CD19-targeted CAR-T Research Articles

Hypophosphatemia correction reduces ICANS incidence and duration in CAR T-cell therapy: a pooled clinical trial analysis.

A common complication of chimeric antigen receptor (CAR)-T cell therapy is immune effector cell associated neurotoxicity syndrome (ICANS), which presents with encephalopathy, aphasia, inattention, somnolence, seizures, weakness, or cerebral edema. Despite its significant morbidity, there are currently no effective targeted treatments. Given the clinical similarities between ICANS and the neurological manifestations of acute hypophosphatemia, we retrospectively reviewed 499 patients treated with CD19-targeted CAR-T cell therapy across multiple clinical trials between 2015 and 2020. In addition to clinical toxicities experienced by the patients, we also interrogated the impact of serum electrolyte data and repletion of corresponding electrolyte deficiencies with ICANS incidence, severity, and duration. Hypophosphatemia was a common occurrence in CAR T-cell recipients, and the only electrolyte derangement associated with a significantly higher cumulative incidence of ICANS. Moreover, phosphorus repletion in hypophosphatemic patients was associated with significantly decreased ICANS incidence and duration. Hypophosphatemia was uniquely associated with encephalopathy neurological adverse events, which also showed the strongest positive correlation with both ICANS and CRS severity. These findings suggest that serum phosphorus could be a reliable biomarker for ICANS, and expeditious, goal-directed phosphorus repletion in response to serum hypophosphatemia could be a safe, inexpensive, and widely available intervention for such patients.

807MO High efficacy and safety of interleukin-6-knockdown CD19-targeted CAR-T cells in relapsed/refractory B-ALL patients

807MO High efficacy and safety of interleukin-6-knockdown CD19-targeted CAR-T cells in relapsed/refractory B-ALL patients

The Kinetics of Inflammation-Related Proteins and Cytokines in Children Undergoing CAR-T Cell Therapy-Are They Biomarkers of Therapy-Related Toxicities?

CD19-targeted CAR-T cell therapy has revolutionized the treatment of relapsed/refractory (r/r) pre-B acute lymphoblastic leukemia (ALL). However, it can be associated with acute toxicities related to immune activation, particularly cytokine release syndrome (CRS) and immune effector cell-associated neurotoxicity syndrome (ICANS). Cytokines released from activated immune cells play a key role in their pathophysiology. This study was a prospective analysis of proinflammatory proteins and cytokines in children treated with tisagenlecleucel. Serial measurements of C-reactive protein, fibrinogen, ferritin, IL-6, IL-8, IL-10, IFNγ, and TNFα were taken before treatment and on consecutive days after infusion. The incidence of CRS was 77.8%, and the incidence of ICANS was 11.1%. No CRS of grade ≥ 3 was observed. All complications occurred within 14 days following infusion. Higher biomarker concentrations were found in children with CRS grade ≥ 2. Their levels were correlated with disease burden and CAR-T cell dose. While cytokine release syndrome was common, most cases were mild, primarily due to low disease burden before lymphodepleting chemotherapy (LDC). ICANS occurred less frequently but exhibited various clinical courses. None of the toxicities were fatal. All of the analyzed biomarkers rose within 14 days after CAR-T infusion, with most reaching their maximum around the third day following the procedure.

Allogeneic CD19-targeted CAR-T therapy in patients with severe myositis and systemic sclerosis

Allogeneic chimeric antigen receptor (CAR)-T cells hold great promise for expanding the accessibility of CAR-T therapy, whereas the risks of allograft rejection have hampered its application. Here, we genetically engineered healthy-donor-derived, CD19-targeting CAR-T cells using CRISPR-Cas9 to address the issue of immune rejection and treated one patient with refractory immune-mediated necrotizing myopathy and two patients with diffuse cutaneous systemic sclerosis with these cells. This study was registered at ClinicalTrials.gov (NCT05859997). The infused cells persisted for over 3months, achieving complete B cell depletion within 2weeks of treatment. During the 6-month follow-up, we observed deep remission without cytokine release syndrome or other serious adverse events in all three patients, primarily shown by the significant improvement in the clinical response index scores for the two diseases, respectively, and supported by the observations of reversal of inflammation and fibrosis. Our results demonstrate the high safety and promising immune modulatory effect of the off-the-shelf CAR-T cells in treating severe refractory autoimmune diseases.

The underlying mechanism of chimeric antigen receptor (CAR)-T cell therapy triggering secondary T-cell cancers: Mystery of the Sphinx?

The U.S. Food and Drug Administration (FDA) has reported cases of T-cell malignancies, including CAR-positive lymphomas, in patients receiving B cell maturation antigen (BCMA)- or CD19-targeted autologous CAR-T cell immunotherapy. These reports were derived from clinical trials and/or post-marketing adverse event data. This finding has attracted widespread attention. Therefore, it is essential to explore the potential mechanisms by which chimeric antigen receptor (CAR)-T cell therapy triggers secondary T-cell cancers to further guarantee the safety of CAR-T cell therapy.

