The ALK inhibitor PF-06463922 is effective as a single agent in neuroblastoma driven by expression of ALK and MYCN.

The first-in-class inhibitor of ALK, c-MET and ROS1, crizotinib (Xalkori), has shown remarkable clinical efficacy in treatment of ALK-positive non-small cell lung cancer. However, in neuroblastoma, activating mutations in the ALK kinase domain are typically refractory to crizotinib treatment, highlighting the need for more potent inhibitors. The next-generation ALK inhibitor PF-06463922 is predicted to exhibit increased affinity for ALK mutants prevalent in neuroblastoma. We examined PF-06463922 activity in ALK-driven neuroblastoma models in vitro and in vivo In vitro kinase assays and cell-based experiments examining ALK mutations of increasing potency show that PF-06463922 is an effective inhibitor of ALK with greater activity towards ALK neuroblastoma mutants. In contrast to crizotinib, single agent administration of PF-06463922 caused dramatic tumor inhibition in both subcutaneous and orthotopic xenografts as well as a mouse model of high-risk neuroblastoma driven by Th-ALK(F1174L)/MYCN Taken together, our results suggest PF-06463922 is a potent inhibitor of crizotinib-resistant ALK mutations, and highlights an important new treatment option for neuroblastoma patients.

Vein visualization: patient characteristic factors and efficacy of a new infrared vein finder technology.

BACKGROUND: We investigated the patient characteristic factors that correlate with identification of i.v. cannulation sites with normal eyesight. We evaluated a new infrared vein finding (VF) technology device in identifying i.v. cannulation sites. METHODS: Each subject underwent two observations: one using the conventional method (CM) of normal, unassisted eyesight and the other with the infrared VF device, VueTek's Veinsite™ (VF). A power analysis for moderate effect size (ß=0.95) required 54 samples for within-subject differences. RESULTS: Patient characteristic profiles were obtained from 384 subjects (768 observations). Our sample population exhibited an overall average of 5.8 [95% confidence interval (CI) 5.4-6.2] veins using CM. As a whole, CM vein visualization were less effective among obese [4.5 (95% CI 3.8-5.3)], African-American [4.6 (95% CI 3.6-5.5 veins)], and Asian [5.1 (95% CI 4.1-6.0)] subjects. Next, the VF technology identified an average of 9.1 (95% CI 8.6-9.5) possible cannulation sites compared with CM [average of 5.8 (95% CI 5.4-6.2)]. Seventy-six obese subjects had an average of 4.5 (95% CI 3.8-5.3) and 8.2 (95% CI 7.4-9.1) veins viewable by CM and VF, respectively. In dark skin subjects, 9.1 (95% CI 8.3-9.9) veins were visible by VF compared with 5.4 (95% CI 4.8-6.0) with CM. CONCLUSIONS: African-American or Asian ethnicity, and obesity were associated with decreased vein visibility. The visibility of veins eligible for cannulation increased for all subgroups using a new infrared device.

Stress-induced enhancement of activity of lymphocyte lysosomal enzymes in pigs of different stress-susceptibility.

To evaluate a possible mechanism of stress-induced lymphopenic effect we assessed the activity of lymphocyte lysosomal enzymes (LE) under immobilization. The effects of immobilization stress on LE (AP, acid phosphatase, cathepsin D and L, beta-N-acetyl-glucosamidase) activity in lymphocytes, number of lymphocytes and plasma cortisol (COR) level in the peripheral blood were examined in the cross-bred Pietrain pigs showing genotypic (presence or lack of RyR1 gene mutation) and phenotypic (reactivity to halothane) differences. It was found that immobilization stress evoked an increase in LE which was concomitant with lymphopenia and a rise of COR level. The most pronounced enhancement of LE, which may reflect a tendency to lymphocyte cytolysis, was found in the recessive homozygotes RyR1 (nn) phenotypically defined as stress/halothane susceptible as well as in the heterozygotes RyR1 (Nn) included in the group of stress/halothane resistant. Despite this individual variability the stress-induced increase in LE activity was present in all the animals. It seems that a possibility of destruction (lysis) of lymphocyte cells should not be excluded as one of the causes of stress lymphopenia.

The insulin, glucose and cholesterol level and activity of lysosomal enzymes in the course of the model alloxan diabetes.

OBJECTIVES: The study was carried out on fifty male rabbits of the New Zealand White breed. Diabetes was caused by a single, intravenous alloxan injection. Rabbits which had glycaemia 7th day after the alloxan administration higher than 11 millimol/litre were selected for the studies. They were divided into 5 groups: I - control (without diabetes); II - 3-week diabetes; III - 6-week diabetes; IV - 3-month diabetes; V - 6-month diabetes. METHODS: In control and experimental rabbits the activity of beta-glucuronidase, N-acetyl-beta-glucosaminidase, lysosomal acid phosphatase, leucine aminopeptidase, cathepsin D, and lysosomal arylesterase was determined in lysosomal fractions of the liver and kidney. RESULTS: Alloxan caused lowering of the activity of all the investigated enzymes in the kidney and liver except lysosomal arylesterase. CONCLUSION: Alloxan injection caused a significant increase in the activity of all the investigated enzymes. The advisable lysosomal enzymes may be useful for the monitoring of the course and effectiveness of diabetes therapy.

