Combining epigenetic drugs with other therapies for solid tumours - past lessons and future promise
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Combining epigenetic drugs with other therapies for
solid tumours - past lessons and future promise
Daphné Morel, Daniel Jeffery, Sandrine Aspeslagh, Geneviève Almouzni,
Sophie Postel-Vinay
To cite this version:
Daphné Morel, Daniel Jeffery, Sandrine Aspeslagh, Geneviève Almouzni, Sophie Postel-Vinay. Com-
bining epigenetic drugs with other therapies for solid tumours - past lessons and future promise. Nature
Reviews Clinical Oncology, Nature Publishing Group, 2019, 17 (2), pp.91-107. �10.1038/s41571-019-
0267-4�. �hal-02371032�
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CopyrightREVIEWS
Combining epigenetic drugs with
other therapies for solid tumours —
past lessons and future promise
Daphné Morel 1,5, Daniel Jeffery 2,5
, Sandrine Aspeslagh3, Geneviève Almouzni 2,5
*
and Sophie Postel-Vinay 1,4,5*
Abstract | Epigenetic dysregulation has long been recognized as a key factor contributing to
tumorigenesis and tumour maintenance that can influence all of the recognized hallmarks of
cancer. Despite regulatory approvals for the treatment of certain haematological malignancies, the
efficacy of the first generation of epigenetic drugs (epi-drugs) in patients with solid tumours has
been disappointing; however, successes have now been achieved in selected solid tumour subtypes,
thanks to the development of novel compounds and a better understanding of cancer biology that
have enabled precision medicine approaches. Several lines of evidence support that, beyond their
potential as monotherapies, epigenetic drugs could have important roles in synergy with other
anticancer therapies or in reversing acquired therapy resistance. Herein, we review the mechanisms
by which epi-drugs can modulate the sensitivity of cancer cells to other forms of anticancer therapy,
including chemotherapy, radiation therapy, hormone therapy, molecularly targeted therapy and
immunotherapy. We provide a critical appraisal of the preclinical rationale, completed clinical
studies and ongoing clinical trials relating to combination therapies incorporating epi-drugs. Finally,
we propose and discuss rational clinical trial designs and drug development strategies, considering
key factors including patient selection, tumour biomarker evaluation, drug scheduling and response
assessment and study end points, with the aim of optimizing the development of such combinations.
1
ATIP–Avenir Group, Over the past decade, remarkable advances in antican- epi-drugs, which are almost exclusively DNA methyl-
UMR981, INSERM (French cer therapy have been made, owing notably to the devel- transferase (DNMT) inhibitors or histone deacetylase
National Institute of Health opment of precision medicine and immunotherapy (HDAC) inhibitors (Box 1). However, success with
and Medical Research),
approaches as well as continuous improvements in the certain third-generation epi-drugs used according to
Gustave Roussy Cancer
Campus, Villejuif, France. available therapeutic options and strategies. Nevertheless, precision medicine paradigms has provided new hope
2
Nuclear Dynamics Unit —
the persistent ability of cancers to resist treatment in the field of epigenetic therapy. For example, the iso
UMR3664, National Centre remains a major challenge that limits the effectiveness citrate dehydrogenase 1 (IDH1) inhibitor ivosidenib
for Scientific Research, all anticancer therapies. received FDA approval on the basis of the results of
Institut Curie, Paris, France. Epigenetic dysregulation has major roles in tumour a phase I trial involving patients with IDH1-mutant
3
University Hospital Brussels, development, progression and acquisition of therapeutic acute myeloid leukaemia (AML), only 4 years after the
Brussels, Belgium.
resistance, in both solid and haematological malignan- first patient was enrolled3. In addition, tazemetostat,
4
Drug Development cies1. To date, however, the efficacy of ‘epi-drugs’ — drugs an inhibitor of the histone-lysine N-methyltransferase
Department (DITEP), Gustave
Roussy Cancer Campus,
that target enzymes involved in epigenetic regulation of enhancer of zeste homologue 2 (EZH2), has demon-
Paris-Saclay University, genome function (Fig. 1) — has mostly been confined to strated efficacy in patients with EZH2-mutant haemato-
Villejuif, France. haematological cancers2, perhaps in part because solid logical malignancies or highly aggressive solid tumours
5
These authors contributed tumours tend to arise from more-differentiated or even harbouring genetic aberrations affecting the SWI/SNF
equally: Daphné Morel, Daniel terminally differentiated cells with a reduced capability chromatin remodelling complex, such as SMARCB1
Jeffery, Geneviève Almouzni for epigenetic reprogramming. Furthermore, biomark- (also known as INI1) or SMARCA4 mutations or dele-
and Sophie Postel-Vinay
ers for patient selection have crucially been lacking in tions4. Furthermore, responses to bromodomain and
*e-mail: genevieve.almouzni@
early phase trials of epi-drugs, which have largely been extra-terminal domain (BET) inhibitors, which tar-
curie.fr; sophie.postel-vinay@
gustaveroussy.fr conducted according to the historical ‘one size fits all’ get certain bromodomain-containing protein (BRD)
https://doi.org/10.1038/ approach. This strategy probably hampered the devel- family members, have been observed in patients with
s41571-019-0267-4 opment of first-generation and second-generation BRD4-rearranged NUT midline carcinoma5.
Nature Reviews | Clinical OncologyReviews
Key points comprise ten biological capabilities acquired during
oncogenesis that drive and/or enable the development
• The first generation of drugs targeting the epigenome (epi-drugs) were developed of cancer, as defined by Hanahan and Weinberg10.
using a ‘one size fits all’ approach and proved to have disappointing efficacy in Various epi-drugs are able to enhance the effective-
patients with solid tumours. ness of four major pillars of the anticancer treatment
• The potential of epi-drugs to modulate the sensitivity of tumours to other anticancer armamentarium, namely genotoxic and/or cytotoxic
drugs and to overcome therapy resistance presents major new avenues of clinical treatments (encompassing chemotherapy and radio-
investigation.
therapy), hormone therapy, molecularly targeted therapy
• A new generation of epi-drugs, which were developed to be more specific for their and immunotherapy11 (Fig. 2). In the following sections of
targets and have promising activity in certain biomarker-selected populations, is
this Review, we describe the synergistic mechanisms
now entering early phase clinical trials and are showing promising efficacy.
of epi-drug combinations stratified according to each of
• Epi-drug development should follow a precision-medicine approach, with further
these four pillars of therapy, first discussing the preclini
identification of robust predictive biomarkers for patient selection and subsequent
implementation of this strategy in clinical trials.
cal evidence, followed by examples of corresponding
clinical trials. The examples provided were selected to
• Sequential treatment schedules and/or dosing at less than the maximum tolerated
dose might improve the efficacy and tolerability of epi-drugs when used in
illustrate the specific challenges to the development
combination with various other anticancer therapies. of epi-drug combinations (including those relating to
patient selection, treatment schedules, toxicities and
efficacy assessments) and to highlight the most recent
Cancers continuously evolve, in part through and most prominent clinical successes and failures.
