Acute Myeloid Leukemia Status Research Articles

Frameshift Mutations in Leukemia-Associated Genes Correlate With Superior Outcomes in Patients Undergoing Allogeneic Stem Cell Transplant for De Novo Acute Myeloid Leukemia.

Allogeneic stem cell transplant (allo-SCT) is a mainstay of treatment for acute myeloid leukemia (AML). Its success depends largely on response of donor T lymphocytes against leukemia cells, known as graft-vs-leukemia (GvL) effect. A key potential driver of GvL is immune response to mutation-derived neoantigens. Previous studies in solid tumors have demonstrated enhanced immunogenicity of frameshift (FS)-derived peptides vs. those from non-synonymous single nucleotide variants (SNVs). We therefore hypothesized that AML cases bearing FS mutations in leukemia-associated genes would be more immunogenic than those with only other types of mutations (non-FS), and thus benefit more from allo-SCT via more robust GvL. We identified AML patients who had undergone allo-SCT between 2010 and 2022 and had next-generation sequencing data available on diagnostic specimens using a 42-gene hot spot panel. We compared the impact of tumor mutations present at diagnosis on overall survival and relapse-free survival based on FS versus non-FS status. Ninety-five AML allo-SCT patients were identified. We observed superior relapse-free survival (P = 0.038, hazard ratio (HR): 0.24) and borderline superior overall survival (P = 0.058, HR: 0.55) post-transplant in de novo AML patients, who had at least one FS mutation (other than NPM1) in one of the 42 assessed genes versus those with only non-FS mutations. Our findings suggest that FS-mutated AML cases may benefit more from allo-SCT than those with only non-FS mutations, possibly due to increased generation of immunogenic neoepitopes. If validated in an expanded study, incorporation of somatic FS mutation status in AML could improve patient selection algorithms for bone marrow transplant and thereby lead to superior outcomes.

Treatment features of adult patients with acute myeloid leukemia during the COVID-19 pandemic

Background. Acute myeloid leukemia (AML) is a highly aggressive oncological disease of the blood and bone marrow, requiring extremely toxic chemotherapy and massive supportive treatment to achieve stable remission. Currently, there is no work to provide medical care to these patients with a high risk of coronavirus infection. This paper presents treatment results of a large AML patient cohort during the COVID-19 pandemic.Aim. To assess the clinical features of coronavirus infection in AML patients.Materials and methods. A retrospective cohort study included patients hospitalized at City Clinical Hospital No. 52 (Moscow) between March 2020 and March 2022. Study inclusion criteria: 1) AML diagnosed within the last 3 years before the development of the COVID-19 pandemic; 2) age over 18 years; 3) laboratory confirmed SARS-CoV-2. AML status (newly diagnosed AML, relapse/refractory disease, remission), age, gender, comorbidity index, previous AML therapy and its outcomes were also assessed.Results. Among 218 patients with acute leukemia, 60 (27.5 %) patients had acute lymphoid leukemia, 158 (72.5 %) had AML. Patients with acute promyelocytic leukemia were allocated to a separate group - 20 (9 %) patients. The statistical data of the remaining 138 (63.5 %) patients with AML, their survival and mortality rates were assessed, and the main prognostic factors influencing the mortality and severity of coronavirus infection were identified. Also, our own results were compared with world statistics.Conclusion. Coronavirus infection in AML patients significantly worsens the prognosis. The main factors influencing the severity of coronavirus infection and survival and mortality rates are age, somatic status of patients due to the presence of concomitant chronic diseases, the development of deep hypoplasia of hematopoiesis, and the active AML status (disease onset or resistant course).

Artificial intelligence-based prediction models for acute myeloid leukemia using real-life data: A DATAML registry study

We designed artificial intelligence-based prediction models (AIPM) using 52 diagnostic variables from 3687 patients included in the DATAML registry treated with intensive chemotherapy (IC, N = 3030) or azacitidine (AZA, N = 657) for an acute myeloid leukemia (AML). A neural network called multilayer perceptron (MLP) achieved a prediction accuracy for overall survival (OS) of 68.5% and 62.1% in the IC and AZA cohorts, respectively. The Boruta algorithm could select the most important variables for prediction without decreasing accuracy. Thirteen features were retained with this algorithm in the IC cohort: age, cytogenetic risk, white blood cells count, LDH, platelet count, albumin, MPO expression, mean corpuscular volume, CD117 expression, NPM1 mutation, AML status (de novo or secondary), multilineage dysplasia and ASXL1 mutation; and 7 variables in the AZA cohort: blood blasts, serum ferritin, CD56, LDH, hemoglobin, CD13 and disseminated intravascular coagulation (DIC). We believe that AIPM could help hematologists to deal with the huge amount of data available at diagnosis, enabling them to have an OS estimation and guide their treatment choice. Our registry-based AIPM could offer a large real-life dataset with original and exhaustive features and select a low number of diagnostic features with an equivalent accuracy of prediction, more appropriate to routine practice.

A High Percentage of DNAM-1+ and Low Percentage of Tactile+ NK Cells at Diagnosis Correlate with Complete Response and Survival in AML: A Sub-Analysis of the Ven-a-Qui Pethema Trial

