Standard Induction Chemotherapy Research Articles

Survival outcomes in patients with high-risk neuroblastoma (HRNB) in remission after relapsed or refractory treatment receiving eflornithine (DFMO) maintenance.

10060 Background: Five-year overall survival in newly diagnosed HRNB patients is around 50%, with relapse as the primary cause of mortality. While survival for newly diagnosed HRNB has improved with new treatment advances, relapsed or refractory (R/R) patients continue to have poor outcomes, with a high risk of subsequent relapse even among those who achieve remission, underscoring the need for treatments to improve EFS in this group. DFMO has been evaluated as a chemopreventative therapy in a single arm phase 2 study designed to compare Event Free Survival (EFS) and Overall Survival (OS) outcomes with published data for HRNB. The results of patients treated after standard therapy have been previously published. We now present an analysis of EFS/OS for patients with R/R disease, who historically have had a two-year EFS of approximately 10%. Methods: A total of 140 HRNB patients were enrolled, prospectively divided into two strata. S1 included patients in initial remission after upfront therapy, whereas S2 included patients who had either relapsed or had disease refractory to standard induction chemotherapy but achieved remission with additional therapy. Patients received 2 years of continuous treatment with oral DFMO 750 ±250 mg/m2 BID and were followed for up to 7 years. Results: A total of 35 patients withR/R disease who had achieved remission after additional therapy were enrolled onto S2, including 28 patients in remission after one or more prior relapses and 7 refractory patients in remission after subsequent therapies. Of the relapsed patients, 23 had one prior relapse, while 5 patients had two or more relapses prior to enrollment; only one previously relapsed patient had received chemoimmunotherapy prior to enrollment in the study. For the entireS2 cohort, two-year EFS was 46% (95% CI 29%, 61%) with the lower confidence interval above the 10% historical control estimate. Four-year EFS was 46% (95% CI 29%, 61%) and OS 62% (95% CI 44%, 76%), with no relapses occurring after 18 months. For patients enrolled with refractory disease only (n=7) the four-year EFS was 85.7%. In the 28 relapsed patients, the four-year EFS was 35.7%, with single relapse patients (n=23) having an EFS rate of 39.1% and multiple relapsed patients (n=5) 20.0%. Conclusions: Patients in remission after chemotherapy for relapsed or refractory HRNB and treated with DFMO had improved EFS compared to the best historical estimate available at the time of the study. Although the number of patients is too small to support clear conclusions, the subgroup of refractory patients had outcomes that trended better than the relapsed subgroup with an EFS rate that was unexpectedly similar to prior reports of DFMO-treated S1 patients in remission after standard upfront therapy. Clinical trial information: NCT02395666 , NCT00026312 .

Tislelizumab versus placebo combined with induction chemotherapy followed by concurrent chemoradiotherapy and adjuvant tislelizumab or placebo for locoregionally advanced nasopharyngeal carcinoma: Interim analysis of a multicenter, randomized, placebo-controlled, double-blind, phase 3 trial.

6001 Background: Patients with locoregionally advanced nasopharyngeal carcinoma (LANPC) have a risk of disease relapse after induction chemotherapy (IC)-concurrent chemoradiotherapy (CCRT). Early phase trials indicated that tislelizumab (PD-1 inhibitor) plus IC followed by CCRT have the potential to deepen responses and extend survival in LANPC. Here, we report the interim results of a phase 3 trial to evaluate the efficacy and safety of tislelizumab plus IC, followed by CCRT and adjuvant tislelizumab therapy in LANPC. Methods: Patients with high-risk LANPC (stage III-IVa, AJCC 8th Staging System, excluding T3N0, and T3N1 with retropharyngeal lymph nodes +) were randomized (1:1) to receive 200 mg tislelizumab or placebo + gemcitabine and cisplatin (GP) every 3 weeks (Q3W) for 3 cycles, followed by CCRT and adjuvant tislelizumab or placebo Q3W for up to an additional 8 cycles. Stratification factors were center and disease stage (III or IVa). Dual primary endpoints were complete response rate (CRR) after induction therapy and progression-free survival (PFS) per RECIST 1.1 by investigator under blinded review. The planned interim analysis (IA) for evaluating the first primary endpoint, CRR, was scheduled at 4 weeks after the completion of induction therapy to test whether addition of tislelizumab to standard IC significantly improved CRR, with allocated alpha at 0.005. Results: Between June 2022, and May 2023, 450 patients were randomized to tislelizumab arm (n = 223) and placebo arm (n = 227). Baseline characteristics were balanced between the two arms. At IA, 93.7% and 93.8% of patients completed 3 cycles of induction therapy in tislelizumab arm and placebo arm, respectively. Significant improvement in CRR was observed in tislelizumab arm vs placebo arm in the intent-to-treat population (ITT) (30.5% vs 16.7%; P = 0.0006). Improvement in CRR in tislelizumab arm vs placebo arm was consistent across key subgroups including disease stage (III [33.8% vs 20.7%]; IVa [28.7% vs 14.5%]), sex (male [28.3 % vs 15.7%]; female [38.0% vs 19.7%]), and ECOG PS (PS of 0 [28.8% vs 16.2%]; PS of 1 [40.6% vs 19.4%]). The ORR was 93.3% in the tislelizumab arm and 90.7% in the placebo arm. In safety population (tislelizumab arm, n=219; placebo arm, n=224), the incidence of Grade ≥3 TEAEs (40.6% vs 39.3%) and SAEs (2.3% vs 1.3%) were similar between two arms. Conclusions: The trial met its first primary endpoint with a statistically significant improvement of CRR with the addition of tislelizumab to standard IC in high-risk LANPC. Induction therapy with tislelizumab plus GP was generally well tolerated with manageable safety profile. Continued follow-up is being conducted to assess long-term efficacy and safety. Clinical trial information: NCT05211232 .

Imatinib with intensive chemotherapy in AML with t(9;22)(q34.1;q11.2)/BCR::ABL1. A DATAML registry study

Acute myeloid leukemia (AML) with t(9;22) (q34.1; q11.2)/BCR::ABL1, a distinct entity within the group of AML with defining genetic abnormalities, belong to the adverse-risk group of the 2022 ELN classification. However, there is little data on outcome since the era of tyrosine kinase inhibitors. Among 5819 AML cases included in the DATAML registry, 20 patients with de novo BCR::ABL1+AML (0.3%) were identified. Eighteen patients treated with standard induction chemotherapy were analyzed in this study. Imatinib was added to chemotherapy in 16 patients. The female-to-male ratio was 1.25 and median age was 54 years. The t(9;22) translocation was the sole chromosomal abnormality in 12 patients. Main gene mutations detected by NGS were ASXL1, RUNX1 and NPM1. Compared with patients with myeloid blast phase of chronic myeloid leukemia (CML-BP), de novo BCR::ABL1+AML had higher WBC, fewer additional chromosomal abnormalities, lower CD36 or CD7 expression and no ABL1 mutations. Seventeen patients (94.4%) achieved complete remission (CR) or CR with incomplete hematologic recovery. Twelve patients were allografted in first remission. With a median follow-up of 6.3 years, the median OS was not reached and 2-year OS was 77% (95% CI: 50–91). Four out of five patients who were not transplanted did not relapse. Comparison of BCR::ABL1+AML, CML-BP, 2017 ELN intermediate (n = 643) and adverse-risk patients (n = 863) showed that patients with BCR::ABL1+AML had a significant better outcome than intermediate and adverse-risk patients. BCR::ABL1+AML patients treated with imatinib and intensive chemotherapy should not be included in the adverse-risk group of current AML classifications.