Cardiotoxic profiles of CAR-T therapy and bispecific T-cell engagers in hematological cancers

BackgroundChimeric antigen receptor (CAR) T-cell therapy and bispecific T-cell engagers, which redirect T-cells to tumor antigens, have immensely benefitted patients with relapsed/refractory B-cell cancers. How these therapies differ in cardiotoxicity is underexplored. We used the World Health Organization pharmacovigilance database, VigiBase, to compare cardiotoxicity profiles between CD19-targeted CAR-T therapy and blinatumomab (a CD19/CD3-targeted bispecific T-cell engager).MethodsSafety reports in VigiBase were filtered for diffuse large B-cell lymphoma (DLBCL, n = 17,479) and acute lymphocytic leukemia (ALL, n = 28,803) for all adverse reactions. Data were further filtered for patients taking CAR-T therapy or blinatumomab. Reporting odds ratios (ROR) and fatality rates were compared between CAR-T cell products (e.g. tisagenlecleucel and axicabtagene ciloleucel), and between CAR-T therapy and blinatumomab.ResultsTisagenlecleucel is associated with cardiac failure (IC025 = 0.366) with fatality rates of 85.7% and 80.0% in DLBCL and pediatric ALL patients respectively. For DLBCL patients, axicabtagene ciloleucel has greater reporting for hypotension than tisagenlecleucel (ROR: 2.54; 95% CI: 1.28–5.03; p = 0.012), but tisagenlecleucel has higher fatality rates for hypotension than axicabtagene ciloleucel [50.0% (tisagenlecleucel) vs 5.6% (axicabtagene ciloleucel); p < 0.001]. Blinatumomab and tisagenlecleucel have similar fatality rates for hypotension in pediatric ALL patients [34.7% (tisagenlecleucel) vs 20.0% (blinatumomab); p = 0.66].ConclusionsTisagenlecleucel is associated with severe and fatal adverse cardiac events, with higher fatality rates for hypotension compared to axicabtagene ciloleucel in DLBCL patients, but similar hypotension fatality rates compared to blinatumomab in pediatric ALL patients. Effective management necessitates experienced physicians, including cardio-oncologists, skilled in interdisciplinary approaches to manage these toxicities.

Consolidation with First and Second Allogeneic Transplants in Adults with Relapsed/Refractory B-ALL Following Response to CD19CAR T Cell Therapy

CD19-targeted chimeric antigen receptor T cell (CAR-T) therapy has led to unprecedented rates of complete remission (CR) in children and adults with relapsed/refractory (r/r) B-cell acute lymphoblastic leukemia (B-ALL), yet the majority of adults relapse after initial response. One proposed method to extend the durability of remission in adults following response to CAR-T therapy is consolidation with allogeneic hematopoietic cell transplantation (alloHCT). Considering the limited published data for the utility of post CAR-T therapy consolidative alloHCT in r/r B-ALL, especially data related to patients receiving a second alloHCT, we sought to describe outcomes of patients with r/r B-ALL at our institution who received their first or second alloHCT following response to CAR-T therapy. We performed a retrospective analysis of adult patients with r/r B-ALL who responded to either investigational or standard of care (SOC) CD19-targeted CAR-T therapy and underwent consolidation with alloHCT while in CR without interim therapy. We identified 45 patients, of whom 26 (58%) and 19 (42%) received their first and second alloHCT as consolidation post CAR-T therapy, respectively. The median age was 31 years (range: 19-67) and 31 (69%) patients were Hispanic. Ph-like was the most common genetic subtype and comprised over half of cases (53%; n = 24). The median number of prior therapies pre-transplant was 5 (range: 2-7), and disease status at the time of alloHCT was CR1, CR2 or ≥CR3 in 7 (16%), 22 (49%) and 16 (35%) patients, respectively. The median time from CAR-T therapy until alloHCT was 93 (range: 42-262) days. The conditioning regimen was radiation-based myeloablative (MAC) in 22 (49%) patients. With a median follow-up of 2.47 years (range: 0.13-6.93), 2-year overall survival (OS), relapse free survival (RFS), cumulative incidence of relapse (CIR) and non-relapse mortality (NRM) were 57.3% (95% CI: 0.432-0.760), 56.2% (95% CI: 0.562-0.745), 23.3% (95% CI: 0.13-0.42), and 20.4% (95% CI: 0.109-0.384), respectively. Two-year OS (52% vs. 68%, P = .641), RFS (54% vs. 59%, P = .820), CIR (33.5% vs. 8.5%, P = .104), and NRM (12.5% vs. 32.2%, P = .120) were not significantly different between patients who underwent their first vs. second transplant, respectively. In univariate analysis, only Ph-like genotype was associated with inferior RFS (P = .03). AlloHCT post CAR-T response is associated with a relatively low early mortality rate and encouraging survival results in high-risk adults with r/r B-ALL, extending to the second alloHCT for fit and eligible patients.

Preclinical assessment of IPH6501, a first-in-class IL2v-armed tetraspecific NK cell engager directed against CD20 for R/R B-NHL, in comparison with a CD20-targeting T cell engager.

