[Digital pathology].

Digitalisation of pathology slides allows pathologists to make diagnoses using a high-resolution computer screen instead of a conventional microscope. In 2020/21, the four pathology departments in the Region of Southern Denmark implemented digital pathology for all histologic samples. Going digital necessitated optimisation of workflows and training of pathologists, avoiding a reduction in diagnostic quality. This review describes the process for realisation of digital pathology and its future perspectives, including artificial intelligence algorithms to be implemented.

On the Road to Digital Pathology in Denmark-National Survey and Interviews.

Digital pathology (DP) is changing pathology departments dramatically worldwide, yet globally, few departments are presently digitalized for the full diagnostic workflow. Denmark is also on the road to full digitalization countrywide, and this study aim to cover experiences during the implementation process in a national context. Thus, quantitative questionnaires were distributed to all pathology departments in Denmark (n = 13) and distributed to all professions including medical clinical directors, medical doctors (MD) and biomedical laboratory scientists (BLS). For a qualitative perspective, we interviewed four employees representing four professions. Data were collected in 2019-2020. From the questionnaire and interviews, we found strategies differed at the Danish departments with regards to ambitions, technological equipment, workflows, and involvement of type of professions. DP education was requested by personnel. Informants were in general positive toward the digital future but mainly had concerns regarding the political pressure to integrate DP before technological advances are sufficient for maintaining rational budgets, workflows, and for sustaining diagnostic quality. This study is a glance on the Danish implementation process in its early stages from personnel's point of view. It shows the complexity when large new workflow processes are to be implemented countrywide and with a large diversity of stakeholders like managers, MD, BLS, IT-professionals, and authorities. To ensure best technological and economical solutions and to maintain-or even optimize-diagnostic quality with DP and workflow alignment, we suggest superior inter- and intradepartmental communication. When implementing DP countrywide, a national working group is warranted with the variety of stakeholders represented.

Utilizing structure based drug design and metabolic soft spot identification to optimize the in vitro potency and in vivo pharmacokinetic properties leading to the discovery of novel reversible Bruton's tyrosine kinase inhibitors.

Bruton's tyrosine kinase (BTK) is an essential node on the BCR signaling in B cells, which are clinically validated to play a critical role in B-cell lymphomas and various auto-immune diseases such as Multiple Sclerosis (MS), Pemphigus, and rheumatoid arthritis (RA). Although non-selective irreversible BTK inhibitors have been approved for oncology, due to the emergence of drug resistance in B-cell lymphoma associated with covalent inhibitor, there an unmet medical need to identify reversible, selective, potent BTK inhibitor as viable therapeutics for patients. Herein, we describe the identification of Hits and subsequence optimization to improve the physicochemical properties, potency and kinome selectivity leading to the discovery of a novel class of BTK inhibitors. Utilizing Met ID and structure base design inhibitors were synthesized with increased in vivo metabolic stability and oral exposure in rodents suitable for advancing to lead optimization.

Discovery of N-(1-Acryloylazetidin-3-yl)-2-(1H-indol-1-yl)acetamides as Covalent Inhibitors of KRASG12C.

KRAS regulates many cellular processes including proliferation, survival, and differentiation. Point mutants of KRAS have long been known to be molecular drivers of cancer. KRAS p.G12C, which occurs in approximately 14% of lung adenocarcinomas, 3-5% of colorectal cancers, and low levels in other solid tumors, represents an attractive therapeutic target for covalent inhibitors. Herein, we disclose the discovery of a class of novel, potent, and selective covalent inhibitors of KRASG12C identified through a custom library synthesis and screening platform called Chemotype Evolution and structure-based design. Identification of a hidden surface groove bordered by H95/Y96/Q99 side chains was key to the optimization of this class of molecules. Best-in-series exemplars exhibit a rapid covalent reaction with cysteine 12 of GDP-KRASG12C with submicromolar inhibition of downstream signaling in a KRASG12C-specific manner.

