AML Transformation Research Articles

Real-World Electronic Medical Records Data Identify Risk Factors for Myelofibrosis and Can Be Used to Validate Established Prognostic Scores.

Myelofibrosis (MF) is a myeloproliferative neoplasia arising de novo as primary myelofibrosis (PMF) or secondary to polycythemia vera or essential thrombocythemia. Patients experience a high symptom burden and a marked reduction in life expectancy. Despite progress in molecular understanding and treatment, the clinical and prognostic heterogeneity of MF complicates treatment decisions. The International Prognostic Scoring System (IPSS) integrates clinical factors for risk stratification in MF. This study leverages the TriNetX database with more than 64,000 MF patients to assess the impact of accessible parameters on survival and complicating events, including AML transformation, cachexia, increased systemic inflammation, thrombosis and hemorrhage. Age over 65 years correlated with increased risks of death, AML transformation, thrombosis and hemorrhage. Anemia (Hb < 10 g/dL), leukocytosis (>25 × 103/µL) and thrombocytopenia (<150 × 103/µL) reduced survival and increased risks across all assessed events. Monocytosis is associated with decreased survival, whereas eosinophilia and basophilia were linked to improved survival. Further, as proof of concept for the applicability of TriNetX for clinical scores, we devised a simplified IPSS, and confirmed its value in predicting outcomes. This comprehensive study underscores the importance of age, anemia, leukocytosis and thrombocytopenia in predicting disease trajectory and contributes to refining prognostic models, addressing the challenges posed by the disease's heterogeneity.

Patterns of lower risk myelodysplastic syndrome progression: factors predicting progression to high-risk myelodysplastic syndrome and acute myeloid leukemia.

The patterns of low-risk myelodysplastic syndrome (MDS) progression and the clinical and molecular features of those patterns have not been well described. We divided our low-risk (LR) MDS patients (N=1,914) into 4 cohorts: 1) patients who remained LR-MDS (LR-LR; N=1,300; 68%), 2) patients who progressed from LR to high-risk (HR) MDS (LR-HR) without transformation into acute myeloid leukemia (AML) (N=317; 16.5%), 3) patients who progressed from LR to HR MDS and then AML (LR-HR-AML; N=124; 6.5%), and 4) patients who progressed from LR MDS directly to AML (LR-AML; N=173; 9%). Risk factors for progression included: male gender, low absolute neutrophil count (ANC), low platelet count, high bone marrow (BM) blasts, ferritin >1000 mcg/L, albumin <3.5 g/dL, multi-lineage dysplasia (MLD), and lack of ring sideroblasts. Among patients with marked BM fibrosis (N=49), 18% progressed directly to AML. Somatic mutations (SM) associated with an increased risk of direct or indirect AML progression included SRSF2 and NRAS. SM in IDH1, IDH2 and NPM1 were more common in patients with direct AML transformation. SM associated with progression to higher risk disease only, without AML transformation, were ASXL1, TP53, RUNX1, and CBL. SF3B1 mutation was associated with less progression. About 171 patients (13.1% of all LR-LR patients) died within two years of diagnosis of LR-MDS without disease progression. Among the 61 cases with documented cause of death, 18 patients (29.5%) died from cytopenia and MDS-related complications. Identifying patterns of disease progression of LR MDS patients and their predictive factors will be crucial to be able to tailor therapy accordingly.

Myelodysplastic Syndromes and Myelodysplastic Syndromes/Myeloproliferative Neoplasms: A Real-World Experience From a Developing Country.

Myelodysplastic syndromes (MDS) include a heterogeneous group of clonal bone marrow disorders characterized by ineffective hematopoiesis. They manifest as dysplasia in bone marrow hemopoietic elements associated with peripheral cytopenias with variable risk of AML transformation. We analyzed retrospectively registry data collected prospectively from patients with primary MDS and patients with MDS/myeloproliferative neoplasm (MPN) in the Jordan University Hospital between January 2007 and September 2021. The registry captured epidemiologic information such as date of diagnosis, age, gender, date of AML transformation, cytogenetics, MDS subtype, risk group according to Revised International Prognostic Scoring System, and survival. The registry also captured baseline ferritin, B12, and lactate dehydrogenase levels. A total of 112 patients with MDS and MDS/MPN were included in the registry. Median age at diagnosis was 59 years. The male-to-female ratio was about 1.2. In a multivariate cox regression model, baseline serum ferritin significantly affected survival as patients with levels exceeding 1,000 μg/L had a risk of death three times higher compared with those with <1,000 μg/L levels (P < .05). To our knowledge, our study is the first comprehensive study examining the epidemiology and prognostic factors in patients with MDS and patients with MDS/MPN in Jordan. Our results show that MDS and MDS/MPN epidemiology in Jordan is different compared with Western countries. Our results also show that baseline serum ferritin levels can be used as a prognostic marker for patients with MDS.

Chronic Myelomonocytic Leukemia (CMML) with AML Typical Mutations ( NPM1, FLT3 or CEBPA) Identify a High-Risk CMML Group Independent of Molecular-Cpss

CMML is rarely associated with AML typical mutations such as NPM1, FLT3, CEBPA and IDH1/2 (mutAML-CMML). It is unknown what its clinical meaning is and if its presence should change our management into an AML-type treatment. The aim was to describe the characteristics, prognosis and treatment of mutAML-CMML patients. We studied 83 CMML patients of Hospital Clinic diagnosed between 1998-2022, with Next Generation Sequencing (NGS). We included 53 patients with mutAML-CMML from other institutions. We identified 31 patients with mutated IDH2 (all c.419G&amp;gt;A p.(Arg140Gln)), 13 with IDH1, 14 with NPM1,7 with CEBPA (4 bZIP) and 11 with FLT3 (3 FLT3-ITD and 7 FLT3-TKD: 6 TKD1 and 2 TKD2 variants, all different from p.(Asp835Tyr) except one). Patient characteristics and outcome of the group with mut IDH1/2 was not significantly different from wt IDH. Those patients with mut NPM1, FLT3 and/or CEBPApresented adverse characteristics and worse prognosis. Therefore, we grouped the patients with these mutations into a category named mutCFN (n=26) (Table 1). The mutCFN incidence in our series was 9%. The mutCFN patients (n=26) vs wtCFN (n=96) were younger (63 vs 71 years, p=0.004), had more leucocytes (16 vs 7.7x10 9/L, p=0.015), monocytes (2.9 vs 1.7x10 9/L, p=0.006), anemia (9.1 vs 11.2 g/L, p&amp;lt;0.001) and bone marrow blasts (11.5 vs 4%, p&amp;lt;0.001). They belonged more frequently to myeloproliferative variant (57.7 vs 30.2%, p=0.012), CMML-2 (69.2 vs 14.7%, p&amp;lt;0.001), high risk CPSS and CPSS-Mol (72 vs 31%, p&amp;lt;0.001; 88.2 vs 44.9%, p=0.001). They showed more transfusion dependence (69.2 vs 22.9%, p&amp;lt;0.001). Co-mutational representation is shown in Figure 1. mutCFN patients have a different co-mutational pattern from wtCFN with more frequently mutated DNMT3A (42.9 vs 6.3%, p&amp;lt;0.001), SF3B1 (15.8 vs 2.3%, p=0.04), WT1 (10 vs 0%, p=0.03), and less TET2 (24 vs 58.3%, p=0.003) . mutCFN patients received treatment more frequently than wtCFN (84.6 vs 36.5%, p&amp;lt;0.001) and earlier (0.97 vs 4.6 months, p=0.005). They were more frequently treated with chemotherapy (53.8 vs 7.3%, p&amp;lt;0.001) and alloSCT (61.5 vs 11.5%, p&amp;lt;0.001). The median follow-up was 9.3 years (95% CI, 8-11). The 2-year cumulative incidence of AML (CIR-AML) transformation was higher in mutCFN patients (44% vs 13%, p&amp;lt;0.001) and overall survival (OS) was shorter (25 vs 38 months, p=0.057). Besides, when those who received and alloSCT were censored at time of alloSCT, mutCFN showed an inferior survival (9.6 vs 35.7 months, p&amp;lt;0.01). Multivariate analysis confirmed mutCFN as an adverse prognostic factor independent of age and CPSS-Mol (HR 2.42, IC 95% 1.23-4.76, p=0.011). Because CFN mutation showed independent prognostic value not captured by CPSS-Mol, we added to CPSS-Mol 1 point if patient had mutCFN. We derived a modified CPSS-Mol-CFN score and identified five groups with different median OS of 85 (95% CI, 54-117) vs 74 (95% CI, 53-94) vs 40 (95% CI, 17-63) vs 22 (95% CI, 11-33) vs 15 (95% CI, 5-25) months, p&amp;lt;0.001with a better predictive capacity than CPSS-Mol (C-index 0.66 vs 0.64). Nowadays, there is no consensus between the two recent 2022 ICC and WHO classifications, but the identification of NPM1 or CEBPA mutations in cases of CMML could define AML. We analysed the group of patients who harboured bZIP- CEBPA or NPM1 mutations (“mutCN”) (n=17) and we found that these patients retained its chemosensitivity (90%CR) and those treated with chemotherapy had better OS (27 vs 9 months, p&amp;lt;0.001). 10 mutCFN patients received alloSCT with a median OS post-alloSCT of 21 months (95% CI, 16-26) and a 2year-CIR-AML of 37.5% (95% CI, 15-61). 5 patients received targeted therapy for a post-transplant relapse: 3 received sorafenib after a molecular relapse with prolonged responses and 2 received an IDH inhibitor, one of them persisting with a complete response after 4 years. We analyzed 5 paired samples (diagnostic CMML-AML transformation). All of the AML related mutations were retained at AML transformation and emerged new mutations in genes of signaling ( FLT3, NRAS), transcription ( RUNX1, CEBPA) and splicing ( STAG2) pathways. Independently of CPSS-Mol, mutCFN CMML patients have a different profile, closer to AML and with worse prognosis. They could benefit from more aggressive AML-treatment based on chemotherapy and alloSCT when possible. Interestingly, some patients may benefit from approved targeted therapy for AML FinancingISCIII PI19/01476 PI22/01660, co-financed EU, Emili-Letang HCPB