dynamic and reversible epigenetic changes that confer We also provide comprehensive lists summarizing past
a fitness advantage. This epigenetic plasticity — a term and ongoing trials of epi-drugs in combination with
used hereinafter in reference to reversible changes in other anticancer therapies (Supplementary Tables 1
epigenetic marks on DNA, histone and non-histone and 2, respectively).
proteins, as well as the functional consequences of these
alterations — is also involved in primary and acquired Combinations with cytotoxic agents
resistance to various anticancer therapies, through mod- Chemotherapies. In preclinical studies assessing the
ulation of tumour cells or their microenvironment6,7 effects of epi-drugs in combination with chemotherapy,
(Supplementary Figure 1). Epi-drugs might therefore synergy has mostly been achieved with DNA-damaging
be most effective when used in combination with other agents. HDAC inhibitors and DNMT inhibitors pro-
treatments and present particularly attractive strate- mote global chromatin relaxation, which facilitates DNA
gies for sensitizing cancer cells to a given therapy and damage by genotoxic agents and interferes with
for overcoming acquired resistance mechanisms in a DNA-damage repair (DDR)12 (Fig. 3a). HDAC inhib-
dynamic fashion. Of note, cancer stem cells, which are itors further impede DDR by disrupting ATM signal-
characterized by their abilities to self-renew and initiate ling, modulating the post-translational modification
cancer8, probably also have important roles in thera- of non-histone substrates of HDACs, including p53
peutic resistance. The inherent phenotypic plasticity and X-ray repair cross-complementing protein 6 (also
of these stem-like cells is key to this resistance and is known as Ku70), and by interfering with the regula-
regulated by both transcription factors and epigenetic tion of histone marks that regulate DDR13. In addition,
changes. Indeed, some third-generation epi-drugs, combined use of HDAC inhibitors and DNMT inhibi
such as lysine-specific histone demethylase 1A (LSD1) tors at low doses might revert resistance to cytotoxic
inhibitors, have been shown to disrupt the molecular agents, in part, by inducing removal of acquired epi
changes underlying this plasticity. However, this topic genetic alterations that drive the resistance phenotype14,15.
has been reviewed elsewhere9 and will not be discussed Furthermore, epigenetic silencing of genes involved in
further herein. shaping the tumour microenvironment (such as TGFBI)
Despite robust preclinical evidence supporting has been associated with resistance to taxanes and can be
epi-drug-containing combination therapies, clinical reversed with DNMT inhibitors, such as 5-azacytidine16,
success with first-generation and second-generation whereas defects in genes encoding components of chro-
epi-drugs has been limited. With the new generation of matin remodelling complexes (such as SMARCA4) are
novel, selective epi-drugs currently entering clinical test- correlated with sensitivity to taxanes and cisplatin17,18.
ing, learning from past successes and failures is crucial to Interestingly, Fillmore et al.19 showed that inhibition of
maximizing future patient benefits. Herein, we provide EZH2 increases the sensitivity of SMARCA4-mutant or
a critical review of preclinical and clinical data support- EGFR-mutant non-small-cell lung cancers (NSCLCs) to
ing the use of epi-drugs in combination with other anti- DNA topoisomerase 2 (TOP2) inhibitors, but conversely
cancer therapies in the treatment of patients with solid promotes resistance in tumours that are wild type for
tumours. We also highlight the high stakes and chal- both genes. These findings highlight the importance
lenges of successful epi-drug development and propose of considering genomic characteristics of the tumour
clinically relevant approaches to optimize this process. when selecting patients for therapy with particular epi-
drugs. Other third-generation epi-drugs, such as BET
An appraisal of epi-drug combinations inhibitors and protein arginine N-methyltransferase 5
Epigenetic regulation influences all of the recognized (PRMT5) inhibitors, also interfere with DDR processes,
hallmarks of cancer, which can each be targeted using raising the possibility of synergy with cytotoxic therapy
various therapeutic approaches. These hallmarks (reviewed in detail elsewhere20,21).
www.nature.com/nrclinoncReviews
Nuclear
phase II studies24,25 conducted to evaluate this strategy
position by combining decitabine with carboplatin in the treat-
ment of women with recurrent and/or progressing ovar-
Nucleus Chromatin
Higher-order remodellers ian cancer illustrate certain challenges associated with
chromatin epi-drug–chemotherapy combinations. In one of these
trials25, low-dose decitabine (10 mg/m2) was admin
istered on days 1–5 of a 28-day cycle, which seemed to
result in better tolerability than high-dose decitabine
(45 or 90 mg/m2) administered on day 1 of each cycle
Histone in the other trial24 (Supplementary Table 1). Differences in
PTM
the trial designs and patient populations precludes any
Ac BRD readers
Structural RNA formal comparison of these trials, although the level of
Nucleosome efficacy — in terms of objective response rate (ORR)
BETi and progression-free survival (PFS) — in the trial of
low-dose decitabine (which included patients with
Me
Ac Histone PTM platinum-refractory disease) also seemed to be superior
DNA writers/ to that observed in the trial of the high-dose regimen
erasers (in which patients with partially platinum-sensitive dis-
HMT HDM
HAT HDAC ease were enrolled), thus supporting the selection of the
DNA
modifiers low-dose regimen for further testing in a subsequent
DNMTi DNMT HATi HDACi HMTi HDMi phase III trial (Supplementary Table 2). Interestingly, both
(SIRTi) (PRMTi) studies showed a high incidence of carboplatin-related
Me hypersensitivity reactions with the combination (35%
Ac Me and 67%), potentially resulting from decitabine-induced
TETi TET
demethylation of genes encoding cytokines involved in
Gene commitment of naive T cells to the T helper 1 (TH1) and
IDH α-KG Trans-acting T helper 2 (TH2) lineages and IgE-mediated hypersensi-
factors tivity reactions24,26. Global decitabine-induced 5-methyl-
IDHi Mutant D-2-HG cytosine demethylation of DNA from peripheral blood
IDH
mononuclear cells (PBMCs) was also noted in both
Fig. 1 | Targeting the epigenome. Genome function is regulated at multiple levels by trials24,25. This effect was dose-dependent27, which is in
>800 epigenetic enzymes. A predominant aspect of this regulation involves variations in contrast to the findings of other clinical trials indicating
the organization of genomic DNA within chromatin structures, the basic unit of which that the pharmacodynamic modulation induced by cer-
is the nucleosome core particle consisting of 146 bp of DNA wrapped around a histone tain epi-drugs reaches a plateau at doses well below the
octamer. Epigenetic modifiers acting at this level comprise four nonexclusive categories: maximum tolerated dose4,28. Notably, Matei et al.25 identi-
the ‘writers’, which add specific marks to DNA or the core histones H2A, H2B, H3, and H4; fied that patients with a PFS duration of >6 months have
‘readers’, which recognize these marks; ‘erasers’, which remove them; and the ‘shapers’, higher numbers of demethylated genes in tumour biopsy
which remodel chromatin, mobilize nucleosomes or mediate histone exchange. Current
samples and in PBMCs, suggesting that PBMC DNA
epigenetic drugs (epi-drugs) target proteins belonging to the first three categories. These
epi-drugs include inhibitors of DNA-modifying enzymes, such as DNA methyltransferases methylation status could provide information on tumour
(DNMTs) that can methylate certain cytosine or adenosine nucleotides of DNA and DNA methylation and serve as a pharmacodynamic
ten-eleven translocation (TET) enzymes that catalyse the oxidation of 5-methylcytosine in or predictive biomarker.
