Michael R. Savona

CD8 T cells are key mediators of anti-tumor immunity, yet their differentiation states and clonal architecture within the acute myeloid leukemia (AML) bone marrow microenvironment remain incompletely defined. We performed high dimensional immune profiling of bone marrow samples from 46 newly diagnosed AML patients using mass cytometry to probe the microenvironment and T cell states. We used single-cell RNA and T cell receptor (TCR) sequencing to define CD8 T cell phenotypes, transcriptional states, and clonal architecture. AML-associated CD8 T cells were skewed toward terminally differentiated effector memory cells re-expressing CD45RA (TEMRAs), while canonically exhausted T cell populations were rare. Functional studies revealed that TEMRA CD8 T cells robustly produced IFNγ and TNFα upon in vitro stimulation. Clonal expansion was concentrated within the TEMRA compartment. Comparison of TCR sequences with known viral epitopes identified a small subset of expanded viral-specific clonotypes, while the majority of highly expanded clonotypes had unknown specificity and may include AML-specific T cells. Differential gene expression analysis revealed shared cytotoxic effector programs during clonal expansion but diverging activation signatures: viral-specific clones upregulated canonical T cell receptor signaling genes, whereas expanded clones of unknown specificity exhibited enrichment of natural killer (NK) receptor-associated genes. These genes comprised the leukemia-associated NK-immune (LaNKi) score which was elevated in an independent AML dataset compared to pan cancer tumor infiltrating lymphocytes. These findings define the CD8 T cell differentiation landscape and clonal architecture in AML marrow at diagnosis and suggest that T cell dysfunction in AML diverges from classical activation and exhaustion paradigms. Key points: CD8 T cells in AML are enriched for terminally differentiated states not defined by traditional checkpoint-inhibitor expression. The LaNKi score marks clonally expanded CD8 T cells of unknown specificity and is validated across independent AML and pan-cancer datasets.

From the 2026 National Summit on Hematologic Cancers

From the 2026 National Summit on Hematologic Cancers

From the 2026 National Summit on Hematologic Cancers

Baseline IPSS‐M risk, response to hypomethylating agent (HMA) therapy, and receipt of allogeneic stem cell transplant (allo‐HCT) have all been individually shown to impact overall survival (OS) in patients with myelodysplastic syndromes (MDS). However, the prognostic impact of response when adjusting for IPSS‐M risk and treatment strategy remains unclear. Hence, we used the VALIDATE database of the International Consortium for MDS (icMDS) to evaluate the impact of International Working Group (IWG) 2023 best response on OS in 715 HMA‐treated, higher‐risk MDS patients stratified by baseline IPSS‐M risk and their treatment strategy (subsequent allo‐HCT vs. medical therapy alone) treating both best response and allo‐HCT as time‐dependent variables. Baseline IPSS‐M risk (hazard ratio (HR): 0.5, p < 0.001) and receipt of allo‐HCT (HR: 0.5, p < 0.001) were the strongest independent predictors of OS, whereas achievement of composite complete response (cCR) had a more modest impact on OS (HR: 0.8, p = 0.004). Among patients treated with medical therapy alone, achieving cCR improved OS significantly (HR: 0.7, p = 0.006). In contrast, among transplanted patients, cCR did not retain independent prognostic value for post‐transplant survival after adjusting for baseline IPSS‐M (HR: 0.9, p = 0.634). Achieving cCR did not fully overcome adverse disease biology as OS continued to segregate according to baseline IPSS‐M risk. In summary, achieving cCR improves outcomes in non‐transplanted patients, but it does not significantly impact post‐transplant OS, suggesting that failure to achieve cCR with HMA may not warrant delay or preclude allo‐HCT. Clinical trials should consider response in the context of IPSS‐M risk distribution and treatment strategy (subsequent allo‐HCT vs. medical therapy alone) to avoid overinterpretation of high response rates.

Targeting oxidative phosphorylation (OXPHOS) represents an attractive therapeutic strategy in acute myeloid leukemia, which exhibits exceptional dependence on mitochondrial respiration compared to normal hematopoietic cells. However, clinical attempts to exploit this vulnerability have been limited by on-target toxicity to healthy tissue. Here, we comprehensively compare the cellular consequences of inhibiting distinct nodes of the electron transport chain in AML. We demonstrate that selective inhibition of the F ₁ subunit of ATP synthase with EB2023 (ammocidin A) delivers an energetic stress to AML cells without the profound redox stress that characterizes complex I inhibition, preventing NAD⁺/NADH imbalance and allowing continued TCA cycling. Further, the duration of OXPHOS inhibition is transient in nature in vivo , a finding revealed through pharmacokinetic and serial pharmacodynamic monitoring of AMPK phosphorylation accompanied by OPA1-mediated mitochondrial structural remodeling that primes AML cells for BCL2 inhibitor synergy. EB2023 in combination with venetoclax demonstrates potent anti-AML activity across cell lines and patient-derived xenograft models at doses that spare normal hematopoietic progenitors and avoid the neuropathy and sustained detrimental systemic metabolic rewiring in healthy tissues associated with prior efforts to target OXPHOS. These findings establish F ₁ -selective ATP synthase inhibition as a clinically actionable therapeutic strategy in AML and establish the duration of OXPHOS inhibition as a critical and previously underappreciated determinant of therapeutic index.

