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Abstract Background: A randomized multicenter trial (BMT CTN 1703) demonstrated that after reduced intensity conditioning HCT, PT-Cy-based GVHD prophylaxis led to significant reductions in acute and chronic GVHD, but was associated with significantly more Grade 2 (moderate) infections and similar overall survival vs Tac/MTX. A linked mechanistic study (BMT CTN 1801) found that PT-Cy led to pan-T cell depletion and reduced T cell receptor (TCR) repertoire diversity vs Tac/MTX. To discover the molecular mechanisms driving the association of PT-Cy with increased infections, we examined pathogen-specific T cell reconstitution in BMT CTN 1801 patients (n=165 PT-Cy, n=159 Tac/MTX). Methods: We performed deep TCR sequencing on 2,369 blood samples collected pre-HCT (recipient baseline) and on Days +7, 14, 28, 63, 100, 180, 270, 365, 730, and from the graft infusion product. We then applied a novel set of TCR classifiers (Adaptive Biotechnologies) capable of identifying pathogen-specific clones associated with CMV, EBV, SARS-CoV-2, HSV-1, HSV-2, parvovirus, RSV, influenza A, rhinovirus, adenovirus, HHV-6B, norovirus and Toxoplasma. Single-cell (sc) TCR/RNA-sequencing on a sample subset (n=26) was performed to determine the phenotype of pathogen-specific T cells. Results. By linking scRNA-seq with pathogen-specific TCR barcoding, we identified CD4 and CD8 memory T cells as their predominant T cell reservoir. We found that the total number of pathogen-specific singleton TCRs was significantly lower with PT-Cy, starting early post-HCT and persisting through 2 years. For example, at Day +14, PT-Cy patients had 7.8-fold fewer pathogen-specific singleton TCRs vs Tac/MTX (mean of 6.9 vs 54.1, p<0.001). Consistent differences between PT-Cy and Tac/MTX were observed for TCRs specific for each pathogen individually. Patients who developed Grade 3 (severe) viral infections had significantly fewer pathogen-specific singleton TCRs at Day +14 compared to those who had no viral infections (mean of 4.0 vs 32.5, p=0.03). Leveraging intrinsic TCR barcoding of the HCT infusion (“graft”) and of the patient's baseline samples (“recipient”), we could assign the graft/recipient origin of the pathogen-specific cells with high fidelity. This analysis revealed pronounced differences in the origin of pathogen-specific T cells with PT-Cy vs Tac/MTX, especially prior to Day +100. The absolute number of pathogen-specific TCRs originating from the graft was significantly higher with Tac/MTX vs PT-Cy. This difference was observed from day +7 through 6 months post-HCT, with up to 7.1-fold more graft-derived pathogen-specific TCRs in Tac/MTX vs PT-Cy (p<0.001 at each timepoint before day +100). Within patients who received Tac/MTX, the balance of graft vs recipient pathogen-specific clones significantly favored graft clones (for example, a mean of 14.9 graft vs 8.6 recipient pathogen-specific TCRs at Day+14, p=0.02). By contrast, with PT-Cy, in addition to having substantially fewer pathogen-specific TCRs overall, the proportion of these TCRs was not significantly different between graft and recipient clones (mean of 2.1 graft vs 3.3 recipient pathogen-specific TCRs at Day+14, p=0.42). These data uncover that, in addition to the reduction in pathogen-specific T cells with PT-Cy, there was a relative skewing of the anti-microbial T cell reservoir towards recipient cells that persisted after conditioning. Conclusions: PT-Cy led to in vivo T cell depletion across all T cell subsets, which prominently included clones tracked from the allograft itself into the HCT recipient, with a relative sparing of recipient T cell clones that survived transplant conditioning. While the mechanism for relative sparing of recipient cells has not yet been determined, these conditioning-exposed cells may be less proliferative than newly infused graft T cells. The depletion of graft T cells included CD4 and CD8 memory cells specific for infections, which may have contributed to relative deficits in protective immunity against infection in patients receiving PT-Cy. Importantly, these data suggest that with PT-Cy, persisting recipient TCRs may play a key role in preserving anti-infectious immunity. Taken together, these results provide a mechanistic basis for key post-transplant clinical outcomes of the landmark BMT CTN 1703 study. They also underscore the importance of recipient immune status with PT-Cy, creating the opportunity for targeted infection risk assessment in these patients.

