High throughput sequencing in acute lymphoblastic leukemia reveals clonal architecture of central nervous system and bone marrow compartments

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Flow cytometry of bone marrow aspirates from neuroblastoma patients is a highly sensitive technique for quantification of low-level neuroblastoma [version 2; peer review: 2 approved]

Background: Bone marrow involvement is an important aspect of determining staging of disease and treatment for childhood neuroblastoma. Current standard of care relies on microscopic examination of bone marrow trephine biopsies and aspirates respectively, to define involvement. Flow cytometric analysis of disaggregated tumour cells, when using a panel of neuroblastoma specific markers, allows for potentially less subjective determination of the presence of tumour cells. Methods: A retrospective review of sequential bone marrow trephine biopsies and aspirates, performed at Great Ormond Street Hospital, London, between the years 2015 and 2018, was performed to assess whether the addition of flow cytometric analysis to these standard of care methods provided concordant or additional information. Results: There was good concurrence between all three methods for negative results 216/302 (72%). Positive results had a concordance of 52/86 (61%), comparing samples positive by flow cytometry and positive by either or both cytology and histology. Of the remaining samples, 20/86 (23%) were positive by either or both cytology and histology, but negative by flow cytometry. Whereas 14/86 (16%) of samples were positive only by flow cytometry. Conclusions: Our review highlights the ongoing importance of expert cytological and histological assessment of bone marrow results. Flow cytometry is an objective, quantitative method to assess the level of bone marrow disease in aspirates. In this study, flow cytometry identified low-level residual disease that was not detected by cytology or histology. The clinical significance of this low-level disease warrants further investigation

Identification of a c-MYB-directed therapeutic for acute myeloid leukemia

A significant proportion of patients suffering from acute myeloid leukemia (AML) cannot be cured by conventional chemotherapy, relapsed disease being a common problem. Molecular targeting of essential oncogenic mediators is an attractive approach to improving outcomes for this disease. The hematopoietic transcription factor c-MYB has been revealed as a central component of complexes maintaining aberrant gene expression programs in AML. We have previously screened the Connectivity Map database to identify mebendazole as an anti-AML therapeutic targeting c-MYB. In the present study we demonstrate that another hit from this screen, the steroidal lactone withaferin A (WFA), induces rapid ablation of c-MYB protein and consequent inhibition of c-MYB target gene expression, loss of leukemia cell viability, reduced colony formation and impaired disease progression. Although WFA has been reported to have pleiotropic anti-cancer effects, we demonstrate that its anti-AML activity depends on c-MYB modulation and can be partially reversed by a stabilized c-MYB mutant. c-MYB ablation results from disrupted HSP/HSC70 chaperone protein homeostasis in leukemia cells following induction of proteotoxicity and the unfolded protein response by WFA. The widespread use of WFA in traditional medicines throughout the world indicates that it represents a promising candidate for repurposing into AML therapy

Novel Glycomimetics Protect Against Glycated Low-Density Lipoprotein-Induced Vascular Calcification In Vitro by Attenuation of the RAGE/ERK/CREB Pathway

Heparan sulphate (HS) can act as a co-receptor on the cell surface and alterations in this process underpin many pathological conditions. We have previously described the usefulness of mimics of HS (glycomimetics) in protection against β-glycerophosphate-induced vascular calcification and in the restoration of the functional capacity of diabetic endothelial colony-forming cells in vitro. This study aims to investigate whether our novel glycomimetic compounds can attenuate glycated low-density lipoprotein (g-LDL)-induced calcification by inhibiting RAGE signalling within the context of critical limb ischemia (CLI). We used an established osteogenic in vitro vascular smooth muscle cell (VSMC) model. Osteoprotegerin (OPG), sclerostin and glycation levels were all significantly increased in CLI serum compared to healthy controls, while the vascular calcification marker osteocalcin (OCN) was down-regulated in CLI patients vs. controls. Incubation with both CLI serum and g-LDL (10 µg/mL) significantly increased VSMC calcification vs. controls after 21 days, with CLI serum-induced calcification apparent after only 10 days. Glycomimetics (C2 and C3) significantly inhibited g-LDL and CLI serum-induced mineralisation, as shown by a reduction in alizarin red (AR) staining and alkaline phosphatase (ALP) activity. Furthermore, secretion of the osteogenic marker OCN was significantly reduced in VSMCs incubated with CLI serum in the presence of glycomimetics. Phosphorylation of cyclic AMP response element-binding protein (CREB) was significantly increased in g-LDL-treated cells vs. untreated controls, which was attenuated with glycomimetics. Blocking CREB activation with a pharmacological inhibitor 666-15 replicated the protective effects of glycomimetics, evidenced by elevated AR staining. In silico molecular docking simulations revealed the binding affinity of the glycomimetics C2 and C3 with the V domain of RAGE. In conclusion, these findings demonstrate that novel glycomimetics, C2 and C3 have potent anti-calcification properties in vitro, inhibiting both g-LDL and CLI serum-induced VSMC mineralisation via the inhibition of LDLR, RAGE, CREB and subsequent expression of the downstream osteogenic markers, ALP and OCN