Background: Natural killer (NK) immune status in Acute Myeloid Leukemia (AML) could influence therapeutic results and natural history of the disease. Objectives: In the phase I/II VEN-A-QUI (Clinical trials NCT04687761) clinical trial for unfit newly diagnosed AML patients (venetoclax+Quizartinib+azacytidine or low-dose cytarabine), it was planned a prospective study of NK populations and their influence in the main clinical outcomes. NK cells were analyzed at the diagnosis in a central laboratory. Material and methods: We analyzed NK cell subpopulations based in the expression of DNAM-1, TIGIT, TACTILE, CD8, PD-1, TIM-3, LAG-3, NKp44, NKp46, NKp30, CD16, CD85j,KIR2D, NKG2C, NKG2A, CD57, CD6, CD244, NKp80, Perforin, Granulolysin and Granzyme B. We compare composite Complete Remission (CRc) results of the Venetoclax/Quizatinib with Aza or Cytarabine vs. not achieving CRc using t Student. MaxStat statistics was used to test the significant limit of those populations with a significant difference (R-statistics version 4.2.2). Survival analysis comparing significant differences between patients in CRc was performed using LogRank test and Kaplan Maier. Cox survival analysis was performed to evaluate if the presence of this populations could be an independent factor against the European Leukemia Net classification of 2017 and p53. Data of molecular characteristics were obtained with an NGS panel of 40 genes. RESULTS: The trial included 76 ineligible patients for intensive chemotherapy (37 and 39 were in Aza and LDAC arms respectively). cCR was obtained in 39 patients (51.3%). ELN2017 risk was favorable in 8, intermediate in 15, and Adverse in 53. In 70 patients NK cell populations were analyzed (4 patients with very low percentage of NK cells, 2 with no sample at diagnosis were excluded from the study). There were significant differences between patients who achieve CRc and those who did not, according to DNAM-1 (74.4% vs 61.9, p=0.0018) and TACTILE (62% vs. 72%, p=0.02) expression on NK cells. The rest of antigens analyzed on NK cells were not significant. We found a cutoff of 62% of DNAM-1+ and less than 81% of TACTILE to differentiate populations in terms of survival using long-rank test. The number of DNAM-1 positive was 47 (60%) and TACTILE- was 55 (78%). The median OS of DNAM-1+ was better than DNAM-1− (18.4 vs 4.7 months, p=0.0001), and between TACTILE− vs TACTILE+ (17,36 vs. 4.6 months, p=0.005). Using an Cox model comparing karyotype, p53 and DNAM-1+ and TACTILE-, only DNAM-1+ was associated with better outcome (HR:0.33 (0.17-0.66) and TACTILE+ with worse (HR: 2.44 (1.20-4.99) in multivariate analysis. Conclusions: DNAM-1(CD226) positive and TACTILE (CD96) negative NK cell populations are associated to CR and better OS. These findings need to be confirmed in larger studies. Interestingly, DNAM-1 is an activating receptor and TACTILE is an inhibitory receptor for NK cells that share the same CD155 ligand but display opposite function.

Mimicking Clinical Trials with Synthetic Acute Myeloid Leukemia Patients Using Generative Artificial Intelligence

Data sharing is often hindered by concerns of patient privacy, regulatory aspects, and proprietary interests thereby impeding scientific progress and establishing a gatekeeping mechanism in clinical medicine since obtaining large data sets is costly and time-consuming. We employed two different generative artificial intelligence (AI) technologies: CTAB-GAN+ and Normalizing Flows (NFlow) to synthesize clinical trial data based on pooled patient data from four previous multicenter clinical trials of the German Study Alliance Leukemia (AML96, AML2003, AML60+, SORAML) that enrolled adult patients (n=1606) with acute myeloid leukemia (AML) who received intensive induction therapy. As a generative adversarial network (GAN), CTAB-GAN+ consists of two adversarial networks: a generator producing synthetic samples from random noise and a discriminator aiming to distinguish between real and synthetic samples. The model converges as the discriminator can no longer reliably differentiate between real or synthetic data. Contrastingly, NFlow consists of a sequence of invertible transformations (flows) starting from a simple base distribution and gradually adding complexity to better mirror the training data. Both models were trained on tabular data including demographic, laboratory, molecular genetic and cytogenetic patient variables. Detection of molecular alterations in the original cohort was performed via next-generation sequencing (NGS) using the TruSight Myeloid Sequencing Panel (Illumina, San Diego, CA, USA) with a 5% variant-allele frequency (VAF) mutation calling cut-off. For cytogenetics, standard techniques for chromosome banding and fluorescence-in-situ-hybridization (FISH) were used. Hyperparameter tuning of generative models was conducted using the Optuna Framework. For each model, we used a total of 70 optimization trials to optimize a custom score inspired by TabSynDex which assesses both the resemblance of the synthetic data to real training data and its utility. Pairwise analyses were conducted between the original and both synthetic data sets, respectively. All tests were carried out as two-sided tests using a significance level α of 0.05. Table 1 summarizes baseline patient characteristics and outcome for both synthetic cohorts compared to the original cohort. Firstly, we found both models to adequately represent patient features, albeit that some individual variables showed a statistically significant deviation from the original cohort. It is important to note that for such a large sample size (n=1606 for each cohort), even miniscule differences can be rendered statistically significant notwithstanding any meaningful clinical difference. Interestingly, variables that deviated from the original distribution were different for both models indicating model architecture to play a vital role in sample representation: While CTAB-GAN+ showed significant deviations for both age and sex, NFlow showed significant deviations for AML status. Complete remission rate was similar between original (70.7%, odds ratio [OR]: 2.41) and CTAB-GAN+ (73.7%, OR: 2.81, p=0.059) and NFlow (69.1%, OR: 2.24, p=0.356). For event-free survival (EFS), which was not included as a target in hyperparameter tuning, both networks deviated significantly from the original cohort (original: median 7.2 months, HR: 1.36; CTAB-GAN+: median 12.8 months, HR 0.74, p<0.001; NFlow: median 9.0 months, HR: 0.87, p=0.001). Overall survival (OS) was well represented by NFlow compared to the original cohort, while CTAB-GAN+ showed a significant deviation (original: median 17.5 months, HR: 1.14; CTAB-GAN+: median 19.5 months, HR 0.88, p<0.001; NFlow: median 16.2 months, HR: 1.00, p=0.055). Both models showed an adequate graph representation in Kaplan-Meier analysis (Figure 1). Here, we demonstrate using two different generative AI technologies that synthetic data generation provides an attractive solution to circumvent issues in current standards of data collection and sharing. It effectively allows for bypassing logistical, organizational, and financial burdens, as well as regulatory and ethical concerns. Ultimately, this enables explorative research inquiries into previously inaccessible data sets and offers the prospect of fully synthetic control arms in prospective clinical trials.

