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ABSTRACT: Relapse after CD19-directed chimeric antigen receptor (CAR)-modified T cells remains a substantial challenge. Short CAR T-cell persistence contributes to relapse risk, necessitating novel approaches to prolong durability. CAR T-cell reinfusion (CARTr) represents a potential strategy to reduce the risk of or treat relapsed disease after initial CAR T-cell infusion (CARTi). We conducted a retrospective review of reinfusion of murine (CTL019) or humanized (huCART19) anti-CD19/4-1BB CAR T cells across 3 clinical trials or commercial tisagenlecleucel for relapse prevention (peripheral B-cell recovery [BCR] or marrow hematogones ≤6 months after CARTi), minimal residual disease (MRD) or relapse, or nonresponse to CARTi. The primary endpoint was complete response (CR) at day 28 after CARTr, defined as complete remission with B-cell aplasia. Of 262 primary treatments, 81 were followed by ≥1 reinfusion (investigational CTL019, n = 44; huCART19, n = 26; tisagenlecleucel, n = 11), representing 79 patients. Of 63 reinfusions for relapse prevention, 52% achieved CR (BCR, 15/40 [38%]; hematogones, 18/23 [78%]). Lymphodepletion was associated with response to CARTr for BCR (odds ratio [OR], 33.57; P = .015) but not hematogones (OR, 0.30; P = .291). The cumulative incidence of relapse was 29% at 24 months for CR vs 61% for nonresponse to CARTr (P = .259). For MRD/relapse, CR rate to CARTr was 50% (5/10), but 0/8 for nonresponse to CARTi. Toxicity was generally mild, with the only grade ≥3 cytokine release syndrome (n = 6) or neurotoxicity (n = 1) observed in MRD/relapse treatment. Reinfusion of CTL019/tisagenlecleucel or huCART19 is safe, may reduce relapse risk in a subset of patients, and can reinduce remission in CD19+ relapse.
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Antígenos CD19 , Imunoterapia Adotiva , Leucemia-Linfoma Linfoblástico de Células Precursoras , Humanos , Criança , Antígenos CD19/imunologia , Antígenos CD19/uso terapêutico , Leucemia-Linfoma Linfoblástico de Células Precursoras/terapia , Imunoterapia Adotiva/métodos , Imunoterapia Adotiva/efeitos adversos , Pré-Escolar , Feminino , Masculino , Receptores de Antígenos Quiméricos/uso terapêutico , Adolescente , Recidiva , Estudos Retrospectivos , Lactente , Receptores de Antígenos de Linfócitos T/uso terapêutico , Resultado do Tratamento , Linfócitos T/imunologiaAssuntos
Predisposição Genética para Doença , Neoplasias Hematológicas , Humanos , Neoplasias Hematológicas/genética , Neoplasias Hematológicas/diagnóstico , Criança , Masculino , Genômica/métodos , Feminino , Adolescente , Pré-Escolar , Síndromes Neoplásicas Hereditárias/genética , Síndromes Neoplásicas Hereditárias/diagnóstico , Perfilação da Expressão Gênica , Sequenciamento de Nucleotídeos em Larga EscalaRESUMO
PURPOSE: CD19-targeted chimeric antigen receptor (CAR)-modified T cells demonstrate unprecedented responses in B-cell acute lymphoblastic leukemia (B-ALL); however, relapse remains a substantial challenge. Short CAR T-cell persistence contributes to this risk; therefore, strategies to improve persistence are needed. METHODS: We conducted a pilot clinical trial of a humanized CD19 CAR T-cell product (huCART19) in children and young adults with relapsed or refractory B-ALL (n = 72) or B-lymphoblastic lymphoma (n = 2), treated in two cohorts: with (retreatment, n = 33) or without (CAR-naive, n = 41) prior CAR exposure. Patients were monitored for toxicity, response, and persistence of huCART19. RESULTS: Seventy-four patients 1-29 years of age received huCART19. Cytokine release syndrome developed in 62 (84%) patients and was grade 4 in five (6.8%). Neurologic toxicities were reported in 29 (39%), three (4%) grade 3 or 4, and fully resolved in all cases. The overall response rate at 1 month after infusion was 98% (100% in B-ALL) in the CAR-naive cohort and 64% in the retreatment cohort. At 6 months, the probability of losing huCART19 persistence was 27% (95% CI, 14 to 41) for CAR-naive and 48% (95% CI, 30 to 64) for retreatment patients, whereas the incidence of B-cell recovery was 15% (95% CI, 6 to 28) and 58% (95% CI, 33 to 77), respectively. Relapse-free survival at 12 and 24 months, respectively, was 84% (95% CI, 72 to 97) and 74% (95% CI, 60 to 90) in CAR-naive and 74% (95% CI, 56 to 97) and 58% (95% CI, 37 to 90) in retreatment cohorts. CONCLUSION: HuCART19 achieved durable remissions with long-term persistence in children and young adults with relapsed or refractory B-ALL, including after failure of prior CAR T-cell therapy.