7030 Background: The therapeutic landscape for relapsed or refractory (R/R) B-cell non-Hodgkin lymphoma (B-NHL) is evolving to include targeted T-cell based immunotherapies, including CD19-targeted CAR-T and CD3xCD20 T-cell engaging (TCE) bispecific antibodies. Yet, there remains an unmet medical need for patients who are refractory to, or ineligible for these treatments. Leveraging natural killer (NK) cells emerges as a promising strategy in hematological malignancies, as shown in a Phase 1 study with IPH6101/SAR’579 in R/R AML (Stein, ASCO 2023; Bajel, ASH 2023). Methods: We developed IPH6501, a tetraspecific antibody-based NK cell engager that simultaneously targets on NK cells the CD16a and NKp46 receptors, the IL-2 receptor with an engineered IL-2 variant (IL2v) and on B-NHL cells the CD20 antigen. This approach boosts NK cell activation and proliferation, cytotoxicity against tumor cells, and cytokine production. The IL-2 variant is designed with mutations that prevent binding to CD25 (IL-2Rα), limiting Treg activation and potential IL-2-related side effects. IPH6501 has been evaluated in mouse, non-human primate, and human-derived models, including cells from R/R B-NHL patients. Results: In vivo studies in non-human primates and tumor mouse models revealed that IPH6501, at well-tolerated doses, significantly boosted peripheral NK cell proliferation, drove their accumulation at tumor locations, and effectively eradicated CD20+ cells in blood and tissues. In human derived-models, IPH6501 significantly induced NK cell proliferation and cytotoxicity. Notably, IPH6501 showed potent activity against a range of B-NHL cell lines, including those with low CD20 expression. In addition, IPH6501 upregulated the expression of activating receptors on NK cells, such as NKG2D, essential for the recognition and killing of malignant cells, thereby introducing another mode of action enabling the elimination of CD20-negative tumor cells. In a comparative analysis with a CD3xCD20 TCE, IPH6501 induced lower cytokine secretion, suggesting a potentially safer profile. Additionally, IPH6501 showed higher killing efficacy compared to the TCE in samples from healthy individuals as well as R/R B-NHL patients, underscoring its therapeutic potential. Conclusions: The preclinical demonstrations of IPH6501 activities across various in vivo models and its effectiveness in ex vivo assays using cells from R/R B-NHL patients provide compelling evidence of its therapeutic potential and tolerability. IPH6501 is emerging as a promising new candidate within the treatment landscape for R/R B-NHL and is currently being investigated in a global, first-in-human phase 1/2 study (NCT 06088654).

Successful Treatment of Concomitant Myasthenia Gravis and Lambert-Eaton Myasthenic Syndrome with Autologous CD19-targeted CAR-T Cells (P4-14.006)

Successful Treatment of Concomitant Myasthenia Gravis and Lambert-Eaton Myasthenic Syndrome with Autologous CD19-targeted CAR-T Cells (P4-14.006)

Abstract CT244: HR001, a novel CD19-targeted CAR-T cell therapy for patients with relapsed/refractory non-Hodgkin's lymphoma: primary results of a phase II study (HRAIN01-NHL01-II)

Abstract Background: HR001 is a CD19-targeted CAR-T cell for patients with relapsed and refractory non-Hodgkin's (R/R NHL). Here, we report the primary results of the phase 2 study (HRAIN01-NHL01-II). Methods: HRAIN01-NHL01-II is an open-label, single-arm, and multicenter (17 centers) study in China. It enrolled R/R NHL patients treated with at least two lines of therapy (must-have anthracycline and CD20-targeted agents) or after autologous hematopoietic stem cell transplantation. All patients received a single infusion of HR001 at the 2.0 × 106 CAR+T cells/kg dose. The disease response was evaluated by the independent review committee with Lugano response criteria [2014]. Results: As of May 16th, 2023, 81 R/R NHL patients with ECOG 0-1 at baseline received the infusion HR001 from the Jinshan manufacturing facility [51.9% male; median age 53.4 years (range 23~74); 3.7% CD19 negative; 91.4% diffuse large B-cell lymphoma; 79.0% III-IV of Ann Arbor classification or Lugano] with a median follow-up of 160 days (range 7 to 454). The median time to best response was 30 days (range 28-358). The 3-month, 6 month and best objective response rate (ORR) was 53.1%(95% CI: 41.7-64.3), 45.7%(95% CI: 34.6-57.1) and 74.1% (95% CI: 63.1-83.2) respectively. The 3-month, 6 month and best complete response rate (CRR) was 32.1%(95% CI: 22.2-43.4), 29.6%(95% CI: 20.0-40.8) and 49.4%(95% CI: 38.1-60.7) respectively. Median duration of response (DOR) and progression-free survival (PFS) were 339 days (95% CI: 149-NE) and 176 days (95% CI: 91-NE) respectively. The overall survival (OS) was not reached. The most common treatment-related adverse events (AEs) happened in the hematologic system. The cytokine release syndrome (CRS) occurred in 95.1% of patients, with ≥3 grade CRS observed in 3.7% of patients. Immune effector cell-associated neurotoxicity syndrome (ICANS) happened in 8.6% of patients, without any ≥ 3 grade ICANS reported. The treatment-related mortality was 1.2%. The median time to CAR T-cell peak expansion was 240 hours (120~672) after infusion, median maximum expansion (Cmax) was 17413.9 copies/μg DNA (124.9~136382.2), and median AUC0-28d was 3201014.0 copies/μg DNA (205082.8~18400016.2). Positive tests for the anti-drug antibody (ADA) were noted in 14.8% of patients, while neutralizing antibody (Nab) tests were positive in 3.7% of patients. Among Nab-positive patients, the complete response was maintained until follow-up. Conclusions: Patients with R/R NHL could benefit from the HR001 treatment with satisfactory objective response, durable remission, and favorable safety profile. Citation Format: Yian Zhang, Yuhua Li, Kai Hu, Wuping Li, Hongwei Xue, Qi Deng, Zheng Ge, Liangquan Geng, Xianmin Song, Kang Yu, Hongmei Jing, Bing Chen, Xin Li, Zhimin Zhai, Jianping Shen, Jiang Cao, Qingming Wang, Peng Liu. HR001, a novel CD19-targeted CAR-T cell therapy for patients with relapsed/refractory non-Hodgkin's lymphoma: primary results of a phase II study (HRAIN01-NHL01-II) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr CT244.

