Identify Acute Myeloid Leukemia Patients Research Articles

Evidence for identification of acute myeloid leukemia using peripheral blood's infrared spectrum and logistic regression

Invasive damage, poor prognosis and survival are problems for the traditional diagnostic and therapeutic methods of acute myeloid leukemia (AML), especially for the middle-aged and elderly. High sensitivity, less pain and fast speed are demands of AML diagnosis or long-term regular monitoring. Infrared (IR) spectroscopy has the potential to compensate for existing deficiencies and is explored in our work. Spectral differences between AML patients and controls are observed on the IR spectra, which indicate the changes in lipids and proteins in patients. The peak shift at 1390 cm−1 occurs on both peripheral blood (PB) and bone marrow (BM) spectra of AML patients making it possible to use PB rather than BM as the diagnostic source, which can relieve the suffering of diagnosis and monitoring. The logistic regression-based classification model can perfectly identify AML patients and controls with an accuracy of 100 % and the area under curve is equal to 1.00. Besides, some markers for identifying AML patients and controls are found. Statistical analysis shows that the peak area ratio A1451/A1390 has a significant difference, and the decision tree-based classification model finds that on the first derivative spectrum, the position of the largest difference is at 1391 cm−1. Furthermore, the proposed criterion based on markers we found can distinguish AML patients and controls with high accuracy of 95 %. Results show that PB’s IR spectrum combined with classification models can monitor the molecular-level biochemical changes and accurately identify AML patients and controls, which is helpful for the effective and efficient diagnosis and prognosis of AML.

C.202G > A/c.376A > G G6PD Polymorphisms Increase the Risk of Fungal Infections in Acute Myeloid Leukemia Patients

Introduction: Patients with acute myeloid leukemia (AML) show a higher risk for several types of infections, including fungal infections (FI), which are one of the main causes of morbidity and mortality. Glucose-6-phosphate dehydrogenase (G6PD) is an enzyme located in all cells that is very necessary in leukocytes for the production of basic and acid proteases that are used to destroy invading microorganisms. Our objective in this study was to evaluate whether polymorphisms in the G6PD gene concomitantly with FI are associated with clinical events and morbidity in patients diagnosed with AML and followed up at the Amazonas State Blood Center (HEMOAM), Manaus, Brazil. Materials and Methods: The study population was randomly constituted of adults and children, of either sex, and any age, with a diagnosis of acute myeloid leukemia, all of whom were undergoing treatment at the HEMOAM. Molecular genotyping was performed using real-time PCR (qPCR) and subsequent Sanger sequencing to confirm the c.202G > A/c.376A > G polymorphisms. Results: A total of 157 patients (91 (58%) males and 66 (42%) females) were involved in the study. The most prevalent AML subtype in the studied group was M3 in 63 patients (40.12%), followed by M5 in 33 patients (21.02%), M2 in 21 patients (13.37%) and M4 in 15 patients (9.55%), with a similar prevalence between genders. The prevalence of fungal infections was identical between genders; however, bruising (p = 0.004), vomiting (p = 0.016) and cardiac alterations (p < 0.001) were higher in females, while persistent cough (p = 0.049) and diarrhea (p < 0.001) were higher in males. A total of eighteen patients presents G6PD polymorphisms, with 8 (5.1%) of these for c.202GA/AA, 18 (11.5%) for c.376AG/GG and 4 (2.5%) for both polymorphisms concomitantly (c.202AA/c.376GG). However, the prevalence of death in patients affected with FI was much higher in those that have these polymorphisms (p < 0.001). Conclusion: We believe that the determination of G6PD polymorphisms will allow the development of monitoring strategies, and aid in early diagnosis and the appropriate and targeted treatment for AML. In addition, evaluating their activity may help to identify AML patients at a higher risk of FI, thus allowing the design of more intensive therapeutic and surveillance strategies.

ELN 2022 Favorable Genetic Risk Acute Myeloid Leukemia Treated with Venetoclax Plus Hypomethylating Agent: Predictors of Response and Survival

