EZH2抑制剂联合地西他滨和利沙托克拉治疗复发难治急性髓系白血病。

Simple Summary Epigenetics is the study of heritable changes in gene expression that do not alter the underlying DNA sequence. In acute myeloid leukemia (AML), these regulatory mechanisms are frequently dysregulated, driving uncontrolled proliferation and blocking normal myeloid differentiation. This review systematically examines the principal layers of epigenetic regulation, including DNA methylation, histone modifications, chromatin remodeling, and RNA-mediated epigenetics. We describe how their disruption contributes to AML pathogenesis, with particular emphasis on how recurrent genetic lesions such as DNMT3A mutations and KMT2A (MLL) rearrangements shape the epigenetic landscape of the disease. We further discuss current therapeutic options for AML, with a focus on approved agents and ongoing clinical trials targeting epigenetic regulators.

Acute myeloid leukemia (AML) is a genetically heterogeneous malignancy for which treatment options have been largely limited to cytotoxic chemotherapy for the past four decades. Next-generation sequencing and other approaches have identified a spectrum of genomic and epigenomic alterations that contribute to AML initiation and maintenance. The key role of epigenetic modifiers and the reversibility of epigenetic changes have paved the way for evaluation of a new set of drug targets, and facilitated the design of novel candidate treatment strategies. More recently, seven new targeted therapies have been FDA-approved demonstrating successful implementation of the past decades' research. In this review, we will summarize the most recent advances in targeted therapeutics designed for a focused group of key epigenetic regulators in AML, outline their mechanism of action and their current status in clinical development. Furthermore, we will discuss promising new approaches for epigenetic targeted treatment in AML which are currently being tested in pre-clinical trials.

Simple Summary Pediatric acute myeloid leukemia (AML) is a complex disease that has traditionally been treated with chemotherapy, but some forms of AML have not responded well to this approach. Recent studies have identified a diverse array of genetic changes that show differences between adult and pediatric AML, resulting in a shift in how clinicians think about the best way to treat this disease. The goal of this review is to provide an overview of key genetic targets and clinical studies that lay the foundation for establishing a framework to bring cutting-edge treatment options to the front line for treating pediatric AML.

There is a desperate need for new and effective therapeutic approaches to acute myeloid leukemia (AML) in both children and adults. Epigenetic aberrations are common in adult AML, and many novel epigenetic compounds that may improve patient outcomes are in clinical development. Mutations in epigenetic regulators occur less frequently in AML in children than in adults. Investigating the potential benefits of epigenetic therapy in pediatric AML is an important issue and is discussed in this review.

Simple Summary Venetoclax–azacitidine is a new standard for elderly or unfit acute myeloid leukemia patients. Nevertheless, resistance remains a matter of concern. The main genetic alterations associated with venetoclax sensitivity are IDH mutations, whereas TP53, signaling mutations, and BAX mutations are associated with venetoclax resistance. Non-genetic resistance mechanisms have also been described, including changes in apoptotic proteins, differentiation states, metabolic status, and mitochondrial machinery. Venetoclax-based triplet therapies including IDH and FLT3 inhibitors or innovative therapies are under investigation to target resistances. Abstract Venetoclax is a BH3-mimetics agent interacting with the anti-apoptotic protein BCL2, facilitating cytochrome c release from mitochondria, subsequent caspases activation, and cell death. Venetoclax combined with azacitidine (VEN-AZA) has become a new standard treatment for AML patients unfit for intensive chemotherapy. In the phase III VIALE-A study, VEN-AZA showed a 65% overall response rate and 14.7 months overall survival in comparison with 22% and 8 months in the azacitidine monotherapy control arm. Despite these promising results, relapses and primary resistance to venetoclax are frequent and remain an unmet clinical need. Clinical and preclinical studies have been conducted to identify factors driving resistance. Among them, the most documented are molecular alterations including IDH, FLT3, TP53, and the newly described BAX mutations. Several non-genetic factors are also described such as metabolic plasticity, changes in anti-apoptotic protein expression, and dependencies, as well as monocytic differentiation status. Strategies to overcome venetoclax resistance are being developed in clinical trials, including triplet therapies with targeted agents targeting IDH, FLT3, as well as the recently developed menin inhibitors or immunotherapies such as antibody–drug conjugated or monoclonal antibodies. A better understanding of the molecular factors driving venetoclax resistance by single-cell analyses will help the discovery of new therapeutic strategies in the future.

