TPS6602 Background: There are currently no approved targeted therapies for newly diagnosed (ND) acute myeloid leukemia (AML) harboring a nucleophosmin-1 mutation ( NPM1 m), which occurs in ~30% of ND adult AML cases. In NPM1 m AML, the NPM1m/XPO1 protein complex binds to DNA to sustain the interaction of menin and wild-type KMT2A that drives upregulation of HOX / MEIS gene expression, resulting in hematopoietic differentiation arrest and leukemogenesis. Revumenib is a first-in-class, oral, potent, and selective inhibitor of the menin-KMT2A interaction. In the phase 1/2 AUGMENT-101 study (NCT04065399), revumenib monotherapy demonstrated clinically meaningful response rates and was generally well tolerated in relapsed/refractory NPM1 m AML, leading to US Food and Drug Administration approval for that patient population on October 24, 2025. Standard AML treatment for younger, non-frail adults is based on intensive chemotherapy (IC) regimens and hematopoietic stem cell transplant (HSCT). The addition of revumenib to standard IC may further improve treatment responses specifically in the ND setting. This study is designed to assess the safety and efficacy of revumenib in combination with IC in patients with ND NPM1 m AML. Methods: REVEAL-ND NPM1 is a phase 3, multicenter, randomized, double-blind, placebo-controlled trial (NCT07211958). Eligible patients are ≥12 years of age, weigh ≥40 kg, and have treatment-naive ND AML with locally determined (centrally confirmed) NPM1 m. Patients will be randomized 1:1 to revumenib or placebo in combination with IC. Induction consists of 1 to 2 cycles of revumenib or placebo (dosed orally) alongside IC with cytarabine and daunorubicin or idarubicin (dosed intravenously). Consolidation consists of 1 to 3 cycles of revumenib or placebo plus cytarabine. HSCT may be performed after initial induction or consolidation. Treatment with revumenib or placebo monotherapy will continue for up to 2 years, with long-term follow-up until death, withdrawal of consent, or study closure. The dual primary endpoints are event-free survival and measurable residual disease-negative complete remission (CR) in the bone marrow, both assessed by independent reviewers. A key secondary endpoint is overall survival. Additional investigator-assessed endpoints include CR rate, composite complete remission (CRc) rate, overall response rate, duration of response (CR, CRc), safety, and quality of life. Overall, ~468 patients will be enrolled. As of January 27, 2026, the study is open to enrollment. Clinical trial information: NCT07211958 .

Acquired resistance to targeted, non-intensive therapies is common in myeloid malignancies. However, the kinetics of selection, the hematopoietic cell compartments where selection occurs, and the molecular mechanisms underlying selection remain open questions. To address this, we studied the kinetics of clonal and transcriptional responses to ivosidenib + venetoclax ± azacitidine combination therapy across hematopoiesis in 8 patients with IDH1-mutant myeloid malignancy. All 8 patients initially responded to treatment but 6 relapsed while 2 remained in sustained remission for >4 years. We performed combined high-sensitivity single-cell (sc) genotyping and scRNA-seq in index-sorted sequential patient samples. In all patients, clonal selection occurred rapidly, within 1-3 treatment cycles. Clonal selection preceded treatment failure by months to years. Relapse was associated with expansion of either clones harboring newly-detected myeloid driver mutations or pre-existing minor clones that underwent differentiation delay upon treatment exposure. In both cases, clonal selection occurred within immature cell populations previously shown to contain leukemic stem cell (LSC) potential. Different genetic alterations within relapse-associated clones converged onto common upregulated transcriptional programs of stemness, branched-chain amino acid catabolism, and genes sensitive to menin inhibition. Importantly, this relapse-associated transcriptional signature was selected within 3 cycles of therapy. In contrast, in both patients remaining in remission, leukemic clones were rapidly eradicated and replaced by clonal and wild-type hematopoiesis. Overall, in patients treated with ivosidenib combination therapy, rapid clonal selection occurs within the first treatment cycles. In those patients destined to relapse, genetically heterogeneous resistant clones are characterized by common transcriptional programs.

