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Abstract BACKGROUND Children exposed to poverty during frontline therapy for B-lymphoblastic leukemia (B-ALL) experience inferior overall survival (OS), higher relapse rates, and earlier relapse than their peers. Real-world data describing treatment patterns and survival after relapse of B-ALL are lacking and, as a result, the impact of poverty following relapse remains poorly understood. This study evaluated the association of poverty exposure, proxied by insurance status, with post-relapse survival and explored potential explanatory mechanisms, including disease biology and therapy utilization. METHODS Data from Retrospective Study of Contemporary Approaches to First Relapse in B-ALL (ReCALL-1), a multicenter retrospective cohort study of patients < 30 years old with first relapse of B-ALL from 2018-2022, were used. The primary exposure was poverty, proxied by insurance status at relapse, categorized as any private insurance (poverty unexposed), public-only insurance (poverty exposed), no insurance, or unknown; the latter two were not included in comparative analyses due to low numbers. The primary outcome was OS. Secondary outcomes included measurable residual disease (MRD)-negative second complete remission (CR2), relapse after CR2, non-relapse mortality (NRM), with relapse treated as a competing risk, and receipt of cellular and immunotherapies. RESULTS Among 462 patients from 31 U.S. institutions, 41% had private insurance, 47% public-only insurance, 2% no insurance, and 10% unknown. Compared to privately insured patients, those with public insurance were more likely to be Hispanic (56% vs 19%, p<0.001) and have non-English preferred language (30% vs 6%, p<0.001). Non-Hispanic Black patients comprised 9% of those with public insurance and 5% of those with private insurance. Disease characteristics that differed by insurance status at relapse included: Ph-like cytogenetics, more common in publicly insured (23% vs 11%, p=0.002), and infant ALL, more common in privately insured patients (10% vs 5%, p=0.06). Publicly insured patients were more likely to have experienced major organ toxicity during frontline therapy (32% vs 18%, p=0.002) and very early relapse (<18m from diagnosis: 43% vs 32%, p=0.07). Other demographic and disease characteristics, including relapse site and cytomolecular features, were similar by insurance status. With a median follow-up of 51 months from relapse, 4-year OS differed by insurance (log-rank p=0.02): 72% for private vs 56% for public insurance. After adjusting for race/ethnicity and language, public insurance increased the hazard of death by 65% (aHR 1.65 95% CI 1.13-2.42, p=0.004). The association remained, but was attenuated, when unbalanced disease characteristics (time to relapse, infant ALL, Ph-like cytogenetics, prior organ dysfunction) were included (aHR 1.47, 95% CI 1.00-2.16, p=0.05). Rates of MRD-negative CR2 were similar (85% public vs 87% private). However, point estimates suggested publicly insured patients had more relapse after CR2 (44% vs 37%, p=0.42) and higher 4-year NRM (16% vs 10%, Gray's test p=0.12), though neither were significant. Treatment choices for first relapse did not differ by insurance status. Utilization of intensive reinduction (91% vs 95% for public vs private, p=0.18) and clinical trial enrollment (22% for both) were similar. Receipt of cellular therapies did not significantly differ by public vs private insurance: Chimeric Antigen Receptor-T cells (CAR-T: 38% vs 34%, p=0.41), hematopoietic cell transplant (HCT: 42% vs 47%, p=0.32). Use of blinatumomab trended higher in those with private insurance (36% vs 45%, p=0.07), and inotuzumab was similar (11% for both, p=0.87). Median time from relapse to CAR-T or HCT was comparable (to CAR-T 102 vs 100 days, to HCT 117 vs 114 days). CONCLUSION This is the first study to demonstrate differential survival by insurance type, a proxy for poverty exposure, after first relapse of B-ALL. In this large multicenter, real-world cohort, patients with public insurance had worse OS after relapse than those with private insurance. The survival disparity existed independent of other SDOH factors. Adjustment for known high-risk disease features only partially accounted for the observed differences in OS. In contrast, rates of MRD-negative CR2 and receipt of CAR-T and HCT were comparable across insurance types. These findings highlight the urgent need to identify effective interventions that will ensure equitable outcomes after relapse.

