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BACKGROUND: More effective incentives are needed to motivate paediatric oncology drug development, uncoupling it from dependency on adult drug development. Although the current European and North-American legislations aim to promote drug development for paediatrics and rare diseases, children and adolescents with cancer have not benefited as expected from these initiatives and cancer remains the first cause of death by disease in children older than one. Drug development for childhood cancer remains dependent on adult cancer indications and their potential market. The balance between the investment needed to execute a Paediatric Investigation Plan (PIP) in Europe and an initial Paediatric Study Plan (iPSP) in the US, coupled with the potential financial reward has not been sufficiently attractive to incite the pharmaceutical industry to develop drugs for rare indications such as childhood cancer. METHODS: We propose changes in the timing and nature of the rewards within the European Paediatric Medicine Regulation (PMR) and Regulation on Orphan Medicinal Products (both currently under review), which would drive earlier initiation of paediatric oncology studies and provide incentives for drug development specifically for childhood indications. RESULTS: We suggest modifying the PMR to ensure mechanism-of-action driven mandatory PIP and reorganization of incentives to a stepwise and incremental approach. Interim and final deliverables should be defined within a PIP or iPSP, each attracting a reward on completion. A crucial change would be the introduction of the interim deliverable requiring production of paediatric data that inform the go/no-go decisions on whether to take a drug forward to paediatric efficacy trials. CONCLUSION: Additionally, to address the critical gap in the current framework where there is a complete lack of incentives to promote paediatric-specific cancer drug development, we propose the introduction of early rewards in the Orphan Regulation, with a variant on the US-Creating Hope Act and its priority review vouchers.
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Motivação , Neoplasias , Adolescente , Adulto , Criança , Humanos , Neoplasias/tratamento farmacológico , Desenvolvimento de Medicamentos , Oncologia , Indústria FarmacêuticaRESUMO
Checkpoint inhibitor (CPI) therapies provide limited benefit to patients with tumors of low immune reactivity. T cell-inducing vaccines hold promise to exert long-lasting disease control in combination with CPI therapy. Safety, tolerability and recommended phase 2 dose (RP2D) of an individualized, heterologous chimpanzee adenovirus (ChAd68) and self-amplifying mRNA (samRNA)-based neoantigen vaccine in combination with nivolumab and ipilimumab were assessed as primary endpoints in an ongoing phase 1/2 study in patients with advanced metastatic solid tumors (NCT03639714). The individualized vaccine regimen was safe and well tolerated, with no dose-limiting toxicities. Treatment-related adverse events (TRAEs) >10% included pyrexia, fatigue, musculoskeletal and injection site pain and diarrhea. Serious TRAEs included one count each of pyrexia, duodenitis, increased transaminases and hyperthyroidism. The RP2D was 1012 viral particles (VP) ChAd68 and 30 µg samRNA. Secondary endpoints included immunogenicity, feasibility of manufacturing and overall survival (OS). Vaccine manufacturing was feasible, with vaccination inducing long-lasting neoantigen-specific CD8 T cell responses. Several patients with microsatellite-stable colorectal cancer (MSS-CRC) had improved OS. Exploratory biomarker analyses showed decreased circulating tumor DNA (ctDNA) in patients with prolonged OS. Although small study size limits statistical and translational analyses, the increased OS observed in MSS-CRC warrants further exploration in larger randomized studies.
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Neoplasias Colorretais , Pan troglodytes , Adenoviridae/genética , Animais , Neoplasias Colorretais/tratamento farmacológico , Febre , Humanos , RNA Mensageiro/uso terapêuticoRESUMO
Rapid evaluation and subsequent regulatory approval of new drugs are critical to improving survival and reducing long-term side-effects for children and adolescents with cancer. The international multi-stakeholder organisation ACCELERATE was created to advance the timely investigation of new anti-cancer drugs. ACCELERATE has enhanced communication and understanding between academia, industry, patient advocates and regulators. It has promoted a mechanism-of-action driven drug development approach and developed Paediatric Strategy Forums. These initiatives have facilitated prioritisation of medicinal products and a focused and sequential strategy for drug development where there are multiple potential agents. ACCELERATE has championed the early assessment of promising drugs in adolescents through their inclusion in adult early phase trials. ACCELERATE has strongly supported alignment between the European Medicines Agency and the US Food and Drug Administration and identification of unmet medical needs through multi-stakeholder collaboration. Early engagement between all stakeholders in the development of new drugs is critical. Innovative clinical trial designs are required, necessitating early discussion with sponsors and regulators. Amplifying the patient advocate voice through inclusion across the drug development continuum will lead to better, patient-centric trials. By these means, children and adolescents with cancer can maximally and rapidly benefit from innovative products to improve outcomes and reduce burdensome sequelae.
