Evolving strategies to overcome barriers in CAR-T cell therapy for acute myeloid leukemia.

INTRODUCTION: Acute myeloid leukemia (AML) is a complex and heterogeneous disease characterized by an aggressive clinical course and limited efficacious treatment options in the relapsed/refractory (R/R) setting. Chimeric antigen receptor (CAR)-modified T (CAR-T) cell immunotherapy is an investigational treatment strategy for R/R AML that has shown some promise. However, obstacles to successful CAR-T cell immunotherapy for AML remain. AREAS COVERED: In analyses of clinical trials of CAR-T cell therapy for R/R AML, complete responses without measurable residual disease have been reported, but the durability of those responses remains unclear. Significant barriers to successful CAR-T cell therapy in AML include the scarcity of suitable tumor-target antigens (TTA), inherent T cell functional deficits, and the immunoinhibitory and hostile tumor microenvironment (TME). This review will focus on these barriers to successful CAR-T cell therapy in AML, and discuss scientific advancements and evolving strategies to overcome them. EXPERT OPINION: Achieving durable remissions in R/R AML will likely require a multifaceted approach that integrates advancements in TTA selection, enhancement of the intrinsic quality of CAR-T cells, and development of strategies to overcome inhibitory mechanisms in the AML TME.

Targeting the membrane-proximal C2-set domain of CD33 for improved CAR T cell therapy.

Current CD33-targeted immunotherapies typically recognize the membrane-distal V-set domain of CD33. Here, we show that decreasing the distance between T cell and leukemia cell membrane increases the efficacy of CD33 chimeric antigen receptor (CAR) T cells. We therefore generated and optimized second-generation CAR constructs containing single-chain variable fragments from antibodies raised against the membrane-proximal C2-set domain, which bind CD33 regardless of whether the V-set domain is present (CD33PAN antibodies). CD33PAN CAR T cells resulted in efficient tumor clearance and improved survival of immunodeficient mice bearing human AML cell xenografts and, in an AML model with limited CD33 expression, forced escape of CD33neg leukemia. Compared to CD33V-set CAR T cells, CD33PAN CAR T cells showed greater in vitro and in vivo efficacy against several human AML cell lines with differing levels of CD33 without increased expression of exhaustion markers. CD33PAN moieties were detected at a higher frequency on human leukemic stem cells, and CD33PAN CAR T cells had greater in vitro efficacy against primary human AML cells. Together, our studies demonstrate improved efficacy with CAR T cells binding CD33 close to the cell membrane, providing the rationale to investigate CD33PAN CAR T cells further toward possible clinical application.

PD-L1+ macrophage and tumor cell abundance and proximity to T cells in the pretreatment large B-cell lymphoma microenvironment impact CD19 CAR-T cell immunotherapy efficacy.

CD19-targeted chimeric antigen receptor T-cell (CAR-T) immunotherapy has transformed the management of relapsed/refractory large B-cell lymphoma (LBCL), yet durable remissions are observed in less than half of treated patients. The tumor microenvironment (TME) is a key and understudied factor impacting CD19 CAR-T therapy outcomes. Using NanoString nCounter transcriptome profiling (n = 24) and multiplex immunohistochemistry (mIHC, n = 15), we studied the TME in pretreatment biopsies from patients with LBCL undergoing CD19 CAR-T therapy. Patients who achieved complete response (CR) after CAR-T therapy demonstrated higher expression of genes associated with T-cell trafficking and function, whereas those who did not achieve CR had higher expression of genes associated with macrophages and T-cell dysfunction. Distinct patterns of immune infiltration and fibrosis in the TME were associated with CAR-T therapy outcomes, and these findings were corroborated using artificial intelligence-assisted image analyses. Patients who achieved CR had a lower proportion of the biopsy occupied by an interspersed immune infiltrate and a higher proportion of hypocellular/fibrotic regions. Furthermore, mIHC revealed lower density of CD4+ T cells and higher densities of both macrophages and tumor cells expressing PD-L1 in non-CR patients. Spatial analysis revealed that PD-1+ T cells were in close proximity to PD-L1+ macrophages or PD-L1+ tumor cells in patients who did not compared to those who did achieve CR after CAR-T therapy. These findings suggest that morphologic patterns in the TME and engagement of the PD-1/PD-L1 axis in pretreatment biopsies may impact CD19 CAR-T immunotherapy response in patients with LBCL.

qPCR assay for detection of Woodchuck Hepatitis Virus Post-Transcriptional Regulatory Elements from CAR-T and TCR-T cells in fresh and formalin-fixed tissue.

