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Purpose: Complete response to induction chemotherapy is observed in ∼60% of patients with newly diagnosed non-M3 acute myelogenous leukemia (AML). However, no methods exist to predict with high accuracy at the individual patient level the response to standard AML induction therapy.Experimental Design: We applied single-cell network profiling (SCNP) using flow cytometry, a tool that allows a comprehensive functional assessment of intracellular signaling pathways in heterogeneous tissues, to two training cohorts of AML samples (n = 34 and 88) to predict the likelihood of response to induction chemotherapy.Results: In the first study, univariate analysis identified multiple signaling “nodes” (readouts of modulated intracellular signaling proteins) that correlated with response (i.e., AUCROC ≥ 0.66; P ≤ 0.05) at a level greater than age. After accounting for age, similar findings were observed in the second study. For patients Conclusions: These data emphasize the value of performing quantitative SCNP under modulated conditions as a basis for the development of tests highly predictive for response to induction chemotherapy. SCNP provides information distinct from other known prognostic factors such as age, secondary AML, cytogenetics, and molecular alterations and is potentially combinable with the latter to improve clinical decision making. Independent validation studies are warranted. Clin Cancer Res; 16(14); 3721–33. ©2010 AACR.

Supplementary Fig. 2. Mutations found in the ASXL family of genes.

Purpose: The Forkhead transcription factors (FOXO) are tumor suppressor genes regulating differentiation, metabolism, and apoptosis that functionally interact with signal transduction pathways shown to be deregulated and prognostic in acute myelogenous leukemia (AML). This study evaluated the level of expression and the prognostic relevance of total and phosphorylated FOXO3A protein in AML.Experimental Design: We used reverse-phase protein array methods to measure the level of total and phosphoprotein expression of FOXO3A, in leukemia-enriched protein samples from 511 newly diagnosed AML patients.Results: The expression range was similar to normal CD34+ cells and similar in blood and marrow. Levels of total FOXO3A were higher at relapse compared with diagnosis. Levels of pFOXO3A or the ratio of phospho to total (PT) were not associated with karyotpe but were higher in patients with FLT3 mutations. Higher levels of pFOXO3A or PT-FOXO3A were associated with increased proliferation evidenced by strong correlation with higher WBC, percent marrow, and blood blasts and by correlation with higher levels of Cyclins B1, D1 and D3, pGSK3, pMTOR, and pStat5. Patients with High levels of pFOXO3A or PT-FOXO3A had higher rates of primary resistance and shorter remission durations, which combine to cause an inferior survival experience (P = 0.0002). This effect was independent of cytogenetics. PT-FOXO3A was a statistically significant independent predictor in multivariate analysis.Conclusions: High levels of phosphorylation of FOXO3A is a therapeutically targetable, independent adverse prognostic factor in AML. Clin Cancer Res; 16(6); 1865–74

Supplementary Data from Dasatinib Inhibits the Growth of Molecularly Heterogeneous Myeloid Leukemias

Purpose: Recent studies suggested that AKT activation might confer poor prognosis in acute myelogenous leukemia (AML), providing the rationale for therapeutic targeting of this signaling pathway. We, therefore, explored the preclinical and clinical anti-AML activity of an oral AKT inhibitor, MK-2206.Experimental Methods: We first studied the effects of MK-2206 in human AML cell lines and primary AML specimens in vitro. Subsequently, we conducted a phase II trial of MK-2206 (200 mg weekly) in adults requiring second salvage therapy for relapsed/refractory AML, and assessed target inhibition via reverse phase protein array (RPPA).Results: In preclinical studies, MK-2206 dose-dependently inhibited growth and induced apoptosis in AML cell lines and primary AML blasts. We then treated 19 patients with MK-2206 but, among 18 evaluable participants, observed only 1 (95% confidence interval, 0%–17%) response (complete remission with incomplete platelet count recovery), leading to early study termination. The most common grade 3/4 drug-related toxicity was a pruritic rash in 6 of 18 patients. Nevertheless, despite the use of MK-2206 at maximum tolerated doses, RPPA analyses indicated only modest decreases in Ser473 AKT (median 28%; range, 12%–45%) and limited inhibition of downstream targets.Conclusions: Although preclinical activity of MK-2206 can be demonstrated, this inhibitor has insufficient clinical antileukemia activity when given alone at tolerated doses, and alternative approaches to block AKT signaling should be explored. Clin Cancer Res; 20(8); 2226–35. ©2014 AACR.

Supplementary Fig. 1. Mutational spectrum of pediatric ALL.

High expression of VEGFC predicts adverse prognosis in acute myeloid leukemia (AML). We therefore explored VEGFC-targeting efficacy as an AML therapy using a VEGFC mAb. VEGFC antibody therapy enforced myelocytic differentiation of clonal CD34+ AML blasts. Treatment of CD34+ AML blasts with the antibody reduced expansion potential by 30% to 50% and enhanced differentiation via FOXO3A suppression and inhibition of MAPK/ERK proliferative signals. VEGFC antibody therapy also accelerated leukemia cell differentiation in a systemic humanized AML mouse model. Collectively, these results define a regulatory function of VEGFC in CD34+ AML cell fate decisions via FOXO3A and serve as a new potential differentiation therapy for patients with AML.Significance: These findings reveal VEGFC targeting as a promising new differentiation therapy in AML. Cancer Res; 78(20); 5940–8. ©2018 AACR.

Supplementary Table 1. Samples used for whole exome and targeted sequencing. Supplementary Table 2. Clinical information of patients. Supplementary Table 3. 560 Genes screened in the targeted sequencing. Supplementary Table 4. Sequences of shRNA or CRISPR-Cas9 sgRNA used in this study. Supplementary Table 5. Sequences of real-time PCR primers used in this study. Supplementary Table 6. Sequences of primers used for Sanger validation of CRISPR-Cas9 indel. Supplementary Table 7. Mutations of KRAS identified in this study have been found in a variety of cancers. Supplementary Table 8. Mutations of PTPN11 in other cancers occurring in the same location as in our ALL cohort. Supplementary Table 9. Recurrent mutation sites of FLT3 in different cancers. Supplementary Table 10. Cell lines or cancer samples harboring either E1099K or T1150A hotspot mutation of WHSC1. Supplementary Table 11. R1446 mutations of CREBBP recurrently occurred in a variety of cancers. Supplementary Table 12. Mutational hotspot (D1399) of EP300 occurring in other cancers occurring in the same location as in our ALL cohort.Supplementary Table 13. Rare mutations occurring in our ALL cohort (mutated in one individual each), but these mutations have been recurrently documented in the COSMIC cancer mutations database. Supplementary Table 14. Mutations of genes involved in DNA repair pathway in relapse samples.

The survival of malignant leukemic cells is dependent on DNA damage repair (DDR) signaling. Reverse Phase Protein Array (RPPA) data sets were assembled using diagnostic samples from 810 adult and 500 pediatric acute myelogenous leukemia (AML) patients and were probed with 412 and 296 strictly validated antibodies, respectively, including those detecting the expression of proteins directly involved in DDR. Unbiased hierarchical clustering identified strong recurrent DDR protein expression patterns in both adult and pediatric AML. Globally, DDR expression was associated with gene mutational statuses and was prognostic for outcomes including overall survival (OS), relapse rate, and remission duration (RD). In adult patients, seven DDR proteins were individually prognostic for either RD or OS. When DDR proteins were analyzed together with DDR−related proteins operating in diverse cellular signaling pathways, these expanded groupings were also highly prognostic for OS. Analysis of patients treated with either conventional chemotherapy or venetoclax combined with a hypomethylating agent revealed protein clusters that differentially predicted favorable from unfavorable prognoses within each therapy cohort. Collectively, this investigation provides insight into variable DDR pathway activation in AML and may help direct future individualized DDR−targeted therapies in AML patients.

SUMMARYB-cell lineage acute lymphoblastic leukemia (B-ALL) is comprised of diverse molecular subtypes and while transcriptional and DNA methylation profiling of B-ALL subtypes has been extensively examined, the accompanying chromatin landscape is not well characterized for many subtypes. We therefore mapped chromatin accessibility using ATAC-seq for 10 B-ALL molecular subtypes in primary ALL cells from 154 patients. Comparisons with B-cell progenitors identified candidate B-ALL cell-of-origin and AP-1-associatedcis-regulatory rewiring in B-ALL.Cis-regulatory rewiring promoted B-ALL-specific gene regulatory networks impacting oncogenic signaling pathways that perturb normal B-cell development. We also identified that over 20% of B-ALL accessible chromatin sites exhibit strong subtype enrichment, with transcription factor (TF) footprint profiling identifying candidate TFs that maintain subtype-specific chromatin architectures. Over 9000 inherited genetic variants were further uncovered that contribute to variability in chromatin accessibility among individual patient samples. Overall, our data suggest that distinct chromatin architectures are driven by diverse TFs and inherited genetic variants which promote unique gene regulatory networks that contribute to transcriptional differences among B-ALL subtypes.HIGHLIGHTSPro-B progenitor cells as the most common cell-of-origin for B-ALLAP-1 TF-associatedcis-regulatory rewiring in B-ALLSubtype-specific accessible chromatin signatures representing 20% of all B-ALL sitesRole for distinct TFs in promoting subtype-specific chromatin architecturesThousands of inherited genetic variants identified impacting chromatin state

Although acute lymphoblastic leukemia (ALL) is the most common childhood cancer, there is limited understanding of the contribution of inherited genetic variation on inter-individual differences in chemotherapy response. Defining genetic factors impacting therapy failure can help better predict response and identify drug resistance mechanisms. We therefore mapped inherited noncoding variants associated with chemotherapeutic drug resistance and/or treatment outcome to ALLcis-regulatory elements and investigated their gene regulatory potential and genomic connectivity using massively parallel reporter assays and promoter capture Hi-C, respectively. We identified 53 variants with reproducible allele-specific effects on transcription and high-confidence gene targets. Subsequent functional interrogation of the top variant (rs1247117) determined that it disrupted a PU.1 consensus motif and PU.1 binding affinity. Importantly, deletion of the genomic interval containing rs1247117 sensitized ALL cells to vincristine. Together, these data demonstrate that noncoding regulatory variation associated with diverse pharmacological traits harbor significant effects on allele-specific transcriptional activity and impact sensitivity to chemotherapeutic agents in ALL.

Contains fulltext : 241347.pdf (Publisher’s version ) (Closed access) Asparaginase (ASNase) therapy has been a mainstay of acute lymphoblastic leukemia (ALL) protocols for decades and shows promise in the treatment of a variety of other cancers. To improve the efficacy of ASNase treatment, we used a CRISPR/Cas9-based screen to identify actionable signaling intermediates that improve the response to ASNase. Both genetic inactivation of Bruton's tyrosine kinase (BTK) and pharmacological inhibition by the BTK inhibitor ibrutinib strongly synergize with ASNase by inhibiting the amino acid response pathway, a mechanism involving c-Myc-mediated suppression of GCN2 activity. This synthetic lethal interaction was observed in 90% of patient-derived xenografts, regardless of the genomic subtype. Moreover, ibrutinib substantially improved ASNase treatment response in a murine PDX model. Hence, ibrutinib may be used to enhance the clinical efficacy of ASNase in ALL. This trial was registered at www.clinicaltrials.gov as # NCT02884453.