Abstract Use of Direct oral anticoagulants (DOAC) has increased exponentially since their introduction in 2010. Over the years they have been used without any systematic oversight, there have been ongoing concerns related to inappropriate or incorrect dosing, incorrect duration, and non-adherence to the drugs leading to adverse patient outcomes. Several factors have been attributed to DOAC non-adherence such as presence or fear of adverse effects, affordability, lack of perceived benefit, or concerns for reversibility. These barriers impact both primary (first time prescription) and secondary adherence (longer term prescription). Poor adherence can lead to adverse outcomes such as stroke, death, and higher medical resource utilization hence increased cost to the patient and payors. This study was conducted as a quality improvement (QI) initiative to improve the adherence to the prescription DOACS. We included patients 18 years old, a diagnosis of Atrial fibrillation and flutter who had a primary care provider writing the prescription for chronic DOAC therapy. Patients with venous thrombosis, pregnancy or who were institutionalized were excluded. The charts were reviewed for primary and secondary adherence using Surescripts functionality which can provide baseline information about prescription fills from the participating pharmacies. In the preimplementation phase we identified 1030 patients meeting inclusion criteria associated with the pilot sites within Mayo Clinic practice in preceding 6 months. For this population, Surescripts data found that the rates of primary and secondary adherence to be 84% and 86% respectively. We then implemented a 6-month pilot where we created workflow for a pharmacy led DOAC management unit. Herein, once a DOAC prescription was made for patients meeting inclusion criteria, they would enter a registry where the pharmacy team would review the charts for primary adherence by utilizing Surescripts data. If adherent these patients would be marked for follow up in the secondary adherence phase. However, if non adherent, the team would initiate intervention including engagement with patient online services which was further escalated to phone calls by the pharmacy team. Once the barriers were identified several remediation approaches were incorporated in the process, these included education about DOACS, medicare education, social work assistance to even switching to warfarin if necessary. The same process was followed for secondary adherence management. In the intervention cohort, we had 411 patients during the pilot, for primary adherence 87.1% of patients were adherent at baseline and an additional 3.4% became adherent with interventions, while 9.5% remain nonadherent. The primary causes of nonadherence, amongst 9.5%, were cost (86%) while 1% were concerned about side effects and 3% were concerned about lack of perceived benefit. The inverntions lead to impoved adherence in most patients and very few patients were transitioned to warfarin. For secondary adherence 92% of the patients were adherent without any intervention, an additional 4% of patients benefited with the interventions of the DOAC management unit with approaches to address cost considerations, while another 4% remained nonadherent where cost remained the primary reason for nonadherence, other reasons being lack of perceived benefit and side effect concerns. In summary, with active intervention, in our patient population we were able to improve the primary adherence from 84% to 91% and secondary adherence from 86% to 96% . Cost remained the biggest barrier for adherence followed by concern for side effects and lack of perceived benefit. Our engaged team of pharmacy staff was able to manage those concerns with the education and cost mitigation strategies leading to improved adherence. Based on previously published literature even a 5% difference in adherence to DOACS can make a huge difference in population-based outcomes. Our work was able to demonstrate a positive impact of the intervention, highlighting the need for a well-managed DOAC unit especially for complex care populations.

