Background: Despite significant advancements in treatment efficacy of acute promyelocytic leukemia (APL), early death (ED) - mainly related to major hemorrhagic and thrombotic events occurring within ...30 days from diagnosis - remains a prominent hurdle to therapeutic success. In this regard, Österroos et al. very recently developed a score which stratifies the risk of ED of patients (pts) with APL in three categories (low score 0-2, high score 3-4, very high score 5-7) according to age (<50 years, 50-59 years, 60-69 years and ≥70 years), white blood cell count (<3.0x10 9/L, 3.0-5.0x10 9/L and >5.0x10 9/L), and platelet count (≥30x10 9/L and <30x10 9/L), all of which are readily available, real-world variables. However, the presence of specific comorbidities, or diseases characteristics, such as the presence of FLT3-ITD mutation, CD2 expression and bcr3 PML::RARA transcript were not evaluated, even if they could potentially further improve the risk stratification ( Österroos et al. Haematologica 2022;107(7):1528-1537). Aims: The aims of this study were to integrate the score proposed by Österroos et al. and evaluate the role of pts comorbidities and disease biological data in modifying risk stratification. Methods: Data were retrospectively collected from 127 consecutive pts diagnosed at our Center from January 2000 to May 2023. Patients had been treated according to the AIDA protocol (n = 76, 60%) or with the combination of all-trans retinoic acid (ATRA) and arsenic trioxide (ATO) (n = 39, 31%) ( Lo-Coco et al. N Engl J Med 2013;369:111-121); 10 pts received ATRA; only 2 pts did not receive any treatment. Results: Table 1 summarizes the main clinical characteristics of the 127 pts included in this study. Overall, ED rate was 11% (n = 14, 12 died within 7 days from diagnosis); the cause of death was a hemorrhagic event in 11 pts. The score identified low- (n = 79, 64%), high- (n = 39, 31%) and very high-risk (n = 6, 5%) categories; ED rates for each category were 3.8%, 20.5% and 0%, respectively. Despite no ED events were registered in our small very-high risk cohort (n = 6), a third of these pts (n = 2) experienced cerebral hemorrhage at disease onset and eventually survived. We confirmed that the score proposed by Österroos et al. was better at predicting ED risk than the Sanz score, with the Area Under the Receiver Operating Characteristic (AUROC) curve of 0.73 (95% CI 0.60-0.87) vs 0.66 (95% CI 0.51-0.81). The AUROC of the reference study cohort was 0.77 (95% CI 0.72-0.83). The presence of specific biological characteristics of the disease, such as FLT3-ITD mutation, bcr3 PML::RARA transcript or CD2 expression were not associated with an increased risk of ED (Table 2). Similarly, the presence of comorbidities (i.e. cardiovascular diseases, hypertension, chronic kidney disease, diabetes mellitus, dyslipidemia, and smoking) were not associated with an increased risk of ED (data not shown). By univariate analysis the presence of fever at diagnosis and male sex showed a trend toward an increased risk of ED (OR 3.77, 95% CI 0.88-19.26, P = 0.08 and OR 3.13, 95% CI 0.92-14.36, P = 0.09, respectively). Conclusions: Our data support the use of the score developed by Ö sterroos et al. to better predict the risk of ED in APL and to select the pts who may benefit from more aggressive supportive care. The presence of fever at diagnosis and male sex seems to be associated with a further increase in the risk of ED but this association needs to be confirmed in a larger study. The integration of biological characteristics of disease and comorbidities does not improve the risk stratification of ED.