Key Points Effector polyfunctional index score of CD4 cells was more associated with responses to combined azacitidine/nivolumab than CD8 cells.Single-cell biomarker assays can be used in predicting responses to immune-based therapies., Acute myeloid leukemia (AML) remains a difficult disease to treat disease. In a phase 2 clinical trial in patients with relapsed/refractory AML, combining the hypomethylating agent, azacitidine, with the PD-1 checkpoint inhibitor, nivolumab, demonstrated encouraging response rates (33%), median event-free, and overall survival, compared with a historical cohort of contemporary patients treated with azacitidine-based therapies, with an acceptable safety profile. Biomarkers of response are yet to be determined. In this study, we leveraged a multiplexed immune assay to assess the functional states of CD4+ and CD8+ cells at a single-cell level in pretherapy bone marrows in 16 patients with relapsed/refractory AML treated with azacitidine/nivolumab. Effector CD4+ but not CD8+ cells had distinct polyfunctional groups and were associated with responses and better outcomes. Further evaluation of the polyfunctional strength index composition across cell types revealed that interferon-gamma (IFN-γ) and tumor necrosis factor-alpha (TNF-α) were the major drivers of enhanced polyfunctionality index of pretherapy CD4+ subset, whereas Granzyme B, IFN-γ, MIP-1b, and TNF-α drove the nonsignificantly enhanced pretreatment Polyfunctional Strength Index of CD8+ subset in the responders. Single-cell polyfunctional assays were predictive of response in AML and may have a potential role as a biomarker in the wider sphere of immunotherapy.

The chronic lymphocytic leukemia (CLL) armamentarium has evolved significantly, with novel therapies that inhibit Bruton Tyrosine Kinase, PI3K delta and/or the BCL2 protein improving outcomes. Still, the clinical course of CLL patients is highly variable and most previously recognized prognostic features lack the capacity to predict response to modern treatments indicating the need for new prognostic markers. In this study, we identified four epigenetically distinct proteomic signatures of a large cohort of CLL and related diseases derived samples (n = 871) using reverse phase protein array technology. These signatures are associated with clinical features including age, cytogenetic abnormalities [trisomy 12, del(13q) and del(17p)], immunoglobulin heavy-chain locus (IGHV) mutational load, ZAP-70 status, Binet and Rai staging as well as with the outcome measures of time to treatment and overall survival. Protein signature membership was identified as predictive marker for overall survival regardless of other clinical features. Among the analyzed epigenetic proteins, EZH2, HDAC6, and loss of H3K27me3 levels were the most independently associated with poor survival. These findings demonstrate that proteomic based epigenetic biomarkers can be used to better classify CLL patients and provide therapeutic guidance.

PURPOSE Sixty percent of newly diagnosed patients with acute myeloid leukemia (ND-AML) receiving frontline therapy attain a complete response (CR), yet 30%-40% of patients relapse. Relapsed or refractory AML (R/R-AML) remains a particularly adverse population necessitating improved therapeutic options. This phase Ib/II study evaluated the safety and efficacy of fludarabine, cytarabine, granulocyte colony-stimulating factor, and idarubicin combined with the B-cell lymphoma-2 inhibitor venetoclax in ND-AML and R/R-AML. MATERIALS AND METHODS The phase IB portion (PIB) enrolled patients with R/R-AML using a 3 + 3 dose escalation and de-escalation algorithm for identification of maximum tolerated dose and dose-limiting toxicities. The phase II portion enrolled patients into two arms to evaluate response and time-to-event end points: phase IIA (PIIA): ND-AML and phase IIB (PIIB): R/R-AML. RESULTS Sixty-eight patients have enrolled to date (PIB, 16; PIIA, 29; PIIB, 23). Median age was 46 years (range, 20-73). Grade 3 and 4 adverse events occurring in ≥ 10% of patients included febrile neutropenia (50%), bacteremia (35%), pneumonia (28%), and sepsis (12%). The overall response rate for PIB, PIIA, and PIIB was 75%, 97%, and 70% with 75%, 90%, and 61%, respectively, achieving a composite CR. Measurable residual disease–negative composite CR was attained in 96% of ND-AML and 69% of R/R-AML patients. After a median follow-up of 12 months, median overall survival (OS) for both PII cohorts was not reached. Fifty-six percent of patients proceeded to allogeneic hematopoietic stem-cell transplantation (ND-AML, 69%; R/R-AML, 46%). In R/R-AML, allogeneic hematopoietic stem-cell transplantation resulted in a significant improvement in OS (median OS, NR; 1-year OS, 87%). One-year survival post-HSCT was 94% in ND-AML and 78% in R/R-AML. CONCLUSION Fludarabine, cytarabine, granulocyte colony-stimulating factor, and idarubicin + venetoclax represents an effective intensive treatment regimen in ND-AML and R/R-AML patients, associated with deep remissions and a high rate of transition to successful transplantation.

Acute myeloid leukemia (AML) is driven by numerous molecular events that contribute to disease progression. Herein, we identify hnRNP K overexpression as a recurrent abnormality in AML that negatively correlates with patient survival. Overexpression of hnRNP K in murine fetal liver cells results in altered self-renewal and differentiation potential. Further, murine transplantation models reveal that hnRNP K overexpression results in myeloproliferation in vivo. Mechanistic studies expose a direct functional relationship between hnRNP K and RUNX1—a master transcriptional regulator of hematopoiesis often dysregulated in leukemia. Molecular analyses show that overexpression of hnRNP K results in an enrichment of an alternatively spliced isoform of RUNX1 lacking exon 4. Our work establishes hnRNP K’s oncogenic potential in influencing myelogenesis through its regulation of RUNX1 splicing and subsequent transcriptional activity.

TP53 mutations in acute myeloid leukemia (AML) are associated with resistance to standard treatments and dismal outcomes. The incidence and prognostic impact of the emergence of newly detectable TP53 mutations over the course of AML therapy has not been well described. We retrospectively analyzed 200 patients with newly diagnosed TP53 wild type AML who relapsed after or were refractory to frontline therapy. Twenty-nine patients (15%) developed a newly detectable TP53 mutation in the context of relapsed/refractory disease. The median variant allelic frequency (VAF) was 15% (range, 1.1% - 95.6%). TP53 mutations were more common after intensive therapy versus lower-intensity therapy (23% versus 10%, respectively; P=0.02) and in patients who had undergone hematopoietic stem cell transplant versus those who had not (36% versus 12%, respectively; P=0.005). Lower TP53 VAF was associated with an increased likelihood of complete remission (CR) or CR with incomplete hematologic recovery (CRi) compared to higher TP53 VAF (CR/CRi rate of 41% for VAF

Lineage-ambiguous leukemias are high-risk malignancies of poorly understood genetic basis. Here, we describe a distinct subgroup of acute leukemia with expression of myeloid, T lymphoid, and stem cell markers driven by aberrant allele-specific deregulation of BCL11B, a master transcription factor responsible for thymic T-lineage commitment and specification. Mechanistically, this deregulation was driven by chromosomal rearrangements that juxtapose BCL11B to superenhancers active in hematopoietic progenitors, or focal amplifications that generate a superenhancer from a noncoding element distal to BCL11B. Chromatin conformation analyses demonstrated long-range interactions of rearranged enhancers with the expressed BCL11B allele and association of BCL11B with activated hematopoietic progenitor cell cis-regulatory elements, suggesting BCL11B is aberrantly co-opted into a gene regulatory network that drives transformation by maintaining a progenitor state. These data support a role for ectopic BCL11B expression in primitive hematopoietic cells mediated by enhancer hijacking as an oncogenic driver of human lineage-ambiguous leukemia. Significance: Lineage-ambiguous leukemias pose significant diagnostic and therapeutic challenges due to a poorly understood molecular and cellular basis. We identify oncogenic deregulation of BCL11B driven by diverse structural alterations, including de novo superenhancer generation, as the driving feature of a subset of lineage-ambiguous leukemias that transcend current diagnostic boundaries. This article is highlighted in the In This Issue feature, p. 2659

In an effort to identify acute myeloid leukemia (AML)-restricted targets for therapeutic development in AML, we analyzed the transcriptomes of 2051 children and young adults with AML and compared the expression profile with normal marrow specimens. This analysis identified a large cohort of AML-restricted genes with high expression in AML, but low to no expression in normal hematopoiesis. Mesothelin (MSLN), a known therapeutic target in solid tumors, was shown to be highly overexpressed in 36% of the AML cohort (range, 5-1077.6 transcripts per million [TPM]) and virtually absent in normal marrow (range, 0.1-10.7 TPM). We verified MSLN transcript expression by quantitative reverse transcription polymerase chain reaction, confirmed cell surface protein expression on leukemic blasts by multidimensional flow cytometry, and demonstrated that MSLN expression was associated with promoter hypomethylation. MSLN was highly expressed in patients with KMT2A rearrangements (P < .001), core-binding factor fusions [inv(16)/t(16;16), P < .001; t(8;21), P < .001], and extramedullary disease (P = .001). We also demonstrated the presence of soluble MSLN in diagnostic serum specimens using an MSLN-directed enzyme-linked immunosorbent assay. In vitro and in vivo preclinical efficacy of the MSLN-directed antibody-drug conjugates (ADCs) anetumab ravtansine and anti-MSLN–DGN462 were evaluated in MSLN+ leukemia cell lines in vitro and in vivo, as well as in patient-derived xenografts. Treatment with ADCs resulted in potent target-dependent cytotoxicity in MSLN+ AML. In this study, we demonstrate that MSLN is expressed in a significant proportion of patients with AML and holds significant promise as a diagnostic and therapeutic target in AML, and that MSLN-directed therapeutic strategies, including ADCs, warrant further clinical investigation.

Bortezomib (BTZ) was recently evaluated in a randomized phase 3 clinical trial by the Children’s Oncology Group (COG) that compared standard chemotherapy (cytarabine, daunorubicin, and etoposide [ADE]) vs standard therapy with BTZ (ADEB) for de novo pediatric acute myeloid leukemia (AML). Although the study concluded that BTZ did not improve outcome overall, we examined patient subgroups benefiting from BTZ-containing chemotherapy using proteomic analyses. The proteasome inhibitor BTZ disrupts protein homeostasis and activates cytoprotective heat shock responses. Total heat shock factor 1 (HSF1) and phosphorylated HSF1 (HSF1-pSer326) were measured in leukemic cells from 483 pediatric patients using reverse phase protein arrays. HSF1-pSer326 phosphorylation was significantly lower in pediatric AML compared with CD34+ nonmalignant cells. We identified a strong correlation between HSF1-pSer326 expression and BTZ sensitivity. BTZ significantly improved outcome of patients with low-HSF1-pSer326 with a 5-year event-free survival of 44% (ADE) vs 67% for low-HSF1-pSer326 treated with ADEB (P = .019). To determine the effect of HSF1 expression on BTZ potency in vitro, cell viability with HSF1 gene variants that mimicked phosphorylated (S326A) and nonphosphorylated (S326E) HSF1-pSer326 were examined. Those with increased HSF1 phosphorylation showed clear resistance to BTZ vs those with wild-type or reduced HSF1-phosphorylation. We hypothesize that HSF1-pSer326 expression could identify patients who benefit from BTZ-containing chemotherapy.

