Therapy For Acute Myeloid Leukemia Patients Research Articles

Aberrant ecotropic viral integration site-1 (EVI-1) and myocyte enhancer factor 2C gene (MEF2C) in adult acute myeloid leukemia are associated with adverse t (9:22) & 11q23 rearrangements.

Acute myeloid leukemia (AML) shows multiple chromosomal translocations & point mutations which can be used to refine risk-adapted therapy in AML patients. Ecotropic viral integration site-1 (EVI-1) & myocyte enhancer factor 2C gene (MEF2C) are key regulatory transcription factors in hematopoiesis and leukemogenesis & both drive immune escape. This prospective study involved 80 adult de novo AML patients recruited from Oncology Center, Mansoura University, between March 2019 and July 2021. The MEF2C and EVI1 expression were measured using a Taqman probe-based qPCR assay. The results revealed that EVI1 and MEF2C expression were significantly elevated in AML patients as compared to control subjects (p = 0.001. 0.007 respectively). Aberrant expressions of EVI1 and MEF2C showed a significant negative correlation with hemoglobin levels (p = 0.034, 0.025 respectively), & bone marrow blasts (p = 0.007, 0.002 respectively). 11q23 translocation was significantly associated with EVI1 and MEF2C (p = 0.004 and 0.02 respectively). Also, t (9;22) was significantly associated with EVI1 and MEF2C (p = 0.01 and 0.03 respectively), higher expression of EVI1 and MEF2C were significantly associated with inferior outcome after induction therapy (p = 0.001 and 0.018 respectively) and shorter overall survival (p = 0.001, 0.014 respectively). In conclusion, EVI1 & MEF2C were significantly expressed in AML cases. EVI1 & MEF2C overexpression were significantly associated with 11q23 rearrangements and t (9;22) and were indicators for poor outcome in adult AML patients; These results could be a step towards personalized therapy in those patients.

Mathematical modelling of clonal reduction therapeutic strategies in acute myeloid leukemia

Over the years, the overall survival of older patients diagnosed with acute myeloid leukemia (AML) has not significantly increased. Although standard cytotoxic therapies that rapidly eliminate dividing myeloblasts are used to induce remission, relapse can occur due to surviving therapy-resistant leukemic stem cells (LSCs). Hence, anti-LSC strategies have become a key target to cure AML. We have recently shown that previously approved cardiac glycosides and glucocorticoids target LSC-enriched CD34+ cells in the primary human AML 8227 model with more efficacy than normal hematopoietic stem cells (HSCs). To translate these in vitro findings into humans, we developed a mathematical model of stem cell dynamics that describes the stochastic evolution of LSCs in AML post-standard-of-care. To this, we integrated population pharmacokinetic-pharmacodynamic (PKPD) models to investigate the clonal reduction potential of several promising candidate drugs in comparison to cytarabine, which is commonly used in high doses for consolidation therapy in AML patients. Our results suggest that cardiac glycosides (proscillaridin A, digoxin and ouabain) and glucocorticoids (budesonide and mometasone) reduce the expansion of LSCs through a decrease in their viability. While our model predicts that effective doses of cardiac glycosides are potentially too toxic to use in patients, simulations show the possibility of mometasone to prevent relapse through the glucocorticoid’s ability to drastically reduce LSC population size. This work therefore highlights the prospect of these treatments for anti-LSC strategies and underlines the use of quantitative approaches to preclinical drug translation in AML.

Improved Treatment Outcomes With Modified Induction Therapy in Acute Myeloid Leukemia (AML): A Retrospective Observational Study From a Regional Cancer Center.

The aggressive, genetically diverse group of malignant illnesses known as acute myeloid leukemia (AML) is characterized by clonally related myeloblast invasion of the bone marrow, blood, and other organs. The treatment regimen plays a crucial role in the management of AML, and it is associated with poor overall survival and enhanced risk of relapse. Induction therapy with a 7+3 DA regimen (daunorubicin + ara-C) has been the treatment of choice for young and fit patients. To evaluate the effect of dose modification in young and fit patients for a modified treatment regimen. This was a retrospective, observational study of AML patients to analyze the outcomes of modified induction therapy in AML patients enrolled at Dr. B. Borooah Cancer Institute, Guwahati, Assam, India, from October 2021 to March 2022. The outcomes of modified induction therapy with intensive chemotherapy (modified 7+3 DA) and low-intensity chemotherapy decitabine (10 days) and venetoclax + azacytidine (seven days) were considered after the first two cycles or 60 days, whichever was earlier. Data from 31 patients with de-novo AML was analyzed; the median age of the patients was 41 years (range: 2-71 years), and the male-to-female ratio was 1.8. There were seven patients in the pediatric age group (2-13 years), and 19%, 65%, and 13% of patients belonged to favorable, intermediate, and high-risk groups, respectively. With regards to modified induction therapy (n=31), 20 (65%) patients received modified "7+3 DA", nine (29%) received hypomethylating agents (HMA, decitabine only), and two patients received HMA (azacitidnie) + venetoclax. Additionally, 23/31 patients completed at least two cycles of induction therapy. Overall, 60 day-induction mortality was 13%, and the complete remission (CR) and partial remission (PR) rates were 48% and 26%, respectively. In patients who received modified "7+3 DA", the CR rate was 55%. The notable reduction in deaths due to infections observed in our study suggests that centers with limited resources for preventing neutropenic complications during induction therapies in AML patients could consider adopting this modified regimen.

