# Mechanisms of Polycomb group protein function in cancer

Victoria Parreno, Anne-Marie Martinez, Giacomo Cavalli

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Victoria Parreno, Anne-Marie Martinez, Giacomo Cavalli. Mechanisms of Polycomb group protein function in cancer. Cell Research, 2022, 32 (3), pp.231-253. 10.1038/s41422-021-00606-6. hal-03805077

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# Mechanisms of Polycomb group protein function in cancer

Victoria Parreno<fnref n="1" />, Anne-Marie Martinez<fnref n="1" />✉ and Giacomo Cavalli ID<fnref n="1" />✉

© The Author(s) 2022

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Cancer arises from a multitude of disorders resulting in loss of differentiation and a stem cell-like phenotype characterized by uncontrolled growth. Polycomb Group (PcG) proteins are members of multiprotein complexes that are highly conserved throughout evolution. Historically, they have been described as essential for maintaining epigenetic cellular memory by locking homeotic genes in a transcriptionally repressed state. What was initially thought to be a function restricted to a few target genes, subsequently turned out to be of much broader relevance, since the main role of PcG complexes is to ensure a dynamically choreographed spatio-temporal regulation of their numerous target genes during development. Their ability to modify chromatin landscapes and refine the expression of master genes controlling major switches in cellular decisions under physiological conditions is often misregulated in tumors. Surprisingly, their functional implication in the initiation and progression of cancer may be either dependent on Polycomb complexes, or specific for a subunit that acts independently of other PcG members. In this review, we describe how misregulated Polycomb proteins play a pleiotropic role in cancer by altering a broad spectrum of biological processes such as the proliferation-differentiation balance, metabolism and the immune response, all of which are crucial in tumor progression. We also illustrate how interfering with PcG functions can provide a powerful strategy to counter tumor progression.

Cell Research (2022) 32:231–253; https://doi.org/10.1038/s41422-021-00606-6

## INTRODUCTION

Polycomb Group (PcG) proteins have first been described as main players in cellular memory known to maintain embryonic chromatin landscapes in a repressed transcriptional state throughout development. Counterintuitively, PcG proteins then appeared to be able to regulate the transcription of developmental genes involved in a wide range of highly dynamic biological processes such as differentiation, stem cell plasticity or cell cycle progression.<fnref n="1" />,² In addition, mutations or dysregulations of PcG proteins have been extensively described in cancer.³ Knowing the importance of PcG proteins in transcriptional regulation, it was not surprising to find a correlation between modification of PcG activities and tumorigenesis. However, an early demonstration of a causal link between the ability of PcG complexes to promote or inhibit the transcription of oncogenes or tumor suppressor genes, respectively, has paved the way for work aimed at studying the different mechanisms by which PcG complexes are involved in the generation and the evolution of cancer cells.

Here, we first describe the molecular mechanisms underlying the recruitment and function of PcG proteins in gene regulation during normal development. We then review the involvement of Polycomb complexes in cancer, highlighting PcG-dependent disturbances of epigenetic processes in tumorigenesis. Next, we focus on the description of the latest discovered mechanisms linking Polycomb to cancer. PcG proteins have been extensively studied in hormone-dependent cancers where hormone-receptors interact directly with PcG proteins, modifying the transcriptional landscape of the affected cells. Furthermore, PcG proteins have been described as capable of modulating the metabolism and the

immune response of the tumor microenvironment, both being hallmarks of cancer. Next, we focus on a new area of research involving mutated histones, also known as oncohistones, and discuss how these mutations can impact PcG behaviour in a tumoral context. Finally, we explain how PcG proteins are able to confer a non-genetic drug-resistance underlying the importance of epigenetics in cancer.

## PCG PROTEINS

PcG proteins are highly conserved throughout metazoan evolution and are essential players in cellular identity. In *Drosophila melanogaster*, mutations in the *Polycomb* gene induce embryonic transformation of anterior segments into posterior segments by inducing ectopic expression of homeotic (Hox) genes.⁴,⁵ Subsequent work identified other mutations triggering derepression of Hox genes, leading to the identification of several genes that were defined as members of the Polycomb group. PcG proteins form two main epigenetic complexes, the Polycomb Repressive Complex 1 and 2 (PRC1 and PRC2), which were later identified as transcriptional regulators targeting a large number of genes in genome-wide studies.⁶

PRC2 is composed of the Embryonic Ectoderm Development (EED), Suppressor of Zeste 12 Homolog Protein (SUZ12) and Enhancer of Zeste Homolog 1/2 (EZH1/2) core constitutive subunits (Fig. 1a). EZH1/2 have a Su(var)3–9, Enhancer-of-zeste and Trithorax (SET) domain with a histone methyltransferase activity that mono-, di- or tri-methylates the lysine 27 of the histone H3 (H3K27me1/2/3).⁷ PRC2 can be divided into two sub-

<footnote n="1">¹Institute of Human Genetics, UMR 9002, CNRS–University of Montpellier, Montpellier, France. ✉email: anne-marie.martinez@igh.cnrs.fr; giacomo.cavalli@igh.cnrs.fr</footnote>
<figure><img src="image_3.png" /><figcaption>**Fig. 1** Composition of PcG proteins in mammals. **a** PRC2 can be sub-divided into PRC2.1 and PRC2.2. The core PRC2 with PCL1/2/3 and the PALI1/2 or EPOP subunits compose the PRC2.1 complex. The association between the core PRC2, JARID2 and AEBP2 constitute PRC2.2. **b** PRC1 complex can be sub-divided into two groups of complexes, namely cPRC1 and ncPRC1. cPRC1 is composed of RING1A/B associated with PCGF2/4 and CBX2/4/6–8. ncPRC1 is composed of either RYBP or YAF2 associated with one of the six PCGF proteins. The graphical representation of each complex is schematic and not aimed to represent the size, shape and relative position of the various subunits. AEBP2, Adipocyte enhancer-binding protein 2; AUTS2, Autism susceptibility candidate 2; BCOR, BCL6 corepressor; CBX2/4/6–8, Chromobox 2/4/6–8; CK2, Casein kinase 2; DCAF7, DDB1 and CUL4 associated factor 7; DP1, E2F dimerization partner 2; E2F, E2F transcription factor 6; EED, Embryonic ectoderm development; EPOP, Elongin B/C and PRC2-associated protein; EZH1/2, Enhancer of zeste homolog 1/2; FBR5, Fibrosin; HDAC1/2, Histone deacetylase 1/2; HP1, Heterochromatin protein 1 gamma (here labeled as HP1, also named CBX3); JARID2, Jumonji AT rich interactive domain 2; KDM2B, Lysine demethylase 2B; L3MBTL2, Lethal(3) malignant brain tumor-like protein 2; MAX, Myc associated factor X; MGA, MAX gene associated protein; PALI1/2, PRC2 associated LCOR isoform 1/2; PCGF1–6, Polycomb group finger 1–6; PCL1–3, Polycomb like protein 1–3; PHC1–3, Polyhomeotic-like protein 1–3; RBBP4/7, Retinoblastoma binding protein 4/7; RING1A/B, Really interesting new gene 1B/A; RYBP, RING1 and YY1 binding protein; SCMH1/2, Sex comb on midleg homolog 1/2; SKP1, S-phase kinase associated protein 1; SUZ12, Suppressor of zeste 12 protein homolog; USP7, Ubiquitin specific peptidase 7; YAF2, YY1-associated factor 2; WDR5, WD repeat domain 5.</figcaption></figure>

complexes, namely PRC2.1 and PRC2.2, characterized by the
association with specific accessory proteins. PRC2.1 contains one
of the three paralogous Polycomb-like (PCL) proteins PCL1/2/3,
also known as PHF1, MTF2, PHF19 respectively, as well as PRC2-
Associated LCOR Isoform 1/2 (PALI1/2) or Elongin B/C and PRC2-

associated Protein (EPOP). In addition of the core subunits, PRC2.2
contains Jumonji and AT-Rich Interaction Domain containing 2
(JARID2) and Adipocyte Enhancer-Binding Protein 2 (AEBP2). Some
PRC2 co-factors can have a negative impact on PRC2 methyl-
transferase activity. The Catalytic Antagonist of Polycomb
(CATACOMB)-PRC2 variant presents a decrease in PRC2 enzymatic activity. Indeed, the *CATACOMB* (also known as *EZHIP* ) gene is poorly expressed in physiological conditions, except in gonads,⁸ due to hypermethylation of its CpG islands (CGIs).⁹ While CATACOMB-PRC2 association does not impact PRC2 recruitment to chromatin, it lessens its ability to associate with sub-stoichiometric co-factors that would otherwise enhance its enzymatic activity.⁸

PRC1 members form an even more diversified combination of variant complexes (Fig. 1b), which can be subdivided into canonical PRC1 (cPRC1) and non-canonical PRC1 (ncPRC1) complexes that all share a core PRC1 comprising one of six Polycomb Group Ring Finger 1–6 (PCGF1–6) proteins and RING1A/ B, an E3 ubiquitin ligase catalyzing the mono-ubiquitination of lysine 119 of histone 2A (H2AK119ub in mammals or H2AK118ub in flies).¹⁰,¹¹ cPRC1.2 and cPRC1.4 are respectively formed by PCGF2 or PCGF4 (also known as MEL-18 or BMI-1), RING1A/B and Sex Comb on Midleg Homolog 1/Like 2 (SCMH1/L2), and can be distinguished from ncPRC1s by the presence of one of Chromo- box 2/4/6–8 (CBX2/4/6–8) proteins as well as one of the Polyhomeotic Homolog 1–3 (PHC1–3).⁶,¹² In addition to RING1A/ B and a PCGF1–6 protein, ncPRC1complexes assemble around RING1 and YY1-Binding Protein (RYBP) or YY1-Associated Factor 2 (YAF2) proteins, which are mutually exclusive homologous proteins able to bind to the same site on the C-terminal domain of RING1B.¹³,¹⁴ Moreover, the ncPRC1 complexes can be further classified by the identity of their PCGF subunit (PCGF1 for PRC1.1, PCGF2 for PRC1.2 and so on). Genome-wide analysis demon- strated that each PRC1 complex has its own chromatin targeting profile suggesting that the recruitment of cPRC1 and ncPRC1 depends on their differential compositions that in turn could contribute to pleiotropic functions.¹⁵

MOLECULAR MECHANISMS MODULATING PRC1 AND PRC2
RECRUITMENT

An important feature of the PRC1 and PRC2 core subunits is the
absence of sequence-specific DNA-binding domains that would
allow their direct recruitment to their target genes. PcG-mediated
gene regulation therefore depends on components that direct
their recruitment to specific chromatin domains. In a classical
model, described in *Drosophila melanogaster*, PRC2 is first
recruited on *cis*-regulatory sequences called Polycomb Response
Elements (PREs) via consensus motifs for sequence-specific DNA-
binding proteins that might interact with PRC2 subunits.¹⁶⁻¹⁸
PRC2, via its E(z) subunit, the *Drosophila* ortholog of EZH2/1,
deposits H3K27me3. This H3K27me3 mark is then recognized by
the cPRC1 PC subunit (ortholog of CBX).⁷,¹⁷,¹⁹,²⁰ Subsequently,
Sce — the ortholog of RING1A/B — ubiquitinates H2AK118¹⁰,¹¹
(Fig. 2a). This model predicts co-occurrence of PRC1 and PRC2 at
their target loci.

However, ncPRC1 complexes do not possess CBX subunits that
recognize H3K27me3 and only a small subset colocalizes with this
PRC2-deposited mark.¹⁵ Moreover, mammalian ncPRC1s can act
upstream of PRC2 by directly recognizing non-methylated DNA in
CGIs leading to the ubiquitination of H2AK119 which is then
recognized by PRC2-JARID2.²¹⁻²⁴ These data suggest that ncPRC1
recruitment to a subset of their targets can act upstream of PRC2
recruitment (Fig. 2a).

The existence of mammalian PREs is still controversial.²⁵,²⁶ The
analysis of PRC2 genome binding identified the enrichment for
CGIs characterized by low levels of DNA methylation, that could
therefore act as PREs in mammals²⁷⁻²⁹ (Fig. 2a). Thanks to their
Polycomb-like (PCL) extended domain, the PCL proteins PHF1,
MTF2 or PHF19 preferentially bind unmethylated CpG-containing
DNA sequences,³⁰ promote PRC2 binding to CGIs³¹,³² and stabilize
the dimerization of PRC2.³³ The accessory subunits — JARID2 and
AEBP2 — are also important for PRC2 recruitment, via recognition

of the H2AK119ub mark, as well as for deposition of H3K27me3 at
specific PcG targets³⁴ (Fig. 2a).

Additional mechanisms involve a PRC1-independent transcrip-
tional repression. Indeed, the proteins Bromo Adjacent Homology
Domain Containing protein 1 (BAHD1) and BAH Domain And
Coiled-Coil Containing 1 (BAHCC1/BAHD2) possess a C-terminal
Bromo Adjacent Homology (BAH) domain which recognizes
H3K27me3.35–38 Moreover, BAHD1 acts as a scaffold protein that
recruits additional co-repressors such as Histone DeAcetylases
(HDACs).35,39 Alternative recruiting mechanisms also involve long
non-coding RNAs (lncRNAs) as well as specific transcription factors
(reviewed respectively in 40,41) (Fig. 2a).

Polycomb recruitment is also modulated by the chromatin
landscape. Indeed, Trithorax Group (TrxG) proteins counteract
Polycomb-mediated gene silencing by decorating chromatin with
active histone marks such as H3K4me1/2/3⁶ and a fine-tuned
balance between these two complexes is critically important. SWI/
SNF and COMPASS complex subunits are the main TrxG proteins,
respectively involved in chromatin remodeling and H3K4 methy-
lation.⁴² Interestingly, MLL2/COMPASS binds specific promoters
and trimethylates H3K4 to promote MLL2-dependent gene
transcription.⁴³ Upon loss of MLL2, H3K27me3 decorates MLL2-
dependent genes and represses them.⁴³ However, in MLL2
depleting context, H3K27me3 spreading is prevented by DNA
methylation at CpG islands.⁴⁴ Dual deletion of MLL2 and DNA
methylation increases the repressive mark spreading while
diluting its level, which ultimately leads to transcription of the
corresponding genes.⁴³ Moreover, spreading of the PRC2 mark is
also counteracted by H3K36me2, which is deposited by NSD1.⁴⁵

In summary, the molecular mechanisms deployed by PcG
complexes to specifically target the genome remain a major area
of interest with important consequences for understanding how
target genes are specified. Coordinating Polycomb action with key
developmental orchestrators, including transcription factors,
involves a wide spectrum of tissue- and time-specific players.
Future studies should provide insight into this complex Polycomb
recruitment network.

PCG PROTEIN FUNCTION IN GENE SILENCING

PcG-mediated transcriptional regulation has been widely por-
trayed as gene silencing and suggested to be mediated by various
mechanisms. First, PcG complexes can mediate chromatin
completion⁴⁶⁻⁴⁸ (Fig. 2b). In *Drosophila melanogaster*, mutations
in cPRC1 genes were shown to induce decompaction of the Hox
clusters, followed by ectopic Hox gene expression which began a
few hours later.⁴⁹ In *Ring1b*-knockout mouse embryonic stem cells
(mESCs) chromatin decompaction and expression of Hox genes
occur even though the H3K27me3 repressive mark is still
present.⁵⁰ Surprisingly, this phenotype is rescued by a catalytic
mutant form of RING1B, suggesting that its E3-ubiquitin ligase is
dispensable for PcG-mediated silencing.⁵⁰ This latter result
 contrasts with research suggesting a role for the H2AK119ub
mark in maintaining PcG-dependent repression.¹¹,²⁴,⁵¹,⁵²

Second, a switch from a transcriptional repressive state to an
active state can be induced by competition between BAF — an
ATP-dependent chromatin remodeling complex part of SWI/SNF
family — and Polycomb complexes.¹² BAF-dependent eviction of
PcG proteins opens chromatin architecture after H3K27me3 and
H2AK119Ub depletion.⁵³ Strikingly, in a dominant-negative BAF
mutant background, accumulation of PRC1 and PRC2 on
chromatin does not necessarily trigger changes in chromatin
landscapes, suggesting that DNA-accessibility to BAF is PcG-
independent.⁵⁴

Third, the maintenance of the repressed state of PcG target
genes also depends on the PcG ability to block initiation and
elongation of transcription (Fig. 2c). In particular, RING1-mediated
ubiquitination maintains RNA polymerase in a poised state.⁵²,⁵⁵,⁵⁶
<figure><img src="image_4.png" /></figure>

**Fig. 2** Polycomb recruitment and action on target genes. **a** First described in *Drosophila melanogaster*, the original pathway of PcG recruitment relies on two sequential steps. First, PRC2 is recruited to chromatin and deposits the repressive H3K27me3 mark via its EZH1/2 subunit. The repressive mark is then recognized by the CBX2/4/6-8 chromodomain, a subunit of cPRC1. Lastly, RING1A/B deposit the ubiquitination on H2AK119 (in *Drosophila*, H2AK118). While in *Drosophila melanogaster* PRC2 recruitment depends on specific transcription factors binding to PREs, in mammals PRC2 recruitment can occur at CGIs or depends on transcription factors or lncRNAs. More recent data suggest an alternative recruitment pathway, in which ncPRC1 complexes are recruited in a KDM2B-dependent manner which deposits the H2AK119 ubiquitination mark. In turn, this mark is recognized by the JARID2 subunit of PRC2.2. Furthermore, PRC2.1 binds the same targets via PCL1/2/3 proteins. Finally, cPRC1 is recruited via CBX2/4/6-8-mediated recognition of H3K27me3. Moreover, the new PcG proteins BAHD1 and BAHD2 have also been found to recognize the H3K27me3 repressive mark. Their interactions with HDACs generate a hypoacetylated chromatin state which participates in transcriptional silencing. **b** Chromatin compaction impairs the transcription of target genes. **c** PcG-mediated silencing depends on the inhibition of the transcriptional machinery while the repressive PRC2 and PRC1 marks are necessary to inhibit the deposition of active histone marks. **d** PRC1 participates in the higher-order 3D chromatin organization via its PHC subunits. The SAM domain of PHC-PRC1 is able to oligomerize which results in the maintenance of the transcriptionally repressed state. BAHD1, Bromo adjacent homology domain containing 1; CBP, CREB binding protein; HDAC, Histone deacetylase; RNA Pol II, RNA Polymerase II; SWI/SNF, Switch/sucrose non-fermentable.

