Peripheral Blood Blast Count Research Articles

Outcome of the Modified St. Jude Total XV Protocol in Turkish Children with Newly Diagnosed Acute Lymphoblastic Leukemia: A Single-Center Retrospective Analysis

The prognostic factors and outcomes of Turkish children with newly diagnosed acute lymphoblastic leukemia (ALL), treated with the Modified St. Jude Total XV Protocol, which was adjusted by adding high-dose methylprednisolone (HDMP) before induction in the original protocol, were assessed in this study. The Modified St. Jude Total XV Protocol was administered to 183 newly diagnosed ALL patients, aged 1-18 years, between 1 January 2008 and 30 January 2016. HDMP was applied at doses of either 10 mg/kg/day (Group A) or 20 mg/kg/day (Group B) for 7 days before induction and then tapered over the next 7 days to 5 or 10 mg/kg/day, and continued at 2 mg/kg/day for 2 weeks during the induction phase. Absolute blast count (ABC) in peripheral blood and minimal residual disease (MRD) in bone marrow were assessed at the end of the initial 7-day HDMP treatment. MRD in the bone marrow was evaluated on day 15 and at the end of the induction period. The follow-up for these patients ended on 15 July 2019. The 5-year event-free (EFS) and overall survival (OS) rates for all patients were 85.6±2.6% and 89.2±2.3%, respectively. The rate of good response to steroids (defined as ABC in peripheral blood of less than 1000/mm3 on day 7) was 88% and 97% of children achieved complete remission after induction. The survival rate and infection frequency did not show statistically significant differences between Group A and B. EFS and OS correlated with initial leukocyte count, age of 10-18 years at diagnosis, CD20 positivity at diagnosis, and gram-negative bacterial infection during remission induction. The remarkable response rates on days 7 and 15, along with the promising EFS and OS results in childhood ALL patients treated with the Modified St. Jude Total XV Protocol, highlight the early and substantial response effect of HDMP. At the onset of induction, short-term HDMP can be initiated, preferably at 10 mg/kg/day for the first 7 days, to minimize potential side effects.

CD74 is expressed in a subset of pediatric acute myeloid leukemia patients and is a promising target for therapy: a report from the Children's Oncology Group.

As curative therapies for pediatric acute myleoid leukemia (AML) remain elusive, identifying potential new treatment targets is vital. We assessed the cell surface expression of CD74, also known as the major histocompatibility complex-II invariant chain, by multidimensional flow cytometry in 973 patients enrolled in the Children's Oncology Group AAML1031 clinical trial (clinicaltrials gov. Identifier: NCT01371981). Thirty-eight percent of pediatric AML patients expressed CD74 at any level and a comparison to normal hematopoietic cells revealed a subset with increased expression relative to normal myeloid progenitor cells. Pediatric AML patients expressing high intensity CD74 typically had an immature immunophenotype and an increased frequency of lymphoid antigen expression. Increased CD74 expression was associated with older patients with lower white blood cells and peripheral blood blast counts, and was enriched for t(8;21), trisomy 8, and CEBPA mutations. Overall, high CD74 expression was associated with low-risk status, however 26% of patients were allocated to high-risk protocol status and 5-year event-free survival was 53%, indicating that a significant number of high expressing patients had poor outcomes. In vitro preclinical studies indicate that anti-CD74 therapy demonstrates efficacy against AML cells but has little impact on normal CD34+ cells. Together, we demonstrate that CD74 is expressed on a subset of pediatric AML at increased levels compared to normal hematopoietic cells and is a promising target for therapy in expressing patients. Given that nearly half of patients expressing CD74 at high levels experience an adverse event within 5 years, and the availability of CD74 targeting drugs, this represents a promising line of therapy worthy of additional investigation.

The Diagnostic Value of Circulating Blasts in Patients with Myeloproliferative Neoplasms in Accelerated or Blast Phase

Introduction: Myeloproliferative neoplasms (MPNs) are progressive cancers with variable propensity to transform to accelerated phase and blast phase (APBP), an acute leukemia-equivalent state. MPN in APBP is associated with very poor prognosis with shortened survival and is notorious for poor response to therapy. The only curative therapy is allogeneic stem cell transplant. Blast phase is defined by 20% blasts or higher in the bone marrow biopsy (BMBx) or peripheral blood (PB), while accelerated phase is defined by blast percentage 10% or higher. The diagnosis, and response to therapy often requires serial BMBx and PB testing. There is recognition of excessive circulating PB blasts in MPNs that occasionally exceeds blast count in BMBs, especially in fibrotic bone marrow. The aim of this study is to investigate the correlation between PB and BMBx blasts, and if PB testing might be a more convenient and lest costly marker for diagnosis and response assessment of MPN in APBP. Methods: This is a cross sectional study, based on single institution retrospective chart review. Subjects included all available subjects over the age of 18, diagnosed with APBP MPN between January 1, 2017, to July 1, 2022. We collected serial blast percentages in simultaneously collected BMBx and PB samples, assessed by the same hematopathologist. We also collected baseline disease characteristics upon transformation to APBP. This is reported descriptively, and statistically analyzed for significance using a paired t-test. Results: 15 subjects and 54 paired BP and BMBx blast assessments were identified. Median age was 59 (range 30 -71) and median duration since MPN diagnosis was 48 months (1-420). Median OS of the cohort since APBP diagnosis was 10 months. BMBx fibrosis was Grade II-III in 13/15 patients. Cytogenetic and mutation profiles, and therapies delivered are listed on Table 1. Based on evaluation of the results, there was no statistical correlation between blasts in the PB and BMBx both prior to therapy, and during therapy (detailed biostatistical analysis will be provided during ASH presentation) Conclusions: Patients with APBP MPN undergo frequent diagnostic testing, including a bone marrow biopsy, to evaluate the percentage of blasts in the bone marrow. Quantification of blast count in peripheral blood should not be as a standalone diagnostic lab test for APBP MPN. BMBx remains essential for this diagnosis, response assessment, as well as cytogenetic and molecular profiling. The outcomes on APBP remain poor despite utilization of modern targeted therapeutics.