Background: Intensive chemotherapy has improved complete remission (CR) rates in T-ALL/LBL. However, long-term overall survival (OS) continues to be suboptimal at about 50%. Nelarabine (NEL) is active in relapsed/refractory T-ALL. PEG-asparaginase (PEG) containing regimens have demonstrated significant activity in the pediatric population (Dunsmore et al. COG AALL0434. J Clin Oncol. 2018 Abstr #10500). We hypothesized that incorporating NEL and PEG in the frontline setting may further improve outcomes in adult T-ALL/LBL (NCT00501826). We had previously reported on the addition of nelarabine to the hyper-CVAD regimen (Abaza Y, Am J Hematol. 2018). Here, we report outcomes with the addition of PEG and longer follow-up. Methods Pts with T-ALL/LBL, either newly diagnosed or minimally treated (no CR with 1 cycle or CR after ≤2 cycles of induction) were eligible. Pts needed to have ECOG performance status (PS) ≤3, creatinine ≤2 mg/dL, bilirubin ≤2 mg/dL and ALT/AST ≤4 x ULN. The objectives were to determine safety, CR rate, progression-free survival (PFS), and OS with this combination. Pts received 8 cycles of with HCVAD (odd cycles 1,3,5,7) alternating with methotrexate (MTX) and cytarabine (ara-C; even cycles 2,4,6,8) at approximately 3-week intervals. Initially pts received NEL 650 mg/m2 IV daily for 5 days after 8 cycles as above. After enrollment of 30 pts, protocol was amended to deliver the 2 courses of NEL after cycle 4 and 5 of therapy in an attempt to reduce risk of early relapse. After enrollment of 79 pts, PEG (1500 IU/m2 IV capped at 3750 IU for pts ≤60 yrs and at 2000 IU for pts >60 yrs) was added to NEL courses on day 5. After completion of intensive phase, pts received 30 months of POMP maintenance (6-MP, vincristine, MTX, prednisone) with NEL/PEG replacing cycle 6 and 7 of POMP as early intensification. Late intensification consisted of MTX/PEG replacing cycle 18 and HCVAD replacing cycle 19 of POMP. CNS prophylaxis included 8 intrathecal doses of MTX alternating with ara-C. Pts with bulky mediastinal disease were considered for local radiation therapy prior to starting POMP maintenance. Results: Between August 16, 2007 and March 6, 2019, we enrolled 96 pts with T-ALL (n=57) and T-LBL (n=39). Median age was 35 yrs (range 18-78), 75 pts were male (78%) and 83 pts (86%) had PS 0-1. In pts with T-ALL, median bone marrow blast % was 79% (range 0-96), 70 pts (73%) had extramedullary disease with mediastinal involvement in 48 pts and CNS involvement in 5 pts. 60 pts (63%) had diploid cytogenetics, 13 pts (14%) had complex cytogenetics, 19 pts had miscellaneous abnormalities and 4 pts had insufficient metaphases. Based on immunophenotype, pts were further characterized as thymic (n=38), early T-cell precursor (ETP; n=38), mature (n=10), not otherwise specified (NOS; n=9), and early-non-ETP (n=3). 12 pts had received prior therapy with HCVAD (n=8), asparaginase-based regimen (n=2), and others (n=2). 30-day mortality was 0%. 9 pts were in CR at enrollment. There were 242 grade 3/4 non-hematological treatment-emergent adverse events (TEAE) with the most frequent grade 3/4 TEAEs being neutropenic infections in 74 pts (77%), hypokalemia in 42 pts (44%), hypophosphatemia in 42 pts (44%) and elevated ALT in 28 pts (29%). Overall response rate was 98% (85/87 pts) with CR in 95% pts (n=83), PR in 2% (n=2), and no response in 2% (n=2). The median no. of cycles to response was 1 (range 1-10) with CR reported based on achieving PET negativity as well as standard response criteria. CR rate in T-ALL was 88% (43/49) and T-LBL was 100% (38/38). At a median follow-up of 58.8 months (mo; 95% CI 42.2, 67.6), 61 pts (63%) are alive with a median PFS and OS of 135.3 and 135.1 mo, respectively (Fig 1a, b) and 55 pts (57%) remain in CR. For pts who received NEL after HCVAD cycle 4, the 7-year OS was 61% compared to 52% who received NEL after cycle 8, hazard ratio 0.91, 95% CI 0.46, 1.82, p=0.7. For pts receiving PEG in addition to NEL, the 2yr OS was 86% (Fig 1c). 30 pts (31%) relapsed and 35 pts (37%) have died including 9 pts who died in CR. 17 pts (18%) pts underwent allogeneic stem cell transplantation, 6 have relapsed, 10 are alive, and 1 pt died in remission. Rate of relapse was 37% with NEL after cycle 8, 33% (16/49) with NEL after cycle 4 and 21% (3/14) with NEL/PEG (p=.38). Conclusion: HCVAD with NEL with or without PEG is safe and effective as frontline therapy of adults with newly diagnosed T-ALL/LBL. Addition of PEG appears to confer a benefit. Figure Disclosures Maiti: Celgene: Research Funding. Jabbour:Adaptive Biotechnologies: Other: Advisory role, Research Funding; AbbVie: Other: Advisory role, Research Funding; Amgen: Other: Advisory role, Research Funding; Genentech: Other: Advisory role, Research Funding; BMS: Other: Advisory role, Research Funding; Takeda: Other: Advisory role, Research Funding; Pfizer: Other: Advisory role, Research Funding. Jain:Servier: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Incyte: Research Funding; BeiGene: Honoraria, Membership on an entity's Board of Directors or advisory committees; TG Therapeutics: Honoraria, Membership on an entity's Board of Directors or advisory committees; Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees; Aprea Therapeutics: Research Funding; Fate Therapeutics: Research Funding; Precision Bioscienes: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Adaptive Biotechnologies: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Verastem: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Cellectis: Research Funding; ADC Therapeutics: Research Funding; Pfizer: Research Funding; BMS: Research Funding; AstraZeneca: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Genentech: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; AbbVie: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Pharmacyclics: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding. Borthakur:FTC Therapeutics: Consultancy; Curio Science LLC: Consultancy; Oncoceutics: Research Funding; Xbiotech USA: Research Funding; Cyclacel: Research Funding; PTC Therapeutics: Consultancy; BioLine Rx: Research Funding; BMS: Research Funding; AstraZeneca: Research Funding; Nkarta Therapeutics: Consultancy; Treadwell Therapeutics: Consultancy; BioLine Rx: Consultancy; Novartis: Research Funding; Abbvie: Research Funding; Jannsen: Research Funding; GSK: Research Funding; PTC Therapeutics: Research Funding; Polaris: Research Funding; BioTherix: Consultancy; Argenx: Consultancy; Incyte: Research Funding. Kadia:Incyte: Research Funding; Abbvie: Honoraria, Research Funding; Astellas: Research Funding; Astra Zeneca: Research Funding; Cyclacel: Research Funding; BMS: Honoraria, Research Funding; Pfizer: Honoraria, Research Funding; Amgen: Research Funding; Celgene: Research Funding; Genentech: Honoraria, Research Funding; JAZZ: Honoraria, Research Funding; Ascentage: Research Funding; Cellenkos: Research Funding; Novartis: Honoraria; Pulmotec: Research Funding. Burger:TG Therapeutics: Research Funding, Speakers Bureau; AstraZeneca: Consultancy; Beigene: Research Funding, Speakers Bureau; Pharmacyclics, an AbbVie company: Consultancy, Research Funding, Speakers Bureau; Gilead Sciences: Consultancy, Research Funding; Janssen Pharmaceuticals: Consultancy, Speakers Bureau. Konopleva:Kisoji: Consultancy; Agios: Research Funding; Genentech: Consultancy, Research Funding; Reata Pharmaceutical Inc.;: Patents & Royalties: patents and royalties with patent US 7,795,305 B2 on CDDO-compounds and combination therapies, licensed to Reata Pharmaceutical; Amgen: Consultancy; Calithera: Research Funding; Cellectis: Research Funding; Ablynx: Research Funding; AbbVie: Consultancy, Research Funding; F. Hoffmann La-Roche: Consultancy, Research Funding; Stemline Therapeutics: Consultancy, Research Funding; AstraZeneca: Research Funding; Ascentage: Research Funding; Rafael Pharmaceutical: Research Funding; Forty-Seven: Consultancy, Research Funding; Eli Lilly: Research Funding; Sanofi: Research Funding. Alvarado:Jazz Pharmaceuticals: Research Funding; MEI Pharma: Research Funding; BerGenBio ASA: Research Funding; Sun Pharma: Research Funding; Daiichi-Sankyo: Research Funding; Tolero Pharmaceuticals: Research Funding; Astex Pharmaceuticals: Research Funding; FibroGen: Research Funding. Short:AstraZeneca: Consultancy; Amgen: Honoraria; Takeda Oncology: Consultancy, Honoraria, Research Funding; Astellas: Research Funding. Faderl:Jazz Pharmaceuticals: Current Employment, Current equity holder in publicly-traded company. O'Brien:Amgen, Astellas, Celgene, GlaxoSmithKline, Janssen Oncology, Aptose Biosciences Inc. Vaniam Group, AbbVie, Alexion, Verastem, Eisai, Juno Therapeutics, Vida Ventures: Consultancy; Gilead, Pharmacyclics, TG Therapeutics, Pfizer, Sunesis: Consultancy, Research Funding; Kite, Regeneron, Acerta: Research Funding. Daver:Genentech: Research Funding; AbbVie: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Astellas: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Novimmune: Research Funding; Gilead: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Amgen: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Trovagene: Research Funding; Fate Therapeutics: Research Funding; ImmunoGen: Research Funding; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees; Celgene: Consultancy, Membership on an entity's Board of Directors or advisory committees; Jazz: Consultancy, Membership on an entity's Board of Directors or advisory committees; Trillium: Consultancy, Membership on an entity's Board of Directors or advisory committees; Syndax: Consultancy, Membership on an entity's Board of Directors or advisory committees; Amgen: Consultancy, Membership on an entity's Board of Directors or advisory committees; KITE: Consultancy, Membership on an entity's Board of Directors or advisory committees; Agios: 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; Bristol-Myers Squibb: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Pfizer: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Karyopharm: Research Funding; Servier: Research Funding. Pemmaraju:Plexxikon: Research Funding; Affymetrix: Other: Grant Support, Research Funding; Pacylex Pharmaceuticals: Consultancy; Celgene: Honoraria; Daiichi Sankyo: Research Funding; DAVA Oncology: Honoraria; Samus Therapeutics: Research Funding; Cellectis: Research Funding; LFB Biotechnologies: Honoraria; Novartis: Honoraria, Research Funding; SagerStrong Foundation: Other: Grant Support; Incyte Corporation: Honoraria; Stemline Therapeutics: Honoraria, Research Funding; Blueprint Medicines: Honoraria; Roche Diagnostics: Honoraria; AbbVie: Honoraria, Research Funding; MustangBio: Honoraria. Bose:Astellas Pharmaceuticals: Research Funding; CTI BioPharma: Honoraria, Research Funding; Celgene Corporation: Honoraria, Research Funding; Kartos Therapeutics: Honoraria, Research Funding; Constellation Pharmaceuticals: Research Funding; Blueprint Medicines Corporation: Honoraria, Research Funding; Pfizer, Inc.: Research Funding; Incyte Corporation: Consultancy, Honoraria, Research Funding, Speakers Bureau; Promedior, Inc.: Research Funding; NS Pharma: Research Funding. Thompson:AbbVie: Research Funding; Pharmacyclics: Research Funding; Janssen-Cilag: Honoraria; Adaptive Biotechnologies: Consultancy, Research Funding; Genentech: Consultancy. Garcia-Manero:Bristol-Myers Squibb: Consultancy, Research Funding; Genentech: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Amphivena Therapeutics: Research Funding; Helsinn Therapeutics: Consultancy, Honoraria, Research Funding; Acceleron Pharmaceuticals: Consultancy, Honoraria; AbbVie: Honoraria, Research Funding; Jazz Pharmaceuticals: Consultancy; Celgene: Consultancy, Honoraria, Research Funding; Astex Pharmaceuticals: Consultancy, Honoraria, Research Funding; Merck: Research Funding; Novartis: Research Funding; H3 Biomedicine: Research Funding; Onconova: Research Funding. Kantarjian:Amgen: Honoraria, Research Funding; Abbvie: Honoraria, Research Funding; Daiichi-Sankyo: Honoraria, Research Funding; Ascentage: Research Funding; Immunogen: Research Funding; Aptitute Health: Honoraria; BioAscend: Honoraria; Delta Fly: Honoraria; Janssen: Honoraria; Oxford Biomedical: Honoraria; Novartis: Honoraria, Research Funding; Pfizer: Honoraria, Research Funding; Sanofi: Research Funding; Actinium: Honoraria, Membership on an entity's Board of Directors or advisory committees; Adaptive biotechnologies: Honoraria; BMS: Research Funding; Jazz: Research Funding. Ravandi:Astellas: Consultancy, Honoraria, Research Funding; Amgen: Consultancy, Honoraria, Research Funding; Macrogenics: Research Funding; Jazz Pharmaceuticals: Consultancy, Honoraria, Research Funding; Xencor: Consultancy, Honoraria, Research Funding; Abbvie: Consultancy, Honoraria, Research Funding; BMS: Consultancy, Honoraria, Research Funding; AstraZeneca: Consultancy, Honoraria; Orsenix: Consultancy, Honoraria, Research Funding; Celgene: Consultancy, Honoraria.

