Search

Two-pore domain potassium (K2P) channels carry background (or leak) potassium current and play a key role in regulating resting membrane potential and cellular excitability. Accumulating evidence points to a role for K2Ps in human pathophysiologies, most notably in pain and migraine, making them attractive targets for therapeutic intervention. However, there remains a lack of selective pharmacological tools. The aim of this work was to apply a “target class” approach to investigate the K2P superfamily and identify novel activators across all the described subclasses of K2P channels. Target class drug discovery allows for the leveraging of accumulated knowledge and maximizing synergies across a family of targets and serves as an additional approach to standard target-based screening. A common assay platform using baculovirus (BacMam) to transiently express K2P channels in mammalian cells and a thallium flux assay to determine channel activity was developed, allowing the simultaneous screening of multiple targets. Importantly, this system, by allowing precise titration of channel function, allows optimization to facilitate the identification of activators. A representative set of channels (THIK-1, TWIK-1, TREK-2, TASK-3, and TASK-2) were screened against a library of Food and Drug Administration (FDA)-approved compounds and the LifeArc Index Set. Activators were then analyzed in concentration–response format across all channels to assess selectivity. Using the target class approach to investigate the K2P channels has enabled us to determine which of the K2Ps are amenable to small-molecule activation, de-risk multiple channels from a technical point of view, and identify a diverse range of previously undescribed pharmacology.

A 45-year-old man appeared to have a central (protrusio) hip dislocation but actually had a transverse posterior wall acetabulum fracture with irreducible posterior dislocation due to impalement of the femoral head on the ischial spine. He underwent urgent open reduction on presentation and subsequent internal fixation in a staged manner. He developed avascular necrosis at 18 months postoperatively.The nondisplaced ilioischial and iliopectineal acetabular radiographic lines were alerts that the dislocation was actually posterior. This led to further imaging before any closed reduction attempts because standard reduction maneuvers would have placed the patient at high risk for iatrogenic femoral head or neck fracture.

TRAAK (KCNK4, K2P4.1) is a mechanosensitive two-pore domain potassium (K2P) channel. Due to its expression within sensory neurons and genetic link to neuropathic pain it represents a promising potential target for novel analgesics. In common with many other channels in the wider K2P sub-family, there remains a paucity of small molecule pharmacological tools. Specifically, there is a lack of molecules selective for TRAAK over the other members of the TREK subfamily of K2P channels. We developed a thallium flux assay to allow high throughput screening of compounds and facilitate the identification of novel TRAAK activators. Using a library of ∼1200 drug like molecules we identified Aprepitant as a small molecule activator of TRAAK. Aprepitant is an NK-1 antagonist used to treat nausea and vomiting. Close structural analogues of Aprepitant and a range of NK-1 antagonists were also selected or designed for purchase or brief chemical synthesis and screened for their ability to activate TRAAK. Electrophysiology experiments confirmed that Aprepitant activates both the 'long' and 'short' transcript variants of TRAAK. We also demonstrated that Aprepitant is selective and does not activate other members of the K2P superfamily. This work describes the development of a high throughput assay to identify potential TRAAK activators and subsequent identification and confirmation of the novel TRAAK activator Aprepitant. This discovery identifies a useful tool compound which can be used to further probe the function of TRAAK K2P channels.

Posttraumatic avascular necrosis (AVN) is osteonecrosis from vascular disruption, commonly encountered after fractures of the femoral neck, proximal humerus, talar neck, and scaphoid. These locations have a tenuous vascular supply; the diagnosis, risk factors, natural history, and treatment are reviewed. Fracture nonunion only correlates with AVN in the scaphoid. In the femoral head, the risk is increased for displaced fractures, but the time to surgery and open versus closed treatment do not seem to influence the risk. Patients with collapse are frequently symptomatic, and total hip arthroplasty is the most reliable treatment. In the humeral head, certain fracture patterns correlate with avascularity at the time of injury, but most do not go on to develop AVN due to head revascularization. Additionally, newer surgical approaches and improved construct stability appear to lessen the risk of AVN. The likelihood of AVN of the talar body rises with increased severity of talar injury. The development of AVN corresponds with a worse prognosis and increases the likelihood of secondary procedures. In proximal pole scaphoid fractures, delays in diagnosis and treatment elevate the risk of AVN, which is often seen in cases of nonunion. The need for vascularized versus nonvascularized bone grafting when repairing scaphoid nonunions with AVN remains unclear.

TREK2 (KCNK10, K2P10.1) is a two-pore domain potassium (K2P) channel and a potential target for the treatment of pain. Like the majority of the K2P superfamily, there is currently a lack of useful pharmacological tools to study TREK2. Here we present a strategy for identifying novel TREK2 activators. A cell-based thallium flux assay was developed and used to screen a library of drug-like molecules, from which we identified the CysLT1 antagonist Pranlukast as a novel activator of TREK2. This compound was selective for TREK2 versus TREK1 and showed no activity at TRAAK. Pranlukast was also screened against other members of the K2P superfamily. Several close analogues of Pranlukast and other CysLT1 antagonists were also tested for their ability to activate K2P channels. Consistent with previous work, structure activity relationships showed that subtle structural changes to these analogues completely attenuated the activation of TREK2, whereas for TREK1, analogues moved from activators to inhibitors. Pranlukast's activity was also confirmed using whole-cell patch clamp electrophysiology. Studies using mutant forms of TREK2 suggest Pranlukast does not bind in the K2P modulator pocket or the BL-1249 binding site. Pranlukast therefore represents a novel tool by which to study the mechanism of TREK2 activation.

