Paola Minetto

Bloodstream Infections Are an Improbable Cause of Positive Serum (1,3)-β-d-Glucan in Hematology Patients

ABSTRACT Ninety-one serum samples from 51 hematology patients with bacteremia infections were tested for (1,3)-β-d-glucan (BG). Eleven samples (15%) from 7 patients (14%) were positive for BG. Of these 7 patients with positive BG results, 4 (8%) had invasive aspergillosis and 3 (6%) had no invasive fungal disease. Bacteremia was an unlikely cause of the false-positive BG results.

High feasibility and antileukemic efficacy of fludarabine, cytarabine, and idarubicin (FLAI) induction followed by risk-oriented consolidation: A critical review of a 10-year, single-center experience in younger, non M3 AML patients.

About 105 consecutive acute myeloid leukemia (AML) patients treated with the same induction‐consolidation program between 2004 and 2013 were retrospectively analyzed. Median age was 47 years. The first induction course included fludarabine (Flu) and high‐dose cytarabine (Ara‐C) plus idarubicin (Ida), with or without gemtuzumab‐ozogamicin (GO) 3 mg/m2 (FLAI‐5). Patients achieving complete remission (CR) received a second course without fludarabine but with higher dose of idarubicin. Patients not achieving CR received an intensified second course. Patients not scheduled for early allogeneic bone marrow transplantation (HSCT) where planned to receive at least two courses of consolidation therapy with Ara‐C. Our double induction strategy significantly differs from described fludarabine‐containing regimens, as patients achieving CR receive a second course without fludarabine, to avoid excess toxicity, and Ara‐C consolidation is administrated at the reduced cumulative dose of 8 g/m2 per cycle. Toxicity is a major concern in fludarabine containing induction, including the recent Medical Research Council AML15 fludarabine, cytarabine, idaraubicin and G‐CSF (FLAG‐Ida) arm, and, despite higher anti‐leukemic efficacy, only a minority of patients is able to complete the full planned program. In this article, we show that our therapeutic program is generally well tolerated, as most patients were able to receive subsequent therapy at full dose and in a timely manner, with a 30‐day mortality of 4.8%. The omission of fludarabine in the second course did not reduce efficacy, as a CR rate of 83% was achieved and 3‐year disease‐free survival and overall survival (OS) were 49.6% and 50.9%, respectively. Our experience shows that FLAI‐5/Ara‐C + Ida double induction followed by risk‐oriented consolidation therapy can result in good overall outcome with acceptable toxicity. Am. J. Hematol. 91:755–762, 2016.

Cardiovascular adverse events complicating the administration of rituximab: report of two cases.

Rituximab is a murine/human chimeric monoclonal antibody directed against the CD20 antigen. It is widely used in combination with polychemotherapy regimens for the treatment of hematological disorders. There is no evidence of direct cardiotoxicity of the drug but a few cases of cardiovascular adverse events have been reported in the literature. We report on two patients affected by stage IV non-Hodgkin lymphoma with bone marrow infiltration and peripheral blood involvement who experienced cardiovascular accidents temporally related to rituximab infusion. In both cases the monoclonal antibody was administered in association with a polychemotherapy regimen but administration was postponed several days later in order to avoid severe cytokine release syndrome because of the high tumor burden. The first case concerns an episode of atrial fibrillation in a patient with a diagnosis of small B-cell lymphoma. The episode happened immediately after rituximab infusion. In the second case there was an episode of chest pain associated with fever and chills during rituximab infusion in a patient with a diagnosis of mantle cell lymphoma. In both cases we noticed an unusual correlation between symptom recurrence and the speed of rituximab infusion. Both patients presented several cardiovascular risk factors but preliminary cardiac function assessment excluded signs of heart dysfunction. The pathogenesis of cardiovascular events during rituximab infusion remains unclear. A key role might be played by cytokine release from B cells as a consequence of rituximab activity. Moreover, pre-existing silent cardiac damage could be co-responsible for the clinical manifestations we reported. We consider our clinical experience relevant because it raises an issue of good clinical practice: despite rituximabs good tolerability profile, patients with cardiovascular risk factors should undergo accurate cardiac assessment so that silent heart disease can be detected. If the suspicion of cardiac damage is high, more extensive cardiac assessment is recommended.

