The Long Noncoding RNA BALR2 Controls Novel Transcriptional Circuits Involved in Chemotherapy Sensitivity of Pediatric Acute Myeloid Leukemia (AML) Blasts

In acute myeloid leukemia (AML), the assessment of post-induction minimal residual disease (MRD) is largely utilized for choosing post-remission therapies aimed at maintaining complete remission (CR) and preventing relapse. This latter is still the major cause of treatment failure in pediatric AML, and even if several efforts have been spent to validate MRD as a prognostic marker, numerous studies demonstrated that MRD negativity cannot be considered a completely reliable surrogate biomarker predicting outcome, since it does not exclude a relapse. The current interpretation is that disease relapse is due to mechanisms leading to therapy resistance mainly depending on driver chimeric or oncogenic protein-coding genes, which are monitored during treatment, and does not consider that chemotherapy resistance may arise from other genetic markers, phenomenon linked to methylation and non-coding RNAs genomic pressure. We, thus, hypothesized that other markers need to be explored to re-interpret leukemia progression. We showed an overall hyper-expression of the lncRNA BALR2 in 132 de novo AML bone marrow samples collected at diagnosis and analyzed the gene expression profile (GEP) of 58 cases. By unsupervised clustering analysis, we produced important advances in identifying BALR2 as a robust novel molecular marker of a new subgroup of AML characterized by a high rate of resistance to induction therapy, independently from the genetic lesions detected at diagnosis and any other prognostic clinical and genetic features. We demonstrated in vitro that BALR2 has a direct role in controlling bi-directionally its own and of its neighbor gene CDK6 promoter activity. This latter finding of high CDK6 expression was shown to sustain its complex with RUNX1 in order to inhibit RUNX1 binding to its target promoters, thus preventing the process of hematopoietic differentiation progression. To support BALR2 as a new proto-oncogene involved in the control of the myeloid differentiation program, we ranked the genes across the expression profile obtaining a signature of 337 transcripts able to cluster CD34+ human stem cell precursors (HSCPs) separately from more mature CD14+ cells. These in silico findings were validated in vitro by showing that, after BALR2 depletion, CD34+ cells had a skewed myeloid differentiation. Furthermore, we found that AML differentiation toward mature myeloid cells with increased phagocytic capacity was obtained through BALR2 level reduction, and enhanced by combinatorial differentiation stimuli. Our findings attribute a distinct role to BALR2 in the block of myeloid stem cell differentiation occurring during leukemogenesis. At the same time, we interrogated GEP ontology, finding that enrichments of genes involved in mitochondrial synthesis pathways were significantly correlated to patients with highest BALR2 levels, and confirmed the same mitochondriogenesis profile in the immature CD34+ HSCPs. We moved to deconvolute this feature and demonstrated that BALR2, by controlling mitochondria gene balance, was directly controlling the mitochondrial mass, which dramatically decreased after BALR2 silencing, this supporting the hypothesis that BALR2 would maintain mitochondrial functions to confer AML resistance to cytotoxicity. Consistently with this line of reasoning, we inhibited mitochondria by tigecycline, demonstrating that its activity was dramatically strengthened in BALR2 depleted cells, when used either alone or in combination with cytosine-arabinoside (Ara-C). Concomitantly, tigecycline treatment in BALR2 silenced AML cells reduced mitochondria depolarization, and increased the number of differentiated M-CFU colonies formation, confirming that BALR2, together with CDK6, forms novel transcriptional networks to create a circuit able to impair myeloid differentiation and to lower chemo-sensitivity in AML. We speculate that a novel therapeutic window of mitochondrial targeting in defined AML subgroups, identified through assessment of BALR2 levels at diagnosis or persistent MRD levels, could be envisaged to optimize the outcome of childhood AML. Disclosures Locatelli: Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees; bluebird bio: Consultancy; Amgen: Honoraria, Membership on an entity's Board of Directors or advisory committees; Bellicum: Consultancy, Membership on an entity's Board of Directors or advisory committees; Miltenyi: Honoraria.

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Expression Of Aldehyde Dehydrogenase Reflected Diverse Stem Cell Properties In Acute Myeloid Leukemia

Blood ◽

10.1182/blood.v122.21.1344.1344 ◽

2013 ◽

Vol 122 (21) ◽

pp. 1344-1344

Author(s):

Van T. Hoang ◽

Eike C. Buss ◽

Isabel Hoffmann ◽

Abraham Zepeda-Moreno ◽

Natalia Baran ◽

...

Keyword(s):

Stem Cells ◽

Acute Myeloid Leukemia ◽

Clinical Study ◽

Board Of Directors ◽

Myeloid Leukemia ◽

Aldh Activity ◽

Advisory Committees ◽

Low Frequencies ◽

Acute Myeloid

Abstract Separation of leukemic stem cells (LSC) and residual hematopoietic stem cells (HSC) from the same individual patient with acute myeloid leukemia (AML) is essential for a proper understanding of the leukemic driving mechanisms. We have studied the role of aldehyde dehydrogenase (ALDH) for this purpose and have defined the functional properties of ALDHbright cells in specific subgroups of AML. We have examined the ALDH activity by flow cytometry in bone marrow samples (BM) from 14 healthy donors and 73 patients with de novo AML. The median frequency of cells with high ALDH activity (ALDHbright cells) in the healthy subjects was 1.92% with a range from 0.58 to 3.16%. For patients with AML, the median number of ALDHbright cells was 0.25% with a broad range from 0.004 to 33.57%. Whereas the majority of patients with AML (n = 56) had low frequencies of ALDHbright cells (median 0.11%; range 0.004 – 1.77%; defined as ALDH-low AML), 17 patients had relatively numerous ALDHbright cells (median 9.01; range 3.54 – 33.57%; defined as ALDH-numerous AML). In both groups, ALDHbright cell populations were highly enriched for CD34+CD38- cells. The ALDHbright cells derived from ALDH-low AML did not contain chromosomal and molecular aberrations characteristic of the original leukemia, and were able to induce multi-lineage hematopoiesis in NSG mouse models. Thus, genetically and functionally normal HSC could be successfully isolated in the ALDHbright subset, whereas LSC were enriched in ALDHdimCD34+CD38- subset for patients with ALDH-low AML. For 17 patients with ALDH-numerous AML, the ALDHbright subset was consistently contaminated with LSC. In clinical follow-ups, patients with ALDH-numerous AML showed resistance to induction chemotherapy and were characterized by a very poor long-term outcome that was comparable to patients with high-risk cytogenetic or molecular genetic markers. In four patients with ALDH-numerous AML we demonstrated that the ALDHbrightCD34+CD38- subset contained chemotherapy-resistant clones with repopulating ability. Furthermore, such ALDHbright cells were characterized by a lower cell-cycle activity and an increased resistance to cytarabine in comparison with ALDHdim blasts in in vitro assays. Our data have provided evidence that LSC and residual HSC can be separated using ALDH in patients with low frequencies of ALDHbright cells. In patients with ALDH-numerous AML, the ALDHbright subset is associated with leukemic features both in vitro and in animal models. Thus our data demonstrated the feasibility of appropriate comparisons of LSC versus HSC from the same patient with specific subtypes of AML and the impact of LSC properties on clinical outcome. Disclosures: Buss: Novartis: Travel support Other; Micromet/Amgen: Reimbursements for participation in a clinical study , Reimbursements for participation in a clinical study Other. Ho:Sanofi-Aventis: Consultancy, Honoraria, Membership on an entity’s Board of Directors or advisory committees; Genzyme: Consultancy, Honoraria, Membership on an entity’s Board of Directors or advisory committees.