CPX-351 is a liposomal formulation of cytarabine/daunorubicin with a 5:1 fixed molar ratio. We investigated the safety and efficacy of escalating doses of CPX-351 in patients with acute myeloid leukemia (AML) at high risk of induction mortality with standard chemotherapy determined through assessment of leukemia and patient-related risk factors for intensive chemotherapy in an open-label, phase II trial. Patients were randomized to receive 50 or 75 units/m2 on days 1, 3, and 5. Once safety was established, a 100 units/m2 arm was opened. Fifty-six patients were enrolled, 16, 24, and 16 in the 50, 75, and 100 units/m2 arms, respectively. The composite complete remission rate (complete remission + complete remission with incomplete blood count recovery) was lowest with 50 units/m2 (19%) compared with 75 units/m2 (38%) and 100 units/m2 (44%) (P = 0.35). The 50 units/m2 arm had a median OS of 4.3 months, compared with 8.6 and 6.2 months for the 75 and 100 units/m2 respectively (P = 0.04). Nonhematologic grade 3/4 treatment-emergent adverse events included febrile neutropenia (34%), pneumonia (23%), and sepsis (16%). CPX-351 at 75 units/m2 has favorable safety and efficacy for AML patients at high risk of induction mortality with some tolerating the standard dose of 100 units/m2.

The value of event-free survival (EFS) as an end point in acute myeloid leukemia (AML) trials has been questioned. We hypothesized that rather than a surrogate for overall survival (OS), improvement in EFS may decrease the use of health care. In this retrospective study, we identified 400 patients with AML who were treated on first-line therapy trials and had OS between 2 and 36 months. We captured health care use from diagnosis until death or until the patient was censored at stem cell transplantation (SCT). We used correlation and regression analysis to determine the relation between health care use and EFS. Among patients with newly diagnosed AML, 35% had adverse-risk AML, 48% received intensive chemotherapy, and 28% received hypomethylating agents. The median EFS censored at SCT was 9.7 months. Longer EFS led to a significant decline in health care use regardless of OS. This held true for all observations, including overall health care use (r = −0.45), sum of clinic visits, emergency room visits, hospitalizations, consultations (r = −0.44), sum of invasive procedures, laboratory and imaging studies (r = −0.51), and blood product transfusions (r = −0.19). These correlations were stronger for patients who achieved a complete remission and held true across age, treatment, and disease risk subgroups. In patients with newly diagnosed AML, improvement in EFS correlates with a decrease in all health care use irrespective of OS duration.

Introduction: The biological heterogeneity of acute myeloid leukemia (AML) complicates personalized medicine. Individual prognosis is typically based on the presence of chromosomal and genetic lesions. Nevertheless, these classifications often lack a priori information about response to therapy. Since the protein expression landscape reflects the functional activity state of cells, we hypothesize that analyzing this can be used for the identification of protein activity markers to provide better risk stratification as well as may provide targeted therapeutic guidance in AML. Areas covered: Herein, we review recently new adopted drugs in the treatment for AML and discuss how quantitative proteomic techniques may contribute to better therapeutic selection in AML. Expert commentary: The net functional state of the cell is defined by the activity of protein within all the pathways that are active in the cell. Recognition of the proteomic profile of the leukemic blast could, therefore, complement current classification systems by providing a better a priori description of what pathways are important within a cell as a guide to the selection of therapy for the patient.

The majority of RUNX1 mutations in acute myeloid leukemia (AML) are missense or deletion-truncation and behave as loss-of-function mutations. Following standard therapy, AML patients expressing mtRUNX1 exhibit inferior clinical outcome than those without mutant RUNX1. Studies presented here demonstrate that as compared with AML cells lacking mtRUNX1, their isogenic counterparts harboring mtRUNX1 display impaired ribosomal biogenesis and differentiation, as well as exhibit reduced levels of wild-type RUNX1, PU.1, and c-Myc. Compared with AML cells with only wild-type RUNX1, AML cells expressing mtRUNX1 were also more sensitive to the protein translation inhibitor homoharringtonine (omacetaxine) and BCL2 inhibitor venetoclax. Homoharringtonine treatment repressed enhancers and their BRD4 occupancy and was associated with reduced levels of c-Myc, c-Myb, MCL1, and Bcl-xL. Consistent with this, cotreatment with omacetaxine and venetoclax or BET inhibitor induced synergistic in vitro lethality in AML expressing mtRUNX1. Compared with each agent alone, cotreatment with omacetaxine and venetoclax or BET inhibitor also displayed improved in vivo anti-AML efficacy, associated with improved survival of immune-depleted mice engrafted with AML cells harboring mtRUNX1. These findings highlight superior efficacy of omacetaxine-based combination therapies for AML harboring mtRUNX1.

INTRODUCTION: Acute leukemia results from a series of mutational events that alter cell growth and proliferation. Mutations result in protein changes that orchestrate growth alterations characteristic of leukemia. Proteomics is a methodology appropriate for study of protein changes found in leukemia. The high-throughput reverse phase protein array (RPPA) technology is particularly well-suited for the assessment of protein changes in samples derived from clinical trials. AREAS COVERED: This review discusses the technical, methodological, and analytical issues related to the successful development of acute leukemia RPPAs. EXPERT COMMENTARY: To obtain representative protein sample lysates, samples should be prepared from freshly collected blood or bone marrow material. Variables such as sample shipment, transit time, and holding temperature only have minimal effects on protein expression. CellSave preservation tubes are preferred for cells collected after exposure to chemotherapy, and incorporation of standardized guidelines for antibody validation is recommended. A more systematic biological approach to analyze protein expression is desired, searching for recurrent patterns of protein expression that allow classification of patients into risk groups, or groups of patients that may be treated similarly. Comparing RPPA protein analysis between cell lines and primary samples shows that cell lines are not representative of patient proteomic patterns.

Acute promyelocytic leukemia (APL) is characterized by the chromosomal translocation t(15;17) and the resulting gene PML-RARA, used for measurable residual disease (MRD) monitoring. Despite highly effective therapy for APL, MRD monitoring practices are not fully established. We aimed to assess the value of MRD monitoring by RT-qPCR in patients with APL treated with ATRA and arsenic trioxide +/- GO. We reviewed 223 patients with APL treated with this regimen. RT-qPCR for PML-RARA was measured every 3 months, and at 12, 18, and 24 months after therapy. Seven patients relapsed. Time to relapse was 7.9-12.4 months in 6 patients, and one patient relapsed after 79.5 months. These data show that MRD monitoring may be important for the detection of relapse in patients treated with this regimen within one year after completing therapy, however, since late molecular relapse is rare, our data suggest a low value of MRD monitoring beyond that first year.

Purpose: The addition of the proteasome inhibitor (PI) bortezomib to standard chemotherapy (ADE: cytarabine [Ara-C], daunorubicin, and etoposide) did not improve overall outcome of pediatric AML patients in the Children's Oncology Group AAML1031 phase 3 randomized clinical trial (AAML1031). Bortezomib prevents protein degradation, including RelA via the intracellular NF-kB pathway. In this study, we hypothesized that subgroups of pediatric AML patients benefitting from standard therapy plus bortezomib (ADEB) could be identified based on pre-treatment RelA expression and phosphorylation status.Experimental design: RelA-total and phosphorylation at serine 536 (RelA-pSer536) were measured in 483 patient samples using reverse phase protein array technology.Results: In ADEB-treated patients, low-RelA-pSer536 was favorably prognostic when compared to high-RelA-pSer536 (3-yr overall survival (OS): 81% vs. 68%, p = 0.032; relapse risk (RR): 30% vs. 49%, p = 0.004). Among low-RelA-pSer536 patients, RR significantly decreased with ADEB compared to ADE (RR: 30% vs. 44%, p = 0.035). Correlation between RelA-pSer536 and 295 other assayed proteins identified a strong correlation with HSF1-pSer326, another protein previously identified as modifying ADEB response. The combination of low-RelA-pSer536 and low-HSF1-pSer326 was a significant predictor of ADEB response (3-yr OS: 86% vs. 67%, p = 0.013).Conclusion and clinical relevance: Bortezomib may improve clinical outcome in a subgroup of AML patients identified by low-RelA-pSer536 and low-HSF1-pSer326.

Acute myeloid leukemia (AML) is a heterogeneous disease with variable responses to therapy. Cytogenetic and genomic features are used to classify AML patients into prognostic and treatment groups. However, these molecular characteristics harbor significant patient-to-patient variability and do not fully account for AML heterogeneity. RNA-based classifications have also been applied in AML as an alternative approach, but transcriptomic grouping is strongly associated with AML morphologic lineages. We used a training cohort of newly diagnosed AML patients and conducted unsupervised RNA-based classification after excluding lineage-associated genes. We identified three AML patient groups that have distinct biological pathways associated with outcomes. Enrichment of inflammatory pathways and downregulation of HOX pathways were associated with improved outcomes, and this was validated in 2 independent cohorts. We also identified a group of AML patients who harbored high metabolic and mTOR pathway activity, and this was associated with worse clinical outcomes. Using a comprehensive reverse phase protein array, we identified higher mTOR protein expression in the highly metabolic group. We also identified a positive correlation between degree of resistance to venetoclax and mTOR activation in myeloid and lymphoid cell lines. Our approach of integrating RNA, protein, and genomic data uncovered lineage-independent AML patient groups that share biologic mechanisms and can inform outcomes independent of commonly used clinical and demographic variables; these groups could be used to guide therapeutic strategies.