DNA to initiate the process of active DNA demethylation. Another category of epi-drugs Overall, despite the preclinical data showing that
with effects on DNA methylation target gain-of-function mutant forms of isocitrate HDAC inhibitors and DNMT inhibitors potentiate
dehydrogenase (IDH) enzymes, which produce the oncometabolite d-2-hydroxyglutarate the activity of cytotoxic chemotherapy, clinical results
(D-2-HG) that competitively inhibits TET hydroxylases and thereby results in aberrant have been disappointing: three out of the five key ran-
DNA methylation. By contrast, wild-type IDH enzymes support DNA demethylation by domized trials of such combinations were halted owing
producing α-ketoglutarate (α-KG), which is a cofactor for TET proteins. Other epi-drugs to a lack of efficacy and/or unfavourable toxicity pro-
inhibit writers or erasers of histone arginine and/or lysine post-translational modifications
files when compared with chemotherapy alone. Such
(PTMs), including histone acetyltransferases (HATs), histone deacetylases (HDACs),
histone methyltransferases (HMTs) and other protein arginine methyltransferases undesired systemic toxicities and limited efficacy might
(PRMTs) and histone demethylases (HDMs), which can also act on non-chromatin factors. warrant investigation of targeted delivery of epi-drugs,
An additional class of epi-drugs inhibits readers of histone PTMs, such as bromodomain for example, as payloads of antibody–drug conjugates
and extra-terminal domain (BET) family proteins (BRDs), which bind acetylated lysine (ADCs) or liposomal particles. Moreover, most — if
residues of histones (as well as other non-chromatin factors). Epi-drugs targeting not all — trials of epi-drug–chemotherapy combina-
chromatin remodelling factors and histone chaperones are also under development, tions have been performed in unselected patient popu
but are beyond the scope of this manuscript. SIRTi, sirtuin inhibitor. Figure adapted from lations (that is, without the use of predictive molecular
ref.161, Springer Nature Limited. biomarkers). Thus, appropriate patient selection using
rational epigenetic biomarkers might produce clinically
Findings of preclinical studies have demonstrated the meaningful improvements in the therapeutic window
potential of DNMT inhibitors to induce the re-expression and new therapeutic opportunities. For example, evalu
of epigenetically silenced genes and thereby restore ating SLFN11 or PCFT promoter hypermethylation
sensitivity of ovarian cancers to platinum-based chemo- using clinical grade assays could potentially enable the
therapy22,23. However, the discordant results of two identification of patients who would benefit most from
Nature Reviews | Clinical OncologyReviews
Box 1 | generations of epi-drugs and radiotherapy was well tolerated in 21 patients with
localized pancreatic ductal adenocarcinoma and led to
Multiple generations of epigenetic drugs (epi-drugs) have been developed. four R0 resections among 11 patients who underwent
First-generation and second-generation epi-drugs include DNA methyltransferase surgical exploration and a median overall survival (OS)
inhibitors (DNMTi) and histone deacetylase inhibitors (HDACi), some of which have
duration of 1.1 years for the entire cohort, which accord-
already been approved for the treatment of certain haematological malignancies2.
ing to the investigators compared favourably with the
DNMTi are pyrimidine analogues that are incorporated into DNA during replication and
form covalent DNMT–drug adducts that trigger the activation of DNA damage response results of other phase I studies of chemoradiotherapy
and can, ultimately, lead to apoptosis, thus explaining the dual epigenetic and cytotoxic in such a patient population38. Second, the combina-
effects of these drugs. With the exception of sirtuin inhibitors, which inhibit nicotinamide tion of vorinostat and vectorized internal radiotherapy
adenine dinucleotide (NAD+)-dependent class III histone deacetylases, HDACi inhibit the with 131I-metaiodobenzylguanidine in children with
Zn2+-dependent catalytic activity of either all (pan-HDACi) or single HDAC enzymes relapsed and/or refractory high-risk neuroblastoma led
(for example, HDAC6i). First-generation DNMTi (azacytidine and decitabine) and HDACi to an ORR of 12% at all dose levels and 17% at the recom
(vorinostat and romidepsin) have unfavourable pharmacokinetic characteristics and mended phase II dose;39 this regimen is now being
target selectivity, which led to the development of second-generation DNMTi (such as evaluated in a randomized phase II trial (Supplementary
zebularine and guadecitabine) and HDACi, (including the hydroxamic acid HDACi Table 2). Why these two studies had promising results
belinostat and panobinostat, the benzamide HDACi tucidinostat (also known as
whereas other clinical trials of epi-drug–radiotherapy
chidamide) and carboxylic acid derivative HDACi (such as valproic acid)), with improved
chemical properties under physiological conditions. combinations did not is unclear, although the appar-
Third-generation epi-drugs include, among others, bromodomain and extra-terminal ently greater permissivity of tumours to epigenetic
domain inhibitors (BETi), histone methyltransferase inhibitors (HMTi) and histone reprogramming in the latter study might be explained
demethylase inhibitors (HDMi) and protein arginine methyltransferase inhibitors (PRMTi). by the paediatric nature of the disease (which is likely to
BETi disrupt the recognition of acetylated lysine residues on histones, a mark associated have arisen during early development, when epigenetic
with active transcription, by BET-containing reader proteins, including certain members processes have key roles in determining cell fate). These
of the bromodomain-containing protein (BRD) family. HMTi (including enhancer of zeste results call for the implementation of biomarkers for
homologue 2 inhibitors and DOT1-like histone-lysine methyltransferase inhibitors), patient selection. For example, a vorinostat sensitivity
HDMi (such as lysine-specific histone demethylase 1A inhibitors) or PRMTi (for example, transcriptional signature (sig-139), which is enriched
targeting PRMT1 or PRMT5) reduce the catalytic activity of the specific target enzyme
for the expression of genes involved in DNA synthe-
or, in the specific case of embryonic ectoderm development (EED) protein inhibitors,
destabilize the enzymatic complex containing the target protein. sis, repair and chromatin remodelling as well as RNA
synthesis, splicing and processing has been identified
using a panel of cell lines, with positive associations
the use of DNMT inhibitors to reverse resistance to with PFS and OS observed in patients with newly diag-
platinum-based agents29 or pemetrexed30, respectively. nosed glioblastoma treated with vorinostat plus standard
temozolomide and radiation therapy40.