Early Peripheral Blood Blast Clearance As a Prognostic Marker for Early Treatment Response in Pediatric Acute Myeloid Leukemia

Background and Significance The cure rate of pediatric acute myeloid leukemia (AML) has improved significantly; however, relapses still occur in up to 30% of the total patient population in high-income countries (HICs), and in an even higher percentage of patients in low- and middle-income countries (LMICs). Early treatment response, cytogenetics and minimal residual disease (MRD) are well-known prognostic factors for treatment outcome; however, sophisticated techniques for identifying prognostic factors are often lacking in LMICs, and are sometimes unsuccessful in HICs. Early treatment response assessed by a bone marrow (BM) aspirate on day 15 or day 22 (D22) (depending on the applicable protocol) after initiation of chemotherapy is a known prognostic factor for outcome in patients with pediatric AML, but does not allow modifications of the first induction chemotherapy course (Creutzig 2014, Abrahamsson 2011, Kern 2003). In both pediatric acute lymphoid leukemia (ALL) and adult AML, early peripheral blood blast clearance (PBBC) is a known early indicator of treatment response (Manabe 2008, Arellano 2012), whereas to the best of our knowledge, this has not yet been reported in pediatric AML. In this study, we evaluated the significance of time to complete PBBC, and rate of PBBC after initiation of chemotherapy, in relation to disease status in BM D22, as measure of early treatment response that has prognostic significance. Study Design and Methods Pediatric patients with AML, aged 0-18 years, that were treated according to the NOPHO-DBH AML-2012 protocol were included. The first 5 days of induction therapy consisted of etoposide, followed by 7 days of low-dose cytarabine and mitoxantrone or liposomal daunorubicin. Peripheral blood (PB) was assessed by morphology at the time of diagnosis, daily in the first week, and thrice weekly until D22 or until occurrence of complete clearance of blasts. BM D22 was assessed by morphology and flow cytometry. A receiver operating characteristic (ROC) analysis was performed to evaluate the predictive power of days from starting chemotherapy to complete PBBC. The correlation between time to complete PBBC and BM D22 results was analyzed using a logistic regression model. Multivariable logistic regression models were applied for the association after adjusting for effects of other covariates. Rate of PBBC was defined as the percentage of the absolute PB blast count on the day of diagnosis that cleared with each day of therapy, on average, until D22 or the day of complete PBBC. Results In total, 319 patients were included, out of whom 241 were eligible for analysis, and 78 not eligible due to missing or incomplete data. Day 9 after initiation of chemotherapy was identified as the most discriminating cut-off point for predicting BM D22 response. Patients were categorized into early PBBC (≤9 days; n=159) and delayed PBBC (>9 days; n=82) groups. Early PBBC was associated with having <5% AML cells in BM D22 as assessed by both morphology (OR 3.11; P=0.005) and flow cytometry (OR 2.73; P=0.041). Early PBBC also showed a much lower likelihood of having >15% AML cells in BM D22 by morphology (OR 0.08; P=0.002). None of the early PBBC patients had >15% AML cells in BM D22 by flow cytometry. For 217 patients (selection due to missing data [n=24]), the rate of PBBC could be calculated, and a discriminative cut-off value of 11% was identified. Patients were categorized into high rate of PBBC (>11%; n=132) and low rate of PBBC (≤11%; n=85). High rate of PBBC was associated with having <5% AML cells in BM D22 by flow cytometry (OR 4.30; P=0.007) and negative MRD status (OR 2.02; P=0.004), while also showing a strong association with a lower likelihood of >15% AML cells in BM D22 (OR 0.11; P=0.007) by morphology. None of the high rate PBBC patients had >15% AML cells in BM D22 by flow cytometry. Conclusion Both early complete PBBC and rate of PBBC after initiation of chemotherapy holds promise as prognostic marker for clinical outcome in pediatric AML, as it correlates significantly with BM D22 response. These findings may offer an early and easily accessible prognostic factor, particularly beneficial for LMICs, as well as for patients with unsuccessful MRD analyses.

Megakaryoblasts in pleural fluid in neonate with transient leukaemia of Down's syndrome.

Twin baby boy born at 35+2 weeks gestation with a birth weight of 2070 grams via emergency cesarean section secondary to abnormal CTG and foetal distress. Full blood count obtained at birth showed haemoglobin 159 g/L, total white cell count 99.54 × 109/L platelet count 779 × 109/L. Manual differential showed an absolute neutrophil count of 2.01 × 109/L absolute lymphocyte count of 45.27 × 109/L and blast count of 50.3 × 109/L (50%). Initial peripheral blood smear morphology review revealed a thrombocytosis, platelet anisocytosis and micromegakaryocytes. It also demonstrated neutropenia, myeloid left shift, large blasts with a high N/C ratio and nucleoli. Additionally, numerous nucleated red blood cells were seen with marked red cell dysplasia including binucleate forms and abnormal haemoglobinisation. There was no prenatal diagnosis of Down's syndrome (DS). The above blood results along with the baby's clinical signs raised suspicion of DS and subsequently transient leukaemia of DS. This baby was then transferred to the regional neonatal intensive care unit on day 2 of life. On admission, he was noted to have bilateral pleural effusion, synthetic liver dysfunction with elevated creatinine and ammonia levels as well as ascites. He was intubated, and ventilated and required inotropic support. Due to adverse features of elevated white cell count, hepatosplenomegaly, pleural effusion as well as ascites [1] and following the multidisciplinary team (MDT) discussion and parental consent, the patient was commenced on low dose cytarabine for 7 days. He tolerated that well, however, did experience mild cytopenias and pyrexic episodes, followed by recovery during the following two weeks with a reduction in pleural effusions, hepatosplenomegaly and reduction in oxygen requirement which enabled successful extubation and commencement of continuous positive airway pressure. Unfortunately, on day 19 of life he was noted to have slowly increasing oxygen requirements with re-accumulation of pleural effusions and increasing hepatosplenomegaly as well as increasing blast count in peripheral blood. He proceeded to require intubation and ventilation again as well as placement of a chest drain to remove pleural fluid to improve respiratory compliance. A review of pleural fluid morphology had shown frequent ‘blast-like’ cells; these were medium-sized with a high N/C ratio with finely condensed chromatin nucleoli and budded basophilic cytoplasm thought to most likely represent megakaryoblasts [2]. As a result of the deterioration, a decision by MDT was made to give another cycle of low-dose cytarabine on days 1 through 5. Following the second cycle of cytarabine, the patient again experienced mild cytopenias and pyrexic episodes, was treated with antibiotics and supportive management and transfusions of packed red cells. He had complete count recovery and no blast population identified morphologically on peripheral blood film on day 26 of the second cycle of cytarabine. His GATA1 mutation was subsequently found to be positive. He continued to improve and was discharged home on day 67 of life Figures 1-3. Karolina Chmielowiec—wrote the manuscript, Gary Beattie—manuscript review, obtained image, obtained and analysed flow cytometry data, Kathryn Clarke—manuscript review, obtained and analysed flow cytometry, Ellen McCloy—manuscript review, morphology review, Bethany Mitchell—manuscript review, morphology review, Christine Macartney—manuscript review, supervising consultant. All authors discussed the results and commented on the manuscript. None. The authors declare no conflict of interest. No funding was required for this submission. Not applicable. Not Applicable. Not Applicable All authors have contributed to the acquisition, analysis and interpretation of data as well as either drafting the paper or revising it critically. All authors approved the submitted and final versions. For original data, please contact [email protected]

