma and Menin inhibitors in leukemia is an important proof-of-concept that epigenetic targeted drugs are effective for cancer treatment. However, dominant oncogenic driver mutations of epigenetic regulators are mostly confined to hematological and rare cancers, therefore the concept of targeting oncogenic epigenetic regulators in solid cancers has been limited. Nonetheless, the success of targeting EZH2 and Menin is an important example of anti-tumor activity and provides important clinical data on toxicity and tolerability for this class of drugs. It is worth mentioning that EZH2 and Menin have a much broader roles in transcriptional regulation, with numerous epigenetic centered hypotheses for their contribution to disease progression in solid cancers. Building on their initial success in lymphoma and leukemia, these agents have now expanded their clinical testing in solid cancers mostly in combination with other drugs.

2.2.2. Synthetic lethality

The cancer genome sequencing project unveiled widespread genetic mutations of epigenetic regulators across all cancers. Loss-of-function and tumor suppressor mutations, including deletions, truncations, or damaging point mutations, are particularly prevalent. Chromatin remodeling proteins such as DNA and histone modifiers are mutated in every cancer type surveyed (Figure 2(a)) [84–87]. In some cases, mutations in epigenetic regulators alone are sufficient to drive tumorigenesis with high penetrance. For example, biallelic inactivation of SMARCB1 occurs in 95% of cases of malignant rhabdoid tumor (MRT) [88,89]. Unlike oncogene drivers which can be directly targeted with inhibitors, tumor suppressors which are lost in cancers cannot be directly drugged. A deep understanding of the downstream biological pathways can provide an avenue to target loss-of-function tumor suppressor pathways. For example, mutations in Tet2 and DNMT3A are known to occur early in hematopoietic progenitor cells, contributing to clonal hematopoiesis [90] and ultimately leukemia development [91–93]. With the understanding of Tet2 function in facilitating cytosine demethylation, Tet2 mutation has been associated with greater likelihood of responses to DNMTi in myeloid dysplastic syndromes (MDS) and chronic myelomonocytic leukemia (CMML) [94]. Similarly, mutations in DNMT3A and IDH1/2 status may predict response in MDS and AML [95]. However, in the absence of such mechanisms, alternative drug discovery strategies are needed to target tumor suppressor mutations, and in recent years, synthetic lethality has been an area of intense focus for epigenetic drug discovery.

Figure 2.

Synthetic lethality approach to target cancers carrying mutations in epigenetic regulators. (a) Across all human cancers, there is a high prevalence of mutations in epigenetic genes. Mutation frequency of top 10 epigenetic regulators (green) were plotted comparing to tumor suppressor TP53 (purple) (source: cBioportal for cancer genomics pan-cancer database). (b) Synthetic lethality (SL) is defined as a circumstance in which the simultaneous mutation or inhibition of two genes is lethal, but the singular mutation or inhibition of either one is viable [80,81]. Identifying synthetic lethality relationships in cancers provides the possibility of developing therapeutics that selectively kill cancer cells harboring specific mutations, whilst sparing the wild-type normal cells. Exploiting synthetic lethality may provide a superior therapeutic index by reducing on-target toxicity. (c) SL relationships can be identified through a variety of experimental approaches. Large scale screening approaches utilizing both genetic tools (CRISPR or RNAi) and small molecule drug libraries can be explored. Such screens can be performed using large panel of cancer cell lines, or in paired isogenic knockout cell lines [82,83]. Created in BioRender. Vatapalli, R. (2024) BioRender.com/g88s208.

The concept of synthetic lethality can be illustrated by a pair of genes A and B, in which damaging mutations or deletion of gene A in cancer cells leads to epigenetic rewiring that results in a selective dependency on gene B, making gene B an attractive therapeutic target for cancers that lack gene A function [96]. Given that normal cells have both gene A and B, they can often tolerate the on-target impact targeting gene B. As such, cancer therapeutics based on the synthetic lethal concept are expected to have favorable therapeutic index by taking advantage of biological redundancy (Figure 2). With the rapid advances in large, genome-wide scale functional genomic screens using shRNA and CRISPR [82,83], together with comprehensive annotation of cancer cell lines by large consortium such as Sanger and Cancer Cell Line Encyclopedia (CCLE), these capabilities have enabled a systematic and comprehensive approach to uncover highly robust and specific synthetic lethal vulnerabilities for tumor suppressor mutations [83,96].