Radiotherapy. In preclinical studies, HDAC, DNMT,
EZH2 or BET inhibition potentiates the antitumour Combinations with hormone therapy
activity of radiotherapy by disrupting DDR and the Oestrogen receptor-positive breast cancer. Oestrogen
cell cycle and increasing oxidative stress31–35 (Fig. 3a). receptor-α (ERα), the pivotal oncogenic driver of 70%
Transient HDAC inhibition with valproic acid (before of breast cancers, is released from heat shock protein 90
and up to 24 hours after irradiation) results in radio- (HSP90) upon binding with its endogenous ligands,
sensitization of human glioma cell lines — a schedule predominantly oestradiol. The ERα–oestrogen complex
that might limit the systemic toxicities associated with subsequently translocates into the nucleus and binds to
chronic HDAC inhibition36. Interestingly, BET inhi- oestrogen-responsive elements — a process tightly regu-
bition can decrease the risk of radiation-induced lung lated by chromatin-remodelling factors41,42. Preclinically,
fibrosis and has a radioprotective effect on irradiated HDAC inhibitors have been shown to increase the anti-
non-malignant lung tissue in mouse models, but con- tumour efficacy of, or reverse resistance to, hormone
versely radiosensitizes thoracic cancer cells (that is, therapies in models of ERα-positive breast cancer43–45.
NSCLC, breast or oesophageal cancer cells) in vitro37. HDAC inhibition interferes with ERα signalling pathway
This potential ability to differentially modulate radio- at multiple levels, including transcriptional suppression
sensitivity according to cell lineage, if recapitulated in of ESR1 (which encodes ERα), reductions in ERα mRNA
humans, could provide new therapeutic opportunities. and protein stability and modulation of the transcrip-
Certain DNMT inhibitors, such as the cytidine tion of ERα targets as well as other genes involved in cell
analogues decitabine and 5-azacytidine, can be incor- proliferation and metastasis46,47 (Fig. 3b). HDAC6 inhibi-
porated into DNA and/or RNA; thus, these agents are tion, specifically, leads to HSP90 hyperacetylation, which
particularly strong radiosensitizers in all tissues and disrupts the HSP90–ERα association and favours ERα
the combination of these epi-drugs with radiotherapy ubiquitination and proteasomal degradation48. Selective
is not recommended owing to concerns regarding toxi HDAC6 inhibitors are, therefore, interesting candidates
city. In clinical trials, combining HDAC inhibitors with for future clinical evaluation48,49.
radiotherapy, usually concurrently and sometimes BET inhibition also suppresses the growth of
in addition to chemotherapy, has mostly resulted in tamoxifen-resistant tumour cells and xenografts in
increased levels of toxicity and minimal patient benefit combination with the anti-oestrogen fulvestrant50,51; the
(Supplementary Table 1). Two phase I studies, however, underlying mechanisms include inhibition of the BRD3
have provided encouraging results. First, the combina- and/or BRD4-dependent recruitment of the histone-
tion of the HDAC inhibitor vorinostat with capecitabine lysine N-methyltransferase NSD2 (also known as WHSC1)
www.nature.com/nrclinoncReviews
Genotoxic and/or cytotoxic
chemotherapy and radiotherapy Targeted therapies
DNMTi and/or HDACi DNMTi and/or HDACi
• Facilitate access to DNA • Revert EMT
• Re-express tumour suppressors
• Increase ROS HDACi
• Promote HIF1α
DNMTi, EZH2i and/or LSD1i degradation
• Differentiate cancer stem cells • Reduce oncoprotein
EZH2i stability
• Sensitive to TOP2i Genome BETi
BETi and/or PRMTi instability • Prevent oncogene
• Decrease DDR capabilities and mutation Activating transcription
Resisting invasion and
cell death metastasis
Enabling Inducing
replicative angiogenesis
immortality
Hallmarks of cancer
Tumour-
Evading growth promoting
suppressors inflammation
Sustaining Avoiding
proliferative immune
signalling response
Deregulating
cellular
energetics
Hormone therapy Immunotherapy
HDACi, HATi and/or BETi DNMTi, HDACi and/or EZH2i
• Reverse endocrine • Increase antigen presentation
resistance and chemokine and/or IFN
expression
DNMTi, HDACi and/or LSDi
• Enable viral mimicry
• M2→M1 macrophage
polarization
BETi
• Anti-inflammatory
Fig. 2 | Epi-drugs can enhance the activity of other anticancer therapies to counteract the hallmarks of cancer.
Current prominent types of anticancer therapeutics have counteracting effects against various hallmarks of cancer
(defined by Hanahan and Weinberg10 in 2011), as indicated by the coloured arcs surrounding the hallmarks of cancer wheel.
Cytotoxic and/or genotoxic chemotherapies and radiotherapy (green arc), hormone therapy (orange arc), immunotherapy
(blue arc) and targeted therapies (purple arc) each counteract specific hallmarks, with some overlap. Epigenetic drugs
(epi-drugs) can enhance the activity of these anticancer therapies through various mechanisms; examples of the relevant
categories of epi-drugs and their augmenting effects observed in preclinical studies are indicated in the figure for each type
of therapy. Accordingly, the combination of epi-drugs with other anticancer therapies can lead to sensitization to treatment
and/or reversal of resistance. BETi, bromodomain and extra-terminal domain inhibitor; DDR, DNA damage repair; DNMTi,
DNA methyltransferase inhibitor; EZH2i, enhancer of zeste homologue 2 inhibitor; EMT, epithelial-to-mesenchymal
transition; HATi, histone acetyltransferase inhibitor; HDACi, histone deacetylase inhibitor; HIF1α, hypoxia-inducible
factor 1α; IFN, interferon; LSD1i, lysine-specific demethylase 1 inhibitor; PRMTi, protein arginine methyltransferase inhibitor;
ROS, reactive oxygen species; TOP2i, DNA topoisomerase 2 inhibitor. Adapted with permission from ref.10, Elsevier.
which monomethylates and/or dimethylates histone H3 a selective inhibitor of the CREB-binding protein (CBP)
lysine 36 to generate H3K36me1 and H3K36me2 marks and histone acetyltransferase p300 (EP300) transcrip-
that are commonly associated with active transcription, tional co-activator proteins, via a mechanism involving
and disruption of elongation-associated phosphory disruption of the core ERα signalling complex54.
