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While chimeric antigen receptor T-cell (CAR-T) therapy has revolutionized the treatment of B-cell malignancies, many patients relapse and therefore strategies to improve antitumor immunity are needed. We previously designed a novel autologous bispecific CAR targeting CD19 and CD22 (CAR19-22), which was well tolerated and associated with high response rates but relapse was common. Interleukin-15 (IL15) induces proliferation of diverse immune cells and can augment lymphocyte trafficking. Here, we report the results of a phase 1 clinical trial of the first combination of a novel recombinant polymer-conjugated IL15 receptor agonist (NKTR-255), with CAR19-22, in adults with relapsed / refractory B-cell acute lymphoblastic leukemia. Eleven patients were enrolled, nine of whom successfully received CAR19-22 followed by NKTR-255. There were no dose limiting toxicities, with transient fever and myelosuppression as the most common possibly related toxicities. We observed favorable efficacy with eight out of nine patients (89%) achieving measurable residual disease negative remission. At 12 months, progression-free survival for NKTR-255 was double that of historical controls (67% vs 38%). We performed correlative analyses to investigate the effects of IL15 receptor agonism. Cytokine profiling showed significant increases in IL15 and the chemokines CXCL9 and CXCL10. The increase in chemokines was associated with decreases in absolute lymphocyte counts and CD8+ CAR T-cells in blood and ten-fold increases in CSF CAR-T cells, suggesting lymphocyte trafficking to tissue. Combining NKTR-255 with CAR19-22 was safe, feasible and associated with high rates of durable responses (NCT03233854).
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H3K27M-mutant diffuse midline gliomas (DMGs) express high levels of the GD2 disialoganglioside and chimeric antigen receptor modified T-cells targeting GD2 (GD2-CART) eradicate DMGs in preclinical models. Arm A of the Phase I trial NCT04196413 administered one IV dose of autologous GD2-CART to patients with H3K27M-mutant pontine (DIPG) or spinal (sDMG) diffuse midline glioma at two dose levels (DL1=1e6/kg; DL2=3e6/kg) following lymphodepleting (LD) chemotherapy. Patients with clinical or imaging benefit were eligible for subsequent intracerebroventricular (ICV) GD2-CART infusions (10-30e6 GD2-CART). Primary objectives were manufacturing feasibility, tolerability, and identification of a maximally tolerated dose of IV GD2-CART. Secondary objectives included preliminary assessments of benefit. Thirteen patients enrolled and 11 received IV GD2-CART on study [n=3 DL1(3 DIPG); n=8 DL2(6 DIPG/2 sDMG). GD2-CART manufacturing was successful for all patients. No dose-limiting toxicities (DLTs) occurred on DL1, but three patients experienced DLT on DL2 due to grade 4 cytokine release syndrome (CRS). Nine patients received ICV infusions, which were not associated with DLTs. All patients exhibited tumor inflammation-associated neurotoxicity (TIAN). Four patients demonstrated major volumetric tumor reductions (52%, 54%, 91% and 100%). One patient exhibited a complete response ongoing for >30 months since enrollment. Eight patients demonstrated neurological benefit based upon a protocol-directed Clinical Improvement Score. Sequential IV followed by ICV GD2-CART induced tumor regressions and neurological improvements in patients with DIPG and sDMG. DL1 was established as the maximally tolerated IV GD2-CART dose. Neurotoxicity was safely managed with intensive monitoring and close adherence to a management algorithm.
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BACKGROUND: Fludarabine in combination with cyclophosphamide (FC) is the standard lymphodepletion regimen for CAR T-cell therapy (CAR T). A national fludarabine shortage in 2022 necessitated the exploration of alternative regimens with many centers employing single-agent bendamustine as lymphodepletion despite a lack of clinical safety and efficacy data. To fill this gap in the literature, we evaluated the safety, efficacy, and expansion kinetics of bendamustine as lymphodepletion prior to axicabtagene ciloleucel (axi-cel) therapy. METHODS: 84 consecutive patients with relapsed or refractory large B-cell lymphoma treated with axi-cel and managed with a uniform toxicity management plan at Stanford University were studied. 27 patients received alternative lymphodepletion with bendamustine while 57 received FC. RESULTS: Best complete response rates were similar (73.7% for FC and 74% for bendamustine, p=0.28) and there was no significant difference in 12-month progression-free survival or overall survival estimates (p=0.17 and p=0.62, respectively). The frequency of high-grade cytokine release syndrome and immune effector cell-associated neurotoxicity syndrome was similar in both the cohorts. Bendamustine cohort experienced lower proportions of hematological toxicities and antibiotic use for neutropenic fever. Immune reconstitution, as measured by quantitative assessment of cellular immunity, was better in bendamustine cohort as compared with FC cohort. CAR T expansion as measured by peak expansion and area under the curve for expansion was comparable between cohorts. CONCLUSIONS: Bendamustine is a safe and effective alternative lymphodepletion conditioning for axi-cel with lower early hematological toxicity and favorable immune reconstitution.