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Antineoplásicos , Neoplasias , Adolescente , Adulto , Antineoplásicos/efeitos adversos , Criança , Desenvolvimento de Medicamentos , Humanos , Neoplasias/tratamento farmacológico , Estados Unidos , United States Food and Drug AdministrationRESUMO
PURPOSE: The HERBY trial evaluated the benefit of the addition of the antiangiogenic agent Bevacizumab (BEV) to radiotherapy/temozolomide (RT/TMZ) in pediatric patients with newly diagnosed non-brainstem high-grade glioma (HGG). The work presented here aims to correlate imaging characteristics and outcome measures with pathologic and molecular data. EXPERIMENTAL DESIGN: Radiological, pathologic, and molecular data were correlated with trial clinical information to retrospectively re-evaluate event-free survival (EFS) and overall survival (OS). RESULTS: One-hundred thirteen patients were randomized to the RT/TMZ arm (n = 54) or the RT/TMZ+BEV (BEV arm; n = 59). The tumor arose in the cerebral hemispheres in 68 patients (Cerebral group) and a midline location in 45 cases (Midline group). Pathologic diagnosis was available in all cases and molecular data in 86 of 113. H3 K27M histone mutations were present in 23 of 32 Midline cases and H3 G34R/V mutations in 7 of 54 Cerebral cases. Total/near-total resection occurred in 44 of 68 (65%) Cerebral cases but in only 5 of 45 (11%) Midline cases (P < 0.05). Leptomeningeal metastases (27 cases, 13 with subependymal spread) at relapse were more frequent in Midline (17/45) than in Cerebral tumors (10/68, P < 0.05). Mean OS (14.1 months) and EFS (9.0 months) in Midline tumors were significantly lower than mean OS (20.7 months) and EFS (14.9 months) in Cerebral tumors (P < 0.05). Pseudoprogression occurred in 8 of 111 (6.2%) cases. CONCLUSIONS: This study has shown that the poor outcome of midline tumors (compared with cerebral) may be related to (1) lesser surgical resection, (2) H3 K27M histone mutations, and (3) higher leptomeningeal dissemination.
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Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Neoplasias Encefálicas/patologia , Quimiorradioterapia/mortalidade , Glioma/patologia , Recidiva Local de Neoplasia/patologia , Procedimentos Neurocirúrgicos/mortalidade , Adolescente , Bevacizumab/administração & dosagem , Neoplasias Encefálicas/diagnóstico por imagem , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/terapia , Criança , Pré-Escolar , Terapia Combinada , Feminino , Glioma/diagnóstico por imagem , Glioma/genética , Glioma/terapia , Histonas/genética , Humanos , Masculino , Mutação , Gradação de Tumores , Recidiva Local de Neoplasia/diagnóstico por imagem , Recidiva Local de Neoplasia/genética , Recidiva Local de Neoplasia/terapia , Estudos Retrospectivos , Taxa de Sobrevida , Temozolomida/administração & dosagemRESUMO
Although outcomes for children with cancer have significantly improved over the past 40 years, there has been little progress in the treatment of some pediatric cancers, particularly when advanced. Additionally, clinical trial options and availability are often insufficient. Improved genomic and immunologic understanding of pediatric cancers, combined with innovative clinical trial designs, may provide an enhanced opportunity to study childhood cancers. Master protocols, which incorporate the use of precision medicine approaches, coupled with the ability to quickly assess the safety and effectiveness of new therapies, have the potential to accelerate early-phase clinical testing of novel therapeutics and which may result in more rapid approval of new drugs for children with cancer. Designing and conducting master protocols for children requires addressing similar principles and requirements as traditional adult oncology trials, but there are also unique considerations for master protocols conducted in children with cancer. The purpose of this paper is to define the key challenges and opportunities associated with this approach in order to ensure that master protocols can be adapted to benefit children and adolescents and ensure that adequate data are captured to advance, in parallel, the clinical development of investigational agents for children with cancer.
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Antineoplásicos , Protocolos Clínicos , Ensaios Clínicos como Assunto , Desenvolvimento de Medicamentos , Criança , Tomada de Decisões , Humanos , Neoplasias/tratamento farmacológico , Projetos de Pesquisa , Participação dos InteressadosRESUMO
Neoantigens, which are expressed on tumor cells, are one of the main targets of an effective antitumor T-cell response. Cancer immunotherapies to target neoantigens are of growing interest and are in early human trials, but methods to identify neoantigens either require invasive or difficult-to-obtain clinical specimens, require the screening of hundreds to thousands of synthetic peptides or tandem minigenes, or are only relevant to specific human leukocyte antigen (HLA) alleles. We apply deep learning to a large (N = 74 patients) HLA peptide and genomic dataset from various human tumors to create a computational model of antigen presentation for neoantigen prediction. We show that our model, named EDGE, increases the positive predictive value of HLA antigen prediction by up to ninefold. We apply EDGE to enable identification of neoantigens and neoantigen-reactive T cells using routine clinical specimens and small numbers of synthetic peptides for most common HLA alleles. EDGE could enable an improved ability to develop neoantigen-targeted immunotherapies for cancer patients.
