He, Rong

ABSTRACT We examined the prognostic value of routine NGS data in newly diagnosed acute myeloid leukemia (AML) treated with intensive (7 + 3 backbone) induction, specified by AML subtype. A contemporary (2015–2025) series of 545 Mayo Clinic patients (median age 56 years, females 44%) was considered. Median follow‐up was 49 months with 341 (63%) allogeneic hematopoietic stem cell transplantations (AHSCT) recorded. AML subtypes included core‐binding factor (CBF; N = 72; 13%), primary non‐CBF ( N = 403; 74%), post‐myelodysplastic syndromes (MDS) or post‐myelodysplastic/myeloproliferative neoplasms (post‐MDS‐MDS/MPN; N = 28; 5%), post‐MPN ( N = 15; 3%), and therapy‐related (t‐AML; N = 27; 5%). Corresponding complete remission rates, with/without count recovery (CR/CRi), were 89%, 76%, 64%, 27%, and 78% ( p < 0.01) and 5‐year transplant‐censored survival rates 68%, 57%, 21%, 0%, and 55% ( p < 0.01). In multivariable analyses, the prognostic value of specific mutations was mostly limited to primary non‐CBF AML where adverse karyotype (OR 1.9; p = 0.04) predicted inferior and FLT3 ‐ITD (OR 0.4; p < 0.01) or NPM1 MUT / FLT3 WT (OR 0.1; p < 0.01) superior CR/CRi while KRAS MUT (HR 8.8; p < 0.01), TP53 MUT (HR 5.4; p < 0.01), and TET2 MUT (HR 2.3; p = 0.01) predicted inferior and NPM1 MUT / FLT3 WT (HR 0.3; p = 0.01) superior survival. Prognostication in intensively‐treated AML should start with subtype specification and recognition of the limited value of NGS in non‐primary AML. Post‐MPN AML is particularly associated with dismal outcomes and should be prognostically distinguished from post‐MDS‐MDS/MPN AML. In primary non‐CBF AML, in addition to previously established risk factors, the favorable impact of FLT3 ‐ITD on achieving CR/CRi and the unfavorable impact of KRAS MUT and TET2 MUT on transplant‐censored survival were noted and require confirmation from additional studies.

6588 Background: TET2 is the most frequently mutated gene in chronic myelomonocytic leukemia (CMML) and has been associated with favorable outcome, with recent studies suggesting a more pronounced prognostic impact in the presence of ≥2 TET2 mutations ( Csizmar et al. leukemia 2025;39:2030; Kynning et al. Br J Haematol. 2025 Nov 25. doi: 10.1111/bjh.70264 ). We sought to determine if this particular observation extended to patients with ≥3 TET2 mutations ( TET2 MUT ) and also held true in the context of recently reiterated genetic risk factors. Methods: The current study was conducted under institutional review board approved minimum risk protocols allowing retrospective patient data collection and analysis. Diagnostic criteria were according to the International Consensus Classification (Arber et al. Blood 2022. 140:1200). All statistical analyses were conducted using JMP 18 software. Results: 536 CMML patients who underwent CLIA-approved NGS testing were stratified according to the number of TET2 MUT : 221 (41%) wild-type, 148 (28%) with one, 153 (29%) 2, and 14 (2%) ≥3 TET2 MUT . Co-mutation frequencies with significant differences between one vs. two TET2 MUT included ASXL1 (46% vs. 29%; p<0.01), KRAS (7% vs. 14%; p=0.02) and SETBP1 (5% vs. 0.6%; p=0.02). Phenotypic comparisons revealed preponderance of prognostically favorable traits associated with wild-type TET2 but no significant differences between one vs. two TET2 MUT . In age-adjusted univariate analysis, transplant-censored overall survival (TCOS), calculated from the time of mutation detection, was superior in patients with one (p<0.01; HR 0.7) or 2 (p<0.01; HR 0.4) but not ≥3 (p=0.4) TET2 MUT , vs. those with wild-type TET2 . TCOS was also superior with 2 vs. 1 (p=0.02; HR 0.7) but not with ≥3 vs. 1 (p=0.7) TET2 MUT ; the results were similar when TCOS was calculated from the time of diagnosis. Multivariable analysis confirmed the survival advantage of exactly two TET2 MUT vs. wild-type TET2 (p<0.01; HR 0.5), one TET2 MUT (p<0.01; HR 0.6), or ≥3 TET2 MUT (p<0.01; HR 0.4), after adjusting for age, sex, anemia, circulating blasts ≥2%, and leukocyte count ≥13×10⁹/L. The favorable impact of exactly two TET2 MUT persisted in CMML-MD and CMML-MP and remained independent of TET2 VAF, mutation type, karyotype, and other prognostically relevant mutations. This association was sustained in the BLAST (p<0.01; HR 0.5) and CPSS-mol (p=0.02; HR 0.7) models, but not in BLAST-mol model (p=0.12). Conclusions: Mechanistic explanation for this novel observation includes the possibility that TET2 mutational categories based on mutation count reflect distinct biological states rather than having a simple linear effect. The persistence of this signal despite adjustment for established molecular and genetic risk factors supports its consideration in the development of future CMML prognostic models.

