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  • Review
  • Open Access

Epigenetic treatment of solid tumours: a review of clinical trials

Clinical EpigeneticsThe official journal of the Clinical Epigenetics Society20157:127

  • Received: 17 May 2015
  • Accepted: 10 November 2015
  • Published:


Epigenetic treatment has been approved by regulatory agencies for haematological malignancies. The success observed in cutaneous lymphomas represents a proof of principle that similar results may be obtained in solid tumours. Several agents that interfere with DNA methylation-demethylation and histones acetylation/deacetylation have been studied, and some (such as azacytidine, decitabine, valproic acid and vorinostat) are already in clinical use.

The aim of this review is to provide a brief overview of the molecular events underlying the antitumour effects of epigenetic treatments and to summarise data available on clinical trials that tested the use of epigenetic agents against solid tumours. We not only list results but also try to indicate how the proper evaluation of this treatment might result in a better selection of effective agents and in a more rapid development.

We divided compounds in demethylating agents and HDAC inhibitors. For each class, we report the antitumour activity and the toxic side effects. When available, we describe plasma pharmacokinetics and pharmacodynamic evaluation in tumours and in surrogate tissues (generally white blood cells).

Epigenetic treatment is a reality in haematological malignancies and deserves adequate attention in solid tumours. A careful consideration of available clinical data however is required for faster drug development and possibly to re-evaluate some molecules that were perhaps discarded too early.


  • Epigenetic treatment
  • Histone deacetylases (HDACs)
  • Histone methyltransferases (HMTs)
  • DNA methylation
  • DNA-methyltransferases
  • ncRNAs
  • Azacytidine
  • Decitabine
  • Valproic acid
  • Suberoylanilide hydroxamic acid (SAHA)


Research on tumour biology has provided conclusive evidence on the primary role of genetic alterations in the initiation and progression of cancer. However, the deregulation of epigenetic processes such as DNA methylation and alterations of “histone code” are equally important oncogenic factors per se [14]. Epigenetic processes affect the packaging of chromatin and direct distinct cellular gene expression programmes. They are heritable through cell division and do not involve changes in the DNA sequence [46]. Operating at the level of chromatin structure, epigenetic mechanisms play a key role during embryogenesis, X-chromosome silencing, cellular proliferation and differentiation and in disease states [2, 46]. They also facilitate a selective readout of the genome, thereby regulating stem cell developmental potential and cell fate. Subtle disturbances of the epigenetic framework in progenitor, differentiating or terminal cells may, besides well-known genetic alterations, promote carcinogenesis [7, 8]. The dynamic and reversible nature of epigenetic mechanisms makes these processes of therapeutic relevance in many diseases including cancer.

Epigenetic processes involve methylation of DNA and post-translational modification of nucleosomal histones, which contribute to a complex “epigenetic code” that superposes the nucleotide sequence to direct gene expression [4, 911] (Fig. 1a).
Fig. 1
Fig. 1

Schematic representation of gene expression regulation by epigenetic drugs, components of the DNA and chromatin-modifying machinery and ncRNAs. a Epigenetic drugs reported to be effective against cancer cells inhibit the activity of DNA methyltransferases (DNMTi) or histone deacetylases (HDACi). DNMTs add a methyl group (CH3) to the 5′ carbon atom of cytosine in DNA CpG dinucleotides. DNMTs also participate in multiprotein chromatin-modifying complexes containing histone deacetylases (HDACs) and histone methyltransferases (HMTs),which induce post-translational modifications of lysine residues in the amino terminal tails of nucleosomal histones, including deacetylation (HDACs), methylation (HMTs) and acetylation (histone acetyltransferases (HAT). Specific molecular modifications on CpGs and nucleosomal histones affect the higher order of chromatin architecture and function by changing the interaction of histones with DNA or the contact between different histones in adjacent nucleosomes. This allows or denies the accessibility of the transcriptional machinery and DNA-binding proteins to specific sites on genome, resulting in activation or silencing of gene transcription. Ac acetylation, Me methylation. b Short and long ncRNA are emerging as novel regulators of chromatin structure, alternative to DNA-binding proteins. They can act as key specificity determinants for epigenetic regulation of gene expression. In the nucleus, both short and long ncRNAs can bind complementary sequences on DNA or nascent RNA transcripts and guide the Argonaute-containing complexes (Ago) to recruit HDACs, HMTs and DNMTs for gene silencing. Nascent lncRNAs can also be tethered to the locus from which they are transcribed through association with RNA polymerase II (Pol II). In the cytosol, microRNAs and siRNAs act as post-transcriptional regulators of the expression of HDAC and DNMTs through their complementarity with mRNA sequences

DNA methylation results from the transfer of a methyl group from a methyl donor substrate, namely S-adenosyl-L-methionine (AdoMet), to the 5′ position of a cytosine in a CpG context. CpG dinucleotides can be sparse or tend to be gathered in repetitive sequences in or around gene promoters in regions known as CpG islands. The methylation status of CpG moieties within regulatory DNA sequences affects the transcription of the related gene [2, 10]. The creation of DNA methylation patterns during the embryogenesis establishes the compartmentalization of genome into transcriptionally active and inactive domains. DNA methylation is fundamental for a correct expression of imprinted genes, chromosomal dosage compensation (X-chromosome inactivation) and tissue-specific gene expression [2, 3, 1214].

The oncogenic effect of DNA methylation is mainly related to the formation of a repressive chromatin structure on promoter regions that impairs the constitutive expression of genes involved in cell cycle regulation, DNA repair, apoptosis, differentiation, drug resistance, angiogenesis and metastasis [14]. A family of enzymes known as DNA methyltransferases (DNMTs) catalyses the DNA methylation reaction. DNMT1 is a maintenance methylase that recognises and methylates hemi-methylated CpG dinucleotides during DNA replication allowing the propagation and conservation of the DNA methylation patterns through the future generations [14, 15]. DNMT3a and 3b generally act as de novo methylases. They are highly expressed in embryonic stem cells, early embryos and developing germ cells and, at a low rate, in somatic tissues or postnatal animals. DNA methyltransferase-3-like (DNMT3-Like) lacks enzymatic activity but may be essential for the establishment of maternal methylation imprints and the appropriate expression of maternally imprinted genes. The inhibitory effect of CpG island methylation on gene expression is mediated by the involvement of proteins with high affinity for methylated CpGs. These methyl CpG-binding proteins (MeCP1, MeCP2, MBDs and Kaiso) [1621] exert their function as transcriptional repressors via chromatin modification. Methyl CpG-binding proteins are often part of large repressor complexes as NuRD, NoRC, mSin3A and SWI-SNF. Repressor complexes recruit histone deacetylases (HDACs) and histone methyltransferases (HMTs) on methylated target promoter sequences. These enzymes catalyse covalent post-translational modifications of specific residues on histone 3 (H3) and 4 (H4) N-terminal tails (e.g. deacetylation of lysine (K) 9, demethylation of K4 and methylation of K9 and K27 of H3), inducing a compacted transcriptionally inactive chromatin structure. Histone acetylation status also depends on the contrasting activities of HDACs and histone acetyltransferases (HAT) group of enzymes. The latter are presumed to induce histone tail modifications (e.g. acetylation of K9 and K14 of H3), resulting in a transcriptionally active chromatin state. As for histone acetylation, histone lysine methylation can be dynamically regulated by the recruitment of members of the histone lysine methyltransferases and demethylase class of enzymes, which impose memory on gene transcription [22]. Other histone tail modifications include phosphorylation, sumoylation, ubiquitination and ADP ribosylation. Overall, DNA methylation and histone modifications work together to assemble a chromatin structure, which dynamically shifts from a transcriptionally permissive state to a transcriptionally inactive state and vice versa [2, 14].