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Importance: Few patient populations are as helpless and in need of advocacy as children with cancer. Pharmaceutical companies have historically faced significant financial disincentives to pursue pediatric oncology therapeutics, including low incidence, high costs of conducting pediatric trials, and a lack of funding for early-stage research. Observations: Review of published studies of pediatric oncology research and the cost of drug development, as well as clinical trials of pediatric oncology therapeutics at ClinicalTrials.gov, identified 77 potential drug development projects to be included in a hypothetical portfolio. The returns of this portfolio were simulated so as to compute the financial returns and risk. Simulated business strategies include combining projects at different clinical phases of development, obtaining partial funding from philanthropic grants, and obtaining government guarantees to reduce risk. The purely private-sector portfolio exhibited expected returns ranging from -24.2% to 10.2%, depending on the model variables assumed. This finding suggests significant financial disincentives for pursuing pediatric oncology therapeutics and implies that financial support from the public and philanthropic sectors is essential. Phase diversification increases the likelihood of a successful drug and yielded expected returns of -5.3% to 50.1%. Standard philanthropic grants had a marginal association with expected returns, and government guarantees had a greater association by reducing downside exposure. An assessment of a proposed venture philanthropy fund demonstrated stronger performance than the purely private-sector-funded portfolio or those with traditional amounts of philanthropic support. Clinical Relevance: A combination of financial and business strategies has the potential to maximize expected return while eliminating some downside risk-in certain cases enabling expected returns as high as 50.1%-that can overcome current financial disincentives and accelerate the development of pediatric oncology therapeutics.
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Oncologia/economia , Modelos Econômicos , Neoplasias/economia , Pediatria/economia , Criança , Custos e Análise de Custo , Humanos , Oncologia/métodos , Neoplasias/diagnóstico , Neoplasias/terapia , Pediatria/métodosRESUMO
Purpose Bevacizumab (BEV) is approved in more than 60 countries for use in adults with recurrent glioblastoma. We evaluated the addition of BEV to radiotherapy plus temozolomide (RT+TMZ) in pediatric patients with newly diagnosed high-grade glioma (HGG). Methods The randomized, parallel group, multicenter, open-label HERBY trial ( ClinicalTrials.gov identifier: NCT01390948) enrolled patients age ≥ 3 years to ≤ 18 years with localized, centrally neuropathology-confirmed, nonbrainstem HGG. Eligible patients were randomly assigned to receive RT + TMZ (RT: 1.8 Gy, 5 days per week, and TMZ: 75 mg/m2 per day for 6 weeks; 4-week treatment break; then up to 12 × 28-day cycles of TMZ [cycle 1: 150 mg/m2 per day, days 1 to 5; cycles 2 to 12: 200 mg/m2 per day, days 1 to 5]) with or without BEV (10 mg/kg every 2 weeks). The primary end point was event-free survival (EFS) as assessed by a central radiology review committee that was blinded to treatment. We report findings of EFS at 12 months after the enrollment of the last patient. Results One hundred twenty-one patients were enrolled (RT+TMZ [n = 59]; BEV plus RT+TMZ [n = 62]). Central radiology review committee-assessed median EFS did not differ significantly between treatment groups (RT+TMZ, 11.8 months; 95% CI, 7.9 to 16.4 months; BEV plus RT+TMZ, 8.2 months; 95% CI, 7.8 to 12.7 months; hazard ratio, 1.44; P = .13 [stratified log-rank test]). In the overall survival analysis, the addition of BEV did not reduce the risk of death (hazard ratio, 1.23; 95% CI, 0.72 to 2.09). More patients in the BEV plus RT+TMZ group versus the RT+TMZ group experienced one or more serious adverse events (n = 35 [58%] v n = 27 [48%]), and more patients who received BEV discontinued study treatment as a result of adverse events (n = 13 [22%] v n = 3 [5%]). Conclusion Adding BEV to RT+TMZ did not improve EFS in pediatric patients with newly diagnosed HGG. Our findings were not comparable to those of previous adult trials, which highlights the importance of performing pediatric-specific studies.
