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1.  Selective targeting of bioengineered platelets to prostate cancer vasculature: new paradigm for therapeutic modalities 
Androgen deprivation therapy (ADT) provides palliation for most patients with advanced prostate cancer (CaP); however, greater than 80% subsequently fail ADT. ADT has been indicated to induce an acute but transient destabilization of the prostate vasculature in animal models and humans. Human re-hydrated lyophilized platelets (hRL-P) were investigated as a prototype for therapeutic agents designed to target selectively the tumour-associated vasculature in CaP. The ability of hRL-P to bind the perturbed endothelial cells was tested using thrombin- and ADP-activated human umbilical vein endothelial cells (HUVEC), as well as primary xenografts of human prostate tissue undergoing acute vascular involution in response to ADT. hRL-P adhered to activated HUVEC in a dose-responsive manner. Systemically administered hRL-P, and hRL-P loaded with super-paramagnetic iron oxide (SPIO) nanoparticles, selectively targeted the ADT-damaged human microvasculature in primary xenografts of human prostate tissue. This study demonstrated that hRL-P pre-loaded with chemo-therapeutics or nanoparticles could provide a new paradigm for therapeutic modalities to prevent the rebound/increase in prostate vasculature after ADT, inhibiting the transition to castration-recurrent growth.
doi:10.1111/jcmm.12515
PMCID: PMC4511351  PMID: 25736582
platelets; androgen deprivation; human prostate xenografts; SPIO nanoparticles
2.  Comparison of [18F]DCFPyL and [68Ga]Ga-PSMA-HBED-CC for PSMA-PET Imaging in Patients with Relapsed Prostate Cancer 
Molecular Imaging and Biology  2015;17(4):575-584.
Purpose
Gallium-68 (Ga-68)-labeled tracers for imaging expression of the prostate-specific membrane antigen (PSMA) such as the [68Ga]Ga-PSMA-HBED-CC have already demonstrated high potential for the detection of recurrent prostate cancer. However, compared to Ga-68, a labeling with fluorine-18 (F-18) would offer advantages with respect to availability, production amount, and image resolution. [18F]DCFPyL is a promising F-18-labeled candidate for PSMA-positron emission tomography (PET) imaging that has been recently introduced. In the current study, we aimed to compare [68Ga]Ga-PSMA-HBED-CC and [18F]DCFPyL for clinical use in biochemically relapsed prostate cancer.
Procedures
In 14 selected patients with PSA relapse of prostate cancer, [18F]DCFPyL PET/X-ray computed tomography (CT) was performed in addition to [68Ga]Ga-PSMA-HBED-CC PET/CT. A systematic comparison was carried out between results obtained with both tracers with regard to the number of detected PSMA-positive lesions, the standardized uptake value (SUV)max and the lesion to background ratios.
Results
All suspicious lesions identified by [68Ga]Ga-PSMA-HBED-CC were also detected with [18F]DCFPyL. In three patients, additional lesions were observed using [18F]DCFPyL PET/CT. The mean SUVmax in the concordant [18F]DCFPyL PSMA-positive lesions was significantly higher as compared to [68Ga]Ga-PSMA-HBED-CC (14.5 vs. 12.2, p = 0.028, n = 15). The mean tumor to background ratios (n = 15) were significantly higher for [18F]DCFPyL compared to [68Ga]Ga-PSMA-HBED-CC using kidney, spleen, or parotid as reference organs (p = 0.006, p = 0.002, p = 0.008), but no significant differences were found using the liver (p = 0.167) or the mediastinum (p = 0.363) as reference organs.
Conclusion
[18F]DCFPyL PET/CT provided a high image quality and visualized small prostate lesions with excellent sensitivity. [18F]DCFPyL represents a highly promising alternative to [68Ga]Ga-PSMA-HBED-CC for PSMA-PET/CT imaging in relapsed prostate cancer.
doi:10.1007/s11307-015-0866-0
PMCID: PMC4493776  PMID: 26013479
Prostate-specific membrane antigen (PSMA); F-18; Ga-68; Positron emission tomography (PET); Prostate cancer
3.  Late INa increases diastolic SR-Ca2+-leak in atrial myocardium by activating PKA and CaMKII 
Cardiovascular Research  2015;107(1):184-196.
Aims
Enhanced cardiac late Na current (late INa) and increased sarcoplasmic reticulum (SR)-Ca2+-leak are both highly arrhythmogenic. This study seeks to identify signalling pathways interconnecting late INa and SR-Ca2+-leak in atrial cardiomyocytes (CMs).