Background: The 2022 European LeukemiaNet (ELN) genetic-risk stratification for acute myeloid leukemia (AML), assigns favorable prognosis to t(8;21)(q22;q22.1)/ RUNX1:: RUNX1T1, inv(16)(p13.1q22) or t(16;16)(p13.1;q22)/ CBFB:: MYH1, NPM1 mutations without FLT3-ITD or adverse karyotype, and bZIP in-frame CEBPA mutations (Döhner , Blood, 2022 ). This risk stratification was derived from intensively treated patients; accordingly, prognostication in ELN favorable genetic risk AML, following venetoclax and hypomethylating agent (VEN-HMA) therapy, remains unclear and was the subject of the current study. Methods: Under an institutional review board approved minimum risk protocol, the Mayo Clinic database was searched to identify AML patients with favorable genetic risk, who received treatment with VEN-HMA in the frontline setting. All patients received at least one cycle of either azacitidine 75 mg/m 2 days 1-7 or decitabine 20 mg/m 2 days 1-5 with VEN dose adjusted based on azole antifungal prophylaxis. Genetic risk stratification and treatment response were assessed according to the 2022 ELN criteria. Survival was estimated from the time of VEN-HMA treatment start to last follow-up or death. Results: Patient characteristics A total of 46 newly-diagnosed AML patients with ELN 2022 favorable genetic risk [ NPM1 mutation, n=36 (78%), CEPBA bZIP mutation, n=7 (15%) and t(8;21)(q22;q22.1)/ inv(16)(p13.1q22), n=4 (9%)] received a median of 4 cycles (range 1-32) of VEN-HMA. Median age was 71 years, with 63% males, and 87% de novo AML. Additional mutations frequently involved TET2 in 17/43 (40%), DNMT3A in 14/45 (31%), SRSF2 in 10/43 (23%), K/NRAS in 10/45 (22%), IDH1 in 8/46 (17%), FLT3-TKD in 7/46 (15%), ASXL1 in 6/43 (14%) and IDH2 in 6/46 (13%),of informative cases.Myelodysplasia (MDS)-related gene mutations ( ASXL1, RUNX1, BCOR, EZH2, SRSF2, U2AF1, SF3B1, STAG2, and/or ZRSR2) were present in 15 of 40 (38%) of patients (93% males). Predictors of response Of the 39 (85%) patients that achieved complete remission, with (CR) ( n=24; 52%), or without count recovery (CRi) ( n=15; 33%); CR/CRi rates were not significantly higher in patients with NPM1 mutations (89%), t(8;21)(q22;q22.1)/ inv(16)(p13.1q22) (75%), or CEPBA bZIP mutations (67%) ( p=0.37). Among patients achieving CR/CRi, measurable residual disease by multiparametric flow cytometry was negative in 15 of 16 (94%) of evaluable cases after cycles 1 or 2. In addition, NPM1 qPCR was negative in 15 of 24 (63%) informative patients. Median time to response was 1.2 months with median response duration of 9.4 months. CR/CRi rates were significantly higher in females vs males (100% vs 76%, p=0.01), and in the absence vs presence of MDS-related gene mutations (96% vs 60%, p=0.004). Multivariable analysis confirmed superior response in the absence of MDS-related gene mutations ( p=0.04). Predictors of survival At a median follow-up of 10.6 months (range; 1.1-51 months), 21 patients (46%) have died, 9 relapsed (23% of those achieving CR/CRi), and 6 (13%) underwent allogeneic hematopoietic stem cell transplant. Median overall survival was 21 months with 1/2/3-year survival rates of 65%/41%/41% (Figure 1a); median relapse-free survival (RFS) was not reached; 1/2/3-year RFS rates were 81%/66%/57% (Figure 1b). Overall survival was superior in patients achieving CR/CRi (23 months vs 5.6 months; p<0.01), in females vs males (not reached vs 9.7 months; p<0.01), and in the absence of MDS-related gene mutations (not reached vs 5.6 months; p=0.01) (Figure 1c). Subsequent multivariable analysis identified female gender ( p=0.01), and absence of MDS-related mutations ( p=0.17) as independent predictors of superior survival. Similar results were obtained when analysis was restricted to NPM1 mutated patients. Conclusions: In the current retrospective study, VEN-HMA therapy in elderly/unfit AML patients with ELN 2022 favorable genetic risk, resulted in response rates that were comparable to those previously appreciated from published reports of patients receiving intensive induction therapy; however, follow-up in the current study was too short to accurately assess long-term overall and relapse-free survival impact. Our study also highlights the adverse prognosis retained by MDS-related gene mutations.

Leveraging machine learning to identify acute myeloid leukemia patients and their chemotherapy regimens in an administrative database.

Administrative datasets are useful for identifying rare disease cohorts such as pediatric acute myeloid leukemia (AML). Previously, cohorts were assembled using labor-intensive, manual reviews of patients' longitudinal chemotherapy data. We utilized a two-step machine learning (ML) method to (i) identify pediatric patients with newly diagnosed AML, and (ii) among the identified AML patients, their chemotherapy courses, in an administrative/billing database. Using 2558 patients previously manually reviewed, multiple ML algorithms were derived from 75% of the study sample, and the selected model was tested in the remaining hold-out sample. The selected model was also applied to assemble a new pediatric AML cohort and further assessed in an external validation, using a standalone cohort established by manual chart abstraction. For patient identification, the selected Support Vector Machine model yielded a sensitivity of 0.97 and a positive predictive value (PPV) of 0.97 in the hold-out test sample. For course-specific chemotherapy regimen and start date identification, the selected Random Forest model yielded overall PPV greater than or equal to 0.88 and sensitivity greater than or equal to 0.86 across all courses in the test sample. When applied to new cohort assembly, ML identified 3016 AML patients with 10,588 treatment courses. In the external validation subset, PPV was greater than or equal to 0.75 and sensitivity was greater than or equal to 0.82 for patient identification, and PPV was greater than or equal to 0.93 and sensitivity was greater than or equal to 0.94 for regimen identifications. A carefully designed ML model can accurately identify pediatric AML patients and their chemotherapy courses from administrative databases. This approach may be generalizable to other diseases and databases.