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Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Neoplasias Cerebelares/tratamento farmacológico , Glioma/tratamento farmacológico , Adolescente , Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Bevacizumab/administração & dosagem , Bevacizumab/efeitos adversos , Neoplasias Cerebelares/patologia , Neoplasias Cerebelares/radioterapia , Neoplasias Cerebelares/cirurgia , Quimiorradioterapia Adjuvante , Criança , Pré-Escolar , Feminino , Glioma/patologia , Glioma/radioterapia , Glioma/cirurgia , Humanos , Masculino , Gradação de Tumores , Intervalo Livre de Progressão , Temozolomida/administração & dosagem , Temozolomida/efeitos adversosRESUMO
An urgent need remains for new paediatric oncology drugs to cure children who die from cancer and to reduce drug-related sequelae in survivors. In 2007, the European Paediatric Regulation came into law requiring industry to create paediatric drug (all types of medicinal products) development programmes alongside those for adults. Unfortunately, paediatric drug development is still largely centred on adult conditions and not a mechanism of action (MoA)-based model, even though this would be more logical for childhood tumours as these have much fewer non-synonymous coding mutations than adult malignancies. Recent large-scale sequencing by International Genome Consortium and Paediatric Cancer Genome Project has further shown that the genetic and epigenetic repertoire of driver mutations in specific childhood malignancies differs from more common adult-type malignancies. To bring about much needed change, a Paediatric Platform, ACCELERATE, was proposed in 2013 by the Cancer Drug Development Forum, Innovative Therapies for Children with Cancer, the European Network for Cancer Research in Children and Adolescents and the European Society for Paediatric Oncology. The Platform, comprising multiple stakeholders in paediatric oncology, has three working groups, one with responsibility for promoting and developing high-quality MoA-informed paediatric drug development programmes, including specific measures for adolescents. Key is the establishment of a freely accessible aggregated database of paediatric biological tumour drug targets to be aligned with an aggregated pipeline of drugs. This will enable prioritisation and conduct of early phase clinical paediatric trials to evaluate these drugs against promising therapeutic targets and to generate clinical paediatric efficacy and safety data in an accelerated time frame. Through this work, the Platform seeks to ensure that potentially effective drugs, where the MoA is known and thought to be relevant to paediatric malignancies, are evaluated in early phase clinical trials, and that this approach to generate pre-clinical and clinical data is systematically pursued by academia, sponsors, industry, and regulatory bodies to bring new paediatric oncology drugs to front-line therapy more rapidly.
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Antineoplásicos/uso terapêutico , Descoberta de Drogas/métodos , Oncologia/métodos , Terapia de Alvo Molecular/métodos , Neoplasias/tratamento farmacológico , Criança , Bases de Dados Factuais , Avaliação de Medicamentos , Indústria Farmacêutica/métodos , HumanosRESUMO
The present work was designed to compare two mechanisms of cellular recognition based on Ab specificity: firstly, when the anti-HER2 mAb trastuzumab bridges target cells and cytotoxic lymphocytes armed with a Fc receptor (ADCC) and, secondly, when HER2 positive target cells are directly recognized by cytotoxic lymphocytes armed with a chimeric antigen receptor (CAR). To compare these two mechanisms, we used the same cellular effector (NK-92) and the same signaling domain (FcεRIγ). The NK-92 cytotoxic cell line was transfected with either a FcγRIIIa-FcεRIγ (NK-92(CD16)) or a trastuzumab-based scFv-FcεRIγ chimeric receptor (NK-92(CAR)). In vitro, the cytotoxic activity against HER2 positive target cells after indirect recognition by NK-92(CD16) was always inferior to that observed after direct recognition by NK-92(CAR). In contrast, and somehow unexpectedly, in vivo, adoptive transfer of NK-92(CD16) + trastuzumab but not of NK-92(CAR) induced tumor regression. Analysis of the in vivo xenogeneic system suggested that the human CH2-CH3 IgG2 used as a spacer in our construct was able to interact with the FcR present at the cell surface of the few NSG-FcR+ remaining immune cells. This interaction, leading to blockage of the NK-92(CAR) in the periphery of the engrafted tumor cells, stresses the critical role of the composition of the spacer domain.
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Antineoplásicos/farmacologia , Citotoxicidade Imunológica/efeitos dos fármacos , Imunoglobulina G/imunologia , Receptores de IgG/imunologia , Proteínas Recombinantes de Fusão/imunologia , Linfócitos T Citotóxicos/imunologia , Transferência Adotiva , Animais , Antineoplásicos/química , Cetuximab/química , Cetuximab/farmacologia , Proteínas Ligadas por GPI/química , Proteínas Ligadas por GPI/genética , Proteínas Ligadas por GPI/imunologia , Expressão Gênica , Humanos , Imunoglobulina G/química , Imunoglobulina G/genética , Células K562 , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Receptor ErbB-2/química , Receptor ErbB-2/genética , Receptor ErbB-2/imunologia , Receptores de IgE/química , Receptores de IgE/genética , Receptores de IgE/imunologia , Receptores de IgG/química , Receptores de IgG/genética , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/genética , Anticorpos de Cadeia Única/química , Anticorpos de Cadeia Única/genética , Anticorpos de Cadeia Única/imunologia , Linfócitos T Citotóxicos/citologia , Transdução Genética , Trastuzumab/química , Trastuzumab/farmacologia , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Neuroblastoma is a predominantly p53 wild-type (wt) tumour and MDM2-p53 antagonists offer a novel therapeutic strategy for neuroblastoma patients. RG7388 (Roche) is currently undergoing early phase clinical evaluation in adults. This study assessed the efficacy of RG7388 as a single-agent and in combination with chemotherapies currently used to treat neuroblastoma in a panel of neuroblastoma cell lines. RG7388 GI50 concentrations were determined in 21 p53-wt and mutant neuroblastoma cell lines of varying MYCN, MDM2 and p14(ARF) status, together with MYCN-regulatable Tet21N cells. The primary determinant of response was the presence of wt p53, and overall there was a >200-fold difference in RG7388 GI50 concentrations for p53-wt versus mutant cell lines. Tet21N MYCN+ cells were significantly more sensitive to RG7388 compared with MYCN- cells. Using median-effect analysis in 5 p53-wt neuroblastoma cell lines, selected combinations of RG7388 with cisplatin, doxorubicin, topotecan, temozolomide and busulfan were synergistic. Furthermore, combination treatments led to increased apoptosis, as evident by higher caspase-3/7 activity compared to either agent alone. These data show that RG7388 is highly potent against p53-wt neuroblastoma cells, and strongly supports its further evaluation as a novel therapy for patients with high-risk neuroblastoma and wt p53 to potentially improve survival and/or reduce toxicity.
