Paludo, Jonas

Abstract Waldenstrom macroglobulinemia (WM) is a lymphoplasmacytic lymphoma associated with an increase in serum immunoglobulin M (IgM) monoclonal protein. The underlying causes of the disease are still not fully understood, but cells in the bone marrow (BM) microenvironment (TME) are anticipated to play a significant role in disease progression. We previously reported that myeloid-derived suppressor cells (MDSCs), a phenotypically heterogeneous group of cells known for their ability to suppress immune responses, are expanded in the bone marrow of patients with WM and that the expansion of MDSCs is particularly notable for increased numbers of polymorphonuclear MDSCs (PMN-MDSCs). In the present study, we evaluated the ability of WM cells to attract MDSCs to the tumor microenvironment (TME). We also evaluated whether PMN-MDSCs suppress the cytotoxic function of T-cells and thereby promote the survival of WM cells. Our study confirms the expansion of MDSCs, notably PMN-MDSCs in BM of WM patients as compared to normals. We also found that PMN-MDSCs expressing CD66b was a predominant population in WM patients. To determine the phenotype and transcriptome of both PMN-MDSCs and monocytic MDSCs (M-MDSCS) in WM, we compared BM aspirates from WM patients to normal bone marrow (NBM) and performed cellular indexing of transcriptomes and epitopes by sequencing (CITEseq) analysis (WM=3, NBM=2) followed by qPCR validation. We found that MDSCs in WM express significantly higher levels of HIF1-α overall (p<0.02) compared to NBM. Furthermore, PMN-MDSCs are more abundant in WM bone marrow than M-MDSCs and express significantly higher levels of CXCR4 (p<0.001), myeloperoxidase (MPO; p<0.0003), and azuricidin (AZU1; p<0.0004), suggesting an activated phenotype. To determine whether WM cells attract MDSCs to the BM, we performed a transwell assay and showed that WM cells (BCWM.1 cell line) attracted MDSCs, particularly PMN-MDSCs, to a greater extent than M-MDSCs (n=3; p<0.001). We next evaluated whether MDSCs prevent cytotoxic T-cells from lysing WM cells. In-vitro co-culture studies with BCWM.1 cells and T-cytotoxic cells (B:CD8-T) showed that T-cytotoxic cells were able to lyse the WM cell line by 46.4% , (median range 46.4 - 47.8%, p=0.001). However, when BCWM.1 cells and T-cytotoxic cells were co-cultured in the presence of MDSCs (i.e B:CD8-T:M), the specific lysis of BCWM.1 cells was reduced by 12.6% respectively (range 2.2- 12.6%, p<0.001). Furthermore, we observed that B:CD8-T:M co-culture significantly reduced CD8-T cell activation as compared to activated T cells alone (17.9%) (median range 17.8- 18%, p<0.001). In contrast, no affect was observed with B: CD8-T coculture by 91% (range 90.6- 91.4% p<.001) confirming that MDSCs suppress CD8-T cell activation. Our study therefore provides evidence that MDSCs in WM have an activated phenotype and that PMN-MDSCs in WM are attracted by malignant cells in the bone marrow. Furthermore, we find that the increased presence of PMN-MDSCs in the WM bone marrow environment suppresses T-cell lysis of malignant cells and may thereby contribute to disease progression. These findings provide a strong rationale for targeting MDSCs as a potential therapeutic strategy in WM.