Targeting CREB-Binding Protein (CREBBP) Overcomes Resistance to Azacitidine and Venetoclax Therapy in Acute Myeloid Leukemia (AML)

Acute myeloid leukemia (AML) is a clonal myeloid malignancy arising from hematopoietic stem or progenitor cells (HSPC) within the bone marrow (BM). Despite advancements in our understanding of AML pathogenesis, patients continue to face grim outcomes, with a five-year relative survival rate of approximately 20%. This highlights the pressing need to identify molecular features of AML that are amenable to therapeutic intervention. The ideal molecular targets would be those broadly dysregulated and integral to multiple signaling pathways implicated in AML pathogenesis. Recent therapeutic regimens have incorporated venetoclax, a small molecule inhibitor of BCL2, in combination with hypomethylating agents, showing moderate success. Venetoclax (VEN) acts as a BH3 mimetic, specifically inhibiting BCL2. While clinical outcomes of VEN combined with hypomethylating agents (decitabine or azacitidine - AZA) have been promising, a substantial subset of patients remains refractory or experiences relapse following treatment with these regimens. Due to the molecular and cellular heterogeneity of AML, identifying ideal mechanisms to circumvent resistance remains elusive. In our investigations, we have observed that patient-derived cells from AML exhibit a higher frequency of dysregulated immune and inflammatory pathways than previously appreciated, referred to as “oncogenic immune signaling states.” Furthermore, immune and inflammatory-related pathways are preferentially expressed in AML patients who are resistant/refractory to VEN/AZA compared to those who respond favorably. To identify critical immune and inflammatory-related pathways contributing to VEN/AZA responses, we developed an oncogenic immune signaling-targeted CRISPR-library, aimed at identifying potential therapeutic targets involved in VEN/AZA resistance in AML. Utilizing this targeted CRISPR-screen approach, we successfully identified CREB-binding protein (CREBBP) as the top candidate linked to resistance to VEN/AZA therapy. Notably, CREBBP alterations have been associated with a poor prognosis in de novo AML, making it a compelling target for further investigation. To validate CREBBP's role in VEN/AZA resistance, we treated a VEN/AZA-resistant AML cell line, THP1 (FAB subtype M5, KMT2A-r), with commercially available pharmacologic inhibitors of CREBBP, A485, and CCS1477, and assessed the response. Remarkably, inhibition of CREBBP with either A485 or CCS1477 significantly decreased cell proliferative ability and clonogenic potential. Importantly, co-treatment with either A485 or CCS1477 and VEN/AZA synergistically abrogated cell proliferation and clonogenic potential of THP1 cells, effectively overcoming resistance (Figure 1). Our ongoing studies include validating CREBBP as a potential therapeutic target for overcoming VEN/AZA resistance in AML. We have created CREBBP-knockdown cell lines through genetic manipulation (shRNA) and will utilize these to confirm the specificity of CREBBP in modifying resistance to VEN/AZA. In addition we will utilize patient-derived xenograft studies in immunocompromised (NRG) mouse models. By investigating CREBBP's role in mediating resistance, we aim to unlock new avenues for enhancing the efficacy of VEN/AZA therapy in AML patients, particularly those facing resistance to current therapies. In conclusion, our research highlights the critical involvement of CREBBP in VEN/AZA resistance in AML. Targeting this molecular pathway holds the promise of offering a potential therapeutic approach to improve treatment outcomes for AML patients, particularly those facing resistance to current therapies. Understanding and harnessing CREBBP's influence could pave the way for developing personalized and effective strategies to combat this challenging hematological malignancy.

Downregulation of DNA methylation enhances differentiation of THP-1 cells and induces M1 polarization of differentiated macrophages

DNA methylation is an epigenetic modification that regulates gene expression and plays an essential role in hematopoiesis. UHRF1 and DNMT1 are both crucial for regulating genome-wide maintenance of DNA methylation. Specifically, it is well known that hypermethylation is crucial characteristic of acute myeloid leukemia (AML). However, the mechanism underlying how DNA methylation regulates the differentiation of AML cells, including THP-1 is not fully elucidated. In this study, we report that UHRF1 or DNMT1 depletion enhances the phorbol-12-myristate-13-acetate (PMA)-induced differentiation of THP-1 cells. Transcriptome analysis and genome-wide methylation array results showed that depleting UHRF1 or DNMT1 induced changes that made THP-1 cells highly sensitive to PMA. Furthermore, knockdown of UHRF1 or DNMT1 impeded solid tumor formation in xenograft mouse model. These findings suggest that UHRF1 and DNMT1 play a pivotal role in regulating differentiation and proliferation of THP-1 cells and targeting these proteins may improve the efficiency of differentiation therapy in AML patients.

Survival benefit of maintenance therapy in AML patients after complete remission in northeast Iran oncology centers with limited access to oral azacitidine.

123 Background: Acute myeloid leukemia (AML) is associated with high relapse rates even in patients who achieve a complete remission with initial treatment. Maintenance therapy with cytotoxic agents has been considered a strategy to prolong disease-free survival (DFS) and overall survival (OS). The lack of availability of oral Azacitidine in some countries limits its use. This study aimed to compare the DFS and OS rates of AML patients with and without maintenance therapy using subcutaneous cytarabine or oral thioguanine. Methods: This retrospective cohort study reviewed the charts of non-m3 AML patients treated in oncology centers in northeast Iran from February 2015 to February 2021. The study included patients aged 15-60 who experienced complete remission following induction and then consolidation chemotherapy and did not receive Allogeneic Hematopoietic Stem Cell Transplantation (allo-HSCT). Patients were categorized based on receiving maintenance therapy (MT) after the first-line treatment and those who were only followed up (FU). Patients’ demographic information, pathology and lab findings, date of relapse and/or death, and first-line and maintenance treatments regimens were extracted. For each of the MT and FU groups, the frequency of main outcomes (death and/or relapse) was calculated, and the Median (95%CI) of DFS and OS was estimated. The log-rank test was used to compare the DFS and OS between groups. Results: A total of 100 patients (50 MT and 50 FU) met the study inclusion criteria. There were no significant differences between the two groups regarding patients’ demographic characteristics, pathology and lab findings, and ELN’s risk category. The relapse rate in the FU and MT groups was 76% (n=38) and 60% (n=30) respectively (p=0.09). 39 individuals (78%) in the FU group and 26 individuals (54.2%) in the MT group died between complete remission and Feb 2022(p=0.01). The Median (95% CI) of DFS in FU and MT groups were 11 (8.6-13.3) and 18 (7.3 -28.6) months respectively (P=0.03). The median OS (95% CI) was 16 (10.8 - 21.1) months in the FU group and 23 (10.8 - 35) months in the MT group (P=0.04). Further comparisons between patients who received subcutaneous cytarabine and oral thioguanine revealed no significant differences regarding DFS and OS. Conclusions: The study results suggest that maintenance therapy with subcutaneous cytarabine or oral thioguanine after complete remission in non-m3 AML patients is associated with improved DFS and OS. Further randomized controlled trials and cost-utility studies are recommended to consider these available cytotoxic agents as maintenance therapy alternatives in regions where Azacitidine is not available.