Methods and results
In murine atrial CMs, SR-Ca2+-leak was increased by the late INa enhancer Anemonia sulcata toxin II (ATX-II). An inhibition of Ca2+/calmodulin-dependent protein kinase II (Autocamide-2-related inhibitory peptide), protein kinase A (H89), or late INa (Ranolazine or Tetrodotoxin) all prevented ATX-II-dependent SR-Ca2+-leak. The SR-Ca2+-leak induction by ATX-II was not detected when either the Na+/Ca2+ exchanger was inhibited (KBR) or in CaMKIIδc-knockout mice. FRET measurements revealed increased cAMP levels upon ATX-II stimulation, which could be prevented by inhibition of adenylyl cyclases (ACs) 5 and 6 (NKY 80) but not by inhibition of phosphodiesterases (IBMX), suggesting PKA activation via an AC-dependent increase of cAMP levels. Western blots showed late INa-dependent hyperphosphorylation of CaMKII as well as PKA target sites at ryanodine receptor type-2 (-S2814 and -S2808) and phospholamban (-Thr17, -S16). Enhancement of late INa did not alter Ca2+-transient amplitude or SR-Ca2+-load. However, upon late INa activation and simultaneous CaMKII inhibition, Ca2+-transient amplitude and SR-Ca2+-load were increased, whereas PKA inhibition reduced Ca2+-transient amplitude and load and additionally slowed Ca2+ elimination. In atrial CMs from patients with atrial fibrillation, inhibition of late INa, CaMKII, or PKA reduced the SR-Ca2+-leak.
Conclusion
Late INa exerts distinct effects on Ca2+ homeostasis in atrial myocardium through activation of CaMKII and PKA. Inhibition of late INa represents a potential approach to attenuate CaMKII activation and decreases SR-Ca2+-leak in atrial rhythm disorders. The interconnection with the cAMP/PKA system further increases the antiarrhythmic potential of late INa inhibition.
doi:10.1093/cvr/cvv153
PMCID: PMC4476413  PMID: 25990311
Late Na current; Antiarrhythmic drugs; Atrial fibrillation; Protein kinases; SR-Ca2+-leak
4.  Strategic Targeting of Multiple Water‐Mediated Interactions: A Concise and Rational Structure‐Based Design Approach to Potent and Selective MMP‐13 Inhibitors 
Chemmedchem  2013;8(9):1457-1461.
Water in the architecture of life: Potent and selective matrix metalloproteinase‐13 (MMP‐13) inhibitors were rationally designed by targeting multiple water‐mediated interactions between the target protein and small‐molecule inhibitors. This structure‐based design concept offers tremendous opportunities for the discovery of unique small molecules with tailored biological activity.imageWILEY-VCH
doi:10.1002/cmdc.201300278
PMCID: PMC4281860  PMID: 23894097
chemical biology; matrix metalloproteinase inhibitors; structure‐based drug design; structure–activity relationships; water‐mediated interactions
5.  Psychometric properties of the Calgary Cambridge guides to assess communication skills of undergraduate medical students  
Objectives:
The aim of this study was to analyse the psychometric properties of the short version of the Calgary Cambridge Guides and to decide whether it can be recommended for use in the assessment of communications skills in young undergraduate medical students.
Methods:
Using a translated version of the Guide, 30 members from the Department of General Practice rated 5 videotaped encounters between students and simulated patients twice. Item analysis should detect possible floor and/or ceiling effects. The construct validity was investigated using exploratory factor analysis. Intra-rater reliability was measured in an interval of 3 months, inter-rater reliability was assessed by the intraclass correlation coefficient.
Results:
The score distribution of the items showed no ceiling or floor effects. Four of the five factors extracted from the factor analysis represented important constructs of doctor-patient communication The ratings for the first and second round of assessing the videos correlated at 0.75 (p < 0.0001). Intraclass correlation coefficients for each item ranged were moderate and ranged from 0.05 to 0.57.
Conclusions:
Reasonable score distributions of most items without ceiling or floor effects as well as a good test-retest reliability and construct validity recommend the C-CG as an instrument for assessing communication skills in undergraduate medical students. Some deficiencies in inter-rater reliability are a clear indication that raters need a thorough instruction before using the C-CG.
doi:10.5116/ijme.5454.c665
PMCID: PMC4277482  PMID: 25480988
undergraduate medical education;  questionnaires;  physician-patient relations;  teaching;  observer variation
6.  Cooperative endocytosis of the endosomal SNARE protein syntaxin-8 and the potassium channel TASK-1 
Molecular Biology of the Cell  2014;25(12):1877-1891.
SNARE proteins can have functions unrelated to membrane fusion. The unassembled form of the SNARE protein syntaxin-8 interacts with the K+ channel TASK-1; both proteins are internalized via clathrin-mediated endocytosis in a cooperative manner. This is a novel mechanism for the control of endocytosis by cargo proteins.