Pracinostat combined with azacitidine in newly diagnosed adult acute myeloid leukemia (AML) patients unfit for standard induction chemotherapy: PRIMULA phase III study

Non-intensive therapies such as the hypomethylating agent (HMA) azacitidine (AZA) have been used in patients with AML ineligible for intensive induction chemotherapy (IC) or stem cell transplant due to advanced age, comorbidities, and/or risk factors. However, response rates and survival remain dismal. Pre-clinical studies indicate the epigenetic combination of HMAs and HDAC inhibitors induce re-expression of silenced genes synergistically. The activity of pracinostat, an oral pan-HDAC inhibitor, has been shown in xenograft tumor models of AML and promising efficacy was seen in a Phase 2 study. This Phase 3 study (NCT03151408) evaluated the efficacy/safety of pracinostat administered with AZA in adult patients with newly diagnosed AML ineligible to receive IC. Patients were randomized to either pracinostat plus AZA or placebo/AZA and stratified by cytogenetic risk and ECOG status. As planned, an interim analysis was performed when 232/390 events (deaths) occurred. A total of 406 patients were randomized (203/group) at the time of the analysis. Median overall survival was 9.95 months for both treatment groups (p=0.8275). There was no significant difference between treatments in secondary efficacy endpoints, reflecting a lack of clinical response. This study did not show a benefit of adding pracinostat to AZA in elderly patients unfit for IC.

Assessment of Measurable Residual Disease on Day 28 of First Induction Chemotherapy in Acute Myeloid Leukaemia Patients

Background: Measurable residual disease (MRD) is an independent, strong, prognostic marker in acute myeloid leukaemia (AML) that is important for risk stratification and treatment planning. This study assessed the MRD on day-28 after first induction chemotherapy in AML patients. Objective: To assess measurable residual disease status on day 28 after the first induction chemotherapy in newly diagnosed acute myeloid leukaemia patients. Methodology: This cross-sectional study was conducted at the Department of Haematology, Bangabandhu Sheikh Mujib Medical University (BSMMU), Dhaka from April 2021 to March 2022 among a total of 25 newly diagnosed AML patients who were admitted and received induction chemotherapy with DA (7+3). On Day-28, bone marrow aspiration was done with standard procedure and first pull of 2ml bone marrow sample was sent to a laboratory for MRD evaluation by multiparameter flow cytometry. MRD was considered positive when value was ?0.1%. Results: On day 28 of post induction chemotherapy most of the patients (92%) achieved morphological remission and among them more than half (52.2%) were MRD positive. The presence of CD14 myeloid marker and baseline higher serum uric acid level was significant in MRD positive patients and cytarabine dose 150 mg/m2 was found significant in MRD negative patients. Conclusion: More than half of the Acute Myeloid Leukaemia patients, treated with standard induction chemotherapy and achieved morphological remission on day 28 of chemotherapy, had positive measurable residual disease assessed by multiparameter flow cytometry. Positive MRD indicates relatively poorer prognosis and may necessitate more cycles of induction followed by more intensive consolidation which may include haematopoietic stem cell transplantation.

Safety and Pharmacokinetics of Quizartinib Combination Therapy With Standard Induction and Consolidation Chemotherapy in Patients With Newly Diagnosed Acute Myeloid Leukemia: Results from Two Phase 1 Trials in Japan and China.

Quizartinib is a potent, oral, second-generation, selective type II FMS-like receptor tyrosine kinase 3 (FLT3) inhibitor. It has shown improved overall survival in a randomized, multinational, Phase 3 (QuANTUM-First) study in patients with FLT3-internal tandem duplication (ITD)-positive newly diagnosed acute myeloid leukemia. We conducted 2 Phase 1b studies in Japan and China to evaluate the safety, pharmacokinetics, and efficacy of quizartinib in combination with standard induction and consolidation chemotherapy in patients with newly diagnosed acute myeloid leukemia. Quizartinib was started at a dose level of 20mg/day and then escalated to 40mg/day, the dose used in the Phase 3 study. Seven patients were enrolled according to the 3 + 3 dose-escalation method in each study, including 3 patients who were FLT3-ITD positive. No dose-limiting toxicities were observed at dose levels up to 40mg/day in both studies. Grade 3 or higher, quizartinib-related, treatment-emergent adverse events included febrile neutropenia, hematologic toxicities, and infections. QT prolongation on electrocardiogram was observed in 5 patients. The pharmacokinetics of quizartinib and its metabolite AC886 were similar between the studies and consistent with previous findings in the United States. We confirmed the tolerability of Japanese and Chinese patients to the dose of quizartinib and chemotherapy regimens used in the QuANTUM-First study.

Combination of nivolumab with standard induction chemotherapy in children and adults with EBV-positive nasopharyngeal carcinoma : Protocol of aprospective multicenter phase2trial.

Treatment of Epstein-Barr virus(EBV)-positive nasopharyngeal carcinoma (NPC) with cisplatin/5-fluorouracil (5-FU) induction chemotherapy, followed by radiochemotherapy and subsequent interferon‑β, has yielded high survival rates in children, adolescents, and young adults. Aprevious study has shown that reduction of radiation dose from 59.4 to 54.0 Gy appears to be safe in patients with complete response (CR) to induction chemotherapy. As immune checkpoint-inhibitors have shown activity in NPC, we hypothesize that the addition of nivolumab to standard induction chemotherapy would increase the rate of complete tumor responses, thus allowing for areduced radiation dose in agreater proportion of patients. This is aprospective multicenter phase2 clinical trial including pediatric and adult patients with their first diagnosis of EBV-positive NPC, scheduled to receive nivolumab in addition to standard induction chemotherapy. In cases of non-response to induction therapy (stable or progressive disease), and in patients with initial distant metastasis, treatment with nivolumab will be continued during radiochemotherapy. Primary endpoint is tumor response on magnetic resonance imaging (MRI) and positron emission tomography (PET) after three cycles of induction chemotherapy. Secondary endpoints are event-free (EFS) and overall survival (OS), safety, and correlation of tumor response with programmed cell death ligand 1 (PD-L1) expression. As cure rates in localized EBV-positive NPC today are high with standard multimodal treatment, the focus increasingly shifts toward prevention of late effects, the burden of which is exceptionally high, mainly due to intense radiotherapy. Furthermore, survival in patients with metastatic disease and resistant to conventional chemotherapy remains poor. Primary objective of this study is to investigate whether the addition of nivolumab to standard induction chemotherapy in children and adults with EBV-positive NPC is able to increase the rate of complete responses, thus enabling a reduction in radiation dose in more patients, but also offer patients with high risk of treatment failure the chance to benefit from the addition of nivolumab. EudraCT (European Union Drug Regulating Authorities Clinical Trials Database) No. 2021-006477-32.

Mitocurcumin utilizes oxidative stress to upregulate JNK/p38 signaling and overcomes Cytarabine resistance in acute myeloid leukemia

Acute myeloid leukemia (AML) is a type of blood cancer that is characterized by the rapid growth of abnormal myeloid cells. The goal of AML treatment is to eliminate the leukemic blasts, which is accomplished through intensive chemotherapy. Cytarabine is a key component of the standard induction chemotherapy regimen for AML. However, despite a high remission rate, 70–80% of AML patients relapse and develop resistance to Cytarabine, leading to poor clinical outcomes. Mitocurcumin (MitoC), a derivative of curcumin that enters mitochondria, leading to a drop in mitochondrial membrane potential and mitophagy induction. Further, it activates oxidative stress-mediated JNK/p38 signaling to induce apoptosis. MitoC demonstrated a preferential ability to kill leukemic cells from AML cell lines and patient-derived leukemic blasts. RNA sequencing data suggests perturbation of DNA damage response and cell proliferation pathways in MitoC-treated AML. Elevated reactive oxygen species (ROS) in MitoC-treated AML cells resulted in significant DNA damage and cell cycle arrest. Further, MitoC treatment resulted in ROS-mediated enhanced levels of p21, which leads to suppression of CHK1, RAD51, Cyclin-D and c-Myc oncoproteins, potentially contributing to Cytarabine resistance. Combinatorial treatment of MitoC and Cytarabine has shown synergism, increased apoptosis, and enhanced DNA damage. Using AML xenografts, a significant reduction of hCD45+ cells was observed in AML mice bone marrow treated with MitoC (mean 0.6%; range0.04%–3.56%) compared to control (mean 38.2%; range10.1%–78%), p = 0.03. The data suggest that MitoC exploits stress-induced leukemic oxidative environment to up-regulate JNK/p38 signaling to lead to apoptosis and can potentially overcome Cytarabine resistance via ROS/p21/CHK1 axis.