FDA approval of tazemetostat for patients with advanced epithelial sarcomas (ES) is the first successful example where synthetic lethality rationale is applied to target loss-of-function of an epigenetic regulator [58,89,97]. During development, gene expression states are tightly regulated during cell fate decision. Two opposing groups of epigenetic regulators, the Polycomb (PcG) protein which generally maintain gene repression and the Trithorax group (TrxG) proteins which generally activate gene expression, are vital to establish and maintain these stable and heritable gene expression states. The SWItch/Sucrose Non-Fermentable (SWI/SNF) chromatin remodeling complex belongs to the crucial TrxG family proteins. Mutations of the SWI/SNF complex proteins, including SMARCB1, ARID1A, SMARCA2/4, occur in ~20% of all cancers [98,99]. It was reported that SWI/SNF complex has an antagonistic relationship with EZH2 or PRC2, which belongs to the PcG group of proteins. Biallelic deletion or mutation of SMARCB1 is a defining hallmark in malignant rhabdoid tumor (MRT) and Ewing sarcoma (ES) [100]. MRT and ES are extremely aggressive cancers that respond poorly to existing therapies. Given the antagonistic relationship of SWI/SNF and PRC2, a series of very elegant preclinical studies [89] demonstrated that EZH2 and/or PRC2 inhibitors have strong anti-tumor activity in MRT and ES. In January 2020, tazemetostat received the FDA accelerated approval for the treatment of adults and children aged 16 years and older with metastatic or locally advanced ES not eligible for surgical resection [52]. The clinical activity of tazemetostat in ES is modest compared to follicular lymphoma, though this represents a major breakthrough in the development of epigenetic therapeutics to leverage the synthetic lethality concept and is the first approval of an epigenetic drug in solid cancers [96].

The Depmap consortium (https://depmap.org/portal/) has now generated CRISPR and shRNA cell line screening data which are publicly available for identification of novel synthetic lethal targets [83]. One exciting finding is the collateral vulnerability to the protein arginine methyltransferase PRMT5 in cancer models that harbor loss of methylthioadenosine phosphorylase (MTAP), which is frequently co-deleted with the tumor suppressor gene CDKN2A [101]. Genetic mutations of MTAP occur in 10–15% of all cancers. MTAP is a critical enzyme that plays a key role in polyamine metabolism and is important for both the methionine and adenine salvage pathway. It is also the only enzyme in mammalian cells known to catalyze the metabolism of methylthioadenosine (MTA). When the MTAP gene is deleted in cancer, MTA accumulates in cells and has partial inhibitory activity on PRMT5, which functions to suppress symmetric dimethylarginine (SDMA) levels. Thus, cancers with MTAP loss are susceptible to PRMT5 inhibition. First-generation PRMT5 inhibitors, including both SAM competitive and uncompetitive inhibitors, were rapidly progressed to clinical trials, though no meaningful clinical efficacy was observed due to dose-limiting hematological toxicities [102]. A major breakthrough has been the development of second generation, MTA cooperative PRMT5 inhibitors that realize the full potential of synthetic lethality in MTAP-deficient cancers. This is now an exciting area with multiple second-generation PRMT5 inhibitors, including MRTX1719, AMG193, TNG908, TNG462 and AZD3470 in clinical trials. MTA is a SAM mimetic and inhibits PRMT5 both in vitro and in cells [102]. When PRMT5 is MTA bound, it causes subtle conformational change of the Glu435 sidechain compared to the SAM-bound PRMT5 state. Remarkably, these subtle structural differences can be leveraged to develop MTA-cooperative PRMT5 inhibitors, which exploit the intracellular concentration differential of MTA and SAM in MTAP-deficient versus MTAP-wildtype cells, selectively killing cancer cells that have lost MTAP protein expression. Normal cells, such as hematopoietic cells, express MTAP protein and maintain a lower level of MTA::SAM ratio and are expected to have minimal levels of PRMT5 inhibited by the MTA cooperative inhibitor, therefore mitigating the hematological toxicities observed with the first-generation inhibitors. Preliminary data from MRTX1719 and AMG193 clinical studies demonstrated very promising safety and efficacy profile. Importantly, no dose limiting hematological toxicities were observed in patients. Altogether, this elegant drug discovery approach for the first time demonstrated that the differential cellular MTA:SAM ratio can be leveraged to create tumor-specific targeting of PRMT5 to maximize selective tumor cell killing. MTAP is widely lost in many solid cancers, including glioblastoma, mesothelioma, lung cancer, bladder cancer, gastric and esophageal cancers, pancreatic cancer, melanoma, and head and neck cancers. An MTA cooperative PRMT5 inhibitor is expected to have an exciting potential in a broad range of solid cancers, hopefully becoming the first targeted therapy for CDKN2A/MTAP negative cancers.