Abstract Relapse and chemoresistance remain major challenges in paediatric acute myeloid leukaemia (PAML), particularly in KMT2A-rearranged (KMT2A-r) subtypes where conventional markers such as CD34 are often absent, complicating measurable residual disease (MRD) detection. Leukaemia stem/regenerating cells (LSC/LRC) drive disease initiation, progression, and relapse, sharing stemness and chemoresistance properties that make them critical therapeutic targets. Using high-dimensional spectral flow cytometry, we identified CD180, a Toll-like receptor-like surface protein, as highly expressed on blasts and stem-like populations in KMT2A-r AML, while near absent on normal haematopoietic stem cells (HSCs). PAML KMT2A-r exhibits an unconventional immunophenotype dominated by CD34 ⁻ CD180 ⁺ populations. Integrated single-cell transcriptomics and functional profiling revealed CD180 high clusters enriched for quiescence, oxidative phosphorylation, and KMT2A/LSC stemness signatures. CD180 ⁺ cells demonstrated robust leukaemia-initiating capacity in xenograft models and persisted through therapy, re-emerging at relapse with phenotypic plasticity. Epigenomic analysis showed CD180 is a direct transcriptional target of the KMT2A::MLLT3 fusion complex, regulated by intragenic enhancers and downregulated by menin and BET inhibitors. Longitudinal single-cell analysis confirmed persistence and clonal evolution of CD180 ⁺ populations during treatment and relapse, underscoring their mechanistic role in chemoresistance and disease progression. In summary, CD180 marks dynamic, relapse-driving populations in KMT2A-r PAML, persists through therapy, and importantly is near absent on normal HSCs, offering a selective therapeutic window. These findings position CD180 as a clinically actionable biomarker for MRD detection and a compelling therapeutic target for eradicating chemoresistant, stem-like cells in paediatric AML. Main Points CD180 marks chemoresistant, relapse-driving stem-like blasts in KMT2A-r paediatric AML, overcoming CD34-based MRD limitations. Absent on normal HSCs, CD180 is a KMT2A::MLLT3 target and actionable for MRD, relapse prediction, and CD180-directed therapies. Novelty This study introduces CD180 as a novel biomarker and therapeutic target in AML, particularly KMT2A-rearranged subtypes where conventional markers are often absent. Unlike MRD strategies focused on bulk blasts, CD180 marks chemoresistant, stem-like populations driving relapse, critical reservoirs poorly defined in paediatric AML. This work fills a major gap in prognostic assessment and therapy by enabling precise detection of relapse-driving cells and offering a selective therapeutic window.

Hypomethylating agents (HMAs) are a mainstay of therapy for myeloid cancers, but genetic biomarkers do not predict who will respond to treatment. Using a variety of single-cell sequencing approaches to define the epigenomic state of responder and nonresponder leukemic cells, we demonstrate that leukemic stem cells (LSC) exist in at least two different epigenomic states: a hematopoietic stem cell (HSC)-or multipotent progenitor (MPP)-like state that is sensitive to HMAs, independent of genetic mutations, or a lymphoid-primed MPP (LMPP)-like nonresponder state. Hypomethylation and chromatin accessibility at ZNF143- and CTCF-binding sites results in activation of HOXB4, which defines the HSC/MPP-like state and HMA-sensitivity. Our study provides evidence that the epigenomic state of the LSC is a major determinant of response to HMAs, and demonstrates that a routine clinical assay can identify patients who will respond.

Bleximenib, a potent and selective menin inhibitor, in combination with intensive chemotherapy (IC) has previously exhibited an acceptable safety profile and efficacy in participants (pts) with newly diagnosed (ND) NPM1-mutated (NPM1m) or KMT2A-rearranged (KMT2Ar) acute myeloid leukemia (AML). We report updated safety and efficacy data (cut-off: July 2025) from this study.In the ALE1002 Phase 1b, multicenter, dose-finding study (NCT05453903), pts received a ‘7+3’ regimen of cytarabine 200 mg/m2/day and daunorubicin 60 mg/m2/day or idarubicin 12 mg/m2/day in combination with bleximenib 30–100 mg twice daily (from Day 4 of induction, including during count recovery). Pts who achieved a complete remission (CR) received consolidation therapy with up to 4 cycles of intermediate-dose cytarabine plus bleximenib. Those not proceeding to allogeneic hematopoietic stem cell transplant could continue bleximenib for up to 12 months. The safety dataset included all pts who received bleximenib 30–100 mg, and the intention-to-treat (ITT) efficacy dataset included those who received bleximenib 100 mg. Investigators assessed response criteria according to European LeukemiaNet (ELN) recommendations.The safety analysis set included 44 pts with ND AML (median age, 57.0 years [range, 19–71]; 52.3% female; 56.8% NPM1m, 43.2% KMT2A; 15.9% FLT3 co-mutations; ELN risk classification: 44.2% favorable, 27.9% intermediate, 27.9% adverse). The median duration of follow-up was 6.3 months (range, 1.31–22.51). All pts had ≥1 treatment-emergent adverse event (TEAE, all grades), the most common being thrombocytopenia (35/44; 79.5%), neutropenia (32/44; 72.7%), diarrhea (31/44; 70.5%), nausea (30/44; 68.2%), anemia, and febrile neutropenia (both 28/44; 63.6%). Most cytopenia TEAEs were Grade 3/4, consistent with an IC backbone. The 30- and 60-day mortality was 0/44 (0%) and 1/44 (2.3%), respectively. No differentiation syndrome was observed. Three TEAEs of QT prolongation were reported, all of which were Grade 1/2 and resolved without bleximenib interruption. Of 24 pts (NPM1m, n=15; KMT2Ar, n=9) in the ITT efficacy dataset, overall response rate (≥partial response) was 95.8%, composite CR (CR + CR with partial hematologic recovery [CRh] + CR with incomplete hematologic recovery rate) was 87.5%, and CR/CRh was 75%. Responses were similar across mutational subtypes. Median (range) time to CR was 28 days (21–36), similar to the median time to first response. Median duration of response was not reached. Among 37 pts achieving composite CR the median (range) time from Day 1 of induction to platelet count recovery was 32.0 days (22.0–82.0), and the median time to neutrophil count recovery was 30.0 days (21.0–71.0).In ND NPM1m or KMT2Ar AML, the safety profile of bleximenib + ‘7+3’ was consistent with a ‘7+3’ IC backbone, which, combined with promising early efficacy data, supports the planned Phase 3 study.