As adoptive cellular therapies become more commonplace in cancer care, there is a growing need to monitor site-specific localization of engineered cells-such as chimeric antigen receptor T (CAR-T) cells and T-cell receptor T (TCR-T) cells-in patients' tissues to understand treatment effectiveness as well as associated adverse events. Manufacturing CAR-T and TCR-T cells involves transduction with viral vectors commonly containing the WPRE gene sequence to enhance gene expression, providing a viable assay target unique to these engineered cells. Quantitative PCR (qPCR) is currently used clinically in fresh patient tissue samples and blood with target sequences specific to each immunotherapy product. Herein, we developed a WPRE-targeted qPCR assay that is broadly applicable for detection of engineered cell products in both fresh and archival formalin-fixed paraffin embedded (FFPE) tissues. Using both traditional PCR and SYBR Green PCR protocols, we demonstrate the use of this WPRE-targeted assay to successfully detect two CAR-T cell and two TCR-T cell products in FFPE tissue. Standard curve analysis reported a reproducible limit of detection at 100 WPRE copies per 20µL PCR reaction. This novel and inexpensive technique could provide better understanding of tissue abundance of engineered therapeutic T cells in both tumor and second-site toxicity tissues and provide quantitative assessment of immune effector cell trafficking in archival tissue.

Stealth transgenes enable CAR-T cells to evade host immune responses.

BACKGROUND: Adoptive cell therapy, such as chimeric antigen receptor (CAR)-T cell therapy, has improved patient outcomes for hematological malignancies. Currently, four of the six FDA-approved CAR-T cell products use the FMC63-based αCD19 single-chain variable fragment, derived from a murine monoclonal antibody, as the extracellular binding domain. Clinical studies demonstrate that patients develop humoral and cellular immune responses to the non-self CAR components of autologous CAR-T cells or donor-specific antigens of allogeneic CAR-T cells, which is thought to potentially limit CAR-T cell persistence and the success of repeated dosing. METHODS: In this study, we implemented a one-shot approach to prevent rejection of engineered T cells by simultaneously reducing antigen presentation and the surface expression of both Classes of the major histocompatibility complex (MHC) via expression of the viral inhibitors of transporter associated with antigen processing (TAPi) in combination with a transgene coding for shRNA targeting class II MHC transactivator (CIITA). The optimal combination was screened in vitro by flow cytometric analysis and mixed lymphocyte reaction assays and was validated in vivo in mouse models of leukemia and lymphoma. Functionality was assessed in an autologous setting using patient samples and in an allogeneic setting using an allogeneic mouse model. RESULTS: The combination of the Epstein-Barr virus TAPi and an shRNA targeting CIITA was efficient and effective at reducing cell surface MHC classes I and II in αCD19 'stealth' CAR-T cells while retaining in vitro and in vivo antitumor functionality. Mixed lymphocyte reaction assays and IFNγ ELISpot assays performed with T cells from patients previously treated with autologous αCD19 CAR-T cells confirm that CAR T cells expressing the stealth transgenes evade allogeneic and autologous anti-CAR responses, which was further validated in vivo. Importantly, we noted anti-CAR-T cell responses in patients who had received multiple CAR-T cell infusions, and this response was reduced on in vitro restimulation with autologous CARs containing the stealth transgenes. CONCLUSIONS: Together, these data suggest that the proposed stealth transgenes may reduce the immunogenicity of autologous and allogeneic cellular therapeutics. Moreover, patient data indicate that repeated doses of autologous FMC63-based αCD19 CAR-T cells significantly increased the anti-CAR T cell responses in these patients.

Unveiling the proteome-wide autoreactome enables enhanced evaluation of emerging CAR T cell therapies in autoimmunity.