The Clinicogenomic Landscape of Induction Failure in Childhood and Young Adult T-Cell Acute Lymphoblastic Leukemia

PURPOSE: Failure to respond to induction chemotherapy portends a poor outcome in childhood acute lymphoblastic leukemia (ALL) and is more frequent in T-cell ALL (T-ALL) than B-cell ALL. We aimed to address the limited understanding of clinical and genetic factors that influence outcome in a cohort of patients with T-ALL induction failure (IF).METHODS: We studied all cases of T-ALL IF on two consecutive multinational randomized trials, UKALL2003 and UKALL2011, to define risk factors, treatment, and outcomes. We performed multiomic profiling to characterize the genomic landscape.RESULTS: IF occurred in 10.3% of cases and was significantly associated with increasing age, occurring in 20% of patients age 16 years and older. Five-year overall survival (OS) rates were 52.1% in IF and 90.2% in responsive patients (P &lt; .001). Despite increased use of nelarabine-based chemotherapy consolidated by hematopoietic stem-cell transplant in UKALL2011, there was no improvement in outcome. Persistent end-of-consolidation molecular residual disease resulted in a significantly worse outcome (5-year OS, 14.3% v 68.5%; HR, 4.10; 95% CI, 1.35 to 12.45; P = .0071). Genomic profiling revealed a heterogeneous picture with 25 different initiating lesions converging on 10 subtype-defining genes. There was a remarkable abundance of TAL1 noncoding lesions, associated with a dismal outcome (5-year OS, 12.5%). Combining TAL1 lesions with mutations in the MYC and RAS pathways produces a genetic stratifier that identifies patients highly likely to fail conventional therapy (5-year OS, 23.1% v 86.4%; HR, 6.84; 95% CI, 2.78 to 16.78; P &lt; .0001) and who should therefore be considered for experimental agents.CONCLUSION: The outcome of IF in T-ALL remains poor with current therapy. The lack of a unifying genetic driver suggests alternative approaches, particularly using immunotherapy, are urgently needed.</p

The Clinicogenomic Landscape of Induction Failure in Childhood and Young Adult T-Cell Acute Lymphoblastic Leukemia

PURPOSE: Failure to respond to induction chemotherapy portends a poor outcome in childhood acute lymphoblastic leukemia (ALL) and is more frequent in T-cell ALL (T-ALL) than B-cell ALL. We aimed to address the limited understanding of clinical and genetic factors that influence outcome in a cohort of patients with T-ALL induction failure (IF). METHODS: We studied all cases of T-ALL IF on two consecutive multinational randomized trials, UKALL2003 and UKALL2011, to define risk factors, treatment, and outcomes. We performed multiomic profiling to characterize the genomic landscape. RESULTS: IF occurred in 10.3% of cases and was significantly associated with increasing age, occurring in 20% of patients age 16 years and older. Five-year overall survival (OS) rates were 52.1% in IF and 90.2% in responsive patients (P < .001). Despite increased use of nelarabine-based chemotherapy consolidated by hematopoietic stem-cell transplant in UKALL2011, there was no improvement in outcome. Persistent end-of-consolidation molecular residual disease resulted in a significantly worse outcome (5-year OS, 14.3% v 68.5%; HR, 4.10; 95% CI, 1.35 to 12.45; P = .0071). Genomic profiling revealed a heterogeneous picture with 25 different initiating lesions converging on 10 subtype-defining genes. There was a remarkable abundance of TAL1 noncoding lesions, associated with a dismal outcome (5-year OS, 12.5%). Combining TAL1 lesions with mutations in the MYC and RAS pathways produces a genetic stratifier that identifies patients highly likely to fail conventional therapy (5-year OS, 23.1% v 86.4%; HR, 6.84; 95% CI, 2.78 to 16.78; P < .0001) and who should therefore be considered for experimental agents. CONCLUSION: The outcome of IF in T-ALL remains poor with current therapy. The lack of a unifying genetic driver suggests alternative approaches, particularly using immunotherapy, are urgently needed

Endovascular strategy or open repair for ruptured abdominal aortic aneurysm: one-year outcomes from the IMPROVE randomized trial.

