Mangaonkar, Abhishek A.

Abstract Allogeneic hematopoietic stem cell transplant (alloHCT) is considered for all patients with myeloid neoplasms (MNs) harboring TP53 mutations (TP53mut). The aim of this international study across 7 transplant centers in the United States and Australia was to identify factors associated with improved post-alloHCT survival. Of 134 TP53mut MN cases who underwent alloHCT, 80% harbored complex karyotype; 94% of TP53 variants were localized to the DNA-binding domain (DBD). Most common comutations were ASXL1 (7%), TET2 (7%), and DNMT3A (6%). Median post-HCT survival was 1.03 years, and overall survival (OS) at 1 year, 2 years, and 3 years was 51.4%, 35.1%, and 25.1%, respectively. A total of 103 cases (76.9%) met the International Consensus Classification (ICC) criteria for MN with mutated TP53 (referred to as ICC-defined TP53mut MN hereafter). The 3-year OS of ICC-defined TP53mut was significantly shorter compared with that of other TP53mut MNs (3-year OS, 16.9% vs 54.9%; P = .002). ICC-defined TP53mut MNs was independently associated with inferior OS (hazard ratio [HR], 2.62; P = .019). The presence of non-DBD TP53mut only (HR, 3.40; P = .005), DNMT3A comutation (HR, 2.64; P = .016), and pre-alloHCT bone marrow blasts ≥5% (HR, 2.76; P = .006) was independently associated with inferior relapse-free survival (RFS), whereas melphalan-based conditioning was associated with superior RFS (HR, 0.52; P = .005). Combining these variables, we constructed a hierarchical model that stratified patients into low-, intermediate-, and high-risk categories with 1-year RFS of 81.3%, 31.3%, and 6.7%, respectively (P < .001). In conclusion, a subset of MN harboring TP53mut who have low blasts pre-alloHCT and received melphalan-based conditioning derived long-term benefit from alloHCT.

Abstract This retrospective analysis aimed to provide evidence-based risk stratification of TP53-mutated (TP53mut) myeloid neoplasms (MNs). Of 580 MNs harboring TP53mut with variant allele frequency (VAF) ≥2%, 219 (37.8%), 194 (33.4%), 92 (15.9%), and 75 (12.9%) were classified as acute myeloid leukemia (AML), myelodysplastic syndrome (MDS) with low blasts (MDS-LB), MDS with excess blasts (MDS-EB)-2, and -EB1 according to the revised fourth edition of the World Health Organization (WHO) classification, respectively. Hierarchical analysis identified the following 4 risk groups with distinct survival: (1) MDS-LB, (2) MDS-EB1/EB2/AML VAF <10%, (3) MDS-EB1/EB2 VAF ≥10%, and (4) AML VAF ≥10%. We next evaluated the impact of allelic status, VAF, and complex karyotype (CK). In our cohort, the significance of biallelic status was limited to MDS with <5% blasts and not for blasts 5% to 9%, as proposed by the International Consensus Classification (ICC), or 5% to 19%, as proposed by the fifth edition of the WHO (WHO-5). MDS-EB1 and -EB2 with VAF ≥10% had comparable survival (9.6 vs 7.2 months; P = .12), regardless of allelic status. Contrary to the ICC proposal, MDS-EB1/EB2 with VAF <10% and CK had poor survival compared with those without CK, comparable to MDS-EB1/EB2 with VAF ≥10% (5.6 vs 26.2 vs 6.3 months; P = .003). Survival of TP53mut AML was poor (median 3.9 months) regardless of allelic/CK status. Thus, using ICC or WHO-5 may underestimate prognosis of MDS with blasts 5% to 19% and 5% to 9%, respectively. Collectively, the hierarchical model acknowledges poor survival of 91.9% TP53mut MDS and AML compared with 36.5% and 80.7% by WHO-5 and ICC, respectively.