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IJS, IMTLJ, KILJ, NLZOH, NUK, SAZU, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Background. The only curative treatment of Myelodysplastic syndromes (MDS) is represented by allogeneic hematopoietic cell transplantation (HCT). The risk scoring system conventionally used in ...clinical practice for transplant decision-making is IPSS-R (Greenberg PL et al, 2012), which has been shown to predict the HCT outcome(Della Porta MG et al, 2014). The recent broader application of next generation sequencing (NGS) in clinical practice provides an abundance of molecular data and poses the challenge of better prognostic stratification in MDS patients. Recently, Bernard et al(Elsa Bernard et al, 2022) developed a molecular prognostic score called IPSS-Molecular (IPSS-M) which defines six risk categories based on the patient's molecular profile combined with clinical and cytogenetic data. In the original study, only 9% of patients underwent HSCT, therefore the utility of this score in the transplant decision making needs to be explored. Aim.The aim of this study is to evaluate the impact of IPSS-M on the outcome of HCT in patients with MDS. Methods and Results. This is a single center study including 60 consecutive MDS patients who received allogeneic HCT from 2004 to 2023 at Papa Giovanni XXIII Hospital, Bergamo. The median age at transplantation was 57 years and 70% were males. HCT related parameters are listed in Table 1. Of note, 53 out of 60 patients (88%) received an upfront HCT. According to IPSS-RA (age-adjusted IPSS-R), patients clustered in Very Low (4/60), Low (11/60), Intermediate (24/60), High (16/60) and Very High (5/60) risk groups. We retrospectively performed a NGS study before HCT by using a panel of 30 genes for all patients who had available DNA and calculated IPSS-M score for each patient. According to IPSS-M, patients clustered in Very Low (2/60), Low (12/60), Moderate Low (15/60), Moderate High (9/60), High (12/60) and Very High (10/60). Re-stratification occurred in 59% of patients (35/60), of which 22% (13/60) were down-staged and 37% (22/60) were upstaged. With a median follow-up time of 7 years after HSCT, the 5-year overall (OS) and relapse-free survival (RFS) was 67% and 64% respectively. In order to evaluate the utility of IPSS-M in estimating OS and RFS in HCT setting compared to IPSS-R, a concordance test was performed. In terms of OS and RFS, IPSS-M resulted respectively in a C-index of 0.772 and 0.729 compared to 0.567 and 0.607 of IPSS-R (table 2). Conclusions. Despite the limitations due to the small sample and the retrospective design of the study, we conclude that IPSS-M performed better compared to the conventional score in predicting RFS and OS of our MDS patients undergoing HSCT, providing data supporting the integration of clinical and molecular information in this setting. Further improvements may be achieved by integrating the molecular data with clinical information regarding HSCT-related parameters such as performance status, donor and conditioning regimen.
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IJS, IMTLJ, KILJ, NLZOH, NUK, SAZU, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Allogeneic hematopoietic stem cell transplantation (HSCT) remains the only curative option for patients with Myelodysplastic syndromes (MDS). For many years, the selection of patients to allogeneic ...HSCT has been largely based on use of the International Prognostic Scoring System-Revised (IPSS-R). However, the recent broader application of next generation sequencing in clinical practice provided an abundance of molecular data and led to the introduction of molecular prognostic scores as IPSS-Molecular (IPSS-M). In this paper, we retrospectively analyzed the outcomes of 57 consecutive MDS patients treated with allogeneic HSCT in our center. Re-stratification from IPSS-R to IPSS-M occurred in almost half of patients. The application of IPSS-M to our cohort demonstrated a stronger prognostic separation compared to IPSS-R and improved the C-index. Very high-risk IPSS-M patients showed worse outcomes following HSCT compared to high-risk patients. This study provides data supporting the need of integrating molecular information in the transplant decision making of patients with MDS. This allows an earlier and better identification of patients to whom the transplant should be advised.