Optimization of novel reversible Bruton's tyrosine kinase inhibitors identified using Tethering-fragment-based screens.

Since the approval of ibrutinib for the treatment of B-cell malignancies in 2012, numerous clinical trials have been reported using covalent inhibitors to target Bruton's tyrosine kinase (BTK) for oncology indications. However, a formidable challenge for the pharmaceutical industry has been the identification of reversible, selective, potent molecules for inhibition of BTK. Herein, we report application of Tethering-fragment-based screens to identify low molecular weight fragments which were further optimized to improve on-target potency and ADME properties leading to the discovery of reversible, selective, potent BTK inhibitors suitable for pre-clinical proof-of-concept studies.

Admission rates in a general practitioner-based versus a hospital specialist based, hospital-at-home model: ACCESS, an open-labelled randomised clinical trial of effectiveness.

BACKGROUND: Hospital at home (HaH) is an alternative to acute admission for elderly patients. It is unclear if should be cared for a primarily by a hospital intern specialist or by the patient's own general practitioner (GP). The study assessed whether a GP based model was more effective than a hospital specialist based model at reducing number of hospital admissions without affecting the patient's recovery or number of deaths. METHODS: Pragmatic, randomised, open-labelled multicentre parallel group trial with two arms in four municipalities, four emergency departments and 150 GPs in Southern Denmark, including + 65 years old patients with an acute medical condition that required acute hospital in-patient care. The patients were randomly assigned to hospital specialist based model or GP model of HaH care. Five physical and cognitive performance tests were performed at inclusion and after 7 days. Primary outcome was number of hospital admissions within 7 days. Secondary outcomes were number of admissions within 14, 21 and 30 days, deaths within 30 and 90 days and changes in performance tests. RESULTS: Sixty seven patients were enrolled in the GP model and 64 in the hospital specialist model. 45% in the hospital specialist arm versus 24% in the GP arm were admitted within 7 days (effect size 2.7, 95% CI 1.3-5.8; p = 0.01) and this remained significant within 30 days. No differences were found in death or changes in performance tests from day 0-7 days between the two groups. CONCLUSIONS: The GP based HaH model was more effective than the hospital specialist model in avoiding hospital admissions within 7 days among elderly patients with an acute medical condition with no differences in mental or physical recovery rates or deaths between the two models. REGISTRATION: No. NCT02422849 Registered 27 March 2015. Retrospectively registered.

Safety at The William Quarrier Scottish Epilepsy Centre.

PURPOSE: We examined the yield from EMFIT bed alarms and staff response time to generalised seizure in a medium term residential assessment unit for epilepsy. METHODS: The Scottish Epilpesy Centre (SEC) has a Video Observation System (VOS) that provides continuous recording of all patient spaces (external and internal) and allows retention of clinically relevant events. A retrospective audit of daily EMFIT test records, nursing seizure record sheets (seizure type and EMFIT alert status), clinical incident reporting systems and the VOS database of retained clinical events was conducted for an 9 month period from April 1st 2016 till December 31st 2016. All generalized tonic clonic seizures (GTCS) were noted by patient, time and location and staff response time to GTCS was calculated. RESULTS: There were 85 people admitted during the audit period who had 61 GTCS. 50 events were in bed and EMFIT alert status was recorded. On 8 occasions the EMFIT did not alert: 5 events were not of sufficient duration or frequency, in 2 the patient fell from the bed early and 1 event the alarm did not trigger. The average response time to GTCS was 23s. The longest response time was 69s (range, 0-69s, sd 15.76.). CONCLUSIONS: The EMFIT bed alarm appears to be a valuable adjunct to safety systems. Within the novel environment of the SEC it is possible to maintain a response time to GTCS that is comparable to hospital based UK video telemetry units.

Neurophysiology versus clinical genetics in Rett syndrome: A multicenter study.