ABSTRACTA total of 142 patients with myeloproliferative neoplasms (MPNs) or acute myeloid leukemia (AML) associated with multihit TP53 mutations (mTP53 MUT) were accessed from the Mayo Clinic database and included (i) chronic phase MPN (MPN‐CP; N = 19), (ii) accelerated phase MPN (MPN‐AP; N = 14), (iii) blast phase MPN (MPN‐BP; N = 28), and (iv) AML (N = 81). Concurrent ASXL1 MUT, EZH2 MUT, IDH1,MUT and IDH2 MUT were more common in MPN‐MUTBP‐mTP53 compared to AML‐mTP53 MUT. At median of 11.6 months follow‐up, 124 (87%) deaths and 19 (13%) allogeneic stem cell transplantations (ASCT) were documented. Overall survival (OS), calculated from the time of mTP53 MUT detection, was similar between MPN‐BP‐mTP53 MUT (median 4.6 months) and MPN‐AP‐mTP53 MUT (5.6 months; p = 0.5) but both were inferior to MPN‐CP‐mTP53 MUT (11.6 months, p < 0.01). OS in MPN‐CP‐mTP53 MUT was similar to that of AML‐mTP53 MUT (median 7.4 months, p = 0.07). In multivariable analysis, OS was favorably affected by ASCT (HR 0.4, p = 0.03) and disease stage (i.e., chronic phase disease) or achieving response to pre‐transplant chemotherapy (HR 0.2, p < 0.01) and unfavorably by the presence of concurrent TET2 MUT or DNMT3A MUT (HR 2.7, p < 0.01). Based on these risk factors, a 3‐tiered risk model was constructed: low (no risk factors; N = 18; median OS 23.8 months); intermediate (one risk factor; N = 44; 11.1 months); and high (two or more risk factors; N = 80; 4 months; p < 0.01). The current study highlights the equally detrimental impact of mTP53 MUT on long‐term survival in MPN and AML and identifies predictors of short‐term survival.

SummaryGermline variants in DDX41 (DDX41MT‐germline predisposition syndrome [GPS]) are associated with predisposition to haematological malignancies (HM), including lymphoid and myeloid neoplasms (MN). We retrospectively analysed the clinical and molecular features of 195 patients diagnosed and treated at Mayo Clinic with DDX41MT‐GPS. Patients with germline DDX41 pathogenic variants (42.3%) and variants of unknown significance (VUS, 57.6%) were included. The median age was 68.6 years (16.2–93.4). Ninety‐two per cent were Caucasian, 64.1% were male and 30.8% had a family history of HM. There were 92 distinct germline variants among our cohort, and the most common was p.Met1? (15.9%), followed by p.Asp140Glyfs*2 (9.2%). Clinical diagnoses included asymptomatic carriers (10.2%), clonal cytopenia of undetermined significance (CCUS, 6.1%), myeloproliferative neoplasms (6.7%), myelodysplastic syndrome (40.5%), acute myeloid leukaemia (20.5%), lymphoid neoplasms (9.2%), plasma cell dyscrasias (6.1%) and solid tumours (22.5%). Patients with MN were older (median age 70 vs. 63.5 years) and more likely to be male (M:F ratio 2.3 vs. 1.0) and most patients (78.8%) with MN had a normal karyotype. The most common somatic variants involved DDX41 (34.4%), followed by TET2 (11.2%), DNMT3A (9.6%) and ASXL1 (9.2%). In summary, we have comprehensively described the spectrum of clinical phenotypes within the Mayo Clinic DDX41MT‐GPS cohort.