Importance Despite the widespread use of systemic antibiotics to prevent infections in surgically treated patients with fracture, high rates of surgical site infection persist. Objective To examine the effect of intrawound vancomycin powder in reducing deep surgical site infections. Design, Setting, and Participants This open-label randomized clinical trial enrolled adult patients with an operatively treated tibial plateau or pilon fracture who met the criteria for a high risk of infection from January 1, 2015, through June 30, 2017, with 12 months of follow-up (final follow-up assessments completed in April 2018) at 36 US trauma centers. Interventions A standard infection prevention protocol with (n = 481) or without (n = 499) 1000 mg of intrawound vancomycin powder. Main Outcomes and Measures The primary outcome was a deep surgical site infection within 182 days of definitive fracture fixation. A post hoc comparison assessed the treatment effect on gram-positive and gram-negative-only infections. Other secondary outcomes included superficial surgical site infection, nonunion, and wound dehiscence. Results The analysis included 980 patients (mean [SD] age, 45.7 [13.7] years; 617 [63.0%] male) with 91% of the expected person-time of follow-up for the primary outcome. Within 182 days, deep surgical site infection was observed in 29 of 481 patients in the treatment group and 46 of 499 patients in the control group. The time-to-event estimated probability of deep infection by 182 days was 6.4% in the treatment group and 9.8% in the control group (risk difference, –3.4%; 95% CI, –6.9% to 0.1%;P = .06). A post hoc analysis of the effect of treatment on gram-positive (risk difference, –3.7%; 95% CI, –6.7% to –0.8%;P = .02) and gram-negative-only (risk difference, 0.3%; 95% CI, –1.6% to 2.1%;P = .78) infections found that the effect of vancomycin powder was a result of its reduction in gram-positive infections. Conclusions and Relevance Among patients with operatively treated tibial articular fractures at a high risk of infection, intrawound vancomycin powder at the time of definitive fracture fixation reduced the risk of a gram-positive deep surgical site infection, consistent with the activity of vancomycin. Trial Registration ClinicalTrials.gov Identifier:NCT02227446

In recent years there has been interest in the use of machine learning in suicide research in reaction to the failure of traditional statistical methods to produce clinically useful models of future suicide. The current review summarizes recent prediction studies in the suicide literature including those using machine learning approaches to understand what value these novel approaches add.Studies using machine learning to predict suicide deaths report area under the curve that are only modestly greater than, and sensitivities that are equal to, those reported in studies using more conventional predictive methods. Positive predictive value remains around 1% among the cohort studies with a base rate that was not inflated by case-control methodology.Machine learning or artificial intelligence may afford opportunities in mental health research and in the clinical care of suicidal patients. However, application of such techniques should be carefully considered to avoid repeating the mistakes of existing methodologies. Prediction studies using machine-learning methods have yet to make a major contribution to our understanding of the field and are unproven as clinically useful tools.

A series of trisubstituted thiazoles have been identified as potent inhibitors of Plasmodium falciparum (Pf) cGMP-dependent protein kinase (PfPKG) through template hopping from known Eimeria PKG (EtPKG) inhibitors. The thiazole series has yielded compounds with improved potency, kinase selectivity and good in vitro ADME properties. These compounds could be useful tools in the development of new anti-malarial drugs in the fight against drug resistant malaria.

Antibody–drug conjugates (ADCs) are a growing class of therapeutics that harness the specificity of antibodies and the cell‐killing potency of small‐molecule drugs. Beyond cytotoxics, there are few examples of the application of an ADC approach to difficult drug discovery targets. Here, we present the initial development of a non‐internalising ADC, with a view to selectively inhibiting an extracellular protein. Employing the wellinvestigated matrix metalloproteinase‐9 (MMP‐9) as our model, we adapted a broad‐spectrum, nonselective MMP inhibitor for conjugation and linked this to a MMP‐9‐targeting antibody. The resulting ADC fully inhibits MMP‐9, and ELISA results suggest antibody targeting can direct a nonselective inhibitor.

Hepatotoxins cause liver damage via many mechanisms but the formation of reactive metabolites and/or damage to liver mitochondria are commonly implicated. We assess 3D human primary hepatocyte microtissues as a platform for hepatotoxicity studies with reactive metabolite-forming and mitochondria-perturbing compounds. We show that microtissues formed from cryopreserved human hepatocytes had bile canaliculi, transcribed mRNA from genes associated with xenobiotic metabolism and expressed functional cytochrome P450 enzymes. Hierarchical clustering was used to distinguish dose-dependent hepatotoxicity elicited by clozapine, fialuridine and acetaminophen (APAP) from control cultures and less liver-damaging compounds, olanzapine and entecavir. The regio-isomer of acetaminophen, N-acetyl-meta-aminophenol (AMAP) clustered with the hepatotoxic compounds. The principal metabolites of APAP were formed and dose-dependent changes in metabolite profile similar to those seen in patient overdose was observed. The toxicological profile of APAP was indistinguishable from that of AMAP, confirming AMAP as a human hepatotoxin. Tissue oxygen consumption rate was significantly decreased within 2h of exposure to APAP or AMAP, concomitant with glutathione depletion. These data highlight the potential utility of perfused metabolically functional human liver microtissues in drug development and mechanistic toxicology.

