Reversal of glucocorticoid resistance in paediatric acute lymphoblastic leukaemia is dependent on restoring BIM expression.

BackgroundAcute lymphoblastic leukaemia (ALL) is the most common paediatric malignancy. Glucocorticoids form a critical component of chemotherapy regimens and resistance to glucocorticoid therapy is predictive of poor outcome. We have previously shown that glucocorticoid resistance is associated with upregulation of the oncogene C-MYC and failure to induce the proapoptotic gene BIM.MethodsA high-throughput screening (HTS) campaign was carried out to identify glucocorticoid sensitisers against an ALL xenograft derived from a glucocorticoid-resistant paediatric patient. Gene expression analysis was carried out using Illumina microarrays. Efficacy, messenger RNA and protein analysis were carried out by Resazurin assay, reverse transcription-PCR and immunoblotting, respectively.ResultsA novel glucocorticoid sensitiser, 2-((4,5-dihydro-1H-imidazol-2-yl)thio)-N-isopropyl-N-phenylacetamide (GCS-3), was identified from the HTS campaign. The sensitising effect was specific to glucocorticoids and synergy was observed in a range of dexamethasone-resistant and dexamethasone-sensitive xenografts representative of B-ALL, T-ALL and Philadelphia chromosome-positive ALL. GCS-3 in combination with dexamethasone downregulated C-MYC and significantly upregulated BIM expression in a glucocorticoid-resistant ALL xenograft. The GCS-3/dexamethasone combination significantly increased binding of the glucocorticoid receptor to a novel BIM enhancer, which is associated with glucocorticoid sensitivity.ConclusionsThis study describes the potential of the novel glucocorticoid sensitiser, GCS-3, as a biological tool to interrogate glucocorticoid action and resistance

Circulating Tumor DNA in Pediatric Cancer

The measurement of circulating tumor DNA (ctDNA) has gained increasing prominence as a minimally invasive tool for the detection of cancer-specific markers in plasma. In adult cancers, ctDNA detection has shown value for disease-monitoring applications including tumor mutation profiling, risk stratification, relapse prediction, and treatment response evaluation. To date, there are ctDNA tests used as companion diagnostics for adult cancers and it is not understood why the same cannot be said about childhood cancer, despite the marked differences between adult and pediatric oncology. In this review, we discuss the current understanding of ctDNA as a disease monitoring biomarker in the context of pediatric malignancies, including the challenges associated with ctDNA detection in liquid biopsies. The data and conclusions from pediatric cancer studies of ctDNA are summarized, highlighting treatment response, disease monitoring and the detection of subclonal disease as applications of ctDNA. While the data from retrospective studies highlight the potential of ctDNA, large clinical trials are required for ctDNA analysis for routine clinical use in pediatric cancers. We outline the requirements for the standardization of ctDNA detection in pediatric cancers, including sample handling and reproducibility of results. With better understanding of the advantages and limitations of ctDNA and improved detection methods, ctDNA analysis may become the standard of care for patient monitoring in childhood cancers

Exploring the Metabolic Landscape of AML: From Haematopoietic Stem Cells to Myeloblasts and Leukaemic Stem Cells

Despite intensive chemotherapy regimens, up to 60% of adults with acute myeloid leukaemia (AML) will relapse and eventually succumb to their disease. Recent studies suggest that leukaemic stem cells (LSCs) drive AML relapse by residing in the bone marrow niche and adapting their metabolic profile. Metabolic adaptation and LSC plasticity are novel hallmarks of leukemogenesis that provide important biological processes required for tumour initiation, progression and therapeutic responses. These findings highlight the importance of targeting metabolic pathways in leukaemia biology which might serve as the Achilles’ heel for the treatment of AML relapse. In this review, we highlight the metabolic differences between normal haematopoietic cells, bulk AML cells and LSCs. Specifically, we focus on four major metabolic pathways dysregulated in AML; (i) glycolysis; (ii) mitochondrial metabolism; (iii) amino acid metabolism; and (iv) lipid metabolism. We then outline established and emerging drug interventions that exploit metabolic dependencies of leukaemic cells in the treatment of AML. The metabolic signature of AML cells alters during different biological conditions such as chemotherapy and quiescence. Therefore, targeting the metabolic vulnerabilities of these cells might selectively eradicate them and improve the overall survival of patients with AML