Combining flow cytometry and WT1 assessment improves the prognostic value of pre-transplant minimal residual disease in acute myeloid leukemia

There is increasing evidence that evaluating minimal residual disease (MRD) at various time points during acute myeloid leukemia (AML) therapy retains a strong prognostic value.[1][1]–[3][2] In particular, the persistence of MRD positivity before hematopoietic stem cell transplantation (HSCT) has

Nucleophosmin gene-based monitoring in de novo cytogenetically normal acute myeloid leukemia with nucleophosmin gene mutations: comparison with cytofluorimetric analysis and study of Wilms tumor gene 1 expression

Abstract We compared the clinical value of minimal residual disease (MRD) monitoring by cytofluorimetric methods, Wilms tumor gene 1 (WT1) expression and the study of nucleophosmin gene (NPM) mutations in a series of 26 patients with NPM-mutated de novo acute myeloid leukemia (NPM-AML) who achieved complete hematological remission after conventional chemotherapy. The relapse risk was significantly lower only in patients achieving a NPM molecular complete response (NPM mol-CR) and confirmed NPM mol-CR (non-detectable NPM mutations in two consecutive marrow samples). The disease-free survival (DFS) of patients achieving a < 4-log or ≥ 4-log reduction in NPM value after induction therapy was 12.6 % and 50%, respectively, at 36 months (p = 0.009). The attainment of a confirmed NPM-CR had a significant influence on overall survival (OS at 36 months was 64.3% and 11.9% in patients obtaining or not obtaining confirmed NPM-CR, respectively, p < 0.03). We confirm that NPM-molecular relapse (NPM-rel) is always followed by hematological relapse (H-rel), but longitudinal studies of NPM mutations may predict an impending H-rel earlier than flow cytometric- or WT1-based methods.

Depletion of SIRT6 enzymatic activity increases acute myeloid leukemia cells vulnerability to DNA-damaging agents

Genomic instability plays a pathological role in various malignancies, including acute myeloid leukemia (AML), and thus represents a potential therapeutic target. Recent studies demonstrate that SIRT6, a NAD+-dependent nuclear deacetylase, functions as genome-guardian by preserving DNA integrity in different tumor cells. Here, we demonstrate that also CD34+ blasts from AML patients show ongoing DNA damage and SIRT6 overexpression. Indeed, we identified a poor-prognostic subset of patients, with widespread instability, which relies on SIRT6 to compensate for DNA-replication stress. As a result, SIRT6 depletion compromises the ability of leukemia cells to repair DNA double-strand breaks that, in turn, increases their sensitivity to daunorubicin and Ara-C, both in vitro and in vivo. In contrast, low SIRT6 levels observed in normal CD34+ hematopoietic progenitors explain their weaker sensitivity to genotoxic stress. Intriguingly, we have identified DNA-PKcs and CtIP deacetylation as crucial for SIRT6-mediated DNA repair. Together, our data suggest that inactivation of SIRT6 in leukemia cells leads to disruption of DNA-repair mechanisms, genomic instability and aggressive AML. This synthetic lethal approach, enhancing DNA damage while concomitantly blocking repair responses, provides the rationale for the clinical evaluation of SIRT6 modulators in the treatment of leukemia.

Combined assessment of WT1 and BAALC gene expression at diagnosis may improve leukemia-free survival prediction in patients with myelodysplastic syndromes

Several genes with relevant pathogenetic and prognostic value have been identified in patients with myelodysplastic syndromes. Overexpression of WT1 at diagnosis has been associated with increased progression to acute myeloid leukemia and reduced leukemia free survival. Conversely, few data are available on the prognostic value of BAALC gene overexpression in AML and MDS patients. We evaluated the prognostic value of the combined expression of WT1 and BAALC genes at diagnosis in 86 MDS patients who had been stratified according to IPSS and R-IPSS scoring systems. Our results suggest that in the whole group of patients, low levels of both WT1 and BAALC were associated with a favorable outcome (3-year LFS 74.5%, median not reached), whereas patients presenting high expression levels of both genes had the worst prognosis (3-year LFS 0%, median 12 months, p<0.001). More specifically, molecular profiling was especially useful for intermediate 1 and intermediate-2/high risk groups. This study suggests that evaluating WT1 and BAALC gene expression at diagnosis may improve standard risk stratification and possibly refine the therapeutic approach for MDS patients.