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: Waldenström Macroglobulinemia (WM) is an indolent B-cell lymphoplasmacytic lymphoma characterized by heterogeneous clinical behavior and variable progression patterns. While MYD88L265Pand CXCR4 mutations are frequently encountered, the broader genomic landscape underpinning disease evolution from smoldering WM (SWM) to active treatment naïve WM (TNWM), post-treatment remission WM (PTWM) and relapsed/refractory WM (RRWM) remains incompletely understood. To address this gap, we studied the mutational profile across the WM disease spectrum. Methods We performed whole exome sequencing (WES) on sorted (CD19 and/or CD138) bone marrow aspirates of patients across the spectrum of WM. Paired germline sequencing was performed using peripheral blood mononuclear cells (PBMCs). Unique molecular identifier-tagged reads were deduplicated using fgbio. WES data were processed through the Sarek pipeline. Any valid mutation from either caller, present on the index or in COSMIC was considered. Copy number variations (CNVs) were inferred via CNVkit and summarized at the gene and arm level (≥50% directional change). Mean tumor and germline coverages were 174X and 74X, respectively. Fisher's exact test was used for enrichment and hypergeometric test for overlap analyses. Results A total of 78 high-quality WM samples [15 SWM, 44 TNWM, 13 RRWM, 6 PTWM in remission (all with paired TNWM samples)] were sequenced. The mutational burden varied across disease states (median SWM=1.48 m/Mb, TNWM= 1.12 m/Mb, RRWM = 1.59 m/Mb; p=0.04). Density plots revealed bimodal VAF distributions, consistent with clonal and subclonal architectures in WM. Overall, 77% harbored a MYD88L265P mutation by WES. Among MYD88^WT^ patients, AS-PCR identified MYD88L265P in 4 (25%) not detected by WES, raising the overall mutation rate to 83%, plus one with a non-L265P variant. Low VAF (<10%) MYD88L265P mutations were seen in 6% of matched PBMCs. MYD88 copy number gain was seen in 2 (17%) MYD88WT TNWM patients. CXCR4 mutations were detected in 38% by WES (18% each frameshift and nonsense). Two (4%) CXCR4WT samples were positive by PCR, giving an overall rate of 40%. CXCR4 mutations were more frequent in RRWM (54%) vs TNWM (41%) and SWM (27%), though not statistically significant. Del(6q) and Del(17p) were noted in 16% (n=12) and 7% (n=5) of patient samples. All del(17p) events were noted in TNWM samples and in one paired post-treatment sample. Overall rate of del(17p) and TP53 mutation was 14% (n=11). RRWM was enriched in ARID1A alterations, a gene implicated in chromatin remodeling and plasma cell identity formation [38% (n=5) in RRWM vs. 11% (n=6) in TNWM and SWM], compared to TNWM and SWM p=0.03]. Alterations in NF-kB signaling gene MAP3K14 were more frequent in RRWM (31%, n=4) compared to the SWM/TNWM cohort [(7%), n=4; p=0.036). We noted an enrichment in the TP53 mutation rate in RRMW [n=3, 23%, OR 7.7] compared to TNWM and SWM cohorts [n=2, 4%), p=0.04]. Mutations in FUBP1 (p=0.03), an upstream regulator of MYC, and CARD11 (p=.043), a B-cell signaling scaffold protein were also more frequent in RRWM. Notably, none of the SWM patients harbored an ARID1A alteration (p=0.057), suggesting acquisition during progression. CNV analysis revealed that MAP3K14 amplifications were frequent in RRWM (n=3, 23%) compared to SWM/TNWM [n=2 (4%), p=0.043]. RRWM samples showed increased CD79B mutations (15%) and significant overlap with the C5/ABC diffuse large B-cell lymphoma (DLBCL) genomic cluster (Chapuy et al., Nat. Med. 2018; p=0.007). This overlap remained significant after excluding MYD88L265P (p=0.019), driven by CARD11, CD79B, and IGLL5, suggesting that a subset of RRWM may acquire an ABC DLBCL-like mutational profile. These findings warrant further investigation using transcriptional profiling. Conclusions This study provides a genomic map of WM evolution from precursor to relapse. Increasing somatic mutations and CNVs support stepwise progression. Enrichment of genes involved in chromatin remodeling and plasma cell identity formation (ARID1A), NF-kB or BCR signaling (CARD11, MAP3K14), and DNA damage repair (TP53) were noted at relapse, all of which have been shown to drive aggressive disease biology. Emergence of an ABC-like genotype in a proportion of RRWM indicates convergent evolution with aggressive lymphomas. Transcriptional studies are warranted to explore this further.