Another form of synthetic lethality commonly seen with epigenetic regulators is paralog synthetic lethality, which is best exemplified within the SWI/SNF chromatin remodeling complex. ARID1A/ARID1B, and SMARCA2/SMARCA4 are two pairs of paralogs which are incorporated into the SWI/SNF complex in a mutually exclusive manner [103,104]. When one of the paralogs is lost, SWI/SNF complexes utilize the remaining paralog to conduct its chromatin remodeling function, driving exclusive sensitivity to the perturbation of the remaining paralog. This paralog dependency relationship has been independently validated by several groups as well as in large-scale functional genomic studies [105–107]. Taking SMARCA2 and SMARCA4 as an example, SMARCA2 and SMARCA4 are ATPase subunits of the SWI/SNF complex. SMARCA4 loss due to mutations and deletions is frequently observed in non-small cell lung cancers and other solid tumors [108]. Oike and colleagues showed that SMARCA2 depletion by shRNA can selectively inhibit the growth of SMARCA4 deficient cells both in vitro and in vivo while sparing SMARCA4 wild-type models [106]. In addition to the ATPase domain, SMARCA2 and SMARCA4 also harbor a druggable bromodomain. Rescue experiments using SMARCA4 expression vectors with either ATPase-dead mutant or bromodomain mutation showed that the bromodomain mutant was able to completely rescue the growth phenotype upon SMARCA2 depletion, but an ATPase dead mutant was not able to. Similarly, bromodomain inhibition was insufficient to inhibit the growth and survival of SMARCA4 deficient models, indicating that the synthetic lethality phenotype is dependent on the ATPase but not the bromodomain function of the proteins [105].

The paralog dependency between SMARCA2 and SMARCA4 has attracted enormous interest to selectively target SMARCA2 for SMARCA4-deficient cancers. One of the efforts is through directly targeting the SMARCA2 ATPase function. However, the highly homologous nature between SMARCA2 and SMARCA4 ATPase domains poses a great challenge. A highly selective SMARCA2 inhibitor is desirable to achieve maximal therapeutic index. Several groups have reported an alternative approach which utilizes a SMARCA2/4 nonselective bromodomain binder and developed a SMARCA2 selective heterobifunctional degrader (also known as PROTACs) [109–111]. Both SMARCA2 selective ATPase inhibitor and selective SMARCA2 PROTACs have been reported [112,113]. Based on very promising pre-clinical data, a few of these molecules have advanced to early stages of clinical development (NCT05639751) [114].

SMARCA2 is among one of the most advanced paralog synthetic lethality-based targets for epigenetic regulators. There are many more synthetic lethal epigenetic targets that are being actively pursued by biotech and pharmaceutical companies, including ARID1A/B, EP300/CBP, DDX3X/3Y [115–117]. The concept of pursuing synthetic lethality for novel epigenetic targets is an area of active research and please refer to Table 2 for agents currently in clinical investigations.

Table 2.

Epigenetic inhibitors targeting synthetic lethal relationships in clinical development.

Epigenetic alteration	Target	Inhibitor	Indication	Rationale	Clinical trial number
ARID1A mutation/LOF	EZH2	Tazemetostat	Ovarian cancer	epigenetic antagonism between SWI/SNF and PcG (Polycomb group protein) [118]	NCT05023655, NCT03348631
BAP1 loss	EZH2	Tazemetostat	Mesothelioma	decrease in H4K20 monomethylation, increase in EZH2 expression [119]	NCT02860286
SMARCA4 mutation/LOF	SMARCA2	PRT-3789 PRT-7732 FHD-909 LY4050784	Solid tumors	paralog dependency in SWI/SNF [106,107]	NCT05639751 NCT06560645 NCT06561685
SYT-SSX	EZH2	Tazemetostat	Synovial sarcoma	epigenetic antagonism between PcG and SWI/SNF [120]	NCT02601937, NCT02601950
SMARCB1	EZH2	Tazemetostat	malignant rhabdoid tumors (MRT) and atypical teratoid/rhabdoid tumors (ATRT)	epigenetic antagonism between PcG and SWI/SNF [97,121]	NCT03213665, NCT02601950
SMARCB1	HDAC	Vorinostat	MRT and ATRT	restore histone acetylation in SMARCB1-null cells [122]	NCT04897880
CREBBP mutation/LOF	p300	CCS-1477	Lung cancer, Heme cancer	paralog dependency [117]	NCT04068597
MTAP	PRMT5	AMG-193, TNG-908, MRTX-1719 TNG-462 AZD-3470	Multiple cancers	preferential PRMT5 inhibition by MTA in MTAP deleted cancer [101,102,123]	NCT05245500, NCT05094336, NCT05275478, NCT05732831 NCT06130553 NCT06137144