Abstract 4018: An innovative in vivo model for CAR-T cell therapy development: Tolerability and efficacy evaluation of CD19-targeting CAR-T cells on human lymphoma using the chicken CAM assay

Abstract BACKGROUND Chimeric antigen receptor (CAR) T cell therapy is a revolutionary approach in cellular immunotherapy, where a patient’s T cells are genetically engineered to recognize and attack cancer cells. In the last decade, several CAR-T cell therapies have been approved by the FDA. However, this encouraging outcome was obtained only with certain blood cancers, like B cell leukemias, lymphomas, etc. Many research/clinical trials are now underway worldwide to extend the CAR T-cell therapy benefits to a large spectrum of cancers, using rodents. This model presents several drawbacks, such as the immunodeficiency of humanized models, ethical constraints, time, and cost. An alternative, pertinent, 3Rs-compliant in vivo model for developing CAR-T therapy is urgently needed. Xenografts on the chicken embryo’s ChorioAllantoic Membrane (CAM) have proven extremely valuable for in vivo cancerology studies. In this work, we evaluated in vivo the anti-tumor efficacy of CD19-targeting CAR-T cells on human lymphoma using Inovotion’s CAM assay. METHODS Human lymphoma Raji cells (ATCC) were grafted on the CAM on embryonic development day (EDD) 9. A 2nd generation of CD19-targeting CAR-T cells (CAR-T-(CD3/4-1BB)) developed by Allogenica (France) were co-injected at EDD9 at E/T (effector CAR-T cell vs target Raji cell) ratio 1:1. One group was injected with CAR-T (CD3/4-1BB) only once at EDD9 and a second group received a first injection of CAR-T (CD3/4-1BB) at EDD9 and a second injection at EDD13. Embryonic viability was checked daily. On EDD18. The efficacy of CAR-T therapy was evaluated basing on tumor weight, metastatic invasion in the lower CAM and CAR-T infiltration in tumors detected by qPCR. Activation of the chicken embryo immune system was detected as well. RESULTS The treatment with CAR-T-(CD3/4-1BB) cells was well tolerated in ovo, no significant embryo mortality/abnormality was observed. CAR-T-(CD3/4-1BB) cells prevented tumor formation (tumor weight was 88.67% to 90.62% lower than control, p &amp;lt; 0.001); and prevented metastasis formation in the lower CAM (86.88% to 81.43% lower than control, p &amp;lt; 0.001). CONCLUSION Different to in vitro models and immunodeficient murine models, chicken embryo possesses a progressively mature immune system. Our results show that CAR-T-(CD3/4-1BB) cells can prevent tumor formation and metastatic invasion when co-injected with tumor cells. Inovotion’s CAM assay is a viable alternative in vivo model for developing CAR-T cell therapy on a large spectrum of cancers. It is fully 3Rs-compliant. Citation Format: Yan Wang, Xavier Rousset, Chloé Prunier, Emilien Dosda, Alejandra Gutierrez-Guerrero, Pauline Abrial, Inna Menkova, Jean Viallet. An innovative in vivo model for CAR-T cell therapy development: Tolerability and efficacy evaluation of CD19-targeting CAR-T cells on human lymphoma using the chicken CAM assay [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4018.

Treatment of secondary CNS lymphoma using CD19-targeted chimeric antigen receptor (CAR) T cells

BackgroundAggressive B cell lymphoma with secondary central nervous system (CNS) involvement (SCNSL) carries a dismal prognosis. Chimeric antigen receptor (CAR) T cells (CAR-T) targeting CD19 have revolutionized the treatment for B cell lymphomas; however, only single cases with CNS manifestations successfully treated with CD19 CAR-T have been reported.MethodsWe prospectively enrolled 4 patients with SCNSL into our study to assess clinical responses and monitor T cell immunity.ResultsTwo of four SNCSL patients responded to the CD19-targeted CAR-T. Only one patient showed a substantial expansion of peripheral (PB) CAR-T cells with an almost 100-fold increase within the first week after CAR-T. The same patient also showed marked neurotoxicity and progression of the SNCSL despite continuous surface expression of CD19 on the lymphoma cells and an accumulation of CD4+ central memory-type CAR-T cells in the CNS. Our studies indicate that the local production of chemokine IP-10, possibly through its receptor CXCR3 expressed on our patient’s CAR-T, could potentially have mediated the local accumulation of functionally suboptimal anti-tumor T cells.ConclusionsOur results demonstrate expansion and homing of CAR-T cells into the CNS in SNCSL patients. Local production of chemokines such as IP-10 may support CNS infiltration by CAR-T cells but also carry the potential of amplifying local toxicity. Future studies investigating numbers, phenotype, and function of CAR-T in the different body compartments of SNSCL patients receiving CAR-T will help to improve local delivery of “fit” and highly tumor-reactive CAR-T with low off-target reactivity into the CNS.

