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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 <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 <5% and PB <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<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 Background Chronic lymphocytic leukemia/small lymphocytic lymphoma (CLL/SLL) is the most common leukemia in US adults and incidence increases with age. Although the treatment (tx) landscape has evolved, it is unclear whether tx selection differs for older patients (pts), given concerns around comorbidities, toxicity, and efficacy. There is a need to better understand pt characteristics, tx patterns, and biomarker use among pts aged ≥65 yrs with CLL/SLL in the current therapeutic era. Methods A retrospective longitudinal study was conducted using the de-identified 100% Medicare Fee-for-Service (FFS) Research Identifiable Files. Included pts were diagnosed with CLL/SLL at age ≥65 and initiated first-line (1L) tx 1/1/2020-9/30/2024 (with ≥12 mo in database prior to 1L tx start). Index date was defined as date of 1L tx start, and pts were followed to death, end of Medicare enrollment, or study end, whichever occurred first. 1L tx was defined as any CLL pharmacologic tx within the first 90 days of index date. Tx utilization was summarized as the number and proportion of pts who received each tx at each line. Descriptive statistics were used to summarize demographic and social characteristics and baseline comorbidities, in all pts and by 1L tx. Proportion and timing of documented biomarker orders any time prior to index were summarized by test type (CLL fluorescence in situ hybridization [FISH], TP53 DNA sequencing and immunoglobulin heavy chain variable region [IGHV] test) and by 1L tx. Results Of the 21,008 pts included, median age was 76.4 yrs at 1L tx, with 43% of pts aged 65-74 yrs, 43% aged 75-84 and 14% aged ≥85. The majority of pts were male (58%) and White (91%). About 8% and 9% of pts had dual eligibility status and low-income subsidy, respectively. Median Charlson Comorbidity Index (CCI) was 4 (range: 1-21), with 40% of pts having CCI ≥5. Covalent Bruton tyrosine kinase inhibitor (cBTKi) monotherapies (mono) were most common at 1L tx (41%), including 10% zanubrutinib (zanu), 15% ibrutinib (ibr), and 16% acalabrutinib (acala). B-cell lymphoma 2 (BCL2)+anti-CD20 monoclonal antibody (mAb) was less common (11%), and chemoimmunotherapy (CIT) was used by 13% of pts, mostly BR (8%) and FCR (3%). Other less common tx included cBTKi+anti-CD20mAb (6%) or cBTKi+BCL2 therapies (1%). Among remaining pts, most received anti-CD20mAb (22%). The use of cBTKi mono increased from 41.7% in 2020 (ibr 30.6%; acala 11.1%) to 47.6% in 2024, driven by increased use of zanu (27.8%; acala 15.7%; ibr 4.2%). BCL2+anti-CD20mAb increased from 9.2% in 2020 to 12.6% in 2024. Pts treated with cBTKi mono were older than those with BCL2+anti-CD20mAb (median age 77.2 vs 74.5), and more likely to have dual eligibility status (10% vs 4%) or low-income subsidy (11% vs 5%). More pts who received zanu mono had CCI ≥5 (39%) versus acala (36%), ibr (35%), or BCL2+anti-CD20mAb (32%). Pts with BCL2+anti-CD20mAb were less likely to have diabetes (23% vs 26%), congestive heart failure (CHF; 16% vs 19%), chronic pulmonary disease (23% vs 27%), and renal diseases (23% vs 27%). Pts with ibr were less likely to have myocardial infarction (29% vs 34%), coronary artery disease (22% vs 25%), CHF (16% vs 19%), and atrial fibrillation (12% vs 18%). Overall, 69% of pts had FISH testing prior to 1L tx, but test orders for TP53 (12%) or IGHV (28%) status were infrequent. Pts with ibr had the lowest rates of TP53 tests (6%), whereas pts with CIT had the lowest rates of IGHV tests (16%). The timing of tests showed a binomial distribution where most pts received FISH, TP53, or IGHV tests either ≤60 days prior to 1L tx (46%, 50%, 33%), or >1 yr before 1L tx (30%, 22%, 43%). Among pts who received ≥2 lines of therapy for CLL (n=4428; 21%), cBTKi mono was the most common in 2L (41%), including 14% zanu, 17% acala, and 10% ibr. CIT and BCL2+anti-CD20mAb were used in 12% and 9% of pts, respectively. Only 634 (3%) pts had 3L+ tx, of whom 33% received cBTKi mono, including 16% zanu, 12% acala, and 5% ibr (5%). Conclusion This large real-world study found that cBTKi mono was the most common 1L tx choice among pts aged ≥65 with CLL/SLL, with increased use of next-generation cBTKi. Pts with more baseline comorbidities were less likely to receive ibr or BCL2-based tx. FISH testing was not performed in nearly one-third of pts, and most pts did not have TP53 or IGVH sequencing prior to tx. Future studies are needed to continue to evaluate evolving tx paradigms in older pts with CLL/SLL.