2.3. Combination therapy

Building on the success of monotherapy activity of HDAC and DNMT inhibitors, combinations of HDAC or DNMT inhibitors with other therapies have been extensively tested in the clinical setting. The most notable success was the combination of DNMTi with cell death agent venetoclax for the treatment of AML. Low-intensity regimen of venetoclax combined with decitabine or azacitidine demonstrated good efficacy and a tolerable safety profile in elderly patients with AML unfit for intensive chemotherapy [124]. There are active clinical trials to explore DNMTi in combination with other agents to further extend the benefit of AML treatment, including Fms-like kinase 3 (FLT3) inhibitor gilteritinib (FLT3 inhibitor) plus azacitidine (NCT02752035) for AML with FLT3 mutation and ADI-PEG 20 (pegylated arginine deiminase) in combination with venetoclax and azacitidine triple combination for AML. Effective use of epigenetic drugs in combination setting is an area of active investigation. In the next session, we will discuss a few combination strategies and current preclinical and clinical activities exploring these opportunities.

2.3.1. Combination therapy with epigenetic agents to inhibit the oncogenic transcriptional programs

One of the most successful concepts in oncology therapy is to target lineage-dependent transcription programs, such as estrogen receptor (ER) mediated transcription in breast cancer and androgen receptor (AR) mediated transcription in prostate cancer. Epigenetic regulation plays a critical role in transcription activation and is often hijacked in the cancer setting to fuel constitutive transcriptional activities driving cancer progression. It is therefore rational to consider combining epigenetic targeted agents with other targeted drugs to suppress oncogenic transcription programs.

Several recent studies reported that EZH2 expression is upregulated in hormone-refractory metastatic prostate cancers and high EZH2 expression levels are correlated with disease stage, and poor outcome in prostate cancer patients [125,126]. In preclinical models, combining EZH2 inhibitors with enzalutamide synergistically inhibits cell proliferation and colony formation and promotes apoptosis in enzalutamide-resistant prostate cancer cells [127]. Mechanistically, EZH2 functions both as a transcription corepressor and coactivator [128]. In mouse models, EZH2 inhibition restrains tumor neuroendocrine differentiation [125,127,129]. EZH2 inhibition increases cytotoxic activity and IFN-γ production of tumor-specific CD8⁺ T lymphocytes [130], highlighting the multi-faceted opportunity to re-sensitize tumors to androgen targeted therapies by inhibiting EZH2. Currently, three EZH2 inhibitors have progressed to clinical testing in Metastatic Castration-Resistant Prostate Cancer for combination with enzalutamide or other androgen deprivation therapies (NCT03480646, NCT04179864, NCT03460977) [131–133].

KAT6A and KAT6B are members of the MYST family of histone acetyltransferases. Dysregulation of MYST family HATs has been implicated in the activation of oncogenic gene expression programs in human cancers [134]. KAT6A was found to be amplified and/or overexpressed in 10–15% of breast cancers and shown to be a significant dependency in breast cancer cell lines overexpressing KAT6A [135]. KAT6A shRNA-mediated knockdown reduced ERα levels in ER+ breast cancer cells, highlighting a role for KAT6A in regulating ERα-mediated growth of breast cancer cells [135]. These findings supported further investigation of the clinical utility of targeting KAT6A in ER+ breast cancers. A first-in-class KAT6A/B inhibitor PF-07248144 is being tested as a single agent and in combination with either fulvestrant or letrozole + palbociclib in advanced solid tumors (NCT04606446) [136].