Melatonin as an immunomodulator in CD19-targeting CAR-T cell therapy: managing cytokine release syndrome

BackgroundChimeric antigen receptor CAR-T cell therapies have ushered in a new era of treatment for specific blood cancers, offering unparalleled efficacy in cases of treatment resistance or relapse. However, the emergence of cytokine release syndrome (CRS) as a side effect poses a challenge to the widespread application of CAR-T cell therapies. Melatonin, a natural hormone produced by the pineal gland known for its antioxidant and anti-inflammatory properties, has been explored for its potential immunomodulatory effects. Despite this, its specific role in mitigating CAR-T cell-induced CRS remains poorly understood.MethodsIn this study, our aim was to investigate the potential of melatonin as an immunomodulatory agent in the context of CD19-targeting CAR-T cell therapy and its impact on associated side effects. Using a mouse model, we evaluated the effects of melatonin on CAR-T cell-induced CRS and overall survival. Additionally, we assessed whether melatonin administration had any detrimental effects on the antitumor efficacy and persistence of CD19 CAR-T cells.ResultsOur findings demonstrate that melatonin effectively mitigated the severity of CAR-T cell-induced CRS in the mouse model, leading to improved overall survival outcomes. Remarkably, melatonin administration did not compromise the antitumor effectiveness or persistence of CD19 CAR-T cells, indicating its compatibility with therapeutic goals. These results suggest melatonin's potential as an immunomodulatory compound to alleviate CRS without compromising the therapeutic benefits of CAR-T cell therapy.ConclusionThe study's outcomes shed light on melatonin's promise as a valuable addition to the existing treatment protocols for CAR-T cell therapies. By attenuating CAR-T cell-induced CRS while preserving the therapeutic impact of CAR-T cells, melatonin offers a potential strategy for optimizing and refining the safety and efficacy profile of CAR-T cell therapy. This research contributes to the evolving understanding of how to harness immunomodulatory agents to enhance the clinical application of innovative cancer treatments.

Management of children and adolescents receiving CAR-T cell therapy for acute lymphoblastic leukemia

Tisagenlecleucel, a commercially available CD19-targeted CAR-T cell product, has dramatically changed the treatment of relapsed/refractory B-cell acute lymphoblastic leukemia (B-ALL). Tisagenlecleucel infusion has been linked to distinct acute adverse events, including cytokine release syndrome, neurotoxicity, hemophagocytic lymphohistiocytosis and prolonged pancytopenia, which are rare with cytocidal chemotherapy. In addition, recent retrospective studies have revealed pre-infusion prognostic factors including high tumor burden (bone marrow leukemia cell fraction ≥5%) and non-response to blinatumomab, another CD19-targeting agent. Not only physicians providing CAR-T cell therapy but also those referring patients for this therapy should thoroughly understand the indications and limitations, characteristic acute complications, pre-treatment factors affecting prognosis, and late complications. This article outlines the current understanding regarding the use of tisagenlecleucel in children and adolescents with B-ALL.

Comment on: Rescue therapy of antisynthetase syndrome with CD19-targeted CAR-T cells after failure of several B-cell depleting antibodies.

Comment on: Rescue therapy of antisynthetase syndrome with CD19-targeted CAR-T cells after failure of several B-cell depleting antibodies.

Real-World Outcomes in Patients with Transformed Follicular Lymphoma Treated with CD19-Targeting CAR-T Therapy