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Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Neoplasias Encefálicas/tratamento farmacológico , Neuroblastoma/tratamento farmacológico , Proteínas Proto-Oncogênicas c-mdm2/antagonistas & inibidores , Pirrolidinas/farmacologia , Proteína Supressora de Tumor p53/antagonistas & inibidores , para-Aminobenzoatos/farmacologia , Neoplasias Encefálicas/metabolismo , Linhagem Celular Tumoral , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Neuroblastoma/metabolismo , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , Pirrolidinas/administração & dosagem , Proteína Supressora de Tumor p53/metabolismo , para-Aminobenzoatos/administração & dosagemRESUMO
Seven years after the launch of the European Paediatric Medicine Regulation, limited progress in paediatric oncology drug development remains a major concern amongst stakeholders - academics, industry, regulatory authorities, parents, patients and caregivers. Restricted increases in early phase paediatric oncology trials, legal requirements and regulatory pressure to propose early Paediatric Investigation Plans (PIPs), missed opportunities to explore new drugs potentially relevant for paediatric malignancies, lack of innovative trial designs and no new incentives to develop drugs against specific paediatric targets are some unmet needs. Better access to new anti-cancer drugs for paediatric clinical studies and improved collaboration between stakeholders are essential. The Cancer Drug Development Forum (CDDF), previously Biotherapy Development Association (BDA), with Innovative Therapy for Children with Cancer Consortium (ITCC), European Society for Paediatric Oncology (SIOPE) and European Network for Cancer Research in Children and Adolescents (ENCCA) has created a unique Paediatric Oncology Platform, involving multiple stakeholders and the European Union (EU) Commission, with an urgent remit to improve paediatric oncology drug development. The Paediatric Oncology Platform proposes to recommend immediate changes in the implementation of the Regulation and set the framework for its 2017 revision; initiatives to incentivise drug development against specific paediatric oncology targets, and repositioning of drugs not developed in adults. Underpinning these changes is a strategy for mechanism of action and biology driven selection and prioritisation of potential paediatric indications rather than the current process based on adult cancer indications. Pre-competitive research and drug prioritisation, early portfolio evaluation, cross-industry cooperation and multi-compound/sponsor trials are being explored, from which guidance for innovative trial designs will be provided.
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Antineoplásicos/uso terapêutico , Oncologia/métodos , Neoplasias/tratamento farmacológico , Pediatria/métodos , Adolescente , Adulto , Criança , Comportamento Cooperativo , Descoberta de Drogas/métodos , Descoberta de Drogas/tendências , Indústria Farmacêutica/métodos , Indústria Farmacêutica/tendências , Tratamento Farmacológico/métodos , Tratamento Farmacológico/tendências , HumanosRESUMO
BACKGROUND: Reversion of chemoresistance by inhibition of P-glycoprotein (P-gp) expression may overcome the chemoresistance observed in many cancer types and may allow for improved therapeutic ratio. We investigated whether siRNA specific for ABCB1 (MDR1) mRNA might restore sensitivity to chemotherapy in tumor cell lines known to overexpress the MDR1 gene. MATERIALS AND METHODS: MDR1-expressing tumor cell lines were transiently transfected with anti-MDR1 silencing RNA (siRNA) before exposure to doxorubicin or methotrexate. The capacity of siRNA to reduce cell proliferation and increase the IC50 of the tested chemotherapies was investigated. RESULTS: siRNA down-regulated MDR1 mRNA expression by 50% in breast carcinoma and osteosarcoma cell lines, and significantly inhibited tumor cell proliferation up to 90% (p<0.01), when co-administered with doxorubicin or methotrexate, despite the known chemoresistance of the cell lines. siRNAs reduced the IC50 of doxorubicin and methotrexate by more than 10-fold (p<0.01). CONCLUSION: These results suggest the potential clinical application of anti-MDR1 siRNA to restore chemosensitivity and possibly improve the therapeutic ratio of these cytotoxic drugs.