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Survey of Serum Aspergillus Antigen in Patients Undergoing Chemotherapy for Acute Myeloid Leukemia or Allogeneic Hematopoietic Stem Cell Transplantation: Role in Predicting Invasive Aspergillosis and in Modifying the Therapeutic Strategy

Blood ◽

10.1182/blood.v126.23.3428.3428 ◽

2015 ◽

Vol 126 (23) ◽

pp. 3428-3428

Author(s):

Van Hung Tran ◽

Florence Persat ◽

Sophie Gardes ◽

Jeremy Monfray ◽

Sophie Ducastelle-Leprêtre ◽

...

Keyword(s):

Acute Myeloid Leukemia ◽

Stem Cell ◽

Board Of Directors ◽

Induction Chemotherapy ◽

Myeloid Leukemia ◽

Advisory Committees ◽

Hematopoietic Stem ◽

Aspergillus Antigen ◽

Acute Myeloid

Abstract Introduction Invasive aspergillosis (IA) remains an important cause of mortality in immunocompromised acute myeloid leukemia (AML) patients receiving induction chemotherapy and in patients undergoing allogeneic hematopoietic stem cell transplantation (allo-HSCT) for hematological malignancies. Early diagnostic is critical and challenging given the efficacy and availability of several new anti-fungal therapies. In this study, we evaluated the performance of different factors in predicting the occurrence of IA, including the Aspergillus antigen galactomannan (GM) detection in sera. Methods We included all AML patients receiving induction chemotherapy and patients undergoing allo-HSCT for any hematological malignancy at our center between April 2006 and April 2014 with available data on Aspergillus antigen GM. Serologic detection of circulating GM fungal biomarker was considered during the 100 days following the first day of induction chemotherapy in AML patients or from the day of allo-HSCT. Usual follow-up included two GM tests per week, only patients with at least three serum GM results were considered. The GM tests have been performed routinely using the ELISA kit (Platelia Aspergillus antigen ELISA, Biorad), giving the results in index values. Demographic, GM index results and diagnostic data were collected. IA cases were classified as proven or probable according to the EORTC criteria. The value of the first antigen test, the delay to positivity, and the slope of the progression of the index value between the first two antigens concentrations were considered as predictors of IA. ROC curves for each predictor and their combination were performed and prognostic scores were established. Results A total of 775 patients were included : i) 292 AML patients, 153 (52%) males with a median age of 62 years (range: 17-79), 15% were classified as favorable, 8% as intermediate I, 18% as intermediate II and 59% as unfavorable according to cytogenetics and molecular markers; ii) 483 allo-HSCT patients, 293 (61%) were males, median age was 48 years (range: 18-70), among them 234 (48%) AML, 66 (14%) multiple myeloma, 46 (10%) Myelodysplastic syndromes, 38 (8%) Non-Hodgkin Lymphoma and the rest of patients had other hematological disorders; 233 (48 %) patients received reduced intensity conditioning and 250 (52%) myeloablative conditioning. The disease status at allo-HSCT was complete remission (CR) in 366 (76%) patients and the rest of patients were in less than CR. HSC source was peripheral blood in 42.2% (90 identical siblings, 150 10/10 matched unrelated, 54 9/10 mismatched unrelated), bone marrow in 42.6% (105 identical siblings, 162 10/10 matched unrelated, 45 9/10 mismatched unrelated) and cord blood in 15.2%. A total of 877 episodes with 16121 GM serum antigen results was considered (median: 18 GM tests per patient). During the follow-up, we identified 121 episodes with at least one positive GM test with a cumulative incidence at day 100 of 13.8%. We also diagnosed 48 IA (2 proven, 46 probable), with a cumulative incidence at day 100 of 5.5% in total, 7.2% in AML and 4.3% in allo-HSCT, respectively. We then classified the GM positive episodes in 82 false-positive (68%) and 39 true-positive episodes (32%) for IA, respectively. A majority of IA events occurred during the first 30 days of follow up, GM positivity showing a positive predictive value of 41% versus a negative predictive value of 99%. The three IA predicting factors had similar independent effects and their combinations were performed, allowing the establishment of an area under ROC of 0.79 (95% CI: 0.70-0.89). Cut off values of the first positive GM serum and slope were equal or higher than 1.04 and 0.04, respectively, and delay to positivity equal or less than 15 days. To simplify the practical use in clinical practice, the prognostic score defining the IA risk probability was defined as the number of predictors present (values from 0 to 3). This score was tested on positive follow-up giving values of 0, 1, 2 or higher for 45 (37%), 39 (32%) and 37 episodes (31%), respectively. A score superior or equal to 2 was indicative of IA in 62% of the cases (figure 1). Conclusion As IA has a significant impact on hematology patient's survival, this GM predictive score combining three predictors (value of the first antigen index, delay of positivity and slope of the index values) may help clinicians to conclude about starting an early preemptive IA treatment. Figure 1. Figure 1. Disclosures Nicolini: Ariad Pharmaceuticals: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Bristol-Myers Squibb: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Novartis: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau.

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Fc Receptor-Dependent Trogocytosis of CD39 Impacts Engraftment and Invasiveness of Acute Myeloid Leukemia Cells

Blood ◽

10.1182/blood-2021-151021 ◽

2021 ◽

Vol 138 (Supplement 1) ◽

pp. 3298-3298

Author(s):

Lili Feng ◽

Haohai Zhang ◽

Paola de Andrade Mello ◽

Dina Stroopinsky ◽

Wenda Gao ◽

...