Lymphopenia Confers Poorer Prognosis in Myelodysplastic Neoplasms (MDS) with Very Low and Low IPSS-M

Introduction: In addition to MDS cell-intrinsic genetic lesions, we and others found increasing evidence of dynamic alterations in both adaptive and innate immunity in the disease genesis and/or progression (Boy et al., Nat Commun 2023;14:588). Although a few reports have suggested a prognostic impact of absolute lymphocyte count (ALC) in both del(5q) and non-del(5q) MDS, lymphopenia is not captured by current prognostic scores. Moreover, the association between lymphopenia and somatic mutations has not been reported. In this work, we describe the prevalence of lymphopenia in MDS patients, the interaction with molecular features, and the impact of lymphopenia on patient outcomes. Methods: We retrospectively included all MDS patients followed in our center (Saint-Louis Hospital, Paris, France) between January 2015 and January 2022 with a molecular evaluation. According to the local laboratory threshold, lymphopenia was defined as ALC &amp;lt;1.5 G/L. Low (LR), intermediate (IR), and high-risk (HR) MDS were defined according to IPSS-M (LR: IPSS-M Low and Very Low, scores &amp;lt; -0.5; IR: IPSS-M Moderate Low and Moderate High, scores -0.5 to 0.5; and HR: IPSS-M High and Very High, scores &amp;gt;0.5 respectively).Survival analyses were estimated between MDS diagnosis until death from any cause (OS) or last follow-up, or until AML transformation (LFS) or last follow-up. Allogeneic stem cell transplantation was considered a censoring event. Results: Our cohort included 372 MDS patients (45% females, median age 72 IQR[65-79]) with 230 (62%), 70 (19%), and 72 (19%) patients with LR, IR and HR-MDS respectively. Compared to IPSS-R, molecular IPSS reclassified 175 patients (47%, with 97 (26%) upstaged and 78 (21%) downstaged). The median ALC in the whole cohort was 1.20 IQR[0.87-1.60] G/L. Lymphopenia was highly prevalent in all MDS subgroups (68%, 79% and 75% in LR, IR and HR-MDS respectively), and ALC inversely correlated with higher IPSS-M (Spearman rho=-0.13, p=0.01). Lymphopenia was associated with lower ANC (median 1.90 IQR[1.1-3.4] versus 2.70 IQR[1.6-4.2], p=0.01) and monocyte count (median 0.31 IQR[0.17-0.46] versus 0.42 IQR[0.25-0.78], p=0.04). Lymphopenia was not significantly associated with an excess of marrow blasts (78/267, 29% versus 21/105, 21% in patients with or without lymphopenia respectively, p=0.09). However, its prevalence was significantly lower in MDS with ringed sideroblasts (RS) (27/49 (55%) versus 240/267 (90%), p&amp;lt;0.01). Consistently, SF3B1 mutations were more prevalent in MDS patients without lymphopenia (29/107 (27%) versus 27/265 (10%), p&amp;lt;0.01), while the rest of the somatic mutational landscape did not differ between the 2 groups. The median follow-up of the entire cohort was 4.1 IQR[3.7-4.6] years. Among IPSS-M categories, lymphopenia adversely impacted OS and LFS in LR-MDS patients (median 8.8 years vs not reached for OS, p=0.02; median 12.3 years versus not reached, p=0.02 for LFS) but not in IR (p=0.34 and p=0.49 for OS and LFS respectively) or in HR-MDS (p=0.82 and p=0.83) patients ( Figure 1). In SF3B1 mutated LR-MDS patients (n=46), the adverse prognostic impact of lymphopenia at diagnosis was still significant for both OS (p=0.03) and LFS (p=0.03). Conclusion: In this single-center retrospective analysis, lymphopenia was highly prevalent at MDS diagnosis in all IPSS-M subgroups and ALC negatively correlated with disease severity. The presence of lymphopenia adversely impacted OS and LFS in the LR-MDS subgroup, potentially suggesting a significant role of host immunity in early disease phase and possibly adding a prognostic parameter to IPSS-M.

Prognostic Characterization of Advanced Mastocytosis Patients Treated with Midostaurin According to Diagnosis and Mutation-Adjusted Risk Score: A Nation-Wide Ceremast Study of 170 Patients

Introduction: Mastocytosis is a spectrum of diseases characterized by the accumulation of atypical mast cells (MC) in tissues. The most frequent form in adult patients is systemic mastocytosis (SM) including indolent SM, bone marrow mastocytosis, smoldering SM and advanced SM (adv-SM). Adv-SM encompasses aggressive SM (ASM), MC leukemia (MCL), SM associated with hematological neoplasms (SM-AHN) and is associated with a poor outcome. Prior the use of tyrosine kinase inhibitors (TKIs), therapeutic options were limited in adv-SM patients. The first TKI developed for adv-SM with KIT D816V mutation was midostaurin, which greatly improved outcomes for patients. However, most patients are primary refractory or relapse early (&amp;lt;12 months) after midostaurin onset. To identify high risk adv-SM patients, a prognostic score the so-called Mutation-Adjusted Risk Score (MARS) have been developed. However, given the heterogeneity of outcomes according to subtype of adv-SM, the identification of high-risk patients within each subtype is critical to optimize therapy. To address this question, we investigated predictive value on outcome of MARS score risk in midostaurin treated Adv-SM patients in a larger group of patients, according to each subtype of adv-SM. Methods: We conducted a nationwide retrospective study of all patients with Adv-SM patients included in the French national mastocytosis reference center registry (CEREMAST). Main inclusion criteria were: (i) adults patients treated with midostaurin in France since 2009 (ii) diagnosis of adv-SM according to WHO 2016 classification (iii) presence of C findings. Patients that have received combination therapy (i.e. azacytidine) or with chronic type of MCL without C finding or lymphoid neoplasm AHN or AML prior midostaurin onset were excluded. Response to midostaurin was assessed according to Valent criteria and MARS score was used to stratify patients into 3 risk groups (high risk, intermediate and low). Results: Overall, 170 patients treated with midostaurin were identified, including 46 ASM patients, 11 MCL patients and 113 SM-AHN patients. Among SM-AHN patients, chronic myelomonocytic leukemia was the most frequent neoplasm (52%). Response to midostaurin was significantly different regarding subtype of adv-SM (73% in ASM vs 27% in MCL vs 50% in SM-AHN, p=0.006). With a median follow-up of 19 months since midostaurin start, median overall survival (OS) was 69, 9.9 and 32 months (p=0.0013) and time to treatment failure (TTF) was 30, and 3.6 and 9 months in ASM, MCL and SM-AHN (p&amp;lt;0.0001), respectively. MARS risk group distribution was heterogeneous between ASM, MCL and SM-AHN (p=0.026). In MARS low risk group, median OS was not reached (NR) and 58 months in ASM and SM-AHN respectively (p=0.034, Figure 1). Median OS for both low risk ASM and low risk SM-AHN were significantly different compared with intermediate/high-risk (int/high-risk) groups (NR vs 33 months for ASM, p=0.003 and 58 months vs 24 months, p=0.017). Int/high-risk SM-AHN was the only group with a median OS not significantly different compared with MCL (24 months vs 9 months p=0.266) Median TTF was 60 months for low risk ASM, 26 months for low risk SM-AHN and only 6.7 months for high/int-risk ASM, 8.4 months for high/int-risk SM-AHN and 3.6 months for MCL. For the 19 patients who underwent hematopoietic stem cell transplantation (HSCT), median OS was 88 months but did not reach statistical significance compared to non-transplanted patients (47 months, p=0.15). In multivariate analysis, MARS and WHO mastocytosis subtype were the only variables independently associated with OS ( figure 2). MARS and midostaurin response were not predictive for both AHN progression and AML transformation in SM-AHN, which occurred in 31% and, 22% of patients respectively. In contrast, presence of portal hypertension/ascites (68% vs 29%, p&amp;lt;0.001) and abnormal karyotype (40 vs 11%, p=0.007) were predictive of AML transformation. Conclusions: We report herein the largest cohort of adv-SM patients treated with midostaurin. We identified portal hypertension/ascites and karyotype abnormalities as risk factors predictive of AML evolution. Subtype of adv-SM are heterogeneous and, together with MARS, are predictive for both OS and TTF. According to these variables, 5 sub-groups of adv-SM have been identified with different outcomes prompting for specific management.