Inhibition of HDACs can be achieved in normal conditions by endogenous molecules [23] explaining the plausibility of this process in the normal regulation of gene expression.

Aberrant DNA methylation and chromatin modifications, altering gene transcription states, are common hallmarks of human tumour cells [24]. Studies on leukaemias have provided paradigmatic examples for the functional implications of genetic and epigenetic alterations in cancer development [25, 26]. These studies underline the possibility of reversing disease-associated aberrant epigenetic states by targeting the catalytic activities of chromatin remodelling enzymes. Thus, these enzymes are attractive targets for therapeutic intervention in cancer [2729]. The possibility of drug development in this field has recently been reviewed [22, 24, 30, 31].

Evidence is growing that non-coding RNAs (ncRNAs) are involved in inducing chromatin modifications and act as additional molecular determinants for epigenetic regulation of gene expression also in human cells [3234] (Fig. 1b). NcRNAs comprise a large and heterogeneous family of RNA molecules differing in length (short, such as microRNAs, and long ncRNAs), which are transcribed from DNA but not translated into proteins. By regulating gene expression at the transcriptional and post-transcriptional level, they affect a broad range of physiologic functions and pathologies such as neoplastic diseases [35, 36]. Both short and long ncRNAs appear to function by guiding the recruitment HDACs, HMTs and DNMTs, and other proteins involved in the epigenetic regulation of transcription, to homology-containing loci on gene promoters and in the genome. Moreover, short ncRNA, as microRNAs and siRNAs, can repress the expression of HDAC, DNMTs and other components of chromatin-modifying complexes at the post-transcriptional level by interacting with their mRNAs [32, 33, 35] (Fig. 1b). Therefore, ncRNAs play direct roles in DNA methylation, heterochromatin formation, histone modification and gene silencing. In turn, they are epigenetically targeted for repression or activation; this can be a valuable way of amplifying changes in the levels of downstream effectors. Knowledge of these emerging regulatory roles of ncRNAs has implication not only in cellular physiology and pathology but also for the development of novel epigenetic drugs that re-establish the correct pattern of gene expression in complex diseases such as cancer.

Currently in the clinical setting, there are two classes of epigenetic drugs, which act through the inhibition of the enzymatic activities responsible for epigenetic transcriptional silencing: DNMTs and HDACs (Fig. 2). DNA methylation inhibitors 5-azacytidine (azacytidine) and 5-aza-2′-deoxycytidine (decitabine) have been approved by the US Food and Drug Administration (FDA) in 2004 and 2006, respectively, for the treatment of patients with myelodysplastic syndromes (MDS). US FDA approved the HDAC inhibitors suberoylanilide hydroxamic acid (SAHA, vorinostat, in 2006) and romidepsin (depsipeptide, in 2009) for the treatment of patients with progressive, persistent or recurrent cutaneous T-cell lymphoma [37]. In 2015, FDA approved panobinostat in combination with bortezomib and dexamethasone for the treatment of patients with multiple myeloma [38] and belinostat for the treatment of patients with peripheral T-cell lymphoma (PTCL) [39].
Fig. 2
Fig. 2

Chemical structures of different classes of DNMT and HDAC inhibitors. Antimetabolites 5-azacytidine and decitabine (5-aza-2′-deoxycytidine) are cytidine analogues; these nucleoside derivatives are incorporated into DNA leading to covalent adduct formation, thus acting as mechanistic inhibitors. The non-nucleoside DNMT inhibitor hydralazine interacts within the binding pocket of the enzyme interfering with the DNA methylation mechanism

The success of epigenetic therapies in inducing clinical responses in MDS and lymphoma not only gave to this kind of treatment high visibility, but it also suggested that similar results might be obtained in solid tumours and that this line of research deserves proper evaluation. The relevance of epigenetic treatment in haematological malignancies (leukaemias, lymphomas, myelodysplastic syndromes, myeloma) have already been described in detail [40].

In clinical studies, epigenetic treatment was administered alone or in combination with standard anticancer therapies (usually chemotherapy, sometimes radiotherapy) to improve their antitumour activities [41]. In some cases, the aim was more directly related to the control of gene activity and to prevent the development of resistance due to the overexpression of a specific gene, for example thymidylate synthase [42].

In several papers [43, 44], particularly those more recently published [45, 46], a detailed explanation for the combination of epigenetic and standard anticancer treatment is provided together with complete and stimulating results.

The present review will focus on the epigenetic treatment of solid tumours: we collected data from clinical studies available as full papers, data on ongoing studies are available at site and have been reviewed [47].

In the text, we discuss data on the pharmacology, pharmacokinetics (PK) and pharmacodynamics (PD), on the toxic side effects and on the antitumour activity of epigenetic treatments. In the tables, we give more details for epigenetic treatment alone (Table 1) or in combination with traditional anticancer agents or radiotherapy (Table 2).
Table 1

Studies on epigenetic treatment alone

Drug (s) and schedule

Study type - histology (number of patients)

Results and data provided


Abexinostat (S78454/PCI-24781)

p.o. 60 mg/m2 bid 4 days on/3 days off

Mixed tumours (15 pts)

PK/PD model predicts thrombocytopenia


Azacytidine + Valproate

Aza s.c. for 10 days q. 28 days.

MTD 75 mg/m2/day

VPA in plasma 75–100 μg/ml (a)

Dose escalation

Mixed tumours (55 pts)

PBMC: DNA methylation decreased. H3 acetylation increased. Patients with stable disease had more H3 acetylation.