Abstract Introduction:A comprehensive integration of significantly mutated genes and pathways with molecular subgroups is lacking in adult B-ALL. Furthermore, the genomic drivers underlying each subgroup remain to be identified.We aimed to perform genomic analysis on a large cohort of patients with adult B-ALL, and to correlate genomic markers with achievement of MRD negativity enabling blinatumomab randomization, and with subsequent response to blinatumomab. Methods:For genomic analysis, we studied 569 adults with newly diagnosed B-ALL registered for initial screening on the ECOG-ACRIN-led E1910 trial (NCT02003222), which evaluated the addition of blinatumomab to standard consolidation chemotherapy. The median age was 52 yrs (range 30 to 71 yrs), with 52% males. Analysis was performed on tumor and matched-normal samples using whole transcriptome sequencing (RNA-seq; tumor only; n=569), whole exome sequencing (n=490), whole genome sequencing (n=131), and single nucleotide polymorphism array (n=445). B-ALL cases were classified into 22 molecular groups. Outcome analysis was limited to patients enrolled and treated on E1910 (n=319). Results: Driver genes (n=268) were identified by the mutation-significance detection tool dNdScv or by the presence of pathogenic variants in known cancer genes, with NUP188 identified as a novel driver gene. Recurrently mutated pathways included: B-cell development (46%), cell cycle (44%), epigenetic regulation (30%), other transcriptional regulation (27%), Ras signaling (25%), RNA machinery (17%) and JAK-STAT signaling (8%). A high frequency of high-risk subtypes was observed including BCR::ABL1 (20%), BCR::ABL1-like (18%), low hypodiploid (14%) and KMT2A (12%). Using tSNE analysis, we identified a new cluster of cases (n=20, 3.5%) lacking a known subgroup driver, with overexpression of CEBPA (n=13) or CEBPB (n=7), distinct from CEBPE/ZEB2, termed “CEBP-altered” ALL (CEBPalt). Five of 13 cases with high CEBPA expression harbored an IGH::CEBPA rearrangement and one harbored a RXRA::CEBPA enhancer hijacking alteration. Of the 7 cases with high CEBPB expression, 5 harbored IGH::CEBPB, and two of these had concomitant clonal BCR::ABL1 fusions. To identify additional mechanisms of CEBP deregulation, in situ Hi-C coupled to H3K27 acetylation immunoprecipitation (HiChIP) was performed on 5 cases. Four cases showed evidence of de novo CEBPA enhancer activity. In two cases, we identified novel insertions (12 and 34nt) ~2kb downstream of CEBPA. A novel translocation and enhancer hijacking event between LINC00426 and the 5' UTR of CEBPB was identified in the remaining case. Overall, we confirmed genomic alterations of CEBPA or CEBPB in 16 of 20 cases with available material. Thus, we have identified a new molecular subgroup of adult B-ALL characterized by genomic alterations that drive enhancer hijacking and oncogenic deregulation of CEBPA/CEBPB. Several subgroups were enriched in patients that failed induction chemotherapy (n=62) or were MRD-positive (n=63) compared to those that achieved MRD-negative status (n=194): BCR::ABL1-like (30 vs 14%, p<0.001), KMT2A (18 vs 9%, p=0.03) and BCL2/MYC (5 vs 1%, p=0.06). Conversely, the following subgroups were enriched in patients that achieved MRD-negativity: PAX5alt (6 vs 18%, p=0.002), TCF3::PBX1 (0 vs 5%, p=0.01) and ZNF384 (2 vs 6%, p=0.08) Within each molecular subgroup we compared the survival of patients who achieved MRD-negative status after induction and were randomized to receive blinatumomab plus chemotherapy (n=93) or chemotherapy only (n=91). Although numbers were low, blinatumomab improved relapse-free survival for patients with hyperdiploid, PAX5alt, BCR::ABL1-like and KMT2A ALL compared to chemotherapy alone. Clonal hematopoiesis of indeterminate potential (CHIP)-related gene mutations were identified in 121 of 417 cases analyzed (29% total: somatic 74%, remission 31%, both 5%). TP53 mutations were most frequent (n=65 patients), with the majority identified in patients with low hypodiploid (n=52). Interestingly, patients with CRLF2 rearrangements collectively harbored the highest number of other CHIP gene mutations (n=13), including DNMT3A (n=4), TET2 (n=3) and ASXL1 (n=3). Conclusions:We provide a comprehensive landscape of genomic alterations in adult B-ALL and identify a new group characterized by deregulation of CEBPA/CEBPB (CEBPalt). We also provide insights into the efficacy of blinatumomab in different molecular subgroups of adult B-ALL.