Clinical Characteristics of Shwachman-Diamond Syndrome in China - Result from Childhood Bone Marrow Failure Diseases Register of China Alliance for Blood Diseases(cBMFR-CABD)

OBJECTIVE This study aimed to identify the clinical features of 73 children diagnosed with SDS from 22 centres in China. METHODS Childhood Bone Marrow Failure Diseases Register of China Alliance for Blood Diseases (cBMFR-CABD) was a bidirectional multicenter cohort study that collected data from 22 paediatric blood disease treatment centres from all over China. Patients were enrolled in the sub-centres from 1.1.2011 that were aged &amp;lt; 18 years at diagnosis and met the diagnostic criteria for an inherited bone marrow failure disorder. The clinical data for the 73 SDS patients analyzed in this study were collected between 1.1.2011 and 31.12.2022. RESULTS Among the 73 SDS children, 38 were male and 35 were female. Short stature (height&amp;lt;-2SD) was found in 53.4 % (39/73) patients. Chronic diarrhea was observed in 24.7 % (18/73) patients. In children evaluated for intellectual and physical malformations, 7 patients (17.3 %,6/52) had mental retardation and 14 (20.5 %, 15/73) patients had physical deformities. Genetic testing was available in 63 patients. seventeen (27.0 %) patients carry bi-allelic c.258+2T&amp;gt;C and c.183-184TA&amp;gt;CT SBDS mutations. Eleven patients (17.5%) are homozygous for the c.258+2T&amp;gt;C mutation. Four patients (6.3%) were compound heterozygous for c.258+2T&amp;gt;C and exon 2 deletions. One patient carries bi-allelic mutation of DNAJC21. This patient had a history of repeated infections and pancytopenia.Bone marrow smear shows multilineage dysplasia. Bone marrow results were available for evaluation in 54 patients including 67 % (36/54) BMF, 30 %(16/54) MDS, 4 % (2/54) AML.TP53 mutation testing was available in 15 patinets with eight (53.3%) patients carry TP53 mutation.Karyotype analysis was available in 36 patinets with 5 patients were complex karyotype. Among the 56 patients available for follow-up, 17 patients underwent haematopoietic stem cell transplantation, 5 patients died (one infection,three transplant-related death and one relapse). Among the 39 non-HSCT patients, 3 patients died of AML transformation. Three-year overall survival of all patients were 91.7%(95%CI 99.5 %~ 83.9% ), BMF patients were 100.0%(95%CI 100.0%~100.0% ), MDS/AML patients were 67.7%(95%CI 94.0%~ 41.4% ). CONCLUSION Our results provide the clinical features of the largest SDS cohort in China. A high incidence of short stature was observed. Bi-allelic c.258+2T&amp;gt;C with c. 183-184 TA &amp;gt; CT SBDS mutations were the most common mutation type. HSCT is an effective treatment modality in SDS. However, AML transformation is still a major risk factor for children with SDS.

Targeting Oncogenic Enhancers in ASXL1-Mutant Chronic Myelomonocytic Leukemia

Introduction: Chronic myelomonocytic leukemia (CMML) is a myeloid neoplasm with limited therapeutic options and dismal outcomes. Truncating mutations in the epigenetic modifier ASXL1 are common and lead to a proliferative disease phenotype, resistance to epigenetic therapies, and higher rates of AML transformation. ASXL1-mutant CMML is characterized by widespread epigenetic remodeling and up-regulation of key leukemogenic driver genes (e.g. HOXA9 and its co-factor MEIS1). There is a genotype-specific activation of enhancers in ASXL1-mutant CMML that is associated with up-regulation of these leukemogenic driver genes. Here, we test the hypothesis that disrupting the oncogenic interactions at these genotype-specific enhancers can be exploited for therapeutic benefit. Methods: We identified activated enhancers specific to ASXL1-mutant CMML as previously described ( Binder et al. Nat Commun 2022;13(1):1434). To disrupt the E2542605 enhancer, we created unique mapped single guide RNAs (confirmed by BLAT). We generated a ribonucleoprotein complex using the Alt-R CRISPR-Cas9 system (IDT Technologies, Coralville IA). We combined the RNP along with Electroporation Enhancer to 3x10^5 cells of the monocytic cell line U937 in 20uL strip-wells provided by the SF Cell Line 4D-Nucleofector X Kit (Lonza, Basel, Switzerland). To deliver the RNP, U937 cells were electroporated via listed cell line specific protocol by the Amaxa 4D-Nucleofector (Lonza). Forty-eight hours post-electroporation, cells were split and sequenced for E2542605 genomic structure disruption. In parallel, RNA was harvested, converted to cDNA and qPCR for HOXA9 and MEIS1 expression was performed using PrimeTime qPCR FAM labeled probe assays (IDT). Gene expression was determined using delta-delta Ct method. For assessment of selective sensitivity of ASXL1 / enhancer interactions, we performed progenitor colony forming assays using primary bone marrow mononuclear cells from CMML patients with or without ASXL1 mutations and healthy normal controls (n=2 in each category), exposed to increasing concentrations of the dual p300/CBP and BET bromodomain inhibitor EP31670. Briefly, we plated cells in methocellulose fortified with recombinant human cytokines supporting hematopoietic growth in duplicate, in a range of EP31670 concentrations. After 10 days, visible hematopoietic colonies were quantified and characterized. IC 50 values were generated using non-linear regression log(EP31670) vs. normalized response. Results: We computationally identified candidate transcriptional regulators enriched at the activated enhancers specific to ASXL1-mutant CMML, including BRD4 (FDR&amp;lt;2.57x10 -370), CBP (FDR=2.56x10 -217), and p300 (FDR=6.36x10 -93). We selected the ASXL1-mutant-specific enhancer E2542605 (predicted to drive HOXA9 expression), given the up-regulation of HOXA9 in ASXL1-mutant CMML (log 2-fold change +2.05, FDR=0.006, n=14). We disrupted E2542605 using two different CRISPR guides, leading to a PCR trace detection of a 14bp deletion and a 26bp indel in the E2542605 enhancer region, respectively. Both enhancer disruptions led to a down-regulation of its putative target gene HOXA9: Mean relative fold-change 0.36 (p=0.001, n=3) and 0.65 (p=0.046, n=3), respectively ( Fig. 1a). We then used a pharmacologic approach (EP31670) to target the candidate transcriptional regulators involved in the oncogenic interaction at these ASXL1-mutant-specific enhancers. We observed selective sensitivity to growth of CMML colonies (particularly ASXL1-mutant colonies) compared to ASXL1-wild type cases and normal bone marrow controls ( Fig. 1b). Conclusions: In CMML patients, truncating ASXL1 mutations are associated with the activation of genotype-specific enhancers and the recruitment of an extended repertoire of transcriptional regulators including BRD4, CBP, and p300. In a model system, the disruption of an enhancer predicted to govern the expression of the leukemogenic driver HOXA9, led to a significant down-regulation of HOXA9 expression. Pharmacologically disrupting the oncogenic interaction at such enhancers with EP31670 in primary patient samples led to significant therapeutic effects at nanomolar drug concentrations in a genotype-specific manner. Oncogenic interactions at activated enhancers are viable therapeutic targets in ASXL1-mutant CMML and warrant further clinical investigation.

