Citations

Abstract Background: Routine laboratory testing prior to chemotherapy administration is a cornerstone of hematology and oncology practice, aimed at ensuring the safety and efficacy of treatment. However, over time, standardized protocols may accumulate excessive or low-yield laboratory tests that offer limited value to clinical decision-making. In 2018, Mayo Clinic implemented standardized treatment protocols using Epic Beacon, an electronic health record module that integrates chemotherapy regimens with associated laboratory testing and clinical workflows. While well-intentioned, these protocols may include redundant tests that delay treatment initiation, increase patient discomfort and anxiety, and burden providers with excessive in-basket communications. We hypothesize that many laboratory tests currently embedded within prechemotherapy workflows are unnecessary and can be safely omitted without compromising patient care. Methods: We conducted a retrospective cohort study evaluating the pre-treatment laboratory testing associated with the R-CHOP (rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisone) chemotherapy protocol within the Mayo Clinic Rochester Division of Hematology. The study included 100 patients diagnosed with diffuse large B-cell lymphoma who received treatment between January 1 and December 31, 2024. Patients were excluded if they had incomplete medical records or received treatment outside the Mayo Clinic system. Data was abstracted via chart review using the Mayo Data Explorer. We analyzed complete blood counts (CBC), which include 12 individual tests, and comprehensive metabolic panels (CMP), which include 16 individual tests, obtained prior to each treatment cycle. For each cycle, we assessed the proportion of abnormal laboratory results and the proportion of abnormal results deemed clinically relevant which were defined as those that prompted a treatment delay, dose reduction or early termination of a cycle. Results: The cohort comprised 100 patients with a mean age of 68.2 years (SD 13.4) and a median age of 71 years (IQR 58.75–77); 57 were male and 43 were female. Based on available documentation, disease stage was classified as limited (stage I/II; n=41) or extensive (stage III/IV; n=59). A total of 519 R-CHOP cycles were administered, including 419 subsequent cycles (cycles 2–6). Fifteen patients discontinued treatment early, although none of the discontinuations were attributable to abnormalities in pre-infusion laboratory testing. Among the 419 subsequent cycles, 34 (8.1%) experienced treatment delays, with only one delay resulting from a clinically significant laboratory abnormality (hyperglycemia). Additionally, 23 patients underwent regimen modification (dose reduction or drug holding), but none of these were due to pre-infusion lab abnormalities.Of the 411 comprehensive metabolic panels (CMPs) obtained prior to these cycles, 315 (76.6%) contained at least one abnormal value, yielding a total of 786 abnormal results. Similarly, 405 of 415 (97.6%) complete blood counts (CBCs) demonstrated at least one abnormality, generating 2,696 abnormal values in total. Despite the high frequency of abnormal findings, the overall clinical yield was minimal, with only 1 of 826 pre-infusion laboratory panels (0.12%) leading to a modification in clinical management. Conclusion: Routine pre-infusion laboratory testing prior to R-CHOP chemotherapy frequently identifies abnormal results. However, these abnormalities are rarely clinically significant and do not affect treatment decisions. These findings support the reconsideration of current testing protocols to reduce unnecessary testing, enhance workflow efficiency, and minimize patient and provider burden without compromising safety.