Abstract 8: Differential flt3 mutational status in acute myeloid leukemia predicts sensitivity to flt3 inhibitor gilteritinib in vitro and in vivo

Abstract Acute Myeloid Leukemia (AML) is a heterogenous hematopoietic cancer characterized by abnormal differentiation and uncontrolled proliferation of hematopoietic progenitor cells. Recent approaches to treatment, which aim to improve outcomes in refractory AML, include strategies to molecularly target specific receptor-mediated signaling pathways which regulate cell growth and proliferation. One target-specific approach is the inhibition of fms-like tyrosine kinase 3 (FLT3). FLT3 is expressed on the surface of hematopoietic progenitor cells and plays an important role in regulating proliferation, survival, and differentiation of multipotent stem cells. The presence of internal tandem duplications (ITDs) at the juxta membrane domain within the FLT3 receptor are common driver mutations in 30% of AML patients. In this study, we utilized 3 luciferase-expressing human AML models, MV4-11, MOLM-13, and, OCI-AML3 to determine the prognostic impact of FLT3 inhibitor Gilteritinib on the allelic ratio of ITD versus wildtype allele. OCI-AML3 expresses wild-type FLT3 whereas MOLM-13 is heterozygous for FLT3-ITD mutations, and MV4-11 possesses a homozygous in-frame tandem repeat insertion in the FLT3 allele. Here we demonstrate that FLT3-ITD mutational status impacts the efficacy of Gilteritinib in both in vitro and in vivo AML models. The in vitro assays showed that treatment with Gilteritinib resulted in significant dose dependent reduction in overall cell viability and bioluminescence in the MV4-11 cell line, while MOLM-13 was only moderately sensitive, and OCI-AML3 was least sensitive to Gilteritinib treatment. Additionally, we evaluated the downstream MAPK and PI3K signaling pathways as FLT3-ITD oncogenic mutations result in ligand-independent constitutive activation of signaling pathways mediating leukemic transformation. For in vivo validation, MV4-11, MOLM-13, and OCI-AML3 luciferase expressing cells were implanted systemically into NCG mice and once tumors were established, we evaluated the survival benefit in response to Gilteritinib and various standard of care agents. In accordance with our in vitro findings, the quantification of tumor progression by bioluminescent imaging showed the highest therapeutic activity of Gilteritinib in MV4-11 tumor bearing mice through selective targeting of FLT3-ITD mutations. Taken together, our in vitro and in vivo results demonstrate that differential FLT3-ITD mutational status in AML models impacts Gilteritinib treatment response and downstream signaling events with the highest activity in MV4-11 followed by MOLM-13, and lack of activity in OCI-AML3 harboring wild-type FLT3. Citation Format: Bincy John, Pasupathi Sundaramoorthy, Elizabeth Rainbolt, Andrew Wong, Zachary Ward, Justin Avery, Savannah Todd, Shasta Kidder, Diana Gietl, Chassidy Hall, David Harris. Differential flt3 mutational status in acute myeloid leukemia predicts sensitivity to flt3 inhibitor gilteritinib in vitro and in vivo [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 8.

Smoking status in acute myeloid leukemia is associated with worse overall survival and independent of prior nonhematopoietic malignancies, cytogenetic abnormalities, and WHO category

Acute myeloid leukemia (AML) is an aggressive hematopoietic malignancy with several patient- and disease-associated variables known to impact prognosis. Tobacco smoking is an environmental factor associated with a greater incidence of AML, but there have been limited studies that evaluated smoking toward overall survival. We retrospectively searched for AML cases and collected clinical and diagnostic data for each case. We also used an independent next-generation sequencing (NGS) data set to assess for a distinct mutational signature associated with smoking. When stratified by smoking status, there was a greater number of males, patients aged ≥60 years, and patients with ≥2 comorbidities within the smoking category (P<.05). Survival analysis demonstrated decreased survival probability in the smokers, male smokers, smokers with 1 other comorbidity, and smokers without a prior history of nonhematopoietic malignancy (P<.05) as compared to nonsmokers. Smoking was associated with a decrease in survival within the World Health Organization categories of AML, not otherwise specified (AML NOS; P=.035) and AML with recurrent genetic abnormalities (AML RGA; P=.002). Multivariate analysis showed that patients who were smokers had a greater hazard ratio than nonsmokers after adjusting for the other covariates. Our findings demonstrated that smoking was independently associated with decreased overall survival after adjusting for other potentially confounding factors. In addition, our results suggest that a mutational signature can be recognized using NGS data in a subset of AML patients who smoke.

TP53 copy number and protein expression inform mutation status across risk categories in acute myeloid leukemia

AbstractMutant TP53 is an adverse risk factor in acute myeloid leukemia (AML), but large-scale integrated genomic-proteomic analyses of TP53 alterations in patients with AML remain limited. We analyzed TP53 mutational status, copy number (CN), and protein expression data in AML (N = 528) and provide a compilation of mutation sites and types across disease subgroups among treated and untreated patients. Our analysis shows differential hotspots in subsets of AML and uncovers novel pathogenic variants involving TP53 splice sites. In addition, we identified TP53 CN loss in 70.2% of TP53-mutated AML cases, which have more deleterious TP53 mutations, as well as copy neutral loss of heterozygosity in 5/32 (15.6%) AML patients who had intact TP53 CN. Importantly, we demonstrate that mutant p53 protein expression patterns by immunohistochemistry evaluated using digital image-assisted analysis provide a robust readout that integrates TP53 mutation and allelic states in patients with AML. Expression of p53 by immunohistochemistry informed mutation status irrespective of TP53 CN status. Genomic analysis of comutations in TP53-mutant AML shows a muted landscape encompassing primarily mutations in genes involved in epigenetic regulation (DNMT3A and TET2), RAS/MAPK signaling (NF1, KRAS/NRAS, PTPN11), and RNA splicing (SRSF2). In summary, our data provide a rationale to refine risk stratification of patients with AML on the basis of integrated molecular and protein-level TP53 analyses.