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Clorhidrato de Bendamustina , Productos Biológicos , Linfoma de Células B Grandes Difuso , Humanos , Clorhidrato de Bendamustina/uso terapéutico , Clorhidrato de Bendamustina/administración & dosificación , Masculino , Femenino , Persona de Mediana Edad , Anciano , Linfoma de Células B Grandes Difuso/tratamiento farmacológico , Productos Biológicos/uso terapéutico , Productos Biológicos/efectos adversos , Adulto , Inmunoterapia Adoptiva/métodos , Inmunoterapia Adoptiva/efectos adversos , Antígenos CD19/inmunología , Antígenos CD19/uso terapéuticoRESUMEN
BACKGROUND: Outcomes are poor for patients with large B-cell lymphoma who relapse after CD19-directed chimeric antigen receptor (CAR) T-cell therapy (CAR19). CD22 is a nearly universally expressed B-cell surface antigen and the efficacy of a CD22-directed CAR T-cell therapy (CAR22) in large B-cell lymphoma is unknown, which was what we aimed to examine in this study. METHODS: In this single centre, open-label, dose-escalation phase 1 trial, we intravenously administered CAR22 at two dose levels (1 million and 3 million CAR22-positive T cells per kg of bodyweight) to adult patients (aged ≥18 years) who relapsed after CAR19 or had CD19-negative large B-cell lymphoma. The primary endpoints were manufacturing feasibility, safety measured by the incidence and severity of adverse events and dose-limiting toxicities, and identification of the maximum tolerated dose (ie, the recommended phase 2 dose). This study is registered with ClinicalTrials.gov (NCT04088890) and is active, but closed for enrolment. FINDINGS: From Oct 17, 2019, to Oct 19, 2022, a total of 41 patients were assessed for eligibility; however, one patient withdrew. 40 patients underwent leukapheresis and 38 (95%) had CAR T-cell products manufactured successfully. The median age was 65 years (range 25-84), 17 (45%) were women, 32 (84%) had elevated pretreatment lactate dehydrogenase, 11 (29%) had refractory disease to all previous therapies, and patients had received a median of four lines of previous therapy (range 3-8). Of the 38 patients treated, 37 (97%) had relapsed after previous CAR19. The identified maximum tolerated dose was 1 million CAR T cells per kg. Of 29 patients who received the maximum tolerated dose, no patients developed a dose-limiting toxicity or grade 3 or higher cytokine release syndrome, immune effector cell-associated neurotoxicity syndrome, or immune effector cell-associated haemophagocytic lymphohistiocytosis-like syndrome. INTERPRETATION: This trial identifies CD22 as an immunotherapeutic target in large B-cell lymphoma and demonstrates the durable clinical activity of CAR22 in patients with disease progression after CAR19 therapy. Although these findings are promising, it is essential to recognise that this is a phase 1 dose-finding study. Further investigations are warranted to establish the long-term efficacy and to delineate the patient subgroups that will derive the most benefit from this therapeutic approach. FUNDING: National Cancer Institute, National Institutes of Health, Stanford Cancer Institute, Leukemia & Lymphoma Society, Parker Institute for Cancer Immunotherapy, Lymph & Co, and the European Hematology Association.
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Antígenos CD19 , Inmunoterapia Adoptiva , Lectina 2 Similar a Ig de Unión al Ácido Siálico , Adulto , Anciano , Femenino , Humanos , Masculino , Persona de Mediana Edad , Antígenos CD19/inmunología , Inmunoterapia Adoptiva/métodos , Inmunoterapia Adoptiva/efectos adversos , Linfoma de Células B Grandes Difuso/terapia , Linfoma de Células B Grandes Difuso/inmunología , Dosis Máxima Tolerada , Receptores Quiméricos de Antígenos/inmunología , Lectina 2 Similar a Ig de Unión al Ácido Siálico/inmunología , Anciano de 80 o más AñosRESUMEN
BACKGROUND: The risk of second tumors after chimeric antigen receptor (CAR) T-cell therapy, especially the risk of T-cell neoplasms related to viral vector integration, is an emerging concern. METHODS: We reviewed our clinical experience with adoptive cellular CAR T-cell therapy at our institution since 2016 and ascertained the occurrence of second tumors. In one case of secondary T-cell lymphoma, a broad array of molecular, genetic, and cellular techniques were used to interrogate the tumor, the CAR T cells, and the normal hematopoietic cells in the patient. RESULTS: A total of 724 patients who had received T-cell therapies at our center were included in the study. A lethal T-cell lymphoma was identified in a patient who had received axicabtagene ciloleucel therapy for diffuse large B-cell lymphoma, and both lymphomas were deeply profiled. Each lymphoma had molecularly distinct immunophenotypes and genomic profiles, but both were positive for Epstein-Barr virus and were associated with DNMT3A and TET2 mutant clonal hematopoiesis. No evidence of oncogenic retroviral integration was found with the use of multiple techniques. CONCLUSIONS: Our results highlight the rarity of second tumors and provide a framework for defining clonal relationships and viral vector monitoring. (Funded by the National Cancer Institute and others.).