Outcome of Intensively Treated Elderly AML Patients Reported to the Harmony Alliance Compares Well to Outcome of Control Patients of the Prospective Randomized HOVON 103 Study in Elderly AML

Introduction: The evaluation of novel drugs in hemato-oncology is hampered by relatively large sample sizes needed for sufficiently powered, randomized controlled trials (RCT). External control data are increasingly applied in prospective phase II and III studies, and might be derived from Real-World Data (RWD) sources, such as national cancer registries. The HARMONY Alliance recently reported on a big data platform, consisting of data that were generated by cooperative European acute myeloid leukemia (AML) study groups ( Lang et al., Trials, 2020). HARMONY or cancer registry data might supplement prospectively collected trial control data. Here, we investigated whether and to what extent data included in the HARMONY AML database and RWD from the Netherlands Cancer Registry (NCR) compare to the control arm of the recently reported prospective randomized HOVON-103 (H103) study in elderly AML ( Ossenkoppele et al., Leukemia, 2020; Janssen et al., Leukemia, 2022). Methods: The H103 trial consecutively randomized newly diagnosed AML patients aged >65 years between standard induction chemotherapy with or without lenalidomide, tosedostat, or selinexor. External control patients concerned newly diagnosed (HARMONY: 2010-2018; NCR: 2014-2018) and intensively treated (intensified cytarabine/anthracycline) elderly AML patients aged >60 years. H103 were matched 1:1 using nearest neighbor propensity scores with NCR and HARMONY patients for age, European Leukemia Net (ELN) 2022 risk classification, and leukocyte count at diagnosis, while HARMONY patients were additionally matched for WHO performance score. Patients with missing values for propensity score factors were excluded from that analysis. The primary endpoint was overall survival (OS), calculated by the Kaplan-Meier method. Results: 67% of HARMONY patients (n=510) concerned trial patients. Median age was lower for HARMONY patients compared with H103-controls (n=279) (67 vs. 69 years, P<0.01), with also a preponderance of male patients in the H103 cohort (54% vs. 62%, P=0.03). Age and male sex did not differ between H103-controls and NCR patients (n=320) (69 vs. 69 years, P=0.46; 62% vs. 62%, P=1.0) (Table 1). WHO performance score of 0 or 1 was reported in 49% and 41% of H103-controls which was less frequent in HARMONY and NCR data (8% and 31%, P<0.01; 26% and 18%, P<0.01, respectively). Unknown WHO status was more prevalent in the external cohorts than in H103, at 43%, 54%, and 1% for the HARMONY, NCR, and H103 cohorts, respectively. ELN 2022 risk (favorable vs. non-favorable) was reported in 24% vs. 76% of H103 patients, which was similar in HARMONY patients (26% vs. 74%; P=0.73). ELN 2022 risk differed, however, for NCR patients (17% vs 83%; P=0.03). The median leukocyte count at diagnosis was lower for H103-controls than for HARMONY and NCR patients (3.65 vs. 17.1, P<0.01; 3.65 vs. 13.1, P<0.01, respectively). Propensity scores matched 278 HARMONY patients and 278 NCR patients to H103-controls. The 2-year OS (estimate±SE) was 40±3% for H103-controls vs. 35±3% for HARMONY patients (P=0.22), and vs. 30±3% for NCR patients (P=0.052), respectively (Figure 1). Conclusion: Characteristics of H103-controls and HARMONY patients appeared largely comparable, while NCR patients were more frequently ELN 2022 non-favorable risk. Following matching, OS was similar between H103-controls and HARMONY patients, while NCR patients showed inferior OS. The inclusion of a large proportion of trial patients in the HARMONY cohort, adhering to strict inclusion and exclusion criteria and fewer comorbidities compared to RWD patients, might explain these observations. These results suggest that matched data of the HARMONY Alliance might supplement prospectively collected control data in studies evaluating intensive therapy in elderly AML.

Assessment of SIRT1-SIRT7 and TP53 Genes Expression in Patients with Acute Myeloid Leukemia

BACKGROUND Sirtuins ( SIRTs) are a family of histone deacetylases with 7 representatives ( SIRT1-7), which affect cell survival and metabolism. They influence gene expression by interacting with histones and transcription factors, or directly modulate pathways associated with cell survival, for instance by downregulating p53, thus potentially contributing to cancerogenesis. On the other hand, SIRTs may function as tumor suppressors depending on cellular context. While the role of SIRTs expression is widely discussed in solid neoplasms, they have not been comprehensively evaluated in acute myeloid leukemia (AML) yet. AIMS To investigate the prognostic importance of initial SIRTs expression in AML patients (pts) and its relationship with the expression of TP53. METHODS The study included 40 newly diagnosed AML pts (19 women, 21 men), with a mean age of 62 years (range: 26-87). Low-, intermediate- or high-risk AML according to European LeukemiaNet (ELN) 2022 stratification was diagnosed in 6 (15%), 13 (32.5%), and 21 (52.5%) of pts respectively. The expression of SIRT1-7 and TP53 was examined by real-time PCR using the TaqMan chemistry and the QuantStudio7 thermal cycler (Applied Biosystems-Thermo Fisher Scientific). The reference genes were selected using GeNorm and NormFinder. The mRNA levels were normalized by the formula ∆Ct = Ct(reference) - Ct(particular mRNA). Group comparisons were performed using Welch's t-test. Pearson correlation evaluated the relationship between individual gene expression. Overall survival (OS) was assessed by the Kaplan-Meier method with the log-rank test. Cox proportional hazard regression of clinical factors and genes expression for predicting OS was performed. RESULTS FLT3 ITDor TKD mutations were present in 10 (25%) of pts, NPM1 in 9 (23%), and IDH1 or IDH2 in 6 (15%). 14 (35%) of the study group were diagnosed with AML with myelodysplasia-related cytogenetic abnormalities, AML progressed from myelodysplastic syndrome, or therapy-related AML (secondary AML, sAML). Standard induction chemotherapy was given in 42.5% of pts, 25% were treated with azacitidine+venetoclax (Aza+Ven), and other non-intensive therapies were administered to 32.5%. SIRT4 expression was detected in 92.5% of pts, while expression of the other genes studied was found in all pts. There were no differences in SIRT 1-7 expression in terms of ELN2022 risk stratification. However, TP53 was downregulated in the high-risk group (fold change - FC=0.6, p=0.004). In pts with FLT3 mutation, SIRT7 was downregulated (FC=0.8, p=0.07), while in both NPM1 and IDH1/2 mutated groups, we showed upregulation of TP53 (FC=1.5, p=0.03, and FC=1.5, p=0.001, respectively). Moreover, in older pts (>60 years old) TP53 was downregulated (FC=0.7, p=0.02). In sAML group SIRT3 and TP53 were downregulated (FC=0.7, p=0.08, and FC=0.7, p=0.05, respectively). Significant positive correlations were shown between the expression of individual SIRTs, and it was most strongly expressed between SIRT2 and SIRT6 (r=0.85, p=0.00), SIRT2 and SIRT7 (r=0.81, p=0.00), and SIRT6 and SIRT7 (r=0.76, p=0.00). The median follow-up was 6.7 months (95% CI: 4.5-9.1). The median OS in the study cohort was not reached. There were no significant differences in treatment response or OS considering each SIRT expression level. However, pts with higher TP53 expression (> median) presented longer OS (median OS not reached vs 5.8 months (95% CI:1.3-5.8, p=0.05) [Figure 1]. In the multivariate Cox proportional hazard model for OS, upregulation of TP53 (HR 0.25, 95%CI: 0.08-0.60, p=0.01), age (HR 1.06, 95%CI: 1.01-1.12, p=0.03), as well as initial albumins (HR 0.01, 95%CI: 0.00-0.07, p=0.01), and LDH levels (HR 1.00, 95%CI: 1.00-1.01, p=0.01) were factors influencing the outcome [Table 1]. CONCLUSIONS The expression of SIRTs in AML presented no evident impact on treatment outcome. However, our study revealed that higher TP53 expression was associated with longer OS. Moreover, TP53 upregulation was found in pts with certain mutational profiles, while downregulation in the elderly and those with sAML. The analysis indicates that SIRTs expression may also vary markedly depending on the clinical and molecular context. The expression of individual SIRTs is closely correlated with each other, but these preliminary results did not show their direct relationship with TP53 expression. Further studies on SIRTs are needed to assess their impact on AML.