Alterations of cohesin complex genes in acute myeloid leukemia: differential co-mutations, clinical presentation and impact on outcome

Functional perturbations of the cohesin complex with subsequent changes in chromatin structure and replication are reported in a multitude of cancers including acute myeloid leukemia (AML). Mutations of its STAG2 subunit may predict unfavorable risk as recognized by the 2022 European Leukemia Net recommendations, but the underlying evidence is limited by small sample sizes and conflicting observations regarding clinical outcomes, as well as scarce information on other cohesion complex subunits. We retrospectively analyzed data from a multi-center cohort of 1615 intensively treated AML patients and identified distinct co-mutational patters for mutations of STAG2, which were associated with normal karyotypes (NK) and concomitant mutations in IDH2, RUNX1, BCOR, ASXL1, and SRSF2. Mutated RAD21 was associated with NK, mutated EZH2, KRAS, CBL, and NPM1. Patients harboring mutated STAG2 were older and presented with decreased white blood cell, bone marrow and peripheral blood blast counts. Overall, neither mutated STAG2, RAD21, SMC1A nor SMC3 displayed any significant, independent effect on clinical outcomes defined as complete remission, event-free, relapse-free or overall survival. However, we found almost complete mutual exclusivity of genetic alterations of individual cohesin subunits. This mutual exclusivity may be the basis for therapeutic strategies via synthetic lethality in cohesin mutated AML.

CD34 negative HLA-DR negative acute myeloid leukaemia: A higher association with NPM1 and FLT3-ITD mutations.

CD34 and HLA-DR negativity is often used as a characteristic immunophenotypic feature of acute promyelocytic leukaemia (APL) that differentiates APL from other subtypes of acute myeloid leukaemia (AML). However, other subtypes of AML, without expression of CD34 and HLA-DR antigens, have also been reported. We analysed the HLA-DR negative de novo non-APL AML cases by dividing HLA-DR negative non-APL group into 2 sub-groups based on CD34 expression and compared the characteristics of CD34 negative HLA-DR negative with CD34 positive HLA-DR negative non-APL AML cases with respect to morphologic, immunophenotypic, molecular and clinical parameters. There were 70 cases (8.54%) which were CD34 negative HLA-DR negative and 52 cases (6.34%) were CD34 positive HLA-DR negative. The median age at diagnosis was higher in CD34 negative HLA-DR negative AML than in CD34 positive HLA-DR negative AML group (38 years vs. 12 years, p < 0.001). DIC rate was higher in CD34 negative HLA-DR negative group than the other group (p < 0.001). Median total leucocyte count was higher with higher blast count in peripheral blood and bone marrow in CD34 negative HLA-DR negative AML cases than the other group (p < 0.05). CD34 negative HLA-DR negative AML was more associated with normal karyotype (96.2% vs. 38.5%; p < 0.001), NPM1 mutation (67.8% vs. 8.3%; p < 0.001) and FLT-ITD mutation (37.3% vs. 13.9%; p < 0.05). In CD34 negative HLA-DR negative group, 16 cases had co-occurrence of NPM1 and FLT3-ITD mutations, whereas no case of CD34 positive HLA-DR negative group had such dual mutation positivity. There was poor median overall survival [3.8months (95%CI: 2.3-7.8months) vs. 20.4months (95% CI: 12.8-25.7months); p=0.0148] in CD34 positive HLA-DR negative AML than CD34 negative HLA-DR negative AML cases. We found that the CD34 negative HLADR negative non APL AML is highly associated with NPM1 and FLT3-ITD mutation, older age at diagnosis, DIC, higher total leucocyte count, higher blast counts and normal karyotype in comparison to CD34 positive HLA-DR negative AML group. Co-occurrence of NPM1 and FLT3-ITD mutation was also exclusively seen in CD34 negative HLA-DR negative group. There was poor overall survival in CD34 positive HLA-DR negative AML than CD34 negative HLA-DR negative AML cases.

CXCR4 (CD184) Expression in Pediatric AML Is Associated with Bone Marrow Retention, Specific Disease Characteristics, and Worse Outcomes: A Report of 1004 Patients from the Children's Oncology Group AAML1031 Protocol