Summary Background Addition of the BCL2 inhibitor venetoclax to lower intensity therapy has been shown to improve overall survival in older (aged 75 years or older) and unfit patients with newly diagnosed acute myeloid leukaemia. The aim of this study was to investigate the activity of venetoclax combined with intensive chemotherapy in patients aged 65 years or younger with acute myeloid leukaemia. Methods This cohort study was done at the MD Anderson Cancer Center in the USA, as part of the single-centre, single arm, phase 2, CLIA trial. Here we report on the independent cohort investigating the safety and activity of venetoclax added to intensive chemotherapy (the CLIA regimen [cladribine, high-dose cytarabine, idarubicin]). Eligible patients were aged 18–65 years with a new diagnosis of acute myeloid leukaemia, mixed phenotype acute leukaemia, or high-risk myelodysplastic syndrome (≥10% blasts or International Prognostic Scoring System ≥2 [intermediate]), who received no previous potentially curative therapy for leukaemia. Patients received cladribine (5 mg/m2) and cytarabine (1·5 g/m2 for patients aged Findings Between Feb 25, 2019, and March 23, 2021, 77 patients were assessed for eligibility, 50 of whom were enrolled. Median age was 48 years (IQR 37–56). 47 (94% [95% CI 83–98]) patients had composite complete response, with the same proportion also having an overall response; two (4% [1–14]) patients did not respond, and one (2% [0–11]) patient died during induction. 37 (82% [95% CI 68–92]) of 45 patients had undetectable measurable residual disease (MRD). At a median follow-up of 13·5 months (IQR 6·4–19·5), the median duration of response, event-free survival, and overall survival were not reached. At 12 months, the estimated duration of response was 74% (95% CI 60–92), event-free survival was 68% (54–85), and overall survival was 85% (75–97). The most common adverse events of grade 3 or worse were febrile neutropenia (42 [84%] patients), infection (six [12%]), and alanine aminotransferase elevations (six [12%]). There was one death during induction in a patient treated with CLIA–venetoclax plus a FLT3 inhibitor. Two patients died of infectious complications while in complete response in consolidation cycles, both of whom had FLT3-mutated acute myeloid leukaemia and were receiving combined therapy with a FLT3 inhibitor. No deaths were deemed to be treatment related. Interpretation Venetoclax added to CLIA was safe and active in patients with newly diagnosed acute myeloid leukaemia or high-risk myelodysplastic syndrome, producing high rates of durable MRD-negative remissions and encouraging event-free survival and overall survival. Funding MD Anderson Cancer Center.

Pediatric acute myeloid leukemia (AML) remains a fatal disease for at least 30% of patients, stressing the need for improved therapies and better risk stratification. As proteins are the unifying feature of (epi)genetic and environmental alterations, and are often targeted by novel chemotherapeutic agents, we studied the proteomic landscape of pediatric AML. Protein expression and activation levels were measured in 500 bulk leukemic patients' samples and 30 control CD34+ cell samples, using reverse phase protein arrays with 296 strictly validated antibodies. The multistep MetaGalaxy analysis methodology was applied and identified nine protein expression signatures (PrSIG), based on strong recurrent protein expression patterns. PrSIG were associated with cytogenetics and mutational state, and with favorable or unfavorable prognosis. Analysis based on treatment (i.e., ADE vs. ADE plus bortezomib) identified three PrSIG that did better with ADE plus bortezomib than with ADE alone. When PrSIG were studied in the context of cytogenetic risk groups, PrSIG were independently prognostic after multivariate analysis, suggesting a potential value for proteomics in combination with current classification systems. Proteins with universally increased (n=7) or decreased (n=17) expression were observed across PrSIG. Certain proteins significantly differentially expressed from normal could be identified, forming a hypothetical platform for personalized medicine.

Background: Measurable residual disease (MRD) by multiparameter flow cytometry (MFC) is highly prognostic for relapse and overall survival (OS) in patients (pts) with acute lymphoblastic leukemia (ALL). However, many pts with apparent "MRD negativity" by MFC still relapse. These relapses are likely due to residual leukemia that is present below the level of detection of MFC (generally ~1x10-4). In light of the limitations of conventional MRD assays, we sought to evaluate the clinical impact of a highly sensitive next-generation sequencing (NGS)-based MRD assay that is capable of detecting residual leukemia at a level of 1x10-6. Methods: We performed a retrospective study of 67 pts with previously untreated Philadelphia chromosome-negative ALL who received frontline therapy between 2/2012 and 7/2018 with a hyper-CVAD-based regimen (n=44) or a lower-intensity hyper-CVD-based regimen (n=23), which was usually combined with inotuzumab ozogamicin, and achieved complete remission (CR). Pts were selected for this study based on availability of banked samples at baseline and at sequential remission time points up to 4.5 months after start of therapy. Six-color MFC was performed as standard of care with an analytic sensitivity of 0.01% (1x10-4). Pretreatment genomic DNA derived from bone marrow was evaluated using a quantitative NGS MRD assay (Adaptive Biotechnologies Co., Seattle, WA) to identify dominant rearrangements within the B-cell receptor (IgH [VDJ and DJ], IgK and IgL). High throughput NGS of remission samples was subsequently performed to assess the target clonal rearrangements at an established analytic sensitivity of 0.0001% (1x10-6). Results: The baseline characteristics of the study population are shown in Table 1. Twenty pts (30%) underwent allogeneic stem cell transplant (alloSCT) in first remission. In the entire cohort, 14 pts relapsed (21%), including 2 with CNS-only relapses. The median duration of follow-up for the entire cohort was 51 months. Among the 109 remission samples, 84 were MRD-negative (MRDneg), 24 were MRD-positive (MRDpos), and 1 was indeterminate by MFC (Figure 1). All 24 MRDpos samples by MFC were also MRDpos by NGS. Among the 84 MRDneg samples by MFC, 32 (38%) were MRDpos by NGS. For these discordant cases, the median level of MRD by NGS was below the expected level of detection by MFC (median 0.002% [range 0.00002%-1.15%]). At the time of CR, 44/67 pts (66%), achieved MRDneg by MFC and 14/38 (37%) achieved MRDneg by NGS. The rates of MRDneg by MFC in pts treated with hyper-CVAD or hyper-CVD regimens were 61% and 74%, respectively, and by NGS were 32% and 46%, respectively. Achieving MRDneg by NGS at the time of CR better predicted the likelihood of durable remission than did achieving MRDneg by MFC. The 5-year cumulative incidence of relapse (CIR) of the 44 pts who achieved MRDneg by MFC and of 14 pts who achieved MRDneg by NGS was 27% and 8%, respectively. Only 1 out of 14 pts who achieved MRDneg by NGS at the time of CR subsequently relapsed. In contrast, MRD status by NGS did not predict for relapse when evaluated at post-CR time points. Among pts treated with a more intensive hyper-CVAD regimen, achievement of MRDneg by NGS at CR provided additional prognostic information compared to MRD status by MFC at the same time point and identified pts with an excellent long-term outcome. The 5-year OS rate for pts who achieved MRDneg by NGS at CR was 100% vs. 55% for those who were MRDpos by NGS (Figure 2A; P Conclusion: Early assessment of MRD using an ultrasensitive NGS assay can identify pts with ALL who have a very low risk of relapse and excellent long-term survival. Prospective studies are warranted to assess whether de-intensification of therapy is feasible for this favorable-risk group of pts who rapidly achieve MRDneg remission. Disclosures Short: Amgen: Honoraria; Astellas: Research Funding; AstraZeneca: Consultancy; Takeda Oncology: Consultancy, Honoraria, Research Funding. Kantarjian:Takeda: Honoraria; Amgen: Honoraria, Research Funding; Actinium: Honoraria, Membership on an entity's Board of Directors or advisory committees; Cyclacel: Research Funding; Pfizer: Honoraria, Research Funding; Agios: Honoraria, Research Funding; Novartis: Research Funding; Immunogen: Research Funding; AbbVie: Honoraria, Research Funding; BMS: Research Funding; Ariad: Research Funding; Jazz Pharma: Research Funding; Daiichi-Sankyo: Research Funding; Astex: Research Funding. Konopleva:F. Hoffmann La-Roche: Consultancy, Research Funding; Eli Lilly: Research Funding; Cellectis: Research Funding; Calithera: Research Funding; Rafael Pharmaceutical: Research Funding; Amgen: Consultancy; Sanofi: Research Funding; Stemline Therapeutics: Consultancy, Research Funding; Ablynx: Research Funding; Agios: Research Funding; AbbVie: Consultancy, Research Funding; Ascentage: Research Funding; AstraZeneca: Research Funding; Forty-Seven: Consultancy, Research Funding; Reata Pharmaceutical Inc.;: Patents & Royalties: patents and royalties with patent US 7,795,305 B2 on CDDO-compounds and combination therapies, licensed to Reata Pharmaceutical; Kisoji: Consultancy; Genentech: Consultancy, Research Funding. Jain:Cellectis: Research Funding; BMS: Research Funding; BeiGene: Honoraria, Membership on an entity's Board of Directors or advisory committees; Verastem: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Fate Therapeutics: Research Funding; Servier: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Pfizer: Research Funding; Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees; Aprea Therapeutics: Research Funding; Pharmacyclics: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Incyte: Research Funding; Genentech: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; AbbVie: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; ADC Therapeutics: Research Funding; TG Therapeutics: Honoraria, Membership on an entity's Board of Directors or advisory committees; Precision Bioscienes: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Adaptive Biotechnologies: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; AstraZeneca: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding. Kebriaei:Kite: Other: Served on advisory board; Amgen: Other: Research Support; Ziopharm: Other: Research Support; Pfizer: Other: Served on advisory board; Novartis: Other: Served on advisory board; Jazz: Consultancy. Yilmaz:Pfizer: Research Funding; Daicho Sankyo: Research Funding; Pint Pharma: Honoraria. Issa:Celegene: Research Funding; Syndax: Research Funding; Novartis: Membership on an entity's Board of Directors or advisory committees. Ravandi:BMS: Consultancy, Honoraria, Research Funding; Abbvie: Consultancy, Honoraria, Research Funding; Amgen: Consultancy, Honoraria, Research Funding; Astellas: Consultancy, Honoraria, Research Funding; Xencor: Consultancy, Honoraria, Research Funding; Macrogenics: Research Funding; Jazz Pharmaceuticals: Consultancy, Honoraria, Research Funding; AstraZeneca: Consultancy, Honoraria; Celgene: Consultancy, Honoraria; Orsenix: Consultancy, Honoraria, Research Funding. Jabbour:Amgen: Other: Advisory role, Research Funding; Adaptive Biotechnologies: Other: Advisory role, Research Funding; BMS: Other: Advisory role, Research Funding; Genentech: Other: Advisory role, Research Funding; AbbVie: Other: Advisory role, Research Funding; Takeda: Other: Advisory role, Research Funding; Pfizer: Other: Advisory role, Research Funding.