Keyword(s):

Acute Myeloid Leukemia ◽

Board Of Directors ◽

Myeloid Leukemia ◽

Research Funding ◽

Bioluminescence Imaging ◽

Lentiviral Vector ◽

Advisory Committees ◽

Acute Myeloid

Abstract Corresponding author: Dr. Simon. C. Robson ([email protected]). Introduction: CD39/ENTPD1 (ectonucleoside triphosphate diphosphohydrolase-1) is the prototypic member of the GDA1-CD39 superfamily of ectonucleotidases and modulates purinergic signaling pathways. CD39 expression has been noted in human acute myeloid leukemia (AML) and likely contributes to chemoresistance [1]. Our study reported here elucidates the impact of Cd39 on engraftment and invasiveness of AML TIB-49 cells using an immunocompetent murine experimental model. Methods: Wild-type (WT) mice and Cd39 -/- mice on C57BL/6 background were bred at Beth Israel Deaconess Medical Center. The syngeneic murine AML cell line TIB-49 (Cd39 negative in vitro) was purchased from American Type Culture Collection. For bioluminescence imaging experiments, TIB-49 cells were transduced with luciferase/mCherry using a lentiviral vector. For AML model, mice were administered with 1×10 6 TIB-49-luciferase cells intravenously via tail vein injection. For chloroma model, mice were subcutaneously inoculated with 1×10 6 TIB-49 cells in the right flank. Bioluminescence imaging of TIB-49-luciferase bearing mice was conducted with the IVIS TM 50 Imaging System. Blood, spleen and bone marrow (BM) were also collected from TIB-49 bearing AML mice for FACS (fluorescence activated cell sorting) analysis. To explore Cd39 in TIB engraftment and invasiveness, TIB-49 cells were further transduced with a lentiviral vector overexpressing mCd39 with TdTomato. WT mice were intravenously inoculated with 1×10 6 of either TIB-49-TdTomato cells or TIB-49-mCd39-TdTomato cells, and the above read-outs were determined. To investigate the potential of CD39 as a therapeutic target, we engineered anti-mouse Cd39 antibodies (αCd39 mAb) with isotype selection and removal of fucose to further promote Fc receptor (FcR) interactions. Results: Bioluminescence imaging results indicated that TIB-49 engraftment was decreased in global Cd39 -/- mice with decreased disease burdens noted relative to WT (Figure 1A). FACS analysis of blood, spleen and BM-derived cells from TIB-49 bearing AML-model mice (day 31) confirmed higher engraftment of TIB-49 cells (TdTomato+) at all sites in WT compared to Cd39 -/- mice (Figure 1B). TIB-49 cells did not express Cd39 in vitro, but TIB-49 cells harvested from spleen and BM of WT but not Cd39 -/- mice displayed high levels of Cd39. This indicated TIB-49 cells acquired Cd39 from host cells, in a process of antibody-independent trogocytosis (Figure 1C), as RT-PCR did not detect Cd39 mRNA expression in TIB-49 cells in vivo. Additionally, circulating TIB-49 cells from the blood of WT mice were Cd39 negative (Figure 1C), suggesting a role for the tumor microenvironment in mediating trogocytosis. TIB-49 cells expressing host Cd39 in WT mice spleen and BM lost Cd39 after being exposed to αCd39 mAb treatment. Cd39 translocated from TIB-49 cells to effector cells, at least in part, dependent on FcR mediated trogocytosis (Figure 1D). When Cd39 was overexpressed on TIB-49 cells (TIB-49-mCd39-TdTomato), the engraftment was boosted in WT mice in vivo when compared to TIB-49-TdTomato cells (day 19, Figure 1E) with higher levels of Cd39 expression than that observed on TIB-49-TdTomato cells in spleen and BM (day 26) (Figure 1F). Moreover, TIB-49-mCd39-TdTomato bearing mice displayed shorter survival times, when compared with TIB-49-TdTomato bearing AML mice (Figure 1G). The αCd39 mAb monotherapy had no effect on TIB-49 chloroma model growth. However, pretreatment with αCd39 mAb effectively boosted daunorubicin chemotherapeutic effects in vivo (Figure 1H and 1I). Conclusions: Our study suggests bidirectional trogocytosis between TIB-49 AML and host immune cells, which is further modulated by FcR interaction. Re-distribution of Cd39 from host to TIB-49 cells or induced high level expression contributes to engraftment and invasiveness, resulting in decreased survival. Targeting CD39 is a potential therapeutic approach, operational not only by boosting chemosensitivity but furthering anti-leukemic effects in experimental models. Disclosures: No relevant conflicts of interest to declare. References: [1] Nesrine Aroua, Emeline Boet, Margherita Ghisi, et al. Extracellular ATP and CD39 Activate cAMP-Mediated Mitochondrial Stress Response to Promote Cytarabine Resistance in Acute Myeloid Leukemia. Cancer Discov. 2020. Figure 1 Figure 1. Disclosures Stroopinsky: The Blackstone Group: Consultancy. Avigan: Celgene: Membership on an entity's Board of Directors or advisory committees, Research Funding; Pharmacyclics: Research Funding; Kite Pharma: Consultancy, Research Funding; Juno: Membership on an entity's Board of Directors or advisory committees; Partner Tx: Membership on an entity's Board of Directors or advisory committees; Karyopharm: Membership on an entity's Board of Directors or advisory committees; Bristol-Myers Squibb: Membership on an entity's Board of Directors or advisory committees; Aviv MedTech Ltd: Membership on an entity's Board of Directors or advisory committees; Takeda: Membership on an entity's Board of Directors or advisory committees; Legend Biotech: Membership on an entity's Board of Directors or advisory committees; Chugai: Membership on an entity's Board of Directors or advisory committees; Janssen: Consultancy; Parexcel: Consultancy; Takeda: Consultancy; Sanofi: Consultancy.

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Dasatinib Inhibits FLT3/ITD and PTPN11mutated Acute Myeloid Leukemia Cells Overexpressing SRC Tyrosine Kinases

Blood ◽

10.1182/blood-2019-128016 ◽

2019 ◽

Vol 134 (Supplement_1) ◽

pp. 1451-1451

Author(s):

Sigal Tavor ◽

Tali Shalit ◽

Noa Chapal Ilani ◽

Yoni Moskovitz ◽

Nir Livnat ◽

...