TPS6600 Background: Optimal treatment for patients (pts) with acute myeloid leukemia (AML) unfit for intensive chemotherapy (IC) remains challenging and depends on pt and disease characteristics and pt preference. In AML with a nucleophosmin-1 mutation ( NPM1 m) or lysine methyltransferase 2A ( KMT2A ) rearrangement ( KMT2A r), menin-KMT2A fusion proteins upregulate HOX/MEIS gene expression, resulting in hematopoietic differentiation arrest and leukemogenesis. Revumenib is a first-in-class, oral, potent, and selective inhibitor of the menin-KMT2A interaction. In the phase 1/2 AUGMENT-101 study (NCT04065399), revumenib monotherapy demonstrated clinically meaningful response rates and was generally well tolerated in pts with relapsed/refractory NPM1 m AML or KMT2A r acute leukemia. To further improve outcomes, combination regimens are being studied as front-line therapy for newly diagnosed (ND) pts. In the phase 1b Beat AML study (NCT03013998), revumenib plus venetoclax/azacitidine (VEN/AZA) demonstrated deep responses in pts ≥60 years (y) of age with ND NPM1 m or KMT2A r AML. EVOLVE-2 will assess whether revumenib plus VEN/AZA prolongs overall survival (OS) and improves complete remission (CR) rates in pts with ND NPM1 m or KMT2A r AML ineligible for IC. Methods: EVOLVE-2 is a phase 3, multicenter, randomized, double-blind, placebo-controlled trial (NCT06652438/EU-CT 2024-512733-32-00). Eligible pts are ≥75 y of age (18–74 y with comorbidities), have ND AML with centrally confirmed NPM1 m or KMT2A r (excluding KMT2A partial tandem duplications/deletions), are ineligible for IC, and are AML treatment-naive. Pts will be randomized 1:1 to revumenib or placebo in combination with VEN/AZA and stratified by age (<75 vs ≥75 y), genotype ( NPM1 m vs KMT2A r), and region (Europe vs Australia vs United States). In Cycle (C) 1, VEN is given daily on Days (D) 1–28 and AZA daily on D1–7, ± revumenib twice daily on D1–28. In C2+, responders (per 2022 European LeukemiaNet criteria) may adjust VEN based on remission status/count recovery. Treatment continues until disease progression, unacceptable toxicity, pt withdrawal, or death. The dual primary endpoints are OS and CR rate. Key secondary endpoints include event-free survival and rate of CR/CR with partial hematologic recovery (CRh). Other endpoints include rates of CRh and CR/CR with incomplete hematologic recovery (CRi), measurable residual disease (MRD) negativity (CR, CR/CRh, CR/CRi with MRD negativity), time to/duration of response, adverse events, time to hematopoietic recovery, and incidence of platelet and red blood cell transfusions. Overall, 448 pts will be enrolled. As of January 27, 2026, the study is open to enrollment. Clinical trial information: NCT06652438 .