Introduction: CXCR4 (CD184) is a G protein-coupled chemokine receptor specific for CXCL12 that is associated with leukocyte chemotaxis. It is expressed on normal hematopoietic cells as well as hematopoietic and epithelial cancer cells. CXCR4 and CXCL12 have been associated with bone marrow retention of AML and ALL blasts and blocking this interaction via small molecule inhibitors has been investigated as a therapeutic target. CXCR4 has also been implicated in other aspects of tumor progression including metastasis, angiogenesis, and survival. As part of Children's Oncology Group protocol AAML1031 for de novo pediatric AML we prospectively evaluated CXCR4 expression by difference from normal (ΔN) flow cytometry and correlated expression with clinical characteristics and outcome. Given the impact of the bone marrow niche on the administration of chemotherapy we hypothesized that high levels of CXCR4 expression would be associated with unfavorable outcomes. Methods: In the AAML1031 phase 3 trial pediatric patients with de novo AML were randomized to receive standard chemotherapy (Arm A) or standard chemotherapy with bortezomib (Arm B), excepting patients with high allelic ratio FLT3-ITD who received standard chemotherapy plus sorafenib (Arm C). Patients were stratified as low-risk (LR) or high-risk (HR) based on cytogenetic and molecular abnormalities, and end of induction (EOI) minimal residual disease (MRD) by flow cytometry. LR patients received four cycles of the standard chemotherapy backbone while HR patients received three followed by best hematopoietic stem cell transplant. Patients who: (1) submitted a bone marrow aspirate for ΔN at diagnosis and (2) provided consent for correlative biology studies were included in this analysis. All diagnostic specimens were centrally and prospectively evaluated for the expression of CXCR4 by ΔN. Results: Surface CXCR4 expression was available for 1004 patients and expression on AML cells varied across the cohort (range: 54-17,793 molecules/cell). For analysis, the study population was divided into quartiles (n=257-258 patients each) based on expression levels. Quartiles showed no association with sex and were distributed as expected between treatment arms. Correlation with clinical characteristics showed that patients in the top quartile of CXCR4 expression were younger (5.9 vs. 11.4 years, P<0.001) and had higher bone marrow (BM) blast counts but lower peripheral blood (PB) blast counts (76% vs. 66%, p<0.001 and 33% vs. 43%, p=0.01, respectively). These patients were highly enriched for HR KMT2A fusions (43% of total fusion positive patients), as well as t(9;11) and NPM1 mutations; while they had a lower frequency of t(8;21), inv(16), and CEBPA mutations (p<0.002 for all). There was no association between CXCR4 expression and protocol risk group assignment or complete remission rates, but patients in the high expressing CXCR4 quartile were more commonly MRD negative at EOI (p=0.03). Flow cytometry also revealed a striking increase in RAM phenotype (p<0.001), with 76% of all RAM patients allocated to the CXCR4 high expressing quartile. Patients in the top quartile of CXCR4 expression had lower 5-year OS (58% vs. 66%, p=0.036) and EFS (38% vs. 47%, p=0.009) compared to other patients. CXCR4 was strongly associated with increased 5-year relapse risk (RR) (54% vs. 39%, p<0.001). Among patients with HR KMT2A fusions high expression of CXCR4 was associated with an increased 5-year RR (85% vs. 57%, p=0.04). Multivariable analysis for 5-year RR including MRD status and prognostic genetic factors indicates CXCR4 expression is independently associated with relapse, with a hazard ratio of 1.33 (95% CI 1.04-1.7; p=0.026). Conclusions: These data demonstrate that CXCR4 expression is associated with individual HR markers (RAM phenotype and HR KMT2A fusions), but not risk status across the entire study. Despite similar risk status, patients in the top quartile of CXCR4 expression had inferior outcomes. These patients had higher BM blast counts but lower PB blast counts, which has been observed in RAM phenotype, likely due to the impact of CXCR4 on bone marrow retention of leukemia cells. This increased retention may also play a role in treatment resistance, suggesting that the addition of small molecule inhibitors of CXCR4/CXCL12 to chemotherapy may help these high expressing patients, especially in subtypes such as HR KMT2A and RAM phenotype patients. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

MYF3001: A Randomized Open Label, Phase 3 Study to Evaluate Imetelstat Versus Best Available Therapy in Patients with Intermediate-2 or High-Risk Myelofibrosis Relapsed/Refractory to Janus Kinase Inhibitor

Myelofibrosis (MF) is a life-threatening myeloproliferative neoplasm. Janus Kinase inhibitors (JAKi) ruxolitinib, pacritinib and fedratinib are the FDA-approved treatment options for MF. Despite benefits reported with ruxolitinib in the frontline setting, a high proportion of patients discontinue treatment (Abdelrahman 2015), and the median overall survival (OS) is 11-16 months (Newberry 2017; Kuykendall 2018; Palandri 2019; Schain 2019; Mascarenhas 2020), highlighting a great unmet need for patients whose disease has relapsed or is non-responsive to JAKi treatment. Imetelstat, a first-in-class telomerase inhibitor, has shown meaningful clinical improvement in IMbark, a Phase 2 study in patients with intermediate-2 (Int2) or high-risk (HR) MF whose disease has relapsed after or was refractory to JAKi (Mascarenhas JCO 2021; NCT02426086). Treatment with 9.4 mg/kg imetelstat resulted in 32.2% symptom response (total symptom score [TSS] reduction ≥50%) at Week 24 and median OS of 29.9 months with median overall study follow-up of 27.4 months (Figure 1). Dose-dependent inhibition of telomerase with imetelstat resulted in on-target activity that correlated with clinical benefits; including dose-dependent reduction in variant allele frequency (VAF) of MF driver mutations and improvement of bone marrow (BM) fibrosis. Furthermore, VAF reduction and BM fibrosis improvement correlated with OS. Imetelstat has an acceptable safety profile, with the most common Grade 3 or higher treatment-emergent adverse events being thrombocytopenia and neutropenia that were manageable and reversible, with minimal clinical consequences. These Phase 2 efficacy and safety results support continued study of imetelstat 9.4 mg/kg in a Phase 3 randomized controlled study. Study MYF3001 (IMpactMF; NCT04576156) is a Phase 3, randomized (2:1), open label, multicenter study of imetelstat compared with best available therapy (BAT) in ~320 adult patients with Int2 or HR MF whose disease has relapsed after or is refractory to JAKi treatment, who are not candidates for further JAKi treatment, and who are not eligible for allogeneic stem cell transplantation. Relapsed and refractory disease will be determined by predefined increases in spleen volume after a period of response and a lack of spleen and/or symptom responses after at least 3 months of JAKi therapy, respectively. Patients will be randomized to receive intravenous imetelstat 9.4 mg/kg every 21 days or investigator-selected BAT that may include hydroxyurea, thalidomide, interferon, danazol, hypomethylating agents, chemotherapy, or other non-JAKi-containing therapy as appropriate (Figure 2). Hematopoietic stem cell transplant and splenectomy are not permitted as BATs. Eligible patients will have an Eastern Cooperative Oncology Group Performance Status score of 0-2 and peripheral blood and marrow blast counts <10%. Patients cannot have clinically significant cardiovascular disease, active systemic hepatitis infection, or chronic liver disease unrelated to underlying MF. At randomization, patients will be stratified based on a) Int2 or HR per Dynamic International Prognostic Scoring System; and b) platelet count at entry (platelets ≥75 and <150 x 109/L vs ≥150 x 109/L). Patients who meet progressive disease criteria and discontinue BAT may be eligible to crossover to imetelstat if they exhibit a ≥25% increase in spleen volume from pretreatment baseline at any time during the study or a palpable increase in splenomegaly following 6 months of BAT, as described in the protocol. The primary endpoint of Study MYF3001 is OS and an interim analysis of safety and efficacy is planned when ~35% of patients planned to be enrolled have died. Secondary endpoints include symptom and spleen response rates at Week 24, progression-free survival, clinical response assessments per modified 2013 International Working Group - Myeloproliferative Neoplasms Research and Treatment criteria, time to and duration of response, reduction in degree of bone marrow fibrosis, safety, pharmacokinetics and patient-reported outcomes. Biomarkers and mutation analyses will be performed to evaluate the impact of imetelstat on reduction/depletion of malignant clones. Approximately 180 sites are planned in North and South America, Europe, Middle East, Australia and Asia. The study is actively enrolling. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Integration of multiparameter flow cytometry score improves prognostic stratification provided by standard models in primary myelofibrosis.