AIMS: To report the longer term outcomes following either a strategy of endovascular repair first or open repair of ruptured abdominal aortic aneurysm, which are necessary for both patient and clinical decision-making. METHODS AND RESULTS: This pragmatic multicentre (29 UK and 1 Canada) trial randomized 613 patients with a clinical diagnosis of ruptured aneurysm; 316 to an endovascular first strategy (if aortic morphology is suitable, open repair if not) and 297 to open repair. The principal 1-year outcome was mortality; secondary outcomes were re-interventions, hospital discharge, health-related quality-of-life (QoL) (EQ-5D), costs, Quality-Adjusted-Life-Years (QALYs), and cost-effectiveness [incremental net benefit (INB)]. At 1 year, all-cause mortality was 41.1% for the endovascular strategy group and 45.1% for the open repair group, odds ratio 0.85 [95% confidence interval (CI) 0.62, 1.17], P = 0.325, with similar re-intervention rates in each group. The endovascular strategy group and open repair groups had average total hospital stays of 17 and 26 days, respectively, P < 0.001. Patients surviving rupture had higher average EQ-5D utility scores in the endovascular strategy vs. open repair groups, mean differences 0.087 (95% CI 0.017, 0.158), 0.068 (95% CI -0.004, 0.140) at 3 and 12 months, respectively. There were indications that QALYs were higher and costs lower for the endovascular first strategy, combining to give an INB of £3877 (95% CI £253, £7408) or €4356 (95% CI €284, €8323). CONCLUSION: An endovascular first strategy for management of ruptured aneurysms does not offer a survival benefit over 1 year but offers patients faster discharge with better QoL and is cost-effective. CLINICAL TRIAL REGISTRATION: ISRCTN 48334791

The effect of aortic morphology on peri-operative mortality of ruptured abdominal aortic aneurysm

Aims To investigate whether aneurysm shape and extent, which indicate whether a patient with ruptured abdominal aortic aneurysm (rAAA) is eligible for endovascular repair (EVAR), influence the outcome of both EVAR and open surgical repair. Methods and results The influence of six morphological parameters (maximum aortic diameter, aneurysm neck diameter, length and conicality, proximal neck angle, and maximum common iliac diameter) on mortality and reinterventions within 30 days was investigated in rAAA patients randomized before morphological assessment in the Immediate Management of the Patient with Rupture: Open Versus Endovascular strategies (IMPROVE) trial. Patients with a proven diagnosis of rAAA, who underwent repair and had their admission computerized tomography scan submitted to the core laboratory, were included. Among 458 patients (364 men, mean age 76 years), who had either EVAR (n = 177) or open repair (n = 281) started, there were 155 deaths and 88 re-interventions within 30 days of randomization analysed according to a pre-specified plan. The mean maximum aortic diameter was 8.6 cm. There were no substantial correlations between the six morphological variables. Aneurysm neck length was shorter in those undergoing open repair (vs. EVAR). Aneurysm neck length (mean 23.3, SD 16.1 mm) was inversely associated with mortality for open repair and overall: adjusted OR 0.72 (95% CI 0.57, 0.92) for each 16 mm (SD) increase in length. There were no convincing associations of morphological parameters with reinterventions. Conclusion Short aneurysm necks adversely influence mortality after open repair of rAAA and preclude conventional EVAR. This may help explain why observational studies, but not randomized trials, have shown an early survival benefit for EVAR. Clinical trial registration: ISRCTN 48334791

Risk assessment models for the development of complications in Maltese type 2 diabetic patients

Introduction: With the IDF Diabetes Atlas 2006 predicting a Type 2 diabetes incidence rate of 11.6% among the Maltese population by 2025, treatment differentiation between high risk and low risk patients is necessary to ensure the sustainability of such a diabetes management program. Objectives: To identify significant predictors and develop local diabetic neuropathy (DNeurM), retinopathy (DRM), nephropathy (DNephrM) and macrovascular (MVM) models which determine complication risk in Maltese diabetic patients. Methods: A cross-sectional retrospective study involving 120 randomly selected patients aged 25-70 years, diagnosed with type 2 diabetes ≤ 1 year and taking metformin 500 mg, perindopril 5 mg and simvastatin 40 mg was carried out at the Endocrine and Diabetes Centre at Mater Dei General Hospital in Malta to collect data for 20 predictors. Complication risk scores were assigned to participants using a developed risk scale. SPSS® 17.0 ANCOVA regression model analyses and backward elimination variable selection method (p<0.05) were used to derive parsimonious models. Results: 12 significant predictors were retained in the models; DNeurM includes body mass index (BMI; p=1×10-4), glycated haemoglobin (HbA1c) level (p=0.00019), serum fasting triglycerides (p=0.002), alcohol abuse (No; p=0.022), systolic blood pressure (BP; p=0.041) and age (p=0.070); DRM includes systolic BP (p=4×10-4), serum fasting triglycerides (p=0.001), HbA1c level (p=0.010), albumin-creatinine ratio (ACR; p=0.040) and waist circumference (p=0.095); DNephrM includes systolic BP (p=3×10-7), urinary glucose (p=3.86×10-4), ACR (p=0.0009), waist circumference (p=0.0012), age (p=0.006), genetic predisposition (No; p=0.026), serum urea (p=0.050) and serum fasting triglycerides (p=0.062); MVM includes waist circumference (p=1×10-6), systolic BP (p=0.0003), total serum cholesterol (p=0.011) and HbA1c level (p=0.060). Conclusion: Twelve significant predictors featured in the parsimonious models: age, genetic predisposition, alcohol abuse, BMI, waist circumference, systolic BP, HbA1c level, serum total cholesterol level, serum fasting triglyceride level, serum urea level, urinary glucose level and ACR.peer-reviewe