2.3.2. Epigenetic therapy in combination with immune checkpoint therapies (ICTs)

There is an increasing amount of the literature supporting the idea of combining epigenetic therapies with ICTs to enhance their antitumor effects. There are many biological rationales proposed, including a) increase in neoantigen expression; b) increase in antigen presentation; c) rejuvenation of exhausted T-cells; d) increase in T cell differentiation and memory; e) induction of other immune cells including dendritic cells and NK cells; f) activation of interferons [96,137–139]. A number of clinical studies have been initiated to explore the benefit of combining epigenetic drugs with ICTs (see Table 3). In this section, we review a few selected examples to demonstrate the key principles and learnings.

Table 3.

Epigenetic inhibitors tested in the clinic in combination with immunotherapies (active trials).

Epi- therapy	Immunotherapy		Indication	Clinical trial Number
	Vaccine	Checkpoint therapy
Demethylation agents (DNMTi) Azacytidine, Decitabine	Allogeneic colon cancer cell vaccine (GVAX) and cyclophosphamide		Metastatic colon cancer	NCT01966289
	DEC-205/NY-ESO-1 Fusion Protein CDX-1401		MDS/AML	NCT03358719
		Pembrolizumab	Metastatic melanoma, NSCLC	NCT02816021, NCT02546986
		Nivolumab and/or Ipilimumab	MDS	NCT02530463
HDACi ACY-241, Entinostat, ZEN-3694		Nivolumab and Ipilimumab	Metastatic unresectable HER2-negative breast cancer, NSCLC, Solids	NCT02453620 NCT02635061 NCT04840589
		Pembrolizumab	Metastatic CRPC	NCT04471974
EZH2i CPI-1205, DS-3201		Pembrolizumab	Advanced urothelial cancer, NSCLC, Head & Neck	NCT03854474 NCT05879484
		Ipilimumab	Metastatis Prostate, Urothelial and Renal cell cancers	NCT04388852
		Atezolizumab and Bevacizumab	Hepatocellular Carcinoma	NCT06294548
		Rituximab	Follicular Lymphoma	NCT05683171
		SHR1701	R/R cHL, Advanced solids R/R BC NHL	NCT05896046 NCT04407741
BETi INCB057643, BMS986158		Pembrolizumab + Epacadostat (IDO1 inhibitor)	Advanced solid cancers	NCT02959437
		Ruxolitinib or Fedratinib	Myelofibrosis	NCT04817007
LSD1i ORY-1001,IMG-7289		Atezolizumab	SCLC	NCT06287775 NCT05191797

Seminal work from Baylin and colleagues demonstrated the role of DNA methylation inhibitors, such as azacytidine, in increasing antigen presentation and major histocompatibility complex I (MHC-I) expression, thereby augmenting antitumor immune responses [140–143]. DNMTi induces double-stranded RNA (dsRNA) production through reactivation of transposable elements like endogenous retroviruses (ERVs), long interspersed retrotransposable elements (LINEs) and short interspersed retrotransposable elements (SINEs) and inverted-repeat Alu elements. Elevated levels of dsRNA are sensed by toll-like receptor 3 (TLR3), RNA helicases retinoic acid-inducible gene protein I (RIG-I) or melanoma differentiation-associated protein 5 (MDA5) mediated pathway to activate Type I interferon response [144]. DNMTis were also shown to reverse immune cell exhaustion and restore anti-tumor responses [145–147]. These preclinical studies have led to the excitement to combine DNMTi with ICTs in clinical trials. When Hodgkin Lymphoma patients were treated with programmed cell death protein 1 (PD-1) antibody camrelizumab in combination with decitabine (NCT02961101), there was a meaningful improvement in both complete response (CR) rates (71%) and progression-free survival (PFS) (100%) when compared to PD-1 monotherapy (32% and 76% respectively) [148]. It will be interesting to see if newer DNMTi will bring broader impacts to enhance ICTs in different cancer types.

Lysine-specific demethylase 1 (LSD1), a histone demethylase, was reported to enhance the efficacy of immunotherapy by acting on both the tumor intrinsic and immune related pathways [149]. In tumor cells, LSD1 inhibition can lead to an increased antigen presentation [150]. More importantly, LSD1 is also involved in the development and regulation of T cells, macrophages, cancer-associated fibroblasts (CAFs) and NK cells [151–154]. With LSD1 inhibitors already in the clinic, these preclinical findings have led to the initiation of several clinical trials that combine LSD1 inhibitors with immune checkpoint inhibitors [155–157].

