DRUG PROFILE

Decitabine in myelodysplastic syndromes

Elias Jabbour, Decitabine (5-aza-2’-deoxycytidine), a cytosine analog, inhibits DNA methylation and has Hagop M Kantarjian, dual effects on neoplastic cells, including the reactivation of silenced genes and Guillermo Garcia-Manero & differentiation at low doses, and cytotoxicity at high doses. Decitabine has promising Jean-Pierre J Issa† clinical efficacy in the treatment of myelodysplastic syndromes (a heterogeneous group of bone marrow malignancies), with evidence of target modulation (hypomethylation) and a †Author for correspondence The University of Texas, favorable toxicity profile. Optimal dosing schedules of decitabine in myelodysplastic Department of Leukemia, syndrome are those that maximize hypomethylation (low dose, high dose intensity, Unit 428, multiple cycles). However, the molecular mechanisms of in vivo response to decitabine are MD Anderson Cancer Center, 1515 Holcombe Blvd, still unclear. Combination therapies that augment decitabine’s epigenetic effect, or take Houston, Texas 77030, USA advantage of gene activation, will likely improve clinical responses and may extend its use Tel.: +1 713 145 2260 to the treatment of other malignancies. Fax: +1 713 794 4297 [email protected] The development of new therapeutic strategies disease progression. The cyclin-dependent for the (MDS) has kinase inhibitor p15 was described as a frequent been the result of extensive understanding of the target of aberrant methylation in MDS and its pathobiology of the disease. Therapeutics target- inactivation is associated with an increased risk ing chromatin structure, angiogenesis and the of progression to microenvironment that nurtures the MDS phe- (AML) [7]. Other similar genes were also notype have demonstrated significant activity described and their aberrant methylation was and offer an opportunity to alter the natural his- associated with resistance to [8,9]. tory of the disease [1]. Chromatin remodeling is a (5-azacitidine) and decitabine (5- powerful mechanism of regulating gene expres- aza-2’-deoxycytidine) are cytosine analogs synthe- sion and protein function [2]. In extreme states, sized in the 1960s with demonstrated in vitro anti- chromatin remodeling can permanently repress leukemic activity [5]. Azacitidine is converted the expression of a gene – a situation termed epi- intracellulary to decitabine. Decitabine in turn is genetic silencing. Such silencing is exploited by converted to decitabine triphosphate which incor- cancers to fully express the malignant porates into DNA, binds to and depletes DNA phenotype [3]. Evidence supporting a role of epi- methyltransferase protein levels, and results in rep- genetic gene silencing in tumorigenesis stems lication-dependent DNA hypomethylation [10,11]. from studies revealing a large number of genes Decitabine appears to have dual effects on treated that are silenced by aberrant DNA methylation cells. At high doses, it causes DNA synthesis arrest in different types of cancers – many of which are and apoptosis due to DNA adducts [12]. At low involved in the control of cell-cycle progression, doses, cells survive but change their gene expression apoptosis, tissue invasion and genomic stability. profile to favor differentiation, reduced prolifera- DNA methylation is remarkably altered in tion, and/or increased apoptosis [10,13]. This dual most malignancies, with concomitant global activity reawakened interest in hypomethylating and localized hypermethylation [4]. This agents as both antineoplastics and biologic response increased methylation affects mainly CpG modifiers [5]. Azacitidine was recently approved by islands located in regulatory regions such as gene the US Food and Drug Administration (FDA) for Keywords: decitabine, promoters, and often suppresses gene expression the treatment of MDS and has been extensively epigenetic therapy, permanently, providing cancers with an alterna- reviewed [14–16]. This article focuses on the role of methylation, tive to mutations or deletions for the inactiva- decitabine in the treatment of MDS. myelodysplastic syndrome tion of tumor-suppressor and other critical genes. Indeed, leukemias and MDS are charac- Mechanism of action terized by the hypermethylation and silencing of Decitabine is a deoxycytidine analog which is multiple genes [5,6]. This process can occur early phosphorylated and incorporated into DNA. Future Drugs Ltd in the disease course and is also associated with Once incorporated, it covalently binds to DNA

