Persistent Acute Myeloid Leukemia Research Articles

Exposure to persistent organic pollutants in newborn dried blood spots and childhood acute myeloid leukemia

Acute myeloid leukemia (AML) is a rare malignancy representing 15–20% of all leukemia diagnoses among children. Maternal exposure to persistent organic pollutants is suggestive of increased risk for childhood AML based on existing evidence. We aimed to evaluate the relationship between persistent organic pollutants and childhood AML using newborn dried bloodspots (DBS) from the Michigan BioTrust for Health. We obtained data on AML cases diagnosed prior to 15 years of age (n = 130) and controls (n = 130) matched to cases on week of birth from the Michigan Department of Health and Human Services. We quantified levels of dichlorodiphenyldichloroethylene (p,p’-DDE), hexachlorobenzene (HCB), and polybrominated diphenyl ether congener 47 (BDE-47) in newborn DBS. We also evaluated other organochlorine pesticides, polychlorinated biphenyls, polybrominated biphenyl congener 153, and polybrominated diphenyl ethers, though these were not further evaluated as >60% of observations were above the limit of detection for these chemicals. To evaluate the association between each chemical and AML, we used multivariable conditional logistic regression. In our multivariable model of HCB adjusted for month of birth, maternal age at delivery, and area poverty, we observed no association with AML (Odds Ratio [OR] per interquartile range increase: 1.17, 95% CI: 0.80, 1.69). For p,p’-DDE, ORs were significantly lower for those exposed to the highest tertile of p,’p-DDE (≥0.29 pg/mL, OR: 0.32, 95% CI: 0.11, 0.95) compared to the first tertile (<0.09 pg/mL). We observed no statistically significant associations between HCB and BDE-47 and AML. We observed a reduced odds of exposure to p,’p-DDE and an increased, though imprecise, odds of exposure to HCB among AML cases compared to controls. Future studies would benefit from a larger sample of AML patients and pooling newborn DBS across multiple states to allow for additional variability in exposures and evaluation of AML subtypes, which may have differing etiology.

Mitophagy As a Novel Cell-Protective Mechanism Against Mitochondrial Ferroptosis in Acute Myeloid Leukemia

Background:Resistance to apoptosis poses a major clinical challenge in the therapy of acute myeloid leukemia (AML), leading to dismal clinical outcomes. Ferroptosis, another mode of regulated cell death that could bypass apoptosis resistance, has recently been investigaed for its therapeutic potential in therapy-resistant cancers. Meanwhile, accumulating evidence suggests that the regulation of ferroptosis is highly context-dependent and thus varies among cancer types. Hence, it is crucial to elucidate unique regulatory mechanisms in each tumor type for developing a therapeutic strategy. We recently found cell survival dependence of AML cells on GPX4, a major negative regulator of ferroptosis, and unique mechanistic involvement of mitochondria in AML ferroptosis. Results: We demonstrate that a selective GPX4 inhibitor ML210 as well as GPX4 knockdown induces ferroptosis, defined by iron- and lipid-peroxidation-dependence, in AML cell lines and primary AML samples, regardless of venetoclax resistance or genetic features including TP53 mutations. The cell sensitivity was negatively correlated with GPX4 protein expressions (i.e., the lower the expression, the higher the sensitivity). Normal bone marrow cells exhibited higher GPX4 levels compared to AML cells and showed significantly less sensitivity to ML210 treatment, suggesting a potential therapeutic window for GPX4 inhibition. Anti-leukemia effects of GPX4 knockdown were also observed in vivo, leading to survival prolongation in AML xenografted mice. Importantly, ferroptosis induction in AML cells is accompanied with and highly dependent on mitochondrial lipid peroxidation. 4-Hydroxynonenal (4-HNE) is one of the most biologically active products of lipid peroxidation, which is known to form adducts with certain amino acid residues of proteins to induce protein aggregates in the cytosol. We thus hypothesized that mitochondrial lipid peroxides induce mitochondrial protein aggregation in AML cell ferroptosis. Intriguingly, confocal microscopy revealed mitochondrial protein aggregation as evidenced by the co-localization of TOMM20 and protein aggregates detected by the PROTEOSTAT assay (Figure 1). Given that mitochondrial stress induced by protein aggregation reportedly induces mitophagy as a cellular stress response, we presumed that AML cells under the stress of ferroptosis may also show increased mitophagy. Indeed, GPX4 inhibition induced mitophagy, particularly in persistent AML cells, which was detected by the Mitophagy dye as mitochondrial acidification when fused with lysosomes, suggesting that mitophagy is a cell-protective response against ferroptosis as a quality control mechanism of malfunctioning mitochondria with lipid peroxidation and protein aggregation. Indeed, among persistent AML cells exposed to GPX4 inhibition, cells with more mitophagy were more resistant to further treatment with ML210 compared to those with less mitophagy. Consistently, an analogue of the ceramide-mediated mitophagy inducer LCL-461 (Dany, et al. 2016 Blood) suppressed GPX4 inhibition-induced ferroptosis in AML cells. On the other hand, inhibition of ULK1-dependent mitophagy through ULK1 knockout or the ULK1 inhibitor SBI0206965 (Bhattacharya, et al. 2021 Molecular Cancer Research) synergistically enhanced the anti-leukemia effects of GPX4 inhibition (Figure 2), suggesting a more effective therapeutic potential in ferroptosis in AML. Conclusion:Our study revealed that ferroptosis in AML cells causes mitochondrial protein aggregation and that mitophagy is induced in ferroptosis-persistent AML cells as a cell-protective mechanism against ferroptosis. This provides novel mechanistic insights into mitochondrial ferroptosis in AML, suggesting the therapeutic potential of co-targeting mitophagy in ferroptosis-based therapy. Further studies are in progress to elucidate the molecular mechanisms involved as well as the in vivo efficacy and safety of the combinatorial treatment.

Outcomes of Patients Treated with Low-Intensity Re-Induction Therapy for Persistent Disease after a Single Intensive Induction Cycle for Newly Diagnosed Acute Myeloid Leukemia (AML)

Introduction: Intensive chemotherapy with 7+3 or CPX-351 remains standard of care for fit adults with newly diagnosed AML. Nearly 50% achieve a remission after one induction cycle, and up to an extra 20% can achieve remission after a second intensive cycle. However, intensive re-induction increases morbidity through significant prolongation of cytopenias. Furthermore, certain patients (pts) may be less likely to respond to intensive re-induction, namely those with adverse risk disease. Recently, with effective non-intensive options, such as venetoclax (Ven) combinations, or IDH inhibitors, we offered low intensity re-induction to select pts with persistent disease after initial intensive therapy as a potential means of reducing morbidity, and potentially improving chance for response. This study describes the characteristics and clinical outcomes of pts with residual disease after intensive induction, treated with either Ven or IDH inhibitor re-induction. Methods: A chart review of ND-AML pts treated between 1/2017 and 2/2022 was performed. We identified pts who either received 7+3 or CPX-351 for initial induction. We then selected pts who had a nadir BM biopsy demonstrating persistent AML (>5% blasts), who then received either a Ven combination or an IDH inhibitor as next treatment. To characterize pt burden, we collected hospitalization duration, PRBC and platelet transfusion burden over 30 days after re-induction, neutropenic fever rates, ICU admission rates, and death within 30 days. We also collected clinical outcomes including CRc after 1 cycle of therapy, best response, proportion of pts proceeding to allogeneic HCT (alloHCT), and leukemia free survival (LFS). Results: A total of 15 pts were identified. Median age of pts was 61 years (22-67), 5 (33%) pts had ELN intermediate risk disease, 10 (67%) pts had ELN adverse risk disease; 5 (33%) pts had mutations in IDH1/2, and 4 (27%) pts had mutations in TP53. Most pts, 11 (73%), received initial induction with 7+3 chemotherapy; on post induction nadir marrow, a median 34% (10-90%) blasts were observed. For re-induction, 11 (73%) pts received a Ven combination (either with decitabine or azacitidine), and 4 (27%) received an IDH1 inhibitor (with azacitidine in 2 pts). Most common reason documented for re-induction choice was adverse risk disease, followed by presence of IDH mutation, followed by loss of fitness for additional intensive therapy. Median hospitalization duration from induction day 1 was 27 days (20-41); median time in hospital after re-induction was initiated was 6 days (0-27). After initiation of re-induction therapy, over the next 30 days, median number of PRBC transfusions administered to these pts was 1 (0-8), median number of platelet transfusions was 2 (0-17). Overall, 2 (13%) pts were re-hospitalized for neutropenic fever, including 1 (8%) pt who was admitted to the ICU and subsequently died (Table 1). In terms of response, 9 (60%) pts achieved a CRc after one cycle of low-intensity therapy. This includes 6 pts who received a Ven combination, and 3 patients who received ivosidenib (with or without azacitidine). Subsequently, 8 (73%) pts on a Ven combination achieved a CRc as best response, and 4 (100%) pts achieved CRc utilizing ivosidenib (Table 1). Ten (67%) pts proceeded to alloHCT. Overall, median leukemia free survival was 377 days (95% CI 235-518), and 7 (47%) pts remain alive at last follow-up (Figure 1). Conclusion: Among pts with residual disease after initial intensive induction, switching to a Ven combination or IDH inhibitor potentially reduces pt burden through shorter hospital stays, and lower burden of neutropenic fevers and transfusions. Furthermore, this strategy has promising efficacy, and serves as a successful bridge to alloHCT. A control cohort of pts receiving intensive re-induction is being composed from pts treated at our institution and will add further context. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