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Antineoplásicos Inmunológicos , Inmunoterapia Adoptiva , Linfoma de Células B Grandes Difuso , Linfoma de Células T , Neoplasias Primarias Secundarias , Receptores Quiméricos de Antígenos , Femenino , Humanos , Persona de Mediana Edad , Productos Biológicos/efectos adversos , Productos Biológicos/uso terapéutico , Hematopoyesis Clonal , Herpesvirus Humano 4/inmunología , Herpesvirus Humano 4/genética , Inmunoterapia Adoptiva/efectos adversos , Linfoma de Células B Grandes Difuso/genética , Linfoma de Células B Grandes Difuso/inmunología , Linfoma de Células B Grandes Difuso/terapia , Linfoma de Células T/etiología , Linfoma de Células T/genética , Linfoma de Células T/inmunología , Linfoma de Células T/terapia , Neoplasias Primarias Secundarias/genética , Neoplasias Primarias Secundarias/etiología , Receptores Quiméricos de Antígenos/inmunología , Receptores Quiméricos de Antígenos/uso terapéutico , Antineoplásicos Inmunológicos/efectos adversos , Antineoplásicos Inmunológicos/uso terapéutico , Integración ViralRESUMEN
Adoptively transferred T cells and agents designed to block the CD47-SIRPα axis are promising cancer therapeutics that activate distinct arms of the immune system1,2. Here we administered anti-CD47 antibodies in combination with adoptively transferred T cells with the goal of enhancing antitumour efficacy but observed abrogated therapeutic benefit due to rapid macrophage-mediated clearance of T cells expressing chimeric antigen receptors (CARs) or engineered T cell receptors. Anti-CD47-antibody-mediated CAR T cell clearance was potent and rapid enough to serve as an effective safety switch. To overcome this challenge, we engineered the CD47 variant CD47(Q31P) (47E), which engages SIRPα and provides a 'don't eat me' signal that is not blocked by anti-CD47 antibodies. TCR or CAR T cells expressing 47E are resistant to clearance by macrophages after treatment with anti-CD47 antibodies, and mediate substantial, sustained macrophage recruitment to the tumour microenvironment. Although many of the recruited macrophages manifested an M2-like profile3, the combined therapy synergistically enhanced antitumour efficacy. Our study identifies macrophages as major regulators of T cell persistence and illustrates the fundamental challenge of combining T-cell-directed therapeutics with those designed to activate macrophages. It delivers a therapeutic approach that is capable of simultaneously harnessing the antitumour effects of T cells and macrophages, offering enhanced potency against solid tumours.
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Antígeno CD47 , Inmunoterapia Adoptiva , Neoplasias , Linfocitos T , Animales , Femenino , Humanos , Masculino , Ratones , Antígenos de Diferenciación/inmunología , Antígenos de Diferenciación/metabolismo , Antígeno CD47/genética , Antígeno CD47/inmunología , Antígeno CD47/metabolismo , Línea Celular Tumoral , Inmunoterapia Adoptiva/métodos , Macrófagos/citología , Macrófagos/inmunología , Neoplasias/inmunología , Neoplasias/metabolismo , Neoplasias/terapia , Receptores de Antígenos de Linfocitos T/genética , Receptores de Antígenos de Linfocitos T/inmunología , Receptores de Antígenos de Linfocitos T/metabolismo , Receptores Quiméricos de Antígenos/genética , Receptores Quiméricos de Antígenos/inmunología , Receptores Quiméricos de Antígenos/metabolismo , Receptores Inmunológicos/inmunología , Receptores Inmunológicos/metabolismo , Linfocitos T/inmunología , Linfocitos T/metabolismo , Linfocitos T/trasplante , Microambiente Tumoral/inmunología , Anticuerpos/inmunología , Anticuerpos/uso terapéutico , Activación de MacrófagosRESUMEN
Chronic graft-versus-host disease (cGVHD) remains a significant source of morbidity and mortality following allogeneic hematopoietic cell transplantation (HCT). Post-transplant, prophylactic rituximab has successfully decreased cGHVD rates in clinical trials, but the durability of this strategy is uncertain. The long-terms effect of post-HCT B cell depletion on immune reconstitution, B cell function, and infectious complications are also unknown. In this study, we provide 10 yr follow-up and correlative analyses on patients given post-HCT, prophylactic rituximab. The objective of the study is to examine the durability of cGVHD protection as well as the long-term effect of rituximab prophylaxis on protective immune reconstitution, B cell function, and alloantibody formation. We analyzed 35 patients