In mESCs, PRC2 can methylate Elongin A to block transcription.⁵⁷ The H2AK119ub and H3K27me3 repressive histone marks respectively repress deposition of the H3K4me2/3 and H3K27Ac active histone marks.⁵⁸,⁵⁹ Moreover, in flies, PRC1-PC binds to CBP and inhibits its H3K27 acetyltransferase activity, contributing to a repressive state⁶⁰ while on the other hand TRX or TRX-related (TRR) association to CBP antagonizes PcG-mediated silencing.⁶¹ Interestingly, while the trimethylated form of H3K27 has been extensively studied, less is known about the importance of H3K27me2 in transcriptional repression. Remarkably, the dimethylated H3K27 mark represents 70% of total histone H3 against 4% for the trimethylated form.⁶² Their distribution is mutually exclusive, indeed, the methylated state of H3K27 correlates with different transcriptional states. It is suggested that H3K27me2 coats most of chromatin in order to protect chromatin changes mediated by Histone Acetyl Transferase (HAT).⁶²

Finally, PcG proteins actively participate in the three-dimensional (3D) organization of the genome, adding a higher-order layer through which they contribute to gene regulation (Fig. 2d). PcG proteins can drive the formation of 3D-loops between regulatory elements such as promoters and

enhancers.⁶³,⁶⁴ Loop formation involves the cPRC1-PH subunit, that oligomerizes via its SAM domain, but is independent of the cPRC1 catalytic activity.⁶⁵⁻⁶⁷ Consistent with a function for 3D architecture in gene regulation, PRC1 knockout in ESCs leads to loss of promoter-promoter contacts resulting in transcriptional upregulation of PRC1 target genes.⁶⁸

## PCG PROTEIN FUNCTION IN TRANSCRIPTIONAL ACTIVATION

Interestingly, an involvement of PcG proteins in transcriptional activation has been suggested in pathological as well as in physiological contexts⁶⁹ and is now better understood at the molecular level (reviewed in⁷⁰). Morey et al. described that only 31% of the cPRC1 and ncPRC1 target genes overlap.⁷¹ While cPRC1 target genes are strongly repressed, ncPRC1 target genes are overall expressed and involved in dynamic processes such as metabolism and cell cycle progression.⁷¹ Thus, H3K27me3-independent PRC1 recruitment appears to be an important feature that favors transcriptionally active states by PcG proteins.⁷²⁻⁷⁴ For instance, PRC1.5 includes the component Autism Susceptibility candidate 2 (AUTS2) that recruits CK2 and p300
<table><caption>Table 1. continued</caption><thead><tr><th>Subunit</th><th>Alterations</th><th>Functions</th><th>Cancer type</th><th>Ref</th></tr></thead><tbody><tr><td rowspan="3">SUZ12</td><td>Y641</td><td>Tumor suppressive function, JAK2-mediated phosphorylation with β-TrCP-mediated ubiquitination leads to EZH2 degradation and hypomethylation of PcG target genes.</td><td>Lymphoma</td><td>250</td></tr><tr><td>K421</td><td>Tumor suppressive function, Smurf2-mediated ubiquitination leads to EZH2 degradation</td><td>T-ALL</td><td>251</td></tr><tr><td>Ubiquitination</td><td>Hypomethylation of PcG target genes, leading to oncogenic-related expression of Notch pathway genes</td><td>T-ALL</td><td>108</td></tr><tr><td rowspan="3">JARID2</td><td>Overexpression</td><td>Oncogenic function by repressing tumor suppressor genes (e.g., HRK), and promoting oncogene expression (e.g., cyclin D1)</td><td>Ovarian, colorectal, HNSCC</td><td>113-115</td></tr><tr><td>Downregulation</td><td>Decreased expression of Suz12 leads to increased activation of the ERK1/2 pathway and increases expression of MMP9 and MMP2 which promotes migration and invasion</td><td>Hepatocellular carcinoma</td><td>252</td></tr><tr><td>Loss-of-functions mutations, deletions</td><td>Impaired EED binding to H3K27me3, decrease in PRC2 catalytic activity</td><td>Myelodysplastic syndrome</td><td>90,253</td></tr><tr><td rowspan="2">EED</td><td>Overexpression</td><td>Promotes EMT by silencing E-cadherin</td><td>Breast cancer, colorectal cancer, hepatocellular carcinoma</td><td>232,254</td></tr><tr><td>Loss-of-functions mutations: I363M</td><td>Oncogenic function by increasing the deposition of the repressive mark at the PTEN promoter which promotes invasion and metastasis</td><td>Ovarian cancer cell lines, rhabdomyosarcomas, hepatocellular carcinoma</td><td>255-257</td></tr><tr><td rowspan="2">JARID2</td><td>Overexpression</td><td>JARID2 plays a tumor suppressive function by repressing self-renewal pathways</td><td>Chronic myeloid disorders</td><td>258</td></tr><tr><td>Deletion</td><td>Oncogenic function and chemoresistance</td><td>Ovarian cancer</td><td>259</td></tr><tr><td>AEBP2</td><td>Overexpression increase protein stability</td><td>Oncogenic function that may depend on its interaction with Elongin BC and USP7 in order to modulate the chromatin landscape</td><td>Breast cancer, colon cancer</td><td>260</td></tr><tr><td rowspan="3">EPOP</td><td>Overexpression</td><td>Might have an oncogenic function by deregulating the gene expression of target genes by altering the chromatin accessibility</td><td>Endometrial stromal sarcoma, ossifying fibromyxoid tumor</td><td>261-263</td></tr><tr><td>PCL1/PHF1</td><td>PHF1, BRD8-PHF1, MEAF6-PHF1</td><td>Oncogenic function, upregulates EZH2 and EED expression levels and the level of H3K27me3, H3K4me2 and H3K9me2</td><td>Gliomas</td><td>264</td></tr><tr><td>PCL2/MTF2</td><td>Overexpression</td><td>Tumor suppressive function, inhibits cell proliferation and promotion of apoptosis by inhibiting MDM2-mediated p53 degradation</td><td>Breast cancer</td><td>265</td></tr><tr><td rowspan="2">PCL3/PHF19</td><td>Overexpression</td><td>Tumor suppressive function, inhibits invasion and angiogenesis by interacting with PRC2</td><td>Prostate cancer, melanoma</td><td>266,267</td></tr><tr><td>Overexpression, genomic amplification</td><td>Oncogenic function, increases PRC2 activity</td><td>Hepatocellular carcinoma, glioblastoma cells, multiple myeloma</td><td>268-270</td></tr><tr><td rowspan="2">CATACOMB</td><td>Fusion genes: MBTD1-CXorf67</td><td>Recurrent fusion of unknown mechanistic function</td><td>Endometrial stromal sarcoma</td><td>209</td></tr><tr><td>Overexpression</td><td>Oncogenic function, mimics H3K27M, oncohistones by binding the SET domain of EZH2 which blocks its catalytic activity leading to the derepression of PRC2 targeted genes</td><td>PFA</td><td>9,271</td></tr></tbody></table>

DLBCL, Diffuse Large B-Cell Lymphoma; GBM, Glioblastoma; HNSCC, Head and Neck Squamous Cell Carcinoma; NKTL, Natural Killer/T-cell Lymphoma; PFA, Posterior Fossa A; T-ALL, T-cell Acute Lymphoblastic Leukemia.
<table><caption>**Table 2.** Alterations of PRC1 components and their mechanistic consequences in cancer.</caption><thead><tr><th>Subunit</th><th>Alterations</th><th>Functions</th><th>Cancer type</th><th>Ref</th></tr></thead><tbody><tr><td rowspan="2">RING1B</td><td>PTMs</td><td>Oncogenic function, promotes the recruitment of demethylase UTX and acetylase p300 at poised promoters (e.g., CCDN2)</td><td>Melanoma</td><td>272</td></tr><tr><td>Phosphorylation</td><td>Oncogenic function by enhancing oncogene expression (e.g., p63)</td><td>AML, breast cancer, gastrointestinal tumors, lymphomas, pancreatic cancers</td><td>138,273,274</td></tr><tr><td></td><td>Overexpression</td><td>Oncogenic function by regulating oncogenic enhancer activity via its association with pioneer factors (e.g., FOXA1) and transcription factors (e.g., ERα)</td><td>Breast cancer, leukemia, hepatocellular carcinoma</td><td>139</td></tr><tr><td rowspan="3">RING1A</td><td>Overexpression</td><td>Oncogenic function by enhancing oncogene expression</td><td>AML</td><td>138</td></tr><tr><td rowspan="2">Overexpression</td><td>Oncogenic function through repression of p21<sup>WIT1/CIP1</sup></td><td>HeLa cells</td><td>275</td></tr><tr><td>Oncogenic function by promoting cancer stem cell self-renewal via the direct repression of RDH16, leading to the decrease in the synthesis of all-trans retinoic acid</td><td>Glioma cells</td><td>276</td></tr><tr><td></td><td>Overexpression</td><td>Oncogenic function by enhancing the expression of CRC stem cell markers (e.g., CD133, CD44, ALDH1A1) via an increase in H3K4me3, while H3K27me3 decreases following upregulation of KMT2A and KDM6A</td><td>Colorectal cancer</td><td>277</td></tr><tr><td>PCGF2/MEL-18</td><td>Overexpression</td><td>Tumor suppressive function by inhibiting the expression of oncogenes (e.g., PI3K/AKT pathway, ZEB1/ZEB2, PCGF4)</td><td>Breast cancer, gastric cancer</td><td>132,134,278,279</td></tr><tr><td>PCGF3</td><td>Overexpression</td><td>Oncogenic function by promoting proliferation via expression of cell cycle-related genes (e.g., CyclinB1, CyclinD1, CDK4), migration-related genes (RhoA, RhoC, CDC42) and by regulation of the PI3K/AKT pathway</td><td>NSCLC</td><td>280</td></tr><tr><td rowspan="2">PCGF4/ BMI-1</td><td>Overexpression</td><td>Oncogenic function, promotes cell immortality by repressing the <i>INK4a-ARF</i> locus</td><td>Breast cancer, NSCLC, gastric carcinoma, pancreatic cancer, hematological malignancies</td><td>124,281–284</td></tr><tr><td>IG-BMI1 fusion</td><td>Oncogenic function</td><td>Chronic lymphocytic leukemia</td><td>285</td></tr><tr><td></td><td>Enhanced protein stability</td><td>Its association with AR inhibits its degradation, leading to the overexpression of AR downstream target genes (PSA, TMPRSS2)</td><td>Prostate cancer</td><td>163</td></tr><tr><td rowspan="2">PTMs</td><td rowspan="2">O-GlcNAcylation</td><td>O-GlcNAcylation increases BMI-1 protein stability, which in turn represses TP53, PTEN and CDKN1A/CDKN2A</td><td>Prostate cancer</td><td>162</td></tr><tr><td>Oncogenic function, promotes cell migration by affecting the EMT</td><td>Breast cancer</td><td>286</td></tr><tr><td>PCGF6</td><td>Frameshift deletion inducing an early stop of translation</td><td>Oncogenic function, promotes cell migration by affecting the EMT</td><td>Hepatocellular carcinoma, breast cancer</td><td>287,288</td></tr><tr><td>CBX1</td><td>Overexpression</td><td>Oncogenic function by activation of the PI3K/AKT pathway</td><td>Breast cancer</td><td>289</td></tr><tr><td rowspan="2">CBX2</td><td>Overexpression</td><td>Oncogenic function by activation of the YAP/β-catenin pathway</td><td>Gastric cancer</td><td>290</td></tr><tr><td></td><td>Oncogenic function by inducing chemoresistance, stem cell-like phenotype</td><td>High-grade serous ovarian carcinoma</td><td>291</td></tr><tr><td rowspan="2">CBX3</td><td>Overexpression</td><td>Oncogenic function via transcriptional regulation of p21 leading to excessive proliferation</td><td>Colon cancer, tongue squamous cell carcinoma</td><td>292,293</td></tr><tr><td></td><td>Oncogenic function, higher expression correlates with poor prognosis</td><td>Breast cancer, NSCLC</td><td>294,295</td></tr><tr><td></td><td>Oncogenic function via FBP1 silencing which positively regulates aerobic glycolysis</td><td>Pancreatic cancer</td><td>296</td></tr><tr><td rowspan="3">CBX4</td><td>Overexpression</td><td>Oncogenic function by increasing angiogenesis via the sumoylation of HIF-1α which enhances VEGF expression</td><td>Hepatocellular carcinoma</td><td>297,298</td></tr><tr><td></td><td>Oncogenic function via interaction with HDAC1 which transcriptionally represses the tumor suppressor KLF6</td><td>Clear cell renal cell carcinoma</td><td>299</td></tr><tr><td></td><td>Tumor suppressive function by repressing Runx2 expression via recruitment of HDAC3 at its promoter, leading to the inhibition of cell migration, invasion and metastasis</td><td>Colorectal carcinoma</td><td>300</td></tr><tr><td>CBX6</td><td>Overexpression</td><td>Oncogenic function by accelerating EMT process in HCC cells via upregulation of Snail and Zeb1</td><td>Hepatocellular carcinoma</td><td>301,302</td></tr></tbody></table>
<table><caption>**Table 2.** continued</caption><thead><tr><th>Subunit</th><th>Alterations</th><th>Functions</th><th>Cancer type</th><th>Ref</th></tr></thead><tbody><tr><td>CBX7</td><td>Downregulation</td><td>CBX6 plays a potential tumor suppressor function by down-regulating BST2. In breast cancer, CBX6 expression is repressed in a PRC2-dependent manner</td><td>Breast cancer</td><td>303</td></tr><tr><td>CBX7</td><td>Overexpression</td><td>Oncogenic function via inhibition of the <i>INK4a-ARF</i> locus</td><td>Prostate cancer, germinal center-derived follicular lymphomas, gastric cancer</td><td>304-306</td></tr><tr><td>CBX7</td><td>Downregulation</td><td>CBX7 plays a tumor suppressive function. Progressive loss of CBX7 expression as <i>INK4a-ARF</i> expression increases</td><td>Thyroid cancer</td><td>307</td></tr><tr><td>CBX7</td><td>Downregulation</td><td>CBX7 plays a tumor suppressive function through inhibition of cyclin E expression</td><td>Lung cancer</td><td>308</td></tr><tr><td>CBX7</td><td>Overexpression</td><td>CBX7 plays an oncogenic function via loss of tumor suppressor miRNAs (<i>miR-125b</i>) and gain of oncogenic miRNAs (<i>miR-182</i> and <i>miR-183</i>)</td><td>Breast cancer</td><td>309</td></tr><tr><td>CBX8</td><td>Overexpression</td><td>Oncogenic function<br>Oncogenic function, binds EGR1 promoter and <i>miR-365-3p</i>, which enhance the AKT/β-catenin pathway<br>In a PRC1-independent manner, cooperates with Wdr5 in order to maintain H3K4me3 at the level of the promoters of Notch pathway genes</td><td>Glioblastoma, breast cancer<br>Hepatocellular carcinoma<br>Breast cancer</td><td>310,311<br>312<br>313</td></tr><tr><td>PHC3</td><td>G201C missense mutation</td><td>Tumor suppressive function</td><td>Osteosarcoma</td><td>314,315</td></tr><tr><td>RYBP</td><td>Cytoplasm-located mutant of RYBP</td><td>Loses tumor suppressive function<br>Tumor suppressive function, more potent ability to bind to caspase 8 which prevents p53 degradation in human tumor cells</td><td>Breast cancer cells, osteosarcoma epithelial cells, colon cancer cells</td><td>316</td></tr><tr><td>RYBP</td><td>Overexpression</td><td>Oncogenic function</td><td>Oligodendroglia tumors, pituitary adenoma, Hodgkin's lymphoma and T cell lymphoma</td><td>317,318<br>317,319,320</td></tr><tr><td>Downregulation</td><td>Overexpression</td><td>RYBP plays a tumor suppressive function by decreasing proliferation, cell migration and metastasis</td><td>Lung cancer, hepatocellular carcinoma, thyroid cancer, breast cancer<br>NSCLC</td><td>321</td></tr><tr><td>YAF2</td><td>Overexpression</td><td>PTMs Phosphorylation</td><td>Breast cancer cells, colon cancer cells</td><td>322</td></tr><tr><td>YAF2</td><td>Overexpression</td><td>S167</td><td>Stabilizes FANK1 and inhibits its degradation, inhibits FANK1-mediated apoptosis</td><td>323</td></tr><tr><td>AutS2</td><td>Overexpression</td><td>The association of <i>AutS2</i> with PRC1.5 favors MSX1 expression, which inhibits the differentiation of T-cell lymphocytes</td><td>T-ALL</td><td>324</td></tr><tr><td>AutS2</td><td>Fusion genes: PAX5-AUTS2</td><td>Recurrent rearrangement between 9p13 and 7q11 regions in B-cell acute lymphoblastic leukemias. Mechanism of action unknown to date.</td><td>B-ALL</td><td>325,326</td></tr><tr><td>BCOR</td><td>Fusion gene: ZC3H7B-BCOR, BCOR-MAML3</td><td>Dysregulation of PRC1-mediated repression which could explain tumorigenesis</td><td>Endometrial stromal sarcomas, clear cell sarcoma of the kidney</td><td>327</td></tr><tr><td>KDM2B</td><td>Loss-of-function</td><td>BCOR plays a tumor suppressive function by inhibiting proliferation and self-renewal mediated by Notch1 target genes</td><td>AML, T-ALL</td><td>328</td></tr><tr><td>KDM2B</td><td>Overexpression</td><td>Tumor suppressive function by suppressing genes of the Notch pathway</td><td>T-ALL</td><td>329,330</td></tr><tr><td>KDM2B</td><td>Overexpression</td><td>Oncogenic function, by regulating cell fate</td><td>AML, Breast cancer</td><td>329,330</td></tr></tbody></table>

AML, Acute Myeloid Leukemia; AR, Androgen Receptor; B-ALL, B-cell Acute Lymphoblastic Leukemia; CRC, Colorectal Cancer; ER, Estrogen Receptor; EMT, Epithelial-Mesenchymal Transition; NSCLC, Non-Small Cell Lung Cancer; PTMs, Post-Translational Modifications; T-ALL, T-cell Acute Lymphoblastic Leukemia.
which, respectively, inhibits the E3-ubiquitin ligase activity by
phosphorylating RING1B and deposits acetylation on histone tails
to facilitate transcription.¹⁵,⁷⁵ For PRC1.5-AUTS2 target genes, the
concomitant enrichment for the H3K4me3 and H4K16Ac active
marks, the presence of the RNA polymerase II and a reduction of
the H3K27me3 repressive mark lead to transcriptional activation.⁷⁵
The transcription factor NRF1 is involved in PRC1.5-AUTS2
recruitment to its target genes, providing an example for
sequence-specific targeting of a PRC1 complex in mammals.⁷⁶

To recapitulate, it is the specific composition of the PcG
complexes, as well as the dynamics of their chromatin binding and
replacement through cell lineages that determine their transcrip-
tional impact.<sup>15,71,77</sup> While canonical PcG proteins maintain
cellular memory, such as in stem cells where they support self-
renewal properties by repressing lineage-specific genes, ncPRC1s
control differentiation in more subtle ways. By fine-tuning
transcription, PcG proteins are master contributors of cell fate
determination,<sup>1</sup> in particular in the control of a balance between
proliferation and differentiation. On the other hand, loss of this
fine balance upon misregulation of PcG-dependent mechanisms
can cause pathogenesis.