Abstract Background The advent of T-cell redirecting bispecific antibodies (BsAb) therapies has led to deeper and durable responses in patients with RRMM. We sought to identify adverse prognostic factors for patients treated with BsAbs. We also explored whether the extent of the non-involved free light chain (niFLC) suppression could serve as a potential marker of T-cell engager activity, given the binding of BsAbs to both clonal and non-clonal plasma cells and the short half-life of serum FLCs. Methods We conducted a retrospective analysis of RRMM patients treated with BsAbs between 2022 and 2025 at Mayo Clinic. Baseline lymphocyte counts and ferritin levels were collected and analyzed both as continuous and dichotomized variables. niFLC was considered undetectable if it was below the quantification limit, while the magnitude of niFLC suppression was defined as the absolute difference between niFLC levels and the lower normal range threshold at the time of best response. In addition to standard statistical methods, interaction models were used to explore the potential effect modification of niFLC dynamics. Results A total of 158 patients were included (teclistamab, 73%; talquetamab, 15%; elranatamab, 12%). The median follow-up is 12 months (range 0 to 28 months). Median age is 71(range 65 to 77); 52% (n=85) were triple-class refractory (refractory to an IMiD, a PI, and CD38-antibody) and 37% (n=60) were penta-refractory. CAR-T therapy was previously administered in 28% (n = 46) patients, and 70% had undergone an autologous stem cell transplant. One or more high risk cytogenetic features including del(17p), gain(1q), amp(1q), del(1p), t(4;14), and t(14;16) were present in 57% (n = 90) of patients. Extramedullary disease was noted at the time of BsAb therapy in 44% of patients. Utilizing IMWG response criteria, 68% (n=107) achieved ≥partial response, and 64% (n=101) had ≥ very good partial response (VGPR) as best response following BsAb treatment. The median time to best response was 1.6 months (range 0.7 to 4.4). On multivariable analyses, baseline immunoparesis (HR 1.96; 95% CI, 1.08–3.58; p = 0.0274) and elevated ferritin levels (≥ 337 ng/mL) (HR 1.96; 95% CI, 1.24–3.09; p = 0.0041) were independent predictors of inferior progression-free survival (PFS). Lower baseline absolute lymphocyte count, analyzed as a continuous variable, was significantly associated with shorter PFS in the univariable analysis (HR 0.66; 95% CI, 0.44–0.98; p = 0.04), but not in the multivariable model (HR 0.75; 95% CI, 0.5–1.13; p = 0.166). Detectable niFLC at the time of best response was also significantly associated with inferior PFS (HR 1.80; 95% CI, 1.14–2.84; p = 0.0114). However, the magnitude of niFLC suppression did not show a statistically significant impact on PFS. The median PFS was 5 months in patients with high ferritin levels (≥ 337 ng/mL) and 21 months in those with lower ferritin (p = < 0.005). Similarly, PFS was 8 months in those with immunoparesis and not reached in those without baseline immunoparesis (p = 0.008). At the time of best response, detectable niFLC was associated with a trend toward shorter PFS (5 vs. 13 months), although the difference was not significant (p = 0.06). Logistic regression analysis supported these findings, with higher ferritin levels (OR 2.28; 95% CI: 1.09–4.96; p = 0.032) and the presence of baseline immunoparesis (OR 2.60; 95% CI: 1.18–6; p = 0.02) being significantly associated with higher likelihood of progression or death. Patients with deep reduction in niFLC achieved a VGPR or better. In contrast, those with detectable or higher-than-normal niFLC levels did worse in terms of response. Notably, niFLC became undetectable in 72% of patients with ≥VGPR (73/101), compared to 52% of those who did not achieve this level of response (30/57). Conclusion Among RRMM patients treated with BsAbs, baseline immunoparesis and elevated ferritin levels emerged as independent predictors of worse PFS. While failure to induce a deep reduction in niFLC at the time of best response suggests a tumor extrinsic mechanism of resistance, and it is also linked to inferior outcomes.