Abstract Introduction While outcomes in patients with R/R FL have improved with T-cell–redirecting treatments (e.g. CAR T-cell therapy, bispecific antibodies), there remains an unmet need for safe, efficacious, and more convenient chemotherapy-free treatment options (Caridà et al, Eur J Haematol. 2025). GOLCA is a potential, first-in-class, oral CELMoD agent designed for the treatment of lymphoma, with preferential distribution to lymphoid organs and enhanced activity in lymphoma cell lines. GOLCA drives the closed, active conformation of cereblon to induce rapid and deep degradation of Ikaros and Aiolos, leading to direct cell killing (agnostic of cell of origin) and immunomodulatory activity. In the two-part, multicenter, first-in-human Phase 1/2 study (CC-99282-NHL-001; NCT03930953), GOLCA was well tolerated and effective in patients with R/R FL (Cordoba et al, ICML 2025, #441). Here, we provide longer follow-up in patients with R/R FL from Part B of the study. Methods Patients with R/R FL with ≥ 2 prior lines of therapy (≥ 1 in Part B if prior anti-CD20 therapy) were included in the study. Patients received GOLCA monotherapy orally, once daily at different dosing schedules in Part A. In Part B, GOLCA was dosed at 0.2 or 0.4 mg (14 days on/14 days off) ± R. Total GOLCA treatment duration was up to 2 years or until progressive disease (PD)/unacceptable toxicity. Primary objectives included safety and recommended Phase 2 dose determination. Results As of April 3, 2025, a total of 72 patients with R/R FL were enrolled: 12 in Part A (GOLCA monotherapy) and 60 in Part B Cohort D (22 received GOLCA 0.2 mg + R; 38 received GOLCA 0.4 mg + R). Patients were heavily pre-treated. Median number of prior treatments was 4.5 (range, 2–6) in Part A and 3 (range, 1–12) in Part B Cohort D. Approximately one-third of the treated patients were exposed to prior T-cell–redirecting therapy, approximately one-third had prior lenalidomide (len) exposure, and approximately one-third were refractory to the last regimen received. At the time of data cutoff, in Part A, 4 patients completed 2 years of GOLCA, remaining in remission at last follow-up; 8 discontinued due to PD. In Part B Cohort D, 41% of patients treated with 0.2 mg and 63% with 0.4 mg were ongoing. PD was the most common reason for discontinuation. In the safety evaluable population (n = 60; GOLCA 0.2 or 0.4 mg + R), neutropenia, an on-target side effect of GOLCA, was the most common grade 3/4 treatment-emergent adverse event (TEAE) and occurred in 65% of patients, followed by anemia (13%) and febrile neutropenia (8%). Serious AEs related to GOLCA occurred in 22% of patients, most commonly infections (17%) and febrile neutropenia (7%); pulmonary embolism occurred in 1 patient with 0.2 mg. The most common causes of GOLCA interruption in the overall safety population were infections (32%) and neutropenia (18%). Dose reductions were mostly due to neutropenia (10%) and febrile neutropenia (5%). No discontinuations or deaths occurred from GOLCA-related TEAEs. Non-hematologic TEAEs were infrequent and mostly low grade, with the most common being gastrointestinal disorders (7%). In the efficacy evaluable patients (n = 56; GOLCA 0.2 or 0.4 mg + R) at a median follow-up of 10.8 months, the overall response rate (ORR) was 89% (complete response rate [CRR], 61%). With 0.2 mg, ORR was 81% (CRR, 43%) and with 0.4 mg, ORR was 94% (CRR, 71%). In the 0.4-mg group, responses were consistent in high-risk subsets, including patients with prior len (ORR, 100%; CRR, 73%) and/or T-cell–redirecting therapy (ORR, 91%; CRR, 64%). With GOLCA 0.4 mg, 10 patients had response lasting > 12 months, including 5 with prior T-cell–redirecting therapy and 6 with prior len. Conclusions With additional follow-up, GOLCA + R continued to show promising efficacy with durable responses and no new safety signals. A higher ORR/CRR and similar tolerability were observed with GOLCA 0.4 mg + R vs GOLCA 0.2 mg + R, including in patients with prior len-based and/or T-cell–redirecting treatment. These data support continued development of GOLCA 0.4 mg + R as a fixed-duration, chemotherapy-free, outpatient option in the ongoing Phase 3 GOLSEEK-4 study in 2L+ FL (NCT06911502).

