RESUMEN
Chimeric antigen receptor (CAR) T cell therapy has demonstrated remarkable efficacy in relapsed/refractory (r/r) B cell malignancies, including in pediatric patients with acute lymphoblastic leukemia (ALL). Expanding this success to other hematologic and solid malignancies is an area of active research and, although challenges remain, novel solutions have led to significant progress over the past decade. Ongoing clinical trials for CAR T cell therapy for T cell malignancies and acute myeloid leukemia (AML) have highlighted challenges, including antigen specificity with off-tumor toxicity and persistence concerns. In T cell malignancies, notable challenges include CAR T cell fratricide and prolonged T cell aplasia, which are being addressed with strategies such as gene editing and suicide switch technologies. In AML, antigen identification remains a significant barrier, due to shared antigens across healthy hematopoietic progenitor cells and myeloid blasts. Strategies to limit persistence and circumvent the immunosuppressive tumor microenvironment (TME) created by AML are also being explored. CAR T cell therapies for central nervous system and solid tumors have several challenges, including tumor antigen heterogeneity, immunosuppressive and hypoxic TME, and potential for off-target toxicity. Numerous CAR T cell products have been designed to overcome these challenges, including "armored" CARs and CAR/T cell receptor (TCR) hybrids. Strategies to enhance CAR T cell delivery, augment CAR T cell performance in the TME, and ensure the safety of these products have shown promising results. In this manuscript, we will review the available evidence for CAR T cell use in T cell malignancies, AML, central nervous system (CNS), and non-CNS solid tumor malignancies, and recommend areas for future research.
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Inmunoterapia Adoptiva , Receptores Quiméricos de Antígenos , Humanos , Inmunoterapia Adoptiva/métodos , Inmunoterapia Adoptiva/efectos adversos , Niño , Receptores Quiméricos de Antígenos/inmunología , Receptores Quiméricos de Antígenos/uso terapéutico , Linfocitos T/inmunología , Linfocitos T/trasplante , Adolescente , Adulto , Receptores de Antígenos de Linfocitos T/inmunología , Receptores de Antígenos de Linfocitos T/uso terapéutico , Microambiente Tumoral/inmunologíaRESUMEN
PURPOSE: Chimeric antigen receptor (CAR) and T-cell receptor (TCR) T-cell therapies are effective in a subset of patients with solid tumors, but new approaches are needed to universally improve patient outcomes. Here, we developed a technology to leverage the cooperative effects of IL15 and IL21, two common cytokine-receptor gamma chain family members with distinct, pleiotropic effects on T cells and other lymphocytes, to enhance the efficacy of adoptive T cells. EXPERIMENTAL DESIGN: We designed vectors that induce the constitutive expression of either membrane-tethered IL15, IL21, or IL15/IL21. We used clinically relevant preclinical models of transgenic CARs and TCRs against pediatric and adult solid tumors to determine the effect of the membrane-tethered cytokines on engineered T cells for human administration. RESULTS: We found that self-delivery of these cytokines by CAR or TCR T cells prevents functional exhaustion by repeated stimulation and limits the emergence of dysfunctional natural killer (NK)-like T cells. Across different preclinical murine solid tumor models, we observed enhanced regression with each individual cytokine but the greatest antitumor efficacy when T cells were armored with both. CONCLUSIONS: The coexpression of membrane-tethered IL15 and IL21 represents a technology to enhance the resilience and function of engineered T cells against solid tumors and could be applicable to multiple therapy platforms and diseases. See related commentary by Ruffin et al., p. 1431.
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Interleucinas , Neoplasias , Receptores Quiméricos de Antígenos , Adulto , Humanos , Ratones , Animales , Niño , Receptores Quiméricos de Antígenos/genética , Receptores Quiméricos de Antígenos/metabolismo , Interleucina-15/genética , Inmunoterapia Adoptiva , Receptores de Antígenos de Linfocitos T , Neoplasias/genética , Neoplasias/terapia , Citocinas/metabolismoRESUMEN
Noradrenergic neuroblastoma is characterized by a core transcriptional regulatory circuitry (CRC) comprised of transcription factors (TF) such as PHOX2B, HAND2, and GATA3, which form a network with MYCN. At normal physiologic levels, MYCN mainly binds to promoters but when aberrantly upregulated as in neuroblastoma, MYCN also binds to enhancers. Here, we investigated how MYCN invades enhancers and whether CRC TFs play a role in this process. HAND2 was found to regulate chromatin accessibility and to assist MYCN binding to enhancers. Moreover, HAND2 cooperated with MYCN to compete with nucleosomes to regulate global gene transcription. The cooperative interaction between MYCN and HAND2 could be targeted with an Aurora A kinase inhibitor plus a histone deacetylase inhibitor, resulting in potent downregulation of both MYCN and the CRC TFs and suppression of MYCN-amplified neuroblastoma tumor growth. This study identifies cooperation between MYCN and HAND2 in neuroblastoma and demonstrates that simultaneously targeting MYCN and CRC TFs is an effective way to treat this aggressive pediatric tumor. SIGNIFICANCE: HAND2 and MYCN compete with nucleosomes to regulate global gene transcription and to drive a malignant neuroblastoma phenotype.
