Aggressive non-Hodgkin Lymphoma Research Articles

Machine Learning-Based Time-Series Clustering Identifies Archetypal Trajectories of Hematotoxicity after CAR T-Cell Therapy

BACKGROUND Severe hematotoxicity has been reported in a subset of patients (pts) after chimeric antigen receptor (CAR) T-cell therapy, leading to severe infections, transfusion dependency, and worse outcomes. Early identification of pts at risk for severe hematotoxicity could inform the consideration of an autologous stem cell boost, allogeneic HCT, and/or additional infectious prophylaxis. While distinct archetypal trajectories of blood count recovery have been proposed (e.g., quick vs. intermittent vs. aplastic), manual trajectory classification is subjective and labor-intensive. To address these limitations, we automated the identification of distinct trajectories of hematotoxicity by applying time-series clustering to longitudinal absolute neutrophil count (ANC) data from >400 pts treated with CAR T-cell therapy. Next, we sought to identify factors associated with the identified trajectories and to assess the predictive ability of the CAR-HEMATOTOX score (Rejeski, Blood 2021). STUDY DESIGN AND METHODS Adults ≥18 years of age who received their first infusion of CAR T cells for hematologic malignancies with commercial or investigational products at the Fred Hutchinson Cancer Center (2013-2023) were included. ANC trajectories were clustered using non-supervised longitudinal k-means based on Euclidean distances using the latrend and kml packages in R 4.1.3. We applied logistic regression to pre-lymphodepletion (pre-LD) variables to predict poor/very poor ANC recovery. Sensitivity and specificity were computed using a probability threshold based on the Youden criteria. RESULTS A total of 509 pts were identified; 106 were excluded due to insufficient data, with 403 pts included for the analysis. The most common disease types were aggressive NHL (n = 161; 40%), indolent NHL (n = 82; 20%), and ALL (n = 74; 18%). CAR T-cell products were axi-cel, n= 101 (25%); brexu-cel, n = 24 (6%); cilta-cel, n = 21 (5%); liso-cel, n = 46 (11%); ide-cel, n = 25 (6%); tisa-cel, 12 (3%); and investigational CD19 or CD20 CAR T-cell products, n = 174 (43%). As shown in Figure 1A, the ANC longitudinal data clustered into four distinct trajectories as follows: 1) high nadir followed by rapid recovery (“very good recovery”), n = 294 (73%); 2) low nadir followed by rapid recovery (“good recovery”), n = 87 (22%); 3) low nadir followed by intermittent recovery (“poor recovery”), n = 13 (3%); 4) aplastic phenotype (“very poor recovery”), n = 9 (2%). In univariate logistic regression, ALL (reference: aggressive NHL; OR = 5.43, 95% CI, 1.71-20.6, p = 0.006), pre-LD lower ANC (OR = 3.33 per log 10 , 95% CI, 1.82-6.25, p < 0.001), lower pre-LD platelet count (OR = 10 per log 10, 95% CI, 3.70-33.3), p < 0.001), and higher pre-LD disease burden as measured by lactate dehydrogenase (LDH) were associated with poor/very poor ANC recovery (OR = 16.6 per log 10, 95% CI, 4.79-59.3, p < 0.001). Next, we assessed the predictive ability of CAR-HEMATOTOX to identify poor/very poor ANC recovery in n = 356 pts with a calculable score. The CAR-HEMATOTOX score high/low showed modest discrimination (C-index: 0.65), which improved when used as a continuous variable (C-index: 0.85). As previously reported, these models had high sensitivity (>90%) but low specificity (high/low: 31%, continuous: 58%). To improve risk prediction of poor/very poor ANC recovery, we built a logistic regression model using restricted cubic splines - allowing for non-linear effects - including pre-LD ANC, platelet, Hb, LDH, CRP, and ferritin, which showed higher discrimination (C-index: 0.91), high sensitivity (88%), and higher specificity (79%) ( Figure 1B). CONCLUSION We introduce an automated, scalable, and disease-agnostic framework to analyze hematologic toxicity after CAR T-cell therapy. K-means clustering of longitudinal ANC data categorized pts into archetypal trajectories of hematologic recovery. A logistic regression model using pre-LD ANC, platelet, Hb, LDH, CRP, and ferritin showed improved discrimination and sensitivity compared to CAR-HEMATOTOX in our training set, but both models had low specificity (poor ability to “rule in” pts at risk of severe hematotoxicity) and thus low clinical utility at this time. We plan to further improve our predictive models by incorporating post-infusion biomarkers of systemic inflammation that we have previously shown to be associated with delayed count recovery after CAR T-cell therapy (Juluri, Blood Advances 2022).

Efficacy and Safety of Valemetostat Monotherapy in Patients with Relapsed or Refractory Peripheral T-Cell Lymphomas: Primary Results of the Phase 2 VALENTINE-PTCL01 Study

Introduction: Peripheral T-cell lymphoma (PTCL) is an aggressive non-Hodgkin lymphoma (NHL), with limited available treatment options for patients (pts) with relapsed or refractory (R/R) disease. Valemetostat tosylate (valemetostat) is a novel and potent dual inhibitor of enhancer of zeste homolog (EZH)2 and EZH1, which is approved in Japan for the treatment of R/R adult T-cell leukemia/lymphoma (ATLL). Here, we report primary results for pts with R/R PTCL treated with valemetostat in the open-label, single-arm, global, phase 2 VALENTINE-PTCL01 study (DS3201-A-U202; NCT04703192). Methods: Pts were ≥ 18 years of age, had a confirmed diagnosis of PTCL, had R/R disease after ≥ 1 prior line of systemic therapy, and pts with anaplastic large cell lymphoma (ALCL) had received prior brentuximab vedotin treatment. Pts received oral valemetostat 200 mg/day in continuous 28-day cycles until disease progression or unacceptable toxicity. The primary endpoint was objective response rate (ORR), assessed by blinded independent central review of PTCL following computed tomography (CT)-based response assessment according to Lugano 2014 criteria. Secondary efficacy endpoints included duration of response (DOR), progression-free survival (PFS), and overall survival (OS). Positron emission tomography (PET)-CT-based response assessment by Lugano 2014 criteria was an exploratory endpoint. Efficacy analyses included all pts who received ≥ 1 dose of valemetostat and had an eligible PTCL subtype confirmed by central hematopathology review; safety analyses included all pts who received ≥ 1 dose of valemetostat. Results: A total of 133 pts with R/R PTCL were enrolled and received ≥ 1 dose of valemetostat. Pts had a median age of 69 years (range 22-85) and 91 pts (68.4%) were male. Pts had received a median of 2 prior lines of therapy (range 1-12) and 35 pts (26.3%) received prior hematopoietic cell transplant (HCT; autologous, n = 32; allogeneic, n = 5). PTCL subtype eligibility was confirmed in 119 pts: 42 pts (31.6%) had angioimmunoblastic T-cell lymphoma (AITL), 41 (30.8%) had PTCL, not otherwise specified (PTCL, NOS), 9 (6.8%) had ALCL (7 [5.3%] ALK-negative and 2 [1.5%] ALK-positive), 8 (6.0%) had nodal PTCL with T follicular helper cell phenotype, and 19 (14.3%) had other PTCL subtypes. As of data cutoff (May 5, 2023), 32 pts (24.1%) were still receiving treatment; reasons for treatment discontinuation included progressive or relapsed disease in 46 pts (34.6%), clinical progression in 19 pts (14.3%), adverse event in 13 pts (9.8%), and 12 pts (9.0%) discontinued study drug to proceed with allogeneic HCT. Median treatment duration was 18 weeks (range 0.3-93.4) and median duration of follow-up was 10.5 months (range 0.2-21.5). Among 119 efficacy-evaluable pts, the CT-based ORR was 43.7% (n = 52) (95% confidence interval [CI], 34.6-53.1), including 17 pts (14.3%) achieving CR and 35 pts (29.4%) achieving PR as best overall response. Median DOR was 11.9 months (95% CI, 7.8-not evaluable [NE]) and median time to response was 8.1 weeks (range 5-37). ORR by PTCL subtype ranged from 31.7% for PTCL, NOS to 54.8% for AITL (Table 1). Using PET-CT-based response assessment, ORR was 52.1% (n = 62) (95% CI, 42.8-61.3), including 32 pts (26.9%) with a complete metabolic response. Median CT-based PFS was 5.5 months (95% CI, 3.5-8.3) and median OS was 17.0 months (95% CI, 13.5-NE). Of 133 pts in the safety analysis set, 128 pts (96.2%) experienced ≥ 1 treatment-emergent adverse event (TEAE) of any grade, 77 pts (57.9%) experienced grade ≥ 3 TEAEs, and 53 pts (39.8%) experienced serious adverse events. The most common all grade and grade ≥ 3 TEAE was thrombocytopenia (Table 2). Overall, 13 pts (9.8%) experienced a TEAE that led to treatment discontinuation, 21 pts (15.8%) had a TEAE that led to dose reduction, and 66 pts (49.6%) had a TEAE that led to dose interruption. Conclusions: Valemetostat demonstrated a high ORR of 43.7% with durable responses (median DOR of 11.9 months) in pts with R/R PTCL, and responses were observed across all PTCL subtypes. A valemetostat dose of 200 mg/day was tolerable; safety analysis showed that the most common TEAEs were cytopenias. These primary results from the VALENTINE-PTCL01 study suggest that valemetostat provides a clinically meaningful benefit for pts with R/R PTCL.