Abstract Introduction: Autologous CD19 CART therapies have been utilized across a wide range of B cell driven autoimmune diseases (AD), including systemic lupus erythematous (SLE), idiopathic inflammatory myopathies (IIM), systemic sclerosis (SSc), & myasthenia gravis (MG). To date, a number of autologous CD19-CART therapies have delivered durable drug free responses in AD patients. However, the safety profile of autologous CD19-CART therapies across different AD has yet to be fully explored. This is especially true for CRS & ICANS. Here, we provide novel insight into the mechanism underlying the safety profile of resecabtagene autoleucel (rese-cel) an autologous CD19 41BBz CART therapy across four separate Phase I/II clinical trials in SLE, IIM, SSc, & MG. Objectives: The primary objective of each trial is safety & tolerability of rese-cel at Day 29. Key secondary objectives include changes in clinical scores & use of immunomodulatory agents. Key translational assessments include CART cell pharmacokinetics (PK), impact on peripheral B cell populations (pharmacodynamics), & serum cytokine levels. Methods: 19 Patients (data cutoff: 6/2/2025 SLE; 5/6/2025 IIM, SSc, and MG), with active disease refractory to standard of care have been treated across 4 Phase I/II clinical trials in SLE (8 patients; NCT06121297), IIM (8 patients; NCT06154252), SSc (2 patients; NCT06328777), & MG (1 patient; NCT06359041). All patients were treated with a single weight-based infusion of rese-cel at a dose of 1 x 106 cells/kg following lymphodepletion (flu 25 mg/m2/d on Days -5, -4, & -3, & cy 1,000 mg/m2/d on Day -3). All non-glucocorticoid immunomodulatory agents were stopped prior to lymphodepletion. Glucocorticoids were tapered post-infusion. Patients were not given prophylactic tocilizumab pre-infusion. Translational assessments were determined as follows: CART PK was evaluated by flow cytometry & dPCR; B-cell enumeration & phenotyping were evaluated by flow cytometry; & serum cytokines were evaluated via immunoassay. Results: Across all indications (19 patients), 7 episodes of CRS were observed (7/19). 6 of 7 patients experienced a Grade 1 CRS, & 1 patient had a Grade 2 CRS. 2 patients developed ICANS; 1 patient had a Grade 4 (previously presented at ACR Convergence 2024), & the other patient had a Grade 3. Both ICANS events were rapidly resolved without sequelae using standard therapy. These CRS and ICANS events differed across indications. In IIM, 4 of 8 patients experienced Grade 1 CRS and no patients experienced ICANs. In SLE, 2 patients experienced Grade 1 CRS and one developed Grade 4 ICANs. In SSC, 1 of 2 patients experienced Grade 2 CRS & the other experienced Grade 3 ICANS. In MG, no CRS or ICANS was observed. Rese-cel infusion products were ~ 66% transduced, with a mean CD4:CD8 ratio of 6. Post-infusion, peak CART expansion was observed on day 12 & the mean peak expansion was 89 cells/µL (SD +/- 214). B cells were rapidly depleted from peripheral blood post-infusion, with nadir occurring at 13 days & repopulation beginning at 62 days post-infusion on average. Re-emergent B cells were largely of the CD24+CD38+ transitional naïve phenotype. Commensurate with B cell depletion, serum BAFF induction was observed, with a mean fold induction of 16 & mean peak level of 56,404 pg/µL (SD: +/- 45,547). Serum cytokines associated with CRS & ICANS; IFNg, IL6, & IL8 reached peak elevation on days 10, 9, & 10 respectively post-infusion. The mean peak elevation for IFNg, IL6, & IL8 were 232 pg/ µL (SD: +/- 320), 44 pg/ µL (SD: +/- 60), & 80 pg/ µL (SD +/- 73), respectively. Conclusion: Rese-cel appears to be well tolerated in autoimmune patients with rates of CRS & ICANS of 37% & 11% across all autoimmune indications, respectively (SLE patients had a 25% CRS rate & 12.5% ICANS rate, IIM patients had a 50% CRS rate & 0% ICANS rate, SSC patients had a 50% CRS & 50% ICANS rate, & MG patients had a 0% CRS & ICANS rate). Notably, the frequency & severity of CRS appears to be low, with no CRS in 63% of patients (12/19) & most CRS events (6/7) being Grade 1. The lower frequency & severity of both CRS & ICANS events are likely correlated with the relatively modest induction of serum IFNg, IL6, & IL8 found in autoimmune patients post rese-cel weight-adjusted dose infusion. High BAFF levels reflect deep systemic B cell depletion. Together, these data support further support late-stage clinical development of rese-cel in autoimmune disease.