… of EZH2 were associated with a prolonged survival compared to … like BCL2 inhibitors and PD-L1 inhibitors. Ultimately, clinical outcome will be significantly improved for elderly AML …

DNA methylation is an enzyme‐driven epigenetic modification that must be precisely regulated to maintain cellular homeostasis. Aberrant methylation status, especially hypermethylation of the promoter sites of tumor‐suppressor genes, is observed in human malignancies and is a proven target for cancer therapy. The first‐generation DNA demethylating agents, azacitidine and decitabine, are widely used for treating several hematological malignancies. In addition, orally bioavailable prodrugs of azacitidine and decitabine have recently been approved by the FDA. We have developed a silylated derivative of decitabine, OR‐2100, which is resistant to degradation by cytidine deaminase and orally bioavailable. It has efficacy against several human hematological malignancies in xenograft mouse models with less hematotoxicity than decitabine. Since DNA demethylating agents are combined with molecularly targeted drugs in clinical use and trials, we think that the less hematotoxic profile of OR‐2100 makes it suitable for use as a combination therapy. In this article, we review the therapeutic approach in hematological malignancies with the DNA demethylating agent OR‐2100.

… In conclusion, decitabine induces global demethylation and … SUZ12, EED, and EZH2 were significantly overrepresented in the genes … BCL2L10 is a member of the Bcl-2 family that has …

The polycomb repressive complex (PRC) 2 contains 3 core proteins, EZH2, SUZ12, and EED, in which the SET (suppressor of variegation–enhancer of zeste-trithorax) domain of EZH2 mediates the histone methyltransferase activity. This induces trimethylation of lysine 27 on histone H3, regulates the expression of HOX genes, and promotes proliferation and aggressiveness of neoplastic cells. In this study, we demonstrate that treatment with the S-adenosylhomocysteine hydrolase inhibitor 3-deazaneplanocin A (DZNep) depletes EZH2 levels, and inhibits trimethylation of lysine 27 on histone H3 in the cultured human acute myeloid leukemia (AML) HL-60 and OCI-AML3 cells and in primary AML cells. DZNep treatment induced p16, p21, p27, and FBXO32 while depleting cyclin E and HOXA9 levels. Similar findings were observed after treatment with small interfering RNA to EZH2. In addition, DZNep treatment induced apoptosis in cultured and primary AML cells. Furthermore, compared with treatment with each agent alone, cotreatment with DZNep and the pan-histone deacetylase inhibitor panobinostat caused more depletion of EZH2, induced more apoptosis of AML, but not normal CD34+ bone marrow progenitor cells, and significantly improved survival of nonobese diabetic/severe combined immunodeficiency mice with HL-60 leukemia. These findings indicate that the combination of DZNep and panobinostat is effective and relatively selective epigenetic therapy against AML cells.

Venetoclax combined with hypomethylating agents is approved for frontline therapy in older/unfit acute myeloid leukemia (AML) patients. However, prospective data on this low intensity therapy in treatment-naive younger AML patients are lacking. This study investigated the efficacy and safety of venetoclax plus decitabine (VEN-DEC) as induction in untreated young fit AML patients in a randomized trial. Patients aged 18-59 years eligible for intensive chemotherapy were randomized 1:1 to receive VEN-DEC or IA-12 (idarubicin and cytarabine). All patients achieving CR/CRi underwent high-dose cytarabine consolidation. The primary endpoint was the composite complete remission rate (CRc) rate after induction therapy. Of 255 screened, 188 were enrolled and randomly assigned, with 94 in each group. In the intention-to-treat population, CRc was 89% (84/94) in the VEN-DEC group versus 79% (74/94) in IA-12 (non-inferiority P = 0.0021). MRD negativity after induction was 80% (67/84) versus 76% (56/74), respectively. VEN-DEC showed superior CRc in patients aged ≥40 years (91% vs. 75%), those with adverse risk (91% vs. 42%) or epigenetic mutations (91% vs. 67%) , but lower CRc in RUNX1::RUNX1T1 fusion cases (44% vs. 88%) compared to IA-12. Patients in the VEN-DEC group experienced fewer grade ≥3 infections (32% vs. 67%) and shorter severe thrombocytopenia duration (median 13 vs. 19 days, P < 0.001). At a median follow-up of 12.1 months, overall and progression-free survival were similar between groups. In conclusion, VEN-DEC demonstrated non-inferior response rates with superior safety over IA-12 in young AML patients. The trial was registered at ClinicalTrials.gov as #NCT05177731.