Functional genomic screens continue to reveal new epigenetic targets that can enhance immune-mediated anti-tumor response. An epigenetic-focused CRISPR/Cas9 screen in chronically stimulated mouse CD8+ T cells revealed that SWI/SNF chromatin remodeling complex components, including ARID1A, DPF2, SMARCC1, and SMARCA4 were among the top scoring hits depleted in the PD1+TIM3+ T cells, highlighting a role of SWI/SNF in regulating T cell exhaustion [158]. Confirmatory studies using single gRNA mediated knockdown of SMARCA4, SMARCC1, and DPF2 validated findings from the primary screen. Pharmacological inhibition of SWI/SNF using SMARCA4/2 ATPase inhibitor or PROTAC degrader resulted in similar decrease in PD1+TIM3+ population and decrease in terminal T cell exhaustion marker CD39+ in multiple donors while increasing the effector T cell pool. Transcriptomics and epigenomic analysis showed that key hallmark genes of exhaustion including TOX, ENTPD1, ITGA2, and TIGIT were directly bound and regulated by the SWI/SNF complex. In an OVA antigen expressing B16 melanoma model, the pretreatment of CD8+ OT-1 T cells with the SMARCA4/2 ATPase inhibitor enhanced their in vivo anti-tumor activity. In another example, genome-wide CRISPR/Cas9 study identified EZH2 as an important player in mediating transcriptional silencing of MHC-I antigen processing pathway to promote immune evasion of T-cell mediated immunity [159]. EZH2 inhibitor can reverse epigenetic silencing of MHC-I antigen processing pathway to re-establish T cell-mediated immunity. Kim et al. has comprehensively reviewed the multi-faceted role of EZH2 in overcoming resistance to immune evasion [160].

In addition to the above examples, there is abundant literature demonstrating that targeting epigenetic regulators, including HDACs, G9a, SETDB1, ADAR1 can enhance ICTs. Here, we also caution that current preclinical models carry major challenges and limitations and might not adequately inform the complex immune-mediated anti-tumor mechanisms nor the complex tumor microenvironment in patients. Therefore, findings from preclinical models might not translate in clinical trials. For example, the HDAC inhibitor Entinostat was shown to have strong anti-tumor effects when combined with ICTs in syngeneic breast and pancreatic cancer mouse models [161]. However, in a trial (NCT02708680) testing atezolizumab in combination with entinostat, no significant benefit was observed [162]. Entinostat also did not show meaningful clinical benefit in the ENCORE 601 trial for non-small cell lung cancer (NSCLC) in combination with PD-1 inhibitor pembrolizumab [163]. To drive the success of rational combination therapies, a thorough mechanistic understanding of each drug’s biological impact on immune and tumor cells is needed.

Reverse translation from clinical experience could be a better path forward to inform biological hypotheses with epigenetic regulation and immune-mediated anti-tumor mechanisms. For example, Goswami et al. [164] observed that ipilimumab increases EZH2 expression in human T cells across various tumor indications and the increased EZH2 expression in T cells inversely correlates with clinical outcome. Following the initial clinical observation, preclinical models were developed and validated the hypothesis that pharmacological inhibition of EZH2 in combination with ipilimumab enhances effector-like T cell responses, providing a strong rationale for testing the combination of EZH2 inhibitor with ipilimumab in the clinic. This led to the initiation of a clinical trial combining EZH1/2 inhibitor (DS3201) with ipilimumab (NCT04388852). In these ongoing clinical trials, comprehensive translational biomarker data needs to be collected to inform how EZH2 inhibition impacts immune cells, tumor cells, and stromal cells. Findings from patient tumors can then be used to develop new hypotheses and assessed in the preclinical setting.

2.3.3. Epigenetics and therapy resistance

The development of drug resistance is a major challenge to the long-term effectiveness of cancer therapies. In addition to on-target genetic mutation causing drug resistance, studies with acquired resistance models also revealed significant contribution of epigenetic reprogramming to the development and maintenance of resistance states, especially in models that lack clear genetic driver mechanisms [165,166]. Drug resistance can be mediated by a subpopulation of cells that persist after treatment, also known as “Drug Tolerant Persisters” (DTPs) [167]. DTPs have stem cell-like features with a high degree of epigenetic plasticity. For instance, EGFR inhibitor-induced DTPs were found to highly express KDM5A. Inhibition of KDM5A or treatment with HDACi led to reduced survival of DTPs upon EGFRi treatment [168]. Profiling of osimertinib DTPs using RNA-seq and ATAC-seq identified consistent activation of the TEAD pathway [169]. These osimertinib DTPs were shown to be sensitive to BRD4 or TEAD inactivation. There is emerging evidence that DTPs adapt to therapeutic intervention through epigenomic modifications, transcriptomic regulation, metabolic remodeling, and altered interactions with the tumor microenvironment [170,171].

