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Supplementary Figure 2. MPFC detection limit.

Supplementary Table 3. MPFC analysis of total AML bone marrow sample according to FLT3 mutational status.

Supplementary Table 4. MPFC analysis of AML bone marrow sample according to FLT3 or NPM1 mutational status.

Supplementary Table 1. anti-human antibodies used.

Supplementary Table 5. Longitudinal analysis of FLT3-ITDs using patient-specific RQPCR in four AML patients.

Supplementary Figure 3. MPFC and molecular analysis of UPN 241 bone marrow samples at diagnosis, during follow-up and at relapse.

Purpose: We evaluated leukemia-associated immunophenotypes (LAIP) and their correlation with fms-like tyrosine kinase 3 (FLT3) and nucleophosmin (NPM1) gene mutational status in order to contribute a better identification of patients at highest risk of relapse in acute myeloid leukemia (AML).Experimental Design: Bone marrow samples from 132 patients with AML were analyzed by nine-color multiparametric flow cytometry. We confirmed the presence of the mutation in diagnostic samples and in sorted cells by conventional RT-PCR and by patient-specific RQ-PCR.Results: Within the CD34+ cell fraction, we identified a discrete population expressing high levels of the IL3 receptor α-chain (CD123) and MIC-2 (CD99) in combination with the IL2 receptor α-chain (CD25). The presence of this population positively correlated with the internal tandem duplications (ITD) mutation in the FLT3 gene (r = 0.71). Receiver operating characteristics showed that, within the CD34+ cell fraction a percentage of CD123/CD99/CD25+ cells ≥11.7% predicted FLT3–ITD mutations with a specificity and sensitivity of >90%. CD34/CD123/CD99/CD25+ clones were also detectable at presentation in 3 patients with FLT3 wild-type/NPM1+ AML who relapsed with FLT3-ITD/NPM1+ AML. Quantitative real-time PCR designed at relapse for each FLT3-ITD in these three cases confirmed the presence of low copy numbers of the mutation in diagnostic samples.Conclusions: Our results suggest that the CD34/CD25/CD123/CD99+ LAIP is strictly associated with FLT3-ITD–positive cells. Clin Cancer Res; 21(17); 3977–85. ©2015 AACR.

Supplementary Table 1 from Sequential Valproic Acid/All-trans Retinoic Acid Treatment Reprograms Differentiation in Refractory and High-Risk Acute Myeloid Leukemia

Epigenetic alterations of chromatin due to aberrant histone deacetylase (HDAC) activity and transcriptional silencing of all-trans retinoic acid (ATRA) pathway are events linked to the pathogenesis of acute myeloid leukemia (AML) that can be targeted by specific treatments. A pilot study was carried out in eight refractory or high-risk AML patients not eligible for intensive therapy to assess the biological and therapeutic activities of the HDAC inhibitor valproic acid (VPA) used to remodel chromatin, followed by the addition of ATRA, to activate gene transcription and differentiation in leukemic cells. Hyperacetylation of histones H3 and H4 was detectable at therapeutic VPA serum levels (≥50 μg/mL) in blood mononuclear cells from seven of eight patients. This correlated with myelomonocytic differentiation of leukemic cells as revealed by morphologic, cytochemical, immunophenotypic, and gene expression analyses. Differentiation of the leukemic clone was proven by fluorescence in situ hybridization analysis showing the cytogenetic lesion +8 or 7q− in differentiating cells. Hematologic improvement, according to established criteria for myelodysplastic syndromes, was observed in two cases. Stable disease and disease progression were observed in five and one cases, respectively. In conclusion, VPA-ATRA treatment is well tolerated and induces phenotypic changes of AML blasts through chromatin remodeling. Further studies are needed to evaluate whether VPA-ATRA treatment by reprogramming differentiation of the leukemic clone might improve the response to chemotherapy in leukemia patients. (Cancer Res 2006; 66(17): 8903-11)