A blastic plasmacytoid dendritic cell neoplasm-like phenotype identifies a subgroup of npm1-mutated acute myeloid leukemia patients with worse prognosis

To the Editor: As widely reported, isolated NPM1 mutations display a positive prognostic value in acute myeloid leukemia (AML) since they are associated with high complete response rate to chemotherapy and with reduced relapse risk, especially if a molecular complete remission (mCR) is achieved. However, a minority of NPM-mut AML patients do not achieve hematological or mCR or display early relapse, irrespectively of FLT3 status. Despite its recognition as a distinct WHO 2016 entity, NPM-mut AML displays indeed a certain degree of clinical and biological heterogeneity. Morphologic spectrum is wide and can involve all the FAB subtypes, with the exception of M3, with blasts frequently showing monocytic differentiation and cup-like nuclei. Even immunophenotype (IF) is not univocal; NPM-mut cells are usually CD34 negative, CD33 and CD13 positive and a “myeloid” or “monocytic” IF can be usually distinguished. No prognostic relevance has been associated to morphological and immunophenotypic features so far. We retrospectively evaluated 38 consecutive young, de novo NPM-mut AML patients diagnosed in our institution between 2006 and 2014 and treated with a fludarabine, high dose cytarabine and idarubicin (FLAI) based induction. Multicolor cytofluorimetric analysis was routinely performed on bone marrow samples obtained at diagnosis, to define lineage according to WHO 2016, and to identify the leukemia associated phenotypes for minimal residual disease (MRD) monitoring. MRD assessment was performed in all patients with both IF and RQ-PCR for NPM1 expression levels quantification, after induction and each of the consolidation courses. In our experience, a greater than 3.5 logarithmic reduction of NPM1 expression after FLAI induction identified patients with the best probability to achieve mCR and best long term outcome. The retrospective review of leukemic immunophenotypes at diagnosis allowed us to identify three different subgroups of patients; 16/38 displayed a myeloid IF [CD33/CD13/CD38/CD117/MPO (1)]; 7/38 a monocytic IF [CD33/CD64/cyLys/CD11b/CD15 (1) with 3/7 patients CD131]; the third group included 10 patients who displayed both myeloid, and monocytic features [CD33/CD13/CD38/CD117/MPO/ CD64/cLys/CD11b/CD15 (1)]. Five patients could not be assigned to any of those groups. FLT3-ITD mutation was detected in 16/38 (42%) patients. Its incidence was significantly higher in the monocytic group, however this did not translate in a worse outcome (data not shown). No statistically significant differences in relapse free survival (RFS) and overall survival (OS) were detectable among the three IF groups. The expression of CD34 did not negatively affect RFS and OS. Interestingly, searching for recurrent aberrant antigen combinations, we identified six patients with [CD56/CD123/CD4 (1)] coexpression; in other seven patients only two of these three markers were present. Since these markers represent part of the typical blastic plasmacytoid dendritic cell neoplasm (BPDCN) IF, we named this phenotype “BPDCNlike”. BPDCN-like IF was equally distributed among the previously described IF subgroups. Three out of the 6 BPDCN-like patients displayed concomitant FLT3 ITD mutation and all patients had normal karyotype. None of these BPDCN-like patients displayed clinical, morphological and biological features generally associated with BPDCN. Overall, the outcome of BPDCN-like patients was poorer compared to those not expressing this antigen combination. Specifically, five out of six BPDCN-like patients achieved CR after induction (83%) but only one patient achieved mCR. Allogeneic stem cell transplantation (HSCT) was scheduled for refractory patients and for those not achieving mCR, with three patients being transplanted. Three out of five patients not obtaining mCR could not be transplanted due to a sudden unresponsive disease relapse. A complete overview of BPDCN-like patients’ features at diagnosis, response to treatment, and long-term outcome is provided in Table 1. Three year RFS was 28 and 72%, respectively, for patient with or without BPDCN-like phenotype (P< .05), whereas 3-year OS was 0 and 63%, respectively (P<0.05). Furthermore, a trend towards an inferior OS was observed even in the seven patients presenting only two of three BPDCN markers. Although the negative impact of each of these antigens has already been described, to the best of our knowledge this is the first report on the prognostic impact of CD123, CD56, and CD4 coexpression in NPM mut AML. CD123 is strongly expressed by plasmacytoid dendritic cells and by their pathological counterpart in BPDCN and it is widely expressed in hematological malignancies. It is also expressed on physiological CD341 hemopoietic progenitors and on leukemic AML stem cells (LSC). The number of CD1231 LSC has been shown to be predictive of clinical outcome. Interestingly, a negative prognostic impact of CD123 expression in NPM-mut AML has already been