Abstract Background: VEXAS (Vacuoles, E1 enzyme, X-linked, Autoinflammatory, Somatic) syndrome is a hemato-inflammatory syndrome due to somatic mutations in UBA1 gene. Recently, DNA methyltransferase inhibitors (specifically, 5-azacitidine) and allogeneic hematopoietic stem cell transplantation (alloHSCT) have shown success in eradication of UBA1 clones and clinical remission in some patients. This study aims to provide a comparative efficacy analysis between DNA methyltransferase inhibitors (hypomethylating agents/HMA) versus alloHSCT in VEXAS syndrome patients. Methods: This is a multi-center, retrospective cohort study. UBA1 mutations were detected either through next generation sequencing [NGS, limit of detection (LOD) 2% variant allele fraction (VAF)] or quantitative droplet digital polymerase chain reaction testing (ddPCR, LOD 0.5% VAF). Outcomes of interest included overall survival (OS), glucocorticoid (GC) discontinuation/reduction in dose (prednisone < 10 mg/day), and complete molecular remission (CMR, defined as an undetectable UBA1 mutation). OS was calculated from the time of diagnosis to death or last follow up. Cumulative incidence of reduction in GC dose was calculated using competing risk analysis after accounting for death. Results: Between years 2020-2024, a total of thirty-nine male patients (median age 67, range 45-82 years) with VEXAS syndrome, necessity for therapy (GC-refractory inflammation and/or progressive marrow failure/myeloid neoplasm) and confirmed UBA1 pathogenic mutation were included. Of these, 18 (46%) received HMA therapy (5-azacitidine 72% and Decitabine 28%) with a median of 4 (range 1-36) cycles. and 21 (54%) underwent alloHSCT (MSD=4, MUD=12, MMUD=2, and haploidentical=3), all received reduced intensity conditioning (Flu/Mel =11, Flu/Bu=5, Flu/Cy/TBI=2, and TBF=3) and 91% received post-transplant cyclophosphamide-based graft-versus-host disease (GVHD) prophylaxis (only 2 patients in alloHSCT group received prior HMA without benefit). Baseline characteristics were well-balanced between HMA and alloHSCT groups; age at treatment (median 70 vs 65 years, p=0.2), and UBA1 M41V subtype (25% vs 40%, p=0.3), cytogenetic and non-UBA1 mutation abnormalities (p=0.4), prior lines of therapy (median 4 in both, p=0.5), time from diagnosis to therapy (median 7 vs 13 months, p=0.15), indication for therapy (p=0.12), Charlson CI scores (median 4 in both, p=0.4) and HCI-CI scores (median 4 in both, p=0.7). At a median follow-up of 45 (range 7-57) months, 11 (28%) deaths (alloHSCT=3 and HMA=8) were documented. Causes of death included infections (n=6), cardiac disease (n=2), poor graft function (n=2) and pulmonary GVHD (n=1). Compared to HMA therapy, alloHSCT group demonstrated higher rates of GC discontinuation (52% vs 6%, p<0.01) and CMR (evaluable= 22, 100% vs 50%, p<0.01). In the HMA group, 50% (n=9) discontinued therapy after a median of 4 months due to infectious complications or lack of benefit. In the alloHSCT group, 3 (14%) patients developed grade 2 acute GVHD (1-skin, 2-gut, no grade 3-4 acute GVHD) and 4 (19%) experienced moderate to severe chronic GvHD (2-lung, 2-mucocutaneous). In multivariable analysis (MVA) adjusted for age and HCT-CI, alloHSCT was identified as the strongest favorable predictor for OS (HR 0.18 95% CI 0.04-0.8) with similar results when analysis was restricted to transplant-eligible patients < 75 years of age (HR 0.1, 95% CI 0.03-0.6). In a 6-month post-treatment landmark analysis (n=31 evaluable), patients who underwent alloHSCT (n=17) exhibited numerically superior GC discontiuation rate (29% vs 14%, p=0.049) and subsequent OS compared to HMA (2-years OS: 84% vs. 61%, median not reached vs 25 months, 95% CI: 0.06–1.7). In a post-treatment competing risk [GC dose reduction (death)] analysis alloHSCT was associated with superior 2-, and 3-year rates of 57% (10%), and 76% (10%), compared to HMA 22% (24%), and 31% (42%) [p<0.01 (0.04)]. AlloHSCT was also associated with higher rate of CMR (evaluable= 21, alloHSCT N=16 and HMA N=5, 100% vs 60%, p=0.01). At the last follow up, none of the patients who achieved CMR had re-emergence of the UBA1 clone (despite early mixed chimerism). Conclusions: This study represents the largest comparative analysis of alloHSCT versus HMA therapy in VEXAS syndrome, and confirms superiority of alloHSCT in achieving molecular remission, GC discontinuation/dose reduction, and overall survival. Prospective clinical trial confirmation is warranted.