DLT: neutropenic fever and thrombocytopenia


Azacytidine +Entinostat

Aza 30–40 mg/m2/day for days 1–6 and 8–10 q 28 days

Entinostat 7 mg/m2 days 3 and 10

Phase I/II trial

NSCLC (45 pts)

Demethylation of 4 epigenetically silenced genes (CDK2a, CDH13, APC, RASSF1a). In plasma DNA was associated with improved progression-free and overall survival


Azacytidine (AC) Phenylbutyrate (PHB)

AC 10-25mg/m2/day for 21 days,

75 mg/m2/day for 7 days

PHB 200–400 mg/m2 days 6, 13 and 20

Phase I

Mixed tumours (27 pts)

Toxicity: neutropenia, anaemia. No PK interaction

“No conclusive statement can be made on histone acetylation or methyltrasferase activity”.


Decitabine + VPA

Dec 5–15 mg/m2/day for 10 days.

VPA 10–20 mg/kg/day for days 5–21 q 28 days

Phase I

NSCLC (8 pts)

Neurological toxicity.

Increase in foetal Hb levels in all pts



1000 mg/m2/day i.v. for days 1–5 q 21 days

Phase II

Ovarian: platinum resistant (18 pts) or micropapillary (14 pts)

Toxicity: thrombosis (3 pts).

Increased H3 acetylation in PBMC and in two tumours



1000 mg/m2/day i.v. for days 1–5 q 21 days

Phase II

Refractory Thymic epithelial tumours (41 pts)

Nausea, vomiting, fatigue

Modest activity

Protein acetylation did not predict outcome



600–1400 mg/m2/day i.v. for days 1–5 q 3 weeks .

Phase I/II

Hepatocarcinoma (60 pts)

PK linear. MTD not reached at 1400 mg/m2.

Toxicity: abdominal pain, liver toxicity, vomiting.

Plasma concentrations higher than effective in vitro levels for 4 hours.

Disease stabilisation. High HR23B associated with more stabilisation.



1000 mg/m2/day i.v. for days 1–5 q 21 days

Mesothelioma (pre-treated) (13 pts)

Not active in terms of RR

toxicitiy: nausea, emesis, fatigue and constipation. One fatal cardiac arrhythmia


Belinostat, (oral formulation)

150–1000 mg/m2/day for days 1–5 q 21 days

Pharmacological evaluation

Mixed tumours (46 pts)

PK and PD: results similar to the parenteral formulation



5–160 mg/day p. o.

RD 40 mg/day p.o.

Phase I

Mixed tumours (39 pts)

DLT: thrombocytopenia, fatigue, atrial fibrillation, ECG alterations, elevated creatinine.

AUC proportional to dose, plasma concentration sufficient for preclinical antitumour activity

Effect on histone acetylation in PBMC



2–8 mg/m2/day

RD 8 mg/m2/day for 8 weeks q 10 weeks

Phase I

Mixed tumours (53 pts)

Toxicity: Thrombocytopenia (DLT).

PK data.



12.5-56 mg/m2/day p.o.

3 times/week for 2 weeks q 3 weeks

RD 45 mg/m2/day

Phase I

Mixed tumours

Inhibition of HDAC activity and induction of acetylation of H3 histones in peripheral WBCs



MTD 10 mg/m2 q14 days

Phase I

Mixed tumours (31 pts)

Toxicities: nausea, vomiting, anorexia, fatigue.

Half-life 39–80 hrs (longer than expected). Linear PK.

Increased H3 acetylation in PBMC. Peak plasma levels higher than effective in vitro concentration.



20 mg p.o. twice/week

Pharmacological study

Mixed tumours (36 pts)

No effect of food on PK parameters



40 mg p.o. three times/week

Sarcoma (47 pts)

Ovarian Sex Cord Tumours (5 pts)

Poorly tolerated. No activity in sarcoma. Activity in OSCT

Toxicity: thrombocytopenia, fatigue, anaemia



20 mg/m2 for days 1 and 8 q 21 days

Prostate (35 pts)

No clinical activity

Toxicity: fatigue, thrombocytopenia, nausea



20 mg

Mixed tumours (4 pts)

PK determined by trace radiolabelled 14C excretion

Rapid oral absorption, liver and renal excretion



2.34 g/m2/day in 6 h for 3 days q 21 days

Phase II

NSCLC (47 pts)

Toxicity: fatigue, nausea, dysgeusia-

3 partial responses (6%)


Quisinostat (JNJ-26481585)

Mixed tumours (92 pts)

Phase I

RD 12 mg days 1,3 and 5

Toxicity: cardiovascular, fatigue, nausea

PD: increased H3Ac in hair follicles, skin and tumour.



RD 600 mg/day p.o. for 5 days q 14 days

Phase I

Head-and-neck refractory

Toxicity: nausea, vomiting, fatigue.

PK data, HDAC inhibition, H4Ac increase in PBMC



13 mg/m2 i.v. in 4 h for days 1, 8 and 15 q 28 days

Phase II

Refractory Prostate (35 pts)

Toxicity: nausea, fatigue, vomiting and anorexia

No antitumour activity



13 mg/m2 in 4 h, for days 1, 8 and 15 q 28 days

Phase II

Head and Neck (14 pts)

Toxicity: nausea, vomiting, constipation, fatigue

H3 hyperacetylation in PBMC

Reduced or stable Ki67

On microarray 641 differentially expressed genes

No consistent change ion methylation of specific genes

Upregulation of p21Waf1/Cip1.



New schedule: 1–9 mg/m2 in 4 h

for days 1, 3, and 5 q 21 days

RD 7 mg/m2

Phase I

Mixed tumours (28 pts)

Increase in 3HAc in PBMC. PK data described.

Toxicity: ECG changes



400 mg/day p.o.

Phase II

Head-and-neck, refractory (13 pts)

No response. Toxicity: anaemia, anorexia, hyperglycemia, thrombocytopenia, dehydration



400 mg/day p.o.

Phase II

Refractory Prostate (27 pts)

IL-6 was higher in patients with toxicity (Fatigue, nausea)



400 mg for 14 days q 21 days

Phase II

Glioblastoma (66 pts)

Analysis of tumour tissue. Increased Acetylation of H2A, H3, H4. up-regulation of e-regulin.

PK influenced by enzyme-inducing drugs.

Toxicity: fatigue, thrombocytopenia. nausea, diarrhoea



400 mg for 14 days q 21 days

Phase II

Breast (14 pts)

No antitumour activity.