Estimation of Relapsed/Refractory Follicular Lymphoma Patients on Therapy in the United States

Background: Follicular lymphoma (FL) represents approximately 20-25% of non-Hodgkin lymphomas (NHL). In contrast to patients with more aggressive NHL, patients with FL have a heterogenous course of disease in which some can spend extended periods in an initial watch-and-wait state and/or within a single line of treatment. As such, these patients may have multiple relapses and a variety of clinical features that may impact disease course (eg, double-refractory disease, increased Follicular Lymphoma International Prognostic Index (FLIPI) scores, high Ann Arbor disease stage, older age, etc.). The objective of this study was to estimate the number of treated patients with FL in each line of therapy (LOT) to better understand the proportion with relapsed or refractory (R/R) disease in the US while accounting for distributions of LOTs within a baseline prevalent population and annual trends in treatment initiation by LOT. Methods: Two companion models were developed from a US perspective: 1) an epidemiologic model that utilized FL incidence and survival data from the Surveillance, Epidemiology, and End Results (SEER) program to project the size of the prevalent population; and 2) a state-transition model describing the proportion of patients with FL initiating treatment and time on treatment by line of therapy. Within the state-transition model, hypothetical incident cohorts were simulated annually to follow FL treatment patterns and survival trajectories until a “steady-state” was reached regarding the distribution of LOTs in the prevalent population. Model inputs were based on data from published literature. Several published sources were identified regarding estimates of treatment rates by line of therapy, with base-case values based on physician survey: 51% for first LOT, 32% for second LOT, and 31% for third and successive LOTs. This distribution was then applied to a baseline prevalent population and subsequent incident cohorts were modeled annually over a 3-year time horizon. Results: In 2022, the estimated FL prevalent population in the US was 154,922. Over a subsequent 3-year period combining the prevalent patients with newly incident patients, the model projected 10,399 patients initiating first-line treatment, 3197 initiating second-line treatment, and 975 initiating third-line treatment. Conclusions: While incidence of FL initial diagnosis in the US is well reported, there are limited data describing prevalence or population makeup with respect to LOT. While many patients can benefit from long remissions after first-line therapy, a portion of patients experience R/R disease and require further treatment. Estimation of the R/R FL population can support with planning of treatment needs as novel therapies emerge in the US.

Nivolumab in Relapsed or Refractory Primary CNS Lymphoma: Multicenter, Retrospective Study

BACKGROUND Primary central nervous system lymphoma (PCNSL) is an aggressive non-Hodgkin's lymphoma confined in the brain and/or spinal cord. While high-dose methotrexate (HDMTX) has been considered the standard of care for newly-diagnosed patients, the optimal treatment strategies for relapsed disease remain challenging, and the outcomes are dismal. Given that 40~50% of PCNSL tissues exhibit aberrancy on 9p24.1 ( PD-L1/PD-L2), immune-checkpoint inhibitors (ICI) may work for the disease. However, there still have been limited data due to the rarity of the disease and the limited accessibility of the drug. To further elucidate the role of ICI in PCNSL, we carried out a nationwide retrospective analysis. METHOD Patients treated with nivolumab for relapsed PCNSL were included in the analysis. Medical records were retrospectively reviewed, and characteristics, including age, sex, prior treatment, risk factors, response, and survival outcomes, were collected. RESULTS From November 2018 to September 2022, 22 consecutive patients were included in the analysis from 4 tertiary institutes in Korea. The median age at diagnosis was 66, and male: female ratio was 1:1. The histologic subtype was diffuse large B-cell lymphoma in all cases, and in 15 cases of which cell-of-origin was identified, 12 cases (80%) were non-GCB subtype. In terms of frontline-treatment, 21 patients (95%) had received HDMTX + procarbazine + vincristine, and rituximab was administered in 73% (16/22) of patients. Nivolumab (at a dose of 3mg/kg every 2 weeks) was treated after median 3 lines (range, 2 - 6) of therapy and at the median age of 67 (range, 37 - 82). Four (18%), twelve (55%), and six (27%) patients were classified as low-, intermediate-, and high-risk groups by IELSG criteria, respectively. Eleven patients (50%) were refractory to the last treatment prior to nivolumab. With a median follow-up duration of 22.3 months (95% CI, 13.1 - 31.5), nine patients (41%) had an objective response (6 complete responses, 3 partial responses). The median time to response was 3.0 months (95% CI, 0.7 - 5.3 months), and the median duration of response was 20.9 months (95% CI, 1.7 - 40.0). The median progression-free survival (PFS) was 2.1 months (95% CI, 0.2 - 4.0), and the 1-yr PFS rate was 36.9% (± 10.9%). The median overall survival (OS) was 18.9 months (95% CI, 5.0 - 32.8), and the 1-yr OS rate was 53.1% (± 13.9%). As expected, responders showed a strong tendency for prolonged OS (36.3 mo. vs. 8.6 mo., p = 0.058). While earlier use (3L vs. 4L or later) of nivolumab was not associated with a higher response rate, it resulted in statistically non-significantly prolonged OS (36.3 months vs. 8.6 months, p = 0.164). Nivolumab was very well-tolerated as no dose reduction was required, and treatment delay was required in two cases (one infection and one seizure). CONCLUSION Given that only restricted treatment strategies are available for relapse PCNSL patients, our study suggests that nivolumab can be a suitable option for the disease. Further studies are warranted to determine predictive biomarkers for nivolumab treatment given the excellent survival duration in responders.

Subcutaneous Mosunetuzumab Is Active and Has a Manageable Safety Profile in Patients with Previously Untreated, Low-Tumor Burden Follicular Lymphoma: Initial Results from the Phase II Morningsun Study

Background: Mosunetuzumab is a CD20xCD3 bispecific antibody that is approved for the intravenous (IV) treatment of patients with relapsed/refractory (R/R) follicular lymphoma (FL) who have received at least two prior lines of systemic therapy, and is suitable for administration in the outpatient setting. In an ongoing Phase I/II study (NCT02500407), IV mosunetuzumab induced durable responses and had a manageable safety profile in patients with heavily pre-treated R/R indolent and aggressive B-cell non-Hodgkin lymphomas (NHL) when administered as a fixed-duration treatment (Budde et al. J Clin Oncol 2022). Subcutaneous (SC) dosing was also evaluated and showed similar efficacy to IV dosing and a manageable safety profile (Budde et al. ASH 2022). MorningSun (NCT05207670) is an ongoing, open-label, multicenter, Phase II study evaluating the efficacy, safety and pharmacokinetics of SC mosunetuzumab in patients with selected B-cell NHLs, including those without prior treatment (Flinn et al. ASCO 2022). For the first time, we present initial efficacy and safety data for patients with previously untreated, low-tumor burden FL who were enrolled into MorningSun at community and academic centers in the United States. Methods: Inclusion criteria include previously untreated Grade (Gr) 1 or 2 FL, low tumor burden by GELF criteria, Ann Arbor stage III or IV disease, and ECOG performance status 0-2. Mosunetuzumab is administered without mandatory hospitalization by SC injection in 21-day cycles. In Cycle (C) 1, mosunetuzumab is given on Day (D) 1 (5mg), D8 (45mg) and D15 (45mg). In C2+, mosunetuzumab is given on D1 only (45mg). Treatment is continued for up to 17 cycles (~1 year). Patients who are in complete metabolic response (CMR) at C8 can discontinue active treatment and enter long-term follow-up. Premedication with dexamethasone (20mg) is mandatory in C1 and C2 and optional thereafter. Acetaminophen and diphenhydramine may also be given. The primary endpoint is the rate of progression-free survival at 24 months. Results: At data cut-off (March 6, 2023), 30 patients had been enrolled (19 at community centers and 11 at academic centers). Median age was 56 years. All patients had ECOG performance status 0-1, and a minority had B symptoms ( Table 1). The median duration of follow-up was 4.2 months (range: 0.2-11.1) and the median number of cycles received was 7 (range: 1-13). Among 24 patients who had at least one post-baseline tumor assessment, 23 achieved an objective response (95.8%; 95% CI: 78.9-99.9), including 20 who achieved a CMR (83.3%; 95% CI: 62.6-95.3). All responses were observed at the first post-baseline tumor assessment in C4 (median time to response: 2.6 months; 95% CI: 2.6-2.8). At cut-off, responses were ongoing in all patients who achieved CMR and 2 of 3 patients who achieved partial metabolic response. The most common adverse event (AE) was injection site reaction (53.3% of patients), all of which were Gr 1 (43.3%) or Gr 2 (10.0%). Cytokine release syndrome (CRS) occurred in 43.3%. Most patients had Gr 1 CRS ( Table 2). No Gr ≥3 CRS events or discontinuations due to CRS were reported. CRS occurred in C1 only. All CRS events resolved. Other common (>20%) AEs were headache (50.0%), nausea (30.0%), fatigue (26.7%), pyrexia (23.3%), chills (23.3%) and alanine aminotransferase increased (23.3%). Gr ≥3 AEs in ≥5% of patients were neutrophil count decreased (2 patients; 6.7%). Serious AEs (SAEs) in ≥5% were CRS (2 patients; 6.7%). No SAEs of infection were reported. AEs potentially consistent with immune effector cell-associated neurotoxicity syndrome (ICANS) occurred in 2 patients (Gr 1 confusional state and Gr 1 memory impairment in one patient each); both events resolved. No mosunetuzumab-related AEs leading to discontinuation occurred and no Gr 5 (fatal) AEs were reported. Conclusions: Initial MorningSun data support that SC mosunetuzumab is active in patients with previously untreated, low-tumor burden FL. Safety data demonstrate a manageable safety profile that is consistent with that seen in patients with R/R B-cell NHLs and is supportive of outpatient administration.