The endosomal SNARE protein syntaxin-8 interacts with the acid-sensitive potassium channel TASK-1. The functional relevance of this interaction was studied by heterologous expression of these proteins (and mutants thereof) in Xenopus oocytes and in mammalian cell lines. Coexpression of syntaxin-8 caused a fourfold reduction in TASK-1 current, a corresponding reduction in the expression of TASK-1 at the cell surface, and a marked increase in the rate of endocytosis of the channel. TASK-1 and syntaxin-8 colocalized in the early endosomal compartment, as indicated by the endosomal markers 2xFYVE and rab5. The stimulatory effect of the SNARE protein on the endocytosis of the channel was abolished when both an endocytosis signal in TASK-1 and an endocytosis signal in syntaxin-8 were mutated. A syntaxin-8 mutant that cannot assemble with other SNARE proteins had virtually the same effect as wild-type syntaxin-8. Total internal reflection fluorescence microscopy showed formation and endocytosis of vesicles containing fluorescence-tagged clathrin, TASK-1, and/or syntaxin-8. Our results suggest that the unassembled form of syntaxin-8 and the potassium channel TASK-1 are internalized via clathrin-mediated endocytosis in a cooperative manner. This implies that syntaxin-8 regulates the endocytosis of TASK-1. Our study supports the idea that endosomal SNARE proteins can have functions unrelated to membrane fusion.
doi:10.1091/mbc.E13-10-0592
PMCID: PMC4055267  PMID: 24743596
7.  Role of late sodium current as a potential arrhythmogenic mechanism in the progression of pressure-induced heart disease 
Aims
To determine the characteristics of the late Na current (INaL) and its arrhythmogenic potential in the progression of pressure-induced heart disease.
Methods and Results
Transverse aortic constriction (TAC) was used to induce pressure overload in mice. After one week the hearts developed isolated hypertrophy with preserved systolic contractility. In patch-clamp experiments both, INaL and the action potential duration (APD90) were unchanged.
In contrast, after five weeks animals developed heart failure with prolonged APDs and the slowed INaL decay time which could be normalized by addition of the INaL inhibitor ranolazine (Ran) or by the Ca/calmodulin-dependent protein kinase II (CaMKII) inhibitor AIP. Accordingly the APD90 could be significantly abbreviated by Ran, tetrodotoxin and the CaMKII inhibitor AIP. Isoproterenol increased the number of delayed afterdepolarizations (DAD) in myocytes from failing but not sham hearts. Application of either Ran or AIP prevented the occurrence of DADs. Moreover, the incidence of triggered activity was significantly increased in TAC myocytes and was largely prevented by Ran and AIP.
Western blot analyses indicate that increased CaMKII activity and a hyperphosphorylation of the Nav1.5 at the CaMKII phosphorylation site (Ser571) paralleled our functional observations five weeks after TAC surgery.
Conclusion
In pressure overload-induced heart failure a CaMKII-dependent augmentation of INaL plays a crucial role in the AP prolongation and generation of cellular arrhythmogenic triggers, which cannot yet be found in early and still compensated hypertrophy. Inhibition of INaL and CaMKII exert potent antiarrhythmic effects and might therefore be of potential therapeutic interest.
doi:10.1016/j.yjmcc.2013.03.021
PMCID: PMC3720777  PMID: 23570977
Heart failure; hypertrophy; arrhythmias; INaL; CaMKII; ranolazine
8.  Phase IB study of the FLT3 kinase inhibitor midostaurin with chemotherapy in younger newly diagnosed adult patients with acute myeloid leukemia 
Leukemia  2012;26(9):2061-2068.
This phase 1b trial investigated several doses and schedules of midostaurin in combination with daunorubicin and cytarabine induction and high-dose cytarabine post-remission therapy in newly diagnosed patients with acute myeloid leukemia (AML). The discontinuation rate on the 50-mg twice-daily dose schedule was lower than 100 mg twice daily, and no grade 3/4 nausea or vomiting was seen. The complete remission rate for the midostaurin 50-mg twice-daily dose schedule was 80% (FLT3–wild-type: 20 of 27 [74%], FLT3-mutant: 12 of 13 [92%]). Overall survival (OS) probabilities of patients with FLT3-mutant AML at 1 and 2 years (0.85 and 0.62, respectively) were similar to the FLT3–wild-type population (0.78 and 0.52, respectively). Midostaurin in combination with standard chemotherapy demonstrated high complete response and OS rates in newly diagnosed younger adults with AML and was generally well-tolerated at 50 mg twice daily for 14 days. A phase III prospective trial is ongoing (CALGB 10603, NCT00651261).