INTRODUCTION. CD19-targeting CAR-T cell therapy has changed the treatment landscape of refractory/relapsed large B cell lymphoma, including transformed from FL (tFL). Axicabtagene ciloleucel (axi-cel), lisocabtagene maraleucel (liso-cel), and tisagenlecleucel (tisa-cel) are now FDA-approved for the treatment of tFL, but limited data are available on their clinical activity in patients (pts) with tFL since outcomes in those pts were not separately reported in registrational studies. Here, we report a single center's experience with CD19-directed CAR-T cell therapy in pts with tFL. METHODS. We performed a retrospective analysis of 54 pts with tFL treated with CAR-T cell therapy from January 2018 to December 2022. Pathological evidence of histologic transformation (HT) was confirmed. The main clinical-biological characteristics of the pts were extracted from the electronic medical records. Cytokine release syndrome (CRS) and neurotoxicity were graded according to ASTCT guidelines. Progression-free (PFS) and overall survival (OS) were measured from the time of CAR-T cell infusion; PFS events were death, relapse, and disease progression. RESULTS. Within our cohort of patients with tFL, the median age was 67 years (range 42-81), 65% were male and 77% had stage III-IV disease. Eight of the pts (15%) had an ECOG Performance Status (ECOG-PS) ≥2. Median time to transformation was 27 months from original FL diagnosis (range 0 to 223), including 6 cases with composite lymphoma (FL + aggressive histology) and 4 with synchronous transformation. Twenty percent presented with high-grade B-cell lymphoma (HGBCL) with MYC and BCL2 and/or BCL6 rearrangements. Using Hans' algorithm, 85% (44/52) were identified as a germinal center B-cell origin (GCB). In 22 cases, targeted sequencing using the MSK-IMPACT panel was performed at the time of HT or prior to CAR-T. TP53 mutation/deletion was present in 50% of the cases (Figure 1). Pts received a median of 3 (range 1-9) prior lines of therapy including those for FL, and 9 pts (17%) had undergone prior autologous stem cell transplantation (SCT). Thirty-four pts (63%) were refractory to the last therapy, including 29 pts (53.7%) with primary refractory disease after HT. Forty-one pts (76%) received bridging therapy. Seventy-two percent of pts received axi-cel, 19% tisa-cel, and 9% liso-cel. CRS of any grade was seen in 43 (80%) pts with grade &amp;gt;3 in 3 (6%). Neurologic toxicity of any grade was observed in 20 (37%) pts with grade &amp;gt;3 in 7 (13%). The median duration of follow-up was 26.2 months. The overall response rate was 87%, including 67% complete response (CR). The median response duration (DOR) was 11 months, with a 2-year DOR of 42% (95% CI: 28-61%). Objective responses were achieved in pts with high-risk features such as HGBCL, TP53 mutation, and chemotherapy-refractory disease. Twenty-nine pts experienced relapse/progression during follow-up, with a median PFS for the entire cohort of 9.8 months and a 2-year PFS of 40% (95% CI: 28.3-56.3%) (Figure 2). Refractory disease after HT and high LDH serum levels were associated with a significantly lower PFS (median PFS not reached vs. 6.4 months, P=0.031; 24.4 vs. 3.2 months, P=0.003 respectively). Notably, all pts who achieved PR eventually progressed. In 23 pts with relapse, a biopsy was obtained, showing DLBCL or HGBCL morphology in all instances. Interestingly, all relapses occurred during the first 24 months. Twenty-six pts received salvage therapy after CAR-T progression, with a median of 1 line (range 1-7). Out of these, eight (31%) pts achieved a CR and went to allogenic SCT as a consolidative strategy. Twenty-two pts died during follow-up, 19 due to progression and 3 of non-lymphoma causes, including 2 of COVID-19 infection. The median OS was 31 months, and the 2-year OS rate was 57% (95% CI: 44.3-73.8%) (Figure 2). Median OS was not reached among the 36 pts who achieved a complete response. CONCLUSIONS. CD19-targeting CAR-T cell therapy is effective for pts with relapsed/refractory tFL, including patients with characteristics of poor prognosis, with efficacy and toxicity profiles comparable to that observed in previous DLBCL studies. Unfortunately, pts who relapse post-CAR-T therapy have limited effective therapeutic options available, and this currently remains an unmet medical need within the field.

Comparative Analysis of Immune Effector Cell-Associated Hematotoxicity (ICAHT) and Infection Incidence Following CD19-Targeted CAR-T Therapy Versus CD7-Targeted CAR-T Therapy

Introduction CD19 CAR-T cell therapy has been revolutionizing the treatment of refractory /relapsed (R/R) B-cell acute lymphoblastic leukemia (B-ALL) over the past decade. While CD7 CAR-T therapy, an innovative approach, has only recently been applied over the last three years in our center to treat R/R T-cell acute lymphoblastic leukemia (T-ALL) and T-cell lymphoblastic lymphoma (T-LBL). While there are studies and guidelines from EBMT addressing hematologic toxicity, specifically Immune Effector Cell-Associated Hematotoxicity (ICAHT), following CD19 CAR-T treatment, the literature is still scarce on ICAHT occurrence post-CD7 CAR-T therapy. Here we conducted a retrospective study aiming at comparing ICAHT and infection incidences following the two distinct CAR-T therapies. Methods Data were extracted from two separate clinical trials. The CD19 CAR is structured with CD19 scFv, CD28-hinge, 4-1BB costimulatory region, and CD3ζ activation domains. T cells were activated by CD3/CD28 microbead 2 days before CD19-CAR lentivirus infection. “Naturally selected” anti-CD7 CAR-T cells (NS7CAR) were transduced an anti-CD7 CAR into bulk peripheral T lymphocytes via a lentivirus vector. NS7CAR is a 2nd generation murine-based CAR-T containing 4-1BB and CD3ζ co-stimulatory domains. All patients received lymphodepletion regimens of fludarabine (30 mg/m 2/d) and cyclophosphamide (300 mg/m 2/d) from Day -5 to Day -3 before CAR-T infusion. A single infusion of either CD19 CAR-T or CD7 CAR-T cells was administrated at a medium dose of 1×10 6/kg. Fitted patients undergo consolidation allogeneic hematopoietic stem cell transplantation (Allo-HSCT) within 1-3 months after CAR-T cell infusion. The hematology toxicity and infection were analyzed during the period from CD19 CAR-T or CD7 CAR-T cell infusion to Allo-HSCT. Hematology toxicity is graded according to “ICAHT: EHA/EBMT Consumption Grading and Best Practice Recommendations” (Table 1). Infection diagnosis includes both etiological diagnosis and imaging diagnosis. Results Between June 1, 2019, and September 22, 2022, 60 patients with R/R B-ALL patients received CD19 CAR-T therapy (https://clinicaltrials.gov NCT04546893, and NCT04792593). Meanwhile, between December 1, 2020, and July 16, 2022, 60 patients with R/R T-ALL/LBL received CD7 CAR-T therapy (NCT04572308, NCT04916860). The CD19 CART group had 19 patients ≤ 14 years old, with a male/female gender ratio of 33:27. The CD7 CART group had 20 patients ≤ 14 years old, with a gender ratio of 49:11. The incidence of severe early ICAHT (Grade III+IV, day 0-30) from CD19 CAR-T cells infusion to allo-HSCT was observed to be 18.3%, which was lower than the incidence of the early ICAHT post CD7 CAR-T therapy at 33.3%, p =0.035. However, the incidence of severe late ICAHT (after day+30) following CD19 CAR-T and the CD7 CAR-T therapy was similar: 27.8% vs. 28.6%, with no statistical difference( p=0.57). There was no statistical difference between CD19 CAR-T and CD7 CAR-T therapy groups in the infection rates with 28.3%vs. 36.7%, p=0.33. In the CD19 CAR-T group, bacterial infections were noted in 10 patients: 6 with Gram-positive bacteria (1 Enterococcus, 1 Streptococcus agalactis, and 4 Staphylococcus), and 4 with Gram-negative bacteria (1 Pseudomonas aeruginosa, 2 Acinetobacter baumannii, and 1 Klebsiella pneumoniae). Viral infections were observed in 7 patients (EBV 2, CMV 2, BKV 3), and 1 had fungal infection. Infection sites included pulmonary infections (7 cases), sepsis (10 cases), and other areas (4 cases). Post-CD7-CAR-T, bacterial infections were observed in 9 cases, 7 with Gram-positive bacteria (6 Staphylococcus, and 1 Streptococcus bradycardia) and 2 with Gram-negative bacteria (1 Pseudomonas aeruginosa, and 1 Klebsiella pneumoniae). Six cases had viral infections (EBV 2, CMV 2, HHV1 type 1, parainfluenza virus 1), and 6 with fungal infections. Infection sites included pulmonary infection (12 cases), sepsis (10 cases), and other areas (5 cases). Conclusions Our study noted a slightly higher incidence of ICAHT within 30 days following CD7 CAR-T infusion compared to CD19 CAR-T therapy. However, there was no statistical difference in the incidence of ICAHT beyond the 30-day. There was no statistical difference for infection rate after CD19 CAR-T and CD7 CAR-T. In both groups, Gram-positive bacteria were primary responsible for bacterial infections.