Mitochondrial Reprogramming By Bcl-2 Inhibitor Venetoclax Enhances αCD19 CAR-T Cell Fitness and Anti-Tumor Efficacy

Introduction Patients undergoing anti-CD19 chimeric antigen receptor T cell (CAR T) therapy for the treatment of aggressive B-cell non-Hodgkin lymphomas (B-NHL) can expect a complete response rate of merely 43-68%. Insubstantial clinical responses to CAR T therapy and a small window of survival for patients facing relapsed/refractory (R/R) disease highlights the critical need for improvement of CAR T therapy in aggressive B-NHL cases. Recent findings suggest that enriching for a CD8 + T Stem Cell Memory (T SCM) population within adoptive cell therapies translates to enhanced antitumor function and persistence in vivo, and thus produces favorable clinical outcomes. T SCM cells display heightened self-renewal, low mitochondrial membrane potential (∆Ψm), and are metabolically dependent upon oxidative phosphorylation (OXPHOS) and fatty acid oxidation (FAO). Selection of CD8 + T SCM subsets is therefore necessary to ensure a robust antitumor response. Previously, we revealed synergy between the Bcl-2 inhibitor, Venetoclax, and CAR T cells in the context of B-NHL. Venetoclax exposure increased CD8 + T SCM cell frequency, decreased ∆Ψm, reduced T Regulatory (T REG) cell frequency, and bolstered effector cytokine production. Though the role of Bcl-2 inhibition within B-NHLs is well characterized (FDA-approved for frontline and relapsed/refractory chronic lymphocytic leukemia [CLL] treatment), further investigation regarding the impact of Venetoclax on the patient T cell compartment is needed. In this study, we demonstrate a role for Venetoclax as a qualitative enhancer of bulk CAR T cell product using both healthy-donor and B-NHL patient-derived T cells, and implicate modulation of mitochondrial pathways as a likely mechanism of action. Methods Whole peripheral blood mononuclear cells were collected from consenting healthy human donors or CLL patients undergoing ramp-up with Venetoclax (BDR 164122) at Roswell Park Comprehensive Cancer Center, and activated with αCD3/αCD28. CLL samples were collected at baseline, 1 day, 7 days, and 1-month timepoints. Transduction was performed with lentiviral vectors encoding CD28 or 4-1BB-based α-CD19 CAR constructs 24 hours post-activation. Transduced cells were cultured in RPMI with hIL-2 and hIL-7. In vivo studies were performed on 7-week-old SCID mice using Raji-Luc, Rec-1, and U2932. Venetoclax was given daily [100mg/kg] via oral gavage. CAR T cells were delivered by single infusion via tail-vein injection at a dose of either 1x10 6 or 3x10 6 cells total.Mice were imaged via IVIS spectrum. Western blot was performed on stimulated untransduced T cells or CAR T cells exposed to Venetoclax. Total protein was quantified, and membranes were probed for apoptotic and mitochondrial protein expression. Patient sample immunophenotyping was performed on CLL patient samples using panels interrogating memory differentiation, exhaustion, and T REG populations. Results Consistent with our hypothesis, a significant increase in mitochondrial proteins associated with FAO and OXPHOS - ACC, PDH, and AMPKα - was observed in exposed CAR T cells, as well increased expression of transcription factor FOXO1 - an essential driver of CD8 + memory differentiation. Voltage-dependent anion channel (VDAC) expression increased, suggesting heightened efflux of respiratory substrates into the tricarboxylic acid cycle. Hexokinase-I, a critical glycolytic enzyme, was significantly downregulated. We next investigated our combination therapy in vivo and observed extended survival outcome by 15 days. Further in vivo studies investigating preclinical models of mantle cell lymphoma and activated B-cell diffuse large B-cell lymphoma are currently ongoing. Within our CLL patient samples, 24-hour and 7-day timepoints reflected a favorable population increase in CD8 + T SCM cells as compared to baseline samples, and marginally restored CD8 + T cell frequencies. Conclusion Metabolic properties guide T cell memory differentiation and fitness. There exists potential to manipulate this mechanism and select the most metabolically fit T cell population for adoptive cell therapies. In addition to answering fundamental questions of CAR T cell biology, the combination of Venetoclax and CAR T cell therapy may provide a solution to the observed clinical gap, in which CAR T therapy as a single agent does not afford favorable response rates in the context of aggressive and R/R B-NHLs.

XPO1 Inhibitor Triggers Autophagy of TP53-Mutated Burkitt's Lymphoma Cells

INTRODUCTION Burkitt lymphoma, a highly aggressive B-cell non-Hodgkin's lymphoma (NHL), has distinct pathological and clinical characteristics. Intensive combination chemotherapy has been widely used in the treatment of Burkitt lymphoma and survival has improved for Burkitt lymphoma patients. TP53 mutation frequency is not age related, and TP53 mutations are significantly associated with higher incidence of relapse. XPO1 inhibitors, novel targeted drugs, reversibly inhibit the nuclear export of tumor-associated proteins and mRNAs by blocking nuclear export protein 1 (XPO1). Preclinical and clinical studies of XPO1 inhibitors have been successfully used in treatment of AML, MM and NHL. METHODS We used CCK8 assays to detect growth inhibition in 4 lymphoma cell lines (Raji, Jeko-1, Daudi, and CA46) under the treatment of different concentrations of KPT330 (XPO1 inhibitor). The 4D-Label-free quantitative proteomics technology was used to determine the changes of protein distribution in the nucleoplasm and cytoplasm of CA46 cells before and after KPT330 treatment (50nM, 48h). The proteomic results were validated by QPCR, westernblotting assay and fluorescent marker analysis. RESULTS The lymphoma cell lines we choosed, Raji, Jeko-1, Daudi, and CA46, all have MYC rearrangement and TP53 mutations. CCK8 assay showed Jeko-1, Daudi and CA46 were relatively sensitive to KPT330 (IC50=86.87nM, 48.34nM and 87.85nM, respectively). The results of the quantitative proteomics suggested proteins with increased nucleo-cytoplasmic ratio after KPT330 treatment were mainly focused on autophagy regulation and p53 signal pathways, including TFEB, USP10, DAP, NRBF2, SQSTM1, etc ( Fig1, a). To confirm the effect of KPT330-triggered autophagy, first, we assessed the apoptosis rates of CA46 after exposure to KPT330 for 24 hours, all of which were below 10%, ruling out apoptosis-induced autophagy ( Fig1, b). Then, westernblotting results showed that the cytoplasm load of autophagy flux related proteins, LC3B-1 and LC3B-II, both increased after KPT330 treatment ( Fig1, c). To confirm whether KPT330 effects the generation or degradation of LC3B-II, we used the late autophagy inhibitor chloroquine (CQ) to cells and observed the p62 protein level reduced and LC3B-II protein level increased with additional use of CQ compared with only KPT330 treatment ( Fig1, d). Furthermore, the ratio of LC3B-II-CQ-presence to LC3B-II-CQ-absence in the KPT330-treated group was higher than that in the control group ( Fig1, d), suggesting that KPT330 may enhance the generation of LC3B-II. We further utilized the pLenti-CMV-mCherry-GFP-LC3B plasmid to establish an LC3B stable-overexpressing CA46 cell line. Flow cytometry analysis showed that the median fluorescence intensity of both GFP and mCherry signals increased after KPT330 treatment( Fig1, e), further confirming the overall elevation of LC3B levels. To investigate the mechanism behind the increased autophagy flux, we utilized westernblotting analysis to validate the results from quantitative proteomics. TFEB protein level showed an increase in nuclear-cytoplasmic ratio upon KPT330 treatment ( Fig2, a). TFEB is known as a transcriptional activation protein, and its nuclear translocation is associated with the upregulated expression of genes involved in autophagosome formation, autolysosome fusion and lysosome biogenesis. QPCR analysis validated that genes associated with the aforementioned processes, such as LC3B, VPS11, CTSD, and CLCN7, exhibited increasing expression levels from 6 to 24 hours after KPT330 treatment ( Fig2, b). More interestingly, USP10, a component of a regulatory loop that controls autophagy and p53/TP53 levels, significantly increased in cell nucleus after drug action ( Fig2, c), which suggested that KPT330 may stabilize p53 function. Finally, we also attempted to discover the potential synergistic therapeutic effect of KPT330 with commonly used chemotherapy drugs in clinical lymphoma treatment. However, none synergistic effect was observed according to KPT330 combined with Cytarabine, Doxorubicin, Azacitidine, Decitabine, Rapamycin, Bortezomib, Duvelisib or Venetoclax, respectively ( Fig2, d). CONCLUSION XPO1 Inhibitor, KPT330 can restrict nuclear export of autophagy-related proteins thus regulate autophagy and stabilize p53 function in TP53-mutated Burkitt's lymphoma cells, which revealed a promising treatment.