Clinicohematological Parameters and Assessment of Post Induction Status in Acute Myeloid Leukemia–Experience at A Tertiary Care Center

Objective: To evaluate the role of clinical, hematological and genetic parameters in acute myeloid leukemia (AML) as the predictors of response to induction chemotherapy.&#x0D; Study Design: Cross-sectional study.&#x0D; Place and Duration of Study: Department of Haematology, Armed Forces Institute of Pathology, from Jun to Jun 2019.&#x0D; Methodology: All the newly diagnosed cases of de-novo AML were included in the study. Clinical, haematological and immunophenotypic parameters were noted. Cytogenetic and molecular analyses were performed. Response to first course of standard induction chemotherapy was assessed.&#x0D; Results: A total of 58 patients were included in the study. The median age was 19 years. Thirty-one (53.4%) were adults while 27 (46.6%) were in the pediatric age group. Thirty-six (62.1%) were males while 22 (37.9%) were females. The most common clinical presentation was fever. The most common French-American-British classification (FAB) subtype was AML-M2. The blast percentage was 78%. Forty-six (79.3%) patients had a normal karyotype. Of the 58 patients, 38 (65.5%) achieved complete remission while 20 (34.5%) did not achieve complete remission after induction chemotherapy.&#x0D; Conclusion: AML was seen in a younger age group in our population. There was statistically significant association of high white blood cell (WBC) count with remission status. Increasing age was associated with a poor response to induction chemotherapy, while translocation t(8;21) was associated with a good response. Assessment of prognostic parameters is vital in the initial diagnostic workup of AML patients to predict response to induction chemotherapy.

Artificial Intelligence-Based Predictive Models for Acute Myeloid Leukemia

Artificial Intelligence-Based Predictive Models for Acute Myeloid Leukemia

Immune and Epigenetic Landscape of TP53-mutated Acute Myeloid Leukemia (AML) and Higher-Risk Myelodysplastic Syndromes (HR-MDS)

Immune and Epigenetic Landscape of TP53-mutated Acute Myeloid Leukemia (AML) and Higher-Risk Myelodysplastic Syndromes (HR-MDS)

Retrospective Analysis of the Outcomes of Patients with Relapsed/Refractory Acute Myeloid Leukemia Included in a Patient Named Program of Gemtuzumab Ozogamicin

▪Introduction: In 2010 the French Health Agency opened a compassionate patient named program of gemtuzumab ozogamicin (GO, Mylotarg®) in relapsed/refractory (R/R) patients with acute myeloid leukemia (AML). Of note, since 2012, it was recommended to use GO at the dose of 3 or 6 mg/m 2 in addition to chemotherapy. We conducted a retrospective trial (NCT03287128) to evaluate the efficacy and the safety of GO-based regimen in R/R adult AML patients.Patients and methods: We retrospectively collected data of patients older than 18 years treated with GO-based regimen for AML in first relapse or for refractory AML, defined by failure after a prior standard intensive chemotherapy, in 18 French centers between December 15, 2011 and November 10, 2016. The primary objective was to assess the response to GO-based regimen. Patients were considered in response if reaching complete remission (CR), CR without platelet recovery (CRp) or CR with incomplete hematological recovery (CRi). Secondary objectives were the cumulative incidence of allogeneic hematopoietic stem cell transplantation (allo-HSCT) and the safety of the use of GO-based regimen.Results: Three hundred and thirty-five adult patients with R/R AML were included. Median age was 58 years (20 to 80 years). At diagnosis, cytogenetics was favorable in 50 (17%) patients, intermediate in 173 (59%) and adverse in 60 (20%). ELN distribution was favorable: 35%, intermediate: 42% and unfavorable: 23%. NPM1 mutation was present in 29% of patients and FLT3 mutation in 23%. Most patients had de novo AML (84%). Two hundred and thirty-eight patients (79%) were in first relapse and 65 (21%) had a refractory AML. The time between first diagnosis of AML and treatment with GO-based regimen was 4 to 16 months (median 9.4 months). Most patients (88%) received GO in combination with various intensive chemotherapy scheme including “7+3” with anthracycline/cytarabine (n=39 patients), intermediate and high-dose cytarabine (n=68), cytarabine in continuous intravenous infusion (n=78), mitoxantrone/cytarabine (n=49) and fludarabine/cytarabine and/or amsacrine and/or etoposide chemotherapy (n=35). Median follow-up time was 11 months.Among the 305 patients, 191 responded to GO-based regimen: 110 (36%) were in CR, 62 (20%) were in CRp and 19 (6%) in CRi for an overall response rate (CR+CRp+CRi, ORR) of 63%. In multivariate analysis, response was associated with age <50 years, de novo AML and relapse status. Among the 191 responders, 110 received additional courses of chemotherapy, 69 with GO. Main reason to not receive additional course (with or without GO) was allo-HSCT project. In the whole population, median overall survival (OS) after day 1 of treatment with GO was 11.2 months. In the population of responders, median OS after response was 20.4 months. In multivariate analysis, longer survival was associated with age < 50 years, de novo AML and favorable ELN group. Cumulative incidence of relapse at 24 months after response was 46%. One hundred and forty-seven patients received allo-HSCT, including 122 responders after GO-based regimen and 25 patients in treatment failure. Cumulative incidence of allo-HSCT at 18 months was 48%. Four-year OS was 48% in transplanted patients versus 19% in non-transplanted patients (Figure 1).Regarding safety of GO-based regimen, early deaths occurred within <30 days after the first dose of GO in 14 patients, and within <60 days in 35 patients. Myelosuppression was observed in all patients. Mean duration of thrombocytopenia <100 G/L was 35 days in responders. Bleeding grade 3 or more was observed in 22 patients (7%). Infection grade 3 or more was observed in 112 patients (30%). Sinusoidal obstruction syndrome (SOS) after GO treatment was reported in 6 patients, resolving in 4 of them. Four cases of fatal SOS were reported after allo-HSCT. Toxic deaths, i.e., not related to worsening leukemia, were reported in 20 patients after the first course of chemotherapy, 3 after additional courses and 33 after allo-HSCT.Conclusion. Our study is the first to report efficacy data in the real-world setting of R/R AML adult patients treated with GO-based regimen. In our cohort of 305 patients, response rate was 63% and GO-based regimen appears as a valuable bridge-to-transplant option. Safety analysis showed toxicities consistent with the known safety profile of GO and chemotherapy. [Display omitted] DisclosuresLambert: ASTELLAS: Consultancy; CELGENE/BMS: Consultancy. Pautas: PFIZER: Consultancy; ABBVIE: Consultancy. Raffoux: ASTELLAS: Consultancy; PFIZER: Consultancy; ABBVIE: Consultancy; CELGENE/BMS: Consultancy. Legrand: Servier: Consultancy. Gastaud: PFIZER: Consultancy; CELGENE/BMS: Consultancy; ABBVIE: Consultancy; GSK: Consultancy. Pigneux: Amgen: Consultancy; Sunesis: Consultancy, Research Funding; BMS Celgene: Consultancy, Research Funding; Roche: Consultancy, Research Funding; Novartis: Consultancy, Research Funding. Mathilde: SERVIER: Consultancy; ABBVIE: Consultancy. Dombret: Amgen: Honoraria, Research Funding; Incyte: Honoraria, Research Funding; Jazz Pharmaceuticals: Honoraria, Research Funding; Novartis: Research Funding; Pfizer: Honoraria, Research Funding; Servier: Research Funding; Abbvie: Honoraria; BMS-Celgene: Honoraria; Daiichi Sankyo: Honoraria. Rousselot: Incyte, Pfizer: Consultancy, Research Funding. Castaigne: PFIZER: Consultancy.