A capability for analyzing complex cellular communication among tissues is important in drug discovery and development, and in vitro technologies for doing so are required for human applications. A prominent instance is communication between the gut and the liver, whereby perturbations of one tissue can influence behavior of the other. Here, we present a study on human gut-liver tissue interactions under normal and inflammatory contexts, via an integrative multi-organ platform comprising human liver (hepatocytes and Kupffer cells), and intestinal (enterocytes, goblet cells, and dendritic cells) models. Our results demonstrated long-term (>2 weeks) maintenance of intestinal (e.g., barrier integrity) and hepatic (e.g., albumin) functions in baseline interaction. Gene expression data comparing liver in interaction with gut, versus isolation, revealed modulation of bile acid metabolism. Intestinal FGF19 secretion and associated inhibition of hepatic CYP7A1 expression provided evidence of physiologically relevant gut-liver crosstalk. Moreover, significant non-linear modulation of cytokine responses was observed under inflammatory gut-liver interaction; for example, production of CXCR3 ligands (CXCL9,10,11) was synergistically enhanced. RNA-seq analysis revealed significant upregulation of IFNα/β/γ signaling during inflammatory gut-liver crosstalk, with these pathways implicated in the synergistic CXCR3 chemokine production. Exacerbated inflammatory response in gut-liver interaction also negatively affected tissue-specific functions (e.g., liver metabolism). These findings illustrate how an integrated multi-tissue platform can generate insights useful for understanding complex pathophysiological processes such as inflammatory organ crosstalk., National Institutes of Health (U.S.) (grant UH3TR00069), United States. Defense Advanced Research Projects Agency (grant Microphysiological Systems Program (W911NF-12-2-00))

Graphical abstract, Focussed studies on imidazopyridine inhibitors of Plasmodium falciparum cyclic GMP-dependent protein kinase (PfPKG) have significantly advanced the series towards desirable in vitro property space. LLE-based approaches towards combining improvements in cell potency, key physicochemical parameters and structural novelty are described, and a structure-based design hypothesis relating to substituent regiochemistry has directed efforts towards key examples with well-balanced potency, ADME and kinase selectivity profiles.

Graphical abstract, The structural diversity and SAR in a series of imidazopyridazine inhibitors of Plasmodium falciparum calcium dependent protein kinase 1 (PfCDPK1) has been explored and extended. The opportunity to further improve key ADME parameters by means of lowering log D was identified, and this was achieved by replacement of a six-membered (hetero)aromatic linker with a pyrazole. A short SAR study has delivered key examples with useful in vitro activity and ADME profiles, good selectivity against a human kinase panel and improved levels of lipophilic ligand efficiency. These new analogues thus provide a credible additional route to further development of the series.

AIM To develop a human in vitro model of non-alcoholic fatty liver disease (NAFLD), utilising primary hepatocytes cultured in a three-dimensional (3D) perfused platform. METHODS Fat and lean culture media were developed to directly investigate the effects of fat loading on primary hepatocytes cultured in a 3D perfused culture system. Oil Red O staining was used to measure fat loading in the hepatocytes and the consumption of free fatty acids (FFA) from culture medium was monitored. Hepatic functions, gene expression profiles and adipokine release were compared for cells cultured in fat and lean conditions. To determine if fat loading in the system could be modulated hepatocytes were treated with known anti-steatotic compounds. RESULTS Hepatocytes cultured in fat medium were found to accumulate three times more fat than lean cells and fat uptake was continuous over a 14-d culture. Fat loading of hepatocytes did not cause any hepatotoxicity and significantly increased albumin production. Numerous adipokines were expressed by fatty cells and genes associated with NAFLD and liver disease were upregulated including: Insulin-like growth factor-binding protein 1, fatty acid-binding protein 3 and CYP7A1. The metabolic activity of hepatocytes cultured in fatty conditions was found to be impaired and the activities of CYP3A4 and CYP2C9 were significantly reduced, similar to observations made in NAFLD patients. The utility of the model for drug screening was demonstrated by measuring the effects of known anti-steatotic compounds. Hepatocytes, cultured under fatty conditions and treated with metformin, had a reduced cellular fat content compared to untreated controls and consumed less FFA from cell culture medium. CONCLUSION The 3D in vitro NAFLD model recapitulates many features of clinical NAFLD and is an ideal tool for analysing the efficacy of anti-steatotic compounds.

Co-crystallisation of the imidazo[1,2-a]pyrazine derivative 15 (3-chloro-N-(4-morpholinophenyl)-6-(pyridin-3-yl)imidazo[1,2-a]pyrazin-8-amine) with Aurora-A provided an insight into the interactions of this class of compound with Aurora kinases. This led to the design and synthesis of potent Aurora-A inhibitors demonstrating up to 70-fold selectivity in cell-based Aurora kinase pharmacodynamic biomarker assays.

A high-throughput screening campaign identified a number of imidazopyridazines as novel inhibitors of the malarial kinase PfPK7. Further synthetic chemistry efforts enabled the preparation of a number of analogues with promising in vitro potencies. Although these compounds show likely broad spectrum inhibitory activity, they represent a useful starting point for further chemical optimisation.

Purpose: To biomechanically compare the Jobe transosseous ulnar collateral ligament (UCL) reconstruction procedure and an interference screw reconstruction (ISR) technique versus the intact elbow UCL. Methods: Intact stiffness of 10 matched cadaveric elbow pairs was tested via submaximal valgus loading at 4 flexion angles. From each pair, a metal ISR and a traditional transosseous Jobe reconstruction was performed with the use of matched hamstring tendon grafts. Initial stiffness (graft tension), overall stiffness, strain, and failure strength of reconstructed elbows were then tested. Results: At each tested flexion angle, Jobe constructs reproduced the initial and overall stiffness of the intact ligament. ISR constructs did not reproduce the overall stiffness of the native ligament at any flexion angle, and they reproduced the initial stiffness only at 30° and 120° of flexion. Jobe constructs were significantly stronger, failing (10° of displacement) at 22.7 Nm after absorbing 1.58 Nm of energy, versus 13.4 Nm and 0.97 Nm for ISR constructs. In all, 40% of bone tunnel reconstructions failed via tunnel fracture, and 70% of interference screw constructs failed via graft slippage. Conclusions: The failure strength and initial and overall stiffness of a traditional Jobe bone tunnel UCL reconstruction are superior to those of an ISR, and only traditional Jobe bone tunnel reconstruction reproduces the initial and overall stiffness of an intact UCL. Clinical Relevance: Many UCL reconstruction techniques have been described, and a paucity of biomechanical data supports their use. This study found the Jobe bone tunnel technique to be biomechanically superior to the ISR technique.

