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Myeloid/lymphoid neoplasm with eosinophilia (MLN-Eo) is a World Health Organization (WHO) established category of hematologic malignancies primarily arising in adults. We discuss an 8-month-old infant who presented with clinical features similar to those of juvenile myelomonocytic leukemia (JMML) but who was diagnosed with MLN-Eo driven by an ETV6-FLT3 fusion. Results of patient-derived leukemia ex vivo studies demonstrated increased sensitivity to type I FLT3 inhibitors as compared with type II inhibitors. Treatment with the type I inhibitor gilteritinib resulted in complete immunophenotypic and cytogenetic remission. This patient subsequently underwent a hematopoietic stem cell transplant and remains in complete remission 1 year later. This is the youngest patient reported with an ETV6-FLT3 fusion and adds to the mounting reports of FLT3-rearranged MLN-Eo, supporting its addition to the WHO classification. Furthermore, this case highlights the clinical utility of ex vivo drug testing of targeted therapies.

Plasmacytoid dendritic cells (pDCs) are the principal natural type I interferon–producing dendritic cells. Neoplastic expansion of pDCs and pDC precursors leads to blastic plasmacytoid dendritic cell neoplasm (BPDCN), and clonal expansion of mature pDCs has been described in chronic myelomonocytic leukemia. The role of pDC expansion in acute myeloid leukemia (AML) is poorly studied. Here, we characterize patients with AML with pDC expansion (pDC-AML), which we observe in ∼5% of AML cases. pDC-AMLs often possess cross-lineage antigen expression and have adverse risk stratification with poor outcome. RUNX1 mutations are the most common somatic alterations in pDC-AML (>70%) and are much more common than in AML without pDC expansion and BPDCN. We demonstrate that pDCs are clonally related to, as well as originate from, leukemic blasts in pDC-AML. We further demonstrate that leukemic blasts from RUNX1-mutated AML upregulate a pDC transcriptional program, poising the cells toward pDC differentiation and expansion. Finally, tagraxofusp, a targeted therapy directed to CD123, reduces leukemic burden and eliminates pDCs in a patient-derived xenograft model. In conclusion, pDC-AML is characterized by a high frequency of RUNX1 mutations and increased expression of a pDC transcriptional program. CD123 targeting represents a potential treatment approach for pDC-AML.

Azacitidine + venetoclax, decitabine + venetoclax, and low-dose cytarabine + venetoclax are now standard treatments for newly diagnosed older or unfit patients with acute myeloid leukemia (AML). Although these combinations are also commonly used in relapsed or refractory AML (RR-AML), clinical and molecular predictors of response and survival in RR-AML are incompletely understood. We retrospectively analyzed clinical and molecular characteristics and outcomes for 86 patients with RR-AML who were treated with venetoclax combinations. The complete remission (CR) or CR with incomplete hematologic recovery (CRi) rate was 24%, and the overall response rate was 31% with the inclusion of a morphologic leukemia-free state. Azacitidine + venetoclax resulted in higher response rates compared with low-dose cytarabine + venetoclax (49% vs 15%; P = .008). Median overall survival (OS) was 6.1 months, but it was significantly longer with azacitidine + venetoclax compared with low-dose cytarabine + venetoclax (25 vs 3.9 months; P = .003). This survival advantage of azacitidine + venetoclax over low-dose cytarabine + venetoclax persisted when patients were censored for subsequent allogeneic stem cell transplantation (8.1 vs 3.9 months; P = .035). Mutations in NPM1 were associated with higher response rates, whereas adverse cytogenetics and mutations in TP53, KRAS/NRAS, and SF3B1 were associated with worse OS. Relapse was driven by diverse mechanisms, including acquisition of novel mutations and an increase in cytogenetic complexity. Venetoclax combination therapy is effective in many patients with RR-AML, and pretreatment molecular characteristics may predict outcomes. Trials that evaluate novel agents in combination with venetoclax therapy in patients with RR-AML that have adverse risk genomic features are warranted.

Key Points Myeloid neoplasm (MPN) clones can evolve from acute myeloid leukemia to gain dominance with isocitrate dehydrogenase inhibition. Pro-differentiation agents such as ivosidenib can unmask MPN sequelae.

We investigated the role of PRMT5 in myeloproliferative neoplasm (MPN) pathogenesis and aimed to elucidate key PRMT5 targets contributing to MPN maintenance. PRMT5 is overexpressed in primary MPN cells, and PRMT5 inhibition potently reduced MPN cell proliferation ex vivo. PRMT5 inhibition was efficacious at reversing elevated hematocrit, leukocytosis, and splenomegaly in a model of JAK2V617F+ polycythemia vera and leukocyte and platelet counts, hepatosplenomegaly, and fibrosis in the MPLW515L model of myelofibrosis. Dual targeting of JAK and PRMT5 was superior to JAK or PRMT5 inhibitor monotherapy, further decreasing elevated counts and extramedullary hematopoiesis in vivo. PRMT5 inhibition reduced expression of E2F targets and altered the methylation status of E2F1 leading to attenuated DNA damage repair, cell-cycle arrest, and increased apoptosis. Our data link PRMT5 to E2F1 regulatory function and MPN cell survival and provide a strong mechanistic rationale for clinical trials of PRMT5 inhibitors in MPN. Significance: Expression of PRMT5 and E2F targets is increased in JAK2V617F+ MPN. Pharmacologic inhibition of PRMT5 alters the methylation status of E2F1 and shows efficacy in JAK2V617F/MPLW515L MPN models and primary samples. PRMT5 represents a potential novel therapeutic target for MPN, which is now being clinically evaluated. This article is highlighted in the In This Issue feature, p. 1611

Chromosome translocations involving the RUNX1 gene at 21q22 are recurring abnormalities in acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS), that is, t(8;21) and t(3;21) and in B-cell acute lymphoblastic leukemia with t(12;21). These translocations result in the fusion of RUNX1 with RUNX1T1, MECOM, and ETV6, respectively, and are implicated in leukemogenesis. Here we describe 10 rare RUNX1 fusion gene partners, including six novel fusions, in myeloid neoplasia. Comprehensive molecular testing revealed the partner genes and features of these fusions in all the tested patients, and detected various recurring myeloid related gene mutations in eight patients. In two patients, RUNX1 mutations were identified. Most of these fusions were detected in patients with high-grade MDS and AML with a relatively short survival. Integration of conventional chromosome analysis, FISH testing and molecular genetic studies allow a comprehensive characterization of these rare RUNX1 fusions. Our study may help define myeloid neoplasms with rare and novel RUNX1 translocations and support appropriate patient management.

Although rare, spontaneous remission (SR) of acute myeloid leukemia (AML) has been reported in the literature, the underlying mechanisms driving remission remain unknown. However, it is most commonly associated with a preceding severe infection. We present a case of a 40-year-old man with no past medical history who presented to our hospital with severe left hip pain and fevers and was found to have AML. Chemotherapy was delayed because the patient required extensive debridement and fasciotomy of his left hip and a prolonged course of antibiotics. After his acute illness had stabilized, a repeat bone marrow biopsy was performed which showed no abnormal myeloid blasts and resolution of his original cytogenetic and molecular abnormalities. At the time of this writing, our patient remains in remission with undetectable minimal residual disease (MRD), now 14 months from his initial diagnosis of AML.

The role of allogeneic hematopoietic cell transplant (allo-HCT) as consolidation after initial venetoclax therapy and the efficacy of venetoclax salvage therapy for relapse after allo-HCT in patients with acute myeloid leukemia (AML) are unclear. We conducted a retrospective study of patients with AML or myelodysplastic syndrome (MDS) who received venetoclax either before or after allo-HCT at Memorial Sloan Kettering Cancer Center and Yale University from 11 August 2016 to 16 November 2020. Among 39 heavily pretreated patients who received venetoclax before allo-HCT, median OS from allo-HCT was not reached after a median follow up of 12.5 months resulting in a 12-month OS estimate of 79.0%. In 37 patients who had received venetoclax-based combinations as salvage therapy after allo-HCT, the overall response rate was 32% with a median OS of 4.7 months (12-month OS estimate: 43.4%). Four patients underwent a second allo-HCT following venetoclax-based salvage therapy suggesting it as a potential salvage treatment option.

Approximately 15% of chronic lymphocytic leukemia/small lymphocytic lymphomas (CLL/SLL) transform into aggressive lymphomas, often diffuse large B cell lymphomas (DLBCL), known as Richter Syndrome ...

Increasing evidence supports the prognostic significance of measurable residual disease (MRD) in acute myeloid leukemia (AML). Dynamic MRD assessment for patients with AML complements baseline patient risk assessment factors in determining patient prognosis. MRD status may also be helpful in informing therapeutic decisions. The European Leukemia Net MRD working party recently issued consensus recommendations for the use of MRD in AML. The Food and Drug Administration also issued advice for using MRD in trials of hematologic malignancies. This article discusses MRD testing, highlights the challenges in adopting MRD testing in clinical practice, and provides insights into the future of the field.

Plasmacytoid dendritic cells (pDC) are the principal natural type I interferon producing dendritic cells. Neoplastic expansion of pDCs and pDC precursors leads to blastic plasmacytoid dendritic cell neoplasm (BPDCN) and clonal expansion of mature pDCs has been described in chronic myelomonocytic leukemia (CMML). The role of pDC expansion in acute myeloid leukemia (AML) is poorly studied. Here we characterize AML patients with pDC expansion (pDC-AML), which we observe in approximately 5% of AML. pDC-AML often possess crosslineage antigen expression and have adverse risk stratification with poor outcome. RUNX1 mutations are the most common somatic alterations in pDC-AML (>70%) and are much more common than in AML without PDC expansion. We demonstrate that pDCs are clonally related to, and originate from, leukemic blasts in pDC-AML. We further demonstrate that leukemic blasts from RUNX1-mutated AML upregulate a pDC transcriptional program, poising the cells towards pDC differentiation and expansion. Finally, tagraxofusp, a targeted therapy directed to CD123, reduces leukemic burden and eliminates pDCs in a patient-derived xenograft model. In conclusion, pDC-AML is characterized by a high frequency of RUNX1 mutations and increased expression of a pDC transcriptional program. CD123 targeting represents a potential treatment approach for pDC-AML.

In patients with multiple myeloma, plasmablastic transformation in the bone marrow is rare and associated with poor outcomes. The significance of discordant extramedullary plasmablastic transformation in patients with small, mature clonal plasma cells in the bone marrow has not been well studied. Here, we report the clinicopathologic, cytogenetic, and molecular features of 10 such patients (male/female: 6/4, median age: 65 y, range: 48 to 76 y) with an established diagnosis of multiple myeloma in the bone marrow composed of small, mature plasma cells in parallel with a concurrent or subsequent extramedullary plasmablastic transformation. Eight patients with available survival data showed an overall aggressive clinical course with a median survival of 4.5 months after the diagnosis of extramedullary plasmablastic transformation, despite aggressive treatment and even in patients with low-level bone marrow involvement. Pathologically, the extramedullary plasmablastic myeloma were clonally related to the corresponding bone marrow plasma cells, showed high levels of CMYC and/or P53 expression with a high Ki-67 proliferation index by immunohistochemistry and harbored more complex genomic aberrations including frequent mutations in the RAS pathway and MYC rearrangements compared with their bone marrow counterparts. In summary, although genetic and immunohistochemical studies were not uniformly performed on all cases due to the retrospective nature of this study, our data suggest that discordant extramedullary plasmablastic transformation of multiple myeloma has an aggressive clinical course and is characterized by frequent mutations in the RAS pathway and more complex genomic abnormalities.