Perturbed epigenetic transcriptional regulation in AML with IDH mutations causes increased susceptibility to NK cells

Isocitrate dehydrogenase (IDH) mutations are found in 20% of acute myeloid leukemia (AML) patients. However, only 30–40% of the patients respond to IDH inhibitors (IDHi). We aimed to identify a molecular vulnerability to tailor novel therapies for AML patients with IDH mutations. We characterized the transcriptional and epigenetic landscape with the IDH2i AG-221, using an IDH2 mutated AML cell line model and AML patient cohorts, and discovered a perturbed transcriptional regulatory network involving myeloid transcription factors that were partly restored after AG-221 treatment. In addition, hypermethylation of the HLA cluster caused a down-regulation of HLA class I genes, triggering an enhanced natural killer (NK) cell activation and an increased susceptibility to NK cell-mediated responses. Finally, analyses of DNA methylation data from IDHi-treated patients showed that non-responders still harbored hypermethylation in HLA class I genes. In conclusion, this study provides new insights suggesting that IDH mutated AML is particularly sensitive to NK cell-based personalized immunotherapy.

Amsacrine-based induction therapy in AML patients with cardiac comorbidities: a retrospective single-center analysis

Intensive chemotherapy is the backbone of induction treatment in patients with acute myeloid leukemia (AML). However, AML patients with concomitant cardiac disease may not be eligible for anthracycline-based therapies. In a small cohort of patients, we have previously shown that anthracycline-free, amsacrine-based chemotherapy TAA (thioguanine, cytarabine, amsacrine) may be as effective as cytarabine/daunorubicin for induction therapy in these patients. In this systematic retrospective single-center analysis, we documented the outcome of 31 patients with significant cardiac comorbidities including coronary heart disease or cardiomyopathy receiving TAA as induction chemotherapy. Median (range) ejection fraction (EF) was 48% (30–67%) in this cohort. Patients with EF below 30% were considered unfit for intensive induction therapy. Event-free survival (EFS), overall survival (OS), and relapse-free survival (RFS) were 1.61, 5.46, and 13.6 months respectively. Poor outcome was primarily related to a high early mortality rate within the first 30 days of therapy, mainly caused by infectious complications. TAA cannot be recommended as a substitute of standard induction for AML patients with significant concomitant cardiac disease. In the era of novel agents, alternative strategies (e.g., hypomethylating agents plus venetoclax) should be considered when anthracycline-based regimens are not suitable.

Leukemic stem cells and advances in hematopoietic stem cell transplantation for acute myeloid leukemia: a narrative review of clinical trials.

The purpose of this literature review is to summarize and provide a brief overview of our current understanding of acute myeloid leukemia (AML) and the role of stem cell transplantation (SCT) in its management. AML is a malignant hematological disorder that is characterized by the uncontrolled proliferation of myeloid blood cells. This disease has been associated with various risk factors such as ionizing radiation, cigarette smoke, pesticides/herbicides, and chemotherapy. SCT remains the most beneficial treatment for medically fit AML patients due to superior survival outcomes. A thorough search was conducted on PubMed, Scopus, ClinicalTrials.gov, Embase and Web of Science using related keywords. Current articles on the uses of stem cell therapy in AML patients were selected. Long term exposure to ionizing radiation and other harmful substances such as benzene, cigarette smoke and chemotherapeutic drugs plays an important role in AML carcinogenesis. Mutations in certain genes (e.g., ASXL1, RUNX1, KIT, TP53, BCR-ABL1) seem to accelerate the process as they affect normal cellular proliferation and cell death. These events may give rise to a small subpopulation of leukemic stem cells (LSC) which continuously sustain tumor development and growth. Patients who are deemed to be medically "fit" should receive an allogenic hematopoietic stem cell transplantation (allo-HSCT) due to improved overall survival (OS) (~50%) and decreased relapsed risk (32% vs. 59%). Several studies have revealed that the medically "unfit" may benefit from more conventional agents such as azacytidine, decitabine, venetoclax or sorafenib.

FLT3 inhibitors as maintenance therapy post allogeneic hematopoietic stem cell transplantation in acute myeloid leukemia patients with FLT3 mutations: A meta-analysis.

Acute myeloid leukemia (AML) patients with a Fms-like tyrosine kinase 3 (FLT3) mutation have a high incidence of relapse despite allogeneic hematopoietic stem cell transplantation (allo-HSCT) and a subsequent poor prognosis. FLT3 inhibitors (FLT3i) have been suggested to reduce the post-transplant relapse risk in recent studies. As more evidence is accumulated, we perform the present meta-analysis to assess the efficacy and safety of FLT3i as post-transplant maintenance therapy in AML patients. Literature search was performed in public databases from inception to December 31, 2021. Overall survival (OS), relapse-free survival (RFS), cumulative incidence of relapse (CIR), non-relapse mortality (NRM), graft-versus-host disease (GVHD) and adverse events were compared between FLT3i and control groups. Pooled hazard ratio (HR) or relative risk (RR) with corresponding 95% confidence interval (CI) were calculated. We identified 12 eligible studies with 2282 FLT3-mutated AML patients who had received HSCT. There was no between-study heterogeneity and a fix-effect model was used. Post-transplant FLT3i maintenance significantly prolonged OS (HR=0.41, 95%CI: 0.32-0.52, p < 0.001) and RFS (HR=0.39, 95%CI 0.31-0.50, p < 0.001), and reduced CIR (HR=0.31, 95%CI 0.20-0.46, p < 0.001) as compared with control. There were no significant risk differences in NRM (RR=0.69, 95%CI 0.41-1.17, p=0.169), acute GVHD (RR=1.17, 95%CI 0.93-1.47, p=0.175), chronic GVHD (RR=1.31, 95%CI 0.91-1.39, p=0.276) and grade ≥3 adverse events between both groups, except for skin toxicity (RR=5.86, 95%CI 1.34-25.57, p=0.019). Post-transplant FLT3i maintenance can improve survival and reduce relapse in FLT3-mutated AML patients and is tolerable.