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Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico , Neuroblastoma , Humanos , Línea Celular Tumoral , Regulación Neoplásica de la Expresión Génica , Proteína Proto-Oncogénica N-Myc/genética , Neuroblastoma/genética , Neuroblastoma/patología , Nucleosomas , Factores de Transcripción/metabolismo , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genéticaRESUMEN
BACKGROUND: Although most patients with newly diagnosed high-risk neuroblastoma (NB) achieve remission after initial therapy, more than 50% experience late relapses caused by minimal residual disease (MRD) and succumb to their cancer. Therapeutic strategies to target MRD may benefit these children. We developed a new chimeric antigen receptor (CAR) targeting glypican-2 (GPC2) and conducted iterative preclinical engineering of the CAR structure to maximize its anti-tumor efficacy before clinical translation. METHODS: We evaluated different GPC2-CAR constructs by measuring the CAR activity in vitro. NOD-SCID mice engrafted orthotopically with human NB cell lines or patient-derived xenografts and treated with human CAR T cells served as in vivo models. Mechanistic studies were performed using single-cell RNA-sequencing. RESULTS: Applying stringent in vitro assays and orthotopic in vivo NB models, we demonstrated that our single-chain variable fragment, CT3, integrated into a CAR vector with a CD28 hinge, CD28 transmembrane, and 4-1BB co-stimulatory domain (CT3.28H.BBζ) elicits the best preclinical anti-NB activity compared with other tested CAR constructs. This enhanced activity was associated with an enrichment of CD8+ effector T cells in the tumor-microenvironment and upregulation of several effector molecules such as GNLY, GZMB, ZNF683, and HMGN2. Finally, we also showed that the CT3.28H.BBζ CAR we developed was more potent than a recently clinically tested GD2-targeted CAR to control NB growth in vivo. CONCLUSION: Given the robust preclinical activity of CT3.28H.BBζ, these results form a promising basis for further clinical testing in children with NB.
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Glipicanos , Neuroblastoma , Receptores Quiméricos de Antígenos , Animales , Niño , Humanos , Ratones , Antígenos CD28 , Gangliósidos , Glipicanos/inmunología , Glipicanos/uso terapéutico , Inmunoterapia Adoptiva/métodos , Ratones Endogámicos NOD , Ratones SCID , Neuroblastoma/metabolismo , Neuroblastoma/terapia , Receptores Quiméricos de Antígenos/genéticaRESUMEN
PURPOSE: Half of the patients with high-risk neuroblastoma who receive GD2-targeted mAb do not achieve long-term remissions. Recently, the antibody hu14.18 has been linked to IL2 (hu14.18-IL2) to enhance its efficacy and shown promising preclinical and clinical activity. We developed two new immunocytokines (IC) by linking two other γc cytokines, IL15 and IL21, to hu14.18. The purpose of this study was to compare hu14.18-IL15 and -IL21 with hu14.18-IL2 in their ability to induce antibody-dependent cell-mediated cytotoxicity (ADCC) against neuroblastoma. EXPERIMENTAL DESIGN: We assessed ADCC of hu14.18-IL15 and -IL2 (human cytokines, cross-reactive to mouse) against GD2low and GD2high neuroblastoma cell lines in vitro. T-cell-deficient mice with orthotopic patient-derived xenografts (PDX) and immunocompetent mice with transplantable orthotopic neuroblastoma were used to test all three ICs, including hu14.18-IL21 (murine IL21, not cross-reactive to human). Mechanistic studies were performed using single-cell RNA-sequencing (scRNA-seq). RESULTS: hu14.18-IL15 and hu14.18-IL2 mediated equivalent in vitro ADCC by human NK cells. When combined with chemotherapy, all three ICs similarly controlled the growth of PDXs in nude mice with murine NK effector cells. However, hu14.18-IL15 and -IL21 outperformed hu14.18-IL2 in immunocompetent mice with syngeneic neuroblastoma, inducing complete tumor regressions and extending survival. scRNA-seq data revealed an increase in CD8+ T cells and M1 tumor-associated macrophages and decreased regulatory T cells and myeloid-derived suppressor cells in the tumor microenvironment. CONCLUSIONS: Hu14.18-IL15 and Hu14.18-IL21 exhibit robust preclinical activity, warranting further consideration for clinical testing in patients with GD2-expressing neuroblastoma.