doi:10.1038/leu.2012.115
PMCID: PMC4118284  PMID: 22627678
FMS-Like Tyrosine kinase 3 receptor (FLT3); acute myeloid leukemia (AML); midostaurin; PKC412; newly diagnosed
9.  Comparison of cerebral blood flow acquired by simultaneous [15O]water positron emission tomography and arterial spin labeling magnetic resonance imaging 
Until recently, no direct comparison between [15O]water positron emission tomography (PET) and arterial spin labeling (ASL) for measuring cerebral blood flow (CBF) was possible. With the introduction of integrated, hybrid magnetic resonance (MR)-PET scanners, such a comparison becomes feasible. This study presents results of CBF measurements recorded simultaneously with [15O]water and ASL. A 3T MR-BrainPET scanner was used for the simultaneous acquisition of pseudo-continuous ASL (pCASL) magnetic resonance imaging (MRI) and [15O]water PET. Quantitative CBF values were compared in 10 young healthy male volunteers at baseline conditions. A statistically significant (P<0.05) correlation was observed between the two modalities; the whole-brain CBF values determined with PET and pCASL were 43.3±6.1 mL and 51.9±7.1 mL per 100 g per minute, respectively. The gray/white matter (GM/WM) ratio of CBF was 3.0 for PET and 3.4 for pCASL. A paired t-test revealed differences in regional CBF between ASL and PET with higher ASL-CBF than PET-CBF values in cortical areas. Using an integrated, hybrid MR-PET a direct simultaneous comparison between ASL and [15O]water PET became possible for the first time so that temporal, physiologic, and functional variations were avoided. Regional and individual differences were found despite the overall similarity between ASL and PET, requiring further detailed investigations.
doi:10.1038/jcbfm.2014.92
PMCID: PMC4126098  PMID: 24849665
arterial spin labeling; cerebral blood flow; magnetic resonance imaging; MR/PET; positron emission tomography; [15O]water PET
10.  The interaction of factor VIIa with rehydrated, lyophilized platelets 
Platelets  2008;19(3):182-191.
The experiments presented here were undertaken to determine if factor VIIa (rFVIIa, the Novo Nordisk product NovoSeven™) will directly bind to rehydrated, lyophilized (RL) platelets for the formation of a catalytic surface with an enhanced ability to generate thrombin. The interaction between rFVIIa and the RL platelet surface was examined by measuring equilibrium and non-equilibrium binding of the coagulation factor to the cells and by following the effects of the surface modification on the kinetics of thrombin generation. The association of rFVIIa with RL platelets was rapid with saturation occurring within minutes. Disassociation was slow, with over half of the coagulation factor remaining bound after two hours. Densities of over one million molecules of rFVIIa per RL platelet were obtained when high concentrations of rFVIIa were incubated with RL platelets. Thrombin generation measurements showed that RL platelet-bound rFVIIa was catalytically active. Thus we can expect that RL platelets, which have been shown to effectively bind to sites of vascular injury, will localize rFVIIa to wounds for an increase in therapeutic index. These studies indicate that rFVIIa-RL platelets are worthy of preclinical and clinical development as an infusion agent for severe bleeding.
doi:10.1080/09537100701493794
PMCID: PMC3925421  PMID: 18432519
Platelet; factor VIIa; coagulation; thrombin; lyophilized
11.  Network processes involved in the mediation of short-term habituation in Aplysia: contribution of intrinsic regulation of excitability and synaptic augmentation 
Short-term habituation (STH) is the decrease in behavioral responding observed during repeated stimulation at regular intervals. For siphon-elicited siphon withdrawal in Aplysia (S-SWR), we previously showed that the amplitude of responses measured in LFS-type siphon motor neurons (LFS MNs) during training is dependent on the stimulus interval used and is training-site specific. The major source of excitation from siphon stimulation onto the LFS MNs comes from the L29 interneurons. Here we examined the role of the L29s in STH by addressing two questions:
(1) What are the relative contributions of intrinsic regulation of excitability and network inhibition on L29 activity during STH training?
By activating L29s with intracellular current injection, we found that intrinsic changes in excitability occur, but only at short training intervals (1 s). We also demonstrated that network inhibition is not required for regulating L29 responses during training, indicating that any expression of inhibition is redundant to the excitability changes.
(2) How does L29 synaptic plasticity contribute to the maintenance of training site-specificity exhibited in LFS MNs?
When training stimuli are delivered 1 s apart [1 s, interstimulus interval (ISI)], L29 responses decrease in both stimulated (trained) and un-stimulated (untrained) pathways, yet site-specificity of training is maintained in the LFS MNs. Our results suggest that activity-dependent synaptic facilitation (augmentation; AUG) expressed by the L29s acts to compensate for the decreased activity in the untrained pathway. First, we demonstrated that the L29-LFS synapse exhibits significant AUG with L29 activation at a 1 s ISI. Second, we showed that the induction of AUG prevents the reduction in siphon-evoked LFS responses that is otherwise observed with decreased L29 activity. Collectively, our results support a role for the L29s in regulating network dynamics during STH training, but only at rapid (1 s ISI) training intervals.
doi:10.3389/fnint.2014.00015
PMCID: PMC3923261  PMID: 24592216
interneuron; excitability; inhibition; reflex; mechanoreceptor; network
12.  Cold shock Y-box protein-1 proteolysis autoregulates its transcriptional activities 
Background
The Y-box protein-1 (YB-1) fulfills pleiotropic functions relating to gene transcription, mRNA processing, and translation. It remains elusive how YB-1 shuttling into the nuclear and cytoplasmic compartments is regulated and whether limited proteolysis by the 20S proteasome releases fragments with distinct function(s) and subcellular distribution(s).