•IPSS-R is commonly used to allocate MDS patients to HSCT.•Novel scores as IPSS-M taking into account the molecular data have been introduced.•This is a retrospective single-center study on MDS patients treated with HSCT.•Re-stratification from IPSS-R to IPSS-M occurred in almost half of patients.•IPSS-M demonstrated a stronger prognostic separation compared to IPSS-R.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Introduction. The outcome of adult patients with acute lymphoblastic leukemia (ALL) has markedly improved following the introduction of a pediatric inspired chemotherapy backbone coupled with minimal ...residual disease (MRD) monitoring at well-defined timepoints (TP), translating into a rational patients' stratification for allogeneic transplantation (Tx). Immunotherapy, in particular blinatumomab, represents a further advancement in the management of ALL. We designed a phase II trial (GIMEMA LAL2317) for adult Ph- CD19+ B-lineage ALL (Ph- B-ALL) consisting of a pediatric chemotherapy backbone - with dose adjustment in patients >55 years - to which 2 cycles of blinatumomab were added, the first after early consolidation cycle 3 (high-dose methotrexate and Ara-C) and the second after late consolidation cycle 6. Tx allocation was based on MRD assessment, carried out in most cases by RQ-PCR and by flow cytometry when a sensitive molecular marker was not identified. The primary objective of the trial was to assess the impact of blinatumomab in increasing the rate of early MRD negativity at end of week 14 (TP3). Patients and methods. The protocol was designed for adults up to the age of 65 years with newly diagnosed Ph- B-ALL; 149 patients were enrolled. Median age was 41 years (18-65), with 28 patients (19%) being older than 55. Eighty-one patients were males, and the median white blood count (WBC) was 4.5x10 9/l (1-474); 12 cases harbored the KMT2A rearrangement, 5 the TCF3:: PBX1 transcript and 31 were classified as Ph-like according to the BCR/ABL1-like predictor . The median follow-up is 37.5 months (0.55-62.8) Results. At the end of induction (TP1, after cycles 1 and 2), 131 patients (88%) were in complete remission (CR), with a significant difference according to age: 94%, 92% and 64% in the 18-40, 40.55 and >55 years cohorts, respectively (p<0.001). All patients with KMT2A and TCF3::PBX1 rearrangements achieved a CR, while 5 Ph-like patients did not. At TP2, i.e. after early consolidation, 85 patients (70%) became MRD-negative (MRD neg). TP3, after the 1 st blinatumomab cycle, 102 patients were MRD neg; the rate of MRD negativity increased to 93% (p<0.001). Imposable results were obtained considering only paired MRD samples. Eight patients remained MRD pos, including 4 with a KMT2A rearrangement and 1 with a E2A::PBX1 rearrangement. OS and DFS at 36 months are 71% and 66% on the whole cohort. OS was significantly different (p=0.00013) according to age cohorts: 76%, 74% and 49% for patients aged 18-40, 40-55 and >55 years, respectively. Likewise, DFS was 71%, 62% and 42% for the same age cohorts, without reaching statistical significance. When patients were stratified for TP2 MRD, OS and DFS were significantly better for MRD neg vs MRD pos cases (p=0.0006 and p<0.0001, respectively). At variance, at TP3, with the caveat of the small sample size of MRD pos patients, the differences between MRD neg and MRD pos patients at TP3 were abolished: indeed, OS and DFS were 82% and 68% for MRD neg, 80% and 70% for MRD pos and 88% and 72% for those lacking MRD evaluation (Figure). Finally, 30 relapses have so far occurred, including 10 Ph-like, 1 KMT2A and 3 TCF3::PBX1 rearranged cases. The overall cumulative incidence of relapse (CIR) was 27.5%; of note, within TP3 MRD neg patients CIR was 42.5% in Ph-like cases vs 17.5% in the remaining patients. By univariate analysis, significant factors for OS were age (p<0.001), CR achievement (p<0.001) and TP2 MRD (p=0.002); TP2 MRD retained significance in multivariate analysis (p=0.001). Univariate analysis for DFS showed as significant the Ph-like signature (p=0.026) and TP2 MRD (p<0.001); both retained statistical significance in multivariate analysis (p=0.021 and <0.001, respectively). Toxicity details will be provided. Conclusions. This phase II trial shows the benefit of adding blinatumomab for the management of Ph- B-ALL, with the primary objective being achieved with 93% of patients being MRD neg after the 1 st cycle of blinatumomab. With a median follow-up of 37.5 months, OS and DFS are 71% and 66%, respectively, better than in historical results, particularly in patients up to the age of 55. Importantly, at TP3 the MRD status no longer impacted, highlighting the therapeutic role of blinatumomab, as observed in Ph+ ALL. Finally, the Ph-like signature retains its negative prognostic impact, suggesting that a combination strategy, similar to that in place for Ph+ ALL, is required.