The AP-1/ETS Transcription Factor Network Regulates Stem Cell Quiescence and Chemotherapy Resistance in Acute Myeloid Leukemia

Despite of the continuous improvements in the treatment for acute myeloid leukemia (AML), therapy resistance and disease relapse have been persistent and attributed to leukemia stem cells (LSCs). Recent single cell RNA sequencing (Galen et al, Bernstein 2019) and deconvoluted bulk RNA sequencing analysis (Zeng et al, Dick 2022) show hierarchical cellular organization in individual AML patient specimens, with LSCs, and quiescent LSCs in particular, at its apex. However, the mechanisms controlling LSCs and their quiescence remain elusive, hindering the development of effective targeted therapies. Here, using human patient AML specimens, we show that activator protein 1 (AP-1) and E26 transformation-specific (ETS) family transcription factors (TFs) coordinately regulate LSC quiescence and disease persistence after chemotherapy. A chemical label tracing technique using carboxyfluorescein succinimidyl ester (CFSE) has emerged as a powerful tool to identify quiescent LSCs, and using this technique, we and others have demonstrated that CFSE-label retaining quiescent LSCs isolated from patient leukemia cells exhibit potent leukemia initiating property, resistance to conventional chemotherapy, and epigenetic plasticity (Takao et al, Kentsis 2018 ASH, Ebinger et al, Jeremias 2016 and 2020). Here, we combined this method with functional genomic approaches to elucidate the mechanisms controlling quiescent LSCs and identify novel therapeutic targets to overcome chemotherapy resistance. First, to identify the candidate genes for LSC quiescence regulators, we performed gene expression and chromatin accessibility profiling of label-retaining quiescent LSCs isolated from human patient leukemia cell-engrafted NSG mice in diverse subtypes of AMLs using mRNA sequencing (RNA-seq) and assay for transposase-accessible chromatin with high-throughput sequencing (ATAC-seq). We identified that several transcription factors important for normal hematopoiesis, which included AP-1 TFs, such as JUN and JUNB, and ETS TFs, such as ETS1 and FLI1, were differentially expressed in quiescent LSCs. Chromatin accessibility profiling further revealed the predominant occupancy of AP-1 and ETS TF family binding motifs in differentially accessible chromatin loci in quiescent LSCs. Importantly, gene expression signatures detected in quiescent LSCs showed significant similarity to gene expression profiles of residual leukemia cells observed in chemotherapy-treated AML patients (GSE116256, Galen et al. 2019), supporting the clinical relevance of our models. Next, to develop a functional screen system to identify the regulators of quiescent LSCs in human patient AML-bearing mice, we designed a lentiviral vector encoding mCherry fluorescent protein and doxycycline-inducible cDNAs marked by specific barcode sequences. cDNA library screening using this lentiviral vector identified several TFs as regulators for AML LSC quiescence, which included JUN as a positive regulator, and ETS1 and FLI1 as negative regulators (Figure 1). Finally, given the evidence of transcriptional activity of JUN to regulate LSC quiescence, we hypothesized that JUN activity may be also associated with chemotherapy resistance and disease persistence in patients. To investigate this, we analyzed JUN expression in individual AML cells in diverse patients before and after chemotherapy treatment (GSE116256, Galen et al. 2019). We found JUN to be highly expressed in persistent AML cells upon induction chemotherapy, especially in leukemia cells with features of hematopoietic stem cells (HSC-like), while ETS1 and FLI1 expression was not (Figure 2). These results argue that JUN activity may be required for chemotherapy resistance, LSC persistence, and ultimate AML relapse. In all, human AML LSC quiescence and therapy response can be dynamically regulated by a distinct transcription factor network. This work defines molecular mechanisms controlling AML LSC quiescence. We anticipate that these findings will lead to the elucidation of essential properties of LSC quiescence and the design of therapeutic strategies for their clinical identification and control. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Clonal Dynamics of IDH1 Mutations in Acute Myeloid Leukemia

Introduction:Isocitrate dehydrogenase 1 (IDH1) gene is mutated in 7-14% of acute myeloid leukemia (AML) patients. IDH1 encodes for an enzyme that catalyzes the conversion of isocitrate to α- ketoglutarate. IDH1 mutation leads to accumulation of the oncometabolite 2-hydroxyglutarate. Clonal evolutionary dynamics of IDH1 mutations in AML have not been clearly characterized. The introduction of targeted small-molecule therapy, Ivosidenib, in AML treatment underscores the significance of understanding the topography of clonal dynamics in IDH1-mutated AML to optimize precision medicine.Methods:We analyzed ~6000 patients with next-generation sequencing (NGS) data and identified 107 patients with IDH1 mutated AML. Disease status was determined for each NGS test date by manual chart review. IDH1 mutation status was characterized during course of AML at diagnosis, remission, relapse, and with persistent disease. Cytogenic risk category was determined using ELN 2017 guidelines. Kaplan Meier survival analysis and log-rank test were used to determine significant differences in overall survival among patient groups.Results:Of the 107 total patients, 39 patients (36%) had AML with myelodysplastic-related changes (AML-MRC) and 39 patients (36%) had AML-NOS. The most frequently co-mutated genes were SRSF2, DNMT3A, ASXL1, RUNX1, NRAS, BCOR, STAG2, NPM1, JAK2, and FLT-3 in order of frequency. Of the total patients, 74 patients (69%) had good cytogenetics, 17 patients (16%) had intermediate cytogenetics, and 16 patients (15%) had poor/very poor cytogenetics. Among the patients with IDH1-mutated AML, 85 patients (79%) were IDH1-positive at initial diagnosis, while 22 patients (21%) were IDH1- negative at diagnosis and acquired the mutation later in disease course. Of those 22 patients, 18 patients gained the mutation in the setting of persistent disease, 3 patients at remission, and 1 patient at relapse. In those with persistent AML (n=42), 30 patients (71%) remained IDH1-positive while 12 patients (29%) lost the mutation. In those achieving remission (n=66), 12 patients (18%) who were IDH1-positive remained IDH1-positive while 51 patients (77%) cleared the mutation. In those with relapsed disease (n= 21), 17 patients (81%) with IDH1-positive AML remained IDH1-positive while 4 patients (19%) lost the mutation at relapse.There were no significant differences in median overall survival in patients who were positive or negative for IDH1 mutations at diagnosis, positive or negative with disease persistence, or among patients who remained IDH1-positive or lost the IDH1 mutation at disease relapse. Patients that were IDH1-positive at diagnosis were more likely to have poorer cytogenetics than patients who were IDH1-negative at diagnosis (p=0.0016).Conclusion:In summary, this study found that IDH1 mutations are unstable throughout the course of AML and periodic genetic testing of AML patients is necessary for optimizing precision medicine approaches. In disease remission, most patients (77%) cleared the IDH1 mutation. In the relapse setting, 81% of patients retained IDH1-positive status. Our study, the largest of its kind to our knowledge, shows that serial genomic profiling for the IDH1 mutation across disease course may be beneficial in helping to tailor targeted therapy for IDH1+ AML. DisclosuresSweet: Bristol Meyers Squibb: Honoraria, Membership on an entity's Board of Directors or advisory committees; AROG: Membership on an entity's Board of Directors or advisory committees; Gilead: Membership on an entity's Board of Directors or advisory committees; Astellas: Consultancy, Membership on an entity's Board of Directors or advisory committees; Novartis: Honoraria, Membership on an entity's Board of Directors or advisory committees. Sallman: Kite: Membership on an entity's Board of Directors or advisory committees; Agios: Membership on an entity's Board of Directors or advisory committees; Aprea: Membership on an entity's Board of Directors or advisory committees, Research Funding; Bristol-Myers Squibb: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Intellia: Membership on an entity's Board of Directors or advisory committees; Magenta: Consultancy; Incyte: Speakers Bureau; AbbVie: Membership on an entity's Board of Directors or advisory committees; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees; Takeda: Consultancy; Shattuck Labs: Membership on an entity's Board of Directors or advisory committees; Syndax: Membership on an entity's Board of Directors or advisory committees. Hussaini: Adaptive: Consultancy, Honoraria, Speakers Bureau; Stemline: Consultancy; Amgen: Consultancy; Seattle Genetics: Consultancy; Celegene: Consultancy; Decibio: Consultancy; Guidepoint: Consultancy; Bluprint Medicine: Consultancy.

