RESUMEN
Recurrence of solid tumors renders patients vulnerable to advanced, treatment-refractory disease state with mutational and oncogenic landscape distinctive from initial diagnosis. Improving outcomes for recurrent cancers requires a better understanding of cell populations that expand from the post-therapy, minimal residual disease (MRD) state. We profile barcoded tumor stem cell populations through therapy at tumor initiation, MRD, and recurrence in our therapy-adapted, patient-derived xenograft models of glioblastoma (GBM). Tumors show distinct patterns of recurrence in which clonal populations exhibit either a pre-existing fitness advantage or an equipotency fitness acquired through therapy. Characterization of the MRD state by single-cell and bulk RNA sequencing reveals a tumor-intrinsic immunomodulatory signature with prognostic significance at the transcriptomic level and in proteomic analysis of cerebrospinal fluid (CSF) collected from patients with GBM. Our results provide insight into the innate and therapy-driven dynamics of human GBM and the prognostic value of interrogating the MRD state in solid cancers.
Asunto(s)
Neoplasias Encefálicas , Glioblastoma , Neoplasias Encefálicas/patología , Glioblastoma/genética , Glioblastoma/patología , Humanos , Recurrencia Local de Neoplasia/genética , Recurrencia Local de Neoplasia/patología , Neoplasia Residual/genética , Células Madre Neoplásicas/patología , ProteómicaRESUMEN
Advances in chimeric antigen receptor (CAR) T cell therapies have led to the modality dominating translational cancer research; however, a standardized protocol for evaluating such therapies in vitro is needed. This protocol details the in vitro preclinical evaluation of CAR-T cell therapies for glioblastoma (GBM), including target cell cytotoxicity and T cell proliferation, activation, and cytokine release assays. For complete details on the use and execution of this protocol, please refer to Vora et al. (2020).
Asunto(s)
Neoplasias Encefálicas , Técnicas de Cocultivo/métodos , Glioblastoma , Inmunoterapia Adoptiva , Modelos Biológicos , Neoplasias Encefálicas/patología , Neoplasias Encefálicas/terapia , Glioblastoma/patología , Glioblastoma/terapia , Humanos , Receptores Quiméricos de Antígenos/inmunología , Linfocitos T/citología , Linfocitos T/inmunología , Células Tumorales CultivadasRESUMEN
Despite aggressive multimodal therapy, glioblastoma (GBM) remains the most common malignant primary brain tumor in adults. With the advent of therapies that revitalize the anti-tumor immune response, several immunotherapeutic modalities have been developed for treatment of GBM. In this review, we summarize recent clinical and preclinical efforts to evaluate vaccination strategies, immune checkpoint inhibitors (ICIs) and chimeric antigen receptor (CAR) T cells. Although these modalities have shown long-term tumor regression in subsets of treated patients, the underlying biology that may predict efficacy and inform therapy development is being actively investigated. Common to all therapeutic modalities are fundamental mechanisms of therapy evasion by tumor cells, including immense intratumoral heterogeneity, suppression of the tumor immune microenvironment and low mutational burden. These insights have led efforts to design rational combinatorial therapies that can reignite the anti-tumor immune response, effectively and specifically target tumor cells and reliably decrease tumor burden for GBM patients.
RESUMEN
Despite a surge in the preclinical development of immunotherapies, current models are unable to predict putative toxicity, particularly the "on-target, off-tumor" effects of these therapeutics. To address this gap, we used a humanized mouse model of hematopoiesis to examine the toxicity profile of CAR-Ts targeting brain tumor-antigens also expressed in the hematopoietic system. In assessing the safety of cell-based therapies, we aim to develop and integrate a preclinical evaluation protocol as a necessary step in the clinical development pathway. For complete details on the use and execution of this protocol, please refer to Vora et al. (2020).