Results
To address these questions, mapping of domains responsible for subcellular targeting was performed. Three nuclear localization signals (NLS) were identified. NLS-1 (aa 149-156) and NLS-2 (aa 185-194) correspond to residues with unknown function(s), whereas NLS-3 (aa 276-292) matches with a designated multimerization domain. Nuclear export signal(s) were not identified. Endoproteolytic processing by the 20S proteasome before glycine 220 releases a carboxy-terminal fragment (CTF), which localized to the nucleus, indicating that NLS-3 is operative. Genotoxic stress induced proteolytic cleavage and nuclear translocation of the CTF. Co-expression of the CTF and full-length YB-1 resulted in an abrogated transcriptional activation of the MMP-2 promoter, indicating an autoregulatory inhibitory loop, whereas it fulfilled similar trans-repressive effects on the collagen type I promoter.
Conclusion
Compartmentalization of YB-1 protein derivatives is controlled by distinct NLS, one of which targets a proteolytic cleavage product to the nucleus. We propose a model for an autoregulatory negative feedback loop that halts unlimited transcriptional activation.
doi:10.1186/1478-811X-11-63
PMCID: PMC3766096  PMID: 24103640
Cold shock protein; DbpB; YBX1; Nuclear localization signal; Post-translational modification; RNA/DNA binding protein
13.  Variable Angle Locking Intercarpal Fusion System for Four-Corner Arthrodesis: Indications and Surgical Technique 
Journal of Wrist Surgery  2012;1(1):73-78.
Four-corner fusion (4CF) is an accepted and regularly performed procedure when managing posttraumatic degenerative disorders in the wrist. This procedure consists of excision of the entire scaphoid in association with midcarpal fusion of the remaining four ulnar carpal bones (hamate, capitate, lunate, and triquetrum). In the majority of cases, the long-term outcome is a functional painless wrist. However, the exact procedure to best achieve a rapid solid bone union of the fusion mass without hardware complications remains controversial. The authors have developed a precise system to ensure precise positioning, firm fixation, and fusion at the midcarpal joint together with an early postoperative recovery, avoiding some of the issues reported with other implants used for 4CF. The described implant is a circular plate accommodating variable angle locking screws as well as compression screws that can firmly fix the plate to the carpal bones. The locking technology produces a very solid construct. A special reaming-distraction-compression guide has also been developed to both countersink the plate on the underlying carpal bone mass and allow distraction of the midcarpal joint for debridement and cancellous bone graft interposition. The features of the implant, its surgical technique, and a relevant case are described.
doi:10.1055/s-0032-1323640
PMCID: PMC3658673  PMID: 23904983
four-corner fusion; SLAC wrist; SNAC wrist; wrist
14.  The cell fate determinant Llgl1 influences HSC fitness and prognosis in AML 
Inactivation of Llgl1 enhances HSC self-renewal and fitness and is associated with unfavorable outcome in human AML.
A unique characteristic of hematopoietic stem cells (HSCs) is the ability to self-renew. Several genes and signaling pathways control the fine balance between self-renewal and differentiation in HSCs and potentially also in leukemia stem cells. Recently, studies have shed light on developmental molecules and evolutionarily conserved signals as regulators of stem cells in hematopoiesis and leukemia. In this study, we provide evidence that the cell fate determinant Llgl1 (lethal giant larvae homolog 1) plays an important role in regulation of HSCs. Loss of Llgl1 leads to an increase in HSC numbers that show increased repopulation capacity and competitive advantage after transplantation. This advantage increases upon serial transplantation or when stress is applied to HSCs. Llgl1−/− HSCs show increased cycling but neither exhaust nor induce leukemia in recipient mice. Llgl1 inactivation is associated with transcriptional repression of transcription factors such as KLF4 (Krüppel-like factor 4) and EGR1 (early-growth-response 1) that are known inhibitors of HSC self-renewal. Decreased Llgl1 expression in human acute myeloid leukemia (AML) cells is associated with inferior patient survival. Thus, inactivation of Llgl1 enhances HSC self-renewal and fitness and is associated with unfavorable outcome in human AML.