Many studies have attempted to establish the genotype-phenotype correlation in Rett syndrome (RTT). Cardiorespiratory measurements provide robust objective data, to correlate with each of the different clinical phenotypes. It has important implications for the management and treatment of this syndrome. The aim of this study was to correlate the genotype with the quantitative cardiorespiratory data obtained by neurophysiological measurement combined with a clinical severity score. This international multicenter study was conducted in four European countries from 1999 to 2012. The study cohort consisted of a group of 132 well-defined RTT females aged between 2 and 43 years with extended clinical, molecular, and neurophysiological assessments. Diagnosis of RTT was based on the consensus criteria for RTT and molecular confirmation. Genotype-phenotype analyses of clinical features and cardiorespiratory data were performed after grouping mutations by the same type and localization or having the same putative biological effect on the MeCP2 protein, and subsequently on eight single recurrent mutations. A less severe phenotype was seen in females with CTS, p.R133C, and p.R294X mutations. Autonomic disturbances were present in all females, and not restricted to nor influenced by one specific group or any single recurrent mutation. The objective information from non-invasive neurophysiological evaluation of the disturbed central autonomic control is of great importance in helping to organize the lifelong care for females with RTT. Further research is needed to provide insights into the pathogenesis of autonomic dysfunction, and to develop evidence-based management in RTT. © 2016 Wiley Periodicals, Inc.

Treating hypoxia in a feeble breather with Rett syndrome.

Rett syndrome (RS) is a unique X-linked dominant neurodevelopmental disorder affecting 1 in 10,000 females. Mutations in the MECP2 gene located on Xq28 have been identified. Many of the characteristic features evolve due to immaturity of the brain in RS. Cardiorespiratory function should be investigated early to characterise the clinical phenotype of the person with RS because each of the three cardiorespiratory phenotypes; apneustic, feeble and forceful breathers have unique and different management strategies. We report a case of a feeble breather showing a correlation between cortical function and tissue pO(2) and pCO(2). We conclude that subtle changes in the levels of blood gases significantly affect cortical function in RS.

Variant of Rett syndrome and CDKL5 gene: clinical and autonomic description of 10 cases.

UNLABELLED: Rett syndrome (RTT) is a severe neurodevelopmental disorder affecting almost exclusively females. The Hanefeld variant, or early-onset seizure variant, has been associated with mutations in CDKL5 gene. AIMS: In recent years more than 60 patients with mutations in the CDKL5 gene have been described in the literature, but the cardiorespiratory phenotype has not been reported. Our aim is to describe clinical and autonomic features of these girls. METHODS: 10 girls with CDKL5 mutations and a diagnosis of Hanefeld variant have been evaluated on axiological and clinical aspects. In all subjects an evaluation of the autonomic system was performed using the Neuroscope. RESULTS: Common features were gaze avoidance, repetitive head movements and hand stereotypies. The autonomic evaluation disclosed eight cases with the Forceful breather cardiorespiratory phenotype and two cases with the Apneustic breather phenotype. CONCLUSIONS: The clinical picture remains within the RTT spectrum but some symptoms are more pronounced in addition to the very early onset of seizures. The cardiorespiratory phenotype was dominated by Forceful breathers, while Feeble breathers were not found, differently from the general Rett population, suggesting a specific behavioral and cardiorespiratory phenotype of the RTT the Hanefeld variant.

Discovery of a potent and highly selective PDK1 inhibitor via fragment-based drug discovery.

We report the use of a fragment-based lead discovery method, Tethering with extenders, to discover a pyridinone fragment that binds in an adaptive site of the protein PDK1. With subsequent medicinal chemistry, this led to the discovery of a potent and highly selective inhibitor of PDK1, which binds in the 'DFG-out' conformation.

Chlamydia pneumoniae IgG and IgA antibody titers and prognosis in patients with coronary heart disease: results from the CLARICOR trial.