Long Non Coding RNA BALR2-CDK6 Axis Influences Cell Differentiation and Metabolic Status of Acute Myeloid Leukemia

Purpose: Non-coding RNA (ncRNAs) genes are at least as frequent as protein-coding genes in the human genome; however, our knowledge on their function is still preliminary. Long ncRNAs are arbitrarily defined as transcripts that are over 200 nucleotides in length, and demonstrate great heterogeneity in their molecular mechanisms. Their emerging role as new players in cancer development and therapy response is widely supported by several scientific reports, and we recently found the LncRNA BALR-2 expression significantly correlated with a decreased response to prednisone treatment in B-cell precursor acute lymphoblastic leukemia (Bcp_ALL). Given its association with mixed lineage leukemia (MLL) rearrangements in Bcp-ALL, we investigated BALR-2 involvement in acute myeloid leukemia (AML) leukemogenic pathways.Methods: We retrospectively analyzed by RQ-PCR bone marrow samples of 132 children with de novo AML harboring different genetic abnormalities (CBF,MLL, and NUP98 rearrangements, FLT3-ITD , other rare translocations and patients without any recurrent molecular abnormality) diagnosed between 2002-2014 in one of the AIEOP centers and treated according to AML-2002/01 protocol. We correlated the expression of BALR-2 with patient event-free survival (EFS) and gene expression profile (GEP, HTA affymetrix 2.0). Using gene set and single sample enrichment analysis (GSEA, ssGSEA), we compared the gene expression signatures of patients with either high (4th quartile) or low expression (1st+2nd+3rd quartiles) of BALR-2. SHI-1 cell line was used to perform in vitro experiments aimed at examining the function of BALR2.Results: The expression of BALR-2 was found higher in all AML samples as compared with those collected from healthy volunteers. Moreover, we did not find any correlation between BALR-2 expression and any specific genetic abnormality. We subdivided patients by using BALR-2 expression quartiles and found that that patients with higher BALR-2 expression (4th quartile, n=32) had worse, although not statistically significant, EFS when compared to that of patients allocated to the 1st+2nd+3rd quartiles (n=100; EFS= 53% versus 61%). We discovered that the 4th quartile was enriched for patients who did not reach complete remission (CR) after induction therapy (28% vs 12%, p=0.03). Supervised clustering analyses of GEP performed in 58 samples showed that cases belonging to the 4th quartile (n=18) clustered separately from the remaining quartiles (n=40), irrespectively of the underlining genetic abnormality shown by these patients. In particular, we found that 57 coding and 12 non-coding RNAs were significantly differentially expressed (Fold Change|2|, p<0.01), and CDK6 , being chromosomally adjacent to BALR2, was overexpressed in the 4th quartile. We further investigated this putative BALR2-CDK6 relationship and found a positive correlation between BALR-2 and CDK6 expression in the whole cohort analyzed (Pearson correlation>0.7, p<0.05). We silenced BALR-2 in vitro and found this induces lower CDK6 mRNA and protein levels, as well as phospho-RB, its direct target. In line with what seen in Bcp-ALL, BALR2-CDK6 knockdown decreased cell proliferation, increased apoptosis, and sensitivity to etoposide treatment. Of note, BALR-2 depletion increased myelomonocytic differentiation, as evidenced by CD11 and PU.1 markers, both of which are Runt-related transcription factor (RUNX1 ) targets. In agreement with studies demonstrating that CDK6 acts as RUNX1 inhibitor, we found that additional 81 RUNX1 target genes showed marked down-regulation in patients with high BALR-2-CDK6 expression. Through ssGSEA, an aberrant regulation of processes regulating mitochondrial mass and activity was found in patients with the 4th quartile of BALR-2 expression (p<0.05).Conclusions: Taken together, our data suggest that BALR-2 expression is associated with worse outcome in AML. Furthermore, our data suggest a cis-regulatory transcriptional relationship between BALR-2 and CDK6, which, in turn, antagonizes RUNX1 and enhances myeloid differentiation. Lastly, in vitro approaches are ongoing to define the mitochondrial signaling events involved in BALR-2's ability to alter the metabolic status in AML, which in turn could lead to novel approaches for inducing anti-leukemic effects. DisclosuresNo relevant conflicts of interest to declare.

