Resistance prediction in AML: analysis of 4601 patients from MRC/NCRI, HOVON/SAKK, SWOG and MD Anderson Cancer Center

Therapeutic resistance remains the principal problem in acute myeloid leukemia (AML). We used area under receiver-operating characteristic curves (AUCs) to quantify our ability to predict therapeutic resistance in individual patients, where AUC=1.0 denotes perfect prediction and AUC=0.5 denotes a coin flip, using data from 4601 patients with newly diagnosed AML given induction therapy with 3+7 or more intense standard regimens in UK Medical Research Council/National Cancer Research Institute, Dutch–Belgian Cooperative Trial Group for Hematology/Oncology/Swiss Group for Clinical Cancer Research, US cooperative group SWOG and MD Anderson Cancer Center studies. Age, performance status, white blood cell count, secondary disease, cytogenetic risk and FLT3-ITD/NPM1 mutation status were each independently associated with failure to achieve complete remission despite no early death (‘primary refractoriness’). However, the AUC of a bootstrap-corrected multivariable model predicting this outcome was only 0.78, indicating only fair predictive ability. Removal of FLT3-ITD and NPM1 information only slightly decreased the AUC (0.76). Prediction of resistance, defined as primary refractoriness or short relapse-free survival, was even more difficult. Our limited ability to forecast resistance based on routinely available pretreatment covariates provides a rationale for continued randomization between standard and new therapies and supports further examination of genetic and posttreatment data to optimize resistance prediction in AML.

Should the standard dimethyl sulfoxide concentration be reduced? Results of a European Group for Blood and Marrow Transplantation prospective noninterventional study on usage and side effects of dimethyl sulfoxide.

BACKGROUND Dimethyl sulfoxide (DMSO) is essential for the preservation of liquid nitrogen-frozen stem cells, but is associated with toxicity in the transplant recipient. STUDY DESIGN AND METHODS In this prospective noninterventional study, we describe the use of DMSO in 64 European Blood and Marrow Transplant Group centers undertaking autologous transplantation on patients with myeloma and lymphoma and analyze side effects after return of DMSO-preserved stem cells. RESULTS While the majority of centers continue to use 10% DMSO, a significant proportion either use lower concentrations, mostly 5 or 7.5%, or wash cells before infusion (some for selected patients only). In contrast, the median dose of DMSO given (20 mL) was much less than the upper limit set by the same institutions (70 mL). In an accompanying statistical analysis of side effects noted after return of DMSO-preserved stem cells, we show that patients in the highest quartile receiving DMSO (mL and mL/kg body weight) had significantly more side effects attributed to DMSO, although this effect was not observed if DMSO was calculated as mL/min. Dividing the myeloma and lymphoma patients each into two equal groups by age we were able to confirm this result in all but young myeloma patients in whom an inversion of the odds ratio was seen, possibly related to the higher dose of melphalan received by young myeloma patients. CONCLUSION We suggest better standardization of preservation method with reduced DMSO concentration and attention to the dose of DMSO received by patients could help reduce the toxicity and morbidity of the transplant procedure.

A novel blood-sparing agent in cardiac surgery? First in-patient experience with the synthetic serine protease inhibitor MDCO-2010: a phase II, randomized, double-blind, placebo-controlled study in patients undergoing coronary artery bypass grafting with cardiopulmonary bypass