lation events on RNA polymerase II that results in stalled The results of two phase II studies demonstrate
transcription, thus ultimately reducing the expression of that combinations of HDAC inhibitors and an anti-
ESR1 and ERα target genes50–53. Similarly, single-agent oestrogen or aromatase inhibitor have acceptable
activity and synergism with anti-oestrogen therapies tolerability, with promising efficacy (Supplementary
has been demonstrated in preclinical studies of CPI-1, Table 1). The first trial, a single-arm study of vorinostat
Nature Reviews | Clinical OncologyReviews
a Irradiation Damage to DNA b
and/or proteins
Hormone precursor
H2O Premetrexed Aromatase
inhibitor
Oxidative Cyt c Microtubule
HDACi Oestrogen
stress or androgen
HDACi
HO• Anti-androgen Anti-oestrogen
DNMTi Taxanes
Proteasome HSP90
Platinum salts PRMTi
HR
Topoisomerase-i EZH2i
PARPi
Chromatin
relaxation DNMTi PCFT HDACi HDACi
PRMTi BETi
DNMTi
Transcription RNAPII
γH2AX Methylated DNA factor
DNA damage repair
BETi
HR target
BETi BRCA1 HR promoter CBPi genes
RAD51
ATM Ku70 RBBP8
Acetylated TOPBP1 HRE
protein Tumour
HDACi proliferation
c d
Regulatory
E-cadherin T cell
VEGFRi
HERi EZH2i M2 macrophage
HDACi
LSD1i
DNMTi
Cytotoxic
T cell
HDACi
RAS
PI3Ki PI3K HDACi
HIF1α CCL5, CXCL9, TCR
RAF BRAFi AKT HIF1α CXCL10, IFNα,
IFNβ M1 macrophage
mTORi mTOR
MHC
MEK MEKi
Proteasome
ERK Type I dsRNA
interferon
response
Oncoproteins
BETi
HDACi EZH2i
AXL, FGFR2, LSD1i
Methylated
HER3 (ERBB3), histone
MET, YAP BIM ISG ERV
TRAIL DNMTi
CDH1
(E-cadherin)
Kinome adaptation
www.nature.com/nrclinoncReviews
◀ Fig. 3 | Rationale for epi-drug combination, with examples of synergy or reversal the 8.3-month OS benefit observed in ENCORE 301
of resistance mechanisms. Overview of epigenetic drug (epi-drug)-induced modulation resulted from beneficial effects on hormone resistance,
of sensitivity to four pillars of anticancer therapy: radiotherapy, cytotoxic chemotherapy cancer stem cells, cell differentiation, anticancer immu-
and DNA damage repair-targeting therapies (part a); hormonotherapy (part b); targeted nity and/or sensitization to subsequent therapies is
therapies (part c); and immunotherapies (part d). Prominent processes are detailed here
unknown. Regardless, this finding led to the designa-
but others exist. Histone deacetylase inhibitors (HDACi) and DNA methyltransferase
inhibitors (DNMTi) synergise with genotoxic and/or cytotoxic therapies by interfering
tion of entinostat plus exemestane as a breakthrough
with the DNA damage response (DDR), notably through increased DNA damage and therapy by the FDA in postmenopausal women with
disruption of the function of DDR proteins. Bromodomain and extra-terminal domain advanced-stage hormone receptor-positive, endocrine-
inhibitors (BETi) globally decrease DDR protein levels, resulting in functional DDR defects. resistant breast cancer, and supported the develop-
Protein arginine methyltransferase 5 inhibitors (PRMTi), among other mechanisms, more ment of the E2112 study58. Results of this randomized,
specifically induce aberrant splicing of DNA repair factors, thereby impairing their activity double-blind, placebo-controlled phase III trial, in which
(part a). HDACi interfere with hormone receptors (HR) at multiple levels, including OS and PFS are co-primary end-points, are expected in
transcriptional regulation, mRNA and protein stability (notably through modulation of 2021 and should shed light on the mechanisms under-
acetylation levels of their chaperone protein HSP90) and transcription of nuclear target lying the benefits of adding entinostat to exemestane
genes. BETi suppress the transcription of the HR themselves or their target genes (part b).
(if confirmed) and on relevant biomarkers for such
HDACi modulate the stability of several tyrosine kinase receptors and downstream
proteins, notably through modulation of acetylation levels of HSP90. HDAC4/6 inhibitors combinations. Importantly and further supporting
more specifically reduce hypoxia-inducible factor 1α (HIF1α) stability and activity, thereby the above strategy, the randomized, double-blind,
synergizing with anti-angiogenic agents. BET inhibitors prevent the transcription of placebo-controlled, phase III ACE study, in which the
alternative tyrosine kinase receptors and proteins that drive resistance through the addition of the HDAC inhibitor tucidinostat to exemes-
‘kinome adaptation’ mechanism (part c). Treatment of cancer cells with enhancer of zeste tane was evaluated in postmenopausal patients with
homologue 2 inhibitors (EZH2i), lysine-specific histone demethylase 1A inhibitors (LSD1i), advanced-stage hormone receptor-positive breast can-
or DNMTi favours chemokine-dependent T cell attraction following enhanced expression cer that had progressed after previous endocrine therapy,
of CC-chemokine ligand 5 (CCL5), CXC-chemokine ligand 9 (CXCL9), CXCL10 or CXCL12. revealed a significant prolongation of PFS in the experi
LSD1i and DNMTi also stimulate type I interferon response through increased expression mental arm (7.5 months versus 3.8 months; HR 0.74,
of endogenous retroviral elements (ERV), which eventually triggers T cell-mediated
95% CI 0.58–0.98; P = 0.03)59.
antitumour immunity following double-strand RNA (dsRNA) stress. Epi-drugs can also
modulate immune cell transcriptional programmes. EZH2i elicit functional alterations in
regulatory T cells and enhance the cytotoxicity of effector T cells, while other epi-drugs, Hormone-sensitive prostate cancer. Observations similar
such as LSD1i, DNMTi and HDACi, decrease myeloid-derived suppressor cell activity to those described in preclinical models of ERα-positive
and favour a M1-like macrophage-mediated antitumour immune response (part d). breast cancer have been reported in models of both
ATM, ataxia telangiectasia mutated; BRAFi, BRAF inhibitor; CBPi, CREB-binding protein hormone-sensitive and castration-resistant prostate can-
inhibitor; Cyt c, cytochrome c; HERi, HER (ErbB) family receptor tyrosine kinase inhibitor; cer models (CRPC)60–63 (Fig. 3b). Additionally, BET inhi-
HRE, hormone response element; ISG, interferon-stimulated genes; MEKi, MEK bition prevents binding of the androgen receptor (AR)
inhibitor; mTORi, mTOR inhibitor; PARPi, poly(ADP-ribose) polymerase inhibitor; PI3Ki, to its target binding sites in the genome and the subse-
PI3K inhibitor; RNAPII, RNA polymerase II; TCR, T cell receptor; VEGFRi, VEGFR inhibitor. quent recruitment of RNA polymerase II; accordingly,
addition of the BET inhibitor JQ1 to the anti-androgen
enzalutamide or of the BET inhibitor OTX-015 to the
combined with tamoxifen in 43 patients with ER-positive, AR-agonist ARN-509 markedly reduced the growth of
endocrine-resistant metastatic breast cancer55, revealed enzalutamide-resistant prostate cancer xenografts in
durable (lasting >24 weeks) objective responses in 19% mice64,65. Early phase clinical trials of BET inhibitors com-
and disease stabilization in 21% of patients. In the second bined with next-generation anti-androgens in patients
trial, ENCORE 301, a randomized placebo-controlled with CRPC are ongoing (Supplementary Table 2).