Factors Associated with in-Hospital Mortality in Patients with Burkitt Lymphoma - Analysis of the National Inpatient Sample (NIS)

Introduction Burkitt Lymphoma (BL) is a highly aggressive B-cell non-Hodgkin Lymphoma, characterized by translocation and dysregulation of the MYC gene. BL comes in three distinct forms: endemic, sporadic, and immunodeficiency-related. BL has become a highly curable disease after the development of intense chemo-immunotherapy regimens, but given the low incidence of the disease, nationally representative studies on outcomes are scarce. We aim to highlight the most recent population-based factors associated with inpatient mortality and morbidity in patients with Burkitt's lymphoma. Methods The National Inpatient Sample database was queried from October 2015 to December 2020. Adults aged >= 18 with a primary diagnosis of BL were identified and stratified into those with and without the outcome of mortality. Complications included were queried using ICD codes most commonly found in the secondary diagnosis variables. Baseline characteristics, inpatient complications, and outcomes were compared using chi-squared and Wilcoxon rank-sum tests. Logistic regression analysis was performed to assess the risk of death before and after adjusting for multiple risk factors. Results We found 3,650 admissions with a primary diagnosis of BL, of which 320 (8.8%) patients died during the hospitalization. Patients who died had higher Charlson Comorbidity Index (CCI) >=3 (69%), tumor lysis syndrome (TLS) (41%), Sepsis (39%), and acute hypoxic respiratory failure (AHRF) (33%). No significant differences were found according to income, hospital bed-size or geographical region. In univariable analysis patients who died tended to be older with age >= 65 years (OR 2.55 (1.23 to 5.26) p=0.039), however, once adjusting for other variables there was no significant difference in mortality by age group, race, or gender. Multivariable analysis identified 3 factors with high association with mortality: acute renal failure (OR 3.07 (1.63 to 5.76), p<0.001), sepsis (OR 7.96 (3.85 to 16.4), p<0.001), and acute hypoxic respiratory failure (OR 2.70 (1.23 to 5.92), p=0.013). Conclusion BL remains a highly aggressive but potentially curable disease in patients able to tolerate intensive chemo-immunotherapy regimens. Our study highlights multiple factors that are independently associated with death include acute renal failure, sepsis, and acute hypoxic respiratory failure. Close monitoring should be placed on patients with the above complications to improve survival.

Real-Word Experience of CAR T-Cells in Patients with Relapsed/Refractory Follicular Lymphoma : A Descart Registry Analysis from the Lysa

Background: Anti-CD19 CAR T-cells have revolutionized the treatment of aggressive B-cell non-Hodgkin lymphomas (NHL) by demonstrating durable responses. Although follow-up remain short, axicabtagene ciloleucel (axi-cel, ZUMA-5) and tisagenlecleucel (tisa-cel, ELARA) demonstrated promising complete response rates (CRR) of 86% and 69%, respectively, in phase II trials including relapsed or refractory (R/R) follicular lymphoma (FL).. Tisa-cel and axi-cel are now approved after at least 2 lines of systemic therapy by the FDA. Few real-word evidence (RWE) data have been reported so far. Methods: Patients were eligible if aged >18 years, had FL histology (grade 1-3A) and had received at least 2 previous treatment lines for tisa-cel and 3 for axi-cel according to the French early access program label. From Dec 2021 to Jan 2023, a total of 112 patients were included in early access program by 21 French centers and in DESCAR-T registry (NCT04328298), among them 87 were infused with a CAR product. At the data cut-off on March 2023, 70 had at least 1 month of follow-up (FU) with PET-CT evaluation, and thus were considered for the safety and efficacy sets (62 tisa-cel and 8 axi-cel,). Results: Patient and disease characteristics were as follows: 46 males (65.7%), median age 62 years (range: 34-79), 3 median prior lines of therapy (range 2-9, including bi-specific antibody in 12.9% and autoSCT in 44.3%), FLIPI 0-2 in 40.5% and 3-5 in 49.5%, bulky disease (>5cm) in 22%, and LDH > N 52.2%. POD24 after 1 st systemic immunochemotherapy (IC) was reported in 44 patients (62.8%). Before CAR T-cell infusion, 58.6% of patients received a bridging therapy (20 (48.8%) chemotherapy, 25 (61%) monoclonal antibodies, 4 (9.8%) kinase inhibitors, 15 (36.6%) lenalidomide, and 3 (7.3%) radiotherapy). All patients but one received a fludarabine and cyclophosphamide-based lymphodepletion. Median time from order to infusion was 48 days (range 34-204), and 41 days (30-328) from leukapheresis to infusion. Median FU was 7.3 months [6.4; 8.2] from product order, and 5.4 months [3.4; 6] from CAR infusion. Best ORR and CRR were 97.5 and 87.5%, respectively with 72.5% of patients in CR at 1 month with projected DOR and DOCR of 72.4% [56.8; 83.1%], and 79.7% [62.7%; 89.6%], respectively. Projected 6-months PFS, OS, were 71.8% [95% CI, 56.6%; 82.4%], 97.4% [83.2%; 99.6%], respectively. Fifteen (21.4%) patients progressed (median time from infusion to first relapse: 3 months (range 1-10 mo). Only one patient died from lymphoma progression. Any-grade CRS and ICANS were reported for 74.3% and 27.1% of patients, but grade 3-4 CRS/ICANS were very unfrequent,1.4% and 4.3%, respectively. No grade 5 was reported. Persisting grade 3-4 hematologic toxicities at 1 and 3 months were: neutropenia (50% and 12.3%), thrombocytopenia (18.6% and 0%), and anemia (8.6% and 0%). Medically relevant bacterial and viral infections were reported in 20% and 14.3% of patients, respectively. Conclusions: Although longer FU is needed to assess disease control, RWE data from the DESCAR-T registry confirm the excellent response rates and safety profile of CAR T-cells in R/R FL after at least 2 lines of previous therapy.

A Phase 1 Study of Subcutaneous Rituximab Hyaluronidase Combined with Local Standard-of-Care Chemotherapy for the Treatment of Diffuse Large B-Cell Lymphoma in Uganda