Background: Acute myeloid leukemia (AML) with CBFB::MYH11 fusion and myeloid/lymphoid neoplasms with eosinophilia and tyrosine kinase gene fusions (MLN-TK) are genetically defined and typically mutually exclusive entities. Case Presentation: We report a unique case of de novo AML harboring two clonal, transcriptionally active class-defining fusions: CBFB::MYH11 and GOLGA4::PDGFRB. A 61-year-old woman presented with leukocytosis with neutrophilia, eosinophilia, and monocytosis; circulating blasts; and a markedly hypercellular marrow. Cytogenetic analysis revealed inv(16)(p13.1q22) and t(3;5)(p21;q32) in all 20 metaphases, and RNA sequencing confirmed expression of both CBFB::MYH11 and GOLGA4::PDGFRB fusions. In addition, an oncogenic WT1 frameshift variant was identified. Hematopathologic findings were largely consistent with AML with CBFB::MYH11 fusion but exhibited features reminiscent of PDGFRB-rearranged MLN-TK. The patient achieved complete remission following the standard 7 + 3 induction chemotherapy regimen for AML with gemtuzumab ozogamicin. Conclusions: This case illustrates the diagnostic challenges posed by concomitant class-defining alterations in hematologic neoplasms and underscores the importance of integrated genomic assessment.

Abstract TP53 and PPM1D are key regulators of DNA damage response and repair, and somatic mutations in these genes often co-occur in hematopoietic cells, expanding under genotoxic stress. Unlike TP53 mutations, where mechanisms of progression are defined, pathways underlying clonal fitness and transformation in PPM1D mutant cells remain unclear. In collaboration with 5 academic institutions, we analyzed the clinical and molecular landscape of 337 patients with clonal hematopoiesis (CH) and clonal cytopenia of undetermined significance (CCUS) across 4 genotypes: PPM1Dmt/TP53wt (n = 170 [50%]), PPM1Dmt/TP53mt (n = 25 [7%]), TP53mt/PPM1Dwt (n = 17 [5%]), and TP53wt/PPM1Dwt (n = 125 [38%]). All PPM1D variants were truncating, located in exon 6 of the gene, with a median variant allele frequency (VAF) of 6% (range, 0.3%-64%). The PPM1Dmt/TP53mt genotype was most frequently encountered in therapy-related CH/CCUS (t-CH/t-CCUS; 80%, 66.5%, 76.5%, and 19%; P ≤ .001) and had a shorter time interval to detection from last genotoxic exposure (6.2, 5.9, 11.25, and 24.5 months; P ≤ .001) compared with PPM1Dmt/TP53wt, TP53mt/PPM1Dwt, and TP53wt/PPM1Dwt genotypes, respectively. Acknowledging the short follow-up duration, rates of malignant transformation were lower in the PPM1Dmt/TP53wt (2%) and PPM1Dmt/TP53mt (4%) groups compared with PPM1Dwt/TP53wt (12%) and PPM1Dwt/TP53mt (17%) groups (P ≤ .001), respectively. In summary, PPM1D mutations are frequently observed in t-CH/t-CCUS, with low median VAFs, and are associated with low rates of progression, even when comutated with TP53.