Characteristics and Outcomes of Younger Adult Patients (Pts) with Myelodysplastic Syndromes (MDS): A Multicenter Retrospective Study

Introduction: MDS are primarily diseases of the elderly, with median age at diagnosis of 72 years. Focused data on clinical and molecular characteristics, as well as outcomes, in younger pts under age 60 years are limited. We conducted this multicenter retrospective study to describe pt characteristics, cytogenetic and molecular profiles, treatment modalities, and outcomes in younger pts with MDS, including adolescent and young adults (AYA). Methods: The study included pts diagnosed with MDS between 2002-2023 at 7 academic centers. We compared baseline characteristics, treatment, and outcomes of AYA (18-39 years) and younger adult (YA) pts (40-59 years). Responses were assessed using the modified IWG 2006 response criteria which included complete remission (CR), marrow CR (mCR), and hematologic improvement (HI). Wilcoxon rank sum test and Fisher's exact (and Chi-square) test were used to compare pt and disease characteristics for continuous and categorical variables, respectively. Overall survival (OS) was estimated using the Kaplan-Meier method and compared using log-rank test. Results: A total of 173 pts were identified (AYA, N=50; YA, N=123). Median age at diagnosis was 50 years (range, 18-59), with 51% females. Baseline characteristics are shown in Table 1. Therapy-related MDS (t-MDS) was observed in 40 pts (23%) with the remaining classified as de novo MDS (N=133; 77%). Inherited bone marrow failure syndromes were documented in 15 pts (9%), with Shwachman-Diamond Syndrome (N=4) and Fanconi anemia (N=4) being the most frequent. Using the WHO 2016 classification, the most common subtypes of MDS in both the AYA and YA cohorts were MDS with multilineage dysplasia (38% vs 19%), MDS with excess blasts-2 (MDS-EB) (24% vs 29%), and MDS-EB-1 (16% vs 20%). Similarly, using the WHO 2022 classification, the most common subtypes were MDS with low blasts (50% vs 21%), MDS with increased blasts-2 (MDS-IB-2) (24% vs 24%), and MDS-IB-1 (16% vs 18%). Stratifying pts using the IPSS-R cytogenetic (CG) risk group, the most common CG risk category identified was good CG (34% vs 52%, p=0.012). Poor/very poor CG were present in 34% and 37% of the AYA and YA groups, respectively. STAG2 and NRAS mutations occurred more frequently in the AYA pts compared to the YA group, while the YA cohort was enriched for the following mutations: TP53, DNMT3A, SF3B1, SRSF2, and TET2 (Figure 1). In the AYA population, 18 pts (38%) had very low/low, 10 (21%) intermediate, and 20 pts (42%) had high/very high risk MDS using IPSS-R. In the YA population, 32 pts (26%) had very low/low, 32 (26%) intermediate, and 57 pts (47%) had high/very high risk MDS using IPSS-R. Twenty-seven percent of pts with very low to intermediate-risk MDS using IPSS-R were upstaged to moderate high to very high-risk MDS using IPSS-M. The majority of pts (AYA: 75%; YA: 63%, p=0.5) were transfusion independent at baseline. Single-agent hypomethylating agents were the most frequently used frontline therapy, used in 17 AYA pts (34%) and 58 YA pts (48%), with more AYA pts receiving upfront allogeneic stem cell transplantation (Allo-SCT) compared to YA pts (28% vs 5%). After frontline therapy (Table 1), an additional 14 AYA (28%) and 52 YA (42%) pts received Allo-SCT. In pts with IPSS-R intermediate to very high-risk, response rates (HI+CR+mCR+mCR with HI) to frontline therapies were 46% and 54.5% in the AYA and YA groups, respectively. After median follow-up of 24 months [range, 1-218], median OS was numerically longer in the AYA versus YA group (155 vs 53 months, p=0.2). AML transformation occurred in 24% and 20% of the pts in the AYA and YA groups, respectively. Median OS was significantly longer in de novo compared to t-MDS pts (56 vs 27 months, p=0.03). Median OS among Allo-SCT compared to non-SCT pts were not reached vs 155 months ( p=0.9) in the AYA group and 53 vs 54 months ( p=0.4) in the YA group. IPSS-R (very low+low vs intermediate vs high+very high) was able to distinguish differences in OS in the YA (133 vs 53 vs 17 months, p=0.0002) and AYA (155 vs not reached vs 18 months, p=0.006) pts, with the exception of the intermediate-risk group in the AYA cohort. Conclusions: In this study, d e novoMDS was seen in the majority of younger pts with MDS (both AYA and YA). Mutations in STAG2 and NRAS were more common in the AYA pts, while YA pts had MDS more enriched for TP53, DNMT3A, TET2 and splicing mutations. This study is still ongoing with a plan to compare the abovementioned groups to older pts (≥60 years) with MDS.

Uproleselan Added to Cladribine Plus Low Dose Cytarabine (LDAC) in Patients with Treated Secondary Acute Myeloid Leukemia (TS-AML)

AML arising from HMA-treated myeloid disorders, also known as “treated-secondary AML” (ts-AML) is associated with an adverse prognosis (Complete Remission [CR] rates 15-30% and median Overall Survival [OS] 6-10 months). E-selectin, a marker of cell survival and resistance to chemotherapy, is highly expressed in the leukemic microenvironment. Exposure of leukemic blasts to HMAs has been shown to increase its expression. Uproleselan is an e-selectin antagonist that has demonstrated antileukemic activity in AML. We sought to study the combination of low-intensity chemotherapy with Cladribine + LDAC (CLAD/LDAC) with uproleselan to overcome local and microenvironmental resistance. This Phase Ib/II clinical trial (NCT04848974) evaluate the safety, tolerability, and efficacy of Uproleselan added to Cladribine and LDAC. A 3+3 dose-escalation approach was implemented to evaluate 2 different dose levels for Cladribine (CLAD)+ LDAC; each 4-week cycle consists of Uproleselan (at fixed dose of 800mg intravenously [IV]). Induction and reinduction if required, included IV uproleselan twice a day from day 1-12; consolidation was given same days but once a day if patients(pts) achieved response (complete response[CR] with incomplete hematological recovery [CRi] or morphologic leukemia-free state [MLFS]). CLAD was IV for 5 days (3.75mg/m2 and 5mg/m2; level -1 and 1 respectively) and subcutaneous LDAC twice daily 10 days (15mg, and 20mg; level -1 and 1 respectively) during induction; consolidation was similar except it was with 3-days of CLAD, for up to 6 cycles, pts aged ≥18 years with a diagnosis of ts-AML with adequate organ function, who have not received therapy for their AML were enrolled. ts-AML is defined as AML arising from previously treated myeloid neoplasm. Presence of the e-selectin was assessed as exploratory analysis. 20 pts have been treated, 18 pts evaluable: 14 (70%) male, median age was 72 yrs (range, 58-86); at the start of therapy, the median bone marrow blasts were 26.5% (20-82%), median WBC was 2.1x10 9/L (0.6-26.4), and median platelets were 26x10 9/L (4-667). Pts had received a median of 1 (1-6) treatments prior to AML transformation. Prior diagnoses were: Chronic Myelomonocytic Leukemia (CMML), MDS, and MDS/MPN in 3 (25%), 14 (70%), and 1 (5%) respectively; all had received HMA, 11 (55%) additionally had received Ven, and 5 (25%) had stem cell transplantation (SCT) prior to enrolling. All pts had unfavorable cytogenetics by ELN 2022. The most frequent mutations were: TET2, TP53, RUNX1 and SRSF2 in 9 (45%), 7 (35%), 6 (30%) and 6 (30%) pts respectively. 18 pts were evaluable for e-selectin; median plasmatic concentration was 26.2ng/mL (15.3-131 ng/mL). Most common AEs were grade(gr) ≥3 neutropenic fever (13 pts, 65%; including 2 gr 5 events), gr 3 bleeding (10%) and gr 2 thrombosis (5%). There were no dose-limiting toxicities observed on dose levels -1 or 1. The chosen dose level was 1 as described above. 2/3 pts who received dose level -1 died during the study follow-up due to sepsis in the first 4-week after induction. Median time to 0.5x10 9/L neutrophil and 50x10 9/L platelets recovery was 30 (17-52) and 38 (20-69) days, respectively. The response rates were 2 (11%) CRi, 1 (6%) CRp, and 4 (22%) MLFS. There was a reduction in BM blasts in 13 pts (72%) (Figure 1). The ORR was 50%(5/10) ( p=0.60) and 20% (1/5) ( p=0.06) among pts who had prior Ven exposure or prior SCT, respectively. The median cycles received was 1.5 (1-4), median cycles at which the best response was achieved was 1 (1-2). 12 pts were taken off protocol due to progression, 3 for death, 3 for allogeneic SCT and 1 continued in remission. The pt who achieved negative MRD-negative by flow cytometry underwent SCT and is still alive. The median follow-up is 8.1 months. Median OS and EFS were 5.3 and 1.4 months respectively; 4-month RFS (CRi, CRp and MLFS) was 30% (Fig 2). In the univariate analysis by Cox regression for plasmatic E-selectin concentration the HR for OS was 1.016 (95% CI 1.002-1.031). Ts-AML, progressing from previously treated antecedent myeloid disorders, is associated with low rates of remission and poor OS. Uproleselan combined with Cladribine + LDAC was well-tolerated with minimal therapy-related AEs - allowing a safe approach to marrow blast reduction and disease control in preparation for a potential allogeneic SCT. We are currently determining the relationship of plasmatic e-selectin concentration and response to treatment.