Synergistic Anti-Tumor Effect on Novel Combination of Selinexor with Azacitidine in AML By Targeting XPO1/eIF4E/C-MYC Signaling

Background: Acute myeloid leukemia (AML) is a heterogeneous hematologic malignancy with high mortality and poor outcome. Highly effective therapy options are always needed for AML. Selinexor (KPT-330) is an oral bioactive selective CRM1 (XPO1) inhibitor, and the clinical trial showed the combination of KPT-330 over standard induction therapy benefits much for AML patients. Azacitidine (AZA), a DNA hypomethylation agent, can induce cell apoptosis and exhibit potential therapeutic efficacy against AML. However, it has not yet determined the anti-tumor effect of KPT-330 with AZA in AML. Here we examined the anti-leukemia effect and potential mechanism of KPT-330 combined with AZA in AML cells to provide evidence for further clinical trials. Methods: A total of 64 AML patients’ BM samples and 64 normal control from healthy volunteers were obtained from Zhongda Hospital Southeast University, in which the 2 patients’ samples were further treated with indicated drugs in vitro. Cell Counting Kit-8 (CCK-8) was used for cell viability assay, Annexin-V/PI staining following flow cytometry analysis for apoptosis, and qPCR and Western Blot for gene expression in AML cell lines and patient samples. CalcuSyn analysis was performed for a synergistic effect. RNA-seq was performed in U937 cells treated with 0.7uM AZA for 48 hrs, and the significantly differentially expressed genes (DEGs) were identified and gene set enrichment analysis (GSEA) was performed. Results: Cell proliferation arrest and apoptosis were significantly observed in U937 and MV4-11 cells treated with the combination of KPT-330 with AZA compared to single drug control (data not shown). Then, we further explored the synergistic effect of the combination in patients’ samples. Results showed that cell growth arrest and apoptosis significantly increased in primary samples from two de novo AML patients compared to the single drug controls, and CalcuSyn analysis showed a synergistic effect of the combination on cell growth arrest (Fig. 1A &1B). To understand the mechanisms underlying the synergy, RNA-seq analysis was performed in U937 cells treated by 0.7uM AZA and 4655 DEGs were identified. XPO1, eIF4E, and MYC are in the top DEGs upon AZA treatment, and GSEA analysis indicated MYC_TARGETS and apoptosis pathway are enriched in DEGs (Fig. 1C &1D ). Moreover, the mRNA level of XPO1 and eIF4E was increased in our AML patient cohort compared with healthy controls (Fig. 1E), and the high expression of XPO1 or elF4E are associated with poor outcome (Fig. 1F). XPO1 is the key component of nuclear-exporting machinery; elF4E, the cargo of XPO1 is translation initiation factor and plays a critical role in translational machinery, which regulates the C-MYC translation. It is also reported that AZA regulates the transcription of C-MYC by affecting methylation. We observed a combination of KPT-330 with AZA significantly down-regulated the mRNA and protein level of oncogene eIF4E and C-MYC in two of the patients’ samples compared to single drug controls (Fig. 1G &1H ). Also, C-MYC knock-down was not only significantly increased the apoptosis in U937 cells compared to scramble shRNA (shCTL) control, but also significantly enhanced the combination-mediated apoptosis compared to that of single drug control in U937 cells (Fig. 1I). these data indicated that XPO1/eIF4E/C-MYC signaling plays critical roles in the combination-mediated anti-leukemia, and the mechanistic model is summarized in Fig.1J. Conclusions: Our data for the first time showed the synergistic effect of a novel combination of KPT-330 with AZA on cell growth arrest and apoptosis in AML patients and also identified the underlying mechanism by targeting XPO1/eIF4E/C-MYC signaling. Our results also highlighted the feasibility of the clinical trial for the combination in the therapy of AML patients. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Trial in Progress: Tagraxofusp-Based Therapy to Eradicate Measurable Residual Disease of Acute Myelogenous Leukemia Prior to Allogeneic Hematopoietic Cell Transplantation