Prognostic modeling in myelofibrosis (MF) has classically pursued the integration of informative clinical and hematological parameters to separate patients' categories with different outcomes. Modern stratification includes also genetic data from karyotype and mutations. However, some poorly standardized variables, as peripheral blood (PB) blast count by morphology, are still included. In this study, we used multiparameter flow cytometry (MFC) with the aim of improving performance of existing scores. We studied 363 MF patients with available MFC files for PB CD34+ cells count determination at diagnosis. We adapted Ogata score to MF context including 2 parameters: absolute CD34+ cells count (/μL) and granulocytes to lymphocytes SSC ratio. A score of 1 was attributed to above-threshold values of each parameter. Accordingly, patients were categorized as MFClow (score=0, 62.0%), MFCint (score=1, 29.5%), and MFChigh (score=2, 8.5%). MFClow had significantly longer median OS (not reached) compared to MFCint (55 months) and MFChigh (19 months). We integrated MFC into established models as a substitute of morphological PB blasts count. Patients were reclassified according to MFC-enhanced scores, and concordance (C-) indexes were compared. As regards IPSS, C-indexes were 0.67 and 0.74 for standard and MFC-enhanced model, respectively (Z score - 3.82; p=0.0001). MFC-enhanced MIPSS70+ model in PMF patients yielded a C-index of 0.78, outperforming its standard counterpart (C-index 0.73; Z score - 2.88, p=0.004). Our data suggest that the incorporation of MFC-derived parameters, easily attainable from standard assay used for CD34+ cells determination, might help to refine the current prognostic stratification models in myelofibrosis.

Conventional and Flowcytometric Minimal Residual Disease Based Risk Stratified Post Remission Therapy Improves Survival in Ph-Negative Pediatric Acute Lymphoblastic Leukemia- Single Centre Experience from South India

Background: Children with Acute Lymphoblastic Leukemia (ALL) in Low Middle income countries(LMIC) face major challenges including treatment abandonment and poor overall survival (OS) and event-free survival (EFS).The preliminary data and results of outcome of Pediatric ALL from our pediatric hematology-oncology center which follows BFM based ALLtreatment protocol has been published in 2015 in ASH forum.Due to non availability of Polymerase chain reaction (PCR)based Minimal residual disease(MRD)analysis, we use multiparametric flowcytometry (FCM) based MRD analysis for remission assessment and risk stratification in our patients. Within resource constraints, we present evidence that outcomes comparable with that seen in high income (HIC)and upper middle income countries(UMIC) can be achieved with a post remission therapy guided by a risk stratification incorporating FCM MRD.Methods: Following IRB approval,an ambidirectional cohort study was performed using clinical information and outcomes of all patients aged 1to 14 years treated for newly diagnosed B- or T-ALL between January 1,2015 and December 31, 2020. AIEOP BFM ALL 2009 protocol with modifications was followed as the institutional protocol in all patients.The treatment algorithm is mentioned in Figure 1. Patients who underwent multiparametric FCM MRD analysis at the end of Induction IA and whose 6 months follow up details were available were included in the analysis. Patients with Ph positive ALL and those who died during Induction IA were excluded. FCM MRD analysis was performed after Induction IA on Day 33.MRD level above 0.01% was considered positive.MRD assessment was repeated following Induction IB on Day 74 in patients who had MRD positivity on Day 33. At the end of Induction IB patients were risk stratified into High risk(HR) and non High risk(Non HR).HR features included Hypodiploidy(<45chromosomes), Positivity for MLL/AF4 or t(4:11),poor prednisolone response(absolute blast count in peripheral blood ≥1 x 10^9/l on Day 8 of initiation of prednisolone) and non remission on Day33. Patients who had persistent MRD positive on Day74 were reallocated to high risk. Treatment with HR protocol was initiated for high risk patients whenever financially manageable.Statistical analysis was done using SPSS version 21 OS and EFS were assessed by Kaplan-Meier method Patients were censored at last follow-up.Results: Median follow-up time was 33.5(4-102) months . Study had 102(n=102) consecutive Ph negative patients who underwent induction therapy. Six patients (5.88%) died during induction IA;96(n=96) children who continued treatment were included in further analysis. The mean age at diagnosis was 6(1-14) years .Forty seven (48.95%)patients were male..B-ALL n=84(87.5%) and T ALL n=12.(12.5%). Four patients(4.16%) had CNS disease at diagnosis. Three children (3.13%) had high risk cytogenetics.Ten children (10.42%)in the cohort had poor prednisolone response. Five patients(5.20%) didnot achieve morphological remission on Day33. Fifteen (15.63%) patients were risk stratified as HR during IA. Four more (4.16%) were reallocated to HR in view of persistent MRD positivity after Induction IB. At median follow-up, the OS was 95.35%±2.65(95% CI 90.16-100.54) and the EFS 94.48%±2.74 (95% CI, 89.11%-99.84%).Female gender predicted better EFS (p=0.042).The EFS of patients without CNS disease at presentation was significantly better(93.5% Vs 23.2% p=0.000). The EFS at median follow up of patients in re HR cohort was 64.19% Vs 97.81% in non HR (p=0.010). EFS by risk stratification is shown in Figure II.Conclusion:Our study suggests that FCM MRD can be successfully incorporated into the treatment algorithm in a resource limited setting. With FCM MRD were able to identify an additional subset of HR patients who otherwise would have been stratified into non HR group. In a prospective cohort , FCM MRD could be tested at an earlier time point in IA induction to facilitate identification of early drug responsive patients with lowest risk of relapse. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Clinical and Genomic Characterization of Secondary Acute Myeloid Leukemia with Mixed Phenotype

Clinical and Genomic Characterization of Secondary Acute Myeloid Leukemia with Mixed Phenotype

Prediction of Complete Remission and Survival in Acute Myeloid Leukemia Using Supervised Machine Learning

Prediction of Complete Remission and Survival in Acute Myeloid Leukemia Using Supervised Machine Learning

Novel Prediction Models for Myelodysplastic Syndromes Using Machine Learning

Novel Prediction Models for Myelodysplastic Syndromes Using Machine Learning

T-MDS Is a Distinct Clinical and Pathological Entity Characterized By Better Survival Compared to t-AML