Ratio of stemness to interferon signalling as a biomarker and therapeutic target of myeloproliferative neoplasm progression to acute myeloid leukaemia.

Progression to aggressive secondary acute myeloid leukaemia (sAML) poses a significant challenge in the management of myeloproliferative neoplasms (MPNs). Since the physiopathology of MPN is closely linked to the activation of interferon (IFN) signalling and that AML initiation and aggressiveness is driven by leukaemia stem cells (LSCs), we investigated these pathways in MPN to sAML progression. We found that high IFN signalling correlated with low LSC signalling in MPN and AML samples, while MPN progression and AML transformation were characterized by decreased IFN signalling and increased LSC signature. A high LSC to IFN expression ratio in MPN patients was associated with adverse clinical prognosis and higher colony forming potential. Moreover, treatment with hypomethylating agents (HMAs) activates the IFN signalling pathway in MPN cells by inducing a viral mimicry response. This response is characterized by double-stranded RNA (dsRNA) formation and MDA5/RIG-I activation. The HMA-induced IFN response leads to a reduction in LSC signature, resulting in decreased stemness. These findings reveal the frequent evasion of viral mimicry during MPN-to-sAML progression, establish the LSC-to-IFN expression ratio as a progression biomarker, and suggests that HMAs treatment can lead to haematological response in murine models by re-activating dsRNA-associated IFN signalling.

Characteristics and survival of secondary acute myeloid leukemia arising from myelodysplastic syndrome (MDS-sAML).

e19075 Background: sAML is a heterogenous category including therapy-related AML, AML arising from MDS or myeloproliferative neoplasm, and other biologically distinct groups. Combining these as one entity likely masks important clinical and therapeutic differences. We defined the clinical profile and survival for MDS-sAML. Methods: The population consisted of patients (pts) with MDS identified in SEER-Medicare (2007-17) by ICD-O-3 histology codes and continuous Medicare parts A+B coverage. Transformation to leukemia was confirmed by at least 2 AML codes during follow up. Established algorithms identified MDS type and risk group, Charlson comorbidity index (CCI), and transfusion dependence for red blood cells (RBC) and platelets. Logistic regression identified risk factors for transformation to MDS-sAML. Kaplan-Meier estimation and multivariable Cox regression were used to analyze overall survival (OS). Results: Of 15,227 MDS pts, 1,871 transformed to AML. MDSsAML incidence varied by MDS risk group and histology. Time to transformation was shorter with higher risk groups. Younger age (odds ratio [OR] = 1.6, 95% confidence interval [95 CI]: 1.4-1.9), lower CCI (OR = 1.6, 95 CI: 1.4-1.9), higher risk (OR = 2.3, 95 CI: 2.0-2.5), transfusion dependence for RBC (OR = 1.2, 95% CI: 1.1-1.3) or platelets (OR = 2.5, 95 CI: 2.1-3.0), were associated with transformation (all respective p &lt; 0.05). Median OS (mOS) after transformation was 3.3 months. The 1- and 2-year OS rates for MDS-sAML were 25% and 12%. MDS-sAML survival varied by antecedent MDS histology and risk group, being longer for pts with antecedent low risk MDS (p &lt; 0.001). Sex, race, and CCI did not affect survival after transformation. Older age (hazard ratio [HR] = 1.5, 95 CI: 1.2-2.0), higher MDS risk (HR = 1.5, 95 CI: 1.2-1.9), RBC transfusion dependence (HR = 1.4, 95 CI: 1.3-1.6), and platelet transfusion dependence (HR = 1.5, 95 CI: 1.3-1.7), were associated shorter MDS-sAML survival (all respective p &lt; 0.05). Conclusions: Risk of MDS-sAML is substantial, with 5% AML transformation even in lower risk MDS histology. Real-world survival of MDS-sAML pts is dismal and lower than reported in clinical trials. Worse MDS risk group before transformation is associated with lower OS after developing AML. These data support a unique pathophysiology of MDS-sAML. [Table: see text]

P062 - Topic: AS04-MDS Biology and Pathogenesis/AS04f-Gene expression profiling: CMML PATIENTS WITH LOW EXPRESSION OF TP53 POINT TO AML TRANSFORMATION

P062 - Topic: AS04-MDS Biology and Pathogenesis/AS04f-Gene expression profiling: CMML PATIENTS WITH LOW EXPRESSION OF TP53 POINT TO AML TRANSFORMATION

Del5q with One Additional Cytogenetic Abnormality Other Than Del20q Identifies a Higher Risk Population of MDS Patients

Background: According to the IPSS-R cytogenetic risk classification for myelodysplastic syndrome (MDS), patients with isolated deletion 5q (del5q), isolated deletion 20q (del20q), and del5q with an additional cytogenetic abnormality are classified as having good cytogenetics, similar to those with a normal karyotype. However the international database study, on which the IPSS-R cytogenetic risk classification is based, has its limitations in the small number of patients with del5q plus one additional cytogenetic abnormality (n=46). We sought to further define the prognostic significance of del5q with one additional cytogenetic abnormality in an evaluation of 86 patients. Methods: We performed a retrospective analysis of 857 MDS patients with a karyotype of isolated del20q (n=160) or any karyotype involving del5q seen at a single tertiary cancer center between 1987-2021. Patients with del5q (n=697) were divided into 3 cohorts: isolated del5q (n=147), del5q with concomitant del20q (del5q+del20q, n=8), and del5q with a non-del20q cytogenetic abnormality (del5q+1, n=78). The remainder of del5q patients with complex karyotypes were excluded. Patient characteristics, laboratory values, and bone marrow data, including blast percentage and cytogenetics, were reviewed. Median OS was estimated using the Kaplan-Meier method and two-tailed log-rank test. Results: The median age of patients was 70 years, with the majority being male (58%). Patients were more likely to present with lower risk disease (IPSS low to int-1, 78% of patients). The median follow-up was 2 years. Patients with del5q+1 had an inferior survival when compared to patients with isolated del5q (median OS 1.78 years vs. 3.87 years, p=0.0047). Del5q+1 patients also had a higher risk of AML transformation (26% vs. 16%, p=0.098, table I) with a shorter median time to AML transformation (1.07 years vs. 3.46 years, p<0.0001, figure 1). A multivariate analysis of the combined del5q and del5q+1 cohort was performed, which evaluated age >60, percent blasts in the bone marrow, IPSS score, and cytogenetics. The only statistically significant prognostic factors for OS with a p-value <0.01 were age>60 (HR: 3.67 (2.24-6.0), p<0.0001 ) and cytogenetics (HR for del5q+1: 1.96 (1.3-2.9), p=0.001). The only prognostic factor for time to AML transformation with a p-value <0.01 was cytogenetic classification (HR for del5q+1: 5.2 (2.0-13.2), p=0.0006). Patients with del20q had an OS between that of del5q (2.63 years vs. 3.87 years, p=0.14) and del5q+1 (2.63 years vs. 1.78 years, p=0.18). Del20q patients were less likely to transform to AML when compared to del5q+1 patients (11% vs. 26%, p=0.0035) with a trend towards longer time to AML transformation (2.1 years vs. 1.07 years, p=0.09). There were no statistically significant differences in AML transformation between the del5q and del20q cohorts (20% vs. 11%, p=0.1; median time to AML transformation: 3.46 years vs. 2.1 years, p=0.1). The evaluation of OS and risk of AML transformation in patients with del5q and a contaminant del20q was limited due to low patient numbers (n=8, 1.1%). The median follow-up for these patients was 2.67 years, and none of the patients transformed to AML during this time. Two patients (25%) expired from the non-disease related causes of presumably underlying heart failure and massive pulmonary embolism following surgery. Interestingly, both deceased patients had >10% bone marrow blasts and were IPSS intermediate risk. The median OS for these 8 patients was not reached. However, the median follow-up time of 2.67 years may suggest their OS is better than those with del5q+1 (median OS ≥2.67 years vs. 1.78 years). Conclusion: Contrary to the suggested IPSS-R cytogenetic risk stratification scheme, this analysis demonstrates a significant difference in terms of OS and risk of AML transformation when patients with isolated del5q or isolated del20q were compared to patients with del5q+1. Patients with del5q+1 had inferior median OS and a shorter time to AML transformation, suggesting more aggressive management may be warranted in this population. Though rare, patients with del5q+del20q may have better clinical outcomes than those with del5q+1, warranting further investigation into this group of patients. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Pediatric Myeloproliferative Neoplasm in Era of Who Classification Systematic Review