study of entinostat added to exemestane in 130 women In a phase II trial in patients with CRPC resistant to
with ER-positive, endocrine-resistant advanced-stage second-line antiandrogen therapy66, the addition of the
breast cancer56, the entinostat combination resulted in HDAC inhibitor panobinostat to the second-generation
numerical but not statistically significant improvements anti-androgen bicalutamide increased the median PFS
in PFS (median 4.3 months compared with 2.3 months in in comparison with that of historical control groups
the placebo group; P = 0.11) but a striking significant (34 weeks versus 16 weeks). However, this potential
improvement in the exploratory OS end-point (median benefit was only observed with high-dose panobino
28.1 months versus 19.8 months; P = 0.036). In particu- stat (40 mg triweekly), which was associated with an
lar, patients with a percentage change in the level of increase in the incidence of severe toxicities: treatment
PBMC pan-protein lysine acetylation above the study interruption and dose reductions were needed in 27.5%
median had a better median PFS than those with a per- and 41% of patients, respectively, versus 11.5% and 4%
centage change below the median (8.6 months versus of patients receiving 20 mg triweekly. This finding sug-
2.8 months; HR 0.32, 95% CI 0.13–0.79). Immunological gests that more selective compounds with an improved
analyses revealed significant depletion of blood granu- therapeutic window might be required for the long-term
locytic and monocytic myeloid-derived suppressor cells clinical development of such combinations.
(MDSCs; P = 0.029 and P = 0.002, respectively), as well
as decreased CD40 expression in immature and mono- Combinations with targeted therapy
cytic MDSCs (P = 0.007 and P = 0.011, respectively) and ErbB-targeted therapies. Preclinical evidence shows that
increased HLA-DR expression in CD14+ monocytes resistance to agents targeting ErbB (also known as HER)
and enrichment of this CD14+HLA-DR hi monocyte family receptor tyrosine kinases (RTKs) can be epige-
subset (P = 0.015 and P = 0.0004, respectively)57. Whether netically driven and thus reverted by epi-drugs (Fig. 3c).
Nature Reviews | Clinical OncologyReviews
Examples include the reversion of epithelial-to- the basis of EGFR mutational status or using epigenetic
mesenchymal transition (EMT) that can occur with biomarkers73. Likewise, in the phase I/II IVORI study
ErbB-targeted therapies through HDAC inhibition8 involving unselected patients with NSCLC74, gefinitib
or the hyperacetylation of HSP90 chaperone proteins plus vorinostat resulted in a median PFS of 9.1 months
resulting from HDAC6 inhibition, which decreases that did not seem to be better than historical data75. In
interactions of HSP90 with stabilization of multiple this regard, an ongoing randomized, placebo-controlled,
RTKs, including EGFR (ErbB-1) 67. Target-specific double-blind phase II/III trial (NCT02416739) of
mechanisms also exist. A deletion polymorphism that gefitinib or erlotinib with or without the HDAC3 inhib-
results in alternative splicing of BCL2L11 (encoding itor nicotinamide in patients with EGFR-mutant NSCLC
the pro-apoptotic BCL-2-like protein 11, also known should provide more relevant data and could potentially
as BIM) is associated with resistance of EGFR-mutant enable the identification of patient selection biomarkers
NSCLC cell lines to EGFR tyrosine kinase inhibi- for this combination (Supplementary Table 2).
tors (TKIs) and shorter PFS durations in series of
patients with EGFR-mutant NSCLC who received such Anti-angiogenic agents. Tumour angiogenesis is mainly
TKIs (6.6 months compared with 11.9 months in those controlled by hypoxia-inducible factor 1α (HIF1α) and
without the polymorphism) 68; notably, alternative its transcriptional target VEGF. Inhibition of HDAC4
splicing of BCL2L11 has been shown to be HDAC3- and/or HDAC6 reduces HIF1α stability and transcrip-
dependent and can be reverted by vorinostat in preclini tional activity by promoting its acetylation, polyubiq-
cal models69. The clinical relevance of this approach is uitination and subsequent proteasome degradation76.
currently being evaluated in a phase I trial of vorino Accordingly, HDAC inhibitors and VEGF inhibitors
stat and gefitinib in patients with EGFR-mutant NSCLC have synergistic anti-angiogenic and antitumour effects
harbouring the BCL2L11 deletion polymorphism in mouse models of various cancers77,78.
(Supplementary Table 2). Multiple early stage clinical trials have been condu
Targeting BRD4 using BET inhibitors also over- cted to evaluate the safety of combinations of HDAC
comes resistance to ErbB-targeted agents in preclinical inhibitors and anti-angiogenic agents, including
models of various tumour types, including anti-EGFR vorinostat with bevacizumab for clear cell renal cell
antibodies in head and neck squamous cell carcinoma carcinoma (RCC)79, resminostat with sorafenib for
(HNSCC) and the dual EGFR and HER2 (ErbB-2) hepatocellular carcinoma 80 and panobinostat with
TKI lapatinib in HER2-positive breast cancer, notably bevacizumab for high-grade glioma81,82 (Supplementary
through the abrogation of the transcription of alterna- Table 1). All of these trials provide evidence that the
tive kinases that drive acquired resistance, such as MET, combinations are tolerable and provided evidence of
AXL, FGFR2 or HER3 (ErbB-3) — a phenomenon clinical benefit79–82. The most recently reported phase I
referred as ‘kinome adaptation’70–72 (Fig. 3c). In in vitro study was conducted to assess the combination of
models of lapatinib-resistant breast cancer, the combi- abexinostat and pazopanib, with durable responses in
nation of JQ1 and lapatinib has even been reported to be seven of ten patients with pazopanib-refractory solid
superior to lapatinib plus other TKIs targeting the bypass tumours, including one response lasting >3.5 years83.
pathways, and removal of lapatinib from the media Among the 22 patients with RCC (including ten and
allowed the cells to regrow despite continued JQ1 mono eight patients with prior progression on pazopanib
therapy, suggesting that BET inhibition re-sensitized and anti-angiogenic agents, respectively), the ORR was
resistant cells rather than simply synergizing with lapa 27% and the median duration of response was 10.5 months.
tinib70,71. RNA sequencing (RNA-seq) revealed that Notably, histone acetylation and HDAC2 expression
the effect of BET inhibition was remarkably specific, in PBMCs were positively correlated with durable
as JQ1 monotherapy resulted in the downregulation responses to treatment83. These promising results led to
of only 8% of all expressed genes, but suppressed 27% of the initiation of the ongoing randomized, double-blind,
lapatinib-induced genes when used in combination placebo-controlled phase III RENAVIV trial of the same
with lapatinib70,71. Importantly, in HNSCC cell lines with combination, with a crossover design, which will ena-
acquired resistance to the anti-EGFR antibody cetuxi- ble a thorough evaluation of clinical benefit in patients
mab, this heterogeneous kinome adaptation is associated with VEGFR TKI-naive advanced-stage RCC and could
with increased levels of BRD4 expression72, which might provide opportunities for biomarker identification
therefore serve as patient selection biomarker for this (Supplementary Table 2).
combination.