Abstract Introduction: CSF3R mutations (CSF3RMUT), most commonly T618I, represent the defining genomic alteration in chronic neutrophilic leukemia (CNL), present in ~80% of cases. Despite emerging real-world data, information on treatment outcomes and prognostic factors remains limited due to rarity of the disease. In this two-center retrospective study, we examined clinical and molecular predictors of treatment response, overall survival (OS), and blast transformation (BT) and outcome following allogeneic stem cell transplant (ASCT). Methods: The study was approved by the institutional review boards of the Mayo Clinic (USA) and the University of Montreal (Canada). Electronic medical records were reviewed to identify patients with CSF3R mutated CNL at both institutions and extract relevant data. Statistical analyses were conducted using JMP Pro 17.0.0 software (SAS Institute, Cary, NC, USA). Results: Forty-four patients (68% male, median age 68 years [range: 26-93]) were included. At diagnosis, 20 (56%) were asymptomatic, 13 (36%) presented with constitutional/non-specific symptoms, and 20 (50%) had antecedent leukocytosis at a median of 13 months(mo) prior. Splenomegaly (palpable or radiological) was observed in 28 (67%) patients, hepatomegaly in 7 (17%) and lymphadenopathy in 4 (9.8%). Median white blood cell count (WBC) was 52×10⁹/L (range: 17-390). CSF3RMUT/variants included 39 (89%) with T618I (7 of the 39 had additional CSF3RMUT), 2 (5%) with M696T, and 1 (2%) each for I598I, Q739* and T640N. Frequent co-mutations included ASXL1 in 25 (63%) patients, SETBP1 in 14 (36%), and SRSF2 in 8 (31%), with overlapping occurrence; 10 (38%) patients had ≥2 of these 3 mutations. Ten patients (median WBC 29 x10⁹/L [range: 19–58]), were initially observed without treatment for a median of 8.6 mo; 2 remained on observation alone at 46 and 77 mo. Hydroxyurea (HU) alone or in combinations excluding ruxolitinib (Ruxo) was the most common first-line treatment (68%). Second-line therapies included Ruxo (39%), HU (25%), and hypomethylating agents (14%). WBC response (≥50% reduction from baseline) was 78% with HU vs. 60% with Ruxo (p=0.2). WBC response to HU was lower with abnormal karyotype (40% vs. 94%; p=0.02). WBC response to Ruxo was lower with SETBP1MUT (0% vs. 72%; p=0.05) and with male sex (40% vs. 100%; p=0.04). Seven patients underwent ASCT, including 2 after BT; four (57%) of the 7 remain alive, including 2 without evidence of disease (both 45 mo post-ASCT; one harbored ASXL1MUT) and the other 2 with active disease (both harbored ASXL1MUT). Three patients died from veno-occlusive disease (Day+34), CNS relapse with AML (at 105 mo post-ASCT), or unrelated causes (at 44 mo post-ASCT). Median OS was longer in transplanted patients (115 vs 23 mo; p=0.03). At a median follow-up of 44 mo for living patients, disease transformation occurred in 10 patients, including 7 with BT (6 with T618I, 1 T640N) and 3 with chronic myelomonocytic leukemia (2 M696T, 1 T618I). BT rate at 2, 3, and 5 years was 15%, 31%, and 43%, respectively. Univariate analysis (UVA) identified male gender (p&lt;0.01), &gt;5% immature WBCs (p=0.02), platelets &lt;160×10⁹/L (p=0.04), higher WBC (p=0.02), and ASXL1 or SETBP1 mutation (p&lt;0.01) as risk factors for BT. On multivariable analysis (MVA), only the latter remained significant. Median OS was 26 mo (95% CI: 21-43) for the entire cohort; 24 mo for patients with T618I CSF3RMUT vs 53 mo with other CSF3RMUT, p=0.1. In UVA, age &gt;60 years (HR 5.0, p=0.03), platelets &lt;160×10⁹/L (HR 3.4, p=0.006), WBC &gt;50×10⁹/L (HR 2.6, p=0.02), and ASXL1 or SETBP1 mutations (HR 4.2, p=0.01) were associated with shorter OS. In MVA, age &gt;60 (HR 6, p=0.03), WBC &gt;50×10⁹/L (HR 12, p&lt;0.01), and the presence of ASXL1MUT or SETBP1MUT (HR 9.5, p=0.01) remained significant. Based on the HRs on MVA, one adverse point was allocated for age &gt;60 years, and two adverse points each for WBC &gt;50×10⁹/L and presence of ASXL1 or SETBP1 mutations. Patients were accordingly stratified into low (0-2 points), intermediate (3) and high (4-5) risk groups with corresponding median OS of “not reached”, 29 mo, and 16 mo (p&lt;0.01). Causes of death included disease progression (82%), infection/bleeding (9%), and ASCT complications (9%). Conclusion: The current study highlights the prognostic relevance of “SETBP1 or ASXL1” mutation for both overall survival and risk of blast transformation. The study also signifies variable natural history and potential benefit from ASCT.