POLYCOMB IN CANCER

Altering the proper functions of PcG can affect cellular identity,
therefore promoting tumorigenesis (Tables 1, 2).

PRC2 in cancer

Polycomb dysregulation in cancer has been the subject of
extensive studies since Varambally et al. demonstrated that
EZH2 overexpression is associated with advanced stage and poor
prognosis in prostate cancer.⁷⁸ Quantitative and qualitative EZH2
dysregulation has been frequently described in solid malignancies
including lung, hepatocellular, breast, colorectal, pancreatic
cancers as well as in several hematologic malignancies.⁷⁹,⁸⁰ EZH2
expression can be regulated by specific transcription factors,
including the MLL-AF9 fusion protein, or by miRNAs that will
induce EZH2 mRNA decay.⁸¹,⁸² Dysregulation of those specific
regulators participates in the tumorigenesis onset. EZH2 over-
expression in patients is associated with a higher risk of relapse.⁷⁸
PRC2 plays a major role in self-renewal of hematopoietic stem
cells;⁸³⁻⁸⁷ its dysregulation is often found in multiple blood
cancers⁸⁷⁻⁹² in which EZH2 can behave both as a tumor
suppressor⁸⁵,⁹³,⁹⁴ or an oncogene⁹⁵⁻⁹⁸ depending on the cell
context (reviewed in⁸⁰) (Fig. 3a).

The onset or cancer progression may be associated with
mutations affecting the catalytic SET-domain of EZH2 that is
essential for H3K27 methylation (Fig. 3b). An EZH2<sup>Y641F</sup>/N gain-of-
function (GOF) mutation affecting the tyrosine 641 (Y641) located
in the SET-domain induces hypermethylation of H3K27.<sup>99,100</sup>
Particularly, EZH2<sup>Y641</sup> has an increased affinity for dimethylated
H3K27 form which causes a widespread redistribution of
H3K27me3 and a decrease in H3K27me2, leading to transcrip-
tional misregulation of affected genes.<sup>100-102</sup> Moreover, the
higher-order chromatin landscape can also be affected. In recent
years, multiple cutting edge approaches have shown that the
genome folds into a hierarchy of structures, from nucleosomes, to
chromatin loops and nanodomains, Topologically Associating
Domain (TADs), chromosome compartments and chromosome
territories.<sup>103</sup> TADs are particularly interesting since they consti-
tute regulatory landscapes for the genes contained within each
TAD.<sup>104</sup> Interestingly, co-repression of several tumor suppressors
was suggested to participate in tumor growth.<sup>105,106</sup> An estab-
lished tumor state can also participate a *posteriori* in the
redistribution of EZH2 on ectopic targets, triggering changes in
cell identity due to misexpression of homeotic genes.<sup>107</sup> In
addition to GOF effects, loss-of-function (LOF) mutations and
deletions affecting EZH2 and SUZ12 in T-cell acute lymphoblastic

leukemia (T-ALL) — a hematopoietic cancer — lead to hypo-
methylation of H3K27 target genes, including *Notch*, a major
player in T-ALL, thereby contributing to oncogenesis¹⁰⁸ (Fig. 3a).
PRC2 LOF is found in around 25% of T-ALL in association with
oncogenic activating mutations of the JAK/STAT signaling path-
way and leads to a global epigenetic remodeling towards
H3K27Ac. This active histone mark is recognized by Bromodomain
and Extraterminal (BET)-domain proteins that act as its specific
readers, allowing reactivation of a BET-dependent transcriptional
network that triggers stem cell-like programs leading to poor
prognosis. PRC2-altered T-ALL being dependent on BET proteins,
BET domain protein inhibition is therefore a promising therapeutic
avenue in PRC2-associated-T-ALL patients.¹⁰⁹

EZH2 post-translational modifications (PTMs) play an additional
role in certain type of cancers¹¹⁰,¹¹¹ (Fig. 3c). In patients with
advanced prostate cancer, H3K36me3 and H3K27me3 levels are
inversely correlated.⁷⁸,¹¹¹ SETD2, the methyltransferase responsi-
ble for H3K36me3 deposition, also monomethylates EZH2 on its
lysine 735 residue, inducing EZH2 degradation and consequently
delaying metastasis. SETD2 is strongly correlated with the
presence of EZH2-K735me1 and particularly found in patients
with prostate cancer with better clinical outcome.¹¹¹ On the
contrary, EZH2-K307 methylation by SMYD2 improves its stability
and participates in the transcriptional repression of pro-apoptotic,
anti-proliferation and anti-invasion target genes¹¹² (Fig. 3c).
Multiple EZH2 PTMs play thus a role in EZH2 function and stability
that will result in an H3K27 hypermethylation or hypomethylation
of the chromatin landscape that favors tumorigenesis (Table 1).

Additionally, SUZ12 is upregulated in a variety of cancers,
including ovarian, colorectal and head and neck squamous cell
carcinoma.<sup>113-115</sup> The knockdown of SUZ12 is able to reverse
tumor growth by inhibiting proliferation and inducing apoptosis
in these contexts.<sup>113,115</sup> On the other hand, SUZ12 loss in T-ALL
disrupts the PRC2 complex, leading to H3K27me3 decrease which
correlates with the opening of chromatin and upregulation of the
corresponding genes involved in oncogenic signaling pathways<sup>92</sup>
(Fig. 3a). Moreover, PRC2 loss induces a genome-wide redistribu-
tion of the H3K27Ac mark and the activation of poised
enhancers.<sup>62</sup> Therefore, similar to EZH2, SUZ12 can act as pro-
oncogenic or tumor suppressor depending on the cancer type.

As previously mentioned, PRC2 can be divided into two sub-
complexes, PRC2.1 and PRC2.2. While their target genes are
overlapping,<sup>34,116</sup> their differences rely on their affinity to
chromatin.<sup>117</sup> Indeed, PRC2.1 tends to have a higher affinity to
chromatin, which leads to an increase in H3K27me3 deposition
and silencing of PcG target genes in the presence of high ratios of
PRC2.1 to 2.2.<sup>117</sup> In leukemia, colon and uterine adenocarcinomas,
missense mutations of SUZ12, SUZ12(R103P/Q), result in JARID2
depletion, leading to an increase in PRC2.1 formation which
enhances PRC2 chromatin occupancy.<sup>117</sup> How PRC2.1 could be
specifically implicated in cancer remains to be determined.

Although PRC2 dysregulation events have been widely docu-
mented in cancer, it is still difficult to decipher whether they are
drivers in tumorigenesis. Even if EZH2 is dispensable for the
progression of prostate and mammary cancer, it is nonetheless
highly expressed.¹¹⁸ In fact, in normally dividing cells, the rate of
EZH2 expression correlates with proliferation rates,¹¹⁸ compensat-
ing the proliferation-dependent dilution of H3K27me3. In these
cancers, even though EZH2 is overexpressed, tumor cells
paradoxically fail to maintain a wild-type dose of H3K27me3.
The use of EZH2 inhibitors for cancer treatment should therefore
carefully take into account the tumor proliferation status.¹¹⁸ With
the aim to identify the cancer types in which treatment using
PRC2 inhibitors could be beneficial, a genomic and transcriptomic
analysis using available databases on clinical tumor samples and a
panel of tumor cell lines has been performed, revealing a
correlation of EZH2, SUZ12 or EED amplifications with poor
prognosis in a subclass of human cancers like renal papillary cell
<figure><img src="image_5.png" /></figure>

<figure><img src="image_6.png" /></figure>

<figure><img src="image_7.png" /></figure>

<figure><img src="image_8.png" /></figure>

<figure><img src="image_9.png" /></figure>

<figure><img src="image_10.png" /></figure>

carcinoma, low-grade glioma and hepatocellular carcinoma.¹¹⁹ Interestingly, GOFs of PRC2 subunits are also anti-correlated with poor prognosis in some cancers like gastric cancer and thymoma, suggesting a tumor suppressor function of PRC2 in those cases.

It remains to be understood why certain tumors are addicted to one specific PRC2 subunits but not the others. Clearly, a better

understanding of the rate-limiting roles and the cell type-specific functions of each of the PRC2 subunits will require future research.

### PRC1 in cancer

Like PRC2, PRC1 components are widely implicated in many types of cancers (Table 2). BMI-1 (PCGF4), a cPRC1.4 subunit, has
**Fig. 3** Multifaceted roles of PRC2 in tumorigenesis. **a** Upregulation of PRC2 components results in H3K27 hypermethylation, which, if present in tumor suppressor genes, induces their downregulation. In contrast, downregulation of PRC2 components at oncogenes leads to H3K27 hypomethylation and a switch to acetylation, contributing to the overexpression of specific oncogenes. **b** GOF mutations (indicated by a star) affecting the SET-domain of EZH2 can lead to overactivation of its H3K27 methyltransferase catalytic activity and to the silencing of tumor suppressor genes. **c** PTMs of EZH2 participate in tumorigenesis. Left: methylation of K307 of EZH2 by SMYD2 enhances its stability, resulting in a H3K27 hypermethylated state of tumor suppressor genes. Right: on the other hand, methylation of its K735 causes EZH2 degradation. The loss of EZH2 induces the replacement of H3K27me3 by H3K27ac, leading to the transcriptional expression of oncogenes. **d** Polycomb-independent roles of EZH2 in transcriptional activation. The gene encoding the AR is a direct target of EZH2-mediated transcriptional activation in Androgen-Dependent and Castration-Resistant Prostate Cancers (ADPC and CRPC, respectively). This mechanism is methylation-independent and escapes EZH2 inhibitors. In CRPC, EZH2 acts as a co-factor of AR. This functional transition of EZH2 from a role of repression to a role of activation of transcription depends on its phosphorylation at the level of Ser21. EZH2 and AR directly interact. This interaction inhibits the degradation of the AR and causes the overexpression of the AR target genes. **e** Under physiological conditions, PRC2 participates in the transcriptional repression of its HOX target genes throughout development. However, oncogenic transformation can redirect PRC2 to new target genes. This PRC2 redistribution, in particular at differentiation-related genes, induces a loss of differentiation and participates in the generation of a pluripotent stem cell-like phenotype. AR, Androgen Receptor; CBP, CREB binding protein; PSA, Prostate-Specific Antigen; SETD2, SET domain-containing 2 (a histone lysine methyltransferase); SMYD2, SET and MYND domain-containing 2.

historically been described as a proto-oncogene that collaborates with the c-Myc oncoprotein to trigger tumorigenesis.<sup>120-123</sup> The *INK4a-ARF* locus, encoding the tumor suppressors p16<sup> Ink4 </sup> and p19<sup> Arf </sup>, is a direct target of PRC1-4.<sup>124</sup> BMI-1 deficiency is associated with overexpression of p16<sup> Ink4 </sup> and p19<sup> Arf </sup> and therefore with cell cycle arrest, senescence and apoptosis (Fig. 4a). In contrast, BMI-1 overexpression triggers cell proliferation by repressing *ink4a-ARF* expression.<sup>124</sup> BMI-1 is involved in gastric, pancreatic, breast and ovarian cancer among others.<sup>125-129</sup> MEL-18 (PCGF2), a BMI-1 homolog, has a tumor suppressing activity.<sup>130-132</sup> BMI-1 and MEL-18 expression levels are inversely correlated in various cancers.<sup>133,134</sup> BMI-1 expression depends on its counterpart MEL-18 (Fig. 4a). c-Myc is a transcriptional activator of BMI-1. *Mel-18* overexpression is linked to *c-myc* downregulation, leading to BMI-1 decrease, p16 upregulation and ultimately to cell senescence.<sup>135</sup> Interestingly, in flies, LOF of cPRC1 members results in upregulation of cancer-related genes, including genes involved in the Notch, JNK and JAK/STAT signaling pathways<sup>74,136,137</sup> (Fig. 4b), a difference that might be due in part to the absence of PcG-mediated repression of the *INK4a-ARF* locus in flies.

Using an in vivo and in vitro approach, ncPRC1.1 was shown to specifically target active genes independently of PRC2.<sup>74,138</sup> At a genome-wide level, the correlation between mammalian RING1B and the H3K27me3 mark decreases during lineage decision processes. While PRC1-RING1B targets are clearly enriched for the repressive H3K27me3 mark in ESCs, this is only the case for ~30% of them in differentiated cells.<sup>74</sup> While gene ontology categories associated with H3K27me3-dependent targets are linked to developmental pathways, H3K27me3-independent targets are linked to cell cycle regulation, cell polarity, metabolism and signaling pathways<sup>74,138</sup> (Fig. 4b). This difference in PRC1 targeting results from major changes in the qualitative and quantitative compositions of the ncPRC1 variant complexes.<sup>15,71</sup>

Unlike PRC2 mutations, PRC1 mutations are not overrepresented in cancer.<sup>139</sup> However, some mutations affecting ncPRC1 have been described.<sup>140,141</sup> In SHH-driven medulloblastoma, the PRC1.1 BCOR scaffold protein is mutated at its C-terminal domain that normally interacts with PCGF1,<sup>141,142</sup> resulting in loss of PRC1.1 recruitment to genes coding for growth factors that would otherwise be repressed<sup>141</sup> (Fig. 4c). Likewise, MGA, a transcription factor that is a member of the Myc network and interacts with ncPRC1.6 subunits, is a tumor suppressor in vivo that acts by recruiting ncPRC1.6 to its target genes.<sup>143</sup> Moreover, BAP1, a component of the Polycomb Repressive complex DeUbiquitinase (PR-DUB), is a tumor suppressor.<sup>144,145</sup> Recent data suggest that this protein prevents widespread H2AK119ub deposition and chromatin condensation at non-target loci, restricting H2AK119ub to Polycomb target genes. BAP1 may thus prevent inappropriate redistribution of Polycomb complexes away from their targets and

play critical roles, particularly by maintaining the appropriate chromatin state of lineage commitment genes.<sup>146-149</sup> It is therefore not surprising that PR-DUB misregulation leads to tumorigenesis. Enhancing deubiquitinase activity leads to a widespread depletion of the H2AK119ub mark.<sup>140</sup> Conversely, disruption of its chromatin recruitment or catalytic activity could result in an increase in H2AK119ub and H3K27me3.<sup>146,150</sup> Depending on the genes targeted, this might switch the transcriptional state of oncogenes or tumor suppressor genes.

In summary, the implication of PcG components in cancer, either by point mutations or by dysregulation of its components, is widely established. Through tumor suppressor or oncogenic activity in a broad type of cancers, PcG members control tumor growth and survival.<sup>151</sup> Targeting PRC2 members or proteins involved in PRC2 stability, either by inhibiting its enzymatic activity or by interfering with PRC2 complex assembly or stability, appears to be a promising strategy to prevent growth of PRC2-dependent tumors<sup>79,152-154</sup> (Table 3). However, since PRC1 can either repress or activate the transcription of its target genes, it is both the downregulation and/or upregulation of tumor suppressors and oncogenes respectively that might participate in tumorigenicity.<sup>69,155-157</sup> The exact role of PRC1 complexes in cancer, and in particular the importance of ncPRC1 complexes, remains to be determined. Future work would be important to better characterize the molecular implication of Polycomb complexes and define appropriate therapeutic approaches to rescue their dysregulation in different types of cancer.

ENVIRONMENTAL CUES AND POLYCOMB-DEPENDENT ONCOGENESIS

Hormone-dependent cancer

PRC1 genes are significantly amplified in hormone-dependent
cancers.¹³⁹ Since hormone receptors are transcription factors, they
might participate in tumorigenesis by triggering ectopic recruit-
ment of Polycomb proteins to a specific set of target genes. In
particular, the androgen receptor (AR) and the estrogen receptor
(ER) can directly recruit PcG proteins at their response elements in
hormone-dependent cancers.¹³⁹,¹⁵⁸⁻¹⁶⁰ In prostate cancer, main-
tenance of AR expression is essential. The overexpression of BMI-1
and its increased protein stability mediated by PTMs, such as O-
GlcNAcylation, participate in the self-renewal of cancer cells and
the progression of prostate cancer¹⁶¹,¹⁶² (Fig. 4d). Furthermore,
the binding of BMI-1 to AR inhibits the ubiquitin–proteasome
degradation pathway.¹⁶³ Surprisingly, the AR interacts with BMI-1
in a PRC1-independent manner.¹⁶³ By coupling ChIP-seq and
CRISPR methodologies, it was found that Androgen Response
Elements (AREs) are located in the *BMI1* locus and enriched for the
H3K27Ac active enhancer mark, suggesting that the AR activates
transcription of BMI-1.¹⁶⁰ Moreover, a positive feedback loop exists
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<figure><img src="image_12.png" /></figure>

<figure><img src="image_13.png" /></figure>

<figure><img src="image_14.png" /></figure>

<figure><img src="image_15.png" /></figure>

<figure><img src="image_16.png" /></figure>

in prostate cancer where BMI-1 overexpression stabilizes AR, which in turn transcriptionally activates BMI-1 expression, leading to tumor progression (Fig. 4e). In addition, a PRC2-independent EZH2 oncogenic function relies on its direct interaction with AR, leading to AR transcription and activation of AR downstream

targets<sup>164–166</sup> (Fig. 3d). This PRC2 genome-wide redistribution also results in ectopic targeting, in particular to tumor suppressor genes, particularly those involved in INF-γ signaling, that are repressed by the H3K27me3 mark in prostate cancer<sup>167,168</sup> (Fig. 3e).
**Fig. 4** Multifaceted roles of PRC1 in tumorigenesis. **a** PCGF2 inhibits the transcription of *c-myc*. Loss of *c-Myc* results in the decrease of PCGF4 expression, and in the derepression of PCG4 target genes, such as the *INK4a-ARF* locus. p19 and p16 participate in proliferation control, respectively, by inhibiting MDM2-mediated degradation of p53 and inhibiting CycD/CDK4-mediated phosphorylation of pRb. **b** PRC1 oncogenic activity may also be PRC2-independent. PRC1 is found on specific targets lacking the H3K27me3 repressive mark. Surprisingly, these genes exhibit active marks such as H3K27Ac and H3K4me1/3. Gene ontology analysis characterized these cancer-related genes as components of cell signaling, like the Notch and JAK/STAT signaling pathways. **c** PRC1 mutations are rarely found in cancer, although some mutations have been found to impact variant PRC1. Indeed, mutations (indicated by a star) in BCOR, a scaffold protein involved in ncPRC1.1, are found in SHH-driven medulloblastoma. The presence of these mutations promotes a neoplastic state of cancer cells by preventing Polycomb recruitment to its target genes. **d** PTM of PRC1 subunits can promote tumorigenesis. The deposition of O-GlcNAcylation on PCGF4 (BMI-1) inhibits its degradation. PCGF4 protein levels are increased and participate in the transcriptional silencing of downstream target genes such as the *INK4a-ARF* locus, thus promoting oncogenic cell proliferation. **e** In hormone-dependent cancers, PRC1 genes are often amplified. Top: in prostate cancer, the AR promotes the expression of PCGF4. Additionally, it can interact with the PCGF4 protein, resulting in inhibition of AR degradation and transcriptional activation of its downstream target genes. Bottom: cPRC1 can also interact with the ER and its pioneer factor FOXA1 in ER⁺ breast cancer cells and bind to enhancers that stimulate transcription of cancer-related genes decorated with active histone marks. AR, Androgen Receptor; Cdk4, 6, Cyclin Dependent Kinase 4, 6; ER, Estrogen Receptor; FOXA1, Forkhead Box A1; Igf2, Insulin-like growth factor 2; MDM2, Murine Double Minute 2; PSA, Prostate Specific Antigen; Rb, Retinoblastoma.