Multiple phase I and/or II studies have shown the PI3K, AKT and/or mTOR inhibitors. Findings of several
feasibility of combining HDAC inhibitors and ErbB preclinical studies indicate the potential of combinations
inhibitors (predominantly EGFR TKIs in patients with of HDAC inhibitors and PI3K–AKT–mTOR path-
NSCLC, but also the anti-HER2 antibody trastuzumab way inhibitors, including the observations that HDAC
in patients with breast cancer) although no clear signs inhibition can destabilize AKT84,85, suppress survivin
of delays or reversal of resistance have been observed expression86, inhibit EMT and invasiveness, or increase
(Supplementary Table 1), perhaps because patients with oxidative stress87,88, and induce autophagy89. Inhibi
epigenetically driven resistance to trastuzumab were not tion of HDACs and the PI3K–AKT–mTOR pathway
selected. For example, patients with NSCLC enrolled in also converges on the upregulation of tumour suppres-
the ENCORE-401 randomized phase II study of erlotinib sor genes, such as FOXO1 in MYC-driven medullo
plus either entinostat or placebo were not selected on blastoma90 (Fig. 3c). Synergy also occurs between mTOR
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inhibitors and BET inhibitors in preclinical models of BET inhibitors also synergize with BRAF inhibi-
triple-negative breast cancer91. Interestingly, BET inhi- tors in preclinical models of BRAF-mutant melanoma,
bition can both prevent and revert resistance to mTOR which leads to increased cytotoxicity and epigenetic
inhibition in such models91. Similar to the aforemen- modulation of dedifferentiated precursor cells102,103.
tioned observations relating to countering the kinome Furthermore, BET inhibition can overcome resistance
adaptation mechanism of resistance, transcriptional to dual MEK and TBK1 inhibition in KRAS-driven
upregulation of RTKs upon PI3K inhibition can be NSCLC models by blocking transcriptional activation
overcome by preventing BRD4 binding at the pro- of YAP1, which is a core component of the Hippo sig-
moter regions of RTK genes using BET inhibitors92. nalling pathway that often contributes to tumorigene-
Furthermore, when single-agent treatments are inef- sis104. Interestingly, the investigators who conducted this
fective, dual PI3K–BET inhibition can induce apoptosis study anticipated that continuous BET inhibition would
across multiple cancer cell lines and mammary tumour not be tolerable, and used in vitro pharmacodynamic
models92. Similar results have been reported for pre- modelling to optimize an intermittent sequential admin-
clinical models of epithelial ovarian cancer and neuro istration schedule that was tolerable and proved to be
blastoma, in which PI3K inhibition overcomes resistance equally effective in mice. Clinical data from studies eval-
to BET inhibitors93,94. These data pose a considerable uating such combinations are not currently available, but
challenge to the drug development dogma that drugs considering that the toxicities have been a major chal-
that do not display single-agent activity should not lenge to the application of epi-drug combinations, with
progress beyond phase I testing95 and suggests that cer- long-term toxicities often being a limiting factor96,105,106,
tain drugs might only be effective when rationally used such intermittent scheduling approaches should be
in combinations. This approach might be especially encouraged and applied more systematically.
pertinent with PI3K inhibitors and first-generation epi-
drugs, the single-agent activity of which has thus far been PARP inhibitors. Preclinical synergy between HDAC
limited in patients with solid tumours. inhibitors and poly(ADP-ribose) polymerase (PARP)
Phase I trials in which mTOR inhibitors were inhibitors has been reported in models of triple-negative
combined with vorinostat in patients with various breast107, prostate108 and ovarian cancer109. Mechanisti
advanced-stage solid tumours have provided some evi- cally, HDAC inhibitors induce a ‘BRCAness’ pheno-
dence of therapeutic activity, but thrombocytopenia type following downregulation of proteins involved in
limited doses to levels below the full dose of each agent homologous recombination, including RAD51, BRCA1
individually, suggesting that synergistic effects also occur and UHRF1. Notably, the combination of olaparib and
in non-malignant cells96,97 (Supplementary Table 1). vorinostat is to be evaluated in patients with metastatic
Again, these findings warrant the evaluation of discon- breast cancer (NCT03742245). Furthermore, in the past
tinuous and/or sequential drug administration sched- 2 years, the results of four independent studies have
ules, targeted drug delivery and/or biomarker-driven shown a strong synergy between PARP inhibitors and
patient selection in order to improve the therapeutic BET inhibitors, notably, in preclinical models of homo
index. Early phase trials of HDAC inhibitors combined logous recombination-proficient ovarian cancer110–113
with mTOR inhibitors are ongoing in patients with (Fig. 3a). Mechanistically, BET inhibitors downregulate
various types of solid tumour, including the combina- various proteins involved in homologous recombina-
tion of everolimus and panobinostat in patients with tion, including BRCA1, BRCA2, RAD51 and RBBP8
H3.3 or H3.1 K27M-mutant glioma (NCT03632317; (also known as CtIP), suggesting broad effects on the
Supplementary Table 2). expression DDR-related genes rather than a targeted
synthetic lethality (as epitomized by PARP inhibition
BRAF and/or MEK inhibitors. In preclinical studies, in cancers associated with germline BRCA mutations).
HDAC inhibitors revert acquired resistance to BRAF Interestingly, a reciprocal observation was made in mod-
and/or MEK inhibitors, notably, by upregulating trans els of BET inhibitor-resistant CRPC, whereby acquired
cription of the caspase 8 inhibitor c-FLIPL98, modulating polycomb repressive complex 2 (PRC2)-mediated silenc-
PI3K signalling99 or downregulating melanocyte line- ing of DDR-related genes led to increased sensitivity to
age pathways (restoration of which drives adaptive drug PARP inhibitors114. Combinations of PARP and BET
resistance in melanoma cells)100 (Fig. 3c). In 2019, HDAC inhibitors have not yet been evaluated in clinical studies.
inhibition was further shown to dramatically enhance We anticipate that rational patient selection and optimal
the effectiveness of BRAF and/or MEK inhibition in dosing and scheduling will be key to maximizing the
preclinical models of both sensitive (BRAF-mutant) therapeutic window and, thus, potential clinical utility,
and resistant (NRAS-mutant or NF1-mutant) RAS considering that thrombocytopenia is a dose-limiting
pathway-driven melanoma101. Specifically, HDAC inhibi toxicity (DLT) of HDAC, PARP and BET inhibitors.