ABSTRACT:Aberrant enhancer usage is a defining feature of oncogenic transcriptional reprogramming. Therapeutic strategies that disrupt enhancer-driven gene regulation may offer new treatment avenues. MYB is a key hematopoietic transcription factor that is frequently dysregulated in a broad range of cancers and plays a critical role in sustaining malignant cell states, including in aggressive leukemia subtypes such as KMT2A-rearranged leukemias. The molecular mechanisms by which it maintains oncogenic transcription remain incompletely understood. Here, we investigate the role of MYB in directing pathological enhancer activity to drive oncogene expression in leukemia. Using high-resolution Micro Capture-C, we show that upon MYB degradation, highly defined enhancer-promoter interactions at MYB binding sites are lost, correlating with the significant downregulation of target gene expression. When anchored to a gene desert region, the Myb transactivation domain (MybTA) is sufficient and necessary for the nucleation of an enhancer-like region. Critically, long-range chromatin interactions are established up to 400 kb away from where MybTA is anchored. This results in the activation of transcription from distal cryptic elements, which is reduced or abolished in the presence of point mutations that disrupt its interaction with the coactivators P300/CBP. All these results indicate that MYB activity alone is sufficient to generate an enhancer, inducing transcription through precise enhancer-promoter cross talk, and identify the MYB-P300/CBP axis as a therapeutically actionable vulnerability in enhancer-driven malignancies.

Inflammation activates blood cells, contributing to ageing and malignancy1-3. Haematopoietic stem cells (HSCs) survive a lifetime of infection to sustain life-long haematopoiesis1-9, but how human HSCs respond and adapt to inflammatory stress is largely unknown. Here, to empirically understand this adaptation, we developed xenograft inflammation-recovery models and performed single-cell multiomics on xenografted human HSCs. Two transcriptionally and epigenetically distinct HSC subsets were identified with one, termed HSC inflammatory memory (HSC-iM), retaining a molecular memory of previous inflammatory treatments. The HSC-iM subset exhibited quiescence and restrained haematopoietic output. Molecularly, the HSC-iM program was enriched in HSCs from adult and paediatric samples across conditions ranging from COVID-19 recovery, sickle cell disease, ageing and clonal haematopoiesis, establishing both the validity of our xenograft models and the physiological relevance of HSC-iM. Clonal haematopoiesis mutations in HSC-iM attenuated the effects of inflammatory stress by promoting HSC activation and differentiation. Moreover, transmission of the pro-inflammatory HSC-iM transcriptional program to differentiated immune progeny was demonstrated in xenograft and physiological settings. Finally, HSC-iM program enrichment in circulating blood cells was associated with a heightened risk score for all-cause mortality in population cohort analyses, underscoring the clinical relevance of this newly identified HSC subset in characterizing heterogeneous health outcomes across a lifetime.

Abstract TP53-mutated (TP53m) Acute myeloid leukemia (AML) represents a clinically intractable and biologically distinct disease. To study the molecular basis of this disease through hemopoiesis, we integrated single-cell RNA-seq (10x and long-read Oxford Nanopore sequencing) with open chromatin profiles (scATAC-seq), whole-genome (WGS) to capture structural variants and copy number alterations, and deep panel DNA sequencing from bone marrow samples taken from a cohort of 49 TP53m AML patients, 7 TP53 wild type (WT) AML patients, and 4 healthy donors. Phased genotyping plus WGS revealed biallelic loss to be associated with intrachromosomal breakage, whereas monoallelic loss favored numerical chromosomal alterations. TP53m clones pervaded stem/progenitors and myeloerythroid lineages but were proportionally depleted in mature lymphocytes. While co-occurring AML driver mutations influence the lineage biases of TP53m clones, we observed expansion of HSC/MPP, LMPP, and late erythroid compartments. In HSC/MPP dominant disease, TP53m cells displayed suppression of translational and mitochondrial-respiratory programs, with enrichment of a “p53-LSC” signature linked to chronic inflammatory stress. This suggests adaptation through metabolic quiescence, conferring persistence under inflammatory pressure. In LMPP-dominant expansion, we observed enriched MYC/E2F targets. Transcriptomic neighborhood analysis with pseudotime showed the erythroid compartment to be heterogeneous, with less mature basophilic cells enriched for TP53m clones, whereas more mature orthochromatic cells were enriched for TP53wt clones, nominating this transition as a tipping point for differentiation delay/arrest. Consistently, in erythroid dominant patients, late-stage erythroid cells demonstrated impaired GATA-1 and KLF1 (erythroid TFs) activities, and TP53m clones showed heightened GATA2, SPI1, and CEBPD regulon activity. TP53m versus TP53wt clones in erythroid differentiation showed heightened transcriptional activity of the EIF2AK1(HRI)→eIF2α mediated integrated stress response (ISR). Differential expression analysis within the aforementioned erythroid compartment showed ISR activation in TP53m clones; SESN2, DDIT4, and DDIT3/CHOP significantly up, with ATF3, and XBP1, up-trending, together with increased GATA2. Replication/mitochondrial-handling components (GINS2, RPA3, ABCB8) were significantly downregulated. These data together support a model in which deficient GATA-1/KLF1 function underpins the intra erythroid arrest of TP53m clones possibly due to activated HRI-ISR circuitry. These detailed molecular differentiation stage specific analyses of TP53m AML provide mechanistic insights and a platform for functional and synthetic lethal studies to specifically target TP53m leukemic cells. Citation Format: Felix A. Radtke, Bijay S. Jaiswal, Gonzalo Lopez, Junfei Zhao, Sagnik Banerjee, Daiane Hemerich Brennan, Yilin Zhao, Verena Körber, Marlen Metzner, Rachel Moore, Bilyana Stoilova, Batchimeg Usukhbayar, David Cruz Hernandez, Maria Ortiz Estevez, Aimee O’Donohue, Daniel Lopes de Menezes, Rajasekhar NVS Suragani, Anita K. Gandhi, Paresh Vyas. Dependent lineage regulatory programs in TP53-mutated acute myeloid leukemia revealed through deep single-cell multi-omic profiling of patient samples [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 7500.