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 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 The ECHELON-2 trial for newly diagnosed CD30+ peripheral T-cell lymphomas (PTCL) demonstrated superior overall survival with brentuximab vedotin plus chemotherapy (BV-CHP) compared to CHOP. However, total enrollment was restricted to only 30% of patients with non-anaplastic large cell lymphoma (ALCL) histologies and was not powered to analyze survival benefits in this cohort. Thus, the benefit of BV-CHP in non-ALCL patients remains less clear. Adding to this uncertainty, CHOP plus etoposide (CHOEP) remains a standard regimen for many non-ALCL PTCL histologies for patients <60 years of age. Which of these three regimens offers superior outcomes for patients with CD30+ non-ALCL PTCL remains to be determined, is a critical knowledge gap in this area, and could meaningfully inform the management of these patients. METHODS We conducted a retrospective analysis of chemotherapy-naïve CD30+ (≥1%) PTCL from 7 centers in the United States. Histologies were limited to PTCL-NOS and PTCL with a T follicular helper phenotype (PTCL-TFH). Frontline regimens were defined as BV-based (BV-CHP), etoposide-based (CHOEP/EPOCH), novel-CHOP (CHOP/CHOEP + novel drug on a clinical trial), or CHOP. CD30 was considered positive if any expression was reported, and when available, stratified as ≥10% or <10% of the total lymphocyte population. RESULTS The CD30+ PTCL cohort consisted of 194 patients with a median age of 68 years at diagnosis and a male (65%) predominance. Nodal PTCL-TFH (65%) was the most common histology. Most had stage III-IV disease (89%) with a high-intermediate to high-risk IPI score of 3-5 (54%). 18% had an ECOG of ≥2, 45% had ≥2 extranodal sites, and 43% had bone marrow involvement, although biopsies were not performed in all patients. CD30 expression in ≥10% of malignant T cells was observed in 55% of all quantitatively reported cases. Frontline therapy consisted of BV-based (36%), etoposide-based (30%), CHOP (27%), and novel-CHOP (7%). Baseline characteristics and CD30 expression were similar between treatment groups. The ORR was 72% (64% CR, 8% PR), with no differences observed by treatment or histology. On multivariate analysis (MVA), a high LDH was associated with lower CR (OR 2.84, p=0.003). For the patients who demonstrated a CR to frontline therapy, 61% underwent ASCT. With a median follow-up of 26.1 months (IQR 12.3-51.6), the median PFS and OS were 11.4 (9.8-14.2) and 47 (32-60.3) months, respectively. The 2-year PFS and OS were 34.9% (28.1-41.7) and 62% (54.5-68.6), respectively. PFS and OS were similar for PTCL-NOS compared to PTCL-TFH, and AITL fared similarly to non-AITL PTCL-TFH. By treatment, novel-CHOP demonstrated the most favorable PFS (median 41.9 months) and OS (median 68.1 months). Survival curves overlapped continuously with all other treatment regimens and was not statistically significant. For patients <60, PFS and OS were equivocal with BV-based (n=17) compared to etoposide-based (n=26) regimens. On MVA, stage III/IV disease conferred an inferior PFS (HR 2.00, p=0.018) as did an IPI score of 3-5 for OS (HR 2.15, p=0.001). Neither histology nor frontline treatment impacted PFS or OS. The chosen frontline treatment also had no survival impact in key subgroup analyses including patients <60 (n=56) or with CD30-expression ≥10% (n=43). For patients with CR to frontline therapy, ASCT demonstrated a significant PFS benefit (HR 0.59, p=0.028) but no statistically significant difference in OS benefit (HR 0.58, p=0.073). CONCLUSION These data provide valuable insight into the lesser-represented patient population from the pivotal ECHELON-2 trial. In this cohort of CD30+ (≥1%) non-ALCL PTCL, there was no clearly superior standard frontline regimen. The number of patients with quantitively reported CD30 expression was too small for definitive conclusions in the ≥10% CD30+ cohort and retrospective pathology review is ongoing. ASCT for patients in CR offered superior outcomes, supporting this strategy regardless of the frontline regimen, recognizing the limitations of non-randomized, retrospective analyses. Novel-CHOP regimens demonstrated encouraging survival, and clinical trials should maintain priority over standard regimens for patients with CD30+ (1-10%) PTCL-NOS and PTCL-TFH.