tors induced chemical synthetic lethality via suppression
of the non-homologous end-joining DDR pathway on Epi-drug–epi-drug combinations. Owing to the interplay
a background of BRAF and MEK inhibitor-mediated among various epigenetic processes, combining several
disruption of the alternative homologous recombina- epi-drugs might be a promising approach to anticancer
tion pathways101. Importantly, MGMT expression was therapy. For example, overexpression of KMT2A (which
found to be characteristic of tumours with inherent encodes lysine N-methyltransferase 2A, also known as
defects in DNA repair and to predict responsiveness to MLL1) drives resistance of EZH2-aberrant cancers to
such combination therapy101. EZH2 inhibition via the interaction of MLL1 with the
Nature Reviews | Clinical OncologyReviews
CBP and EP300 co-activators, which results in losses by divergent results obtained with EZH2 inhibitors.
of H3K27 methylation and reciprocal gains in H3K27 Inhibition of EZH2 in T cells leads to enhanced anti-
acetylation, thereby triggering oncogenic transcriptional tumour responses with ICIs in preclinical models by
reprogramming; however, BET inhibition disrupts sig- eliciting functional alterations in regulatory T (Treg)
nalling mediated by H3K27 acetylation and re-sensitizes cells and by increasing the cytotoxicity of effector
cancer cells to EZH2 inhibitors115. Similarly, BET inhi- T cells 134. In line with these results, a patient with
bition can reverse resistance to DNMT inhibitors in SMARCB1-negative chordoma and disease progression
preclinical models of colorectal cancer (CRC)116. on EZH2 inhibition with tazemetostat had a durable
and exceptional abscopal response to radiotherapy last-
Combinations with immunotherapies ing >2 years; comparisons of tumour biopsy samples
Antitumour immunity involves the complex interplay obtained prior to treatment and during treatment with
among immune, cancer and stromal cells. Specific tazemetostat revealed a substantial increase in intra
DNA-modifying or histone-modifying enzymes and tumoural and stromal infiltrates of CD8+ cytotoxic T cells
histone chaperones, including DNMTs, KMT1A (also and FOXP3+ Treg cells, as well as enhanced expression
known as SUV39H1) and chromatin assembly factor 1 of the immune-checkpoint proteins programmed
(CAF-1), respectively, contribute to both the immuno cell death 1 (PD-1) and lymphocyte activation gene 3
genicity of cancer cells and the lineage commitment (LAG3) on T cells135. These findings suggest that EZH2
and/or maturation of immune cells117,118; thus, epi-drugs inhibition can promote sustained antitumour immune
can potentially be used to modulate antitumour responses and lead to immune-checkpoint activa-
immunity119,120. Several mechanisms of resistance to tion135, although the consequences of these effects,
immune-checkpoint inhibitors (ICIs)121 can be reverted, particularly with regard to ICI sensitivity, remain
in preclinical models, by epigenetic manipulations unclear at present. Contradictory pro-tumour effects
(Fig. 3d) — a topic that we have reviewed elsewhere120. have been reported preclinically with the EZH2 inhib-
Here, we review important new data that have been itor GSK126, which resulted in increased numbers of
reported in the past few months. MDSCs but decreased numbers of CD4+ T cells and
Preclinical modulation of adaptive immunity has IFNγ+CD8+ T cells in the tumour microenvironment136.
been described with the ablation of LSD1 or inhibition Together, these observations suggest that the cellular
of HDACs or DNMTs, which can enhance antigen pres- composition of the tumour and drug administration
entation, in part through the induction of dsRNA pro- schedule can influence the immunomodulatory effects
duction from endogenous retrovirus (ERV) genes and the of EZH2 inhibition.
resultant type I interferon response, leading to increased Some emerging clinical findings underscore the
T cell infiltration of tumours and synergistic effects with promise of combinations of epi-drugs and ICIs, notably
ICIs122–124. Similarly, DNMT inhibitors can further syner- in tumours that are resistant and/or refractory to ICIs
gize with HDAC inhibitors to induce ERV transcription, (Supplementary Table 1). For example, the combination
promote CC-chemokine ligand 5 (CCL5)-dependent of the anti-PD-1 antibody pembrolizumab and vorino
T cell infiltration and favour T cell memory and effector stat in a phase I/Ib study resulted in one confirmed partial
phenotypes, and to downregulate immunosuppressive response (PR) and eight stable disease responses among
MYC signalling, thereby reverting immune evasion in 24 patients with ICI-resistant metastatic NSCLC137.
NSCLC models125. Interestingly, such an effect has also Moreover, preliminary results from the ENCORE-601
been described with CDK4/6 inhibitors, which indirectly phase Ib/II trial evaluating the combination of pembroli-
reduce the activity of DNMT1 and thereby increase ERV zumab and entinostat demonstrate a PR in one patient
expression126; this observation highlights how some with microsatellite-stable CRC (MSS-CRC), PRs in nine
non-epigenetic drugs can have clinically relevant indi- patients and a complete response in one patient with
rect epigenetic effects. Moreover, chemokine-dependent ICI-resistant melanoma and PRs in seven patients
T cell attraction has also been described following with ICI-resistant NSCLC (among 16, 53 and 67 patients,
enhanced expression of CXCL12 resulting from DNMT respectively)138–140; in the NSCLC expansion cohort, a
inhibition127 or of CCL5, CXCL9 and CXCL10 from high level of peripheral CD14+CD16− HLA-DRhi clas-
LSD1128 or EZH2 inhibition129. Finally, the development sic monocytes at baseline (defined as ≥13.1% of live
of selective clinical-grade inhibitors of the histone-lysine PBMCs per millilitre of blood) was correlated with
N-methyltransferase KMT1A might bring future clini- a better ORR (21.1% versus 6.5%) and median PFS
cal benefits via enhancement of the long-term memory (5.3 months versus 2.7 months)138. Contrary to these
reprogramming capacity of CD8+ T cells118. single-arm phase I–II trials of HDAC inhibitors, how-
With regard to innate immunity, LSD1, DNMT and ever, the only randomized study with results reported
HDAC inhibitors decrease the activity of MDSCs to date revealed an increased risk of toxicities and no
and promote an M1-like macrophage-mediated anti- therapeutic benefit from combining pembrolizumab
tumour immune response in mouse models of breast with the second-generation DNMT inhibitor CC-486
or pancreatic cancer130,131. HDAC inhibition can also (oral azacytidine)141. The choice of full-dose CC-486 and
enhance the cytolytic activity of NK cells and TLR9 the absence of patient selection, either for likely sensi-
agonist-induced abscopal responses (that is, in lesions tivity to pembrolizumab or CC-486, might at least par-
beyond the one treated with radiotherapy)132,133. tially explain these results. Results from other studies
Of note, modulation of immunity using epi-drugs of epi-drug plus ICI combinations are difficult to inter-
will probably require some fine-tuning, as illustrated pret at present because most data are preliminary and
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