BACKGROUND Antifibrinolytics have been used for 2 decades to reduce bleeding in cardiac surgery. MDCO-2010 is a novel, synthetic, serine protease inhibitor. We describe the first experience with this drug in patients. METHODS In this phase II, double-blind, placebo-controlled study, 32 patients undergoing isolated primary coronary artery bypass grafting with cardiopulmonary bypass were randomly assigned to 1 of 5 increasing dosage groups of MDCO-2010. The primary aim was to evaluate pharmacokinetics (PK) with assessment of plasmatic concentrations of the drug, short-term safety, and tolerance of MDCO-2010. Secondary end points were influence on coagulation, chest tube drainage, and transfusion requirements. RESULTS PK analysis showed linear dosage-proportional correlation between MDCO-2010 infusion rate and PK parameters. Blood loss was significantly reduced in the 3 highest dosage groups compared with control (P = 0.002, 0.004 and 0.011, respectively). The incidence of allogeneic blood product transfusions was lower with MDCO-2010 4/24 (17%) vs 4/8 (50%) in the control group. MDCO-2010 exhibited dosage-dependent antifibrinolytic effects through suppression of D-dimer generation and inhibition of tissue plasminogen activator-induced lysis in ROTEM analysis as well as anticoagulant effects demonstrated by prolongation of activated clotting time and activated partial thromboplastin time. No systematic differences in markers of end organ function were observed among treatment groups. Three patients in the MDCO-2010 groups experienced serious adverse events. One patient experienced intraoperative thrombosis of venous grafts considered possibly related to the study drug. No reexploration for mediastinal bleeding was required, and there were no deaths. CONCLUSIONS This first-in-patient study demonstrated dosage-proportional PK for MDCO-2010 and reduction of chest tube drainage and transfusions in patients undergoing primary coronary artery bypass grafting. Antifibrinolytic and anticoagulant effects were demonstrated using various markers of coagulation. MDCO-2010 was well tolerated and showed an acceptable initial safety profile. Larger multi-institutional studies are warranted to further investigate the safety and efficacy of this compound.

Associations of ambulatory blood pressure with urinary caffeine and caffeine metabolite excretions.

Intake of caffeinated beverages might be associated with reduced cardiovascular mortality possibly via the lowering of blood pressure. We estimated the association of ambulatory blood pressure with urinary caffeine and caffeine metabolites in a population-based sample. Families were randomly selected from the general population of Swiss cities. Ambulatory blood pressure monitoring was conducted using validated devices. Urinary caffeine, paraxanthine, theophylline, and theobromine excretions were measured in 24 hours urine using ultrahigh performance liquid chromatography tandem mass spectrometry. We used mixed models to explore the associations of urinary excretions with blood pressure although adjusting for major confounders. The 836 participants (48.9% men) included in this analysis had mean age of 47.8 and mean 24-hour systolic and diastolic blood pressure of 120.1 and 78.0 mm Hg. For each doubling of caffeine excretion, 24-hour and night-time systolic blood pressure decreased by 0.642 and 1.107 mm Hg (both P values <0.040). Similar inverse associations were observed for paraxanthine and theophylline. Adjusted night-time systolic blood pressure in the first (lowest), second, third, and fourth (highest) quartile of paraxanthine urinary excretions were 110.3, 107.3, 107.3, and 105.1 mm Hg, respectively (P trend <0.05). No associations of urinary excretions with diastolic blood pressure were generally found, and theobromine excretion was not associated with blood pressure. Anti-hypertensive therapy, diabetes mellitus, and alcohol consumption modify the association of caffeine urinary excretion with systolic blood pressure. Ambulatory systolic blood pressure was inversely associated with urinary excretions of caffeine and other caffeine metabolites. Our results are compatible with a potential protective effect of caffeine on blood pressure.

Influencing factors on the length of stay in lumbar spine surgery: analysis of the German spine registry

INTRODUCTION Spinal disc herniation, lumbar spinal stenosis and spondylolisthesis are known to be leading causes of lumbar back pain. The cost of low back pain management and related operations are continuously increasing in the healthcare sector. There are many studies regarding complications after spine surgery but little is known about the factors predicting the length of stay in hospital. The purpose of this study was to identify these factors in lumbar spine surgery in order to adapt the postoperative treatment. MATERIAL AND METHODS The current study was carried out as a post hoc analysis on the basis of the German spine registry. Patients who underwent lumbar spine surgery by posterior surgical access and with posterior fusion and/or rigid stabilization, whereby procedures with dynamic stabilization were excluded. Patient characteristics were tested for association with length of stay (LOS) using bivariate and multivariate analyses. RESULTS A total of 356 patients met the inclusion criteria. The average age of all patients was 64.6 years and the mean LOS was 11.9 ± 6.0 days with a range of 2-44 days. Independent factors that were influencing LOS were increased age at the time of surgery, higher body mass index, male gender, blood transfusion of 1-2 erythrocyte concentrates and the presence of surgical complications. CONCLUSION Identification of predictive factors for prolonged LOS may allow for estimation of patient hospitalization time and for optimization of postoperative care. In individual cases this may result of a reduction in the LOS.