Abstract Background: Undetectable minimal residual disease (uMRD) at the end of first-line (1L) treatment (tx) with 12-cycles (C) of venetoclax (ven) + obinutuzumab (obin) is associated with prolonged survival for patients (pts) with chronic lymphocytic leukemia (CLL, Al Sawaf et al. Blood 2024). We hypothesized that MRD may be used to guide ven tx duration for CLL pts treated with 1L ven + obin. Methods In this ongoing phase II, multicenter investigator-initiated study, pts receive ven + obin for 1L tx of CLL/small lymphocytic lymphoma (NCT04447768). Eligible pts have a CIRS score of ≤ 6 and require 1L tx per iwCLL 2018 criteria. Pts receive 6 x 28-day C of obin and initiate ven on C1D22. Ven duration is dependent on peripheral blood (PB) MRD status measured by ClonoSeq next generation sequencing. At C7, pts undergo MRD testing. Pts uMRD6 (<10-6 sensitivity) at both C7 and C9 dc ven after 9C and enter treatment free observation (TFO). Pts with detectable MRD6 (dMRD6) at C7 undergo repeat testing at C12; if uMRD5 (<10-5) pts dc ven after 12C. Pts dMRD5 at C12 continue ven for an additional 12C (24C total) prior to TFO. The primary study endpoint is 36-month PFS; the unpromising rate is set at 82% and will be tested by a nonparametric survival estimate (Kaplan-Meier) along with Greenwood's formula. RNA sequencing (RNAseq) was performed pre-tx on patient bone marrow (or PB if inaspirable) samples. Unsupervised clustering using machine learning (ML) based on consensus nonnegative matrix factorization was applied to the most variable genes to identify patient clusters. Clusters were then biologically characterized by gene signature scoring using xCell algorithm and assessed for association with uMRD6 at C7 using a Fisher exact test. Results Baseline characteristics of the 100 pts who initiated tx include: median age 58 (range 34-81); 71% male, 90% White, 3% Black and 2% Asian, 5% unknown; 49% IGHV unmutated, 16% TP53 mutation, 8% del17p (previously reported, Roeker et al. ASH 2024). 3 pts stopped tx prior to C7. 51 pts were uMRD6 at C7 and C9 and completed 9C ven. 46 pts were dMRD6 at C7; 6 pts stopped tx outside of meeting protocol requirements and are included in intention-to-treat (IIT) analyses. 32 pts were uMRD5 at C12 and dc ven after 12C. 8 pts were dMRD5 at C12: 5 continued ven per protocol (3 completed 24C ven, 2 ongoing tx as of 15 May 2025 data cut-off), 2 stopped ven outside of protocol requirements (included in ITT analyses), and 1 pt withdrew consent. At a median follow-up of 35 months, the 36-month PFS rate is 90% (95% CI: 84%, 97%). 36-month OS is 94% (95% CI: 89%, 99%). Landmark PFS analyses from EOT were performed for pts who dc ven after 9C due to uMRD6 (24-month PFS 92%, 95% CI: 85, 100%, n=51) and pts who dc ven after 12C due to uMRD5 (24-month PFS 96%, 95% CI: 89%, 100%, n=32). For pts dc ven after 9C due to uMRD6 (n=51), PB MRD at 12C after EOT was: 59% uMRD6, 12% uMRD5, 4% uMRD4, 8% dMRD4 and 18% unknown. For pts dc ven after 12C due to uMRD5 status (n=32), PB MRD at 12C following EOT was: 9% uMRD6, 31% uMRD5, 9% uMRD4, 25% dMRD4 and 25% unknown. Of the 5 pts with PD, 3 pts have had Richter Transformation to diffuse large B-cell lymphoma (all uMRD6 at C7 / 9). Two pts with PD have not met iwCLL tx criteria and remain in TFO. Five pts have died (suicide, West Nile encephalitis, COVID-19, complications of glioblastoma and RT DLBCL). Five C12 dMRD5 pts who continued ven have reached C18, with only 1 of 5 converting to uMRD5 in PB. However, all 5 maintained uMRD4 (<10-4). Of 3 pts that completed 24C ven (2 ongoing tx), 1 pt had deepening of response to uMRD5 at C24, and 2 pts had rising MRD on ven (dMRD4 at C24). To investigate patient heterogeneity at baseline and its relationship to uMRD, we clustered patient RNAseq samples into two clusters using artificial intelligence/ML based unsupervised clustering. The clusters associated with uMRD6 at C7 (Fisher exact test p=0.038), and patients in the cluster with significantly higher immunosuppressive cells such as T-regulatory cells and M2 macrophages were less likely to be uMRD6 at C7. Conclusions At a median follow-up of 35 months, the estimated 36-month PFS rate is 90%. RNAseq transcriptomic analyses identified that patients with enriched immunosuppressive cells were less likely to be uMRD6 at C7. Additional follow-up is required to confirm the durability of this MRD-guided ven + obin tx strategy.