Abstract Introduction Histiocytic neoplasms (HN) are a rare, heterogeneous group of hematologic disorders derived from myeloid-dendritic cell lineages. The most common forms include Langerhans cell histiocytosis (LCH), Erdheim-Chester disease (ECD), and Rosai-Dorfman disease (RDD). These disorders are driven by recurrent activating mutations in the MAPK pathway, most frequently involving BRAF, MAP2K1, PIK3CA, KRAS, and NRAS. This discovery enabled a major therapeutic advance with the use of BRAF and MEK inhibitors, resulting in durable responses. For patients harboring KRAS mutations, MEK inhibitors (MEKi) remain the primary therapeutic approach. However, the impact of KRAS mutation subtypes on treatment response remains poorly understood. Our study aimed to characterize the spectrum of KRAS mutations in LCH, ECD, and RDD and evaluate the responses to MEKi, providing insights to guide future treatment strategies. Methods We conducted a retrospective review of patients with LCH, ECD, and RDD who were evaluated at Mayo Clinic or University of Alabama at Birmingham. Patients were included if next-generation sequencing identified a KRAS mutation. The response assessment was based on radiographic evaluation under the most recent Histiocyte Society consensus guideline. Results We identified 23 patients with a KRAS mutation: 17 females (73.9%) and 6 males (26.1%), with a median age at diagnosis of 61 years (range, 20–78). RDD accounted for 73.9% (17/23) and ECD for 26.1% (6/23); no KRAS mutations were detected in LCH patients. The most frequent KRAS variants were p.K117N (34.8%), p.G12D (17.4%), p.A146T (13.0%), p.G13C (8.7%), and p.G13D (8.7%), with additional variants including p.T58I, p.A59G, p.A146P, and p.A146V. Mutation distribution differed by disease group: p.G12D was more common in ECD (50%) than RDD (5.9%) (p=0.01), whereas p.K117N occurred in 47.1% of RDD and was absent in ECD. Regarding treatment, 57% (13/23) received MEKi, all initially cobimetinib, with 1 patient switched to trametinib due to progression. Of the 10 patients who did not receive MEKi, 1 was treated with vemurafenib for a concurrent BRAF p.V600E mutation; 3 remained on observation; 2 underwent surgery without recurrence; 3 received other systemic therapies including corticosteroid, methotrexate, rituximab, lenalidomide, or sirolimus; and 1 developed diffuse large B-cell lymphoma, treated with R-CHOP. The overall response rate in the 12 patients on MEKi (excluding 1 patient too early for assessment) was 75%—3 (25%) CR and 6 (50%) PR (including 1 who initially progressed on cobimetinib but responded to trametinib). All 3 non-responders had RDD. Among the 5 patients with p.K117N, 3 had no response to cobimetinib, 1 achieved PR, and 1 achieved CR; notably, 1 non-responder switched to trametinib and achieved PR in 2 months. All 3 patients with codon 146 mutations achieved PR. Of the 2 patients with p.G12D, 1 achieved CR with a co-occurring MAP2K1 mutation, and 1 had no response. No additional genotype–response patterns were observed. Similar heterogeneity has been reported in prior studies. In a phase II cobimetinib trial, an ECD patient with p.R149G achieved PR, whereas another ECD patient with p.G12R and an LCH patient with p.G13C achieved CR in the presence of co-occurring ARAF and NRAS mutations, respectively. A retrospective cohort described an ECD patient with p.K117N achieving PR on dabrafenib and trametinib with a concurrent BRAF mutation and another with p.G12A achieving PR on trametinib. In a multicenter analysis of trametinib, a patient with p.A59T (co-occurring BRAF and NRAS mutations) had no response, whereas another with p.A146P achieved PR. Collectively, these findings, together with our data, underscore that KRAS-mutated histiocytoses exhibit heterogeneous responses to MEKi, likely influenced by variant type and co-mutations. Discussion KRAS mutations represent an important subset of MAPK pathway alterations in HN, although their prevalence varies by disease type and appears extremely rare in LCH, with no cases identified in our cohort. The most common variants include p.K117N and p.G12D, which may be linked to suboptimal response to MEKi in the absence of concurrent MAPK pathway alterations. As MEKi acts downstream, efficacy likely depends on the level of pathway activation conferred by specific KRAS variants. Larger studies are needed to confirm these observations and identify mutation-specific predictors to guide personalized treatment.

Abstract Introduction The 2022 WHO/ICC classifications of myeloid neoplasms (MNs) incorporated genomics into the existing clinicopathologic framework, defining new (mono)genetic entities. However, this iterative approach resulted in a mix of pathologically and genomically-defined entities, reliant on arbitrary thresholds and historically-derived boundaries between entities. We hypothesised that integrating genomics with other features, de novo from first principles, might better incorporate modern molecular knowledge to define the biological continuum, evolutionary links and overlapping features across MN. Methods The discovery cohort comprised 6484 thoroughly annotated pts. Contemporary diagnoses included 1817 AML, 1695 MDS, 737 MPN, and 1101 MDS/MPN. Non-malignant pts (AA/CHIP/CCUS/ICUS) were included to explore beyond current neoplastic boundaries. 