Toxicity: Fatigue, nausea, diarrhoea, and lymphopenia



400 mg for 14 days q 21 days

Phase II

Ovarian (27 pts)

No antitumour activity

Toxicity: Neutropenia, Leukopenia, Thrombocytopenia, Constitutional, Gastrointestinal, Metabolic



400 mg for 14 days q 21 days

Phase II

NSCLC second line (16 pts)

No antitumour activity

Toxicity: fatigue, dehydration, hyperglycemia, mild




400–800 mg for 14 days q 21 days

RD 400 mg for 14 days q 21 days

Breast, colorectal, NSCLC (16 pts)

No antitumour activity. Toxicity anorexia, asthenia, nausea, thrombocytopenia, vomiting, weight loss



400 mg for 14 days q 21 days

Thyroid (19 pts)

No antitumour activity

Toxicity: fatigue, dehydration, ataxia, pneumonia, bruises, thrombosis, thrombocytopenia



600 mg bid days 1–3 q 7 days or 400 mg for 14 days q 21days

Phase I

Gastrointestinal (16 pts)

DLT thrombocytopenia.

Some PK data: AUC μM/h 7.75±2.79 for 400 mg; 3.94±1.56 with 300 mg.

t ½ 1.05±0.32 – 1.49±0.82 hours



100–500 mg once or twice daily for 14 days q 21 days

Phase I

Mixed tumours (18 pts)

MTD not reached. Recommended dose 500 for once, 200 for twice daily.

Some PK data: AUC linear with dose



300 or 400 mg bid days 1–3 q 7 days

Mesothelioma (pretreated) (13 pts)

2 PR.

Toxicity: fatigue, anorexia, dehydration, diarrhea,

nausea, and vomiting



2 h i.v. infusion

75–900 mg/m2/day days 1–3 q 21 days

or 300–900 mg/m2/day days 1–5 q 21 days

Phase I

Mixed tumours (37 pts)

Toxicity: myelotoxicity, fatigue, anorexia, hyperglicemia

Increase in acetylated histones in PBMC and in tumour cells. PK data.


SAHA oral

MTD 400 mg/day or 600 mg/day days 1–3 q 7 days

Phase I

Mixed tumours (73 pts)

Toxicity: anorexia, dehydration, diarrhea, and fatigue. In PBMC acetylation increased 2 hrs after dose, back to basal levels at 8 hours



300 mg tid

Breast (25 pts)

Decrease of proliferation-associated genes.

No effect on methylation



400 mg daily continuously

Melanoma (39 pts)

Toxicity fatigue, nausea, lymphopenia, and hyperglycemia.

Some biochemical correlative data presented.



300 mg tid days 1–3, 8–10, 15–17 q 21 days

Mesothelioma (pretreated) (329 pts)

Randomised phase III: no benefit

Toxicity: fatigue or malaise



10–80 mg/day p. o. 3 times/week for 3 weeks q 4 weeks

RD 60 mg/day

Phase I

Mixed tumours (30 pts)

DLT: fatigue, hypokalemia, ECG alterations.

AUC proportional to dose.

HDAC increases at doses 60 mg.



10–90 mg daily five times a week q 2 wks

RD 60 mg/day

Phase I

Mixed tumours (38 pts)

PK data. No correlation of AcH3 and response.

Toxicity: fatigue, nausea, vomiting.



intravenous infusion in 1 h

30-250 mg/kg/day for days 1–5 q 21 days

RD 60 mg/kg/day

Phase I

Mixed tumours (26 pts)

Toxicity: neurological.

HDAC2 decreased; H3 Acetylation increased;

VPA plasma levels 0.3-0.9 mM.



p. o. 20-40mg/kg/day for 5 days

Phase I

Cervical cancer (12 pts)

VPA in plasma 73–170 μg/ml. (0.4-1 mM)

No correlation of H3 acetylation in tumour biopsies and plasma VPA. Toxicity: Depressed consciousness



500 mg p. o. tid (target concentration 50–100 μg/ml)

(0.3-0.6 mM)

Phase II

Low-grade Neuroendocrine (8 pts)

Two tumours had a 2-4-fold increase in Notch-1 mRNA, 3 had a decrease.


The references are included at the end of the text

5FU 5-Fluorouracil, 5mC 5-methyl Cytosine, AUC area under the curve (also a dosing calculation for Carboplatin), Bid bis in die (twice a day), DLT dose-limiting toxicity, FEC combination of Fluorouracil, Epirubicin, Cyclophosphamide, FolFOx combination chemotherapy of Folinic acid, 5-Fluorouracil and Oxaliplatin, GI gastrointestinal, i.v. intravenously, MTD maximum tolerated dose, NSCLC non-small cell lung cancer, PBMC peripheral blood mononuclear cells, PD pharmacodynamic, PFS progression-free survival, PK pharmacokinetics, p.o. per os (orally), PR partial response, Pt patient, q every (Latin “quaque”), RA rapid acetylator (Hydralazyne metabolism), RD recommended dose, RR response rate, SA slow acetylator (Hydralazyne metabolism), SAHA Vorinostat, Zolinza ®, TS thymidylate Synthetase, target enzyme for 5FU activity, VPA Valproic Acid, WBC white blood cells

(1) Oral dose of VPA titrated in each patient to obtain adequate plasma concentrations.

Table 2

Epigenetic treatment associated with a conventional anticancer agent

Epigenetic drug

Tumour type and chemo

Results and data provided



RD 75 mg/m2/day

days 1–4 and 15–18 q 28 days

Phase I

Erlotinib 150 mg/day

Mixed tumours (30 pts)

Aza: increasing dose (75–100) and days of treatment (2–8)

Toxicity: rash, diarrhoea, nausea, and fatigue



Ovarian cancer, platinum insensitive (30 pts)

Toxicity: fatigue, myelosuppression

DR4 methylation in PBMC related to activity



75 mg/m2/day for days 1-5

Prostate cancer (22 pts)

Docetaxel (day6) , prednisone

Toxicity: myelosuppression

Reduction in GADD-45 methylation (peripheral DNA) on day 5

Only pts that had a reduction had a response.



15–45 mg tid days 1-5

Sarcoma 22 pts

Doxorubicin 75 mg/m2 day 4

Neutropenia (growth factors required), fatigue, thrombocytopenia, and anemia.

PK of Abexinostat described. HDAC levels inhibited in PBMC



1000 mg/m2/day for 5 days

Carboplatin AUC 5 day 3

Resistant ovarian cancer (29 pts)

Toxicity: neutropenia, thrombocytopenia, vomiting

No effect, study closed



1000 mg/m2, 48 h c.i.