In acute myeloid leukemia (AML), therapy resistance frequently occurs, leading to high mortality among patients. However, the mechanisms that render leukemic cells drug resistant remain largely undefined. Here, we identified loss of the histone methyltransferase EZH2 and subsequent reduction of histone H3K27 trimethylation as a novel pathway of acquired resistance to tyrosine kinase inhibitors (TKIs) and cytotoxic drugs in AML. Low EZH2 protein levels correlated with poor prognosis in AML patients. Suppression of EZH2 protein expression induced chemoresistance of AML cell lines and primary cells in vitro and in vivo. Low EZH2 levels resulted in derepression of HOX genes, and knockdown of HOXB7 and HOXA9 in the resistant cells was sufficient to improve sensitivity to TKIs and cytotoxic drugs. The endogenous loss of EZH2 expression in resistant cells and primary blasts from a subset of relapsed AML patients resulted from enhanced CDK1-dependent phosphorylation of EZH2 at Thr487. This interaction was stabilized by heat shock protein 90 (HSP90) and followed by proteasomal degradation of EZH2 in drug-resistant cells. Accordingly, inhibitors of HSP90, CDK1 and the proteasome prevented EZH2 degradation, decreased HOX gene expression and restored drug sensitivity. Finally, patients with reduced EZH2 levels at progression to standard therapy responded to the combination of bortezomib and cytarabine, concomitant with the re-establishment of EZH2 expression and blast clearance. These data suggest restoration of EZH2 protein as a viable approach to overcome treatment resistance in this AML patient population.

Chemotherapy resistance is the main impediment in the treatment of acute myeloid leukaemia (AML). Despite rapid advances, the various mechanisms inducing resistance development remain to be defined in detail. Here we report that loss-of-function mutations (LOF) in the histone methyltransferase EZH2 have the potential to confer resistance against the chemotherapeutic agent cytarabine. We identify seven distinct EZH2 mutations leading to loss of H3K27 trimethylation via multiple mechanisms. Analysis of matched diagnosis and relapse samples reveal a heterogenous regulation of EZH2 and a loss of EZH2 in 50% of patients. We confirm that loss of EZH2 induces resistance against cytarabine in the cell lines HEK293T and K562 as well as in a patient-derived xenograft model. Proteomics and transcriptomics analysis reveal that resistance is conferred by upregulation of multiple direct and indirect EZH2 target genes that are involved in apoptosis evasion, augmentation of proliferation and alteration of transmembrane transporter function. Our data indicate that loss of EZH2 results in upregulation of its target genes, providing the cell with a selective growth advantage, which mediates chemotherapy resistance.

Acute myeloid leukemia (AML), a malignant disease of the bone marrow, is characterized by the clonal expansion of myeloid progenitor cells and a block in differentiation. The high heterogeneity of AML significantly impedes the development of effective treatment strategies. Enhancer of zeste homolog 2 (EZH2), the catalytic subunit of the polycomb repressive complex 2 (PRC2), regulates the expression of downstream target genes through the trimethylation of lysine 27 on histone 3 (H3K27me3). Increasing evidence suggests that the dysregulation of EZH2 expression in various cancers is closely associated with tumorigenesis. In the review, we examine the role of EZH2 in AML, highlighting its crucial involvement in regulating stemness, proliferation, differentiation, immune response, drug resistance and recurrence. Furthermore, we summarize the application of EZH2 inhibitors in AML treatment and discuss their potential in combination with other therapeutic modalities. Therefore, targeting EZH2 may represent a novel and promising strategy for the treatment of AML.