Given the global surge in autoimmune diseases, it is critical to evaluate emerging therapeutic interventions. Despite numerous new targeted immunomodulatory therapies, comprehensive approaches to apply and evaluate the effects of these treatments longitudinally are lacking. Here, we leveraged advances in programmable-phage immunoprecipitation methodology to explore the modulation, or lack thereof, of autoantibody profiles, proteome-wide, in both health and disease. Using a custom set of over 730,000 human-derived peptides, we demonstrated that each individual, regardless of disease state, possesses a distinct and complex constellation of autoreactive antibodies. For each individual, the set of resulting autoreactivites constituted a unique immunological fingerprint, or "autoreactome," that was remarkably stable over years. Using the autoreactome as a primary output, we evaluated the relative effectiveness of various immunomodulatory therapies in altering autoantibody repertoires. We found that therapies targeting B cell maturation antigen (BCMA) profoundly altered an individual's autoreactome, while anti-CD19 and anti-CD20 therapies had minimal effects. These data both confirm that the autoreactome comprises autoantibodies secreted by plasma cells and strongly suggest that BCMA or other plasma cell-targeting therapies may be highly effective in treating currently refractory autoantibody-mediated diseases.

A stochastic framework for evaluating CAR T cell therapy efficacy and variability.

Based on a deterministic and stochastic process hybrid model, we use white noises to account for patient variabilities in treatment outcomes, use a hyperparameter to represent patient heterogeneity in a cohort, and construct a stochastic model in terms of Ito stochastic differential equations for testing the efficacy of three different treatment protocols in CAR T cell therapy. The stochastic model has three ergodic invariant measures which correspond to three unstable equilibrium solutions of the deterministic system, while the ergodic invariant measures are attractors under some conditions for tumor growth. As the stable dynamics of the stochastic system reflects long-term outcomes of the therapy, the transient dynamics provide chances of cure in short-term. Two stopping times, the time to cure and time to progress, allow us to conduct numerical simulations with three different protocols of CAR T cell treatment through the transient dynamics of the stochastic model. The probability distributions of the time to cure and time to progress present outcome details of different protocols, which are significant for current clinical study of CAR T cell therapy.

Timing of anti-PD-L1 antibody initiation affects efficacy/toxicity of CD19 CAR T-cell therapy for large B-cell lymphoma.

ABSTRACT: More than half of the patients treated with CD19-targeted chimeric antigen receptor (CAR) T-cell immunotherapy for large B-cell lymphoma (LBCL) do not achieve durable remission, which may be partly due to PD-1/PD-L1-associated CAR T-cell dysfunction. We report data from a phase 1 clinical trial (NCT02706405), in which adults with LBCL were treated with autologous CD19 CAR T cells (JCAR014) combined with escalating doses of the anti-PD-L1 monoclonal antibody, durvalumab, starting either before or after CAR T-cell infusion. The addition of durvalumab to JCAR014 was safe and not associated with increased autoimmune or immune effector cell-associated toxicities. Patients who started durvalumab before JCAR014 infusion had later onset and shorter duration of cytokine release syndrome and inferior efficacy, which was associated with slower accumulation of CAR T cells and lower concentrations of inflammatory cytokines in the blood. Initiation of durvalumab before JCAR014 infusion resulted in an early increase in soluble PD-L1 (sPD-L1) levels that coincided with the timing of maximal CAR T-cell accumulation in the blood. In vitro, sPD-L1 induced dose-dependent suppression of CAR T-cell effector function, which could contribute to inferior efficacy observed in patients who received durvalumab before JCAR014. Despite the lack of efficacy improvement and similar CAR T-cell kinetics early after infusion, ongoing durvalumab therapy after JCAR014 was associated with re-expansion of CAR T cells in the blood, late regression of CD19+ and CD19- tumors, and enhanced duration of response. Our results indicate that the timing of initiation of PD-L1 blockade is a key variable that affects outcomes after CD19 CAR T-cell immunotherapy for adults with LBCL.

Factors associated with long-term outcomes of CD19 CAR T-cell therapy for relapsed/refractory CLL.