A Phase II Study of CPX-351 Plus Venetoclax in Patients with Relapsed/Refractory (R/R) or Newly Diagnosed Acute Myeloid Leukemia (AML)

A Phase II Study of CPX-351 Plus Venetoclax in Patients with Relapsed/Refractory (R/R) or Newly Diagnosed Acute Myeloid Leukemia (AML)

Use of High Dose Cytarabine (HiDAC) for Post-Remission or Re-Induction Therapy Is Safe and Effective in Select Older AML Patients

IntroductionA standard therapy for fit older adults (≥60 years) with acute myeloid leukemia (AML) being treated with curative intent consists of induction chemotherapy with cytarabine and an anthracycline followed by 1-4 cycles of shorter course post-remission therapy with cytarabine +/- anthracycline. Historically, HiDAC has been reserved for younger patients due to the high incidence of cytarabine-induced neurotoxicity, febrile neutropenia, and hospital re-admissions in older patients (Mayer et al. 1994). In select older adults (e.g., those with good-risk cytogenetic/molecular abnormalities, or requiring reinduction for persistent AML), the risk/benefit for HiDAC may favor its use. Real world evidence is lacking on the safety of full dose HiDAC (total 18 grams/cycle) in fit older adults in the 60-75 age range.MethodsWe performed a retrospective analysis of AML patients ≥60 years who received at least one cycle of HiDAC, defined as 3g/m2 every 12 hours for 6 doses, either days 1-3 or days 1, 3, 5, as post-remission therapy or primary re-induction (i.e., after failure of primary induction with 7+3 or CPX-351) between July 1, 2014 and May 28, 2021. AML risk was defined by European LeukemiaNet (ELN) guidelines. All patients had daily neurologic exams including prior to each dose of HiDAC and at the beginning of each nursing shift.The primary endpoint was tolerability, defined as average dose per cycle, rate of dose reductions, incidence of febrile neutropenia, cerebellar toxicity, and rate of hospital re-admissions. Secondary endpoints were overall survival (OS), composite complete remission (CCR) for patients treated with HiDAC re-induction, and duration of relapse-free survival (RFS) (from day 1 of treatment) in all patients.ResultsFrom July 2014 to May 2021, 34 patients ≥60-years-old were treated with HiDAC at our center. Median age at HiDAC administration was 64 (60-73) [Table 1]. Approximately half the patients were Hispanic and male and almost all (97%) had ECOG performance scores of 0-1. HiDAC was used as post-remission therapy (following 7+3-type induction in 80%) in 25 patients and as re-induction in 14. Eleven patients (32%) were ELN favorable risk, 8 (25%) were intermediate, and 15 (44%) were poor risk.For post-remission therapy, the average total dose of cytarabine per cycle was 16.6 gm/m2 (6-18 gm/m2) and the median number of cycles was 4 (1-4) [Table 2]; 6 patients (24%) required dose reductions, the most common reason being upcoming allogeneic stem cell transplant (n=4). No cerebellar toxicity was observed. Duration of neutropenia was on average 15 days (7-53) in the re-induction group and 7.5 days (1-29) in the post-remission group, with 23 post-remission patients (92%) receiving granulocyte colony stimulating factors (G-CSF). Eleven hospital readmissions occurred, most commonly (73%) for febrile neutropenia. Early mortality rates were low, with one patient on the post-remission arm and one patient on the re-induction arm dying within 30 and 60 days, respectively, both due to sepsis. Median OS was 15.8 months (95% CI 0-31.8) in patients who received post-remission HiDAC, with 44% of patients still alive with a median follow up of 17.2 months (95% CI 7.2-55.8). Favorable risk patients (n=11) receiving post-remission HiDAC had median OS of 15.8 months (95% CI 0-43.1), while intermediate/poor risk patients (n=14) had median OS of 11.3 months (95% CI 9.3-13.2) [p=0.53]. Median OS was 9.8 months (95% CI 2.5-17.1) in patients who received HiDAC re-induction, with 29% of patients still alive with a median follow up of 28.6 months (95% CI 16.6-39.9) [Table 3]. Five patients (36%) achieved CCR after HiDAC re-induction, all of whom went on to receive post-remission HiDAC. Median RFS was 8.5 months for re-induction and 8.8 months for post-remission patients [Table 3].ConclusionsHiDAC (18 grams) can be safely given to select patients over age 60. Critical to tolerability is rigorous screening of “fitness,” limiting the upper age receiving HiDAC to 75, ensuring adequate renal function and euvolemia, and advances in supportive care. Encouraging survival outcomes were observed in older adults receiving post-remission or re-induction HiDAC, independent of ELN risk. [Display omitted] DisclosuresBradley: AbbVie: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees. Sekeres: Takeda/Millenium: Membership on an entity's Board of Directors or advisory committees; BMS: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees. Watts: Rafael Pharmaceuticals: Consultancy; Genentech: Consultancy; Bristol Myers Squibb: Consultancy; Takeda: Consultancy, Research Funding; Jazz Pharmaceuticals: Consultancy; Aptevo Therapeutices: Research Funding.