The association observed between coronary heart disease (CHD) and Chlamydia (Chlamydophila) pneumoniae antibodies prompted, during the 1990s, several primary and secondary prevention trials with various antibiotics. In our CLARICOR trial, a randomized placebo-controlled trial in 4372 patients with stable CHD, a brief clarithromycin regimen was followed, unexpectedly, by increased long-term mortality. We now compare C. pneumoniae antibody levels at entry with population levels, with the patients' individual histories, and with their subsequent outcomes. IgG antibody levels were somewhat raised, but elevated IgA and IgG titers were unrelated to entry data (including prior acute myocardial infarction), except for an association with smoking and with not using statins. Hazards of mortality and of other outcomes tended to slightly increase with IgA and decrease with IgG titers, but the unfavorable clarithromycin effect was unrelated to antibody levels and remains unexplained. Smoking-related lung disease probably underlies the link between heart disease and increased IgG titers.

Psychophysiological reactivity to sleep-related emotional stimuli in primary insomnia.

The present study examined psychophysiological reactivity to emotional stimuli related and non-related to sleep in people with primary insomnia (PPI) and in good sleepers (GS). Twenty-one PPI and 18 GS were presented with five blocks of neutral, negative, positive, sleep-related negative and sleep-related positive pictures. During the presentation of the pictures, facial electromyography (EMG) of the corrugator and the zygomatic muscles, heart rate (HR) and cardiac vagal tone (CVT) were recorded. Subjective ratings of the stimuli were also collected. We found that only PPI exhibited greater inhibition of the corrugator activity in response to sleep-related positive stimuli compared to the other blocks of stimuli. Furthermore, PPI rated the sleep-related negative stimuli as more unpleasant and arousing and showed higher CVT in response to all stimuli as compared to GS. Results were interpreted as indicating that PPI exhibit craving for sleep-related positive stimuli, and also hyper-arousability in response to sleep-related negative stimuli, as compared to GS. Our results suggest that psychological treatment of insomnia could benefit by the inclusion of strategies dealing with emotional processes linked with sleep processes.

Statin treatment prevents increased cardiovascular and all-cause mortality associated with clarithromycin in patients with stable coronary heart disease.

In the CLARICOR trial, significantly increased cardiovascular (CV) and all-cause mortality in stable patients with coronary heart disease were observed after a short course of clarithromycin. We report on the impact of statin treatment at entry on the CV and all-cause mortality. The multicenter CLARICOR trial randomized patients to oral clarithromycin (500 mg daily; n = 2172) versus matching placebo (daily; n = 2201) for 2 weeks. Patients were followed through public databases. In the 41% patients on statin treatment at entry, no significant effect of clarithromycin was observed on CV (hazard ratio [HR], 0.68, 95% confidence interval [CI], 0.38-1.22; P = 0.20) or all-cause mortality (HR, 1.08; 95% CI, 0.71-1.65; P = 0.72) at 2.6-year follow up. In the patients not on statin treatment at entry, clarithromycin was associated with a significant increase in CV (HR, 1.90; 95% CI, 1.34-2.67; P = 0.0003; statin-clarithromycin interaction P = 0.0029) and all-cause mortality (HR, 1.33; 95% CI, 1.05-1.67; P = 0.016; statin-clarithromycin interaction P = 0.41). Multivariate analysis and 6-year follow up confirmed these results. Concomitant statin treatment in stable patients with coronary heart disease abrogated the observed increased CV mortality associated with 2 weeks of clarithromycin.

SNS-314, a pan-Aurora kinase inhibitor, shows potent anti-tumor activity and dosing flexibility in vivo.