Endotoxin lipopolysaccharide (LPS) is known to cause liver injury primarily involving inflammatory cells such as Kupffer cells, but few in vitro culture models are applicable for investigation of inflammatory effects on drug metabolism. We have developed a three-dimensional human microphysiological hepatocyte-Kupffer cell coculture system and evaluated the anti-inflammatory effect of glucocorticoids on liver cultures. LPS was introduced to the cultures to elicit an inflammatory response and was assessed by the release of proinflammatory cytokines, interleukin 6 and tumor necrosis factor α. A sensitive and specific reversed-phase-ultra high-performance liquid chromatography-quadrupole time of flight-mass spectrometry method was used to evaluate hydrocortisone disappearance and metabolism at near physiologic levels. For this, the systems were dosed with 100 nM hydrocortisone and circulated for 2 days; hydrocortisone was depleted to approximately 30 nM, with first-order kinetics. Phase I metabolites, including tetrahydrocortisone and dihydrocortisol, accounted for 8-10% of the loss, and 45-52% consisted of phase II metabolites, including glucuronides of tetrahydrocortisol and tetrahydrocortisone. Pharmacokinetic parameters, i.e., half-life, rate of elimination, clearance, and area under the curve, were 23.03 hours, 0.03 hour(-1), 6.6 × 10(-5) l⋅hour(-1), and 1.03 (mg/l)*h, respectively. The ability of the bioreactor to predict the in vivo clearance of hydrocortisone was characterized, and the obtained intrinsic clearance values correlated with human data. This system offers a physiologically relevant tool for investigating hepatic function in an inflamed liver.

Graves' disease (GD) is an autoimmune thyroid disorder that is inherited as a complex trait. We have genotyped 77 affected sib-pairs with autoimmune thyroid disease for eight polymorphic markers spanning the cytotoxic T lymphocyte antigen-4 ( CTLA-4 ) region of chromosome 2q31-q33, and for five markers spanning the major histocompatibility complex ( MHC ) region of chromosome 6p21. Non-parametric analysis showed linkage of GD to the CTLA-4 region with a peak non-parametric linkage (NPL) score of 3.43 ( P = 0.0004) at the marker D2S117. The proportion of affected full-sibs sharing zero alleles (z0) reached a minimum of 0.113 close to D2S117, giving a locus-specific lambdas for this region of 2.2. Families with brother-sister sib-pairs showed a peak NPL of 3.46 ( P = 0.0003, lambdas > 10) at D2S117, compared with 2.00 ( P = 0.02, lambdas = 1.9) in the families with only affected females, suggesting a stronger influence in families with affected males. Association between GD and the G allele of the Thr17Ala polymorphism within the CTLA-4 gene ( CTLA4A/G ) was observed using unaffected sib controls ( P = 0.005). Lesser evidence for linkage was found at the MHC locus, with a peak NPL score of 1.95 ( P = 0.026), between the markers D6S273 and TNFalpha. We demonstrate that the CTLA-4 locus (lambdas = 2.2) and the MHC locus (lambdas = 1.6) together confer approximately 50% of the inherited susceptibility to GD disease in our population.

We describe three patients with trophic ulceration and blistering of the fingertips associated with carpal tunnel syndrome. One of the patients also had non-insulin-dependent diabetes mellitus. Autonomic neuropathy distal to the carpal tunnel was probably present in all subjects at the time of presentation; in the patient with recent symptoms the skin was warm, and sweating was virtually absent, whilst the other two patients described cold skin, consistent with prolonged autonomic neuropathy.

Background We hypothesized that internal fixation procedures performed on trauma intensive care unit (ICU) patients with systemic infections, some also febrile, would be at increased risk for deep infection. Methods A total of 128 patients (mean age, 37.4 years; mean Injury Severity Score [ISS], 34.7) admitted to the ICU with 179 femur or tibia fractures developed systemic infections. Systemic infections included sepsis, pneumonia, urinary tract infections, abdominal infections, and wound infections remote to the fracture. Of the fractures, 33 open and 146 closed underwent 150 intramedullary and 29 plate fixation procedures. Data were gathered regarding antibiotic use, systemic infection timing in relation to the date of fixation, and whether fever (>38.2°C) was present within 24 hours of fixation. Patients were followed up for a mean of 491 days. Results Twenty-eight procedures were performed a mean of 4.7 days after the diagnosis of a systemic infection, and 151 were performed a mean of 9.3 days before the diagnosis. Forty-five procedures were performed in patients who were febrile within 24 hours. Of the 179 procedures, 10 (5.6%) developed a deep infection. Four patients' implant infection was potentially hematogenously seeded with the same organism as their systemic infection. Neither the timing of the systemic infection in relation to the fixation procedure nor the presence of fever within 24 hours of fixation, days of preoperative antibiotics, location of the fracture, type of fixation (intramedullary nail vs. plate fixation), or type of systemic infection was significantly associated with the development of an infection. The only significant risk factor for developing an orthopedic infection was an open fracture (p Conclusion Internal fixation performed in ICU patients with fever or in close conjunction to the diagnosis of systemic infection led to a 5.6% infection rate, which compares favorably with historic infection rates for fixation of open or closed tibia and femur fractures. Level of evidence Therapeutic, level IV.