Background: Despite effective front-line therapies, DLBCL & MM have high relapse rates. Novel therapies like CAR-T & bispecific antibodies (BsAb) offer deep & durable remission compared to salvage therapies, but have a unique toxicity profile (TP) that requires monitoring. With no head-to-head trials, the rationale for therapy selection may vary. This study explores provider choices, sequencing preferences, & access barriers. Methods: Two anonymous surveys (MM and DLBCL) of US-based Hematologists were conducted from 2/2024 to 6/2024, on practice settings, CAR-T & BsAb availability, treatment preferences & barriers. Responses were analyzed to assess preferences across clinical scenarios. Results: Surveys were completed by 37 MM & 45 DLBCL providers. Most were at centers offering both CAR-T & BsAb. Most gave CAR-T inpatient, & BsAb outpatient. CAR-T was given by 73% MM providers & 66% DLBCL (92% used Cilta-cel, 96% used Ide-cel; 100% used Axi-cel, 90% used Liso-cel & 73% Tisa-cel). In MM, 78% used Teclistamab, 60% Talquetamab & 24% Elranatamab. In DLBCL, 62% used Epcoritamab & 40% Glofitamab. Providers preferred CAR-T over BsAb in MM after 4+ prior lines of therapy (LOT) (89%), & in DLBCL after 2+ prior LOT (93%). CAR-T was favored more among academic providers & those at CAR-T centers. Practice volume did not impact selection. Among providers at centers with BsAb-only, CAR-T was preferred by 60% (MM) and 50% (DLBCL). Providers at centers without both favored CAR-T (MM: 75%, DLBCL: 86%). CAR-T was chosen for better outcomes, short treatment, center choice, & prior experience. Other factors that influenced decisions were lower incidence of cytopenia/infection & “BsAb effective post-CAR-T, less so vice versa” (MM), & “curative potential of CAR-T” (DLBCL). Logistical barriers & institutional constraints influenced BsAb selection: CAR-T unavailability, access to specialized centers, referral complexity, insurance delay, manufacturing delay & hospitalization needs. In MM, 78% providers preferred Cilta-cel over Ide-cel, especially academic providers & those with CAR-T experience. CAR-T volume did not impact selection. Cilta-cel was preferred due to perceived better response rate (RR), more experience, “progression-free survival (PFS) benefit”, center choice, & logistical issues with Ide-cel. Ide-cel was selected due to center choice, favorable TP & insurance preference. Teclistamab was preferred by 78% due to more experience, better RR, TP & center choice, Elranatamab by 11% (better TP, RR, & “PFS”), & Talquetamab by 11% due to having a non-BCMA target. Academic providers preferred Teclistamab (89% vs 50%), & nonacademic providers preferred Talquetamab (38% vs 4%) and Elranatamab (13% vs 8%). In DLBCL after 2+ LOT, 69% favored Axi-cel due to prior experience, RR, center choice, & logistical issues with other products. Liso-cel & Tisa-cel were chosen by 27% & 2%, respectively, due to favorable TP. Axi-cel was also preferred 2nd line by 73% for similar reasons, while 24% favored Liso-cel for its favorable TP. Epcoritamab was preferred by 56% (subcutaneous injection, more experience & center choice) & Glofitamab by 42% (shorter treatment duration, easy administration, more experience, & center choice). Practice setting & volume did not impact selection between products in DLBCL. InBCMA-naïve MM pts, majority (97%) chose an anti-BCMA agent over GPRC5D due to better RR, TP, prior experience, & center choice. InBCMA-exposed pts, most (92%) favored an anti-GPRC5D agent. In contrast in DLBCL, 80% chose CD19 CAR-T after prior CD19-directed therapy, & 18% chose CD20 BsAb. Therefore, antigen escape was less of a concern in DLBCL vs MM. For relapse after 2nd line CAR-T in DLBCL, 78% preferred BsAb over other salvage options due to better RR, TP, & more experience. Providers prioritized RR, survival, & TP in decision making. Treatment logistics, duration, frequency, & cost were less crucial. MM providers also prioritized quality of life, which was less so in DLBCL. There were no differences in priority based on practice setting & experience with CAR-T. Conclusion: The study highlights factors influencing therapy choices in MM & DLBCL. Despite CAR-T's potential for better outcomes, significant access barriers exist. Efforts to streamline referral pathways, logistics, & expanding availability are crucial to improve outcomes. Understanding these preferences & barriers can guide future management strategies.