1407 pts had ≥1 follow-up sample/s. Validation was performed in an independent international cohort (n=2012). An unsupervised random forest algorithm was used to integrate status (and VAF) for 28 mutations, karyotypes, CBC features, sex and age (total parameters 144). Features were selected for potential to be independent of BM sampling. Proximity-based multidimensional scaling/PCA were used to visualize latent structure and identify variance-driving features. Results The model projected pts onto a continuous 3-dimensional space. We first applied clustering seeking 1st-order structure, revealing 4 major clusters (C1–C4). These were further subdivided into 58 distinct subclusters (SCs) by an unsupervised iterative clustering approach. All SCs were enriched for distinctive features, with diverse associations linking genomics to clinicopathological/outcome features. Notably, contemporary diagnoses were distributed widely throughout this landscape, with 9/58 SCs containing cases spanning multiple WHO/ICC MN categories. C1 displayed the most internal heterogeneity, with admixed AML, MDS and MDS/MPN pts. It naturally subdivided down to 4th-order subclustering. Ten SCs were dominated by splicing factor mutations. For example, SF3B1MT segregated into 3 SCs, alone or with high-VAF TET2/ASXL1 or DNMT3A mutations. A separate TET2/SRSF2 SC associated with elevated monocyte%, reflecting a discrete CMML-phenotype genomic entity. NPM1MT formed a distinct SC with FLT3-ITD, distinguished further by VAFs of FLT3, TET2, and NRAS. Current genomically-defined AML subtypes separated in the 4th subclustering. C2 encompassed classical MPN driver mutations across 6 SCs, including a distinct CALRMT group and others defined by variable JAK2 VAFs, co-mutation patterns and other CBC features: eg high-VAF SF3B1MT correlated with PB blast%, and concurrent TET2MT with higher Hb/monocyte counts. C3 was characterized by TP53 alterations, with 5 SCs differing by TP53MT burden, 17p lesions, karyotypic complexity and age. Two SCs, one marked by high TP53 VAF+complex karyotype, the other by ch17 lesions, were associated with the worst OS in the whole cohort. C4 contained most known non-malignant pts, dividing into 4 SCs: 2 largely comprising CCUS and AA, while 2 reflect overlaps/convergence between AA and MDS and MPN, respectively. Repurposing single-cell pseudotime analysis suggested multiple disease origins, with various progression trajectories converging on C3. Projecting them onto the model longitudinal samples moved along these derived paths, consistent with their known progression/remission changes, validating the model's biological relevance and predictive utility. Remarkably, the 58 SCs displayed distinct risks of clonal progression and survival. To maximize potential clinical utility we created a pan-MN prognostic score integrating these features. It performed robustly (c-index 0.84), matching/exceeding existing prognostic tools and validated in an independent dataset. Conclusion Integrating clinicopathologic and genomic data we developed ab initio a continuum model of MNs that transcends historical diagnostic boundaries, captures disease evolution and stratifies clinical outcomes. It may better reflect the core biology and natural histories of specific cases, placing more rational boundaries between discrete entities. Potential advantages extend to treatment selection/trial design (in the era of molecularly-targeted therapies) and personalised prognostication throughout each disease course. Expanded validation is needed to resolve complexity in some areas (notably in C2/C4) and validate/refine this prototype framework.

Abstract Background The prognostic relevance of fluorescence in-situ hybridization (FISH) abnormalities is well-established in multiple myeloma (MM), where high-risk (HR) abnormalities are associated with poor survival. In AL amyloidosis, however, the cytogenetic landscape and its implications might be different. For example, t(11;14), common in AL amyloidosis, has historically been linked to inferior outcomes in the pre-daratumumab era. HR FISH abnormalities are less common and less well-defined in this setting, underscoring the need to evaluate their significance in this disease. Methods We retrospectively analyzed 698 newly diagnosed AL amyloidosis patients seen at Mayo Clinic, Rochester, between 2012–2021 who had baseline FISH testing. HR FISH was defined as del(17p)/monosomy 17, t(4;14), t(14;16) or t(14;20). Baseline characteristics and treatment patterns were compared between the HR and the standard-risk (SR) groups. Staging used the Mayo 2004 system with European modification. Hematologic responses were assessed at 2 and 6 months, and cardiac/renal responses at 3, 6, 12, and 24 months per consensus criteria. Overall response rate (ORR) for hematologic and organ responses included patients who achieved partial response (PR), very good partial response (VGPR), and complete response (CR). Overall survival (OS) was estimated via Kaplan-Meier; comparisons used log-rank testing. Cox regression was used for univariate and multivariable analyses. Results Among 698 patients, 73 (10.4%) had HR FISH. The most common HR abnormality was t(14;16) (n=34, 4.9%), followed by del(17p)/monosomy 17 (n=21, 3%), and t(4;14)/t(14;20) (n=12 each, 1.7%). Across the cohort, t(11;14) was most frequent (51.3%), followed by del(13q) (36.2%), hyperdiploidy/tetraploidy (26.3%), and 1q gain/amp (23.6%). Compared to SR group, HR FISH group had fewer males (43% vs. 69%, p<0.0001), with no difference in age (62.5 vs. 65, p=0.44), cardiac (73% each, p=1) or renal involvement (52% vs. 54%, p=0.71). More HR patients had early cardiac stage (69% vs. 55.9%, p=0.04). Both groups were predominantly lambda isotype (82% vs. 74%, p=0.15), with similar baseline dFLC (21.6 vs. 28.4 mg/dL; p=0.81). Intact immunoglobulin secretion was more common in the HR group (68% vs. 45%, p=0.004). The HR group more often underwent advanced bone imaging (44% vs. 29%, p = 0.015) and had a higher incidence of lytic lesions or fractures (27% vs. 15%, p = 0.046). Use of bortezomib-based therapy, primarily CyBorD, was similar between the groups (67% each). Daratumumab-based therapy (16% vs. 11%, p=0.25) and autologous stem cell transplantation (ASCT) (33% vs. 25%, p=0.16) were numerically higher in the HR group, though not statistically significant. Post-ASCT maintenance was more frequent in the HR group (17% vs. 7%, p=0.006). At 2 months, hematologic ORR was higher in the HR group (90% vs. 72%, p=0.007), with higher ≥VGPR (67% vs. 32%, p<0.0001) and CR rates (24% vs. 8%, p=0.0004). At 6 months, the HR group maintained higher ≥VGPR (89% vs. 71%, p=0.008) and CR rates (41% vs. 26%, p=0.04), with no difference in ORR (98% vs. 92%, p=0.18). Cardiac responses at 6 months favored the HR group (VGPR or better: 29% vs. 12%, p=0.013; CR: 13% vs. 3%, p=0.007). At 12 months, cardiac ≥VGPR remained higher (50% vs. 29%, p=0.01), though CR was not significantly different (18% vs. 9%, p=0.1). Renal ≥VGPR at 6 months was more common in the HR group (47% vs. 22%, p=0.004), but not renal CR (9% vs. 7%, p=0.95). At 12 months, renal ≥VGPR remained higher in HR (61% vs. 40%, p=0.004). With a median follow-up of 87 months, median OS was not reached in HR (95% CI: 72.6–NR) vs. 88.9 months (95% CI: 72.8–110.5) in SR (p=0.03). In multivariable analysis, age ≥65 (HR 1.56, p=0.0001) and >2 organ involvement (HR 1.46, p=0.002) were associated with worse OS, while ASCT (HR 0.35, p<0.0001) and cardiac stage I/II (HR 0.48, p<0.0001) predicted better OS. HR FISH was not independently associated with OS (HR 0.74, p=0.14). Conclusions HR FISH abnormalities in AL amyloidosis are not independently associated with OS, unlike in MM. Patients with HR FISH abnormalities more often presented with earlier cardiac stage and demonstrated deeper hematologic and organ responses than patients with SR FISH abnormalities. This study underscores how clonal characteristics can potentially aid in more personalized treatment approaches for patients.

Abstract BackgroundWarm autoimmune hemolytic anemia (wAIHA) is clinically characterized by autoantibody-mediated (typically IgG) erythrocyte destruction at body temperature. While oral corticosteroids remain first-line therapy, rituximab is frequently used as a second-line agent, mostly supported with concomitant background medications, yet real-world evidence is limited and utilization is driven by published retrospective data and expert opinion. Real-world evidence characterizing its utilization patterns and outcomes is needed to inform clinical practice. AimsThis study aimed to: 1) describe treatment patterns among patients with newly diagnosed wAIHA, 2) evaluate rescue therapy use and healthcare resource utilization (HCRU) with current standard of care (SOC), 3) characterize patterns of rituximab use during the first year. MethodsNewly diagnosed wAIHA patients were identified from multiple US databases (MarketScan® Databases, IQVIA PharMetrics, Optum Clinformatics®, and Optum PanTher EHR). Patients with ≥1 wAIHA diagnosis code in any position (ICD-10-CM D59.11) from October 2020 to December 2023 and 12-month post-index follow-up were pooled together. Secondary wAIHA was defined with evidence of hematologic malignancy, solid tumors, immunodeficiencies, or autoimmune diseases in the 6 months prior to diagnosis. Patients on treatment were defined by use of oral corticosteroids (OCS), immunosuppressants (IST) (including azathioprine, mycophenolate mofetil, cyclosporine, cyclophosphamide, or tacrolimus) or rituximab. Initial therapy was defined as the first treatment occurring ≥ 30 days prior to diagnosis. Therapies were identified as combination when initialized within 21 days of each other. The regimen is considered active until all therapies are discontinued or a new agent is initiated. Rituximab regimens were classified as standard (4 weekly doses), alternative (2 doses given 2 weeks apart), single infusion, or extended (>4 infusions). Results Among 2,028 wAIHA patients (median age 62.0 years, 59% female), 55% had secondary wAIHA. Among patients on treatment (n=1,439), 64% received OCS monotherapy; rituximab and IST, alone or in combination with OCS were received by an additional 28% and 5% of patients, respectively. Overall, 80% of treated patients received OCS in some combination during initial therapy. Within 12 months following diagnosis, 1 in 4 patients required ≥ 1 blood transfusion, 1 in 8 required either erythropoiesis-stimulating agents (ESA) or intravenous immunoglobulin (IVIG). Within 12 months of diagnosis, 35% of patients required ≥ 1 hospitalization, average length of stay 3.8 days; AIHA was the principal diagnosis for 65% cases. Greater than 1/3 of patients (38%) had ≥ 1 emergency department visit. Almost all wAIHA patients (98%) had an