Thymic epithelial (26 pts)

Cisplatin, doxorubicin, cyclophosphamide

Toxicity: nausea, diarrhea, neutropenia, thrombocytopenia,

Immunomodulatory effect observed


Belinostat 1000 mg/m2 i.v. for days 1–3 then p.o. 2000 mg for days 4-5

Unknown primary (44 pts)

Paclitaxel, carboplatin

Randomised phase II. No clinical benefit



6 mg/m2/day for days 1–21, 28-day cycle

Phase II Pancreas.

Gemcitabine 1000 mg/m2 days 1, 8 and 15 (174 pts)

Increased incidence of neutropenia and thrombocytopenia

No improvement of gemcitabine activity



4–10 mg/m2/day

RD 6 mg/m2/day for days 1–14 q 21 days

Phase I

Mixed tumours (54pts)

Capecitabine 1650–2000 mg/m2/day for 14 days q 21 days

PK not altered by capecitabine.

Toxicity: Thrombocytopenia



45–135 mg/m2 6 h infusion for day 1

RD 90mg/m2

Phase I

Carboplatin (AUC 5 or 6) day 8

Mixed tumours (33 pts)

Dose dependent, reversible demethylation in PBMC maximally at day 10. Demethylation of the MAGE1A gene

Toxicity: myelosuppression



10 mg/m2/day for 5 days

Carboplatin AUC 5 day 8

Ovarian cancer (17 pts)

35% RR 10.2 months PFS

In PBMC and tumours global and gene-specific demethylation.

Demethylation of MLH1, RASSF1A, HOXA10, HOXA11 correlated with PFS



0.15 mg/kg i.v. daily × 5 days/wk for 2 wks

Phase I-II


p.o. 75 mg/m2 daily for weeks 2–5 of a 6-week cycle

Refractory Melanoma (35 pts)

Toxicity: mainly haematological

No effect on promoters of DNA repair genes

Excellent PK and PD data (also in tumours)



days 1–5 q 28 days

10–20 mg/m2/day

Phase I

Ovarian, recurrent (10 pts)

Carboplatin AUC 5 day 5

Toxicity: myelosuppression, nausea, fatigue

global and gene-specific DNA methylation


Decitabine i.v. day 1

45 mg/m2

Carboplatin AUC 6 day 8

Ovarian cancer (15 pts)

Patients with methylated hMLH1 tumour DNA in plasma

Decitabine appears to reduce the efficacy of carboplatino

Decrease in global levels of methylation with Decitabine.



01–0.2 mg/kg 3 days weekly


10-30mg q 4 days

Temozolomide 150–200 mg/m2/day

Resistant melanoma

Toxicity: myelosuppression, fatigue, nausea

No antitumour acitivity.



10 mg p.o. day 1 and 15 q 28 days

Randomised phase II

NSCLC (132 pts)

Erlotinib 150 mg/day

Toxicity: rash, fatigue, diarrhoea, nausea

High E-caderin levels associated with longer PFS



1–5 mg/kg days 1,8,15 q 28 days

Phase I

Mixed tumours (19pts)

13-cis retinoic acid 1 mg/kg

Toxicity: hyponatremia, neutropenia, anaemia.

PD: Increased histone acetylation



5 mg/week

Breast (64pts)

Examestane 25 mg/day

Randomised phase II. Patients had progressed with AI.

Protein lysine hyperacetylation associated with prolonged PFS



10 mg/2 weeks

Phase I

Mixed tumours (31 pts)

Sorafenib (400 mg tid)

Toxicity: Handfoot syndrome, nausea/vomiting, and fatigue


Hydralazine (182 mg RA; 83 mg SA)

+ Valproate (40 mg)

Phase II (17 pts)

Mixed tumours: re-treatment of resistant patients with same chemo as before

Toxicity: mainly haematological

Reduction in global DNA methylation, histone deacetylase

activity, and promoter demethylation



(182 mg RA; 83 mg SA)

+ Valproate (30 mg)

Phase II

Progressive Cervical cancer (36 pts)

Cisplatin + Topotecan

Advantage in PFS (10 vs. 6 months)

Molecular correlates pending.



(182 mg RA; 83 mg SA)

+ Valproate (30 mg)

Phase II

Breast (16 pts)

Doxorubicin, cyclophosphamide

Decrease in 5mC content and HDAC activity.

Up- and down-regulation of many genes.



alone: 20 mg for days 1,3 and 5 for 2 weeks q 3weeks

with chemo: 15 mg

Prostate (pretreated) (16 pts)

Docetaxel 75 mg/m2 q 21 days

Toxicity: dyspnea, neutropenia

Increase in histone acetylation in PBMC

No relevant antitumour actitvity



30 mg/day, days 1,3 and 5 q 14 days

Recurrent glioma (12 pts)

Bevacizumab 10 mg/kg q 14 days

Toxicity: thrombocytopenia, hypophosphatemia, hemorrhage, thrombosis.



10 mg days 1,3 and 5 q 14 days

Phase I

Mixed tumours (12 pts)

Bevacizumab 10 mg/kg q 14 days

Everolimus 5 or 10 mg

Toxicity: Mucositis, arrhythmia.

No consistent change in HDAC activity in PBMC



30 mg days 1 and 4


Erlotinib 100 mg/day

DLT: cardiac, nausea. Fatigue.

PK and PD data.



20 mg

Gleevec-resistant GIST (12 pts)

No actibvity but evidence of 3HAc increase in PBMC


Panobinostat 10 mg days 1, 3 and 5

Paclitaxel, Carbopaltin AUC=5

Phase I

Miscellaneous tumours (12 pts)

Toxicity: diarrhea, fatigue, and vomiting




RD 300 mg

GI carcinoma (14 pts)


30 Gy in 3 Gy/day over 2 weeks

Toxicity: fatigue

Diarrhoea proportional to the volume of intestine irradiated.

[41, 138]


200–800 mg/day 1week q 2 weeks

RD 600 mg/day

Refractory colorectal

FolFOx (21 pts)

Toxicity: fatigue, anorexia, dehydration

no consistent modulation of TS expression



days 1–3 q 14 days 600–2000 mg/day

RD 1700 mg once 600 mg tid

Phase I

Refractory colorectal (43 pts)


toxicity: neutropenia, thrombocytopenia, fatigue, nausea or vomiting, anorexia, mucositis.

No consistent effect on biopsies.