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: Targeted therapy yields deep and durable response in histiocytic neoplasms.However, relapse rate following interruption is high at 75-77%, with a median time to relapse 6-9 months in previous studies. Outcomes after discontinuation in those who have already achieved a durable response are not well understood and are the focus of this study. Methods: We performed a multicenter retrospective study, including 4 US centers, including patients with histiocytic disorders treated with BRAF inhibitor (BRAFi), MEK inhibitor (MEKi), or combination BRAF/MEKi, and whose therapy was interrupted after ≥12 months of therapy following a best response of partial (PR) or complete response (CR). Progression-free survival (PFS) and overall survival (OS) were analyzed with Kaplan-Meier method. Cox regression was performed to identify clinical and molecular factors that are associated with PFS. Results: 26 patients were included (median age 55 [range 12-72]; male 53.8%; Caucasian 69.2%). Most predominant histology was Erdheim-Chester Disease (ECD) (n=15, 57.5%), followed by mixed histiocytosis (n=5, 19.2%). Rosai-Dorfman Disease (RDD) (n=4, 15.4%), and Langerhans Cell Histiocytosis (LCH) (n=2, 7.7%). All had BRAF V600E analysis and 10 (38.5%) were positive. 19 patients underwent additional molecular testing; alternative mutations identified included MAP2K1/2 (n=6, 23.1%), NRAS (n=1, 3.8%), RAF1 (n=1, 3.8%), ASXL1 (n=1, 3.8%), and SRSF2 (n=1, 3.8%). Targeted therapy was given as frontline therapy (n=8, 30.8%) or in the relapsed refractory setting (n=18, 69.2%) with median 1 prior line of therapy (range 0-7). Targeted therapies include BRAFi (n=5, 19.2%; 2 dabrafenib, 3 vemurafenib), MEKi (n=17, 65.4%; 14 cobimetinib, 3 trametinib), and BRAF/MEKi (n=4, 15.4%; 3 dabrafenib/trametinib, 1 vemurafenib/cobimetinib). PR and CR occurred in 8 (30.8%) and 18 (69.2%) patients, respectively. The median duration of treatment was 32 months (range 13-60) (12-24 months: 9 [34.6%]; 24-36 months: 7 [26.9%]; >36 months: 10 [38.5%]). Median time from initiation of targeted therapy to best response was 6 months (<6 months: 14 [53.8%]; 6-12 months: 5 [19.2%]; >12 months: 7 [26.9%]). Median duration from best response to drug discontinuation was 19 months (12-24 months: 14 [53.8%]; 24-36 months: 7 [26.9%]; >36 months: 5[19.2%]). Reasons for drug discontinuation include toxicity (n=8, 30.8%), patient preference (n=13, 50.0%), cost prohibition (n=1, 3.8%), and other critical medical conditions (n= 4,15.4%). Eleven patients (42.3%) experienced disease progression after discontinuation of targeted therapy, 5 patients progressed within 6 months. Systemic and CNS progression were captured by PET/CT and MRI brain, respectively. At median follow-up of 33.0 months (95%CI 24.3-47.1), median PFS, defined by time from targeted therapy discontinuation to progression or death, was 29 months (95% confidence interval [CI] not estimable). Median OS was not reached. In Cox regression model, BRAF V600E mutation (hazard ratio [HR] 4.16, 95% CI 1.10-15.70, p 0.036) and CNS involvement (HR 4.64, 95% CI 1.21-17.67, p 0.025) were associated with inferior PFS. MEKi containing regimen trended toward improved PFS (HR 0.35, 95%CI 0.10-1.21, p 0.097). There was no difference in PFS when assessing demographics, histology, prior therapies, best response to targeted therapy, treatment duration, or time from best response to discontinuation. Of the 11 patients with progression, 9 were re-challenged with targeted therapy (7 switched class), 2 did not receive next line therapy (1 transitioned to hospice and passed from progression, 1 has pending evaluation for next line therapy). Of 6 with response assessments, all recaptured response (1 CR, 5 PR), and none had progressed by data cutoff. Conclusions: In patients with histiocytic neoplasm and prior durable response to targeted therapy, ~40% experienced disease progression after treatment discontinuation. BRAF V600E mutation and CNS disease are associated with inferior PFS, suggesting these subgroups may not be appropriate for limited duration of treatment. Novel approaches are needed in these high-risk patients.

