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Abstract Introduction: Chronic lymphocytic leukemia (CLL) is marked by impaired T cell responses that limit the efficacy of immunotherapies. CD19-directed chimeric antigen receptor T cell therapy (CART19) received FDA approval to treat CLL in 2024. However, itsefficacy in CLL patients remains low, with durable remission rate of ~20%. Ibrutinib, a Bruton tyrosine kinase inhibitor, has shown synergy with CART19 in preclinical and early clinical studies. We aimed to study the impact of ibrutinib on CART19 utilizing clinical samples from the pivotal clinical trial E1912 (NCT02048813) of ibrutinib/rituximab (IR) vs fludarabine/cyclophosphamide/rituximab (FCR). This trial led to the FDA approval of first line ibrutinib in CLL. Methods: We generated CART19 incorporating either CD28 (28ζ) or 41BB (BBζ) costimulatory domains from T cells isolated from patients with CLL treated with IR or FCR on the E1912 trial. IR samples were collected at baseline (pretreatment), 6- or 12-months during treatment, or at progression. FCR samples were collected at baseline, 15-18 months during treatment, or at progression. For animal experiments, NOD-SCID-γ-/- (NSG) mice were intravenously (i.v.) engrafted with 1×10⁶ luciferase⁺ CD19⁺ JeKo-1 cells. Upon tumor establishment (~10⁸ photons/sec by bioluminescence imaging), mice were randomized and treated i.v. with 1×10⁶ 28ζ or BBζ derived from longitudinal IR- or FCR-treated patient samples. To further assess the impact of prolonged ibrutinib exposure on CART19 function, we chronically stimulated healthy donor-derived 28ζ or BBζ in vitro for 22 days with JeKo-1 cells and 100nM ibrutinib or vehicle control and then tested CART19 functions. Finally, we performed single cell RNA sequencing (scRNAseq) of longitudinal patient T cell samples treated with IR. Results: Ex vivo T cell expansion was significantly enhanced following IR compared to FCR. BBζ generated from IR-treated patients after 6 months of ibrutinib exhibited enhanced in vitro expansion (p=0.016) and antigen-specific proliferation (p=0.038) in the presence of JeKo-1 cells. This corresponded with significantly improved antitumor efficacy and prolonged survival in vivo compared to cells generated at disease progression (p=0.017). In contrast, 28ζ showed no significant differences in efficacy across IR studied treatment timepoints. BBζ from FCR-treated patients did not show differences in tumor control based on treatment timepoints but demonstrated shorter survival compared to BBζ from IR-treated patients. 28ζ from FCR-treated patients generated at disease progression demonstrated significantly reduced tumor control and shorter survival compared to 28ζ produced at baseline (p=0.0002), indicating detrimental effects of FCR treatment on the therapeutic potential of CART products. To further investigate the impact of prolonged ibrutinib exposure, we utilized in vitro models of CART exhaustion. We found that, after repeated antigen stimulation with JeKo-1 cells, ibrutinib-treated BBζ had significantly increased antigen-specific degranulation (p=0.005), and cytokine production (IL-2, p=0.045; TNF-α, p=0.003; IFN-γ, p=0.001), compared to BBζ treated with vehicle control. In contrast, 28ζ did not show improved degranulation or cytokine production in cocultures with ibrutinib. Prolonged ibrutinib exposure led to downregulation of FOXO3, a transcription factor associated with T cell exhaustion which has been shown to cause dysfunction specifically in BBζ. These results further indicate that ibrutinib may be more beneficial to BBζ. Indeed, scRNAseq revealed that after ≥ 6 months of ibrutinib, T cells showed downregulation of inflammatory, oxidative stress, and apoptosis pathways, paralleled by modest activation of MYC and E2F targets. However, at disease progression, T cells under ibrutinib treatment displayed marked upregulation of interferon signaling, inflammation responses, and stress-related programs, indicating a shift toward immune reactivation or dysfunction.Conclusion: Our study shows that BBζ, but not 28ζ, generated 6 months during ibrutinib treatment has superior antitumor activity and persistence compared to similar later timepoints or FCR-treated counterparts. Prolonged ibrutinib exposure enhances BBζ function by modulating stress and apoptotic pathways. These findings highlight the importance of pre-CART treatment timing and support ibrutinib as a potential enhancer of CART cell fitness in CLL.