Background The ZBTB16‐RARA fusion gene, resulting from the reciprocal translocation between ZBTB16 on chromosome 11 and RARA genes on chromosome 17 [t(11;17)(q23;q21)], is rarely observed in acute myeloid leukemia (AML), and accounts for about 1% of retinoic acid receptor‐α (RARA) rearrangements. AML with this rare translocation shows unusual bone marrow (BM) morphology, with intermediate aspects between acute promyelocytic leukemia (APL) and AML with maturation. Patients may have a high incidence of disseminated intravascular coagulation at diagnosis, are poorly responsive to all‐trans retinoic acid (ATRA) and arsenic tryoxyde, and are reported to have an overall poor prognosis. Aims The mutational profile of ZBTB16‐RARA rearranged AML has not been described so far. Materials and methods We performed targeted next‐generation sequencing of 24 myeloid genes in BM diagnostic samples from seven ZBTB16‐RARA+AML, 103 non‐RARA rearranged AML, and 46 APL. The seven ZBTB16‐RARA‐positive patients were then screened for additional mutations using whole exome sequencing (n = 3) or an extended cancer panel including 409 genes (n = 4). Results ZBTB16‐RARA+AML showed an intermediate number of mutations per patient and involvement of different genes, as compared to APL and other AMLs. In particular, we found a high incidence of ARID1A mutations in ZBTB16‐RARA+AML (five of seven cases, 71%). Mutations in ARID2 and SMARCA4, other tumor suppressor genes also belonging to SWI/SNF chromatin remodeling complexes, were also identified in one case (14%). Discussion and conclusion Our data suggest the association of mutations of the ARID1A gene and of the other members of the SWI/SNF chromatin remodeling complexes with ZBTB16‐RARA+AMLs, where they may support the peculiar disease phenotype., This is the first report showing the mutational landscape of the rare ZBTB16‐RARA rearranged acute myeloid leukemia (AML). We identified a high frequency of ARID1A mutations, suggesting the involvement of the SWI/SNF chromatin remodeling complexes in the clinical presentation, acute promyelocytic leukemia‐like morphology and all‐trans retinoic acid/arsenic trioxide treatment resistance of ZBTB16‐RARA rearranged AMLs.

Obesity correlates with hematologic malignancies including leukemias, but risk of specific leukemia subtypes like acute promyelocytic leukemia and underlying molecular mechanisms are poorly understood. We explored multiple datasets for correlation between leukemia, body mass index (BMI) and molecular features. In a population-based study (n=5.2 million), we correlated BMI with promyelocytic leukemia, and other acute myeloid, lymphoid or other leukemias. In cross-sectional studies, we tested BMI deviation in promyelocytic leukemia trial cohorts from that expected based on national surveys. We explored The Cancer Genome Atlas for transcriptional signatures and mutations enriched in promyelocytic leukemia and/or obesity, and confirmed a correlation between body mass and FLT3 mutations in promyelocytic leukemia cohorts by logistic regression. In the population-based study, hazard ratio per 5 kg/m2 increase was: promyelocytic leukemia 1.44 (95%CI: 1.0-2.08), non-promyelocytic acute myeloid leukemias 1.17 (95%CI: 1.10-1.26), lymphoid leukemias 1.04 (95%CI: 1.0-1.09), other 1.10 (95%CI: 1.04-1.15). In cross-sectional studies, body mass deviated significantly from that expected (Italy: P

Acute myeloid leukemia (AML) is a hematological malignancy with an undefined heritable risk. Here we perform a meta-analysis of three genome-wide association studies, with replication in a fourth study, incorporating a total of 4018 AML cases and 10488 controls. We identify a genome-wide significant risk locus for AML at 11q13.2 (rs4930561; P = 2.15 × 10

Acute myeloid leukemia (AML) is a hematological malignancy with an undefined heritable risk. Here we perform a meta-analysis of three genome-wide association studies, with replication in a fourth study, incorporating a total of 4018 AML cases and 10488 controls. We identify a genome-wide significant risk locus for AML at 11q13.2 (rs4930561; P = 2.15 × 10−8; KMT5B). We also identify a genome-wide significant risk locus for the cytogenetically normal AML sub-group (N = 1287) at 6p21.32 (rs3916765; P = 1.51 × 10−10; HLA). Our results inform on AML etiology and identify putative functional genes operating in histone methylation (KMT5B) and immune function (HLA)., Genome wide association studies in cancer are used to understand the heritable genetic contribution to disease risk. Here, the authors perform a genome wide association study in European patients with acute myeloid leukemia and identify loci associated with risk of developing the disease.