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Antígenos CD19/metabolismo , Leucemia-Linfoma Linfoblástico de Células Precursoras/tratamento farmacológico , Receptores de Antígenos de Linfócitos T/metabolismo , Receptores de Antígenos Quiméricos/metabolismo , Adolescente , Adulto , Criança , Pré-Escolar , Feminino , Humanos , Masculino , Projetos Piloto , Adulto JovemRESUMO
The pioneer transcription factor (TF) PU.1 controls hematopoietic cell fate by decompacting stem cell heterochromatin and allowing nonpioneer TFs to enter otherwise inaccessible genomic sites. PU.1 deficiency fatally arrests lymphopoiesis and myelopoiesis in mice, but human congenital PU.1 disorders have not previously been described. We studied six unrelated agammaglobulinemic patients, each harboring a heterozygous mutation (four de novo, two unphased) of SPI1, the gene encoding PU.1. Affected patients lacked circulating B cells and possessed few conventional dendritic cells. Introducing disease-similar SPI1 mutations into human hematopoietic stem and progenitor cells impaired early in vitro B cell and myeloid cell differentiation. Patient SPI1 mutations encoded destabilized PU.1 proteins unable to nuclear localize or bind target DNA. In PU.1-haploinsufficient pro-B cell lines, euchromatin was less accessible to nonpioneer TFs critical for B cell development, and gene expression patterns associated with the pro- to pre-B cell transition were undermined. Our findings molecularly describe a novel form of agammaglobulinemia and underscore PU.1's critical, dose-dependent role as a hematopoietic euchromatin gatekeeper.
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Agamaglobulinemia/genética , Cromatina/genética , Proteínas Proto-Oncogênicas/genética , Transativadores/genética , Adolescente , Adulto , Linfócitos B/fisiologia , Diferenciação Celular/genética , Linhagem Celular , Criança , Pré-Escolar , Células Dendríticas/fisiologia , Feminino , Regulação da Expressão Gênica no Desenvolvimento/genética , Células HEK293 , Hematopoese/genética , Células-Tronco Hematopoéticas/fisiologia , Humanos , Lactente , Linfopoese/genética , Masculino , Mutação/genética , Células Precursoras de Linfócitos B/fisiologia , Células-Tronco/fisiologia , Adulto JovemRESUMO
PURPOSE: Neurotrophic tyrosine receptor kinase (NTRK) fusions have been described as oncogenic drivers in a variety of tumors. However, little is known about the overall frequency of NTRK fusion in unselected pediatric tumors. Here, we assessed the frequency, fusion partners, and clinical course in pediatric patients with NTRK fusion-positive tumors. PATIENTS AND METHODS: We studied 1,347 consecutive pediatric tumors from 1,217 patients who underwent tumor genomic profiling using custom-designed DNA and RNA next-generation sequencing panels. NTRK fusions identified were orthogonally confirmed. RESULTS AND DISCUSSION: NTRK fusions were identified in 29 tumors from 27 patients with a positive yield of 2.22% for all patients and 3.08% for solid tumors. Although NTRK2 fusions were found exclusively in CNS tumors and NTRK1 fusions were highly enriched in papillary thyroid carcinomas, NTRK3 fusions were identified in all tumor categories. The most canonical fusion was ETV6-NTRK3 observed in 10 patients with diverse types of tumors. Several novel NTRK fusions were observed in rare tumor types, including KCTD16-NTRK1 in ganglioglioma and IRF2BP2-NTRK3 in papillary thyroid carcinomas. The detection of an NTRK fusion confirmed the morphologic diagnosis including five cases where the final tumor diagnosis was largely based on the discovery of an NTRK fusion. In one patient, the diagnosis was changed because of the identification of an ETV6-NTRK3 fusion. One patient with infantile fibrosarcoma was treated with larotrectinib and achieved complete pathologic remission. CONCLUSION: NTRK fusions are more frequently seen in pediatric tumors than in adult tumors and involve a broader panel of fusion partners and a wider range of tumors than previously recognized. These results highlight the importance of screening for NTRK fusions as part of the tumor genomic profiling for patients with pediatric cancer.