Abstract Introduction Matrix assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF MS, mass fix) represents a sensitive and specific alternative to electrophoresis and immunofixation for quantifying monoclonal antibodies in patients with multiple myeloma (MM). Here we report real world data utilizing mass fix and compare its test characteristics to a gold standard of minimal residual disease (MRD) quantification utilizing next generation sequencing (NGS). Results All patients who had obtained mass fix and MRD testing by NGS as part of routine clinical care at Mayo Clinic Arizona were screened for this study. In total, 343 pairs of mass fix and NGS testing within 50 days of each other representing 269 unique patients with secretory multiple myeloma were included. Cases of IgD myeloma and those secreting only free light chains were excluded. The threshold of MRD negativity was set at <1x10-6(less than one malignant cell in one million) and was assessed utilizing the clonoSEQ NGS assay. The median age of our cohort is 67, it is 44% female, and primarily Caucasian (88%) with a median follow-up of 7.3 months from testing date. FISH testing was performed in 96% of patients and at least one high risk cytogenetic abnormality (t(4;14), t(14;16), del(17p)) was found in 23% of patients. The t(11;14) translocation was found in 18%, +1q in 26%, and amp(1q) in 6% of evaluated patients. Notably, patients at all stages of treatment or disease status were included in our study. Induction treatment for newly diagnosed MM (NDMM, 133 patients) was mostly daratumumab-containing quadruplet (63%) or triplet drug regimens (36%). Autologous stem cell transplant (ASCT) was performed in 79% of NDMM and 80% of relapsed refractory MM patients (RRMM, 138 patients). Of the RRMM patients, 94% were triple class exposed and 41% were penta-exposed. Overall agreement between NGS and mass fix results was 65% (Kappa statistic: 0.31, 95% CI: 0.22 – 0.41). Most discordant cases were instances of mass fix negativity and NGS positivity (n = 89, 26% of all testing pairs). Only 7% of cases (n= 30) represented mass fix positivity with NGS negativity. Mass fix was able to define therapeutic monoclonal antibody by its unique mass spectrometric profile in 38 of the total test pairs that otherwise would have been considered disease associated. Of the 30 patients testing mass fix positive and NGS negative, 25 (83%) represent post-treatment evaluations. Thus, ongoing mass fix positivity represents monoclonal protein washout kinetics. All of these cases are noted to have declining mass fix levels at last follow up and 40% had undetectable monoclonal protein. The discordant mass fix negative and NGS positive cases were evenly distributed across NDMM and RRMM representing 32% and 20% of test pairs in each group, respectively. Amongst these 89 total test pairs, the median abundance of malignant clone by NGS was 10 cells per million (range 0.06 to 297,895). This is in contrast to 136 concordantly positive test pairs where the median was 2960 cells per million (range 0.353 to 1,017,360). The median monoclonal protein identified (in those with a quantifiable M-spike) was 0.30 g/dL. Receiver operating characteristic curves using NGS as gold standard found the optimal mass fix threshold to be 0.010g/dL (specificity 95%, sensitivity 51%) for NDMM patients and 0.046g/dL (specificity 84%, sensitivity 55%) in RRMM patients. Conclusions Development of a MRD assay based on peripheral blood sampling could greatly improve the ease and speed of obtaining these assessments. Our experience highlights both the great potential and need to identify appropriate clinical contexts for optimal use of mass fix MRD assessment. A significant limitation of our data is that low level monoclonal proteins without a previous reference above a threshold of 20mg/dL are not reported as positive and may account for a significant fraction of discordant cases reported here. All clonoSEQ NGS samples are referenced to a positive control greatly increasing specificity. The currently reported dataset is biased against mass fix as not all test pairs reported are known to have a positive reference. Further delineation of these cases is forthcoming. Previous reports have also noted that mass fix test characteristics can improve with time from treatment which will be assessed with future follow-up.

Abstract Background Erdheim-Chester disease (ECD) is a very rare histiocytic/dendritic cell neoplasm driven by mutations in genes of the MAPK pathway, such as BRAFV600E. Its clinical presentation is highly heterogeneous, and data on its natural history and prognosis in large cohorts are limited. Here, we investigated the clinical features and outcomes of a very large international cohort of patients with ECD. Methods We included patients diagnosed with ECD between 1990 and 2024, followed at 13 referral centers in Europe and the Americas, who had available clinical and follow-up data (with a minimal follow-up duration of 6 months). We analyzed the clinical presentation, treatment, and outcomes, and sought to identify predictors of survival. The study was conceived and conducted within the ECD Global Alliance network. Results A total of 1044 patients with ECD were included. The median age at diagnosis was 57 years (IQR 46-66), and the male-to-female ratio was 2.2. The median age at the first ECD manifestation was 55 years (IQR 43-65). Bone pain (20%), kidney and urinary manifestations (18%), and diabetes insipidus (11%) were the most frequently reported initial features. The median number of involved sites was 4 (IQR 3-6), most frequently including the long bones (n=768, 74%), the retroperitoneum, (n=605, 58%), the periaortic space (n=449, 43%), the CNS (n=368, 35%), the facial/orbit area (n=347, 33%), and the heart (n=334, 32%). Among 955 tested patients, 572 (60%) harbored the BRAFV600E mutation, and 86 (9%) a MAP2K1 mutation. Clinical and/or pathology overlap with Langerhans cell histiocytosis and Rosai-Dorfman-Destombes disease was observed in 115 (11%) and 42 (4%) patients, respectively. Clonal hematopoiesis was identified in 217 of 556 tested patients (39%), and a history of second cancer was reported in 272 patients (26%), including both solid (14%) and hematologic (12%) neoplasms. First-line treatment mostly included interferon-alpha (n=342, 33%) and BRAFi/MEKi (n=314, 30%), while second- and third-line treatments mostly consisted of BRAFi/MEKi (55% and 63%, respectively). After a median follow-up of 51 months (IQR 20-89), 222 (21%) patients died; causes of death were related to ECD in almost half of the patients (n=101). A multivariate Cox regression model identified the following factors as significantly associated with increased mortality: older age (HR 1.058, 95%CI 1.033-1.083; p<0.001), pseudo-neurodegenerative lesions (HR 2.280, 95%CI 1.337-3.891; p=0.022), elevated CRP at diagnosis (HR 2.574, 95%CI 1.024-6.467; p=0.044), and second hematologic cancer (HR 2.639, 95%CI 1.602-4.348; p<0.001). Conclusions Despite advances in treatments over the last decade, ECD remains associated with substantial mortality. Age, neurodegenerative lesions, elevated CRP levels, and the presence of a second hematologic cancer are the main predictors of poor survival.