Acute myeloid leukemia (AML) is a hematological malignancy with an undefined heritable risk. Here we performed a meta-analysis of three genome-wide association studies, with replication in a fourth study, incorporating a total of 4018 AML cases and 10488 controls. We identified a genome-wide significant risk locus for AML at 11q13.2 (rs4930561; P = 2.15 × 10-8; KMT5B). We also identified a genome-wide significant risk locus for the cytogenetically normal AML sub-group (N=1287) at 6p21.32 (rs3916765; P = 1.51 × 10-10; HLA). Our results inform on AML etiology by identifying putative functional genes operating in histone methylation (KMT5B) and immune function (HLA).

Despite the high probability of cure of patients with acute promyelocytic leukemia (APL), mechanisms of relapse are still largely unclear. Mutational profiling at diagnosis and/or relapse may help to identify APL patients needing frequent molecular monitoring and early treatment intervention. Using an NGS approach including a 31 myeloid gene-panel, we tested BM samples of 44 APLs at the time of diagnosis, and of 31 at relapse. Mutations in PML and RARA genes were studied using a customized-NGS-RNA panel. Patients relapsing after ATRA-chemotherapy rarely had additional mutations (P = .009). In patients relapsing after ATRA/ATO, the PML gene was a preferential mutation target. We then evaluated the predictive value of mutations at APL diagnosis. A median of two mutations was detectable in 9/11 patients who later relapsed, vs one mutation in 21/33 patients who remained in CCR (P = .0032). This corresponded to a significantly lower risk of relapse in patients with one or less mutations (HR 0.046; 95% CI 0.011-0.197; P < .0001). NGS-analysis at the time of APL diagnosis may inform treatment decisions, including alternative treatments for cases with an unfavorable mutation profile.

We present a case report of a patient with acute myeloid leukemia (AML) characterized by the simultaneous presence of nucleophosmin 1 (NPM1) mutation and the breakpoint cluster region-Abelson (BCR-ABL) fusion oncogene. Our findings emphasize the importance of routinely including BCR-ABL in the diagnostic workup of AML in order to offer to the patients the most appropriate risk category and treatment options.

Since the comprehensive recommendations for the management of acute promyelocytic leukemia (APL) reported in 2009, several studies have provided important insights, particularly regarding the role of arsenic trioxide (ATO) in frontline therapy. Ten years later, a European LeukemiaNet expert panel has reviewed the recent advances in the management of APL in both frontline and relapse settings in order to develop updated evidence- and expert opinion–based recommendations on the management of this disease. Together with providing current indications on genetic diagnosis, modern risk-adapted frontline therapy, and salvage treatment, the review contains specific recommendations for the identification and management of the most important complications such as the bleeding disorder APL differentiation syndrome, QT prolongation, and other all-trans retinoic acid– and ATO-related toxicities, as well as recommendations for molecular assessment of the response to treatment. Finally, the approach to special situations is also discussed, including management of APL in children, elderly patients, and pregnant women. The most important challenges remaining in APL include early death, which still occurs before and during induction therapy, and optimizing treatment in patients with high-risk disease.

Acute myeloid leukemia (AML) with FLT3-ITD mutations (FLT3-ITDmut) remains a therapeutic challenge, with a still high relapse rate, despite targeted treatment with tyrosine kinase inhibitors. In this disease, the CD34/CD123/CD25/CD99+ leukemic precursor cells (LPCs) phenotype predicts for FLT3-ITD-positivity. The aim of this study was to characterize the distribution of FLT3-ITD mutation in different progenitor cell subsets to shed light on the subclonal architecture of FLT3-ITDmut AML. Using high-speed cell sorting, we sequentially purified LPCs and CD34+ progenitors in samples from patients with FLT3-ITDmut AML (n = 12). A higher FLT3-ITDmut load was observed within CD34/CD123/CD25/CD99+ LPCs, as compared to CD34+ progenitors (CD123+/−,CD25−,CD99low/−) (p = 0.0005) and mononuclear cells (MNCs) (p FLT3-ITDmut burden was also observed in LPCs of AML patients with a small FLT3-ITDmut clones at diagnosis. On the contrary, the mutation burden of other myeloid genes was similar in MNCs, highly purified LPCs and/or CD34+ progenitors. Treatment with an anti-CD99 mAb was cytotoxic on LPCs in two patients, whereas there was no effect on CD34+ cells from healthy donors. Our study shows that FLT3-ITD mutations occur early in LPCs, which represent the leukemic reservoir. CD99 may represent a new therapeutic target in FLT3-ITDmut AML.