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 Allogeneic hematopoietic cell transplantation (alloHCT) is indicated in patients (pts) with FLT3-ITD mutated AML. The randomized BMT CTN 1506 trial (Levis et al. JCO) demonstrated that maintenance with FLT3 inhibitor gilteritinib (gilt) significantly improved relapse-free survival (RFS) among pts with detectable peri-alloHCT minimal residual disease (MRD) but was also associated with a greater degree of myelosuppression and infections. In older pts, aged >60 years, increasing post-HCT toxicity may adversely impact outcomes. On this basis, we performed a post-hoc analysis of pts aged > 60 years enrolled onto BMT CTN 1506 to examine the impact of gilteritinib maintenance in older pts. METHODS Pts ≥60 years randomized on BMT CTN 1506 (gilt=47, placebo=57; N=104) were included. The primary endpoint of this post-hoc analysis was RFS. We additionally assessed overall survival (OS), incidence of relapse, non-relapse mortality (NRM) in all pts and in pts with detectable MRD. RESULTS Baseline characteristics were balanced. Age was similar (65.2 vs 64.7 years; p = 0.12), as was prior TKI exposure, and NPM1 mutation status. Myeloablative conditioning (MAC) was used in 8 (17%) pts randomized to gilt and 18 (32%) pts randomized to placebo (p=0.08). Median f/u of survivors was 43.8 months (range:1-61 mo.). Estimated RFS and OS at 48 months were not different between groups (RFS: gilt: 57.0%, 95%CI 44.1-73.7%, placebo: 57.1%, 95%CI 45.4-71.8%; OS: gilt: 56.9%, 95%CI 42.9-46.7%, placebo: 58.4%, 95%CI 46.7-73.1%). While gilt assignment did not affect survival, several predictors of poorer survival were identified including use of reduced intensity conditioning (RIC) (HR = 3.46; p = 0.014), use of calcineurin inhibitor/MTX for graft-vs-host disease (GVHD) prophylaxis (HR = 2.29; p = 0.041), high-risk cytogenetics (HR = 2.64; p = 0.021), and MRD (+) pre-HCT (HR = 2.29; p = 0.018). Finally, relative to gilt assignment, opposing rates of relapse and NRM were observed. The incidence of relapse was lower with gilt (16%, 95%CI 6.8-28% vs 32%, 95%CI 20-45%, p=0.046) but NRM was higher (27%, 95%CI 15-41% vs 11%, 95% CI 4.3-21%, p=0.033). We next analyzed the 49 pts (gilt=22, placebo=27) with detectable peri-HCT MRD. RFS and OS again were not different (RFS: gilt: 44.4%, 95%CI 27.1-72.7%, placebo: 43.8%, 95%CI 28.4-67.5%, OS: gilt: 49.2%,95%CI, 31.3-77.4%, placebo: 55.6%, 95%CI 39.4-77.8%). The incidence of relapse trended lower in gilt pts (25%, 95%CI, 5.3-44.7% vs 45% (95%CI, 25-63%), p=0.2) while NRM trended higher (31%, 95%CI 12-52% vs 11%, 95%CI, 2.7-26%, p=0.12). Seventeen (36%) patients completed planned 2 year maintenance with gilt compared to 29 (51%) completing therapy with placebo. Median time on gilt was 13.2 months. The most common reason for discontinuation was adverse events (n=19, 40%), of which cytopenias (n=8) and infections (n=4) were most frequent. Non-relapse causes of death among gilt treated pts included infections (n=8), intracranial hemorrhage related to thrombocytopenia (n=1), second cancer (n=1), and Guillain-Barre syndrome (n=1). Most infections (n=5) were attributed to GVHD as an underlying cause. CONCLUSION Among pts aged ≥60 years, gilt maintenance did not improve RFS or OS. Gilt use was associated with reduced relapse regardless of peri-HCT MRD status demonstrating disease effect; however, higher NRM limited overall benefit. In older FLT3 mutated pts, employing strategies to mitigate HCT related toxicity, e.g. alternative GVHD prophylaxis strategies, may better accommodate use of maintenance therapy including gilt. Lower doses of gilt (particularly with azole co-use) may be worth exploring to mitigate drug AEs, improve compliance, thus further improving benefit.