TET2 and DNMT3A mutations are frequently identified in T-cell lymphomas of T follicular helper cell origin (TCL-TFH), clonal hematopoiesis (CH), and myeloid neoplasms (MNs). The relationships among these 3 entities, however, are not well understood. We performed comprehensive genomic studies on paired bone marrow and tissue samples as well as on flow cytometry–sorted bone marrow and peripheral blood subpopulations from a cohort of 22 patients with TCL-TFH to identify shared CH-type mutations in various hematopoietic cell compartments. Identical mutations were detected in the neoplastic T-cell and myeloid compartments of 15 out of 22 patients (68%), including TET2 (14/15) and DNMT3A (10/15). Four patients developed MNs, all of which shared CH-type mutations with their TCL-TFH; additional unique genetic alterations were also detected in each patient’s TCL-TFH and MN. These data demonstrate that CH is prevalent in patients with TCL-TFH and that divergent evolution of a CH clone may give rise to both TCL-TFH and MNs.

While the majority of NK/T-cell neoplasms are unfavorable in clinical outcome, a few pathologic variants demonstrate relatively indolent behavior. These include primarily four types of tissue-based T-cell or NK-cell lymphoproliferative disorders and two leukemic forms of T-cell or NK-cell lymphoproliferative disorder. The diagnostic criteria and other clinicopathologic features of these entities are discussed as question and answer format.

No abstract available Keywords: AML; CTLA-4 antagonist; P2RY8-CRLF2 fusion; immunotherapy.

Key Points Core-binding factor AML can evolve from good-risk disease into aggressive disease through the gain of additional genomic aberrations. In this unique case, an AML patient died of hypereosinophilic syndrome with solid organ infiltration of differentiated eosinophils.

INTRODUCTION Mixed phenotype (MP) is characteristic for de novo mixed phenotype acute leukemia (dnMPAL) but can also be seen in blast phase of myeloproliferative neoplasms (MPN-BP), myeloid/lymphoid neoplasms with eosinophilia & rearrangements, acute myeloid leukemia (AML) with recurrent cytogenetic abnormalities (AML-RCA) and secondary AML (sAML) including AML with myelodysplasia-related changes (AML-MRC) and therapy-related AML (t-AML). Although WHO classification excludes sAML with MP (sAML-MP) from dnMPAL, the significance of MP in the setting of sAML and their genetic landscape has not been studied. METHODS The MSKCC pathology data base was searched from 01/2014 to 09/2020 and a cohort of 125 patients with MP as defined per WHO 2016 classification was obtained. The clinical, morphologic, immunophenotypic, and cytogenetic/molecular results were reviewed. Patients with a diagnosis of AML-RCA, myeloid/lymphoid neoplasms with eosinophilia & rearrangements, MPN-BP, B-ALL with isolated MPO and myelodysplastic syndrome (MDS) were excluded (Fig 1). RESULTS 50 cases of sAML-MP (14 t-AML and 36 AML-MRC) were retrieved and compared to 42 dnMPAL cases and 100 sAML without MP (37 t-AML and 63 AML-MRC) (Table 1). The median age at diagnosis was 66 years which was higher than dnMPALs (44.5 yrs, p value Molecular studies (Table 2) revealed that the most commonly mutated genes in sAML-MP were RUNX1, DNMT3A, TP53 which were all present in a significantly higher frequency in comparison to dnMPAL. The frequency of RUNX1 mutation was higher in sAML-MP even in comparison to sAML without MP. Conversely, mutations in PHF6 frequently noted in dnMPALs were uncommon in sAML cohorts. Cluster analysis based on immunophenotyping of the 3 cohorts revealed 6 clusters (Fig 2) with separation between the 3 cohorts. Most of the dnMPAL with B/M phenotype formed a distinct tight cluster (cluster 3). Most of the sAML-MP were seen clustering together in cluster 4 whereas sAML without MP predominated in cluster 5 and 6. Cluster 1 revealed a mixture of dnMPAL and sAML-MP with predominantly T/M phenotype. The overall survival (OS) was inferior in the sAML-MP cohort in comparison to dnMPAL cohort (median survival: 6.77 vs 36.99 mths, p value 0.00005; Fig 3A) and was similar to the sAML without MP cohort (median survival: 6.77 vs 4.96 mths, p value 0.33; Fig 3A). In a multivariate analysis, model adjusted for the age and allotransplant did not explain difference in OS between these groups. Comparison of OS between the 6 clusters (Fig 3B) revealed better OS in clusters 2 and 3 that were enriched for dnMPAL. Cluster1 in spite of having a mixture of dnMPAL and sAML-MP did poorly. This could be driven by enrichment in DNMT3A mutations in the dnMPAL with T/M phenotype. Immunophenotypically distinct blast populations were flow cytometrically sorted in 4 cases of sAML-MP (2 each with T/M and B/M phenotype). While the majority of the genetic abnormalities were shared between populations with different lineages, a case with multiple mutations showed divergent KRAS/NRAS mutations showing clonal divergence as possible late event driving distinct lineage maturation (Table 3). CONCLUSION sAML-MP and sAML without MP are clinically similar and have biological overlaps with frequent somatic mutations in TP53, chromatin modifying genes and spliceosome-complex genes, which is different from dnMPAL. However, the frequency of RUNX1 mutation is higher in sAML-MP than sAML without MP suggesting its role in lineage infidelity. Though, cluster analysis based on immunophenotype separates the 3 cohorts into enriched clusters, there are still overlaps between sAML-MP and dnMPAL especially with a T/M phenotype as well as between sAML-MP and sAML without MP especially in TP53 mutated cases. Additional RNA seq and ATAC seq are currently being performed on flow cytometrically sorted distinct blast populations of the sAML-MP cases. Figure 1 Figure 1. Disclosures Galera: Paige.AI: Research Funding. Dogan: Physicians' Education Resource: Honoraria; Seattle Genetics: Consultancy; Takeda: Consultancy, Research Funding; EUSA Pharma: Consultancy; Roche: Consultancy, Research Funding; Peer View: Honoraria. Tallman: Syros: Membership on an entity's Board of Directors or advisory committees; NYU Grand Rounds: Honoraria; Kura: Membership on an entity's Board of Directors or advisory committees; Innate Pharma: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees; Biosight: Membership on an entity's Board of Directors or advisory committees; Roche: Membership on an entity's Board of Directors or advisory committees; Jazz Pharma: Membership on an entity's Board of Directors or advisory committees; Oncolyze: Membership on an entity's Board of Directors or advisory committees; KAHR: Membership on an entity's Board of Directors or advisory committees; Orsenix: Membership on an entity's Board of Directors or advisory committees; Daiichi-Sankyo: Membership on an entity's Board of Directors or advisory committees; Abbvie: Membership on an entity's Board of Directors or advisory committees; Amgen: Research Funding; Rafael Pharmaceuticals: Research Funding; Glycomimetics: Research Funding; Biosight: Research Funding; Orsenix: Research Funding; Abbvie: Research Funding; Mayo Clinic: Honoraria; UC DAVIS: Honoraria; Northwell Grand Rounds: Honoraria; NYU Grand Rounds: Honoraria; Danbury Hospital Tumor Board: Honoraria; Acute Leukemia Forum: Honoraria; Miami Leukemia Symposium: Honoraria; New Orleans Cancer Symposium: Honoraria; ASH: Honoraria; NCCN: Honoraria. Levine: Celgene: Research Funding; Incyte: Consultancy; Roche: Honoraria, Research Funding; Morphosys: Consultancy; Imago: Membership on an entity's Board of Directors or advisory committees; Lilly: Honoraria; QIAGEN: Membership on an entity's Board of Directors or advisory committees; Astellas: Consultancy; Amgen: Honoraria; Gilead: Honoraria; Zentalis: Membership on an entity's Board of Directors or advisory committees; Ajax: Membership on an entity's Board of Directors or advisory committees; Prelude: Membership on an entity's Board of Directors or advisory committees; Isoplexis: Membership on an entity's Board of Directors or advisory committees; Janssen: Consultancy; C4 Therapeutics: Membership on an entity's Board of Directors or advisory committees; Mission Bio: Membership on an entity's Board of Directors or advisory committees; Auron: Membership on an entity's Board of Directors or advisory committees. Roshal: Physicians' Education Resource: Other: Provision of services; Celgene: Other: Provision of services; Auron Therapeutics: Other: Ownership / Equity interests; Provision of services. Glass: GLG: Consultancy. Xiao: Stemline Therapeutics: Research Funding.

Genomic studies in acute myeloid leukemia (AML) have generated a near complete catalogue of genes mutated at varying frequencies both across patients and in individual leukemias. The high variability of mutation burden within a given leukemia is suggestive of a stepwise evolutionary process composed of early, clonal, mutations and subsequent subclonal events. The receptor tyrosine kinase, FLT3, is the most commonly mutated gene in AML, with mutations frequently manifesting as internal tandem duplications (ITDs) in the juxtamembrane domain leading to constitutive kinase activation. Although FLT3 is commonly a subclonal mutational event, FLT3 ITD mutations portend a poor prognosis particularly when combined with DNMT3A and NPM1, earlier mutations that drive clonal expansion. Notwithstanding its role as a subclonal driver, previous preclinical FLT3 models have utilized retroviral overexpression or germline mutant expression at the endogenous locus precluding accurate temporal modeling of disease. These efforts have prohibited evaluation of FLT3 mutational acquisition in the context observed in AML patients. Here, we report the development of an endogenously targeted, Flp inducible, Flt3 ITD mouse allele which can be somatically activated subsequent to cooperating disease alleles. When activated with a tamoxifen inducible FlpoER, Flt3 mutant mice developed rapid leukocytosis peaking at 4-6 weeks post activation and resolving by 8-10 weeks, a finding not previously observed in constitutive models. This leukocytosis was disproportionately monocytic and accompanied by pronounced anemia and thrombocytopenia. Long term, these mice develop a myeloproliferative disease , reminiscent of previously reported constitutive alleles. In competitive transplantation studies, Flt3 mutant cells initiated disease and outcompeted wild-type cells. Despite this competitive advantage, disease was incapable of transplanting into secondary recipients. We further observed a non-cell autonomous depletion of SLAM+ LSKs suggesting the Flt3 mutant cells cannot propagate disease in self-renewing stem cells. To evaluate how this allele influenced leukemic evolution we crossed this Flt3 ITD allele to a Flp inducible Npm1 c mouse where a pulse of tamoxifen simultaneously activated both alleles. The combination of mutant Npm1 and Flt3 resulted in progressive leukocytosis which did not resolve. Within 6 weeks of mutational activation, these mice developed a lethal AML with robust anemia, thrombocytopenia, leukocytosis and expanded cKIT+ blasts in the blood. RNA-sequencing and immunophenotyping by CyTOF revealed distinct patterns of differentiation, gene-expression and downstream signaling.In an effort to model sequential mutational acquisition, we crossed the Flp Flt3 ITD allele to a Cre-inducible Dnmt3a R878H. Cre mRNA was electroporated into lineage negative bone marrow cells to activate the Dnmt3a R878H allele and transplanted into lethally irradiated recipients. Four weeks post engraftment, Flt3 ITD was activated with a pulse of tamoxifen. In contrast to the Flt3-Npm1 model, we observed an increase and subsequent decrease in WBC similar to the kinetics observed in Flt3 ITD only mice. However, by 20 weeks we observed a robust and consistent increase in WBC accompanied by an emergence of cKIT+ cells in the blood. Histopathology indicated that >50% of mice expressing both alleles in sequence developed AML marked by increased blasts in the marrow, with moderate anemia and thrombocytopenia compared to the Flt3-Npm1 models. Critically, in contrast to Flt3 ITD only mice, acquisition of the Flt3 ITD in Npm1 or Dnmt3a mutant HPSCs induced fully transplantable AML with immunophenotypic characteristics seen in human AML with these same genotypes. Collectively these results demonstrate that different co-occurring mutations are capable of transforming Flt3 ITD mutant cells, albeit with distinct latencies and mechanisms of cooperativity. In summary, our studies utilizing novel multi-recombinase models of leukemogenesis reveal new insights into the early phase of oncogene activation, and how cooperating alleles influence this response. This inducible Flt3 ITD allele represents a significant advance in modeling clonal evolution in myeloid malignancies and provides a critical isogenic platform for preclinical development of novel leukemia therapeutic regimens. Figure 1 Figure 1. Disclosures Bowman: Mission Bio: Honoraria, Speakers Bureau. Xiao: Stemline Therapeutics: Research Funding. Miles: Mission Bio: Honoraria, Speakers Bureau. Trowbridge: Fate Therapeutics: Patents & Royalties; H3 Biomedicine: Research Funding. Levine: Amgen: Honoraria; Lilly: Honoraria; Mission Bio: Membership on an entity's Board of Directors or advisory committees; Imago: Membership on an entity's Board of Directors or advisory committees; Celgene: Research Funding; Ajax: Membership on an entity's Board of Directors or advisory committees; QIAGEN: Membership on an entity's Board of Directors or advisory committees; Gilead: Honoraria; Zentalis: Membership on an entity's Board of Directors or advisory committees; Isoplexis: Membership on an entity's Board of Directors or advisory committees; Roche: Honoraria, Research Funding; Janssen: Consultancy; Astellas: Consultancy; Morphosys: Consultancy; Incyte: Consultancy; Auron: Membership on an entity's Board of Directors or advisory committees; Prelude: Membership on an entity's Board of Directors or advisory committees; C4 Therapeutics: Membership on an entity's Board of Directors or advisory committees.