Background: Patients with Diffuse Large B-Cell Lymphoma (DLBCL) who are treated in low-resource settings have inferior outcomes compared to those in high-resource settings, with 12-month overall survival (OS) rates typically under 50% with standard chemotherapy (Gopal et al. [PMID:26934054]). Rituximab, an anti-CD20 monoclonal antibody, when combined with chemotherapy, improves OS for DLBCL and is the standard of care for patients with CD20+ DLBCL in resource-rich settings. Despite receiving FDA approval over 25 years ago and its subsequent inclusion in the World Health Organization's List of Essential Medicines, IV rituximab is not readily available or routinely used in sub-Saharan Africa (SSA). This is in part due to cost and the limited availability of infusion centers in SSA. Subcutaneous rituximab hyaluronidase (sqR) is a potential solution, but its safety is not established in these settings. Here, we evaluated the safety and efficacy of a fixed dose of 1400 mg sqR in combination with standard-of-care chemotherapy in Ugandan adults with CD20+ DLBCL. Methods: This open-label phase I study, conducted at the Uganda Cancer Institute [(UCI) Kampala, Uganda], enrolled patients age ≥ 18 with newly diagnosed CD20+ DLBCL with measurable disease, and an ECOG performance status of < 2. Patients living with HIV were eligible if they were receiving antiretroviral therapy and had CD4+ T-cell count >200 cells/uL. The first cohort (n=6) received IV rituximab, 375mg/m 2 on day 1 of cycle 1 plus CHOP chemotherapy (IV cyclophosphamide 750 mg/m 2, IV doxorubicin 50 mg/m 2, IV vincristine 1·4 mg/m 2 (maximum 2 mg/m 2) on day 1, and oral prednisone 100 mg on days 1-5). This cohort received sqR (1400 mg) for subsequent cycles. The second cohort (n=12) received sqR plus CHOP for all cycles. Safety and tolerability were evaluated by monitoring adverse events and graded by Common Terminology Criteria for Adverse Events (CTCAE) version 5; secondary outcomes included response rates and treatment completion. We used Kaplan-Meier methods to estimate OS and progression-free survival (PFS), defined as disease progression or death, with 95% confidence intervals (95% CIs) at 12 months from initiation of treatment. Results: Between October 25 th, 2019 and October 7, 2022, 38 patients were assessed for eligibility; 19 patients met the criteria and were enrolled. One patient was not given the study treatment due to UCI-mandated COVID-19 restrictions. Of the 18 participants, median age was 36.5 years (IQR 25.2-50.0), 10 (56%) were male and 10 (56%) presented with advanced stage (III/IV) disease. Three (16.7%) participants were living with HIV and their median CD4 count was 311. Participants completed a median of 6 cycles of therapy. Grade 3 or 4 hematologic toxicities were reported in 11 (61%) participants; grade 3 or 4 non-hematologic toxicities were reported in 4 (22%). The most common hematologic toxicity was neutropenia (n=9, 50%); the most common non-hematologic toxicities were anorexia (n=7; 39%) and hyponatremia (n=7; 39%). Fifteen of the 18 participants completed treatment; 3 died during treatment. Of the 15 who completed treatment, 14 (93.3%) patients achieved a complete response (CR) and 1 patient (6.7%) had a partial response (PR). The patient who had a PR subsequently developed progressive disease, as did 2 patients who initially had a CR after therapy (Figure 1). The OS and PFS at 12 months was 83% (95% CI, 68-100%) and 66% (95% CI 47-92%) respectively. Conclusion: As demonstrated in other parts of the world, sqR together with CHOP chemotherapy was safe, well-tolerated, and efficacious among Ugandan patients with DLBCL. The very high CR, PFS, and OS rates are nearly double that of historical controls at the UCI, and comparable to outcomes expected in resource-rich settings. Colleagues have previously demonstrated the safety and efficacy of an oral chemotherapy regimen for aggressive non-Hodgkin's lymphoma in East Africa (Mwanda et al. [PMID: 19470940]). Given the results of this study, the potential for a safe and effective regimen, which precludes the need for IV therapy, for the treatment of DLBCL exists. Additionally, this trial strengthened the clinical trial research infrastructure in Uganda; trials conducted locally are critical to determining pragmatic and optimal treatment strategies. We anticipate that this research will improve care for DLBCL patients not only in Uganda but in other resource-limited settings as well.

Trial in Progress: Phase 1 Trial Evaluating the Safety and Tolerability of Odronextamab in Combination with Cemiplimab in Relapsed/Refractory Aggressive B-Cell Non-Hodgkin Lymphoma

Background and Significance CD20×CD3 bispecific antibodies (bsAbs) have emerged as an important immunotherapeutic approach for relapsed/refractory (R/R) B-cell non-Hodgkin lymphoma (B-NHL). These agents have shown compelling efficacy in indolent B-NHL, with the CD20×CD3 bsAb odronextamab demonstrating 75% complete response (CR) rates and encouraging durability of responses (median duration of response [DoR], 20.5 months) in heavily pretreated patients with R/R follicular lymphoma in the ELM-2 study (Kim TM, et al. ASH. 2022). In comparison, patients with R/R diffuse large B-cell lymphoma who were treated with odronextamab in the same study demonstrated a CR rate of 31%, and median DoR was 10.2 months (Kim WS, et al. ASH. 2022). These data indicate an unmet need for treatment strategies that can improve the depth and durability of responses in patients with aggressive R/R B-NHL. Combination therapy with drugs that complement the mechanism of action of CD20×CD3 bsAbs may provide an opportunity to address this unmet need. Cemiplimab is a human immunoglobulin G4 (IgG4) monoclonal antibody directed against programmed cell death protein 1 (PD-1), blocking PD-1- and programmed cell death ligand 1 (PD-L1)-driven inhibitory T-cell signaling. Combining odronextamab and cemiplimab provides an attractive therapeutic opportunity to improve anti-tumor efficacy while restricting T-cell exhaustion, and preliminary results for this combination provided proof of concept, with evidence of clinical activity and manageable tolerability in patients with B-lymphoid malignancies (n=12; Topp MS, et al. ASH. 2017). Here, we investigate whether the addition of cemiplimab to odronextamab can improve the depth and durability of response in patients with previously treated aggressive B-NHL. Study Design and Methods This is a Phase 1, open-label, multicenter, dose-escalation and -expansion study that will take place at up to 28 sites globally (NCT02651662). Intravenous (IV) odronextamab is administered in 21-day cycles, initially as monotherapy and with step-up dosing during Cycle (C) 1 to mitigate the risk of cytokine release syndrome (Figure). Full-dose odronextamab is administered on Day (D) 1, D8 and D15 of C2-C4, and then on D15 of each cycle during maintenance (C5 onwards) until disease progression or treatment discontinuation. IV cemiplimab starts on C2D15 at the specified dose level (DL1-DL7) and is administered on D15 of subsequent cycles. The dose-limiting toxicity (DLT) period is 28 days from the start of cemiplimab administration. At the end of dose escalation, a regimen will be selected for further investigation in dose expansion. Approximately 60 patients will be enrolled. Patients will be aged ≥18 years with aggressive CD20+ B-NHL that is R/R after ≥2 lines of systemic therapy containing an anti-CD20 antibody and an alkylating agent. Patients must have measurable disease, Eastern Cooperative Oncology Group performance score (ECOG PS) ≤1, and adequate bone marrow and organ function. Exclusion criteria include primary central nervous system lymphoma, prior allogeneic stem cell transplantation, autoimmune disease, prior adverse events from anti-PD-1/PD-L1 therapy, and uncontrolled infections. Primary endpoints are safety, tolerability, DLTs, and to determine the recommended Phase 2 dose of the combination regimen of odronextamab and cemiplimab. Secondary endpoints include pharmacokinetics, immunogenicity, and preliminary anti-tumor activity (overall response rate, CR rate, and DoR, according to the Lugano classification). There are no formal hypotheses for this study and analyses will be descriptive in nature.

Impact of “Cell-of-Origin” on Outcome after Axicabtagene-Ciloleucel CAR-T Cell Therapy in Relapsed/Refractory Aggressive Non-Hodgkin B-Cell Lymphomas

Introduction: Despite significant advances in characterizing aggressive B-cell non-Hodgkin lymphomas (aNHL) based on molecular and genetic signatures, the Hans algorithm, determined by immunohistochemistry, remains the most commonly used approach to differentiate aNHLs based on the cell-of-origin (COO) into germinal center B-cell (GCB) and non-GCB types. Although the Hans algorithm has demonstrated reliability as a predictor of better outcomes in newly diagnosed GCB cases, conflicting evidence has emerged, particularly in the relapsed/refractory (R/R) setting, and there are no definitive data about the impact of COO in patients (pts) receiving CAR-T cells. Thus, we conducted a retrospective analysis to evaluate the impact of COO extended to R/R aNHLs who underwent treatment with axicabtagene-ciloleucel (axi-cel). Methods: We retrospectively collected data from pts with R/R aNHLs treated with axi-cel at Fred Hutchinson Cancer Center between February 2018 and August 2022. The Hans algorithm, using CD10, BCL6 and MUM1 markers, was employed to stratify the pts into two cohorts: GCB and non-GCB. Paired biopsies of diagnosis and relapse before CAR-T eligibility were reviewed at our institution. Response to CAR-T was assessed at one month per Lugano PET/CT criteria. Univariate cox proportional hazard regression was used to estimate associations between baseline factors and outcomes. Statistically significant factors ( p<.05) in univariate analysis were further evaluated by multivariate analysis. Continuous variables were compared by Mann-Whitney U test and categorical variables by Fischer's exact test. Progression-free survival (PFS) was calculated from the date of CAR-T cell infusion to the date of death from any cause, disease progression or relapse. Overall survival (OS) was calculated from the date of CAR-T cell infusion to the date of death from any cause. Results: Sixty-four pts were included in this analysis, with an equal distribution of pts in the two cohorts: 32 with GCB subtype and 32 with non-GCB subtype. Baseline pts' characteristics and histology subtypes are listed in Table 1. Transformed follicular lymphoma and primary mediastinal B-cell lymphoma were not included in this analysis. The presence of bulky disease (defined as >5cm diameter) was higher in the GCB subtype (56% vs 22%, p<.001). Although primary refractory cases were more frequent in the GCB cohort (87.5 vs 66%), statistical significance was not reached (NR)( p=.07). The same proportion of pts received bridging therapy in the two groups (53% vs 53%, p=.99). Pts evaluable for response assessment were 62. Median time to best response was 29 days (range, 21-186) for GCB pts and 31 days (range, 22-182) for non-GCB pts ( p=.91). The overall response rate was 70% for GCBs (34.5% complete response [CR], 34.5% partial response [PR]) and 75% for non-GCBs (44% CR, 31% PR). After a median follow-up of 24.5 months (range, 9-121), comparable outcomes were observed for both the COO groups: despite the estimated median OS of 14 months (95% CI, 4.73-NR) for GCB vs 40 months (95% CI, 12.55-67.46) for non-GCB, we did not find any significant differences ( p=.73) (Figure 1A). Estimated median PFS was 10 months (95% confidence interval [CI], 3.5-16) for GCB and 11 months (95% CI, 0-24) for non-GCB ( p=.57) (Figure 1B). The 2-year OS and PFS were: GCB, 45% (95% CI, 30-69) and 44% (95% CI, 29-67) respectively; non-GCB, 55% (95% CI, 38-79) and 29% (95% CI, 16-55), respectively. Treatment-related toxicities did not differ between GCB and non-GCB, with cytokine-release syndrome rates of 97% vs 94% ( p=.99) and neurotoxicity rates of 53% vs 62.5% ( p=.61), respectively. The incidence of infectious complications was 25% vs 19% ( p=.76). On univariate analysis, the impact of COO on OS (HR=0.82, 95% CI 0.39-1.73, p=.60) and PFS (HR=1.18, 95% CI 0.61-2.28, p=.62) was undetermined. After adjusting for bulky disease, elevated LDH, absolute lymphocyte count≤1000/uL, performance status≥2, >2 lines of therapy and response status before axi-cel infusion, the impact of COO on OS (adjusted HR [aHR]=1.73, 95% CI 0.71-4.22, p=.23) and PFS (aHR=2.01, 95% CI 0.93-4.35, p=.08), remained undetermined. Conclusion: We could not confirm an association between COO, as determined by Hans algorithm, and outcomes in R/R aNHLs who received axi-cel at our center. Prospective analyses on larger datasets and other types of CAR-T products are needed.