Acute promyelocytic leukaemia (APL) represents a modern success of precision medicine. However, fatalities occurring within the first 30 days of induction treatment, in particular intracranial haemorrhage (ICH), remain the main causes of death. We studied the clinico-biological characteristics of 13 patients with APL who experienced ICH. Compared to 85 patients without this complication, patients with ICH were older and more frequently had high-risk APL. Moreover, positivity for the 'swirl' sign at neuroradiological imaging and hydrocephalus were predictors of a fatal outcome, together with lower fibrinogen, prolonged international normalized ratio (INR) and higher lactate dehydrogenase levels.

Non-T cell activation linker (NTAL) is a lipid raft-membrane protein expressed by normal and leukemic cells and involved in cell signaling. In acute promyelocytic leukemia (APL), NTAL depletion from lipid rafts decreases cell viability through regulation of the Akt/PI3K pathway. The role of NTAL in APL cell processes, and its association with clinical outcome, has not, however, been established. Here, we show that reduced levels of NTAL were associated with increased all-trans retinoic acid (ATRA)-induced differentiation, generation of reactive oxygen species, and mitochondrial dysfunction. Additionally, NTAL-knockdown (NTAL-KD) in APL cell lines led to activation of Ras, inhibition of Akt/mTOR pathways, and increased expression of autophagy markers, leading to an increased apoptosis rate following arsenic trioxide treatment. Furthermore, NTAL-KD in NB4 cells decreased the tumor burden in (NOD scid gamma) NSG mice, suggesting its implication in tumor growth. A retrospective analysis of NTAL expression in a cohort of patients treated with ATRA and anthracyclines, revealed that NTAL overexpression was associated with a high leukocyte count (P = 0.007) and was independently associated with shorter overall survival (Hazard Ratio: 3.6; 95% Confidence Interval: 1.17–11.28; P = 0.026). Taken together, our data highlights the importance of NTAL in APL cell survival and response to treatment.

The results from the RATIFY trial (ClinicalTrials.gov: NCT00651261; CALGB 10603) showed that midostaurin combined with standard chemotherapy significantly improved outcomes in patients with FMS-like tyrosine kinase 3 (FLT3)–mutated acute myeloid leukemia (AML), compared with placebo. In this post hoc subgroup analysis from the trial, we evaluated the impact of midostaurin in 163 patients with FLT3-tyrosine kinase domain (TKD) mutations. At a median follow-up of 60.7 months (95% CI, 55.0-70.8), the 5-year event-free survival (EFS) rate was significantly higher in patients treated with midostaurin than in those treated with placebo (45.2% vs 30.1%; P = .044). A trend toward improved disease-free survival was also observed with midostaurin (67.3% vs 53.4%; P = .089), whereas overall survival (OS) was similar in the 2 groups. Patients with AML and NPM1mut/FLT3-TKDmut or core binding factor (CBF)–rearranged/FLT3-TKDmut genotypes had significantly prolonged OS with or without censoring at hematopoietic cell transplantation (HCT), compared with NPM1WT/CBF-negative AMLs. The multivariable model for OS and EFS adjusted for allogeneic HCT in first complete remission as a time-dependent covariable, revealed NPM1 mutations and CBF rearrangements as significant favorable factors. These data show that NPM1 mutations or CBF rearrangements identify favorable prognostic groups in patients with FLT3-TKD AMLs, independent of other factors, also in the context of midostaurin treatment.