Background Addition of the BCL2 inhibitor venetoclax to lower intensity therapy improved survival (OS) for older and unfit patients with newly diagnosed AML. We have previously reported the safety and efficacy of the nucleoside analogue cladribine combined with high dose ara-C and idarubicin (ida) as an intensive induction regimen (CLIA) in patients with newly diagnosed AML. Here, we report the results of the combination of venetoclax with the CLIA regimen for younger, fit patients with newly diagnosed AML. Methods Patients < 65 years of age with newly diagnosed AML or high-risk MDS were enrolled (NCT02115295). Induction was cladribine 5 mg/m2 IV on D1-5, ara-C 1.5 g/m2 IV on D1-5, and idarubicin 10 mg/m2 IV on D1-3; venetoclax was given at an effective dose of 400mg PO on D2-8 without ramp up with every cycle. Consolidation consisted of 3d of CLIA (2d of ida). AML patients with FLT3-ITD or FLT3 point mutations could receive midostaurin on D6-19 or gilteritinib on D1-14 for induction, and continuously thereafter. Venetoclax dose modifications for CYP3A4 inhibitors were made. All patients underwent baseline next generation sequencing and MRD testing by multiparameter flow cytometry at the time of response. Results A total of 31 patients have been enrolled. The median age was 48 years (range, 18-64). Baseline characteristics are shown in Table 1. Median follow up is 8.8 months. All patients were evaluable for response, with 28 of 31 (90%) achieving a remission, including 23 (74%) complete remission (CR) and 5 (16%) CR with incomplete count recovery (CRi) (Table 1). Of the 2 patients that did not respond, one was ELN adverse risk with TP53 mutation and complex karyotype, the other was ELN favorable risk with a t(8;21) translocation and mutations in IDH1 and JAK2. At time of initial response assessment 19 of 30 (63%) had undetectable MRD, with another 4 patients (77%) becoming MRD negative overall. Median cycles until response was 1 (range: 1 - 2), and patients have received a median of 2 cycles (range: 1 - 5). Median time to absolute neutrophil count ≥ 1000/µL was 32 days (range: 18 - 49) and platelet count ≥ 50k and 100k/µL were 29 days (range: 17 - 61) and 32 days, respectively after induction. 8 patients received FLT3 inhibition (FLT3i) in addition to CLIA and venetoclax, 7 (88%) with gilteritinib and 1 (12%) with midostaurin. Within this subset, there were 6/8 (75%) CR and 1 CR (12%) (Table 2). Median overall survival (OS) for those receiving FLT3i is not yet reached (95% CI: 0.79 - NE months). Median duration of response for the entire cohort is not reached. At 6 and 12 months 87% of responders had an ongoing response (SE=9%). 18 of 28 (64%) of responders received an allogeneic stem cell transplant. Median OS has not been reached, with 6- and 12-month rates of OS 93% (SE=5%) and 81% (SE=9%), respectively (Figure 1). Median OS in all ELN risk categories has not yet been reached, with 6 month OS of 91% (SE=9%), 100% (SE=NA), and 83% (SE=15%) in favorable, intermediate, and adverse risk groups, respectively (Figure 2). Median event free survival (EFS) has not been reached, with 6- and 12-month rates of EFS are 90% (SE=5%) and 79% (SE=9%), respectively. Overall the regimen was well tolerated, with an acceptable toxicity profile. Tumor lysis syndrome was not seen. The most common Grade >/= 3 adverse events were neutropenic fever, pneumonia, nausea, and liver transaminitis. There was one induction death in the FLT3i cohort and none with CLIA + venetoclax. Conclusion Venetoclax added to CLIA was safe and highly effective in newly diagnosed pts with AML and high risk MDS, including patients with FLT3-mutated AML and adverse risk by ELN or IPSS-R. The addition of FLT3i may be associated with delayed count recovery. CLIA plus venetoclax provided high rates of durable MRD negative remissions and encouraging event-free and overall survival across prognostic subgroups. Disclosures Kantarjian: BMS: Research Funding; Actinium: Honoraria, Membership on an entity's Board of Directors or advisory committees; Adaptive biotechnologies: Honoraria; Aptitute Health: Honoraria; BioAscend: Honoraria; Daiichi-Sankyo: Honoraria, Research Funding; Immunogen: Research Funding; Jazz: Research Funding; Novartis: Honoraria, Research Funding; Pfizer: Honoraria, Research Funding; Sanofi: Research Funding; Delta Fly: Honoraria; Ascentage: Research Funding; Janssen: Honoraria; Abbvie: Honoraria, Research Funding; Amgen: Honoraria, Research Funding; Oxford Biomedical: Honoraria. Borthakur:Abbvie: Research Funding; BioLine Rx: Consultancy; FTC Therapeutics: Consultancy; BioTherix: Consultancy; Jannsen: Research Funding; GSK: Research Funding; Nkarta Therapeutics: Consultancy; Treadwell Therapeutics: Consultancy; Argenx: Consultancy; AstraZeneca: Research Funding; Cyclacel: Research Funding; PTC Therapeutics: Consultancy; PTC Therapeutics: Research Funding; Novartis: Research Funding; BioLine Rx: Research Funding; BMS: Research Funding; Incyte: Research Funding; Curio Science LLC: Consultancy; Oncoceutics: Research Funding; Xbiotech USA: Research Funding; Polaris: Research Funding. Yilmaz:Daicho Sankyo: Research Funding; Pint Pharma: Honoraria; Pfizer: Research Funding. DiNardo:ImmuneOnc: Honoraria; AbbVie: Consultancy, Honoraria, Research Funding; Agios: Consultancy, Honoraria, Research Funding; Celgene: Consultancy, Honoraria, Research Funding; Calithera: Research Funding; Daiichi Sankyo: Consultancy, Honoraria, Research Funding; Notable Labs: Membership on an entity's Board of Directors or advisory committees; Takeda: Honoraria; Jazz: Honoraria; MedImmune: Honoraria; Syros: Honoraria; Novartis: Consultancy. Andreeff:Centre for Drug Research & Development; Cancer UK; NCI-CTEP; German Research Council; Leukemia Lymphoma Foundation (LLS); NCI-RDCRN (Rare Disease Clin Network); CLL Founcdation; BioLineRx; SentiBio; Aptose Biosciences, Inc: Membership on an entity's Board of Directors or advisory committees; Amgen: Research Funding; Daiichi-Sankyo; Breast Cancer Research Foundation; CPRIT; NIH/NCI; Amgen; AstraZeneca: Research Funding; Daiichi-Sankyo; Jazz Pharmaceuticals; Celgene; Amgen; AstraZeneca; 6 Dimensions Capital: Consultancy. Daver:Bristol-Myers Squibb: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Pfizer: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Karyopharm: Research Funding; Servier: Research Funding; Genentech: Research Funding; AbbVie: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Astellas: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Novimmune: Research Funding; Gilead: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Amgen: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Trovagene: Research Funding; Fate Therapeutics: Research Funding; ImmunoGen: Research Funding; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees; Celgene: Consultancy, Membership on an entity's Board of Directors or advisory committees; Jazz: Consultancy, Membership on an entity's Board of Directors or advisory committees; Trillium: Consultancy, Membership on an entity's Board of Directors or advisory committees; Syndax: Consultancy, Membership on an entity's Board of Directors or advisory committees; Amgen: Consultancy, Membership on an entity's Board of Directors or advisory committees; KITE: Consultancy, Membership on an entity's Board of Directors or advisory committees; Agios: 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. Jain:Aprea Therapeutics: Research Funding; Servier: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees; Pharmacyclics: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Fate Therapeutics: Research Funding; BeiGene: Honoraria, Membership on an entity's Board of Directors or advisory committees; Adaptive Biotechnologies: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; BMS: Research Funding; Pfizer: Research Funding; AstraZeneca: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; ADC Therapeutics: Research Funding; TG Therapeutics: Honoraria, Membership on an entity's Board of Directors or advisory committees; Incyte: Research Funding; Cellectis: Research Funding; Verastem: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Genentech: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; AbbVie: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Precision Bioscienes: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding. Jabbour:Takeda: Other: Advisory role, Research Funding; Genentech: Other: Advisory role, Research Funding; Pfizer: Other: Advisory role, Research Funding; BMS: Other: Advisory role, Research Funding; Amgen: Other: Advisory role, Research Funding; Adaptive Biotechnologies: Other: Advisory role, Research Funding; AbbVie: Other: Advisory role, Research Funding. Short:Astellas: Research Funding; Amgen: Honoraria; AstraZeneca: Consultancy; Takeda Oncology: Consultancy, Honoraria, Research Funding. Alvarado:Daiichi-Sankyo: Research Funding; BerGenBio ASA: Research Funding; Tolero Pharmaceuticals: Research Funding; Astex Pharmaceuticals: Research Funding; Jazz Pharmaceuticals: Research Funding; FibroGen: Research Funding; MEI Pharma: Research Funding; Sun Pharma: Research Funding. Bose:Constellation Pharmaceuticals: Research Funding; Celgene Corporation: Honoraria, Research Funding; CTI BioPharma: Honoraria, Research Funding; Blueprint Medicines Corporation: Honoraria, Research Funding; Pfizer, Inc.: Research Funding; Astellas Pharmaceuticals: Research Funding; NS Pharma: Research Funding; Promedior, Inc.: Research Funding; Incyte Corporation: Consultancy, Honoraria, Research Funding, Speakers Bureau; Kartos Therapeutics: Honoraria, Research Funding. Garcia-Manero:Merck: Research Funding; Bristol-Myers Squibb: Consultancy, Research Funding; AbbVie: Honoraria, Research Funding; H3 Biomedicine: Research Funding; Amphivena Therapeutics: Research Funding; Genentech: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Acceleron Pharmaceuticals: Consultancy, Honoraria; Onconova: Research Funding; Novartis: Research Funding; Astex Pharmaceuticals: Consultancy, Honoraria, Research Funding; Helsinn Therapeutics: Consultancy, Honoraria, Research Funding; Jazz Pharmaceuticals: Consultancy; Celgene: Consultancy, Honoraria, Research Funding. Konopleva:Sanofi: Research Funding; Stemline Therapeutics: Consultancy, Research Funding; Ablynx: Research Funding; Genentech: Consultancy, Research Funding; Ascentage: Research Funding; Amgen: Consultancy; Agios: Research Funding; AstraZeneca: Research Funding; Eli Lilly: Research Funding; F. Hoffmann La-Roche: Consultancy, Research Funding; Rafael Pharmaceutical: Research Funding; Cellectis: Research Funding; Calithera: Research Funding; Kisoji: Consultancy; AbbVie: Consultancy, Research Funding; Reata Pharmaceutical Inc.;: Patents & Royalties: patents and royalties with patent US 7,795,305 B2 on CDDO-compounds and combination therapies, licensed to Reata Pharmaceutical; Forty-Seven: Consultancy, Research Funding. Ravandi:Jazz Pharmaceuticals: Consultancy, Honoraria, Research Funding; Macrogenics: Research Funding; AstraZeneca: Consultancy, Honoraria; Xencor: Consultancy, Honoraria, Research Funding; Celgene: Consultancy, Honoraria; Astellas: Consultancy, Honoraria, Research Funding; Amgen: Consultancy, Honoraria, Research Funding; BMS: Consultancy, Honoraria, Research Funding; Abbvie: Consultancy, Honoraria, Research Funding; Orsenix: Consultancy, Honoraria, Research Funding. Kadia:Amgen: Research Funding; Cellenkos: Research Funding; Cyclacel: Research Funding; Genentech: Honoraria, Research Funding; Abbvie: Honoraria, Research Funding; Novartis: Honoraria; BMS: Honoraria, Research Funding; Astra Zeneca: Research Funding; Celgene: Research Funding; Ascentage: Research Funding; Astellas: Research Funding; Pulmotec: Research Funding; Incyte: Research Funding; Pfizer: Honoraria, Research Funding; JAZZ: Honoraria, Research Funding.