Acquisition of IDH2 Mutations in Relapsed/Refractory AML Is Associated with Worse Patient Outcomes

Introduction Isocitrate dehydrogenase 2 (IDH2) is mutated in ~10% of acute myeloid leukemia (AML). However, the clonal evolution of IDH2 mutations through the course of AML has not been clearly elucidated. The presence of targeted therapy, Enasidenib, for the treatment of IDH2 mutated AML underscores the importance of understanding the clonal dynamics of IDH2 mutations. Methods IRB approval was obtained. In this study, we analyzed ~6000 patients with NGS results to identify 120 AML patients with IDH2 mutations and longitudinal next generation sequencing (NGS) testing. Disease status was determined for each NGS test date by chart review. IDH2 mutation status was chronicled for each of the following disease states: diagnosis, remission, relapse, and persistent disease. Cytogenetic risk category was based on ELN 2017 guidelines. Statistical analyses were performed using SPSS. Results Of the 120 patients, there were 62 patients (51.67%) with AML-NOS and 58 patients (48.33%) with AML with myelodysplasia-related changes (AML-MRC). The most commonly co-occurring mutated genes included DNMT3A, SRSF2, RUNX1, ASXL1, NRAS, BCOR, NPM1, STAG2, FLT3, and PHF6 in order of frequency. Concurrent IDH1 and IDH2 mutations were seen in 2 patients, although IDH1/2 mutations were previously reported to be mutually exclusive. Of the total patients with IDH2 mutations, 105 patients (88%) were IDH2-positive at the initial diagnosis and 15 patients (12%) were IDH2-negative at diagnosis and acquired the mutation later in disease. Of those 15 patients, 7 patients gained the mutation during persistent disease, 6 during relapse, and 2 at remission (neither of whom relapsed). Forty-eight patients (40%) who were IDH2-positive in a prior test were found to be IDH2-positive with persistent AML, while 11 patients (9%) with IDH2-positive AML lost the IDH2 mutation despite the presence of persistent AML. Twenty-one patients (18%) who were IDH2-positive in a prior test were found to remain IDH2-positive in remission, while 49 patients (41%) cleared the IDH2 mutation. Twenty-four patients (20%) with IDH2-positive AML were found to be IDH2-positive at disease relapse, while 7 patients (6%) lost the IDH2 mutation at relapse. Kaplan-Meier survival analysis and the log-rank test were used to analyze overall survival (OS) to control for confounding factors of AML category (AML-MRC vs AML-NOS) and cytogenetic risk (Figure 1). Patients who were IDH2-positive at diagnosis had significantly better survival than patients who gained the IDH2 mutation later in disease (Figure 1A, p=0.024). Patients who were IDH2-negative at remission had significantly improved survival compared to patients who were IDH2-positive at remission (Figure 1B, p=0.002). Patients who had lost the IDH2 mutation with persistent disease had significantly greater overall survival than those who remained IDH2-positive with persistent AML (Figure 1C, p=0.035). No significant difference in OS was found based on IDH2 mutation status at relapse. Conclusion In summary, in the largest study of IDH2 clonal dynamics to date, we found that IDH2 mutations are not stable during AML disease course and frequent genetic testing of AML patients in necessary to tailor personalized therapy. Most patients (70%) cleared IDH2 in disease remission. In those with refractory disease, 18% of IDH2+ AMLs lose IDH2. In the relapse setting, 22% of IDH2+ AML show loss of IDH2. Overall, 12% of patients gained IDH2 mutation later in disease course usually in the setting of refractory/relapsed AML. These patients, along with those who remained IDH2+ in remission and during refractory disease, fared worse than their counterparts. Thus, the longitudinal IDH2 mutation testing at different disease stages may be helpful in prognostic stratification. Figure 1 Disclosures Sallman: Agios, Bristol Myers Squibb, Celyad Oncology, Incyte, Intellia Therapeutics, Kite Pharma, Novartis, Syndax: Consultancy; Celgene, Jazz Pharma: Research Funding. Padron:BMS: Research Funding; Novartis: Honoraria; Kura: Research Funding; Incyte: Research Funding. Talati:Pfizer: Honoraria; BMS: Honoraria; Astellas: Speakers Bureau; Jazz: Speakers Bureau; AbbVie: Honoraria. Hussaini:Boston Biomedical: Consultancy; Stemline: Consultancy; Adaptive: Honoraria.

IAMP21 in acute myeloid leukemia is associated with complex karyotype, TP53 mutation and dismal outcome.

Acute myeloid leukemia (AML) with intrachromosomal amplification of chromosome 21 (iAMP21) is rare and has not been well characterized. We report 13 patients, 7 men and 6 women, with a median age of 65 years. Eleven patients presented with AML with myelodysplasia-related changes, and two patients had therapy-related AML. Cytopenias were detected in all patients (11 pancytopenia and two bi-lineage cytopenia). Myelodysplastic changes were observed in all 11 patients with adequate cells to evaluate. Myelofibrosis was present in ten patients. All patients had a complex karyotype, including abnormalities of chromosomes 5, 7, 17, and hsr(21)(q22), and ten patients showed TP53 deletion and/or mutation. Eleven patients received AML-based chemotherapy, one of whom also received hematopoietic stem cell transplant. By the end of the last follow-up, eight patients died with median survival of 3.2 months, four patients were alive with persistent AML, and one was in complete remission. The median overall survival was 6 months for all patients. We conclude that AML with iAMP21 is often associated with cytopenias, myelodysplasia, a complex karyotype, TP53 mutation/deletion, and a poor prognosis despite current therapies.

S910 A NEW MEMBRANE‐BOUND IL‐21/4–1BBL‐EXPANDED NK PRODUCT EXHIBITS ACTIVITY AGAINST ACUTE MYELOID LEUKEMIA ACTIVITY IN VIVO

Background:Adoptive NK cells infusion is a promising immunotherapy for acute myeloid leukemia (AML) patients.Aims:The aim of this study was to explore that two weeks of clinical grade expansion could provide sufficient numbers and purity of NK cells for clinical application with an enhanced ant‐leukemia response in vitro.Methods:NK cells were expanded in the large‐scale expansion platform based on membrane‐bound interleukin‐21/4–1BBL K562 feeder cells (mbIL21/4‐1BBL). The phenotype as well as responsitveness of NKcells were evaluated before and post NK cells expansion. NCG mouse model were used to assess the in vivo persistence and in vivo anti‐leukemia effect of NK products. Finally, 20 minimal residual disease persistent or relapse (MRD+) AML were enrolled to evaluate the safety of adoptive expanded NK products infusion.Results:In this study, using the large‐scale expansion platform, we first demonstrated that 2 weeks of expansion could satisfy clinical use in number (∼4.1 × 109 NK cell) and purity (median 81%) from 2 × 107 peripheral blood mononuclear cells based on membrane‐bound interleukin‐21/4‐1BBL K562 feeder cells (mbIL21/4‐1BBL), with expansion folds of 1318 and 2876 at 2 and 3 weeks, respectively. The CD107α production as well as TNF‐α secretion capacity of NK products against AML cell lines, or primary AML blasts, were comparable between 2 weeks and 3 weeks expanded NK products, both of which were higher than those resting NK cells before expansion. The MbIL21/4‐1BBL expanded NK product xenografted into immunodeficient leukemia mice could be found in bone marrow, spleen, liver, lung and peripheral blood, persisting for at least 2 weeks and reducing the AML burden in vivo. NK cells infusions once, or three times every other days exhibited similar tumor control. Compared with the case‐paired 20 minimal residual disease persistent or relapse (MRD+) AML patients with regular consolidation therapy, 20 AML MRD+ patients who accepted the expanded NK treatment showed better leukemia‐free survival and overall survival.Summary/Conclusion:In conclusion, this study demonstrated that mbIL‐21/4‐1BBL expanded NK cells exhibited anti‐leukemia activity against AML in vitro and in a mouse model, and the infusion of expanded NK product was potentially efficacious for AML patient without adverse effects.