High response rates have been reported after CD19-targeted chimeric antigen receptor-modified (CD19 CAR) T-cell therapy for relapsed/refractory (R/R) chronic lymphocytic leukemia (CLL), yet the factors associated with duration of response in this setting are poorly characterized. We analyzed long-term outcomes in 47 patients with R/R CLL and/or Richter transformation treated on our phase 1/2 clinical trial of CD19 CAR T-cell therapy with an updated median follow-up of 79.6 months. Median progression-free survival (PFS) was 8.9 months, and the 6-year PFS was 17.8%. Maximum standardized uptake value (hazard ratio [HR], 1.15; 95% confidence interval [CI], 1.07-1.23; P < .001) and bulky disease (≥5 cm; HR, 2.12; 95% CI, 1.06-4.26; P = .034) before lymphodepletion were associated with shorter PFS. Day +28 complete response by positron emission tomography-computed tomography (HR, 0.13; 95% CI, 0.04-0.40; P < .001), day +28 measurable residual disease (MRD) negativity by multiparameter flow cytometry (HR, 0.08; 95% CI, 0.03-0.22; P < .001), day +28 MRD negativity by next-generation sequencing (HR, 0.21; 95% CI, 0.08-0.51; P < .001), higher peak CD8+ CAR T-cell expansion (HR, 0.49; 95% CI; 0.36-0.68; P < .001), higher peak CD4+ CAR T-cell expansion (HR, 0.47; 95% CI; 0.33-0.69; P < .001), and longer CAR T-cell persistence (HR, 0.56; 95% CI, 0.44-0.72; P < .001) were associated with longer PFS. The 6-year duration of response and overall survival were 26.4% and 31.2%, respectively. CD19 CAR T-cell therapy achieved durable responses with curative potential in a subset of patients with R/R CLL. This trial was registered at www.clinicaltrials.gov as #NCT01865617.

γ-Secretase inhibitor in combination with BCMA chimeric antigen receptor T-cell immunotherapy for individuals with relapsed or refractory multiple myeloma: a phase 1, first-in-human trial.

BACKGROUND: γ-Secretase inhibitors (GSIs) increase B cell maturation antigen (BCMA) density on malignant plasma cells and enhance antitumour activity of BCMA chimeric antigen receptor (CAR) T cells in preclinical models. We aimed to evaluate the safety and identify the recommended phase 2 dose of BCMA CAR T cells in combination with crenigacestat (LY3039478) for individuals with relapsed or refractory multiple myeloma. METHODS: We conducted a phase 1, first-in-human trial combining crenigacestat with BCMA CAR T-cells at a single cancer centre in Seattle, WA, USA. We included individuals aged 21 years or older with relapsed or refractory multiple myeloma, previous autologous stem-cell transplant or persistent disease after more than four cycles of induction therapy, and Eastern Cooperative Oncology Group performance status of 0-2, regardless of previous BCMA-targeted therapy. To assess the effect of the GSI on BCMA surface density on bone marrow plasma cells, participants received GSI during a pretreatment run-in, consisting of three doses administered 48 h apart. BCMA CAR T cells were infused at doses of 50 × 106 CAR T cells, 150 × 106 CAR T cells, 300 × 106 CAR T cells, and 450 × 106 CAR T cells (total cell dose), in combination with the 25 mg crenigacestat dosed three times a week for up to nine doses. The primary endpoints were the safety and recommended phase 2 dose of BCMA CAR T cells in combination with crenigacestat, an oral GSI. This study is registered with ClinicalTrials.gov, NCT03502577, and has met accrual goals. FINDINGS: 19 participants were enrolled between June 1, 2018, and March 1, 2021, and one participant did not proceed with BCMA CAR T-cell infusion. 18 participants (eight [44%] men and ten [56%] women) with multiple myeloma received treatment between July 11, 2018, and April 14, 2021, with a median follow up of 36 months (95% CI 26 to not reached). The most common non-haematological adverse events of grade 3 or higher were hypophosphataemia in 14 (78%) participants, fatigue in 11 (61%), hypocalcaemia in nine (50%), and hypertension in seven (39%). Two deaths reported outside of the 28-day adverse event collection window were related to treatment. Participants were treated at doses up to 450 × 106 CAR+ cells, and the recommended phase 2 dose was not reached. INTERPRETATIONS: Combining a GSI with BCMA CAR T cells appears to be well tolerated, and crenigacestat increases target antigen density. Deep responses were observed among heavily pretreated participants with multiple myeloma who had previously received BCMA-targeted therapy and those who were naive to previous BCMA-targeted therapy. Further study of GSIs given with BCMA-targeted therapeutics is warranted in clinical trials. FUNDING: Juno Therapeutics-a Bristol Myers Squibb company and the National Institutes of Health.