Abstract Purpose To assess incidence and risk factors for anthracycline-induced left ventricular dysfunction, and associated mortality in order to inform patients and providers of the risk, as well as for clinical trial endeavours. Background Anthracyclines (ACs) are very effective cancer therapeutics, but their use is confronted by the risk of cardiotoxicity. Estimates of AC-induced cardiotoxicity based on systematic analyses are limited and have not recently been defined. We present here a systematic review and meta-analysis of the incidence of anthracycline induced subclinical (AC-LVD) and clinical LV dysfunction (AC-CHF) and AC-CHF related mortality in non-liposomal AC recipients over the last 3 decades. Methods We searched Ovid MEDLINE(R), Embase, Cochrane Database and other major databases from their inception until June 22, 2023 for studies examining the rate of AC induced cardiomyopathy in patients receiving ACs and anthraquinones for haematologic, breast (BC) and sarcomatous malignancies. Covidence was used to aid with the management of our systematic review. Screening of abstracts and data extraction was independently performed by 2 reviewers. Results 6,365 abstracts were reviewed, of which 74 were included in the final analysis. 72 studies (n=41,138) reported data on AC-LVD, and 47 studies (n=27,012) reported data on AC-CHF. The average age was 53 years and 50% of the patients were female. Data on cardiac function at baseline was available in only 70% of the studies. Overall, AC-LVD occurred in 8.2% (95% CI 6.8%-10.2% I2 97%) (Fig 1). AC-CHF was noted in 2.2% (95% CI 1.6% to 2.9%, I2 91%) (Fig 2A), mortality related to decompensated AC-CHF was 0.5% (95% CI 0.3%-0.9%, I2 88%) (Fig 2B) while total mortality rate among AC recipients was 24.9% (95% CI 14.8-38.7%, I2 99%). Incidence of AC-LVD was higher in doxorubicin recipients (vs other ACs 10.5% vs 5-5.6%), studies from North America & Asia (vs other regions; 9.3-9.6% vs 6.7%), and prospective studies (vs retrospective studies; 8.7% vs 7.5%). AC-LVD was more common in BC patients (vs other cancers; 8.7% vs 3.5-7.2%) while AC-CHF was more common in lymphoma patients (8.5% vs 1.6-4.1%). AC-LVD was 4.6 times (95% CI 1.2-17.1, p=0.02) more likely in patients with a mean age >70 (vs <70). 21 definitions for AC-LVD were used across the studies. The most common definition (15 studies; n=7904) was "any decline in LVEF from baseline" resulting in a 5.8% rate of AC-LVD. Conclusions AC-LVD and AC-CHF has been reported in up to 10% and 3% of AC recipients, respectively. AC-CHF related mortality occurs in <1% of anthracycline recipients and has a 2-3% share in overall mortality among these patients. The type of malignancy and anthracycline can influence the rate of AC-LVD, AC-CHF and AC-CHF related mortality. Global region, age of the patient population and study design also seem to influence incidence rates. There is noteworthy heterogeneity in definitions for cardiotoxicity across studies.

Abstract Background Hypomagnesemia has been correlated with inferior outcomes in patients with large B cell lymphoma (LBCL) undergoing stem cell transplants. As T-cell and myeloid cell dysfunction have been associated with low magnesium conditions, we investigated whether serum magnesium (Mg) levels could predict clinical outcomes in LBCL patients who received chimeric antigen receptor T-cell therapy. Methods Patients with LBCL who received axi-cel under the ZUMA-1 trial or as FDA approved therapy at Mayo Clinic were examined. Serum samples were obtained at specified time points and cytokine analysis was performed. Single cell RNA sequencing was performed on peripheral blood mononuclear cells. The Student T-test, Kruskal Wallis, or Fisher's Exact Tests were used to compare differences in demographics across Mg levels. Survival curves were plotted using the Kaplan-Meier methodology and compared using the Wilcoxon test. Results We found that hypomagnesemia before lymphodepletion chemotherapy predicted for inferior progression-free and overall survival in the pivotal study ZUMA-1 (NCT02348216). These results were validated in an independent cohort of LBCL patients receiving axicabtagene ciloleucel (axi-cel) at Mayo Clinic. Hypomagnesemia correlated with increased inflammatory serum markers and cytokine levels including ferritin, IL-6, IL1Ra, IL-8, and MIP1a. scRNAseq analysis unveiled altered immune interactions between monocytes and T cells with a concordant immune suppressive transcriptome. Conclusions Hypomagnesemia at the time of CAR-T infusion associates with an unfavorable inflammatory profile and decreased response and survival in LBCL patients receiving axi-cel. These findings suggest a potentially actionable prognostic factor for patients with large cell lymphoma undergoing CAR-T.