Keyword(s):

Gene Expression ◽

Acute Myeloid Leukemia ◽

Board Of Directors ◽

Myeloid Leukemia ◽

Drug Sensitivity ◽

Kinase Inhibitors ◽

Advisory Committees ◽

Acute Myeloid

Background: Recent advances in acute myeloid leukemia(AML) targeted therapy improve overall survival. While these targeted therapies can achieve prolonged remissions, most patients will eventually relapseunder therapy. Our recent studies suggest that relapse most often originates from several sub-clones of leukemic stem cells (LSCs), present before therapy initiation, and selected due to several resistance mechanisms. Eradication of these LSCs during treatment induction /remission could thus potentially prevent relapse. The overall goal of the current study was to identify drugs which can be safely administrated to patients at diagnosis and that will target LSCs. Since simultaneously testing multiple drugs in vivo is not feasible, we used an in vitrohigh throughput drug sensitivity assay to identify new targets in primary AML samples. Methods: Drug sensitivity and resistance testing (DSRT) was assessed in vitro (N=46 compounds) on primary AML samples from patients in complete remission (N=29). We performed whole exome sequencing and RNAseq on samples to identify correlations between molecular attributes and in vitro DSRT. Results:Unsupervised hierarchical clustering analysis of in vitro DSRT, measured by IC50, identified a subgroup of primary AML samples sensitive to various tyrosine kinase inhibitors (TKIs). In this subgroup, 52% (9/17) of AML samples displayed sensitivity to dasatinib (defined as a 10-fold decrease in IC50 compared to resistant samples). Dasatinib has broad TKI activity, and is safely administered in the treatment of leukemia. We therefore focused our analysis on predicting AML response to dasatinib, validating our results on the Beat AML cohort. Enrichment analysis of mutational variants in dasatinib-sensitive and resistant primary AML samples identified enrichment of FLT3/ITD (p=0.05) and PTPN11(p=0.05) mutations among dasatinib responders. Samples resistant to dasatinib were enriched with TP53 mutations (p=0.01). No global gene expression changes were observed between dasatinib-sensitive and resistant samples in our cohort, nor in the Beat AML cohort. Following this, we tested the differential expression of specific dasatinib-targeted genes between dasatinib-responding and resistant samples. No significant differences were identified. However, unsupervised hierarchical clustering of dasatinib targeted genes expression in our study and in the Beat AML cohort identified a subgroup of AML samples (enriched in dasatinib responders) that demonstrated overexpression of three SRC family tyrosine kinases:FGR, HCK and LYN as well as PTK6, CSK, GAK and EPHB2. Analysis of the PTPN11 mutant samples revealed that the IC50 for dasatinib in 23 carriers of the mutant PTPN11 was significantly lower compared to the IC50 of PTPN11 wild type samples (p=0.005). LYN was also upregulated (p<0.001) in the mutant samples. We therefore hypothesized that gene expression of dasatinib-targeted genes could be used as a predictive biomarker of dasatinib response among FLT3/ITD carriers. We found that among FLT3/ITD AML carriers in the Beat AML cohort LYN, HCK, CSK and EPHB2 were significantly over-expressed in the dasatinib responding samples (N=27) as compared to the dasatinib resistant samples (N=35). To predict response to dasatinib among FLT3/ITD carriers we used a decision tree classifier based on the expression levels of these four genes. Our prediction model yielded a sensitivity of 74% and specificity of 83% for differentiating dasatinib responders from non-responders with an AUC of 0.84. Based on our findings, we selected FLT3/ITD AML samples and injected them to NSG-SGM3 mice. We found that in a subset of these samples, dasatinib significantly inhibited LSCs engraftment. This subset of FLT3/ITD AML samples expressed higher levels of LYN, HCK,FGR and SRC as compared to the FLT3/ITD samples that were not sensitive to dasatinib therapy in vivo. In summary, we identified a subgroup of AML patients sensitive to dasatinib, based on mutational and expression profiles. Dasatinib has anti-leukemic effects on both blasts and LSCs. Further clinical studies are needed to demonstrate whether selection of tyrosine kinase inhibitors, based on specific biomarkers, could indeed prevent relapse. Disclosures Tavor: Novartis: Membership on an entity's Board of Directors or advisory committees; Abbvie: Membership on an entity's Board of Directors or advisory committees; Astellas: Membership on an entity's Board of Directors or advisory committees; Pfizer: Membership on an entity's Board of Directors or advisory committees; BMS companies: Membership on an entity's Board of Directors or advisory committees.

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A Novel Proteomic Profiling of the Bone Marrow Microenvironment Reveals Elevated Levels of the Chemokine CCL23 Isoforms in Acute Myeloid Leukemia

Blood ◽

10.1182/blood-2019-122121 ◽

2019 ◽

Vol 134 (Supplement_1) ◽

pp. 2709-2709

Author(s):

Haydar Celik ◽

Katherine E. Lindblad ◽

Bogdan Popescu ◽

Giovanna Fantoni ◽

Gege Gui ◽

...

Keyword(s):

Bone Marrow ◽

Acute Myeloid Leukemia ◽

Board Of Directors ◽

Myeloid Leukemia ◽

Research Funding ◽

Serum Samples ◽

Advisory Committees ◽

Extracellular Compartment ◽

Acute Myeloid

The bone marrow (BM) microenvironment is increasingly recognized as an important contributor to acute myeloid leukemia (AML) pathogenesis. However, despite growing interest in characterizing different components and cellular architecture of the BM niche and their biological significance in leukemogenesis, the proteomic constitution of the BM extracellular compartment that distinguishes a leukemic niche from its normal counterpart has not yet been fully described. We therefore performed a quantitative, large-scale proteomic analysis of 1,305 human proteins of the non-cellular compartment of BM (plasma) samples from ten relapsed or refractory AML patients and from ten age- and sex-matched healthy donors (HDs) using an aptamer-based, highly multiplexed, affinity proteomics platform (SOMAscan). This screen identified a total of 168 differentially abundant proteins, of which 91 were significantly more and 77 proteins significantly less abundant in leukemic BM compared with healthy marrow (FC ≥ 1.5, FDR ≤ 0.05). Comparative analysis of BM plasma and peripheral blood (PB) serum samples from the same AML patients and HDs revealed 65 similarly regulated proteins (37 up-regulated vs. 28 down-regulated) and 1 differently regulated protein between the two compartments. Out of the total 168 proteins, 102 proteins were specifically dysregulated only in the BM compartment. TruSeq Stranded Total RNA-sequencing (Illumina) was also performed using paired-end 75bp sequencing on a HiSeq 3000. RNA was isolated from PAXgene BM RNA tubes (Qiagen) collected in parallel with samples for proteomic analysis. Results of analysis of differentially expressed transcripts only partially overlapped with those candidates identified from our validated proteomic approach, indicating that sequencing of RNA derived from cellular sources of BM may be a suboptimal screening strategy to determine the true proteomic composition of the extracellular compartment of the AML marrow microenvironment. In addition to several previously reported proteins, our proteomics screen discovered numerous aberrantly expressed proteins in leukemic marrow whose role in AML pathogenesis is currently unknown. Using pathway analysis, we identified sets of proteins enriched for specific biological pathways including RAS, ephrin, PDGF, PI3K/AKT, MAPK, Notch, TLR, JAK-STAT, NFκB, Rap1, and Tie2 signaling pathways. A systems biology analysis approach revealed the highly connected network of cytokines and chemokines as the most striking AML-associated proteomic alteration in the BM. We identified IL-8 as a differentially expressed and key central molecule of this network in AML, consistent with recent reports. Importantly, we also identified significantly elevated levels of CKβ8 and CKβ8-1, alternatively spliced isoforms of the myelosuppressive chemokine CCL23 also known as myeloid progenitor inhibitory factor 1 (MPIF-1) or CKβ8, in both leukemic marrow and PB serum samples (Figure 1). Given the critical importance of cytopenias, often disproportional to the degree of leukemic marrow involvement, in the morbidity and mortality of patients with myelodysplastic syndrome (MDS) and AML, we subsequently confirmed this striking finding by performing orthogonal validation in a larger cohort of MDS and AML patients using an ELISA-based immunoassay. This novel finding suggests the possibility that CCL23 may play a role in suppression of normal hematopoiesis in MDS and AML. In support of this hypothesis, we demonstrated in vitro myelosuppressive effects of CCL23 isoforms on colony formation by human CD34+ hematopoietic stem and progenitor cells (HSPCs) in an in vitro colony forming unit assay, resulting in an approximately 2.5-fold decrease in CFU-GM and an evident decrease in CFU-GEMM counts. In summary, our broad and quantitative proteomic dataset of extracellular factors present in leukemic and normal aging bone marrow has already provided novel mechanistic insights into AML pathogenesis and should serve, together with paired RNA-sequencing information, as a useful public resource for the research community. Disclosures Lai: Jazz Pharma: Membership on an entity's Board of Directors or advisory committees; Jazz Pharma: Speakers Bureau; Astellas: Speakers Bureau; Daiichi-Sankyo: Membership on an entity's Board of Directors or advisory committees; Agios: Membership on an entity's Board of Directors or advisory committees. Hourigan:SELLAS Life Sciences Group AG: Research Funding; Merck, Sharpe & Dohme: Research Funding.