Patient-Specific Circulating Tumor DNA for Monitoring Response to Menin Inhibitor Treatment in Preclinical Models of Infant Leukemia

Background: In infant KMT2A (MLL1)-rearranged (MLL-r) acute lymphoblastic leukemia (ALL), early relapse and treatment response are currently monitored through invasive repeated bone marrow (BM) biopsies. Circulating tumor DNA (ctDNA) in peripheral blood (PB) provides a minimally invasive alternative, allowing for more frequent disease monitoring. However, a poor understanding of ctDNA dynamics has hampered its clinical translation. We explored the predictive value of ctDNA for detecting minimal/measurable residual disease (MRD) and drug response in a patient-derived xenograft (PDX) model of infant MLL-r ALL. Methods: Immune-deficient mice engrafted with three MLL-r ALL PDXs were monitored for ctDNA levels before and after treatment with the menin inhibitor SNDX-50469. Results: The amount of ctDNA detected strongly correlated with leukemia burden during initial engraftment prior to drug treatment. However, following SNDX-50469 treatment, the leukemic burden assessed by either PB leukemia cells through flow cytometry or ctDNA levels through droplet digital polymerase chain reaction (ddPCR) was discrepant. This divergence could be attributed to the persistence of leukemia cells in the spleen and BM, highlighting the ability of ctDNA to reflect disease dynamics in key leukemia infiltration sites. Conclusions: Notably, ctDNA analysis proved to be a superior predictor of MRD compared to PB assessment alone, especially in instances of low disease burden. These findings highlight the potential of ctDNA as a sensitive biomarker for monitoring treatment response and detecting MRD in infant MLL-r ALL

The Promise of Single-cell Technology in Providing New Insights into the Molecular Heterogeneity and Management of Acute Lymphoblastic Leukemia

Drug resistance and treatment failure in pediatric acute lymphoblastic leukemia (ALL) are in part driven by tumor heterogeneity and clonal evolution. Although bulk tumor genomic analyses have provided some insight into these processes, single-cell sequencing has emerged as a powerful technique to profile individual cells in unprecedented detail. Since the introduction of single-cell RNA sequencing, we now have the capability to capture not only transcriptomic, but also genomic, epigenetic, and proteomic variation between single cells separately and in combination. This rapidly evolving field has the potential to transform our understanding of the fundamental biology of pediatric ALL and guide the management of ALL patients to improve their clinical outcome. Here, we discuss the impact single-cell sequencing has had on our understanding of tumor heterogeneity and clonal evolution in ALL and provide examples of how single-cell technology can be integrated into the clinic to inform treatment decisions for children with high-risk disease

Inhibition of the NLRP3 inflammasome using MCC950 reduces vincristine-induced adverse effects in an acute lymphoblastic leukemia patient-derived xenograft model

Vincristine is one of the most important chemotherapeutic drugs used to treat acute lymphoblastic leukemia (ALL). Unfortunately, vincristine often causes severe adverse effects, including sensory–motor neuropathies, weight loss, and overall decreased well-being, that are difficult to control and that decrease the quality of life and survival of patients. Recent studies demonstrate that sensory–motor adverse effects of vincristine are driven by neuroinflammatory processes, including the activation of the Nod-like receptor 3 (NLRP3) inflammasome. In this study, we aimed to test the effects of MCC950, a specific NLRP3 inhibitor, on the prevention of vincristine-induced adverse effects as well as tumor progression and vincristine efficacy in NOD/SCID/interleukin-2 receptor γ-negative mice patient-derived xenografts of ALL. We demonstrate that co-administration of MCC950 effectively prevented the development of mechanical allodynia, motor impairment, and weight loss and significantly improved the overall well-being of the animals without negatively impacting the in vivo efficacy of vincristine as a single agent or in combination with standard-of-care drugs. These results provide proof of principle that the adverse effects of vincristine chemotherapy can be prevented using NLRP3 inflammasome inhibitors and provide new options for the development of effective treatment strategies

A Social Identity Approach to Sport Psychology: Principles, Practice, and Prospects.