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CD19-directed chimeric antigen receptor (CAR) T cells show characteristic proliferation kinetics after infusion that correlate with response. Clearance of circulating disease, B-cell aplasia (BCA), and cytokine release syndrome (CRS) are used to observe CAR T-cell function, given the lack of commercial CAR T-cell measurement assays. We investigated the utility of common hematology laboratory parameters in 166 patients with B-cell acute lymphoblastic leukemia (B-ALL) who were treated with CAR T-cell therapy targeting CD19. CAR T-cell infusion was followed by disappearance of circulating blasts in 86% of patients at a median of 6 days. After a lag phase, there was a rapid expansion in absolute lymphocyte count (ALC) in the second week that coincided with the appearance of atypical lymphocytes. The expansion phase was followed by a contraction phase with a concomitant decrease in atypical lymphocytes. In vitro CAR T-cell studies showed similar kinetics and morphological changes. Peak ALC and overall expansion was greater in sustained responders compared with that in nonresponders. Patients with early loss of BCA and those with eventual CD19+ minimal residual disease/relapse showed lower overall lymphocyte expansion compared with the controls. Pleomorphic lymphocytosis was noted in the cerebrospinal fluid at post-CAR time points. We conclude that lymphocyte counts and differential can also be used to evaluate CAR T-cell expansion after infusion, along with BCA and CRS. This is the first report to characterize the morphology of CAR T cells and determine the utility of lymphocyte kinetics.
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Imunoterapia Adotiva , Receptores de Antígenos de Linfócitos T , Proliferação de Células , Humanos , Cinética , Contagem de Linfócitos , Recidiva , Linfócitos TRESUMO
PURPOSE: To prospectively evaluate the effectiveness of risk-adapted preemptive tocilizumab (PT) administration in preventing severe cytokine release syndrome (CRS) after CTL019, a CD19 chimeric antigen receptor T-cell therapy. METHODS: Children and young adults with CD19-positive relapsed or refractory B-cell acute lymphoblastic leukemia were assigned to high- (≥ 40%) or low- (< 40%) tumor burden cohorts (HTBC or LTBC) based on a bone marrow aspirate or biopsy before infusion. HTBC patients received a single dose of tocilizumab (8-12 mg/kg) after development of high, persistent fevers. LTBC patients received standard CRS management. The primary end point was the frequency of grade 4 CRS (Penn scale), with an observed rate of ≤ 5 of 15 patients in the HTBC pre-defined as clinically meaningful. In post hoc analyses, the HTBC was compared with a historical cohort of high-tumor burden patients from the initial phase I CTL019 trial. RESULTS: The primary end point was met. Seventy patients were infused with CTL019, 15 in the HTBC and 55 in the LTBC. All HTBC patients received the PT intervention. The incidence of grade 4 CRS was 27% (95% CI, 8 to 55) in the HTBC and 3.6% (95% CI, 0.4 to 13) in the LTBC. The best overall response rate was 87% in the HTBC and 100% in the LTBC. Initial CTL019 expansion was greater in the HTBC than the LTBC (P < .001), but persistence was not different (P = .73). Event-free and overall survival were worse in the HTBC (P = .004, P < .001, respectively). In the post hoc analysis, grade 4 CRS was observed in 27% versus 50% of patients in the PT and prior phase I cohorts, respectively (P = .18). CONCLUSION: Risk-adapted PT administration resulted in a decrease in the expected incidence of grade 4 CRS, meeting the study end point, without adversely impacting the antitumor efficacy or safety of CTL019.