Acute promyelocytic leukemia (APL) is a subtype of acute leukemia characterized by a unique t(15;17) translocation generating the PML/RARA fusion gene and hybrid oncoprotein. Besides its critical role in leukemogenesis, this genetic aberration serves as a disease-specific biomarker for rapid diagnosis and monitoring of minimal residual disease (MRD). Moreover, PML/RARA is specifically targeted by All-trans retinoic acid (ATRA) and arsenic trioxide (ATO), two agents that synergistically act to induce degradation of the oncoprotein. Large clinical studies including two randomized trials conducted in newly diagnosed APL patients have shown that the ATRA-ATO combination is superior to conventional ATRA and chemotherapy both in terms of efficacy and safety. Preliminary studies using oral formulations of arsenic and ATRA suggest that oral arsenic is as effective and manageable as intravenous ATO. Following early retrospective studies indicating the prognostic relevance of PML/RARA monitoring, several prospective studies were conducted in large cohorts of APL patients enrolled in clinical trials with the aim of better assessing the prognostic value of longitudinal PCR testing. The results consistently showed that molecular remission (defined as negativization of the PCR test for PML/RARA) correlates with a significantly decreased risk of relapse, whereas persistence of PCR positivity for PML/RARA after consolidation or conversion from negative to positive during follow-up is strongly associated with hematologic relapse. Based on these data, various groups started using pre-emptive salvage therapy for patients who persisted PCR-positive after frontline consolidation or converted from negative to positive PCR during follow-up. Finally, several expert panels have recommended that molecular remission should be considered a therapeutic objective in APL, and molecular response has been adopted as a study endpoint in modern clinical trials.

Prolonged therapy with all-trans-retinoic acid (ATRA) and arsenic trioxide (ATO) is highly effective in newly diagnosed acute promyelocytic leukemia (APL) but there is limited data on the efficacy of this regimen in the relapse setting. We report here on 22 APL patients treated with prolonged ATRA-ATO therapy at the time of disease relapse. Twenty patients obtained molecular complete remission (CRm) after 2 cycles (90%). Of these, two patients underwent hematopoietic stem cell transplant (HSCT) while the remaining proceeded to receive additional cycles (up to a total of 5) of ATRA-ATO. With a median follow-up of 58 months from the time of relapse (range: 21-128 months), the 4-year OS probability was 0.85 (95% CI 0.61-0.94), DFS was 0.74 (95% CI 0.49-0.88), and EFS 0.68 (95% CI 0.45-0.83). Two patients were resistant to ATRA-ATO salvage and five relapsed at a median of 19 months. Of these, four died due to progressive disease while three relapsed achieved a new CRm after further salvage therapy. This experience confirms the potentially curative effect of prolonged ATRA-ATO therapy in relapsed APL, especially in patients with long first complete remission.

Response to therapy is affected by the genetic heterogeneity of acute myeloid leukemia (AML) and persistence of leukemic cells below the threshold of morphological complete remission (mCR). Such persistence is called minimal (or measurable) residual disease (MRD). Areas covered: MRD assessment allows early identification of patients who are at high risk of relapse and who should timely receive aggressive therapy (e.g. allogeneic stem cell transplantation) and of those with a good quality mCR in whom an aggressive front-line therapy can be spared, avoiding the harm of excessive treatment toxicity. The most exploited methods to assess MRD are multiparameter flow cytometry (via identification of immunophenotypic aberrancies) or PCR-based assays (via identification of cytogenetic/molecular abnormalities). Expert commentary: A growing body of evidences demonstrates that positive MRD-testing at various time-points throughout the treatment course identifies patients at high risk of relapse. We will focus on the role of MRD as a biomarker to refine risk assessment and to prospectively direct treatment choices in adult with AML.

Vitamin C (Vit C or Ascorbate) is essential for many fundamental biochemical processes. Vit C is an essential nutrient with redox functions at normal physiologic concentrations. The main physiologic function of this vitamin is related to its capacity to act as a co-factor for a large family of enzymes, collectively known as Fe and 2-oxoglutarate-dependent dioxygenases. It also modulates epigenetic gene expression through the control of TET enzymes activity. Vit C also has several biological properties allowing to restore the deregulated epigenetic response observed in many tumors. High-dose Vit C has been investigated as a treatment for cancer patients since the 1969. Pharmacologic ascorbate acts as a pro-drug for hydrogen peroxide formation (H2O2) and, through this mechanism, kills cancer cells. To achieve high in vivo concentrations, Ascorbate must be injected by i.v. route. Initial clinical studies of Ascorbate cancer treatment have provided encouraging results, not confirmed in subsequent studies. Recent clinical studies using i.v. injection of high-dose Ascorbate have renewed the interest in the field, showing that significant anti-tumor activity. Pre-clinical studies have led to identify tumors sensitive to Ascorbate that could potentially benefit from this treatment either through an epigenetic modulator effect or through tumor killing by oxidative stress.