Introduction Philadelphia-negative MPNs are most prevalent in the older population (median age at the diagnosis above 60 years) and are rarely diagnosed in pediatrics. It is estimated that the overall incidence of MPNs in pediatric patients is around 0.82 per 100,000 patients yearly, and MPNs are approximately 100 times more common in adults than children. Methods: We performed a systematic review following the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines. English literature (PubMed, SCOPUS, and Google Scholar) was searched for studies, reviews, case series, and case reports of patients with Philadelphia negative MPNs (including essential thrombocythemia, polycythemia vera, primary myelofibrosis) in the pediatric age group (< 18 years). Only studies that fulfilled WHO-2008 or 2016 criteria for MPN were included. Results: We reviewed 33 articles of available published literature from 2008 to 2022 and collected data on a total of 196 patients in the pediatric population. Among the cohort of patients, 139 had essential thrombocythemia (ET), 20 had polycythemia vera (PV), and 37 had myelofibrosis (MF). Patients' clinical and biological characteristics are addressed in Table 1. The median age at the time of diagnosis for each disease varied, with 8.8 years for ET, 10 years for PV, and 3.6 years for MF. There was a variety in the blood counts at the time of diagnosis between all three diseases (Table 1). Discussion: Based on our review, the presence of splenomegaly pediatrics with PV was not mentioned in most cases, yet around a third of the reported data have splenomegaly, matching the percentage found in adults. On the other hand, hemorrhage was not observed in pediatrics with PV, thrombosis complicated in 2 (10%) cases, and leukemic transformation was documented in 2 (10%) cases. The number of PV cases in this review is small, with only 20 reported cases, so it is challenging to get conclusions, yet the presentation and complications between adults and pediatrics are comparable, apart from a possibly higher chance for AML transformation in the pediatric population. In adults with PV, around 98% of the patients were positive for either JAK2V617F in 95% or other JAK2 mutations in 3%. While in this pediatrics review, JAK2V617F mutation was found in only 45% of the cases and JAK2 exon 12 mutation in 15%. 36% of the ET cases were asymptomatic at presentation, a number lower than that observed in adults (45%). However, thrombosis is less common in pediatrics than in adults (9% versus 21%), and bleeding as a complication from ET in pediatrics was equal to adults (9%). JAK2V617F was the most common mutation (20% of the ET patients). Based on our review, MF cases in pediatrics are more common than PV cases in comparison with adults, in which MF is the least common. Data for pediatric patients with PMF is scarce; based on this sample of 37 patients, the percentage of symptomatic patients is higher than adults, but unlike adults, most pediatrics are presented with symptoms of cytopenia rather than constitutional symptoms, same for palpable splenomegaly where it was more reported compared to adults. The pediatric population with PMF has an entirely different mutation profile, with none of the patients in our data having JAK2V617F or MPL mutations. Interestingly in our collected data, none of the patients with PMF developed thrombosis or leukemic transformation, compared with 7% of adult patients who have documented leukemic transformation. Conclusion: Given the rarity of MPNs in pediatrics and different characteristics compared with adults, international collaborative efforts to study MPNs in this age group are needed to understand these disorders better. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Single-Cell Multiomics Reveal Hypomethylating Agent Partially Restores Hematopoiesis in a LR-MDS Patient at the Cost of Leukemic Transformation

Background Myelodysplastic syndromes (MDS) are a heterogeneous group of diseases characterized by ineffective hematopoiesis. Hypomethylating agents (HMA) such as decitabine and azacytidine (AZA) induce clinical responses in both lower-risk (LR) and higher-risk groups; however, the treatment eventually fails and can be followed by leukemic transformation. The mechanism underlying transient clinical benefit from low-dose HMA (restoration of hematopoiesis) is not fully understood. Here we used cutting-edge single-cell multi-omic approaches to describe the abnormal clonal architecture and transcriptome in LR-MDS and how they are altered following HMA therapy. Results First, we defined genomic differences between aged and young healthy hematopoietic stem and progenitor cells (HSCPs) by building a single-cell atlas of human HSCP with molecularly-defined concentrations of relevant CITE seq antibodies. 350k high-quality bone marrow nucleated cells were obtained from four young donors; varying race and gender. A 6-fold molecular titration of 278 CITE-seq antibodies determined those which are functional and which contribute to clustering. Next, we repeated the captures on samples from aged healthy people (n=3), MDS patients (n=4) and conducted differential analysis on cell states and transcriptomes. Compared to young marrow, healthy marrow from elderly donors revealed global upregulation of inflammatory signals and innate immunity. In erythroid progenitors (ERP) we identified abnormally high expression of the transcription factors KLF1 and GATA1, but lower than normal expression of their hemoglobin target genes. At the same time altered erythropoiesis in the elderly appeared to be compensated by increased frequency of multilineage progenitors and more mature erythroblasts mitigating clinical anemia. Comparing MDS to aged marrow, while most cell populations from MDS showed significant gene expression changes, the amplitude was dwarfed by abnormal mRNA processing gene expression in ERP and erythroblasts. Moreover, in de novo LR-MDS samples and an anemia sample (without known mutations) further downregulation of hemoglobin genes in the erythroid lineage was accompanied by decreased erythroblast numbers leading to anemia. To study the impact of AZA on LR-MDS, we longitudinally profiled a single MDS patient (pre-HMA, post-HMA, sAML transformation, cycle 2 VEN+AZA) (Panel 1). We found that 6 cycles of AZA treatment depleted HSC; whereas lymphoid-primed multipotential progenitors (LMPP), megakaryocyte-erythroid progenitors (MEP), and less mature erythroblasts were enriched after AZA therapy (Panel 2 left). Using genotyping of the transcriptome (GoT) (Panel 2 right), we defined cells with mutations to find that not only SRSF2-relevant pathways (eg. mRNA processing, and mRNA splicing) are molecularly corrected (downregulated) but also there are fewer mutant cells among primitive stem cells and lineage-committed progenitors. However, mutant ERP and erythroblasts accumulated and pathways were more active suggesting the blockade of differentiation at terminal erythropoiesis is not rescued, consistent with the clinical observation that the patient was still RBC transfusion-dependent despite platelet recovery. Notably, these mutant erythroid cells, while defective, never transformed to leukemia. The expansion of mutant multilineage progenitors and LMPP preceded AML transformation and were eliminated (as evidenced by flow and CITE-seq) following two cycles of VEN+AZA. Conclusions: Age-matched references are critical in differential analysis of hematological diseases like AML or MDS. With matched elderly donor data sets, we were able to molecularly isolate functional disruption of erythropoiesis and megakaryopoiesis underlying anemia and thrombocytopenia, respectively, observed in an LR-MDS patient. AZA rescued the dysregulated signals in SRSF2-mutated LR-MDS patient HSPCs, but not in ERP and erythroblasts, and therefore only partially restored hematopoiesis. However, as AZA drove differentiation of both wildtype and mutant HSCP, accumulation of mutant multilineage progenitors and LMPPs contributed to progression to AML. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Final Results of Ruxopeg, a Phase 1/2 Adaptive Randomized Trial of Ruxolitinib (Rux) and Pegylated Interferon Alpha (IFNa) 2a in Patients with Myelofibrosis (MF)