Expansion, Persistence and Pharmacodynamic Profile of ADI-001, a First-in-Class Allogeneic CD20-Targeted CAR Gamma Delta T Cell Therapy, in Patients with Relapsed/Refractory Aggressive B-Cell Non-Hodgkin's Lymphoma

Introduction ADI-001 is a first-in-class allogeneic gamma delta (γδ) CAR T cell therapy targeting the B-cell antigen, CD20. Expansion and persistence of cell therapy products and release of functional cytokines have historically correlated with patient outcomes. Here we report cellular kinetics and pharmacodynamic correlates from a phase 1, multicenter, open-label, dose escalation study to evaluate ADI-001 in R/R B cell NHL. Methods Cellular kinetics of ADI-001 were measured using three orthogonal methods, including quantitative SNP profiling of cell product (AlloCell), flow-cytometry for CAR+ Vδ1 γδ T cells, and droplet digital PCR (ddPCR) quantification of CAR transgene copies. Using these methods, expansion of ADI-001 was assessed in the peripheral blood for 24 evaluable patients across four dose levels in association with a phase 1 dose-escalation trial (NCT04735471). Relationships between ADI-001 cellular kinetics and radiographic clinical responses were also examined. Serum biomarkers related to host immune cell recovery during lymphodepletion and cytokine release were monitored for pharmacodynamic purposes. Humoral immunogenicity was assessed by Luminex™-based anti-HLA antibody screening. Other correlative characteristics were also evaluated, including degree of HLA mismatching between patient and ADI-001 product in relation to response and/or ADI-001 expansion and persistence. Results As of the May 2023 cutoff for reporting, ADI-001 was detected at all dose levels using ddPCR and flow cytometry to quantify CAR transgene copies and CAR+ Vδ1 γδ T cells, respectively. Treatment at the highest dose level, achieved a notable mean C max of 201,666 copies/µg or 483 CAR+ cells/µl, and mean day 28 persistent exposure of 16,553 copies/µg or 21 CAR+ cells/µL. Additional measures of ADI-001 exposure (AlloCell) further demonstrated a robust dose-dependent expansion profile of ADI-001 in the peripheral blood. Subjects with a BOR of CR or PR were observed to have a mean peak of 180,107 CAR copies/µg versus a mean peak of 20,950 CAR copies/µg in subjects with a BOR of SD or PD. Additionally, as expected following lymphodepletion, a transient increase in the homeostatic cytokine IL-15, coinciding with ADI-001 expansion and host-mediated immune cell recovery, was observed. Using high-resolution HLA typing, the degree of HLA mismatch between allogeneic ADI-001 product and patients did not associate with degree of cellular expansion, day 28 persistence, or response. Finally, emergence of donor-specific anti-drug antibodies following treatment was not observed. Conclusions Using three orthogonal measures of exposure, we show for the first time robust dose-dependent expansion and persistence of an allogeneic CAR γδ T cell therapy, ADI-001, irrespective of extensive degree of HLA mismatch. ADI-001 C max and D28 persistent exposure were comparable to, or exceeded, those demonstrated by alternative allogeneic CAR T therapies and approved autologous CD19 CAR T therapies. Favorable clinical responses were also associated with higher mean and median peak of ADI-001 cells.

Trial-in-Progress: A Phase 1/2 Multi-Center Study of Onct-808, a ROR1-Specific CAR T, in Adult Patients with Relapsed/Refractory Aggressive B Cell Lymphoma

BACKGROUND: The therapeutic landscape for B-cell lymphomas (BCL) leverages several modalities, including chemotherapy, targeted agents, adoptive cell therapy, and immunotherapy. While these approaches have shown clinical benefit, challenges remain, such as variable response rates, treatment resistance, and toxicities. ROR1-specific CAR T-cell therapy represents a promising potential advancement in the treatment of aggressive BCL in patients (pts) who have failed previous CD19 CAR T therapy. ROR1-targeted CAR T therapy offers several advantages that could address these challenges. By directing CAR T cells to specifically recognize and eliminate ROR1-expressing tumor cells, it offers a highly targeted approach with the potential for enhanced efficacy. Moreover, ROR1 is often highly expressed on aggressive BCL, making it a suitable therapeutic target. This focused approach may help overcome the limitations associated with current treatments and improve response rates. After birth ROR1 is predominantly expressed on malignant cells; therefore, this selective expression pattern may reduce the risk of toxicities, including off-tumor target elimination and non-specific activation of immune cells. The development of ROR1-specific CAR T holds promise in enhancing the treatment outcomes for patients with aggressive BCL. With targeted action and potential to reduce toxicity, ROR1-specific CAR T cell therapy offers an innovative and novel strategy for improving patient outcomes in BCL. The ROR1 binding moiety for ONCT-808 is derived from zilovertamab. Zilovertamab vedotin showed preliminary evidence of efficacy and no evidence of on-target off-tumor toxicity in patients with advanced B cell malignancies (Wang 2022). METHODS: This is a Phase 1/2, single-arm, open-label, multi-center trial to evaluate the safety and efficacy of ONCT-808 in pts with aggressive BCL, including large B-cell lymphoma (LBCL) and mantle cell lymphoma (MCL) with no approved therapy available. The study comprises two distinct phases: Phase 1 for dose escalation followed by Phase 2 for dose expansion. In Phase 1, a standard 3+3 dose escalation design will be used with 3 provisional dose levels and expected sample size of up to 18 patients. Subjects will receive a conditioning regimen of intravenous (IV) cyclophosphamide and fludarabine, followed by ONCT-808 IV infusion. The dose of ONCT-808 will be sequentially escalated to determine the recommended Phase 2 dose (RP2D) for LBCL and MCL cohorts. Bridging therapy is permitted if clinically indicated. Phase 2 will enroll LBCL and MCL patients into separate dose expansion cohorts at the RP2D determined in Phase 1. ONCT-808 will be administered as a single IV infusion of autologous CAR T cells. The primary outcomes include assessing the incidence, severity, and relationship of Dose Limiting Toxicities (DLT), Treatment Emergent Adverse Events (TEAE), and RP2D selection of ONCT-808. Secondary outcomes include evaluating the Overall Response Rate (ORR), Complete Response (CR) rate, Duration of Response (DOR), pharmacokinetics of ONCT-808, expansion, persistence of ROR1 CAR-positive T cells, overall safety and tolerability. Key inclusion criteria include histologically confirmed aggressive B-cell NHL, measurable lesions per Lugano criteria (Cheson, 2014), and relapsed/refractory (R/R) disease without available therapy (must be previously treated with systemic therapy, progressed after or ineligible for autologous HSCT, and received prior approved anti-CD19 CAR-T at least 6 months prior to enrollment unless refused/ineligible). Key exclusion criteria include previous ROR1-targeted therapy, ongoing immunosuppression, and CNS involvement. The trial is currently active and patients have been treated in the phase 1 dose escalation. Clinical Trial Identifier: NCT05588440

A CD19/CD20-Directed Bispecific CAR-T Cell Therapy in Relapsed or Refractory Aggressive B-Cell Non-Hodgkin Lymphoma (NHL)