Background Numerous studies convincingly demonstrate that measurable residual disease (MRD) in AML at time of allogeneic cell transplantation (alloHCT) is a powerful, independent predictor of subsequent relapse. Attempts to eradicate AML MRD pre-transplant with consolidation chemotherapy have been unsuccessful, with reported MRD conversion rates as low as 20% (Appelbaum et al., Blood 2019). Tagraxofusp is an IL-3 truncated diphtheria toxin recombinant protein that binds CD123 with high affinity. Preliminary data using tagraxofusp as consolidation therapy in AML patients has demonstrated good tolerability (Lane et al., Blood 2017). Pathways of resistance to tagraxofusp have been identified preclinically as loss of the intracellular target for diphtheria toxin, diphthamide, through methylation of an enzyme involved in diphthamide synthesis. Treatment with a hypomethylating agent has the potential to restore expression of diphthamide and increase tagraxofusp sensitivity in AML cells (Togami et al., J Clin Invest 2019). Design This is a single-arm, open-label, multi-center phase Ib/II clinical trial to evaluate the safety and effectiveness of tagraxofusp +/- azacitidine in AML patients who are in first or second remission with detectable MRD and are planned to undergo alloHCT. MRD will be measured by multi-dimensional flow cytometry with sensitivity of 0.02% (Hematologics). Patients will be enrolled in a modified 3+3 dose escalation design with two cohorts: cohort A) tagraxofusp 12 mcg/kg IV qday on days 1-5; and cohort B) azacitidine 75 mg/m2 subq qday on days 1-5 plus tagraxofusp 12 mcg/kg qday on days 8-12. Patients achieving MRD-negativity after cycle 1 will proceed to alloHCT. If a patient remains MRD-positive, an additional cycle of investigational therapy will be administered. All patients will proceed to alloHCT after no more than 2 cycles of investigational therapy. Both cohorts contain a dose escalation and a dose expansion phase. Cohort B will open if cohort A is not efficacious. Up to 44 participants will be enrolled in this trial at University of California Los Angeles and University of California San Francisco (two constituent sites of the University of California Hematologic Malignancies Consortium). Objectives The primary objective is to estimate the rate of conversion from MRD-positive pre-investigational therapy to MRD-negative after 1 or 2 cycles of investigational therapy. Key secondary objectives include to determine the safety of tagraxofusp +/- azacitidine in this patient population and to evaluate if adverse effects of this investigational therapy lead to delays in alloHCT. Exploratory objectives will involve use of single cell RNA sequencing to investigate if response to investigational therapy is predicted by leukemia-specific biomarkers and to evaluate if putative cell surface markers (inferred from RNA detection) are associated with MRD persistence. Main Outcomes The primary endpoint is MRD status after 1 or 2 cycles of investigational therapy. Key secondary endpoints include dose-limiting toxicities (any grade 4 or 5 non-hematologic toxicity or any grade 3 or greater hematologic toxicity at day 42) and number of days between investigational therapy and alloHCT. Conclusions There is an unmet need to improve outcomes in AML with detectable MRD pre-transplant. The strategy in this study is to employ tagraxofusp as an alternative mechanism of action to eradicate chemo-resistant AML cells responsible for subsequent relapse. In order to mitigate any potential resistance the residual leukemia population may have against tagraxofusp, azacitidine will be added to optimize the likelihood for MRD negativity and the possibility for improved outcomes after alloHCT. This study was sponsored by the ASH CRTI. References Appelbaum J, Loeb A, Gardner K, et al. Additional Cytotoxic Chemotherapy Is Unlikely to Eliminate Measurable Residual Acute Myeloid Leukemia (AML). Blood. 2019;134(Supplement_1):260-260. Lane AA, Sweet KL, Wang ES, et al. Results from Ongoing Phase 1/2 Trial of SL-401 As Consolidation Therapy in Patients with Acute Myeloid Leukemia (AML) in Remission with Minimal Residual Disease (MRD). Blood. 2017;130(Supplement 1):2583-2583. Togami K, Pastika T, Stephansky J, et al. DNA methyltransferase inhibition overcomes diphthamide pathway deficiencies underlying CD123-targeted treatment resistance. J Clin Invest. 2019;129(11):5005-5019. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

First-in-human study of WT1 recombinant protein vaccination in elderly patients with AML in remission: a single-center experience

Wilms’ tumor 1 (WT1) protein is highly immunogenic and overexpressed in acute myeloid leukemia (AML), consequently ranked as a promising target for novel immunotherapeutic strategies. Here we report our experience of a phase I/II clinical trial (NCT01051063) of a vaccination strategy based on WT1 recombinant protein (WT1-A10) together with vaccine adjuvant AS01B in five elderly AML patients (median age 69 years, range 63–75) receiving a total of 62 vaccinations (median 18, range 3–20) after standard chemotherapy. Clinical benefit was observed in three patients: one patient achieved measurable residual disease clearance during WT1 vaccination therapy, another patient maintained long-term molecular remission over 59 months after the first vaccination cycle. Interestingly, in one case, we observed a complete clonal switch at AML relapse with loss of WT1 expression, proposing suppression of the original AML clone by WT1-based vaccination therapy. Detected humoral and cellular CD4+ T cell immune responses point to efficient immune stimulation post-vaccination, complementing hints for induced conventional T cell infiltration into the bone marrow and a shift from senescent/exhausted to a more activated T cell profile. Overall, the vaccinations with WT1 recombinant protein had an acceptable safety profile and were thus well tolerated.To conclude, our data provide evidence of potential clinical efficacy of WT1 protein-based vaccination therapy in AML patients, warranting further investigations.

Expression Changes of CX3CL1 and Interleukin-6 Genes During Remission Induction Therapy in Patients with Acute Myeloid Leukemia

Background: Acute myeloid leukemia (AML) syndrome is a hematologic malignancy due to the extensive clonal proliferation of leukemic precursor cells and is rapidly fatal unless treated or in response to chemotherapy. Cytogenetic findings have an important role in the prognosis and categorization of AML. This study aimed to investigate the expression changes in CX3CL1 and Interleukin-6 (IL-6) genes before and after chemotherapy as remission induction therapy in AML patients. Materials and Methods: In this study, 69 patients (36 males, 33 female) with AML were selected from tertiary medical health centers. A quantitative polymerase chain reaction was performed for mRNA expression of CX3CL1 and IL-6 genes before and after induction chemotherapy using the 2-ΔΔCT method. Results: The expression of CX3CL1 and IL-6 were significantly increased after induction chemotherapy. Also, the ΔCt mean of CX3CL1 and IL-6 mRNA was not significant between AML subtype groups. Conclusion: We showed that chemotherapy significantly increases the expression of CX3CL1 and IL-6, which can be used as a prognostic factor of AML. [GMJ.2021;10:e2288]

Intensified Cytarabine Dose during Consolidation Therapy in AML Patients Under 65 Years Is Not Associated with Survival Benefit