doi:10.1084/jem.20120596
PMCID: PMC3549713  PMID: 23277453
15.  Imaging and Elastometry of Blood Clots Using Magnetomotive Optical Coherence Tomography and Labeled Platelets 
Improved methods for imaging and assessment of vascular defects are needed for directing treatment of cardiovascular pathologies. In this paper, we employ magnetomotive optical coherence tomography (MMOCT) as a platform both to detect and to measure the elasticity of blood clots. Detection is enabled through the use of rehydrated, lyophilized platelets loaded with superparamagnetic iron oxides (SPIO-RL platelets) that are functional infusion agents that adhere to sites of vascular endothelial damage. Evidence suggests that the sensitivity for detection is improved over threefold by magnetic interactions between SPIOs inside RL platelets. Using the same MMOCT system, we show how elastometry of simulated clots, using resonant acoustic spectroscopy, is correlated with the fibrin content of the clot. Both methods are based upon magnetic actuation and phase-sensitive optical monitoring of nanoscale displacements using MMOCT, underscoring its utility as a broad-based platform to detect and measure the molecular structure and composition of blood clots.
doi:10.1109/JSTQE.2011.2162580
PMCID: PMC3700352  PMID: 23833549
Elasticity; magnetic forces; nanoparticle; optical imaging; thrombosis
16.  Rapid induction of complete molecular remission by sequential therapy with LDAC and sorafenib in FLT3-ITD-positive patients unfit for intensive treatment: two cases and review of the literature 
Treatment of acute myeloid leukemia remains a therapeutic challenge. Even in younger patients with a low rate of co-morbidities less than 50% of patients can be cured. For older patients or patients with significant co-morbidities, the situation appears even worse. In patients not eligible for intensive treatment approaches - e.g. due to underlying medical conditions - therapeutic approaches remain almost exclusively palliative. However, even with less intense treatment approaches, temporary remission can be achieved and this contributes to prolonged survival and improved quality of life of the respective patient. Targeted therapies have been widely used as palliative treatment in- and outside clinical trials as single agents. Combination with low-dose cytarabine (LDAC) potentially improves remission rates and can be safely administered in an outpatient setting.
Previous studies showed that additive hematologic toxicity of combinatory therapeutic approaches may arise from simultaneous treatment (e.g. chemotherapy plus targeted therapies). However, sequential therapies have already proven their feasibility in clinical trials. Here, we report two cases of rapid induction of complete molecular remission by sequential therapy with LDAC and sorafenib in patients unfit for intensive chemotherapy without significant long-term toxicity.
doi:10.1186/1756-8722-6-39
PMCID: PMC3686641  PMID: 23759001
AML; Sorafenib; Small molecule; LDAC; Targeted therapy
17.  S-Nitrosothiol-Modified Dendrimers as Nitric Oxide Delivery Vehicles 
Biomacromolecules  2008;9(3):834-841.
The synthesis and characterization of two generation-4 polyamidoamine (PAMAM) dendrimers with S-nitrosothiol exteriors are reported. The hyperbranched macromolecules were modified with either N-acetyl-D,L-penicillamine (NAP) or N-acetyl-L-cysteine (NACys) and analyzed via 1H and 13C-NMR, UV absorption spectroscopy, MALDI-TOF mass spectrometry, and size exclusion chromatography. Treatment of the dendritic thiols with nitrite solutions yielded the corresponding S-nitrosothiol nitric oxide (NO) donors (G4-SNAP, G4-NACysNO). Chemiluminescent NO detection demonstrated that the dendrimers were capable of storing ~2 μmol NO·mg−1 when exposed to triggers of S-nitrosothiol decomposition (e.g., light and copper). The kinetics of NO-release were found to be highly dependent on the structure of the nitrosothiol (i.e., tertiary vs. primary) and exhibited similar NO-release characteristics to classical small molecule nitrosothiols reported in the literature. As demonstration of utility, the ability of G4-SNAP to inhibit thrombin-mediated platelet aggregation was assayed. At equivalent nitrosothiol concentrations (25 μM), the G4-SNAP dendrimer resulted in a 62% inhibition of platelet aggregation, compared to only 17% for the small molecule NO donor. The multivalent NO storage, the dendritic effects exerted on nitrosothiol stability and reactivity, and the utility of dendrimers as drug delivery vehicles highlight the potential of these constructs as clinically useful S-nitrosothiol-based therapeutics.
doi:10.1021/bm7011746
PMCID: PMC3564213  PMID: 18247567
18.  Optimizing magnetomotive contrast of SPIO-labeled platelets for thrombosis imaging in optical coherence tomography 
Proceedings of SPIE  2012;8213:82131N.