Abstract Background: The primary goal of induction therapy in acute myeloid leukemia (AML) is to induce complete remission with (CR) or without (CRi) hematologic recovery. In order to accomplish this, re-induction might be necessary based on D14-residual AML. The current study examines the impact of re-induction on relapse, transplant-censored survival, and post-transplant survival in a consecutive series of AML patients who have achieved CR/CRi following intensive chemotherapy. The study also examined the impact of variations in consolidation chemotherapy including the use of anthracycline-based chemotherapy (AC) in the first consolidation cycle and the use of high (HiDAC) vs. intermediate (IDAC) dose cytarabine. Methods: The current study was conducted under IRB-approved minimum risk protocols that allowed retrospective collection and analysis of patient data. Formal criteria were used for AML diagnosis, genetic risk stratification, and adjudication of response (Blood 2022;140:1345). Patients who received less intensive induction regimens or failed to achieve CR/CRi were excluded. Relapse-free (RFS) and overall survival (OS) analysis was calculated from the date of documented CR/CRi. Patients who underwent allogeneic stem cell transplant (ASCT) were censored at the time of transplant. Conventional statistical methods were utilized (JMP Pro 17.1.0, SAS Institute, Cary, NC, USA). Results: A total of 279 AML patients diagnosed between 2004 and 2020 were included: median age 58 years, males 60%, de novo 75%, post-chronic myeloid neoplasm (post-CMN) 16%, and therapy-related 9%. Induction regimens included idarubicin (Ida) + cytarabine (Cyt) in 183 (66%) patients, daunorubicin (Da) 60 mg/m² + Cyt in 76 (27%), and Da 90 mg/m2 + Cyt in 19 (7%). Re-induction was utilized to achieve CR/CRi in 72 (27%) at a median of 18 days from start of first induction regimen. The indication for re-induction was based on residual bone marrow blast (median 30%; range 4-90) on day14 (N=53) or day 21-28 (N=15), persistent extramedullary disease (N=2) or the presence of circulating blast (N=2). At a median transplant censored follow-up of 26 months (mos), 93 (33%) deaths, 117 (42%) relapses, and 134 (48%) ASCTs were documented; ASCTs were performed in CR1 (50%) or CR2 (50%). Median RFS was 18 mos, with respective 1-, 2-, 3- year RFS of 63%, 42% and 35%. In multivariable analysis (MVA), age ≥65 years (HR 1.5, p=0.04), European LeukemiaNet (ELN) adverse karyotype (vs. intermediate HR 1.7, p=0.02; vs. favorable HR 2.8, p&lt;0.01), and the need for re-induction to achieve CR/CRi (HR 1.7, p=0.01) were independently predictive of relapse. RFS was not impacted by NPM1 mutation ( HR 0.5, p=0.09) or FLT3-ITD (HR 1.3, p=0.3). The median OS was 80 mos, with 5-year OS of 53%. In MVA, age ≥65 years (HR 2, p=0.01), ELN adverse karyotype (vs. intermediate HR 3.3, p&lt;0.01; vs. favorable HR 4.7, p&lt;0.01), FLT3-ITD (HR 1.9, p=0.04), and the need for re-induction (HR 1.9, p=0.03) independently predicted inferior OS. RFS was similar among patients who received AC (N=84; Idarubicin 9 mg/m2 or Da 60 mg/m2 days 1 to 2/3 + 100 mg/m2 Cyt days 1 to 5/7; median 18 mos) as their first consolidation cycle vs. those who received HiDAC (N=99; 3 g/m2 q12 hours on days 1,3 and 5; median 16 