Pre-transplant minimal residual disease assessment and transplant-related factors predict the outcome of acute myeloid leukemia patients undergoing allogeneic stem cell transplantation.

We studied pretransplant minimal residual disease (MRD) in 224 patients (median age 44years; range 17-65) with acute myeloid leukemia (AML) undergoing allogeneic stem cell transplant (HSCT) in complete remission. MRD was evaluated on marrow samples using multicolor flow cytometry and assessment of WT1 gene expression. Both methods showed a strong prognostic value and their combination allowed the identification of three groups of patients with different risk of relapse. In multivariate analysis, combined MRD was the only predictor of cumulative incidence of relapse, regardless of donor type, conditioning regimen, first or second CR at HSCT, HSCT year, and ELN risk group. Multivariate regression model showed that only negative combined MRD status (P<.001) and myeloablative conditioning (P=.004) were independently associated with better OS. Among MRD-positive patients, a reduced incidence of relapse was observed in patients receiving haplo transplant (P<.05) and in patients who showed grade II-IV aGVHD (P<.03). In patients with negative combined MRD, the intensity of conditioning regimen did not affect the overall favorable outcome. We suggest that pretransplant MRD evaluation combined with transplant-related factors can identify AML patients at higher risk for relapse and might help in defining the overall transplant strategy.

ELN 2017 Genetic Risk Stratification Predicts Survival of Acute Myeloid Leukemia Patients Receiving Allogeneic Hematopoietic Stem Cell Transplantation

European LeukemiaNet (ELN) 2017 risk stratificationbygenetics is prognostic of outcomes in patients with acute myeloid leukemia (AML). However, the prognostic impact of the 2017 ELN genetic risk stratification after allogeneic hematopoietic cell transplantation (alloHCT) is not well established. We examined the effect of 2017 ELN genetic risk stratification on alloHCT outcomes of AML. We included 500 adult (≥18 years) AML patients in first (n=370) or second (n=130) complete remission receiving alloHCT from 2005 to 2016. Patients were classified into favorable (12%), intermediate (57%), and adverse (32%) 2017 ELN risk groups. The Cox proportional hazard model was used to conduct the multivariable analyses of leukemia-free survival (LFS) and overall survival (OS). Relapse and nonrelapse mortality were analyzed by the Fine-Gray regression model. OS at 2 years was 72% in the favorable versus 60% in the intermediate versus 45% in the adverse risk groups (P < .001). In multivariable analyses, the 2017 ELN classifier was an independent predictor of OS after alloHCT with significantly higher overall mortality in the intermediate (hazard ratio [HR]=1.68, 95% confidence interval [CI], 1.06-2.68; P=.03) and adverse (HR=2.50, 95% CI, 1.54-4.06; P < .001) risk groups compared to the favorable risk group. Similarly, LFS was worse in the intermediate (HR=1.63, 95%, CI 1.06-2.53; P=.03) and adverse (HR 2.23, 95% CI, 1.41-3.54; P < .001) risk groups while relapse was higher in the adverse risk group (HR=2.36, 95% CI, 1.28-4.35; P=.006) as compared to the favorable risk group. These data highlight the prognostic impact of the 2017 ELN genetic risk stratification on the survival of AML patients after alloHCT. Patients in the adverse risk group had the highest risk of relapse and worst survival. Thus the 2017 ELN prognostic system can help identify AML patients who may benefit from clinical trials offering relapse mitigation strategies to improve transplant outcomes.

Interleukin-27 and interleukin-35 in de novo acute myeloid leukemia: expression and significance as biological markers.

Background and objectivesIL27 and IL35 are regulatory T cells (T-regs) related cytokines; they were accused in eukemogenesis of acute myeloid leukemia (AML). This study aimed to assess the expression of these cytokines in de novo AML and investigate their role as biomarkers.Subjects and methodsSeventy newly diagnosed patients with primary AML and 30 matched healthy volunteers were recruited. AML diagnosis was confirmed with flowcytometric and immunophenotypic analyses, while ELISA was used to assess serum levels of IL27 and IL35 in patients and controls. Receiver operating characteristic curve analysis was used to estimate IL27 and IL35 optimum cutoff values for predicting AML.ResultsSerum levels of both cytokines were significantly higher in AML patients than controls (P<0.001), with no effect of gender or French-American-British subtypes. Significant correlations of IL27 and IL35 with poor prognostic factors and with each other were detected in patients only. IL27 optimum cutoff for predicting AML was >43, AUC (0.926) with a sensitivity 74% and specificity 96.6% (P<0.001), while for IL35>27.8, AUC (0.972) with 88% and 98% sensitivity and specificity, respectively (P<0.001).ConclusionConclusively, this study proved that IL27and IL35 could identify AML patients from healthy subjects, and their overexpression denotes poor prognosis. Based on the simplicity and wide availability of their detection technique we recommend the inclusion of IL27 and IL35 in the diagnostic/prognostic workup of AML; however, further longitudinal research is needed to prove their exact prognostic value.