Three CD19-directed chimeric antigen receptor (CAR) T cell products are approved in the US for patients with relapsed or refractory (R/R) aggressive B cell lymphoma. Most patients treated in the third or later line will experience disease progression. Resistance to CD19-directed CAR T cell therapy may be due to tumor, T cell, or microenvironmental factors. Tumor-intrinsic causes of relapse include CD19 antigen loss, which is observed in approximately one third of relapsed cases(Spiegel, Dahiya, et al., 2021). Lower pretreatment CD19 antigen density has also been associated with treatment failure(Spiegel, Patel, et al., 2021). Conversely, enrichment of memory CD8+ T cells within the infusion product has been associated with achievement of complete remission (CR)(Deng et al., 2020). IMPT-314 is an autologous, naïve/memory enriched T cell product transduced by a lentivirus to express a tandem, OR-gate CD19/CD20-directed CAR with a 4-1BB co-stimulatory domain(Zah et al., 2016). IMPT-314 incorporates the same CAR and a highly similar manufacturing process as CART19/20, which was evaluated in a single-institution, Phase 1 study (NCT04007029)(Larson et al., 2023; Puliafito et al., 2023). Eleven patients with R/R B cell NHL have been treated at two dose levels (50 x 10 6 ± 30%, n=8; 200 x 10 6 ± 30%, n=3). Ten patients were evaluable for safety, and eleven patients were evaluable for response. Cytokine release syndrome (CRS) occurred in 60% of patients, however there were no CRS events above Grade 1. No patients experienced immune effector cell associated neurotoxicity syndrome (ICANS) of any grade. The overall response rate (ORR) and CR rate were 91% and 73%, respectively. As of March 6, 2023, with a median follow-up of 20.7 months, median progression-free survival (PFS) was 18.2 months (2.6-not estimable), and the median overall survival (OS) has not been reached (5.7-not estimable). This study of CART19/20 demonstrates the potential of simultaneous targeting of CD19 and CD20 with a bispecific CAR. Study Design The MPCT-012L study (NCT05826535) is a Phase 1/2 multicenter, open label study to evaluate the safety and efficacy of IMPT-314 in participants with relapsed/refractory aggressive B-cell NHL, defined as diffuse large B cell lymphoma (DLBCL), transformed follicular lymphoma (tFL), primary mediastinal large B cell lymphoma (PMBCL), and high-grade B-cell lymphoma (HGBL), after 2 or more lines of systemic therapy. Participants who have progressed after prior CD19 CAR-T cell therapy are eligible and are included in a separate cohort. All participants must have received prior anti-CD20 monoclonal antibody and an anthracycline. Patients with tFL must have received at least one line of therapy after transformation to DLBCL. Subjects with active (symptomatic or untreated) CNS involvement are excluded. There is no requirement for demonstration of CD19 or CD20 tumor expression at baseline. Phase 1 will be conducted using a modified 3+3 design with a starting dose of 100 x 10 6 CAR-T cells. Dose limiting toxicities (DLTs) will be evaluated for 28 days post-dosing. If a dose level is determined to be safe based on the DLT incidence, a total of up to 15 total subjects may be backfilled at that dose, per cohort. The primary objectives for Phase 1 are to evaluate the safety of IMPT-314 and to determine the recommended Phase 2 dose. The primary objective of Phase 2 is to estimate the efficacy of IMPT-314 as measured by CR rate. Secondary endpoints include evaluation of pharmacokinetics and time to event outcomes.

Results of the Astral Study: A Prospective Phase II Clinical Study of the German Lymphoma Alliance to Assess the Efficacy and Toxicity of High-Dose Chemotherapy Followed By Allogeneic Stem Cell Transplantation As Treatment of Primary Progressive and Relapsed Aggressive Non-Hodgkin Lymphoma

Introduction Patients with relapsed and refractory (r/r) aggressive B- or T-cell lymphoma have a dismal prognosis. Registry analyses as well as one preceding prospective clinical study demonstrated durable disease control through graft-versus-lymphoma activity conferred by allogeneic stem cell transplantation (alloSCT) in this population and suggests this modality being a preferred treatment option after failing of less toxic approaches. Recently, allogeneic SCT has been shown to result in long term PFS in r/r large B-cell lymphoma (LBCL) with preceding CAR-T treatment. Despite the current achievements in treatment of r/r LBCL by introducing anti CD19 CAR-T as a treatment modality appproximately 60% need further approaches to achieve long term progression free survival (PFS). Here we represent the results of a prospective clinical trial of allogeneic SCT in r/r aggressive B- and T-cell lymphoma using a thiotepa based conditioning regimen. Methods ASTRAL is a national, multicenter, prospective phase II study of the German Lymphoma Alliance (GLA). Eligible were patients aged 18 or older with relapsed or primary progressive aggressive B- or T-cell lymphoma who had a fully matched (10/10) related or unrelated donor available. Patients underwent myeloablative conditioning regimen with Fludarabine 125 mg/m², Thiotepa 15 mg/kg and Cyclophosphamide 120 mg/kg (FTC). GVHD prophylaxis and posttransplant immunosuppression was done by local standards. The primary objective was to determine the efficacy and toxicity of the defined conditioning therapy followed by allogeneic stem cell transplantation. The primary endpoint was progression free survival at 1 year. Key secondary efficacy endpoints included rate of complete remissions (CR), rate of partial remissions (PR), overall response rate (OR), rate of progressive disease (PD), relapse rate, event free survival (EFS) and overall survival (OS) at 1 year and non-relapse mortality (NRM). Results From June 2019 until December 2021 in total 60 patients were enrolled. The demographic data are presented in table 1. The majority of patients (70%) had B-cell lymphomas. Patients had received a median of 3 prior lines (range 1-6), including autoSCT in 42% of all patients and CAR-T in 29% of the B-cell patients. The rate of primary progressive disease before salvage therapy was 27% (29% B-cell and 22% T-cell). 83% (50/60) still had measurable disease (no CR) prior to FTC, thereof 81% with B-cell (34/42) and 89% with T-cell lymphoma (16/18) and 13 (22%) had high intermediate and high IPI. After alloSCT, CR was achieved in 45% of patients with B-cell and in 67% with T-cell lymphoma, respectively. The overall response rate was 53%, 48% for B-cell lymphoma and 67% T-cell lymphoma. After a median observation time of 30 months, 1-year PFS was 40% [95%-CI: 28%-52%] (38% [23%-53%] (B-cell) vs. 44% [21%-67%] (T-cell) (figure 1). EFS was identical to PFS. After one year 15 patients (25%) had progressive disease or relapse, thereof 13 patients (29%) with B-cell lymphoma and two with T-cell lymphoma (11%). Thirty-seven patients died. Main cause of death was treatment related 19/37 (51%), including 8/19 deaths due to toxicity of the conditioning therapy (6 infections, 1 respiratory failure and 1 neurotoxicity). 12/37 (32%) patients died lymphoma related, 4/37 (11%) patients died of a SARS-Cov-2 infection, and two causes of death were unknown. 1-year OS was 43% [31%-56%] (40% [26%-55%] (B-cell) vs. 50% [27%-73%] (T-cell)). 1-year NRM was 35% [23%-47%] (31% [17%-45%] (B-cell) vs. 44% [20%-68%] (T-cell)). Most of the patients with grade 3-5 adverse events had infections 40/60 (67%), renal disorders 17/60 (28%) and oral mucositis 14/60 (23%). Conclusions This is the second prospective trial on alloSCT for r/r aggressive lymphoma ever performed, demonstrating that alloSCT can provide sustained disease control in heavily pretreated patients. Also in the modern era, including patients who failed CARTs the strong anti-lymphoma activity of allogeneic SCT is shown in the low relapse/progression rate and the long term PFS. FTC conditioning avoids the toxicities of busulfan-based regimens but fosters neuro- and other toxicities, especially when given at 15mg/kg. Still, NRM remains a challenge and warrants exploration of novel transplant strategies, such as modulation of conditioning intensity and GVHD prophylaxis, e. g. post-transplant cyclophosphamide.