Prognostic Significance of Minimal Residual Disease in Patients with Secondary Acute Myeloid Leukemia

BACKGROUNDSecondary acute myeloid leukemia (AML) is defined as AML that develops from a prior myelodysplastic or myeloproliferative disorder, or from exposure to chemotherapy, radiation, or environmental toxins. Patients with secondary AML have inferior prognosis compared to those with de novo AML,1 and are more likely to be elderly with increased comorbidities and worse performance status.2 The detection of minimal residual disease (MRD) also has independent prognostic value in patients with AML.3-6 In elderly AML patients, MRD predicts worse outcomes independent of the poor treatment risk factors associated with increased age.7Due to the similar disease pathogenesis between elderly AML and secondary AML patients, it is likely that MRD testing will have similar implications. However, there is limited data on the prognostic role of MRD testing in secondary AML. METHODSWe retrospectively analyzed de novo and secondary AML patients at Washington University Medical Center who received post-induction, post-consolidation, pre-transplant, or post-transplant MRD testing during their treatment. MRD testing was performed with multiparameter flow cytometry testing using the “different from normal” method at one reference laboratory,8,9 with positive MRD defined as detection of an abnormal myeloblast population above 0.1%. A total of 180 patients were included. Any patient with at least one positive MRD was considered “MRD positive.” Baseline pretreatment characteristics were compared between groups with Mann-Whitney, Chi-square, or Fisher's exact test, when appropriate. We evaluated overall survival (OS) and relapse rates in de novo and secondary AML patients stratified by MRD status using Kaplan-Meier curves and the log rank test, or cumulative incidence function, as appropriate. Multivariate Cox proportional hazards model was also fitted to determine if MRD testing has similar prognostic value across de novo and secondary AML patients. RESULTSOur analysis included 133 patients with de novo and 47 patients with secondary AML. Secondary AML patients were significantly older at age of diagnosis (mean age 59.85 versus 51.17, p<0.001), however had similar rates of MRD positivity (28% versus 29%, p=0.830). There was a trend toward poorer cytogenetic risk profiles and worse Karnofsky Performance Status in secondary AML (Table 1). In all patients, we confirmed that MRD positivity confers significantly worse OS (45.5% versus 70.8%, p=0.020) and relapse rates at 2 years (53% versus 25.5%, p=0.029) (Figure 1). In de novo AML patients, MRD positivity continued to demonstrate significantly worse 2-year OS (44.1% versus 76.3%, p=0.006) and relapse rates (57.6% versus 20.7%, p=0.008). However, in secondary AML patients, there were no differences in OS (p=0.69) or relapse rates (p=0.98) between MRD positive and negative patients. Multivariate analysis demonstrated significant interaction (p=0.042) between MRD status and AML subtypes. MRD positivity emerged as an independent prognostic indicator of worse OS in de novo patients (HR 2.8, 95% CI 1.524-5.405) but not in secondary patients (HR 1.5, 95% CI 0.188-2.336). DISCUSSIONWe confirm that MRD status is a poor prognostic indicator in all AML patients, and this also holds true in patients with de novo AML. However, we found that MRD status has less prognostic value in secondary AML patients. This may be due to the overall poor prognosis seen in secondary AML patients, suggesting that MRD monitoring in this group of patients may be less useful for risk stratification. However, our study is limited by the small sample size, particularly in the number of secondary AML patients who displayed MRD positivity (N=13). In addition, we defined MRD positivity across all time points of testing, which may decrease patient homogeneity. A larger study with increased follow-up is necessary to confirm our findings, as these results have important implications in risk-stratification and treatment of patients with secondary AML. [Display omitted] DisclosuresDiPersio:BMS, Asterias, Amphivena, Bluebird: Other: Travel, Accomodations, Expenses; Hemedicus, DAVA Oncology: Speakers Bureau; Celgene, Bioline, Vasculox, Cellworks, Rivervest: Consultancy; Magenta Therapeutics: Equity Ownership.

Antigen Expression Varies Significantly between Molecular Subgroups of Acute Myeloid Leukemia Patients: Clinical Applicability Is Hampered by Establishment of Relevant Cutoffs

Introduction: In this single-center study of 268 acute myeloid leukemia (AML) patients, we have tested if a subset of 4 routinely employed immunophenotypic stem cell-associated markers correlated with the presence of recurrently mutated genes and if the markers were predictive for mutational status. Methods: Immunophenotypic data from 268 diagnostic AML samples obtained in 2009–2018 were analyzed retrospectively for the antigens CD34, CD117, CD123, and CLEC12A. Correlation between immunophenotypes and mutations was analyzed by Fischer’s exact test. Clinical applicability of the markers for predicting mutational status was evaluated by receiver operating characteristics analyses, where an area under the curve (AUC) of at least 0.85 was accepted as clinically relevant. Results: For a number of genes, the antigen expression differed significantly between mutated and wild-type gene expression. Despite low AUCs, CD123 and CLEC12A correlated with FLT3+NPM1− and FLT3+NPM1+. Three subsets met the AUC requirements (CD34+, CD34+CD117+, and CD34−CD117+) for predicting FLT3−NPM1+ or FLT3+NPM1+. Conclusion: The value of immunophenotypes as surrogate markers for mutational status in AML seems limited when employing CD123 and CLEC12A in combination with CD34 and CD117. Defining relevant cutoffs for given markers is challenging and hampered by variation between laboratories and patient groups.