Limited data are available on the use of internal fixation in combat zone hospitals. The authors performed a retrospective review of 713 surgical cases during 2 Operation Enduring Freedom deployments to a Level III theater hospital in 2007 and 2009 to 2010. The epidemiology and short- to intermediate-term outcomes of patients treated with internal fixation devices were studied. The authors found that, with judicious use, internal fixation under a damage control protocol in a combat theater hospital can be performed with acceptable complication rates.

Ribonuclease H2 (RNase H2) is a nuclease that specifically hydrolyzes RNA residues in RNA-DNA hybrids. Mutations in the RNase H2 enzyme complex have been identified in the genetic disorder Aicardi-Goutieres syndrome (AGS), which has similarities to the autoimmune disease systemic lupus eryrthrematosis (SLE). The RNase H2 enzyme has also been recently implicated as a key genome surveillance enzyme. Therefore, small-molecule modulators of RNase H2 activity may have utility in therapeutics and as tools to investigate the cellular functions of RNase H2. A fluorescent quench assay, measuring cleavage of an RNA-DNA duplex substrate by recombinant RNase H2, was developed into a high-throughput format and used to screen a 48 560 compound library. A hit validation strategy was subsequently employed, leading to the identification of two novel inhibitor compounds with in vitro nanomolar range inhibition of RNase H2 activity and >100-fold selectivity compared with RNase H type 1. These compounds are the first small-molecule inhibitors of RNase H2 to be reported. They and their derivatives should provide the basis for the development of tool compounds investigating the cellular functions of the RNase H2 enzyme, and, potentially, for pharmacological manipulation of nucleic acid-mediated immune responses.

To estimate the strength of the associations between the suicide of psychiatric in-patients and demographic, historical, symptomatic, diagnostic and treatment factors.A systematic review and meta-analysis of controlled studies of the suicide of psychiatric in-patients including suicides while on approved or unapproved leave.Factors that were significantly associated with in-patient suicide included a history of deliberate self-harm, hopelessness, feelings of guilt or inadequacy, depressed mood, suicidal ideas and a family history of suicide. Patients suffering from both schizophrenia and depressed mood appeared to be at particular risk. The association between suicidal ideas and in-patient suicide was weak and did not reach statistical significance after a quantitative correction for publication bias. A high-risk categorization as defined by a combination of retrospectively determined individual risk factors was strongly statistically associated with in-patient suicide (OR=10.9), with a sensitivity of 64% and a specificity of 85%.Despite the apparently strong association between high-risk categorization and subsequent suicide, the low base rate of in-patient suicide means that predictive value of a high-risk categorization is below 2%. The development of safer hospital environments and improved systems of care are more likely to reduce the suicide of psychiatric in-patients than risk assessment.

Lead optimization studies using 7 as the starting point led to a new class of imidazo[4,5-b]pyridine-based inhibitors of Aurora kinases that possessed the 1-benzylpiperazinyl motif at the 7-position, and displayed favorable in vitro properties. Cocrystallization of Aurora-A with 40c (CCT137444) provided a clear understanding into the interactions of this novel class of inhibitors with the Aurora kinases. Subsequent physicochemical property refinement by the incorporation of solubilizing groups led to the identification of 3-((4-(6-bromo-2-(4-(4-methylpiperazin-1-yl)phenyl)-3H-imidazo[4,5-b]pyridin-7-yl)piperazin-1-yl)methyl)-5-methylisoxazole (51, CCT137690) which is a potent inhibitor of Aurora kinases (Aurora-A IC(50) = 0.015 +/- 0.003 muM, Aurora-B IC(50) = 0.025 muM, Aurora-C IC(50) = 0.019 muM). Compound 51 is highly orally bioavailable, and in in vivo efficacy studies it inhibited the growth of SW620 colon carcinoma xenografts following oral administration with no observed toxicities as defined by body weight loss.

Several recent studies have reported that serious violence towards self and others is more common in the first episode of psychosis than after treatment.To estimate the proportion of survivors of suicide attempts during psychotic illness by jumping from a height who had not previously received treatment with antipsychotic medication.An audit of the medical records of patients admitted to nine designated trauma centres in New South Wales, Australia, after surviving a jump of more than 3 m. Jumping was defined using routine hospital ascribed International Classification of Diseases (ICD) codes. The height of the jump and all clinical data were extracted from case notes.The files of 160 survivors of jumps of more than 3 m were examined, which included 70 who were diagnosed with a psychotic illness (44%). Thirty-one of the 70 diagnosed with a psychotic illness (44%, 95% confidence interval [CI] 32-56%) had never received treatment for psychosis and hence were in the first episode of psychosis. One in five (19.4%) of all survivors of a suicide attempt by jumping had an undiagnosed and untreated psychosis that was often characterized by frightening delusional beliefs.A large proportion of the survivors of suicide attempts by jumping were diagnosed with a psychotic illness, which confirms an association between psychosis and suicide by jumping. Some suicides might not have been linked to psychosis had the patient not survived the suicide attempt, suggesting that the contribution of schizophrenia to suicide mortality might have been underestimated in psychological autopsy studies. The finding that nearly half of the survivors diagnosed to have a psychotic illness had never received treatment with antipsychotic medication indicates a greatly increased risk of suicide by jumping in the first episode of psychosis when compared to the annual risk after treatment.