Asunto(s)
Neoplasias Encefálicas/terapia , Hematopoyesis/inmunología , Inmunoterapia Adoptiva/métodos , Animales , Neoplasias Encefálicas/inmunología , Hematopoyesis/fisiología , Células Madre Hematopoyéticas/inmunología , Humanos , Inmunoterapia/métodos , Ratones , Modelos Animales , Receptores Quiméricos de Antígenos/inmunologíaRESUMEN
Glioblastoma (GBM) is the most common malignant adult brain tumor that is resistant to the standard care therapy. Advances in chimeric antigen receptor (CAR) T cell therapies have spurred renewed interest in developing CAR T cell therapies to target chemoradiotherapy-resistant brain tumor-initiating cells. This protocol shows how to isolate peripheral blood mononuclear cells from healthy donors and generate CAR T cells for the antigens of interest, and how to intracranially inject the CAR T cells into a patient-derived xenograft mouse model of GBM. For complete details on the use and execution of this protocol, please refer to Vora et al. (2020).
Asunto(s)
Glioblastoma/inmunología , Receptores Quiméricos de Antígenos/metabolismo , Linfocitos T/inmunología , Ensayos Antitumor por Modelo de Xenoinjerto/métodos , Puntos Anatómicos de Referencia , Animales , Proliferación Celular , Células HEK293 , Humanos , Lentivirus/fisiología , Activación de Linfocitos/inmunología , Ratones , Células Madre Neoplásicas/patología , Plásmidos/metabolismoRESUMEN
BACKGROUND: Central nervous system tumors remain the leading cause of cancer-related mortality amongst children with solid tumors, with medulloblastoma (MB) representing the most common pediatric brain malignancy. Despite best current therapies, patients with recurrent MB experience have an alarmingly high mortality rate and often have limited therapeutic options beyond inadequate chemotherapy or experimental clinical trials. Therefore, a systematic review of the literature regarding treatment strategies employed in recurrent pediatric MB will evaluate previous salvage therapies in order to guide future clinical trials. The aim of this systematic review will be to investigate the efficacy and safety of salvage therapies for the management of children with progressive, treatment-refractory, or recurrent MB. METHODS: We will conduct literature searches (from 1995 onwards) in MEDLINE, EMBASE, ClinicalTrials.gov, WHO International Clinical Trials Registry Platform, and Cochrane Central Register of Controlled Trials. Studies examining the survival and toxicity of therapies administered to treatment-refractory pediatric MB patients will be included. Two reviewers will independently assess the search results based on predefined selection criteria, complete data abstraction, and quality assessment. The primary outcomes of this review will be overall and progression-free survival. Secondary outcomes will include safety and toxicity of each therapy administered. The study methodological quality (or bias) will be appraised using an appropriate tool. Due to the nature of the research question and published literature, we expect large inter-study heterogeneity and therefore will use random effects regression analysis to extract the combined effect. In additional analyses, we will investigate the role of re-irradiation and mono- vs. poly-therapy in recurrent disease, and whether molecular subgrouping of MB influences salvage therapy. DISCUSSION: This systematic review will provide an overview of the current literature regarding salvage therapies for relapsed MB patients. Investigation of clinically tested therapies for children with recurrent MB has significant implications for clinical practice. By reviewing the efficacy and toxicity of MB salvage therapies, this study will identify effective therapeutic strategies administered to recurrent MB patients and can inform future clinical trials aimed to improve patient survivorship and quality of life. SYSTEMATIC REVIEW REGISTRATION: PROSPERO CRD42020167421.
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Neoplasias Encefálicas , Neoplasias Cerebelosas , Meduloblastoma , Neoplasias Cerebelosas/terapia , Niño , Humanos , Meduloblastoma/terapia , Calidad de Vida , Terapia Recuperativa , Revisiones Sistemáticas como AsuntoRESUMEN
Differentiation is a central key capability of stem cells. Their ability to be multipotent and undergo self-renewal are key identifying features of stem cells. A differentiation assay allows for study of one of the essential features of stem cells, the ability to differentiate into all of the cell types of its lineage, in order to ensure that the cells cultured and utilized in key experiments indeed have stem cell properties. Neural stem cells when plated in differentiation media, differentiate into all three neural lineages: Neurons, Astrocytes, and Oligodendrocytes. Brain tumor initiating cells (BTICs) are cells present in brain tumors that possess stem cell properties and are able to self-renew and differentiate into neural lineages. In the current chapter, we discuss protocols involved in immunofluorescence staining and identification of differentiated cells from BTIC populations.