mos) or IDAC (N=58; IDAC; 1-2 g/m2 q12 hours on days 1, 3 and 5; median 24 mos; p=0.9); the corresponding median OS were 125, 85, and 40 mos (p=0.4). In patients aged ≥65 years, but not younger patients, both RFS (p&lt;0.01; median 23 (N=36) vs. 8 (N=24) mos) and OS (median 40 vs. 12 mos, p&lt;0.01) were significantly longer with IDAC vs. AC as their first consolidation cycle. In the AC consolidation cohort, patients who received 2-3 additional HiDAC/IDAC cycles displayed longer OS (p&lt;0.01) and RFS (p=0.047), compared to those who received less. In both the HiDAC and IDAC cohorts, RFS and OS were not significantly different between those who received 3-4 versus 1-2 consolidation cycles. The need for re-induction did not affect post-transplant survival (p=0.5), which was superior in patients receiving Ida + Cyt induction (vs. Da 60mg/m2 + Cyt, HR 0.6, p=0.04) and in those transplanted in CR1 (HR 0.5, p&lt;0.01). Conclusions: The current study signals the need for re-induction to achieve CR1 as a negative predictor of outcome in AML patients achieving CR/CRi following intensive induction chemotherapy. The particular information is potentially actionable in terms of the choice of consolidation strategy.

Abstract Background: The recently published BLAST clinical and BLAST molecular (BLAST-Mol) risk models for chronic myelomonocytic leukemia (CMML) incorporate clinical and molecular variables, respectively, for prediction of overall survival (OS), while BLAST-BT (blast transformation) risk model was predictive of BT free survival (BTFS) [Blood 2025;doi:]. In the current study, the Mayo Clinic (USA) CMML cohort used in the aformentioned study was expanded from 457 to 600 patients and merged with an external cohort of 501 patients from the Humanitas Research Hospital (HRH; Italy) in order to validate the BLAST, BLAST-Mol and BLAST-BT risk models. Methods: CMML diagnosis and sub-categorization were according to the International Consensus Classification, which included CMML-1 (BM/PB blasts &lt;10%/&lt;5%) vs. CMML-2 (BM/PB blasts ≥10%/≥5%) and CMML-MP (leukocytes ≥13 x 10⁹/L) vs. CMML-MD (leukocytes &lt;13 x 10⁹/L) [Blood 2022;140:1200]. The BLAST score is based on clinical risk factors including PB blasts ≥2%, leukocytes ≥13 x 109/L, and severe or moderate anemia resulting in BLAST low, intermediate, and high risk groups. BLAST-Mol also accounts for mutations and karyotype and identifies PHF6 and TET2 as “favorable” and DNMT3A, U2AF1, BCOR, SETBP1, ASXL1, NRAS, PTPN11, RUNX1, TP53, as “unfavorable” mutations with adverse karyotype also considered unfavorable. BLAST-BT variables include absence of PHF6MUT, BM blasts ≥10%, presence of DNMT3AMUT, PB blasts ≥2%, leukocyte count ≥13 x 109 /L, and ASXL1MUT. OS analyses were censored for allogeneic stem cell transplant (ASCT). Conventional statistical methods were employed, using JMP Pro 18.0.0 software SAS Institute, Cary, NC, USA. Results: A total of 1,101 patients (median age 73 years; 67% males) with newly-diagnosed CMML were considered: Mayo cohort 600 and HRH cohort 501. Compared to the HRH cohort, the Mayo cohort was more likely to include CMML-MP (44% vs. 28%; p&lt;0.01), CMML-1 (87% vs. 81%; p&lt;0.01), ASXL1MUT (48% vs. 35%; p&lt;0.01), SRSF2MUT (43% vs. 33%; p&lt;0.01), NRASMUT (15% vs 