Early minimal residual disease assessment after AML induction with fludarabine, cytarabine and idarubicin (FLAI) provides the most useful prognostic information

Despite intensive therapy, relapse occurs in about half of acute myeloid leukaemia (AML) patients and cannot be accurately predicted by the risk group at diagnosis. The probability of relapse is closely correlated with the persistence of morphologically undetectable leukaemic cells after therapy (minimal residual disease; MRD). The clinical relevance of evaluating MRD with multicolour flow cytometry (MFC) during conventional AML treatment has been widely reported (Buccisano et al, 2012; Terwijn et al, 2013). We recently reported the high feasibility and efficacy of the fludarabine-containing regimen FLAI-5 (fludarabine, cytarabine, idarubicin) followed by risk-oriented consolidation for young AML patients (Guolo et al, 2016). In the present study, we evaluated the prognostic impact of longitudinal MRD assessment in the same cohort of patients by identifying the time-points with the highest prognostic impact. One hundred and twenty consecutive AML patients were treated between 1 January 2004 and 31 December 2015. Median age at diagnosis was 47 years (range 18–67). The distribution of patients according to the European LeukaemiaNet (ELN) 2017 risk score (D€ ohner et al, 2017) was: good risk N. 25 (21%), intermediate risk N. 65 (54%), and highrisk N. 30 (25%). Median follow-up was 60 months (95% confidence interval: 43 6–76 4). Only patients who achieved complete remission (CR) after FLAI-5 and for whom MRD data were available were included in the MRD study. MFC-MRD was assessed at 3 time points (TPs): TP1, after FLAI-5 induction; TP2, after the second induction course (idarubicin plus high dose cytarabine); TP3, at the end of consolidation therapy for patients who did not undergo allogeneic stem cell transplantation (HSCT), and at the time of transplantation for patients undergoing transplant (see Fig S1). For MFC analysis, bone marrow samples obtained at diagnosis were analysed with a broad panel of monoclonal antibodies. A positive MFC-MRD in the follow-up samples was defined by the presence of no less than 25 clustered leukaemic cells/10 total events (threshold of 0 025% residual leukaemic cells) (Guolo et al, 2017). Patients presenting WT1 overexpression at diagnosis, defined as a number of WT1 copies/10 000 ABL1 greater than 1000, were evaluated for WT1-MRD at TP1. A cut-off of WT1 copies/10 000 ABL1 lower than 250 was chosen to define MRD negativity (Cilloni et al, 2009; Guolo et al, Table I. Factors affecting cumulative incidence of relapse.

Intesive fludarabine-high dose cytarabine-idarubicin combination as induction therapy with risk-adapted consolidation may improve treatment efficacy in younger Acute Myeloid Leukemia (AML) patients: Rationales, evidences and future perspectives

Acute Myeloid Leukemia (AML) is the commonest form of leukemia in the adults, with an incidence of 3-4 cases per 100,000 people/year. After the first description of the effective cytarabine + antracycline (3+7) induction regimen, in the last 3 decades, no effective targeted drug has been included in the standard treatment of AML. Many efforts of modifying 3+7 adding a third drug or increasing the dose of anthracycline, cytarabine or both did not lead to substantial improvements, mainly due to increased toxicity. Many in vitro and in vivo evidences suggested that fludarabine may increase efficacy of cytarabine through a synergistic effect. Considering the continuous improvements in supportive care and management of infectious complications the feasibility of more intensive induction strategies have increased and a renewed interest in fludarabine-containing induction strategies arose. The recent MRC AML 15 trial has shown that a fludarabine-containing induction, FLAG-Ida, resulted superior to conventional 3+7 in terms of complete remission rates, relapse incidence and survival, although only a minority of patients could complete the whole planned consolidation program due to an excessive hematological toxicity. Our group recently published a 10-year experience with a fludarabine-containing induction that slightly differed from the MRC one and resulted in good efficacy and higher feasibility. In this commentary we review the major evidences supporting the employ of a fludarabine-containing induction in AML, and discuss the future perspectives.