PURPOSE: Chronic myelomonocytic leukemia (CMML) is a rare myeloid neoplasm characterized by clinical heterogeneity and is associated with poor outcomes. To date, limited molecular information has been incorporated into disease classification and risk stratification. We aimed to integrate genomic features into the clinical decision-making process for CMML. PATIENTS AND METHODS: We analyzed a retrospective cohort of 3013 patients with CMML (training set) and a prospective population of 516 patients (validation set). Using an innovative framework for multimodal data analysis, we developed molecular-based disease taxonomy and prognostication. RESULTS: Unsupervised clustering identified nine entities with distinct genomic features and outcomes ( P < .001), including splicing machinery, transcription factors, signal transduction and tyrosine kinase pathways aberrations, and high-risk molecular signatures. Notably, 15% of patients showed molecular/clinical overlap with other myeloid neoplasms. We integrated molecular and clinical information to build the international CMML Prognostic Scoring System (iCPSS), incorporating mutations in nine genes together with hematologic parameters and cytogenetic abnormalities. The iCPSS identified five groups with distinct probability of overall and leukemia-free survival in both training and validation cohorts ( P < .001), outperforming existing prognostic models. Importantly, 55% of patients were reassigned to higher or lower risk groups by the iCPSS. Decision analysis demonstrated that iCPSS could refine the optimal timing of allogeneic transplantation at the individual level; compared with conventional prognostic tools, iCPSS-based decision modeling changed transplantation strategy in 31% of cases, resulting in a significant gain-in-life expectancy for eligible patient population ( P < .001). A federated learning platform was implemented to enable continuous, privacy-preserving model update across multiple centers. CONCLUSION: Molecular information improves CMML classification and prognostication, supports more effective clinical decision making, and potentially refines the design of clinical trials.

Clonal hematopoiesis of indeterminate potential (CHIP) driven by somatic mutations in TET2 and DNMT3A is present in >10% of adults over 60 and confers substantial risk for hematologic malignancy and cardiovascular disease, yet the majority of patients with CHIP do not progress to disease. Analyzing 1,020,538 individuals across three biobanks (UK Biobank, All of Us, BioVU), we show that a discrete subset of enzymatically disruptive mutations — TET2 loss-of-function variants and the DNMT3A R882 hotspot — account for the majority of clinical risk in these genes and exhibit the strongest clonal fitness advantage. Because DNMT3A and TET2 encode enzymes that modulate DNA methylation, we reasoned that peripheral blood methylation patterns should reflect the functional impact of individual mutations, enabling a direct readout of enzymatic dysfunction in CHIP patients. We developed and validated methylation-based activity scores for TET2 and DNMT3A as patient specific biomarkers that quantify enzymatic activity. These scores capture functional heterogeneity across mutation subtypes, predict disease risk comparably to clinical risk scores such as the Clonal Hematopoiesis Risk Score and the AHA PREVENT cardiovascular risk model. Integrating the activity score with the clinical models substantially improves prediction of incident cytopenia, myeloid neoplasm, and major adverse cardiovascular events. These findings establish that TET2 and DNMT3A CHIP pathogenicity is proportional to the degree of enzymatic disruption conferred by specific variants, and nominate methylation-based activity scores as a functional biomarker for individualized CHIP risk stratification and monitoring therapeutic response.

Spirocyclic β-lactones (SβLs) are ring-strain activated natural products produced by actinomycetota, possessing potent and selective cytotoxicity, and with an unknown pharmacological mechanism. To further the understanding of their mechanism of action and the discovery of new SβLs, we obtained a series of known and previously not isolated SβLs from Streptomyces platensis , which contained a cryptic SβL-producing gene cluster. Utilizing single cell Multiplexed Activity Metabolomics to assess a panel of regulated cell injury and cell death markers in metabolomic arrays tested against MV-4-11 cells, phosphoprotein-S6 phosphorylation inhibition was observed for several metabolomic features, prompting us to prioritize these metabolites for structure elucidation. Several compounds were isolated including one new variant, permitting correlation of a cryptic biosynthetic gene cluster entry for oxazolomycin D (OxD) family metabolites. Cytotoxicity and flow cytometric structure–activity relationships for isolated analogs were determined using an expanded phospho-flow metabolism marker panel suggesting that SβLs possess two pharmacophores: a mixed polyketide component, involved in target engagement and inhibition, and a nonribosomal peptide synthetase-derived covalent β-lactone warhead, contributing to cytotoxicity. Global cellular thermal proteome profiling analysis of OxD treated cells implicated potent interaction with and modulation of spliceosome associated proteins. Spliceosome-associated functional changes were validated by alternate splicing analysis upon induction by OxD. As aberrant RNA splicing occurs in nearly all cancer types, the spliceosome-associated cellular response profiles of OxD define an unexplored opportunity for therapeutic development in cancers dependent upon alternative splicing.