Abstract BTK degraders promote proteasomal degradation of BTK and can overcome resistance to covalent and noncovalent BTK inhibitors in patients with relapsed CLL. Currently the molecular basis for BTK degrader resistance is unknown. We therefore set out to understand BTK degrader resistance based on observations from the phase 1 trial of zelebrudomide (NX-2127) (NCT04830137), the first BTK degrader to enter clinical studies. Targeted 500-gene sequencing was performed on blood and/or tissue from four CLL patients treated with zelebrudomide. In one patient previously treated with chemoimmunotherapy, ibrutinib, venetoclax, and pirtobrutinib, BTK A428D was detected in the baseline blood sample with variant allele frequency (VAF) of <2%, along with BTK V416L (13%) and mutations in TP53 (60%) and SF3B1 (37%). Following daily treatment with 100mg zelebrudomide and subsequent dose escalation to 200mg, at 6 months the VAF of all four mutations increased, with BTK A428D VAF increasing to 6% in the blood and 29% in a lymph node. BTK A428D was detected at baseline but not at progression in a second patient. To evaluate the impact of BTK A428D on drug response, we transduced BTK-dependent TMD8 cells with BTK WT, C481S, L528W, and A428D, and found that BTK A428D uniquely conferred resistance to all FDA-approved BTK inhibitors (ibrutinib, acalabrutinib, zanubrutinib, pirtobrutinib) as well as three BTK degraders under clinical evaluation (zelebrudomide, bexobrutideg, BGB-16673). To model the subclonal nature of BTK A428D we conducted a competition assay in which TMD8 cells expressing GFP + BTK A428D (5%) were mixed with TMD8 cells expressing mCherry + BTK WT (95%); over 9 days the GFP + BTK A428D expanded from 5% to 88-97% with all four BTKi and all three BTK degraders. In vitro kinase assays revealed that the BTK A428D mutant protein lacks kinase activity yet facilitates calcium flux and ERK/AKT signaling upon B-cell receptor activation, suggesting a scaffolding effect similar to other kinase deficient BTK mutations. To determine whether BTK A428D affects BTK degradation, we developed a reporter that fuses the BTK kinase domain to GFP. BTK A428D showed minimal degradation following 20 hours of treatment with a titration of all three BTK degraders. Similarly, we knocked BTK A428D into the endogenous BTK locus of HBL1 cells and confirmed a lack of degradation at doses up to 1 µM of all 3 degraders. Consistent with these findings, surface plasmon resonance revealed that the FDA-approved BTK inhibitors as well as zelebrudomide, bexobrutideg, BGB-16673, and AbbVie-1 (Compound 1 from WO 2023183811) are unable to bind to BTK A428D. Additionally, an in-cell NanoBRET assay demonstrated that zelebrudomide was unable to mediate the interaction between cereblon and BTK A428D. Strikingly, NanoBRET also demonstrated that a panel of promiscuous kinase-binding compounds was unable to bind to BTK A428D. The lack of binding of both selective and promiscuous kinase binders to BTK A428D led us to speculate that the mutation disrupted the ATP binding pocket to an unusual degree. We therefore solved the crystal structure of apo BTK A428D and found that, in addition to its predicted steric clash with ligand binding in the hinge region, the mutation led the protein to crystallize with its activation loop occupying the catalytic cleft, rendering it inaccessible. This is the first apo BTK structure observed in this autoinhibited conformation, and explains the observed binding profile. We hypothesized that the addition of a second targeted agent such as venetoclax might mitigate expansion of the A428D subclone. To test this, we introduced the BTK A428D mutation into the endogenous BTK gene in 3% of TMD8 cells and then exposed them to 0.1µM of zelebrudomide, bexobrutideg, or BGB-16673 +/- venetoclax 0.1µM. Combining the BTK degraders with venetoclax slowed the emergence of the BTK A428D resistant population and, in the case of zelebrudomide, eradicated both the wild-type and A428D clones. In summary, we identified and validated BTK A428D as a clinically relevant acquired pan-resistance mutation that confers resistance by blocking access to the ATP binding pocket. This mechanism differs from other kinase dead BTK mutations in which the kinase's enzymatic pocket remained accessible.These data also provide rationale for combining BTK degraders with BCL2 inhibitors, particularly in patients with BTK A428D mutations, or high-risk mutations such as in TP53.