given prophylactic rituximab on phase II clinical trial. Clinical outcomes included cGVHD development, relapse and survival outcomes, and infectious outcomes. Correlative analyses included B cell subset analysis, development of antibodies to infectious antigens, and, for male patients receiving female donor grafts, development of antibodies to HY antigens. To further investigate the effect of rituximab on immune reconstitution and function, we also analyzed 43 similarly transplanted patients who did not receive post- or peri-HCT rituximab as a comparator group. For patients who received rituximab, the 8-yr cumulative incidence of cGHVD and freedom from immunosuppression were 20.0% and 76.2%, respectively. Importantly, no late incidences of cGVHD developed beyond 14 mo post-HCT. Relative to patients who did not receive rituximab, post-HCT rituximab was associated with increased B cell aplasia at 1 yr post-HCT (42.9% versus 11% of patients, P = .037); by 3 yr post-HCT, this aplasia resolved. Patients who received rituximab also had a significantly lower proportion of IgD+/CD38+ transitional B cells at 3 yr post-HCT (78.8% versus 89.9%, P = .039); at 10 yr post-HCT, this percentage remained markedly decreased at 50.7%. Rituximab prophylaxis altered B cell function. In male patients receiving female donor grafts, fewer patients developed HY antibodies at 3 yr post-HCT (20% versus 78%, P = .04). At 10 yr post-HCT, HY antibody production remained decreased at 33%. Rituximab prophylaxis was also associated with significantly lower antibody response to tetanus and EBV infectious antigens as well as lower IgG levels. Despite these changes, post-HCT was not associated with increased infections, although patients who received rituximab required intravenous immunoglobulin (IVIG) supplementation more frequently than those who did not (62.9% versus 32.6% of patients, P = .01). Prior data on the efficacy and feasibility of rituximab prophylaxis are durable, with persistent reduction in cGVHD. Rituximab prophylaxis also results in lasting B cell immunologic changes, with altered B cell subset composition and decreased alloantibody formation. Associated infectious risks were not increased, perhaps mitigated by high IVIG use.
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Linfocitos B , Enfermedad Injerto contra Huésped , Trasplante de Células Madre Hematopoyéticas , Reconstitución Inmune , Rituximab , Humanos , Rituximab/uso terapéutico , Femenino , Masculino , Trasplante de Células Madre Hematopoyéticas/efectos adversos , Linfocitos B/inmunología , Linfocitos B/efectos de los fármacos , Persona de Mediana Edad , Enfermedad Injerto contra Huésped/prevención & control , Enfermedad Injerto contra Huésped/inmunología , Adulto , Trasplante Homólogo/efectos adversos , Resultado del Tratamiento , Anciano , Estudios de SeguimientoRESUMEN
ABSTRACT: Chimeric antigen receptor (CAR) T cells directed against CD19 (CAR19) are a revolutionary treatment for B-cell lymphomas (BCLs). CAR19 cell expansion is necessary for CAR19 function but is also associated with toxicity. To define the impact of CAR19 expansion on patient outcomes, we prospectively followed a cohort of 236 patients treated with CAR19 (brexucabtagene autoleucel or axicabtagene ciloleucel) for mantle cell lymphoma (MCL), follicular lymphoma, and large BCL (LBCL) over the course of 5 years and obtained CAR19 expansion data using peripheral blood immunophenotyping for 188 of these patients. CAR19 expansion was higher in patients with MCL than other lymphoma histologic subtypes. Notably, patients with MCL had increased toxicity and required fourfold higher cumulative steroid doses than patients with LBCL. CAR19 expansion was associated with the development of cytokine release syndrome, immune effector cell-associated neurotoxicity syndrome, and the requirement for granulocyte colony-stimulating factor 14 days after infusion. Younger patients and those with elevated lactate dehydrogenase (LDH) had significantly higher CAR19 expansion. In general, no association between CAR19 expansion and LBCL treatment response was observed. However, when controlling for tumor burden, we found that lower CAR19 expansion in conjunction with low LDH was associated with improved outcomes in LBCL. In sum, this study finds CAR19 expansion principally associates with CAR-related toxicity. Additionally, CAR19 expansion as measured by peripheral blood immunophenotyping may be dispensable to favorable outcomes in LBCL.