The infusion autograft absolute number of inhibitory killer immunoglobulin-like receptor (KIR) 2DL2 and activating natural killer (NK)p30 cells are predictors of clinical outcomes in lymphoma patients undergoing autologous peripheral blood hematopoietic stem cell transplantation (APBHSCT). To assess if the long-term recovery of these NK cell subsets still holds clinical relevance, we set up to investigate their prognostic ability at day 100 post-APBHSCT. This was a retrospective single-institution study including 107 patients from our prior phase III trial who had a clinical assessment at day 100 post-APBHSCT. The median follow-up from day 100 was 168.19 months (interquartile range: 156.85–181.28 months). Patients with day 100 inhibitory KIR2DL2 < 0.08 cells/µL and activating NKp30 ≥ 0.19 cells/µL experienced superior overall survival (OS) and progression-free survival (PFS). A multivariate analysis revealed both the day 100 inhibitory KIR2DL2 [OS: HR = 1.449, 95%CI, 1.231–1.895, p < 0.013; and PFS: HR = 2.069, 95%CI, 1.134–3.775, p < 0.021] and activating NKp30 [OS: HR = 4.985, 95%CI, 2.614–9.506, p < 0.0001; and PFS: HR = 4.661, 95%CI, 2.598–8.393, p < 0.0001] were independent predictors for OS and PFS. Inhibitory KIR2DL2 and activating NKp30 NK cells at day 100 are prognostic immune biomarkers in lymphoma patients treated with APBHSCT.

Abstract Unirradiated patients with relapsed/refractory (R/R) B-cell non-Hodgkin lymphoma (NHL) who undergo anti-CD19 chimeric antigen receptor T-cell therapy (CART) have a predominant localized pattern of relapse, the significance of which is heightened in individuals with limited/localized disease before CART. This study reports on the outcomes of patients with R/R NHL and limited (&lt;5 involved sites) disease bridged with or without radiotherapy. A multicenter retrospective review of 150 patients with R/R NHL who received CART with &lt;5 disease sites before leukapheresis was performed. Bridging treatment, if any, was administered between leukapheresis and CART infusion. Study end points included relapse-free survival (RFS), event-free survival (EFS), and overall survival. Before CART infusion, 48 patients (32%) received bridging radiotherapy (BRT), and 102 (68%) did not. The median follow-up was 21 months. After CART infusion, BRT patients had higher objective response (92% vs 78%; P = .046) and sustained complete response rates (54% vs 33%; P = .015). Local relapse in sites present before CART was lower in the BRT group (21% vs 46%; P = .003). BRT patients had improved 2-year RFS (53% vs 44%; P = .023) and 2-year EFS (37% vs 34%; P = .039) compared with patients who did not receive BRT. The impact of BRT was most prominent in patients who had ≤2 pre-CART involved disease sites, with 2-year RFS of 62% in patients who received BRT compared with 42% in those who did not (P = .002). BRT before CART for patients with limited (&lt;5 involved disease sites) R/R NHL improves response rate, local control, RFS, and EFS without causing significant toxicities.