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Membro 1 da Subfamília B de Cassetes de Ligação de ATP/genética , Neoplasias da Mama/tratamento farmacológico , Resistencia a Medicamentos Antineoplásicos/genética , Osteossarcoma/tratamento farmacológico , RNA Interferente Pequeno/genética , Subfamília B de Transportador de Cassetes de Ligação de ATP , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Sequência de Bases , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Proliferação de Células , Doxorrubicina/farmacologia , Doxorrubicina/uso terapêutico , Feminino , Humanos , Concentração Inibidora 50 , Metotrexato/farmacologia , Metotrexato/uso terapêutico , Osteossarcoma/patologiaRESUMO
The identification of the optimal T-cell effector subtype is a crucial issue for adoptive cell therapy with chimeric receptor-modified T cells. The ideal T cell population must be able to home toward tumor site, exert prolonged antitumoral activity, and display minimal toxicity against normal tissues. Therefore, we characterized the in vitro antitumoral properties of three effector T-cell populations: Epstein-Barr virus-specific cytotoxic T lymphocytes (EBV-CTLs), cytokine-induced killer (CIK) cells, and γ9δ2 T (GDT) cells, after transduction with a chimeric receptor specific for the CD33 antigen, broadly expressed on acute myeloid leukemia cells. EBV-CTLs, CIK, and GDT cells were generated and transduced with high efficiency with a retroviral vector coding for an anti-CD33-ζ chimeric receptor without alterations of their native phenotype. Anti-CD33-ζ chimeric receptor-redirected T cells displayed analogous in vitro chemotactic activity toward CXCL12. In addition, anti-CD33-ζ chimeric receptor-expressing EBV-CTLs, CIK, and GDT cells showed potent and similar cytotoxicity against several CD33⺠leukemic targets both in short-term 4-hours-5¹chromium-release assays (mean killing vs primary leukemic cells at effector:target ratio of 5:1; 50%, 61%, and 50% for EBV-CTLs, CIK, and GDT cells, respectively) and in long-term assays, where they were cocultured with leukemic cells for 6 days on stromal mesenchymal cells (mean survival of primary leukemic cells at effector:target ratio of 1:100; 18%, 16%, and 29% for EBV-CTLs, CIK, and GDT cells, respectively). Moreover, all effector cells acquired consistent capability to proliferate in vitro after contact with CD33⺠cells and to release high and comparable levels of immunostimulatory cytokines, while secreting similar low amount of immunoregulatory cytokines as the unmanipulated counterpart. Our results indicate that expression of an anti-CD33-ζ chimeric receptor potently and similarly increase the antileukemic functions of different effector T-cell subtypes, underlying the impossibility to identify a more potent T-cell population through in vitro analysis, and consistently with recent observations that have emerged from clinical trials with chimeric receptor-modified T cells, suggesting the need to perform such type of studies in the human setting.
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Terapia Baseada em Transplante de Células e Tecidos , Quimerismo , Leucemia/imunologia , Leucemia/terapia , Receptores de Antígenos de Linfócitos T , Linfócitos T/imunologia , Linfócitos T/metabolismo , Complexo CD3/genética , Complexo CD3/imunologia , Complexo CD3/metabolismo , Linhagem Celular Tumoral , Quimiotaxia de Leucócito/imunologia , Criança , Vetores Genéticos/genética , Células HL-60 , Herpesvirus Humano 4/genética , Herpesvirus Humano 4/imunologia , Humanos , Células K562 , Ativação Linfocitária/genética , Ativação Linfocitária/imunologia , Receptores de Antígenos de Linfócitos T/genética , Receptores de Antígenos de Linfócitos T/metabolismo , Linfócitos T Citotóxicos/imunologia , Transdução GenéticaRESUMO
BACKGROUND: Graft contamination has been blamed for causing relapse in children with high-risk neuroblastoma (HRNB) after autologous hematopoietic stem cell transplantation (HSCT). PROCEDURE: We report the long-term results of hematopoietic reconstitution, post-transplant complications, and clinical outcome of 44 children with HRNB treated with busulfan/melphalan high-dose chemotherapy followed by transplantation of purged CD34+ immunoselected autologous peripheral HSCT. Minimal residual disease (MRD) of grafts was evaluated by anti-GD2 immunofluorescence or tyrosine hydroxylase reverse transcriptase-polymerase chain reaction (RT-PCR). RESULTS: Contaminating neuroblasts were found in 19/38 grafts (50%) before CD34+ positive selection, and none after (technique sensitivity of one cell in 10(5)). A median of 6.5 × 10(6) CD34+ cells/kg (range 0.8-23.7) were transplanted with only 2% of TRM. Neutrophils and platelet recovery occurred within a median of 12 days (range 9-47) and 44 days (range 12-259), respectively, without any secondary graft failure. Twenty-three percents of patients experienced a sepsis (10/44) and 14% a pyelonephritis (6/44). Recurrence of varicella zoster virus occurred in 21% of patients (9/44). Negative RT-PCR MRD within the leukapheresis product and cis-retinoic acid therapy were significantly and independently associated to a better survival (P < 0.05). Overall and event-free survivals at 5 years post-transplant were at 59.3% and 48.3% respectively. CONCLUSIONS: Besides high rates of manageable infections due to late immune recovery, transplantation with CD34+ immunoselected grafts in HRNB children was feasible and did not affect long-term hematopoiesis.