<table><caption>**Table 3.** PCG inhibitors and ongoing clinical trials.</caption><thead><tr><th>Target</th><th>Agent</th><th>Cancer</th><th>Status</th><th>Clinical study (NCT#)</th><th>Ref</th></tr></thead><tbody><tr><td rowspan="5">EZH2</td><td rowspan="5">Tazemetostat (formerly known as: EPZ-6438, E7438)</td><td>B-cell NHL</td><td>Phase 2</td><td>NCT03456726</td><td rowspan="2">331</td></tr><tr><td>MRT, RTK, ATRT, synovial sarcoma, malignant rhabdoid tumor of ovary, renal medullary carcinoma, epitheloid sarcoma, solid tumor with an EZH2 GOF mutation</td><td>Phase 2</td><td>NCT02601950</td></tr><tr><td>Malignant mesothelioma</td><td>Phase 2</td><td>NCT02860286</td><td rowspan="2">332,333</td></tr><tr><td>B-cell lymphomas, advanced solid tumors DLBCL, follicular lymphoma</td><td>Phase 1<br>Phase 2</td><td>NCT01897571</td></tr><tr><td rowspan="2">SHR2554</td><td>Lymphoid neoplasm</td><td>Phase 1</td><td>NCT03603951</td><td rowspan="4">334</td></tr><tr><td>Solid tumor, lymphoma</td><td>Phase 2</td><td>NCT04407741</td></tr><tr><td rowspan="2">CPI-1205</td><td>B-cell lymphoma</td><td>Phase 1</td><td>NCT02395601</td></tr><tr><td>mCRPC</td><td>Phase 2</td><td>NCT03480646</td></tr><tr><td rowspan="4">Valemetostat Tosylate (DS-3201b)</td><td>T-cell leukemia/lymphoma</td><td>Phase 2</td><td>NCT04703192</td><td rowspan="4"></td></tr><tr><td>Small cell lung cancer</td><td>Phase 1</td><td>NCT03879798</td></tr><tr><td>Lymphoma, non-Hodgkin lymphoma</td><td></td><td>NCT02732275</td></tr><tr><td></td><td></td><td></td></tr></tbody></table>

ATRT, Atypical Teratoid Rhabdoid Tumors; DLBCL, Diffuse Large B-cell Lymphoma; GOF, Gain-Of-Function; mCRPC, Metastatic Castration Resistant Prostate Cancer; MRT, Malignant Rhabdoid Tumors; NHL, Non-Hodgkin's Lymphoma; RTK, Rhabdoid Tumors of the Kidney.

The redistribution of PCG-targets is an important mechanism participating in tumorigenesis and cancer progression. Surprisingly, in breast cancer, ERα, β-catenin and EZH2 interact and target oncogenes, such as *c-Myc* and *Cyclin D1*, acting as transcriptional co-activators.¹⁶⁹ Furthermore, the redistribution of PRC1 leads to its association with active enhancers enriched for the H3K4me1 mark.¹³⁹ RING1B was proposed to facilitate ERα recruitment to enhancers and super-enhancers, as well as to promoters of cancer-related genes¹³⁹,¹⁷⁰ (Fig. 4e). However, how RING1B is recruited to open chromatin sites and how it selectively binds to a subset of them is still unclear.

## Metabolism

Proliferation and growth of cancer cells are known to be associated with an extensive rewiring of metabolism and energy production networks where Polycomb complexes are clearly involved. As already mentioned, changes in methylation of H3K27 participate in tumor progression.⁹⁴,¹⁰⁸ Tight regulation of the methyl group available for EZH2 activity is essential to maintain a proper chromatin landscape. The catalytic activity of EZH2 depends on the methyl donor S-adenosylmethionine (SAM)¹⁷¹ (Fig. 5e). SAM is formed by the combination of a methionine, which crosses the cell membrane via the LAT1 transporter, and an ATP molecule. Cancer cells with higher levels of LAT1 expression have a more aggressive phenotype.¹⁷² Upon

LAT1 depletion, the SAM pool is significantly reduced, correlating with a decrease in H3K27me3 deposition even if EZH2 protein concentration is constant.¹⁷² In addition, repression of RXRα, a known negative regulator of LAT1, by the PRC2 complex maintains a positive feedback loop between LAT1 and EZH2, enhancing EZH2 methyltransferase activity¹⁷² (Fig. 5a). Indeed, EZH2 inhibition via competition with SAM has a potent anti-tumor effect.¹⁷³

PCG proteins are also involved in the regulation of branched-chain amino acids (BCAAs),¹⁷⁴ key regulatory components for protein synthesis and energy production, both of which are also the fuel of cancer progression.¹⁷⁵ Enzymes required for BCAA catabolism, known as BCAA aminotransferases (BCATs), are often overexpressed in cancer cells.¹⁷⁶ In myeloproliferative neoplasms (MPNs), the combination of partial loss of PRC2 and expression of the constitutively active oncoprotein NRASG12D — a member of the Ras GTPase family — has been shown to lead to BCAT1 expression, which is normally repressed in hematopoietic stem cells.¹⁷⁴ The increase in BCAT1 results in a larger pool of BCAAs that activates mTOR, a protein kinase known to participate in tumor growth and proliferation.¹⁷⁷ It should be noted that in patients with Acute Myeloid Leukemia (AML), the expression of EZH2 and *BCAT1* is inversely correlated, a high expression of BCAT1 being associated with a poor survival outcome.¹⁷⁴ In glioblastoma cancer cells, rather than modulating BCAT
<figure><img src="image_17.png" /></figure>

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<figure><img src="image_19.png" /></figure>

<figure><img src="image_20.png" /></figure>

expression, it is the BCAA pool that increases. In this case, EZH2 represses EAF2 which inhibits Hypoxia-Inducible Factor 1 (HIF1).<sup>178</sup> HIF-1 overexpression participates in the Warburg effect by supporting glycolytic metabolism and upregulating expression of LAT1, the main transporter of BCAAs, which results in an increase in BCAA pool<sup>178,179</sup> (Fig. 5a, right).

The Warburg effect is the most well-known cancer metabolic alteration, whereby malignant cells use glycolysis rather than oxygen-dependent metabolism. A tight regulation of glucose homeostasis is essential to counter the proliferation of cancer cells. An important node in this pathway is the reaction catalyzed by the enzyme Fructose-1,6-biphosphatase (FBP1). Low FBP1
**Fig. 5** Environment-dependent oncogenic activities of PcG proteins. **a** Left: in a physiological condition, the membrane transporter LAT1 participates in the transport of methionine which reacts with ATP to produce SAM. SAM can in turn be used by PRC2 to induce trimethylation of H3K27, resulting in a PcG-mediated silencing of its targets genes. *Lat1* expression depends on RXRα. Right: in cancer cells, *Lat1* is overexpressed, enhancing SAM production and inducing H3K27 hypermethylation of the chromatin landscape. The *Lat1* negative regulator, RXRα, is thus repressed resulting in a positive feedback loop whereby EAF2 transcriptional silencing dependent on PRC2 results in overexpression of HIF1, which can in turn stimulate *Lat1* expression. Therefore, an excess of LAT1 at the cellular membrane increases the transport of BCAAs, thereby enhancing protein synthesis. **b** Controlling the immune system is of a major importance in cancer. Cancer cells use different mechanisms to do this. First, PRC1 is able to increase the transcriptional expression of CCL2, which will dampen Treg immune response. In addition, PRC2-mediated silencing of the MHC-I antigen processing pathway results in MHC-I absence at the cell membrane, concealing cancer cells from cytotoxic T cells. Finally, PCGF4 overexpression in cancer cells stimulates the expression of GATA2, which will inhibit MICA/B transcription and reduces its presence at the membrane. This prevents the recognition of cancer cells by NK cells. These mechanisms enhance the immunosuppressive response and inhibit the cytotoxic response that would otherwise kill the cancer cells. **c** Oncohistones are a new line of research, analyzing the effect of mutations on histone genes that could have an impact on tumorigenesis. H3K27M has a dominant negative effect on EZH2 catalytic activity. Left: in a wild-type condition, PRC2 is recruited to nucleation sites that present unmethylated CGIs. Trimethylation of H3K27 occurs and spreads around the nucleation site. The boundaries of Polycomb domains are decorated with H3K36me2. Right: in the presence of the H3K27M oncohistone, that represents 10% of all H3, an epigenetic remodeling occurs. The spreading of H3K27me3 is inhibited and active histone marks, such as H3K27ac, are present on the oncohistone. BCAAs, Branched-chain amino acids; CCL2, C-C motif chemokine ligand 2; CCR2, C-C motif chemokine receptor; EAF2, ELL associated factor 2; GATA2, GATA binding protein 2; HIF1, Hypoxia inducible factor 1; IDH1, Isocitrate dehydrogenase 1; KDM6A/B, Lysine demethylase 6A/B; LAT1, L-type amino acid transporter 1; MHC-I, Major histocompatibility complex I; MHC-I APP, Major histocompatibility complex I antigen processing pathway; MICA/B, MHC I polypeptide-related sequence A/B; RXRα, Retinoid X receptor-alpha; SAM, S-adenosylmethionine; SAH, S-adenosylhomocysteine; TCR, T-cell receptor; SETD2, SET domain containing 2 (histone lysine methyltransferase).

enzyme activity correlates with higher production of pyruvate, the downstream product of the glycolysis pathway. An overproduction of pyruvate corresponds to a greater store of energy available for cancer cell growth. In hepatocellular carcinoma and clear cell renal cell carcinoma, the mRNA levels of *Ezh2* and *FBP1* are inversely correlated due to the presence of the EZH2-dependent H3K27me3 repressive mark at the promoter of the gluconeogenic enzyme-coding gene.¹⁸⁰ Tumor growth was shown to be thwarted either by a short-hairpin RNA (shRNA) directed against *Ezh2*, or by the reintroduction of FBP1. Interestingly, FBP1 and EZH2 interact directly. In doing so, FBP1 is able to reduce the methyltransferase activity of EZH2 by dissociating the PRC2 complex. This double negative feedback loop provides new insights into the involvement of Polycomb in "oncometabolism".

Metabolic reprogramming during tumorigenesis is required to better sustain the energy necessary for cancer progression and survival.¹⁸¹,¹⁸² PcG proteins have been implicated in the regulation of metabolic genes involved in metabolism of fatty acids and pyruvates among others.¹³⁹,¹³⁸ While the link between PRC2 function and metabolism in physiology and cancer is certain, much work remains to be done in order to understand the molecular underpinnings of this link in different cancer types and to harness them to design effective therapeutic strategies. Noteworthy, with most of the current research focusing on the link between PRC2 and metabolism,¹⁸⁴ it might also be of interest to examine the involvement of PRC1 in future work.

Immune system

The immune system has a wide array of cells that protect from foreign bodies, also known as *non-self*. Innate immune cells provide a rapid and nonspecific response while adaptive immune cells have a slower response that relies on a memory process that will be specific to a known foreign object.¹⁸⁵ In principle, both innate and adaptive immune cells exert an anti-tumor function.

However, cancer cells can develop multiple mechanisms to evade recognition and destruction by the immune system and become resistant to therapy. In prostate cancer, elevated PRC1 levels and activity coincide with epithelial-to-mesenchymal transition (EMT) and stemness signatures. PRC1 directly promotes metastasis at metastatic initiation sites by controlling self-renewal and both cPRC1 and ncPRC1.1 components directly induce transcriptional expression of CCL2 and other pro-metastatic genes that encode cytokines, which suppress the immune response and promote a pro-angiogenic environment¹⁸⁶ (Fig. 5b). CCL2

expression has an oncogenic function by recruiting immune cells such as M2-type Tumor-Associated Macrophages (TAMs) and T-regulatory cells (Tregs), promoting an immunosuppressive microenvironment favorable to tumor progression. Moreover, Natural Killer (NK) cells are also involved in the innate immune response. Upon recognition of MICA/B by NK cells, an immune cytotoxic response is displayed. However, BMI-1 stimulates GATA2 expression which in turn directly inhibits MICA/B expression (Fig. 5b). Reduction of MICA/B expression on the surface of cancer cells prevents NK cell activation and the cytotoxic response.¹⁸⁷ The combination of these two escape mechanisms promotes cancer cell progression and metastasis. Pharmacological treatment using a catalytic inhibitor of PRC1 suppresses metastasis by reverting the immunosuppressive microenvironment and promoting the recruitment of NK cells and T effector cells.¹⁸⁶,¹⁸⁷

Cytotoxic T cells (CD8⁺ T) identify cancer cells presenting foreign antigens by their Major Histocompatibility Complex I (MHC-I). An IFN-γ response is then induced to kill the cancer cells. In order to survive, cancer cells downregulate the MHC-I antigen processing pathway (MHC-I APP), resulting in decreased presentation of foreign antigens to CD8⁺ T¹⁸⁸ (Fig. 5b). PRC2 represses transcription of various MHC-I APP components, participating in cancer cell immunosurveillance escape.¹⁸⁹ Furthermore, PRC2 inhibits anti-tumor immunity by altering the transcriptional landscape of Tregs. Indeed, immunocompetent mice bearing tumors treated with an EZH2 inhibitor show a significant decrease in tumor volume compared to mice deficient in T cells, suggesting an interplay between EZH2 and the T cell immune response.¹⁹⁰ Tregs promote tumor progression in an EZH2-dependent manner by producing immunosuppressive cytokines and preventing recruitment of T CD8⁺.¹⁹⁰,¹⁹¹ Pharmacological EZH2 inhibition induces a change in the production of pro-inflammatory cytokines which promotes anti-tumor activity and significantly increases the ratio between CD8⁺ T and Tregs in the tumor microenvironment.¹⁹⁰

Cancer immunotherapy has revolutionized the clinical approach in the field of oncology. However, anti-CTLA4, the first monoclonal antibody used in cancer therapy as an immune checkpoint, induces an upregulation of EZH2 expression¹⁹¹ which may prevent anti-tumor immunity by inducing an immunosuppressive tumor microenvironment.¹⁹⁰,¹⁹¹ A synergistic strategy coupling anti-CTLA4 and an EZH2 inhibitor reverses cancer resistance to the immune system.¹⁹¹,¹⁹² Moreover, considering the involvement of PcG proteins in pluripotency, it is not surprising that PcG proteins are also involved in cancer stem cell (CSC) development and
resistance to treatment. Although anti-PD1 immunotherapy is sufficient to recruit CD8⁺T cells into the tumor microenvironment, it is not sufficient to kill BMI-1⁺CSCs.¹⁹³ Inhibition of BMI-1 represses H2AK119ub-decorated target genes and increases DNA-damage, stimulating the inflammatory response and CD8⁺T cells recruitment.¹⁹³ In summary, joint targeting of immune checkpoints and PcG proteins appears to be a new promising therapeutic approach to efficiently counter cancer progression by stimulating the immune response.¹⁹¹⁻¹⁹³

## Oncohistones

As already mentioned, the catalytic activities of "writers" and "erasers" enzymes that modify histone PTMs are often dysregulated in cancer where chromatin landscapes are modified, resulting in aberrant transcription of the corresponding genes.⁹⁹,¹⁹⁴ In addition, the lack of recognition of H3K27me3 by the CBX7 "reader" results in a transcriptional de-repression of tumor suppressor genes.¹⁹⁵ Likewise, the BAHCC1 mutation in its BAH domain leads to upregulation of tumor suppressor genes that dampen tumor progression.³⁸

These data point to a direct involvement of histone modifications in tumorigenesis. Indeed, somatic mutations in histone genes occur at high frequency in cancer, and they can exhibit oncogenic properties.¹⁹⁶ K-to-M/I missense substitutions in histone variants, analyzed from available sequenced genomes of several human cancer types of ~3000 patients, further argue for a driver or contributor effects of the known N-terminal tail mutations affecting H3.¹⁹⁶ These mutations are particularly frequent in rare malignancies such as glioma and chondroblastoma. This analysis allowed detection of previously unappreciated situations where histones are mutated at low frequency in common cancers, like H3K27M in melanoma and AML.¹⁹⁶

One of the most studied cancer-associated "oncohistones" carries the H3K27M substitution, whereby H3 lysine 27 is mutated to methionine, a missense mutation showing high genetic penetrance in pediatric glioblastomas¹⁹⁷,¹⁹⁸ (Fig. 5c). It is noteworthy that different H3 mutants are found in distinct locations. Indeed, H3F3A mutations such as H3.3K27M or H3.3G34R/V are found respectively in midline pediatric high-grade gliomas and cortex, whereas HIST1H3B mutations affecting the canonical H3.1 are restricted to the brainstem.¹⁹⁹ H3F3A which encodes the histone variant H3.3 is found mutated in 60% of Diffuse Intrinsic Pontine Glioma (DIPG) cases¹⁹⁸ and this mutation is suggested to be the first hit in DIPG tumorigenesis.²⁰⁰ This driver mutation is associated with obligate partner mutations throughout tumor progression,²⁰⁰ in particular in the cell cycle regulatory gene TP53 or the chromatin remodeler ATRX. Interestingly, while H3.3K27M represents less than 10% of total H3, this level is sufficient to induce a significant decrease in the trimethylated state of H3K27, leading to a decrease in PcG-dependent transcriptional silencing.²⁰¹,²⁰²

The epigenome is drastically altered in an H3K27M context. Indeed, while H3K27me3 is specifically restricted to unmethylated CGIs and H3K27me3/2 levels are significantly decreased, the monomethylation level of H3K27 remains unchanged.²⁰³ Intriguingly, H3K27me1 distribution is completely rewired in an H3K27M context.²⁰³ Moreover, just like in a wild-type H3 context, H3K36me2 restricts the spreading of H3K27me2/3.²⁰³ Furthermore, H3K27Ac levels are globally increased at the H3K27M location.²⁰¹,²⁰⁴ This suggests that H3K27M has a dominant negative effect on the catalytic activity of the EZH2 methyltransferase.²⁰¹,²⁰²