PURPOSE: The Aurora family of serine/threonine kinases (Aurora-A, Aurora-B, and Aurora-C) plays a key role in cells orderly progression through mitosis. Elevated expression levels of Aurora kinases have been detected in a high percentage of melanoma, colon, breast, ovarian, gastric, and pancreatic tumors. We characterized the biological and pharmacological properties of SNS-314, an ATP-competitive, selective, and potent inhibitor of Aurora kinases. METHODS: We studied the biochemical potency and selectivity of SNS-314 to inhibit Aurora kinases A, B, and C. The inhibition of cellular proliferation induced by SNS-314 was evaluated in a broad range of tumor cell lines and correlated to inhibition of histone H3 phosphorylation, inhibition of cell-cycle progression, increase in nuclear content and cell size, loss of viability, and induction of apoptosis. The dose and administration schedule of SNS-314 was optimized for in vivo efficacy in mouse xenograft models of human cancer. RESULTS: In the HCT116 human colon cancer xenograft model, administration of 50 and 100 mg/kg SNS-314 led to dose-dependent inhibition of histone H3 phosphorylation for at least 10 h, indicating effective Aurora-B inhibition in vivo. HCT116 tumors from animals treated with SNS-314 showed potent and sustained responses including reduction of phosphorylated histone H3 levels, increased caspase-3 and appearance of increased nuclear size. The compound showed significant tumor growth inhibition in a dose-dependent manner under a variety of dosing schedules including weekly, bi-weekly, and 5 days on/9 days off. CONCLUSIONS: SNS-314 is a potent small-molecule inhibitor of Aurora kinases developed as a novel anti-cancer therapeutic agent for the treatment of diverse human malignancies.

Clarithromycin for 2 weeks for stable coronary heart disease: 6-year follow-up of the CLARICOR randomized trial and updated meta-analysis of antibiotics for coronary heart disease.

OBJECTIVES: We have reported increased 2.6-year mortality in clarithromycin- versus placebo-exposed stable coronary heart disease patients, but meta-analysis of randomized trials in coronary heart disease patients showed no significant effect of antibiotics on mortality. Here we report the 6-year mortality of clarithromycin- versus placebo-exposed patients and updated meta-analyses. METHODS: Centrally randomized, placebo controlled multicenter trial. All parties were blinded. Analyses were by intention to treat. Meta-analyses followed the Cochrane Collaboration methodology. RESULTS: We randomized 4,372 patients with stable coronary heart disease to clarithromycin 500 mg (n = 2,172) or placebo (n = 2,200) once daily for 2 weeks. Mortality was followed through public register. Nine hundred and twenty-three patients (21.1%) died. Six-year mortality was significantly higher in the clarithromycin group (hazard ratio 1.21, 95% confidence interval 1.06-1.38). Adjustment for entry characteristics (sex, age, prior myocardial infarction, center, and smoking) did not change the results (1.18, 1.04-1.35). Addition of our data to that of other randomized trials on antibiotics for patients with coronary heart disease versus placebo/no intervention (17 trials, 25,271 patients, 1,877 deaths) showed a significantly increased relative risk of death from antibiotics of 1.10 (1.01-1.20) without heterogeneity. CONCLUSIONS: Our results stress the necessity to consider carefully the strength of the indication before administering antibiotics to patients with coronary heart disease.

QT intervals and QT dispersion determined from a 12-lead 24-hour Holter recording in patients with coronary artery disease and patients with heart failure.

BACKGROUND: QT dispersion is considered an index of spatial inhomogeneity of repolarization duration and increased dispersion of ventricular repolarization is supposed to increase the risk of ventricular arrhythmia. Circadian variation in QT dispersion was investigated. METHODS: Three different modes of lead selection was used: all 12-leads (QTdisp 12), only precordial leads (QTdisp 6), and one pair of preselected leads (QTdisp 2) in a 24-hour Holter recording every fourth hour each comprising 10 consecutive measurements in 54 healthy subjects, 29 patients with coronary artery disease (CAD), and 29 patients with heart failure (HF). RESULTS: A significant circadian variation was observed in healthy subjects when modes QTdisp 12 and QTdisp 6 were used (Mean +/- SD 35.58 +/- 16.48 ms; P < 0.0001; and 28.82 +/- 16.02 ms; P < 0.0001, respectively), and in patients with CAD (Mean +/- SD 37.86 +/- 17.87 ms; P < 0.01; and 28.72 +/- 17.06 ms; P < 0.0001, respectively), whereas no circadian variation was observed in QTdisp 2. No circadian variation was observed in patients with HF irrespectively of lead selection. Patients with CAD without myocardial infarction (MI) had a circadian variation in QTdisp 12 (Mean +/- SD 33.13 +/- 14.86 ms; P < 0.05), whereas no circadian variation was observed in patients with MI (Mean +/- SD 40.35 +/- 18.80 ms; P = NS). CONCLUSIONS: Circadian variation of QT dispersion was detected in healthy subjects and in patients with uncomplicated CAD, but not in those who had suffered a previous MI and in patients with HF. The number of leads among which selection of the longest and shortest QT intervals took place was critical for the disclosure of circadian variation of QT dispersion.