9%; p&lt;0.01), PHF6MUT (7% vs. 4%; p&lt;0.01), and less likely to harbor TET2MUT (50% vs. 58%; p&lt;0.01), and SF3B1MUT (4% vs. 10%; p&lt;0.01) [Table 1]. At a median follow-up of 55 months (range 0-202), 686 (62%) deaths, 218 (20%) BTs, and 142 (13%) ASCTs were documented. CPSS-mol risk categories at diagnosis included low (N=190; median OS 67 months), intermediate-1(N=271; median 38 months), intermediate-2 (N=407; median 26 months), and high (N=231; median 16 months; p&lt;0.01) [AUC at 5 years 0.75]. BLAST clinical risk distribution was low (N=356; median OS 65 months), intermediate (N=393; median 30 months), and high (N= 352; median 16 months; p&lt;0.01) [Figure 1; AUC at 5 years 0.79]. The corresponding AUCs calculated for the Mayo and HRH cohorts, separately, were 0.81 and 0.76, respectively. BLAST-Mol risk assignments included low (N=357; median OS 66 months), intermediate (N=512; median 27 months) and high (N=232; median 15 months; p&lt;0.01) [Figure 2; AUC at 5 years 0.79]. The corresponding AUCs for the Mayo and HRH cohorts were 0.81 and 0.77, respectively. The 3-year cumulative incidence of BT (accounting for death from another cause) stratified by BLAST-BT risk model included 8% for very low risk (N=54), 11% for low risk (N=332), 20% for intermediate-risk (N=281), 27% for high risk (N=250), and 34% for very high risk (N=176) [p&lt;0.01; Figure 3]. Multivariable analysis of the combined Mayo-Humanitas cohort (N=1,101) identified age &gt;65 years (HR 1.7, 95% CI 1.3-2.1), CMML-2 subtype (HR 2.0; 95% CI 1.6-2.5), and a number of other mutations or co-mutations with BLAST-Mol-independent and CPSS-mol-independent prognostic contribution (to be elaborated further at the time of meeting presentation). Conclusions: The current study validates the predictive performance of the BLAST, BLAST-Mol and BLAST-BT risk models in a large two-center international cohort of patients with CMML. The possibility of further enhancement of the BLAST-Mol model is suggested by the demonstration of independent prognostic relevance from additional mutations and co-mutations.

Abstract Background: TP53-mutated myeloid neoplasms (MNs) represent one of the most lethal subgroups of blood cancers. The World Health Organization (WHO-5) recognize TP53-mutated (TP53mut) myelodysplastic syndrome (MDS) and International Consensus Classification (ICC) recognize TP53mut MDS and acute myeloid leukaemia (AML) as distinct disease entities, acknowledging their dismal prognosis and the urgent need for novel therapeutic strategies. However, key discrepancies exist between the classifications, creating uncertainty in diagnosis, influencing therapeutic decisions, and complicating clinical trial enrolment for patients with uniformly poor survival. Importantly, neither classifications included TP53-mutated myelofibrosis (MF), other myeloproliferative neoplasms (MPN), and MDS/MPN overlap syndromes, despite growing evidence that these cases carry similar adverse outcomes. To address these critical gaps, we conducted a comprehensive analysis across the full spectrum of TP53-mutated MN. Methods: We included TP53mut cases with variant allele frequency (VAF ≥2%) of AML, MDS, MDS/MPN overlap, myelofibrosis and other MPN managed at the Mayo clinic (Rochester, USA) and Local Health Network