Rehydratable, lyophilized platelets loaded with superparamagnetic iron oxides (SPIOs) has the potential to provide magnetomotive imaging contrast to sites of vascular damage, including thrombosis complicating atherosclerosis and hemorrhage. Magnetomotive optical coherence tomography (MMOCT) contrasts SPIO-platelets based on their nanoscale, magnetically-induced motion. We report improvements in MMOCT imaging contrast and sensitivity by optimizing the magnetic properties and SPIO loading of the platelets. SPIO-platelets have been shown to specifically adhere to sites of vascular damage in porcine arteries ex vivo. This may lead to new methods for detecting internal bleeding and monitoring the formation of blood clots using infused SPIO-platelets.
doi:10.1117/12.906837
PMCID: PMC3324306  PMID: 22506092
Magnetomotive Optical Coherence Tomography; Platelets; Superparamagnetic iron oxide
19.  Detection of Unstable Carotid Plaque by Tissue Doppler Imaging and Contrast-Enhanced Ultrasound in a Patient with Recurrent Amaurosis Fugax 
Ultrasound (US) is one of the most important diagnostic tools available for the detection and evaluation of carotid stenosis. The case of a 70-year-old woman with recurrent right-sided amaurosis fugax presented here highlights the way in which tissue Doppler imaging (TDI) and contrast-enhanced US (CEUS) may aid in the diagnosis of carotid plaque vulnerability. Furthermore, the novel inverse fly-through technique was used for the three-dimensional visualization of the carotid stenosis.
doi:10.1155/2013/354382
PMCID: PMC3556432  PMID: 23365782
20.  Attentional dynamics during free picture viewing: Evidence from oculomotor behavior and electrocortical activity 
Most empirical evidence on attentional control is based on brief presentations of rather abstract stimuli. Results revealed indications for a dynamic interplay between bottom-up and top-down attentional mechanisms. Here we used a more naturalistic task to examine temporal signatures of attentional mechanisms on fine and coarse time scales. Subjects had to inspect digitized copies of 60 paintings, each shown for 40 s. We simultaneously measured oculomotor behavior and electrophysiological correlates of brain activity to compare early and late intervals (1) of inspection time of each picture (picture viewing) and (2) of the full experiment (time on task). For picture viewing, we found an increase in fixation duration and a decrease of saccadic amplitude while these parameters did not change with time on task. Furthermore, early in picture viewing we observed higher spatial and temporal similarity of gaze behavior. Analyzing electrical brain activity revealed changes in three components (C1, N1 and P2) of the eye fixation-related potential (EFRP); during picture viewing; no variation was obtained for the power in the frontal beta- and in the theta activity. Time on task analyses demonstrated no effects on the EFRP amplitudes but an increase of power in the frontal theta and beta band activity. Thus, behavioral and electrophysiological measures similarly show characteristic changes during picture viewing, indicating a shifting balance of its underlying (bottom-up and top-down) attentional mechanisms. Time on task also modulated top-down attention but probably represents a different attentional mechanism.
doi:10.3389/fnsys.2013.00017
PMCID: PMC3671178  PMID: 23759704
eye fixation-related potentials; saccadic eye movements; top-down attention; bottom-up attention; sustained attention; EEG
21.  Do communication training programs improve students’ communication skills? - a follow-up study 
BMC Research Notes  2012;5:486.
Background
Although it is taken for granted that history-taking and communication skills are learnable, this learning process should be confirmed by rigorous studies, such as randomized pre- and post-comparisons. The purpose of this paper is to analyse whether a communication course measurably improves the communicative competence of third-year medical students at a German medical school and whether technical or emotional aspects of communication changed differently.
Method
A sample of 32 randomly selected students performed an interview with a simulated patient before the communication course (pre-intervention) and a second interview after the course (post-intervention), using the Calgary-Cambridge Observation Guide (CCOG) to assess history taking ability.
Results
On average, the students improved in all of the 28 items of the CCOG. The 6 more technically-orientated communication items improved on average from 3.4 for the first interview to 2.6 in the second interview (p < 0.0001), the 6 emotional items from 2.7 to 2.3 (p = 0.023). The overall score for women improved from 3.2 to 2.5 (p = 0.0019); male students improved from 3.0 to 2.7 (n.s.). The mean interview time significantly increased from the first to the second interview, but the increase in the interview duration and the change of the overall score for the students’ communication skills were not correlated (Pearson’s r = 0.03; n.s.).
Conclusions
Our communication course measurably improved communication skills, especially for female students. These improvements did not depend predominantly on an extension of the interview time. Obviously, “technical” aspects of communication can be taught better than “emotional” communication skills.
doi:10.1186/1756-0500-5-486
PMCID: PMC3495627  PMID: 22947372
22.  The ryanodine receptor leak: how a tattered receptor plunges the failing heart into crisis 
Heart Failure Reviews  2012;18(4):475-483.