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Interleucina-2 , Neuroblastoma , Animales , Humanos , Interleucina-15/genética , Interleucina-15/uso terapéutico , Interleucinas , Ratones , Ratones Desnudos , Neuroblastoma/tratamiento farmacológico , Neuroblastoma/genética , Microambiente TumoralRESUMEN
Neuroblastoma accounts for 15% of cancer-related deaths in children, highlighting an unmet need for novel therapies. Selinexor is a small molecule inhibitor of XPO1. XPO1 shuffles cargo proteins with a nuclear export sequence from the nucleus to the cytosol, many of which are essential for cancer growth and cell maintenance. We systematically tested the effect of selinexor against neuroblastoma cells in vitro and in vivo and used an advanced proteomic and phosphoproteomic screening approach to interrogate unknown mechanisms of action. We found that selinexor induced its cytotoxic effects in neuroblastoma through the predominantly nuclear accumulation of p53 and global activation of apoptosis pathways. Selinexor also induced p53 phosphorylation at site S315, which is one initiating step for p53 degradation. Since this phosphorylation step is undertaken mostly by aurora kinase A (AURKA), we used the clinically available AURKA inhibitor, alisertib, and found p53-mediated lethality could be further augmented in three orthotopic xenograft mouse models. These findings suggest a potential therapeutic benefit using selinexor and alisertib to synergistically increase p53-mediated cytotoxicity of high-risk neuroblastoma.
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Neuroblastoma , Proteína p53 Supresora de Tumor , Animales , Apoptosis , Azepinas , Línea Celular Tumoral , Humanos , Hidrazinas , Carioferinas/metabolismo , Ratones , Neuroblastoma/tratamiento farmacológico , Proteómica , Pirimidinas , Receptores Citoplasmáticos y Nucleares/metabolismo , Receptores Citoplasmáticos y Nucleares/farmacología , Triazoles , Proteína p53 Supresora de Tumor/metabolismo , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
The translation of chimeric antigen receptor (CAR) T cell therapy for pediatric solid tumors is limited by the lack of preclinical models that fully recapitulate solid tumor biology. We describe steps to implement neuroblastoma metastatic and orthotopic mouse models. We delineate an analysis pipeline to quantify the efficacy and determine the immunological characteristics of both CAR T and tumor cells in these models. Both mouse models can be applied to evaluate other experimental therapies for neuroblastoma. For complete details on the use and execution of this protocol, please refer to Li et al. (2021).
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Técnicas de Cultivo de Célula , Inmunoterapia Adoptiva , Neuroblastoma , Receptores Quiméricos de Antígenos , Animales , Modelos Animales de Enfermedad , Femenino , Masculino , Ratones , Neuroblastoma/metabolismo , Neuroblastoma/terapia , Linfocitos T/citologíaRESUMEN
Targeting solid tumors must overcome several major obstacles, in particular, the identification of elusive tumor-specific antigens. Here, we devise a strategy to help identify tumor-specific epitopes. Glypican 2 (GPC2) is overexpressed in neuroblastoma. Using RNA sequencing (RNA-seq) analysis, we show that exon 3 and exons 7-10 of GPC2 are expressed in cancer but are minimally expressed in normal tissues. Accordingly, we discover a monoclonal antibody (CT3) that binds exons 3 and 10 and visualize the complex structure of CT3 and GPC2 by electron microscopy. The potential of this approach is exemplified by designing CT3-derived chimeric antigen receptor (CAR) T cells that regress neuroblastoma in mice. Genomic sequencing of T cells recovered from mice reveals the CAR integration sites that may contribute to CAR T cell proliferation and persistence. These studies demonstrate how RNA-seq data can be exploited to help identify tumor-associated exons that can be targeted by CAR T cell therapies.
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Anticuerpos Monoclonales/farmacología , Glipicanos/genética , Neoplasias del Sistema Nervioso/terapia , Neuroblastoma/terapia , Receptores de Antígenos de Linfocitos T/genética , Receptores Quiméricos de Antígenos/genética , Animales , Anticuerpos Monoclonales/química , Anticuerpos Monoclonales/genética , Anticuerpos Monoclonales/metabolismo , Línea Celular Tumoral , Proliferación Celular , Exones , Femenino , Expresión Génica , Glipicanos/antagonistas & inhibidores , Glipicanos/química , Glipicanos/inmunología , Humanos , Inmunoterapia Adoptiva/métodos , Ratones , Ratones Desnudos , Modelos Moleculares , Neoplasias del Sistema Nervioso/genética , Neoplasias del Sistema Nervioso/mortalidad , Neoplasias del Sistema Nervioso/patología , Neuroblastoma/genética , Neuroblastoma/mortalidad , Neuroblastoma/patología , Unión Proteica , Conformación Proteica , Receptores de Antígenos de Linfocitos T/inmunología , Receptores Quiméricos de Antígenos/inmunología , Análisis de Secuencia de ARN , Análisis de Supervivencia , Linfocitos T/inmunología , Linfocitos T/metabolismo , Carga Tumoral , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Immune checkpoint inhibitors (ICIs) have improved overall survival for cancer patients, however, optimal duration of ICI therapy has yet to be defined. Given ICIs were first used to treat patients with metastatic melanoma, a condition that at the time was incurable, little attention was initially paid to how much therapy would be needed for a durable response. As the early immunotherapy trials have matured past 10 years, a significant per cent of patients have demonstrated durable responses; it is now time to determine whether patients have been overtreated, and if durable remissions can still be achieved with less therapy, limiting the physical and financial toxicity associated with years of treatment. Well-designed trials are needed to identify optimal duration of therapy, and to define biomarkers to predict who would benefit from shorter courses of immunotherapy. Here, we outline key questions related to health, financial and societal toxicities of over treating with ICI and present four unique clinical trials aimed at exposing criteria for early cessation of ICI. Taken together, there is a serious liability to overtreating patients with ICI and future work is warranted to determine when it is safe to stop ICI.