FMS-like tyrosine kinase 3 (FLT3) mutations show variable detectability in relapse/refractory acute myeloid leukemia (AML) with unclear clonal evolution dynamics. This prospective noninterventional study examined clonal evolution and outcomes from AML diagnosis to relapse/refractory disease occurrences. Of 650 patients included, 172 were FLT3-positive (FLT3pos) and 472 were FLT3-negative (FLT3neg; 99.1

Abstract TP53-mutated (TP53m) AML is a distinct biological entity with adverse prognosis, characterized by prominent clonal heterogeneity; cells within a patient may exhibit wild type, monoallelic, or biallelic TP53 status, alongside co-occurring genomic alterations and transcriptomic heterogeneity. This complexity poses challenges in accurately aligning genotypes with transcriptomic and epigenomic profiles at single-cell resolution. To address this, we integrated single-cell RNA-seq (10x and long-read Oxford Nanopore, ONT) with open chromatin profiles (scATAC-seq) and whole-genome sequencing (WGS) to capture structural variants (SV), copy number alterations (CNV), and single nucleotide variants (SNV) from bone marrow samples taken from a cohort of 47 TP53m, 7 TP53 wild-type (wt) AML patients, and 4 healthy donors.Cell type annotation for scRNA-seq (335K cells) utilized a bone marrow reference dataset, while scATAC-seq (130K cells) was annotated using the ArchR label-transfer method. Cell type proportions were concordant across modalities (median correlation, ρ = 0.76). SV and CNV profiles showed that TP53m samples displayed frequent loss of Chr5q (72.5%), Chr17p (52.5%), and Chr7q (40%); chromothripsis was present in 16 (40%) samples. All these events were reported to occur in under 3% of TP53wt AMLs. In contrast, TP53m samples showed relative paucity of SNV co-mutations.We leveraged CNV profiles to map genetic gain/loss effect in scRNA and scATAC-seq compared to healthy donors, providing a clear separation of cells with abnormal CNV profiles. In TP53m AML, apart from mature lymphoid populations, >80% of cells including lymphoid progenitors (LMPP), displayed abnormal CNV status. Abnormal CNV cell fractions by sample strongly correlated with orthogonal tumor purity estimations from WGS for scRNA (ρ = 0.81) and ATAC-seq (ρ = 0.84).Single-cell SNV mapping was attempted with the standard 10x pipeline, where we detected driver hotspot mutations in only ∼3% of non-lymphoid cells. To improve mutation calling, we developed a new SNV pipeline using long-read ONT chemistry on full-length cDNA. SNV calling rates rose to 15% in non-lymphoid cells. Mature lymphoid cells showed low alteration rates: SNVs (1-2%), CNVs (5%), while expressing lineage-specific markers, confirming cell-type assignment. Long-read TP53 transcript sequencing also enabled phased variant genotyping. Combined with WGS SV and CNV profiles, we observed that TP53 loss-of-function dosage distinctly associates with the genomic architecture: biallelic loss of TP53 was associated with extensive intrachromosomal breaks and chromothripsis, whereas monoallelic loss primarily induced numerical chromosomal changes. This multi-omic framework enables high-resolution characterization of somatic genomic alterations at the single-cell level in TP53m AML, providing a powerful platform for mechanistic interrogation of disease biology. Citation Format: Gonzalo Lopez Garcia, Felix Andreas Radtke, Sagnik Banerjee, BIJAY JAISWAL, Daiane Hemerich Brennan, Yilin Zhao, Verena Körber, Marlen Metzner, Rachel Moore, Bilyana Stoilova, Junfei Zhao, Bettina Nadorp, David Cruz Hernandez, Batchimeg Usukhbayar, Aimee O’Donohue, Maria Ortiz Estevez, Daniel Lopes de Menezes, Rajasekhar NVS Suragani, Paresh Vyas, Anita Gandhi. High-resolution multi-omic dissection of bone marrow in TP53-mutant acute myeloid leukemia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 5938.