Non-O Blood Group and Pretreatment Risk of Venous Thromboembolism in Patients with Diffuse Large B-Cell Lymphoma

Introduction: Diffuse large B-cell lymphoma (DLBCL) is the most frequent aggressive non-Hodgkin lymphoma (NHL). It has been extensively documented that the risk of venous thromboembolism (VTE) is augmented in patients with aggressive lymphomas. The timing of VTE occurrence in this group of patients is highly variable and depends on diverse risk factors. Lately, it has been observed that ABO blood group type can have a prognostic impact on both thrombosis development and survival outcomes in patients with cancer. Therefore, the study aimed to examine the ABO blood group as a risk factor for VTE development in patients with DLBCL, to examine the relation between ABO blood groups and the timing of VTE development, and to investigate the association between ABO group type and treatment and survival outcomes. Methods: A total of 340 patients with DLBCL (newly diagnosed and relapsed) have been included in the study and all of them have been treated at the Lymphoma Center, Clinic for Hematology, University Clinical Center of Serbia. Data regarding VTE events were collected for all the patients included in the study, from the time of diagnosis to 3 months after the last cycle of therapy. ABO blood group has been determined before the treatment initialization. VTE was diagnosed objectively based on clinical examination, laboratory evaluation, and radiographic studies. Results: The median patients' age was 60 years (range, 19-89 years) and 52.9% were males. Newly diagnosed patients were dominant in the study population (94.4%). The majority of patients had advanced stage (stage III Ann Arbor 22.3% and stage IV 30.7%, respectively) of the disease. The median international prognostic index (IPI) was 2 (range, 0-5). Complete remission (CR) and partial remission (PR) were most commonly recorded (51.7% and 28.8%, respectively). The rate of VTE was 12.6% (43 patients), with the highest frequency of lower and upper extremity deep vein thrombosis (12 patients each). VTE was diagnosed before the initiation of specific hematological treatment in 54.5% of DLBCL patients, whilst 45.5% of DLBCL patients have been diagnosed with VTE following the hematological treatment initiation. The following distribution of ABO blood groups among DLBCL patients was detected: A blood group 39.4%, B blood group 17.6%, AB blood group 9.1%, and O blood group 33.8%. No significant association between VTE development and ABO group type was found (p=0.706) nor between VTE development and time of VTE development (p=0.895). The rate of VTE in DLBCL patients with A blood type was 4.7%, in B 2.1%, in AB 1.8%, and in O 4.1%, respectively (p=0.706). DLBCL patients with the non-O blood group developed VTE significantly more frequently prior to the hematological treatment commencement than after the treatment initiation (6.2% vs. 2.6%, p=0.003). Univariate logistic regression analysis showed that non-O blood type in DLBCL patients is a prognostic factor for VTE development prior to hematological treatment initiation (odds ratio (OR)=8.56, 95% confidence interval (CI): 1.92-38.21, p=0.005). Multivariate logistic regression model demonstrated that non-O blood type is an independent predictor of pretreatment VTE development in DLBCL patients (OR=18.33, 95% CI: 1.51-222.87, p=0.022). Therapy response of DLBCL patients was significantly associated with the ABO blood group (p=0.012) in the way that A and AB blood types were associated with unsatisfactory treatment response (no statistically significant association was found between ABO blood type and DLBCL patients' Ann Arbor stage, mediastinal involvement, bulky disease, extranodal disease, IPI, central nervous system involvement). Univariate regression analysis showed that A blood type is a predictor of unsatisfactory therapy response in patients with DLBCL (OR=1.85, 95% CI: 1.04-3.29, p=0.035). However, in multivariate regression model, this finding lost statistical significance. No significant association was observed regarding ABO blood type, progression-free survival, and overall survival in patients with DLBCL (p=0.398 and p=0.593, respectively). Conclusion: In DLBCL patients, non-O blood group is an independent predictor of VTE occurrence prior to initiation of a specific hematologic treatment. DLBCL patients with A blood group could be at risk of insufficient treatment response.

A hybrid few-shot multiple-instance learning model predicting the aggressiveness of lymphoma in PET/CT images

Background and objectivePatients with aggressive non-Hodgkin lymphoma (NHL) undergo distinct therapy strategies compared with indolent NHL patients. However, it is challenging to estimate NHL aggressiveness based on visual inspection of positron emission tomography (PET) or computed tomography (CT) images. Since diffuse large B-cell lymphoma (DLBCL) and Follicular lymphoma (FL) are the most typical and dominant aggressive and indolent NHL, respectively, this study aims to develop an artificial-intelligence-enabled model to distinguish DLBCL from FL in PET/CT images as the first step to tackle this challenge. MethodsWe propose a hybrid few-shot multiple-instance learning model to predict the aggressiveness of the NHL. First, rotation-based self-supervision learning (SSL) has been employed to train the encoder on a large-scale, publicly available CT image dataset. Second, hybrid instance-level features are obtained for each NHL lesion by combining deep features with the radiomics features from both PET and CT modalities. Third, instance-level features are transformed into bag-level (or patient-level) representations. Finally, bag-level representations are fed into a distance-based classifier through few-shot learning to predict NHL aggressiveness. ResultsOur model achieves an accuracy of 0.751 ± 0.008, a sensitivity of 0.787 ± 0.012, a specificity of 0.715 ± 0.013, an F1-score of 0.753 ± 0.009, and an area under the curve (AUC) of 0.795 ± 0.009 at the bag level. It outperforms the typical counterparts that use the radiomic features, random forest for feature selection, and support vector machines (SVMs) as classifiers. The three counterparts yield accuracies of 0.714 ± 0.023, 0.705 ± 0.008, and 0.698 ± 0.008, respectively. Moreover, settings of the SSL training dataset (Deep lesion) and task (rotation), hybrid CT and radiomic PET features, the pool-layer strategy of maximum, and distance-based classifier generate the best model. ConclusionsA hybrid few-shot multiple-instance learning model can predict lymphoma aggressiveness in PET/CT images and could be a potential tool for determining therapy strategies. Hybrid features and the combination of SSL, few-shot learning, and weakly supervised learning are the two powerful pillars of the model, and these can be expanded to other medical applications with limited samples and incomplete annotations.

Effect of T-regulatory cells and interleukin-35, interleukin-10, and transforming growth factor-beta on diffuse large B-cell lymphoma.

Diffuse large B-cell lymphoma (DLBCL) is an aggressive non-Hodgkin lymphoma that affects B lymphocytes. It can develop in the lymph nodes and can be localized or generalized. Despite DLBCL being considered potentially curable, little research has been conducted on the relationship between the body's immune response and DLBCL. To study the expression and significance of T-regulatory cells (Tregs) interleukin (IL)-35, IL-10, and transforming growth factor-beta (TGF-β) in DLBCL. Data from 82 patients with DLBCL who were initially admitted to The First Affiliated Hospital of Ningbo University (Zhejiang Province, China) between January 2017 and June 2022 and treated with standard first-line regimens were reviewed. Three patients were lost to follow-up; thus, 79 patients were included in the statistical analysis and then divided into three groups according to the evaluation of clinical efficacy: Incipient (new-onset and treatment-naïve), effectively treated, and relapsed-refractory. Thirty healthy individuals were included in the control group. The expression of peripheral blood T lymphocytes and their associated factors IL-35, IL-10, and TGF-β in the four groups were observed. In contrast to the successfully treated and normal control groups, both the incipient and relapse-refractory groups exhibited greater proportions of CD4-positive (+) Tregs (P < 0.05), whereas the proportion of CD8+ Tregs did not differ substantially between the groups. Serum levels of IL-35 and IL-10 in the incipient and relapsed-refractory groups were higher than those in the effectively treated and normal control groups (P < 0.05). There was no statistically significant distinction in the expression level of TGF-β between the groups (P > 0.05). The correlation between IL-35 and IL-10 concentrations was significantly positive, with a correlation coefficient of 0.531 (P < 0.05). The correlation between IL-35 and TGF-β concentration was significantly positive, with a correlation coefficient of 0.375 (P < 0.05). The correlation between IL-10 and TGF-β concentration was significantly positive, with a correlation coefficient of 0.185 (P < 0.05). The expression concentrations of IL-35, IL-10 and TGF-β were apparently and positively correlated (P < 0.05). Tregs IL-35, and IL-10 may be closely associated with the occurrence and development of DLBCL and the detection of related indices may be helpful in the analysis of disease prognosis.

The influence of immunodeficiency, disease features, and patient characteristics on survival in plasmablastic lymphoma

AbstractPlasmablastic lymphoma (PBL) is a rare and aggressive non-Hodgkin lymphoma associated with immunodeficiency, characterized by uncertain treatment approaches and an unfavorable prognosis. We conducted a multicenter, international, retrospective cohort study, aiming to characterize the clinical features, risk factors, and outcomes of patients with PBL. Data were collected from 22 institutions across 4 countries regarding patients diagnosed with PBL between 1 January 1999 and 31 December 2020. Survival risk factors were analyzed using both univariate and multivariate regression models. Overall survival (OS) was calculated using Kaplan-Meier statistics. First-line treatment regimens were stratified into standard- and higher-intensity regimens, and based on whether they incorporated a proteasome inhibitor (PI). A total of 281 patients (median age, 55 years) were included. Immunodeficiency of any kind was identified in 144 patients (51%), and 99 patients (35%) had HIV-positive results. The 5-year OS for the entire cohort was 36% (95% confidence interval, 30%-42%). In multivariate analysis, inferior OS was associated with Epstein-Barr virus–negative lymphoma, poor performance status, advanced stage, and bone marrow involvement. In an independent univariate analysis, the international prognostic index was associated with OS outcomes. Neither immunosuppression nor HIV infection, specifically, influenced OS. Among patients treated with curative intent (n = 234), the overall response rate was 72%. Neither the intensity of the treatment regimen nor the inclusion of PIs in first-line therapy was associated with OS. In this large retrospective study of patients with PBL, we identified novel risk factors for survival. PBL remains a challenging disease with poor long-term outcomes.