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Inhibidores de Puntos de Control Inmunológico/administración & dosificación , Neoplasias/tratamiento farmacológico , Ensayos Clínicos como Asunto , Esquema de Medicación , Medicina Basada en la Evidencia , Humanos , Inhibidores de Puntos de Control Inmunológico/efectos adversos , Neoplasias/inmunología , Neoplasias/mortalidad , Neoplasias/patología , Seguridad del Paciente , Medición de Riesgo , Factores de Riesgo , Factores de Tiempo , Resultado del TratamientoRESUMEN
Faithful tumor mouse models are fundamental research tools to advance the field of immuno-oncology (IO). This is particularly relevant in diseases with low incidence, as in the case of pediatric malignancies, that rely on pre-clinical therapeutic development. However, conventional syngeneic and genetically engineered mouse models fail to recapitulate the tumor heterogeneity and microenvironmental complexity of human pathology that are essential determinants of cancer-directed immunity. Here, we characterize a novel mouse model that supports human natural killer (NK) cell development and engraftment of neuroblastoma orthotopic patient-derived xenograft (O-PDX) for pre-clinical antibody and cytokine testing. Using cytotoxicity assays, single-cell RNA-sequencing, and multi-color flow cytometry, we demonstrate that NK cells that develop in the humanized mice are fully licensed to execute NK cell cytotoxicity, permit human tumor engraftment, but can be therapeutically redirected to induce antibody-dependent cell-mediated cytotoxicity (ADCC). Although these cells share phenotypic and molecular features with healthy controls, we noted that they lacked an NK cell subset, termed activated NK cells, that is characterized by differentially expressed genes that are induced by cytokine activation. Because this subset of genes is also downregulated in patients with neuroblastoma compared to healthy controls, we hypothesize that this finding could be due to tumor-mediated suppressive effects. Thus, despite its technical complexity, this humanized patient-derived xenograft mouse model could serve as a faithful system for future testing of IO applications and studies of underlying immunologic processes.
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Anticuerpos Monoclonales Humanizados/farmacología , Antineoplásicos Inmunológicos/farmacología , Neuroblastoma/inmunología , Animales , Citotoxicidad Celular Dependiente de Anticuerpos/inmunología , Trasplante de Médula Ósea , Estudios de Casos y Controles , Línea Celular Tumoral , Terapia Combinada , Modelos Animales de Enfermedad , Femenino , Humanos , Células Asesinas Naturales/inmunología , Células Asesinas Naturales/metabolismo , Masculino , Ratones , Neuroblastoma/tratamiento farmacológico , Neuroblastoma/patología , Resultado del Tratamiento , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
PURPOSE OF REVIEW: In the era of immune-oncology, a breakthrough in the field of pediatric solid tumor research has been the demonstration that immunotherapy for patients with high-risk neuroblastoma improves the event-free and overall survival. Immunotherapeutic approaches including a monoclonal antibody targeting the cell surface glycosphingolipid disialoganglioside and cytokines successfully eliminate minimal residual disease. RECENT FINDINGS: Since this seminal discovery, clinical trials evaluating immunotherapy in combination with chemotherapy and cellular therapies have begun to demonstrate effectiveness in treatment of bulky disease. Broader knowledge has also been gained regarding immunotherapy-limiting side-effects. Furthermore, biologic studies in actively treated patients have contributed to our growing understanding of the underlying immunologic processes and mechanisms of tumor response and immune evasion. SUMMARY: The example of neuroblastoma is beginning to demonstrate that various immunotherapies combined with more conventional anticancer treatments can be synergistic. These advancements pose new challenges to both clinical researchers and medical provider and herald a new era in pediatric cancer therapy.