▪Background: Acute myeloid leukemia (AML) is characterized by a high relapse rate, indicating insufficient clearance of leukemia-initiating cells. Depending on genetic risk stratification, consolidating chemotherapy proves to significantly reduce the risk of relapse. In particular, in younger AML patients higher dosage of cytarabine appears to improve long-term outcome, while there is no apparent benefit of multiagent combination, compared to cytarabine monotherapy. However, to this end the optimal dosage of single agent cytarabine in consolidation therapy after 7+3 remission induction remains elusive.Methods: Here, we retrospectively assessed the impact of different dosages of cytarabine consolidation on outcome in a large real-world data set from the German Study Alliance Leukemia-Acute Myeloid Leukemia (SAL-AML) registry. Patients below 65 years of age, registered between April 2005 and September 2020 with non-acute promyelocytic leukemia, who attained complete remission after intensive induction and received at least one consolidation cycle with intermediate (IDAC) or high dose cytarabine (HiDAC) were selected. To account for differences in patient and disease characteristics between both groups, the average treatment effect was estimated by propensity score weighting.Results: 642 patients received HiDAC consolidation with a median dosage of 5794.88 (IQR, 4745.48-5971.56) mg/m 2/d with a median number of 3 cycles (IQR, 2-3), whereas 178 patients received IDAC consolidation with 1946.16 (IQR, 1869.51-2469.15) mg/m 2/d with a median of 2 cycles (IQR, 1-3). IDAC-treated patients showed in average a higher age (median (IQR) 58.5 (49-62) years vs. 50 (41-56) years) and more comorbidities with 43.8% having an HCT-CI score of 2-4, compared to 22.3% among HiDAC-treated patients. Alongside, significantly more secondary (5.1% vs. 3.1%) and therapy-related (12.4% vs. 4.1%) AML as well as more adverse (14.5% vs. 6.5%) and less favorable (40.6% vs. 56%) genetic risk features according to ELN 2017 risk classification were found among IDAC-treated patients. After propensity score weighting for differences in patient and disease characteristics, overall survival after 5 years was comparable between HiDAC-treated (71.1 %) and IDAC-treated (67.7%) patients. Moreover, no significant differences in relapse-free survival were observed after 5 years (47.4 vs. 45.2%). Notably, more patients treated with IDAC received allogeneic stem cell transplantation in first remission (37.6 vs. 19.8%) while significantly more HiDAC-treated patients underwent allogeneic stem cell transplantation in relapse (30.8 vs. 20.2%). Censoring for allogeneic stem cell transplantation in first remission revealed no significant survival difference with regard to cytarabine dosage. Considering only ELN favorable risk AML patients, there was no difference in 5-years overall (80.5% vs. 83.9%) nor relapse-free (57.7% vs. 56.8%) survival. Of note, significantly more patients treated with HiDAC suffered from ≥3 CTCAE infectious complications (56.7 vs. 44.1%), which was more striking in patients above 50 years of age. The rate of other ≥3 CTCAE non-hematological toxicities and secondary malignancies was comparable in both treatment groups.Conclusion: This retrospective analysis suggests no significant benefit of high dose cytarabine compared to intermediate dosages in consolidation for AML patients under 65 years of age, independent of ELN risk group. DisclosuresKrause: Siemens: Research Funding; Takeda: Honoraria; Pfizer: Honoraria; art-tempi: Honoraria; Kosmas: Honoraria; Gilead: Other: travel support; Abbvie: Other: travel support. Schliemann: Philogen S.p.A.: Consultancy, Honoraria, Research Funding; Abbvie: Consultancy, Other: travel grants; Astellas: Consultancy; AstraZeneca: Consultancy; Boehringer-Ingelheim: Research Funding; BMS: Consultancy, Other: travel grants; Jazz Pharmaceuticals: Consultancy, Research Funding; Novartis: Consultancy; Roche: Consultancy; Pfizer: Consultancy. Haenel: Jazz: Consultancy, Honoraria; GSK: Consultancy; Bayer Vital: Honoraria; Takeda: Consultancy, Honoraria; Roche: Consultancy, Honoraria; Novartis: Consultancy, Honoraria; Amgen: Consultancy; Celgene: Consultancy, Honoraria. Brummendorf: Takepart Media: Honoraria; Repeat Diagnostics: Research Funding; Pfizer: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Novartis: Honoraria, Patents & Royalties, Research Funding; Janssen: Honoraria; Bristol Myers: Research Funding. Fransecky: Abbvie: Honoraria, Research Funding; Medac: Honoraria; Novartis: Honoraria; Amgen: Honoraria; Takeda: Honoraria. Einsele: Janssen, Celgene/BMS, Amgen, GSK, Sanofi: Consultancy, Honoraria, Research Funding. Held: MSD: Honoraria, Membership on an entity's Board of Directors or advisory committees; Amgen: Research Funding; Roche: Research Funding; Acortech Biopharma: Research Funding; BMS: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Roche: Honoraria, Membership on an entity's Board of Directors or advisory committees. Platzbecker: Janssen: Honoraria; Celgene/BMS: Honoraria; AbbVie: Honoraria; Geron: Honoraria; Takeda: Honoraria; Novartis: Honoraria. Baldus: Amgen: Honoraria; Celgene/BMS: Honoraria; Novartis: Honoraria; Jazz: Honoraria. Mueller-Tidow: Janssen Cilag: Consultancy, Research Funding; Pfizer: Consultancy, Research Funding; Bioline: Consultancy, Research Funding.