Abstract INTRODUCTION: Warm autoimmune hemolytic anemia (wAIHA) is a rare, autoimmune-mediated hematologic disorder associated with significant morbidity and mortality and may occur as a primary condition or secondary to an underlying disorder. It is the most common subtype of autoimmune hemolytic anemia (AIHA), accounting for 75-80% of AIHA cases. Older adults with wAIHA are particularly vulnerable, as they often have multiple comorbidities and are already at elevated risk of death. This study aimed to assess mortality associated with wAIHA. METHODS: Adults with wAIHA (wAIHA cohort) and without AIHA (non-AIHA cohort) were identified from the 100% Medicare Fee-for-Service database (10/01/2019–12/31/2023). Evidence of wAIHA was defined as one of the following: 1) a primary wAIHA diagnosis (International Classification of Diseases, Tenth Revision, Clinical Modification [ICD-10-CM]: D59.11) in the inpatient setting (first instance of wAIHA diagnosis defined the index date), 2) ≥2 wAIHA diagnosis in any setting ≥30 days apart (second instance of wAIHA diagnosis defined the index date), or 3) a wAIHA diagnosis preceded ≥30 days by any AIHA diagnosis (ICD-10-CM: D59.10, D59.19, D59.8, D59.9; first instance of wAIHA diagnosis defined the index date). Patients in the wAIHA cohort were excluded if they had a diagnosis for cold or mixed-type AIHA (ICD-10-CM: D59.12, D59.13) during the baseline period (12 months before index). The non-AIHA cohort had no diagnoses for any AIHA during the study period; a random index date during continuous Medicare eligibility was selected and patients were matched 1:10 with the wAIHA cohort based on age on index date. Using overlap weighting, age-matched cohorts were further weighted on sex, race, region, index year, and baseline comorbidities associated with mortality, excluding conditions defining secondary wAIHA. Mortality during the follow-up period was evaluated between the weighted cohorts using the Kaplan-Meier method and Cox proportional hazard models. Patients whose death was not observed were censored at the earlier of the end of continuous Medicare eligibility or data availability. RESULTS: 3,112 and 31,120 patients were included in the wAIHA and non-AIHA cohorts, respectively. After weighting, among the wAIHA vs non-AIHA cohorts, mean [standard deviation; SD] age was 74.4 [11.6] vs 74.6 [11.2] years, the proportion of female were 56.1% vs 56.1%, mean [SD] Quan-CCI without secondary wAIHA-defining conditions was 1.9 [2.0] vs 1.9 [2.1], and mean [SD] follow-up time was 17.2 [11.7] vs 14.9 [11.4] months, respectively. In addition, 53.7% of the wAIHA cohort had secondary wAIHA. The most common secondary wAIHA-defining conditions were autoimmune and inflammatory diseases (wAIHA cohort: 27.8%; non-AIHA cohort: 11.0%), hematologic and lymphoproliferative disorders (wAIHA cohort: 23.8%; non-AIHA cohort: 1.3%), and solid tumors (wAIHA cohort: 16.8%; non-AIHA cohort: 12.6%). The most common comorbidities for both cohorts were hypertension (wAIHA cohort: 77.4%; non-AIHA cohort: 78.3%) and dyslipidemia (wAIHA cohort: 72.2%; non-AIHA cohort: 74.7%). Survival rates in the wAIHA and non-AIHA cohorts were 80.3% vs 84.0% at 12 months, 70.7% vs 77.6% at 24 months, and 63.7% vs 73.1% at 36 months, respectively (all p<0.001). Patients with wAIHA were at 20%, 27%, and 30% higher risk of death at 12, 24, and 36 months, respectively, compared to patients without AIHA (hazard ratios: 1.20, 1.27, and 1.30; all p<0.001). CONCLUSIONS: This large retrospective study is the first to assess real-world incremental mortality associated with wAIHA among Medicare beneficiaries. Patients with wAIHA were at significantly greater risk of death following the initial diagnosis. These results may point to potential gaps in early diagnosis and effective disease management, which could contribute to increased mortality. Future research should focus on specific subgroups within the wAIHA cohort to identify differences in mortality as well as characteristics that may be associated with death.