Development of a method for analysis of ketamine and norketamine enantiomers in equine brain and cerebrospinal fluid by capillary electrophoresis

Ketamine and norketamine are being transported across the blood brain barrier and are also entering from blood into cerebrospinal fluid (CSF). Enantioselective distributions of these compounds in brain and CSF have never been determined. The enantioselective CE based assay previously developed for equine plasma was adapted to the analysis of these compounds in equine brain via use of an acidic pre-extraction of interferences prior to liquid/liquid extraction at alkaline pH. CSF can be treated as plasma. With 100 mg of brain tissue and 0.5 mL of CSF or plasma, assay conditions for up to 30 nmol/g and 6 μM, respectively, of each enantiomer with LOQs of 0.5 nmol/g and 0.1 μM, respectively, were established and the assays were applied to equine samples. CSF and plasma samples analyzed stemmed from anesthetized patient horses and brain, CSF and plasma were obtained from anesthetized horses that were euthanized with an overdose of pentobarbital. Data obtained indicate that ketamine and norketamine enantiomers are penetrating into brain and CSF with those of ketamine being more favorably transported than norketamine, whereas metabolites of norketamine are hindered. More work is required to properly investigate possible stereoselectivities of the ketamine metabolism and transport of metabolites from blood into brain tissue and CSF.

Milk fatty acids as possible biomarkers to early diagnose elevated concentrations of blood plasma nonesterified fatty acids in dairy cows

Most cows encounter a state of negative energy balance during the periparturient period, which may lead to metabolic disorders and impaired fertility. The aim of this study was to assess the potential of milk fatty acids as diagnostic tools of detrimental levels of blood plasma nonesterified fatty acids (NEFA), defined as NEFA concentrations beyond 0.6 mmol/L, in a data set of 92 early lactating cows fed a glucogenic or lipogenic diet and subjected to 0-, 30-, or 60-d dry period before parturition. Milk was collected in wk 2, 3, 4, and 8 (n = 368) and blood was sampled weekly from wk 2 to 8 after parturition. Milk was analyzed for milk fatty acids and blood plasma for NEFA. Data were classified as "at risk of detrimental blood plasma NEFA" (NEFA ≥ 0.6 mmol/L) and "not at risk of detrimental blood plasma NEFA" (NEFA <0.6 mmol/L). Concentrations of 45 milk fatty acids and milk fat C18:1 cis-9-to-C15:0 ratio were subjected to a discriminant analysis. Milk fat C18:1 cis-9 revealed the most discriminating variable to identify detrimental blood plasma NEFA. A false positive rate of 10% allowed us to diagnose 46% of the detrimental blood plasma NEFA cases based on a milk fat C18:1 cis-9 concentration of at least 230 g/kg of milk fatty acids. Additionally, it was assessed whether the milk fat C18:1 cis-9 concentrations of wk 2 could be used as an early warning for detrimental blood plasma NEFA risk during the first 8 wk in lactation. Cows with at least 240 g/kg of C18:1 cis-9 in milk fat had about 50% chance to encounter blood plasma NEFA values of 0.6 mmol/L or more during the first 8 wk of lactation, with a false positive rate of 11.4%. Profit simulations were based on costs for cows suffering from detrimental blood plasma NEFA, and costs for preventive treatment based on daily dosing of propylene glycol for 3 wk. Given the relatively low incidence rate (8% of all observations), continuous monitoring of milk fatty acids during the first 8 wk of lactation to diagnose detrimental blood plasma NEFA does not seem cost effective. On the contrary, milk fat C18:1 cis-9 of the second lactation week could be an early warning of cows at risk of detrimental blood NEFA. In this case, selective treatment may be cost effective.