400 mg/day

Tamoxifen (43 pts)

Hormone-resistant breast

Histone hyperacetylation and higher baseline HDAC2 levels that correlated with response



RD 400 mg/day 14 days q 21 days

600 mg/day bid q 21days

Phase I (28 pts)

Mixed tumours

Paclitaxel (200 mg/m2) (Carboplatin (AUC 6)

Toxicity: emesis, neutropaenia, fatigue



400 mg/day

Randomised Phase II

vs. placebo

NSCLC 94 pts

Paclitaxel Carboplatin

Toxicity thrombocytopenia, nausea, emesis, fatigue.

RR 34% vs. 12%



400 mg/day

Phase I-II

resistant colorectal


Failed to establish an MTD

Toxicity: fatigue, thrombocytopenia and mucositis.

Intratumoral TS downregulation in one patient. Acetylation of H3 in PBMCs



200–300 mg bid days 1–3 q 7 days

Phase I-II

Breast (54 untreated pts)

Paclitaxel 90 mg/m2/week Bevacizumab 10 mg/kg

Increased diarrhoea with the addition of SAHA

Increased acetylation of Hsp90 and α-tubulin



400 mg/day 14 days

Phase II

Glioblastoma (37 pts)

Bortezomib 1.3 mg/m2/day days 1,4,8 and 11 q 21days

Toxicity: Fatigue.

No therapeutic advantage



100–200 mg/day for days 1–14 q 21 days

Phase I

Mixed tumours

12 patients

Docetaxel 50–75 mg/m2 day 4 q 21 days

Excessive toxicity: neutropenic fever, cardiac, bleeding

No PK interaction



for days 1–2 400–100 mg/day

RD 800 mg/day

Phase I (32 pts)

Mixed tumours

Doxorubicin on day 3

20 mg/m2 weekly

Toxicity: fatigue, nausea

HDAC2 expression in PBMC similar to tumours

no correlation of SAHA levels with acetylation



300 mg/day for 16 days q 28 days

Phase I

Mixed tumours (22pts)


0.15-0.7 mg/m2 i.v.

days 1, 8 and 15 q 28 days

Toxicity: Fatigue, nausea, diarrhea, vomiting,

PK data. Data on proteasome inhibition in PBMC



300 mg for days 1–3 q 8 days


1.3 mg/m2 days1,8 and 15 q 21 days

NSCLC (21 pts)

Preoperative treatment.

Toxicity: fatigue and hypophosphatemia



200–300 mg tid for days 1–4 and 8-11

Bortezomib 1–1.3 mg/m2 for day 9

Phase I (60 pts)

Comparison in PBMC and biopsies after SAHA and SAHA-Bort. Dcreased Nur77 expression.



400 mg p.o.

for days 1–7 and 15–21 q 28 days

NSCLC (33 pts) Erlotinib-resistant

Erlotinib 150 mg/day

No clinical activity

Toxicity: anemia, fatigue and diarrhoea.



300–400 mg/day for days 1-14

Gastric (30 pts)

Capecitabine, Cisplatin

Toxicity: thrombocytopenia, fatigue, stomatitis, anorexia

H3Ac correlated with SAHA dose



200–400 mg p.o. for days 1–14 q 21 days

Mixed tumours (35 pts)

Sorafenib 400 mg p.o. bid

Recommended dose for SAHA 300 mg/die, but not tolerated.

Toxicity: hand-foot syndrome. No tumour response.



tid for days 1–4 and 8–11 q 21 days

Mixed tumours (29 pts)

Bortezomib 1.3 mg/m2 i.v. for days 1, 4, 8 and 11

Toxicity thrombocytopenia, fatigue, increased ALT, elevated INR, and diarrhea.

PK data provided.



p.o. for days 1–14

MTD 400 mg

Mixed tumours (23 pts)

Bortezomib i.v. for days 1, 4, 8 and 11 q 21 days.

MTD 1.3 mg/m2

Toxicity: fatigue, hyponatremia, nausea, anorexia

Some PK data



300 mg daily

Mixed tumours (78 pts)

Pazopanib 600 mg daily

Toxicity: thrombocytopenia, neutropenia, fatigue, hypertension, diarrhea, vomiting


SAHA 400 mg daily

Gefitinib 250 mg

NSCLC pretreated (52 pts)

No clinical benefit

Toxicity: anorexia, diarrhea, fatigue, anemia



30–90 mg/kg/day

for days 1–5 q 21 days

MTD 75 mg/kg/day

Karenitecin i.v. 0.8-1 mg/m2/day

for days 3–7 q 21 days

Melanoma: xenografts

Phase I-II (39 pts)

Toxicity: somnolence, fatigue

VPA levels at MTD 1.28 mMol

Histone hyperacetylation was observed in PBMC. No effect of valproate on Karenitecin PK



15–160 mg/kg/day for days 1–3

RD 120 mg/kg/day

Phase I (44 pts)

Mostly breast

FEC day 3

Toxicity: somnolence, myelosuppression

Histone acetylation in tumour samples and in PBMCs correlated with valproic acid levels and was further linked to baseline HDAC2 but not to HDAC6 expression



10–90 mg/kg/day

Melanoma (32 pts)

Dacarbazine 800 mg/m2 q 21 days,

interferon-α 600.000 IU twice daily

Toxicity: neurological, myelosuppression

Acetylation in PBMC measured.

“casting some doubts on the clinical use of VPA in this setting”.



Dose escalated to obtain active plasma concentration

Mesothelioma resistant to cisplatin (45 pts)

Doxorubicin 60 mg/m2 q 21 days

Toxicity: myelosuppression

16% partial response rate



15–160 mg/kg/day for days 1–3

MTD 140

Phase I

Mixed tumours (48pts)

Epirubicin 100 mg/m2 for day 3

Toxicity: somnolence, confusion, neutropenia

VPA levels correlate with acetylation in PBMC

Plasma VPA higher than in vitro effective concentrations


References are included at the end of the text

A.I. aromatase inhibitor, 5FU 5-Fluorouracil, 5mC 5-methyl Cytosine, AUC area under the curve (also a dosing calculation for Carboplatin), Bid bis in die (twice a day), DLT dose-limiting toxicity, FEC combination of Fluorouracil, Epirubicin, Cyclophosphamide, FolFOx combination chemotherapy of Folinic acid, 5-Fluorouracil and Oxaliplatin, GI gastrointestinal, GIST gastrointestinal stromal tumour, HNC head-and-neck carcinoma, i.v. intravenously, MTD maximum tolerated dose, NSCLC non-small cell lung cancer, PBMC peripheral blood mononuclear cells, PD pharmacodynamic, PFS progression-free survival, PK pharmacokinetics, p.o. per os (orally), PR partial response, Pt patient, q every (Latin “quaque”), RA rapid acetylator (Hydralazyne metabolism), RD recommended dose, RR response rate, SA slow acetylator (Hydralazyne metabolism), SAHA Vorinostat, Zolinza ®, TS Thymidylate Synthetase, target enzyme for 5FU activity, VPA Valproic Acid, WBC white blood cells

Pharmacological aspects

An effective treatment, based on a sound scientific rationale, requires the identification of the relevant target(s) and the demonstration that the target(s) can be inhibited without excessive toxicity, and that the duration of this effect is sufficient to interfere with cell growth.