all-cause outpatient visit with a median of 31 visits per patient. Among patients receiving rituximab in any line (n=613), median time from diagnosis to first administration was 59.0 days (interquartile range: 42.0-171.0). For the first course of rituximab, 62.2% of patients received a complete course (standard, alternative, or extended courses). Among patients with a completed course, 39.8% had >4 infusions without a 90-day gap between administrations with a median of 8 administrations within 12 months. Of the nearly 38% of patients that did not receive a complete course of rituximab, 33% received a single infusion. The use of rescue medications in the period prior to rituximab treatment was compared to the period after treatment (defined as 60 days to 12-months post final rituximab infusion). Use of blood transfusions was observed to increase from 9.0% to 32.8%, while the use of ESA and IVIG were stable at approximately 2% and 4%, respectively.Conclusion With current wAIHA SOC, US HCRU remains high, posing a financial burden in terms of the rate of outpatient and emergency department visits, and inpatient stays. Rituximab is an early line therapy post-diagnosis, typically in combination with OCS. No changes in rescue treatments, and an increase in blood transfusions after rituximab infusion suggest treatment gaps remain. Analysis reveals heterogeneity in rituximab dosing patterns including high deviation from standard dosing practices; this variance from published expert opinion along with common use of rescue therapies suggests continued need for standardized guidelines and development of targeted therapies in wAIHA.

Abstract Introduction: Langerhans Cell Histiocytosis (LCH) is a rare histiocytic neoplasm, with approximately 90% of patients exhibiting alterations in the MAPK pathway. BRAF and MEK inhibitors have shown promising results in treating MAPK pathway-altered LCH. However, treatment interruptions due to adverse effects are common, and limited access to targeted therapies complicates care globally. While chemotherapy remains a common treatment for LCH, the outcomes for patients with MAPK pathway-altered LCH specifically receiving chemotherapy are still unclear. Methods: This multi-institutional international retrospective cohort study analyzed adult patients with BRAF or MAP2K (gain-of-function, GOF) altered LCH that were consecutively seen at Mayo Clinic, MN, AZ, and FL, University of Alabama at Birmingham, and Peking Union Medical College Hospital in China between 2007 and 2024. All patients who were included in this study were treated with chemotherapy, and those who have received targeted therapies or other modalities of frontline treatment were excluded. All time-to-event analyses were performed from the frontline therapy initiation, using the Kaplan-Meier method. Progression-free survival (PFS) included death and progression as events and was measured from the time of treatment initiation. The response assessment was based on radiographic evaluation in accordance with the most recent Histiocyte Society consensus guideline. Risk organ involvement was defined as disease involving the liver, spleen, bone marrow or central nervous system. Results: A total of 63 patients with LCH were included. The median age of the population was 34 years (range: 27-43), and males were 59%. Risk organ involvement was seen in 44 (70%) of patients and multisystemic LCH was observed in 48 (76%) patients. Among the 63 patients, BRAF gain-of-function alterations were observed in 79%, MAP2K mutations in 19%, and BRAF fusions in 2%. Of the BRAF gain-of-function alterations, BRAFV600E mutation was seen in 64% patients and [p.N486_P490del] deletion in 36%. There was no significant difference in organ involvement (single-system versus multisystem) or risk organ involvement between patients with BRAF and MAP2K alterations. All 63 patients received systemic therapy with chemotherapy and the regimens included Cytarabine (n=43), Cladribine (n=5), Etoposide (n=1), Methotrexate (n=1), and combined Methotrexate and Cytarabine (n=13). The median follow-up for the entire cohort was 3.8 years (range 3.2–5.0 years), with similar durations observed in patients with single-system vs. multisystem involvement and those with risk-organ involvement vs. no risk-organ involvement. The overall response rate among patients who received chemotherapy was 89%, with complete responses in 15 patients (26%) and partial responses in 40 patients (64.5%). The median progression-free survival (PFS) for the entire cohort was 2.61 years (95% CI: 2.06–NA). The 3-year PFS did not significantly differ based on risk organ involvement: 50.6% (95% CI: 36–71) for patients with risk organ involvement versus 47.4% (95% CI: 28–81) for those without, p=0.52. Patients with single-system disease experienced longer PFS compared to those with multisystem disease; the 3-year PFS was 80.2% (95% CI: 58.7–100) versus 42.2% (95% CI: 29–61.5), respectively, p=0.04. Conclusions: Our results indicate that LCH patients with a GOF alteration in the MAPK pathway derive significant clinical benefit from chemotherapy, achieving deep and durable responses. Therefore, patients should not be excluded from treatment due to the absence of targeted therapies, as they can still attain substantial responses and clinical improvement with chemotherapy, even in the context of MAPK pathway-driven disease.