There is a strong interest in understanding the molecular mechanisms by which oncohistone mutations change the epigenome and impact gene expression. PRC2 was proposed to have a higher affinity for the mutated histone, which binds the EZH2 enzymatic domain, inhibiting its methyltransferase activity.²⁰¹,²⁰⁵,²⁰⁶ However, the mechanism by which H3K27M oncohistones inhibits PRC2 activity is still under debate.²⁰⁷ The finding that PRC2 appears

to be excluded from the H3K27M-K27Ac domains²⁰⁸ argues against the model of PRC2 sequestration by H3K27M. Moreover, while it is suggested that gliomagenesis is dependent on PRC2 inhibition,²⁰¹ it has been demonstrated that loss of PRC2 disables growth and colony formation in H3K27M-positive DIPG cells, underlying the importance of PRC2 in tumor maintenance.²⁰⁸

Interestingly, CATACOMB/EZHIP, a PRC2 co-factor, either via its overexpression or a chromosomal translocation inducing its fusion with the NuA4 subunit gene MBTD1, described in low-grade endometrial stromal sarcoma,²⁰⁹ decreases PRC2-dependent methyltransferase activity.⁹ CATACOMB/EZHIP-dependent hypo-methylation is due to a conserved methionine residue M406 which inhibits EZH2, mimicking the H3K27M oncohistone.⁹ Moreover, H3K27M and CATACOMB/EZHIP are mutually exclusive in gliomas, specifically in Posterior Fossa A (PFA) ependymomas.²¹⁰ Both are suggested to decrease H3K27 trimethylation by blocking the spreading of the repressive mark from CGIs.²¹¹

In Giant Cell Tumor of the bone (GCT), the oncohistone H3.3G34W is encountered in 90% of cases.²¹² Interestingly, this residue is not post-translationally modified but its impact on the epigenome is undeniable. This mutation leads to loss of H3K36me3 which counteracts H3K27me3 deposition by PRC2.²¹² As a consequence, a redistribution of the H3K27me3 repressive mark occurs from intergenic to genic regions, resulting in perturbation in Polycomb-mediated silencing and in the maintenance of a progenitor state of the mutated cells.²¹²

As previously mentioned, a crosstalk exists between H3K36me2/3 and H3K27me3 and this interplay remains in the presence of the H3K36M oncohistones, in which lysine 36 of the histone 3 is replaced by a methionine. This mutation is found in 95% of chondroblastomas and 92% of GCT, respectively in the H3F3B and H3F3A genes.²¹³ Following the oncohistone paradigm, the H3K36me2/3 PTMs are reduced due to the inability of specific methyltransferases, namely SETD2, NSD1-NSD3, to deposit their marks.²¹⁴,²¹⁵ H3K36M reduces H3K36 methylation and increases nucleosome availability for PRC2 to deposit H3K27me3.²¹⁴ The genome-wide increase in this repressive mark then induces a PRC1 redeployment which overall dilutes PRC1 at its canonical binding sites, leading to de-repression of self-renewal genes²¹⁴,²¹⁶ (Fig. 5c). Similarly, in human papillomavirus (HPV)-negative head and neck squamous cell carcinomas (HNSCCs),²¹⁷ the H3K36 methylation state is involved in oncogenic promotion.²¹⁷ NSD1 writer mutations, similarly to H3K36M, remodel the chromatin landscape by decreasing H3K36me2 levels. Considering the interplay between H3K36me2 and H3K27me3, Polycomb components would be expected to be involved in HNSCCs. However, the precise mechanism at play is yet to be characterized.

All these data show that the emerging oncohistone field is an important area of oncology, but much remains to be done and a current strong focus is on the investigation of how histone mutations contribute to epigenome reprogramming and whether these mutations are primarily drivers or contributors of tumorigenesis in a wide range of human cancers.¹⁹⁴

## Non-genetic drug resistance in cancer

The ability of cancer cells to adapt to or resist anti-cancer therapies may be inherently of a genetic nature or may be acquired during treatment.²¹⁸ Alongside an undergoing genetic evolution of cancer genomes, cancer cells can also be modified in their epigenetic landscapes and this non-genetic contribution can play a major role in cancer resistance. In fact, relapsing patients often do not present specific mutations that would explain a lower efficiency for the same therapy.²¹⁹,²²⁰

Cancer cells are actually able to evolve and change completely their transcriptional landscape to adapt to treatment-induced stress. Polycomb implication in cancer drug-resistance depends on PRC2 and its catalytic activity as well as on other concomitant mechanisms that can induce transcriptional plasticity.²²¹⁻²²³
In multiple myeloma (MM), cell adhesion-mediated drug resistance (CAM-DR) develops when malignant plasma cells interact with stromal cells in the bone marrow and become less sensitive to chemotherapy.²²⁴ In an in vitro system that recapitulates CAM-DR, anti-MM treatment results in an increase and redistribution of H3K27me3 in a dose-dependent manner in cultured MM cells only when they do not adhere to stromal cells.²²⁴ CAM-DR counteracts drug-induced H3K27 hypermethylation via phosphorylation of EZH2 at serine 21, leading to overexpression of anti-apoptotic genes which participate in survival and drug-resistance.²²⁴ In addition, *mir-15a* downregulation triggers PHF19 upregulation in relapsed MM patients.²²⁵ The involvement of PHF19 in drug-resistance might depend on its ability to stimulate proliferation by promoting EZH2 serine 21 phosphorylation, which inhibits the H3K27me3 deposition and leads to upregulation of genes linked to cell growth.²²⁵

In Testicular Germ Cell Tumors (TGCT), resistance to cisplatin is accompanied by a global decrease in H3K27me3 and H2AK119Ub levels, leading to upregulation of Polycomb target genes.²²³ Inhibition of the UTX and JMD3 enzymes, responsible for H3K27 demethylation, is sufficient to increase H3K27me3 and make TGCT cells more sensitive to the initial chemotherapy.²²³

In AML, therapeutic resistance can arise in the apparent absence of new genetic mutations and is antagonized by inhibiting Lsd1, a demethylase chromatin modulator involved in the regulation of enhancer activity.²¹⁹ Inhibition of Lsd1 creates enhancer switching, generating new binding sites for pioneer factors that ultimately activate the enhancers of key drug resistance genes. Inhibition of a key chromatin modulator in AML then makes it possible to resensitize cells to the primary treatment.²¹⁹

Unlike mutations, failed or disrupted epigenetic mechanisms can be quite easily reverted using epidrugs to overcome cancer progression by rewiring malignant epigenomes, either to resensitize tumor cells resistant to conventional therapy or to sensitize them to new therapies. Given the importance of PcG proteins in transcriptional regulation, it will therefore be of great interest to further characterize the mechanisms by which PcG proteins contribute to drug-resistance. In particular, it will be important to expand research aimed at understanding Polycomb functions at enhancers,¹³⁹,²²⁶ since they might be involved in various cancer types and stages.

## CONCLUSIVE REMARKS: POLYCOMB EPIGENETICS IN CANCER

Although it is commonly assumed that cancer arises from a set of multiple mutations, a pan-cancer analysis established that about 5% of cancer cases did not have driver mutations that could explain tumorigenesis, pointing out that genetics might not be the only player in cancer.²²⁷ Non-genetic alterations appear to represent an alternative path toward the development, progression and drug-resistance of cancer cells. In pancreatic ductal adenocarcinoma, metastases do not show driver gene mutations but rather follow drastic epigenomic reprogramming,¹⁸¹,²²⁸ suggesting that epigenetic modifiers are mainly involved. Additionally, ependymomas — a childhood brain tumor — are characterized by a very low mutation rate,²²⁹ suggesting that cancer is not only a consequence of DNA mutations, but rather emerges and evolves from a crosstalk between genetic and non-genetic processes. In an extreme view, cancer has been defined as an "epigenetic disease".²³⁰ It would therefore not be surprising to find misregulated Polycomb proteins as epi-drivers in tumorigenesis.

PcG proteins have imposed themselves in a wide range of biological processes. Clearly, they are landmark components in the field of cancer research and we have only started to understand the extent of their oncogenic functions. As most research focuses on the EZH2 catalytic subunit of PRC2, it will be interesting to better characterize the involvement of the different PRC2 subunits as well as on the many flavors of PRC1 complexes. Nonetheless, a

fascinating part of the oncogenic function of PcG components relies on the fact that some of them can act both in a manner dependent or independent on Polycomb complexes.²³¹ It will be interesting to better characterize these PcG functions at the molecular level in order to have a complete picture of their mode of action. While it is clear that the overexpression or down-regulation of PcG proteins is involved in cancer, it will be important to characterize how modifying protein stability by either PTMs and/or interaction with yet unidentified partners might be implicated in tumorigenesis. Finally, context is of paramount importance: misregulation of Polycomb proteins results in different, sometimes even opposing results in different cancer types. Identifying molecular pathways leading to these context-dependent effects will be crucial in order to improve cancer diagnosis, prognosis and therapy.

## REFERENCES

1. Loubiere, V., Martinez, A. M. & Cavalli, G. Cell fate and developmental regulation dynamics by polycomb proteins and 3D genome architecture. *BioEssays* **41**, 1–15 (2019).

2. Chan, H. L. & Morey, L. Emerging roles for polycomb-group proteins in stem cells and cancer. *Trends Biochem. Sci.* **44**, 688–700 (2019).

3. Piunti, A. & Shilatifard, A. The roles of polycomb repressive complexes in mammalian development and cancer. *Nat. Rev. Mol. Cell Biol.* **22**, 326–345 (2021).

4. Lewis, E. B. A gene complex controlling segmentation in Drosophila. *Nature* **276**, 565–570 (1978).

5. Levine, S. S. et al. The core of the polycomb repressive complex is compositionally and functionally conserved in flies and humans. *Mol. Cell. Biol.* **22**, 6070–6078 (2002).

6. Schuettengruber, B., Bourbon, H. M., Di Croce, L. & Cavalli, G. Genome regulation by polycomb and trithorax: 70 years and counting. *Cell* **171**, 34–57 (2017).

7. Kuzmichev, A., Nishioka, K., Erdjument-Bromage, H., Tempst, P. & Reinberg, D. Histone methyltransferase activity associated with a human multiprotein complex containing the Enhancer of Zeste protein. *Genes Dev* **16**, 2893–2905 (2002).

8. Ragazzini, R. et al. EZHIP constrains Polycomb Repressive Complex 2 activity in germ cells. *Nat. Commun.* **10**, 3858 (2019).

9. Piunti, A. et al. Catacomb: An endogenous inducible gene that antagonizes H3K27 methylation activity of Polycomb repressive complex 2 via an H3K27M-like mechanism. *Sci. Adv.* **5**, eaax2887 (2019).

10. Wang, H. et al. Role of histone H2A ubiquitination in Polycomb silencing. *Nature* **431**, 873–878 (2004).

11. Cohen, I., Bar, C. & Ezhkova, E. Activity of PRC1 and histone H2AK119 monoubiquitination: revising popular misconceptions. *BioEssays* **42**, 1–8 (2020).

12. Shao, Z. et al. Stabilization of chromatin structure by PRC1, a Polycomb complex. *Cell* **98**, 37–46 (1999).

13. Garcia, E., Marcos-Gutiérrez, C., Del Mar Lorente, M., Moreno, J. C. & Vidal, M. RYBP, a new repressor protein that interacts with components of the mammalian Polycomb complex, and with the transcription factor YY1. *EMBO J* **18**, 3404–3418 (1999).

14. Wang, R. et al. Polycomb group targeting through different binding partners of RING1B C-terminal domain. *Structure* **18**, 966–975 (2010).

15. Gao, Z. et al. PCGF homologs, CBX proteins, and RYBP define functionally distinct PRC1 family complexes. *Mol. Cell* **45**, 344–356 (2012).

16. Simon, J., Chiang, A., Bender, W., Shimell, M. J. & O'Connor, M. Elements of the Drosophila bithorax complex that mediate repression by Polycomb group products. *Dev. Biol.* **158**, 131–144 (1993).

17. Van Kruijsbergen, I., Hontelez, S. & Veenstra, G. J. C. Recruiting polycomb to chromatin. *Int. J. Biochem. Cell Biol* **67**, 177–187 (2015).

18. Kassis, J. A. & Brown, J. L. Polycomb group response elements in Drosophila and vertebrates. *Adv. Genet.* **81**, 83–118 (2013).

19. Pengelly, A. R., Copur, Ö., Jäckle, H., Herzig, A. & Müller, J. A histone mutant reproduces the phenotype caused by loss of histone-modifying factor Polycomb. *Science* **339**, 698–699 (2013).

20. Min, J., Zhang, Y. & Xu, R. M. Structural basis for specific binding of Polycomb chromodomain to histone H3 methylated at Lys 27. *Genes Dev* **17**, 1823–1828 (2003).

21. He, J. et al. Kdm2b maintains murine embryonic stem cell status by recruiting PRC1 complex to CpG islands of developmental genes. *Nat. Cell Biol.* **15**, 373–384 (2013).

22. Blackledge, N. P. et al. Variant PRC1 complex-dependent H2A ubiquitylation drives PRC2 recruitment and polycomb domain formation. *Cell* **157**, 1445–1459 (2014).
23. Cooper, S. et al. Jarid2 binds mono-ubiquitylated H2A lysine 119 to mediate crosstalk between Polycomb complexes PRC1 and PRC2. *Nat. Commun.* **7**, 13661 (2016).

24. Tamburri, S. et al. Histone H2AK119 mono-ubiquitination is essential for Polycomb-mediated transcriptional repression. *Mol. Cell* **77**, 840–856.e5 (2020).

25. Sing, A. et al. A vertebrate Polycomb response element governs segmentation of the posterior hindbrain. *Cell* **138**, 885–897 (2009).

26. Woo, C. J., Kharchenko, P. V., Daheron, L., Park, P. J. & Kingston, R. E. A region of the human HOXD cluster that confers polycomb-group responsiveness. *Cell* **140**, 99–110 (2010).

27. Ku, M. et al. Genomewide analysis of PRC1 and PRC2 occupancy identifies two classes of bivalent domains. *PLoS Genet* **4**, e1000242 (2008).

28. Mendenhall, E. M. et al. GC-rich sequence elements recruit PRC2 in mammalian ES cells. *PLoS Genet* **6**, e1001244 (2010).

29. Riising, E. M. et al. Gene silencing triggers polycomb repressive complex 2 recruitment to CpG Islands genome wide. *Mol. Cell* **55**, 347–360 (2014).

30. Li, H. et al. Polycomb-like proteins link the PRC2 complex to CpG islands. *Nature* **549**, 287–291 (2017).

31. Hunkapiller, J. et al. Polycomb-like 3 promotes polycomb repressive complex 2 binding to CpG islands and embryonic stem cell self-renewal. *PLoS Genet* **8**, e1002576 (2012).

32. Perino, M. et al. MTF2 recruits Polycomb Repressive Complex 2 by helical-shape-selective DNA binding. *Nat. Genet.* **50**, 1002–1010 (2018).

33. Chen, S., Jiao, L., Liu, X., Yang, X. & Liu, X. A dimeric structural scaffold for PRC2-PCL targeting to CpG Island chromatin. *Mol. Cell* **77**, 1265–1268.e7 (2020).

34. Hofjeldt, J. W. et al. Non-core subunits of the PRC2 Complex are collectively required for its target-sitespecificity. *Mol. Cell* **76**, 423–436.e3 (2019).

35. Bierne, H. et al. Human BAHD1 promotes heterochromatic gene silencing. *Proc. Natl. Acad. Sci. USA* **106**, 13826–13831 (2009).

36. Zhao, D. et al. The BAH domain of BAHD1 is a histone H3K27me3 reader. *Protein Cell* **7**, 222–226 (2016).

37. Fan, H. et al. A conserved BAH module within mammalian BAHD1 connects H3K27me3 to Polycomb gene silencing. *Nucleic Acids Res* **49**, 4441–4455 (2021).

38. Fan, H. et al. BAHCC1 binds H3K27me3 via a conserved BAH module to mediate gene silencing and oncogenesis. *Nat. Genet.* **52**, 1384–1396 (2020).

39. Lebreton, A. et al. A bacterial protein targets the BAHD1 chromatin complex to stimulate type III interferon response. *Science* **331**, 1319–1321 (2011).

40. Achour, C. & Aguilo, F. Long non-coding RNA and Polycomb: an intricate partnership in cancer biology. *Front. Biosci.* **23**, 2106–2132 (2018).

41. Yu, M. et al. Direct recruitment of polycomb repressive complex 1 to chromatin by core binding transcription factors. *Mol. Cell* **45**, 330–343 (2012).

42. Cenik, B. K. & Shilatifard, A. COMPASS and SWI/SNF complexes in development and disease. *Nat. Rev. Genet.* **22**, 38–58 (2021).

43. Douillet, D. et al. Uncoupling histone H3K4 trimethylation from developmental gene expression via an equilibrium of COMPASS, Polycomb and DNA methylation. *Nat. Genet.* **52**, 615–625 (2020).

44. Brinkman, A. B. et al. Sequential ChIP-bisulfite sequencing enables direct genome-scale investigation of chromatin and DNA methylation cross-talk. *Genome Res* **22**, 1128–1138 (2012).

45. Streubel, G. et al. The H3K36me2 methyltransferase Nsd1 demarcates PRC2-mediated H3K27me2 and H3K27me3 domains in embryonic stem cells. *Mol. Cell* **70**, 371–379 (2018).

46. Francis, N. J., Kingston, R. E. & Woodcock, C. L. Chromatin compaction by a polycomb group protein complex. *Science* **306**, 1574–1577 (2004).

47. Trojer, P. et al. L3MBTL2 protein acts in concert with PcG protein-mediated monoubiquitination of H2A to establish a repressive chromatin structure. *Mol. Cell* **42**, 438–450 (2011).

48. Grau, D. J. et al. Compaction of chromatin by diverse polycomb group proteins requires localized regions of high charge. *Genes Dev* **25**, 2210–2221 (2011).

49. Cheutin, T. & Cavalli, G. Loss of PRC1 induces higher-order opening of Hox loci independently of transcription during Drosophila embryogenesis. *Nat. Commun.* **9**, 3898 (2018).

50. Eskeland, R. et al. Ring1B compacts chromatin structure and represses gene expression independent of histone ubiquitination. *Mol. Cell* **38**, 452–464 (2010).

51. Blackledge, N. P. et al. PRC1 catalytic activity is central to polycomb system function. *Mol. Cell* **77**, 857–874.e9 (2020).

52. Zhou, W. et al. Histone H2A monoubiquitination represses transcription by inhibiting RNA polymerase II transcriptional elongation. *Mol. Cell* **29**, 69–80 (2008).

53. Kadoch, C. et al. Dynamics of BAF-polycomb complex opposition on heterochromatin in normal and oncogenic states. *Nat. Genet.* **49**, 213–222 (2017).

54. Hodges, H. C. et al. Dominant-negative SMARCA4 mutants alter the accessibility landscape of tissue-unrestricted enhancers. *Nat. Struct. Mol. Biol.* **25**, 61–72 (2018).

55. Dellino, G. I. et al. Polycomb silencing blocks transcription initiation. *Mol. Cell* **13**, 887–893 (2004).

56. Stock, J. K. et al. Ring1-mediated ubiquitination of H2A restraints poised RNA polymerase II at bivalent genes in mouse ES cells. *Nat. Cell Biol.* **9**, 1428–1435 (2007).