Drawing on social identity theory and self-categorization theory, we outline an approach to sport psychology that understands groups not simply as features of sporting contexts but rather as elements that can be, and often are, incorporated into a person's sense of self and, through this, become powerful determinants of their sport-related behavior. The underpinnings of this social identity approach are outlined, and four key lessons for sport that are indicative of the analytical and practical power of the approach are presented. These suggest that social identity is the basis for sports group (1) behavior, (2) formation and development, (3) support and stress appraisal, and (4) leadership. Building on recent developments within sport science, we outline an agenda for future research by identifying a range of topics to which the social identity approach could fruitfully contribute

In vivo activity of the dual PI3Kδ and PI3Kγ inhibitor duvelisib against pediatric acute lymphoblastic leukemia xenografts

Background: Acute lymphoblastic leukemia (ALL) remains one of the most common causes of cancer-related mortality in children. Phosphoinositide 3-kinases (PI3Ks) are a family of lipid kinases, and aberrations in the PI3K pathway are associated with several hematological malignancies, including ALL. Duvelisib (Copiktra) is an orally available, small molecule dual inhibitor of PI3Kδ and PI3Kγ, that is Food and Drug Administration (FDA) approved for the treatment of relapsed/refractory chronic lymphocytic leukemia and small lymphocytic lymphoma. Here, we report the efficacy of duvelisib against a panel of pediatric ALL patient-derived xenografts (PDXs). Procedures: Thirty PDXs were selected for a single mouse trial based on PI3Kδ (PIK3CD) and PI3Kγ (PIK3CG) expression and mutational status. PDXs were grown orthotopically in NSG (NOD.Cg-PrkdcscidIL2rgtm1Wjl/SzJAusb) mice, and engraftment was evaluated by enumerating the proportion of human versus mouse CD45+ cells (%huCD45+) in the peripheral blood. Treatment commenced when the %huCD45+ reached greater than or equal to 1%, and events were predefined as %huCD45+ greater than or equal to 25% or leukemia-related morbidity. Duvelisib was administered per oral (50 mg/kg, twice daily for 28 days). Drug efficacy was assessed by event-free survival and stringent objective response measures. Results: PI3Kδ and PI3Kγ mRNA expression was significantly higher in B-lineage than T-lineage ALL PDXs (p-values <.0001). Duvelisib was well-tolerated and reduced leukemia cells in the peripheral blood in four PDXs, but with only one objective response. There was no obvious relationship between duvelisib efficacy and PI3Kδ or PI3Kγ expression or mutation status, nor was the in vivo response to duvelisib subtype dependent. Conclusions: Duvelisib demonstrated limited in vivo activity against ALL PDXs

Systemic AAV vectors for widespread and targeted gene delivery in rodents

We recently developed adeno-associated virus (AAV) capsids to facilitate efficient and noninvasive gene transfer to the central and peripheral nervous systems. However, a detailed protocol for generating and systemically delivering novel AAV variants was not previously available. In this protocol, we describe how to produce and intravenously administer AAVs to adult mice to specifically label and/or genetically manipulate cells in the nervous system and organs, including the heart. The procedure comprises three separate stages: AAV production, intravenous delivery, and evaluation of transgene expression. The protocol spans 8 d, excluding the time required to assess gene expression, and can be readily adopted by researchers with basic molecular biology, cell culture, and animal work experience. We provide guidelines for experimental design and choice of the capsid, cargo, and viral dose appropriate for the experimental aims. The procedures outlined here are adaptable to diverse biomedical applications, from anatomical and functional mapping to gene expression, silencing, and editing