Role of IL27 and IL35 as biomarkers and potential targets in acute myeloid leukemia.

e18516 Background: Acute myeloid leukemia (AML) is one of the most common leukemia affecting adult. Regulatory T cells (Tregs) are a subset of CD4 (+) T cells that plays a central role immune tolerance. Objective: We aimed to explore the role of interleukin- 27 (IL27) &amp; interleukin- 35 (IL35) produced by T regs and its associations with clinical characteristics and prognostic markers of AML patients. Methods: The study included 70 patients with acute myeloid leukemia (AML) and 30 healthy controls. We used flowcytometry immunophenotyping to confirm the diagnosis of AML. Serum level of interleukin- 27 (IL 27) &amp; interleukin- 35 (IL 35) were measured at diagnosis by enzyme-linked immunosorbentassay (ELISA). Results: Higher levels of IL27 and IL35 were detected in AML patients than the control group (p ≤ 0.001). We used Receiver operating characteristic (ROC) analysis for detection of optimum IL27 &amp; IL35 cut off level needed for predicting AML. ROC curve analysis demonstrated IL27 optimum cutoff is &gt; 43, AUC 0.926 with a sensitivity 74% and Specificity 96.6% (P-value &lt;0.00). IL35 optimum cutoff was &gt; 27.8 AUC 0.972 with 88% and 98% sensitivity and specificity respectively (P-value &lt;0.001). Serum IL27 &amp; IL35 could well identify AML patients from healthy individuals. More importantly IL27 &amp; IL35 correlated with clinical variable, and could be served as an independent prognostic factor. Conclusions: IL27 &amp; IL 35 might have an application as a diagnostic and prognostic indicator for AML patients and could be served as an independent prognostic factor.

ELN 2017 Risk Classification Predicts Survival of AML Patients Receiving Allogeneic Hematopoietic Stem Cell Transplantation

European LeukemiaNet (ELN) 2017 updated prognostic system for acute myeloid leukemia (AML), which incorporates both cytogenetic and molecular risks, stratifying patients into 3 distinct genetic risk groups: favorable, intermediate and adverse. While most patients in favorable risk group can be cured with chemotherapy alone, the majority of patients in intermediate and adverse genetic risk groups are considered for curative allogeneic hematopoietic stem cell transplantation (alloHCT). However, the impact of ELN 2017 risk classification on survival of AML patients after alloHCT remains unclear. We examined the prognostic significance of ELN 2017 classification on 503 AML patients in first or second complete remission (373 CR1/ 130 CR2) receiving alloHCT at Moffitt Cancer Center) from 2005-2016. Patients were classified into favorable (n=58, 11.6%), intermediate (n=284, 56.8%) and adverse (n=158, 31.6%) ELN risk groups. The median age at alloHCT for the entire cohort was 55 years (range, 18-76 years) and 29% of all patients had many comorbidities (HCT ≥3) at transplant. Disease risk index (DRI) was low in 38 (7.6%), intermediate in 393 (78.6%) and high in 69 (13.8%) patients, and 22% (n=109) of all patients had secondary AML. Myeloablative conditioning was used in 77% (n=389) of patients, while 23% (n=114) received reduced-intensity conditioning regimen. HLA-matched unrelated donor (47%) was the most commonly used donor type, followed by matched related (27.7%) and mismatched donors (n=75, 9.8%). For the entire cohort, 2-year overall survival (OS) was 56% (52-60%) and leukemia-free survival (LFS) was 51% (47-55%). Patients with CR1 and CR2 status at alloHCT had similar OS (58% vs. 53%, p=0.17). We identified that ELN 2017 genetic risk is prognostic of OS after alloHCT in patients with AML: 72% (61-84%) in favorable risk vs. 60% (54-66%) in intermediate risk vs. 45% (38-53%) in adverse risk groups (p Our findings highlight the prognostic impact of ELN 2017 genetic risk on survival of AML patients undergoing alloHCT in CR1/CR2. Patients in the adverse risk group had higher risk of relapse and worse survival. Thus, ELN 2017 prognostic system can help identify AML patients in CR1/CR2 who can benefit from clinical trials offering relapse reduction strategies in order to improve their survival.

High Expression of SPAG1 Is Associated with a Worse Clinical Outcome in Intermediate Risk Acute Myeloid Leukemia That Can be Partially Overcome By Hematopoietic Stem Cell Transplantation