57. Ardehali, M. B. et al. Polycomb repressive complex 2 methylates elongin A to regulate transcription. *Mol. Cell* **68**, 872–884.e6 (2017).

58. Nakagawa, T. et al. Deubiquitylation of histone H2A activates transcriptional initiation via trans-histone cross-talk with H3K4 di- and trimethylation. *Genes Dev* **22**, 37–49 (2008).

59. Tie, F. et al. CBP-mediated acetylation of histone H3 lysine 27 antagonizes Drosophila Polycomb silencing. *Development* **136**, 3131–3141 (2009).

60. Tie, F. et al. Polycomb inhibits histone acetylation by CBP by binding directly to its catalytic domain. *Proc. Natl. Acad. Sci. USA* **113**, E744–E753 (2016).

61. Tie, F. et al. Trithorax monomethylates histone H3K4 and interacts directly with CBP to promote H3K27 acetylation and antagonize polycomb silencing. *Development* **141**, 1129–1139 (2014).

62. Ferrari, K. J. et al. Polycomb-dependent H3K27me1 and H3K27me2 regulate active transcription and enhancer fidelity. *Mol. Cell* **53**, 49–62 (2014).

63. Lanzuolo, C., Roure, V., Dekker, J., Bantignies, F. & Orlando, V. Polycomb response elements mediate the formation of chromosome higher-order structures in the bithorax complex. *Nat. Cell Biol.* **9**, 1167–1174 (2007).

64. Ogiyama, Y., Schuettengruber, B., Papadopoulos, G. L., Chang, J. M. & Cavalli, G. Polycomb-dependent chromatin looping contributes to gene silencing during Drosophila development. *Mol. Cell* **71**, 73–88.e5 (2018).

65. Wani, A. H. et al. Chromatin topology is coupled to polycomb group protein subnuclear organization. *Nat. Commun.* **7**, 10291 (2016).

66. Kundu, S. et al. Polycomb repressive complex 1 generates discrete compacted domains that change during differentiation. *Mol. Cell* **65**, 432–446.e5 (2017).

67. Bonev, B. et al. Multiscale 3D genome rewiring during mouse neural development. *Cell* **171**, 557–572.e24 (2017).

68. Schoenfelder, S. et al. Polycomb repressive complex PRC1 spatially constrains the mouse embryonic stem cell genome. *Nat. Genet.* **47**, 1179–1186 (2015).

69. Pherson, M. et al. Polycomb repressive complex 1 modifies transcription of active genes. *Sci. Adv.* **3**, e1700944 (2017).

70. Giner-Laguarda, N. & Vidal, M. Functions of polycomb proteins on active targets. *Epigenomes* **4**, 17 (2020).

71. Morey, L., Aloia, L., Cozzuto, L., Benitah, S. A. & Di Croce, L. RYBP and Cbx7 define specific biological functions of polycomb complexes in mouse embryonic stem cells. *Cell Rep* **3**, 60–69 (2013).

72. Frangini, A. et al. The aurora B kinase and the polycomb protein ring1B combine to regulate active promoters in quiescent lymphocytes. *Mol. Cell* **51**, 647–661 (2013).

73. Schaaf, C. A. et al. Cohesin and polycomb proteins functionally interact to control transcription at silenced and active genes. *PLoS Genet* **9**, e1003560 (2013).

74. Loubiere, V. et al. Coordinate redeployment of PRC1 proteins suppresses tumor formation during Drosophila development. *Nat. Genet.* **48**, 1436–1442 (2016).

75. Gao, Z. et al. An AUTS2-Polycomb complex activates gene expression in the CNS. *Nature* **516**, 349–354 (2014).

76. Liu, S *et al.* NRF1 association with AUTS2-Polycomb mediates specific gene activation in the brain. *Mol. Cell* **81**, 4663–4676.e8 (2021).

77. Morey, L. et al. Polycomb regulates mesoderm cell fate-specification in embryonic stem cells through activation and repression mechanisms. *Cell Stem Cell* **17**, 300–315 (2015).

78. Varambally, S. et al. The polycomb group protein EZH2 is involved in progression of prostate cancer. *Nature* **419**, 624–629 (2002).

79. Kim, K. H. & Roberts, C. W. M. Targeting EZH2 in cancer. *Nat. Med.* **22**, 128–134 (2016).

80. Herviou, L., Cavalli, G., Cartron, G., Klein, B. & Moreaux, J. EZH2 in normal hematopoiesis and hematological malignancies. *Oncotarget* **7**, 2284–2296 (2016).

81. Yamaguchi, H. & Hung, M. Regulation and role of EZH2 in cancer. *Cancer Res. Treat.* **46**, 209–222 (2014).

82. Safaei, S., Baradaran, B., Hagh, M. F. & Alivand, M. R. Double sword role of EZH2 in leukemia. *Biomed. Pharmacother.* **98**, 626–635 (2018).

83. Xie, H. et al. Polycomb repressive complex 2 regulates normal hematopoietic stem cell function in a developmental-stage-specific manner. *Cell Stem Cell* **14**, 68–80 (2014).

84. Lee, S. C. W. et al. Polycomb repressive complex 2 component Suz12 is required for hematopoietic stem cell function and lymphopoiesis. *Blood* **126**, 167–175 (2015).

85. Mochizuki-Kashio, M. et al. Ezh2 loss in hematopoietic stem cells predisposes mice to develop heterogeneous malignancies in an Ezh1-dependent manner. *Blood* **126**, 1172–1183 (2015).
86. Yu, W. et al. Depletion of polycomb repressive complex 2 core component EED impairs fetal hematopoiesis. *Cell Death Dis* **8**, e2744 (2017).

87. Brecqueville, M. et al. Mutations and deletions of the SUZ12 polycomb gene in myeloproliferative neoplasms. *Blood Cancer J* **1**, e33 (2011).

88. Béguelin, W. et al. EZH2 is required for germinal center formation and somatic EZH2 mutations promote lymphoid transformation. *Cancer Cell* **23**, 677–692 (2013).

89. Okosun, J. et al. Integrated genomic analysis identifies recurrent mutations and evolution patterns driving the initiation and progression of follicular lymphoma. *Nat. Genet.* **46**, 176–181 (2014).

90. Ueda, T. et al. Propagation of trimethylated H3K27 regulated by polycomb protein EED is required for embryogenesis, hematopoietic maintenance, and tumor suppression. *Proc. Natl. Acad. Sci. USA* **113**, 10370–10375 (2016).

91. Béguelin, W. et al. EZH2 and BCL6 cooperate to assemble CBX8-BCOR complex to repress bivalent promoters, mediate germinal center formation and lymphomagenesis. *Cancer Cell* **30**, 197–213 (2016).

92. Broux, M. et al. Suz12 inactivation cooperates with JAK3 mutant signaling in the development of T-cell acute lymphoblastic leukemia. *Blood* **134**, 1323–1336 (2019).

93. Muto, T. et al. Concurrent loss of Ezh2 and Tet2 cooperates in the pathogenesis of myelodysplastic disorders. *J. Exp. Med.* **210**, 2627–2639 (2013).

94. Wang, C. et al. Ezh2 loss propagates hypermethylation at T cell differentiation-regulating genes to promote leukemic transformation. *J. Clin. Invest.* **128**, 3872–3886 (2018).

95. Yan, J. et al. EZH2 overexpression in natural killer/T-cell lymphoma confers growth advantage independently of histone methyltransferase activity. *Blood* **121**, 4512–4520 (2013).

96. D'Angelo, V. et al. EZH2 is increased in paediatric T-cell acute lymphoblastic leukemia and is a suitable molecular target in combination treatment approaches. *J. Exp. Clin. Cancer Res.* **34**, 83 (2015).

97. Zhang, H., Gu, H., Li, L., Ren, Y. & Zhang, L. EZH2 mediates ATO-induced apoptosis in acute myeloid leukemia cell lines through the Wnt signaling pathway. *Tumor Biol* **37**, 5919–5923 (2016).

98. Papakonstantinou, N. et al. The histone methyltransferase EZH2 as a novel prosurvival factor in clinically aggressive chronic lymphocytic leukemia. *Oncotarget* **7**, 35946–35959 (2016).

99. Sneeringer, C. J. et al. Coordinated activities of wild-type plus mutant EZH2 drive tumor-associated hypertrimethylation of lysine 27 on histone H3 (H3K27) in human B-cell lymphomas. *Proc. Natl. Acad. Sci. USA* **107**, 20980–20985 (2010).

100. Yap, D. B. et al. Somatic mutations at EZH2 Y641 act dominantly through a mechanism of selectively altered PRC2 catalytic activity, to increase H3K27 trimethylation. *Blood* **117**, 2451–2459 (2011).

101. Souroulas, G. P. et al. An oncogenic EZH2 mutation induces tumors through global redistribution of histone 3 lysine 27 trimethylation. *Nat. Med.* **22**, 632–640 (2016).

102. Oricchio, E. et al. Genetic and epigenetic inactivation of SESTRIN1 controls mTORC1 and response to EZH2 inhibition in follicular lymphoma. *Sci. Transl. Med* **9**, eaak9969 (2017).

103. Jerkovic', I & Cavalli, G Understanding 3D genome organization by multidisciplinary methods. *Nat. Rev. Mol. Cell Biol.* **511–528 (2021)**

104. Szabo, Q., Bantignies, F. & Cavalli, G. Principles of genome folding into topologically associating domains. *Sci. Adv* **5**, eaaw1668 (2019).

105. Cai, Y. et al. H3K27me3-rich genomic regions can function as silencers to repress gene expression via chromatin interactions. *Nat. Commun.* **12**, 719 (2021).

106. Donaldson-Collier, M. C. et al. EZH2 oncogenic mutations drive epigenetic, transcriptional, and structural changes within chromatin domains. *Nat. Genet.* **51**, 517–528 (2019).

107. Mortimer, T. et al. Redistribution of EZH2 promotes malignant phenotypes by rewiring developmental programmes. *EMBO Rep* **20**, e48155 (2019).

108. Ntziachristos, P. et al. Genetic inactivation of the PRC2 complex in T-cell acute lymphoblastic leukemia. *Nat. Med.* **18**, 298–301 (2012).

109. Andrieu, G. P. et al. PRC2 loss of function confers a targetable vulnerability to BET proteins in T-ALL. *Blood* **138**, 1855–1869 (2021).

110. Li, Z. et al. Methylation of EZH2 by PRMT1 regulates its stability and promotes breast cancer metastasis. *Cell Death Differ* **27**, 3226–3242 (2020).

111. Yuan, H. et al. SETD2 restricts prostate cancer metastasis by integrating EZH2 and AMPK signaling pathways. *Cancer Cell* **38**, 350–365.e7 (2020).

112. Zeng, Y. et al. Regulation of EZH2 by SMYD2-mediated lysine methylation is implicated in tumorigenesis. *Cell Rep* **29**, 1482–1498.e4 (2019).

113. Li, H. et al. SUZ12 promotes human epithelial ovarian cancer by suppressing apoptosis via silencing HRK. *Mol. Cancer Res.* **10**, 1462–1472 (2012).

114. Liu, Y. L. et al. Expression and clinicopathological significance of EED, SUZ12 and EZH2 mRNA in colorectal cancer. *J. Cancer Res. Clin. Oncol.* **141**, 661–669 (2015).

115. Wu, Y. et al. SUZ12 is a novel putative oncogene promoting tumorigenesis in head and neck squamous cell carcinoma. *J. Cell. Mol. Med.* **22**, 3582–3594 (2018).

116. Healy, E. et al. PRC2.1 and PRC2.2 synergize to coordinate H3K27 trimethylation. *Mol. Cell* **76**, 437–452.e6 (2019).

117. Youmans, D. T., Gooding, A. R., Dowell, R. D. & Cech, T. R. Competition between PRC2.1 and 2.2 subcomplexes regulates PRC2 chromatin occupancy in human stem cells. *Mol. Cell* **81**, 488–501 (2021).

118. Wassef, M. et al. Impaired PRC2 activity promotes transcriptional instability and favors breast tumorigenesis. *Genes Dev* **29**, 2547–2562 (2015).

119. Erokhin, M. et al. Clinical correlations of Polycomb Repressive Complex 2 in different tumor types. *Cancers (Basel)* **13**, 3155 (2021).

120. Haupt, Y., Alexander, W. S., Barri, G., Peter Klinken, S. & Adams, J. M. Novel zinc finger gene implicated as myc collaborator by retrovirally accelerated lymphomagenesis in Eμ-myc transgenic mice. *Cell* **65**, 753–763 (1991).

121. van Lohuizen, M. et al. Identification of cooperating oncogenes in Eμ-myc transgenic mice by provirus tagging. *Cell* **65**, 737–752 (1991).

122. Haupt, Y., Bath, M. L., Harris, A. W. & Adams, J. M. BMI-1 transgene induces lymphomas and collaborates with myc in tumorigenesis. *Oncogene* **8**, 3161–3164 (1993).

123. Gunster, M. J. et al. Identification and characterization of interactions between the vertebrate polycomb-group protein BMI1 and human homologs of polyhomeotic. *Mol. Cell. Biol.* **17**, 2326–2335 (1997).

124. Jacobs, J. L., Kieboom, K., Marino, S., DePinho, R. A. & Van Lohuizen, M. The oncogene and Polycomb-group gene bmi-1 regulates cell proliferation and senescence through the ink4a locus. *Nature* **397**, 164–168 (1999).

125. Li, W., Li, Y., Tan, Y., Ma, K. & Cui, J. Bmi-1 is critical for the proliferation and invasiveness of gastric carcinoma cells. *J. Gastroenterol. Hepatol.* **25**, 568–575 (2010).

126. Wang, M. C. et al. Polycomb complex protein BMI-1 promotes invasion and metastasis of pancreatic cancer stem cells by activating PI3K/AKT signaling, an ex vivo, in vitro, and in vivo study. *Oncotarget* **7**, 9586–9599 (2016).

127. Althobiti, M. et al. The prognostic significance of BMI1 expression in invasive breast cancer is dependent on its molecular subtypes. *Breast Cancer Res. Treat* **182**, 581–589 (2020).

128. Zhao, Q. et al. Role of BMI1 in epithelial ovarian cancer: Investigated via the CRISPR/Cas9 system and RNA sequencing. *J. Ovarian Res.* **11**, 31 (2018).

129. Wang, M. C. et al. BMI-1, a promising therapeutic target for human cancer. *Oncol. Lett.* **10**, 583–588 (2015).

130. Tetsu, O. et al. Mel-18 negatively regulates cell cycle progression upon B cell antigen receptor stimulation through a cascade leading to c-myc/cdc25. *Immunity* **9**, 439–448 (1998).

131. Won, H. Y. et al. Loss of Mel-18 enhances breast cancer stem cell activity and tumorigenicity through activating Notch signaling mediated by the Wnt/TCF pathway. *FASEB J* **26**, 5002–5013 (2012).

132. Lee, J. & Kong, G. Mel-18, a tumor suppressor for aggressive breast cancer. *Oncotarget* **6**, 15710–15711 (2015).

133. Guo, W. J. et al. Mel-18 acts as a tumor suppressor by repressing BMI-1 expression and down-regulating Akt activity in breast cancer cells. *Cancer Res* **67**, 5083–5089 (2007).

134. Zhang, X. W. et al. BMI1 and Mel-18 oppositely regulate carcinogenesis and progression of gastric cancer. *Mol. Cancer* **9**, 40 (2010).

135. Guo, W.-J., Datta, S., Band, V. & Dimri, G. P. Mel-18, a polycomb group protein, regulates cell proliferation and senescence via transcriptional repression of Bmi-1 and c-Myc oncogenes. *Mol. Biol. Cell* **18**, 536–546 (2007).

136. Martinez, A.-M. et al. Polyhomeotic has a tumor suppressor activity mediated by repression of Notch signaling. *Nat. Genet.* **41**, 1076–1082 (2009).

137. Classen, A. K., Bunker, B. D., Harvey, K. F., Vaccari, T. & Bilder, D. A tumor suppressor activity of Drosophila Polycomb genes mediated by JAK-STAT signaling. *Nat. Genet.* **41**, 1150–1155 (2009).

138. van den Boom, V. et al. Non-canonical PRC1.1 targets active genes independent of H3K27me3 and is essential for leukemogenesis. *Cell Rep* **14**, 332–346 (2016).

139. Chan, H. L. et al. Polycomb complexes associate with enhancers and promote oncogenic transcriptional programs in cancer through multiple mechanisms. *Nat. Commun.* **9**, 3377 (2018).

140. Balasubramani, A. et al. Cancer-associated ASXL1 mutations may act as gain-of-function mutations of the ASXL1-BAP1 complex. *Nat. Commun.* **6**, 7307 (2015).

141. Kutscher, L. M. et al. Functional loss of a noncanonical BCOR-PRC1.1 complex accelerates SHH-driven medulloblastoma formation. *Genes Dev* **34**, 1161–1176 (2020).

142. Waszak, S. M. et al. Germline elongator mutations in Sonic Hedgehog medulloblastoma. *Nature* **580**, 396–401 (2020).

143. Mathsyaraja, H. et al. Loss of MGA mediated Polycomb repression promotes tumor progression and invasiveness. *Elife* **10**, e64212 (2021).

144. Harbour, J. W. et al. Frequent mutation of BAP1 in metastasizing uveal melanomas. *Science* **330**, 1410–1413 (2010).

145. Wang, S. S. et al. Bap1 is essential for kidney function and cooperates with Vhl in renal tumorigenesis. *Proc. Natl. Acad. Sci. USA* **111**, 16538–16543 (2014).
146. Lafave, L. M. et al. Loss of BAP1 function leads to EZH2-dependent transformation. *Nat. Med.* **21**, 1344–1349 (2015).

147. Kuznetsov, J. N. et al. BAP1 regulates epigenetic switch from pluripotency to differentiation in developmental lineages giving rise to BAP1-mutant cancers. *Sci. Adv* **5**, eaax1738 (2019).

148. Campagne, A. et al. BAP1 complex promotes transcription by opposing PRC1-mediated H2A ubiquitylation. *Nat. Commun.* **10**, 348 (2019).

149. Conway, E. et al. BAP1 enhances Polycomb repression by counteracting widespread H2AK119ub1 deposition and chromatin condensation. *Mol. Cell* **81**, 1–16 (2021).

150. Xia, Y. K. et al. Tumor-derived neomorphic mutations in ASXL1 impairs the BAP1-ASXL1-FOXK1/K2 transcription network. *Protein Cell* **12**, 557–577 (2021).

151. Pasini, D. & Di Croce, L. Emerging roles for Polycomb proteins in cancer. *Curr. Opin. Genet.* **36**, 50–58 (2016).

152. Zhou, M., Sun, J., Liu, Y. & Ma, J. Suppressor of Zeste 12 homolog RNA interference inhibits retinoblastoma cell invasion. *Oncol. Lett.* **8**, 1933–1936 (2014).