Psychiatrists and psychologists acting as expert witnesses in court cases are often accused of bias or error. We examined the level of agreement and factors influencing agreement between expert reports admitted into evidence during adversarial civil proceedings. The inter-rater reliability of the psychiatric diagnosis was examined in 51 pairs of civil medicolegal reports written by experts engaged by the same side and 97 pairs of experts engaged by opposite sides. Reports written by experts engaged by the same adversarial side had good agreement about the presence of a mental disorder (kappa = .74) but had only fair agreement about the specific psychiatric diagnosis (average kappa = .31). Reports written by experts engaged by opposing adversarial sides had poor agreement about the presence of any mental disorder and also the specific psychiatric diagnosis. Experts were more likely to agree about the presence of a mental disorder if the plaintiff was involved in a fatal accident. The agreement of treating doctors and experts was similar to that of pairs of experts.

Fifty periprosthetic supracondylar femur fractures above a total knee arthroplasty were reviewed. Fractures were closed Lewis and Rorabeck type II with a stable prosthesis. Twenty-nine patients (group I), were treated with locked condylar plating. Twenty-one patients (group II) were treated with nonlocked plating systems or intramedullary fixation. Minimum follow-up was 1.7 years. There were 5 malunions (20%) in group I and 9 (47%) in group II (P < .05). There were no nonunions in group I and 3 (16%) in group II. Complication rates were 12% in group I, compared to 42% in group II. Group I patients had less operative blood loss, healed in better alignment, and had greater knee motion. All 7 patients treated with a retrograde intramedullary nail developed a malunion or nonunion. Locked plating is a reliable treatment for periprosthetic supracondylar femur fractures. We experienced a lower complication, revision, malunion, and nonunion rate with locked plating versus conventional treatment options.

We report an unusual case of recurrent, painful, unilateral gynaecomastia (GM) in an elderly male with relapsing Graves' hyperthyroidism and co-existing primary hypogonadism. This patient presented to the Breast Clinic with a 4-month history of painful, right GM. Malignancy was excluded but T3 was noted to be raised at 7.3 pmol l(-1) (normal 3.5-5.5) with a suppressed thyroid-stimulating hormone. Testosterone, luteinizing hormone and follicle-stimulating hormone were consistent with primary hypogonadism. He was later referred to physicians with night sweats and painful right GM. FT3 was 7.4 and carbimazole was commenced. Within 4 months, the night sweats and right GM had resolved but he became hypothyroid. When carbimazole was stopped, right GM recurred together with hyperthyroidism. The male breast, which is sensitive to subtle changes in T/E2 ratio, is more likely to be stimulated in an elderly male with hyperthyroidism and pre-existing hypogonadism, and hence recurrence of GM with relapsing hyperthyroidism. Recognition of this association is clinically relevant to avoid unnecessary investigations and undue patient anxiety, and to facilitate appropriate early diagnosis and treatment.

Compartment syndrome results from elevated tissue pressure within an osseofascial compartment that is caused either by an increase in the contents of the compartment, such as that produced by an accumulation of blood or other fluids, or by a decrease in the volume of the compartment, such as that occurring after the application of a constrictive cast or dressing1,2. The pathophysiology of compartment syndrome is well defined: as compartment pressure rises above arteriolar pressure, capillary flow decreases while venous outflow is obstructed, thus impairing perfusion to compartment structures1. Permanent losses of neural and muscular function are the feared outcomes. Compartment syndromes occur most frequently after lower-extremity trauma, often after a high-energy injury. We report the cases of two children in whom compartment syndrome of the leg developed after a low-energy fracture of the femoral shaft was treated with an early sitting spica cast. The parents of both children were informed that data concerning their cases would be submitted for publication. Case 1. A twenty-month-old boy fell and sustained a spiral midshaft femoral fracture. The fracture was not angulated, the limb was shortened by 1 cm, and the patient had normal circulation and neurologic function distal to the fracture. There were no signs of injury to the leg. A long-leg splint was applied in the emergency department and, four hours after the injury, a spica cast was applied in the operating room with the patient under general anesthesia. The casting technique, which was described by Czertak and Hennrikus3 in 1999 and is a modification of the technique described by Kasser4, involved three steps: (1) placement of a below-the-knee fiberglass cast on the affected leg (including the foot) with felt padding along the calf and the popliteal fossa, (2) application of a …

Most illnesses caused by Shiga toxin-producing Escherichia coli (STEC) have been attributed to E. coli serotype O157:H7, but non-O157 STEC infections are now increasingly recognized as public health problems worldwide. The O121:H19 serotype is being isolated more frequently from clinical specimens and has been implicated in one waterborne outbreak. We used multilocus virulence gene profiling, a PCR-based assay, to characterize the virulence gene content of 24 isolates of serotype O121:H19 and nonmotile variants. We also performed multilocus enzyme electrophoresis and multilocus sequencing to establish the clonal relatedness of O121 isolates and to elucidate the relationship of O121 to common STEC clones. The 24 isolates were found to represent a single bacterial clone, as there was no allelic variation across 18 enzyme loci among the isolates. The complete nucleotide sequence of the intimin gene differed by four substitutions from that of the epsilon (Int-ε) allele of O103:H2 strain PMK5. The typical O121 virulence gene profile was similar to the profiles of enterohemorrhagic E. coli (EHEC) clones of E. coli : it included a Shiga toxin 2 gene ( stx 2 ), two genes on the EHEC plasmid ( toxB and ehxA ), and the gene encoding intimin ( eae ). Despite the similarities, putative virulence genes distributed on O islands—large chromosomal DNA segments present in the O157:H7 genome—were useful for discriminating among STEC serotypes and the O121:H19 clone had a composite profile that was distinct from the profiles of the other major EHEC clones of pathogenic E. coli . On the basis of sequencing analysis with 13 housekeeping genes, the O121:H19 clone did not fall into any of the four classical EHEC and enteropathogenic E. coli groups but instead was closely related to two eae -negative STEC strains.