Molecular Predictors and Current Management of Minimal Residual Disease (MRD) Following Induction Chemotherapy for Acute Myeloid Leukemia (AML)

▪Background: MRD is a powerful prognostic factor in AML. Emerging data indicate that allogeneic stem cell transplant (alloSCT) with MRD results in outcomes equivalently poor to alloSCT with morphologic AML (Araki et al., JCO 2016). Genomic predictors of MRD are unclear, and relative efficacy of therapies for MRD remains elusive.Objectives: Here we provide an integrated analysis of responses for 163 patients (pts) who underwent induction chemotherapy with baseline next-generation sequencing (NGS) followed by serial immunophenotypic monitoring for MRD.Methods:163 patients starting in April 2014 who underwent induction chemotherapy at Memorial Sloan Kettering Cancer Center were retrospectively studied. All received anthracycline + cytarabine, with or without investigational agents. Immunophenotypic MRD was identified in bone marrow aspirates (BMA) by multiparameter flow cytometry. Any level of residual disease was considered MRD+. Molecular analysis was obtained from pre-induction BMA by NGS using 28 or 49 gene panels. Cytogenetics/FISH were performed using standard techniques.Results: Patient characteristics are in Table 1. 7/163 (4.9%) died within 30 days of induction.153 pts had BM biopsy after induction prior to further therapy. 124/153 underwent flow after induction. 65/124 (52.4%) achieved CR/CRi after induction alone, 31/124 (25%) MRD+CR/CRi, and 34/124 (27.4%) MRD-CR/CRi. Pre-induction molecular analysis from 126 suggests that certain cytogenetic and molecular abnormalities correlate with achievement of MRD-CR. (Figure 1) Only 2/25 (8%) with RUNX1, 0/13 with SF3B1, and 0/11 with TP53 mutations achieved MRD-CR/CRi as best response after 1 cycle of induction. Only 3 additional RUNX1, 2 SF3B1, and 0/11 TP53 achieved MRD-CR/CRi as best response after a second cycle of therapy. In contrast, 7/8 with CBF AML (inv16 and no KIT mutation, n=4) or (t(8;21), n=3) achieved MRD-CR/CRi (n=5) or CR without flow (n=2) after 1 cycle of induction. 91/163 (55.8%) underwent alloSCT following induction or additional therapy. Post-alloSCT follow-up indicates potential value in converting MRD+ to MRD-. 84/91 were evaluable for MRD with flow cytometry prior to alloSCT. 41/84 (48.8%) were MRD-, 30/84 (35.7%) MRD+, and 13/84 (15.4%) persistent AML. 13/41 (31.7%) MRD-pre-alloSCT were MRD- post-induction. 28/41 (68.2%) MRD+ or persistent AML converted to MRD- prior to alloSCT following additional therapy. 23/29 MRD+CR/CRi pts after induction were intermediate/unfavorable and therefore transplant candidates. 19/23 MRD+CR/CRi intermediate/unfavorable underwent transplant (9 without post-induction therapy, 10 after consolidation), while 4 did not proceed to transplant due to relapse after induction (n=1), relapse after consolidation (n=2), and patient preference. There was no significant difference in post-transplant OS between early MRD-CR immediately following induction and later conversion to MRD-CR prior to alloSCT (Figure 1B). Post-transplant analysis reveals that most pts who enter transplant with persistent AML (n=13) or MRD+ (n=30) clear MRD (30/43, 69.7%) by the first post-transplant BM (median 32 days, Figure 1C). Despite initial post-transplant MRD clearance, pts who entered alloSCT with persistent AML or MRD+ had poorer post-transplant OS compared to pts who entered alloSCT with MRD- (p=0.02, Figure 1D).Conclusion: Our data show that AML pts with specific molecular mutations (RUNX1, SF3B1, and TP53) are unlikely to achieve MRD-CR/CRi after induction chemotherapy. We further show that additional therapy such as consolidation may be advantageous for some MRD+ pts to achieve MRD-CR prior to alloSCT, although others remain resistant to MRD clearance. Post-transplant OS is improved in pts who are MRD- at time of transplant, regardless of whether they required additional therapy beyond induction to achieve this state. Our results suggest that development of MRD-eradicating therapies after AML induction has the potential to improve post-transplant outcomes. [Display omitted] DisclosuresGoldberg:AROG: Research Funding; Pfizer: Research Funding; Celgene: Consultancy. Arcila:Invivoscribe, Inc.: Consultancy, Honoraria. Perales:Takeda: Other: Personal fees; Merck: Other: Personal fees; Abbvie: Other: Personal fees; Incyte: Membership on an entity's Board of Directors or advisory committees, Other: Personal fees and Clinical trial support; Novartis: Other: Personal fees. Tallman:ADC Therapeutics: Research Funding; Daiichi-Sankyo: Other: Advisory board; Orsenix: Other: Advisory board; Cellerant: Research Funding; BioSight: Other: Advisory board; AROG: Research Funding; AbbVie: Research Funding.

Single-Cell Mass Cytometry Characterizes Phenotypic and Functional Heterogeneity in Acute Myeloid Leukemia at Diagnosis, in Remission and Relapse

Single-Cell Mass Cytometry Characterizes Phenotypic and Functional Heterogeneity in Acute Myeloid Leukemia at Diagnosis, in Remission and Relapse

Mesenchymal stromal cells contribute to quiescence of therapy-resistant leukemic cells in acute myeloid leukemia.

Persistence of leukemic cells after induction therapy has been shown to correlate with poor survival in acute myeloid leukemia (AML). In this study, we tested if human mesenchymal stromal cells (hMSCs) have protective effects on leukemic cells undergoing chemotherapy. Persistent disease was used as marker to identify cases with therapy-resistant leukemic cells in 95 patients with AML. Immunophenotyping, cell cycle, and apoptosis assays were assessed by flow cytometry. AML coculture studies were performed with hMSC of healthy donors. Samples from patients with persistent disease had increased fractions of CD34+ CD38- and quiescent leukemic cells. Comparison of sample series collected at time points of diagnosis and blast persistence showed a relative therapy resistance of quiescent leukemic cells. Consistent with these observations, relapsed disease always displayed higher proportions of quiescent cells compared to samples of first diagnosis suggesting that quiescence is an important therapy escape mechanism of resistant cells. Co-culture studies demonstrated that hMSC protect leukemic cells from the effect of AraC treatment by enriching for quiescent cells, mimicking the effects observed in patients. This effect was even detectable when no direct stromal contact was established. Our data suggest that hMSC contribute to quiescence and therapy resistance of persistent AML cells.

Remissions after Third Induction Chemotherapy for Primary Non-Responders with Acute Myeloid Leukemia (AML) Are Uncommon and Short-Lived

The outcome of adult patients with AML who are primary non-responders to two courses of induction chemotherapy is poor. However, the utility of a 3rd induction for a select subgroup of these patients is uncertain. Here, we evaluated the rates of response and survival after a 3rd course of induction chemotherapy for primary non-responders with AML.We identified 98 patients from the Princess Margaret Cancer Centre between May 1999 and March 2015 who were non-responders to induction and reinduction chemotherapy. No-response to re-induction chemotherapy was defined according to the Revised Recommendations of the International Working Group for AML (JCO, 2003) as patients who survived > 7 days post re-induction and had persistent AML in blood or bone marrow (>5%). Median age was 58.3 years [range: 20-76.6]. 50 (51%) were male. 2% had favorable, 18% normal, 18% intermediate, and 48% adverse cytogenetics. 50% had de novo AML, 23% had AML secondary to MDS or MPN, and 17% had therapy-related AML. Induction chemotherapy consisted of "7+3" (n =88), Nove-HiDAC (n=1), Flag-Ida (n= 2), or similar variants (n=7). Reinduction chemotherapy consisted of Nove-HiDAC (n=70), Flag-Ida (n=7), "7+3" (n=1) or other similar variants (n =20). No patients received the same regimen for both induction and reinduction.Of the 98 primary non-responders, 15 received a 3rd induction regimen, while the others received supportive/palliative care ± low-dose chemotherapy (57 pts), or a non-induction clinical trial (26 pts). Average age was 56.4 (sd: 12.9) for patients who received supportive/palliative care and 47.0 (sd: 17.5) for patients who received a 3rd induction (p=0.008). Other baseline characteristics including gender, cytogenetic risk, marrow blast count post 2nd induction, and time between 1st and 2nd induction, did not differ between patients who did and did not receive a 3rd induction. Time to 3rd induction was a median of 54 days [range:36-126] from the start of the 2nd induction. Of the 15 third inductions, 7 were clinical trials evaluating novel agents in combination with induction chemotherapy, while the other 8 were combinations of standard chemotherapeutics (Flag-Ida n=1), AMSA+HiDAC (n=2), Daunorubicin+ HiDAC (n=1), Nove-HiDAC (n=4).Of the 15 patients who received a 3rd induction, 3 (20%) achieved a CR following Nove-HiDAC and Flag-Ida or AMSA+HiDAC chemotherapy, where the Ara-C was given as continuous infusion. 1 patient underwent allogeneic stem cell transplant (SCT) approximately 3.7 months after 3rd induction and remains alive 4.6 years post CR. 2 patients relapsed 2.3 and 4.7 months post CR without having received alloSCT. None of the 12 other patients responded to the 3rd induction and none had prolonged aplasia. 2 of 15 (13%) died during 3rd induction. Among the 83 patients who did not receive a 3rdinduction, 1 achieved a CR after a phase 1 clinical trial (MDM2 inhibitor) and remains in CR 3.6 years following an alloSCT.For patients who survived the immediate post induction period and were discharged from hospital median overall survival from the start of the 2nd induction did not differ between patients who did and did not receive a 3rd induction (276 days [range: 78-1304] vs 181.5 days [range: 47-1855] respectively p= 0.14). Median duration of hospital stay (including subsequent admissions) was longer for patients receiving a 3rd induction compared to those who did not (94 days following start of the 2nd induction [range: 47-169] vs 57 days [range: 51-181], respectively;(p= 0.003)).In summary, remissions after 3rd inductions for primary non-responders are uncommon, and short-lived, suggesting that 3rd inductions should be considered with caution and only when an SCT strategy is in place. DisclosuresGupta:Incyte Corporation: Consultancy, Research Funding; Novartis: Consultancy, Honoraria, Research Funding. Schuh:Amgen: Membership on an entity's Board of Directors or advisory committees. Yee:Novartis Canada: Membership on an entity's Board of Directors or advisory committees, Research Funding. Schimmer:Novartis: Honoraria.