In Patients with Acute Myeloid Leukemia, Hidac Administered on Days 1, 2 and 3 of Consolidation Course Is As Efficacious As Hidac Given on Days 1, 3 and 5, but Is Better Tolerated

Introduction: High-dose Ara-C (HiDAC) is a post-remission consolidation protocol commonly used in patients with acute myeloid leukemia (AML). The optimal regimen of HiDAC administration is not fully established. Recent studies suggest that a short and condensed schedule of HiDAC consolidation therapy given on days 1, 2 and 3 (HiDAC123) may be superior to the traditional days 1, 3 and 5 regimen (HiDAC135). The current retrospective study compared the impact of these two therapeutic approaches on the outcomes of AML patients treated at the Rambam Leukemia Unit.Methods: This retrospective cohort analysis included patients diagnosed with AML between the years 2015-2020 who were candidates for aggressive chemotherapy. Fifty-seven patients received a total of 76 courses of HiDAC135 mainly between 2015 and 2017, while 77 other patients received a total of 135 courses of HiDAC123 mainly between 2017 and 2020. The HiDAC dose in the two groups was 3 g/m 2 for patients aged less than 55 years, and 1.5 g/m 2 for patients aged 55-65 years. Patient demographics, the ELN leukemia risk category and NPM1 status were compared. The analysis also included data on complications per course of HiDAC therapy, need for hospitalization, the number of days of hospitalization if needed, the occurrence of neutropenic fever per course, the need for blood products (packed red blood cells and platelets), neurotoxicity and death within the 28 days of the course.Results: The HiDAC123 and HiDAC135 groups were comparable in terms of the ELN risk category and NPM1 status. The groups significantly differed in age (median of 49.8 years and 55.9 years, respectively; p=0.01), which was most likely to be related to the use of venetoclax and azacitidine during the period when HiDAC123 was prescribed. Similar percentage of patients achieved complete remission (CR) after induction chemotherapy (93.5% and 94.7%, in the HiDAC123 and HiDAC135 groups, respectively). There were no differences in the overall survival (OS) and event-free survival (EFS) between the two groups (Figure 1). Significantly more patients in the HiDAC135 group were hospitalized for more than 4 days compared with the HiDAC123 group (51.3% vs 31.9%; p=0.008) and significantly more patients in the HiDAC135 group needed more than 2 units of packed red blood cells than in the HiDAC123 group (38.2% vs 21.5%, p=0.011). There was no difference in the percentage of patients in need of platelet transfusion. Likewise, there was no difference between the two groups in either the number of hospitalizations post-HiDAC therapy, the occurrence of neutropenic fever events or the incidence of death within the first 28 days after the HiDAC therapy.Conclusions: The present analysis demonstrates that HiDAC123 is as safe and efficacious as HiDAC135 in terms of OS and EFS and is associated with shorter hospitalization and lower requirement for blood products. These results show that HiDAC123 could be the regimen of choice as consolidation therapy in AML patients. [Display omitted] DisclosuresZuckerman: Orgenesis Inc.: Honoraria; BioSight Ltd: Honoraria; AbbVie: Honoraria; Janssen: Honoraria; Novartis: Honoraria; Gilead Sciences: Honoraria, Speakers Bureau; Cellect Biotechnology: Honoraria. Ofran: Pfizer: Consultancy; AbbVie: Consultancy; Medison Israel: Consultancy; Astellas: Consultancy; Janssen: Consultancy.

In Vivo crispr Screening Identifies GLUT1 As a Key Metabolic Regulator of MLL-AF9-Driven Acute Myeloid Leukemia

Disease relapse in patients with acute myeloid leukemia (AML) is associated with a failure of current treatments to eradicate leukemia stem cells (LSCs), a self-renewing population of cells responsible for disease progression and maintenance. Thus, novel therapeutic strategies designed to specifically target LSCs while sparing normal hematopoietic stem cells are needed. To identify dependencies in LSCs that may reveal new treatment opportunities, we performed an in vivo CRISPR/Cas9 dropout screen in the widely used MLL-AF9-driven AML murine model. The pooled lentiviral CRISPR library was designed to target 960 genes encoding cell surface proteins expressed on MLL-AF9 AML cells as these are accessible for therapeutic targeting.The facilitated glucose transporter member 1(GLUT1), a major mediator of cellular glucose uptake, emerged as the highest ranked dependency in the screen, with all 6 sgRNAs depleted more than 10-fold in vivo. Consistent with the results from the screen, validation experiments confirmed that sgRNA-mediated GLUT1 disruption in c-Kit +Cas9 +dsRed +MLL-AF9 cells led to a 5-fold reduction in the establishment of leukemia in both the bone marrow and spleen of recipient mice. In line with these in vivo observations, leukemia cells expressing GLUT1 sgRNAs were rapidly depleted over time in an ex vivo competition assay (p<0.0001). GLUT1 disruption also led to a marked increase in mean survival from 28 to 73 days in mice transplanted with sorted GLUT1 sgRNA-expressing leukemia cells relative to controls. Notably, while GLUT1 loss did not affect apoptosis or cell-cycle state, it led to a more than two-fold increase in the surface expression of the myeloid differentiation marker Gr-1 (p=0.0002). Interestingly, knockdown of GLUT1 lead to reduced mRNA expression levels of key downstream genes of MLL-driven leukemia Meis1 (p<0.0001) and Hoxa9 (p=0.0013) , both of which are commonly downregulated upon differentiation. These findings suggest that GLUT1 ablation arrests AML cell growth at least in part via accelerated differentiation and attenuated cell proliferation.Given GLUT1-mediated glucose transfer constitutes the first rate-limiting step for glucose metabolism, we assessed the metabolic profile of MLL-AF9 AML cells following loss of GLUT1. Bioenergetic profiling revealed that the rate of glycolysis was significantly decreased upon GLUT1 knockdown, as measured by a decrease in extracellular acidification rate (ECAR), glucose uptake, hexokinase activity and extracellular lactate production.To further assess the feasibility of GLUT1 inhibition as a therapy for AML patients, we treated murine cKit +MLL-AF9 leukemia cells with BAY-876, a potent and highly selective GLUT1 inhibitor. BAY-876 impaired tumor growth following 24hr (IC 50 60.3 nM) and 48hr (IC 50 68.8 nM) treatment ex vivo in a dose-dependent manner. Interestingly, the inhibitory effect on the counterpart healthy bone marrow c-Kit + cells was significantly weaker (24hr IC 50 347.7 nM; 48hr IC 50 258.4nM), indicating selective targeting of LSCs. To test the efficacy of BAY-876 as an anti-leukemic agent in vivo, sublethally irradiated mice were transplanted with c-Kit +MLL-AF9 AML cells and 3 days post-injection, were randomised into two groups (Veh n=4; BAY-876 n=6) and orally treated with either vehicle or 4mg/kg of BAY-876 daily. Following 10 days of treatment, mice were sacrificed and leukemia burden was assessed. Notably, substantially lower levels of leukemia cells in the bone marrow (p=0.0095), spleen (p=0.0095), and peripheral blood (p=0.036) were observed in the BAY-876 treatment group with no significant loss of body weight. Consistent with these findings, the average spleen weight was reduced by 66% upon BAY-876 treatment (p=0.0136).Collectively, we demonstrate that MLL-AF9-driven AML cells are dependent on GLUT1 for continued growth and survival. Targeting of GLUT1 downregulates glycolysis and induces cellular differentiation. We report that genetic or pharmacological inhibition of GLUT1 is sufficient to impair leukemic growth in vitro and in vivo, highlighting a potential therapeutic opportunity for disarming intrinsic metabolic dependencies of LSCs. Ongoing studies are aimed at translating these findings to the human disease and exploring combinatorial therapies that may act synergistically to overcome mechanisms of therapy resistance and metabolic plasticity. DisclosuresNo relevant conflicts of interest to declare.