To investigate the changes in provision of hospital-based services for patients with diabetes in an English region over a 10-year period.Questionnaires were completed by lead clinicians in hospitals in the Northern region of England in 1988 and repeated in 1998. Information was sought on diabetes service provision including the staff and their working practices. Data are presented to demonstrate changes during the 10 years.During a 10-year period the number of consultants providing specialized diabetes services increased from 16 to 25 (to become one per 126 240 population). Their outpatient sessions changed from 34 to 55.5 per week, with a decrease in nonspecialists providing diabetes services. Reductions occurred in registrar numbers providing sessions from 23 to 15 and senior house officers from 16 to 14. Increases occurred in other health care professionals: diabetes specialist nurses from 19 to 30.3 whole time equivalents (WTEs); dieticians from 16 to 32.3 WTEs and chiropodists from 8 to 23 WTEs. The numbers of specialized clinics and units providing services from diabetes care centres increased. Improved facilities in clinics and access to laboratory tests were available to all units. Diabetes registers came into use in 12 of 16 units, but there have been difficulties in providing funding. 'Out-of-hours' advice has moved towards advising their patients to see their general practitioners or the accident and emergency department of the hospitals.The number of diabetes professional staff and the provision of specialized diabetes services have increased during a 10-year period in the Northern region of England. However, they still fall far short of recommended staffing levels and services are far from comprehensive in most districts.

The present study investigates human visual processing of simple two‐colour patterns using a delayed match to sample paradigm with positron emission tomography (PET). This study is unique in that we specifically designed the visual stimuli to be the same for both pattern and colour recognition with all patterns being abstract shapes not easily verbally coded composed of two‐colour combinations. We did this to explore those brain regions required for both colour and pattern processing and to separate those areas of activation required for one or the other. We found that both tasks activated similar occipital regions, the major difference being more extensive activation in pattern recognition. A right‐sided network that involved the inferior parietal lobule, the head of the caudate nucleus, and the pulvinar nucleus of the thalamus was common to both paradigms. Pattern recognition also activated the left temporal pole and right lateral orbital gyrus, whereas colour recognition activated the left fusiform gyrus and several right frontal regions. Hum. Brain Mapping 13:213–225, 2001. © 2001 Wiley‐Liss, Inc.

In this study, sustained, selective, divided, and switching attention, and reloading of working memory were investigated in schizophrenia by using a newly developed Visual Attention Battery (VAB). Twenty-four outpatients with schizophrenia and 24 control participants were studied using the VAB. Performance on VAB components was correlated with performance of standard tests. Patients with schizophrenia were significantly impaired on VAB tasks that required switching of attention and reloading of working memory but had normal performance on tasks involving sustained attention or attention to multiple stimulus features. Switching attention and reloading of working memory were highly correlated with Trails (B-A) score for patients. The decline in performance on the switching-attention task in patients with schizophrenia met criteria for a differential deficit in switching attention. Future research should examine the neurophysiological basis of the switching deficit and its sensitivity and specificity to schizophrenia.

Summary We report the case of a teenage boy with thrombocytopenia complicating varicella infection, which responded promptly to treatment with intravenous immunoglobulin therapy. Such treatment is well recognized in the management of immune thrombocytopenia, particularly where steroids are contra-indicated, but we have not found any previous case reports of the use of immunoglobulin in this condition.

Graves disease (GD) is a common autoimmune thyroid disorder that is inherited as a complex multigenic trait. By using a single microsatellite marker at each locus, we screened the type 1 diabetes loci IDDM4, IDDM5, IDDM6, IDDM8, and IDDM10 and the fucosyltransferase-2 locus for linkage in sib pairs with GD. This showed a two-point nonparametric linkage (NPL) score of 1.57 (P=.06) at the IDDM6 marker D18S41, but NPL scores were

Four families with familial hypocalciuric hypercalcaemia were studied. The probands presented with abdominal pain, which in three was due to acute pancreatitis; in two the condition was life threatening. Serum concentrations of calcium, magnesium, phosphate, and immunoassayable parathyroid hormone, urinary calcium excretion, and the rate of renal tubular reabsorption of phosphate were measured; the findings were compared with results in 10 patients with primary hyperparathyroidism matched for serum calcium concentration to establish differences between the diseases. Familial hypocalciuric hypercalcaemia should be suspected in patients with hypercalcaemia in whom daily urinary calcium excretion is below 5 mmol (200 mg) provided renal insufficiency, vitamin D deficiency, and ingestion of drugs that reduce calcium excretion have been excluded. Most cases appear to run a benign course, but some may suffer considerable morbidity. Surgical treatment should be reserved for patients with severe complications, when all parathyroid tissue should be removed.

1. The development of effective drugs targeted at the underlying causative agent of AIDS (eg, HIV) is paramount to reducing morbidity and mortality. 2. Antiretroviral drugs may either inhibit HIV replication at the reverse transcriptase phase of the replication cycle or they may attack the cycle before HIV enters the healthy target cell. 3. AZT has been found to reduce the mortality of patients with AIDS and AIDS-related complex, decrease the frequency and severity of opportunistic infections, and may improve neurologic functioning.

A 25-year-old male with Gorlin's syndrome associated with an intracardiac lesion, presented with jaw cysts requiring surgical treatment. One of the cysts contained unusual histological components including fibrous tissue, myxomatous stroma and abnormal epithelial hyperplasia. Metabolic investigation showed normal parathyroid function and normal tissue responsiveness to parathyroid hormone, despite several clinical and radiological features suggestive of pseudo-hypoparathyroidism. The principal features of the syndrome are reviewed.