Introduction: Therapy-related myeloid neoplasm (t-MN) includes acute myeloid leukemia (AML), myelodysplastic syndrome (MDS), and myelodysplastic/myeloproliferative neoplasms (MDS/MPN) that occur as a complication of DNA-damaging therapies. The World Health Organization recommends considering t-MN as a single entity. Whether t-MDS and t-AML have distinct characteristics and outcomes is not known. The aim of this study was to compare clinicopathological characteristics and outcomes of t-MDS and t-AML.Methods: All patients diagnosed with t-MN based on the World Health Organization criteria were identified. Bone marrow biopsies, cytogenetic, and next-generation sequencing (NGS) were obtained at the treating physician's discretion. Pathogenic/likely pathogenic variants (PV) were called based on the standard criteria. Overall survival (OS) was calculated from the time of t-MN diagnosis until the last follow up using Kaplan-Meier analysis using Wilcoxon test. For survival analysis comparing chemotherapy to best supportive care (BSC) only, patients were censored at the time of allogeneic stem cell transplant (SCT). Multivariate analysis was performed using Cox proportional hazard method and corrected using false discovery rate (FDR). Statistical analysis was performed using JMP (v14.1, SAS Institute) and significance was defined as P<0.05.Results: We identified 554 patients, of which 180 (32.4%), 365 (65.8%), and 9 (1.6%) presented as t-AML, t-MDS, and t-MDS/MPN respectively. t-MDS/MPN patients were excluded from further analysis due to a small number. Clinical and laboratory characteristics of the t-AML and t-MDS cohorts is shown in Table 1. t-AML patients were significantly more anemic and thrombocytopenic at presentation. As expected, t-AML had a higher peripheral blood and BM blast count. There was no difference in proportion of patients with chromosomal abnormalities, though a statistically significantly higher proportion of t-MDS patients had chromosome 5 abnormality, 5q deletion, monosomy 5, chromosome 7 abnormality, monosomy 7 compared to t-AML patients. On other hand, 11q23 (mixed lineage leukemia, MLL) rearrangement was more common in t-AML compared to t-MDS (9.3% vs. 2.8%, P=0.005).A higher proportion of t-MDS patients had PV detected by NGS compared to t-AML (92.9% vs. 85.6%, P=0.038). A higher proportion of t-MDS patients had PV in TP53 (37.6% vs. 21.4%, P=0.004) and ASXL1 (23% vs. 11.7%, P=0.016) genes; whereas a higher proportion of t-AML patients had PV in RAS (18.9% vs. 9.1%, P=0.013) and WT1 (8.1% vs. 2.9%, P=0.05) genes.One hundred twenty-eight (35%) of 365 t-MDS patients progressed to t-AML. The presence of any chromosomal abnormality at t-MDS diagnosis predicted a higher risk of transformation to t-AML (χ 2 3.9, P=0.03).t-AML patients had a significantly shorter OS compared to t-MDS (9.2 vs. 19.7 months, P<0.0001, Figure A). This difference persisted when stratified by no disease modifying therapies (BSC only) 2 vs. 17 months (P<0.0001, Figure B), as well as among those who received at least one line of chemotherapy (14 vs. 24.6 months, P<0.001). Finally, a higher proportion of patients with t-AML underwent SCT and there was a trend towards improved survival for t-AML patients (vs. t-MDS 52.9 vs. 20.7 months, P=0.07) from the time of transplant.Multivariate analysis for OS performed to control for all the variables that were different at presentation (except for blast count), showed that t-MDS (as opposed to t-AML) phenotype at diagnosis, and undergoing SCT were independent predictors of improved survival (Table 2).Conclusion: t-MDS and t-AML have distinct clinical, cytogenetic, and genetic features at presentation. In the absence of disease modifying therapies, t-AML is a more aggressive phenotype, consistently associated with a shorter survival. Even after controlling adverse risk features, t-AML phenotype had a shorter survival compared to t-MDS. [Display omitted] DisclosuresLitzow: AbbVie: Research Funding; Astellas: Research Funding; Amgen: Research Funding; Actinium: Research Funding; Pluristem: Research Funding; Jazz: Other: Advisory Board; Omeros: Other: Advisory Board; Biosight: Other: Data monitoring committee. Hiwase: Novartis: Membership on an entity's Board of Directors or advisory committees; AbbVie: Membership on an entity's Board of Directors or advisory committees.

Mebendazole Mediates Proteasomal Degradation of GLI Transcription Factors in Acute Myeloid Leukemia.

The prognosis of elderly AML patients is still poor due to chemotherapy resistance. The Hedgehog (HH) pathway is important for leukemic transformation because of aberrant activation of GLI transcription factors. MBZ is a well-tolerated anthelmintic that exhibits strong antitumor effects. Herein, we show that MBZ induced strong, dose-dependent anti-leukemic effects on AML cells, including the sensitization of AML cells to chemotherapy with cytarabine. MBZ strongly reduced intracellular protein levels of GLI1/GLI2 transcription factors. Consequently, MBZ reduced the GLI promoter activity as observed in luciferase-based reporter assays in AML cell lines. Further analysis revealed that MBZ mediates its anti-leukemic effects by promoting the proteasomal degradation of GLI transcription factors via inhibition of HSP70/90 chaperone activity. Extensive molecular dynamics simulations were performed on the MBZ-HSP90 complex, showing a stable binding interaction at the ATP binding site. Importantly, two patients with refractory AML were treated with MBZ in an off-label setting and MBZ effectively reduced the GLI signaling activity in a modified plasma inhibitory assay, resulting in a decrease in peripheral blood blast counts in one patient. Our data prove that MBZ is an effective GLI inhibitor that should be evaluated in combination to conventional chemotherapy in the clinical setting.