Abstract Background People of color (POC) and patients (pts) from socioeconomically disadvantaged backgrounds often face financial, geographic, and systemic barriers, limiting access to advanced treatments (Tx). A survey was conducted to better understand how race and socioeconomic factors affect pt awareness of and access to bispecific antibodies (BsAbs) and chimeric antigen receptor T-cell (CAR-T) therapies in the US. Methods This study used 30-minute online surveys (March-June 2024) of 1301 pts with RRMM across 7 countries. Results were analyzed using descriptive statistics and χ2 tests, with P<.01 for all comparisons unless specified; these are presented for the 305 US pts. Results The 305 US pts identified as White (n=227) or POC (n=78), which included Black/African American (n=42); Hispanic/Latino (n=24); and Asian, Pacific Islander, or Arab (n=12). Pts self-reported as having easy/very easy (n=92), neutral (n=117), or difficult/very difficult (n=96) financial burden, being employed (n=142) or not employed (n=163), and having ≤secondary (n=72) or >secondary education (n=232). In the US, more POC reported difficult/very difficult financial circumstances than White pts (47% v 26%). Fewer POC stated they were in remission (28% v 49%) or had consulted an MM specialist (41% v 59%, P=.028), even after adjusting for self-reported good financial circumstances. Among pts with neutral-very easy financial situations, stated remission rates were still lower for POC (n=41) v White pts (n=168) (34% v 59%, P=.026), as were MM specialist visits (41% v 66%, P=.022). Pts with >secondary education (n=232) and those who were employed (n=142) had higher stated remission rates than those with ≤secondary education (n=72) and those who were not employed (n=163) (47% v 32%, P=.025 and 66% v 24%, respectively). Physical burden was experienced by more POC v White pts (91% v 66%), even after adjusting for financial circumstances (90% v 65%, P=.011, for POC v White pts in neutral/very easy financial circumstances). POC and pts with difficult financial circumstances (including POC and White pts) perceived their Tx as less effective than expected across several measures, including emotional and mental health, overall impact on daily life, side effects, Tx costs, and the effect of Tx schedule on everyday life. Awareness of BsAbs and CAR-T was lower among POC v White pts (BsAb: 49% v 77%; CAR-T: 67% v 81%, P=.032), pts who were not employed v employed (BsAb: 63% v 77%, P=.015; CAR-T: 74% v 81%, P=.046), pts with ≤secondary v >secondary education (CAR-T only: 75% v 78%, P=.021), and those in difficult/very difficult v easy/very easy financial circumstances (BsAb: 54% v 90%; CAR-T: 64% v 90%). Among pts aware of BsAbs, 18% (n=39) reported that the Tx was offered by their physician. More employed v not employed pts were offered BsAbs (26% v 10%). There were non-statistically significant differences in the proportion offered BsAbs based on race (POC 8% v White pts 21%, P=.065), financial status (difficult/very difficult 15% v easy/very easy 23%, P=.288), or level of education (≤secondary education 15% v >secondary education 20%, P=.428). Among pts aware of CAR-T, 24% (n=56) reported that the Tx was offered by their physician. Fewer POC than White pts were offered CAR-T (6% v 29%), even after adjusting for financial status (10% v 32%, P=.04 for pts with good finances). Among those aware of CAR-T, fewer pts not employed v employed (12% v 37%), pts with ≤secondary v >secondary education (11% v 27%, P=.015), and pts with financial burden (difficult/very difficult 10% v easy/very easy 31%) were offered CAR-T. Overall, fewer POC received CAR-T than White pts (1.3% v 10.7%, P=.03). Additionally, more employed v not employed pts received CAR-T (15.2% v 2.5%) or BsAbs (23.5% v 4.4%). Conclusions Consistent with existing health equity inequalities (Chen et al, Int J Equity Health, 2025), POC and pts in poorer socioeconomic circumstances had less awareness of and access to BsAbs and CAR-T. To address these disparities, partnering with trusted messengers and pt advocacy groups supporting Black, Hispanic, and Asian pts and those with socioeconomic hardships can enhance healthcare infrastructure, support initiatives to help pts navigate advanced Tx options, and develop pt- and physician-targeted education. These strategies can improve awareness and promote Tx equity, ultimately ensuring that all pts with MM receive optimal care regardless of their socioeconomic status and race.