It has been persuasively shown in the last two decades that the development of heart failure is closely linked to distinct alterations in Ca2+ cycling. A crucial point in this respect is an increased spontaneous release of Ca2+ out of the sarcoplasmic reticulum during diastole via ryanodine receptors type 2 (RyR2). The consequence is a compromised sarcoplasmic reticulum Ca2+ storage capacity, which impairs systolic contractility and possibly diastolic cardiac function due to Ca2+ overload. Additionally, leaky RyR2 are more and more regarded to potently induce proarrhythmic triggers. Elimination of spontaneously released Ca2+ via RyR2 in diastole can cause a transient sarcolemmal inward current and hence delayed after depolarisations as substrate for cardiac arrhythmias. In this article, the pathological role and consequences of the SR Ca2+-leak and its regulation are reviewed with a main focus on protein kinase A and Ca2+-calmodulin-dependent kinase II. We summarise clinical consequences of “leaky RyR2” as well as possible therapeutic strategies in order to correct RyR2 dysfunction and discuss the significance of the available data.
doi:10.1007/s10741-012-9339-6
PMCID: PMC3677980  PMID: 22932727
Heart failure; Ryanodine receptor; SR Ca2+ leak; PKA; CaMKII
23.  Poly-N-Acetylglucosamine Fibers Amplify the Effectiveness of Recombinant Factor VIIA on Clot Formation in Hemophilia B Canine Blood 
The Journal of Trauma  2011;71(2 Suppl 1):S171-S175.
Background
Achieving hemostasis in anticoagulated patients is an increasingly important clinical issue. Poly-N-acetylglucosamine (pGlcNAc) nanofibers activate platelets by β3 subunit (CD61) and the von Willebrand receptor GP1b (CD42b) integrin signaling for generation of a prothrombotic surface membrane. Recombinant coagulation factor VIIa (rFVIIa) functions in hemophilia A and B by catalyzing formation of the Xa/Va complex on the surface of activated platelets. These observations suggest that pGlcNAc nanofibers may amplify the activity of rFVIIa in hemophilic blood.
Methods
The activity of rFVIIa on platelets was tested by performing thromboelastographic analysis with blood from hemophilia B dogs in the presence of pGlcNAc nanofibers and increasing concentrations of rFVIIa. Mechanisms for hemostatic system activation were investigated with inhibitors of tissue factor, factor XIIa, and platelet function.
Results
Recombinant FVIIa was observed to partially restore the ability of the hemophiliac blood to form fibrin clots in a dose-dependent manner with thromboelastographic analysis. The addition of pGlcNAc nanofibers amplified the rFVIIa effect. The activity of rFVIIa and the amplification effect of pGlcNAc were dependent on platelet integrin function but independent of FXIIa and tissue factor activities.
Conclusions
The pGlcNAc nanofibers amplify rFVIIa activity in hemophilia B canine blood by activating platelets through integrin-dependent mechanisms.
doi:10.1097/TA.0b013e31822556f4
PMCID: PMC3339183  PMID: 21814114
Factor VIIa; Poly-N-acetyl glucosamine nanofibers; Hemophilia B; Thromboelastogram; Corn trypsin inhibitor
24.  Active Interfacial Shear Microrheology of Aging Protein Films 
Physical review letters  2010;104(1):016001.
The magnetically driven rotation of 300 nm diameter rods shows the surface viscosity of albumin at an air-water interface increases from 10−9 to 10−5 Ns/m over two hours while the surface pressure saturates in minutes. The increase in surface viscosity is not accompanied by a corresponding increase in elasticity, suggesting that the protein film anneals with time, resulting in a more densely packed film leading to increased resistance to shear. The nanometer dimensions of the rods provide the same sensitivity as passive microrheology with an improved ability to measure more viscous films.
PMCID: PMC3407536  PMID: 20366371
25.  Intracellular Retention of ABL Kinase Inhibitors Determines Commitment to Apoptosis in CML Cells 
PLoS ONE  2012;7(7):e40853.
Clinical development of imatinib in CML established continuous target inhibition as a paradigm for successful tyrosine kinase inhibitor (TKI) therapy. However, recent reports suggested that transient potent target inhibition of BCR-ABL by high-dose TKI (HD-TKI) pulse-exposure is sufficient to irreversibly commit cells to apoptosis. Here, we report a novel mechanism of prolonged intracellular TKI activity upon HD-TKI pulse-exposure (imatinib, dasatinib) in BCR-ABL-positive cells. Comprehensive mechanistic exploration revealed dramatic intracellular accumulation of TKIs which closely correlated with induction of apoptosis. Cells were rescued from apoptosis upon HD-TKI pulse either by repetitive drug wash-out or by overexpression of ABC-family drug transporters. Inhibition of ABCB1 restored sensitivity to HD-TKI pulse-exposure. Thus, our data provide evidence that intracellular drug retention crucially determines biological activity of imatinib and dasatinib. These studies may refine our current thinking on critical requirements of TKI dose and duration of target inhibition for biological activity of TKIs.
doi:10.1371/journal.pone.0040853
PMCID: PMC3397954  PMID: 22815843

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