Background CD19-targeted chimeric-antigen receptor-modified T-cells (CAR-T) are promising in the treatment of refractory B-lymphoblastic leukemia (B-ALL). Minimal residual disease (MRD) detection by multicolor flow cytometry (FCM) is critical to distinguish B-ALL MRD from regenerating, non-neoplastic B-cell populations. Methods FCM was performed on samples from 9 patients with B-ALL treated with CAR-T. Results All 9 patients showed response to CAR-T. Additionally, FCM revealed circulating CD10 + B cells, potentially mimicking MRD. Circulating CD10+ B-cells were detected in blood from 3 days to 3 months after CAR-T, comprising 73% (median) of B-cells (52–83%, 95%CI). They expressed CD19, CD10, CD20, bright CD9, CD22, CD24, moderate CD38 and dim CD58, but were CD34 (–), with bright CD45 and polyclonal surface light chain immunoglobulin (sIg) expression. A similar CD10 + B-cell subpopulation was detected by marrow FCM, amidst abundant B-cell precursors. Conclusions These circulating CD10 + B-cells are compatible with immature B-cells, and are a reflection of B-cell recovery within the marrow. They are immunophenotypically distinguishable from residual B-ALL. Expression of light chain sIg and key surface antigens characterizing regenerating B-cell precursors can distinguish immature B-cells from B-ALL MRD and prevent misdiagnosis. © 2017 International Clinical Cytometry Society

Background: Venetoclax combined with azacitidine (aza/ven), decitabine (dec/ven), and low-dose cytarabine (LDAC/ven) is commonly used off-label in relapsed/refractory (RR)-AML patients; however, predictors of response and survival are incompletely understood. Objectives: To report clinical outcomes of RR-AML patients (pts) treated with venetoclax combination therapy and to analyze molecular predictors of these outcomes. Methods: All pts with RR-AML, who received treatment with aza/ven, dec/ven or LDAC/ven from 08/2016 to 02/2020 at Memorial Sloan Kettering Cancer Center in New York City were included. The best response to therapy was determined using the 2017 European LeukemiaNet (ELN) response criteria. The overall response rate (ORR) was defined as the combination of the complete response (CR) + complete response with incomplete count recovery (CRi) + morphologic leukemia free state (MLFS) rate. Measurable residual disease (MRD) was assessed by multiparameter flow cytometric analysis of bone marrow aspirate samples and any level of residual disease was considered MRD+. Overall survival (OS) was calculated from cycle 1 day 1 of therapy until death or time of last follow-up. Results: A total of 86 pts were treated with venetoclax-based combination therapy (A). Median age was 67 years. More than half of the pts had received prior HMA therapy, 17% had received a prior allogeneic stem cell transplant. The majority of pts received aza/ven (41%) while 23% and 31% received dec/ven and LDAC/ven, respectively. Treatment trajectories of all patients are shown in the Swimmers' plot in panel (B). While the ORR was 31% and 24% of patients achieved a CR/CRi, 60% of pts were refractory to venetoclax therapy; 45% of responding pts eventually relapsed with a median follow up of 12 months (B, C). Oncoprint showing associations between molecular patterns and response to venetoclax combination therapy is shown in panel (D). Mutations in NPM1 (CR/CRi 46%, ORR 62%; OR 4.53, 95% CI 1.31-15.66, p=0.02) and IDH1 (CR/CRi 56%, ORR 67%; OR 5.3, 95% CI 1.21-23.24, p=0.03) but not in IDH2 (CR/CRi 40%, ORR 40%; OR 1.57, 95% CI 0.49-4.99, p=0.54) were associated with statistically significantly increased response rates. Adverse cytogenetics predicted lower odds of response (CR/CRi 11%, ORR 20%; OR 0.32, 95% CI 0.11-0.9, p=0.03). While TP53 (0%), NRAS/KRAS (20%/0%), SF3B1 (0%), ASXL1 (17%) and EZH2 (0%) mutations were associated with low response rates, due to small sample size, these associations were not statistically significant. Median OS for all pts was 6.1 months (95%CI: 4.9-10 month) (E). Pts who were treated with aza/ven had a median OS of 25 months (95% CI 5.8 months-NR), as compared to 5.4 months for patients treated with dec/ven (95% CI 3.9 months-NR; HR 1.63, p=0.2) and 3.9 months for patients treated with LDAC/ven (OS 3.9 months, 95% CI 2.5-8.3 months; HR 2.71, p=0.002). Presence of adverse cytogenetics, TP53 and KRAS/NRAS mutations was associated with poor OS (4.1 months, 95% CI 3.2-6.1 months) compared to a favorable OS when these features were absent (15 months, 95% CI 7.1 months-NA, p=0.001; F). In contrast, presence of mutations in IDH1/2, NPM1 or STAG2 was associated with prolonged OS (15 months, 95% CI 7.1 months-NA) whereas absence of these favorable mutations predicted poor OS (4.4 months, 95% CI 3.5-6.4 months, p=0.0051). Additionally, EZH2 (HR 4.13, p=0.01; H) and SF3B1 (HR 2.5, p=0.02; I) mutations predicted worse OS. Conclusion: Response rates to venetoclax combination therapy in RR-AML pts appeared lower than what has been reported for AML pts who receive venetoclax therapy as first line treatment. However, response rates were high and survival favorable for RR-AML pts with mutations in NPM1, IDH1 and STAG2. In contrast, RR-AML pts with high-risk molecular features such as adverse cytogenetics and mutations in TP53, KRAS/NRAS, EZH2 and SF3B1 had poor outcomes. Clinical trials combining aza/ven with novel therapeutics targeting specific adverse molecular characteristics are urgently needed to improve the outcomes of venetoclax therapy in RR-AML patients. Figure Disclosures Xiao: Stemline Therapeutics: Research Funding. Glass:Gerson Lehman Group: Consultancy. King:Abbvie: Other: advisory board. Abdel-Wahab:Merck: Consultancy; Janssen: Consultancy; Envisagenics Inc.: Current equity holder in private company; H3 Biomedicine Inc.: Consultancy, Research Funding. Levine:Novartis: Consultancy; Celgene: Consultancy, Honoraria, Research Funding; Roche: Consultancy, Honoraria, Research Funding; Lilly: Consultancy, Honoraria; Janssen: Consultancy; Astellas: Consultancy; Morphosys: Consultancy; Amgen: Honoraria; Isoplexis: Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees; Gilead: Honoraria; Imago: Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees; C4 Therapeutics: Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees; Loxo: Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees; Prelude Therapeutics: Research Funding; Qiagen: Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees. Stein:Amgen: Consultancy; Astellas Pharmaceuticals: Consultancy, Membership on an entity's Board of Directors or advisory committees; Agios Pharmaceuticals: Consultancy, Membership on an entity's Board of Directors or advisory committees; Celgene Pharmaceuticals: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Biotheryx: Consultancy; Abbvie: Consultancy; Seattle Genetics: Consultancy; Syndax: Consultancy, Research Funding; Genentech: Consultancy, Membership on an entity's Board of Directors or advisory committees; Bayer: Research Funding; Syros: Membership on an entity's Board of Directors or advisory committees; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; PTC Therapeutics: Membership on an entity's Board of Directors or advisory committees; Daiichi-Sankyo: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding. Tallman:UpToDate: Patents & Royalties; Novartis: Membership on an entity's Board of Directors or advisory committees; Roche: Membership on an entity's Board of Directors or advisory committees; Jazz Pharma: Membership on an entity's Board of Directors or advisory committees; Oncolyze: Membership on an entity's Board of Directors or advisory committees; Delta Fly Pharma: Membership on an entity's Board of Directors or advisory committees; Rigel: Membership on an entity's Board of Directors or advisory committees; KAHR: Membership on an entity's Board of Directors or advisory committees; Daiichi-Sankyo: Membership on an entity's Board of Directors or advisory committees; Bioline rx: Membership on an entity's Board of Directors or advisory committees; Amgen: Research Funding; Rafael: Research Funding; Glycomimetics: Research Funding; BioSight: Membership on an entity's Board of Directors or advisory committees, Research Funding; ADC Therapeutics: Research Funding; Orsenix: Research Funding; Cellerant: Research Funding; Abbvie: Research Funding. Goldberg:Aptose: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Celgene: Consultancy; Daiichi Sankyo: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Genentech: Consultancy, Membership on an entity's Board of Directors or advisory committees; ADC Therapeutics: Research Funding; Aprea: Research Funding; AROG: Research Funding; AbbVie: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Celularity: Research Funding; Pfizer: Research Funding; Dava Oncology: Honoraria. OffLabel Disclosure: venetoclax therapy for the treatment for realpsed and treatment refractory AML