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Inmunoterapia , Neuroblastoma , Anticuerpos Monoclonales/uso terapéutico , Niño , Citocinas , Humanos , Factores Inmunológicos , Neuroblastoma/terapiaRESUMEN
Interleukin (IL)-2 and IL-21 dichotomously shape CD8+ T cell differentiation. IL-2 drives terminal differentiation, generating cells that are poorly effective against tumors, whereas IL-21 promotes stem cell memory T cells (TSCM) and antitumor responses. Here we investigated the role of metabolic programming in the developmental differences induced by these cytokines. IL-2 promoted effector-like metabolism and aerobic glycolysis, robustly inducing lactate dehydrogenase (LDH) and lactate production, whereas IL-21 maintained a metabolically quiescent state dependent on oxidative phosphorylation. LDH inhibition rewired IL-2-induced effects, promoting pyruvate entry into the tricarboxylic acid cycle and inhibiting terminal effector and exhaustion programs, including mRNA expression of members of the NR4A family of nuclear receptors, as well as Prdm1 and Xbp1 While deletion of Ldha prevented development of cells with antitumor effector function, transient LDH inhibition enhanced the generation of memory cells capable of triggering robust antitumor responses after adoptive transfer. LDH inhibition did not significantly affect IL-21-induced metabolism but caused major transcriptomic changes, including the suppression of IL-21-induced exhaustion markers LAG3, PD1, 2B4, and TIM3. LDH inhibition combined with IL-21 increased the formation of TSCM cells, resulting in more profound antitumor responses and prolonged host survival. These findings indicate a pivotal role for LDH in modulating cytokine-mediated T cell differentiation and underscore the therapeutic potential of transiently inhibiting LDH during adoptive T cell-based immunotherapy, with an unanticipated cooperative antitumor effect of LDH inhibition and IL-21.
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Linfocitos T CD8-positivos/inmunología , Inhibidores Enzimáticos/farmacología , Interleucinas/metabolismo , L-Lactato Deshidrogenasa/antagonistas & inhibidores , Melanoma Experimental/terapia , Células Madre/inmunología , Animales , Linfocitos T CD8-positivos/efectos de los fármacos , Linfocitos T CD8-positivos/metabolismo , Linfocitos T CD8-positivos/trasplante , Diferenciación Celular/efectos de los fármacos , Diferenciación Celular/inmunología , Línea Celular Tumoral/trasplante , Humanos , Memoria Inmunológica , Inmunoterapia Adoptiva/métodos , Interleucina-2/inmunología , Interleucina-2/metabolismo , Interleucinas/inmunología , L-Lactato Deshidrogenasa/metabolismo , Melanoma Experimental/inmunología , Ratones , Cultivo Primario de Células , Células Madre/efectos de los fármacos , Células Madre/metabolismoRESUMEN
BACKGROUND: Natural killer (NK) cells are one of the main effector populations of immunotherapy with monoclonal antibody and cytokines, used in combination with chemotherapy to treat children with high-risk neuroblastoma on this phase II trial. However, the impact of chemoimmunotherapy on NK cell kinetics, phenotype, and function is understudied. METHODS: We prospectively examined NK cell properties from 63 children with newly diagnosed neuroblastoma enrolled in a phase II trial (NCT01857934) and correlated our findings with tumor volume reduction after 2 courses of chemoimmunotherapy. NK cell studies were conducted longitudinally during chemoimmunotherapy and autologous hematopoietic cell transplantation (autoHCT) with optional haploidentical NK cell infusion and additional immunotherapy. RESULTS: Chemoimmunotherapy led to significant NK cytopenia, but complete NK cell recovery reliably occurred by day 21 of each therapy course as well as after autoHCT. Haploidentical NK cell infusion elevated the NK cell count transiently during autoHCT. NK cell cytotoxicity increased significantly during treatment compared with diagnosis. In addition, NK cells maintained their ability to respond to cytokine stimulation in culture longitudinally. Unsupervised cluster analysis of CD56bright NK cell count and tumor volume at diagnosis and after two courses of chemoimmunotherapy identified two patient groups with distinct primary tumor sizes and therapy responses. CONCLUSION: After profound NK cytopenia due to chemoimmunotherapy, endogenously reconstituted NK cells exhibit enhanced NK cytotoxicity compared with pretherapy measurements. Our data suggest a relationship between CD56bright expression and tumor size before and after two courses of chemoimmunotherapy; however, future studies are necessary to confirm this relationship and its predictive significance. TRIAL REGISTRATION NUMBER: NCT01857934.