10.1586/14750708.2.6.835 © 2005 Future Drugs Ltd ISSN 1475-0708 Therapy (2005) 2(6), 835-842 835 DRUG PROFILE – Jabbour, Kantarjian, Garcia-Manero & Issa

methyltransferases and traps the enzymes to of decitabine produces a loss of clonogenicity in DNA, acting as an irreversible inhibitor of their the same range of cells in the S phase (30–50%). enzymatic activity. As a result, decitabine pro- A longer exposure time of 24 h showed a mark- duces marked DNA hypomethylation in vitro edly higher antineoplastic activity, with greater and in vivo [17]. Through hypomethylation than 95% loss of clonogenicity, using a dose of induction, decitabine restores silenced gene 1µm[25]. In plasma and cerebrospinal fluid (CSF) expression. The precise molecular mechanism of pharmacokinetic assays, the half-life of decitabine this phenomenon is increasingly being deci- was between 39 and 144 min depending on the phered. A number of studies have illustrated a animal model, and decitabine concentrations in cascade of biochemical events triggered by pro- the CSF were 27 to 58% of the plateau plasma moter DNA methylation that involves initial concentration [11]. However, the intracellular half- DNA binding proteins, which attract histone life of decitabine, particularly after DNA incorpo- deacetylases and histone methylases that eventu- ration, is not known and maybe considerably ally modify histones into a silenced chromatin higher. Oral administration of decitabine is not state [18,19]. Moreover, a feedback loop appears to optimal due to rapid decomposition of this analog be evident between DNA methylation and his- in acid [11]. tone methylation whereby each of these bio- chemical modifications at a given gene triggers Clinical trials the other, thus creating a self-reinforcing silenc- Phase I studies ing loop [19]. This silencing loop is interrupted by Decitabine has been in clinical trials for over two decitabine. Thus, treatment of neoplastic cell decades [5]. The original studies were classical lines with decitabine has been shown to induce Phase I trials which identified the maximally toler- hypomethylation and reverse the silenced histone ated dose (MTD) as 1500 to 2250 mg/m2 [26,27]. code rapidly at tumor-suppressor gene loci [20–22]. The dose-limiting toxicity was primarily hemato- This dual effect (hypomethylation–histone logic. Phase II studies were disappointing in solid changes) explains the superiority of decitabine on tumors [28], but more promising in AML, MDS gene expression activation, compared with the and CML. histone-deacetylase inhibitors [23]. Decitabine also The first clinical studies of decitabine in has significant effects on the expression of genes hematologic malignancies used 1500 to that are not silenced by CpG island methylation. 2500 mg/m2/course. Response rates with decit- Decitabine induces the expression of p21, a gene abine as a single agent or in combination with that shows no DNA hypermethylation in other therapies were 30 to 60% [13,26,27,29–31] cancer [24]. At a molecular level, the effects of (Table 1). Despite promising activity, high-dose decitabine on the histone code are not limited to decitabine regimens were not pursued because of genes showing silencing by promoter-associated delayed and prolonged myelosuppression. At methylation [22], for example the histone H3- these doses, the drug is likely to be working as a lysine 9 acetylation:methylation ratio was signifi- cytotoxic cytosine nucleoside analog, and its cantly increased by decitabine treatment of neo- superiority in that regard to was not plastic cells at genes showing no DNA clear. Decitabine at a lower dose schedule hypermethylation. This silencing-independent (15 mg/m2 three times a day for 3 days) was activity of decitabine remains incompletely reported to have encouraging activity in understood. Some of the changes could be reac- MDS [32]. An even lower dose (0.15 mg/kg/day tive, related to the stresses of exposing cells to a over 1 h for 10 days) was reported to have bio- potentially cytotoxic agent. Nevertheless, the ulti- logical efficacy in reactivating hemoglobin F in mate antineoplastic mechanism of action for patients with sickle cell disease, with relatively decitabine could be very pleiotropic and deserves little toxicity [33]. These observations, combined further investigation. with the short half-life of the drug and its abso- lute requirement for DNA synthesis for activity, Pharmacology & led to a novel Phase I trial of decitabine in Given that the antineoplastic action of decitabine patients with relapsed or refractory leukemia, is a result of its incorporation into newly synthe- testing low-dose longer exposure schedules with sized DNA, it is an S phase-specific agent [11]. At the intent of finding an ‘optimal dose’ for low doses, it does not block progression responses other than the MTD [34]. A total of 50 of G1-phase cells into the S phase. In clonogenic patients (44 with AML/MDS, five with CML assays, a 1 h exposure at a concentration of 10 µm and one with acute lymphoblastic leukemia

836 Therapy (2005) 2(6) Decitabine – DRUG PROFILE

Table 1. Selected decitabine Phase I and II clinical trials in myeloid malignancies. Decitabine therapy n Response Nonhematologic toxicity Ref.