Abstract Currently, the role of DNA methylation in the immunoglobulin M (IgM) monoclonal gammopathy disease spectrum remains poorly understood. In the present study, a multiomics prospective analysis was conducted integrating DNA methylation, RNA sequencing (RNA-seq), and whole-exome sequencing data in 34 subjects (23 with Waldenström macroglobulinemia [WM], 6 with IgM monoclonal gammopathy of undetermined significance [MGUS], and 5 normal controls). Analysis was focused on defining differences between IgM gammopathies (WM/IgM-MGUS) compared with controls, and specifically between WM and IgM-MGUS. Between groups, genome-wide DNA methylation analysis demonstrated a significant number of differentially methylated regions that were annotated according to genomic region. Next, integration of RNA-seq data was performed to identify potentially epigenetically deregulated pathways. We found that pathways involved in cell cycle, metabolism, cytokine/immune signaling, cytoskeleton, tumor microenvironment, and intracellular signaling were differentially activated and potentially epigenetically regulated. Importantly, there was a positive enrichment of the CXCR4 signaling pathway along with several interleukin (interleukin 6 [IL-6], IL-8, and IL-15) signaling pathways in WM compared with IgM-MGUS. Further assessment of known tumor suppressor genes and oncogenes uncovered differential promoter methylation of several targets with concordant change in gene expression, including CCND1 and CD79B. Overall, this report defines how aberrant DNA methylation in IgM gammopathies may play a critical role in the epigenetic control of oncogenesis and key cellular functions.

9018 Background: Comprehensive hematology education during fellowship is paramount for providing high-quality care, especially in busy practices. Variability in fellow educational experiences is expected. However, fellowship programs must ensure a well-rounded, equitable educational experience for all fellows to prepare them for practice and board exams in the increasingly dynamic field of hematology. Methods: We administered an e-mail survey to hematology/oncology fellows (n=30) at Mayo (Rochester) to explore inpatient hematology education experiences and the Fellow Hematology Curriculum (FHC) during 2021-2022. Based on results, we designed a fellow-led, consultant-supervised quality improvement (QI) initiative to improve FHC. Responses were in a five-point Likert scale and open-ended format. Results: Twenty fellows completed the survey (66% response rate). Respondents were well distributed (35% PGY4, 25% PGY5, 40% PGY6). Resources most used for self-education were ASH SAP (85%), UpToDate (80%), and NCCN guidelines (50%). 100% of respondents agreed/strongly agreed that the FHC could be improved. 95% agreed/strongly agreed that there is a high degree of variability among consultants regarding engagement in fellows’ education. 45% agreed that education is adequately paired with relevant clinical duties to reinforce key concepts. 25% agreed that FHC had well-defined structure and objectives. In the open-ended responses, a structured curriculum covering a set list of topics paired with the relevant rotation was the most common recommendation by the fellows for improving the FHC and the inpatient hematology educational experience. This prompted a QI initiative to implement a structured, evidence-based curriculum covering essential topics required for hematology practice. Each week of a four-week rotation has a relevant theme (eg, “Marrow Failure week” during the inpatient leukemia service). Fellows selected a topic of interest and created a lecture to be used by all faculty and learners during the relevant rotation (eg, “Aplastic Anemia” during the Marrow Failure). Consultants and pharmacists in respective disease groups reviewed the topics and presentations. These lectures are now provided to fellows and faculty via e-mail prior to relevant rotations. Conclusions: Our survey demonstrated that trainees’ individual educational experiences on inpatient hematology services are widely variable and largely unsatisfying without some form of structured, relevant learning. We identified the need for deliberate, coordinated education as a gap to bridge toward achieving the goal of equitable, comprehensive learning. We intend to perform a post-implementation survey after one year of practicing a structured, relevantly paired FHC. This assessment will be used to make improvements in subsequent iterations of the FHC using the Plan-Do-Study-Act methodology.