in South Australia (Australia). Cases were classified according to the revised 4th edition WHO classification (WHO-4R). Biallelic TP53 loss was defined as: (i) ≥2 mutations with VAF ≥2%; (ii) single TP53mut with VAF ≥2% plus 17p loss on karyotype, FISH, or SNP array; or (iii) single TP53mut with VAF ≥50% with or without 17p loss. Monoallelic loss was defined as a single TP53mut with VAF &lt;50% without 17p loss or copy neutral loss of heterozygosity. The primary endpoint was overall survival (OS) from diagnosis, censored at allogeneic transplantation. Conditional inference tree analysis was performed to define the effect of individual predictors on OS. Results: We identified 853 TP53mutMN cases including WHO-4R-defined AML (BM/PB blasts ≥20%, n=314; 36.8%), MDS with excess blasts-2 (EB-2; BM 10–19% or PB 5–19%, n=129; 15.1%), MDS-EB-1 (BM 5–9% or PB 2–4%, n=100; 11.7%), and MDS with low blasts (LB; BM &lt;5% and PB &lt;1%, n=228; 26.7%). MF included chronic phase (MF-CP, n=48; 5.6%), accelerated phase (MF-AP, n=7; 0.8%), and blast phase (MF-BP, n=17; 2.0%). Additional cases comprised other MPN-chronic phase (n=3, 0.4%) and MDS/MPN overlap syndromes (n=7, 0.8%). Of 848 evaluable TP53mut MN, 616 (72.6%) harbored biallelic TP53 loss and 232 (27.4%) had monoallelic loss, with 42.4% (n=98) of monoallelic cases harboring a complex karyotype (CK). Biallelic loss was highly prevalent in AML (83.1%), MDS (70.4%), and MDS/MPN overlap (71.4%), but was less frequent in cases with MPN (33.3%) and MF-chronic phase (25.0%). Across the spectrum, the frequency of biallelic loss increased stepwise with disease progression, rising from MDS-LB (62.8%) to MDS-EB1 (76.0%), MDS-EB2 (79.5%), AML (83.1%), and MF-AP/BP (79.2%) (P&lt;0.0001). In multivariable analysis, WHO-4R category, TP53mut VAF, biallelic loss and CK were independent predictors of poor survival in TP53mut MN. Conditional inference tree analysis stratified the cohort into seven prognostic groups with distinct median OS: (1) AML, MF-BP/AP, and MDS/MPN overlap regardless of mono/biallelic and CK status (n=335, 39.7%; median OS 4.0 months); (2) MDS-EB1/EB2 with biallelic loss or monoallelic plus CK (n=212, 25.1%; 7.2 months); (3) MDS-LB, MF-CP and MPN with CK (n=156, 18.5%, OS 10.2 months); (4) biallelic MDS-LB (n=28, 3.3%; OS 15.6 months), (5) monoallelic MDS-EB1/EB2 without CK (n=20, 2.4%; OS 26.2 months), (6) monoallelic MDS-LB without CK (n=57, 6.7%; OS 33.4 months), and (7) MF-CP and other MPN without CK (n=36, 4.3%; OS 37.0 months). This analysis defines a clear stepwise risk gradient, identifying subsets with extremely poor prognosis versus a relatively indolent disease. Conclusions: This is the first study to evaluate the prognostic impact of TP53mut MN across the entire MN spectrum, providing a framework for future refinement of TP53-driven disease classification. Key findings include: (i) MF-AP/BP and MDS/MPN overlap exhibit survival outcomes as dismal as AML, irrespective of the TP53 allele status; and (ii) complex karyotype is a surrogate for “biallelic equivalent”, refining risk interpretation in monoallelic cases. Overall, our study defines seven prognostic subgroups spanning all TP53mut MN, offering a foundation for risk-adapted clinical trial design and future classification updates.