In the case of epigenetic treatment, it is necessary to prove that one of the different mechanisms that finely regulate gene expression (see above) is altered. Even if this mechanism is well established for many molecules, for some compounds such as CI-994 (N-acetyl-dinaline), an inhibitor of class I HDACs, the actual relevance of an epigenetic mode of action remains uncertain [27, 48, 49].

The list of epigenetic drugs is becoming longer every day and it also includes “old” drugs: Hydralazine, for example, has long been used as an anti-hypertensive agent, but it has recently been recognised as a demethylating agent [50, 51]. This also happens for HDAC inhibitors that include old molecules (such as valproic acid (VPA) [52] and a dozen compounds that have been recently synthesised [53].


To obtain an authentic effect on epigenetic mechanisms, the choice of the dose and schedule of administration is crucial since, especially for demethylating agents, if the drug concentration is too high this may result in cell toxicity or in a “traditional” antiproliferative effect. Furthermore, when a combination is used, the epigenetic effect must persist during chemotherapy (and possibly also later) in order to obtain an adequate synergism. This concept is guiding recent studies on new schedules of drug administration, as is the case for romidepsin [54].

Detailed data on pharmacokinetics have been published for several drugs (see Tables 1 and 2 for a complete list). The pharmacology of demethylating agents has been originally described in the past, and recent analyses mostly concern their interaction with other agents used in combination, such as temozolomide [55].

The pharmacology of HDAC inhibitors is particularly complex since many of these compounds act as enzyme inducers and may therefore modify their own kinetics when repeated dosing is used, or the pharmacology of associated drugs.

Drug concentrations in plasma are generally low, and a very sensitive assay, such as HPLC coupled to mass spectrometry, is required to obtain reliable data [56].

Individual characteristics may influence the kinetic parameters (pharmacogenetics): in the case of hydralazine, fast or slow metabolism is genetically determined, and in some studies, doses have been escalated or de-escalated according to individual metabolic parameters [5759].

Pharmacodynamic effect

The most interesting part of the evaluation of anticancer drugs, particularly when dealing with an innovative mechanism of action, is the study of their effect in tumour cells. Several technical problems arise when trying to quantify the effect of epigenetic drugs, especially in solid tumours: it is difficult to decide what to measure, when and where.

Concerning demethylating agents, the effect is generally measured by evaluating global DNA methylation, but some authors determined the methylation status of specific genes that had previously been selected [6062] and the level of expression of foetal haemoglobin has also been used as a PD marker [63]. The subject of DNA methylation may also be related to drug resistance caused by the activity of O(6)-methylguanine DNA methyltransferase (MGMT) that has been described in cerebral tumours [3, 64]. Decitabine has been used in an attempt to reduce methylation of genes involved in DNA repair in melanoma patients treated with temozolomide [55] but no such effect was evident even if decitabine caused hypomethylation of the HbF gene promoter.

The situation is more complex for HDAC inhibitors since not all drugs inhibit the different enzyme classes that are present in eukaryotic cells to the same extent. Some molecules have a wide inhibiting effect [65]; some are more restricted, and class-specific inhibitors such as CHR-3996, specific for class I HDACs enzymes [45], are being introduced in the clinics. Furthermore, the inhibition of de-acetylating enzymes may result in acetylation not only of histones, but also of other proteins, such as tubulin and Hsp90 that are involved in anticancer drug activity/resistance or in unrelated cellular pathways [66, 67].

An additional difficulty derives from the definition of a cut-off value: some authors, for example, required a doubling of histone acetylation to consider a result as “relevant” [68] but this was not mandatory in many other studies.

The effect of HDAC inhibitors was generally determined in terms of enzymatic activity, but in some cases histone acetylation, particularly H3 and H4, has been used as marker of activity. A consistent increase of H3 acetylation in peripheral blood mononuclear cells (PBMC) at effective doses has been observed with several agents, even if large inter-patient variations were often reported [69], and, more importantly, intratumoral H3 acetylation increase did not always correlate with response [70].

The effect of epigenetic treatment has also been evaluated by looking at specific genes in terms of expression [57, 62] or re-activation [71]. It is becoming clear that many elements are involved, and that it may be difficult to identify a consistent pattern in gene activation-inactivation [72] to be used as a marker of epigenetic activity.

The gold standard of pharmacodynamic studies is the evaluation of the effects in tumours. The study of malignant cells is certainly more complicated in patients with solid tumours than in leukaemia since cancer cells are more difficult to obtain, especially at multiple time points. Some reports on the evaluation of epigenetic therapy in solid tumour tissues have been published and deserve special consideration. The number of evaluable samples was often low, but informative results were obtained and reported [41, 73]. In ovarian cancer, it has been possible to evaluate the activity of demethylating agents on cells obtained from ascitic fluid and a gene-specific reduction of DNA methylation was evident [74]. Serial biopsies were obtained from patients with in head-and-neck carcinomas [70], glioblastoma [75], cervical and breast cancer and other tumours [57, 58, 66].

To overcome the difficulty in obtaining tumour samples, several groups have focused on the identification of surrogate markers. PBMC represent the most commonly used alternative. It is possible to measure HDAC activity and histone acetylation or DNA methylation. PBMC have also been used, in studies of drug combinations, to measure the target of non-epigenetic drug [76]. When PBMC and tumour biopsies were compared, however, results were not always consistent [66]. PBMC represent a “surrogate” tissue, and further improvement is required in order to make results obtained in these cells more representative of what is actually taking place in cancer.

A promising technique is the evaluation of circulating cell-free DNA [77] on which several specific analyses to identify epigenetic modifications can be performed [78]. The activity of demethylating agents has been evaluated by measuring the methylation status of circulating cell-free DNA in plasma in patients with refractory advanced non-small cell lung cancer (NSCLC) [62]. This is an interesting alternative with applications in many aspects of medicine, but further studies are required before it can be considered a reliable marker of epigenetic activity.