MPN-161: A Phase I Study of TP-3654, an Oral PIM Kinase Inhibitor, in Patients with Intermediate-2 or High-Risk Primary or Secondary Myelofibrosis (MF)

Context: MF is characterized by bone marrow fibrosis (BMF), ineffective hematopoiesis, splenomegaly, and debilitating symptoms. Expression of PIM-1 is significantly upregulated in MF hematopoietic cells, indicating that PIM-1 is a potential therapeutic target in MF. TP-3654, an oral small-molecule investigational PIM-1 kinase inhibitor with favorable selectivity against other kinases, has been shown to inhibit proliferation and increase apoptosis in murine and human hematopoietic cells expressing the JAK2V617F mutation. TP-3654 alone and in combination with ruxolitinib normalizes leukocyte counts and reduces spleen size and BMF in JAK2V617F and MPLW515L murine MF models. Objective: To evaluate TP-3654 monotherapy in patients with MF. Design: Phase I, open-label, dose-escalation study (NCT04176198). Setting: Multicenter, US only. Patients or Other Participants: Key eligibility criteria include primary or secondary MF (post-polycythemia vera-MF/post-essential thrombocythemia-MF); intermediate-2/high-risk MF per Dynamic International Prognostic Scoring System (DIPSS); previously failed or are ineligible to receive Janus kinase inhibitors (e.g., ruxolitinib, fedratinib); grade ≥2 BMF; platelet count ≥50x109/L; absolute neutrophil count ≥1x109/L; peripheral blood blast counts Interventions: Patients receive oral TP-3654. Dose escalation, performed using a Bayesian Logistic Regression Model with Escalation with Overdose Control, will continue until identification of the maximum tolerated dose (MTD) and/or recommended phase II dose (RP2D) is achieved. Dose expansion will follow. Main Outcomes Measures: Primary objectives: to determine MTD and/or identify the RP2D of TP-3654 monotherapy. Secondary objectives: to assess preliminary activity (spleen volume reduction, symptom improvement, and BMF reduction), determine QT interval changes, and establish the pharmacokinetic profile. Exploratory objectives: to study pharmacodynamic markers in peripheral blood and BM biopsies. This study is currently recruiting.

The Outcome of Patients with High Risk MDS Achieving Stable Disease after Treatment with 5- Azacitidine. A Retrospective Analysis of the Hellenic (Greek) Mds Study Group

INTRODUCTIONThe demethylating factor 5-Azacytidine (AZA) mainly improves survival in high-risk MDS patients (intermediate-2 and high risk, according to IPSS) who responded to the treatment, but the outcome of patients who achieved stable disease (SD) is unclear. This retrospective study of the Hellenic MDS Study Group was conducted to investigate the significance of achieving stable disease as best response at 6 months in terms of overall survival and time to trasformation to acute myeloid leukemia (AML) METHODSResponse categories were “overall response” (defined as complete remission, partial remission, or any hematologic improvement), “non-response” (disease progression or treatment failure), and “stable disease” (none of the above). This retrospective study of the Hellenic MDS Study Group included 405 intermediate-2 or high risk MDS patients treated with AZA. Extensive biostatistical analysis performed in this study included Kaplan-Meier survival analysis. The level of statistical significance was set at a probability value of less than 0.050 (P <0.050). RESULTSThe median age of the patients was 74 years (range: 33 - 89). Baseline patient characteristics are presented in Table 1. SD patients had lower lymphocyte count and higher platelets count than responders and non-responders. Additionally, responders had higher hemoglobin levels and blast count in peripheral blood than non-responders. Patients achieving an overall response had longer time to transformation to AML and better OS, compared to SD patients (estimated median time to transformation to AML 30 months, 95% CI = 25-35 vs. 18 months, 95% CI = 13.6-22.4, P <0.001; estimated median OS: 26 months, 95% CI = 23-29 vs. 18 months, 95% CI = 15.2-20.8, P <0.001). Furthermore, 46 out of 95 patients (48.4%) who achieved SD and who continued treatment with AZA for a median time of 14 months (range: 7 - 58) showed a lower risk for transformation to AML and increased OS, compared to SD patients who discontinued the treatment (estimated median time to transformation to AML 23 months, 95% CI = 11.8-34.2 vs. 13 months, 95% CI = 8.2-17.8, P=0.002; estimated median OS 20 months, 95% CI = 5.5-34.5 vs. 11 months, 95% CI = 5.7-16.3, P <0.001). Moreover, SD patients who continued treatment with AZA had no differences in time to transformation to AML (Figure 1a) and in OS (Figure 1b), compared to patients showing response to AZA (estimated median time to transformation to AML 23 months, 95% CI = 11.8-34.2 vs. 30 months, 95% CI = 25-35, P=0.52; estimated median OS 20 months, 95% CI = 5.5 - 34.5 vs. 26 months, 95% CI = 23-29, P=0.50). CONCLUSIONSMDS patients achieving SD in the first six months of treatment with AZA as best response should continue receiving AZA as they may benefit from prolonged treatment. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Phase I/Ib Trial of the MUC1 Inhibitor GO-203-2C Alone and in Combination with Decitabine for Acute Myeloid Leukemia