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IJS, IMTLJ, KILJ, NLZOH, NUK, SAZU, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Background: CEBPA-mutated acute myeloid leukemia (AML) is a separate entity in the updated WHO and ICC classifications. Previously identified by double mutations, the clinical profile and favorable ...outcome were correlated with in-frame mutations in bZIP domain. Beyond mutations recurrent in myeloid neoplasms (involving TET2, FLT3, NRAS), CEBPA-mutated AML is enriched in gene mutations (i.e., GATA2, CSF3R, WT1) rarely observed in CEBPA-wild type AML. No conclusive findings have been drawn for the prognostic impact of additional mutations. Of note, somatic GATA2 mutations have been associated with invasive fungal infections in myeloid neoplasms. Although overall correlating with favorable prognosis, an unexpectedly high rate of treatment-related mortality (TRM) in CR has been described in a large multicenter study on intensively treated CEBPA-mutated patients (pts) (Schlenk et al Blood 2013;122(9):1576). Aims: The aim of our study was to correlate baseline and treatment characteristics with the outcome and hematological toxicity in an intensively treated cohort of CEBPA-bZIP mutated AML pts. Methods: Pts entering the study had a diagnosis of CEBPA-bZIP AML confirmed according to 2022 WHO and ICC criteria. Pts were characterized by next-Generation deep amplicon sequencing with Ion Torrent platform (ThermoFisher Scientific) covering a panel of 40 genes. Sequence alignment and filtering were performed by NextGENe version 2.4.2.1 (SoftGenetics). The first two chemotherapy cycles were categorized according to the delivery of an anthracycline (ANTHRAC) and cytarabine (ARA-C) at high dose (HDAC). The study was approved by the local institutional review board. Results: From 2004 to 2023, 49 pts with CEBPA-bZIP mutated AML met the inclusion criteria at project study sites (Firenze, Bergamo). Their features are detailed in Table 1. CR rate was achieved in 45 of 49 pts (91.8%), with a DFS and OS of 74.3% and 82.4% at 5 y, respectively. Based on NGS, at least one additional mutation was identified in 18 (36.7%) pts (NGS+), the majority of them involving GATA2 (n=14, 28.6%). As per treatment, 44 pts received at least two cycles, of which no (n=2, 4.6%), one (n=14, 31.8%), or two (n=28, 63.6%) ANTHRAC-containing cycles. As regarded HDAC, 19 (43.2%), 17 (38.6%) and 8 (18.2%) pts received no, one or two HDAC-containing cycles, respectively. No significant differences emerged for DFS or OS between NGS+ and NGS- groups (P=0.33 and 0.52, respectively). To gain insight into potential causes of toxicity during CR, we focused on hematopoietic recovery after induction and consolidation. NGS+ pts showed slower neutrophil (ANC) recovery (median, 22 days to >500/uL after induction and 25 days after consolidation) compared to NGS- group (19 days, P=0.018; 18 days P=0.058, respectively). Such effect was enhanced by treatment with ANTHRAC-containing cycles: NGS+ pts receiving two cycles required significantly longer time to recover from first consolidation cycle, both for ANC (30 days) and platelet count (34 days) vs 22 (P=0.012) and 27 (P=.010) days, respectively, of other categories (Figure 1). A proportion of 45.5% (5/11) NGS+/ANTHRAC-2 pts could not complete the planned chemotherapy program due to hematological toxicity compared to 19.3% (6/31, P=0.09) in other categories. No significant prolongation of recovery was observed for HDAC. Conclusions: CEBPA-bZIP mutated AML is featured by high response rate and favorable outcome. Delayed hematopoietic recovery was observed in pts bearing additional mutations (especially GATA2) when treated with anthracyclines in first two cycles. Our findings suggest the cumulative dosage of anthracyclines should be limited in CEBPA-mutated/NGS+ patients to spare hematological toxicity on turn impairing the therapeutic plan.