Myelodysplasia-Related Gene Mutations for NGS-Based MRD Assessment in Acute Myeloid Leukemia

Introduction: Measurable residual disease (MRD) plays an increasing role for treatment management of AML patients. While next-generation-sequencing (NGS) based MRD assessment can be well standardized, there is uncertainty about the type of gene mutations that associate with prognosis and are useful for MRD assessment. The 2022 ELN recommendations assign patients with at least one mutation in the genes ASXL1, BCOR, EZH2, RUNX1, SF3B1, SRSF2, STAG2, U2AF1 or ZRSR2 to the subgroup of myelodysplasia-related gene mutations (MRGM), which is associated with poor prognosis. We aimed to define the prognostic effect of MRD after induction chemotherapy in AML patients with MRGM using MRG mutations for MRD assessment by NGS. Methods: We retrospectively collected bone marrow and peripheral blood samples from AML patients in complete remission (CR) or CR with incomplete hematologic recovery (CRi) after one to two cycles of 7+3-based standard induction chemotherapy (IC) with at least one MRGM detected at diagnosis by myeloid panel sequencing covering 48 AML associated genes. Patients treated with one course of IC were included if they proceeded to allogeneic hematopoietic cell transplantation (alloHCT) already after one cycle. Molecular MRD status was assessed using targeted NGS and bioinformatics error-correction with 0.01% sensitivity as described before using only MRGM as MRD markers. Results: MRD was assessed on 104 adult newly-diagnosed AML patients (median age 58.6) with a total of 166 MRGM, resulting in a median of one MRGM per patient (range 1-4). 43 (41.3%) patients had secondary or therapy-related AML and 20 (19.2%) had myelodysplasia-related cytogenetic abnormalities. 68 (65.4%) patients were MRD positive for at least one MRGM after one (18 patients (17.3%)) or two (86 patients (82.7%)) cycles of intensive chemotherapy, while 36 (34.6%) became MRD negative. Measured variant allele frequencies (VAF) of positive samples ranged from 0.02-43.1% (median VAF 1.34%).MRD positive patients were older and more likely to be male compared to MRD negative patients, while cytogenetic risk, WBC count, Hgb, platelet count and ECOG status at diagnosis were similar to MRD negative patients. 53 (77.9%) MRD positive and 23 (63.9%) MRD negative patients underwent alloHCT in first CR/CRi.Median follow-up was 3.35 years. OS (P = 0.92), RFS (P = 0.99) and CIR (P = 0.98) were similar in MRD positive (at least one MRGM detectable) and negative (no MRGM detectable) patients (Figure 1). OS, RFS and CIR were also similar between MRD positive and negative patients, when MRD negativity was defined by conversion of at least one MRGM to negativity (55 (52.9%) MRD positive, 49 (47.1%) MRD negative), and when ASXL1 was excluded from the analysis. As variants with VAF >5% in CR/CRi samples may indicate clonal hematopoiesis, we excluded all variants with VAF >5% and repeated the analysis with the 88 (84.6%) remaining patients. Again, OS, RFS and CIR were similar for MRD positive and negative patients. We next reasoned that reduction of VAF correlates with the sensitivity of leukemic cells and possibly with depth of remission and long-term outcome. The fold-reduction of VAFs from diagnosis to the time of MRD analysis was calculated for all variants. For patients with several MRGMs, the variant with the largest fold-reduction was considered including undetectable variants. Patients with more or less than a 2-log, 3-log or 4-log reduction (log 10) had similar OS, RFS and CIR, respectively. Evaluating each MRGM individually, only RUNX1-MRD associated with RFS and by trend CIR (comparing MRD positive to negative patients for OS, HR 3.41, 95% CI 0.43-26.82, P = 0.24; RFS, HR 4.37, 95% CI 1.07-17.79, P = 0.04; CIR, HR 3.30, 95% CI 0.79-13.82, P = 0.1). Conclusion: 65.4% of AML-MRGM patients in CR/CRi remained MRD positive when MRGMs were used for MRD assessment by NGS. MRD positivity was associated with age above 60 years and male sex.Using all MRG mutations for MRD assessment at a threshold of 0.01% or by various degrees of log-reduction was not predictive of outcome after intensive induction chemotherapy. This limits the number of potential MRD markers for NGS analysis before alloHCT, although individual markers like RUNX1 may be of prognostic value.

Fms-like Tyrosine Kinase 3 Ligand Kinetic Profile Is the Strongest Factor Predicting Refractoriness after Induction and Event-Free Survival in Adults with AML: A Filo Prospective Multicentric Study

Background: Recently, a monocentric study has shown in adults with a non-previously treated acute myeloid leukemia (AML) that the stagnation of low levels of Fms-like tyrosine kinase 3 ligand concentration (FLc) in plasma, up to day 22 after an intensive induction, was associated with refractoriness and lower survivals (Peterlin et al, 2021). The present study wanted to confirm these results prospectively on a larger cohort of patients and in a multicentre way. Methods: This was a prospective non-interventional multicentre investigator sponsored trial (FLAMVAL study, clinicaltrials.gov: NCT04641910. One of the main objective was to study the impact on various outcomes of FL levels, evaluated during an intensive induction, in adults (>=18 year (y) old) with a non-previously treated non-M3 AML. Patients could receive a standard intensive induction chemotherapy combining anthracycline + cytarabine as part of a trial or not. Outcomes to be studied were responses, 1y overall survival (OS) and event-free survival (EFS), and 1-y leukemia-free survival (LFS) and relapse in responders. Response was evaluated according to ELN2017 criteria at day+30 after induction. Using a new cytokine multiplex assay from Milliplex (MILLIPLEX®), FL levels (expressed in pg/mL) were evaluated at Day (D) 1, D8, D15 and D22 from Day 1 of induction. Results: Between June 2021 and August 2022, 226 patients from 16 FILO centres were included but 204 with full evaluation of FL levels (816 samples) were considered for analyses. Their characteristics are reported in Table 1. At the time of analysis (July 2023), 1-y OS, EFS and LFS were 87+-5%, 78.3+-6% and 59.8+-8%, respectively. Median [Q1-Q3] FLc were 1.22 [0-4.19] at D1, 13.83 [2.26-72.46] at D8, 128.55 [13.06-293.43] at D15 and 24.14 [4.60-155.14] at D22. The number of refractory patients was significantly higher for those patients with a FLc < median at Day 8 (30.4% vs 13.7%, p=0.01), D15 (35.6% vs 8%, p<0.001) and D22 (31.1% vs 12.9%, p=0.007). 1-y OS and LFS were similar between patients with a FLc < or >= median at the four times considered. This was true also for 1y-EFS at D1 and D22. However, 1-y EFS was significantly worse for those with a FLc < median at D8 (48.5 % (41-55) vs 31 % (25-37), p=0.004) and at D15 (47.6 (40-55) vs 31% (25-37), p=0.002). As reported before (Peterlin et al, 2021), all patients but 4 (with a decrease of levels all along) can be categorized according to three FLc kinetic profiles: i) sustained increase from days 1 to 22 (FLI group n=39, 19% of the cohort), ii) increase from days 1 to 15, then decrease at day 22 (FLD group, n=86, 42% of the cohort) and iii) stagnation of low levels all along (<100 pg/mL from days 1 to 22, FLL group, n=75, 37% of the cohort). OS and LFS were similar between these three groups while 1-y EFS was significantly lower for the FLL group (45% [33; 57]) compared to FLI and FLD groups (64% [49; 79] and 71% [ 61; 80], p<0.001) ( Figure 1). This difference was due to a higher proportion of refractory patients in the FLL group (n=45/75 42.7% vs FLI =5/39, 12.8% vs FLD n=8/86, 9.3%, p<0.001, 2 patients not evaluated for response). Similar incidence of relapse was observed for responders comparing the three groups. Interestingly, only 8.8% (n=11/125) of the FLI/FLD patients had a FLc level <100 pg/ml at both times D8 and D15, suggesting that a FLL status can be predicted as soon as D15. For multivariate analyses, we compared FLc kinetic profiles (FLL vs FLI/FLD) on one hand and FLc medians (D1 D8 D15 and D22) on the other hand with other factors associated with a p value <0.20 by univariate analyses. In both models, only FLc kinetic profile and medians together with ELN2017 classification were found to be independent factors predicting response and EFS. However, FLL status was the strongest factor associated with refractoriness (OR: 6.2, 95%CI: 2.94-13.73, p<0.001, ELN favorable vs Intermediate p=0.003, favorable vs adverse p=0.002) and lower EFS (OR: 2.58, 95%CI: 1.40-4.79, p=0.002, ELN p=0.01 and 0.004). Conclusion: This large prospective multicentre trial confirms that a stagnation of low levels of FLc during a first-line induction in adults with non-M3 AML is the strongest factor (above ELN classification) associated with refractoriness and lower EFS in these patients. The study suggests also that this simple test could be useful to envisage salvage therapy including allotransplant just after D15 of induction. Funding This study was funded by Astellas Pharma Inc.