0.75–80 mg/kg i.v. for 8–44 h 30 10% marrow Mild diarrhea, alopecia [27] response 45–100 mg/kg i.v. for 40–90 h 27 22% CR Mild nausea, vomiting, [26] mucositis 45–270 mg/m2 i.v. over 27 15% CR Grade III hepatic, GI, renal, [50] 9–12 h × 3 days pulmonary 270–360 mg/m2 i.v. 12 25% CR No grade III-IV [29] over 3–4h × 3days 45–50 mg/m2 i.v. × 3 days 10 50% HI Nausea/vomiting, peritonitis [30] 250 mg/m2 × 6 days with 22 59% CR Nausea/vomiting, diarrhea, [31] 120 mg/m2 on days 6–7, or peritonitis, CNS toxicity, 12 mg/m2 on days 5–7 weight loss, GI bleed 5–20 mg/m2 i.v. over 1 h × 50 18% CR, Hepatotoxicity [34] 10–20 days 14% PR CR: Complete response; GI: Gastrointestinal; HI: Hematologic improvement; i.v.: Intravenous; PR: Partial response.

[ALL]) were treated with increasing doses of increment after at least two cycles [38]. The decitabine (5, 10, 15 and 20 mg/m2) intrave- median duration of response was 9 months and nously over 1hr daily, 5 days aweek for 2 the median survival 15 months, with a 2-year consecutive weeks. The starting dose per course survival rate of 34%. Clinical complete remis- in this study was thus 30 times less than the sions were also associated with cytogenetic remis- MTD. The duration was then increased to 15 sions [39]. Survival was better for patients and 20 days. The treatment was well tolerated, achieving a cytogenetic response compared with with myelosuppression being the major side those who did not. Preliminary results of another effect. Responses were seen at all dose levels eval- Phase II trial of decitabine in MDS were recently uated. Overall, there were nine complete reported in abstract form [40]. The study was a responses (CR), one partial response (PR) and randomized Phase II trial of decitabine, testing three hematologic improvements (HI). In that both the dose intensity and subcutaneous route study, the most striking finding was that patients of administration. Patients received a total dose treated at higher cumulative doses had a lower of 100 mg/m2/course, and were randomized in a response rate. This low response rate at high doses Bayesian design to three groups consisting of: was consistent with earlier studies. Recent studies •10mg/m2/day intravenously over 1 h for have also evaluated decitabine as a continuous 10 days infusion in MDS [35] as well as in solid tumors [36]. •20mg/m2/day intravenously over 1 h for These dose schedules were generally found to be 5days less effective and/or more toxic (myelosuppression) 2 than bolus intravenous schedules. •20mg/m/day subcutaneously for 5 days (two doses) Phase II studies Cycles were administered every 4 weeks and Two relatively large Phase II studies of decitabine responses evaluated after 3 cycles. A total of 63 in MDS were recently reported (Table 2). Wijer- evaluable patients were reported on, with a mans and colleagues reported on several studies median age of 66 years (range 39–90 years), 21% conducted in Europe of decitabine in had secondary MDS and 57% had unfavorable MDS [32,35,37]. In these studies, 169 older cytogenetics. The overall response rate was patients (median 70 years) with intermediate or reported to be 82% (CR: 37%; PR: 8%; marrow high-risk MDS were treated with a relatively low CR: 20%; clinical benefit: 16%). Based upon the dose of decitabine (135 mg/m2 total treatment schedule, 47% of patients in the intra- dose/course). The overall response rate was 49% venous (5-day treatment group) achieved a CR and the induction death rate was 7%. A remarka- compared with 29%, and 24% of patients rand- ble response in the platelet count was seen, with omized to the subcutaneous group and intra- 63% of the patients showing a significant platelet venous (10-day treatment) groups, respectively, www.future-drugs.com 837 DRUG PROFILE – Jabbour, Kantarjian, Garcia-Manero & Issa