Several novel mechanism-based therapeutic modalities are currently being clinically investigated for the treatment of patients with acute myelogenous leukemia (AML), including agents that exploit genomic vulnerabilities, attenuate leukemia stem cell populations and/or or synergize with anti-leukemic cytotoxic/epigenetic therapies. Lysine-specific demethylase 1 (LSD1) is an enzyme that functions in transcriptional repression by catalyzing the removal of histone H3 lysine 4 methylation, a histone modification associated with transcriptionally competent gene enhancers and transcriptional start sites. Small molecule mediated inhibition of LSD1 alters the chromatin state and the transcriptional output of LSD1 target genes. Transcriptional 'reprogramming' by LSD1 inhibitors either causes a direct impact on cell fate and/or renders malignant cells more susceptible to the treatment with other cancer therapeutic agents. LSD1 inhibitors have shown encouraging phenotypic effects in myelogenous leukemia (AML) models but the key molecular determinants governing LSD1 inhibitor sensitivity remain to be further investigated. Here, we explored the in vitro sensitivity of 350 cancer cell lines to our LSD1 inhibitor CPI-482 to identify potential hyper-responder cell contexts. Four AML cell lines showed high sensitivity with low nanomolar concentration GI50s, each of which contained either a JAK2V617F mutation or a genetic aberration that resulted in JAK-STAT pathway activation. Oral administration of LSD1 inhibitor CPI-482 on a once daily or a once weekly dosing schedule resulted in significant tumor growth inhibition in SET-2 and HEL 92.1.7 JAK2 mutant AML xenograft mouse models. Given the unmet need and poor prognosis in post-MPN secondary AML (sAML) we then explored CPI-482 in a tertiary transplant post-MPN AML retroviral transduction murine model (Jak2V617F retrovirus transduced intoTp53 null bone marrow). Jak2V617F/Tp53 null spleen cells were transplanted into lethally irradiated recipient mice along with wild-type donor support whole bone marrow cells. Mice were randomized to treatment with vehicle, Ruxolitinib (60mg/kg twice daily) or CPI-482 (60mg/kg weekly). Once-weekly treatment with CPI-482 significantly improved survival compared to vehicle (p Treatment of the JAK2V617F mutant AML cell lines SET-2 and HEL with CPI-482 resulted in specific transcriptional effects, including increased expression of the myeloid differentiation markers LY96 and CD86 and inflammatory response genes. CPI-482 also resulted in upregulation of genes that are repressed by the HOXA9 oncogene in other leukemia contexts. The induction of specific CPI-482 mediated gene expression and phenotypic changes was recapitulated by knockdown of the transcription factor GFI1B, suggesting that, consistent with prior findings in other leukemia contexts, LSD1 functionally cooperates with GFI1B in JAK2V617F mutant AML cells. These data provide support for a potential therapeutic impact of the LSD1 inhibitor CPI-482 in AML and sAML in the context of the JAK2V617F mutation, and thus extend the previous findings that LSD1 inhibitors may have utility in JAK2V617F mutated malignant proliferative neoplasms. Given the pressing need for new therapies for sAML which evolves from a pre-existing MPN, we believe these data form the rationale for a mechanism based clinical trial in this adverse risk myeloid malignancy. Figure Disclosures Rampal: Pharmaessentia: Consultancy; Galecto: Consultancy; Abbvie: Consultancy; Stemline: Consultancy, Research Funding; Constellation: Research Funding; Incyte: Consultancy, Research Funding; Celgene: Consultancy; Promedior: Consultancy; CTI Biopharma: Consultancy; Jazz Pharmaceuticals: Consultancy; Blueprint: Consultancy. McGrath:Constellation Pharmaceuticals: Current Employment, Current equity holder in publicly-traded company. Xiao:Stemline Therapeutics: Research Funding. Nikom:Constellation Pharmaceuticals: Current Employment, Current equity holder in publicly-traded company. Wang:Constellation Pharmaceuticals: Current Employment, Current equity holder in publicly-traded company. Levine:Imago: Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees; C4 Therapeutics: Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees; Isoplexis: Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees; Celgene: Consultancy, Honoraria, Research Funding; Roche: Consultancy, Honoraria, Research Funding; Lilly: Consultancy, Honoraria; Janssen: Consultancy; Astellas: Consultancy; Morphosys: Consultancy; Novartis: Consultancy; Amgen: Honoraria; Gilead: Honoraria; Prelude Therapeutics: Research Funding; Qiagen: Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; Loxo: Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees. Trojer:Constellation Pharmaceuticals: Current Employment, Current equity holder in publicly-traded company.

Background: Venetoclax (ven) combined with azacitidine (aza/ven), decitabine (dec/ven), and low-dose cytarabine (LDAC/ven) is approved as frontline therapy for older or otherwise unfit AML patients (pts) and also frequently used for pts with relapsed/refractory (RR)-AML. However, clinical outcomes of AML pts who receive an allogeneic stem cell transplant (alloSCT) after ven combination (ven combo) therapy are unclear. Methods: All AML pts who received treatment with aza/ven, dec/ven or LDAC/ven as either initial induction or for RR disease at Memorial Sloan Kettering Cancer Center from 08/2016 to 02/2020 and underwent a subsequent allo-SCT were included. The response to ven therapy prior to alloSCT was determined using the 2017 European LeukemiaNet response criteria. The overall response rate was defined as the combination of the complete response (CR) + complete response with incomplete count recovery (CRi) + morphologic leukemia free state (MLFS) rate. Measurable residual disease (MRD) was assessed by multiparameter flow cytometric analysis of bone marrow aspirate samples. Any level of residual disease was considered MRD+. Overall survival (OS) after alloSCT was calculated from the day of graft infusion until death or time of last follow-up. Results: A total of 130 pts were treated with ven combo therapy with 18 pts (13.8% of all pts) receiving a subsequent alloSCT. Median age was 65 years (A). While 3 pts received a ven combo as the first treatment for AML, 15 pts had RR-AML. Aza/ven was most commonly used (72%) and the majority of pts (83%) received 1-2 cycles of ven therapy prior to alloSCT. For 7, 6, 4 and 1 pts the donor was a matched related, a matched unrelated, a mismatched unrelated, or haploidentical; only one pt had received a prior alloSCT for AML (B). Unmodified peripheral blood stem cells (66%) and reduced intensity conditioning regimens (77%) were most commonly used. In pts who proceeded to an alloSCT, 12/18 pts achieved a response after ven combination therapy: 4/18 MRD-CR/CRi, 4/18 MRD+CR/CRi, 4/18 MLFS; 6/18 pts had persistent disease (C and D). It is important to note that these response rates were specific to patients who received an alloSCT after ven combination therapy. While pts with DNMT3A, NPM1, IDH1/2 and FLT3 mutations had a high response rate to ven therapy prior to alloSCT, none of the pts with TP53 or NRAS mutations achieved a response prior to alloSCT (E). Only DNMT3A mutations were statistically significantly associated with a high response rate prior to alloSCT (ORR 100%, CR/CRi 63%, p=0.01). AlloSCT was able to convert pts with MRD+ or persistent disease into an MRD- state in 50% of cases (F). With a median follow up of 10 months, the median OS was not reached; 56% of pts relapsed after alloSCT. Median time to relapse for all pts was 18 months (95% CI 4 months-not reached [NR]). Disease status prior to alloSCT was a powerful predictor of post-transplant outcomes. Pts who achieved CR, CRi, or MLFS with ven therapy prior to alloSCT had significantly prolonged OS (median OS not reached) compared with pts who had persistent disease prior to alloSCT (median OS 7 months, 95% CI 2.14 months-NR; p=0.029; G). The poor OS for pts with persistent disease prior to alloSCT was mainly driven by a high incidence of relapse for these pts compared to pts who achieved a response to ven combos (H). The median time to relapse after alloSCT was 17.9 months (95% CI 6.1 months-NR) for pts who achieved CR, CRi, or MLFS prior to alloSCT, and only 3.3 months (95% CI 1.9 months-NR, p=0.052) for pts with persistent disease prior to alloSCT. While 67% of pts experienced grade I-II acute graft versus host disease (aGVHD), only 1 pt was found to have grade III aGVHD and no pt experienced grade IV aGVHD. Median time to aGVHD after alloSCT was 1.4 months (95% CI 0.89 months-NR). Conclusion: Venetoclax combination therapy can bridge some AML pts to subsequent alloSCT. Most pts who underwent alloSCT received reduced-intensity conditioning given their advanced age and comorbidities. Pts who achieved CR, CRi, or MLFS with ven-based therapy prior to alloSCT had more favorable outcomes after alloSCT. With more widespread use of venetoclax, we anticipate that alloSCT following good responses to venetoclax combination therapy will become more common in pts with AML. Longer-term studies are needed to determine durability of post-alloSCT responses following pre-alloSCT venetoclax combination therapy. Figure Disclosures Ponce: Kadmon: Membership on an entity's Board of Directors or advisory committees; Generon: Membership on an entity's Board of Directors or advisory committees; Ceramedix: Membership on an entity's Board of Directors or advisory committees; Takeda: Research Funding. Young:Amgen: Other: Common stock; Merck: Other: Common stock; Pfizer: Other: Common stock. Gyurkocza:Actinium: Research Funding. Chan:AbbVie: Current equity holder in publicly-traded company; Bristol-Myers Squibb: Current equity holder in publicly-traded company. Xiao:Stemline Therapeutics: Research Funding. Glass:Gerson Lehman Group: Consultancy. King:Abbvie: Other: advisory board. Cai:Imago Biosciences, Inc.: Consultancy, Current equity holder in private company; DAVA Oncology: Honoraria. Stein:Celgene Pharmaceuticals: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Agios Pharmaceuticals: Consultancy, Membership on an entity's Board of Directors or advisory committees; Amgen: Consultancy; Abbvie: Consultancy; Seattle Genetics: Consultancy; Syndax: Consultancy, Research Funding; Genentech: Consultancy, Membership on an entity's Board of Directors or advisory committees; Bayer: Research Funding; Biotheryx: Consultancy; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; PTC Therapeutics: Membership on an entity's Board of Directors or advisory committees; Syros: Membership on an entity's Board of Directors or advisory committees; Astellas Pharmaceuticals: Consultancy, Membership on an entity's Board of Directors or advisory committees; Daiichi-Sankyo: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding. Giralt:GSK: Membership on an entity's Board of Directors or advisory committees, Research Funding; Celgene: Membership on an entity's Board of Directors or advisory committees, Research Funding; Actinuum: Other: Advisory board, Research Funding; Amgen: Other: Advisory Board, Research Funding; OMEROS: Research Funding; Jensenn: Membership on an entity's Board of Directors or advisory committees, Research Funding; Johnson & Johnson: Membership on an entity's Board of Directors or advisory committees, Research Funding; Novartis: Membership on an entity's Board of Directors or advisory committees; TAKEDA: Membership on an entity's Board of Directors or advisory committees, Research Funding; JAZZ Pharmaceutical: Membership on an entity's Board of Directors or advisory committees; MILTENYI: Research Funding; PFIZER: Membership on an entity's Board of Directors or advisory committees, Research Funding; Sanofi: Membership on an entity's Board of Directors or advisory committees, Research Funding; BMS: Membership on an entity's Board of Directors or advisory committees, Research Funding; SPECTRUM Pharma: Membership on an entity's Board of Directors or advisory committees; KITE: Membership on an entity's Board of Directors or advisory committees. Perales:Omeros: Honoraria, Membership on an entity's Board of Directors or advisory committees; Medigene: Membership on an entity's Board of Directors or advisory committees, Other; NexImmune: Membership on an entity's Board of Directors or advisory committees; Servier: Membership on an entity's Board of Directors or advisory committees, Other; Cidara Therapeutics: Other; Takeda: Honoraria, Membership on an entity's Board of Directors or advisory committees; Novartis: Honoraria, Membership on an entity's Board of Directors or advisory committees; Nektar Therapeutics: Honoraria, Membership on an entity's Board of Directors or advisory committees; Abbvie: Honoraria, Membership on an entity's Board of Directors or advisory committees; MolMed: Membership on an entity's Board of Directors or advisory committees; Merck: Consultancy, Honoraria; Miltenyi Biotec: Research Funding; Kite/Gilead: Honoraria, Research Funding; Incyte Corporation: Honoraria, Research Funding; Bristol Myers Squibb: Honoraria, Membership on an entity's Board of Directors or advisory committees; Celgene: Honoraria; Bellicum: Honoraria, Membership on an entity's Board of Directors or advisory committees. Tallman:Novartis: Membership on an entity's Board of Directors or advisory committees; Cellerant: Research Funding; Orsenix: Research Funding; ADC Therapeutics: Research Funding; UpToDate: Patents & Royalties; Abbvie: Research Funding; Roche: Membership on an entity's Board of Directors or advisory committees; Glycomimetics: Research Funding; Rafael: Research Funding; Amgen: Research Funding; Bioline rx: Membership on an entity's Board of Directors or advisory committees; Daiichi-Sankyo: Membership on an entity's Board of Directors or advisory committees; KAHR: Membership on an entity's Board of Directors or advisory committees; Rigel: Membership on an entity's Board of Directors or advisory committees; Delta Fly Pharma: Membership on an entity's Board of Directors or advisory committees; Oncolyze: Membership on an entity's Board of Directors or advisory committees; BioSight: Membership on an entity's Board of Directors or advisory committees, Research Funding; Jazz Pharma: Membership on an entity's Board of Directors or advisory committees. Goldberg:Genentech: Consultancy, Membership on an entity's Board of Directors or advisory committees; Daiichi Sankyo: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Celgene: Consultancy; Aptose: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; AbbVie: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; ADC Therapeutics: Research Funding; Aprea: Research Funding; AROG: Research Funding; Celularity: Research Funding; Pfizer: Research Funding; Dava Oncology: Honoraria.