Abstract Introduction: The 5th edition of the World Health Organization (WHO-5) and the International Consensus Classification (ICC) recognize TP53-mutated myeloid neoplasms (MN) as a distinct disease category, reflecting their uniformly dismal outcomes and the urgent need for novel therapeutic strategies. However, both classifications do not include TP53-non-mutatedcases with 17p (TP53 locus) loss, despite emerging evidence that such lesions may confer biologic and clinical behaviour similar to TP53-mutated disease. Our preliminary work demonstrated that 17p loss without TP53 mutation in therapy-related MN (t-MN) is associated with poor survival, approximating that of TP53-mutated cases (Hiwase et al., Blood 2023). Yet, the true frequency and prognostic impact of 17p loss in TP53-non-mutatedMN remains largely undefined, representing a critical gap in current disease classification and risk stratification. To address this critical knowledge gap, we aim to define the frequency and prognostic impact of 17p loss across the TP53 locus in TP53-non-mutatedMN, including both myelodysplastic syndromes (MDS) and acute myeloid leukemia (AML). Finally, we compared the survival outcomes of this cohort with those of TP53-mutated MN. Method This international collaborative study was designed to evaluate the frequency and prognostic significance of 17p loss in TP53-non-mutatedMN. We included MDS and AML patients who had conventional cytogenetic analysis and did not have TP53-mutation(TP53-non-mutated)by next-generation sequencing (NGS) with sensitivity of ≥2% variant allele frequency (VAF). These cases were systematically reviewed for 17p loss using karyotype data, and where available, fluorescence in situ hybridization (FISH) and SNP array were used to confirm the 17p/TP53 locus deletion. To assess the clinical impact, we compared the overall survival (OS) of TP53-non-mutatedcases with 17p loss against a reference cohort of TP53-mutated MNs with VAF ≥2% (n=603) (South Australian Myeloid Neoplasm Registry (SA-MN) and Mayo Clinic, Rochester, USA). Results We screened 8876 cases of TP53-non-mutatedMN cases across an international cohort comprising the SA-MN (Australia; n=1046), the Düsseldorf MDS Registry (Germany; n=545), Munich Leukaemia Laboratory (Germany; n=5785), and King's College Hospital, (United Kingdom; n=1500). We identified 85 (1%) cases with 17p loss across the TP53 locus, representing the largest international cohort to define the frequency of 17p loss in TP53-non-mutatedMN. Complex karyotype was present in 40% (n=34) of cases, frequently co-occurring with del(5q) and del(7q). Clinically, most cases were MDS (n=69; 81.2%), including MDS-IB1 (n=19), MDS-IB2 (n=15), MDS-LB (n=16), t-MDS (n=8), MDS/MPN overlap (n=3), CCUS (n=1), or unspecified MDS (n=7), while AML accounted for 16 cases (18.8%). Among evaluable cases, the majority were male (78.6%) and had a median age of 66.5 years at diagnosis. The most frequent cytogenetic abnormality causing 17p loss across the TP53 locus was monosomy 17 (39%), followed by i(17q) (29%), add (17p) (14%), derivative chromosome 17 from unbalanced translocations (11%), and del(17p) (7%). The majority represented de-novo MN (60%), with t-MN in 40% of cases. Median OS of evaluable 17p loss in TP53-non-mutatedcases (n=28; 10.2 months) was poor and comparable to TP53-mutated MN (n=603, 8.7 months; P=0.42). Similarly, survival censored for a stem cell transplant was also equally poor (10.4 vs. 8.7 months; P=0.34) in both groups. Complex vs. non-complex karyotype (7.7 vs. 10.7 months; P=0.68) did not significantly influence the OS of 17p loss. Median OS of MDS cases tended to be longer compared to AML (16.7 vs. 9.1 months; P=0.11), although not statistically significant. Conclusion: In this largest international study, 1% of TP53-non-mutatedMNs harbor 17p loss across the TP53 locus, displaying poor survival equivalent to TP53-mutated disease. Exclusive reliance on NGS to identify TP53 mutations misses these high-risk structural TP53 abnormalities, which are readily detectable by conventional karyotyping. These findings provide compelling evidence to include TP53-non-mutatedcases with 17p loss in the next revision of the classifications, enabling accurate prognostication and equitable access to curative and clinical trials of novel therapies.