Deep sequencing has revealed that epigenetic modifiers are the most mutated genes in acute myeloid leukemia (AML). Thus, elucidating epigenetic dysregulation in AML is crucial to understand disease mechanisms. Here, we demonstrate that metal response element binding transcription factor 2/polycomblike 2 (MTF2/PCL2) plays a fundamental role in the polycomb repressive complex 2 (PRC2) and that its loss elicits an altered epigenetic state underlying refractory AML. Unbiased systems analyses identified the loss of MTF2–PRC2 repression of MDM2 as central to, and therefore a biomarker for, refractory AML. Thus, immature MTF2-deficient CD34+CD38− cells overexpress MDM2, thereby inhibiting p53 that leads to chemoresistance due to defects in cell-cycle regulation and apoptosis. Targeting this dysregulated signaling pathway by MTF2 overexpression or MDM2 inhibitors sensitized refractory patient leukemic cells to induction chemotherapeutics and prevented relapse in AML patient-derived xenograft mice. Therefore, we have uncovered a direct epigenetic mechanism by which MTF2 functions as a tumor suppressor required for AML chemotherapeutic sensitivity and identified a potential therapeutic strategy to treat refractory AML. Significance: MTF2 deficiency predicts refractory AML at diagnosis. MTF2 represses MDM2 in hematopoietic cells and its loss in AML results in chemoresistance. Inhibiting p53 degradation by overexpressing MTF2 in vitro or by using MDM2 inhibitors in vivo sensitizes MTF2-deficient refractory AML cells to a standard induction-chemotherapy regimen.

Our understanding of the significance of epigenetic dysregulation in the pathogenesis of myeloid malignancies has greatly advanced in the past decade. Enhancer of Zeste Homolog 2 (EZH2) is the catalytic core component of the Polycomb Repressive Complex 2 (PRC2), which is responsible for gene silencing through trimethylation of H3K27. EZH2 dysregulation is highly tumorigenic and has been observed in various cancers, with EZH2 acting as an oncogene or a tumor-suppressor depending on cellular context. While loss-of-function mutations of EZH2 frequently affect patients with myelodysplastic/myeloproliferative neoplasms, myelodysplastic syndrome and myelofibrosis, cases of chronic myeloid leukemia (CML) seem to be largely characterized by EZH2 overexpression. A variety of other factors frequently aberrant in myeloid leukemia can affect PRC2 function and disease pathogenesis, including Additional Sex Combs Like 1 (ASXL1) and splicing gene mutations. As the genetic background of myeloid malignancies is largely heterogeneous, it is not surprising that EZH2 mutations act in conjunction with other aberrations. Since EZH2 mutations are considered to be early events in disease pathogenesis, they are of therapeutic interest to researchers, though targeting of EZH2 loss-of-function does present unique challenges. Preliminary research indicates that combined tyrosine kinase inhibitor (TKI) and EZH2 inhibitor therapy may provide a strategy to eliminate the residual disease burden in CML to allow patients to remain in treatment-free remission.

Mutations in chromatin-regulating genes play a critical role in the pathogenesis of myelodysplastic neoplasia (MDS) and acute myeloid leukemia (AML), as genetic mutations affecting chromatin structure and function are key drivers of these hematologic malignancies. Central to the discussion are key emerging genes such as ASXL1, SRSF2, and EZH2, which are recognized as adverse prognostic markers. Mutations in these genes, coupled with subsequent alterations in epigenetic mechanisms, disrupt normal gene expression by impairing histone modification and RNA splicing processes. Specifically, mutations in ASXL1 enhance removal of ubiquitylation at histone H2AK119, leading to altered gene expression and impaired hematopoietic stem cell differentiation. Mutations in SRSF2, an RNA splicing factor, alter RNA-binding specificity, inducing aberrant splicing of key genes such as EZH2. Loss-of-function mutations in EZH2 disrupt PRC2-mediated transcriptional repression, promoting leukemic progression. However, while the effects of these mutations are understood, treatment options for high-risk patients remain limited. Emerging strategies, such as venetoclax combined with hypomethylating agents, showing promise in mitigating the poor prognosis associated with these mutations. This review consolidates recent findings on these epigenetic regulators and their interactions, providing insights into the multifaceted mechanisms of leukemogenesis in the interest of inspiring targeted therapeutic strategies and bridging extant treatment gaps for MDS/AML.