de Miguel et al. profiled chromatin accessibility in five pairs of parental and osimertinib resistant lung cancer models and showed that osimertinib-resistant cells harbor a differentiated chromatin accessibility pattern compared to parental cells [172]. Chromatin accessibility gains are enriched in promoters and enhancer regions for target genes associated with upregulation of EMT and receptor tyrosine kinase (RTK) signaling pathways, and downregulation of epithelial cell differentiation and cell–cell adhesion. A diverse set of resistance-associated transcriptional programs were observed, underscoring the heterogeneity of the resistant state. In search of chromatin associated factors that govern these changes, SWI/SNF chromatin remodeling complex component SMARCA4 and SWI/SNF associated transcription factors including TP53, SOX2 were identified. The contribution of SMARCA4 in driving the resistance states was further substantiated by concordant change in SWI/SNF chromatin occupancy in the enhancer regions of the differentially expressed genes. Osimertinib resistance can be reversed in a subset of models by SMARCA4 inhibition. Moreover, SMARCA4 perturbation can deepen the response of patient-derived xenograft models to Osimertinib.

Epigenetic alterations can drive pro-survival signaling, DNA repair, expression of drug efflux transporters, promote EMT, and reprogram metabolic pathways to cause drug resistance [173]. In such cases, utilizing a mechanism-based combinatorial approach including epigenetic therapies to overcome drug resistance is an attractive approach. The concept of epigenetic “priming” to re-sensitize resistant tumors to conventional chemotherapy targeted therapies or immunotherapies has been gaining great attention. One of the mechanisms by which pretreatment or “epigenetic-priming” increases the efficacy of the combination partner is through the reactivation of tumor-suppressor genes. For example, DLBCL patients treated with low-dose azacytidine showed reactivation of tumor suppressor SMAD1 and were re-sensitized to doxorubicin treatment leading to a higher remission rate [174].

While there are ample preclinical examples supporting the rationale of combining with epigenetic therapy to combat or prevent resistance, we are not aware of any successes in clinical trials. The main challenge is that unlike genetic-based resistance mechanisms, it is still not clear if epigenetic mechanisms of resistance such as DTPs occur in human cancer. With the advance of single-cell sequencing and improved ability to perform biomarker analysis using clinical samples, it will be important to clearly demonstrate the specific “epigenetic alterations” driving therapy resistance in cancer patients.

2.4. Novel modalities for targeting epigenetic factors

In the last decade, targeted protein degradation including PROTAC and Molecular Glue (MG) degraders has gained interest as novel drugging modalities [175,176]. These molecules are able to induce target protein degradation mimicking the effects of genetic knockdown and have demonstrated differential and in some cases superior activity compared to traditional catalytic inhibitors. These approaches are particularly relevant to epigenetic drug discovery as many epigenetic regulators have protein complex scaffolding functions beyond enzymatic activities.

There is increasing evidence from the studies of Menin/MLL inhibitors and EED/EZH2 inhibitors that protein degraders can further improve therapeutic efficacy [177–179]. Since 2021, multiple protein degraders have entered clinical trials, including SMARCA2 (PRT3789 [NCT05639751] and PRT7732 [NCT06560645]) degraders. In the case of selective SMARCA2 PROTACs, a nonselective SMARCA2/4 dual binder against the bromodomain was linked to an E3 ubiquitin ligase warhead. Selectivity toward SMARCA2 was engineered by optimizing the ternary structure between target protein and E3 ligase. Utilizing the PROTAC modality to improve selectivity and druggability is an exciting avenue for the development of more effective epigenetic therapeutics.

Another novel targeting modality that holds great potential is molecular glue approach. Molecular glues can promote protein–protein interaction, and in many cases molecular glues are designed to enhance the interaction between a target protein and an E3 ligase to promote target protein degradation. In 2019, Blake et al. reported the development of a BRD4 monovalent degrader GNE-0011, which recruited DCAF16 as the E3 ligase to mediate its degradation activity [180,181]. By applying a transposable covalent handle to existing binders, Toriki et al. reported HDAC1/3 and SMARCA2/4 molecular glues that recruit RNF126 to mediate the degradation effects. While no molecular glue targeting an epigenetic driver has progressed into clinical testing yet, we expect breakthroughs in this field can lead to more effective epigenetic drugs with improved potency and selectivity in the near future.