High Levels of Circulating Granulocytic Myeloid-Derived Suppressor Cells (G-MDSCs) Predict Failure of CD19-Targeting CAR-T Cell Therapy

Background The mechanisms of resistance to CAR-T cell therapy may depend on the physical and biological characteristics of the tumor itself, on CAR-T cells in terms of efficiency and fitness, or on the microenvironment. Myeloid-derived suppressor cells (MDSCs) play a crucial role in hematological tumors, promoting immunosuppression and tumor progression. Their presence hampers the anti-tumor immune response, posing challenges for effective therapeutic interventions. The predictive role of myeloid-derived suppressor cells (MDSCs) of monocytic and granulocytic origin (M-MDSCs and G-MDSCs, respectively) in the context of CAR-T cell therapy is largely unknown. Aim and methods Our aim was to quantify circulating M-MDSC and G-MDSC in patients (n=26) receiving approved CAR-T cell products for high-grade B cell lymphoma or acute lymphoblastic leukemia. Whole blood samples were collected at enrolment (time point 1), after leukapheresis (time point 2) and just after Cy/Flu lymphodepletion (time point 3). Stringent criteria for multicolour flow cytometry identification of M-MDSC (CD14/HLA-DR/CD124/CD11b/CD45 and light scattering) and G-MDSC (CD15/CD11b/CD66b/oxidized low-density lipoprotein receptor -1, LOX-1, /CD45 and light scattering) were used. Blood samples were examined immediately after collection to prevent artefactual G-MDSC changes. Results M-MDSC number, although very variable between patients, remained essentially constant from time point 1 to time point 3 in each individual (range 1.2-947 cells/ml at the latest time point) and was not associated with a higher risk of relapse. Conversely, G-MDSC number, which was extremely low at time points 1 and 2, markedly increased at time point 3 (range 1.5-85.5 cells/ml). According to our preliminary data, at time point 3, mean G-MDSC were 39.3, 44.5, and 12.3 cells/ml in tisacel, axi-cel and brexu-cel, respectively. Mean G-MDSC were significantly lower in patients treated to receive brexu-cel (Figure 1). Responses were evaluated one, three, and six months after receiving CAR-T cells infusion. As we aimed at finding if G-MDSC could predict prognosis, we set a cut-off value of 30 G-MDSC/ml at time-point 3 (immediately after the completion of lymphodepletion); overall, approximately 35% of the patients had G-MDSC &amp;gt;30/ml at that moment. Kaplan-Meier survival analysis revealed that G-MDSC &amp;gt;30/ml at time point 3 associated with a higher risk of relapse, with a PFS of 80% vs 10% six months after CAR-T infusion ( p= 0.0002) (Figure 1). The sudden appearance of G-MDSC following the lymphodepletion is likely to reflect emergency granulopoiesis that favors an increased release of G-MDSC to the bloodstream and may be ideally customized with a tailored lymphodepletion. Conclusion In conclusion, here we show that the high circulating levels of G-MDSC induced by the lymphodepletion are associated with a lack of response to CAR-T cell therapy, thus representing a promising predictive biomarker of CAR-T cell therapy efficacy and possibly a hypothetical point of intervention for ameliorating CAR-T efficacy.