The methylation of lysine 27 on histone H3 (H3K27me3) is a chromatin mark associated with nucleosome condensation and gene expression silencing. EZH2 is a lysine methyltransferase that catalyzes H3K27me3. In this issue of Cancer Research, Porazzi and colleagues report that pretreatment with EZH2 inhibitors opened up the H3K27me3-marked chromatin of acute myeloid leukemia (AML) cells, which enhanced DNA damage and apoptosis induced by chemotherapeutic agents, in particular the topoisomerase II inhibitors, doxorubicin and etoposide. The EZH2 inhibitor/doxorubicin combination also enabled the expression of proapoptotic genes, potentially contributing to the death of AML cells. This study has significant implications for improving the efficacy of DNA-damaging cytotoxic agents in AML, thereby enabling lower chemotherapy doses and reducing treatment-related side effects. See related article by Porazzi et al., p. 458

Chemotherapy resistance and disease relapse are major determinants of treatment failure in acute myeloid leukemia (AML). Therapy-induced senescence (TIS) is one outcome of chemotherapy, but its immunological consequences in AML remain unclear. Here we show that ex vivo chemotherapy induces senescence in a subset of therapy-naïve AML samples. TIS is marked by elevated interferon signaling, upregulation of human leukocyte antigen (HLA) class I and II molecules, and increased presentation of leukemia- and senescence-associated peptides, conferring AML cells antigen-presenting cell-like features. These changes enhance autologous CD4+ and CD8+ T cell responses against AML, both ex vivo and in patient-derived xenograft models. TIS also restores AML sensitivity to immune checkpoint blockade therapy. Mechanistically, we identify reduced Polycomb Repressive Complex 2 (PRC2) activity as central to TIS induction and its immunogenicity. PRC2 inhibition reactivates senescence-related genes and HLA expression in non-senescent AML cells, enabling T cell activation. These findings uncover a senescence-driven immune mechanism with potential to improve therapy outcomes in AML. Acute myeloid leukemia responses to chemotherapy vary, and the effects of therapy induced senescence on anti-tumor immunity remain unclear. Here, the authors show that chemotherapy-induced senescence increases antigen presentation and T-cell recognition of leukemia cells and enhances responses to immune checkpoint blockade, while EZH2 inhibition can restore these immune features in less responsive leukemias.

Epigenetic dysregulation plays important roles in leukemogenesis and the progression of acute myeloid leukemia (AML). Histone acetyltransferases (HATs) and histone deacetylases (HDACs) reciprocally regulate the acetylation and deacetylation of nuclear histones. Aberrant activation of HDACs results in uncontrolled proliferation and blockade of differentiation, and HDAC inhibition has been investigated as epigenetic therapeutic strategy against AML. Cell growth was assessed with CCK-8 assay, and apoptosis was evaluated by flow cytometry in AML cell lines and CD45 + and CD34 + CD38- cells from patient samples after staining with Annexin V-fluorescein isothiocyanate (FITC)/propidium iodide (PI). EZH2 was silenced with short hairpin RNA (shRNA) or overexpressed by lentiviral transfection. Changes in signaling pathways were detected by western blotting. The effect of chidamide or EZH2-specific shRNA (shEZH2) in combination with adriamycin was studied in vivo in leukemia-bearing nude mouse models. In this study, we investigated the antileukemia effects of HDAC inhibitor chidamide and its combinatorial activity with cytotoxic agent adriamycin in AML cells. We demonstrated that chidamide suppressed the levels of EZH2, H3K27me3 and DNMT3A, exerted potential antileukemia activity and increased the sensitivity to adriamycin through disruption of Smo/Gli-1 pathway and downstream signaling target p-AKT in AML cells and stem/progenitor cells. In addition to decreasing the levels of H3K27me3 and DNMT3A, inhibition of EZH2 either pharmacologically by chidamide or genetically by shEZH2 suppressed the activity of Smo/Gli-1 pathway and increased the antileukemia activity of adriamycin against AML in vitro and in vivo. Inhibition of EZH2 by chidamide has antileukemia activity and increases the chemosensitivity to adriamycin through Smo/Gli-1 pathway in AML cells (Fig. 5). These findings support the rational combination of HDAC inhibitors and chemotherapy for the treatment of AML.