Improved relapse-free survival after autologous stem cell transplantation does not translate into better quality of life in chronic lymphocytic leukemia: lessons from the randomized European Society for Blood and Marrow Transplantation-Intergroup study

In chronic lymphocytic leukemia (CLL) medical progress is driven by clinical studies with relapse-free survival (RFS) as the primary endpoint. The randomized EBMT-Intergroup trial compared high-dose therapy and autologous stem cell transplantation (ASCT) to observation and demonstrated a substantial improvement of RFS without showing improved overall survival for the transplant arm. Here we report quality of life (QoL) information of the first 3 years following randomization from that study. The main objective was to assess the impact of treatment on QoL over time. Two secondary analyses were performed to further investigate the impact of ASCT and relapse on QoL. In the primary analysis, we demonstrate an adverse impact of ASCT on QoL which was largest at 4 months and continued throughout the first year after randomization. Further, we demonstrated a sustained adverse impact of relapse on QoL which worsened over time. Despite better disease control by ASCT the side effects thus turned the net effect towards inferior QoL in the first year and comparable QoL in the following 2 years after randomization. This study emphasizes the importance of information concerning QoL impacts when patients are counseled about treatments aimed at improving RFS in the absence of a survival benefit.

K-t GRAPPA-accelerated 4D flow MRI of liver hemodynamics: influence of different acceleration factors on qualitative and quantitative assessment of blood flow.

OBJECTIVE We sought to evaluate the feasibility of k-t parallel imaging for accelerated 4D flow MRI in the hepatic vascular system by investigating the impact of different acceleration factors. MATERIALS AND METHODS k-t GRAPPA accelerated 4D flow MRI of the liver vasculature was evaluated in 16 healthy volunteers at 3T with acceleration factors R = 3, R = 5, and R = 8 (2.0 × 2.5 × 2.4 mm(3), TR = 82 ms), and R = 5 (TR = 41 ms); GRAPPA R = 2 was used as the reference standard. Qualitative flow analysis included grading of 3D streamlines and time-resolved particle traces. Quantitative evaluation assessed velocities, net flow, and wall shear stress (WSS). RESULTS Significant scan time savings were realized for all acceleration factors compared to standard GRAPPA R = 2 (21-71 %) (p < 0.001). Quantification of velocities and net flow offered similar results between k-t GRAPPA R = 3 and R = 5 compared to standard GRAPPA R = 2. Significantly increased leakage artifacts and noise were seen between standard GRAPPA R = 2 and k-t GRAPPA R = 8 (p < 0.001) with significant underestimation of peak velocities and WSS of up to 31 % in the hepatic arterial system (p <0.05). WSS was significantly underestimated up to 13 % in all vessels of the portal venous system for k-t GRAPPA R = 5, while significantly higher values were observed for the same acceleration with higher temporal resolution in two veins (p < 0.05). CONCLUSION k-t acceleration of 4D flow MRI is feasible for liver hemodynamic assessment with acceleration factors R = 3 and R = 5 resulting in a scan time reduction of at least 40 % with similar quantitation of liver hemodynamics compared with GRAPPA R = 2.

Impact of local endothelial shear stress on neointima and plaque following stent implantation in patients with ST-elevation myocardial infarction: A subgroup-analysis of the COMFORTABLE AMI-IBIS 4 trial.