Anakinra for Refractory Cytokine Release Syndrome or Immune Effector Cell-Associated Neurotoxicity Syndrome after Chimeric Antigen Receptor T Cell Therapy.

Chimeric antigen receptor-engineered (CAR)-T cell therapy remains limited by significant toxicities, including cytokine release syndrome (CRS) and immune effector cell-associated neurotoxicity syndrome (ICANS). The optimal management of severe and/or refractory CRS/ICANS remains ill-defined. Anakinra has emerged as a promising agent based on preclinical data, but its safety and efficacy in CAR-T therapy recipients are unknown. The primary objective of this study was to evaluate the safety of anakinra to treat refractory CRS and ICANS after CAR-T therapy. The secondary objective was to evaluate the impact of key treatment-, patient-, and disease-related variables on the time to CRS/ICANS resolution and treatment-related mortality (TRM). We retrospectively analyzed the outcomes of 43 patients with B cell or plasma cell malignancies treated with anakinra for refractory CRS or ICANS at 9 institutions in the United States and Spain between 2019 and 2022. Cause-specific Cox regression was used to account for competing risks. Multivariable cause-specific Cox regression was used to estimate the effect of anakinra dose on outcomes while minimizing treatment allocation bias by including age, CAR-T product, prelymphodepletion (pre-LD) ferritin, and performance status. Indications for anakinra treatment were grade ≥2 ICANS with worsening or lack of symptom improvement despite treatment with high-dose corticosteroids (n = 40) and grade ≥2 CRS with worsening symptoms despite treatment with tocilizumab (n = 3). Anakinra treatment was feasible and safe; discontinuation of therapy because of anakinra-related side effects was reported in only 3 patients (7%). The overall response rate (ORR) to CAR-T therapy was 77%. The cumulative incidence of TRM in the whole cohort was 7% (95% confidence interval [CI], 2% to 17%) at 28 days and 23% (95% CI, 11% to 38%) at 60 days after CAR-T infusion. The cumulative incidence of TRM at day 28 after initiation of anakinra therapy was 0% in the high-dose (>200 mg/day i.v.) recipient group and 47% (95% CI, 20% to 70%) in the low-dose (100 to 200 mg/day s.c. or i.v.) recipient group. The median cumulative incidence of CRS/ICANS resolution from the time of anakinra initiation was 7 days in the high-dose group and was not reached in the low-dose group, owing to the high TRM in this group. Univariate Cox modeling suggested a shorter time to CRS/ICANS resolution in the high-dose recipients (hazard ratio [HR], 2.19; 95% CI, .94 to 5.12; P = .069). In a multivariable Cox model for TRM including age, CAR-T product, pre-LD ferritin level, and pre-LD Karnofsky Performance Status (KPS), higher anakinra dose remained associated with lower TRM (HR, .41 per 1 mg/kg/day increase; 95% CI, .17 to .96; P = .039. The sole factor independently associated with time to CRS/ICANS resolution in a multivariable Cox model including age, CAR-T product, pre-LD ferritin and anakinra dose was higher pre-LD KPS (HR, 1.05 per 10% increase; 95% CI, 1.01 to 1.09; P = .02). Anakinra treatment for refractory CRS or ICANS was safe at doses up to 12 mg/kg/day i.v. We observed an ORR of 77% after CAR-T therapy despite anakinra treatment, suggesting a limited impact of anakinra on CAR-T efficacy. Higher anakinra dose may be associated with faster CRS/ICANS resolution and was independently associated with lower TRM. Prospective comparative studies are needed to confirm our findings.

Anti-HLA antibodies in recipients of CD19 versus BCMA-targeted CAR T-cell therapy.