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Antígenos CD19 , Inmunofenotipificación , Inmunoterapia Adoptiva , Humanos , Masculino , Antígenos CD19/inmunología , Persona de Mediana Edad , Inmunoterapia Adoptiva/efectos adversos , Inmunoterapia Adoptiva/métodos , Femenino , Anciano , Receptores Quiméricos de Antígenos/inmunología , Adulto , Linfoma de Células del Manto/inmunología , Linfoma de Células del Manto/sangre , Anciano de 80 o más Años , Productos BiológicosRESUMEN
Chimeric antigen receptor (CAR) T cells targeting CD22 (CD22-CAR) provide a therapeutic option for patients with CD22+ malignancies with progression after CD19-directed therapies. Using on-site, automated, closed-loop manufacturing, we conducted parallel Phase 1b clinical trials investigating a humanized CD22-CAR with 41BB costimulatory domain in children and adults with heavily treated, relapsed/refractory (r/r) B-ALL. Of 19 patients enrolled, 18 had successful CD22-CAR manufacturing, and 16 patients were infused. High grade (3-4) cytokine release syndrome (CRS) and immune effector-cell-associated neurotoxicity syndrome (ICANS) each occurred in only one patient; however, three patients experienced immune-effector-cell-associated hemophagocytic lymphohistiocytosis-like syndrome (IEC-HS). Twelve of 16 patients (75%) achieved CR with an overall 56% MRD-negative CR rate. Duration of response was overall limited (median 77 days), and CD22 expression was downregulated in 4/12 (33%) available samples at relapse. In summary, we demonstrate that closed-loop manufacturing of CD22-CAR T cells is feasible and is associated with a favorable safety profile and high CR rates in pediatric and adult r/r B-ALL, a cohort with limited CD22-CAR reporting.
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Inmunoterapia Adoptiva , Leucemia-Linfoma Linfoblástico de Células Precursoras B , Receptores Quiméricos de Antígenos , Lectina 2 Similar a Ig de Unión al Ácido Siálico , Humanos , Lectina 2 Similar a Ig de Unión al Ácido Siálico/inmunología , Niño , Adulto , Femenino , Masculino , Adolescente , Inmunoterapia Adoptiva/métodos , Inmunoterapia Adoptiva/efectos adversos , Adulto Joven , Receptores Quiméricos de Antígenos/inmunología , Leucemia-Linfoma Linfoblástico de Células Precursoras B/terapia , Leucemia-Linfoma Linfoblástico de Células Precursoras B/inmunología , Preescolar , Persona de Mediana Edad , Linfocitos T/inmunología , Linfocitos T/metabolismoRESUMEN
Chimeric antigen receptor T cells (CAR-Ts) have remarkable efficacy in liquid tumors, but limited responses in solid tumors. We conducted a Phase I trial (NCT02107963) of GD2 CAR-Ts (GD2-CAR.OX40.28.z.iC9), demonstrating feasibility and safety of administration in children and young adults with osteosarcoma and neuroblastoma. Since CAR-T efficacy requires adequate CAR-T expansion, patients were grouped into good or poor expanders across dose levels. Patient samples were evaluated by multi-dimensional proteomic, transcriptomic, and epigenetic analyses. T cell assessments identified naive T cells in pre-treatment apheresis associated with good expansion, and exhausted T cells in CAR-T products with poor expansion. Myeloid cell assessment identified CXCR3+ monocytes in pre-treatment apheresis associated with good expansion. Longitudinal analysis of post-treatment samples identified increased CXCR3- classical monocytes in all groups as CAR-T numbers waned. Together, our data uncover mediators of CAR-T biology and correlates of expansion that could be utilized to advance immunotherapies for solid tumor patients.
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Neuroblastoma , Receptores Quiméricos de Antígenos , Niño , Adulto Joven , Humanos , Receptores Quiméricos de Antígenos/genética , Receptores de Antígenos de Linfocitos T/genética , Proteómica , Inmunoterapia Adoptiva/efectos adversos , Inmunoterapia Adoptiva/métodos , Linfocitos T , Neuroblastoma/patología , Tratamiento Basado en Trasplante de Células y TejidosRESUMEN
The central role of target antigen density on chimeric antigen receptor T cell potency highlights the need for accurate measurement of antigen levels on clinical tumor samples. Here, we present a protocol for quantifying antigen density for six cell-surface antigens on neuroblastoma cells metastatic to bone marrow. We describe steps for patient sample acquisition, flow cytometry panel development, instrument setup, and compensation and detail procedures for running clinical samples and data analysis. For complete details on the use and execution of this protocol, please refer to Heitzeneder et al. (2022).1.