Trial in Progress: A Phase I/II Study of Tafasitamab Plus Lenalidomide in Patients with Relapsed CNS Lymphoma

Background and Significance The development of novel, more effective biological strategies to prevent secondary CNS lymphoma (CNSL) and to treat primary and secondary CNSL would have significant impact. It is well established that the CD19 transmembrane glycoprotein is expressed by B-cells throughout differentiation, by the majority of B cell malignancies, and by follicular dendritic cells. Given the high incidence of neurotoxicity associated with blood-brain barrier disruption with CD19-directed CAR-T cells and bi-specific T-cell engager antibodies, Parker et al. recently investigated the possibility of CD19 expression by previously unrecognized cell population(s) in the brain. Single-cell RNA sequencing data demonstrated CD19 expression in brain mural cells, an integral component of the neurovascular unit that regulates integrity of the blood-brain barrier.(Cell, 2020; 183:126-42). Given these observations and the compelling problem of relapsed primary and secondary CNS lymphoma, we are evaluating tafasitamab, an anti-CD19 humanized monoclonal antibody, as a novel therapeutic for refractory CNS lymphomas. We hypothesize that tafasitamab may modulate blood-brain barrier integrity by perturbation of CD19 expressed on mural cells; but without recruitment of cytotoxic T cells and resultant cytokine release syndrome. Our rationale is supported by the Phase II L-MIND study, in which tafasitamab plus lenalidomide exhibited synergistic efficacy against relapsed/refractory aggressive non-Hodgkin lymphoma, with overall response rate of 60%, complete response rate of 43%, and a favorable toxicity profile compared to CAR-T without neurotoxicity. Notably, the L-MIND trial excluded patients previously treated with immunomodulatory drugs as well as CNS lymphoma. (Lancet Oncology 2020: 21:978-88). We also recently reported that tafasitamab plus lenalidomide induced brain parenchymal regressions in two consecutive CNS lymphoma patients with disease refractory to lenalidomide, pomalidomide and rituximab. (British Journal of Haematology 2023 201(1):154-157). Study Design and Methods This single-arm, open-label, multicenter Phase I/II study (NCT05351593) assesses safety, CNS pharmacokinetics and preliminary efficacy of tafasitamab plus lenalidomide in patients with relapsed primary and secondary CNSL. The Phase I portion of the study will determine tolerated dose (MTD) and recommended Phase II dose (RP2D) of lenalidomide in combination with tafasitamab. We are applying 3+3 dose escalation rules to examine lenalidomide 10mg, 15mg and 20 mg plus tafasitamab at 12 mg/kg. After the RP2D is identified, the Phase II portion of the study will begin. The primary objective of the Phase II expansion is to evaluate the clinical benefit rate (stable disease or better) via Simon's minimax two-stage design. Exploratory endpoints include assessment of effect of tafasitamab on blood-brain barrier permeability, correlation of the relationships between tumor response or resistance with tumor mutational profile, and changes in CSF and blood immune cell phenotypes. The study plans to enroll up to 35 patients. Eligibility mandates age ≥ 18 years, anticipated survival &amp;gt;2 months, ECOG PS 0-1, and receipt of at least one prior systemic therapy. Patients with prior exposure to lenalidomide or pomalidomide or concomitant systemic lymphoma are allowed. Patients with HIV infection, CNS PTLD, and those who previously received a CD19-targeting CAR-T product are ineligible. The study is currently enrolling at the University of California San Francisco, with plans to expand enrollment to other institutions within the United States.

Effects of B Cell Depletion by CD19-Targeted Chimeric Antigen Receptor T Cells in a Murine Model of Systemic Sclerosis.

Our goal was to study the tolerance and efficacy of two B cell depletion strategies, including one with CD19-targeted chimeric antigen receptor (CAR) T cells, in a preclinical model mimicking the severe lung damages observed in systemic sclerosis. B cell depletion strategies were evaluated in the Fra-2 transgenic (Tg) mouse model. We considered a first group of 16 untreated mice, a second group of 15 mice receiving a single dose of anti-CD20 monoclonal antibody (mAb), and a third group of 8 mice receiving CD19-targeted CAR-T cells in combination with anti-CD20 monoclonal antibody. After six weeks of clinical evaluation, different validated markers of inflammation, lung fibrosis, and pulmonary vascular remodeling were assessed. CD19-targeted CAR-T cells infusion in combination with anti-CD20 mAb resulted in a deeper B cell depletion than anti-CD20 mAb alone in the peripheral blood and lesional lungs of Fra-2 Tg mice. CAR-T cell infusion worsened the clinical score and increased mortality in Fra-2 Tg mice. In line with the above findings, CAR-T cell infusion significantly increased lung collagen content, the histological fibrosis score, and right ventricular systolic pressure. CAR-T cells accumulated in lesional lungs and promoted T activation and inflammatory cytokine production. Treatment with anti-CD20 mAb in monotherapy had no impact on lung inflammation-driven fibrosis and pulmonary hypertension. B cell therapies failed to show efficacy in the Fra2 Tg mice. The exacerbated Fra-2 lung inflammatory burden stimulated accumulation and expansion of activated CD19-targeted CAR-T cells, secondarily inducing T cell activation and systemic inflammation, finally leading to disease worsening.