MUC1, an oncogenic protein aberrantly expressed on AML and leukemic stem cells, supports critical aspects of the malignant phenotype including resistance to apoptosis, cell proliferation and autonomous self renewal. GO-203-2C is a novel cell-penetrating peptide that blocks homodimerization of the MUC1-C subunit required for nuclear translocation and downstream signaling. In murine AML models, treatment with GO-203-2C resulted in specific eradication of myeloid leukemia. GO-203-2C is synergistic with hypomethylating agents in pre-clinical models of AML. Based on these findings, we are conducting a phase I/Ib study of GO-203-2C alone and in combination with decitabine in patients with AML.Patients with relapsed or refractory (R/R) AML or those with untreated AML who were not candidates for cytotoxic therapy were eligible. Study design consisted of a dose escalation phase with 3 patients each treated with 6.25mg/m2, 12.5mg/m2 and 18mg/m2 intravenous GO-203-2C alone on days 1-5, 8-12 and 15-19 of a 28 day treatment cycle. A subsequent dose escalation with a total of 7 patients was conducted with G0-203-2C at 12.5mg/m2 and 18mg/m2 GO-203-2C administered on day 1-5 and 8-12 in conjunction with decitabine at the standard dose of 20mg/m2 on days 8-12 of each cycle. The study is currently enrolling an expansion cohort; outcomes for 20 patients will be available once the accrual goal is met.Median age was 66 (45 - 81) years with equal gender representation. ECOG performance status was 1 or 2 in all patients. 3 patients were treated upfront and 13 had R/R disease after one to five prior therapies including 4 post-allogeneic transplant. AML prognostic risk was determined according to ELN strata with 12% favorable, 44% intermediate and 44% high risk. Adverse events were primarily CTCAE grade 1 or 2. There was an episode of grade 3 hypertriglyceridemia in one patient and grade 2 pancreatitis in another for which study drug was held and grade 3 hypertension for which dose was reduced. 5 episodes of treatment delay occurred amongst 3 patients due to grade 3-4 neutropenia or thrombocytopenia. Median time to absolute neutrophil >1.0 x 10^9/uL or platelet >100 x 10^9/uL was 8 days with cessation of therapy required in one case that failed to recover by day 50. Another patient experienced atrial fibrillation leading to 4 missed doses of GO-203-2C in cycle 1 and was not evaluable for dose-limiting toxicity but continued on study receiving 9 of 10 doses in cycle 2 and was included in response assessment.Response was defined by IWG criteria as complete remission (CR), complete remission with incomplete count recovery (CRi), or partial remission (PR). Patients in the cohort with GO-203-2c alone received up to 2 cycles of therapy. All 9 patients treated with GO-203-2c alone had resistant disease on marrow assessment but 1 patient had sustained clearance of peripheral blasts. In the combination cohort, response was achieved in 57% (4 of 7). One patient achieved CR and remains on therapy cycle 11. Two patients achieved CRi, although one was taken off study for hematologic toxicity, the other relapsed prior to cycle 3. One patient had partial remission after cycle 2 but later progressed. 1 patient had resistant disease after 3 cycles and two had progressive disease (PD) defined as doubling of peripheral blood blast count or marrow blast percentage, after 1 or 2 cycles. Amongst patients who had response, 2 were upfront and 2 R/R; 2 patients had favorable risk, 1 intermediate and 1 high risk disease.We have previously reported that suppression of the immune response by MUC1 in AML is associated with expansion of myeloid-derived suppressor cells (Pyzer AR. Blood 2017) and up-regulation of PD-L1 expression (Pyzer AR, Leukemia 2017). Of note, response to GO-203-2C treatment was associated with decreased MUC1 expression on CD34+CD38-lineage- leukemia stem cells from 3 evaluable patients. MUC1 inhibition also resulted in decreased PD-L1 expression in leukemia cells and reduction in myeloid derived suppressor cells as demonstrated by multichannel flow cytometric analysis of CD11B+HLA-DR-CD33+ cells.In summary, the results demonstrate that the combination of GO-203-2C and decitabine is safe, and importantly, demonstrates both anti-leukemia and immunomodulatory effects. DisclosuresStone:Ono: Consultancy; Roche: Consultancy; Juno Therapeutics: Consultancy; Jazz,: Consultancy; Novartis: Consultancy; DSMN: Consultancy; Pfizer: Consultancy; Janssen: Consultancy; Cornerstone: Consultancy; Astellas: Consultancy; Amgen: Consultancy; Agios: Consultancy; Celgene: Consultancy, Membership on an entity's Board of Directors or advisory committees; AbbVie: Consultancy, Membership on an entity's Board of Directors or advisory committees; Sumitomo Dainippon: Consultancy. DeAngelo:Immunogen: Honoraria, Research Funding; Novartis Pharmaceuticals Corporation: Consultancy, Honoraria, Research Funding; Pfizer Inc.: Consultancy, Honoraria, Research Funding; Amgen: Consultancy, Research Funding; BMS: Consultancy; ARIAD: Consultancy, Research Funding; Incyte: Consultancy, Honoraria; Blueprint Medicines: Honoraria, Research Funding; Shire: Honoraria; Glycomimetics: Research Funding; Takeda Pharmaceuticals U.S.A., Inc.: Honoraria; Celgene: Research Funding. Neuberg:Synta Pharmaceuticals: Other: Stock shares. Avigan:Genus Oncology: Research Funding.

Validation of the Who-Defined 20% Circulating Blasts Threshold for Diagnosis of Acute Leukemia in Myelofibrosis: A Single Center Study of 144 Patients with Blast or Chronic Phase Disease with Excess Blasts

Background:The World Health Organization (WHO)-defined criteria for acute myeloid leukemia (AML), including blast phase myeloproliferative neoplasms (MPN-BP), includes the presence of ≥20% blasts in the peripheral blood (PB), regardless of blast content in the bone marrow (BM) (Blood 2016;127:2391) . It is currently unclear if this 20% threshold is valid for primary myelofibrosis (PMF), in terms of distinguishing PB-defined MPN-BP from either chronic phase disease with excess blasts (PMF-EB; defined by the presence of 5-19% circulating blasts) or BM-defined MPN-BP.Methods:Diagnoses of MPN, including PMF and MPN-BP, were according to WHO criteria (Blood 2016;127:2391). The study population included chronic phase PMF patients with 5-19% circulating blasts (PMF-EB) and MPN patients with blast phase disease (MPN-BP), defined by the presence of ≥20% blasts in PB only (PB-defined MPN-BP) or BM (BM-defined MPN-BP). Statistical analyses for PMF-EB considered clinical and laboratory data collected at the time of documented PB blast count of ≥5% and for MPN-BP, date of leukemic transformation. Phenotypic and prognostic comparisons considered four distinct categories: i) chronic phase disease with 5-9% circulating blasts (PMF-EB-1), ii) chronic phase disease with 10-19% circulating blasts (PMF-EB-2), iii) MPN-BP defined by ≥20% BM blasts (BM-defined) and iv) MPN-BP defined by ≥20% blasts in PB only (PB-defined).Results:The total number of study patients was144 and included 41 patients with PMF-EB and 103 with MPN-BP; the 41 patients with PMF-EB included 28 with PMF-EB-1 (median age 69 years; 61% males) and 13 with PMF-EB-2 (median age 64 years; 54% males); the 103 patients with MPN-BP included 71 BM-defined (median age 68 years; 69% males) and 32 PB-defined (median age 66 years; 72% males) MPN-BP (p=0.6 for age and 0.6 for gender distribution). The four operational groups were also similar in their need for red cell transfusions (p=0.4), hemoglobin level (p=0.9), leukocyte count (p=0.9) and driver mutational status (p=0.3), including incidence of type 1/like CALR mutations (11% for PMF-EB-1 vs 15% for PMF-EB-2 vs 17% for PB-defined MPN-BP vs 19% for BM-defined MPN-BP). Significant differences were noted for platelet count (lower for both PB and BM defined MPN-BP; p=0.004) and incidence of abnormal karyotype (p=0.02) and unfavorable karyotype (22% for PMF-EB-1, 62% PMF-EB-2, 61% PB-defined MPN-BP and 49% BM-defined MPN-BP; p=0.03). Survival data showed significant difference in favor of PMF-EB, compared to both BM- and PB-defined MPN-BP, while they were similar for PMF-EB-1 vs PMF-EB-2 and for PB- vs BM-defined MPN-BP (Figure 1).Conclusions:The observations from the current study support the prognostic validity of current WHO criteria in distinguishing chronic phase PMF with excess blasts (PMF-EB) from PB-defined MPN-BP. The current study also confirms the validity of using circulating blast percentage to define MPN-BP, regardless of BM blast percentage. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.