Antibodies against foreign human leukocyte antigen (HLA) molecules are barriers to successful organ transplantation. B cell-depleting treatments are used to reduce anti-HLA antibodies but have limited efficacy. We hypothesized that the primary source for anti-HLA antibodies is long-lived plasma cells, which are ineffectively targeted by B cell depletion. To study this, we screened for anti-HLA antibodies in a prospectively enrolled cohort of 49 patients who received chimeric antigen receptor T-cell therapy (CARTx), targeting naïve and memory B cells (CD19-targeted, n = 21) or plasma cells (BCMA-targeted, n = 28) for hematologic malignancies. Longitudinal samples were collected before and up to 1 year after CARTx. All individuals were in sustained remission. We identified 4 participants with anti-HLA antibodies before CD19-CARTx. Despite B cell depletion, anti-HLA antibodies and calculated panel reactive antibody scores were stable for 1 year after CD19-CARTx. Only 1 BCMA-CARTx recipient had pre-CARTx low-level anti-HLA antibodies, with no follow-up samples available. These data implicate CD19neg long-lived plasma cells as an important source for anti-HLA antibodies, a model supported by infrequent HLA sensitization in BCMA-CARTx subjects receiving previous plasma cell-targeted therapies. Thus, plasma cell-targeted therapies may be more effective against HLA antibodies, thereby enabling improved access to organ transplantation and rejection management.

A novel polymer-conjugated human IL-15 improves efficacy of CD19-targeted CAR T-cell immunotherapy.

Chimeric antigen receptor (CAR)-modified T-cell therapies targeting CD19 represent a new treatment option for patients with relapsed/refractory (R/R) B-cell malignancies. However, CAR T-cell therapy fails to elicit durable responses in a significant fraction of patients. Limited in vivo proliferation and survival of infused CAR T cells are key causes of failure. In a phase 1/2 clinical trial of CD19 CAR T cells for B-cell malignancies (#NCT01865617), low serum interleukin 15 (IL-15) concentration after CAR T-cell infusion was associated with inferior CAR T-cell kinetics. IL-15 supports T-cell proliferation and survival, and therefore, supplementation with IL-15 may enhance CAR T-cell therapy. However, the clinical use of native IL-15 is challenging because of its unfavorable pharmacokinetic (PK) and toxicity. NKTR-255 is a polymer-conjugated IL-15 that engages the entire IL-15 receptor complex (IL-15Rα/IL-2Rßγ) and exhibits reduced clearance, providing sustained pharmacodynamic (PD) responses. We investigated the PK and immune cell PDs in nonhuman primates treated with NKTR-255 and found that NKTR-255 enhanced the in vivo proliferation of T cells and natural killer cells. In vitro, NKTR-255 induced dose-dependent proliferation and accumulation of human CD19 CAR T cells, especially at low target cell abundance. In vivo studies in lymphoma-bearing immunodeficient mice demonstrated enhanced antitumor efficacy of human CD19 CAR T cells. In contrast to mice treated with CAR T cells alone, those that received CAR T cells and NKTR-255 had markedly higher CAR T-cell counts in the blood and marrow that were sustained after tumor clearance, without evidence of persistent proliferation or ongoing activation/exhaustion as assessed by Ki-67 and inhibitory receptor coexpression. These data support an ongoing phase 1 clinical trial of combined therapy with CD19 CAR T cells and NKTR-255 for R/R B-cell malignancies.

Unveiling the autoreactome: Proteome-wide immunological fingerprints reveal the promise of plasma cell depleting therapy.

The prevalence and burden of autoimmune and autoantibody mediated disease is increasing worldwide, yet most disease etiologies remain unclear. Despite numerous new targeted immunomodulatory therapies, comprehensive approaches to apply and evaluate the effects of these treatments longitudinally are lacking. Here, we leverage advances in programmable-phage immunoprecipitation (PhIP-Seq) methodology to explore the modulation, or lack thereof, of proteome-wide autoantibody profiles in both health and disease. We demonstrate that each individual, regardless of disease state, possesses a distinct set of autoreactivities constituting a unique immunological fingerprint, or "autoreactome", that is remarkably stable over years. In addition to uncovering important new biology, the autoreactome can be used to better evaluate the relative effectiveness of various therapies in altering autoantibody repertoires. We find that therapies targeting B-Cell Maturation Antigen (BCMA) profoundly alter an individual's autoreactome, while anti-CD19 and CD-20 therapies have minimal effects, strongly suggesting a rationale for BCMA or other plasma cell targeted therapies in autoantibody mediated diseases.