Three different forms of testosterone (T) replacement therapy were compared; they were the intramuscular injection of mixed testosterone esters 250 mg; the subcutaneous implantation of 6 X 100 mg pellets of fused testosterone; and the oral administration of testosterone undecanoate (TU) 80 mg twice daily. Six hypogonadal males were treated with oral TU for an eight week period, during which time serial serum hormonal estimations were performed over 10 h at the initiation and after four and eight weeks of therapy. Serum T levels showed marked variability both between subjects and within the same subject on different occasions. We attribute this to variability in absorption of TU, which is formulated in oleic acid. The overall mean T level calculated from the areas under the profiles of TU was 12.0 nmol/l. Hormone responses to injected T esters were studied in nine hypogonadal males. Serum T rose to supraphysiological peak concentrations (mean 71 nmol/l) 24-48 h after an injection, followed by an exponential decay to reach baseline concentrations after 2-3 weeks. The overall calculated mean T level in subjects receiving testosterone esters 250 mg every three weeks was 27.7 nmol/l. Subcutaneous implantation of testosterone in six hypogonadal men produced a gradual rise in serum T followed by a slow decline, with T levels remaining within the normal range for 4-5 months. The calculated overall mean T level over 21 weeks after implantation was 17.0 nmol/l. Serum oestradiol (E2) levels remained within the normal male range throughout the study periods on both TU and T implant therapy but showed a supraphysiological peak (mean 347 pmol/l) 24-48 h after a T injection. 5 alpha-dihydrotestosterone (DHT) levels appeared to parallel those of T on the three forms of therapy, with DHT:T ratios being highest for TU therapy. This was also true for the target organ metabolite 5 alpha-androstane-3 alpha,17 beta-diol. At the doses studied drug costs were similar for T implantation (every 5 months) and T ester injections (every 3 weeks), but were 7-8 times higher for TU (80 mg twice a day). We conclude that T implantation remains overall the most physiological form of androgen replacement therapy, is generally well accepted and attended by few side effects; TU may have a useful role in the initial phases of therapy.

Summary Two patients with insulin-dependent diabetes mellitus (Type I), developed severe, life-threatening hyperkalaemia, the first following treatment with spironolactone, the second during treatment for staphylococcal septicaemia when glucose-induced hyperkalaemia occurred. Investigations demonstrated co-existing hyporeininaemic hypoaldosteronism. Prompt recognition of this combined hormone-deficiency syndrome led to appropriate treatment and recovery. The biochemical features and clinical importance of hyporeninaemic hypoaldosteronism are discussed.

Summary A case of unilateral thyroid swelling due to actinomycosis in a 27 year old farm worker is described. Diagnosis was made by fine needle biopsy under ultrasound control.

Twenty-four hour profiles of circulating prolactin have been documented in eight boys with simple delayed puberty, eleven with gynaecomastia, three of whom were retested following its spontaneous resolution, and two normal adult men. Mean 24 h prolactin levels in four boys with delayed puberty and ten with gynaecomastia exceeded the mean levels for the two adult men. A sleep-associated rise in prolactin levels occurred at all stages of puberty irrespective of the presence or absence of gynaecomastia, and in some subjects peaks also occurred during the daytime. Boys with gynaecomastia had higher 24 h means levels of prolactin (P less than 0.05), higher daytime levels (P less than 0.05) and higher sleep-associated levels (P less than 0.05) than did control subjects. These were not related to the degree or duration of the gynaecomastia, but 24 h mean levels of prolactin and oestradiol were positively correlated. In one subject who had had transient galactorrhoea, high levels of circulating prolactin, oestrone and oestradiol fell following spontaneous resolution of the gynaecomastia. We believe that oestrogen: androgen imbalance during the daytime is the major cause of pubertal gynaecomastia, with hyperprolacinaemia (which may cause galactorrhoea) sometimes occurring as a response to relative hyperoestrogenaemia.

We have studied 30 women with symptomatic benign breast disease in order to test the hypothesis that in some it might be caused by an enzyme variant leading to excessive adrenal production of oestrone. Synthetic 1-24 ACTH (Synacthen) was given intramuscularly after overnight suppression with dexamethasone, during the early follicular phase of the menstrual cycle. In no subjects did plasma oestrone levels show a consistent and significant response to ACTH despite the expected consistent rises in androstenedione and cortisol. We conclude that it is unlikely that adrenal oestrogens play a significant role in the pathophysiology of even a minority of patients with benign breast disease.

We describe the clinical and biochemical features of six men with male pseudohermaphroditism due to androgen resistance. Each of the subjects had male-gender behavior but incomplete virilization. The underlying defects in androgen metabolism were defined by studies of the 5 alpha-reductase enzyme and the androgen receptor in fibroblasts cultured from biopsies of genital skin. Four of the six have 5 alpha-reductase deficiency, and two have defects of the androgen receptor (the Reifenstein syndrome). The responses of these men to androgen treatment were assessed by monitoring nitrogen balance, plasma luteinizing hormone (LH) values, and clinical parameters of virilization including penile growth, potency and ejaculatory volume, muscle bulk, and growth of body and facial hair. In all of the subjects with 5 alpha-reductase deficiency and one man with the Reifenstein syndrome significant response occurred, as evidence by nitrogen retention, lowered plasma LH levels, and improved virilization, with doses of parenteral testosterone esters that raised plasma testosterone levels above the normal male range and brought plasma dihydrotestosterone levels into the normal male range. The subject who did not respond with clinical virilization nevertheless showed nitrogen retention in response to acute testosterone administration. This patient had a profound deficiency of the androgen receptor, whereas the man with a receptor defect who did respond clinically to therapy had normal amounts of a qualitatively abnormal receptor. We conclude that high dose androgen therapy may be of benefit in improving virilization, self-image, and sexual performance in subjects with 5 alpha-reductase deficiency who have male-gender behavior and in some subjects with defects of the androgen receptor.