Introduction: Acute myeloid leukemia (AML) is a heterogeneous disease with variable responses to therapy and clinical outcomes. Cytogenetics and molecular analyses help to stratify patients and to select therapy, especially regarding indication of hematopoietic stem cell transplant (HSCT) after achieving complete remission (CR). Patients in the intermediate cytogenetic risk category (~40%) represent a clinical dilemma regarding consolidation because of the high rate of relapse with chemotherapy alone and high rate of morbidity/mortality with HSCT. Consequently, we aimed to identify new prognostic markers to refine the risk stratification of this patients' subgroup and to identify which patients are most likely to benefit from HSCT.Methods: We analyzed RNA sequencing data of 263 specimens from patients with de novo AML treated with curative intent including 165 patients with intermediate risk cytogenetics. Data from 24 586 genes were normalized as RPKM values with logarithmic transformation and standardization. Cox proportional hazard models were used to assess the prognostic impact of gene expression (GE) on overall survival (OS) and relapse-free survival (RFS). We performed univariate analyses (UVA) and multivariate analyses (MVA) adjusted for age and white blood cell count (WBC) at diagnosis, mutations in NPM1, FLT3, CEBPA, RUNX1, ASXL1, TP53 and DNMT3A and HSCT as a time-dependent (TD) covariate. Interaction between GE and TD-HSCT was tested in MVA for OS and RFS. Genes with a significant interaction between GE and TD-HSCT (p < 0.10) were retained for further analyses. GE of candidate markers were dichotomized using a bioinformatic method assessing hazard ratios (HR) and p values for all potential cut-offs to identify the most optimal threshold. The markers were finally reassessed as dichotomic variables for association with covariates, CR rates, RFS and OS. All statistical tests were two-sided with p values < 0.05 considered significant.Results: We identified SPAG1 (Sperm Associated Antigen 1) as the gene with the highest HR for OS and RFS in the intermediate cytogenetic risk group. SPAG1 expression was dichotomized on the median RPKM value in the global cohort of 263 de novo AML specimens (RPKM cut-off 2.06). Using this cut-off, 79 patients had low expression of SPAG1 and 86 patients had high expression of SPAG1 in the intermediate cytogenetic risk cohort. Median age, sex, WBC at diagnosis and HSCT in CR1 rates were similar between the two groups. Patients with high expression of SPAG1 had a higher frequency of FLT3-ITD (51.2% vs 32.9%, p = 0.02) and DNMT3A mutations (51.2% vs 32.9%, p = 0.02). SPAG1-high patients were enriched in the NPM1/FLT3-ITD/DNMT3A triple positive mutation population (SPAG1-high 24/33 vs SPAG1-low 9/33, OR 3.00, p = 0.01). The frequencies of all other analyzed mutations were similar between both groups. CR rates did not differ between the two groups (SPAG1-high 77.9% vs SPAG1-low 79.7%, p = 0.77). In UVA with censorship at time of HSCT, SPAG1-high patients had worse OS (5-year estimates 14.2% vs 28.1%, HR 1.75, 95% CI 1.16-2.63, p < 0.01) and RFS (5-year estimates 9.3% vs 27.1%, HR 1.90, 95% CI 1.20-3.01, p < 0.01) (Figure 1). In MVA with censorship at time of HSCT, high expression of SPAG1 remained significantly associated with OS (HR 1.78, 95% CI 1.12-2.83, p = 0.01) and RFS (HR 2.34, 95% CI 1.38-3.96, p < 0.01). Furthermore, there was a significant interaction between SPAG1 GE and TD-HSCT (p = 0.09) in the RFS model. Importantly, SPAG1 had a lower prognostic impact for RFS in the model including TD-HSCT (HR 1.65, 95% CI 1.07-2.55, p = 0.02) compared with the model in which survival was censored at time of HSCT (HR 2.34, p < 0.01). High SPAG1 expression was also associated with worse OS in the TCGA AML dataset which is enriched in intermediate cytogenetic risk samples (p < 0.001).Conclusion: In patients with intermediate cytogenetic risk AML, high expression of SPAG1 is independently associated with worse OS and RFS. The prognostic impact of SPAG1 expression on RFS is lower when adjusted for TD-HSCT indicating that the adverse prognosis conferred by high expression of this gene may be partially overcome by HSCT in CR1. Consequently, SPAG1 expression might help identify AML patients with intermediate cytogenetic risk who are most likely to benefit from HSCT in CR1. These results need to be validated in other independent cohorts and prospective studies before implementation into clinics. [Display omitted] DisclosuresSauvageau:ExCellThera: Employment, Equity Ownership.