Mitoxantrone, Etoposide, and Cytarabine (MEC) Following Epigenetic Priming with Decitabine in Adults with Relapsed/Refractory Acute Myeloid Leukemia (AML) or High-Risk Myelodysplastic Syndrome (MDS): Final Results from a Phase 1/2 Study

Background: Standard chemotherapies for relapsed or refractory acute myeloid leukemia (AML) or high-risk myelodysplastic syndrome (MDS) are often unsuccessful. Pre-treatment ("priming") with hypomethylating agents such as decitabine has been shown to sensitize AML cells to chemotherapeutics, prompting a phase 1/2 study (NCT01729845) of MEC preceded by decitabine priming (d/MEC) in relapsed/refractory AML/MDS.Methods: Patients ≥18 years with relapsed/refractory AML or high-risk MDS (>10% blasts) requiring first or subsequent salvage therapy were eligible if they had adequate organ function and a treatment-related mortality (TRM) score of <9.2 (corresponding to an expected TRM of <9.2% with standard induction chemotherapy). Prior treatment with hypomethylating agents or MEC (but not combined) was allowed. Patients with prior hematopoietic stem cell transplant (HCT) were eligible. Excluded were patients with concomitant illness with expected survival <1 year and uncontrolled infection. In phase 1, cohorts of 6-12 patients were assigned to 1 of 3 dose levels of decitabine (20mg/m2 for 5, 7, or 10 days) followed 5 days later by MEC (mitoxantrone 8mg/m2/day x 5 days; etoposide 100 mg/m2/day x 5 days; cytarabine 1 g/m2/day x 5 days). Patients with persistent AML were eligible for re-induction. Patients achieving a CR or CR with incomplete platelet recovery (CRp) could receive 2 more cycles of similarly dosed d/MEC. Dose-limiting toxicity (DLT) was defined as: 1) a grade 3 non-hematologic toxicity lasting >48 hours resulting in a >7-day delay of the subsequent treatment cycle; 2) a grade ≥4 non-hematologic toxicity, if recovery to grade ≤2 within 14 days. Both excluded febrile neutropenia/infection and constitutional symptoms. During phase 2, a Simon minimax 2-stage design was used to monitor whether a response rate of 0.25 was reached, with 80% power and a 1-sided alpha of 7%, assuming a historical CR rate of 10-15%.Results: Fifty-two patients, median age 55 (range: 19-72) years, with primary refractory (n=19) or relapsed disease (n=33; median duration of prior CR: 5 [1-19] months) and a median TRM score of 3.15 (range: 0.07-9.05) were enrolled. They received a median of 2 (1-7) prior therapies; 14 had prior HCT. During dose escalation, 1 DLT occurred at each the 2nd and 3rd dose level, identifying a 10-day course of decitabine with MEC as MTD. Since no differences in response rates were noted between the 7 and 10-day course of decitabine, a 7-day course of decitabine was used in phase 2 for logistical reasons. Overall, 11/51 evaluable patients achieved a CR (21.6%; 95% confidence interval [CI]: 11.3-35.3%) of whom 9 had no measurable residual disease (MRD) by flow cytometry. This CR rate compared favorably to a historic control population with patient matching based on duration of prior remission and number of prior salvage therapies (Estey et al. Blood 1996; 88:756), with an observed/expected CR ratio of 1.77. Seven further patients achieved a CRp, and 2 had a CR with incomplete count recovery (CRi) for an overall response rate (ORR) of 20/51 (39.2%; 95% CI: 25.8-53.9%). Among the 45 evaluable patients treated with a 7 or 10-day course of d/MEC, the ORR was 37.8% (95% CI: 23.8-53.5%). Five patients achieved a morphologic leukemia-free state, 21 had resistant disease, and 5 died before response assessment. Median response duration (CR, CRi or CRp) was 130 (15-1,229) days, with 3 of these responses ongoing. Overall survival of these 20 patients was longer (median of 332 [62-1,000 days]) than that for patients who failed to achieve remission but did not experience TRM (median 119 [45-530] days). Inclusion of 6 eligible patients that were treated with this regimen outside the clinical trial did not change these results. Nine patients died within 28 days of treatment initiation for a TRM rate of 17.6% due to infection/septic shock (5), intracranial hemorrhage (1), respiratory failure (2), and progressive disease (1). Besides grade 3-4 cytopenias, infection/neutropenic fever, nausea, and mucositis were the most common adverse events.Conclusion: D/MEC is feasible and has anti-leukemic activity in heavily pretreated relapsed/refractory AML and high-risk MDS. A comparative analysis of these results with other contemporary intensive salvage regimens is ongoing. A follow-up study is also being done to explore the relative value of pre-treatment vs. concomitant use of decitabine with intensive salvage chemotherapy. DisclosuresOthus:Glycomimetics: Consultancy; Celgene: Consultancy. Scott:Celgene: Consultancy, Membership on an entity's Board of Directors or advisory committees.

Tetraploidy/near-tetraploidy acute myeloid leukemia

A tetraploid/near tetraploid (T/NT) karyotype is a rare finding in acute myeloid leukemia (AML). Here we report 38 AML patients with a T/NT karyotype, including 26 men and 12 women with a median age of 65 years. T/NT karyotype was detected at initial diagnosis of AML in 16 patients, and acquired during the course of disease in 22 patients. All patients showed large blasts with frequent prominent nucleoli, cytoplasmic vacuoles and/or inclusions, nuclear irregularity and/or budding. Eleven patients had a non-complex and 27 had a complex T/NT karyotype; 21 patients also had pseudodiploid and/or triploid clones. After T/NT karyotype detection, 32 patients received chemotherapy and 10 also received stem cell transplant. After a median of 6.2 months follow-up, 32 patients died of disease or complications, 5 were alive with complete remission, and 1 alive with persistent AML. Median overall survival (OS) was 5 months. Patients with a non-complex T/NT karyotype had a significantly superior OS compared to those with a complex T/NT karyotype (10.7 vs. 3.4 months, p=0.0280). We conclude that T/NT karyotype in AML is often associated with distinctive morphologic features and conveys generally poor outcomes. Distinction of complex versus non-complex T/NT karyotype provides further prognostic information.