Assuntos
Anticorpos Monoclonais Humanizados/uso terapêutico , Antígenos CD19/imunologia , Síndrome da Liberação de Citocina/tratamento farmacológico , Resistencia a Medicamentos Antineoplásicos , Imunoterapia Adotiva/efeitos adversos , Leucemia-Linfoma Linfoblástico de Células Precursoras B/terapia , Terapia de Salvação , Adolescente , Adulto , Criança , Pré-Escolar , Síndrome da Liberação de Citocina/etiologia , Síndrome da Liberação de Citocina/patologia , Feminino , Seguimentos , Humanos , Lactente , Masculino , Recidiva Local de Neoplasia/tratamento farmacológico , Recidiva Local de Neoplasia/patologia , Projetos Piloto , Leucemia-Linfoma Linfoblástico de Células Precursoras B/imunologia , Leucemia-Linfoma Linfoblástico de Células Precursoras B/patologia , Prognóstico , Estudos Prospectivos , Estudos Retrospectivos , Fatores de Risco , Taxa de Sobrevida , Adulto JovemRESUMO
Idiopathic multicentric Castleman disease (iMCD) is a rare and poorly understood hematologic disorder characterized by lymphadenopathy, systemic inflammation, cytopenias, and life-threatening multiorgan dysfunction. Interleukin-6 (IL-6) inhibition effectively treats approximately one-third of patients. Limited options exist for nonresponders, because the etiology, dysregulated cell types, and signaling pathways are unknown. We previously reported 3 anti-IL-6 nonresponders with increased mTOR activation who responded to mTOR inhibition with sirolimus. We investigated mTOR signaling in tissue and serum proteomes from iMCD patients and controls. mTOR activation was increased in the interfollicular space of iMCD lymph nodes (N = 26) compared with control lymph nodes by immunohistochemistry (IHC) for pS6, p4EBP1, and p70S6K, known effectors and readouts of mTORC1 activation. IHC for pS6 also revealed increased mTOR activation in iMCD compared with Hodgkin lymphoma, systemic lupus erythematosus, and reactive lymph nodes, suggesting that the mTOR activation in iMCD is not just a product of lymphoproliferation/inflammatory lymphadenopathy. Further, the degree of mTOR activation in iMCD was comparable to autoimmune lymphoproliferative syndrome, a disease driven by mTOR hyperactivation that responds to sirolimus treatment. Gene set enrichment analysis of serum proteomic data from iMCD patients (n = 88) and controls (n = 42) showed significantly enriched mTORC1 signaling. Finally, functional studies revealed increased baseline mTOR pathway activation in peripheral monocytes and T cells from iMCD remission samples compared with healthy controls. IL-6 stimulation augmented mTOR activation in iMCD patients, which was abrogated with JAK1/2 inhibition. These findings support mTOR activation as a novel therapeutic target for iMCD, which is being investigated through a trial of sirolimus (NCT03933904).
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Biomarcadores Tumorais/metabolismo , Hiperplasia do Linfonodo Gigante/patologia , Regulação Neoplásica da Expressão Gênica , Interleucina-6/metabolismo , Proteoma/análise , Proteínas Quinases S6 Ribossômicas/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Adolescente , Adulto , Idoso , Estudos de Casos e Controles , Hiperplasia do Linfonodo Gigante/metabolismo , Criança , Pré-Escolar , Estudos de Coortes , Feminino , Seguimentos , Humanos , Lactente , Masculino , Pessoa de Meia-Idade , Prognóstico , Proteoma/metabolismo , Transdução de Sinais , Adulto JovemRESUMO
RUNX1 is frequently mutated in myeloid and lymphoid malignancies. It has been shown to negatively regulate Toll-like receptor 4 (TLR4) signaling through nuclear factor κB (NF-κB) in lung epithelial cells. Here we show that RUNX1 regulates TLR1/2 and TLR4 signaling and inflammatory cytokine production by neutrophils. Hematopoietic-specific RUNX1 loss increased the production of proinflammatory mediators, including tumor necrosis factor-α (TNF-α), by bone marrow neutrophils in response to TLR1/2 and TLR4 agonists. Hematopoietic RUNX1 loss also resulted in profound damage to the lung parenchyma following inhalation of the TLR4 ligand lipopolysaccharide (LPS). However, neutrophils with neutrophil-specific RUNX1 loss lacked the inflammatory phenotype caused by pan-hematopoietic RUNX1 loss, indicating that dysregulated TLR4 signaling is not due to loss of RUNX1 in neutrophils per se. Rather, single-cell RNA sequencing indicates the dysregulation originates in a neutrophil precursor. Enhanced inflammatory cytokine production by neutrophils following pan-hematopoietic RUNX1 loss correlated with increased degradation of the inhibitor of NF-κB signaling, and RUNX1-deficient neutrophils displayed broad transcriptional upregulation of many of the core components of the TLR4 signaling pathway. Hence, early, pan-hematopoietic RUNX1 loss de-represses an innate immune signaling transcriptional program that is maintained in terminally differentiated neutrophils, resulting in their hyperinflammatory state. We hypothesize that inflammatory cytokine production by neutrophils may contribute to leukemia associated with inherited RUNX1 mutations.