Children with Down syndrome (DS) frequently develop transient abnormal myelopoiesis that can evolve to the myeloid leukemia of DS (ML-DS). TAM spontaneously resolves in most cases but progresses to ML-DS with additional mutations, most commonly in the cohesin complex. However, the mechanisms by which these alterations promote leukemia are unknown. We leveraged the RAD21-mutant CMY cell line and RAD21-corrected CMY isogenic clones, as well as patient data, to investigate the effect of cohesin mutations during leukemia progression. RNA-sequencing revealed that HLA-class II genes were significantly down-regulated with cohesin mutations. Furthermore, HLA-DR was found to be lower in ML-DS relative to TAM, and these decreased levels were associated with increased risk of leukemia progression. Multi-omic analyses revealed that haploinsufficiency of RAD21 altered chromatin accessibility and impaired the occupancy of GATA1s and CIITA, the master regulator of HLA-class II gene expression. Chromatin binding of CIITA was increased with RAD21 correction, providing a mechanism by which restoration of cohesin improves HLA-class II expression. Finally, decreased levels of RAD21 or STAG2 expression in other subtypes of AML also exhibited reduced expression of HLA-class II genes. Thus, cohesin may contribute to leukemia by altering HLA-class II gene expression.

PURPOSE:To evaluate the efficacy and safety of the cluster of differentiation 47-targeted antibody magrolimab plus azacitidine (Magro/Aza) versus azacitidine alone in treatment-naïve patients with higher-risk myelodysplastic syndromes (MDS) in the phase III ENHANCE study (ClinicalTrials.gov identifier: NCT04313881). METHODS:Based on the Revised International Prognostic Scoring System, patients with intermediate- to very-high-risk MDS were randomly assigned to receive Magro (1 mg/kg on days [D]1 and 4; 15 mg/kg on D8; 30 mg/kg on D11 and D15, and then once per week for five doses, followed by 30 mg/kg maintenance doses once every 2 weeks)/Aza (75 mg/m2 daily on D1-7 or on D1-5 and 8-9 in 28-day cycles) or matched placebo plus azacitidine (Placebo/Aza). Dual primary end points were complete remission (CR) rate (per 2006 International Working Group criteria) and overall survival (OS). RESULTS:At final analysis, 539 patients were randomly assigned to Magro/Aza (n = 268) or Placebo/Aza (n = 271) arms. Baseline characteristics were generally well balanced between treatment arms. In the Magro/Aza versus Placebo/Aza arms, the CR rate was 21.3% versus 23.6% (odds ratio, 0.876 [95% CI, 0.585 to 1.312]; P = .5218), and median OS was 15.9 versus 18.6 months (hazard ratio, 1.203 [95% CI, 0.947 to 1.528]; P = .1299). Magro/Aza had a higher incidence of grade ≥3 adverse events (AEs; 92.8% v 79.2%), AE-associated study drug discontinuations (24.0% v 12.1%), serious AEs (71.9% v 51.5%), and fatal AEs (15.2% v 9.8%) versus Placebo/Aza. CONCLUSION:ENHANCE did not meet the primary end points of CR rate and OS, and showed more frequent severe AEs in patients treated in the Magro/Aza arm.