Predictors of response to axicabtagene-ciloleucel CAR T cells in aggressive B cell lymphomas: A real-world study.

Chimeric antigen receptor T-cell (CAR T) therapy has shown promising efficacy in relapsed and refractory diffuse large B cell lymphoma (DLBCL). While most patients undergo CAR T infusion with active disease, the impact of some clinical variables, such as responsiveness to the pre-CAR T chemotherapy on the response to CAR T, is unknown. In this single-institution study, we studied the impact of several pre-CAR T variables on the post-CAR outcomes. Sixty patients underwent apheresis for axicabtagene-ciloleucel (axi-cel) and 42 of them (70.0%) had primary refractory disease. Bridging therapy between apheresis and lymphodepletion was given in 34 patients (56.7%). After axi-cel, the overall response rate was 63.3%. Responsiveness to the immediate pre-CAR T therapy did not show a significant association with response to axi-cel, progression-free (PFS) or overall (OS) survival. Multivariable analysis determined that bulky disease before lymphodepletion was independently associated with inferior outcomes, and patients that presented with high-burden disease unresponsive to immediate pre-CAR T therapy had a dismal outcome. This data supports proceeding with treatment in CAR T candidates regardless of their response to immediate pre-CAR T therapy. Interim therapeutic interventions should be considered in patients who have known risk factors for poor outcomes (bulky disease, high LDH).

Predictors of cytopenias after treatment with axicabtagene ciloleucel in patients with large B-cell lymphoma.

Cytopenias are important but less studied adverse events following chimeric antigen receptor-engineered T cell (CAR-T) therapy. In our analysis of patients with large cell lymphoma who received axicabtagene ciloleucel (axi-cel), we sought to determine the rate and risk factors of clinically significant short term cytopenias defined as grade ≥3 neutropenia, anemia, or thrombocytopenia, or treatment with growth factors or blood product transfusions between days 20-30 after axi-cel. Fifty-three pts received axi-cel during the study period and severe cytopenias were observed in 32 (60%) pts. Significant cytopenias were more common in non-responders (stable or progressive disease) vs. responders (partial or complete response) (100% vs. 70%; p = .01). In the multivariable model, platelet transfusion within a month before leukapheresis, number of red blood cell and platelet transfusions between leukapheresis to lymphodepletion, pre-lymphodepletion absolute neurophil count, pre-lymphodepletion lactate dehydrogenase, and number of dexamethasone treatments after CAR-T were significantly associated with severe cytopenias after axi-cel.

Impact of CD19 CAR T-cell product type on outcomes in relapsed or refractory aggressive B-NHL.

CD19-targeted chimeric antigen receptor-engineered (CD19 CAR) T cells are novel therapies showing great promise for patients with relapsed or refractory (R/R) aggressive B-cell non-Hodgkin lymphoma (B-NHL). Single-arm studies showed significant variations in outcomes across distinct CD19 CAR T-cell products. To estimate the independent impact of the CAR T-cell product type on outcomes, we retrospectively analyzed data from 129 patients with R/R aggressive B-NHL treated with cyclophosphamide and fludarabine lymphodepletion followed by either a commercially available CD19 CAR T-cell therapy (axicabtagene ciloleucel [axicel] or tisagenlecleucel [tisacel]), or the investigational product JCAR014 on a phase 1/2 clinical trial (NCT01865617). After adjustment for age, hematopoietic cell transplantation-specific comorbidity index, lactate dehydrogenase (LDH), largest lesion diameter, and absolute lymphocyte count (ALC), CAR T-cell product type remained associated with outcomes in multivariable models. JCAR014 was independently associated with lower cytokine release syndrome (CRS) severity compared with axicel (adjusted odds ratio [aOR], 0.19; 95% confidence interval [CI]; 0.08-0.46), with a trend toward lower CRS severity with tisacel compared with axicel (aOR, 0.47; 95% CI, 0.21-1.06; P = .07). Tisacel (aOR, 0.17; 95% CI, 0.06-0.48) and JCAR014 (aOR, 0.17; 95% CI, 0.06-0.47) were both associated with lower immune effector cell-associated neurotoxicity syndrome severity compared with axicel. Lower odds of complete response (CR) were predicted with tisacel and JCAR014 compared with axicel. Although sensitivity analyses using either positron emission tomography- or computed tomography-based response criteria also suggested higher efficacy of axicel over JCAR014, the impact of tisacel vs axicel became undetermined. Higher preleukapheresis LDH, largest lesion diameter, and lower ALC were independently associated with lower odds of CR. We conclude that CD19 CAR T-cell product type independently impacts toxicity and efficacy in R/R aggressive B-NHL patients.