Assuntos
Antígenos CD34 , Transplante de Células-Tronco Hematopoéticas/métodos , Neuroblastoma/terapia , Bussulfano/uso terapêutico , Criança , Seguimentos , Hematopoese , Transplante de Células-Tronco Hematopoéticas/efeitos adversos , Células-Tronco Hematopoéticas/citologia , Humanos , Separação Imunomagnética , Infecções/induzido quimicamente , Leucaférese , Melfalan/uso terapêutico , Neoplasia Residual/diagnóstico , Neuroblastoma/complicações , Neuroblastoma/mortalidade , Análise de Sobrevida , Transplante AutólogoRESUMO
BACKGROUND: Cytokine-induced killer cells are ex vivo-expanded cells with potent antitumor activity. The infusion of cytokine-induced killer cells in patients with acute myeloid leukemia relapsing after allogeneic hematopoietic stem cell transplant is well tolerated, but limited clinical responses have been observed. To improve their effector functions against acute myeloid leukemia, we genetically modified cytokine-induced killer cells with chimeric receptors specific for the CD33 myeloid antigen. DESIGN AND METHODS: SFG-retroviral vectors coding for anti-CD33-ζ and anti-CD33-CD28-OX40-ζ chimeric receptors were used to transduce cytokine-induced killer cells. Transduced cells were characterized in vitro for their ability to lyse leukemic targets (4-hour (51)chromium-release and 6-day co-cultures assays on human stromal mesenchymal cells), to proliferate ((3)H-thymidine-incorporation assay) and to secrete cytokines (flow cytomix assay) after contact with acute myeloid leukemia cells. Their activity against normal CD34(+) hematopoietic progenitor cells was evaluated by analyzing the colony-forming unit capacity after co-incubation. RESULTS: Cytokine-induced killer cells were efficiently transduced with the anti-CD33 chimeric receptors, maintaining their native phenotype and functions and acquiring potent cytotoxicity (up to 80% lysis after 4-hour incubation) against different acute myeloid leukemia targets, as also confirmed in long-term killing experiments. Moreover, introduction of the anti-CD33 chimeric receptors was accompanied by prominent CD33-specific proliferative activity, with the release of high levels of immunostimulatory cytokines. The presence of CD28-OX40 in chimeric receptor endodomain was associated with a significant amelioration of the anti-leukemic activity of cytokine-induced killer cells. Importantly, even though the cytokine-induced killer cells transduced with anti-CD33 chimeric receptors showed toxicity against normal hematopoietic CD34(+) progenitor cells, residual clonogenic activity was preserved. CONCLUSIONS: Our results indicate that anti-CD33 chimeric receptors strongly enhance anti-leukemic cytokine-induced killer cell functions, suggesting that cytokine-induced killer cells transduced with these molecules might represent a promising optimized tool for acute myeloid leukemia immunotherapy.
Assuntos
Terapia Baseada em Transplante de Células e Tecidos/métodos , Células Matadoras Induzidas por Citocinas/imunologia , Leucemia Mieloide/terapia , Doença Aguda , Linhagem Celular Tumoral , Proliferação de Células , Células Cultivadas , Criança , Técnicas de Cocultura , Células Matadoras Induzidas por Citocinas/citologia , Células Matadoras Induzidas por Citocinas/metabolismo , Citotoxicidade Imunológica/imunologia , Citometria de Fluxo , Células HEK293 , Células HL-60 , Humanos , Leucemia Mieloide/imunologia , Leucemia Mieloide/patologia , Receptores Citoplasmáticos e Nucleares/genética , Receptores Citoplasmáticos e Nucleares/imunologia , Proteínas Recombinantes de Fusão/imunologia , Lectina 3 Semelhante a Ig de Ligação ao Ácido Siálico/imunologia , Fatores de TempoRESUMO
Tropomyosin-related kinase receptor C (TrkC) is a neurotrophin receptor with tyrosine kinase activity that was expected to be oncogenic. However, it has several characteristics of a tumor suppressor: its expression in tumors has often been associated with good prognosis; and it was recently demonstrated to be a dependence receptor, transducing different positive signals in the presence of ligand but inducing apoptosis in the absence of ligand. Here we show that the TrkC ligand neurotrophin-3 (NT-3) is upregulated in a large fraction of aggressive human neuroblastomas (NBs) and that it blocks TrkC-induced apoptosis of human NB cell lines, consistent with the idea that TrkC is a dependence receptor. Functionally, both siRNA knockdown of NT-3 expression and incubation with a TrkC-specific blocking antibody triggered apoptosis in human NB cell lines. Importantly, disruption of the NT-3 autocrine loop in malignant human neuroblasts triggered in vitro NB cell death and inhibited tumor growth and metastasis in both a chick and a mouse xenograft model. Thus, we believe that our data suggest that NT-3/TrkC disruption is a putative alternative targeted therapeutic strategy for the treatment of NB.
