miR-126 identifies a quiescent and chemo-resistant human B-ALL cell subset that correlates with minimal residual disease.

Complete elimination of B-cell acute lymphoblastic leukemia (B-ALL) by a risk-adapted primary treatment approach remains a clinical key objective, which fails in up to a third of patients. Recent evidence has implicated subpopulations of B-ALL cells with stem-like features in disease persistence. We hypothesized that microRNA-126, a core regulator of hematopoietic and leukemic stem cells, may resolve intratumor heterogeneity in B-ALL and uncover therapy-resistant subpopulations. We exploited patient-derived xenograft (PDX) models with B-ALL cells transduced with a miR-126 reporter allowing the prospective isolation of miR-126(high) cells for their functional and transcriptional characterization. Discrete miR-126(high) populations, often characterized by MIR126 locus demethylation, were identified in 8/9 PDX models and showed increased repopulation potential, in vivo chemotherapy resistance and hallmarks of quiescence, inflammation and stress-response pathway activation. Cells with a miR-126(high) transcriptional profile were identified as distinct disease subpopulations by single-cell RNA sequencing in diagnosis samples from adult and pediatric B-ALL. Expression of miR-126 and locus methylation were tested in several pediatric and adult B-ALL cohorts, which received standardized treatment. High microRNA-126 levels and locus demethylation at diagnosis associate with suboptimal response to induction chemotherapy (MRD > 0.05% at day +33 or MRD+ at day +78).

Longitudinal single-cell profiling of chemotherapy response in acute myeloid leukemia.

Acute myeloid leukemia may be characterized by a fraction of leukemia stem cells (LSCs) that sustain disease propagation eventually leading to relapse. Yet, the contribution of LSCs to early therapy resistance and AML regeneration remains controversial. We prospectively identify LSCs in AML patients and xenografts by single-cell RNA sequencing coupled with functional validation by a microRNA-126 reporter enriching for LSCs. Through nucleophosmin 1 (NPM1) mutation calling or chromosomal monosomy detection in single-cell transcriptomes, we discriminate LSCs from regenerating hematopoiesis, and assess their longitudinal response to chemotherapy. Chemotherapy induced a generalized inflammatory and senescence-associated response. Moreover, we observe heterogeneity within progenitor AML cells, some of which proliferate and differentiate with expression of oxidative-phosphorylation (OxPhos) signatures, while others are OxPhos (low) miR-126 (high) and display enforced stemness and quiescence features. miR-126 (high) LSCs are enriched at diagnosis in chemotherapy-refractory AML and at relapse, and their transcriptional signature robustly stratifies patients for survival in large AML cohorts.

Posttransplantation Cyclophosphamide- and Sirolimus-Based Graft-Versus-Host-Disease Prophylaxis in Allogeneic Stem Cell Transplant.

Post-transplantation cyclophosphamide (PTCy) has emerged as a promising graft-versus-host-disease (GVHD) prophylaxis in the setting of allogeneic hematopoietic stem cell transplantation (HSCT) from haploidentical donors and more recently in matched donor transplants. Herein, we describe our real-life experience on 249 adult patients undergoing allogeneic HSCT, from HLA-matched related (MRD), HLA-matched unrelated (MUD), or mismatched related donors (MMRD). Patients received unmanipulated peripheral blood stem cells (PBSCs), using a GVHD prophylaxis with PTCy and sirolimus. Mycophenolate mofetil was added in MUD or MMRD. In the HLA-matched donor group (MRD, n = 48, MUD, n = 50), the cumulative incidence of grades II-IV and III-IV acute GvHD was 23% and 9% at 100 days, respectively. The cumulative incidence of chronic GvHD was 25% at 2 years, severe only for 5% of the patients. The cumulative incidences of relapse and transplant-related mortality (TRM) were 31% and 9% at 2 years, respectively. The 2-year overall survival (OS) was 72% and progression-free survival (PFS) 60%; the composite endpoint of GvHD/relapse-free survival (GRFS) was 52% at 2 years. In the haploidentical donor group (n = 151), we documented a cumulative incidence of grades II-IV and III-IV acute GVHD of 35% and 20% at 100 days, respectively, and a cumulative incidence of chronic GvHD of 39% at 2 years. We observed severe chronic GVHD in 15% of the patients. The cumulative incidence of relapse and TRM was 32% and 25% at 2 years, respectively. The 2-year OS was 48%, whereas PFS was 43%; GRFS was 28% at 2 years. However, more patients in the haploidentical group presented high/very high disease risk index (DRI) and higher HCT-comorbidity index. In patients classified in the low-intermediate DRI, 2-year GRFS was 53% in MRD, 65% in MUD, and 46% in haploidentical HSCT (P = .33). Sirolimus-PTCy platform deserves further investigation as an alternative to calcineurin-inhibitor-based GVHD prophylaxis for all donor sources. In patients presenting a low-intermediate DRI, this strategy translates in relevant survival independently from the transplant source.

Ruxolitinib for chronic steroid-refractory graft versus host disease: a single center experience.