ABSTRACT:Patients with acute myeloid leukemia (AML) ineligible for intensive chemotherapy (IC) have limited treatment options. The phase 3 ENHANCE-3 study aimed to determine whether magrolimab (magrolimab arm) was superior to placebo (control arm) when either was combined with venetoclax and azacitidine. Adults with previously untreated AML who were ineligible for IC were randomized to receive magrolimab (1 mg/kg on days 1 and 4, 15 mg/kg on day 8, 30 mg/kg on days 11 and 15, then weekly for 5 weeks, and then every 2 weeks) or placebo, venetoclax (100 mg on day 1, 200 mg on day 2, and 400 mg daily thereafter), and azacitidine (75 mg/m2 days 1-7) in 28-day cycles. The primary end point was overall survival (OS); key secondary end points included complete remission (CR) rate and safety. After randomization of 378 patients, the trial was stopped at a prespecified interim analysis owing to futility. At final analysis, with median follow-up of 7.6 months (magrolimab arm) vs 7.4 months (control arm), median OS was 10.7 vs 14.1 months (hazard ratio, 1.178; 95% confidence interval, 0.848-1.637). The CR rate within 6 cycles was 41.3% vs 46.0%. Addition of magrolimab to venetoclax and azacitidine resulted in more fatal adverse events (19.0% vs 11.4%), primarily driven by grade 5 infections (11.1% vs 6.5%) and respiratory events (2.6% vs 0%). There were similar incidences of any-grade infections, febrile neutropenia, and neutropenia between arms. These results highlight the difficulty in improving outcomes for patients with AML who were ineligible for IC. This trial was registered at www.clinicaltrials.gov as #NCT05079230.

ABSTRACT:Myeloid leukemia of Down syndrome (DS) is preceded by a transient neonatal preleukemia driven by somatic mutations in the chromosome X gene GATA1, resulting in a shorter protein isoform (GATA1s). GATA1s mutations occur at high frequency in DS, but beyond trisomy 21, risk factors for this preleukemia are unknown. We investigated whether germline genetic variation influences development of GATA1s mutations in DS. Whole-genome sequencing was performed on 434 children with DS from the Oxford DS Cohort Study previously screened for GATA1s mutations. After quality control, association tests were conducted separately for disomic autosomes, trisomic chromosome 21, and chromosome X. Regression tests were performed for mutation variant allele frequency or the binary trait (103 GATA1s-positive cases, 326 controls), adjusting for sex and ancestry-related principal components. Genetic ancestry of each participant was inferred and tested for association with GATA1s mutations. We identified 3 genome-wide significant (P < 5 × 10-8) loci associated with GATA1s mutations. However, these may be false positives because few linked variants showed evidence of association at each locus. No significant associations were detected on chromosome 21 or the GATA1 region on chromosome X. Increasing proportions of South Asian genetic ancestry were associated with an increased risk of GATA1s mutations, with each 10% increase in ancestry associated with a 1.11-fold higher risk of developing GATA1s mutations (P = .031). Our genetic epidemiology study of somatic GATA1s mutations in DS did not identify strong germ line genetic effects. The association with genetic ancestry may relate to unmeasured genetic or nongenetic effects, such as fetal exposures, and warrants further investigation.