Long Non-Coding RNA LINC00XPRR Is Upregulated in Newly Diagnosed Multiple Myeloma Promoting Poor Response to High-Dose Melphalan with Autologous Stem Cell Transplantation and Three-Drug Regimen

Abstract Multiple myeloma is an incurable malignant hematopoietic blood cancer. It develops from genetic disposition and abnormal plasma cells with over 30,000 new cases diagnosed in the United States every year. The 5-year relative survival rate for myeloma patients if diagnosed at early stage is 79.5%, however majority of patients are diagnosed once the cancer has already metastasized which decreases the 5-year relative survival rate to only 59%. The standard care for treating newly diagnosed multiple myeloma (NDMM) is three-drug induction therapy followed by high-dose melphalan (HDM) and autologous stem cell transplantation (ASCT). While there is no cure for multiple myeloma, diagnosis and standard therapy leads to prolonged remission and improved quality of life. Identifying genetic predictors for positive treatment responses and survival represents an unmet medical need. Long non-coding RNAs (lncRNAs) play important regulatory roles in promoting cancer growth and metastasis and due to their tissue-specificity they are becoming important players for development of personalized therapies. In addition, lncRNA-protein interactions have shown to be vital for biological processes associated with cancer, but currently there is a lack in our understanding of their role in initiation or progression of myeloma. Further, the roles of lncRNAs as biomarkers or targets for myeloma therapies have not yet been defined. In order to address these issues, we performed transcriptome sequencing from NDMM patients which identified 244 differentially expressed lncRNAs in poor responders (indicated as <24 months progression free survival (PFS)) to three-drug regimen, HDM and ASCT (n=38) compared to standard responders (indicated as >24 months PFS) to three-drug regimen, HDM and ASCT (n=78). Our analysis revealed LINC00XPRR (lncRNA00Xpoor responder-related) as one of the top-most upregulated novel lncRNA associated with poor responders to standard myeloma care. We next validated LINC00XPRR high expression in single cell RNA-sequencing data in NDMM patient samples. Multiplexed fluorescent RNA in situ hybridization (mFISH) analysisshowed that LINC00XPRR expression increased in melphalan treated myeloma cell lines and is highly expressed in bone marrow aspirates from patient samples. We further determined higher levels of LINC00XPRR in the cytoplasmic compartment of the cells compared to nuclear localization through QuPath analysis. LINC00XPRR mechanistically and epigenetically modulates cytotoxic and apoptotic phenotypes in myeloma cell lines when overexpressed in U266B1 and MM1.S cells or using RPMI8226 CRISPR/Cas9 knockdown system and in appropriate multiple myeloma-associated mouse models. Additionally, we used RNA immunoprecipitation (RIP) combined with qPCR to determined that LINC00XPRR directly interacts with CELF2, RNA-binding protein. Lastly, using target specific LINC00XPRR designed locked-nucleic acid antisense oligonucleotides (LNA ASOs) to knockdown LINC00XPRR, we saw an increase in apoptotic phenotypes indicating clinical potential of LNA ASOs-mediated LINC00XPRR knockdown. Overall, we are the first to identify lncRNAs associated with poor response in NDMM patients to standard three-drug induction chemotherapy followed by HDM and ASCT and show that LINC00XPRR enhanced expression may be both an important biomarker and target for future therapeutic interventions. Our current studies focus on the biology of lncRNAs in myeloma and the mechanism by which increased expression of LINC00XPRR is associated with poor outcomes in patients with NDMM receiving standard induction therapy, HDM and ASCT. Topics: multiple myeloma, long non-coding RNAs, LNA ASOs, RNA therapeutics, treatment response, RNA sequencing, apoptosis

Picalm::MLLT10 can Identify a New Subgroup of Acute Leukemias of Ambiguous Lineage with Unique Extramedullary Disease and Treatment Response

Background: Mixed-phenotype acute leukemias (MPAL), which is included in acute leukemias of ambiguous lineage (ALAL), can be classified as four different subtypes based on recurring genetic alterations including BCR::ABL, KMT2A and ZNF384 gene rearrangement as well as BCL11B activation. The 2022 WHO classification has already mentioned PICALM::MLLT10 fusions are also enriched in MPAL but need more data. In 2022 ELN classification, PICALM::MLLT10 was identified as acute myeloid leukemia (AML) with other rare recurring translocations but not ALAL. In addition, there is no separate classification for PICALM::MLLT10 in the new ICC classification. Therefore, it is still confused whether PICALM::MLLT10 corresponds to AML or acute lymphoblastic leukemia (ALL) or ALAL. Methods: Fourteen PICALM::MLLT10 positive patients of 390 AL patients (14/390, 3.6%) were identified by RNA sequencing (RNA-seq) in our center from 2020-2022, including 8 female and 6 male, with a median age of 33 years (16-50 years). These patients were newly diagnosed AL according to bone marrow morphology and immunology including 5 ALAL, 5 T-ALL [ 4 early T-cell precursor ALL (ETP-ALL), 1 cortical T-ALL], 2 AML, 1 B-ALL with aberrant expression of myeloid antigen, and 1 B/T MPAL. Results: The mean white blood cell counts of these patients was 8.25×10 9/L (2.34-51×10 9/L) and platelet counts was 168.5×10 12/L (39-429×10 12/L) individually. It should be mentioned that extramedullary disease (EMD) was found in 7 cases (7/14), including mediastinum, tonsil, and skin. In terms of immunotyping, CD7 was identified in all patients (14/14) and CD33 in 71.4% (10/14) patients. The major concurrence mutations were PHF6 mutation (8/14), JAK3 mutation (5/14), and SUZ12 mutation (4/14). Characteristic cytogenetic abnormality t(10;11)(p12.3;q14.2) was only found in three cases (3/14) (Table 1). The PICALM breakpoints are mainly concentrated in exon 17 (n=6) and exon 19 (n=8). Exon 4 (n=8), exon 6 (n=2), exon 9 (n=2) and exon 10 (n=2) are the most commonbreakpointsof MLLT10. To our knowledge, this is the first report about the breakpoints and fusion gene forms of PICALM::MLLT10. For initial treatment, these patients individually received standard ALL induction chemotherapy (VDPCP, vincristine, idarubicin, pegaspargase, cyclophosphamide, prednisone) and AML induction chemotherapy (3+7 IA regimen including idarubicin, cytarabine). The initial complete remission (CR) rate was only 35.7% (5/14), and 9 patients showing no remission (NR). These NR patients subsequently received salvage chemotherapy. It is worth mentioning that 6 of them received combined chemotherapy regimen including low doses of cytarabine (LDAC), granulocyte colony-stimulating factor (G-CSF) and anthracyclines such as aclarubicin or idarubicin or homoharringtonine (CAG or IAG or HAG), and 2 of 6 patients also further received combination therapy with venetoclax (CAG or IAG +VEN). Finally, 5 of them dramatically achieved CR (5/6). In the other 3 patients, 1 patient who was failure to initial Hyper-CVAD A/B regimen also dramatically achieved CR after the more Hyper-CVAD A regimen combined with venetoclax, and 1 patient achieved CR while another still NR with both receiving CLAG (cladribine, cytarabine, G-CSF) regimen. Subsequently 11 patients (9 CR, 2 with refractory disease) received allogenic hematopoietic stem cell transplantation (allo-HSCT), and all the CR patients survived well after HSCT (follow up 3-24 months, median 15 months) without relapse, while 2 NR patients died soon after transplantation because of severe complications (Table 2). Conclusions: Our data suggested PICALM::MLLT10 positive AL should be more appropriately recognized as an independent ALAL entity and may benefit from LDAC, G-CSF and anthracyclines combination chemotherapy as well as venetoclax. Sequential HSCT after chemotherapy combined with venetoclax may further improve long-term survival in AL patients with CR even MRD positive.