Abstract Background: Multiple myeloma (MM) is always preceded by monoclonal gammopathy of undetermined significance (MGUS) and smoldering multiple myeloma (SMM). While these precursor conditions are asymptomatic, they are not entirely benign and carry a lifelong risk of progression to MM. Unlike other cancers defined by pathology, malignant transformation from MGUS or SMM to MM has so far relied on demonstration of clinical end-organ damage or high tumor cell burden (i.e. SLiM criteria, SV Rajkumar Lancet Onc 2014), as morphology and cytogenetics cannot reliably distinguish them. Methods: To identify genomic features underlying clinical behavior in MGUS and SMM, a total of 374 patients with SMM (n= 290) or MGUS (n=84) with available whole exome (whole exome sequencing, WES, n=190) or whole genome sequencing (whole genome sequencing WGS, n=184) were included in the study. Overall, none of the patients had evidence of SLiM or CRAB criteria at the time of the biopsy. Based on the center of origin and availability of WES and WGS, 227 and 97 patients were used to form our study training and validation set. The study was designed in two parts: 1) defining a new classification for MM precursor conditions; 2) integrating genomics to existing prognostic score to improve accuracy in identifying patients at imminent risk of progression. Overall, 68 patients in the training set were either lost to follow-up or enrolled in a clinical trial and they were only included in the classification part of the study. Results: By leveraging the distribution of established MM-defining genomic events (Maura et al. JCO 2024) in the training set, we developed a workflow that differentiates MGUS and SMM into two genomically distinct entities: one with evidence of neoplastic transformation (i.e., genomic MM) and one without (i.e., genomic MGUS). Overall, 39% of MGUS and 91% of SMM cases had genomic evidence of neoplastic transformation in the training set (i.e., genomic MM). When we applied the same workflow and classification to the validation set, 46% and 83% of MGUS and SMM had evidence of neoplastic transformation, respectively. Importantly, none of the patients classified as genomic-MGUS progressed to MM in either the training or validation cohorts, with a median follow-up of 40 and 61 months, respectively. Moreover, all patients with IMWG 2/20/20 high risk SMM were classified as genomic MM in both the training and validation set. Interestingly, 5 out of 62 (8%) patients enrolled in early intervention trials for high risk SMM were genomically classified as genomic-MGUS. Overall, these data demonstrated that the MM genomic background and neoplastic transformation can be acquired very early in time, even before the SMM phase, in line with what observed in solid tumors. To avoid overestimating the prognostic impact of myeloma-defining genomic events—which are, by definition, enriched in the genomic-MM group compared to genomic-MGUS—we restricted our prognostic analysis to patients in whom transformation has already occurred (genomic-MM). In fact, distinct genomic drivers may predict malignant transformation but not necessarily the risk of imminent progression to MM once transformation has occurred. After running 5-fold cross validations corrected for IMWG 2/20/20, only presence of RAS mutations, genomic events involving MYC, presence of neoplastic copy number variant (CNV) signatures and APOBEC mutagenesis were predictive of early progression in the training set. Adding the presence of high-risk myeloma-defining genomic events to the IMWG 2/20/20 model (i.e., genomic-IMWG) significantly improves the accuracy of predicting MM progression in both the training (c-index 0.69 vs. 0.74) and validation sets (c-index 0.74 vs. 0.79), compared to the IMWG 2/20/20 alone. Conclusion: By leveraging comprehensive genomic profiling integrated with the clinical IMWG 2/20/20 risk score, we propose a novel classification and prognostication system for MM precursor conditions, which will better define patients who may benefit from early therapeutic intervention versus watch-and-wait monitoring.