Minimal Residual Disease (MRD) By Either Flow Cytometry or Cytogenetics Prior to an Allogeneic Hematopoietic Cell Transplant (allo-HCT) Predicts Poor Acute Myeloid Leukemia (AML) Outcomes

Introduction: Persistent AML is a known risk factor for poor outcomes after allo-HCT. The impact of MRD in patients who achieve complete remission (CR) or CR with incomplete count recovery (CRi) has been less well studied.Methods: We retrospectively reviewed the records of AML patients who underwent allo-HCT in morphological remission (<5% myeloblasts and normal marrow cellularity) with or without blood count recovery between January, 2000 and January, 2014. Data was collected for variables known to impact the prognosis of AML patients (Table 1). MRD was defined as evidence of abnormalities associated with AML by either flow cytometry, cytogenetics or Fluorescence in situ hybridization (FISH). The impact of MRD identified at the time of allo-HCT on cumulative incidence of relapse (CIR), progression free survival (PFS), and overall survival (OS) was assessed in MRD+ and MRD- patients.Results: A total of 166 eligible patients were identified. The median follow-up among living patients is 46 months (range, 13-103).Thirty seven (22%) patients had evidence of MRD (13 by flow cytometry only, 17 by cytogenetics/FISH only and 7 by both). MRD was more common in patients with poor risk karyotype at diagnosis and CRi at the time of allo-HCT (Table 1). PFS (P= 0.0016), OS (P=0.002), and CIR (P=0.02) were all significantly worse in MRD+ patients (Figures 1& 2). In univariate analysis, MRD+ patients, assessed by flow cytometry had worse PFS (P=0.0216) and OS (P=0.0314) compared to MRD- patients. Similarly patients with evidence of MRD+ by cytogenetics/FISH had worse PFS (P=0.007) and OS (P=0.0031). In a multivariate cox proportional hazards model 1) any MRD positivity prior to allo-HCT, 2) poor-risk karyotype at diagnosis, and 3) CRi at allo-HCT independently predicted significantly poor PFS and OS. Only poor-risk karyotype was associated with a significant increase in CIR, while MRD positivity showed a trend towards higher CIR.Conclusion: MRD positivity prior to HCT by either flow cytometry or by cytogenetics/FISH independently predicts adverse AML outcomes.Table 1Comparison of pre-transplant variablesCovariateLabelMRD + (N=37)MRD - (N=129)P-ValueAge(years)< 408 (21%)20 (16%)0.70840 - 5920 (53%)69 (54%)≥ 6010 (26%)39 (30%)Karyotype riskFavorable/ Intermediate19 (53%)95 (74%)0.011Poor18 (47%)33 (26%)Timing of Allo-HCT1st remission (CR1)28 (74%)97 (76%)0.792> CR110 (26%)31 (24%)Allo-HCT after1st relapse(>CR1): duration of CR1> 12 mo31 (82%)113 (88%)0.285≤ 12 mo7 (18%)15 (12%)Secondary AMLNo23 (60%)78 (61%)0.964Yes15 (40%)50 (39%)Complete remission vs CRiCR28 (74%)110 (86%)0.077CRi10 (26%)18 (14%)Conditioning RegimenAblative24 (63%)72 (56%)0.449Other14 (37%)56 (44%)Donor TypeMatched sibling donor12 (32%)42 (33%)0.887Other26 (68%)86 (67%)Female donor: male recipient (FDMR)Other28 (80%)91 (78%)0.844FDMR7 (20%)25 (22%) [Display omitted] [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Phase I Study of Non-Engrafting Allogeneic, Mismatched, Unmanipulated Peripheral Blood Mononuclear Cell Infusions to Treat Poor-Prognosis Acute Myeloid Leukemia

BACKGROUND: Non-engrafting or microchimeric allogeneic immunotherapies might dramatically benefit patients with acute myeloid leukemia (AML). Mismatched G-CSF-mobilized peripheral blood (PB) and NK cells administered after chemotherapy increase complete remission (CR) and overall survival rates in poor-prognosis AML, without significant risk of graft-versus-host disease (GVHD). For example, as first-line therapy in elderly patients, the G-CSF-mobilized approach increased disease-free survival from 10% to 39% in a randomized trial (Guo et al, Blood 2011). By contrast, this trial addresses the safety of unprimed, unselected HLA-mismatched cell therapy in chemorefractory or relapsed AML, to avoid exposing donors to G-CSF and further minimize the risk of GVHD. Secondary objectives include describing the incidence of CR, GVHD, prolonged aplasia, and infusion reactions. METHODS/PATIENTS: Seven patients (median 63, range 57-68 years) with primary refractory AML (n = 1) or 1st (n = 4) or 2nd (n = 2) marrow relapse at a median of 2.9 (range, 1.6-8.0) months after completing consolidation were enrolled on this ongoing dose-escalation trial (NCT 1793025). Four had intermediate-risk caryotypes or FLT3 -ITD, 3 high-risk - including 2 with extremely complex and monosomal caryotypes and deleted or mutated TP53. One had therapy-related AML, 1 likely evolved from MDS, while 1 relapsed with bilineage leukemia (myeloid + T cell). Participants were ineligible for allotransplant due to comorbidities or adverse AML genetics. They had 14%-92% marrow blasts before re-induction, with either NoVE (n = 5), high-dose ara-C/VP16 (n = 1), or high-dose ara-C/mitoxantrone (n = 1). Donors were mismatched to recipients in varying degrees (n = 3 haploidentical and 4 with 0-3/10 allele-level matches). Target CD3+ cell dose was 1 x 107/kg for the 1st 4 patients and 5 x 107/kg for the next 3. Unmanipulated PB mononuclear cells were infused 24-48 hours after the last dose of re-induction chemotherapy (''day 0''). Marrow was evaluated on day 14 and upon hematopoietic recovery. RESULTS: SAFETY: Two patients experienced neutropenic fever from several hours to 5 days after cell infusion, but we observed no definite acute or delayed infusion reactions and no GVHD. One patient with cirrhosis developed septic shock with liver/renal failure prior to the infusion and declined dialysis; his long-term endpoints could not be assessed but he tolerated the infusion well. Sustained platelets &gt; 100 × 109/L and neutrophils &gt; 1 × 109/L occurred at +16-31 days and +14-39 days respectively in patients who achieved CR. RESPONSES: One patient achieved near morphologic CR (≤6% marrow blasts, normal PB) at day +29 and was consolidated with an HLA-identical sibling transplant at day +49; she remains in CR 28 months later. Two achieved CR lasting 77 and 164 days in the absence of further therapy. Three had persistent AML. BIOLOGY:Products contained mainly T cells (72%-80%) with varying proportions of NK (9%-17%), NK/T (0.6%-6%), B (7%-14%), and CD14+ (10%-17%) cells. Two broad patterns of total lymphocyte counts were observed: dramatic bursts from ≤ 0.2 to ≥ 1.0 (up to 1.4) × 109/L peaking days 5-8 post infusion, vs. stability/gradual recovery; numbers are too small to draw conclusions about any association with CR (p = 0.4 Fisher exact test). Lymphocyte bursts were not seen in 45 historical controls treated with chemotherapy alone (p = 0.02 for association with cell therapy). No patient-donor pairs were predicted to exhibit NK alloreactivity mediated by C1 or C2 KIR-ligand mismatching, and only 1 pair each had predicted Bw4-mediated donor-vs-recipient and recipient-vs-donor reactivity, respectively. When tested, marrow macrochimerism was undetected (n = 2). CONCLUSIONS: The incidence of life-threatening infections and duration of aplasia in the heavily-pretreated patients observed thus far do not exceed that expected with chemotherapy alone. Cell infusion is well-tolerated. The 42% (95% CI, 25%-84%) CR rate warrants continued dose escalation per protocol, especially because (1) we administer only 1 infusion and (2) our patients are treated with relapsed/refractory AML baring extremely poor prognostic features. Our results suggest that rejection of donor cells protects against engraftment and GVHD, and might sometimes be associated with a lymphocyte burst. Biologic correlations to identify ways to optimize anti-leukemic effects will be performed. Disclosures No relevant conflicts of interest to declare.