BACKGROUND Numerous studies have demonstrated an association between endothelial shear stress (ESS) and neointimal formation after stent implantation. However, the role of ESS on the composition of neointima and underlying plaque remains unclear. METHODS Patients recruited in the Comfortable AMI-IBIS 4 study implanted with bare metal stents (BMS) or biolimus eluting stents (BES) that had biplane coronary angiography at 13month follow-up were included in the analysis. The intravascular ultrasound virtual-histology (IVUS-VH) and the angiographic data were used to reconstruct the luminal surface, and the stent in the stented segments. Blood flow simulation was performed in the stent surface, which was assumed to represent the luminal surface at baseline, to assess the association between ESS and neointima thickness. The predominant ESS was estimated in 3-mm segments and was correlated with the amount of neointima, neointimal tissue composition, and with the changes in the underlying plaque burden and composition. RESULTS Forty three patients (18 implanted with BMS and 25 with BES) were studied. In both stent groups negative correlations were noted between ESS and neointima thickness in BMS (P<0.001) and BES (P=0.002). In BMS there was a negative correlation between predominant ESS and the percentage of the neointimal necrotic core component (P=0.015). In BES group, the limited neointima formation did not allow evaluation of the effect of ESS on its tissue characteristics. ESS did not affect vessel wall remodeling and the plaque burden and composition behind BMS (P>0.10) and BES (P>0.45). CONCLUSIONS ESS determines neointimal formation in both BMS and BES and affects the composition of the neointima in BMS. Conversely, ESS does not impact the plaque behind struts irrespective of stent type throughout 13months of follow-up.

CD47 protein expression in acute myeloid leukemia: A tissue microarray-based analysis

Binding of CD47 to signal regulatory protein alpha (SIRPα), an inhibitory receptor, negatively regulates phagocytosis. In acute myeloid leukemia (AML), CD47 is overexpressed on peripheral blasts and leukemia stem cells and inversely correlates with survival. Aim of the study was to investigate the correlation between CD47 protein expression by immunohistochemistry (IHC) in a bone marrow (BM) tissue microarray (TMA) and clinical outcome in AML patients. CD47 staining on BM leukemia blasts was scored semi-quantitatively and correlated with clinical parameters and known prognostic factors in AML. Low (scores 0-2) and high (score 3) CD47 protein expression were observed in 75% and 25% of AML patients. CD47 expression significantly correlated with percentage BM blast infiltration and peripheral blood blasts. Moreover, high CD47 expression was associated with nucleophosmin (NPM1) gene mutations. In contrast, CD47 expression did not significantly correlate with overall or progression free survival or response to therapy. In summary, a BM TMA permits rapid and reproducible semi-quantitative analysis of CD47 protein expression by IHC. While CD47 expression on circulating AML blasts has been shown to be a negative prognostic marker for a very defined population of AML patients with NK AML, CD47 expression on AML BM blasts is not.

Simultaneous analysis of nuclear and mitochondrial DNA, mRNA and miRNA from backspatter from inside parts of firearms generated by shots at "triple contrast" doped ballistic models

When a firearm projectile hits a biological target a spray of biological material (e.g., blood and tissue fragments) can be propelled from the entrance wound back towards the firearm. This phenomenon has become known as "backspatter" and if caused by contact shots or shots from short distances traces of backspatter may reach, consolidate on, and be recovered from, the inside surfaces of the firearm. Thus, a comprehensive investigation of firearm-related crimes must not only comprise of wound ballistic assessment but also backspatter analysis, and may even take into account potential correlations between these emergences. The aim of the present study was to evaluate and expand the applicability of the "triple contrast" method by probing its compatibility with forensic analysis of nuclear and mitochondrial DNA and the simultaneous investigation of co-extracted mRNA and miRNA from backspatter collected from internal components of different types of firearms after experimental shootings. We demonstrate that "triple contrast" stained biological samples collected from the inside surfaces of firearms are amenable to forensic co-analysis of DNA and RNA and permit sequence analysis of the entire mtDNA displacement-loop, even for "low template" DNA amounts that preclude standard short tandem repeat DNA analysis. Our findings underscore the "triple contrast" method's usefulness as a research tool in experimental forensic ballistics.