demonstrating that, within low-dose schedules, 3 months of therapy. The median survival of the most dose-intensive regimens were clinically responders (CR and PR) was 678 days compared superior. The side-effect profile was favorable and with 406 days in nonresponders (p = 0.038). included primarily myelosuppression. Overall, as an intent-to-treat analysis, there was a nonsignificant trend for longer time-to-AML Phase III studies transformation or death (338 days in the decit- A Phase III study of decitabine versus supportive abine group vs. 263 days in the supportive care care in patients with advanced MDS was recently group, p = 0.2). Subgroup analysis indicated a reported in abstract form (Table 2) [41]. A total of greater benefit in IPSS INT-2/high risk patients. 170 patients were randomized to decitabine ver- All patients who responded to decitabine had sus supportive care. Decitabine was administered higher quality of life scores, and myelosuppres- as a 3-h infusion of 15 mg/m2 every 8 h for 3 sion was the primary toxicity reported. A days, with cycles repeated every 6 weeks for up to Phase III multicenter trial comparing the same ten cycles (135 mg/m2/cycle). The groups were dose/schedule of decitabine with supportive care comparable for several risk factors, including: in elderly patients (age > 60 years) with MDS is •Age currently being performed by the European • Cytogenetics Organization for Research and Treatment of Cancer (EORTC). • Time from diagnosis • International prognostic scoring system In vivo molecular effects of decitabine (IPSS) score Global hypomethylation after decitabine therapy • Secondary MDS (14%) in vivo was observed in early trials, thus confirm- Decitabine resulted in a higher OR rate ing target modulation [42]. This has been studied (17%; CR: 9%; PR: 8 vs. 0%; p < 0.001), with in more detail in recent studies in AML and median response duration of 9 months. CML [17,34]. Hypomethylation after decitabine Responses were obtained after a median of was dose dependent, peaked 10 to 15 days after a

Table 2. Clinical results of single-agent decitabine in patients with MDS. Dose Patient characteristics Response Ref. Median n IPSS > 1 Rate Median Median age (range) (%) (%) duration survival (years) 50–75 mg/m2/day CI x 3 days every 6 weeks 72 29 NR OR: 54 ≥31 weeks 46 weeks [35] (n = 21); (58–82) (CR: 29; 40 mg/m2/day CI x 3 days every 6 weeks PR: 18; HI: 7) (n = 8) 15 mg/m2 t.i.d. i.v. 68 66 76 OR: 49 31 weeks 15 months [32] over 4 h x 3 days every 6 weeks (38–84) (CR: 20; PR: 5; HI:24) 45–50 mg/m2/day i.v. over 4 h x 3days every 70 169 72 OR: 49 40 weeks 15 months *[37] 6 weeks (38–89) 20 mg/m2/day i.v. over 1 h x 5 days every 4 66 63 52 OR: 82 NR NR [40] weeks (n = 32) (39–90) (CR: 37; vs.10 mg/m2/day i.v. over 1 h x 10 days every PR: 8; HI:36) 4 weeks (n = 14) vs.10 mg/m2 b.i.d. sc. x 5 days every 4 weeks (n = 17) 15 mg/m2 i.v. NR 89 69 OR: 17 ≥9 months NR [41] over 3 h t.i.d. x 3 days every 6 weeks (CR: 9; PR:8; HI: NR) * Includes updated results of patients enrolled in the two previously reported trials. b.i.d.: Twice daily; CI: Confidence interval; CR: Complete response; HI: Hematologic improvement; IPSS: International prognostic scoring system; i.v.: Intravenous; MDS: Myelodysplastic syndrome; NR: Not reported; OR: Overall response; PR: Partial response; sc.: Subcateous t.i.d.: Three-times daily.