// Emiliano Fabiani 1, * , Giulia Falconi 1, * , Nelida Ines Noguera 1, 2 , Ernestina Saulle 3 , Laura Cicconi 1 , Mariadomenica Divona 1 , Cristina Banella 2 , Alessandra Picardi 1 , Anna Maria Cerio 4 , Letizia Boe 5 , Massimo Sanchez 5 , Elvira Pelosi 4 , Ugo Testa 4 , Francesco Lo-Coco 1, 2 and Maria Teresa Voso 1 1 Universita di Roma Tor Vergata, Dipartimento di Biomedicina e Prevenzione, Rome, Italy 2 Fondazione Santa Lucia, Laboratorio di Neuro-Oncoematologia, Rome, Italy 3 Istituto Superiore di Sanita, Centro Nazionale per la Ricerca e la Valutazione Preclinica e Clinica dei Farmaci, Rome, Italy 4 Istituto Superiore di Sanita, Dipartimento di Ematologia ed Oncologia, Rome, Italy 5 Istituto Superiore di Sanita, Grandi Strumentazioni e Core Facilities, Rome, Italy * These authors contributed equally to this work Correspondence to: Maria Teresa Voso, email: voso@med.uniroma2.it Keywords: acute promyelocytic leukemia, forkhead box C1, ATRA, decitabine, epigenetic regulation Received: April 27, 2017 Accepted: August 31, 2017 Published: September 20, 2017 ABSTRACT Forkhead box (FOX) genes encode transcription factors, which regulate embryogenesis and play an important role in hematopoietic differentiation and in mesenchymal niche maintenance. Overexpression of the family member FOXC1 has been reported in solid tumors and acute myeloid leukemia (AML). We studied FOXC1 expression and function in acute promyelocytic leukemia (APL) and normal hematopoietic progenitors. FOXC1 mRNA and protein levels were significantly lower in primary marrow samples from 27 APL patients, as compared to samples obtained from 27 patients with other AML subtypes, and 5 normal CD34+ hematopoietic cells. FOXC1 expression significantly increased in APL samples at the time of remission following consolidation treatment. In cell lines overexpressing PML-RARA, and in the NB4 t(15;17)-positive cell line, FOXC1 expression was lower than in other non-APL cell lines, and increased following treatment with all-trans retinoic acid (ATRA), due to functional binding of ATRA to the FOXC1 promoter region. Reduced FOXC1 expression was also associated to DNA hypermethylation of the +354 to +568 FOXC1 region, both in primary APL, and in NB4 cells. Treatment of NB4 cells with decitabine demethylated FOXC1 and upregulated its expression. Our findings indicate that FOXC1 is consistently repressed in APL due to hypermethylation and the presence of the PML-RARA rearrangement. A potential role of hypomethylating treatment in advanced APL remains to be established.

Purpose The initial results of the APL0406 trial showed that the combination of all- trans-retinoic acid (ATRA) and arsenic trioxide (ATO) is at least not inferior to standard ATRA and chemotherapy (CHT) in first-line therapy of low- or intermediate-risk acute promyelocytic leukemia (APL). We herein report the final analysis on the complete series of patients enrolled onto this trial. Patients and Methods The APL0406 study was a prospective, randomized, multicenter, open-label, phase III noninferiority trial. Eligible patients were adults between 18 and 71 years of age with newly diagnosed, low- or intermediate-risk APL (WBC at diagnosis ≤ 10 × 109/L). Overall, 276 patients were randomly assigned to receive ATRA-ATO or ATRA-CHT between October 2007 and January 2013. Results Of 263 patients evaluable for response to induction, 127 (100%) of 127 patients and 132 (97%) of 136 patients achieved complete remission (CR) in the ATRA-ATO and ATRA-CHT arms, respectively ( P = .12). After a median follow-up of 40.6 months, the event-free survival, cumulative incidence of relapse, and overall survival at 50 months for patients in the ATRA-ATO versus ATRA-CHT arms were 97.3% v 80%, 1.9% v 13.9%, and 99.2% v 92.6%, respectively ( P < .001, P = .0013, and P = .0073, respectively). Postinduction events included two relapses and one death in CR in the ATRA-ATO arm and two instances of molecular resistance after third consolidation, 15 relapses, and five deaths in CR in the ATRA-CHT arm. Two patients in the ATRA-CHT arm developed a therapy-related myeloid neoplasm. Conclusion These results show that the advantages of ATRA-ATO over ATRA-CHT increase over time and that there is significantly greater and more sustained antileukemic efficacy of ATO-ATRA compared with ATRA-CHT in low- and intermediate-risk APL.