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Neoplasias de la Médula Ósea , Neuroblastoma , Humanos , Médula Ósea , Citometría de Flujo , Inmunoterapia , Neuroblastoma/terapia , Neoplasias de la Médula Ósea/terapiaRESUMEN
Most relapsed/refractory large B cell lymphoma (r/rLBCL) patients receiving anti-CD19 chimeric antigen receptor (CAR19) T cells relapse. To characterize determinants of resistance, we profiled over 700 longitudinal specimens from two independent cohorts (n = 65 and n = 73) of r/rLBCL patients treated with axicabtagene ciloleucel. A method for simultaneous profiling of circulating tumor DNA (ctDNA), cell-free CAR19 (cfCAR19) retroviral fragments, and cell-free T cell receptor rearrangements (cfTCR) enabled integration of tumor and both engineered and non-engineered T cell effector-mediated factors for assessing treatment failure and predicting outcomes. Alterations in multiple classes of genes are associated with resistance, including B cell identity (PAX5 and IRF8), immune checkpoints (CD274), and those affecting the microenvironment (TMEM30A). Somatic tumor alterations affect CAR19 therapy at multiple levels, including CAR19 T cell expansion, persistence, and tumor microenvironment. Further, CAR19 T cells play a reciprocal role in shaping tumor genotype and phenotype. We envision these findings will facilitate improved chimeric antigen receptor (CAR) T cells and personalized therapeutic approaches.
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Linfoma de Células B Grandes Difuso , Receptores Quiméricos de Antígenos , Humanos , Receptores Quiméricos de Antígenos/genética , Recurrencia Local de Neoplasia/tratamiento farmacológico , Linfoma de Células B Grandes Difuso/terapia , Linfoma de Células B Grandes Difuso/tratamiento farmacológico , Inmunoterapia Adoptiva/métodos , Linfocitos T , Antígenos CD19/genética , Microambiente TumoralRESUMEN
Approximately 60% of patients with large B cell lymphoma treated with chimeric antigen receptor (CAR) T cell therapies targeting CD19 experience disease progression, and neurotoxicity remains a challenge. Biomarkers associated with resistance and toxicity are limited. In this study, single-cell proteomic profiling of circulating CAR T cells in 32 patients treated with CD19-CAR identified that CD4+Helios+ CAR T cells on day 7 after infusion are associated with progressive disease and less severe neurotoxicity. Deep profiling demonstrated that this population is non-clonal and manifests hallmark features of T regulatory (TReg) cells. Validation cohort analysis upheld the link between higher CAR TReg cells with clinical progression and less severe neurotoxicity. A model combining expansion of this subset with lactate dehydrogenase levels, as a surrogate for tumor burden, was superior for predicting durable clinical response compared to models relying on each feature alone. These data credential CAR TReg cell expansion as a novel biomarker of response and toxicity after CAR T cell therapy and raise the prospect that this subset may regulate CAR T cell responses in humans.
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Síndromes de Neurotoxicidad , Receptores Quiméricos de Antígenos , Antígenos CD19 , Humanos , Inmunoterapia Adoptiva/efectos adversos , Inmunoterapia Adoptiva/métodos , Lactato Deshidrogenasas , Síndromes de Neurotoxicidad/etiología , Proteómica , Receptores de Antígenos de Linfocitos TRESUMEN
Idecabtagene vicleucel (ide-cel) was FDA-approved in March 2021 for the treatment of relapsed/refractory multiple myeloma after 4 lines of therapy. On the KarMMa trial, grade ≥ 3 cytopenias and infections were common. We sought to characterize cytopenias and infections within 100 days after ide-cel in the standard-of-care (SOC) setting. This multi-center retrospective study included 52 patients who received SOC ide-cel; 47 reached day-90 follow-up. Data were censored at day 100. Grade ≥ 3 cytopenia was present among 65% of patients at day 30 and 40% of patients at day 90. Granulocyte colony stimulating factor (G-CSF) was administered to 88%, packed red blood cell transfusions to 63%, platelet transfusions to 42%, thrombopoietin (TPO) agonists to 21%, intravenous immunoglobulin to 13%, and CD34+ stem cell boosts to 8%. At day 100, 19% and 13% of patients had ongoing use of TPO agonists and G-CSF, respectively. Infections occurred in 54% of patients and were grade ≥ 3 in 23%. Earlier infections in the first 30 days were typically bacterial (68%) and severe (50%). Later infections between days 31 and 100 were 50% bacterial and 42% viral; only 13% were grade ≥ 3. On univariate analysis, high pre-CAR-T marrow myeloma burden (≥ 50%), circulating plasma cells at pre-lymphodepletion (LD), and grade ≥ 3 anemia at pre-LD were associated with grade ≥ 3 cytopenia at both days 30 and 90. Longer time from last bridging treatment to LD was the only significant risk factor for infection.