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IJS, IMTLJ, KILJ, NLZOH, NUK, SAZU, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Leukemia is a complex disease and the molecular mechanisms of malignant transformation are not fully understood yet. Cohesins are essential proteins, forming a complex which plays critical roles in ...various cellular processes, not only by canonical DNA binding and chromosome segregation but also in non-canonical regulation of gene expression in both proliferating and post-mitotic cells. Mutations or gene expression alterations of genes encoding the components of the cohesins' complex may affect chromosomal stability and potentially contribute to the pathogenesis of leukemia. Somatic cohesins mutations have been already reported to occur in myeloproliferative disorders and pediatric Acute Lymphoblastic Leukemia (ALL), while on the other hand, germline mutations are causative of Cohesinopathies such as Cornelia de Lange Syndrome (CdLS). Our previous first report of a patient affected by both CdLS and ALL suggested a potential involvement of cohesin germline mutations in the pathogenesis of ALL. Moreover, we demonstrated that germline mutations in STAG1 gene could play a crucial role in predisposition to pediatric hematological malignancies, including ALL. These findings suggest that both somatic and germline alterations of cohesin genes may be associated with the development of ALL. This study aims to understand the molecular mechanisms affected by Cohesin genes alterations, that could potentially pave the way for the development of targeted therapies and improved treatment strategies for leukemia patients. By employing a DNA-targeted next-generation sequencing (NGS) screening of a cohort of 120 consecutive pediatric ALL cases (enrolled in the AIEOP-BFM protocol at various Italian centers), we discovered a total of 11 germline cohesin mutations, further categorized as follows: 6 out of 11 were missense, 4 out of 11 were located in the 3' or 5' untranslated regions (UTR) and 1 out of 11 was a splice-acceptor variant. To investigate the functional implications of selected germline mutations in STAG1 gene, we took advantage of the CRISPR/Cas9 technique to generate in vitro models of two Lymphoblastoid Cell Lines (LCLs) harboring specific variants already found in two patients of our cohort. Additionally, we established control LCLs from healthy donors' samples using the same CRISPR/Cas9 approach. Subsequently, we performed RNA sequencing (RNAseq) analysis on these cell lines, and our preliminary findings revealed a total of 619 differentially expressed genes (DEGs) compared to the control group. Many of them are involved in critical intracellular pathways, like ribosomal transcription, suggesting potential disruptions of essential cellular processes caused by germline STAG1 mutations. Moreover, we have successfully developed an advanced computational analysis pipeline with the primary objective of detecting fusion transcripts from RNAseq data. This pipeline was applied to a dataset of 529 consecutive diagnose and 95 relapse samples of pediatric B/T-ALL patients and in five B-ALL cases a fusion transcript involving the STAG1 (n=1), STAG2 (n=3), and NIPBL (n=1) genes was identified. The significance of these findings was further reinforced by additional cases from both pediatric and adult cohorts through an international collaborative effort and five more fusion transcripts of STAG2 (n=2) and NIPBL (n=3) were uncovered. Investigation to clarify the underlying functional consequences of these fusion genes in pediatric ALL are ongoing. Collectively, these observations endeavor to elucidate the role of germline and somatic cohesin events in pediatric ALL. In particular, the investigation seeks to unveil the non-canonical functions of these proteins, thus paving the way for the identification and characterization of a novel and distinct subcategory of leukemia within the pediatric population.
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IJS, IMTLJ, KILJ, NLZOH, NUK, SAZU, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
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).
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EMUNI, FIS, FZAB, GEOZS, GIS, IJS, IMTLJ, KILJ, KISLJ, MFDPS, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
Among acute myeloid leukemia (AML) patients with a normal karyotype (CN-AML), NPM1 and CEBPA mutations define World Health Organization 2008 provisional entities accounting for approximately 60% of ...patients, but the remaining 40% are molecularly poorly characterized. Using whole-exome sequencing of one CN-AML patient lacking mutations in NPM1, CEBPA, FLT3-ITD, IDH1, and MLL-PTD, we newly identified a clonal somatic mutation in BCOR (BCL6 corepressor), a gene located on chromosome Xp11.4. Further analyses of 553 AML patients showed that BCOR mutations occurred in 3.8% of unselected CN-AML patients and represented a substantial fraction (17.1%) of CN-AML patients showing the same genotype as the AML index patient subjected to whole-exome sequencing. BCOR somatic mutations were: (1) disruptive events similar to the germline BCOR mutations causing the oculo-facio-cardio-dental genetic syndrome; (2) associated with decreased BCOR mRNA levels, absence of full-length BCOR, and absent or low expression of a truncated BCOR protein; (3) virtually mutually exclusive with NPM1 mutations; and (4) frequently associated with DNMT3A mutations, suggesting cooperativity among these genetic alterations. Finally, BCOR mutations tended to be associated with an inferior outcome in a cohort of 422 CN-AML patients (25.6% vs 56.7% overall survival at 2 years; P = .032). Our results for the first time implicate BCOR in CN-AML pathogenesis.
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IJS, IMTLJ, KILJ, NLZOH, NUK, SAZU, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