Abstract Introduction Obecabtagene autoleucel (obe-cel) and brexucabtagene autoleucel (brexu-cel) are CD19 targeted chimeric antigen receptor T cell (CAR-T) therapies, approved for the treatment of adults with relapsed/refractory (r/r) B-cell acute lymphoblastic leukemia (ALL). Mechanistic differences between obe-cel and brexu-cel including differing costimulatory domains (4-1BB vs. CD28), CD19 binding domains (intermediate vs. high affinity) and split dose (Days 1 and 10) vs. single infusion may impact in-vivo cellular kinetics that translate into variant clinical outcomes. In pivotal trials, obe-cel resulted in lower rates of severe cytokine release syndrome (CRS) and immune effector cell associated neurotoxicity syndrome (ICANS) compared to brexu-cel with comparable efficacy (Roddie et al., NEJM 2025; Shah et al., Lancet 2021). Real-world utilization and outcomes with obe-cel are unknown given the relatively recent approval. Methods The ROCCA database, comprising real world data from patients (pts) with r/r ALL treated at 40 North American institutions was utilized in this analysis. Pts with r/r ALL were eligible if they were apheresed for obe-cel since its approval (11/8/2024) or brexu-cel over a comparable period (since 8/1/24) and had at least 30 days of follow up. Data cut off was 7/15/2025. CRS/ICANS were graded per ASTCT criteria. Measurable residual disease (MRD) was assessed by flow cytometry and/or next generation sequencing per institutional standards. Results 38 pts have undergone apheresis for obe-cel (36 infused, all received both infusions) and 54 (53 infused) for brexu-cel over the study period. Pts in the obe-cel and brexu-cel cohorts had the following baseline characteristics: median age, 47 vs. 39 years; ECOG 2+, 15% vs. 26%; Ph+ disease, 24% vs 13%; KMT2Ar, 0 vs. 4%; TP53m, 11% vs 11%, and hypodiploid karyotype, 5% vs 6%. The following prior treatment patterns were observed: median lines of therapy before CAR-T 3 vs. 3; prior SCT 24% vs. 30%, prior blinatumomab 68% vs. 46%, prior inotuzumab 45% vs. 30%. At apheresis, active disease (>5% bone marrow blasts) was present in 47% vs. 55% of obe-cel and brexu-cel pts, respectively; the remainder were in CR (with 18% vs. 22% in MRD- CR, respectively). Among obe-cel (n= 35 [92% of cohort]) and brexu-cel (21 [38% of cohort] pts with disease re-assessment prior to lymphodepletion (LD), 31% vs. 43% had active BM disease, respectively; few pts (24% vs. 17%) had >20% marrow disease burden prior to LD. CAR-mediated toxicity differed significantly between the cohorts. CRS occurred in 56% of obe-cel pts compared to 94% of brexu-cel pts (p < 0.0001). There were no Gr3+ CRS events among the obe-cel pts; 3 (6%) brexu-cel pts had Gr3+ CRS (p = 0.27). ICANS occurred in 17% of obe-cel pts vs. 51% of brexu-cel pts (p = 0.001). Gr3+ ICANS occurred in 6% of obe-cel vs. 32% of brexu-cel pts (p = 0.0027). Among the obe-cel pts, CRS occurred in 31% after the first infusion and 46% after the second; ICANS occurred in 3% after the first infusion and 15% after the second. Prolonged Gr4 neutropenia (ANC < 500 cells/uL beyond day 30 from infusion) occurred in 24% of obe-cel vs. 28% of brexu-cel pts (p = 0.73). Deaths within the first 28 days of infusion occurred in 0 obe-cel pts and 4 brexu-cel pts (2 of infection, 1 of infection/brain bleed, and 1 of liver failure in the setting of Gr4 CRS and HLH). Response rates were high and did not significantly differ between cohorts (p = 0.85). Among the 31 obe-cel pts with available response data, 25 (81%) achieved an MRD- CR/CRi, 3 (10%) had an MRD+ CR/CRi, 2 (6%) had a CR/CRi with unknown MRD, and 1 (3%) did not respond. Among 41 brexu-cel pts with available response data, 33 (80%) had an MRD- CR/CRi, 4 (10%) an MRD+ CR/CRi, and 4 (10%) CR/CRi with unknown MRD. Conclusion Pts selected for obe-cel apheresis were similar to those for brexu-cel over the study period (noting that not all centers had access to obe-cel during this time). Similar to clinical trial results, obe-cel was associated with lower rates of CRS/ICANS. Rates of MRD-negative CR were high and did not differ between cohorts. A larger sample and longer follow up are required for further analyses; we anticipate a cohort of ~75 obe-cel treated pts by the annual meeting and will provide updated data.