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Inmunoterapia/métodos , Células Asesinas Naturales/metabolismo , Neuroblastoma/genética , Adolescente , Niño , Preescolar , Femenino , Humanos , Lactante , Recién Nacido , Cinética , Masculino , Estudios ProspectivosRESUMEN
Aggressive cancers often have activating mutations in growth-controlling oncogenes and inactivating mutations in tumor-suppressor genes. In neuroblastoma, amplification of the MYCN oncogene and inactivation of the ATRX tumor-suppressor gene correlate with high-risk disease and poor prognosis. Here we show that ATRX mutations and MYCN amplification are mutually exclusive across all ages and stages in neuroblastoma. Using human cell lines and mouse models, we found that elevated MYCN expression and ATRX mutations are incompatible. Elevated MYCN levels promote metabolic reprogramming, mitochondrial dysfunction, reactive-oxygen species generation, and DNA-replicative stress. The combination of replicative stress caused by defects in the ATRX-histone chaperone complex, and that induced by MYCN-mediated metabolic reprogramming, leads to synthetic lethality. Therefore, ATRX and MYCN represent an unusual example, where inactivation of a tumor-suppressor gene and activation of an oncogene are incompatible. This synthetic lethality may eventually be exploited to improve outcomes for patients with high-risk neuroblastoma.
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Proteína Proto-Oncogénica N-Myc/genética , Neuroblastoma/metabolismo , Proteína Nuclear Ligada al Cromosoma X/genética , Animales , Preescolar , Estudios de Cohortes , Femenino , Amplificación de Genes , Humanos , Lactante , Masculino , Ratones , Mitocondrias/genética , Mitocondrias/metabolismo , Mutación , Proteína Proto-Oncogénica N-Myc/metabolismo , Neuroblastoma/genética , Especies Reactivas de Oxígeno/metabolismo , Proteína Nuclear Ligada al Cromosoma X/metabolismoRESUMEN
PURPOSE: Immunotherapy with IL2, GM-CSF, and an anti-disialoganglioside (GD2) antibody significantly increases event-free survival in children with high-risk neuroblastoma. However, therapy failure in one third of these patients and IL2-related toxicities pose a major challenge. We compared the immunoadjuvant effects of IL15 with those of IL2 for enhancing antibody-dependent cell-mediated cytotoxicity (ADCC) in neuroblastoma. EXPERIMENTAL DESIGN: We tested ADCC against neuroblastoma patient-derived xenografts (PDX) in vitro and in vivo and examined the functional and migratory properties of NK cells activated with IL2 and IL15. RESULTS: In cell culture, IL15-activated NK cells induced higher ADCC against two GD+ neuroblastoma PDXs than did IL2-activated NK cells (P < 0.001). This effect was dose-dependent (P < 0.001) and was maintained across several effector-to-tumor ratios. As compared with IL2, IL15 also improved chemotaxis of NK cells, leading to higher numbers of tumorsphere-infiltrating NK cells in vitro (P = 0.002). In an orthotopic PDX model, animals receiving chemoimmunotherapy with an anti-GD2 antibody, GM-CSF, and a soluble IL15/IL15Rα complex had greater tumor regression than did those receiving chemotherapy alone (P = 0.012) or combined with anti-GD2 antibody and GM-CSF with (P = 0.016) or without IL2 (P = 0.035). This was most likely due to lower numbers of immature tumor-infiltrating NK cells (DX5+CD27+) after IL15/IL15Rα administration (P = 0.029) and transcriptional upregulation of Gzmd. CONCLUSIONS: The substitution of IL15 for IL2 leads to significant tumor regression in vitro and in vivo and supports clinical testing of IL15 for immunotherapy in pediatric neuroblastoma.
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Anticuerpos Monoclonales/inmunología , Citotoxicidad Celular Dependiente de Anticuerpos/inmunología , Inmunoterapia/métodos , Interleucina-15/inmunología , Células Asesinas Naturales/inmunología , Neuroblastoma/patología , Animales , Anticuerpos Monoclonales/administración & dosificación , Niño , Femenino , Gangliósidos/inmunología , Factor Estimulante de Colonias de Granulocitos y Macrófagos/inmunología , Humanos , Interleucina-2/inmunología , Neuroblastoma/inmunología , Neuroblastoma/terapia , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Consolidation therapies for children with intermediate- or high-risk acute myeloid leukemia (AML) are urgently needed to achieve higher cure rates while limiting therapy-related toxicities. We determined if adoptive transfer of natural killer (NK) cells from haploidentical killer immunoglobulin-like receptor (KIR)-human leukocyte antigen (HLA)-mismatched donors may prolong event-free survival in children with intermediate-risk AML who were in first complete remission after chemotherapy. Patients received cyclophosphamide (Day - 7), fludarabine (Days - 6 through - 2), and subcutaneous interleukin-2 (Days - 1, 1, 3, 5, 7, and 9). Purified, unmanipulated NK cells were infused on Day 0, and NK cell chimerism and phenotyping from peripheral blood were performed on Days 7, 14, 21, and 28. As primary endpoint, the event-free survival was compared to a cohort of 55 patients who completed chemotherapy and were in first complete remission but did not receive NK cells. Donor NK cell kinetics were determined as secondary endpoints. Twenty-one patients (median age at diagnosis, 6.0 years [range, 0.1-15.3 years]) received a median of 12.5 × 106 NK cells/kg (range, 3.6-62.2 × 106 cells/kg) without major side effects. All but 3 demonstrated transient engraftment with donor NK cells (median peak donor chimerism, 4% [range, 0-43%]). KIR-HLA-mismatched NK cells expanded in 17 patients (81%) and contracted in 4 (19%). However, adoptive transfer of NK cells did not decrease the cumulative incidence of relapse (0.393 [95% confidence interval: 0.182-0.599] vs. 0.35 [0.209-0.495]; P = .556) and did not improve event-free (60.7 ± 10.9% vs. 69.1 ± 6.8%; P = .553) or overall survival (84.2 ± 8.5% vs. 79.1 ± 6.6%; P = .663) over chemotherapy alone. The lack of benefit may result from insufficient numbers and limited persistence of alloreactive donor NK cells but does not preclude its potential usefulness during other phases of therapy, or in combination with other immunotherapeutic agents. TRIAL REGISTRATION: www.clinicaltrials.gov , NCT00703820 . Registered 24 June 2008.