838 Therapy (2005) 2(6) Decitabine – DRUG PROFILE

10-day course and recovers to baseline at 4 to MDS. However its potential as an epigenetic 6 weeks. Hypomethylation after cycle 1 drug has just begun to be investigated and several showed an inconsistent association with questions remain unanswered. response, with a positive correlation in AML, The issue of optimal dosing of this agent is but an inverse correlation in CML. It was under evaluation and still unclear, given that hypothesized that the inverse correlation decitabine has dual activity (hypomethylating at between hypomethylation and response could low doses, cytotoxic at high doses). Here, the be due to a cell death mechanism of response classical MTD route to drug development is not and resistance, whereby resistant cells can appropriate, and may have hindered the full eval- withstand more hypomethylation. uation of the drug. Indeed favorable responses Daskalakis and colleagues studied p15 meth- were reported at doses ten to 30-times lower ylation in DNA extracted from bone marrow than the MTD in patients with MDS, and it is mononuclear cells from patients with MDS not clear that higher doses are beneficial clini- treated with decitabine [43]. They found demeth- cally. Correlative studies suggest that the in vitro ylation in serial samples in nine out of 12 observation of rapid saturation of the hypometh- patients treated, and evidence of p15 gene reacti- ylation effect (and loss of the differentiation vation by immunohistochemistry in four effect) with increasing doses is also true patients with low baseline expression. This reac- in vivo [10]. Moreover, it was recently reported tivation was observed in responding patients, that greater hypomethylation and greater clinical and indeed, the authors show evidence of gene efficacy were the result of a better dose intensity reactivation in morphologically dysplastic cells of decitabine. The sum total of dose-finding in patients who were not in complete remission studies suggest that: at the time of examination, demonstrating • Short bolus infusions are better than continuous in vivo the potential of this drug. However, this infusion schedules effect may require multiple cycles of the drug. In • Lower doses are better than high doses a recent study, p15 hypomethylation after one cycle of decitabine was observed, however there • Dose intensity results in higher responses was no correlation between p15 methylation at Pharmacologically, these data could be baseline or after therapy, and response [34]. explained by a correlation between peak levels of The effects of decitabine on gene expression the drug and responses – an issue that deserves in vivo remain to be well characterized. In addi- investigation. Simply put, a high dose of the tion to effects on silenced tumor-suppressor drug is required for intracellular incorporation, genes, it will be important to also look at a after which the intracellular half-life of the drug broad variety of potential targets. In vitro, decit- may be long enough to achieve a therapeutic abine has been shown to activate genes that do effect. It remains to be seen whether 20 mg/m2 not show promoter CpG island methylation over 1 h is optimal to achieve hypomethylation, (e.g., p21) [24], and gene expression microarrays and how many days are required. The issue of have also revealed activation of a number of optimal treatment duration and maintenance genes that do not have promoter-associated therapy should also be investigated. CpG islands [44]. It is distinctly possible that the It is not entirely clear whether responses to therapeutic effects of this drug involve more decitabine in vivo are related to hypomethylation than simple induction of tumor-suppressor or cytotoxicity. The observation of decreasing gene hypomethylation and such studies will responses with increasing dose favors hypo- help elucidate these issues. methylation, but this question is far from being definitively answered. Moreover, even if Expert commentary & outlook hypomethylation is the mechanism mediating Given that MDS is primarily a disease of older responses, events downstream remain to be individuals, aggressive therapies such as combina- defined. Possibilities include: tion chemotherapy and stem cell transplantation • Direct cell death signaling by hypomethylation, are simply not realistic for most patients. There is perhaps through reactivation of retrotransposons much interest therefore in exploring less toxic • Induction of differentiation agents in this disease, and learning how to inte- grate them in a multiagent therapeutic approach. • Induction of senescence It is obvious from the above data that decitabine, • Induction of apoptosis through reactivation of especially at lower doses, has significant effects in proapototic molecules [45] www.future-drugs.com 839 DRUG PROFILE – Jabbour, Kantarjian, Garcia-Manero & Issa