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GDF1 Is a Regulator of Sphingolipid Metabolism in Acute Myeloid Leukemia

Blood ◽

10.1182/blood-2019-130964 ◽

2019 ◽

Vol 134 (Supplement_1) ◽

pp. 1245-1245

Author(s):

Weiyuan Wang ◽

Emily Sullivan ◽

Vasiliki Papakotsi ◽

Andrea Cote ◽

Paul Toran ◽

...

Keyword(s):

Stem Cell ◽

Board Of Directors ◽

New Hampshire ◽

Myeloid Leukemia ◽

Stem Cell Differentiation ◽

Sphingolipid Metabolism ◽

Advisory Committees ◽

University Of New Hampshire ◽

Real Time Qpcr ◽

Acute Myeloid

BACKGROUND: Mutations to epigenetic and spliceosome regulators frequently occur in acute myeloid leukemia (AML) and contribute to the underlying pathobiology of the disease. In addition, recent studies have sought to improve the anti-AML efficacy of sphingolipid-based therapeutics by identifying the underlying mechanisms responsible for dysfunctional sphingolipid metabolism. Sphingolipids are an extensive classification of lipids that play profound roles in membrane structure as well as regulation of cellular function and fate. Ceramide, as the hydrophobic moiety of sphingolipids, serves as the hypothetical center of sphingolipid metabolism. It regulates cellular stress responses and apoptosis, whereas many of its metabolites regulate opposing processes such as proliferation and survival. Growth/differentiation factor 1 (GDF1) is a TGF-beta superfamily member, of which other members have been attributed roles in stem cell biology and hematopoiesis. Mutations in GDF1 have been found to be associated with congenital cardiovascular malformations, yet little is known about the role of GDF1 outside of cardiac development. GDF1 is encoded from a bicistronic gene that also encodes for ceramide synthase 1 (CERS1), which is responsible for generating the C18:0 species of ceramide. Recently, an anti-AML role was attributed to CERS1 in FLT3-ITD-mutated AML (Dany et. al. Blood 2016). However, a potential role for GDF1 was not evaluated in this study. Intriguingly, in preliminary work using FLT3ITD transgenic mice we observed an inverse correlation between the expression of GDF1 and UGCG which encodes for the ceramide detoxifying enzyme glucosylceramide synthase. Therefore, our present study tested the hypothesis that GDF1, encoded from the bicistronic CERS1-GDF1 gene, exerts anti-AML efficacy by downregulating ceramide neutralization and promoting stem cell differentiation METHODS & RESULTS: Studies were carried out using AML cell lines as well as primary cells harvested from transgenic murine models of myelodysplastic syndrome (MDS) and AML (FLT3ITD, Nup98-HoxD13, Srsf2P95H-mutant, Tet2-deficient, Asxl1-deficient, and Tert-deficient). The expression of GDF1 was evaluated by real time qPCR in hematopoietic cells isolated from these models of MDS and AML. GDF1 expression was greatest in Srsf2P95H-mutant (MDS) samples, which corresponded to enhanced sensitivity to ceramide-based therapies. We had recently reported this MDS-specific sensitivity for nanoliposomal ceramide (Barth et. al. Blood Advances 2019), which is a ceramide-based therapy currently in a clinical trial for solid tumor malignancies (ClinicalTrials.gov identifier: NCT02834611). Next, cell lines and models were treated with recombinant GDF1. Real time qPCR revealed that GDF1 treatment downregulated the expression of the genes in the ceramide neutralization pathway including UGCG. In addition, flow cytometry was used to show that GDF1 treatment promoted hematopoietic stem cell differentiation. Lastly, C57BL/6J mice engrafted with C1498 AML cells was used to show that GDF1 enhanced the therapeutic efficacy of cytarabine and nanoliposomal ceramide. CONCLUSIONS: Overall, this study provided evidence for differential expression of GDF1 in subtypes of MDS and AML and showed that GDF1 can regulate sphingolipid metabolism by downregulating ceramide neutralization. Importantly, we have demonstrated that GDF1 exerts anti-AML efficacy in combination with either standard care therapy or ceramide-based therapy. Therefore, GDF1-elevating strategies are well-positioned as novel therapeutic approaches for the treatment of AML and related myeloid hematological disorders. This work was funded by NIH/NCI K22 CA190674 (B.M.B.) and University of New Hampshire COBRE Pilot Project Grant NIH/NIGMS P20 GM113131 (B.M.B.). The authors acknowledge US Provisional Patent 62/602,437, issued to B.M.B. and the University of New Hampshire. Disclosures Loughran: Bioniz: Membership on an entity's Board of Directors or advisory committees; Keystone Nano: Membership on an entity's Board of Directors or advisory committees. Barth:University of New Hampshire: Patents & Royalties: US Provisional Patent 62/602,437; NIH (NCI and NIGMS): Research Funding.