Immune Senescence and Exhaustion Correlate with Response to Flotetuzumab, an Investigational CD123×CD3 Bispecific Dart® Molecule, in Acute Myeloid Leukemia

Immune Senescence and Exhaustion Correlate with Response to Flotetuzumab, an Investigational CD123×CD3 Bispecific Dart® Molecule, in Acute Myeloid Leukemia

Factors Associated with Clinical Outcomes after Venetoclax-Based Combination Therapy in Acute Myeloid Leukemia and High-Grade Myeloid Neoplasms

Background Outcomes of older patients (pts) with acute myeloid leukemia (AML) ineligible for intensive chemotherapy (IC) remain poor with an overall survival (OS) of &amp;lt;12 months (mo). The combination of venetoclax (ven) and azacitidine (aza) for such newly diagnosed AML pts in the confirmatory VIALE-A trial demonstrated an improvement in OS compared with aza alone. However, this trial excluded pts with prior hypomethylating (HMA) therapy, as well as pts with relapsed/refractory (R/R) disease. Therefore, the outcomes of those pts remain unknown. We thus investigated the outcomes of pts treated with ven-based therapy in the newly diagnosed and R/R settings. Methods Pts diagnosed with AML or high-grade myeloid neoplasms (MN) between 2/2018 - 5/2020 who completed at least one cycle of ven-based therapy were included. Clinical data were collected and targeted next generation sequencing evaluating a panel of ~80 genes commonly mutated in myeloid malignancies was performed. Responses were assessed using the IWG criteria, and OS was calculated from cycle one day one until the date of last follow-up/death. Categorical variables were compared using Fisher's exact test. Univariable and multivariable Cox- and logistic-regression were used to assess factors associated with response and OS. Results Fifty pts were prescribed ven at our center, and eight were excluded as they received less than one full cycle of therapy. Of the 42 pts included, the majority (91%) had AML and 9% had high-grade R/R MDS or CMML. Median age at diagnosis was 66 years (28-88) and 23 pts (55%) were female. Per ELN risk stratification, most (81%) pts had poor, 14% had intermediate, and 5% had favorable-risk disease. Forty percent had de novo-, 52% had secondary/therapy-related AML (t-AML) and 8% had R/R MN. Twenty one (50%) pts received venetoclax for newly diagnosed AML, and of those nine (43%) had prior HMA therapy. Of the other 21 (50%) pts who received venetoclax in the R/R setting, 10 (48%) received prior HMA therapy. Median duration of prior HMA therapy was 6 cycles (1-24). Ven was given in combination with HMA, low dose cytarabine (LDAC), or IC in 35 (83.3%), 6 (14.3%), and 1 (2.4%) pt respectively. Forty pts were evaluable for response. Complete response (CR) or CR with incomplete count recovery (CRi) rate was 47.5%. CR/CRi rates were significantly different based on ven combination (IC&amp;gt; HMA &amp;gt; LDAC, p=.001), AML status (de novo &amp;gt; t-AML &amp;gt; s-AML, p=.025), ELN risk and cytogenetic group (favorable &amp;gt; intermediate &amp;gt; poor, p=.001 and p=.004 respectively). CR/CRi rates were lower for pts receiving prior HMA, p=.012. CR/CRi rates were higher for pts with FLT3 (p=.040) and NPM1 mutations (p=.04). There was a trend for lower CR/CRi for pts with TP53 mutations, p=.095. [Table 1] Newly diagnosed vs R/R disease status, number of mutations (≤3 vs &amp;gt;3), and prior treatment with IC for AML had no impact on CR/CRi rates. There was no difference in median OS (mOS) between newly diagnosed vs R/R pts: 10.9mo vs 9.5mo respectively, p=.61. OS was significantly different based on response (CR vs CRi vs MLFS/PR vs PD with mOS not reached, 13.2, 9.5, and 2.3 mo respectively, p&amp;lt;.001. OS was longer for pts with de-novo AML&amp;gt; t-AML&amp;gt; s-AML with mOS of 15.6, 10.9, and 2.8 mo respectively, p=.007. Lastly, mOS was significantly shorter for pts with prior HMA exposure vs not at 5.0 vs 15.6 mo respectively, p=.003. [Figure 1] Multivariable analysis (MVA) for response demonstrated ELN cytogenetic intermediate vs high risk to be the only factor associated with CR/CRi, p=.004. MVA for OS showed prior HMA exposure and ELN poor cytogenetic risk to be the only two factors associated with shorter OS, p=.022 and p=.007 respectively. Conclusions In summary, this study of real world outcomes with ven combination therapy in AML has revealed disease features predictive of clinical outcomes. Remarkably, we observed no difference in response rates or OS between pts with newly diagnosed AML or R/R disease, which may be in part reflective of a population with high disease risk less likely to respond to upfront therapy. Additionally, we identify in a MVA disease factors predictive of response to therapy such as ELN cytogenetic risk and prior HMA exposure, the latter of which was not previously evaluated in clinical trials. Together, these observations highlight the clinical potential of ven combinations for R/R AML, as well as the need for novel therapeutic strategies to overcome the poor outcomes of pts with prior HMA exposure. Disclosures Patel: Celgene: Consultancy, Speakers Bureau; DAVA Pharmaceuticals: Honoraria; France Foundation: Honoraria; Agios: Consultancy.