Mutations in IDH1 occur in 6–10% of patients (pts) with AML. Ivosidenib (IVO), an oral, targeted, small-molecule inhibitor of mIDH1, has been approved in combination with azacitidine (AZA) for pts with mIDH1 AML who are ineligible for IC, on the basis of results from the phase 3 AGILE study. AGILE demonstrated the long-term clinical benefit of IVO+AZA, with a median overall survival (OS) of 29 months. Here, we report preliminary results from ALIDHE (NCT05907057), an ongoing postapproval study designed in collaboration with the Acute Leukemia Advocates Network to evaluate safety and efficacy of IVO+AZA in a real-world setting inclusive of all pts with mIDH1 AML ineligible for IC managed as per local clinical practice. A pt steering committee ensures that pt needs, experiences, and perspectives are at the forefront of the study. Methods This international, single-arm, open-label phase 3b study enrolls adult pts (≥18 years) with newly diagnosed mIDH1 AML ineligible for IC and ECOG PS ≤2. Pts are treated with 500 mg oral IVO once daily and 75 mg/m2 SC or IV AZA for 7 days in 28-day cycles. Data are collected every 4 weeks while pts are on treatment for ≥112 weeks, and then every 12 weeks for OS. Primary endpoints include treatment-emergent adverse events (TEAEs), serious AEs (SAEs), AEs of special interest (AESIs), and AEs leading to IVO and/or AZA discontinuation, interruption, dose reduction, or death. Secondary endpoints include event-free survival, disease response, and time to response. Measurable residual disease (MRD) and biomarker correlations with clinical outcomes are exploratory endpoints under optional consent. MRD is assessed centrally by flow cytometry on fresh bone marrow (BM) aspirates. mIDH1 allele subtype and co-mutations are analyzed centrally on baseline peripheral blood samples using NGS. Results As of 5 May 2025 data cutoff, 92 pts from 7 countries (Europe + Canada) were enrolled; 89 were included in safety and efficacy analyses. Median age was 75 years (min-max: 51–84; Q1-Q3: 72–80); 66.3% of pts had ECOG PS 0–1 and 58.4% had de novo AML. Median baseline BM blast level was 39.8% (min-max: 0–100; Q1-Q3: 23.8–56.3; n=76). Median turnaround time for mIDH1 local testing was 7 days and median time from testing to study drug start was 20 days. Locally, direct molecular profiling methods were used in 73.0% of pts, including NGS (56.2%). In central assessment by NGS, distribution of the 5 most common mIDH1 variants (R132C, G, H, L, and S) was 51.1%, 7.6%, 18.5%, 3.3%, and 8.7%, respectively. Most frequent co-mutated genes included DNMT3A (40.7%), ASXL1 (28.8%), RUNX1 (28.8%), SRSF2 (27.1%), U2AF1 (20.3%), TET2 (15.3%), STAG2 (15.3%), CEBPA (10.2%), and NPM1 (10.2%). TP53 and JAK2 mutations occurred in 8.5% and 6.8% of pts, respectively; 25.4% of pts had mutations in the RTK/RAS pathway. At the cutoff date, 68 pts (76.4%) were still on treatment; median treatment duration was 5.1 months (min-max: 0.2–14.9; Q1-Q3: 2.9–7.8). Any-grade (G) TEAEs were reported in 93.3% of pts, G≥3 TEAEs in 80.9%, and SAEs in 57.3%. TEAEs occurring in >20% of pts were neutropenia (32.6%), nausea (29.2%), QT prolongation (23.6%), and anemia (20.2%). AESIs occurred in 22.5% of pts, including G≥3 QT prolongation in 9% and G≥2 differentiation syndrome in 13.5%. TEAEs led to any study drug discontinuation in 11.2% of pts, any dose reduction in 7.9%, and any study drug interruption in 75.2%. In total, 6 (6.7%) pts had TEAEs leading to death, none of which were considered treatment related. Preliminary efficacy showed CR in 37 (42%) pts, CR/CRh in 40 (45%), and an ORR of 52/89 (58%). As of 7 April 2025, MRD was assessed in 16 pts in CR/CRh/CRi after ≥6 cycles of IVO+AZA; 5 (31.3%) reached MRD negativity per flow cytometry, all (45.5%) among the 11 pts in CR at the time of assessment. Conclusions Outcomes in clinical practice can differ from those reported in pivotal trials. However, preliminary findings from ALIDHE are consistent with the AGILE pivotal trial. IVO+AZA had similar safety results in ALIDHE as in AGILE and efficacy outcomes were promising. The data will enrich knowledge on the IVO+AZA safety profile and help translate efficacy to effectiveness, bridging the gap between clinical trials and clinical practice, fulfilling the objectives of this real-world postapproval study. The study is still enrolling; future analyses will focus on effectiveness and QOL.

INTRODUCTION:Ivosidenib is an inhibitor of mutant isocitrate dehydrogenase 1 (mIDH1) that is approved alone or in combination with azacitidine in patients with mIDH1 acute myeloid leukemia (AML) that are ineligible to receive intensive chemotherapy. AIMS:Here we describe the design of ALIDHE, an international, multicenter, single-arm, open-label Phase 3b study of ivosidenib + azacitidine for treatment of newly diagnosed mIDH1 AML in usual clinical practice. METHODS:The primary endpoints are adverse events (AEs), clinical laboratory anomalies assessed as AEs, patients requiring transfusion and number of units transfused, and infection rate. The impact of ivosidenib + azacitidine treatment on health-related quality of life, healthcare resource utilization and measurable residual disease will also be assessed. CLINICAL TRIAL REGISTRATION NUMBER:NCT05907057 (ClinicalTrials.gov).

Supplementary protocol for "Phase Ib/II Investigator Initiated Study of the IDH1-mutant inhibitor ivosidenib (AG120) with the BCL2 inhibitor venetoclax +/- azacitidine in IDH1-mutated hematologic malignancies"

Studying the consequences of somatic mutations in pre-malignant and cancerous tissues is challenging due to noise in single-cell transcriptome data and difficulty in identifying the clonal identity of single cells. We optimized TARGET-seq to develop TARGET-seq+, which combines RNA sequencing (RNA-seq), the analysis of cell surface protein expression, and genotyping in single cells with improved sensitivity. We describe the steps for cell isolation, the preparation of single-cell RNA-seq (scRNA-seq) and genotyping libraries, and sequencing. We also provide guidance on the analysis of single-cell genotyping, transcriptome pre-processing, and data integration. For complete details on the use and execution of this protocol, please refer to Jakobsen et al.1.