To obtain an adequate effect, sufficient drug concentrations must persist in the target cells for an adequate time. If there is a relationship between plasma kinetics and tissue effect (PK/PD relationship), drug dosing may be adapted on the basis of PK parameters that are easier to obtain. The permanence of a sufficient drug concentration or of a measurable effect has been evaluated in different tumour types and provides a rationale for the antitumour activity observed (see Tables 1 and 2 for details). Concerning the PK/PD relationship, data are difficult to interpret: no correspondence was found between plasma levels of valproate and histone acetylation in cervical cancer [79] so that PD assays may be required until we can devise more efficient PK models.

Toxic side effects

One of the most interesting characteristics of many epigenetic drugs is that toxicity, at doses sufficient to achieve effective plasma concentrations, is generally very mild and has been described in detail in many studies of epigenetic agents used alone or in combination with standard anticancer agents (see Tables 1 and 2 for details).

Toxicity of demethylating agents

The best known agents that interfere with DNA methylation are decitabine and 5-azacytidine. More limited data are available for zebularine [46]. Since these agents also have traditional antiproliferative activity, the dose used plays a key role. At high doses, 5-azacytidine can cause neutropenia [68], similar to what is described for traditional anticancer drugs. It is generally assumed that in order to exploit the epigenetic action, it is necessary to use very low doses that are insufficient to cause any antiproliferative effect.

Toxicity of HDAC inhibitors

VPA is the best known molecule in this class since it has been used for many years as an antiepileptic drug: it is extremely well tolerated by patients, and also its long-term effects are well known. The most commonly reported complaints are neurological symptoms (such as dizziness) that are generally transitory and reversible [79, 80]. Neurological symptoms may become excessive when VPA is combined with other agents [63].

When vorinostat (SAHA) was tested in mice, relevant systemic toxicity was observed only at high doses [81]. In patients, anaemia, anorexia, hyperglycaemia, thrombocytopenia, fatigue and nausea have frequently been reported [82]. Similar toxic side effects were described for SB939 [83], for rosminostat [69] and romidepsin [70]. ECG abnormalities of different severity are the most concerning toxic side effects of CHR3996 [45] and of romidepsin [54].

Antitumour activity

In a phase III trial of chemotherapy ± epigenetic treatment in cancer of the uterine cervix [59], there was an increase in progression-free survival, and the analysis of molecular correlates is pending.

Concerning phase II studies, that evaluated the antitumour activity of epigenetic agents alone or in combination with standard anticancer treatment, several studies have been reported. In many cases, results were described in terms of reduction of tumour volume in advanced disease resistant to several lines of anticancer treatments, similarly to what is standard for the evaluation of traditional antiproliferative agents [74, 82, 84, 85]. It is probably not surprising that several trials were reported as negative (see Tables 1 and 2 for details). Responses were evaluated according to Response Evaluation Criteria In Solid Tumors (RECIST): these may be useful for conventional anticancer therapies but do not seem adequate for epigenetic treatments that may result in disease stabilisation rather than in tumour shrinkage.

The evaluation of an epigenetic treatment, which is strictly connected to gene expression, can be performed more accurately in diseases where the genetic influence on the activity of the antitumour treatment is known. For this reason, the choice to evaluate the combination of entinostat and erlotinib in NSCLC was very sound [86]. The addition of the epigenetic agent, however, only improved progression-free survival (PFS) in tumours with high levels of e-cadherin, suggesting that this may represent a selection criterion for further studies.

The neo-adjuvant treatment of breast cancer represents a unique possibility in order to evaluate the activity of new agents. Not only new anticancer agents but also epigenetic therapy (hydralazine and valproate) have been tested in this setting [57], and the analysis of tumour biopsies confirmed that an epigenetic effect in terms of demethylation and of histone acetylation was detectable.

An interesting tactic was to evaluate the addition of epigenetic treatment in order to prevent or to reverse resistance due to the overexpression of a specific gene. This is the case, for example, of the increase in thymidylate synthase induced by fluorouracil treatment [87]. A clinical trial performed to demonstrate that this can be obtained in patients proved that treatment was feasible even if no clinically meaningful effect was obtained and no data were available for PD evaluation [88]. In a different study, patients with tumours progressing during standard chemotherapy were treated with the same regimen with the addition of hydralazine and valproate [58]. Data were analysed in detail, and even if tumour response was limited, there was evidence that adequate plasma concentrations were achieved and that an epigenetic effect was present. A similar approach was used in melanoma patients treated with temozolomide and decitabine: there was no antitumour activity even in the presence of a measurable PD effect [55].


Epigenetic therapy is being more and more recognised as an effective and well-tolerated treatment of cancer. Data in leukaemias and myelodysplastic syndromes are now consistent, and the success obtained in cutaneous lymphomas represents a proof of principle that solid tumours may also respond. This is also supported by preclinical data; still clinical results fall short of expectations: several reasons may explain this discrepancy.

We are convinced that, similarly to what has been observed for tyrosine-kinase inhibitors in cancer [89], we need a better selection of tumours and of patients that may benefit from these treatments. It has already been stated that epigenetic drugs, and HDAC inhibitors in particular, “might be useful only in those tumours in which HDACs are directly involved in the pathogenesis” [27].

It is not surprising that clinical results have generally been disappointing: standard evaluation of anticancer activity, mostly based on tumour volume reduction, may not be an adequate index of activity for epigenetic treatment. Epigenetics is a complex mechanism of gene regulation: it will take time before we can exploit it at its best. We definitely need more appropriate tests to select potentially responding tumours, but we also need agents with demonstrated epigenetic activity and solid data in order to choose the most effective dose and schedule.

Several technical issues remain to be solved and this will keep researchers busy, both in preclinical and clinical settings, for a long time.





DNA methyltransferase-3-like


DNA methyltransferases


Food and Drug Administration


histone 3


histone 4


histone acetyltransferases


histone deacetylases


histone methyltransferases


myelodysplastic syndromes


O(6)-methylguanine DNA methyltransferase


non-coding RNAs


non-small cell lung cancer


peripheral blood mononuclear cells




progression-free survival




Response Evaluation Criteria In Solid Tumors


suberoylanilide hydroxamic acid (vorinostat)


valproic acid



We apologise to the researchers whose works are not cited here due to space limitations. The work was supported in part by the Italian Association for Cancer Research (AIRC IG11949 to C.N.), Italian Foundation for Cancer Research (Fellowship for Italy 15142 to E.D.M) and University of Rome “La Sapienza”.

This work is dedicated to the late Professor Carlo Nervi, for us an example of how, in Oncology, clinical practice should be combined with scientific curiosity.

Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License (, which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver ( applies to the data made available in this article, unless otherwise stated.

Authors’ Affiliations

Department of Medical and Surgical Sciences and Biotechnology, University of Rome “la Sapienza”, Corso della Repubblica, 97, 04100 Latina, Italy


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