Abstract Background The treatment (tx) landscape for chronic lymphocytic leukemia (CLL) has evolved, with National Comprehensive Cancer Network (NCCN) guideline-preferred novel therapies (NTs), such as acalabrutinib, zanubrutinib, and venetoclax-based regimens, now preferred over ibrutinib and chemoimmunotherapy (CIT) in front-line (1L) settings. We previously showed that patients (pts) from different racial/ethnic groups may not have equitable access to NTs. Here, we examined racial/ethnic inequities in 1L NT utilization among US pts with CLL and potential social determinants of health (SDOH) drivers of these inequities. Methods This retrospective cohort study utilized the US-based, electronic health record-derived de-identified Flatiron Health Research Database, linked to neighborhood (US Census track or block group) data from American Community Survey and Agency for Healthcare Research and Quality datasets. Eligible pts included adults with CLL who started 1L tx 01/01/2019-07/31/2024. The primary outcome was receipt of 1L therapy (CIT vs preferred NTs, ibrutinib vs preferred NTs), with NCCN guideline-preferred NTs (v2.2023-v3.2025) as a reference. Associations between race/ethnicity (White, Black, Hispanic) and 1L tx types were assessed using logistic regressions, adjusting for age, sex, year of 1L start, immunoglobulin heavy chain variable region status, and del17p/TP53 status. To better understand which SDOH factors contribute to racial and ethnic inequities in tx utilization, we performed mediation analysis using Multiple Mediation Analysis implemented through nonlinear multiple additive regression tree models. Individual-level (Eastern Cooperative Oncology Group status, practice type, insurance) and 20 area-level SDOH factors measuring social deprivation (defined as limited access to economic, social, neighborhood, physical, or healthcare resources) were assessed as potential mediators. Results Of the 4452 pts included in the study, 3717 were White (83%), 371 were Black (8%), 209 were Hispanic (5%), and 155 were Other race (4%). Black and Hispanic pts were more likely to live in neighborhoods with higher social deprivation, such as residential segregation (predominant race/ethnicity in the area: White, Black, Hispanic, Diverse), with no internet access, no vehicle ownership, and no health insurance coverage. Overall, 37% of pts received a preferred NT, highest among White (38%) pts, followed by Black (35%) and Hispanic (26%) pts. Compared with White pts, Hispanic pts were more likely to receive CIT than preferred NTs (adjusted odds ratio [aOR] 2.12; 95% confidence interval [CI] 1.46-3.09); Black (aOR 1.44; 95% CI 1.05-1.97) and Hispanic pts (aOR 1.83; 95% CI 1.17-2.85) were more likely to receive ibrutinib than preferred NTs. Pts living in areas with the highest social deprivation were less likely than their counterparts in lowest social deprivation neighborhoods to receive preferred NTs. Specifically, residence in predominantly Black or Hispanic neighborhoods (26% or 27% vs 39% predominantly White), areas with lowest socioeconomic status (32% vs 40%), and neighborhoods with the lowest levels of internet access (32% vs 39%), vehicle ownership (32% vs 39%), and health insurance coverage (34% vs 41%) was associated with lowest receipt of preferred NTs. In the mediation analysis, individual- and area-level SDOH factors explained 94.4% (95% CI 2.9-185.9) of the observed Black-White inequity in receipt of ibrutinib versus preferred NTs; residential segregation appeared to be the most important mediator (59.7%; 95% CI −3.5 to 123.0). Similarly, the Hispanic-White inequity in receipt of preferred NTs was driven by the combined effect of area-level SDOH and individual-level variables (88.5%; 95% CI 45.6-131.3), with residential segregation as the main driver (48.3%; 95% CI 10.7-85.9), followed by practice setting (9.8%; 95% CI −3.5 to 23.1). In contrast, measured SDOH factors did not explain the inequity in receipt of CIT versus preferred NTs between Hispanic and White pts (21.5%; 95% CI −8.3 to 51.4), suggesting that other factors may contribute to the observed inequities. Conclusion Black and Hispanic pts with CLL were less likely than White pts to receive preferred NTs. A significant proportion of these disparities was explained by area-level SDOH, particularly residential segregation. These findings underscore the need to address structural barriers to ensure equitable access to emerging, guideline-recommended treatments.