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Protocolos de Quimioterapia Combinada Antineoplásica/administración & dosificación , Células Asesinas Naturales/trasplante , Leucemia Mieloide Aguda/terapia , Trasplante Haploidéntico/métodos , Adolescente , Traslado Adoptivo , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Niño , Preescolar , Quimioterapia de Consolidación , Ciclofosfamida/administración & dosificación , Ciclofosfamida/uso terapéutico , Supervivencia sin Enfermedad , Esquema de Medicación , Femenino , Humanos , Lactante , Interleucina-2/administración & dosificación , Interleucina-2/uso terapéutico , Masculino , Vidarabina/administración & dosificación , Vidarabina/análogos & derivados , Vidarabina/uso terapéuticoRESUMEN
The use of bisphosphonates for pain control in children with cancer is not extensively studied. We retrospectively evaluated 35 children with cancer treated with intravenous bisphosphonates for pain management at a single institution from 1998 through 2015. We analyzed pain scores and opioid and adjuvant medication consumption before bisphosphonate administration, daily for 2 weeks, and at 3 and 4 weeks after administration. We also determined the time interval between diagnosis and first administration of bisphosphonates and duration of life after bisphosphonate administration. Mean pain scores were 2.45 (±2.96) and 0.75 (±1.69) before and 14 days after bisphosphonate administration, respectively ( P = .25), and morphine equivalent doses of opioids were 5.52 (±13.35) and 5.27 (±9.77), respectively ( P = .07). Opioid consumption was significantly decreased at days 4 to 8, days 11 to 12, and week 3 after first bisphosphonate administration. The median duration of life after first bisphosphonate administration was 80 days, indicating its use late in the course of treatment. Bisphosphonates did not significantly improve pain outcomes at 2 weeks, but opioid consumption was reduced at several time points during the first 3 weeks. The use of bisphosphonates earlier in the course of pediatric oncological disease should be evaluated in prospective investigations.
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Neoplasias Óseas/tratamiento farmacológico , Dolor en Cáncer/tratamiento farmacológico , Difosfonatos/uso terapéutico , Manejo del Dolor/métodos , Administración Oral , Adolescente , Factores de Edad , Edad de Inicio , Analgésicos Opioides/administración & dosificación , Niño , Preescolar , Femenino , Gabapentina/administración & dosificación , Humanos , Masculino , Dimensión del Dolor , Grupos Raciales , Factores Sexuales , Factores de Tiempo , Adulto JovenRESUMEN
BACKGROUND: Neoadjuvant chemotherapy is given to children with unresectable hepatoblastoma to increase the rate and safety of curative complete surgical resection. Elevated levels of serum alpha-fetoprotein (sAFP) decline with tumor shrinkage. In this single-institution retrospective study, we determined early dynamic changes of sAFP levels and tumor volume in children during therapy for unresectable hepatoblastoma. METHODS: We correlated early dynamic changes of sAFP levels and tumor volume and the sum of the longest primary tumor and measurable metastatic disease diameters as per RECIST 1.1 criteria with patient outcome. RESULTS: There were 34 patients, 7 of whom died of disease. Patients with ≥ 90% (≥ 1 log10) decrease in sAFP levels after two chemotherapy courses had a better event-free survival (P = 0.039) and overall survival (OS; P = 0.045) than those with < 90% decrease. During this treatment interval, average tumor volume decreased from 481 mL (± 254 mL) to 268 mL (± 258 mL; P < 0.001) which was associated with OS (P = 0.029). Relative change in sAFP levels or tumor volume in between course 2 and pre-surgery or response as per RECIST 1.1 was not associated with OS. CONCLUSION: Early decline of sAFP levels and tumor volume, but not response as per RECIST 1.1 may predict survival in children with unresectable hepatoblastoma. This finding could be useful to identify therapy non-responders for whom alternative interventions may be required for cure. Confirmation of the finding using larger patient cohorts will be necessary before this strategy is incorporated into prospective trials.