Predictive Response Biomarkers for BET Inhibitors in AML

BACKGROUND: Despite mounting knowledge, the management of patients with acute myeloid leukemia (AML) can be challenging, particularly for relapsed/refractory patients who lack innovative, effective treatments. Recent preclinical studies of AML have demonstrated promissory therapeutic activity of the novel class of inhibitors of bromodomain and extra terminal (BET) family proteins. To identify AML patients that may benefit from BET inhibitor treatment, we sought to discover and evaluate predictive molecular biomarkers for sensitivity towards BET inhibition, focusing on the prototype BET inhibitor JQ1.METHODS: Following written informed consent and according to the Declaration of Helsinki, bone marrow, peripheral blood, and skin biopsy samples were obtained prospectively from AML patients (n=139) and analyzed for their whole exome profiles and ex vivo responses to JQ1 exposure, as part of an Individualized Systems Medicine (ISM) study of AML (Pemovska et al., 2013). Consequently, associations between molecular features and drug responses were investigated using logistic regression models for data integration. Novel signals emerging from our cohort of patients were carried forward for reproducibility assessment in publicly available cell line datasets (Cancer Cell Line Encyclopedia and Catalogue of Somatic Mutations in Cancer for genomic data, and Cancer Therapeutics Response Portal and Genomics of Drug Sensitivity in Cancer for drug response data). In addition, literature-reported indicators of sensitivity were examined in the AML cohort in order to evaluate their clinical translatability.RESULTS: Enabling a genome-wide search for predictive biomarkers for BET inhibitor response, our bioinformatics pipeline returned a set of signals significantly associated with JQ1 treatment.Frequently mutated in AML, isocitrate dehydrogenases IDH1 and IDH2, tet methylcytosine dioxygenase TET2, and Wilms' tumor WT1 each appeared to confer resistance upon mutation to the samples harboring them. Participating in a common biochemical pathway, the 4 genes have been shown to be mutated following a mutual exclusivity pattern in AML (Scourzic et al., 2015), which was successfully replicated in our cohort. Accordingly, grouping of samples with mutation in any of the genes yielded a significantly more resistant group to JQ1 treatment in comparison to wild-type samples. Incorporation of TET2 and WT1 deletions from the samples' copy number variation data supported this hypothesis. Importantly, this observation was also successfully validated in cell line drug sensitivity data (Figure 1A).Moreover, FLT3-mutated samples in our cohort returned significantly higher sensitivity scores to JQ1 compared to wild-type samples, irrespective of their NPM1 mutation status. The superior response trend observed within this group was maintained independently of whether the mutation was an internal tandem duplication (FLT3-ITD) or in the tyrosine kinase domain (FLT3-TKD) of the gene (Figure 1B).Finally, and in contrast to other studies, AML samples harboring mutation to NPM1 were not significantly more sensitive to BET inhibition with respect to NPM1 wild-type samples.CONCLUSION: Taken together, the results suggest novel insights for the stratification of AML patients for treatment using BET inhibition therapy, taking this malignancy's heterogeneity into account. Particularly, while FLT3 mutations are to be considered as sensitizing factors to BET inhibitors, samples with mutations in IDH1, IDH2, TET2, or WT1 were resistant, and thus, patients harboring these gene mutations may not benefit. These observations also propose drug partners that are worth investigating in combination with BET inhibitors in the defined patient subpopulations, namely FLT3 inhibitors and hypomethylating agents (or IDH1/2 inhibitors), respectively. Furthermore, the variability of responses observed among samples harboring NPM1 mutations highlights the molecular heterogeneity existing within this group of patients, stressing the need for identifying additional companion diagnostics for a more effective stratification of this population. Although our findings require additional validation in a larger cohort and through clinical investigations, this study provides evidence of molecular indicators that could help identify AML patients that would benefit from BET inhibitor therapy. [Display omitted] DisclosuresTsallos:Novartis: Research Funding. Wennerberg:Novartis: Research Funding. Porkka:Celgene: Honoraria, Research Funding; Novartis: Honoraria, Research Funding. Heckman:Novartis: Research Funding; Orion Pharma: Research Funding; Celgene: Research Funding.

Emerging Real-World Midostaurin Treatment Patterns and Outcomes in FLT3-Mutated Acute Myeloid Leukemia at a Comprehensive Cancer Center