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 INTRODUCTION Immune checkpoint inhibitors (ICIs) have emerged as a cornerstone of cancer therapy. However, despite their efficacy, ICIs can cause a unique spectrum of autoimmune toxicities known as immune-related adverse events (irAEs). In contrast to irAEs involving other organ systems, hematologic irAEs such as ICI-associated immune thrombocytopenia (ICI-ITP) are less common. As a result, existing data on ICI-ITP are largely confined to isolated case reports, single-center case series, and pharmacovigilance databases lacking granular clinical data. To address this key knowledge gap, we sought to comprehensively characterize the incidence, risk factors, clinical features, treatments, and outcomes associated with ICI-ITP. METHODS We performed a multicenter cohort study of adult patients who initiated ICI therapy at 29 hospitals across 7 major cancer centers in 11 geographically diverse states in the US between 2016-2023. ICI-ITP was defined as a decline in platelet count to <100 x109/L in the absence of an alternative more plausible etiology of thrombocytopenia. Patients with hematologic malignancies were excluded, as were those who received cytotoxic chemotherapy within 30 days prior to thrombocytopenia onset. For each patient identified as having ICI-ITP, we collected data on demographics, comorbidities, medications, laboratory values, treatments, and outcomes. All data were extracted by manual chart review. To identify risk factors for ICI-ITP and to examine its association with death, for each case of ICI-ITP we also collected data on two control patients who initiated ICI therapy at the same site and in the same year but without development of ICI-ITP. We used multivariable logistic regression to identify independent risk factors for ICI-ITP and its recovery (the latter defined according to criteria established by the ITP International Working Group). We used a multivariable Cox model to examine the association between ICI-ITP and its severity with all-cause mortality. RESULTS We identified a total of 214 patients who developed ICI-ITP among 86,648 examined (0.25%). The incidence was highly consistent across sites (range, 0.15–0.30%). Risk factors for ICI-ITP included simultaneous receipt of two different classes of ICIs, additional irAEs, and a lower baseline platelet count. ICI-ITP developed at a median of 8 weeks (IQR, 4–18) following ICI initiation. The median nadir platelet count was 41 x109/L (IQR, 17–64). Six out of 214 patients (2.8%) developed a major bleed, and 16 patients (7.5%) developed a clinically relevant non-major bleed. Cell-bound anti-platelet antibody testing was positive in 6 of 8 patients (75.0%) for whom it was assessed, and median circulating thrombopoietin levels (201 pg/mL [IQR, 178–295]) were similar to those seen in primary ITP. Treatments used included glucocorticoids (49.5%), immune globulin (18.2%), and thrombopoietin receptor agonists (13.6%). Overall, 161 patients (75.2%) recovered from ICI-ITP at a median of 2 weeks (IQR, 1–5) following ICI-ITP onset. Predictors of recovery included additional irAEs and a higher nadir platelet count. Treatment response was observed in 57.0%, 56.4%, and 58.3% of patients within 60 days following initiation of glucocorticoids, immune globulin, and thrombopoietin receptor agonists, respectively. Of 76 patients rechallenged with an ICI, 23 (30.3%) developed recurrent ICI-ITP at a median of 9 weeks (IQR, 2–15); 17 of these 23 patients (73.9%) recovered. Finally, ICI-ITP and its severity were monotonically associated with a higher risk of death (adjusted hazard ratio 2.96 [95% CI, 2.14–4.08] for those with severe ICI-ITP compared to those without ICI-ITP). CONCLUSION In this multicenter study involving patients treated with ICIs at 29 hospitals across the US, we identified 214 patients with ICI-ITP. For the first time, we characterize in detail the incidence, risk factors, clinical features, treatments, and outcomes of ICI-ITP, a relatively novel disease entity. We found that more than half of patients responded to conventional ITP-directed therapies, and that most patients rechallenged did not develop recurrent ICI-ITP. We also found that ICI-ITP was independently associated with an increased risk of death, including a nearly 3-fold increased risk in those with severe ICI-ITP. These findings highlight ICI-ITP as a rare but clinically significant complication of ICI therapy that hematologists must be equipped to recognize and manage.

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 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.