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Anemia , Mieloma Múltiple , Receptores Quiméricos de Antígenos , Trombocitopenia , Humanos , Mieloma Múltiple/terapia , Estudios Retrospectivos , Nivel de Atención , Factor Estimulante de Colonias de GranulocitosRESUMEN
The increasing use of mass cytometry for analyzing clinical samples offers the possibility to perform comparative analyses across public datasets. However, challenges in batch normalization and data integration limit the comparison of datasets not intended to be analyzed together. Here, we present a data integration strategy, CytofIn, using generalized anchors to integrate mass cytometry datasets from the public domain. We show that low-variance controls, such as healthy samples and stable channels, are inherently homogeneous, robust against stimulation, and can serve as generalized anchors for batch correction. Single-cell quantification comparing mass cytometry data from 989 leukemia files pre- and post normalization with CytofIn demonstrates effective batch correction while recapitulating the gold-standard bead normalization. CytofIn integration of public cancer datasets enabled the comparison of immune features across histologies and treatments. We demonstrate the ability to integrate public datasets without necessitating identical control samples or bead standards for fast and robust analysis using CytofIn.
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Algoritmos , Conjuntos de Datos como Asunto , Citometría de Flujo/métodos , Melanoma/tratamiento farmacológico , Biología Computacional/métodos , Humanos , Inhibidores de Puntos de Control Inmunológico/farmacología , Inhibidores de Puntos de Control Inmunológico/uso terapéutico , Linfocitos Infiltrantes de Tumor/efectos de los fármacos , Linfocitos Infiltrantes de Tumor/inmunología , Melanoma/inmunología , Melanoma/patología , Neoplasias/tratamiento farmacológico , Neoplasias/inmunología , Neoplasias/patología , Análisis de la Célula Individual , Neoplasias Cutáneas/tratamiento farmacológico , Neoplasias Cutáneas/inmunología , Neoplasias Cutáneas/patologíaRESUMEN
Diffuse intrinsic pontine glioma (DIPG) and other H3K27M-mutated diffuse midline gliomas (DMGs) are universally lethal paediatric tumours of the central nervous system1. We have previously shown that the disialoganglioside GD2 is highly expressed on H3K27M-mutated glioma cells and have demonstrated promising preclinical efficacy of GD2-directed chimeric antigen receptor (CAR) T cells2, providing the rationale for a first-in-human phase I clinical trial (NCT04196413). Because CAR T cell-induced brainstem inflammation can result in obstructive hydrocephalus, increased intracranial pressure and dangerous tissue shifts, neurocritical care precautions were incorporated. Here we present the clinical experience from the first four patients with H3K27M-mutated DIPG or spinal cord DMG treated with GD2-CAR T cells at dose level 1 (1 × 106 GD2-CAR T cells per kg administered intravenously). Patients who exhibited clinical benefit were eligible for subsequent GD2-CAR T cell infusions administered intracerebroventricularly3. Toxicity was largely related to the location of the tumour and was reversible with intensive supportive care. On-target, off-tumour toxicity was not observed. Three of four patients exhibited clinical and radiographic improvement. Pro-inflammatory cytokine levels were increased in the plasma and cerebrospinal fluid. Transcriptomic analyses of 65,598 single cells from CAR T cell products and cerebrospinal fluid elucidate heterogeneity in response between participants and administration routes. These early results underscore the promise of this therapeutic approach for patients with H3K27M-mutated DIPG or spinal cord DMG.
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Astrocitoma , Neoplasias del Tronco Encefálico , Gangliósidos , Glioma , Histonas , Inmunoterapia Adoptiva , Mutación , Receptores Quiméricos de Antígenos , Astrocitoma/genética , Astrocitoma/inmunología , Astrocitoma/patología , Astrocitoma/terapia , Neoplasias del Tronco Encefálico/genética , Neoplasias del Tronco Encefálico/inmunología , Neoplasias del Tronco Encefálico/patología , Neoplasias del Tronco Encefálico/terapia , Niño , Gangliósidos/inmunología , Perfilación de la Expresión Génica , Glioma/genética , Glioma/inmunología , Glioma/patología , Glioma/terapia , Histonas/genética , Humanos , Inmunoterapia Adoptiva/métodos , Receptores Quiméricos de Antígenos/inmunología , Neoplasias de la Médula Espinal/genética , Neoplasias de la Médula Espinal/inmunología , Neoplasias de la Médula Espinal/patología , Neoplasias de la Médula Espinal/terapiaRESUMEN
Pediatric cancers often mimic fetal tissues and express proteins normally silenced postnatally that could serve as immune targets. We developed T cells expressing chimeric antigen receptors (CARs) targeting glypican-2 (GPC2), a fetal antigen expressed on neuroblastoma (NB) and several other solid tumors. CARs engineered using standard designs control NBs with transgenic GPC2 overexpression, but not those expressing clinically relevant GPC2 site density (â¼5,000 molecules/cell, range 1-6 × 103). Iterative engineering of transmembrane (TM) and co-stimulatory domains plus overexpression of c-Jun lowered the GPC2-CAR antigen density threshold, enabling potent and durable eradication of NBs expressing clinically relevant GPC2 antigen density, without toxicity. These studies highlight the critical interplay between CAR design and antigen density threshold, demonstrate potent efficacy and safety of a lead GPC2-CAR candidate suitable for clinical testing, and credential oncofetal antigens as a promising class of targets for CAR T cell therapy of solid tumors.