A Phase II Study of 5-Azacytidine (AZA) and Venetoclax As Maintenance Therapy in Patients with Acute Myeloid Leukemia (AML) in Remission

A Phase II Study of 5-Azacytidine (AZA) and Venetoclax As Maintenance Therapy in Patients with Acute Myeloid Leukemia (AML) in Remission

AML-294: Maintenance Therapy After Allogeneic Hematopoietic Transplant for Acute Myeloid Leukemia: A Systematic Review and Meta-Analysis

Context: Allogeneic hematopoietic stem cell transplant (HSCT) is a standard therapy for patients with acute myeloid leukemia (AML), when the risk for relapse overweighs projected transplant-related morbidity and mortality. Disease relapse remains the most common reason for transplant failure and patient death, and treatment of relapse remains extremely challenging. Reducing the risk of post-transplant relapse without unwanted toxicity by means of post-transplant maintenance therapy has recently been studied in several prospective studies. Objectives: We aimed to evaluate the efficacy and safety of maintenance therapy for AML after allogeneic HSCT. Design, Setting and Patients: Systematic review and meta-analysis of randomized controlled trials which compared maintenance therapy with observation or placebo in AML patients after allogeneic HSCT. The Cochrane Library, PubMed, conference proceedings, and references were searched until February 2021. Primary outcome was overall survival (OS). Secondary outcomes included relapse free survival (RFS), relapse rate, and safety (including adverse events and GVHD). Results: Our search yielded five trials conducted between the years 2009 and 2018, including 736 patients, all published in peer-reviewed journals. Maintenance therapy comprised of tyrosine kinase inhibitors (TKI) – i.e., sorafenib - 2 studies and midostaurin - 1 study as well as hypomethylating agents (HMAs), namely decitabine and azacytidine - one study each. Maintenance after allogeneic HSCT was associated with an improved OS, HR=0.61 (95% CI 0.47-0.80). Subgroup analyses by type of maintenance therapy revealed advantage in OS with either TKI or HMA maintenance [HR=0.50 (95% CI 0.33-0.77) and HR=0.69 (95% CI 0.49-0.98), respectively]. Data from five trials was available for RFS analysis and showed improved RFS in the maintenance group compared to the control arm HR=0.51 [95% CI 0.40 - 0.66]. Relapse rate was significantly decreased in the maintenance arm compared to the control arm, RR=0.41 (95% CI 0.20-0.88, 4 trials). Regarding safety, there was no difference between the arms concerning adverse events (AE), including grade 3-4 AE, rate of infections and acute and chronic GVHD. Conclusions: Our meta-analysis shows that post-transplant maintenance therapy in AML patients is effective in decreasing relapse rate and improving RFS and OS, with a satisfactory safety profile.

Maintenance therapy after allogeneic hematopoietic transplant for acute myeloid leukemia: a systematic review and meta-analysis

Background For patients undergoing allogeneic hematopoietic stem cell transplant (HSCT) for acute myeloid leukemia (AML), disease relapse remains the most common reason for transplant failure and patient death. Recent randomized controlled trials (RCTs) have aimed to reduce the risk of relapse by means of post-transplant maintenance therapy. Methods We performed a systematic review and meta-analysis of RCTs comparing the efficacy and safety of maintenance with observation or placebo in patients with AML after allogeneic HSCT. We searched Cochrane Library, PubMed and conference proceedings up to Febuary 2021. Results Our search yielded five trials including 736 patients. Maintenance therapy consisted of tyrosine kinase inhibitors (TKIs) in 3 studies (sorafenib 2 studies; midostaurin 1 study) and hypomethylating agents (HMAs) in 2 studies (decitabine and azacytidine 1 study each). Maintenance therapy was associated with an improved overall survival (OS), HR = 0.61 (95% CI 0.47–0.80). Subgroup analysis revealed advantage in OS with either TKI or HMA maintenance. Relapse free survival (RFS) was also improved in the maintenance arm compared with the control arm HR = 0.51(95% CI 0.40 − 0.66). There was no difference between the two arms in overall grade 3/4 adverse events or overall infections, in grade 3/4 infections, or in acute and chronic graft versus host disease. Conclusions Our meta-analysis shows that post-transplant maintenance therapy in AML patients is effective in improving RFS and OS, with a satisfactory safety profile.