Assuntos
Apoptose , Comunicação Autócrina , Regulação Neoplásica da Expressão Gênica , Proteínas de Neoplasias/biossíntese , Neuroblastoma/metabolismo , Neurotrofina 3/biossíntese , Receptor trkC/biossíntese , Animais , Linhagem Celular Tumoral , Sobrevivência Celular , Galinhas , Técnicas de Silenciamento de Genes , Humanos , Camundongos , Camundongos Nus , Proteínas de Neoplasias/genética , Transplante de Neoplasias , Neuroblastoma/genética , Neuroblastoma/patologia , Neuroblastoma/terapia , Neurotrofina 3/genética , Receptor trkC/genética , Transplante Heterólogo , Regulação para Cima/genéticaRESUMO
UNLABELLED: Assessment of osteosarcoma response to neoadjuvant chemotherapy is performed by histopathologic analysis after surgical resection of the primary tumor. The purpose of this study was to evaluate whether (18)F-FDG PET could be a noninvasive surrogate to histopathologic analysis and allow for earlier response evaluation to neoadjuvant chemotherapy in osteosarcoma. METHODS: Metabolic response to neoadjuvant chemotherapy was assessed in immunocompetent rats with a preestablished orthotopic osteosarcoma using (18)F-FDG PET before and after receiving 2 doses of ifosfamide. Comparison was then made by assessing histologic responses on euthanized animals. RESULTS: Maximum standardized uptake value (SUVmax) measured by (18)F-FDG PET after 2 doses of chemotherapy was correlated to histologic classification (P < 0.01). An SUVmax less than 15 corresponded to good responders, whereas an SUVmax greater than 15 but less than 20 and an SUVmax greater than 20 corresponded to partial responders or nonresponders, respectively. A 40% decrease in SUVmax between the first and second (18)F-FDG PET scans distinguished between partial and good response to chemotherapy. CONCLUSION: Determination of SUVmax using semiquantitative (18)F-FDG PET predicts response to neoadjuvant chemotherapy earlier than does histologic analysis.
Assuntos
Antineoplásicos/administração & dosagem , Neoplasias Ósseas/tratamento farmacológico , Neoplasias Ósseas/patologia , Fluordesoxiglucose F18/farmacocinética , Osteossarcoma/tratamento farmacológico , Osteossarcoma/patologia , Tomografia por Emissão de Pósitrons/métodos , Animais , Neoplasias Ósseas/diagnóstico por imagem , Quimioterapia Adjuvante , Modelos Animais de Doenças , Avaliação Pré-Clínica de Medicamentos , Humanos , Taxa de Depuração Metabólica , Terapia Neoadjuvante , Osteossarcoma/diagnóstico por imagem , Compostos Radiofarmacêuticos/farmacocinética , Ratos , Ratos Sprague-Dawley , Estatística como Assunto , Resultado do TratamentoRESUMO
Neuroblastoma (NB), the most frequent solid tumor of early childhood, is diagnosed as a disseminated disease in >60% of cases, and several lines of evidence support the resistance to apoptosis as a prerequisite for NB progression. We show that autocrine production of netrin-1, a multifunctional laminin-related molecule, conveys a selective advantage in tumor growth and dissemination in aggressive NB, as it blocks the proapoptotic activity of the UNC5H netrin-1 dependence receptors. We show that such netrin-1 up-regulation is a potential marker for poor prognosis in stage 4S and, more generally, in NB stage 4 diagnosed infants. Moreover, we propose that interference with the netrin-1 autocrine loop in malignant neuroblasts could represent an alternative therapeutic strategy, as disruption of this loop triggers in vitro NB cell death and inhibits NB metastasis in avian and mouse models.
Assuntos
Fatores de Crescimento Neural/genética , Neuroblastoma/genética , Neuroblastoma/patologia , Proteínas Supressoras de Tumor/genética , Morte Celular , Linhagem Celular Tumoral , Sobrevivência Celular , Intervalo Livre de Doença , Regulação Neoplásica da Expressão Gênica , Humanos , Lactente , Recém-Nascido , Estadiamento de Neoplasias , Netrina-1 , Neuroblastoma/mortalidade , Prognóstico , RNA Interferente Pequeno/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Sobreviventes , Regulação para CimaRESUMO
BACKGROUND: MYCN oncogene amplification has been defined as the most important prognostic factor for neuroblastoma (NB), the most common solid extracranial neoplasm in children. High copy numbers are strongly associated with rapid tumor progression and poor outcome, independently of tumor stage or patient age, and this has become an important factor in treatment stratification. PROCEDURE: By real-time quantitative PCR analysis, we evaluated the clinical relevance of circulating MYCN DNA of 267 patients with locoregional or metastatic NB in children less than 18 months of age. RESULTS: For patients in this age group with INSS stage 4 or 4S NB and stage 3 patients, serum-based determination of MYCN DNA sequences had good sensitivity (85%, 83%, and 75% respectively) and high specificity (100%) when compared to direct tumor gene determination. In contrast, the approach showed low sensitivity patients with stages 1 and 2 disease. CONCLUSION: Our results show that the sensitivity of the serum-based MYCN DNA sequence determination depends on the stage of the disease. However, this simple, reproducible assay may represent a reasonably sensitive and very specific tool to assess tumor MYCN status in cases with stage 3 and metastatic disease for whom a wait and see strategy is often recommended.