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Glipicanos/inmunología , Inmunoterapia Adoptiva , Neuroblastoma/tratamiento farmacológico , Receptores de Antígenos de Linfocitos T/metabolismo , Animales , Línea Celular Tumoral , Glipicanos/metabolismo , Humanos , Inmunoterapia/métodos , Neuroblastoma/patología , Receptores de Antígenos de Linfocitos T/inmunología , Receptores Quiméricos de Antígenos/inmunología , Linfocitos T/efectos de los fármacos , Linfocitos T/inmunología , Ensayos Antitumor por Modelo de Xenoinjerto/métodosRESUMEN
Alterations of the gut microbiota after allogeneic hematopoietic cell transplantation (allo-HCT) are a key factor in the development of transplant-related complications such as graft-versus-host disease (GVHD). Interventions that preserve the gut microbiome hold promise to improve HCT-associated morbidity and mortality. Murine models demonstrate that prebiotics such as fructo-oligosaccharides (FOSs) may increase gut levels of short-chain fatty acids (SCFAs) such as butyrate and consequently induce proliferation of immunomodulatory FOXP3+CD4+ regulatory T cells (Tregs), which impact GVHD risk. We conducted a pilot phase I trial to investigate the maximum tolerated dose of FOS in patients undergoing reduced-intensity allo-HCT (n = 15) compared with concurrent controls (n = 16). We administered the FOS starting at pretransplant conditioning and continuing for a total of 21 days. We characterized the gut microbiome using shotgun metagenomic sequencing, measured stool short-chain fatty acids (SCFAs) using liquid chromatography-mass spectrometry, and determined peripheral T cell concentrations using cytometry by time-of-flight. We found that FOS was safe and well-tolerated at 10 g/d without significant adverse effects in patients undergoing allo-HCT. Community-level gut microbiota composition differed significantly on the day of transplant (day 0) between patients receiving FOS and concurrent controls; however, FOS-associated alterations of the gut microbiota were not sustained after transplant. Although the impact of FOS was fleeting, transplantation itself impacted a substantial number of taxa over time. In our small pilot trial, no significant differences were observed in gut microbial metabolic pathways, stool SCFAs, or peripheral Tregs, although Tregs trended higher in those patients who received FOS. A marker of CD4+ T cell activation (namely, CTLA4+) was significantly higher in patients receiving FOS, whereas a non-significant trend existed for FOP3+CD4+ Treg cells, which were higher in those receiving FOS compared with controls. FOS is well tolerated at 10 g/d in patients undergoing reduced-intensity allo-HCT. Although the alterations in gut microbiota and peripheral immune cell composition in those receiving FOS are intriguing, additional studies are required to investigate the use of prebiotics in HCT recipients.
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Microbioma Gastrointestinal , Enfermedad Injerto contra Huésped , Trasplante de Células Madre Hematopoyéticas , Animales , Enfermedad Injerto contra Huésped/prevención & control , Humanos , Ratones , Oligosacáridos , PrebióticosRESUMEN
PURPOSE: The Cancer Immune Monitoring and Analysis Centers - Cancer Immunologic Data Commons (CIMAC-CIDC) Network is supported by the NCI to identify biomarkers of response to cancer immunotherapies across clinical trials using state-of-the-art assays. A primary platform for CIMAC-CIDC studies is cytometry by time of flight (CyTOF), performed at all CIMAC laboratories. To ensure the ability to generate comparable CyTOF data across labs, a multistep cross-site harmonization effort was undertaken. EXPERIMENTAL DESIGN: We first harmonized standard operating procedures (SOPs) across the CIMAC sites. Because of a new acquisition protocol comparing original narrow- or new wide-bore injector introduced by the vendor (Fluidigm), we also tested this protocol across sites before finalizing the harmonized SOP. We then performed cross-site assay harmonization experiments using five shared cryopreserved and one lyophilized internal control peripheral blood mononuclear cell (PBMC) with a shared lyophilized antibody cocktail consisting of 14 isotype-tagged antibodies previously validated, plus additional liquid antibodies. These reagents and samples were distributed to the CIMAC sites and the data were centrally analyzed by manual gating and automated methods (Astrolabe). RESULTS: Average coefficients of variation (CV) across sites for each cell population were reported and compared with a previous multisite CyTOF study. We reached an intersite CV of under 20% for most cell subsets, very similar to a previously published study. CONCLUSIONS: These results establish the ability to reproduce CyTOF data across sites in multicenter clinical trials, and also highlight the importance of quality control procedures, such as the use of spike-in control samples, for tracking variability in this assay.