Introduction: Fms-like tyrosine kinase 3 (FLT3) gene mutation occurs in approximately 25-30% of acute myeloid leukemia (AML) cases and is associated with poor prognosis. Decreased overall survival is reported in FLT3-mutated vs. FLT3-wildtype. Midostaurin, a pan-targeted kinase inhibitor that inhibits activated FLT3 received FDA approval in April 2017 for adult patients with newly diagnosed FLT3-mutated AML in combination with standard cytarabine and daunorubicin induction and cytarabine consolidation. We report descriptive US clinical treatment patterns and outcomes in FLT3-mutated early midostaurin-users, historical FLT3-mutated patients prior to midostaurin approval, and historical FLT3-wildtype AML patients all treated with 7+3 induction therapy at an academic cancer specialty hospital to support the development of a larger database across several comprehensive cancer centers.Methods: This retrospective, observational study utilized ICD codes, tumor registry data, and pharmacy records from the Huntsman Cancer Institute (HCI) to identify AML patients treated with 7+3 induction chemotherapy from 2007 to July 2018. FLT3-mutated midostaurin-users treated with 7+3 induction including midostaurin from May 2017 to July 2018 comprise Group 1. Historical FLT3-mutated patients prior to midostaurin approval (non-users) and FLT3-wildtype patients comprise Groups 2 and 3, respectively. Complete response (CR), relapse rates, overall survival and treatment patterns were described.Results: A total of 105 patients met eligibility for inclusion in the study. Groups 1, 2, and 3 included five, 39 and 61 patients, respectively. Off-label midostaurin use in Group 1 after induction therapy was observed in one patient with post-consolidation monotherapy and salvage therapy in combination with ATRA (tretinoin) and CLAG (cladribine, cytarabine, and filgrastim). Following midostaurin approval, two FLT3-mutated patients received induction therapy without midostaurin due to enrollment in clinical trials that excluded midostaurin use for induction and consolidation therapy. These two patients were excluded from the study. Descriptive results of the comparative groups are summarized in Table 1. CR rate from induction therapy was 100% for Group 1, 90% for Group 2, and 77% for Group 3. The proportion of patients who received consolidation therapy was 60%, 74%, and 67%, and patients who maintained CR during consolidation therapy was 100%, 83%, and 49% for Groups 1, 2, and 3, respectively. Sixty-six percent of eligible Group 1 patients, 74% of Group 2 patients, and 54% of Group 3 patients received transplant. Median time from diagnosis to transplant was 81, 99, and 145 days for Groups 1, 2, and 3, respectively. The proportion of patients who received salvage therapy in Groups 1, 2, and 3 was 20%, 38%, and 56% respectively. Median follow-up was 6 months for Group 1, 14 months for Group 2, and 24 months for Group 3. After CR, 20% of Group 1, 49% of Group 2, and 59% of Group 3 relapsed. All Group 1 patients were alive at time of analysis while four Group 2, and eight Group 3 patients died during the study period.Discussion: Similar CR and relapse rates were observed between the comparative groups, although early use observations indicate improved response rates of induction therapy and consolidation therapy in a limited Group 1 sample. Patients in Group 1 and Group 2 underwent transplant earlier and more frequently than patients in Group 3, which may explain the higher relapse rate in Group 3. As this data resource is expanded across similar institutions, statistical comparisons of FLT3-mutated AML patients treated with and without midostaurin can be made.Sponsorship: Funding for this study was provided by Novartis Pharmaceuticals. DisclosuresUnni:Novartis: Research Funding. Ndife:Novartis: Employment. Joseph:Novartis Pharmaceuticals Corporation: Employment; Amgen: Equity Ownership; Pfizer: Equity Ownership; Express Scripts: Equity Ownership. Bonifacio:Novartis: Employment. Stein:Novartis: Consultancy; Daiichi Sankyo: Consultancy; Agios: Consultancy; Bayer: Consultancy; Celgene: Consultancy; Pfizer: Consultancy. Shami:Lone Star Biotherapies: Equity Ownership; Pfizer: Consultancy; JSK Therapeutics: Employment, Equity Ownership, Membership on an entity's Board of Directors or advisory committees; Baston Biologics Company: Membership on an entity's Board of Directors or advisory committees. Kovacsovics:Amgen: Honoraria, Research Funding; Abbvie: Research Funding. Brixner:BD: Consultancy; Abbott: Consultancy; AbbVie: Consultancy; AstraZeneca: Consultancy; Gilead: Consultancy; Millcreek Outcomes Group: Equity Ownership; University of Utah: Research Funding. Stenehjem:Novartis: Research Funding.

Quantitative Monitoring By RT-PCR of Molecular Markers on Day +100 after Allogeneic Stem Cell Transplantation Predicts Outcome in Patients with Acute Myeloid Leukemia

Introduction:Monitoring of minimal residual disease (MRD) is commonly used after allogeneic stem cell transplantation (SCT) in acute myeloid leukemia (AML) patients. Flow cytometry of leukemia-associated aberrant immunophenotypes (LAIP) and chimerism analyses are frequently used tools to monitor MRD. These methods have lower sensitivities and lower relapse detection rates in contrast to molecular markers monitored by quantitative RT-PCR. However, their clinical implications are still limited and only little evidence of single molecular markers is available. We report on a retrospective study of 144 patients with molecular MRD markers transplanted at the Ludwig-Maximilans-University Hospital of Munich-Grosshadern, and analyzed the prognostic values of molecular MRD Monitoring after SCT, baseline risk factors, and follow-up (FU) markers (e.g. chimerism analyses and LAIP).Patients and Methods:Between January 2000 and August 2012, 495 AML patients underwent SCT at our institution. 164 patients had molecular MRD markers and 144 patients had at least one sample with quantitative results prior and/or after SCT, and were included into the analyses. At transplantation, patients had a median age of 48 years (range, 18 -73), and 49 patients were in complete remission, while 95 patients were in a more advanced status of their disease (>CR2). 106 patients received SCT from a HLA matched (10/10) and 38 patients from a HLA mismatched donor. Grafts were obtained either from related (n=60) or unrelated (n=84) donors. Most patients received reduced intensity conditioning (n=142). In 338 bone marrow samples MRD was monitored prior to SCT, on day +30 and on day +100. During the FU period after day +100, MRD was monitored at individual intervals in 429 peripheral blood samples. Quantitative RT-PCR was performed for NPM1 mutation (n=52), MLL-PTD (n=31), RUNX1-RUNX1T1 (n=12), CBFß-MYH11 (n=14), MLL rearrangements (n=20), MDS-EVI1/EVI1 (n=10), and DEK-CAN (n=5). Sensitivities of the different RT-PCRs assays ranged between 10-4 and 10-6.Results:After a median FU of 41 months (range, 4 – 115), 43 patients (30%) relapsed. The MRD levels monitored by RT-PCR prior to SCT and on day +30 after SCT showed no significant impact on relapse-free survival (RFS) and overall survival (OS). At day +100 after SCT, MRD positivity was strongly associated with worse RFS (HR 3.1, p=0.001) and OS (HR 3.2, p=0.004). This was also reflected in a cumulative two-year incidence of relapse of 61% for MRD positive patients versus 15% for MRD negative patients (p<0.0001). In addition, a change to MRD positivity during the FU period was strongly associated with a worse RFS (Mantel-Byar, p<0.0001). Furthermore, in univariate regression models we analyzed SCT baseline factors and FU markers. The remission status prior to SCT, the NPM1 mutation status, a HLA matched donor, the platelet count at day +100 (cut-off <50 G/l), and the European LeukemiaNet risk stratification (favorable versus others) were prognostic relevant risk factors. In the multivariate model only MRD positivity on day +100 (HR 3.7, p=0.013) and a low platelet count on day +100 (HR 5.0, p=0.005) were significantly associated with worse RFS. LAIP analyses (3-colors flow-cytometry, cut-off 0.1%) and a change in chimerism (STR-PCR and XY-FISH, cut-off 80% and 90%) of sorted (CD3+) and unsorted cells at the respective time points had no influence on RFS and OS.Conclusions:Molecular MRD monitoring after SCT might be a useful tool to identify AML patients at high risk for relapse, in contrast to other FU markers, e.g. chimerism analyses and LAIP. In particular, MRD positivity on day +100 after SCT and the switch to MRD positivity during the FU period were significantly associated with worse RFS. As perspective, the individual molecular MRD course might be used as prognostic factor for the guidance of treatment post SCT. Patients with MRD positivity on day +100 after SCT or with a switch to MRD positivity in the FU period may be considered for donor lymphocyte infusions (DLI) or chemotherapeutic interventions, such as hypomethylating agents. DisclosuresSubklewe:AMGEN Inc.: Research Funding.