• Induction of immune responses through • Combinations that augment its epigenetic modulation of tumor antigens [46] or the host’s effect immune system • Combinations that take advantage of its As mentioned earlier, loss of methylation of epigenetic effects. the p15 tumor-suppressor gene after multiple cycles was observed in patients with MDS Combinations of decitabine and histone responding to decitabine, but p15 hypomethyl- deacetylase inhibitors (HDACi) are synergistic in ation acutely after cycle 1 did not correlate with reactivating gene expression [23], and combina- response in separate studies. Multiple cycles tions with inhibitors of methylated-DNA binding may be needed to achieve enough tumor-sup- proteins or histone H3 lysine 9 methyltransferases pressor gene hypomethylation – a molecular are also attractive possibilities. Based on in vitro property that may explain the clinical patterns synergistic activity, trials combining decitabine of response. Overall, the molecular mediators of with the HDACi’s valproic acid, depsipeptide and decitabine effect (beyond hypomethylation) suberoylanilide hydroxamic acid are ongoing. remain to be clarified. Separately, decitabine has been shown in vitro to A key issue in the management of MDS is to sensitize cells to the effects of biologic therapy reduce its heterogeneity using molecular profiles, such as retinoic acid [48] and to increase the which will then be essential to select patients for expression of pro-apoptotic molecules [45], which therapy. The IPSS uses a combination of clinical may enhance the efficacy of classic chemothera- data with cytogenetics to achieve a certain degree peutic agents. It has also been demonstrated to of selection [47], but it remains imperfect. More reverse drug resistance in selected cases [49]. Clini- importantly, the IPSS does not predict who is cal trials investigating such combination therapies going to respond to decitabine or other treatment are underway. modalities in MDS. A concerted effort to supple- The next frontier for decitabine is to move ment the IPSS using gene-expression profiles is beyond MDS. There is no evidence for particu- essential to making progress in this disease. lar methylation profiles in MDS that would In moving forward, it is important to consider make this disease uniquely sensitive to the relative benefits and properties of the two hypomethylating agents [5]. It may simply be that hypomethylating agents – azacitidine and decit- responses in MDS were made possible by the abine. While both agents inhibit methylation, need to use low doses of the agent in this disease they are not identical or interchangeable. Azaci- and the possibility of treating patients with mul- tidine incorporates into RNA and inhibits RNA tiple cycles. It is imperative to now test this translation [15]. The contribution of this property approach (lower doses, multiple cycles) in other to clinical efficacy may be important but has not malignancies, including solid tumors. In fact, been studied. Only after inefficient conversion there is substantial evidence for decitabine activ- to decitabine does azacitidine become a ity in other hematologic malignancies such as . Clinically, comparing AML [34] and CML [17]. Renewed interest in this the Phase III studies of azacitidine and decitab- agent led to ongoing trials (alone or in combina- ine the response rates are similar, although the tion) in various malignancies, and the results of population of patients treated with decitabine these should be available within the next was more advanced (higher IPSS score, longer few years. As a single agent, these include trials time from treatment). Of note, responses seem in previously untreated older patients with to occur earlier with decitabine than azacitidine. AML, in imatinib-resistant CML, in relapsed or Noncomparative Phase II studies suggest a refractory CLL, as well as solid tumor trials in higher CR rate with decitabine, though this , prostate cancer, melanoma, lym- observation may have related to persistent efforts phoma and others. Combinations involving at optimizing decitabine schedules, effects that decitabine and either classical chemotherapeutics are still to be done for azacitidine. Overall, direct or HDACi are also being tested in AML, ALL, comparative studies of the two drugs are needed, CML and various solid tumors such as refractory but the distinction of their mechanisms of action ovarian cancer and breast cancer. indicate that there is a likely role for both drugs in MDS. Conclusion The future use of decitabine will likely be in Hypomethylating agents reverse gene silencing, combination with other agents. This could be which appears critical to maintaining the neo- thought of in two types of combinations: plastic phenotype. Decitabine shows significant

840 Therapy (2005) 2(6) Decitabine – DRUG PROFILE

promise in MDS in clinical trials, and its use will are already apparent, and more impressive become widespread, at least for myeloid malig- benefits will come from a combination of thera- nancies. Although many questions regarding this peutic approaches. In the long run, it is hoped agent remain to be answered, including dose and that such therapy will contribute to achieving in vivo mechanisms of action, clinical benefits actual cures of human malignancies.

Highlights

• DNA methylation is an epigenetic modification responsible for silencing of gene transcription. • Inappropriate inhibition of the transcription of certain genes, such as tumor-suppressor genes and genes involved in DNA repair, can lead to unregulated growth and proliferation of cells in both solid tumors and hematologic malignancies. • Decitabine (5-aza-2’-deoxycytidine) is a potent and specific inhibitor of DNA methyltransferase and is a hypomethylating agent. It has dual effects on neoplastic cells, including reactivation of silenced genes and differentiation at low doses, and cytotoxicity at high doses. • Decitabine at low doses is an effective treatment in advanced MDS, with manageable toxicity (primarily myelosuppression). • Decitabine in vivo mechanisms of action, optimal dosing, relative efficacy compared to azacitine (5- azacitidine) and efficacy in other malignancies (including solid tumors) remain to be clarified • Clinical trials exploiting combination epigenetic therapy or making use of gene reactivation are currently ongoing.

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