Background: Mixed phenotype acute leukemia is a rare disease characterized by an expanded blast population exhibiting multiple lineage features. It is a diagnosis of exclusion, and we and others have observed that AML-MRC or therapy-related AML cases may also exhibit similar features. We therefore sought to determine whether multi-lineage expression is present in de-novo and therapy-related MDS and whether it has an impact on clinical outcomes. Methods: We reviewed pathology, flow cytometry, cytogenetic, and molecular reports from patients seen at Memorial Sloan Kettering Cancer Center between 1996 and 2020 and identified 472 patients diagnosed with MDS using a combination of custom natural language processing tools and manual review. Cox proportional hazards modeling was performed to assess the contribution of patient characteristics, pathology, flow cytometric, cytogenetic, and molecular characteristics to overall survival (OS). Fisher's exact testing was used to assess the association of individual features. Results: We found that an abnormal myeloid lineage signature was associated with poorer OS after adjusting for age, cytogenetics, and molecular features (HR=2.3, p Conclusion: Routine flow cytometric and molecular assessment of diagnostic bone marrows can provide added prognostic value in MDS. Identification of an abnormal myeloid lineage is associated with higher risk disease, while identification of abnormal plasma cell features is associated with lower risk disease. SRSF2 mutation is also associated with abnormal plasma cell features, but does not contribute independently to OS. Abnormal T-cell features may also provide prognostic value, although assessment in a larger cohort will be necessary. We confirm previous findings indicating that TP53 mutation is associated with higher risk disease while SF3B1 mutation is associated with better outcomes characteristic of MDS-RARS. In addition, our data suggests that IDH1 and EZH2 are associated with poorer outcomes and that IDH2 is associated with lower risk disease. Figure 1 Disclosures Tallman: Bioline rx: Membership on an entity's Board of Directors or advisory committees; Delta Fly Pharma: Membership on an entity's Board of Directors or advisory committees; Amgen: Research Funding; Abbvie: Research Funding; Cellerant: Research Funding; Orsenix: Research Funding; ADC Therapeutics: Research Funding; BioSight: Membership on an entity's Board of Directors or advisory committees, Research Funding; Glycomimetics: Research Funding; Rafael: Research Funding; Oncolyze: Membership on an entity's Board of Directors or advisory committees; UpToDate: Patents & Royalties; Novartis: Membership on an entity's Board of Directors or advisory committees; Roche: Membership on an entity's Board of Directors or advisory committees; Jazz Pharma: Membership on an entity's Board of Directors or advisory committees; Daiichi-Sankyo: Membership on an entity's Board of Directors or advisory committees; KAHR: Membership on an entity's Board of Directors or advisory committees; Rigel: Membership on an entity's Board of Directors or advisory committees. Levine:Morphosys: Consultancy; Astellas: Consultancy; Janssen: Consultancy; Lilly: Consultancy, Honoraria; Isoplexis: Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees; C4 Therapeutics: Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees; Imago: Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees; Loxo: Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees; Qiagen: Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; Prelude Therapeutics: Research Funding; Gilead: Honoraria; Novartis: Consultancy; Amgen: Honoraria; Roche: Consultancy, Honoraria, Research Funding; Celgene: Consultancy, Honoraria, Research Funding. Xiao:Stemline Therapeutics: Research Funding. Glass:Gerson Lehman Group: Consultancy.

TAFRO syndrome, a clinical subtype of idiopathic multicentric Castleman disease (iMCD), consists of a constellation of symptoms/signs including thrombocytopenia, anasarca, fever, reticulin fibrosis/renal dysfunction, and organomegaly. The etiology of iMCD-TAFRO and the basis for cytokine hypersecretion commonly seen in iMCD-TAFRO patients has not been elucidated. Here, we identified a somatic MEK2P128L mutation and a germline RUNX1G60C mutation in two patients with iMCD-TAFRO, respectively. The MEK2P128L mutation, which has been identified previously in solid tumor and histiocytosis patients, caused hyperactivated MAP kinase signaling, conferred IL-3 hypersensitivity and sensitized the cells to various MEK inhibitors. The RUNX1G60C mutation abolished the transcriptional activity of wild-type RUNX1 and functioned as a dominant negative form of RUNX1, resulting in enhanced self-renewal activity in hematopoietic stem/progenitor cells. Interestingly, ERK was heavily activated in both patients, highlighting a potential role for activation of MAPK signaling in iMCD-TAFRO pathogenesis and a rationale for exploring inhibition of the MAPK pathway as a therapy for iMCD-TAFRO. Moreover, these data suggest that iMCD-TAFRO might share pathogenetic features with clonal inflammatory disorders bearing MEK and RUNX1 mutations such as histiocytoses and myeloid neoplasms.

Hodgkin/Reed-Sternberg (HRS) cells in the setting of chronic lymphocytic leukemia (CLL) exist in 2 forms: type I with isolated HRS cells in a CLL background (Hodgkin-like lesion) and type II with typical classic Hodgkin lymphoma, a variant of Richter transformation (CHL-RT). The clinical significance of the 2 morphological patterns is unclear, and their biological features have not been compared. We retrospectively reviewed 77 cases: 26 of type I and 51 of type II CHL-RT; 3 cases progressed from type I to type II. We examined clinical features, Epstein-Barr virus (EBV) status, and clonal relatedness after microdissection. Median age for type I was 62 years versus 73 years for type II (P=.01); 27% (type I) versus 73% (type II) had a history of CLL. HRS cells were positive for EBV in 71% (55/77), similar in types I and II. Clonality analysis was performed in 33 cases (type I and type II combined): HRS cells were clonally related to the underlying CLL in 14 and unrelated in 19. ZAP-70 expression of the CLL cells but not EBV status or morphological pattern was correlated with clonal relatedness: all 14 clonally related cases were ZAP-70 negative, whereas 74% (14/19) of clonally unrelated cases were ZAP-70 positive. Overall median survival (types I and II) after diagnosis was 44 months. Advanced age was an adverse risk factor for survival, but not histologic pattern, type I versus type II. HRS-like cells in a background of CLL carries a similar clinical risk to that of CHL-RT and may progress to classic Hodgkin lymphoma in some cases.

Alpha-synuclein is highly expressed in the central nervous system and plays an important role in pathogenesis of neurodegenerative disorders such as Parkinson's disease and Lewy body dementia. Previous studies have demonstrated the expression of α-synuclein in hematopoietic elements and peripheral blood mononuclear cells, although its roles in hematopoiesis and adaptive immunity are not studied. Using an α-synuclein knock out (KO) mouse model, we have recently shown that α-synuclein deficiency is associated with a mild defect in late stages of hematopoiesis. More importantly, we demonstrated a marked defect in B lymphocyte development and IgG, but not IgM production in these mice. Here we show a marked defect in development of T lymphocytes in α-synuclein KO mice demonstrated by a significant increase in the number of CD4 and CD8 double negative thymocytes and significant decreases in the number of CD4 single positive and CD8 single positive T cells. This resulted in markedly reduced peripheral T lymphocytes. Interestingly, splenic CD4(+) and CD8(+) T cells that developed in α-synuclein KO mice had a hyperactivated state with higher expression of early activation markers and increased IL-2 production. Moreover, splenic CD4(+) T cells from α-synuclein KO mice produced lower levels of IL-4 upon antigenic stimulation suggesting a defective Th2 differentiation. Our data demonstrate an important role for α-synuclein in development of T lymphocytes and regulation of their phenotype and function.

Measurable residual disease is associated with inferior outcomes in patients with acute myeloid leukemia (AML). Measurable residual disease monitoring enhances risk stratification and may guide therapeutic intervention. The European LeukemiaNet working party recently came to a consensus recommendation incorporating leukemia associated immunophenotype-based different from normal approach by multi-color flow cytometry for measurable residual disease evaluation. However, the analytical approach is highly expertise-dependent and difficult to standardize. Here we demonstrate that loss of plasmacytoid dendritic cell differentiation after 7+3 induction in AML is highly specific for measurable residual disease positivity (specificity 97.4%) in a uniformly treated patient cohort. Moreover, loss of plasmacytoid dendritic cell differentiation as determined by a blast-to-plasmacytoid dendritic cell ratio >10 was strongly associated with inferior overall and relapse-free survival (RFS) [Hazard ratio 2.79, 95% confidence interval (95%CI): 0.98-7.97; P=0.077) and 3.83 (95%CI: 1.51-9.74; P=0.007), respectively), which is similar in magnitude to measurable residual disease positivity. Importantly, measurable residual disease positive patients who reconstituted plasmacytoid dendritic cell differentiation (blast/ plasmacytoid dendritic cell ratio

Traditionally low-grade B-cell lymphomas have been excluded from the category of monomorphic posttransplant lymphoproliferative disorders. However, recent reports identified Epstein-Barr virus-positive (EBV) extranodal marginal zone lymphomas (MZL), almost exclusively seen in the posttransplant setting. Some reported cases responded to reduced immunosuppression, suggesting that they should be considered as a form of posttransplant lymphoproliferative disorders. We identified 10 cases of EBV MZL, 9 in extranodal sites and 1 presenting in lymph node. Two cases arose following solid organ transplantation, but other settings included iatrogenic immunosuppression for rheumatoid arthritis (2); prior chemotherapy (2); congenital immune deficiency (1); and increased age (3), as the only potential cause of immune dysfunction. There were 4 males and 6 females; age range 18 to 86. The atypical plasmacytoid and/or monocytoid B cells were positive for EBV in all cases, with either latency I or II in all cases tested. Monotypic light chain expression was shown in all with 6 cases positive for IgG, and 2 for IgM, undetermined in 2. Clonal immunoglobulin gene rearrangement was positive in all cases with successful amplification. MYD88 L265P was wild type in the 6 cases tested. We show that EBV MZLs can arise in a variety of clinical settings, and are most often extranodal. Treatment varied, but most patients had clinically indolent disease with response to reduction of immune suppression, or immunochemotherapy.