ABSTRACTTransplant‐associated thrombotic microangiopathy (TA‐TMA) is an endothelial dysfunction syndrome observed after allogeneic hematopoietic cell transplant (alloHCT). Our aim was to externally validate the impact of high‐risk features on the clinical outcomes of adult patients meeting the updated TA‐TMA harmonizing criteria. Between 2005 and 2022, 99 patients were diagnosed with TA‐TMA at Mayo Clinic Rochester (incidence 6.2%) after a median of 137 days post alloHCT (IQR: 34–283 days). The development of TA‐TMA was associated with an inferior overall survival posttransplant (HR: 3.8, 95% CI: 2.97–4.72). High‐risk features, including concomitant infection, acute graft‐versus‐host disease (GVHD), and organ dysfunction, were associated with poor survival, while LDH elevation was not associated with inferior outcomes. The most common treatment strategy for TA‐TMA was discontinuation of calcineurin or mTOR inhibitors in 80 (81%) patients. Thirty (37.5%) patients experienced worsening of GVHD with this strategy, of which 26 (86.7%) patients had died at last follow‐up. The most common cause of death among these patients was worsening GVHD (69%; n = 18), followed by infection (11%; n = 3), disease relapse (8%; n = 2), other/unknown causes (8%; n = 2), or TA‐TMA (4%; n = 1). Objective response rate (ORR) to initial treatment for the cohort was 56.6%. Eculizumab was used in 11 patients with an observed ORR of 70%, including 5 complete responses. In conclusion, TA‐TMA remains a significant contributor to non‐relapse mortality and is associated with worse survival following alloHCT. Not all high‐risk features, particularly LDH elevation, have consistently demonstrated a negative impact in adult cohorts. Patients with TA‐TMA may benefit from immune suppression dose adjustment, rather than a discontinuation, and the addition of complement‐directed therapy, particularly among high‐risk patients.

Abstract Background: DNMT3A and TET2 are epigenetic regulator genes commonly mutated in age-related clonal hematopoiesis (CH). Despite having opposed epigenetic functions, these mutations are associated with increased all-cause mortality and a low risk for hematological neoplasms. In this study, we make use of a natural inflammatory response occurring during COVID-19 to define the impact of these mutations on inflammation and outcomes. Methods: A cohort of 243 community-based patients (Olmsted County, MN) with COVID-19 were identified. CH status was determined on PBMC using error corrected sequencing (lower detection limit 0.5% VAF). In a subset of patients, we use multi-omics interrogation of the genome, transcriptome, and epigenome at single cell (sc) resolution along with DNA methylation, in PBMC, from patients with COVID-19 and CH. Results: We detected 97 CH mutations in 72 (29.6%) of 243 patients with COVID-19. The most frequent being DNMT3A (n=30, 30%) and TET2 (n=26, 28%). The presence of CH negatively impacted overall survival when restricted to TET2 and DNMT3A mutations. However, only DNMT3Amt CH retained age and comorbidity independent prognostication (HR 2.84, p = 0.022). We then assessed for changes in DNA methylation using Illumina MethylationEPIC array on patients with COVID-19 and DNMT3A (n=3) or TET2mt (n=4) CH. Site specific DNA methylation changes revealed significant loss of methylation in DNMT3Amt CH in comparison to TET2mt patients, with 10,944 hypomethylated and 1,160 hypermethylated sites. We found that actively transcribed states (Tx, TxWk) were more commonly hypomethylated in DNMT3Amt CH while enhancers (Enh) and promoters (TssA, TssAFlnk) were more commonly hypomethylated in TET2mt CH. We then conducted