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Biomarcadores de Tumor/sangre , Hepatoblastoma/tratamiento farmacológico , Neoplasias Hepáticas/tratamiento farmacológico , alfa-Fetoproteínas/análisis , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Niño , Preescolar , Femenino , Hepatoblastoma/mortalidad , Hepatoblastoma/patología , Humanos , Lactante , Neoplasias Hepáticas/mortalidad , Neoplasias Hepáticas/patología , Masculino , Terapia Neoadyuvante , Estudios Retrospectivos , Resultado del Tratamiento , Carga TumoralRESUMEN
Although anti-disialoganglioside (GD2) antibodies are successfully used for neuroblastoma therapy, a third of patients with neuroblastoma experience treatment failure or serious toxicity. Various strategies have been employed in the clinic to improve antibody-dependent cell-mediated cytotoxicity (ADCC), such as the addition of interleukin (IL)-2 to enhance natural killer (NK) cell function, adoptive transfer of allogeneic NK cells to exploit immune surveillance, and retinoid-induced differentiation therapy. Nevertheless, these mechanisms are not fully understood. We developed a quantitative assay to test ADCC induced by the anti-GD2 antibody Hu14.18K322A in nine neuroblastoma cell lines and dissociated cells from orthotopic patient-derived xenografts (O-PDXs) in culture. IL-2 improved ADCC against neuroblastoma cells, and differentiation with all-trans retinoic acid stabilized GD2 expression on tumor cells and enhanced ADCC as well. Degranulation was highest in licensed NK cells that expressed CD158b (P < 0.001) and harbored a killer-cell immunoglobulin-like receptor (KIR) mismatch against the tumor-specific human leukocyte antigen (HLA; P = 0.016). In conclusion, IL-2 is an important component of immunotherapy because it can improve the cytolytic function of NK cells against neuroblastoma cells and could lower the antibody dose required for efficacy, thereby reducing toxicity. The effect of IL-2 may vary among individuals and a biomarker would be useful to predict ADCC following IL-2 activation. Sub-populations of NK cells may have different levels of activity dependent on their licensing status, KIR expression, and HLA-KIR interaction. Better understanding of HLA-KIR interactions and the molecular changes following retinoid-induced differentiation is necessary to delineate their role in ADCC.
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Anticuerpos Monoclonales/administración & dosificación , Citotoxicidad Celular Dependiente de Anticuerpos/inmunología , Protocolos de Quimioterapia Combinada Antineoplásica/administración & dosificación , Gangliósidos/inmunología , Células Asesinas Naturales/inmunología , Neuroblastoma/terapia , Terapia Combinada , Humanos , Interleucina-2/administración & dosificación , Neuroblastoma/inmunología , Neuroblastoma/patología , Tretinoina/administración & dosificación , Células Tumorales CultivadasRESUMEN
PURPOSE: We previously reported the cases of 3 children with diffuse intrinsic pontine glioma (DIPG) in whom noncontiguous treatment-related abnormalities (NCTRAs) developed in the brain after expanded-field radiation therapy (RT). To investigate the occurrence and putative mechanism of NCTRAs, we reviewed brain magnetic resonance imaging studies of patients with DIPG treated in 2 consecutive phase I clinical trials (trials 1 and 2). METHODS AND MATERIALS: The 55 children included in these trials received small-molecule inhibitors: vandetanib in trial 1 (n=32; mean age 6.4 years) and vandetanib and dasatinib in trial 2 (n=23; mean age 5.8 years). The patients also received conformal 3-dimensional RT (cumulative dose 54 Gy). For patients enrolled in trial 1, the clinical target volume (CTV) was expanded by 1 cm from the gross tumor volume. In trial 2, the expansion to form the CTV was 2 to 3 cm. A review of imaging studies was performed from the initial diagnosis through the end of progression-free survival. The imaging findings were grouped into 5 categories according to the presence, absence, location, extent, and putative mechanism of NCTRAs. Statistical analysis was performed to evaluate the association between covariates and NCTRA, cohort characterization, and survival comparisons. RESULTS: Overall survival was similar in both studies (P=.74). NCTRAs developed in 9 patients (39%) treated in trial 2 but in none treated in trial 1. The NCTRAs included T2-weighted hyperintensities with (n=3; radiation necrosis) or without (n=5) contrast uptake, supratentorial leukoencephalopathy (n=2), and ischemic stroke (n=1). All NCTRAs, except for 1, occurred within the CTV. Compared with nonaffected patients, patients with a NCTRA were younger (P=.003) and had had larger relative brain volumes exposed to doses >20 Gy. CONCLUSIONS: The imaging features of NCTRAs suggest that their development is secondary to synergistic steno-occlusive vascular effects induced by the combination of RT, an expanded CTV, potent antiangiogenic therapy, young age, and, in 1 case, a genetic predisposition.