153. Ciarapica, R. et al. Pharmacological inhibition of EZH2 as a promising differentiation therapy in embryonal RMS. *BMC Cancer* **14**, 139 (2014).

154. Ramakrishnan, S. et al. Inhibition of EZH2 induces NK cell-mediated differentiation and death in muscle-invasive bladder cancer. *Cell Death Differ* **26**, 2100–2114 (2019).

155. Liu, M. et al. The polycomb group protein PCGF6 mediates germline gene silencing by recruiting histone-modifying proteins to target gene promoters. *J. Biol. Chem.* **295**, 9712–9724 (2020).

156. Cohen, I. et al. PRC1 fine-tunes gene repression and activation to safeguard skin development and stem cell specification. *Cell Stem Cell* **22**, 726–739.e7 (2018).

157. Scelfo, A. et al. Functional landscape of PCGF proteins reveals both RING1A/B-dependent-and RING1A/B-independent-specific activities. *Mol. Cell* **74**, 1037–1052.e7 (2019).

158. Wang, H. et al. Estrogen receptor α-coupled Bmi1 regulation pathway in breast cancer and its clinical implications. *BMC Cancer* **14**, 122 (2014).

159. Bhan, A. et al. Histone methyltransferase EZH2 is transcriptionally induced by estradiol as well as estrogenic endocrine disruptors bisphenol-a and diethyl-stilbestrol. *J. Mol. Biol.* **426**, 3426–3441 (2014).

160. Zhu, S. et al. BMI1 is directly regulated by androgen receptor to promote castration-resistance in prostate cancer. *Oncogene* **39**, 17–29 (2020).

161. Bansal, N. et al. BMI-1 targeting interferes with patient-derived tumor-initiating cell survival and tumor growth in prostate cancer. *Clin. Cancer Res.* **22**, 6176–6191 (2016).

162. Li, Y. et al. O-GlcNAcylation modulates Bmi-1 protein stability and potential oncogenic function in prostate cancer. *Oncogene* **36**, 6293–6305 (2017).

163. Zhu, S. et al. BMI1 regulates androgen receptor in prostate cancer independently of the polycomb repressive complex 1. *Nat. Commun.* **9**, 500 (2018).

164. Xu, K. et al. EZH2 oncogenic activity in castration-resistant prostate cancer cells is Polycomb-independent. *Science* **(80–)** **338**, 1465–1469 (2012).

165. Kim, J. et al. Polycomb- and methylation-independent roles of EZH2 as a transcription activator. *Cell Rep* **25**, 2808–2820.e4 (2018).

166. Liu, Q. et al. Polycomb group proteins EZH2 and EED directly regulate androgen receptor in advanced prostate cancer. *Int. J. Cancer* **145**, 415–426 (2019).

167. Shin, Y. J. & Kim, J. H. The role of EZH2 in the regulation of the activity of matrix metalloproteinases in prostate cancer cells. *PLoS One* **7**, e30393 (2012).

168. Wee, Z. N. et al. EZH2-mediated inactivation of IFN-γ-JAK-STAT1 signaling is an effective therapeutic target in MYC-driven prostate cancer. *Cell Rep* **8**, 204–216 (2014).

169. Shi, B. et al. Integration of estrogen and Wnt signaling circuits by the polycomb group protein EZH2 in breast cancer cells. *Mol. Cell. Biol.* **27**, 5105–5119 (2007).

170. Zhang, Y. et al. Estrogen induces dynamic ERα and RING1B recruitment to control gene and enhancer activities in luminal breast cancer. *Sci. Adv* **6**, eaaz7249 (2020).

171. Pascual, G., Domínguez, D. & Benitah, S. A. The contributions of cancer cell metabolism to metastasis. *DMM Dis. Model. Mech* **11**, dmm032920 (2018).

172. Dann, S. G. et al. Reciprocal regulation of amino acid import and epigenetic state through Lat1 and EZH2. *EMBO J* **34**, 1773–1785 (2015).

173. Liu, T. P., Lo, H. L., Wei, L. S., Hsiao, H. H. Y. & Yang, P. M. S-adenosyl-L-methionine-competitive inhibitors of the histone methyltransferase EZH2 induce autophagy and enhance drug sensitivity in cancer cells. *Anticancer. Drugs* **26**, 139–147 (2015).

174. Gu, Z. et al. Loss of EZH2 reprograms BCAA metabolism to drive leukemic transformation. *Cancer Discov* **9**, 1228–1247 (2019).

175. Sivanand, S. & Vander Heiden, M. G. Emerging roles for branched-chain amino acid metabolism in cancer. *Cancer Cell* **37**, 147–156 (2020).

176. Ananieva, E. A. & Wilkinson, A. C. Branched-chain amino acid metabolism in cancer. *Curr. Opin. Clin. Nutr. Metab. Care* **21**, 64–70 (2018).

177. Tian, T., Li, X. & Zhang, J. mTOR signaling in cancer and mTOR inhibitors in solid tumor targeting therapy. *Int. J. Mol. Sci.* **20**, 755 (2019).

178. Pang, B. et al. EZH2 promotes metabolic reprogramming in glioblastomas through epigenetic repression of EAF2-HIF1α signaling. *Oncotarget* **7**, 45134–45143 (2016).

179. Zhang, B. et al. Regulation of branched-chain amino acid metabolism by hypoxia-inducible factor in glioblastoma. *Cell. Mol. Life Sci.* **78**, 195–206 (2021).

180. Liao, K. et al. A feedback circuitry between polycomb signaling and fructose-1, 6-bisphosphatase enables hepatic and renal tumorigenesis. *Cancer Res* **80**, 675–688 (2020).

181. McDonald, O. G. et al. Epigenomic reprogramming during pancreatic cancer progression links anabolic glucose metabolism to distant metastasis. *Nat. Genet.* **49**, 367–376 (2017).

182. Faubert, B., Solmonson, A. & DeBerardinis, R. J. Metabolic reprogramming and cancer progression. *Science* **368**, eaaw5473 (2020).

183. Brookes, E. et al. Polycomb associates genome-wide with a specific RNA polymerase II variant, and regulates metabolic genes in ESCs. *Cell Stem Cell* **10**, 157–170 (2012).

184. Zhang, T., Gong, Y., Meng, H., Li, C. & Xue, L. Symphony of epigenetic and metabolic regulation - Interaction between the histone methyltransferase EZH2 and metabolism of tumor. *Clin. Epigenetics* **12**, 72 (2020).

185. Dranoff, G. Cytokines in cancer pathogenesis and cancer therapy. *Nat. Rev. Cancer* **4**, 11–22 (2004).

186. Su, W. et al. The Polycomb repressor complex 1 drives double-negative prostate cancer metastasis by coordinating stemness and immune suppression. *Cancer Cell* **36**, 139–155.e10 (2019).

187. Duan, Q. et al. High glucose promotes pancreatic cancer cells to escape from immune surveillance via AMPK-Bmi1-GATA2-MICA/B pathway. *J. Exp. Clin. Cancer Res.* **38**, 192 (2019).

188. Burr, M. L. et al. An evolutionarily conserved function of polycomb silences the MHC class I antigen presentation pathway and enables immune evasion in cancer. *Cancer Cell* **36**, 385–401.e8 (2019).

189. Kim, H. J., Cantor, H. & Cosmopoulos, K. Overcoming immune checkpoint blockade resistance via EZH2 inhibition. *Trends Immunol* **41**, 948–963 (2020).

190. Wang, D. et al. Targeting EZH2 reprograms intratumoral regulatory T cells to enhance cancer immunity. *Cell Rep* **23**, 3262–3274 (2018).

191. Goswami, S. et al. Modulation of EZH2 expression in T cells improves efficacy of anti-CTLA-4 therapy. *J. Clin. Invest.* **128**, 3813–3818 (2018).

192. Zingg, D. et al. The histone methyltransferase Ezh2 controls mechanisms of adaptive resistance to tumor immunotherapy. *Cell Rep* **20**, 854–867 (2017).

193. Jia, L., Zhang, W. & Wang, C.-Y. BMI1 inhibition eliminates residual cancer stem cells after PD1 blockade and activates antitumor immunity to prevent metastasis and relapse. *Cell Stem Cell* **27**, 238–253.e6 (2020).

194. Bannister, A. J. & Kouzarides, T. Regulation of chromatin by histone modifications. *Cell Res* **21**, 381–395 (2011).

195. Ren, C. et al. Small-molecule modulators of methyl-lysine binding for the CBX7 chromodomain. *Chem. Biol.* **22**, 161–168 (2015).

196. Nacev, B. A. et al. The expanding landscape of 'oncohistone' mutations in human cancers. *Nature* **567**, 473–478 (2019).

197. Schwartzentruber, J. et al. Driver mutations in histone H3.3 and chromatin remodeling genes in paediatric glioblastoma. *Nature* **482**, 226–231 (2012).

198. Wu, G. et al. Somatic histone H3 alterations in paediatric diffuse intrinsic pontine gliomas and non-brainstem glioblastomas. *Nat. Genet.* **44**, 251–253 (2012).

199. Castel, D. et al. Histone H3F3A and HIST1H3B K27M mutations define two subgroups of diffuse intrinsic pontine gliomas with different prognosis and phenotypes. *Acta Neuropathol* **130**, 815–827 (2015).

200. Nikbakht, H. et al. Spatial and temporal homogeneity of driver mutations in diffuse intrinsic pontine glioma. *Nat. Commun.* **7**, 11185 (2016).

201. Lewis, P. W. et al. Inhibition of PRC2 activity by a gain-of-function H3 mutation found in pediatric glioblastoma. *Science* **340**, 857–861 (2013).

202. Chan, K. M. et al. The histone H3.3K27M mutation in pediatric glioma reprograms H3K27 methylation and gene expression. *Genes Dev* **27**, 985–990 (2013).

203. Harutyunyan, A. S. et al. H3K27M in gliomas causes a one-step decrease in H3K27 methylation and reduced spreading within the constraints of H3K36 methylation. *Cell Rep* **33**, 108390 (2020).

204. Herz, H. et al. Histone H3 lysine-to-methionine mutants as a paradigm to study chromatin signaling. *Science* **345**, 1065–1070 (2014).

205. Justin, N. et al. Structural basis of oncogenic histone H3K27M inhibition of human polycomb repressive complex 2. *Nat. Commun.* **7**, 1–11 (2016).

206. Fang, D. et al. H3.3K27M mutant proteins reprogram epigenome by sequencing the PRC2 complex to poised enhancers. *Elife* **7**, e36696 (2018).

207. Bender, S. et al. Reduced H3K27me3 and DNA hypomethylation are major drivers of gene expression in K27M mutant pediatric high-grade gliomas. *Cancer Cell* **24**, 660–672 (2013).

208. Piunti, A. et al. Therapeutic targeting of polycomb and BET bromodomain proteins in diffuse intrinsic pontine gliomas. *Nat. Med.* **23**, 493–500 (2017).

209. Dewaele, B. et al. Identification of a novel, recurrent MBTD1-CXorf67 fusion in low-grade endometrial stromal sarcoma. *Int. J. cancer* **134**, 1112–1122 (2014).
210. Pajtler, K. W. et al. Molecular heterogeneity and CXorf67 alterations in posterior fossa group A (PFA) ependymomas. *Acta Neuropathol* **136**, 211–226 (2018).

211. Jain, S. U. et al. H3K27M and EZHIP impede H3K27-methylation spreading by inhibiting allosterically stimulated PRC2. *Mol. Cell* **80**, 726–735.e7 (2020).

212. Khazaei, S. et al. H3.3 G34W promotes growth and impedes differentiation of osteoblast-like mesenchymal progenitors in giant cell tumor of bone. *Cancer Discov* **10**, 1968–1987 (2020).

213. Behjati, S. et al. Distinct H3F3A and H3F3B driver mutations define chondroblastoma and giant cell tumor of bone. *Nat. Genet* **45**, 1479–1482 (2013).

214. Lu, C. et al. Histone H3K36 mutations promote sarcomagenesis through altered histone methylation landscape. *Science* **352**, 844–849 (2016).

215. Abe, S., Nagatomo, H., Sasaki, H. & Ishiuchi, T. A histone H3.3K36M mutation in mice causes an imbalance of histone modifications and defects in chondrocyte differentiation. *Epigenetics* **16**, 1123–1134 (2020).

216. Fang, D. et al. The histone H3.3K36M mutation reprograms the epigenome of chondroblastomas. *Science* **352**, 1344–1348 (2016).

217. Papillon-cavanagh, S. et al. Impaired H3K36 methylation defines a subset of head and neck squamous cell carcinomas. *Nat. Genet* **49**, 180–185 (2017).

218. Rueff, J. & Rodrigues, A. S. Cancer drug resistance: A brief overview from a genetic viewpoint. *Methods Mol. Biol* **1395**, 1–18 (2016).

219. Bell, C. C. et al. Targeting enhancer switching overcomes non-genetic drug resistance in acute myeloid leukaemia. *Nat. Commun.* **10**, 2723 (2019).

220. Bell, C. C. & Gilan, O. Principles and mechanisms of non-genetic resistance in cancer. *Br. J. Cancer* **122**, 465–472 (2020).

221. Hu, S. et al. Overexpression of EZH2 contributes to acquired cisplatin resistance in ovarian cancer cells in vitro and in vivo. *Cancer Biol. Ther.* **10**, 788–795 (2010).

222. Hirukawa, A. et al. Reduction of global H3K27me3 enhances HER2/ErbB2 targeted therapy. *Cell Rep* **29**, 249–257.e8 (2019).

223. Singh, R. et al. Epigenetic remodeling through downregulation of polycomb repressive complex 2 mediates chemotherapy resistance in testicular germ cell tumors. *Cancers (Basel)* **11**, 796 (2019).

224. Kikuchi, J. et al. Phosphorylation-mediated EZH2 inactivation promotes drug resistance in multiple myeloma. *J. Clin. Invest.* **125**, 4375–4390 (2015).

225. Yu, T. et al. Polycomb-like protein 3 induces proliferation and drug resistance in multiple myeloma and is regulated by miRNA-15a. *Mol. Cancer Res.* **18**, 1063–1073 (2020).

226. Loubiere, V., Papadopoulos, G. L., Szabo, Q., Martinez, A. M. & Cavalli, G. Widespread activation of developmental gene expression characterized by PRC1-dependent chromatin looping. *Sci. Adv* **6**, eaax4001 (2020).

227. Campbell, P. J. et al. Pan-cancer analysis of whole genomes. *Nature* **578**, 82–93 (2020).

228. Makohon-Moore, A. P. et al. Limited heterogeneity of known driver gene mutations among the metastases of individual patients with pancreatic cancer. *Nat. Genet.* **49**, 358–366 (2017).

229. MacK, S. C. et al. Epigenomic alterations define lethal CIMP-positive ependymomas of infancy. *Nature* **506**, 445–450 (2014).

230. Feinberg, A. P. The key role of epigenetics in human disease prevention and mitigation. *N. Engl. J. Med.* **378**, 1323–1334 (2018).

231. Cavalli, G. EZH2 Goes Solo. *Science* **338**, 1430–1431 (2012).

232. Veneti, Z., Gkouskou, K. K. & Eliopoulos, A. G. Polycomb repressor complex 2 in genomic instability and cancer. *Int. J. Mol. Sci.* **18**, 1–16 (2017).

233. Wan, J. et al. PCAF-primed EZH2 acetylation regulates its stability and promotes lung adenocarcinoma progression. *Nucleic Acids Res* **43**, 3591–3604 (2015).

234. Li, B. et al. MELK mediates the stability of EZH2 through site-specific phosphorylation in extranodal natural killer/T-cell lymphoma. *Blood* **134**, 2046–2058 (2019).

235. Li, Z. et al. Macrophages-stimulated PRMT1-mediated EZH2 methylation promotes breast cancer metastasis. *Biochem. Biophys. Res. Commun.* **533**, 679–684 (2020).

236. Lo, P. W. et al. O-GlcNAcylation regulates the stability and enzymatic activity of the histone methyltransferase EZH2. *Proc. Natl. Acad. Sci. USA* **115**, 7302–7307 (2018).

237. Chu, C. S. et al. O-GlcNAcylation regulates EZH2 protein stability and function. *Proc. Natl. Acad. Sci. USA* **111**, 1355–1360 (2014).

238. Cha, T.-L. et al. Akt-mediated phosphorylation of EZH2 suppresses methylation of lysine 27 in histone H3. *Science* **310**, 306–310 (2005).

239. Kim, E. et al. Phosphorylation of EZH2 activates STAT3 signaling via STAT3 methylation and promotes tumorigenicity of glioblastoma stem-like cells. *Cancer Cell* **23**, 839–852 (2013).

240. Yan, J. et al. EZH2 phosphorylation by JAK3 mediates a switch to noncanonical function in natural killer/T-cell lymphoma. *Blood* **128**, 948–958 (2016).

241. Jin, X. et al. CDK5/FBW7-dependent ubiquitination and degradation of EZH2 inhibits pancreatic cancer cell migration and invasion. *J. Biol. Chem.* **292**, 6269–6280 (2017).

242. Wan, L. et al. Phosphorylation of EZH2 by AMPK suppresses PRC2 methyltransferase activity and oncogenic function. *Mol. Cell* **69**, 279–291.e5 (2018).

243. Wu, S. C. & Zhang, Y. Cyclin-dependent kinase 1 (CDK1)-mediated phosphorylation of enhancer of zeste 2 (Ezh2) regulates its stability. *J. Biol. Chem.* **286**, 28511–28519 (2011).

244. Kaneko, S. et al. Phosphorylation of the PRC2 component Ezh2 is cell cycle-regulated and up-regulates its binding to ncRNA. *Genes Dev* **24**, 2615–2620 (2010).

245. Wei, Y. et al. CDK1-dependent phosphorylation of EZH2 suppresses methylation of H3K27 and promotes osteogenic differentiation of human mesenchymal stem cells. *Nat. Cell Biol.* **13**, 87–94 (2011).

246. Ko, H. W. et al. GSK3β inactivation promotes the oncogenic functions of EZH2 and enhances methylation of H3K27 in human breast cancers. *Oncotarget* **7**, 57131–57144 (2016).

247. Anwar, T et al. P38-mediated phosphorylation at T367 induces EZH2 cytoplasmic localization to promote breast cancer metastasis. *Nat. Commun.* **9**, (2018).

248. Nie, L. et al. CDK2-mediated site-specific phosphorylation of EZH2 drives and maintains triple-negative breast cancer. *Nat. Commun.* **10**, 1–15 (2019).

249. Yang, C. C. et al. Phosphorylation of EZH2 at T416 by CDK2 contributes to the malignancy of triple negative breast cancers. *Am. J. Transl. Res.* **7**, 1009–1020 (2015).

250. Sahasrabuddhe, A. A. et al. Oncogenic Y641 mutations in EZH2 prevent Jak2/β-TrCP-mediated degradation. *Oncogene* **34**, 445–454 (2015).