Francesco Lo-Coco speaks to Sebastian Dennis-Beron, Commissioning Editor: Francesco Lo-Coco is currently a full professor of hematology and head of the Laboratory of Integrated Diagnosis of Oncohematologic Diseases at the Department of Biomedicine and Prevention of the University Tor Vergata of Roma. He obtained his MD degree from the University of Pisa in 1981, and his specialization in clinical and laboratory hematology is from the University La Sapienza of Rome in 1985. From 1992 to 1994, he has trained on molecular genetics of lymphomas at Columbia University, New York. His main scientific interest and research activities include genetic characterization, monitoring and treatment of hematologic tumors, particularly acute myeloid leukemia and acute promyelocytic leukemia (APL). He has published over 390 internationally peer-reviewed articles, mainly focused on molecular diagnosis and follow-up of leukemia as well as on treatment of APL. He chairs at present the APL subcommittee of the Italian National Cooperative Group, GIMEMA. He served as president of the Italian Society of Experimental Hematology, chairman of the Education Committee of the European Hematology Association, board member of the Italian Foundation for Cancer Research, member of the Committe on Health Research of the Italian Ministry of Health and member of the Editorial Board of the journals Leukemia, Journal of Clinical Oncology and Haematologica.

In the context of precision medicine, assessment of minimal residual disease (MRD) has been used in acute myeloid leukemia (AML) to direct individual treatment programs, including allogeneic stem cell transplantation in patients at high-risk of relapse. One of the limits of this approach has been in the past the paucity of AML markers suitable for MRD assessment. Recently, the number of biomarkers has increased, due to the identification of highly specific leukemia-associated immunophenotypes by multicolor flow-cytometry, and of rare mutated gene sequences by digital droplet PCR, or next-generation sequencing (NGS). In addition, NGS allowed unraveling of clonal heterogeneity, present in AML at initial diagnosis or developing during treatment, which influences reliability of specific biomarkers, that may be unstable during the disease course. The technological advances have increased the application of MRD-based strategies to a significantly higher number of AML patients, and the information deriving from MRD assessment has been used to design individual post-remission protocols and pre-emptive treatments in patients with sub-clinical relapse. This led to the definition of MRD-negative complete remission as outcome definition in the recently published European Leukemianet MRD guidelines. In this review, we summarized the principles of modern technologies and their clinical applications for MRD detection in AML patients, according to the specific leukemic markers.

Although targeted therapies have proven effective and even curative in human leukaemia, resistance often ensues. IDH enzymes are mutated in similar to 20% of human AML, with targeted therapies under clinical evaluation. We here characterize leukaemia evolution from mutant IDH2 (mIDH2)-dependence to independence identifying key targetable vulnerabilities of mIDH2 leukaemia that are retained during evolution and progression from early to late stages. Mechanistically, we find that mIDH2 leukaemia are metastable and vulnerable at two distinct levels. On the one hand, they are characterized by oxidative and genotoxic stress, in spite of increased 1-carbon metabolism and glutathione levels. On the other hand, mIDH2 leukaemia display inhibition of LSD1 and a resulting transcriptional signature of all-trans retinoic acid (ATRA) sensitization, in spite of a state of suppressed ATRA signalling due to increased levels of PIN1. We further identify GSH/ROS and PIN1/LSD1 as critical nodes for leukaemia maintenance and the combination of ATRA and arsenic trioxide (ATO) as a key therapeutic modality to target these vulnerabilities. Strikingly, we demonstrate that the combination of ATRA and ATO proves to be a powerfully synergistic and effective therapy in a number of mouse and human mIDH1/2 leukemic models. Thus, our findings pave the way towards the treatment of a sizable fraction of human AMLs through targeted APL-like combinatorial therapies.