Outcome of Patients with Acute Myeloid Leukemia Treated with Salvage High-Dose Cytarabine Monotherapy

IntroductionCytarabine in high doses (> 1 g/m2) is commonly combined with other cytotoxic chemotherapy agents to treat acute myeloid leukemia (AML) in the initial and salvage settings. Single-agent high dose cytarabine (HiDAC) is traditionally reserved for consolidation therapy for patients in remission. At our institution we also use single agent HiDAC to treat patients with active disease. We reviewed our experience to assess the outcomes.MethodsWe identified patients (pts) with AML diagnosed from 2009-2014 and treated at the Hospital of The University of Pennsylvania with anthracycline plus infusional cytarabine induction (7+3) who received subsequent single agent HiDAC for active disease. Pts included were those with evidence of residual leukemia on nadir biopsy after one cycle of therapy or those with refractory disease (positive nadir after 2 cycles of therapy or disease at the time of or within 30 days of count recovery). Comparisons of categorical covariates by subgroup were evaluated using the Fisher's exact test.PatientsIn total, 44 pts with median age of 54 (range 23-69) years were identified. Twenty-five (56%) pts had de novo AML, and 19 (43%) had AML secondary to prior myelodysplasia (MDS) or a myeloproliferative neoplasm (MPN). Twenty-two pts (50%) had unfavorable cytogenetics, 8 (18%) were FLT3 -ITD positive, and 4 (9%) were NPM1 mutant (Table 1). Indications for HiDAC salvage included positive nadir marrow or frankly progressive disease at day 14 of 1 (n=28) or refractory disease (n=16)OutcomeSix pts (14%) died during index hospital admission for HiDAC therapy (5 from sepsis, 1 from multi-organ failure). Twelve (27%) pts achieved CR following HiDAC salvage and 4 (9%) achieved CRp. Pts with de novo AML were more likely to achieve a CR/CRp (11/24; 46%) compared to those with secondary AML (antecedent myeloid disorder or prior chemotherapy/radiation therapy) (5/20, 25%; p=0.037). CR/CRp rates were higher for pts with a white blood cell (WBC) count ≤10K/µL at initial diagnosis (50% vs. 20%; p=0.039). Pts treated for residual disease at first nadir were more likely to respond to HiDAC (n=13/28, 46%) than pts treated for refractory disease (3/16, 19%; p=0.104). Cytogenetic risk, age, presence of extramedullary disease, and FLT3 -ITD/NPM1 mutation status were not associated with response in this small sample (Table 2). No features predicted early mortality.ConclusionOur data suggest that pts with AML who received HiDAC therapy for active disease after 1-2 cycles of anthracycline plus infusional cytarabine induction have CR rates of 36% with single agent HiDAC with a 14% rate of early mortality. De-novo disease, initial WBC count < 10K/µL and treatment for residual disease at first nadir predict for a better response. Pts presenting with refractory disease are less likely to respond to this approach, consistent with previously published data with HiDAC combination regimens. HiDAC monotherapy for persistent AML following initial induction is a reasonable therapeutic option. [Display omitted] [Display omitted] DisclosuresLoren:Merck: Research Funding. Mangan:Incyte Corporation: Membership on an entity's Board of Directors or advisory committees. Frey:Novartis: Research Funding. Porter:Novartis: Patents & Royalties, Research Funding.

A Phase 1 Study of Lenalidomide in Combination with Mitoxantrone, Etoposide, and Ara-C in Patients with Relapsed or Refractory Acute Myeloid Leukemia

Background: The prognosis for patients with relapsed acute myeloid leukemia (AML) remains extremely poor. Standard reinduction regimens, such as mitoxantrone, etoposide, and ara-C (MEC), have been associated with complete remission rates in the 23 to 30% range. Lenalidomide (Len), an immunomodulatory drug, has efficacy in relapsed AML and has been shown to augment the effects of standard chemotherapies (anthracyclines and ara-C) in preclinical AML studies. These data prompted the current phase 1 study of Len in combination with MEC in pts with relapsed AML.Methods: The trial included pts (≥ 18 yrs) with relapsed or refractory AML. The primary objective was determination of the maximum tolerated dose (MTD) of Len when given in combination with MEC. Secondary objectives included safety, efficacy and time to count recovery. The study examined escalating doses (5-10 mg) of Len given daily for the first 14 days in combination with standard MEC doses of mitoxantrone (8 mg/m2/d), etoposide (100 mg/m2/d), and ara-C (1000 mg/m2/d) on days 4 through 8. Due to observations of prolonged count recovery, the Len dosing schedule was amended by reducing the duration of treatment to 10 days starting on Day 1. The dose of len was then re-escalated starting at 5 mg/d (5-10-25-50). A standard 3+3 dose-escalation design was used. Dose limiting toxicity was defined as Grade IV rash or Grade IV neuropathy during the first 28 day period, or delayed neutrophil (ANC <500/mL) or platelet (platelet count <20,000/mL) recovery beyond Day 45 after start of re-induction chemotherapy in the absence of persistent AML. Pts achieving a complete remission (CR) or complete remission with incomplete blood count recovery (CRi) went on to consolidation chemotherapy or hematopoietic stem cell transplantation at the discretion of the treating physician.Results: A total of 33 pts have been enrolled in the study (25 escalation phase [5 mg days 1-14, n=3; 10 mg days 1-14, n=6; 5 mg days 1-10, n=3; 10 mg days 1-10, n=3; 25 mg days 1-10, n=3; 50 mg days 1-10, n=6] and 8 expansion phase pts at 50 mg/d days 1-10). One patient was enrolled, found to be ineligible, and not treated. The median age was 62 years (range, 28-74), 45% were male and the median number of prior treatments was 2. Three dose limiting toxicities, all due to delayed count recovery past day 45, were observed in the escalation phase (n = 2/6 pts in the 10 mg/d day1-14 dose group and n = 1/6 in the in the 50 mg/d day1-10 dose group). The RP2D of Len in combination with MEC was 50 mg/d on days 1-10. Among the 14 pts treated at the RP2D, common grade 3/4 adverse events (AEs) regardless of causality were mainly hematologic including febrile neutropenia (36%). Overall 4 pts including none treated at the RP2D died during treatment. All of the on-treatment deaths, (grade 5 sepsis, n =1; infection, n=1; respiratory failure, n =2), were determined to be unrelated to the study drug. 32 pts were evaluable for response (one pt too early for assessment) with 12 pts achieving a CR and 1 pt a CRi with a total CR/CRi rate of 41% (95% CI: 24-59%). The median time to a neutrophil count of 1500/mL was 30 days and to a platelet count of 100,000/mL was 22 days.Conclusions: Len in combination with MEC re-induction chemotherapy for pts with relapsed or refractory AML was generally well tolerated and associated with an improved response rate as compared to historical controls. The safety profile was consistent with reported events in other MEC chemotherapy trials. Overall, these data suggest further exploration of high-dose Len in combination with MEC. DisclosuresDeAngelo:Celgene: Consultancy; Agios: Consultancy; Incyte: Consultancy; Amgen: Consultancy; Pfizer: Consultancy; Bristol Myers Squibb: Consultancy; Ariad: Consultancy; Novartis: Consultancy. Off Label Use: lenalidomide in AML. Fathi:Takeda Pharmaceuticals International Co.: Research Funding; Merck: Membership on an entity's Board of Directors or advisory committees; Agios: Membership on an entity's Board of Directors or advisory committees; Exelexis: Research Funding; Seattle Genetics: Membership on an entity's Board of Directors or advisory committees, Research Funding; Ariad: Consultancy. Steensma:Incyte: Consultancy; Amgen: Consultancy; Celgene: Consultancy; Onconova: Consultancy. Attar:Agios: Employment. Stone:Abbvie: Consultancy; Amgen: Consultancy; Agios: Consultancy; Celator: Consultancy; Merck: Consultancy; Karyopharm: Consultancy; Novartis: Research Funding; Celgene: Consultancy; Sunesis: Consultancy, Other: DSMB for clinical trial; Roche/Genetech: Consultancy; Pfizer: Consultancy; AROG: Consultancy; Juno: Consultancy.