3. Conclusions

Cancer epigenetic drug discovery is a rapidly evolving field that holds great promise for the development of more effective and targeted therapies. Advances in cancer epigenetic research have already led to the discovery of epigenetic biomarkers that can be used for early cancer detection, risk assessment, and monitoring of treatment response. With the advancement of novel drugging approaches, diverse types of small-molecule inhibitors, including catalytic inhibitors, protein–protein interaction inhibitors, and protein degraders, can be designed for specific targets. The FDA approval of DNMT, HDAC, EZH2, and IDH1/2 inhibitors has demonstrated that targeting epigenetic mechanisms can be effective in cancer therapy. We are particularly encouraged by the increased pace and interest in the clinical development of many epigenetic agents (Tables 1–3). Due to space limitations, we have focused the discussion on epigenetics targeted agents under active clinical investigation, but we expect that the next wave of epigenetic agents to enter the clinic will have broad impacts in many if not all cancers. Epigenetic drugs when used in combination with other treatment modalities, such as chemotherapy, radiation therapy, or immunotherapy, could enhance treatment response and overcome drug resistance. The exciting field of cancer epigenetics holds tremendous potential for revolutionizing the cancer treatment landscape in the coming years.

4. Future perspectives

The future of cancer epigenetic drug discovery is poised to benefit significantly from advancements in drug discovery and epigenomic sequencing technologies. First, the development of new drugging modalities, such as PROTACs, molecular glue and covalent drugging strategies, holds great promise to develop more effective drugs targeting epigenetic regulators. These new capabilities have the potential to target previously “undruggable” epigenetic regulators, expanding the repertoire of targets. Second, by mapping the epigenomic landscape of different cancer types, we can now more precisely define different cancer subtypes by combining genetic and epigenomic annotations. The integration of single-cell epigenomics and multi-omics approaches will further enhance our understanding of the dynamic nature of the cancer cell states and reveal additional novel cancer-specific aberrations and vulnerabilities. This, together with combinatorial CRISPR screens in preclinical models, will help uncover the next wave of epigenetic targets for cancer therapies. In addition, cancer-specific metabolic states should be considered during drug design and optimization to maximize cancer targeting while sparing non-cancerous tissues. Last but not the least, reverse translation using clinical observations to inform preclinical research will play a pivotal role in developing mechanism-based combination strategies with epigenetic drugs. Here, the ability to apply new epigenomic sequencing technologies, including single-cell sequencing, to patient tumor samples will transform our understanding of tumor epigenetic state in “real-time.” This deeper insight into epigenomic dysregulation in cancer patients will provide more precision in guiding mechanism-based combination strategies.

Funding Statement

This paper was not funded.

Article highlights

Cancer epigenetic drug discovery

• Dysregulated epigenetic landscape is a hallmark of all cancers and can be therapeutically targeted.

• FDA approval of inhibitors against DNMTs, HDACs, and EZH2 showed initial success in several cancer indications.

Precision targeted epigenetic therapeutics

• Precision-targeted epigenetic therapeutics will have broader impact to all cancers.

• Genetic mutations of epigenetic regulators are common in all cancers and offer new opportunities to develop epigenetic drugs targeting these newly defined cancer segments.

• New generation of targeted epigenetic drugs based on epigenetic oncogene drivers and synthetic lethality, such as inhibitors against PRMT5 and Menin, is showing great promise in clinical studies.

Combination Therapy

• Mechanism-based combination strategies will realize the full potential of epigenetic drugs.

• Epigenetic therapy can enhance and deepen the therapeutic benefits of immune checkpoint therapies.

• Epigenetic therapy can be used to combat therapy resistance.

Disclosure statement

All authors are employees of AstraZeneca. Alex Rossi is now an employee of Flare therapeutics.

The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript apart from those disclosed.

No writing assistance was utilized in the production of this manuscript.

Author contributions

Rajita Vatapalli, Ho Man Chan, Alex P. Rossi, Jingwen Zhang wrote the main manuscript text. Rajita Vatapalli and Alex P. Rossi made the tables and figures.