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

Abstract Background: Myeloproliferative neoplasms (MPNs) are associated with an elevated risk of thrombotic and hemorrhagic complications. Selective serotonin reuptake inhibitors (SSRIs), commonly prescribed for major depressive disorder and various anxiety disorders, exert a dual effect on platelet function, displaying both prothrombotic and pro hemorrhagic tendencies. However, their impact on vascular outcomes in patients with concomitant MPNs remains unclear. The current study examines the association between SSRI use and the risk of thrombotic and bleeding events in patients with MPNs. Methods: We conducted a retrospective, propensity matched study using the TrinetX network database. Patients >18 years of age, diagnosed with any of the following MPNs: Polycythemia Vera (PV), Essential Thrombocythemia (ET), or Primary Myelofibrosis (PMF) were included. The cohorts were then compared between MPN patients prescribed SSRIs after this diagnosis to those who were never on SSRIs. Patients who were ever prescribed Serotonin Norepinephrine Reuptake Inhibitors (SNRIs) were excluded. Propensity score matching was applied to match demographics and comorbidities. The primary outcomes were all-cause mortality and risk of thrombotic events like deep vein thrombosis (DVT), pulmonary embolism (PE), stroke and myocardial infarction (MI). Secondary outcomes were risk of gastrointestinal (GI) bleeding and intracranial hemorrhage (ICH). Results: Propensity score-matching resulted in 10,920 patients in each group, with MPNs, with and without SSRI use respectively. All-cause mortality was higher in those with SSRI use, Hazard Ratio 1.094, (95% CI 1.006, 1.19). The risk of thromboembolism was higher in those with SSRI use: venous thromboembolism, OR 1.968 (1.666, 2.324), PE, OR 2.225 (1.754, 2.824), Stroke OR 3.019 (2.556, 3.566) and MI OR 2.72 (2.288, 3.233). The risk of bleeding was as follows: GI bleeding OR 2.123 (1.877, 2.401) and ICH OR 2.893 (2.268, 3.691). Conclusion: The current study suggests that SSRI use in patients with MPNs is significantly associated with increased risk of thrombosis and bleeding, particularly stroke and ICH. SSRIs predispose to thrombosis by transiently elevating serotonin levels, causing a pro inflammatory state, impaired fibrinolysis, and vascular endothelial dysfunction. However, with chronic use, SSRIs can deplete intraplatelet serotonin levels, impairing platelet aggregation causing an increased risk of bleeding, especially in conjunction with antiplatelet or cytoreductive agent use. Given the high prevalence of SSRI use, our study underscores the need for further prospective studies, heightened clinical vigilance when prescribing SSRIs in patients with MPNs and potential consideration of alternate agents, using individualized treatment strategies.

Abstract Introduction: The real-world outcomes in patients (pts) with PCALCL in the era of targeted therapies like Brentuximab Vedotin (BV) is not well defined due to lack of large multicenter studies. We conducted a multicenter retrospective analysis (RWA) to assess patient and disease characteristics, treatment patterns, survival outcomes and impact of novel therapies in this rare type of cutaneous T cell lymphoma (CTCL). Methods: The analysis included 167 adult (≥18 years) patients (pts) with PCALCL across 9 academic institutions in the United States diagnosed between 01/2012 to 06/2022. The study was approved by the Institutional Review Boards at the respective sites. Demographic and disease factors were summarized using descriptive statistics. Overall survival (OS) and relapse-free survival (RFS) were estimated using Kaplan-Meier method and compared via log rank tests. Prognostic models included factors such as age, sex, race, performance status (PS), disease stage, type of initial lesion, presence of B symptoms, previous history of lymphomatoid papulosis and prior transplant. Prognostic factors for survival were assessed using both univariable and multivariable Cox proportional hazard models. Results: The median age at diagnosis was 61 (range 18-93) years. The majority of pts were male (55%) and white (82%), with good PS as indicated by an ECOG 0-1 (71%). The initial skin presentation consisted of tumors in 62% of the pts, while 34% presented with plaques or patches. Notably, no pts presented with erythroderma. Most pts underwent PET scan for staging at initial diagnosis (83%) with the majority having early-stage (IA-IIA) disease (86%). Furthermore, palpable lymph nodes (11%) and the presence of B symptoms (4%) were rare at diagnosis, and no pts had blood involvement. The most common sites of involvement were extremities (33%) followed by the head and neck (H&N) and trunk (31% each). Immunophenotyping revealed CD4 positivity in 59% of pts and CD8 positivity in 25%. The T cell receptor (TCR) rearrangement was positive in 28% of pts, though it was not completed in the majority of cases (72%). Most common initial treatments included localized radiation (61%) and surgical excision (19%). A small subset of pts received frontline systemic chemotherapy regimens such as cytotoxic chemotherapy (11%), BV (8%) and oral methotrexate (MTX) (8%). Approximately 7% of the pts had a stem cell transplant. Overall response rate (ORR) for radiation, oral MTX, BV and cytotoxic chemotherapy were 78% (CR:78% 74/100), 42% (CR:63% 6/12) and 39% (CR:67% 6/12), 89% (CR: 88% 15/18) respectively. At a median follow-up of 3.8 years (range 0.04-10.23), the median RFS and OS were 2.7 years (95% CI:1.86, 3.43) and 12 years (95% CI:10.6- NA) respectively. The estimated 3- and 5-year RFS are 46% (95% CI:0.38, 0.55) and 37% (95% CI:0.28, 0.46) respectively. The