Patient-Reported Outcomes in Acute Myeloid Leukemia Patients with FLT3-ITD Mutation Receiving Quizartinib Vs. Standard Chemotherapy: Results from the Quantum-First Trial

Introduction: QuANTUM-First (NCT02668653) is a multicenter, global, randomized, double-blind phase 3 clinical trial evaluating the efficacy and safety of the novel oral fms-like tyrosine kinase 3 inhibitor (FLT3i) quizartinib in combination with standard induction and consolidation chemotherapy, and as maintenance monotherapy for up to 36 cycles in patients 18-75 years old with FLT3-ITD positive acute myeloid leukemia (AML). Quizartinib demonstrated a clinically meaningful and statistically significant improvement in overall survival (OS) versus placebo. An exploratory endpoint of the trial was to assess the impact of quizartinib on patient-reported outcomes (PROs). This is the first report of the longitudinal results of the PRO measures (PROMs) collected in the QuANTUM-First trial. Methods: Patients were randomized to receive chemotherapy + quizartinib or chemotherapy + placebo. PROMs were the European Organization for Research and Treatment of Cancer QLQ-C30 (EORTC QLQ-C30) and EuroQol EQ-5D-5L. Baseline measurement was collected on day 8 of the first induction cycle, and repeated measurements were collected on day 28 of induction cycles 1-2, day 6 and 28 of consolidation cycles 1-4, and day 1 of continuation cycles 1-34 at 3-cycle intervals. EORTC QLQ-C30 and EQ-5D-5L scores were calculated as per their respective scoring manuals and included mean (95% CI, p-value) score for each domain of the EORTC QLQ-C30 and EQ-5D-5L index score, as well as mean change from baseline score at each timepoint. A minimal clinically important difference (MCID) score ≥10 points for each domain of the EORTC QLQ-C30, was considered clinically meaningful. A mixed-effect model for repeated measures (MMRM) and time until definitive deterioration (TUDD) was used to assess the longitudinal impact of treatment on PROs. TUDD was defined as time from baseline PRO score to first deterioration of the score beyond the MCID as compared to the baseline without further improvement of more than one MCID as compared to the reference score or without any further available score. Results: A total of 509 out of 539 patients from the trial (254 quizartinib; 255 placebo) were included for PRO analysis. Patient compliance to questionnaire completion at the beginning of each phase was high and similar for both arms and most scales (Global Health Status [GHS] completion rate (%) on induction cycle 1, consolidation cycle 1, and continuation cycle 1: 99.2, 95.3, 93.4, respectively) Baseline PRO scores were comparable between arms and were lower than the general population norm EORTC QLQ-C30 and EQ-5D-5L scores for EU and US (Table 1). Per MMRM, there were clinically meaningful improvements from baseline in GHS (Figure 1) and fatigue in both arms. However, there was no significant difference between arms for the change from baseline score although placebo arm had numerically better scores specifically during the maintenance phase (treatment difference for quizartinib - placebo was -2.0 [95% CI: -4.8, 0.7], p=0.1479 for GHS and 3.0 [95% CI: -0.1, 6.1], p=0.0600 for fatigue). Similar trends towards clinically meaningful improvement in score over time for both arms and no significant differences between arms were observed in most of the functional and symptom scales of EORTC QLQ-C30 and EQ-5D-5L index scores. By TUDD analysis, quizartinib arm showed slower deterioration in several scales (specifically GHS, cognitive function, appetite loss, and constipation) but the differences did not reach statistical significance. Conclusions: QuANTUM-First is the first study to explore the impact of quizartinib on PROs. Quizartinib showed improvement in OS without any detrimental impact on quality of life and symptoms when added to standard chemotherapy followed by maintenance monotherapy in patients with newly diagnosed FLT3-ITD AML.

Targeting CDK8/CDK19 to Disrupt Leukemic Stem Cell-like Population in Acute Myeloid Leukemia: Exploring RVU120 As a Promising Frontline Therapy

Leukemic stem cells (LSCs) are implicated in the initiation, maintenance, and relapse of leukemia. The failure of current standard induction chemotherapy, which primarily targets dividing cells, has been linked to the quiescence of LSCs, leading to resistance to treatment. Consequently, there is an urgent need for novel therapies specifically aimed at eradicating LSCs to achieve more effective and durable treatment responses. CDK8 and its paralog CDK19 associate with the mediator complex and regulate gene expression programs linked to cancer and stem cell pluripotency. RVU120, a highly selective and potent CDK8/19 inhibitor, is currently undergoing clinical trials for AML and high-risk myelodysplastic syndrome (HR-MDS) (NCT04021368). Previous studies have demonstrated its potential mechanism of action on LSCs, wherein inhibition of CDK8/CDK19 led to the activation of lineage commitment genes in sensitive AML cells. Through recent gene expression analysis, it was discovered that inhibition of CDK8/CDK19 in transformed CD34+ cord blood cells resulted in the downregulation of stemness and quiescence-associated genes, while simultaneously upregulating genes associated with hematopoietic lineage-committed progenitors. Building on these findings, this study further investigates the potential of CDK8/CDK19 inhibition as a means to target LSCs, providing a rationale for RVU120 as a frontline treatment option in AML. The impact of RVU120 on LSCs has been investigated using an in vitro hierarchical AML-8227 model. This primary model maintains distinct populations in culture conditions, including CD34+CD38- cells with quiescent LSC potential, CD34+CD38+ cells enriched in progenitors, and CD34- more mature blasts (Lechman et al., 2016, Cancer Cell). The CD34+CD38- LSC population was further characterized by flow cytometry showing the expression of LSC surface markers, namely CD45RA, CD123, CD44, and negative expression for CD90, rendering it a suitable model for mechanistic studies. Next, AML-8227 cells were subjected to RVU120 treatment, resulting in a noticeable reduction in cell proliferation and viability. Flow cytometric analysis demonstrated a significant decrease in the percentage and total cell number of LSC-enriched/progenitor populations within the AML hierarchical model upon inhibition of CDK8/CDK19. Furthermore, RVU120 treatment induced the differentiation of cells, as evidenced by a sequential increase in the percentage of early myeloid differentiation marker, CD64, and late myeloid differentiation marker, CD14 (Fig. 1). In order to deconvolute the activity and mechanism of action of RVU120 in different cell populations, single-cell RNAseq analysis on AML-8227 cells has been performed at 24h and 3 days of treatment using technology from 10xGenomics. Additionally, analyzed AML-8227 cells have been enriched in the CD34+ population to evaluate the effect of RVU120 on stem and progenitor fraction at the single-cell level. The bioinformatic analysis of the performed experiment is ongoing and its results will be integrated into the data described above. CDK8 activity can be conveniently tracked by measuring specific phosphorylation of its target S726 on STAT5 protein. Dose and exposure-dependent inhibition of this marker has been established in cells exposed to RVU120. Levels of STAT5 S726 have been measured in different cell populations in patients from the first-in-human study of RVU120 in AML/HR-MDS. Consistent with established roles of CDK8 in LSCs, significantly elevated levels of pSTAT5 were observed in CD34+ stem/progenitor-enriched population when compared to CD34- blast cells measured at baseline before the start of the treatment (p<0.0001; ratio paired t-test; updated data from ASH 2022; Fig. 2). In summary, RVU120 displays cytotoxic and differentiating effects on well-characterized LSC populations in various models. Additionally, the CDK8 activity marker is directly associated with LSC-enriched bone marrow cells in AML patients, highlighting RVU120's potential as a promising therapeutic for targeting AML LSCs. Moreover, single-cell studies offer insights into its inhibitory effects on LSC-enriched populations and capacity for inducing differentiation. Overall, these results support RVU120 as a frontline candidate in AML therapy, countering therapeutic failure caused by persistent LSCs.