Dear Editor, Despite recent therapeutic advances, overall survival of acute myeloid leukemia (AML) patients remains poor. Cytarabine (Ara-C), in combination with anthracyclines, induces complete remissions in 60 – 80% of AML patients, with only a fraction of them achieving long-term survival, and 60 – 90% of refractory AML patients succumb to the disease. Treatment failure is mainly attributed to the insuf fi cient eradication of cell-kinetically quiescent leukemia stem/progenitor cells (LSPCs) that reside in the bone marrow (BM) niche. Therefore, overcoming the dormancy of LSPCs should improve survival of AML patients [1]. Quiescent LSPCs are dependent on enhancer of zeste homolog (EZH)2 and EZH1, the key catalytic subunits of the polycomb repressive complex 2 (PRC2). Genetic ablation of EZH2/1 in an AML model depleted quiescent LSPCs [2]. In this study, we observed that inhibition of EZH2/1 by valemetostat tosylate (DS-3201b or valemetostat) monotherapy in a phase 1 clinical trial induced proliferation and mobilization of immature blasts into circulation in AML patients. Preclinical experiments in vitro and in vivo recapitulated the fi ndings and indicated the therapeutic potential of valemetostat in combination with Ara-C. We assessed blast counts in AML patients enrolled in the U-102 study before and after valemetostat treatment (patient characteristics are described in Supplementary Table S1). Circulating blasts increased 0.14-to 474.1-fold with a mean increase of 65.42 ± 51.23 (SEM) fold (Fig. 1A, Supplementary Fig. S1A) after valemetostat administration. To study cell kinetics

Summary Background Activating mutations of EZH2, an epigenetic regulator, are present in approximately 20% of patients with follicular lymphoma. We investigated the activity and safety of tazemetostat, a first-in-class, oral EZH2 inhibitor, in patients with follicular lymphoma. Methods This study was an open-label, single-arm, phase 2 trial done at 38 clinics or hospitals in France, the UK, Australia, Canada, Poland, Italy, Ukraine, Germany, and the USA. Eligible patients were adults (≥18 years) with histologically confirmed follicular lymphoma (grade 1, 2, 3a, or 3b) that had relapsed or was refractory to two or more systemic therapies, had an Eastern Cooperative Oncology Group performance status of 0–2, and had sufficient tumour tissue for central testing of EZH2 mutation status. Patients were categorised by EZH2 status: mutant (EZH2mut) or wild-type (EZH2WT). Patients received 800 mg of tazemetostat orally twice per day in continuous 28-day cycles. The primary endpoint was objective response rate based on the 2007 International Working Group criteria for non-Hodgkin lymphoma, assessed by an independent radiology committee. Activity and safety analyses were done in patients who received one dose or more of tazemetostat. This study is registered with ClinicalTrials.gov, NCT01897571, and follow-up is ongoing. Findings Between July 9, 2015, and May 24, 2019, 99 patients (45 in the EZH2mut cohort and 54 in the EZH2WT cohort) were enrolled in the study. At data cutoff for the analysis (Aug 9, 2019), the median follow-up was 22·0 months (IQR 12·0–26·7) for the EZH2mut cohort and 35·9 months (24·9–40·5) for the EZH2WT cohort. The objective response rate was 69% (95% CI 53–82; 31 of 45 patients) in the EZH2mut cohort and 35% (23–49; 19 of 54 patients) in the EZH2WT cohort. Median duration of response was 10·9 months (95% CI 7·2–not estimable [NE]) in the EZH2mut cohort and 13·0 months (5·6–NE) in the EZH2WT cohort; median progression-free survival was 13·8 months (10·7–22·0) and 11·1 months (3·7–14·6). Among all 99 patients, treatment-related grade 3 or worse adverse events included thrombocytopenia (three [3%]), neutropenia (three [3%]), and anaemia (two [2%]). Serious treatment-related adverse events were reported in four (4%) of 99 patients. There were no treatment-related deaths. Interpretation Tazemetostat monotherapy showed clinically meaningful, durable responses and was generally well tolerated in heavily pretreated patients with relapsed or refractory follicular lymphoma. Tazemetostat is a novel treatment for patients with follicular lymphoma. Funding Epizyme.

Epigenomic dysregulation is associated with several noncommunicable diseases (NCDs). Chromatin-modifying drugs have the ability to re-establish normal epigenetic regulation, thereby fine-tuning the epigenome. The biochemical and pharmacological mechanisms of these drugs, which target histone lysine and arginine methyltransferases, DNA methyltransferases, histone deacetylases and histone acetyltransferases, histone demethylases and bromodomain and extra-terminal motifs, serve as ideal tools for understanding the modes of action of these epigenetic regulators and the resulting biological events within chromatin. The present review provides an overview of these biochemical mechanisms and some downstream effects of the aforementioned chromatin-modifying drugs.

Background and Objectives: Acute myeloid leukemia (AML) is characterized not only by its heterogeneity but also by its high relapse rate. This results in limited treatment options, especially in elderly or therapy-refractory patients. It is known that inhibiting anti-apoptotic BCL-2 family proteins can be effective; however, cellular resistance mechanisms often limit the efficacy of this treatment. We studied the effects of the BCL-2 inhibitor ABT-737, the MCL-1 inhibitor S63845, and their combination on AML cell lines and primary AML patient cells. Materials and Methods: To analyze the effects of ABT-737 and S63845 treatment on cells, cell energy phenotype, apoptosis, and cell cycle were assessed, and gene expression by RT-qPCR and protein levels by Western blot analysis were measured. Results: Treatment with the BCL-2 inhibitor ABT-737, the MCL-1 inhibitor S63845, and their combination reduced AML cell viability and induced apoptosis. Dual treatment also altered the expression of epigenetic regulators, as the levels of DNMT1, EZH2, SUZ12, and HDAC1 were reduced, while histone acetylation was increased. An increase in pro-apoptotic markers (PARP cleavage, caspase-9) was observed, and the expression of oncogenes (MYC, WT1) was reduced in model cell lines and primary AML patient cells. Conclusions: BCL-2 and MCL-1 inhibition, alone or in combination, induced apoptosis and altered the expression of epigenetic regulators and oncogenes in AML cell lines and primary patient cells, with no consistent advantage of combined treatment over single agents. BCL-2/MCL-1 inhibition remains a promising approach for AML, and further work should clarify which patients or disease subtypes are most likely to benefit from combined versus single-agent treatment.

… (DNMT) inhibitors such as decitabine. This review describes the pharmacology of decitabine and the clinical evidence supporting its use across the AML treatment spectrum. …

BACKGROUND The hypomethylating agent Decitabine (DAC) is a valuable treatment option in acute myeloid leukemia (AML), particularly in elderly patients (pts) not suitable for intensive chemotherapy (CHT). However, limited data are available about efficacy and safety of DAC in clinical practice. PATIENTS AND METHODS We retrospectively reviewed data of 104 AML pts treated with DAC in eight Italian Hematological Centers from 2015 to 2017. The objective of this study was to evaluate the efficacy and safety of DAC in older AML pts outside of clinical trial. Seventy-five (75%) pts received DAC as first line treatment (Cohort 1) and 29 pts as salvage therapy (Cohort 2). All pts received a DAC schedule of 20 mg/sqm IV for 5-days, every 28 days. The median age was 72.5 years (74 in cohort 1 and 66 in cohort 2) and 16% of pts had an ECOG performance status >2 at the start of DAC treatment (with non-significant difference in the two cohorts). The cumulative illness rating scale (CIRS) was > 6 in 27% of pts. Forty-five pts (43%) had secondary AML. Bone marrow blast count was > 30% in 64% of patients (67/104). In the relapsed cohort 17/29 (59%) patients were treated with DAC after conventional CHT, 5/29 (17%) after allo-SCT and 7/29 (24%) after azacitidine therapy. RESULTS A total of 469 DAC cycles were given to the 104 pts with a median of 3 cycles (range 1-21) and 45/104 (43%) pts received > 4 cycles. The Overall Response Rate (ORR = Complete Remission-CR plus Partial Remission-PR) was 33%, significantly higher in Cohort 1 (42%) compared to Cohort 2 (14%) (p = 0.009). The median duration of response was 6 months (range 1-20). In Cohort 1 the best response (CR or PR) was obtained between 3th and 6th cycle. In multivariate Cox regression analysis, achievement of CR or PR (HR = 0.78; p = 0.0004), CIRS < 6 (HR = 0.9; p = 0.04) and complex karyotype (HR = 0.8; p = 0.03) were significant predictors of better overall survival (OS). Median OS from the start of DAC therapy was 11 months for the whole population with a significant OS advantage in Cohort 1 (median OS 12.7 mths vs 6.3 mths; p = 0.003); median OS was significantly longer in responders compared to non-responders (22.6 mths vs 5.7 mths; p < 0.0001). At the last follow-up, 56 patients (54%) are still alive and 48 (46%) are dead (71% due to disease progression). The most common toxicities were myelosuppression and documented infectious complications that occurred mainly during the first 4 cycles. CONCLUSION These data confirm the efficacy (ORR 33%) and the acceptable safety profile of DAC in the real life management of AML in elderly pts unsuitable for intensive CHT, with a significant better performance in first line therapy (ORR 42%, median OS 12.7 mths). The efficacy of DAC, both in first line and as salvage therapy, may probably be improved with combined treatment strategies and/or with different DAC schedules that could increase its anti-leukemic effect.

Mechanisms-of-resistance to decitabine and 5-azacytidine, mainstay treatments for myeloid malignancies, require investigation and countermeasures. Both are nucleoside analog pro-drugs processed by pyrimidine metabolism into a deoxynucleotide analog that depletes the key epigenetic regulator DNA methyltranseferase 1 (DNMT1). Here, upon serial analyses of DNMT1 levels in patients’ bone marrows on-therapy, we found DNMT1 was not depleted at relapse. Showing why, bone marrows at relapse exhibited shifts in expression of key pyrimidine metabolism enzymes in directions adverse to pro-drug activation. Further investigation revealed the origin of these shifts. Pyrimidine metabolism is a network that senses and regulates deoxynucleotide amounts. Deoxynucleotide amounts were disturbed by single exposures to decitabine or 5-azacytidine, via off-target depletion of thymidylate synthase and ribonucleotide reductase respectively. Compensating pyrimidine metabolism shifts peaked 72–96 h later. Continuous pro-drug exposures stabilized these adaptive metabolic responses to thereby prevent DNMT1-depletion and permit exponential leukemia out-growth as soon as day 40. The consistency of the acute metabolic responses enabled exploitation: simple treatment modifications in xenotransplant models of chemorefractory leukemia extended noncytotoxic DNMT1-depletion and leukemia control by several months. In sum, resistance to decitabine and 5-azacytidine originates from adaptive responses of the pyrimidine metabolism network; these responses can be anticipated and thus exploited.

Treatment of relapsed or refractory acute myeloid leukemia (AML) has presented challenges for hematologists for decades. Despite numerous clinical studies, outcomes are consistently disappointing with 5-year overall survival rates of approximately 10%. Allogeneic hematopoietic cell transplantation at the time of second complete remission remains the only reliable option with curative potential. However, recent approval of several new agents has transformed treatment paradigms in AML that had been in place for almost half a century. This new therapeutic landscape provides the opportunity to revisit the approach to relapsed or refractory AML. Through illustrative cases we describe our approach, which increasingly relies on specific disease biology. We focus on treatment outside the context of clinical trials since such trials are not available in most parts of the world. Primarily we consider age, fitness to tolerate intensive chemotherapy, remission duration, and presence of a targetable mutation to guide treatment. It is inevitable that the coming years will bring new targets and agents which may prove most effective when combined with each other and/or chemotherapy. Future studies are needed to determine how best to implement this evolving armamentarium of treatment options, to elucidate mechanisms of resistance, and to continue to pursue novel drug discovery.