The B/T subtype of mixed phenotype acute leukemia (B/T MPAL) is defined by co-expression of antigens of both B- and T-cell lineages on leukemic blasts. Although it has been suggested that multilineage antigen expression portends poor response to chemotherapy, the clinical characteristics and driver mutations that underlie the pathogenesis of this rare subtype of acute leukemia are scarcely known. We identified nine cases of B/T MPAL from multiple institutions and correlated clinical and immunophenotypic findings with next-generation sequencing data. We report that B/T MPAL commonly presents with lymphadenopathy in adolescence and young adulthood. While the tumors have diverse cytogenetic and genomic perturbations, recurrent acquired aberrations include mutations in the putative transcriptional regulator PHF6 and the JAK-STAT and Ras signaling pathways. Alterations were also identified in genes encoding hematopoietic transcription factors, cell cycle regulators/tumor suppressors, and chromatin modifying enzymes. The genomic landscape of B/T MPAL strongly resembles that of T-ALL subgroups associated with early developmental arrest, while genetic alterations that are common in B-ALL were rarely seen. Two-thirds of the patients responded to ALL-based chemotherapy with or without stem cell transplantation. Our observations lay the groundwork for further study of the unique biology and clinical trajectory of B/T MPAL.

Genomic analysis has greatly influenced the diagnosis and clinical management of patients affected by diverse forms of hematologic malignancies. Here, we review how genetic alterations define subclasses of patients with acute leukemias, myelodysplastic syndromes (MDS), myeloproliferative neoplasms (MPNs), non-Hodgkin lymphomas, and classical Hodgkin lymphoma. These include new subtypes of acute myeloid leukemia defined by mutations in RUNX1 or BCR-ABL1 translocations as well as a constellation of somatic structural DNA alterations in acute lymphoblastic leukemia. Among patients with MDS, detection of mutations in SF3B1 define a subgroup of patients with the ring sideroblast form of MDS and a favorable prognosis. For patients with MPNs, detection of the BCR-ABL1 fusion delineates chronic myeloid leukemia from classic BCR-ABL1− MPNs, which are largely defined by mutations in JAK2, CALR, or MPL. In the B-cell lymphomas, detection of characteristic rearrangements involving MYC in Burkitt lymphoma, BCL2 in follicular lymphoma, and MYC/BCL2/BCL6 in high-grade B-cell lymphomas are essential for diagnosis. In T-cell lymphomas, anaplastic large-cell lymphoma is defined by mutually exclusive rearrangements of ALK, DUSP22/IRF4, and TP63. Genetic alterations affecting TP53 and the mutational status of the immunoglobulin heavy-chain variable region are important in clinical management of chronic lymphocytic leukemia. Additionally, detection of BRAFV600E mutations is helpful in the diagnosis of classical hairy cell leukemia and a number of histiocytic neoplasms. Numerous additional examples provided here demonstrate how clinical evaluation of genomic alterations have refined classification of myeloid neoplasms and major forms of lymphomas arising from B, T, or natural killer cells.

TAFRO syndrome is a rare clinicopathologic variant of idiopathic multicentric Castleman disease characterized by Thrombocytopenia, Ascites (anasarca), myeloFibrosis, Renal dysfunction, and Organomegaly. Here, we report a case of TAFRO syndrome in an HIV-negative young Caucasian male who presented with fever, normocytic anemia, thrombocytopenia, and acute renal insufficiency. The serum interleukin-6 (IL-6) level was elevated. Chest and abdominal CT revealed bilateral pleural effusion, ascites, splenomegaly, and multiple mildly enlarged lymph nodes. An excisional biopsy of inguinal lymph node showed a few atrophic follicles and expansion of interfollicular areas by marked vascular proliferation and polytypic plasmacytosis. HHV-8 was negative. Subsequent bone marrow biopsy was normocellular with moderately increased megakaryocytes and occasional megakaryocytic emperipolesis. His signs and symptoms improved after treatment with methylprednisolone and tocilizumab (anti-IL-6 receptor antibody). Our study confirms the distinctive nature of this syndrome, which should allow for better recognition and appropriate therapy. Keywords: Castleman-Kojima disease, TAFRO syndrome, Idiopathic multicentric Castleman disease, IL-6, HHV-8

Background Thrombotic thrombocytopenic purpura (TTP) is a life-threatening diagnosis requiring prompt initiation of therapeutic plasma exchange (TPE). Measurement of immature platelet (PLT) fraction (%-IPF) differentiates PLT consumption or destruction from hypoproduction. Study Design and Method Our study evaluated %-IPF changes over the course of TTP treated with TPE and as a measure of treatment efficacy. Eleven idiopathic TTP patients, two human immunodeficiency virus (HIV)-associated TTP patients, and five non-TTP patients with thrombocytopenia were enrolled into our study. All patients were treated with TPE and had ADAMTS13 activity measured. Results All idiopathic TTP patients had a significantly increased %-IPF and decreased absolute immature PLT count (A-IPC) and PLT count at presentation. An A-IPC value of less than 5 × 109/L at presentation has 84.6% sensitivity, 80% specificity, and 91.7% positive predictive value for diagnosing TTP. A concurrent steady decline in %-IPF and increased PLT counts toward normal was observed in TTP patients undergoing TPE. The A-IPC, however, showed an increase and decrease curve that was not seen in the two HIV-associated TTP patients with no response to TPE and the five non-TTP patients. More importantly, reaching an A-IPC ratio of 3 compared to baseline value during TPE can readily differentiate idiopathic TTP from the other two groups and is correlated with good clinical responses to TPE. An abrupt increase of A-IPC during TPE was also noted in a TTP patient who relapsed 3 days before PLT count decrease. A-IPC is positively correlated with ADAMTS13 activity at presentation but negatively correlated with ADAMTS13 activity during recovery. Conclusion A-IPC should be routinely analyzed for diagnosing and monitoring TTP patients.

CRABP-II, a retinoic acid binding protein, shuffles retinoic acid from cytoplasm into nucleus and forms a complex with nuclear retinoic acid receptor to facilitate transcriptional activities of retinoic acid. In this study, we studied the expression patterns of CRABP-II in pancreatic ductal adenocarcinoma (PDAC) compared with those in normal pancreas, chronic pancreatitis, and precancerous lesions. We showed no detectable expressions of CRABP-II in normal pancreatic parenchyma, normal ductal epithelium, and chronic pancreatitis. In contrast, the expression of CRABP-II was readily detected in all PDACs including metastatic PDACs. CRABP-II staining was also observed and progressively increased from pancreatic intraepithelial neoplasia 1 to 3. In addition, when fine needle aspiration specimens were evaluated from patients with PDAC, CRABP-II was positive in 55.6% cases if cytology diagnosis was "atypia," and in 87.5% cases, if "malignancy." Our study suggests that CRABP-II is highly and specifically expressed in PDAC and is more commonly expressed in high-grade precursor cancerous lesions than in low-grade lesions. Therefore, overexpression of CRABP-II is a late event of pancreatic carcinogenesis, and it could be used as a diagnostic marker to distinguish PDAC from other benign pancreatic conditions in both resection and cytology specimens.

The histological diagnosis of luetic lymphadenitis remains challenging for the pathologists particularly when the pertinent clinical information is lacking. Here we report a case of a 23-year-old male, an immigrant from South America, who had a history of epithelioid hemangioma, presented with a painless enlarged left inguinal lymph node. In addition to the characteristic follicular hyperplasia and capsular thickening, lymphohistiocytic nodules are also seen. Recognition of such rare nodular proliferation can be helpful in diagnosing luetic lymphadenitis.