Single-Center Experience of Decitabine/Venetoclax in Newly Diagnosed Older AML Patients with Non-Favorable Genetics: Bridging to Allo-HCT or Treatment Continuation

Background DNA-hypomethylating agents (HMAs) have improved the prognosis of patients (pts) with acute myeloid leukemia (AML) unfit for standard induction chemotherapy. AML pts aged >60 years eligible for allografting (HCT) can be effectively bridged to HCT by HMA monotherapy. This was demonstrated in the EORTC randomized phase III trial AML21 (NCT02172872) comparing 10-day decitabine (DEC) to “7+3” induction (Lübbert et al., EHA 2022, S125), with similar transplant rates and post-HCT outcomes in both arms. Also the combination of an HMA with venetoclax (VEN) is not only effective in pts unfit for more intensive treatment (DiNardo et al., New Engl. J. Med. 2020) but also in pts fit for HCT (Pollyea et al., BMT 2021, Maiti et al., ASH 2022, #7725). A recent analysis demonstrated that HMA+VEN as pretransplantation therapy can attain post-HCT outcomes comparable to intensive chemotherapy regimens (Winters et al., TCT 2022). Patients and Methods We retrospectively analyzed 47 consecutive older AML pts receiving firstline treatment with DEC+VEN. DEC was administered on five consecutive days per cycle. VEN was administered up to 28 days per cycle (see Table 1). All pts who had received at least one dose of DEC and VEN were included. Data on all pts were retrospectively collected as reported in the medical charts. Written informed consent of all pts to analysis of their records for scientific purposes was provided. Key parameters captured and analyzed were: age, ECOG performance status (PS), ELN 2017 risk, number of cycles of DEC+VEN administered, response rates and overall survival (OS). Common safety parameters (e.g., prolonged neutropenia, tumor lysis syndrome, etc.) were assessed. Results Between 2/2019 and 2/2023, 47 pts with AML received DEC+VEN as firstline treatment. Of those, 18 pts (38%) proceeded to HCT, 29 (62%) did not (ineligibility for HCT, wish of pt, intercurrent events etc.). Median follow-up was 25.0 (HCT) and 14.8 months (no HCT). Key pt characteristics in the two groups were i. median age: 69 and 79 years, ii. ECOG PS 0: 50 and 27%, iii. de novo AML: 39 and 72%, iv. ELN 2017 adverse risk: 67 and 59%, median number of treatment cycles administered were 2.0 and 3.0. Safety data are shown in Table 1. A hematologic response before HCT (CR, CRi, MLFS) was attained in 8/18 pts (44%), with only 2/18 receiving >2 cycles. Median time from treatment start to HCT was 2.5 months (for transplant details, refer to Table 1). For non-transplanted pts, CR, CRi, MLFS as best response were attained in 16/29 (55%), 13/29 receiving >4 cycles. In pts receiving HCT, graft-versus-host-disease occurred in 10/18 pts (56%), median OS was not reached (16/18 pts alive; Fig. 1). In pts not receiving HCT, median OS was 9.0 months. A direct comparison of treatment outcomes between the two groups is not possible, given the different selection factors operative for pts attaining HCT or not. Conclusions Among these older AML pts with non-favorable genetics, administration of a limited number of DEC+VEN cycles followed by HCT was well feasible in almost 40% of pts (median age 69 years) and resulted in very encouraging post-HCT OS in this first analysis. In pts not able to receive HCT (median age 79 years), DEC+VEN resulted in response rates, treatment continuation and OS comparable to “real-world” older AML pts treated with AZA+VEN, suggesting equivalent efficacy of both regimes.

Oral Decitabine/Cedazuridine Vs Intravenous Decitabine for Acute Myeloid Leukemia: Final Results of a Randomized, Crossover, Registration-Enabling, Pharmacokinetics Study

Background: Parenterally administered hypomethylating agents (HMAs), decitabine (DEC) and azacitidine (AZA), are approved in Europe for adult patients with acute myeloid leukemia (AML) who are not candidates for standard induction chemotherapy as single agent or in combination with venetoclax. Cedazuridine(C) is a novel, potent, and safe inhibitor of cytidine deaminase (CDA) and when given in combination with decitabine, cedazuridine enables efficient oral availability of decitabine.ASTX727 (DEC-C) is a fixed-dose combination (FDC) tablet of 35 mg DEC and 100 mg cedazuridine, DEC-C has been approved for MDS in the US, Canada, and Australia. Here we present final results using DEC-C in an AML population appropriate for single-agent decitabine treatment. Aims: Describe the final clinical outcomes of patients with AML with additional analysis of genetic profiles. Methods: The ASCERTAIN study used a randomized two-period, two-sequence, two-treatment, crossover study design. Patients were randomized 1:1 to either Sequence A: DEC-C (35 mg DEC/100 mg cedazuridine) in Cycle 1 followed by IV-DEC at 20 mg/m 2 in Cycle 2, or Sequence B: receiving IV-DEC in Cycle 1 followed by DEC-C on Cycle 2 to compare pharmacokinetics (PK) [primary endpoint Area Under the Curve (AUC) equivalence over 5 days of dosing]. All patients received DEC-C from Cycle 3 until treatment discontinuation to assess safety and clinical efficacy. Patients were eligible as per the EMA-approved decitabine label (newly diagnosed AML who are not candidates for standard induction chemotherapy). Clinical responses were assessed according to modified International Working Group (IWG) 2003 response criteria. Pre-treatment peripheral blood was used for DNA isolation from leukocytes, and molecular abnormalities identified using a NGS hematologic malignancy panel. Cox-regression analysis on the various factors (eg binary mutational status, CR, etc.) was used for analyses of risk-factors for overall survival (OS). Results: 89 patients were randomized, of whom 87 were treated. The median age of patients was 78.0 years (range, 61 to 92) with 31 (35.6%) males and 56 (64.4%) females. Cytogenetic risk classification was poor-risk in 33 (37.9%), intermediate-risk in 45 (51.7%), and 9 (10.4%) not evaluated patients. For the primary endpoint, preliminary PK data was available from 69 patients who successfully completed PK assessments for both IV-DEC and DEC-C cycles, and the DEC AUC 0-24 (h*ng/mL) 5-Day geometric mean estimate was 904 for DEC-C and 907 for IV-DEC resulting in an oral/IV geometric LSM AUC ratio of 99.64% (90% CI of 91.23-108.8%). Safety findings were consistent with those anticipated for IV-DEC (related Grade ≥ 3 AEs in more than 10% were thrombocytopenia, neutropenia, anaemia, and febrile neutropenia). As of the database lock (25May2023), median follow up was 24.5 (20.2, 25.0) months (25% of 19.7 and 75% of 25.2 months). The best response was complete response (CR) in 19/87 (21.8%, 95% CI: 13.7, 32.0%); CR with incomplete blood cell count recovery (CRi) in 5 patients (5.7%); and CR with incomplete platelet recovery (CRp) in 2 (2.3%); resulting in composite response rate [CR + CRi] of 27.6%. The median overall survival was approximately 8.9 months (95% CI: 6.0, 13.1).These results obtained with DEC-C are consistent with those observed for IV-DEC. Multiple genes, including ASXL1, TP53, RUNX1, and SRSF2 genes were present in more than 20% of the AML patients. TP53 mutation was associated with worse survival. Summary/Conclusion: This randomized phase 3 study in AML patients not candidates for standard induction chemotherapy demonstrates that Daily X 5 dosing of the oral FDC of DEC-C resulted in an equivalent DEC AUC exposure to IV-DEC at 20 mg/m 2 over 5 days. Pharmacodynamic data showed similar demethylation rates (≤1.1% difference). In addition, safety findings and observed clinical activity (response rates and OS) are consistent with published data from IV-DEC, suggesting that DEC-C has the potential to be an oral alternative to the standard IV-DEC Daily×5 regimen. These findings support the use of DEC-C in patients with AML who are not candidates for standard induction chemotherapy.