BACKGROUND: Chronic Graft versus Host Disease (GvHD) is a serious complication of allogeneic hematopoietic stem cell transplant that severely impacts quality of life and long-term survival. About 50-to-60 % of patients treated with steroids require a further line of therapy due to lack of sustained response. Ruxolitinib, a JAK1/2 inhibitor, has recently been approved for the treatment of acute GvHD. METHODS: We aimed to retrospectively evaluate ruxolitinib efficacy and safety in a cohort of patients diagnosed with moderate (25 %) or severe (75 %) steroid-refractory or steroid-dependent chronic GvHD. Response evaluation was performed at three and six months. RESULTS: Thirty-six patients received ruxolitinib after a median of three previous lines (range, r 1-11) for a median of 8.6 months (r 1-51.6). Cutaneous GvHD was the most frequent presentation. We observed an overall response of 59 % (CR 9%, PR 50 %) at three months and 62 % (CR 15 %, PR 46 %) at six months. Two patients had hematologic disease recurrence and were censored at relapse; no other permanent discontinuation due to adverse events were documented. Cutaneous, oral, genital and ocular GvHD significantly improved after treatment. 2-year overall survival and 2-year transplant related mortality were 74 % and 19 % respectively. Ruxolitinib was associated with a significant reduction of steroid dose. CONCLUSION: Ruxolitinib was confirmed to be a safe and effective option as salvage treatment also for advanced stages of chronic GvHD. Longer follow up is needed to evaluate durability of response. Prospective analyses on larger cohorts are ongoing.

The place of ceftazidime/avibactam and ceftolozane/tazobactam for therapy of haematological patients with febrile neutropenia.

OBJECTIVES: To evaluate ceftazidime/avibactam (C/A) and ceftolozane/tazobactam (C/T) use in haematological patients with febrile neutropenia receiving high-dose chemotherapy and haematopoietic stem cell transplantation (HSCT). METHODS: A retrospective study was conducted to assess C/A and C/T efficacy through infection-related mortality (IRM) and bacteraemia clearance for carbapenem-resistant Gram-negative bacteria (CR-GNB) pre-engraftment blood-stream infections (PE-BSIs) between January-December 2018. RESULTS: Seventy patients underwent allogeneic HSCT: C/A and C/T were dispensed in 13% and 3%, respectively. C/A was administered as definite therapy for carbapenem-resistant Klebsiella pneumoniae (CR-Kp) PE-BSI in four carriers (bacteraemia clearance in 5 days), empirical therapy for a clinically documented infection in two patients (one carrier with pneumonia and one non-carrier with shock) and empirical therapy for fever of unknown origin in three CR-Kp carriers. C/T was administered as definite therapy for carbapenem-resistant Pseudomonas aeruginosa (CR-Pa) intra-abdominal infection in one carrier and empirical therapy for a clinically documented infection (pneumonia) in one non-carrier. Among patients without PE-BSIs and with Gram-positive bacteria PE-BSIs, IRM was 0% at +30 days; conversely, it was 30% in GNB PE-BSIs (two CR-Kp and one CR-Pa C/T-resistant). Thirty-nine patients underwent autologous HSCT: C/A and C/T were administered, respectively, as definite therapy for CR-Kp PE-BSI in one carrier (bacteraemia clearance in 3 days) and for Pa PE-BSI (three strains, one CR-Pa) in one non-carrier (bacteraemia clearance in 2 days). Overall, IRM at +30 days was 0%. CONCLUSIONS: Monitoring multidrug-resistant GNB colonisation enabled selection of carriers who benefit from prompt administration of new antibiotics, improving HSCT outcomes in a high-risk population. C/A and C/T were effective in bacteraemia clearance; unfortunately, multidrug-resistant GNB PE-BSIs were still a burden to IRM.

CPX-351 treatment in secondary acute myeloblastic leukemia is effective and improves the feasibility of allogeneic stem cell transplantation: results of the Italian compassionate use program.

Secondary acute myeloid leukemia (sAML) poorly responds to conventional treatments and allogeneic stem cell transplantation (HSCT). We evaluated toxicity and efficacy of CPX-351 in 71 elderly patients (median age 66 years) with sAML enrolled in the Italian Named (Compassionate) Use Program. Sixty days treatment-related mortality was 7% (5/71). The response rate at the end of treatment was: CR/CRi in 50/71 patients (70.4%), PR in 6/71 (8.5%), and NR in 10/71 (19.7%). After a median follow-up of 11 months relapse was observed in 10/50 patients (20%) and 12 months cumulative incidence of relapse (CIR) was 23.6%. Median duration of response was not reached. In competing risk analysis, CIR was reduced when HSCT was performed in first CR (12 months CIR of 5% and 37.4%, respectively, for patients receiving (=20) or not (=30) HSCT, p = 0.012). Twelve-months OS was 68.6% (median not reached). In landmark analysis, HSCT in CR1 was the only significant predictor of longer survival (12 months OS of 100 and 70.5%, for patients undergoing or not HSCT in CR1, respectively, p = 0.011). In conclusion, we extend to a real-life setting, the notion that CPX is an effective regimen for high risk AML patients and may improve the results of HSCT.

SETBP1 induces transcription of a network of development genes by acting as an epigenetic hub.

SETBP1 variants occur as somatic mutations in several hematological malignancies such as atypical chronic myeloid leukemia and as de novo germline mutations in the Schinzel-Giedion syndrome. Here we show that SETBP1 binds to gDNA in AT-rich promoter regions, causing activation of gene expression through recruitment of a HCF1/KMT2A/PHF8 epigenetic complex. Deletion of two AT-hooks abrogates the binding of SETBP1 to gDNA and impairs target gene upregulation. Genes controlled by SETBP1 such as MECOM are significantly upregulated in leukemias containing SETBP1 mutations. Gene ontology analysis of deregulated SETBP1 target genes indicates that they are also key controllers of visceral organ development and brain morphogenesis. In line with these findings, in utero brain electroporation of mutated SETBP1 causes impairment of mouse neurogenesis with a profound delay in neuronal migration. In summary, this work unveils a SETBP1 function that directly affects gene transcription and clarifies the mechanism operating in myeloid malignancies and in the Schinzel-Giedion syndrome caused by SETBP1 mutations.