scDNA-seq in DNMT3Amt (n=3) and TET2mt (n=1) CH in COVID-19 and observed that while in TET2mt CH, mutations were largely restricted to classical and intermediate monocytes, in DNMT3Amt CH, mutations were seen in myeloid and lymphoid lineage cells. We performed scRNA-seq on patients with COVID-19 and DNMT3A (n=3) or TET2mt (n=6) CH. Differential gene expression analysis identified 1,569 upregulated genes in DNMT3Amt patients and 205 downregulated. Pathway analysis demonstrated an upregulation of genes involved in T cell survival, function, and inflammation in DNMT3Amt patients compared to TET2mt patients with a significant overexpression of GIMAP1, GIMAP4, IRF2, IL32 in multiple cell types. We correlated transcriptomic changes with chromatin accessibility using the 10X Genomics Multiome platform to profile both gene expression and open chromatin from the same samples. Analysis of global distribution of cut sites and differentially accessible peaks showed increased chromatin accessibility in DNMT3Amt CH, especially CD4+ T lymphocytes and NK cells. Conclusion: We demonstrate an age, sex, and comorbidity-independent adverse impact on mortality associated with DNMT3Amt CH in the context of COVID-19 and highlight transcriptomic and epigenetic differences between DNMT3A and TET2mt CH, with single cell resolution. Citation Format: Jenna A Fernandez, Wazim Mohammed Ismail, Moritz Binder, Terra Lasho, Susan M. Geyer, Amelia Mazzone, Christy M. Finke, Abhishek Mangaonkar, Jeong-Heon Lee, Liguo Wang, Vernadette A. Simon, Fariborz Rakhshan Rohakthar, Amik Munankarmy, Jonathan J. Harrington, Melissa R. Snyder, Keith D. Robertson, Alexandre Gaspar-Maia, Mrinal Patnaik. Divergent impacts of DNMT3A and TET2 mutant clonal hematopoiesis on COVID-19 related outcomes [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: DNA Methylation, Clonal Hematopoiesis, and Cancer; 2025 Feb 1-4; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2025;85(3 Suppl):Abstract nr A012.

Abstract Background: Age-related clonal hematopoiesis (ARCH) is a precursor state to blood cancers and is associated with epigenetic age acceleration (EpiAgeAccel). While ARCH is associated with increased all-cause mortality and cardiovascular disease, not much is known about these risks with therapy-related clonal hematopoiesis (t-CH) – CH occurring in the context of prior cytotoxic chemotherapy/radiation exposure. Methods: Patients were prospectively recruited for CH and EpiAge assessment from Mayo Clinic between 2022 and 2024. CH status was determined using error corrected sequencing on peripheral blood (PB) DNA (sensitivity 0.5% VAF). Patients with CH were classified as t-CH if they had received chemotherapy or radiation prior to CH detection. All other CH cases were classifed as ARCH. For EpiAge analysis, PB DNA was analyzed using the Illumina MethylationEPIC array. The resulting beta values were used to obtain epigentic age estimates using Horvath’s clock, Hannum’s Clock, PhenoAge, DunedinPACE, and estimated telomere lengths (DNAmTL). Results: 106 (62%) of 170 patients assessed were found to have CH. CH patients were classified into ARCH-63 (59%) and t-CH-43 (41%). While the median chronological age (69 vs 67 years) was similar between ARCH and t-CH, t-CH patients had lower Hgb [10.9g/dL vs. 13g/dL (p=0.0001)], lower platelet [146x109/L vs. 192x109/L (p=0.0132)] and lower WBC [4.4x109/L vs. 5.2x109/L (p=0.0345)]. The 3 most common ARCH mutations were TET2 (20%), DNMT3A (19%), and ASXL1 (14%), while PPM1D (27%), TET2 (18%), and DNMT3A (14%) were most frequent in t-CH. There was no significant difference in median EpiAge via Horvath (68 vs 67.5years p=0.2826), Hannum (65 vs 66 years p=0.9333), or PhenoAge (55 vs 56 years p=0.618) between ARCH and t-CH. However, DunedinPACE of aging was higher in t-CH (1.27 vs 1.18 bioyear/chronoyear p=0.0148) and median DNAmTL was shorter in t-CH (6.35 vs 6.49kb p=0.0247), compared to ARCH (Note patients with telomere biology disorders were excluded). Multivariable regression analysis adjusting for chronologic age, smoking status, and male sex showed that both ARCH and t-CH had EpiAgeAccel via Horvath(5.8years p=0.0001, 3.4 years p=0.051), Hannum(5years p=0.001, 4.5 years p=0.007), and PhenoAge (6.7years p=0.002, 7.6 years p=0.002) clocks. t-CH more than doubled DunedinPACE (0.2 bioyear/chronoyear, p&lt;0.001) of aging compared to ARCH (0.08 bioyear/chronoyear, p=0.026). t-CH also demonstrated a significant decrease in DNAmTL (0.34kb, p&lt;0.001) in comparison to ARCH (0.17kb, p=0.015). With a median follow-up of 20 months, t-CH was associated with worse overall survival in comparison to ARCH, and non-CH patients (p=0.013, 93% vs. 90% alive at 10 months, 87 vs. 72% alive at 20 months). Conclusion: Our prospective CH cohort study validates the association between CH and EpiAgeAccel using several different methylation clocks and for the first time highlights an increased pace of aging and shorter DNAmTL in the context of inferior survival outcomes in t-CH. Citation Format: Marissa Li, Jenna Fernandez, Kristina Kirschner, Tamir Chandra, Kristen McCullough, Sarah Aug, Terra Lasho, Christy Finke, James Foran, Cecilia Arana Yi, Yael Kusne, Talha Badar, Abhishek Mangaonkar, Naseema Gangat, Rachel Simon, Alejandro Ferrer, Mrinal Patnaik. A prospective cohort study of biological aging and epigenetic age acceleration in 106 patients with age and therapy-related clonal hematopoiesis [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: DNA Methylation, Clonal Hematopoiesis, and Cancer; 2025 Feb 1-4; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2025;85(3 Suppl):Abstract nr A015.

ABSTRACT Patients with newly diagnosed acute myeloid leukemia (ND‐AML) derive variable survival benefit from venetoclax + hypomethylating agent (Ven‐HMA) therapy. The primary objective in the current study was to develop genetic risk models that are predictive of survival and are applicable at the time of diagnosis and after establishing treatment response. Among 400 ND‐AML patients treated with Ven‐HMA at the Mayo Clinic, 247 (62%) achieved complete remission with (CR) or without (CRi) count recovery. Multivariable analysis–derived hazard ratios (HR), including 1.8 for European LeukemiaNet (ELN) adverse karyotype, 4.7 for KMT2Ar , 1.7 for TP53 MUT , 2.6 for KRAS MUT , and 2.1 for IDH2 WT were applied to develop an HR‐weighted risk model: low, intermediate, and high; respective median survival censored for allogeneic stem cell transplant (ASCT) (3‐year survival) were “not reached” (67%), 19.1 (33%), and 7.1 months (0%). In patients achieving CR/CRi, adverse karyotype, KMT2Ar , KRAS MUT , IDH2 WT predicted inferior survival, allowing for a complementary response‐stratified risk model. The model was externally validated and was shown to be superior to the ELN 2024 risk model (AIC 179 vs. 195 and AUC 0.77 vs. 0.69). Survival was inferior with failure to achieve CR/CRi or not receiving ASCT; 3‐year survival for high‐risk with or without ASCT was 42% versus 0% ( p < 0.01); intermediate 72% versus 43% ( p = 0.06); and low‐risk 88% versus 78% ( p = 0.53). The Mayo genetic risk models offer pre‐treatment and response‐based prognostic tools for ND‐AML treated with Ven‐HMA. The current study underscores the prognostically indispensable role of achieving CR/CRi and ASCT.