Kinetics of Complete Remission (CR) and CR Duration and Its Impact on Overall Survival (OS) and Event-Free Survival (EFS) in Quantum-First

Background Results from the phase 3 QuANTUM-First study (NCT02668653) showed that in patients (pts) with newly diagnosed (nd) acute myeloid leukemia (AML) and positive for FMS-like tyrosine kinase 3-internal tandem duplication ( FLT3-ITD+), the addition of the oral, highly potent, and selective, type 2 FLT3 inhibitor quizartinib (Quiz) to standard chemotherapy ± allogeneic hematopoietic cell transplantation (allo-HCT), followed by continuation monotherapy for up to 3 years, decreased the relative risk of death by 22.4% vs placebo (PBO) (PMID: 37116523). We assessed 1) the impact of CR duration on OS and EFS and 2) the kinetics of CR achievement over time after induction. Methods QuANTUM-First enrolled pts aged 18-75 years with FLT3-ITD+ nd AML, randomized to receive standard induction chemotherapy with either Quiz (40 mg/d) or PBO, and stratified by region (Europe, North America, or Asia/Australia/South America), age (<60 years, ≥60 years), and white blood cell count (WBC; <40×10 9/L, ≥40×10 9/L) at diagnosis. Pts who achieved CR or CR with incomplete hematologic recovery (CRi) received up to 4 cycles of high-dose cytarabine combined with Quiz (40 mg/d) or PBO and/or allo-HCT followed by up to 3 years (36 cycles) of continuation therapy with Quiz (30-60 mg/d) or PBO. OS was the primary endpoint, EFS and rates of CR and composite complete remission (CRc) were secondary endpoints; CR duration was an exploratory endpoint. The impact of CR duration on OS was analyzed by extended Cox regression model stratified by region, age, and WBC count at diagnosis and included treatment group and CR duration status (time-dependent covariate). CR duration status was 0 for pts who did not achieve CR, 1 for CR pts throughout the duration of sustained CR, and 0 for CR pts after they relapsed. Results Of the 539 randomized pts enrolled in QuANTUM-First (Quiz, n=268; PBO, n=271), there were 297 CR pts (Quiz, n=147; PBO, n=150) based on independent review committee assessment after 1 to 2 courses of induction. Among 533 treated pts, 20% had 2 induction cycles. Baseline characteristics in CR pts were well balanced between arms: median age in years (range), 56.0 (23-75) vs 55.5 (20-74); females, 55.8% vs 60.0%; ECOG ≥1, 64.6% vs 63.3%; mutated NPM1 / CEBPA,66.7% vs 64.7% / 25.9% vs 23.3%; variant allele frequency (VAF) >25%, 68.7% vs 66.7%; and WBC count at diagnosis ≥40×10 9/L, 55.8% vs 49.3%. Median OS in CR pts was not reached (95% CI 48.6 months-NE) in the Quiz arm vs 35.4 (95% CI 15.1-NE) months in the PBO arm (HR 0.572, 95% CI 0.400-0.816; Figure 1). Although CR rates were similar between arms (55% in both), median CR duration was longer with Quiz vs PBO (38.6 months, 95% CI 21.9-NE vs 12.4 months, 95% CI 8.8-22.7; HR 0.621, 95% CI 0.451-0.857). Based on the extended Cox model, CR duration status covariate was strongly predictive for OS (HR 0.156, 95% CI 0.113-0.216; nominal P<0.0001; Figure 2). Importantly, the HR for Quiz vs PBO effect on OS in the model with the time-dependent covariate CR duration status was higher (HR 0.851, 95% CI 0.672-1.076) vs the HR of the primary OS analysis (HR 0.776, 95% CI 0.615-0.979), implying that a substantial fraction of OS benefit was mediated through its effects on achieving a durable CR (Figure 2). EFS favored Quiz over PBO when induction treatment failure (ITF) was defined as not achieving CRc or CR, by the end of induction up to day 56 from the last induction cycle, with HR 0.729 (95% CI 0.592-0.897; nominal P=0.0031) and 0.818 (95% CI 0.669-0.999; nominal P=0.032), respectively. EFS was similar between arms when ITF was defined, according to FDA guidance, as not achieving CR by day 42 from the start of the last induction cycle (HR 0.916, 95% CI 0.754-1.114; P=0.24; primary EFS analysis). Between day 42 and the end of the induction phase, there were 51 pts who achieved CR (Quiz, n=33; PBO, n=18), and they were considered as ITF with EFS event on day 1 in the primary analysis of EFS. Among these 51 pts, 9 (Quiz, n=5; PBO, n=4) had CRi by day 42 and became CR after day 42 in the induction phase. Conclusions These analyses demonstrated that in QuANTUM-First: 1) a substantial fraction of the estimated effect of Quiz on OS was mediated through its effect on achieving a durable CR and 2) more pts in the Quiz arm vs the PBO arm achieved CR after day 42 of the last induction cycle, suggesting that there was a delay in CR achievement with Quiz. Longer EFS was observed in Quiz over PBO based on the EFS definition including CRs between day 42 and end of induction.

Decider-2: Prospective Randomized Multicenter Phase III Trial of Decitabine and Venetoclax Administered in Combination with All- Trans Retinoic Acid (ATRA) or Placebo in Patients with Acute Myeloid Leukemia Who Are Ineligible for Induction Chemotherapy

Background and Significance DNA-hypomethylating agent (HMA) based therapy is the treatment backbone for older, medically non-fit AML/MDS pts, and studies have been performed with multiple combination partners over the last years (reviewed by Stomper et al., Leukemia 2021). Recently, the combination of azacitidine or decitabine (DEC) with venetoclax (VEN) has become the novel standard for many of these pts. Based on preclinical evidence of priming of AML cells to all- trans retinoic acid (ATRA) by DEC, we have conducted a randomized phase II trial in 200 elderly and medically non-fit AML pts on the effect of ATRA as add-on to DEC (DECIDER-1, Lübbert et al., J. Clin. Oncol 2020). Median overall survival (OS) was 8.2 months with ATRA v 5.1 months without ATRA (hazard ratio, 0.65; 95% CI, 0.48 to 0.89; P = .006). Notably, ATRA did not add toxicity, the combination was active also in AML with adverse genetics, and it appeared to prolong time to secondary treatment resistance. A randomized open-label phase II trial (DEC -/+ ATRA) is also being conducted in higher-risk MDS pts (Zhou et al., ASH 2021, 539). Investigating a triple combination of HMA+VEN+ATRA is a logical next step. We could demonstrate cooperativity between these 3 drugs in AML cell lines irrespective of TP53 mutation status on cell growth inhibition and apoptosis (Bresser et al., ASH 2022, 11537). Study Design and Methods This academic study is a randomized, prospective, double-blind, placebo-controlled, parallel group, multicenter trial. It is planned to randomize 256 pts in a ratio of 1:1 to the treatment arms at ~35 sites in Germany. The primary objective is to compare the efficacy of ATRA v placebo as add-on to the standard treatment DEC+VEN with respect to OS. Secondary objectives are to compare ATRA v placebo as add-on to DAC+VEN with respect to objective best response (i.e. CR/CRi/MLFS/PR), CR with negative measurable residual disease (MRD), probability of survival with objective best response, health-related quality of life and safety. Sample size calculation is based on the primary endpoint OS. The study should have a power of 80% to demonstrate superiority of ATRA vs. placebo with respect to OS at one-sided level alpha of 2.5% under the assumption of a hazard ratio of 0.67. This requires the observation of 191 deaths. With a recruitment period of 3 years and an additional one year follow-up period, it is necessary to include 230 pts. To account for the possibility of up to 10% of the pts with incomplete follow-up, 230/0.9 = 256 pts will be randomized in total (ratio 1:1). Interim safety analyses will be performed after the 2nd cycle of the 10th randomized pt, and after the 3rd cycle of the 50th, the 100th and the 150 th randomized pt, and monitored by an Independent Data Monitoring Committee. Key inclusion criteria are: age ≥18 years; previously untreated AML (WHO 2016); ineligibility for standard induction chemotherapy; ECOG ≤ 2; WBC <25×109/L (prior cytoreduction allowed). Key exclusion criterion is: acute promyelocytic leukemia. Treatment All pts will be treated with DAC+VEN as standard treatment and will additionally either receive ATRA or placebo (Fig. 1). Study treatment is continued until relapse/progression, hematopoietic cell transplantation, death or unacceptable toxicity (whichever occurs first). Correlative investigations include serial MRD measurement, NGS profiling and apoptosis studies. Registration EUDRACT 2020-005495-36; DRKS 00023646 (P001516). Funding Funded by the German Research Foundation (DFG, project no. 367848349/LU 429/13-1); ATRA is provided free of charge by Cheplapharm (Germany). Figure 1 Overview of the study treatment