Background: For patients (pts) with Philadelphia chromosome (Ph)-negative B-cell acute lymphoblastic leukemia (ALL), combination chemotherapy achieves complete remission rates of 80-90%; however, many pts ultimately relapse, leading to a cure rate of only 40-50%. Blinatumomab is highly effective in both the relapsed/refractory setting and for eradication of measurable residual disease (MRD). We hypothesized that early incorporation of blinatumomab in pts with newly diagnosed Ph-negative B-cell ALL would decrease the need for intensive chemotherapy, lead to deeper and more durable responses, and improve survival. Methods: Pts 14-59 years of age with newly diagnosed Ph-negative pre-B-cell ALL, including pts who had received no more than 1 prior cycle of chemotherapy, were eligible. Pts were required to have a performance status of ≤3, total bilirubin ≤2 mg/dl and creatinine ≤2 mg/dl. Pts received hyper-CVAD alternating with high-dose methotrexate and cytarabine for up to 4 cycles, followed by 4 cycles of blinatumomab at standard doses. Pts with CD20+ disease (≥1% cells) received 8 doses of ofatumumab (2000 mg) or rituximab (375 mg/m2). Eight administrations of prophylactic IT chemotherapy were given in the first 4 cycles. Maintenance was with alternating blocks of POMP (given in maintenance cycles 1-3, 5-7, 9-11, and 13-15) and blinatumomab (given in maintenance cycles 4, 8, and 12). After 2 pts with high-risk features experienced early relapse prior to blinatumomab administration, for pts #10+ the protocol was amended so that pts with high-risk disease features (e.g. CRLF2+ by flow cytometry, complex karyotype, KMT2A rearranged, low-hypodiploidy/near triploidy, TP53 mutation, or persistent MRD) started blinatumomab after 2 cycles of hyper-CVAD. Results: 39 pts have been treated, 34 of whom are evaluable for efficacy (5 too early for assessment). 6 pts were in complete remission (CR) at enrollment and unevaluable for morphologic response. Pt characteristics of the 34 evaluable pts are summarized in Table 1. Median age was 36 years (range, 17-59 years). At least one high-risk feature was present in 19 pts (56%), including TP53 mutation in 27%, CRLF2+ in 20%, and an adverse-risk karyotype in 26%. 82% of pts received ofatumumab or rituximab. Among 28 pts with active disease at study entry, 100% achieved CR, with 82% achieving CR after the first cycle. MRD negativity by 6-color flow cytometry was achieved in 20/23 responding pts (87%) after 1 cycle and 33/34 pts (97%) overall. There were no early deaths, and the 60-day mortality rate was 0%. With a median follow-up of 22 months (range, 1-40 months), the 2-year continuous remission and OS rates were 79% and 86%, respectively (Figure 1). Overall, 5 pts (15%) relapsed, 12 (35%) underwent allogeneic SCT in first remission (including 1 additional pt who relapsed post-SCT), and 17 (50%) remain in continuous remission and are currently on treatment or have completed maintenance. All relapses occurred in pts with established poor-risk features, including 2 pts with KMT2A rearrangement, 2 pts with TP53 mutation (1 of whom was low hypodiploid), and 1 pt with baseline WBC count of 32 x 109/L. Two of these relapses occurred during hyper-CVAD cycles before the amendment allowing for earlier integration of blinatumomab for pts with high-risk disease features. Treatment was overall well-tolerated. Four pts developed grade 2-3 cytokine release syndrome (grade 2, n=3; grade 3, n=1) which resolved with corticosteroids and interruption of blinatumomab. Overall, 14 (41%) pts had a neurological AE of any grade due to blinatumomab. Only one pt discontinued blinatumomab due to blinatumomab-related adverse event (grade 2 encephalopathy and dysphasia). Conclusion: Sequential combination of hyper-CVAD and blinatumomab is highly effective as frontline treatment of Ph-negative B-cell ALL, with a CR rate of 100% and 97% of pts achieving MRD negativity. Survival data are promising with an estimated 2-year OS of 86%, which compares favorably to historical controls. This study continues to accrue pts. Disclosures Short: Astellas: Research Funding; Amgen: Honoraria; AstraZeneca: Consultancy; Takeda Oncology: Consultancy, Honoraria, Research Funding. Kantarjian:Jazz: Research Funding; Novartis: Honoraria, Research Funding; Pfizer: Honoraria, Research Funding; Daiichi-Sankyo: Honoraria, Research Funding; Oxford Biomedical: Honoraria; Aptitute Health: Honoraria; BioAscend: Honoraria; Delta Fly: Honoraria; Janssen: Honoraria; Actinium: Honoraria, Membership on an entity's Board of Directors or advisory committees; Immunogen: Research Funding; BMS: Research Funding; Adaptive biotechnologies: Honoraria; Abbvie: Honoraria, Research Funding; Sanofi: Research Funding; Amgen: Honoraria, Research Funding; Ascentage: Research Funding. Ravandi:Astellas: Consultancy, Honoraria, Research Funding; Amgen: Consultancy, Honoraria, Research Funding; Macrogenics: Research Funding; AstraZeneca: Consultancy, Honoraria; Jazz Pharmaceuticals: Consultancy, Honoraria, Research Funding; Xencor: Consultancy, Honoraria, Research Funding; Orsenix: Consultancy, Honoraria, Research Funding; Abbvie: Consultancy, Honoraria, Research Funding; Celgene: Consultancy, Honoraria; BMS: Consultancy, Honoraria, Research Funding. Kadia:Abbvie: Honoraria, Research Funding; BMS: Honoraria, Research Funding; Cyclacel: Research Funding; Astellas: Research Funding; Cellenkos: Research Funding; Pfizer: Honoraria, Research Funding; Celgene: Research Funding; Pulmotec: Research Funding; Amgen: Research Funding; Genentech: Honoraria, Research Funding; Ascentage: Research Funding; Incyte: Research Funding; JAZZ: Honoraria, Research Funding; Novartis: Honoraria; Astra Zeneca: Research Funding. Thompson:Pharmacyclics: Research Funding; Adaptive Biotechnologies: Consultancy, Research Funding; Genentech: Consultancy; Janssen-Cilag: Honoraria; AbbVie: Research Funding. Alvarado:Tolero Pharmaceuticals: Research Funding; Daiichi-Sankyo: Research Funding; Astex Pharmaceuticals: Research Funding; Jazz Pharmaceuticals: Research Funding; Sun Pharma: Research Funding; BerGenBio ASA: Research Funding; MEI Pharma: Research Funding; FibroGen: Research Funding. Jain:Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees; Precision Bioscienes: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Fate Therapeutics: Research Funding; AstraZeneca: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Pharmacyclics: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; AbbVie: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Genentech: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; BMS: Research Funding; Pfizer: Research Funding; ADC Therapeutics: Research Funding; Incyte: Research Funding; Servier: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Cellectis: Research Funding; Verastem: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Adaptive Biotechnologies: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; TG Therapeutics: Honoraria, Membership on an entity's Board of Directors or advisory committees; BeiGene: Honoraria, Membership on an entity's Board of Directors or advisory committees; Aprea Therapeutics: Research Funding. Yilmaz:Pint Pharma: Honoraria; Pfizer: Research Funding; Daicho Sankyo: Research Funding. Konopleva:Calithera: Research Funding; Genentech: Consultancy, Research Funding; Amgen: Consultancy; Reata Pharmaceutical Inc.;: Patents & Royalties: patents and royalties with patent US 7,795,305 B2 on CDDO-compounds and combination therapies, licensed to Reata Pharmaceutical; Agios: Research Funding; AstraZeneca: Research Funding; Cellectis: Research Funding; Stemline Therapeutics: Consultancy, Research Funding; Forty-Seven: Consultancy, Research Funding; Rafael Pharmaceutical: Research Funding; Eli Lilly: Research Funding; AbbVie: Consultancy, Research Funding; Ascentage: Research Funding; F. Hoffmann La-Roche: Consultancy, Research Funding; Kisoji: Consultancy; Sanofi: Research Funding; Ablynx: Research Funding. Garcia-Manero:AbbVie: Honoraria, Research Funding; Celgene: Consultancy, Honoraria, Research Funding; Bristol-Myers Squibb: Consultancy, Research Funding; Astex Pharmaceuticals: Consultancy, Honoraria, Research Funding; Acceleron Pharmaceuticals: Consultancy, Honoraria; Amphivena Therapeutics: Research Funding; Jazz Pharmaceuticals: Consultancy; Merck: Research Funding; H3 Biomedicine: Research Funding; Helsinn Therapeutics: Consultancy, Honoraria, Research Funding; Novartis: Research Funding; Genentech: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Onconova: Research Funding. O'Brien:Amgen, Astellas, Celgene, GlaxoSmithKline, Janssen Oncology, Aptose Biosciences Inc. Vaniam Group, AbbVie, Alexion, Verastem, Eisai, Juno Therapeutics, Vida Ventures: Consultancy; Kite, Regeneron, Acerta: Research Funding; Gilead, Pharmacyclics, TG Therapeutics, Pfizer, Sunesis: Consultancy, Research Funding. Jabbour:Takeda: Other: Advisory role, Research Funding; AbbVie: Other: Advisory role, Research Funding; Pfizer: Other: Advisory role, Research Funding; Genentech: Other: Advisory role, Research Funding; BMS: Other: Advisory role, Research Funding; Adaptive Biotechnologies: Other: Advisory role, Research Funding; Amgen: Other: Advisory role, Research Funding. OffLabel Disclosure: Blinatumomab - frontline therapy for ALL