Background: Myeloid and lymphoid leukemias each have distinct diagnostic algorithms, treatment paradigms, and therapeutic options. In mixed phenotype acute leukemia (MPAL), myeloid and lymphoid immunophenotypes are expressed simultaneously. MPAL is rare, accounting for fewer than 5% of all leukemias, and carries a poor prognosis. Currently, there is no standard of care for treatment of this disease and therapeutic options tailored to this group of patients are lacking. Additionally, the biological mechanisms underlying lineage infidelity are poorly understood. Here we seek to establish more precise biological characterization of this disease and have identified a new subgroup of patients with Mixed Phenotype Acute Myeloid Leukemia (AML-MP). Methods: We developed a novel natural language processing pipeline and used it to review 830 patient records, representing all acute leukemia patients treated at Memorial Sloan Kettering Cancer Center since 2015. We identified those with ambiguous lineages based on multiparameter flow cytometry data and integrated next-generation sequencing, cytogenetics, and clinical information. Statistical methods are outlined with the associated results below. Results: Among the 830 patient records reviewed, 54 (6.5%) patients with mixed lineage characteristics were identified. Of these, 26 (48%) carried a formal diagnosis of MPAL while 28 (52%) carried a diagnosis of AML with myelodysplasia related changes (AML-MRC) or therapy related AML (t-AML). Among the cases expressing multiple lineages, 34 (25%) had B/Myeloid features (11 MPAL, 23 AML-MRC), 17 (13%) had T/Myeloid features (13 MPAL, 4 AML-MRC), 3 (0.7%) had B/T/Myeloid lineage (2 MPAL, 1 AML-MRC). Only 8 patients received prior chemotherapy at our institution > 1 year prior to diagnosis and were classified as t-AML. As a control group, we also identified 79 patients with AML-MRC exhibiting myeloid lineage alone. We pooled the mixed-lineage and myeloid-only AML-MRC cases and performed k-means clustering into 2 groups using all available molecular features (Figure 1a). Although TP53 was enriched within AML cases (p Among patients with mixed lineage characteristics, we performed a cox proportional hazards analysis and found that RUNX1 and SRSF2 mutations were predictors of better overall survival (OS) after adjusting for age, cytogenetics, and diagnosis type (age > 60: p < 0.01, high risk cytogenetics: p Among AML-MRC myeloid only cases, ASXL1 mutation was associated with improved OS and TET2 mutation showed a trend toward poorer OS after adjusting for age and cytogenetic risk (age>60: p=0.01, ASXL1: p=0.02, TET2: p=0.06; overall: p < 0.01, Figure 1b, right). Cytogenetic risk did not independently contribute to the model (high risk: p=0.3, intermediate risk: p=0.55). We also manually reviewed flow cytometry results from 64 patients diagnosed with MPAL or diagnosed with t-AML or AML-MRC despite expressing markers of multiple lineages (Mixed Phenotype AML). Using a k-nearest neighbor approach, we found that immunophenotype identified two distinct patient populations - one largely Mixed Phenotype AML, and the other with a mixture of MPAL and Mixed Phenotype AML cases (p Conclusions: Genomic characteristics improve prognostication for patients with MPAL, with RUNX1 and SRSF2 predicting a more favorable OS and TP53 not predictive of worse OS; this is in contrast to current stratifications of patients with de novo AML. While TP53 is enriched among AML cases, it does not distinguish AML-MRC from MPAL. Additionally, AML-MRC patients with ASXL1 mutations had improved overall survival after adjusting for cytogenetic risk. Some cases that are categorized as AML-MRC and t-AML by WHO criteria should be considered Mixed Phenotype AML and may be better classified into one of two distinct immunophenotypic subsets. Our study and model provide a more refined biological classification and prognostic schema for patients with MPAL and AML-MP; further validation of these observations is required in other data sets. Figure 1 Disclosures Roshal: Auron Therapeutics: Equity Ownership, Other: Provision of services; Celgene: Other: Provision of Services; Physicians' Education Resource: Other: Provision of services. Levine:Prelude Therapeutics: Research Funding; Roche: Consultancy, Research Funding; Celgene: Consultancy, Research Funding; Gilead: Consultancy; Qiagen: Membership on an entity's Board of Directors or advisory committees; C4 Therapeutics: Membership on an entity's Board of Directors or advisory committees; Novartis: Consultancy; Loxo: Membership on an entity's Board of Directors or advisory committees; Isoplexis: Membership on an entity's Board of Directors or advisory committees; Imago Biosciences: Membership on an entity's Board of Directors or advisory committees; Lilly: Honoraria; Amgen: Honoraria. Tallman:Indy Hematology Review: Honoraria; Hematology Oncology of Indiana: Honoraria; Salzberg Weill Cornall MSKCC Seminar in Hematologic Malignancies: Honoraria; AbbVie: Membership on an entity's Board of Directors or advisory committees, Research Funding; International Conference in Leukemia: Honoraria; 14th Annual Miami Cancer Meeting: Honoraria; Amgen: Consultancy; Orsenix: Membership on an entity's Board of Directors or advisory committees, Research Funding; UpToDate: Patents & Royalties; Mayo Clinic: Honoraria; New Orleans Summer Cancer Conference: Honoraria; ADC Therapeutics: Research Funding; Danbury Hospital Tumor Board: Honoraria; Arog Pharmaceuticals: Research Funding; Nohla: Membership on an entity's Board of Directors or advisory committees, Research Funding; KAHR: Membership on an entity's Board of Directors or advisory committees; Daiichi-Sankyo: Membership on an entity's Board of Directors or advisory committees; BioSight: Membership on an entity's Board of Directors or advisory committees, Research Funding; Bioline: Membership on an entity's Board of Directors or advisory committees, Research Funding; University of Oklahoma Medical Center: Honoraria; Cellerant Therapeutics: Research Funding; Rigel: Membership on an entity's Board of Directors or advisory committees.

Dysregulation of the protein arginine methyltransferase, PRMT5, occurs in a variety of malignancies and is associated with increased cancer cell proliferation. We investigated the role of PRMT5 in MPN pathogenesis through use of C220, a potent and selective small molecule PRMT5 inhibitor, and aimed to elucidate key PRMT5 targets contributing to the maintenance of MPN cells. We found that PRMT5 is overexpressed in MPN, most highly in PV. C220 dose-dependently reduced the proliferation of MPN cell lines and BaF3 cells expressing JAK2V617F in vitro. Human CD34+ cells from PV/PMF patients showed a high degree of sensitivity to C220 ex vivo. Consistent with the in vitrostudies, C220 treatment of SET2 xenografts in vivoled to a 71% reduction of median tumor volume, greater than that observed with JAK inhibition (59%). We assessed the efficacy of C220 in Jak2V617F- and MPLW515L-driven murine MPN models.In the conditional Jak2V617Fmodel of PV, mice treated with C220 displayed significant lower hematocrits (median 50.1% vs. 77.4 %, p We next assessed the efficacy of combined PRMT5 and JAK inhibition. Dual targeting of JAK and PRMT5 displayed additive effects on SET2 cell proliferation in vitro and was superior to monotherapy with either agent in vivo. In the Jak2V617Fmodel, dual JAK/PRMT5 inhibition further reduced key features such as splenomegaly (median weight 39 mg vs. 68 mg or 127 mg, p=0.0004 and p To further understand the mechanism by which C220 leads to therapeutic efficacy in MPN, we performed RNA-seq in sorted MEP from the Jak2V617FBMT model and from SET2 xenografts after vehicle or C220 in vivotherapy. GSEA revealed E2F targets as one of the major gene sets down regulated upon treatment with C220. Consistent with the potent suppression of E2F1 target gene expression in vivo, we found that E2F1 and PRMT5 physically interact in JAK2-mutant MPN cells. Moreover, PRMT5 inhibition altered the methylation status of E2F1, leading to reduced expression of E2F1 downstream targets, including genes involved in cell cycle and DNA damage repair. Functional investigation of key downstream targets is ongoing. Our data provide evidence that PRMT5 is a credentialed therapeutic target in MPN, alone and in combination with JAK inhibition. Moreover, the therapeutic effects of PRMT5 inhibition in MPN include attenuated expression of E2F targets, which our data suggests is mediated by PRMT5 methylation of E2F1. These data demonstrate a novel link between PRMT5, E2F1 gene regulatory function, and the survival of MPN cells and provide a strong mechanism-based rationale for therapeutic studies of PRMT5 inhibitors in MPN. Based on these studies and others, PRT543, a novel and selective PRMT5 inhibitor, is currently being evaluated in a Phase I clinical trial in advanced cancers, including myelofibrosis (NCT03886831). Disclosures Bhagwat: Prelude Therapeutics: Employment. Viny:Hematology News: Membership on an entity's Board of Directors or advisory committees; Mission Bio: Other: Sponsored travel. Grego:Prelude Therapeutics: Employment. Mehta:Prelude Therapeutics: Employment. Scherle:Prelude Therapeutics: Employment. Vaddi:Prelude Therapeutics: Employment. Levine:Novartis: Consultancy; Roche: Consultancy, Research Funding; Imago Biosciences: Membership on an entity's Board of Directors or advisory committees; Prelude Therapeutics: Research Funding; Celgene: Consultancy, Research Funding; Isoplexis: Membership on an entity's Board of Directors or advisory committees; C4 Therapeutics: Membership on an entity's Board of Directors or advisory committees; Loxo: Membership on an entity's Board of Directors or advisory committees; Qiagen: Membership on an entity's Board of Directors or advisory committees; Gilead: Consultancy; Lilly: Honoraria; Amgen: Honoraria.

Background Myelodysplastic syndrome (MDS) is a clonal, pre-leukemic stem cell disorder characterized by decreased blood counts due to ineffective hematopoiesis. Plasmacytoid dendritic cells (PDC) are stem cell derived, type I interferon producing dendritic cells, that are readily identifiable by flow cytometry (FC) using expression of CD123 and HLA-DR. PDCs have been shown to be decreased in acute myeloid leukemia (AML). Our study uses FC to evaluate PDC in MDS, their relationship to the Revised International Prognostic Scoring System (IPSS-R) for MDS, as well as their relationship to clinical outcomes. Methods We identified 197 patients with new MDS diagnoses and examined FC data of bone marrow aspirates (BMA) at first presentation to our institution for blast and PDC percentages. Patients who presented with an outside diagnosis of MDS greater than 1 year before presentation to our institution were excluded. MDS with excess blasts 1 and 2 (MDS-EB1/EB2) were designated as high grade, while MDS with isolated del5q (MDS-d5q), single lineage dysplasia (MDS-SLD), and multilineage dysplasia (MDS-MLD) were designated as low grade, with or without ring sideroblasts. 163 patients had sufficient data to calculate their IPSS-R risk categories. Sixteen patients with a history of solid tumor malignancies undergoing BMA for cytopenias were used as controls. The bone marrow of these control patients showed no evidence of morphologic dysplasia, they had normal karyotypes, and no pathogenic variants were detected on a 28 gene next generation sequencing based myeloid panel. We used CD34 and CD117 to quantify blasts, and CD123 and HLA-DR to quantify PDCs. Results The proportion of PDCs expressed as a percentage of total WBCs was significantly lower in higher risk MDS diagnoses (p Discussion This study demonstrates that PDC proportions decrease with MDS disease progression and are progressively lower as IPSS-R risk category increases. We also demonstrate that quantification of PDC in MDS can aid in predicting outcomes, although this may be due to a strong association with IPSS-R categories. FC is a useful and clinically feasible tool for quantitating PDCs in bone marrow aspirates, and this measurement is correlated with risk in MDS. Disclosures Arcila: Invivoscribe, Inc.: Consultancy, Honoraria. Roshal:Celgene: Other: Provision of Services; Auron Therapeutics: Equity Ownership, Other: Provision of services; Physicians' Education Resource: Other: Provision of services.

Clonal heterogeneity and evolution of mantle cell lymphoma (MCL) remain unclear despite the progress in our understanding of its biology. Here, we report a 71-yr-old male patient with an aggressive MCL and depict the clonal evolution from initial diagnosis of typical MCL to relapsed blastoid MCL. During the course of the disease, the patient was diagnosed with classic Hodgkin lymphoma (CHL) and received a CHL therapeutic regimen. Molecular analysis by next-generation sequencing of both MCL and CHL demonstrated clonally related CHL with characteristic immunophenotype and PDL1/2 gains. Moreover, our data illustrate the clonal heterogeneity and acquisition of additional genetic aberrations including a rare fusion of SEC22B-NOTCH2 in the process of clonal evolution. Evidence obtained from our comprehensive immunophenotypic and genetic studies indicates that MCL and CHL can originate from a common precursor by divergent clonal evolution, which may pose a therapeutic challenge.