Specific cellular signal-transduction responses to in vivo combination therapy with ATRA, valproic acid and theophylline in acute myeloid leukemia

Acute myeloid leukemia (AML) frequently comprises mutations in genes that cause perturbation in intracellular signaling pathways, thereby altering normal responses to growth factors and cytokines. Such oncogenic cellular signal transduction may be therapeutic if targeted directly or through epigenetic regulation. We treated 24 selected elderly AML patients with all-trans retinoic acid for 2 days before adding theophylline and the histone deacetylase inhibitor valproic acid (ClinicalTrials.gov NCT00175812; EudraCT no. 2004-001663-22), and sampled 11 patients for peripheral blood at day 0, 2 and 7 for single-cell analysis of basal level and signal-transduction responses to relevant myeloid growth factors (granulocyte-colony-stimulating factor, granulocyte/macrophage-colony-stimulating factor, interleukin-3, Flt3L, stem cell factor, erythropoietin, CXCL-12) on 10 signaling molecules (CREB, STAT1/3/5, p38, Erk1/2, Akt, c-Cbl, ZAP70/Syk and rpS6). Pretreatment analysis by unsupervised clustering and principal component analysis divided the patients into three distinguishable signaling clusters (non-potentiated, potentiated basal and potentiated signaling). Signal-transduction pathways were modulated during therapy and patients moved between the clusters. Patients with multiple leukemic clones demonstrated distinct stimulation responses and therapy-induced modulation. Individual signaling profiles together with clinical and hematological information may be used to early identify AML patients in whom epigenetic and signal-transduction targeted therapy is beneficial.

Relationship Between CD56 Antigen Expression and Quantification of MDR1 Gene Expression in Patients with De Novo Acute Myeloid Leukemia(AML).

Abstract 4677 IntrodutionThe expression of CD56 antigen and over-expression of the multidrug resistance gene 1 (MDR1) seems to confer the poor therapeutic outcome to AML. The aim of this study is to investigate the relationship between CD56 antigen expression and MDR1 gene expression in patients with de novo AML and explore the indicating effect of these two factors on clinical drug resistance. Patients and methodsA real-time quantitative reverse transcriptase polymerase chain reaction method was established for detecting MDR1 expression levels and a three-color flow cytometry analysis with CD45/SSC gating was used to examine CD56 antigen expression in 79 patients with de novo AML. ResultsCD56 antigen was recorded in 19 out of 79 cases (24.1%) and particularly in those with M5 subtype and t(8;21) AML. Moreover, CD56 expression was significantly associated with unfavorable cytogenetic abnormalities (P< 0.05). Significantly higher percentage (57.1%, 4/7) of patients with t(8;21) demonstrated CD56 expression than those with favorable cytogenetic abnormalities (P< 0.05). CD56+ AML patients had higher incidence of splenohepatomegalia and level of lactate dehydrogenase than CD56- patients (P< 0.05). The median expression level of MDR1 was statistically higher in CD56+ AML patients than that in CD56- cases (P< 0.001) and 89.5% (17/19) CD56+ AML patients were found with high MDR1 expression. The CR rate in high MDR1 / CD56+ AML patients was significantly lower than that in low MDR1/ CD56- cases. (58.8% vs 89.2%, P< 0.01). ConclusionsThere is a linear correlation between MDR1 and CD56 expression in AML. This relationship may explain why CD56 expression is related to a poor prognosis in AML. Therefore both with high MDR1 expression level and CD56 antigen expression can identify AML patients with unfavorable outcome. Disclosures:No relevant conflicts of interest to declare.