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Decitabine Super-Responders: Challenging the Dogma of Long-Term Remission for Acute Myeloid Leukemia

Blood ◽

10.1182/blood-2018-99-119916 ◽

2018 ◽

Vol 132 (Supplement 1) ◽

pp. 5190-5190 ◽

Cited By ~ 1

Author(s):

Joseph Maakaron ◽

Gabriel N. Mannis ◽

Alison R. Walker ◽

Alice S. Mims

Keyword(s):

Acute Myeloid Leukemia ◽

Stem Cell ◽

Board Of Directors ◽

Myeloid Leukemia ◽

Cell Transplant ◽

Advisory Committees ◽

Term Survival ◽

Aggressive Approach ◽

Acute Myeloid

Abstract Introduction Decitabine and hypomethylating agents are considered non-curative treatments for patients with acute myeloid leukemia (AML) and are a standard approach for those patients not fit for intensive induction chemotherapy, usually due to age and multiple comorbidities. A phase 2 study reported a better overall response rate of 64% with a 10-day schedule of decitabine (Blum, Garzon et al. 2010). A second study reported on a higher rate of response among patients with TP53 mutations (Welch, Petti et al. 2016). However, these responses were not considered durable with median duration of response of about 11 months. We herein describe four patients with longer than expected response duration to decitabine, two of which were able to discontinue therapy with sustained multi-year remission and an apparent cure of their AML. Methods We sought to evaluate the clinical and pathological characteristics of patients with a particular long-term response to decitabine. We screened patients at two institutions who have been in complete remission (CR) for greater than 2 years with single-agent decitabine therapy. Current efforts are on-going to identify patients from five other institutions that also meet these criteria and will be added for the final presentation of this data. Results Four patients have been identified who have experienced CR for over 2 years with average age at diagnosis of 70 years (range 60-76). Three patients had cytogenetically normal (CN) AML and one had core-binding factor (CBF) with t(8;21). Time to treatment response for CR was as follows: 2 patients required 2 cycles, 1 required 1 cycle, and 1 required 4 cycles of decitabine. Two CN patients received 27 cycles of decitabine in total before therapy discontinuation because of trial termination [1]. One patient has remained in CR for 4 years while the second CR lasted for 8 years after cessation of therapy. Both patients were lost to follow-up after those time periods. The other CN patient remains in CR and continues on therapy (30 cycles). The CBF patient relapsed after 28 cycles of therapy and unfortunately succumbed to complications of relapsed AML. Molecular testing was performed on 2 of 4 patients as listed in Table 1 with no clear correlation to outcomes with this limited number of patients. Discussion Currently, the only acceptable long-term cure for patients with AML is intensive induction followed by consolidation with high-dose chemotherapy or allogeneic stem cell transplant. Elderly patients and patients with multiple comorbid conditions are not candidates for this aggressive approach. Here we present 4 patients with prolonged responses to 10-day decitabine induction therapy with 2 patients who appear to be cured of their AML with this approach. Further study is warranted to better characterize these super-responders and determine if there is a particular subpopulation of patients with AML that can achieve long-term survival without the perils of intensive chemotherapy or stem-cell transplantation. Disclosures Mannis: NKarta: Membership on an entity's Board of Directors or advisory committees; Agios: Research Funding; AbbVie: Membership on an entity's Board of Directors or advisory committees. Mims:Abbvie Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees; Novartis: Consultancy; Agios Pharmaceuticals: Consultancy, Membership on an entity's Board of Directors or advisory committees.

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RORA Is a Potential Prognostic Biomarker and Therapeutic Target for Patients with Acute Myeloid Leukemia

Blood ◽

10.1182/blood-2019-126311 ◽

2019 ◽

Vol 134 (Supplement_1) ◽

pp. 2696-2696

Author(s):

Christina A. Snider ◽

Kevin Fung ◽

Francesca Gould ◽

Vera Adema ◽

Cassandra M Kerr ◽

...

Keyword(s):

Acute Myeloid Leukemia ◽

Growth Inhibition ◽

Mrna Expression ◽

Board Of Directors ◽

Therapeutic Target ◽

Myeloid Leukemia ◽

Research Funding ◽

Advisory Committees ◽

Acute Myeloid

One strategy to improve precision medicine in acute myeloid leukemia (AML) is to further our understanding of the biological factors that influence pharmacologic efficacy. In a recent study, whole transcriptome analysis was conducted using pre- and post-treatment samples from a cohort of relapsed or refractory (R/R) AML patients treated with mitoxantrone, etoposide, cytarabine (MEC), and ixazomib. Logistic regression and linear discriminant analysis identified RORa as a predictor of response to treatment (Advani, et al., Clin Cancer Res, 2019, 4231-4237). RORa is not frequently mutated in AML but is described to be a tumor suppressor in patients with solid tumors. In the latter patient population, increased RORa expression is also associated with improved survival. We first retrospectively evaluated the prognostic significance of RORa in a larger cohort of R/R patients with AML (BeatAML). Second, we characterized RORa expression in cellular models of AML and sought to determine whether a commercially available RORa/RORγ agonist, SR1078, has anti-proliferative capacity in leukemia cell lines. For the correlative survival analyses, RORa mRNA expression and clinical information was downloaded from the BeatAML cohort (Tyner, et al., Nature, 2018, 526-531). Patients were categorized into high expressers (≥ median) and low expressers (<median). In total, 121 R/R patients were investigated. The mean age of diagnosis was 62 ± 12 years, and 58% of patients were male. The most common specific diagnosis at inclusion was AML with myelodysplasia related changes (28%), followed by AML with mutated NPM1 (18%) and therapy-related myeloid dysplasia (13%). Over half of the analyzed samples were peripheral blood (58%), and the remaining samples were either bone marrow aspirate (58%) or from leukapharesis (2%). As a whole, the median mRNA expression levels of RORa in patients with R/R AML (n=121) compared to healthy subjects (n=21) were 6.6 log2 CPM vs. 2.5 log2 CPM (P<.0001). After grouping patients into low RORa expresser and high RORa expresser groups, patients with above median expression of RORa were found to have significantly longer overall survival (19 mo. vs. 13 mo.; P=.0052; Figure 1). During normal hematopoiesis, we observed that RORa expression decreased with the stages of cellular maturation and is highly expressed in cells of AML patients with complex karyotype (BloodSpot, 2018). We noted that RORa mRNA expression varied across patients suggesting differences between AML subtypes. Expression levels were also confirmed to be different across AML subtypes in vitro by using cell line models (K-562, KG-1, OCI-AML3, U-937, THP-1, MOLM-13). In particular, analysis of TCGA data showed higher mean RORa expression in DNMT3A mutant (MT) (626 RPKM) compared to mean RORa expression in CEBPAMT (243 RPKM; P=0.026), NPM1MT (167 RPKM; P=0.012), and FLT3MT (236 RPKM; P=.003) AML patients. We then analyzed the sensitivity of cell lines to the commercially available synthetic RORa /RORγ ligand, SR1078. Myeloid lineage cells U-937 and KG-1 were used as a cellular model. Cells in logarithmic phase were treated with increased concentrations of the RORa /RORγ agonist SR1078 (from 3 nM to 30 mM) for 24 hours. Cell viability was measured by MTT tetrazolium reduction assay. Maximal growth inhibition was reached at 30 µM for U-937 (80%) and KG-1 (75%), respectively. We then combined SR1078 with MEC to evaluate whether in vitro SR1078 increased MEC growth inhibition. The addition of SR1078 to MEC significantly decreased cell viability in KG-1 cells compared to MEC alone (82% vs. 19%, P=.029). Our study suggests that increased RORa expression may be associated with improved survival in patients with R/R AML and that RORa may be a potential therapeutic target in AML. Figure 1 Disclosures Gerds: Celgene Corporation: Consultancy, Research Funding; Imago Biosciences: Research Funding; Pfizer: Consultancy; CTI Biopharma: Consultancy, Research Funding; Incyte: Consultancy, Research Funding; Roche: Research Funding; Sierra Oncology: Research Funding. Mukherjee:Bristol-Myers Squibb: Speakers Bureau; Takeda: Membership on an entity's Board of Directors or advisory committees; Pfizer: Honoraria; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Celgene Corporation: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Partnership for Health Analytic Research, LLC (PHAR, LLC): Consultancy; McGraw Hill Hematology Oncology Board Review: Other: Editor; Projects in Knowledge: Honoraria. Nazha:Jazz Pharmacutical: Research Funding; Incyte: Speakers Bureau; Novartis: Speakers Bureau; MEI: Other: Data monitoring Committee; Tolero, Karyopharma: Honoraria; Abbvie: Consultancy; Daiichi Sankyo: Consultancy. Maciejewski:Alexion: Consultancy; Novartis: Consultancy. Sekeres:Millenium: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees; Syros: Membership on an entity's Board of Directors or advisory committees. Advani:Abbvie: Research Funding; Pfizer: Honoraria, Research Funding; Glycomimetics: Consultancy, Research Funding; Amgen: Research Funding; Macrogenics: Research Funding; Kite Pharmaceuticals: Consultancy.