Abstract Introduction The European Leukemia Net (ELN) 2022 stratified acute myeloid leukemia (AML) patients (pts) treated with intensive chemotherapy (IC) into three groups based on genetic features. ELN 2024 (Döhner et al, Blood) and Beat AML 2024 (Hoff et al, Blood Advances) proposed revised systems to better capture the impact of genetic features in pts receiving less-intensive therapy (LIT). Real-world validation across treatment settings, including allogeneic hematopoietic stem cell transplantation (allo-HCT), is warranted. Methods We evaluated the performance of ELN 2022, ELN 2024, and Beat AML 2024 among AML pts treated at Mayo Clinic from 2016 to 2025. Only treated pts with available molecular data included. Response was assessed according to the ELN 2022 criteria. Overall survival (OS) was calculated from diagnosis. Multivariable Cox proportional hazard (CPH) models were used to adjust for covariates. Model performances were assessed using Harrell's c-index. Results 426 AML pts (median age 68 years; 60% male) were included; 195 (46%) received IC and 231 (54%) received LIT, while Allo-HCT was performed in 133 (31%) pts. Complex karyotype was present in 126 (30%) and normal cytogenetics in 143 (34%) pts. Molecular mutations included TP53 (20%), TET2 (18%), DNMT3A (16%), SRSF2 (16%), RUNX1 (16%), ASXL1 (16%), and NPM1 (15%). Among evaluable pts (83%), complete remission (CR) after first-line therapy was achieved in 47%. Risk group assignments varied: ELN 2022 vs. ELN 2024 classified pts as favorable (20% vs. 55%), intermediate (20% vs. 24%), and adverse (60% vs. 20%); Beat AML 2024 categorized pts as favorable (41%), intermediate (21%), and adverse (38%). ELN 2024 reclassified 229 (54%) pts of whom 5% upstaged and 49% downstaged, primarily affecting ELN 2022 adverse (67%) and intermediate (46%) groups. Among LIT-treated, CR rates did not differ across groups in any classification. Per ELN 2024, CR was achieved in 43% (favorable), 67% (intermediate), and 29% (adverse) of the LIT-treated pts. However, among IC-treated pts, CR was higher in favorable/intermediate vs. adverse groups across all models (p&lt;0.0001). Allo-HCT was most common in ELN 2024 intermediate (41%) and favorable (32%) vs. adverse (16%) groups. Median OS (months; 95% CI) for the entire cohort was 15.7 (12.5–20.9) with median follow-up time of 49.8 (40.9-55). By ELN 2022, OS of the entire cohort was 49.4 (30.7–NA) for favorable, 50.0 (29.2–NA) for intermediate, and 8.8 (7.7–11.2) for adverse (p&lt;0.0001). Per ELN 2024, OS of the entire cohort was 25.4 (18.5–37.3), 20.3 (13.4–39.8), and 5.7 (3.4–8.2), respectively (p&lt;0.0001). Beat AML 2024 stratified OS of the entire cohort as 42.5 (30.7–NA), 11.4 (9.3–16.7), and 5.6 (3.5–8.0) for favorable, intermediate, and adverse groups (p&lt;0.0001). ELN 2024 stratified the OS of the pts treated with LIT as 15.7 (10.1-24.9), 8.0 (4.9-13.9), and 6.8 (3.4-10.2) for favorable, intermediate and adverse risk groups, (p &lt;0.0001). Among ELN 2022 adverse-risk pts, those reclassified as favorable per ELN 2024 had superior OS (15.0, 9.8–22.7) compared to intermediate (7.8, 5.0–9.3) or adverse (5.4, 3.3–8.0) risk groups, p&lt;0.0001. C-index (95% CI) for OS in the entire cohort was highest for Beat AML 2024 (0.658, 0.629–0.688), followed by ELN 2022 (0.627, 0.599–0.655), and ELN 2024 (0.600, 0.568–0.631). In LIT-treated pts, ELN 2024 (0.597, 0.558–0.637) and Beat AML 2024 (0.586, 0.545–0.627) outperformed ELN 2022 (0.559, 0.521–0.596). Among allo-HCT pts, regardless of treatment type, ELN 2022 (0.610, 0.539–0.681) performed better than ELN 2024 (0.568, 0.487–0.648). In multivariable CPH models adjusted for age, bone marrow blasts, treatment intensity, and complex karyotype, ELN 2022 and ELN 2024 adverse-risk groups had higher hazard of death (HR 2.4, 1.6–3.5 and HR 1.7, 1.1–2.5, respectively, vs. favorable). No significant difference was observed between intermediate and favorable risk for ELN 2022 (HR 1.4, 0.9–2.2) or ELN 2024 (HR 1.4, 0.9–1.9). Allo-HCT was associated with a lower hazard of death (HR 0.4, 0.3–0.5). Conclusions In this real-world AML cohort, ELN 2024 and Beat AML 2024 improved OS prediction in LIT-treated pts compared to ELN 2022. This advantage was not observed in allo-HCT pts. Multivariable analysis confirmed the adverse-risk group as having the worst prognosis across all models. These findings highlight the importance of aligning molecular risk stratification with treatment modalities to enhance prognostication in AML.