Effect of melatonin administration on activities of some lysosomal enzymes in the mouse.

OBJECTIVES: Changes in the activity of beta-glucuronidase, N-acetyl-beta-glucosaminidase, cathepsin D and L, alanine aminopeptidase and lysosomal acid lipase in lysosomal fractions of the liver and kidneys of mice, which were administered 20 mg/kg b.w. of exogenous melatonin (N-acetyl-5-methoxytryptamine) for 7 and 14 days were investigated. METHODS: The slices of the liver and kidney were homogenized in 0.1M phosphate buffer, pH 7.0. Homogenates were subjected to differentiated centrifuging and determination of studied enzymes. RESULTS: Melatonin caused lowering of the activity of all the investigated lysosomal enzymes in the liver and kidney. CONCLUSION: Administration of melatonin was caused the lowering of the activity of the investigated lysosomal enzymes in comparison with values in control groups.

Epidermal growth factor and pro-inflammatory cytokines regulate the expression of components of plasminogen activation system in U373-MG astrocytoma cells.

Cytokines and growth factors that influence both secretion of the extracellular matrix (ECM) proteins and migration of the cells decide about the final outcome of tissue remodelling. We have examined expression of the components of the plasminogen activation system in human astrocytoma U373-MG cells and found that interleukin 1beta (IL-1beta), tumour necrosis factor alpha TNF-alpha), interferon gamma (INF-gamma) and epidermal growth factor (EGF) specifically regulate the expression of tissue-type plasminogen activator (t-PA), urokinase-type plasminogen activator (u-PA), plasminogen activator inhibitor type 1 (PAI-1) and protease nexin-1 (PN-1). We conclude that EGF and IFN-gamma are new important regulators of the plasminogen activation system in astrocytoma cells and, therefore, may influence turnover of extracellular matrix and migration of cells within the brain.

Reprogramming of TIMP-1 and TIMP-3 expression profiles in brain microvascular endothelial cells and astrocytes in response to proinflammatory cytokines.

Cytokine-dependent regulation of tissue inhibitors of metalloproteinases (TIMPs) expression provides an important mechanism for controlling the activity of matrix metalloproteinases. We present data indicating that during inflammatory processes TIMP-1 and TIMP-3 may be involved in the proteolytic remodeling of subendothelial basement membrane of the brain microvascular system, a key step during leukocyte migration into the brain perivascular tissue. In brain endothelial cells the expression of TIMP-1 is dramatically up-regulated by major proinflammatory cytokines, with the combination of interleukin-1beta (IL-1beta) and tumor necrosis factor-alpha (TNF alpha) exhibiting the strongest synergistic stimulation. Simultaneously, IL-1beta/TNF alpha almost completely blocks TIMP-3 expression. Both synergistic effects are dose-dependent within the concentration range 0.05-5 ng/ml of both cytokines and correlate with the expression of inducible nitric oxide synthase, an endothelial cell activation marker. Down-regulation of TIMP-3 expression is also detected in astrocytes treated with TNF alpha or IFN-gamma whereas oncostatin M as well as TNF alpha up-regulate TIMP-1 mRNA level. We propose that the cytokine-modified balance between TIMP-1 and TIMP-3 expression provides a potential mechanism involved in the regulation of microvascular basement membrane proteolysis.

Effect of ethanol administration on activities of some lysosomal hydrolases in the mouse.

1. The studies were carried on 98 random 6-week-old male mice. The mice were divided into control and experimental groups. The experimental animals were given 10% and 20% ethanol solution daily for 7, 14 and 28 days. 2. In the lysosomal fractions of the liver, kidneys, muscle and brain, the activities of beta-galactosidase, beta-glucuronidase, N-acetyl-beta-glucosaminidase, alanyl-aminopeptidase, leucyl-aminopeptidase, lysosomal esterase and lysosomal lipase were affected. 3. The changes in enzyme activities in the investigated tissue were related to the time and concentration of the administered ethanol.

The effect of small doses of mercury on the level of selected lysosomal enzymes in the plasma and lymphocytes of sheep.

1. The 12-month-old ewes, Slovak Merinos breed, were given 40 mg of mercury daily for 28 days in the form of HgCl2. 2. Administration of mercury had a significant influence on the activity of the investigated lysosomal enzymes in the plasma and lymphocytes of sheep.

Activity of some lysosomal enzymes in plasma and leucocytes of rabbits exposed to effect of retinol and hydrocortisone.

Observing activity of some lysosomal enzymes in blood serum and leucocytes of rabbits subjected to injection of 200,000 units of retinol and 25 mg of hydrocortisone/kg of body weight it was found that: 1. In the effect of retinol administration there was an increase in the activity AP, BGAL, BGLU, AspAT and lipase in blood serum after 72 hours and NAGL after 168 hours while in leucocytes BGAL and NAGL after 72 hours and AGAL after 168 hours. 2. As a result of hydrocortisone injection the activity of all the enzymes examined (except Ala-Na) in blood serum increased markedly already after 24-48 hours. 3. In leucocytes hydrocortisone caused a significant increase in the activity of AP, BGRD, NAGL, BGAL, AGAL and cathepsin D. 4. The glucose level in blood plasma decreased after 48 hours and 120 hours after hydrocortisone injection and 168 hours after retinol injection.