Autologous transplant vs chimeric antigen receptor T-cell therapy for relapsed DLBCL in partial remission.

The relative efficacy of autologous hematopoietic cell transplant (auto-HCT) vs chimeric antigen receptor T-cell (CAR-T) therapy in patients with diffuse large B-cell lymphoma (DLBCL) who achieve a partial remission (PR) after salvage chemotherapy is not known. Using the Center for International Blood & Marrow Transplant Research registry database, we identified adult patients with DLBCL who received either an auto-HCT (2013-2019) or CAR-T treatment with axicabtagene ciloleucel (2018-2019) while in a PR by computed tomography or positron emission tomography scan. We compared the clinical outcomes between the 2 cohorts using univariable and multivariable regression models after adjustment for relevant baseline and clinical factors. In the univariable analysis, the 2-year progression-free survival (52% vs 42%; P = .1) and the rate of 100-day nonrelapse mortality (4% vs 2%; P = .3) were not different between the 2 cohorts, but consolidation with auto-HCT was associated with a lower rate of relapse/progression (40% vs 53%; P = .05) and a superior overall survival (OS) (69% vs 47%; P = .004) at 2 years. In the multivariable regression analysis, treatment with auto-HCT was associated with a significantly lower risk of relapse/progression rate (hazard ratio = 1.49; P = .01) and a superior OS (hazard ratio = 1.63; P = .008). In patients with DLBCL in a PR after salvage therapy, treatment with auto-HCT was associated with a lower incidence of relapse and a superior OS compared with CAR-T. These data support the role of auto-HCT as the standard of care in transplant-eligible patients with relapsed DLBCL in PR after salvage therapy.

Severe cytokine release syndrome is associated with hematologic toxicity following CD19 CAR T-cell therapy.

CD19-targeted chimeric antigen receptor (CAR) T-cell therapy has demonstrated remarkable efficacy in patients with relapsed/refractory B-cell malignancies; however, it is associated with toxicities including cytokine release syndrome (CRS), neurotoxicity, and impaired hematopoietic recovery. The latter is associated with high-grade cytopenias requiring extended growth factor or transfusional support, potentially leading to additional complications such as infection or hemorrhage. To date, the factors independently associated with hematologic toxicity have not been well characterized. To address this deficit, we retrospectively analyzed 173 patients who received defined-composition CD19 CAR T-cell therapy in a phase 1/2 clinical trial (https://clinicaltrials.gov; NCT01865617), with primary end points of absolute neutrophil count and platelet count at day-28 after CAR T-cell infusion. We observed cumulative incidences of neutrophil and platelet recovery of 81% and 75%, respectively, at 28 days after infusion. Hematologic toxicity was noted in a significant subset of patients, with persistent neutropenia in 9% and thrombocytopenia in 14% at last follow-up. Using debiased least absolute shrinkage selector and operator regression analysis for high-dimensional modeling and considering patient-, disease-, and treatment-related variables, we identified increased CRS severity as an independent predictor for decreased platelet count and lower prelymphodepletion platelet count as an independent predictor of both decreased neutrophil and platelet counts after CD19 CAR T-cell infusion. Furthermore, multivariable models including CRS-related cytokines identified associations between higher peak serum concentrations of interleukin-6 and lower day-28 cell counts; in contrast, higher serum concentrations of transforming growth factor-ß1 were associated with higher counts. Our findings suggest that patient selection and improved CRS management may improve hematopoietic recovery after CD19 CAR T-cell therapy.