251. Yu, Y. L. et al. Smurf2-mediated degradation of EZH2 enhances neuron differentiation and improves functional recovery after ischaemic stroke. *EMBO Mol. Med.* **5**, 531–547 (2013).

252. Xue, C. et al. The down-regulation of SUZ12 accelerates the migration and invasion of liver cancer cells via activating ERK1/2 pathway. *J. Cancer* **10**, 1375–1384 (2019).

253. Suh, J. L. et al. Discovery of selective activators of PRC2 mutant EED-I363M. *Sci. Rep.* **9**, 1–10 (2019).

254. Yu, H. et al. PRC2/EED-EZH2 complex is up-regulated in breast cancer lymph node metastasis compared to primary tumor and correlates with tumor proliferation in situ. *PLoS One* **7**, 1–8 (2012).

255. Cao, J., Li, H., Liu, G., Han, S. & Xu, P. Knockdown of JARID2 inhibits the proliferation and invasion of ovarian cancer through the PI3K/Akt signaling pathway. *Mol. Med. Rep.* **16**, 3600–3605 (2017).

256. Lei, X. et al. JARID2 promotes invasion and metastasis of hepatocellular carcinoma by facilitating epithelial-mesenchymal transition through PTEN/AKT signaling. *Oncotarget* **7**, 40266–40284 (2016).

257. Walters, Z. S. et al. JARID2 is a direct target of the PAX3-FOXO1 fusion protein and inhibits myogenic differentiation of rhabdomyosarcoma cells. *Oncogene* **33**, 1148–1157 (2014).

258. Celik, H. et al. JARID2 Functions as a tumor suppressor in myeloid neoplasms by repressing self-renewal in hematopoietic progenitor cells. *Cancer Cell* **34**, 741–756.e8 (2018).

259. Zhang, Q., Wang, W. & Gao, Q. β-TrCP-mediated AEBP2 ubiquitination and destruction controls cisplatin resistance in ovarian cancer. *Biochem. Biophys. Res. Commun.* **523**, 274–279 (2020).

260. Liefke, R., Karwacki-Neisius, V. & Shi, Y. EPOP interacts with elongin BC and USP7 to modulate the chromatin landscape. *Mol. Cell* **64**, 659–672 (2016).

261. Hofvander, J. et al. PHF1 fusions cause distinct gene expression and chromatin accessibility profiles in ossifying fibromyxoid tumors and mesenchymal cells. *Mod. Pathol. an Off. J. United States Can. Acad. Pathol. Inc* **33**, 1331–1340 (2020).

262. Gebre-Medhin, S. et al. Recurrent rearrangement of the PHF1 gene in ossifying fibromyxoid tumors. *Am. J. Pathol.* **181**, 1069–1077 (2006).

263. Micci, F., Panagopoulos, I., Bjerkhagen, B. & Heim, S. Consistent rearrangement of chromosomal band 6p21 with generation of fusion genes JAZF1/PHF1 and EPC1/PHF1 in endometrial stromal sarcoma. *Cancer Res* **66**, 107–112 (2006).

264. Wang, F. et al. Polycomb-like 2 regulates PRC2 components to affect proliferation in glioma cells. *J. Neurooncol.* **148**, 259–271 (2020).

265. Liang, Y. et al. PCL2 regulates p53 stability and functions as a tumor suppressor in breast cancer. *Sci. Bull.* **63**, 629–639 (2018).

266. Jain, P., Ballare, C., Blanco, E. & Vizan, P. & Di Croce, L. PHF19 mediated regulation of proliferation and invasiveness in prostate cancer cells. *Elife* **9**, 1–26 (2020).

267. Ghislin, S., Deshayes, F., Middendorp, S., Boggetto, N. & Alcaide-Loridan, C. PHF19 and Akt control the switch between proliferative and invasive states in melanoma. *Cell Cycle* **11**, 1634–1645 (2012).

268. Ren, Z. et al. PHF19 promotes multiple myeloma tumorigenicity through PRC2 activation and broad H3K27me3 domain formation. *Blood* **134**, 1176–1189 (2019).

269. Deng, Q. et al. PHF19 promotes the proliferation, migration, and chemosensitivity of glioblastoma to doxorubicin through modulation of the SIAH1/β-catenin axis. *Cell Death Dis* **9**, 1049 (2018).

270. Xu, H. et al. MicroRNA-195-5p acts as an anti-oncogene by targeting PHF19 in hepatocellular carcinoma. *Oncol. Rep.* **34**, 175–182 (2015).
271. Hübner, J. M. et al. EZHIP/CXorf67 mimics K27M mutated oncohistones and functions as an intrinsic inhibitor of PRC2 function in aggressive posterior fossa ependymoma. *Neuro. Oncol.* **21**, 878–889 (2019).

272. Rai, K. et al. Dual roles of RNF2 in melanoma progression. *Cancer Discov* **5**, 1314–1327 (2015).

273. Bosch, A. et al. The polycomb group protein RING1B is overexpressed in ductal breast carcinoma and is required to sustain FAK steady state levels in breast cancer epithelial cells. *Oncotarget* **5**, 2065–2076 (2014).

274. Sánchez-Beato, M. et al. Variability in the expression of polycomb proteins in different normal and tumoral tissues. A pilot study using tissue microarrays. *Mod. Pathol.* **19**, 684–694 (2006).

275. Gong, Y. et al. NSPc1 is a cell growth regulator that acts as a transcriptional repressor of p21Waf1/Cip1 via the RARE element. *Nucleic Acids Res* **34**, 6158–6169 (2006).

276. Hu, P.-S. et al. NSPc1 promotes cancer stem cell self-renewal by repressing the synthesis of all-trans retinoic acid via targeting RDH16 in malignant glioma. *Oncogene* **36**, 4706–4718 (2017).

277. Ji, G. et al. PCGF1 promotes epigenetic activation of stemness markers and colorectal cancer stem cell enrichment. *Cell Death Dis* **12**, 633 (2021).

278. Lee, J. Y. et al. Loss of the polycomb protein Mel-18 enhances the epithelial-mesenchymal transition by ZEB1 and ZEB2 expression through the down-regulation of miR-205 in breast cancer. *Oncogene* **33**, 1325–1335 (2014).

279. Park, J. H. et al. Loss of Mel-18 induces tumor angiogenesis through enhancing the activity and expression of HIF-1α mediated by the PTEN/PI3K/Akt pathway. *Oncogene* **30**, 4578–4589 (2011).

280. Hu, Y. et al. PCGF3 promotes the proliferation and migration of non-small cell lung cancer cells via the PI3K/AKT signaling pathway. *Exp. Cell Res.* **400**, 112496 (2021).

281. Srinivasan, M. et al. Downregulation of Bmi1 in breast cancer stem cells suppresses tumor growth and proliferation. *Oncotarget* **8**, 38731–38742 (2017).

282. Takamatsu-Ichihara, E. & Kitabayashi, I. The roles of Polycomb group proteins in hematopoietic stem cells and hematological malignancies. *Int. J. Hematol.* **103**, 634–642 (2016).

283. Guo, B. H. et al. Bmi-1 promotes invasion and metastasis, and its elevated expression is correlated with an advanced stage of breast cancer. *Mol. Cancer* **10**, 10 (2011).

284. Vonlanthen, S. et al. The bmi-1 oncoprotein is differentially expressed in non-small cell lung cancer and correlates with INK4A-ARF locus expression. *Br. J. Cancer* **84**, 1372–1376 (2001).

285. Rouhigharabaei, L. et al. BMI1, the polycomb-group gene, is recurrently targeted by genomic rearrangements in progressive B-cell leukemia/lymphoma. *Genes, Chromosom. Cancer* **52**, 928–944 (2013).

286. Lee, J.-H. et al. Integrative analysis of mutational and transcriptional profiles reveals driver mutations of metastatic breast cancers. *Cell Discov* **2**, 16025 (2016).

287. Yang, Y. F., Pan, Y. H., Tian, Q. H., Wu, D. C. & Su, S. G. CBX1 indicates poor outcomes and exerts oncogenic activity in hepatocellular carcinoma. *Transl. Oncol.* **11**, 1110–1118 (2018).

288. Lee, Y.-H. et al. HP1β is a biomarker for breast cancer prognosis and PARP inhibitor therapy. *PLoS One* **10**, e0121207 (2015).

289. Zheng, S. et al. Overexpression of CBX2 in breast cancer promotes tumor progression through the PI3K/AKT signaling pathway. *Am. J. Transl. Res.* **11**, 1668–1682 (2019).

290. Zeng, M., Li, B., Yang, L. & Guan, Q. CBX2 depletion inhibits the proliferation, invasion and migration of gastric cancer cells by inactivating the YAP/β-catenin pathway. *Mol. Med. Rep.* **23**, 137 (2021).

291. Wheeler, L. J. et al. CBX2 identified as driver of anoikis escape and dissemination in high grade serous ovarian cancer. *Oncogenesis* **7**, 92 (2018).

292. Zhang, H., Chen, W., Fu, X., Su, X. & Yang, A. CBX3 promotes tumor proliferation by regulating G1/S phase via p21 downregulation and associates with poor prognosis in tongue squamous cell carcinoma. *Gene* **654**, 49–56 (2018).

293. Fan, Y., Li, H., Liang, X. & Xiang, Z. CBX3 promotes colon cancer cell proliferation by CDK6 kinase-independent function during cell cycle. *Oncotarget* **8**, 19934–19946 (2017).

294. Chang, S.-C. et al. CBX3/heterochromatin protein 1 gamma is significantly upregulated in patients with non-small cell lung cancer. *Asia. Pac. J. Clin. Oncol.* **14**, e283–e288 (2018).

295. Liang, Y. K., Lin, H. Y., Chen, C. F. & Zeng, D. Prognostic values of distinct CBX family members in breast cancer. *Oncotarget* **8**, 92375–92387 (2017).

296. Chen, L.-Y. et al. CBX3 promotes proliferation and regulates glycolysis via suppressing FBP1 in pancreatic cancer. *Biochem. Biophys. Res. Commun.* **500**, 691–697 (2018).

297. Jiao, H. K. et al. Prognostic significance of Cbx4 expression and its beneficial effect for transarterial chemoembolization in hepatocellular carcinoma. *Cell Death Dis* **6**, e1689 (2015).

298. Li, J. et al. Cbx4 governs HIF-1α to potentiate angiogenesis of hepatocellular carcinoma by its SUMO E3 ligase activity. *Cancer Cell* **25**, 118–131 (2014).

299. Jiang, N. et al. CBX4 transcriptionally suppresses KLF6 via interaction with HDAC1 to exert oncogenic activities in clear cell renal cell carcinoma. *EBioMedicine* **53**, 102692 (2020).

300. Wang, X. et al. CBX4 suppresses metastasis via recruitment of HDAC3 to the Runx2 promoter in colorectal carcinoma. *Cancer Res* **76**, 7277–7289 (2016).

301. Wang, J., He, H., Jiang, Q., Wang, Y. & Jia, S. CBX6 promotes HCC metastasis via transcription factors Snail/Zeb1-mediated EMT mechanism. *Onco. Targets. Ther.* **13**, 12489–12500 (2020).

302. Zheng, H. et al. CBX6 overexpression contributes to tumor progression and is predictive of a poor prognosis in hepatocellular carcinoma. *Oncotarget* **8**, 18872–18884 (2017).

303. Deng, H. et al. CBX6 is negatively regulated by EZH2 and plays a potential tumor suppressor role in breast cancer. *Sci. Rep.* **9**, 197 (2019).

304. Ma, R. G., Zhang, Y., Sun, T. T. & Cheng, B. Epigenetic regulation by polycomb group complexes: Focus on roles of CBX proteins. *J. Zhejiang Univ. Sci. B* **15**, 412–428 (2014).

305. Scott, C. L. et al. Role of the chromobox protein CBX7 in lymphomagenesis. *Proc. Natl. Acad. Sci. USA* **104**, 5389–5394 (2007).

306. Bernard, D. et al. CBX7 controls the growth of normal and tumor-derived prostate cells by repressing the Ink4a/Arf locus. *Oncogene* **24**, 5543–5551 (2005).

307. Pallante, P. et al. Loss of the CBX7 gene expression correlates with a highly malignant phenotype in thyroid cancer. *Cancer Res* **68**, 6770–6778 (2008).

308. Forzati, F. et al. CBX7 is a tumor suppressor in mice and humans. *J. Clin. Invest.* **122**, 612–623 (2012).

309. Hannafon, B. N., Sebastiani, P., de las Morenas, A., Lu, J. & Rosenberg, C. L. Expression of microRNA and their gene targets are dysregulated in preinvasive breast cancer. *Breast Cancer Res* **13**, R24 (2011).

310. Lee, S. H., Um, S. J. & Kim, E. J. CBX8 suppresses Sirtolin-induced premature senescence in human breast cancer cells via cooperation with SIRT1. *Cancer Lett* **335**, 397–403 (2013).

311. Li, G. et al. Altered expression of polycomb group genes in glioblastoma multiforme. *PLoS One* **8**, e08970 (2013).

312. Zhang, C. Z. et al. CBX8 exhibits oncogenic activity via AKT/β-Catenin activation in hepatocellular carcinoma. *Cancer Res* **78**, 51–63 (2018).

313. Chung, C.-Y. et al. CBX8 acts non-canonically with Wdr5 to promote mammary tumorigenesis. *Cell Rep* **16**, 472–486 (2016).

314. Iwata, S. et al. Polycomb group molecule PHC3 regulates polycomb complex composition and prognosis of osteosarcoma. *Cancer Sci* **101**, 1646–1652 (2010).

315. Deshpande, A. M. et al. PHC3, a component of the hPRC-H complex, associates with E2F6 during G0 and is lost in osteosarcoma tumors. *Oncogene* **26**, 1714–1722 (2007).

316. Tan, K. et al. Tumor suppressor RYBP harbors three nuclear localization signals and its cytoplasm-located mutant exerts more potent anti-cancer activities than corresponding wild type. *Cell. Signal.* **29**, 127–137 (2017).

317. Zhan, S., Wang, T., Ge, W. & Li, J. Multiple roles of Ring 1 and YY1 binding protein in physiology and disease. *J. Cell. Mol. Med.* **22**, 2046–2054 (2018).

318. Seitz, V. et al. Classical hodgkin's lymphoma shows epigenetic features of abortive plasma cell differentiation. *Haematologica* **96**, 863–870 (2011).

319. Zhou, H. et al. RING1 and YY1 binding protein suppresses breast cancer growth and metastasis. *Int. J. Oncol.* **49**, 2442–2452 (2016).

320. Tong, A.-H. et al. Overexpression of RYBP inhibits proliferation, invasion, and chemoresistance to cisplatin in anaplastic thyroid cancer cells via the EGFR pathway. *J. Biochem. Mol. Toxicol.* **33**, e22241 (2019).

321. Zhuang, X.-F. et al. miR-34b inhibits the migration/invasion and promotes apoptosis of non-small-cell lung cancer cells by YAF2. *Eur. Rev. Med. Pharmacol. Sci.* **23**, 2038–2046 (2019).

322. Zhang, S. et al. YAF2 exerts anti-apoptotic effect in human tumor cells in a FANK1- and phosphorylation-dependent manner. *Biochem. Biophys. Res. Commun.* **554**, 99–106 (2021).

323. Nagel, S. et al. Deregulation of polycomb repressor complex 1 modifier AUTS2 in T-cell leukemia. *Oncotarget* **7**, 45398–45413 (2016).

324. Coyaud, E. et al. PAX5-AUTS2 fusion resulting from t(7;9)(q11.2;p13.2) can now be classified as recurrent in B cell acute lymphoblastic leukemia. *Leukemia research* **34**, e323–5 (2010).

325. Astolfi, A. et al. BCOR involvement in cancer. *Epigenomics* **11**, 835–855 (2019).

326. Panagopoulos, I. et al. Fusion of the ZC3H7B and BCOR genes in endometrial stromal sarcomas carrying an X;22-translocation. *Genes. Chromosomes Cancer* **52**, 610–618 (2013).

327. Tanaka, T. et al. Internal deletion of BCOR reveals a tumor suppressor function for BCOR in T lymphocyte malignancies. *J. Exp. Med.* **214**, 2901–2913 (2017).

328. Isshiki, Y. et al. KDM2B in polycomb repressive complex 1.1 functions as a tumor suppressor in the initiation of T-cell leukemogenesis. *Blood Adv* **3**, 2537–2549 (2019).
329. Kottakis, F. et al. NDY1/KDM2B functions as a master regulator of Polycomb complexes and controls self-renewal of breast cancer stem cells. *Cancer Res* **74**, 3935–3946 (2014).

330. He, J., Nguyen, A. T. & Zhang, Y. KDM2b/JHDM1b, an H3K36me2-specific demethylase, is required for initiation and maintenance of acute myeloid leukemia. *Blood* **117**, 3869–3880 (2011).

331. Gounder, M. et al. Tazemetostat in advanced epithelioid sarcoma with loss of INI1/SMARCB1: an international, open-label, phase 2 basket study. *Lancet. Oncol.* **21**, 1423–1432 (2020).

332. Morschhauser, F. et al. Tazemetostat for patients with relapsed or refractory follicular lymphoma: an open-label, single-arm, multicentre, phase 2 trial. *Lancet. Oncol.* **21**, 1433–1442 (2020).

333. Italiano, A. et al. Tazemetostat, an EZH2 inhibitor, in relapsed or refractory B-cell non-Hodgkin lymphoma and advanced solid tumors: a first-in-human, open-label, phase 1 study. *Lancet. Oncol.* **19**, 649–659 (2018).

334. Vaswani, R. G. et al. Identification of (R)-N-((4-Methoxy-6-methyl-2-oxo-1,2-dihydropyridin-3-yl)methyl)-2-methyl-1-(1-(1-(2,2,2-trifluoroethyl)piperidin-4-yl)ethyl)-1H-indole-3-carboxamide (CPI-1205), a potent and selective inhibitor of histone methyltransferase EZH2, Suitab. *J. Med. Chem* **59**, 9928–9941 (2016).

## ACKNOWLEDGEMENTS

Research in the G.C. laboratory was supported by grants from the European Research Council (Advanced Grant 3DEpi, No. 788972), the EU (CHROMDESIGN, No. 813327), the Fondation pour la Recherche Médicale (DEI20151234396), the MSDAVENIR foundation (Project GENE-IGH), the INSERM and the French National Cancer Institute (INCa PLBIO18-362). A-M.M. is supported by a grant of the Fondation ARC pour la Recherche contre le Cancer (No. 216574). V.P. was supported by the Laboratory of Excellence EpiGenMed.

## AUTHOR CONTRIBUTIONS

All authors have written and edited the manuscript.

## COMPETING INTERESTS

The authors declare no competing interests.

## ADDITIONAL INFORMATION

**Correspondence** and requests for materials should be addressed to Anne-Marie Martinez or Giacomo Cavalli.

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