Hepatitis B viral load in dried blood spots: A validation study in Zambia

Background: Access to hepatitis B viral load (VL) testing is poor in sub-Saharan Africa (SSA) due toeconomic and logistical reasons.Objectives: To demonstrate the feasibility of testing dried blood spots (DBS) for hepatitis B virus (HBV)VL in a laboratory in Lusaka, Zambia, and to compare HBV VLs between DBS and plasma samples.Study design: Paired plasma and DBS samples from HIV-HBV co-infected Zambian adults were analyzedfor HBV VL using the COBAS AmpliPrep/COBAS TaqMan HBV test (Version 2.0) and for HBV genotypeby direct sequencing. We used Bland-Altman analysis to compare VLs between sample types and bygenotype. Logistic regression analysis was conducted to assess the probability of an undetectable DBSresult by plasma VL.Results: Among 68 participants, median age was 34 years, 61.8% were men, and median plasma HBV VLwas 3.98 log IU/ml (interquartile range, 2.04–5.95). Among sequenced viruses, 28 were genotype A1 and27 were genotype E. Bland–Altman plots suggested strong agreement between DBS and plasma VLs. DBSVLs were on average 1.59 log IU/ml lower than plasma with 95% limits of agreement of −2.40 to −0.83 logIU/ml. At a plasma VL ≥2,000 IU/ml, the probability of an undetectable DBS result was 1.8% (95% CI:0.5–6.6). At plasma VL ≥20,000 IU/ml this probability reduced to 0.2% (95% CI: 0.03–1.7).

Postmortem quantitative 1.5-T MRI for the differentiation and characterization of serous fluids, blood, CSF, and putrefied CSF.

The purpose of the present study was to investigate whether serous fluids, blood, cerebrospinal fluid (CSF), and putrefied CSF can be characterized and differentiated in synthetically calculated magnetic resonance (MR) images based on their quantitative T 1, T 2, and proton density (PD) values. Images from 55 postmortem short axis cardiac and 31 axial brain 1.5-T MR examinations were quantified using a quantification sequence. Serous fluids, fluid blood, sedimented blood, blood clots, CSF, and putrefied CSF were analyzed for their mean T 1, T 2, and PD values. Body core temperature was measured during the MRI scans. The fluid-specific quantitative values were related to the body core temperature. Equations to correct for temperature differences were generated. In a 3D plot as well as in statistical analysis, the quantitative T 1, T 2 and PD values of serous fluids, fluid blood, sedimented blood, blood clots, CSF, and putrefied CSF could be well differentiated from each other. The quantitative T 1 and T 2 values were temperature-dependent. Correction of quantitative values to a temperature of 37 °C resulted in significantly better discrimination between all investigated fluid mediums. We conclude that postmortem 1.5-T MR quantification is feasible to discriminate between blood, serous fluids, CSF, and putrefied CSF. This finding provides a basis for the computer-aided diagnosis and detection of fluids and hemorrhages.

Phosphatidylethanol (PEth) in blood samples from "driving under the influence" cases as indicator for prolonged excessive alcohol consumption.

Phosphatidylethanol (PEth) is considered as specific biomarker of alcohol consumption. Due to accumulation after repeated drinking, PEth is suitable to monitor long-term drinking behavior. To examine the applicability of PEth in "driving under the influence of alcohol" cases, 142 blood samples with blood alcohol concentrations (BAC) ranging from 0.0-3.12 ‰ were analyzed for the presence of PEth homologues 16:0/18:1 (889 ± 878 ng/mL; range analysis, PEth thresholds were evaluated to differentiate moderate and excessive alcohol consumption with acceptable sensitivity and specificity in accordance with the 1.6 ‰ BAC limit. With a threshold of 700 ng/mL for PEth 16:0/18:1, prolonged excessive alcohol consumption was detected in 65.9 % of drunk drivers with a BAC ≥ 1.6 ‰ and in 31.6 % of the samples with a BAC < 1.6 ‰. Similar results were obtained for PEth 16:0/18:2 with a threshold of 300 ng/mL. Both criteria, PEth 16:0/18:1 and PEth 16:0/18:2, were conform in the evaluation of drinking habits in 88.7 % of blood samples. These results show the possibility to detect prolonged excessive alcohol consumption, even if the BAC is below the legal threshold of 1.6 ‰ for driving aptitude assessment. As a consequence, concentrations of PEth 16:0/18:1 ≥ 700 ng/mL and of PEth 16:0/18:2 ≥ 300 ng/mL may be considered as indicators for the necessity of driving aptitude assessment in addition to BAC.