estimated 3- and 5-year OS were 89% (95% CI:0.82, 0.93) and 82% (95% CI:0.73, 0.88) respectively. At the time of the last follow-up, 79% were alive, while 21% were deceased. The cause of death (COD) was unknown in 50% of the cases and only 10% (3/of deaths were attributed to lymphoma. Other known CODs included infection (20%) and second malignancies 7%. On univariate analysis (UVA), age (p=.007), female sex (p=.04), ECOG PS 1-2 compared to 0 (p=.007) and prior transplant (p=.009) predicted for worse RFS. On multivariate analysis (MVA), only ECOG PS 1-2 (HR 2.31, 95 CI:1.06, 1.18, p=.009) and prior transplant (HR 5.38, 95 CI:2.30, 12.57, p=.0001) had a significant impact on RFS. Age and presence of B symptoms were strong predictors of OS on both UVA (age p=<.0001; B symptoms p=<.0001) and MVA (age: HR 1.12, 95 CI:1.06, 1.18, p=<.0001; B symptoms: HR 16.48, 95 CI: 3.69, 73.60, p=.0002). However, the impact of B symptoms should be interpreted with caution as only 4% of pts presented with this at baseline. On UVA, each one-year increase in age was associated with an 8.5% rise in the risk of death. ECOG PS 1-2 was associated with worse OS on univariate analysis (HR 2.62, 95% CI 0.99, 6.93, p=.05), but was not significant on MVA (HR 1.05, 95% CI 0.368, 2.974, p=.93). Conclusions: In this large RWA of PCALCL, excellent long-term survival was observed with predominantly skin directed frontline therapies. The use of systemic treatments, including BV, was infrequent as first line therapy. Older age and poor PS were significant predictors for reduced survival.

Abstract Background Previous studies in aggressive B-cell lymphoma demonstrate that patients with shorter diagnosis-to-treatment intervals (DTI) represent those with higher risk disease. This suggests that patients enrolled in clinical trials may carry a selection bias as they often have longer DTI than patients in the general population. This has important implications for clinical research design and the generalizability of clinical trials in aggressive lymphomas. However, the prognostic implications of DTI have not been evaluated in peripheral T-cell lymphomas (PTCL). Therefore, we examined DTI and its association with clinical factors and outcomes in a prospective observational cohort of patients with PTCL from the United States within the LEO (Lymphoma Epidemiology and Outcomes) and MER (University of Iowa and Mayo Clinic Specialized Programs of Research Excellence Molecular Epidemiology Resource). Patients and Methods We evaluated patients enrolled to the MER (2002-2015) and LEO (2015-2020) cohorts with the following forms of PTCL: PTCL-not otherwise specified (PTCL-NOS, n=217), nodal T-follicular helper (TFH) cell lymphoma (nTFHL, n=153), anaplastic large cell lymphoma (ALCL), ALK positive (ALCL, ALK+, n=74), ALCL, ALK negative (ALCL, ALK-, n=95), enteropathy associated T-cell lymphoma (n=18), and monomorphic epitheliotropic intestinal T-cell lymphoma (n=2). Associations of DTI with clinical factors and outcomes were examined using Kaplan Meier and Cox models. Event free survival (EFS) was defined as the time from start of treatment to relapse/progression, start of 2nd line therapy, or death from any cause. Overall survival (OS) was defined as the time from start of treatment until death from any cause. DTI was defined as the number of days between the first diagnostic biopsy and the start of therapy. Patients who were initially observed or had DTI > 100 days were excluded. Results In 559 patients (229 MER and 330 LEO), median age was 60, 34% had IPI>2 and 71% were stage III/IV. 472 (84%) patients initially received anthracycline-based chemotherapy. Median DTI was 23 days (interquartile range (IQR): 12-39) and was similar in MER (21 days, IQR: 11-33) and LEO (25 days, IQR: 14-42). 168 (30%) patients had DTI 0-14 days, 178 (32%) 15-28 days, and 213 (38%) 29-100 days. Overall in the cohort, shorter DTI was associated with a worse 5-year OS (DTI 0-14 days: 43%; DTI 15-28 days: 53%; DTI 29-100 days: 58%, p<0.001) and 5-year EFS (DTI 0-14 days: 31%; DTI 15-28 days: 39%; DTI 29-100 days: 45%, p<0.001). In multivariable analyses, the association of DTI with outcomes remained consistent after adjusting for age and International Prognostic Index (IPI). Shorter DTI was strongly associated with adverse clinical factors, including elevated lactate dehydrogenase levels, poor performance status, B-symptoms, and higher IPI in both cohorts (all P < .003). DTI was not associated with age, sex, or distance from the LEO/MER center (all p>0.3). 6% patients with DTI of 0-14 days received 1L treatment on a clinical trial, compared to 11% with DTI 15-28 days and 13% with DTI 29-100 days, underscoring that patients with lower DTI may be underrepresented in clinical trials. When examining within PTCL subtypes, the association between short DTI (0-14 days) and inferior outcomes was most prominent in PTCL, NOS (OS HR=2.13, 95% CI: 1.52-2.94; EFS HR =1.49, 95% CI: 1.19-1.85). Associations between short DTI and outcomes were weaker in other subtypes: nTFHL (OS HR=1.28, 95% CI: 0.83-2.00; EFS HR =1.22, 95% CI: 0.82-1.81), ALCL, ALK-, (OS HR=1.30, 95% CI: 0.59-2.86; EFS HR =1.32, 95% CI: 0.68-2.56), ALCL, ALK+, (OS HR=1.05, 95% CI: 0.26-4.17; EFS HR =2.12, 95% CI: 0.84-5.26). Conclusion Shorter DTI is associated adverse prognostic clinical factors (LDH, stage, high IPI) and inferior EFS and OS in newly diagnosed PTCL. Patients with DTI ≤ 14 days, which represents approximately 1/3 patients with PTCL, may be underrepresented in clinical trials. Understanding of this selection bias should inform clinical trial design and interpretation. Future studies should consider strategies to minimize barriers to enrollment for patients with low DTI.