e14535 Background: Cytokine release syndrome (CRS) is the most common complication of chimeric antigen receptor T cells (CAR-T) for hematologic malignancies. We hypothesized that patients undergoing CAR-T therapy who are on metformin will experience a lower incidence or severity of CRS than patients not on this medication. Methods: We evaluated all patients treated with CAR-T at Mayo Clinic Rochester between 1/1/2018 and 1/31/2023, and abstracted relevant clinical and laboratory characteristics to determine the incidence and severity of CRS, PFS as well as OS in patients who were, or were not, on metformin before and during CAR-T therapy for Type 2 diabetes mellitus. We used the Chi-Square test, Mann-Whitney test and the Kaplan-Meier method to analyze data. Results: In the 5-year interval under consideration 237 patients received CAR-T, 168 for non-Hodgkin lymphoma (NHL), and 69 for multiple myeloma (MM). 23 patients (15 NHL and 8 MM) were on metformin at the time of CAR-T. 15 patient developed CRS while on metformin and 175 patients not on metformin (p=0.05). Grade 3 or 4 CRS incidence was 4% in both groups (p=0.41). Of the 153 patients with NHL not on metformin, 121 developed any grade of CRS, while 10 of 15 patients on metformin developed CRS (p=0.268). 4 NHL patients developed CRS Grade ≥3 or higher, while 0 of 15 on metformin developed Grade ≥3 CRS (p=0.237). For the MM patients, the incidence of CRS was 49 patients not on metformin and 5 patients on metformin (p<0.001). 3 MM patients developed CRS Grade ≥3 or higher, while 0 of 8 on metformin developed Grade ≥3 CRS (p=0.346). 12 of 15 patients on metformin with NHL achieved CR compared to 92 of 153 patients not on metformin (p=0.1305). 3 of 8 patients with MM on metformin compared to 24 of 61 patients with MM not on metformin achieved CR or better post CAR-T (p=0.92). Tocilizumab was used in 40% of patients on metformin but in 66% of patients not on metformin (p=0.0016). In NHL patients, PFS from CAR-T infusion was not reached (range: 6.3 to NR) for patients on metformin versus 9.9 months (range: 6.3–19.5) in patients not on metformin (p = 0.3332). In the MM patients, progression-free survival (PFS) from CAR-T infusion was 8.4 months (95% CI: 7.9–not reached [NR]) for patients on metformin and 22.6 months (13.7–NR) for those not on metformin (p = 0.07). Median OS for NHL patients on metformin was not reached (9.766–NR) versus 32.6 months (18.6–NR) for patients not on metformin (p = 0.7022). Median OS was 17 months (95% CI: 8.3 to NR) for patients with MM on metformin vs NR (95% CI: 9.4 to NR) for patients not on metformin (p = 0.2559). Conclusions: In this retrospective study, the co-administration of metformin during CAR-T cell therapy for NHL and MM appears to be associated with a numerically reduced incidence of CRS. The effect appears to be particularly marked in patients with multiple myeloma. Metformin therapy did not seem to affect the response to CAR-T therapy in either NHL or MM.

1564 Background: Physical function (PF) is a core outcome in cancer trials that can inform treatment tolerability. PF can be assessed across modalities including by patient (pt) report, clinician (clin) report, performance tests and digital sensors. The optimal approach for PF measurement has not been defined. Methods: In4M is a prospective observational study evaluating PF in pts initiating chemotherapy for breast cancer or lymphoma followed over 9 months. We used an electronic platform (Hugo) to aggregate serially assessed pt-reported PF, clin- and pt-reported ECOG performance status (PS), 6-minute walk test (6MWT) and wearable (Fitbit) data. We evaluated feasibility in all pts; associations among PF measures and the ability of PF measures to detect change were evaluated in pts with both baseline (BL) and 3-month (3M) data. We used Pearson correlations to assess the relationship of average daily steps with measures of PF. To assess change, we compared each PF measure from BL to 3M using a 2-sided α=.05 paired t-test. Results: Of the208 pts enrolled, 193 (93%) and 159 (76%) completed surveys, and 121 (58%) and 124 (60%) provided wearable data (evaluable: ≥1 day data) at BL and 3M, respectively. When requiring a stringent BL with 4+ days of data with ≥1 weekend day to be evaluable, 88 (42%) and 120 (58%) provided wearable data at BL and 3M. At both BL and 3M, pt-reported PF was generally more highly correlated with wearable-derived daily steps than clin-reported PF and 6MWT were (Table). Significant declines in PF were observed for pt- and clin-reported PF (all p<0.05), but not other modalities. Conclusions: Longitudinalassessment of multi-modal PF data including pragmatic wearable data collection was feasible in pts receiving chemotherapy. Pt-reported PF was the only modality that was both correlated with step count and able to detect change. Additional wearable metrics, correlations among PF measurements and outcomes, and clinically meaningful change scores will be reported. Optimal PF measurement may improve future reporting of treatment tolerability. Clinical trial information: NCT05214144 . [Table: see text]