Abstract Introduction High disease burden has been associated with reduced efficacy and resistance to BCMA CAR-T therapy in relapsed/refractory multiple myeloma (van de Donk, Blood Cancer Discov, 2021; Martin, J Clin Oncol, 2023). While CAR-T cells can still induce remissions in these patients, outcomes are often worse compared to those with a lower tumor load. In addition to tumor debulking, novel strategies are needed to improve responses and durability of CAR-T cells in these patients. We previously demonstrated that immunomodulatory drugs (IMiDs) improve in vivo responses to T cell engagers (TCE) in high tumor burden setting by boosting T cell activation (Meermeier, Blood Cancer Discov, 2021; Meermeier, Blood, in press). IMiDs have also been shown to promote CAR-T cell antitumor activity in moderate to low tumor burden conditions (E:T ratios ≥1:1) (Yan, J Transl Med, 2024; Wang, Clin Cancer Res, 2018). We hypothesized that combining CAR-T with an IMiD may improve CAR-T function in high tumor burden setting. Methods We generated murine anti-BCMA.CD28 CAR-T cells by transducing Con-A stimulated T lymphocytes extracted from hCRBN+ mice, expanded with IL-7 and IL-15 to express a memory phenotype. We then tested CAR-T, or untransduced T cells, ability to kill in vitro IMiD resistant, hCRBN-, Vk*MYC myeloma cells in low (E:T 1:1) and high (E:T 1:≥5) tumor burden, with and without pomalidomide (POM) and IL-2. Tumor killing and T cell phenotype were assessed by flow cytometry at 24-72 hours. This controlled system allowed us to focus on putative CAR-T cell intrinsic impact of IMiDs. Results In low tumor burden setting, CAR-T effectively killed tumor at all timepoints, with or without POM; addition of POM increased T cell number, confirming that IMiDs support CAR-T expansion. Unexpectedly, in high tumor burden (E:T 1:≥5), adding POM diminished CAR-T mediated killing over time and hindered CAR-T expansion. Moreover, combination with POM decreased granzyme B, while increased expression of exhaustion marker TOX, ultimately leading to CAR-T apoptosis. This phenotype was further intensified by increasing concentrations of IL-2 in the co-culture, resulting in increased expression of CAR-T expressing LAG-3, TIM-3, and TIGIT, suggesting that CAR-T cells may be overactivated by POM and IL-2. Our manufacturing protocol preferentially expands memory CAR-T cells, favored for their durability, proliferative capacity, and considered to rapidly expand upon encounter with antigen. We reasoned that CAR-T are already sufficiently activated by CAR/CD3z stimulation, especially with a CD28 co-stimulatory domain (versus 4-1BB), known promote rapid, effector memory expansion upon activation (Kawalekar, Immunity, 2016). Activation is intensified by high tumor burden where antigen is highly abundant. We hypothesize under intense, sustained antigen exposure in high tumor burden, the addition of POM paradoxically triggers activation induced cell death (AICD). Conclusion Although it is well accepted that POM can boost T cell function and proliferation, we demonstrate that in high tumor burden, POM drives memory CAR-T to increase exhaustion markers and apoptotic cell death by overactivation, further exacerbated by exogenous IL-2. Interestingly, we have shown opposite effects of preclinically combining POM with TCE therapy in high tumor burden, based on dosing schedule. Concomitant administration transiently boosted T cell activation improving response rates but rapidly led to T cell exhaustion (Meermeier, Blood Cancer Discov, 2021). In contrast, IMiD pretreatment in the context of step-up dosing TCE administration improved T cell fitness and longevity (Meermeier, Blood, in press).The effect of IMiDs on T cells transcriptional program is likely to differ depending on differentiation stage and mode of T cell activation. Memory cells have more sensitive TCR signaling (Kumar, Immunity, 2011) and can be hyperactivated, leading to AICD. Thus, timing of IMiD administration, when tumor burden is low, may be a critical consideration as different T cell subsets require different levels of activation. Conversely, TCE provide only initial activation via TCR signaling, therefore POM may help in providing necessary co-stimulation in this setting. Future studies will evaluate different co-stimulatory domains and test in vivo effects of POM on CAR-T expansion and persistence, particularly as CAR-T cells diminish over time as tumor burden increases.