Abstract Myeloproliferative neoplasms (MPNs) are typically ascribed to hyperactive JAK-STAT signaling via mutations in JAK2, CALR, or MPL. Here, we characterize how activating mutations in the calcium sensor STIM1, previously established as the cause of Stormorken Syndrome (SS), a rare congenital disorder, are sufficient to drive development of myelofibrosis (MF). We further uncover distinct interactions between STIM1 activity and JAK2 versus CALR driven MPNs. We initially identified two SS patients with activating mutations in STIM1 (R304W and S88G) and features of MF, including marked reticulin fibrosis with megakaryocytic hyperplasia and atypia. This led us to hypothesize that altered calcium signaling may play a role in the development of MF in both SS and MPNs. Single cell RNA sequencing performed on samples from both patients demonstrated elevated NFκB inflammatory signaling in monocyte populations, suggesting a link between altered STIM1 and store operated calcium entry (SOCE) activity and aberrant inflammation, thereby contributing to MF development. To corroborate these findings from our SS patients, we generated a knock-in mouse model of the heterozygous R304W mutation. Induction of hematopoietic-specific expression of mutant Stim1 with Vav-Cre recapitulated the hematological features of SS, including elevated SOCE activity and thrombocytopenia. We also recapitulated MF features including the development of bone marrow fibrosis and severe osteosclerosis, with the mice developing splenomegaly as young as 9 weeks old. In ex vivo megakaryocyte differentiation assays with our Stim1 R304W knock-in mice, we also observed aberrant cytoskeletal organization and elevated TGFβ mRNA expression. When induction was driven universally through CMV-Cre, we also validated that mice recapitulated multiple SS features, including thrombocytopenia, tubular aggregate myopathy, growth deficiency, and the development of bone marrow fibrosis as young as 10 weeks. We additionally identified a separate cohort of 9 related individuals with SS and STIM1 mutations (L92V) as well as a separate patient with the R304W mutation. In ex vivo megakaryocytic differentiation assays, cells from SS patients demonstrated defective thrombopoiesis with significantly decreased proplatelet formation, and displayed aberrant cytoskeletal formation within the pro-platelet termini, in conjunction with increased expression of TGFβ. Further supporting a role for STIM1 in MPNs, we identified upregulation of STIM1 expression in MPN patient CD34+ hematopoietic stem/progenitors' cells (HSPCs), we well as megakaryocyte progenitors and platelets, including those with CALR or JAK2 mutations. To evaluate the functional role of STIM1 in MPNs, we generated patient-derived xenograft (PDX) models with CALR and JAK2-mutantpatient samples subjected to CRISPR ablation of STIM1. In an initial CALR-mutant PDX experiment, abrogation of STIM1 led to decreased human CD45+ cell engraftment in conjunction with prolonged survival. In a second CALR-mutant PDX experiment, STIM1 targeting did not overtly impact engraftment or survival. In contrast, CRISPR ablation of STIM1 in JAK2-mutant PDX models led to exacerbated disease phenotypes, including increased human CD45+ cell engraftment, increased splenomegaly, and early lethality. Altogether, this study represents the first demonstration of MF development in patients with Stormorken Syndrome and reveals a previously unrecognized role of aberrant SOCE underlying SS and MPNs. We recapitulated these findings in a novel conditional mouse model. These findings also illustrate unique interactions between STIM1 and JAK2 and CALR driven MPNs. These observations have important implications for the development of targeted therapeutic approaches for these disorders.