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Subunidade alfa 2 de Fator de Ligação ao Core , Neutrófilos , Subunidade alfa 2 de Fator de Ligação ao Core/genética , NF-kappa B/metabolismo , Neutrófilos/metabolismo , Transdução de Sinais , Receptores Toll-LikeRESUMO
PURPOSE: The diagnosis of cancer predisposition in pediatric patients with cancer is vital for treatment decisions, surveillance, and management of at-risk family members. Somatic tumor testing can identify potential underlying constitutional variants that confer increased cancer risk. Here, we report the characteristics of constitutional variants identified through tumor testing. MATERIALS AND METHODS: Data were abstracted from medical record review of 1,023 patients who received inhouse somatic tumor testing over a 28-month period. Patients were identified for testing using referral criteria developed as a collaboration between genomic diagnostics, pathology, and oncology. Characteristics of patients who underwent constitutional testing, including family history and variant loss of heterozygosity, were tracked. RESULTS: From 1,023 patients who underwent somatic tumor sequencing in a 28-month period, 210 variants were identified in 141 patients (13.8%) that were concerning for cancer predisposition syndromes requiring intervention. A total of 73 variants in 41 patients have undergone clinical confirmatory testing thus far. Of these, 26 variants were confirmed to be constitutionally present (35.6%). Among patients tested, 23 (56.1%) of 41 total patients were diagnosed with a cancer predisposition syndrome. CONCLUSION: Our data demonstrate that more than one third of variants in tumor somatic sequencing that were concerning for underlying cancer predisposition were constitutionally confirmed. Overall, somatic tumor testing identified potential cancer predisposition syndromes in pediatric patients, and some would not have been identified on the basis of clinical history alone.
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Patient-derived xenotransplantation models of human myeloid diseases including acute myeloid leukemia, myelodysplastic syndromes and myeloproliferative neoplasms are essential for studying the biology of the diseases in pre-clinical studies. However, few studies have used these models for comparative purposes. Previous work has shown that acute myeloid leukemia blasts respond to human hematopoietic cytokines whereas myelodysplastic syndrome cells do not. We compared the engraftment of acute myeloid leukemia cells and myelodysplastic syndrome cells in NSG mice to that in NSG-S mice, which have transgene expression of human cytokines. We observed that only 50% of all primary acute myeloid leukemia samples (n=77) transplanted in NSG mice provided useful levels of engraftment (>0.5% human blasts in bone marrow). In contrast, 82% of primary acute myeloid leukemia samples engrafted in NSG-S mice with higher leukemic burden and shortened survival. Additionally, all of 5 injected samples from patients with myelodysplastic syndrome showed persistent engraftment on week 6; however, engraftment was mostly low (<2%), did not increase over time, and was only transiently affected by the use of NSG-S mice. Co-injection of mesenchymal stem cells did not enhance human myelodysplastic syndrome cell engraftment. Overall, we conclude that engraftment of acute myeloid leukemia samples is more robust compared to that of myelodysplastic syndrome samples and unlike those, acute myeloid leukemia cells respond positively to human cytokines, whereas myelodysplastic syndrome cells demonstrate a general unresponsiveness to them.
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Citocinas/metabolismo , Sobrevivência de Enxerto/imunologia , Hospedeiro Imunocomprometido , Leucemia Mieloide Aguda/imunologia , Leucemia Mieloide Aguda/metabolismo , Síndromes Mielodisplásicas/imunologia , Síndromes Mielodisplásicas/metabolismo , Animais , Transplante de Medula Óssea , Citocinas/sangue , Modelos Animais de Doenças , Feminino , Humanos , Leucemia Mieloide Aguda/terapia , Masculino , Células-Tronco Mesenquimais/metabolismo , Camundongos , Síndromes Mielodisplásicas/terapia , Transplante HeterólogoRESUMO
Utility of flow cytometry in the evaluation of pediatric hematopoietic neoplasms and the differences from adult hematopoietic neoplasms are discussed in this review. Distinction of hematogones from B-lymphoblasts, detection of residual/relapsed disease after novel targeted therapies, and evaluation of pediatric myeloid neoplasms are discussed.
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Citometria de Fluxo , Neoplasias Hematológicas , Leucemia , Linfoma , Criança , Neoplasias Hematológicas/diagnóstico , Neoplasias Hematológicas/patologia , Humanos , Leucemia/diagnóstico , Leucemia/patologia , Linfoma/diagnóstico , Linfoma/patologiaRESUMO
Prolonged exit from quiescence by hematopoietic stem cells (HSCs) progressively impairs their homeostasis in the bone marrow through an unidentified mechanism. We show that Rb proteins, which are major enforcers of quiescence, maintain HSC homeostasis by positively regulating thrombopoietin (Tpo)-mediated Jak2 signaling. Rb family protein inactivation triggers the progressive E2f-mediated transactivation of Socs3, a potent inhibitor of Jak2 signaling, in cycling HSCs. Aberrant activation of Socs3 impairs Tpo signaling and leads to impaired HSC homeostasis. Therefore, Rb proteins act as a central hub of quiescence and homeostasis by coordinating the regulation of both cell cycle and Jak2 signaling in HSCs.