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Donor Cell Acute Myeloid Leukemia after Transplant for Follicular Lymphoma

Blood ◽

10.1182/blood-2019-122932 ◽

2019 ◽

Vol 134 (Supplement_1) ◽

pp. 5697-5697

Author(s):

Lacey S. Williams ◽

Catherine E. Lai

Keyword(s):

Bone Marrow ◽

Acute Myeloid Leukemia ◽

Follicular Lymphoma ◽

Board Of Directors ◽

Myeloid Leukemia ◽

Donor Cell ◽

Cell Leukemia ◽

Advisory Committees ◽

Donor Cell Leukemia ◽

Acute Myeloid

Donor cell leukemia is postulated to account for up to 5% of all leukemia "relapses" after hematopoietic stem cell transplant (SCT), though in many cases this is the first leukemia diagnosis for the patient if their transplant was for non-leukemia primary diseases. The rarity of the condition and heterogeneity of disease create challenges in diagnosis and management. In the present case, donor cell leukemia (DCL) developed in a 68-year-old female after allogeneic SCT 18 years earlier for follicular lymphoma. Only one other case of DCL after transplantation for follicular lymphoma has been reported (Boulton-Jones et al., Bone Marrow Transplantation, 2005). Furthermore, this case is atypical in that the presentation occurred many years after transplantation, since very few cases of DCL occur more than 15 years after original transplant. Case In 1993, the patient was diagnosed with stage IIIA follicular lymphoma at age 50. She achieved a complete remission with CHOP (cyclophosphamide, doxorubicin, vincristine, and prednisone) for 4 years. She relapsed in 1998 and received treatment with fludarabine and mitoxantrone. In 1999, she enrolled in a toxitumomab clinical trial (NCT00268203) but discontinued therapy secondary to side effects. Due to persistent disease, she proceeded with SCT and received EPOCH-F (etoposide, prednisone, vincristine, cyclophosphamide, doxorubicin, and fludarabine) prior to allogeneic SCT from her brother in 2000 (6/6 HLA match), augmented with TH2 cells. She received graft versus host disease (GVHD) prophylaxis with cyclosporine, however her post transplant course was complicated by engraftment syndrome and gastrointestinal and skin GVHD. In 2019, she presented to hematology for evaluation of worsening chronic neutropenia and thrombocytopenia persistent for three years, noted during work-up for symptomatic cholelithiasis. Bone marrow biopsy revealed acute myeloid leukemia (AML) with a hypocellular marrow with 30% blasts and myelodysplasia related changes. Her cytogenetics showed 46XY, +1, der(1;7)(q10;p10)/47,sl,+8/46,XY. FISH analyses demonstrated deletion 7q31 D7S486 locus in 156/200 cells (78%). NGS panel showed IDH1 (VAF16%) and U2AF1 (VAF 26%) mutations. Based on cytogenetics and chimerism studies showing 100% donor, the patient was diagnosed with donor-derived AML secondary to allogeneic SCT from her brother. The brother currently has no known hematologic problems. The patient was treated with CPX-351 (liposomal cytarabine and daunorubicin) and achieved a complete remission, followed by consolidation with CPX-351. Given her complex cytogenetics and poor prognosis, the patient proceeded to non-myeloablative haploidentical peripheral blood SCT from her son, with post-transplant cyclophosphamide. She subsequently had complications of neutropenic fever and C. dificile colitis, with progressive colitis leading to her death on day 22 after SCT. Discussion Though cytogenetic and molecular studies along with functional status assist clinicians in treatment decisions for DCL patients, the benefits and risks of treatment remain difficult to balance for this unique subset of leukemia. Of patients that achieve remission for greater than 18 months, many undergo second allogeneic SCT, however a similar number of patients have remissions of at least 18 months treated with chemotherapy alone (Wiseman, Biology of Blood and Bone Marrow Transplantation, 2011). In 15 reported cases that went to SCT, approximately 50% lived longer than 12 months after their DCL diagnosis. Second allogeneic SCT is often favored after initial remission in patients with good performance status due to high risk for relapse. This case illustrates the challenge in management of donor cell leukemia, a rather rare entity with very few cases in the literature developing greater than 15 years after transplant. Limited robust evidence favoring a particular treatment supports the need for further prospective studies. Disclosures Lai: Agios: Membership on an entity's Board of Directors or advisory committees; Daiichi-Sankyo: Membership on an entity's Board of Directors or advisory committees; Jazz Pharma: Membership on an entity's Board of Directors or advisory committees; Jazz Pharma: Speakers Bureau; Astellas: Speakers Bureau.