Circadian variation in QT dispersion determined from a 12-lead Holter recording: a methodological study of an age- and sex-stratified group of healthy subjects.

BACKGROUND: QT dispersion is considered to reflect inhomogeneity of myocardial repolarization. METHOD: The circadian variation of QT interval dispersion was examined in 95 healthy subjects using 24-hour Holter monitoring. Three different methods of lead selection were applied: all 12 leads (QTdisp 12), only precordial leads (QTdisp 6), and the pair of leads selected at 3 a.m. in which the longest and shortest QT intervals were found in each individual subject (QTdisp 2). RESULTS: A preliminary methodological study including measurements from every minute in 10 subjects revealed no significant circadian variation using mean values of QTdisp 12, QTdisp 6, or QTdisp 2 obtained every hour, every 2, or every 4 hours, except in QTdisp 6, which demonstrated a significant circadian variation (P < 0.01) in 1-hour measurements. Analysis of all 95 subjects using measurements obtained every 4 hours revealed a significant circadian variation in QTdisp 12 and QTdisp 6 (P < 0.0001), whereas no circadian variation was seen in QTdisp 2. A subdivision into 10-year age groups revealed that subjects at age >50 years had a significant circadian variation in QTdisp 12 and QTdisp 6, but not in QTdisp 2. Only in males a significant circadian variation was seen in QTdisp 12 (P < 0.0001), whereas QTdisp 6 demonstrated a circadian variation both in females (P < 0.001) and in males (P < 0.0001). CONCLUSIONS: Selection of leads is of crucial importance for repetitive measurements of QT dispersion. Circadian variation was detected in subjects over 50 years of age, when all 12 or only the 6 precordial leads were taken into account.

Diurnal variation and dispersion in QT interval in cirrhosis: relation to haemodynamic changes.

BACKGROUND/AIMS: A long QT(C) interval has been described in a substantial fraction of patients with cirrhosis, but information on QT variation and dispersion is sparse. The aim was to determine QT variation with time and QT dispersion (QT(disp)). METHODS: The study population comprised 23 patients with cirrhosis, undergoing a haemodynamic investigation. 24-h 12 lead Holter monitoring provided information on QT and heart rate variability. RESULTS: Mean QT(C) was above upper normal limit (440 ms(1/2)) in eleven patients (47%) and significantly higher than in controls (441 vs 400 ms(1/2), p<0.01). The minimum value of QT(C) (but not the maximum value) showed a significant diurnal variation both in cirrhosis and controls. QT(disp) in cirrhosis and controls was similar (33 vs 36 ms, ns), but related to indicators of liver dysfunction, central circulation time, and arterial blood pressure (r=0.44-0.58, p=0.03-0.001). No diurnal variation of QT(disp) was found in cirrhosis. Heart rate variability was reduced with a significant relation to central hypovolaemia (r=0.55, p=0.01). CONCLUSIONS: Twenty-four hours QT(C) is prolonged in a substantial fraction of patients with cirrhosis, but with normal diurnal variation. The combination of long QT(C) and normal QT(disp) suggests delayed myocyte repolarisation on the cellular level, rather than temporal and spatial heterogeneity in the myocardial wall.