Background: Inotuzumab ozogamicin (INO) and blinatumomab both improve overall survival (OS) in relapsed or refractory B-cell acute lymphoblastic leukemia (ALL). Use of these effective monoclonal antibodies in the frontline setting may lead to deep and durable remissions in older adults with newly diagnosed Philadelphia chromosome (Ph)-negative B-cell ALL. Methods: Patients (pts) ≥60 years of age with newly diagnosed Ph-negative pre-B-cell ALL, including pts who had received no more than 1 prior cycle of chemotherapy, were eligible. Pts were required to have a performance status of ≤3, total bilirubin ≤1.5 mg/dl, AST/ALT ≤3x ULN and creatinine ≤2 mg/dl. Pts received mini-hyper-CVD (cyclophosphamide and dexamethasone at 50% dose reduction, no anthracycline, methotrexate at 75% dose reduction, cytarabine at 0.5 g/m2 x 4 doses) for up to 8 cycles. INO was initially given at a dose of 1.3-1.8mg/m2 on day 3 of cycle 1 and 0.8-1.3mg/m2 on day 3 of cycles 2-4. Rituximab (if CD20+) and prophylactic IT chemotherapy were given for the first 4 cycles. Responding pts received POMP maintenance for up to 3 years. In order to decrease the risk of veno-occlusive disease (VOD), the protocol was amended in 3/2017 (pts 50+) to give INO in fractionated doses each cycle (0.6 mg/m2 on day 2 and 0.3 mg/m2 on day 8 of cycle 1; 0.3 mg/m2 on day 2 and 8 of cycles 2-4) and to administer 4 cycles of blinatumomab following 4 cycles of hyper-CVD plus INO, followed by maintenance with 12 cycles of POMP and 4 cycles of blinatumomab (1 cycle of blinatumomab after every 3 cycles of POMP). The cumulative dose of INO given before and after this most recent amendment was 4.3 mg/m2 and 2.7 mg/m2, respectively. Results: 73 pts have been treated, 70 of whom are evaluable for efficacy (3 pts too early for response assessment). 6 pts were in complete remission (CR) at enrollment and unevaluable for morphological response. Pt characteristics of the 70 evaluable pts are summarized in Table 1. Median age was 68 years (range, 60-81 years); 29 pts (41%) were ≥70 years. 41% were positive for TP53 mutation, 18% were CRLF2 positive by flow cytometry, and 27% had adverse-risk karyotype. 38/64 pts (59%) were CD20+ and received rituximab. Among 64 pts evaluable for morphologic response, 63 (98%) responded (CR, n=56; CRp, n=6; CRi, n=1). MRD negativity by flow cytometry was achieved in 53/66 pts (80%) after 1 cycle and 65/68 pts (96%) overall. There were no early deaths, and the 30-day and 60-day mortality rates were 0% and 3%, respectively. Among 69 pts who achieved remission, 9 (13%) relapsed, 3 (4%) underwent allogeneic SCT in first remission (1 of whom subsequently relapsed), 35 (51%) remain on treatment or have completed therapy, and 21 (32%) died in CR/CRp. Notably, 6 pts (9%) developed VOD, 1 after subsequent allogeneic SCT. The rate of VOD was 6/70 (9%) with no difference in rate of VOD in pts who did or did not receive fractionated INO and blinatumomab. With a median follow-up of 45 months (range, 2-98 months), the 4-year continuous remission and OS rates were 78% and 50%, respectively (Figure 1A). Age and cytogenetic risk were the primary factors associated with OS. The 4-year OS rate was 61% in pts 60-69 years vs. 34% in pts ≥70 years (P=0.06), driven by higher rates of death in remission in the older pts (13/29 [45%] vs. 8/41 [20%] in pts 60-69 years of age; P=0.03). These remission deaths in pts ≥70 years were primarily due to infection (n=7) or development of MDS/AML (n=3). Pts with high-risk cytogenetic features (e.g. KMT2A rearranged, low hypodiploidy/near triploidy, complex cytogenetics) had a 4-year OS rate of 22% vs. 57% for patients without high-risk cytogenetic features (Figure 1B; P=0.009). Neither CRLF2 positivity by flow cytometry nor the presence of a TP53 mutation significantly impacted OS. Conclusion: Reduced-intensity chemotherapy with hyper-CVD plus INO, with or without blinatumomab, is safe and effective in older adults with newly diagnosed Ph-negative ALL, with an overall response rate of 98% and 4-year OS rate of 50%. This novel regimen leads to durable remissions and apparent cure in the majority of pts age 60-69 years of age and in those without poor-risk cytogenetic features. To decrease treatment-related mortality, the protocol has been amended to eliminate chemotherapy for pts ≥70 years of age. Disclosures Short: Amgen: Honoraria; Takeda Oncology: Consultancy, Honoraria, Research Funding; Astellas: Research Funding; AstraZeneca: Consultancy. Kantarjian:Janssen: Honoraria; Abbvie: Honoraria, Research Funding; Immunogen: Research Funding; Oxford Biomedical: Honoraria; Delta Fly: Honoraria; Adaptive biotechnologies: Honoraria; Aptitute Health: Honoraria; Daiichi-Sankyo: Honoraria, Research Funding; BioAscend: Honoraria; Actinium: Honoraria, Membership on an entity's Board of Directors or advisory committees; Ascentage: Research Funding; Amgen: Honoraria, Research Funding; Jazz: Research Funding; Sanofi: Research Funding; Pfizer: Honoraria, Research Funding; BMS: Research Funding; Novartis: Honoraria, Research Funding. Ravandi:Celgene: Consultancy, Honoraria; Orsenix: Consultancy, Honoraria, Research Funding; Astellas: Consultancy, Honoraria, Research Funding; AstraZeneca: Consultancy, Honoraria; Jazz Pharmaceuticals: Consultancy, Honoraria, Research Funding; Xencor: Consultancy, Honoraria, Research Funding; Macrogenics: Research Funding; Amgen: Consultancy, Honoraria, Research Funding; BMS: Consultancy, Honoraria, Research Funding; Abbvie: Consultancy, Honoraria, Research Funding. Jain:Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees; Fate Therapeutics: Research Funding; Aprea Therapeutics: Research Funding; Servier: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Cellectis: Research Funding; Verastem: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Adaptive Biotechnologies: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Precision Bioscienes: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Pfizer: Research Funding; BMS: Research Funding; AstraZeneca: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Genentech: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; ADC Therapeutics: Research Funding; Incyte: Research Funding; Pharmacyclics: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; BeiGene: Honoraria, Membership on an entity's Board of Directors or advisory committees; TG 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, Research Funding. Kadia:JAZZ: Honoraria, Research Funding; Pfizer: Honoraria, Research Funding; BMS: Honoraria, Research Funding; Abbvie: Honoraria, Research Funding; Novartis: Honoraria; Astra Zeneca: Research Funding; Cyclacel: Research Funding; Ascentage: Research Funding; Genentech: Honoraria, Research Funding; Amgen: Research Funding; Cellenkos: Research Funding; Celgene: Research Funding; Astellas: Research Funding; Incyte: Research Funding; Pulmotec: Research Funding. Alvarado:FibroGen: Research Funding; Astex Pharmaceuticals: Research Funding; Daiichi-Sankyo: Research Funding; Sun Pharma: Research Funding; MEI Pharma: Research Funding; Tolero Pharmaceuticals: Research Funding; BerGenBio ASA: Research Funding; Jazz Pharmaceuticals: Research Funding. Burger:Janssen Pharmaceuticals: Consultancy, Speakers Bureau; TG Therapeutics: Research Funding, Speakers Bureau; AstraZeneca: Consultancy; Pharmacyclics, an AbbVie company: Consultancy, Research Funding, Speakers Bureau; Beigene: Research Funding, Speakers Bureau; Gilead Sciences: Consultancy, Research Funding. Daver:Trillium: Consultancy, Membership on an entity's Board of Directors or advisory committees; Syndax: Consultancy, Membership on an entity's Board of Directors or advisory committees; Bristol-Myers Squibb: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Pfizer: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Karyopharm: Research Funding; Servier: Research Funding; Genentech: Research Funding; AbbVie: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Astellas: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Novimmune: Research Funding; Gilead: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Amgen: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Trovagene: Research Funding; Fate Therapeutics: Research Funding; ImmunoGen: Research Funding; Daiichi Sankyo: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Amgen: Consultancy, Membership on an entity's Board of Directors or advisory committees; KITE: Consultancy, Membership on an entity's Board of Directors or advisory committees; Agios: Consultancy, Membership on an entity's Board of Directors or advisory committees; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees; Celgene: Consultancy, Membership on an entity's Board of Directors or advisory committees; Jazz: Consultancy, Membership on an entity's Board of Directors or advisory committees. Borthakur:Abbvie: Research Funding; Jannsen: Research Funding; Xbiotech USA: Research Funding; Polaris: Research Funding; BMS: Research Funding; Oncoceutics: Research Funding; Curio Science LLC: Consultancy; AstraZeneca: Research Funding; Novartis: Research Funding; Incyte: Research Funding; PTC Therapeutics: Consultancy; Argenx: Consultancy; BioTherix: Consultancy; Nkarta Therapeutics: Consultancy; Treadwell Therapeutics: Consultancy; PTC Therapeutics: Research Funding; GSK: Research Funding; FTC Therapeutics: Consultancy; Cyclacel: Research Funding; BioLine Rx: Consultancy; BioLine Rx: Research Funding. DiNardo:Novartis: Consultancy; ImmuneOnc: Honoraria; Syros: Honoraria; Daiichi Sankyo: Consultancy, Honoraria, Research Funding; MedImmune: Honoraria; Celgene: Consultancy, Honoraria, Research Funding; Calithera: Research Funding; Notable Labs: Membership on an entity's Board of Directors or advisory committees; Agios: Consultancy, Honoraria, Research Funding; AbbVie: Consultancy, Honoraria, Research Funding; Takeda: Honoraria; Jazz: Honoraria. Konopleva:Forty-Seven: Consultancy, Research Funding; AbbVie: Consultancy, Research Funding; Cellectis: Research Funding; Stemline Therapeutics: Consultancy, Research Funding; AstraZeneca: Research Funding; Amgen: Consultancy; Ablynx: Research Funding; Agios: Research Funding; Ascentage: Research Funding; Eli Lilly: Research Funding; Calithera: Research Funding; Sanofi: Research Funding; Genentech: Consultancy, Research Funding; Kisoji: Consultancy; Rafael Pharmaceutical: Research Funding; Reata Pharmaceutical Inc.;: Patents & Royalties: patents and royalties with patent US 7,795,305 B2 on CDDO-compounds and combination therapies, licensed to Reata Pharmaceutical; F. Hoffmann La-Roche: Consultancy, Research Funding. Pemmaraju:Plexxikon: Research Funding; Cellectis: Research Funding; Daiichi Sankyo: Research Funding; Samus Therapeutics: Research Funding; Incyte Corporation: Honoraria; Pacylex Pharmaceuticals: Consultancy; Celgene: Honoraria; AbbVie: Honoraria, Research Funding; Blueprint Medicines: Honoraria; Affymetrix: Other: Grant Support, Research Funding; SagerStrong Foundation: Other: Grant Support; Stemline Therapeutics: Honoraria, Research Funding; LFB Biotechnologies: Honoraria; DAVA Oncology: Honoraria; Novartis: Honoraria, Research Funding; Roche Diagnostics: Honoraria; MustangBio: Honoraria. Garcia-Manero:Jazz Pharmaceuticals: Consultancy; Bristol-Myers Squibb: Consultancy, Research Funding; Acceleron Pharmaceuticals: Consultancy, Honoraria; Helsinn Therapeutics: Consultancy, Honoraria, Research Funding; Celgene: Consultancy, Honoraria, Research Funding; Amphivena Therapeutics: Research Funding; Merck: Research Funding; Genentech: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; AbbVie: Honoraria, Research Funding; Astex Pharmaceuticals: Consultancy, Honoraria, Research Funding; Novartis: Research Funding; H3 Biomedicine: Research Funding; Onconova: Research Funding. O'Brien:Gilead, Pharmacyclics, TG Therapeutics, Pfizer, Sunesis: Consultancy, Research Funding; Amgen, Astellas, Celgene, GlaxoSmithKline, Janssen Oncology, Aptose Biosciences Inc. Vaniam Group, AbbVie, Alexion, Verastem, Eisai, Juno Therapeutics, Vida Ventures: Consultancy; Kite, Regeneron, Acerta: Research Funding. Jabbour:Amgen: Other: Advisory role, Research Funding; BMS: Other: Advisory role, Research Funding; Genentech: Other: Advisory role, Research Funding; AbbVie: Other: Advisory role, Research Funding; Adaptive Biotechnologies: Other: Advisory role, Research Funding; Takeda: Other: Advisory role, Research Funding; Pfizer: Other: Advisory role, Research Funding. OffLabel Disclosure: Inotuzumab ozogamicin and blinatumomab - frontline treatment of ALL