Background MF is characterized by splenomegaly, anemia, debilitating symptoms and bone marrow (BM) fibrosis. Rux is the current standard of care improving symptoms and splenomegaly but with little impact on the malignant clone and BM fibrosis. IFNa can reduce mutant allele burden and fibrosis but is poorly tolerated in highly symptomatic patients (pts). The RUXOPEG study was designed to assess the efficacy and safety of different doses combinations of Rux + pegylated IFNa 2a in both JAK inhibitor and IFNa-naïve MF pts (NCT02742324). Methods RUXOPEG is a multi-center Bayesian Phase 1/2 adaptive trial. Phase 1 (P1) evaluated 9 possible combinations of doses of Rux, and IFNa (10, 15 and 20 mg BID, and 45, 90 and 135 mcg/week, respectively) and included 6 cohorts of 3 pts. Phase 2 (P2) randomized the 2 best dose combinations selected from P1. Primary tolerance criterion was the occurrence of dose limiting toxicities (DLT) within 45 days in P1. Primary efficacy criterion was >50% reduction in spleen length (SL) by palpation within 24 weeks (W). Secondary objectives included molecular response, quality of life and symptoms evolution, reduction of BM fibrosis, event-free and overall survival. Key inclusion criteria: diagnosis of MF (WHO), Intermediate (Int) or high risk (HR) (IPSS), need of active therapy. Key exclusion criteria: prior treatment with Rux or IFNa, eligibility for stem cell transplantation, autoimmune disease, history of depression. Enrolment in P1 was completed in Nov 2018 and in P2 in Nov 2020. We report the data collected after last pt last visit after 12 months in the study (Nov 2021). Results A total of 37 pts (18 in P1, 19 in P2) were included from 6 centers. Median (range) age was 63.5 years (37-71), 21 had primary MF, 16 post ET or post PV MF. Median (range) SL was 9 cm (0-20) by palpation and 19.5 cm (10.6-30) by imaging. Mean (range) hemoglobin was 12.2 g/dL (7.8-16.2), WBC 12.2 G/L (3.5- 36.1), platelets 372 G/L (150-1304). IPSS at inclusion: Int-1 in 23, Int-2 or HR in 14 pts. Mean number of mutations (mut) per pt was 3.4.; 21 pts had JAK2V617F, 11 CALR and 4 MPL mut; NGS identified 90 additional mut in 17 genes in 33 pts, including mut in HR genes: ASXL1 (n= 21), EZH2 (5), SRSF2 (2), IDH1 (1), and TP53 (5). Safety: No DLT occurred among the 18 pts included in 6 cohorts of P1 (primary safety criterion), and 19 pts were randomized between the 2 dose combinations selected for P2: IFNa 135 mcg/w + Rux 15 (level 6, 9 pts) or 20 (level 9, 10 pts) mg BID. Over 24W, a total of 363 AEs (59% gr1, 33% gr2, 7.5% gr3, 0.5% gr4) including 6 SAE (2 skin cancers, 2 anemias, 1 urinary infection, 1 Raynaud's phenomenon) were reported. Only 2.5% of them resulted in treatment interruption. Most frequent AEs were anemia (18.7%), thrombocytopenia (13.7%), GI (7%), musculoskeletal (6.8%) and asthenia (6.6%). During the 12-month period of the study, 1 patient died of AML transformation after 10.4 months. Efficacy: Primary efficacy endpoint (reduction by at least 50% of SL within 24 W) was reached in 70% (IC95%: 53;84) of pts (26/37 pts in ITT, 67% in P1, 74% in P2) with a clear decrease in SL at W24 (median 3.5 cm by palpation, range 0-12) (Fig 1), maintained at 12 months (median SL of 3 cm, range 0-12) (Fig 2) including complete resolution of palpable splenomegaly in 38% of pts. The best response rate (83%) was seen in patients receiving dose level 6. JAK2V617F allele burden decreased from a median (range) of 84% (23-96) at baseline to 65% (16-95), and 53% (16-92) after 6, and 12 months. Genotyping of progenitor-derived colonies performed in 10 patients showed that the combination of drugs clearly targets in 12 months committed (CD34+CD38+), immature (CD34+CD38-) and HSC-enriched progenitors (CD90+CD34+CD38-) carrying homozygous or heterozygous JAK2V617F. BM biopsies at inclusion and after 12 months are currently centrally reviewed (preliminary results show reduction of BM cellularity and grade of fibrosis in selected pts), and results of additional exploratory endpoints including evolution of clonal architecture with single cell and NGS sequencing, cytokine array and changes in immune effector cells will be presented. Conclusion RUXOPEG is the first study to formally assess the safety and efficacy of Rux + pegylated IFNa 2a combination in patients with MF never exposed to either drug before. Final results show that the combination provided high rates of important reductions in SL and JAK2V617F allele burden due to selective targeting of mutated progenitors. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Hypoplastic MDS: Validation of the New World Health Organization 2022 Proposal

Introduction: Myelodysplastic syndromes (MDS) are a heterogeneous group of hematologic neoplasms that is characterized by ineffective hematopoiesis. The morphological diagnosis is essential and reveals a subset of patients with hypoplastic marrow, referred to as hypoplastic MDS (MDS-h) that is more recently being proposed as a distinct entity in the World Health Organization (WHO) 2022 proposal for myeloid neoplasms. MDS-h shares several features with aplastic anemia (AA) and paroxysmal nocturnal hemoglobinuria (PNH), thus, it needs a careful diagnostic approach, but also, the natural history of the disease is distinct than those with non-hypoplastic MDS (MDS-nh) warranting consideration for further subclassification. Here, we aim to study the characteristics and molecular profiling of MDS-h as opposed to the (MDS-nh), in addition to treatment and disease outcomes, and overall survival. Methods: We conducted a retrospective analysis of all patients diagnosed with MDS at the H. Lee Moffitt Cancer Center & Research Institute from our MDS database. Patients were divided into two groups: those with MDS-h, defined as bone marrow cellularity of less than 30%, and those with MDS-nh, with cellularity greater than 30%. We further subdivided patients based on the marrow blasts (<5% vs >=5%). Patients with MDS/MPN were excluded. Results: A total of 4037 patients with MDS were identified, of those, 403 (10%) were MDS-h. The median age at diagnosis was 65 and 68 for MDS-h and MDS-nh respectively (P= 0.03). In females, MDS-h was observed more compared to MDS-nh (41% vs 35%, P= 0.008). There was no difference in terms of IPSS-R, LGL clones, PNH clones, and history of autoimmune diseases. MDS-RS was more observed in MDS-nh. Interestingly, therapy-related MDS (t-MDS) was more observed in MDS-h (29% vs 20%, P <0.005). Patients with MDS-h had more thrombocytopenia, neutropenia and were more likely to be platelet transfusion dependent (31% vs 23% P=0.001). The following somatic mutations were found more frequently in MDS-nh compared to MDS-h: SF3B1 (P<0.005), SRSF2 (P<0.005), TET2 (P=0.004), and ASXL1 (P=0.02). There was a better response to ATG/CSA among MDS-h compared to MDS-nh (46% vs 21% P=0.019). On the contrary patients with MDS-nh were more likely to respond to lenalidomide when compared to patients with MDS-h (46% vs 22% P=0.001). There was no statistically significant difference in response to hypomethylating agents. The median overall survival (OS) for MDS-h was 41.6 months compared to 35 months in patients with MDS-nh (P=0.05). The HR for OS was 0.88 (P =0.057) adjusted for IPSS-R. There was no difference in the rate of AML transformation. Then, we further categorized patients based on the marrow blasts. Among MDS-h, 43% (170/400) had myeloblasts >=5% (MDS-EB), The median OS was 53 months for MDS-h with < 5% blasts (n=231) compared to 27 months for those with >= 5% blasts (n=170) (P < 0.005). Among all patients with MDS-EB, those with MDS-h (n= 170) had median OS of 27 month compared to 19 months for MDS-EB with MDS-nh (n=1465) (P=0.02). Conclusions: In summary, we conclude that MDS-h accounted for 10% of MDS cases. The OS for MDS-h is better than MDS-nh. Additionally, MDS-h tended to have lower frequency of splicing and epigenetic somatic mutations. Among MDS-h, patients with marrow myeloblasts >=5% clearly had inferior outcomes compared to those MDS-h with blasts <5%. Therefore, our findings support the new WHO 2022 proposal for MDS-h as a distinct entity with restriction to patients with marrow cellularity of <30% and marrow myeloblasts <5%.