Abstract Introduction: Inherited & acquired bone marrow failure syndromes (BMFS) have “stress erythropoiesis” with macrocytic anemia, elevated fetal hemoglobin (HbF) & erythropoietin (EPO) levels. The degree of HbF elevation & its capacity to discriminate between patients with inherited & acquired BMFS needs to be systemically studied. Genetic testing is often delayed & it is important to establish an early clinical diagnosis to make clinical decisions such as initiation of immunosuppressive therapy. Methods: After institutional review board approval, we compiled a clinically & genetically well-annotated cohort of patients with BMFS evaluated at Mayo Clinic. All patients were required to have at least cytopenia(s) & evidence of low or variable bone marrow cellularity upon a bone marrow evaluation. Patients with pre-clonal or malignant disorders were excluded. Clinical diagnoses were independently verified for accuracy. Detailed clinical & laboratory features were recorded. Germline genetic testing was performed as clinically appropriate on peripheral blood or bone marrow samples through targeted panels. Distribution of continuous variables was statistically compared using nonparametric (Mann-Whitney or Kruskal-Wallis) tests, while nominal variables were compared using the Chi-Square test. Kaplan-Meier estimate of overall survival (OS) was computed from the date of diagnosis to date of death or last follow-up (also censored at the time of allogeneic hematopoietic stem cell transplant). Results: Between years 1998 - 2023, 129 patients met criteria for inclusion in the study. Among these, clinical diagnoses included immune-mediated aplastic anemia (AA, n=56, 43%, 21% severe or very severe as per the Camitta criteria), telomere biology disorders (TBD, n=37, 29%), Fanconi anemia (n=10, 8%), GATA2 haploinsufficiency (n=8, 6%), Diamond-Blackfan anemia (DBA, n=2, 2%), deficiency of adenosine deaminase-2 (DADA2, n=2, 2%), congenital dyserythropoietic anemia (CDA, n=1, 1%), congential amegakaryocytic thrombocytopenia (CAMT, n=1, 1%), Schwachman-Diamond syndrome (n=1, 1%) & undefined (n=11, 9%). Among the inherited group, 63% had an identifiable germline pathogenic variant (most commonly, TERT/TERC 30%, FANC group 25% & GATA2 15%) with the rest diagnosed on a clinical basis. Elevated HbF >1% was noted in 79% of all BMFS patients; 83% AA, 70% TBD, 50% in DADA2 & 67% in undefined patients (no data was available for other inherited subtypes). In patients with an elevated HbF, absolute monocyte count (p=0.04) was significantly higher; however, there were no other significant associations with variables at diagnosis such as hemoglobin (p=0.5), white blood cell count (WBC, p=0.14), absolute neutrophil count (p=0.5), platelet count (p=0.7), mean corpuscular volume (MCV, p=0.2), absolute lymphocyte count (p=0.1), absolute reticulocyte count (p=0.4), EPO level (p=0.3), presence of a PNH clone (p=0.2), or presence of a germline variant (p=0.3). When diseases were grouped into inherited & acquired (AA) subgroups, median HbF was not significantly different (median%, 4.5 versus 2.3, p=0.4), with elevated HbF (>1%) in 75% versus 83% (p=0.6) patients respectively. On the other hand, minor PNH clone (>0.1% in at least two cell types) was detectable in the acquired group only (0 versus 55%, p<0.0001). Similarly, acquired group had a shorter (<3 months) onset of cytopenias preceding diagnosis (P<0.0001), with no significant differences in MCV (p=0.2), marrow dysplasia (p=0.09) or atypia (p=0.6), & splenomegaly (p=0.9). At median follow-up of 7 years, Kaplan-Meier estimate of transplant-censored median OS was lower in inherited versus acquired group (16 years versus median not reached, p=0.02). Elevated HbF did not significantly predict adverse OS in either group. Conclusions: Fetal hemoglobin is elevated in both patients with inherited & acquired (aplastic anemia) BMFS. When compared to established variables, elevated HbF has neither diagnostic utility nor prognostic relevance in predicting adverse outcomes. On the other hand, onset of cytopenias and presence of a minor PNH clone are both useful clinical variables in differentiating patients with inherited from acquired BMFS. Our data offers practical clinical utility in making early treatment decisions for patients with BMFS.