Arsenic trioxide and all-trans retinoic acid have become the frontline treatments for patients with acute promyelocytic leukemia (APL). Despite the long wait for an oral arsenic drug, a commercially available agent, realgar–indigo naturalis formula (RIF), was not launched in China until 2009. Since then, over 5000 APL patients have been treated with oral RIF in China. Oral arsenic not only shows a clinical efficacy comparable to that of IV formulations but also displays a better safety profile, improved quality of life, and lower medical costs for patients. The promising results promote incorporating an outpatient postremission therapy model into clinical practice for both low-risk and high-risk APL patients in China. In this review, we discuss the evolution of oral arsenic RIF in the treatment of APL, with a special focus on how to address the related complications during induction therapy.

Acute myeloid leukaemia (AML) is a highly heterogeneous disease characterized by uncontrolled proliferation, block in myeloid differentiation and recurrent genetic abnormalities. In the search of new effective therapies, identification of synthetic lethal partners of AML genetic alterations might represent a suitable approach to tailor patient treatment. Genetic mutations directly affecting DNA repair genes are not commonly present in AML. Nevertheless, several studies indicate that AML cells show high levels of DNA lesions and genomic instability. Leukaemia-driving oncogenes (e.g., RUNX1-RUNXT1, PML-RARA, TCF3-HLF, IDH1/2, TET2) or treatment with targeted agents directed against aberrant kinases (e.g., JAK1/2 and FLT3 inhibitors) have been associated with reduced DNA repair gene expression/activity that would render leukaemia blasts selectively sensitive to synthetic lethality induced by poly(ADP-ribose) polymerase inhibitors (PARPi). Thus, specific oncogenic chimeric proteins or gene mutations, rare or typically distinctive of certain leukaemia subtypes, may allow tagging cancer cells for destruction by PARPi. In this review, we will discuss the rationale for using PARPi in AML subtypes characterized by a specific genetic background and summarize the preclinical and clinical evidence reported so far on their activity when used as single agents or in combination with classical cytotoxic chemotherapy or with agents targeting AML-associated mutated proteins.

Malignant disease and its treatment carry huge burdens for patients. Some are immediate, in that the disease itself presents as a life threatening event, or the treatment may result in immediate and devastating toxicity. More often the treatment of cancer is associated with more subtle or late events, yet these may impact the quality of life for cancer survivors in a variety of ways. In addition to the physical sequelae of cancer or its treatment, cancer survivors often experience consequences in the form of social or mental incapacity. Session III of this Colloquium on Cardio-Oncology focuses on some of these concerns, both from the perspective of health care providers who strive to minimize the burdens, but also from the viewpoint of the patient him or herself who must deal with the price that must often be paid for increased survival or cure.

The objective of this study was to investigate health-related quality of life (HRQOL), symptom burden, and comorbidity profile in long-term acute promyelocytic leukemia (APL) survivors treated with standard chemotherapy. Overall, 307 long-term APL survivors were invited to participate. HRQOL was assessed with the Medical Outcomes Study 36-Item Short-Form Health Survey (SF-36) and compared with that of age and sex-matched controls from the general population. Symptom burden was assessed with the MD Anderson Symptom Inventory (MDASI) questionnaire and comorbidity profile was also investigated. Median follow-up time since diagnosis was 14.3 years (interquartile range: 11.1-16.9 years). APL survivors had a statistically and clinically meaningful worse score for the role physical scale of the SF-36 (-9.5; 95% CI, -15.7 to -3.2, P = 0.003) than their peers in the general population. Fatigue was reported as moderate to severe by 29% of patients and 84.4% reported at least one comorbidity. Prevalence of comorbidity in APL survivors was higher than that reported by the general population. Also, marked variations were found in the HRQOL profile by number of comorbidities. Even many years after treatment ends, APL survivors treated with standard chemotherapy do not fully recover as they report HRQOL limitations and a substantial burden of symptoms.