Understanding What Matters to Myelodysplastic Syndrome Patients - a Study of Preferences for Treatments with Hypomethylating Agents

Introduction/Background: Hypomethylating agents (HMAs) are the standard of care for the treatment of myelodysplastic syndrome (MDS). Understanding how patients with MDS perceive and value different HMA characteristics provides critical treatment selection insight for healthcare providers. Objectives: To understand HMA preferences for United States (US) and Canadian patients with MDS. Given HMAs’ treatments’ similar efficacy and safety profiles, we also investigated how HMA administration burden can play a role in patients’ preferences when all other factors are equal. Materials and Methods: We used the discrete-choice experiment (DCE) method to conduct a preference study in collaboration with patient organizations in the US and Canada (i.e., MDS Foundation, the Aplastic Anemia and MDS International Foundation, and the Aplastic Anemia and Myelodysplasia Association of Canada). The study was comprised of two Phases: (1) a qualitative Phase (i.e., DCE survey design) and (2) a quantitative Phase (i.e., DCE survey implementation). In Phase 1, literature review, interviews with clinician, patient and caregiver, and input from the patient organizations informed the DCE survey design. Qualitative interviews were analyzed through content analysis. In Phase 2, survey participants answered a series of DCE choice tasks to indicate their preference between different hypothetical HMA profiles that varied in characteristics (i.e., attributes). Respondents were also asked to indicate their preferences by choosing between HMA profiles mirroring real-life options, all associated with the same risk of AML transformation and level of fatigue but differing in mode of and frequency administration: IV HMAs requiring 5 days or 7 days of treatment session each month, SC HMA with 5 sessions each month, oral HMA and no treatment. Survey results were analyzed using random parameter logit models. The relative importance of attributes (RAI) was calculated, which is the contribution (expressed in percentages) of each attributes relative to all attributes in patient preferences. A larger RAI percentage score shows that the attribute has a larger effect on how the respondents make their decisions. All patient respondents were recruited through the patient organizations’ networks. Results: 16 Phase 1 respondents completed the interview and 184 Phase 2 respondents (including 158 patients and 26 caregivers serving as patient proxies) completed the DCE survey. Interview respondents in Phase 1 valued efficacy attributes (i.e., risk of AML transformation and fatigue) and expressed the treatment burdens associated with injectable treatments (such as injection site reaction, stressful experience of damaged veins, burden of travelling to clinic, etc.). The Phase 2 survey participants had a mean age of 67.2 years old and 88% were from the US. About one-third of the survey participants did not know their MDS risk, 35% were low risk and 31% were high risk. Most (56%) participants were currently receiving treatment, while 24% had received treatment but discontinued and the remaining 20% had not received treatment. The attributes’ importance rankings (via RAI) were risk of AML transformation (40%), fatigue level (33%), number of visits (12%), mode of administration (6%), visit duration (5%), and administration frequency (4%). When HMA treatments were assumed to be associated with the same risk of AML and level of fatigue but to differ in terms of mode and frequency administration, 77% of respondents were predicted to switch to an oral pill if available to them. When considering real-life profiles, 77% and 88% of participants considered the oral option to be the best choice and no treatment to be the worst one, respectively. Conclusion: MDS patients demonstrated strong preferences for HMA with lesser treatment burden (i.e., oral pill) when such agents are with similar treatment efficacy and safety profiles. Understanding patients’ treatment preferences can facilitate shared decision making towards optimal management of MDS.

Uproleselan Added to Cladribine Plus Low Dose Cytarabine (LDAC) in Patients with Treated Secondary Acute Myeloid Leukemia (TS-AML)

Background Treated secondary AML (TS-AML), arising after prior HMA-treated MDS, is associated with very poor prognosis (Complete Remission [CR] rates 15-30% and median Overall Survival [OS] 6-8 months). E-selectin ligand is highly expressed on AML blasts in the leukemic microenvironment and may be a marker of cell survival and resistance to chemotherapy. Exposure of leukemic blasts to HMAs has been shown to increase their expression of E-selectin ligand. Uproleselan is an E-selectin antagonist that overcomes resistance to chemotherapy in AML (Barbier, Nat Commun 2020). We sought to study the combination of low-intensity chemotherapy with Cladribine + LDAC (CLAD/LDAC) with uproleselan to overcome local and microenvironmental resistance and improve outcomes in this difficult subset. Methods This is Phase Ib/II clinical trial (NCT04848974) to evaluate the safety, tolerability, and explore the efficacy of Uproleselan added to Cladribine and LDAC. A 3+3 dose-escalation approach was implemented to evaluate 2 different dose levels for Cladribine (CLAD)+ LDAC; each 4-week cycle consists of Uproleselan (at a fixed dose of 800mg intravenously [IV]) added to IV CLAD 5 days (3.75mg/m2 and 5mg/m2; level -1 and 1, respectively) and subcutaneous LDAC twice daily 10 days (15mg, and 20mg; level -1 and 1, respectively) during induction; consolidation was similar except it was with 3-days of CLAD, for up to 6 cycles. Pts aged ≥18 years with a diagnosis of TS-AML with adequate organ function, who have not received therapy for their AML were enrolled. TS-AML is defined as AML arising from a previously treated myeloid neoplasm. Presence of the E-selectin ligand was assessed using Flow Cytometry (FC). Results 10 pts have been treated, with 9 pts currently evaluable: 6 (67%) were male and the median age was 68 years (range, 58-80); at the start of therapy, the median bone marrow blasts were 33% (1-78%), median WBC was 2.2x109/L (0.6-20.1), median platelets were 18x109/L (4-305), and median creatinine was 0.98mg/dL (0.67-1.52). Pts had received a median of 1 (1-2) treatments prior to AML transformation. Prior diagnoses were: therapy-related Myelodysplastic Syndrome (t-MDS), Chronic Myelomonocytic Leukemia (CMML), MDS and MDS/MPN in 3 (33%), 3 (33%), 2 (22%) and 1 (11%) respectively; all had received HMA, 5 (56%) additionally had Ven and 3 (33%) had stem cell transplantation (SCT) prior to enrolling. All pts had unfavorable features by ELN 2017. The most frequent mutations were:SXL1, TP53 and TET2 in 4 pts each (44%), SRSF2 and NRAS in 3 patients each (33%) and SETBP1, RUNX1 and EZH2 in 2 patients each (22%). 6 pts were evaluable for E-selectin ligand expression; the median expression was 59% (42%-95%) and median MFI was 20.5 (13-262). The most common SAEs were ≥ grade 3 neutropenic fever (70%), (including 2 grade 5 events), grade 3 bleeding (10%), and grade 2 thrombosis (5%) (Table 1). There were no dose-limiting toxicities observed on dose levels -1 or 1. Two pts treated on dose level -1 die during the study follow-up due to sepsis within the first 4-weeks during induction. Median time to 0.5x109/L neutrophil and 50x109/L platelets recovery was 29 (17-39) and 38 (33-48) days respectively. The median follow-up is 4+ months. 8 pts were evaluable for response at the time of analysis. The ORR was 62% (5/8), including 2 (25%) PR, 1 (13%) CRi, 1 (13%) CRp and 1 (13%) MLFS. There was a reduction in BM blasts in 6 pts (75%) (Figure 1). 5 pts were taken off protocol due to progression, 2 for death, 1 for allogeneic SCT and 1 continued onto maintenance in remission. The one pt who achieved negative MRD, underwent SCT and is still alive. Median OS and EFS were 4.7 and 1.3 months respectively; 4-month RFS (CRi, CRp, and MLFS) was 67%. The median cycles received was 1 (1-3), median cycles at which the best response was achieved was 1 (1-2). The 4-month OS were 100% and 60% among responders vs. non-responders, respectively (p=0.27), and the 4-month EFS were 50% and 0% respectively (p=0.01). The ORR was 40% (2/5) (p=0.85) and 33% (1/3) (p=0.85) among pts who had prior Ven exposure or prior SCT, respectively. Conclusions The combination of Cladribine + LDAC with Uproleselan was overall well tolerated with few treatment-related AEs. The combination produced an ORR of 62% in a high-risk, refractory population whose prognosis is very dismal. The relationship of E-selectin ligand expression, response to treatment, and outcomes is being analyzed. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal