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1.
Br J Cancer ; 131(4): 763-777, 2024 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-38942989

RESUMO

BACKGROUND: Certain paediatric nervous system malignancies have dismal prognoses. Retinoic acid (RA) is used in neuroblastoma treatment, and preclinical data indicate potential benefit in selected paediatric brain tumour entities. However, limited single-agent efficacy necessitates combination treatment approaches. METHODS: We performed drug sensitivity profiling of 76 clinically relevant drugs in combination with RA in 16 models (including patient-derived tumouroids) of the most common paediatric nervous system tumours. Drug responses were assessed by viability assays, high-content imaging, and apoptosis assays and RA relevant pathways by RNAseq from treated models and patient samples obtained through the precision oncology programme INFORM (n = 2288). Immunoprecipitation detected BCL-2 family interactions, and zebrafish embryo xenografts were used for in vivo efficacy testing. RESULTS: Group 3 medulloblastoma (MBG3) and neuroblastoma models were highly sensitive to RA treatment. RA induced differentiation and regulated apoptotic genes. RNAseq analysis revealed high expression of BCL2L1 in MBG3 and BCL2 in neuroblastomas. Co-treatments with RA and BCL-2/XL inhibitor navitoclax synergistically decreased viability at clinically achievable concentrations. The combination of RA with navitoclax disrupted the binding of BIM to BCL-XL in MBG3 and to BCL-2 in neuroblastoma, inducing apoptosis in vitro and in vivo. CONCLUSIONS: RA treatment primes MBG3 and NB cells for apoptosis, triggered by navitoclax cotreatment.


Assuntos
Apoptose , Sinergismo Farmacológico , Meduloblastoma , Neuroblastoma , Tretinoína , Peixe-Zebra , Humanos , Animais , Tretinoína/farmacologia , Meduloblastoma/tratamento farmacológico , Meduloblastoma/patologia , Meduloblastoma/metabolismo , Meduloblastoma/genética , Apoptose/efeitos dos fármacos , Neuroblastoma/tratamento farmacológico , Neuroblastoma/patologia , Neuroblastoma/metabolismo , Neuroblastoma/genética , Linhagem Celular Tumoral , Compostos de Anilina/farmacologia , Ensaios Antitumorais Modelo de Xenoenxerto , Sulfonamidas/farmacologia , Proteína bcl-X/genética , Proteína bcl-X/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Camundongos , Neoplasias Cerebelares/tratamento farmacológico , Neoplasias Cerebelares/patologia , Neoplasias Cerebelares/metabolismo , Neoplasias Cerebelares/genética , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Proteínas Proto-Oncogênicas c-myc/genética , Proteínas Proto-Oncogênicas c-myc/metabolismo , Proteína Proto-Oncogênica N-Myc
2.
J Neurooncol ; 168(2): 317-332, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38630384

RESUMO

INTRODUCTION: Patients with pediatric low-grade gliomas (pLGGs), the most common primary brain tumors in children, can often benefit from MAPK inhibitor (MAPKi) treatment. However, rapid tumor regrowth, also referred to as rebound growth, may occur once treatment is stopped, constituting a significant clinical challenge. METHODS: Four patient-derived pediatric glioma models were investigated to model rebound growth in vitro based on viable cell counts in response to MAPKi treatment and withdrawal. A multi-omics dataset (RNA sequencing and LC-MS/MS based phospho-/proteomics) was generated to investigate possible rebound-driving mechanisms. Following in vitro validation, putative rebound-driving mechanisms were validated in vivo using the BT-40 orthotopic xenograft model. RESULTS: Of the tested models, only a BRAFV600E-driven model (BT-40, with additional CDKN2A/Bdel) showed rebound growth upon MAPKi withdrawal. Using this model, we identified a rapid reactivation of the MAPK pathway upon MAPKi withdrawal in vitro, also confirmed in vivo. Furthermore, transient overactivation of key MAPK molecules at transcriptional (e.g. FOS) and phosphorylation (e.g. pMEK) levels, was observed in vitro. Additionally, we detected increased expression and secretion of cytokines (CCL2, CX3CL1, CXCL10 and CCL7) upon MAPKi treatment, maintained during early withdrawal. While increased cytokine expression did not have tumor cell intrinsic effects, presence of these cytokines in conditioned media led to increased attraction of microglia cells in vitro. CONCLUSION: Taken together, these data indicate rapid MAPK reactivation upon MAPKi withdrawal as a tumor cell intrinsic rebound-driving mechanism. Furthermore, increased secretion of microglia-recruiting cytokines may play a role in treatment response and rebound growth upon withdrawal, warranting further evaluation.


Assuntos
Neoplasias Encefálicas , Citocinas , Glioma , Microglia , Mutação , Inibidores de Proteínas Quinases , Proteínas Proto-Oncogênicas B-raf , Humanos , Proteínas Proto-Oncogênicas B-raf/genética , Proteínas Proto-Oncogênicas B-raf/metabolismo , Microglia/metabolismo , Microglia/efeitos dos fármacos , Glioma/metabolismo , Glioma/tratamento farmacológico , Glioma/patologia , Glioma/genética , Citocinas/metabolismo , Animais , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/tratamento farmacológico , Neoplasias Encefálicas/patologia , Neoplasias Encefálicas/genética , Inibidores de Proteínas Quinases/farmacologia , Ensaios Antitumorais Modelo de Xenoenxerto , Criança , Camundongos , Proliferação de Células/efeitos dos fármacos , Linhagem Celular Tumoral , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos
3.
Mol Oncol ; 17(1): 37-58, 2023 01.
Artigo em Inglês | MEDLINE | ID: mdl-36181342

RESUMO

Chemotherapy resistance is a persistent clinical problem in relapsed high-risk neuroblastomas. We tested a panel of 15 drugs for sensitization of neuroblastoma cells to the conventional chemotherapeutic vincristine, identifying tariquidar, an inhibitor of the transmembrane pump P-glycoprotein (P-gp/ABCB1), and the ERBB family inhibitor afatinib as the top resistance breakers. Both compounds were efficient in sensitizing neuroblastoma cells to vincristine in trypan blue exclusion assays and in inducing apoptotic cell death. The evaluation of ERBB signaling revealed no functional inhibition, that is, dephosphorylation of the downstream pathways upon afatinib treatment but direct off-target interference with P-gp function. Depletion of ABCB1, but not ERRB4, sensitized cells to vincristine treatment. P-gp inhibition substantially broke vincristine resistance in vitro and in vivo (zebrafish embryo xenograft). The analysis of gene expression datasets of more than 50 different neuroblastoma cell lines (primary and relapsed) and more than 160 neuroblastoma patient samples from the pediatric precision medicine platform INFORM (Individualized Therapy For Relapsed Malignancies in Childhood) confirmed a pivotal role of P-gp specifically in neuroblastoma resistance at relapse, while the ERBB family appears to play a minor part.


Assuntos
Membro 1 da Subfamília B de Cassetes de Ligação de ATP , Neuroblastoma , Animais , Humanos , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/genética , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/metabolismo , Vincristina/farmacologia , Afatinib , Peixe-Zebra/metabolismo , Resistencia a Medicamentos Antineoplásicos , Neuroblastoma/genética , Subfamília B de Transportador de Cassetes de Ligação de ATP/genética , Receptores ErbB/metabolismo , Recidiva , Linhagem Celular Tumoral
4.
Cancers (Basel) ; 14(3)2022 Feb 08.
Artigo em Inglês | MEDLINE | ID: mdl-35159116

RESUMO

The survival rate among children with relapsed tumors remains poor, due to tumor heterogeneity, lack of directly actionable tumor drivers and multidrug resistance. Novel personalized medicine approaches tailored to each tumor are urgently needed to improve cancer treatment. Current pediatric precision oncology platforms, such as the INFORM (INdividualized Therapy FOr Relapsed Malignancies in Childhood) study, reveal that molecular profiling of tumor tissue identifies targets associated with clinical benefit in a subgroup of patients only and should be complemented with functional drug testing. In such an approach, patient-derived tumor cells are exposed to a library of approved oncological drugs in a physiological setting, e.g., in the form of animal avatars injected with patient tumor cells. We used molecularly fully characterized tumor samples from the INFORM study to compare drug screen results of individual patient-derived cell models in functional assays: (i) patient-derived spheroid cultures within a few days after tumor dissociation; (ii) tumor cells reisolated from the corresponding mouse PDX; (iii) corresponding long-term organoid-like cultures and (iv) drug evaluation with the corresponding zebrafish PDX (zPDX) model. Each model had its advantage and complemented the others for drug hit and drug combination selection. Our results provide evidence that in vivo zPDX drug screening is a promising add-on to current functional drug screening in precision medicine platforms.

5.
NPJ Precis Oncol ; 6(1): 94, 2022 Dec 27.
Artigo em Inglês | MEDLINE | ID: mdl-36575299

RESUMO

The international precision oncology program INFORM enrolls relapsed/refractory pediatric cancer patients for comprehensive molecular analysis. We report a two-year pilot study implementing ex vivo drug sensitivity profiling (DSP) using a library of 75-78 clinically relevant drugs. We included 132 viable tumor samples from 35 pediatric oncology centers in seven countries. DSP was conducted on multicellular fresh tumor tissue spheroid cultures in 384-well plates with an overall mean processing time of three weeks. In 89 cases (67%), sufficient viable tissue was received; 69 (78%) passed internal quality controls. The DSP results matched the identified molecular targets, including BRAF, ALK, MET, and TP53 status. Drug vulnerabilities were identified in 80% of cases lacking actionable (very) high-evidence molecular events, adding value to the molecular data. Striking parallels between clinical courses and the DSP results were observed in selected patients. Overall, DSP in clinical real-time is feasible in international multicenter precision oncology programs.

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