Abstract Purpose. Malignant histiocytoses (MHs) including Langerhans cell sarcoma (LCS), interdigitating dendritic cell sarcoma (IDCS), and histiocytic sarcoma (HS) are rare, aggressive neoplasms of the macrophage-dendritic cell lineage. Prognosis is generally poor, particularly in disseminated disease. While mutation-specific therapies are increasingly used in indolent histiocytic neoplasms, their role in MH remains unclear due to the frequent presence of complex mutational profiles. Our understanding of the genomic landscape of MH is limited. This study aimed to comprehensively characterize the molecular alterations in MH.Methods. We conducted a retrospective chart review of MH patients seen at Mayo Clinic and the University of Alabama at Birmingham who underwent comprehensive next-generation sequencing (NGS), including DNA ± RNA analysis. A systematic literature review using PubMed identified additional MH cases with available molecular data.Results. We identified 17 institutional MH cases (3 IDCS, 3 LCS, 11 HS) and 72 cases from the literature (6 IDCS, 11 LCS, 55 HS) with NGS data. Among the 89 total patients, 26 (29%) had DNA-only sequencing and 63 (71%) had combined DNA/RNA sequencing. Twelve patients (n=12; 13%) had a single mutation; the majority had multiple alterations. The most frequently mutated genes were: RAS (n=26; 29%), CDKN1/2 (n=26; 29%), MAP2K1 (n=15; 17%), PTEN (n=11; 12%), BRAF non-V600E (n=8; 9%), PTPN11 (n=8; 9%), CREBBP (n=8; 9%), TET2 (n=8; 9%), DNMT3A (n=7; 8%), fusions (n=7; 8%), BRAFV600E (n=6; 7%), and NF1 (n=4; 4%). Fusions included BRAF::NRF11, BRAF::CLIP2, BRAF::MBP, EGFR::CYTOR, CDKN2A::ADGRG6, IGH::BCL6, and CLIP2-BRAF. Two patients had no alteration detected. Non-V600E BRAF mutations included D594G (n=3), G596R (n=1), G466A (n=1), N581T (n=1), N581I (n=1), and N581S (n=1). Common co-mutations with BRAF included CDKN2A/B (n=9), TP53 (n=6), KRAS/NRAS (n=5), and PDGFR (n=1). Only 2 of 15 MAP2K1-mutated patients (13%) had isolated mutations; the rest had co-mutations, most frequently TP53 (n=3) and BCL2 (n=3). MAP2K1 mutations included F53I (n=2), C121S (n=2), I103N (n=1), Q56P(n=3), E203K (n=1), F53-K57del (n=1), and F53-Q58del (n=1).NRAS mutations (n=6) included Q16K (n=2), G12C (n=1), Q61R (n=1), and 2 unspecified. KRAS mutations (n=20) included G13D (n=3), Q61H (n=3), G12V (n=2), K117N (n=1), V14I (n=1), and 10 unspecified. The most frequent co-mutational pattern (24%) was RAS plus another gene: BRAF (n=3), TP53 (n=6), MAP2K1 (n=2), PTEN (n=4), and CDKN2A/B (n=6).Twelve patients (12/26; 46%) were treated with targeted therapies. Responses included 6 partial and 5 complete responses. Median duration of response was 27.5 months (range 7–54). The sole non-responder had co-occurring NRASand TP53 mutations and received trametinib. Responders received a range of agents matched to their mutations: trametinib (BRAFV600E, MAP2K1F53I, KRASQ16H/PTEND24G/EPHB1R327C, MAP2K1F53I/BCL2/CDKN1B/KIT, and CALR), dabrafenib + trametinib(BRAFG596R/KRASQ61H), imatinib (MAP2K1C121S/METG28881T/DUSP2G137D/HIST1H3BC97Y/GRIN2AS1216C and PTPN11/STK11/GNA11/JAK2), chidamide + sintilimab(IDH2G515A/RHOAG50T/TET23344delC), sirolimus(PTEN/FGFR3/SETD2) and sorafenib (BRAFD594G/KRASK117N/TP53). Most responders (8/11) had multiple mutations; only 3 had single alterations, suggesting multi-mutant MH may still be targetable. Common complex patterns included BRAF+CDKN2A + CDKN2B (n=4), BRAF + CDKN2A + TP53 (n=2), and BRAF + CDKN2A + KRAS (n=2).Conclusion. The MAPK pathway (BRAF, MAP2K1, RAS) is the most altered signaling axis in MH, often accompanied by mutations in tumor suppressors and cell cycle regulators such as TP53, CDKN2A/B, and PTEN. Most MH patients exhibit multiple genomic alterations. Despite this complexity, durable clinical responses to targeted agents are achievable, even in heavily mutated cases. These findings support the use of comprehensive genomic profiling in MH to identify therapeutic targets and guide precision treatment strategies.