Abstract Background: Covalent Bruton tyrosine kinase inhibitors (cBTKi) have dramatically changed the treatment (tx) landscape of chronic lymphocytic leukemia/small lymphocytic lymphoma (CLL/SLL), especially in the first-line setting. However, cBTKi resistance or intolerance eventually occurs. Pirtobrutinib is a highly selective, non-covalent BTKi that inhibits BTK with low nM potency throughout the daily dosing interval. Pirtobrutinib demonstrated safety and efficacy in the phase 1/2 BRUIN study among patients (pts) with R/R CLL/SLL, including pts previously treated with a BTKi. Pirtobrutinib is approved for treating CLL in adults in the EU after prior tx with a BTKi, and in the USA for adults with CLL/SLL who have received at least 2 prior lines of therapy, including a BTKi and a BCL-2i. Here, we report the final results from the phase 1/2 BRUIN study (NCT03740529) with more than 5 years (yrs) follow-up (f/u), focusing on efficacy and safety of pirtobrutinib in pts with CLL/SLL in the post-cBTKi setting. Methods: Pts with prior cBTKi-exposed CLL/SLL were eligible for tx with pirtobrutinib. Endpoints included overall response rate (ORR) per 2018 iwCLL criteria, ORR including partial response with lymphocytosis (PR-L), duration of response (DOR), and progression-free survival (PFS), assessed by independent review committee (IRC; presented herein) and investigator, overall survival (OS), and safety. Data cut was 27 January 2025, providing a median study f/u of 34.1 months (mos; range, 0.5-69.8). Results: Among the 282 pts with CLL/SLL who were cBTKi-exposed and treated with pirtobrutinib, of which 84.4% (238/282) received 200 mg as the starting dose, the median age was 69 yrs (range, 36-88). The median number of prior therapies was 4 (range, 1-11), with 45.4% of pts having received prior BCL2i, and 77.0% having discontinued prior cBTKi due to progressive disease (PD), 19.5% due to toxicity, and 3.5% due to other reasons/missing. High-risk features were frequent: unmutated IGHV in 85.8% (193/225), mutated TP53 in 40.4% (99/245), and del(17p) in 29.2% (59/202). The ORR was 72.3% (95% CI, 66.7-77.5). ORR including PR-L was 81.6% (95% CI, 76.5-85.9), which was consistent across subgroups including BCL2i-exposed (80.5%; 95% CI, 72.5-86.9), BCL2i-naive (82.5%; 95% CI, 75.5-88.1), unmutated IGHV (82.9%; 95% CI, 6.8-87.9), del(17p) (88.1%; 95% CI, 77.1-95.1), and prior cBTKi discontinuation due to PD (79.7%; 95% CI, 73.8, 84.9) or toxicity (87.5%; 95% CI, 76.8-94.4). The median DOR was 18.4 mos (95% CI, 14.82-20.27), with 36-mos DOR rate of 28.0% (21.6-34.7). The median PFS was 18.7 mos (95% CI, 16.6-21.8) among all pts and the 36-mos PFS rate was 25.0% (95% CI, 19.4-31.0) with 44 mos of f/u. Further, the median PFS was 22.3 mos (95% CI, 19.3-27.6) for BCL2i-naive pts, and 15.9 mos (95% CI, 13.6-17.5) for BCL2i-exposed pts; 16.6 mos (95% CI, 13.8-19.4) among those who discontinued prior cBTKi therapy due to PD, and not estimable (NE) (95% CI, 25.2-NE) in those who discontinued due to toxicity. Among all pts with CLL/SLL, with a median f/u of 46.5 mos, the median OS was NE (95% CI, 47.8-NE), with the 36-mos OS rate of 62.0% (95% CI, 55.8-67.7). Median time on pirtobrutinib was 20.0 mos (range, 0.2-69.8), and 43 (15.2%) pts were still on tx at data cutoff. The most frequent TEAEs, regardless of attribution, were fatigue (38.7%), neutropenia/neutrophil count decreased (35.8%), diarrhea (30.5%), cough (29.8%), COVID-19 (28.4%), and contusion (27.7%). The most frequent grade ≥3 TEAEs were infection (36.5%) and neutropenia/neutrophil count decreased (29.8%). Low rates of grade ≥3 TEAE of hypertension (5.3%), hemorrhage/hematoma (2.5%), and atrial fibrillation/flutter (2.1%) were observed. Grade 5 TEAEs, regardless of attribution, occurred in 35 (12.4%) pts. Overall, 11 (3.9%) pts had a TRAE leading to a dose reduction, and 9 (3.2%) pts had a TRAE leading to pirtobrutinib discontinuation. Conclusion: In heavily pretreated R/R post-cBTKi CLL/SLL pts, pirtobrutinib continues to show favorable and durable efficacy. Pirtobrutinib is well-tolerated, with low rates of hypertension, hemorrhage/hematoma, and atrial fibrillation/flutter, which can be frequently observed with cBTKi. Additionally, dose reductions/discontinuations due to pirtobrutinib TRAEs were low. No new pirtobrutinib safety signals were identified with more than 5 yrs of f/u.