Background Measurable residual disease (MRD) is associated with inferior outcomes in patients with acute myeloid leukemia (AML). MRD monitoring enhances risk stratification and may guide therapeutic intervention. Post-induction MRD is frequently cleared with further therapy and the clearance may lead to better outcomes. In contrast, persistent MRD is associated with poor outcomes. At present it is not possible to predict which patients are likely to clear MRD with further therapy. Here we report a simple, objective, widely applicable and quantitative MFC approach using the ratio of blast/PDC to predict persistent MRD and poor outcomes in AML. Patients and Methods A cohort of 136 adult patients with a confirmed diagnosis of AML by WHO criteria who underwent standard induction therapy at a single center between 4/2014 and 9/2017 was initially included. 69 patients achieved complete morphologic remission (36 MRD-neg. and 33 MRD-pos.). MRD status was assessed by MFC using a different from normal (DfN) approach. PDC were quantified as the percent of total WBC by flow cytometry based on low side scatter, moderate CD45, CD303, bright CD123 and HLA-DR expression. Results The proportion of PDC was markedly decreased in patients with AML (≥20% blasts) (N=136) with a median of 0.016% (interquartile range IQR: 0.0019%-0.071%, Figure 1A), more than 10-fold lower than observed in normal controls (median 0.23%, IQR 0.17%-0.34%) (N=20). While there was no difference between MRD-neg. and normal control groups (median 0.31%, IQR: 0.17%-0.49%; vs. 0.28%, IQR: 0.17%-0.34%), MRD-pos. group had significantly reduced PDC proportion compared to the control (median 0.074%, IQR: 0.022%-0.33%, Wilcoxon rank sum, p=0.019). In an attempt to achieve better separation and to eliminate possible effects of hemodilution, the ratio of blast/PDC was calculated by using the proportions of blasts and PDCs out of total WBCs as quantitated by flow cytometry. A cut-off threshold of the blast/PDC ratio of 10 was chosen to separate each group (Figure 1B). Importantly, a ratio cut-off of 10 had a corresponding specificity of 97.4% for predicting MRD positivity status. MRD positivity was significantly associated with inferior overall survival (OS) and relapse-free survival (RFS) in our study cohort (OS HR 4.11 (95% CI: 1.30-13.03), p=0.016; RFS HR 4.20 (95% CI: 1.49-11.82), p=0.007, Figure 1C and D). The 2-year cumulative incidence of relapse in the MRD-neg. group compared to MRD-pos. group was 10% (95% CI: 2-24%) vs. 37% (95% CI: 18-56%, p=0.014). Importantly, blast/PDC ratio ≥10 was also strongly associated with inferior OS and RFS (OS HR 3.12 (95% CI: 1.13-8.60), p= 0.028; RFS HR 4.05 (95% CI: 1.63-10.11), p=0.003, Figure 1E and F), which is similar in magnitude to MRD positivity. Furthermore, MRD-pos. patients with blast/PDC ratio Conclusion We have established an objective and quantitative MFC method to risk stratify post induction AML patients by risk for relapse, MRD clearance and likelihood of survival. Loss of PDC correlates with residual leukemia, is highly specific for MRD positivity in post-induction patients, and strongly predicts poorer overall survival and higher likelihood of relapse. Loss of PDC also predicts persistent MRD in post-induction MRD-pos. patients despite further therapy, suggesting that MRD-pos. patients with normal PDC may benefit from further therapy prior to transplant, while MRD-pos. patients with loss of PDC may not. Figure 1. Figure 1. Disclosures Goldberg: AROG: Research Funding; Pfizer: Research Funding; Celgene: Consultancy. Geyer:Dava Oncology: Honoraria. Levine:Isoplexis: Equity Ownership; C4 Therapeutics: Equity Ownership; Gilead: Honoraria; Qiagen: Equity Ownership, Membership on an entity's Board of Directors or advisory committees; Prelude: Research Funding; Imago: Equity Ownership; Roche: Consultancy, Research Funding; Loxo: Consultancy, Equity Ownership; Celgene: Consultancy, Research Funding; Novartis: Consultancy; Epizyme: Patents & Royalties; Janssen: Consultancy, Honoraria. Tallman:BioSight: Other: Advisory board; AROG: Research Funding; AbbVie: Research Funding; Cellerant: Research Funding; ADC Therapeutics: Research Funding; Orsenix: Other: Advisory board; Daiichi-Sankyo: Other: Advisory board.

Introduction Plasmacytoid dendritic cells (PDC) are the major natural type I interferon producing dendritic cells that play critical roles in the immune response. PDC are differentiated from hematopoietic stem cells under the control of multiple transcription factors including GATA-2, IKAROS, IRF8, TCF4, ID2 and RUNX1. PDC can be easily identified by flow cytometry due to high-level expression of CD123, HLA-DR and CD303/BDCA2 in the context of lack of lineage markers. In healthy subjects, PDC are Patients and Methods AML patients were searched from the patient database at the MSKCC between 5/2014 and 12/2017. Flow cytometric data obtained on the bone marrow aspirate at the diagnosis were reviewed and cases with PDC proportion greater than 3% of WBC (defined by CD45 positive cells) were included. All the patients had genomic sequencing studies (a targeted NGS panel comprising 30 genes with mutations enriched in AML) performed at diagnosis. PDC and leukemic blasts were sorted by flow cytometry from a subset of patients (N=6) and mutational profiles were compared after sequencing studies. Histopathologic assessment was also performed. Results 24 patients were identified (20 male and 4 female) with a median age of 69 years old (Interquartile range IQR: 56-76). The prevalence was approximately 5% among total new AML patients diagnosed during the same time period. 21 (88%) patients were secondary AML (18 AML with myelodysplasia-related changes (AML-MRC) and 3 therapy-related AML) and only 3 (12%) were de novo AML. No skin presentation was seen. 17 (71%) patients had cytogenetic abnormalities but only 1 with complex karyotype, 1 del(5p) and 3 del(7q). 14 (58%) had pathogenic RUNX1 aberrations (13 mutations and 1 rearrangement, Figure 1A). Other recurrent mutations included ASXL1 (N=7, 29%), IDH1 (N=4, 17%), SRSF2 (N=4, 17%), DNMT3A (N=3, 13%), TET2 (N=3, 13%), NRAS (N=3, 13%), and TP53 (N=3, 13%). The median of PDC proportion was 8.1% (IQR: 4-12.7%). In contrast to BPDCN, these PDC were negative for CD56, TCL-1 and TdT (Figure 1B). The leukemic blasts from 17 (71%) patients showed cross-lineage antigen expression including CD19 (N=4), CD2 and/or CD7 (N=9), and both CD7/CD19 (N=4). The sorted leukemic blasts and PDC shared identical mutations in 6 examined patients confirming the clonal relationship. Unlike CMML with increased PDC, PDC in AML did not form aggregates, showed leukemic distribution pattern and were also increased in peripheral blood at diagnosis. 17 patients received Daunorubicin based induction therapy and 6 had hypomethylating agents. 13 achieved complete remission (5 MRD negative, 6 MRD positive, 2 unknown MRD status). 11 received allo-HSCT with 5 relapsed post transplant. With a median follow-up of 17 months, 18 patients have died. The median survival was 20 months (Figure 1C). Conclusion AML with plasmacytoid dendritic cell differentiation has unique clinicopathologic features including predominantly secondary AML, enriched for RUNX1 mutations, frequent cross-lineage antigen expression and poor prognosis. The PDC are clonally related to the leukemic blasts. CMML with PDC proliferation, AML with PDC differentiation and BPDCN might represent a spectrum of myeloid stem cell neoplasm with aberrant PDC differentiation. Further studies are needed to investigate the molecular mechanism of particularly the role of RUNX1 in PDC differentiation in AML. Figure 1 Figure 1. Disclosures Goldberg: Celgene: Research Funding; Abbvie: Research Funding; AROG: Research Funding; Pfizer: Research Funding. Tallman:ADC Therapeutics: Research Funding; Orsenix: Other: Advisory board; AbbVie: Research Funding; BioSight: Other: Advisory board; Cellerant: Research Funding; Daiichi-Sankyo: Other: Advisory board; AROG: Research Funding. Arcila:Invivoscribe, Inc.: Consultancy, Honoraria.

Introduction Secondary acute myeloid leukemia (s-AML) includes AML with myelodysplasia-related changes (AML-MRC) transformed from an antecedent diagnosis of myelodysplastic syndrome (MDS) or from myeloproliferative and myelodysplastic overlap syndrome (MPN/MDS), as well as therapy-related AML (t-AML) arising in patients with prior exposure to leukemogenic therapies. S-AML has a dismal prognosis. Although the genomic profiling of AML has been well studied, few studies have focused on the molecular aberrations of s-AML. A recent study has demonstrated shared genomic aberrations by AML-MRC and t-AML including mutations in TP53, SRSF2, SF3B1, U2AF1, ZRSR2, ASXL1, EZH2, BCOR, or STAG2. Another recent study has shown that AML patients with mutations in TP53 and chromatin-splicesosome confer poor prognosis and likely represent s-AML. As such, further characterization of the genomic landscape of s-AML is needed. Recurrent somatic PHF6 mutations are rare in de novo AML (~3%), but the frequency and characteristics of PHF6 mutated s-AML are largely unknown. Patients and Methods AML-MRC and t-AML patients were searched from the patient database at the MSKCC between 1/2014 and 12/2016. All the patients were reviewed and the diagnosis was confirmed. Only patients with genomic sequencing studies (a targeted NGS panel comprising 30 genes with mutations enriched in AML) performed at diagnosis were included. Results 152 patients were identified. There were 7 t-AML patients with favorable cytogenetic abnormalities that likely represent a distinct biological subgroup; therefore these cases were excluded from further analysis. AML-MRC (N=75) and t-AML (N=70) patients had similar clinical characteristics including ages, gender distribution, cytogenetic risk, treatment and survival (median survival: 8 months in t-AML and 10 months in AML-MRC, p=0.4); The mutational profiles of AML-MRC and t-AML cohorts were also comparable with recurrent TP53, PHF6, RAS, and RUNX1 mutations, albeit with different frequencies (Figure 1; Red represents t-AML and blue AML-MRC). Therefore we combined AML-MRC and t-AML cohorts for further analysis. 43 (29.7%) patients had TP53 mutations, 17 (11.7%) patients had PHF6 mutations, and 85 (58.6%) patients had neither mutation (non-TP53/non-PHF6). TP53 and PHF6 mutations were mutually exclusive. TP53 mutations were nearly exclusively associated with adverse cytogenetic abnormalities including alterations involving chromosomes (chr) 5p and/or 7q (41/43, 95.3%) with the majority being complex karyotype (30/43, 70%). By contrast, only 18% PHF6 mutated s-AML patients had chr 5p/7q abnormalities and none of them had complex karyotype. Among the patients with non-TP53/non-PHF6 mutations, 30% had chr 5p/7q abnormalities and 6% had complex karyotype. Although secondary AML has very poor prognosis, PHF6 mutated patients had better overall survival than the other two groups (Log rank test, p=0.0095; median survival: PHF6 vs TP53 vs. non-TP53/non-PHF6: 24 vs. 7 vs. 9 months). Immunophenotypically, the blasts from PHF6 mutated patients showed a high frequency of loss of CD38 (44% vs 17%, p=0.04) and CD33 expression (69% vs 28%, p=0.004) compared to PHF6 unmutated cases, indicative of a less mature, stem-cell derived phenotype Conclusion PHF6 mutations define a unique subset of s-AML that is associated with a more primitive stem/progenitor immunophenotype, absent complex karyotype and relatively better outcomes. PHF6 mutations are mutually exclusive to TP53 mutations. Studies are needed to further define the genomic classification of s-AML patients in an unbiased way and to elucidate the role of PHF6 mutations in s-AML. Figure 1 Figure 1. Disclosures Goldberg: AROG: Research Funding; Abbvie: Research Funding; Pfizer: Research Funding; Celgene: Research Funding. Tallman:Orsenix: Other: Advisory board; AbbVie: Research Funding; Cellerant: Research Funding; BioSight: Other: Advisory board; AROG: Research Funding; Daiichi-Sankyo: Other: Advisory board; ADC Therapeutics: Research Funding. Arcila:Invivoscribe, Inc.: Consultancy, Honoraria. Levine:Loxo: Consultancy, Equity Ownership; Gilead: Honoraria; Roche: Consultancy, Research Funding; Epizyme: Patents & Royalties; Imago: Equity Ownership; Qiagen: Equity Ownership, Membership on an entity's Board of Directors or advisory committees; Celgene: Consultancy, Research Funding; C4 Therapeutics: Equity Ownership; Novartis: Consultancy; Janssen: Consultancy, Honoraria; Isoplexis: Equity Ownership; Prelude: Research Funding.