Introduction: Post-transplant cyclophosphamide (PTCy) has emerged as the new standard of care for graft-versus-host disease (GVHD) prophylaxis following reduced-intensity conditioning (RIC) allogeneic stem cell transplant (alloSCT). The BMT CTN 1703 phase III trial demonstrated superior GVHD/relapse or progression-free survival (GRFS) in patients receiving PTCy, without affecting relapse risk. The primary objective of this study was to compare the risk of relapse in high-risk myeloid neoplasms post RIC/Nonmyeloablative (NMA) transplant using PTCy vs Tacrolimus/Methotrexate (TAC/MTX) as GVHD prophylaxis (PPX). Methods: We retrospectively reviewed patients with high-risk myeloid neoplasms following RIC/NMA alloSCT who received PTCy or TAC/MTX for GVHD PPX between Jan 2018 and June 2023. Criteria for high-risk myeloid neoplasms included acute myeloid leukemia (AML) with ≥1 of the following (complex or monosomal karyotype, TP53, WT1, FLT3 ITD+/NPM1-, active disease, MRD+, secondary AML), Myelodysplastic syndrome (MDS) with ≥1 of the following (complex karyotype, monosomal karyotype, TP53, RAS pathway mutation, marrow blast of 10% or more) or chronic myelomonocytic leukemia (CMML). Kaplan-Meier and log-rank tests were used to estimate overall survival (OS) and disease-free survival (DFS). Non-relapse mortality (NRM) and relapse incidence (RI) were calculated using competing risk analysis. Factors with P&lt;0.10 in univariate analysis were included in multivariate analysis Results: Out of 552 alloSCTs, 108 (19.5%) patients met the inclusion criteria. Seventy-two (66.6%) received TAC/MTX, and 36 (33.3%) received PTCy. The median follow-up time was 26 months (range 3-68). The median age at transplant was 65 years (range 62-69) with age comparable between the two groups (p=0.6). Haploidentical transplant accounted for 33.3% of PTCy, while the remainder had matched related or matched unrelated transplant (MRD/MUD). Compared to TAC/MTX group, the PTCy group had more frequent NRAS mutation (19.4% vs 8.3%, p=0.094), but fewer patients with hematopoietic cell transplantation-specific comorbidity index (HCT-CI) ≥ 3(41.7% vs 61.1%, p=0.056). AML MRD+ was more prevalent in the PTCy group (22.2% vs 8.3%, p=0.098). The two groups were otherwise well-balanced in terms of disease distribution, disease risk index, TP53 mutation, and complex monosomal karyotype (CK and MK). The 1-yr OS, DFS, and GRFS for PTCy vs TAC/MTX were 78.7% vs 72.8% (p=0.9), 53.6% vs 67.9% (p=0.2), 38% vs 40%, (p=0.4), respectively. The 1-yr cumulative incidence of relapse (CIR) was higher in PTCy compared to TAC/MTX (37.6% vs 11.1%, p=0.01). Both haploidentical and MRD/MUD in the PTCy group had similar 1-yr RI (34.7 vs 38.3% p=0.7). Limiting the analysis to MRD/MUD, the PTCy group had a higher risk of relapse compared to TAC/MTX (34.7% vs 10.2%, P=0.007). Sub-analysis showed a higher 1-yr-CIR in the PTCy cohort for secondary AML (40% vs 0.0%, p=0.01), TP53 mutation (44.4% vs 18.1%, p=0.02), CK/MK (71.4% vs 22.2%, P=0.008), and chromosome 17 abnormality (60% vs 10%, P=0.001), compared to TAC/MTX. Despite of the increased CIR in the PTCy group, only patients with chromosome 17 abnormality receiving PTCy had an inferior DFS compared to TAC/MTX (median 5 vs. 14 months, p=0.008). In multivariate analysis for 1-yr CIR that included TP53, chromatin modifiers mutations, complex karyotype, high/very high DRI, and PTCy, both PTCy (HR 4.56, 95% CI 1.1-18.07, p=0.031) and high/very high DRI (HR 12.745, 95% CI 1.5-103.4, p=0.017) were independently predictive of post-transplant relapse. However, these factors were not significantly associated with 1-year OS. Conclusion:In patients with high-risk MN following RIC/NMA alloSCT, those receiving PTCy as GVHD PPX had a higher risk of relapse compared to TAC/MTX, irrespective of the donor source. Notably, an inferior DFS was observed only in patients with chromosome 17 abnormality who received PTCy. Despite of the increased relapse risk in the PTCy group, OS, DFS, and GRFS were comparable to TAC/MTX, likely due to higher risk of GVHD and NRM in the latter group.