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Phase I Study of Ixazomib with Conventional Chemotherapy in the Treatment of Acute Myeloid Leukemia in Older Adults

Blood ◽

10.1182/blood-2020-140584 ◽

2020 ◽

Vol 136 (Supplement 1) ◽

pp. 7-8

Author(s):

Philip C. Amrein ◽

Eyal C. Attar ◽

Geoffrey Fell ◽

Traci M. Blonquist ◽

Andrew M. Brunner ◽

...

Keyword(s):

Acute Myeloid Leukemia ◽

Board Of Directors ◽

Myeloid Leukemia ◽

Research Funding ◽

Remission Rate ◽

Conventional Chemotherapy ◽

Advisory Committees ◽

Secondary Aml ◽

Grade 3 ◽

Acute Myeloid

Introduction: Outcomes for acute myeloid leukemia (AML) among older patients has remained largely unchanged for decades. Long-term survival for patients aged >60 years is poor (median survival 10.5 months). Targeting the proteasome in AML is attractive, since leukemia stem cells have demonstrated sensitivity to proteasome inhibition in preclinical models, perhaps through down regulation of nuclear NF-KB (Guzman, Blood 2001). AML cell lines are susceptible to synergistic cytotoxicity when bortezomib, a proteasome inhibitor, is combined with daunorubicin and cytarabine. We have shown that adding bortezomib to standard treatment in AML results in a high remission rate, although grade 2 sensory neurotoxicity was noted in approximately 12% of treated patients. A newer generation proteasome inhibitor, ixazomib, is less frequently associated with neurotoxicity, and, therefore, was selected for combination with conventional chemotherapy in this phase I trial. The primary objective of this study was to determine the maximum tolerated dose (MTD) of ixazomib in combination with conventional induction and consolidation chemotherapy for AML. Herein are the initial results of this trial. Methods: Adults >60 years of age with newly diagnosed AML were screened for eligibility. Patients with secondary AML were eligible, including those with prior hypomethylating agent therapy for myelodysplastic syndromes (MDS). We excluded those with promyelocytic leukemia. There were 2 phases in this study. In the first phase (A), the induction treatment consisted of the following: cytarabine 100 mg/m2/day by continuous IV infusion, Days 1-7; daunorubicin 60 mg/m2/day IV, Days 1, 2, 3, and ixazomib was provided orally at the cohort dose, Days 2, 5, 9, and 12. Consolidaton or transplant was at the discretion of the treating physician in phase A. In the second phase (B), induction was the same as that with the determined MTD of ixazomib. All patients were to be treated with the following consolidation: cytarabine at 2 g/m2/day, days 1-5 with ixazomib on days 2, 5, 9, and 12 at the cohort dose for consolidation. A standard 3 + 3 patient cohort dose escalation design was used to determine whether the dose of ixazomib could be safely escalated in 3 cohorts (1.5 mg/day, 2.3 mg/day, 3.0 mg/day), initially in induction (phase A) and subsequently in consolidation (phase B). The determined MTD of ixazomib in the first portion (A) of the trial was used during induction in the second portion (B), which sought to determine the MTD for ixazomib during consolidation. Secondary objectives included rate of complete remission, disease-free survival, and overall survival (OS). Results: Thirty-six patients have been enrolled on study, and 28 have completed dose levels A-1 through A-3 and B1 through B-2. Full information on cohort B-3 has not yet been obtained, hence, this report covers the experience with the initial 28 patients, cohorts A-1 through B-2. There were 12 (43%) patients among the 28 with secondary AML, either with prior hematologic malignancy or therapy-related AML. Nineteen patients (68%) were male, and the median age was 68 years (range 61-80 years). There have been no grade 5 toxicities due to study drug. Three patients died early due to leukemia, 2 of which were replaced for assessment of the MTD. Nearly all the grade 3 and 4 toxicities were hematologic (Table). There was 1 DLT (grade 4 platelet count decrease extending beyond Day 42). There has been no grade 3 or 4 neurotoxicity with ixazomib to date. Among the 28 patients in the first 5 cohorts, 22 achieved complete remissions (CR) and 2 achieved CRi, for a composite remission rate (CCR) of 86%. Among the 12 patients with secondary AML 8 achieved CR and 2 achieved CRi, for a CCR of 83%. The median OS for the 28 patients has not been reached (graph). The 18-month OS estimate was 65% [90% CI, 50-85%]. Conclusions: The highest dose level (3 mg) of ixazomib planned for induction in this trial has been reached safely. For consolidation there have been no serious safety issues in the first 2 cohorts with a dose up to 2.3 mg, apart from 1 DLT in the form of delayed platelet count recovery. The recommended phase 2 dose of ixazomib for induction is 3 mg. Accrual to cohort B-3 is ongoing. Notably, to date, no grade 3 or 4 neurotoxicity has been encountered. The remission rate in this older adult population with the addition of ixazomib to standard chemotherapy appears favorable. Figure Disclosures Amrein: Amgen: Research Funding; AstraZeneca: Consultancy, Research Funding; Takeda: Research Funding. Attar:Aprea Therapeutics: Current Employment. Brunner:Jazz Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees; Forty-Seven Inc: Membership on an entity's Board of Directors or advisory committees; AstraZeneca: Research Funding; Takeda: Research Funding; Celgene: Membership on an entity's Board of Directors or advisory committees, Research Funding; Novartis: Research Funding. Hobbs:Constellation: Honoraria, Research Funding; Novartis: Honoraria; Incyte: Research Funding; Merck: Research Funding; Bayer: Research Funding; Jazz: Honoraria; Celgene/BMS: Honoraria. Neuberg:Celgene: Research Funding; Madrigak Pharmaceuticals: Current equity holder in publicly-traded company; Pharmacyclics: Research Funding. Fathi:Blueprint: Consultancy; Boston Biomedical: Consultancy; BMS/Celgene: Consultancy, Research Funding; Novartis: Consultancy; Kura Oncology: Consultancy; Trillium: Consultancy; Amgen: Consultancy; Seattle Genetics: Consultancy, Research Funding; Abbvie: Consultancy; Pfizer: Consultancy; Newlink Genetics: Consultancy; Forty Seven: Consultancy; Trovagene: Consultancy; Kite: Consultancy; Daiichi Sankyo: Consultancy; Astellas: Consultancy; Amphivena: Consultancy; PTC Therapeutics: Consultancy; Agios: Consultancy, Research Funding; Takeda: Consultancy, Research Funding; Jazz: Consultancy. OffLabel Disclosure: Ixazomib is FDA approved for multiple myeloma. We are using it in this trial for acute myeloid leukemia.

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