Ivosidenib Improves Overall Survival Relative to Standard Therapies in Relapsed or Refractory Mutant IDH1 AML: Results from Matched Comparisons to Historical Controls

Ivosidenib Improves Overall Survival Relative to Standard Therapies in Relapsed or Refractory Mutant IDH1 AML: Results from Matched Comparisons to Historical Controls

Abstract 1920: Response to hypomethylating agents based on cytidine deaminase expression, genetic polymorphisms, and NPM1 mutation status in acute myeloid leukemia

Abstract Response to hypomethylating agents (HMAs) in patients (pts) with acute myeloid leukemia (AML) is variable. Data on biological predictors of response is limited. Cytidine deaminase (CDA) inactivates HMAs. Increased CDA activity may lead to HMA resistance. Using core pathway analysis, we identified nucleophosmin 1 (NPM1) indirectly influences CDA expression. We hypothesized that responses to HMAs occur in the setting of decreased CDA expression regulated by NPM1 and investigated the relationship between NPM1 status, CDA expression, single nucleotide polymorphisms (SNPs) in CDA, and HMA response in pts with AML. AML pts with banked samples who received HMA-based therapy between 1/2014 to 12/2018 were reviewed. Responses following at least 2 cycles of HMA were categorized as responders (R) or non-responders (NR). Pt, disease, NPM1 status, and treatment characteristics were summarized. CDA gene and protein expression was examined in bone marrow and peripheral blood samples at diagnosis using qRT-PCR and CDA sandwich ELISA, respectively. CDA gene expression levels were normalized to the housekeeping gene, 18s, and the comparative CT method was used to assess expression. Comparisons based on response and NPM1 mutation status were performed using the Mann-Whitney U test. 17 SNPs previously shown to alter CDA activity were selected for analysis. SNPs were determined using real-time PCR with allele specific probes; longer insertions/deletions were identified by sanger sequencing. Univariate logistic regression analysis was performed to discern the association between SNPs in CDA and response to HMAs. 54 pts had available blood, marrow, or buccal samples available for analysis. 33 pts provided blood or marrow samples for gene and protein analysis prior to HMA. 22 pts (67%) were classified as R in this cohort. 35 pts had available buccal swabs for genotyping, and 28 pts (80%) were classified as R. Median OS was 21 months (mo) for all pts, 23 mo among R, and 18 mo in NR. CDA expression was significantly decreased in NPM1 wild type pts compared to NPM1 mutant pts (p=0.02) but did not differ in R compared to NR. No significant differences were identified in CDA protein expression based on NPM1 status or response. No SNPs were significantly associated with response. Baseline CDA gene expression in bulk tumor cells was significantly lower in NPM1 wild type pts compared to NPM1 mutant pts but no different between pts responding to HMAs compared to NR. There was no clear correlation between CDA protein expression and NPM1 status or response. None of the CDA SNPs were predictive of response to HMAs. This analysis reveals feasibility of assessing CDA activity in this population. Our small sample size limits our ability to determine CDA activity as a biological predictor of response, and ongoing pt accrual will allow for further exploration of the role of CDA in HMA response. Citation Format: Brittany Knick Ragon, Issam S. Hamadeh, C Greer Vestal, Alicia Hamilton, Mathew L. Smith, Danielle Boselli, Jing Ai, Thomas G. Knight, Michael R. Grunwald, Jonathan M. Gerber, Edward A. Copelan, Lawrence J. Druhan, Belinda R. Avalos, Nury M. Steuerwald, Jai N. Patel. Response to hypomethylating agents based on cytidine deaminase expression, genetic polymorphisms, and NPM1 mutation status in acute myeloid leukemia [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 1920.

Post-remission clonal hematopoiesis; Practical implications for measurable residual disease assessment in acute myeloid leukemia (AML).

7529 Background: Clonal Hematopoiesis may persist following complete remission (CR) in patients with acute myeloid leukemia (AML) but does not necessarily indicate residual AML and may represent persistence of pre-leukemic stem cells. Post-remission CH identified by NGS has not been systemically studied in parallel with measurable residual disease (MRD) detection by flow cytometric immunophenotyping (FCI). Methods: We studied bone marrow sample from AML patients at baseline and CR by targeted deep NGS of 295 genes (median 403x depth) and compared the results to FCI. Measurable residual disease (MRD) detection by FCI was performed by comparing the phenotype at CR to baseline and by detection of leukemia associated immunophenotype (LAIP) and derivation from normal (DFN) (sensitivity: 0.1%). Post-CR CH was defined as presence of mutations originally detected in AML with variant allele frequency &gt; 2.5%. FCI results were categorized into 4 groups: a) AML MRD negative by LAIP or DFN b) AML MRD+ (similar to baseline) c) AML MRD+ (different from baseline), d) Negative for AML MRD, but aberrant phenotype suggestive of pre-leukemic cells. We correlated FCI and NGS results. Results: 101 patients were included in the study. 45 (45%) had persistent post-CR clonal hematopoiesis; 23 (51%) had phenotypic alterations detected by FCI including AML MRD+ in 18 (40%) and pre-leukemic cells in 5 (10%). Among patient with no detectable mutations by NGS (n = 56; 55%), 14 (25%) had FCI aberrancies including AML MRD+ in 4 (7%) and pre-leukemic cells in 10 (18%). CH was significantly more common in samples with residual phenotypic aberrancies detected by FCI (p = 0.004). There was no significant correlation between FCI group d and persistent CH (p = 0.4). Persistent ASXL1 (p = 0.024, OR = 7.2 ) and RUNX1 (p = 0.016; OR = 17.3) mutations were significantly associated with FCI abnormalities. The correlation coefficient between FCI abnormalities and RUNX1 mutations inferred from a Bayesian network structure was 0.66. Conclusions: NGS and FCI are complementary in evaluating post treatment disease status in AML. Post CR-CH is associated with phenotypic abnormalities that either represent residual AML or pre-leukemic cells. The latter may not have the same prognostic implications as AML MRD; however, the association with outcome needs to be elucidated. Single cell DNA sequencing technologies may be helpful in more accurately deciphering the association of individual gene mutations and their contribution to phenotypic aberrations.