Abstract
Necrotizing enterocolitis (NEC) is a severe, currently untreatable intestinal disease that predominantly affects preterm infants and is driven by poorly characterized inflammatory pathways. ...Here, human and murine NEC intestines exhibit an unexpected predominance of type 3/T
H
17 polarization. In murine NEC, pro-inflammatory type 3 NKp46
−
RORγt
+
Tbet
+
innate lymphoid cells (ILC3) are 5-fold increased, whereas ILC1 and protective NKp46
+
RORγt
+
ILC3 are obliterated. Both species exhibit dysregulation of intestinal TLR repertoires, with TLR4 and TLR8 increased, but TLR5-7 and TLR9-12 reduced. Transgenic IL-37 effectively protects mice from intestinal injury and mortality, whilst exogenous IL-37 is only modestly efficacious. Mechanistically, IL-37 favorably modulates immune homeostasis, TLR repertoires and microbial diversity. Moreover, IL-37 and its receptor IL-1R8 are reduced in human NEC epithelia, and IL-37 is lower in blood monocytes from infants with NEC and/or lower birthweight. Our results on NEC pathomechanisms thus implicate type 3 cytokines, TLRs and IL-37 as potential targets for novel NEC therapies.
Systemic relapses remain a major problem in locally advanced rectal cancer. Using short-course radiotherapy followed by chemotherapy and delayed surgery, the Rectal cancer And Preoperative Induction ...therapy followed by Dedicated Operation (RAPIDO) trial aimed to reduce distant metastases without compromising locoregional control.
In this multicentre, open-label, randomised, controlled, phase 3 trial, participants were recruited from 54 centres in the Netherlands, Sweden, Spain, Slovenia, Denmark, Norway, and the USA. Patients were eligible if they were aged 18 years or older, with an Eastern Cooperative Oncology Group (ECOG) performance status of 0–1, had a biopsy-proven, newly diagnosed, primary, locally advanced rectal adenocarcinoma, which was classified as high risk on pelvic MRI (with at least one of the following criteria: clinical tumour cT stage cT4a or cT4b, extramural vascular invasion, clinical nodal cN stage cN2, involved mesorectal fascia, or enlarged lateral lymph nodes), were mentally and physically fit for chemotherapy, and could be assessed for staging within 5 weeks before randomisation. Eligible participants were randomly assigned (1:1), using a management system with a randomly varying block design (each block size randomly chosen to contain two to four allocations), stratified by centre, ECOG performance status, cT stage, and cN stage, to either the experimental or standard of care group. All investigators remained masked for the primary endpoint until a prespecified number of events was reached. Patients allocated to the experimental treatment group received short-course radiotherapy (5 × 5 Gy over a maximum of 8 days) followed by six cycles of CAPOX chemotherapy (capecitabine 1000 mg/m2 orally twice daily on days 1–14, oxaliplatin 130 mg/m2 intravenously on day 1, and a chemotherapy-free interval between days 15–21) or nine cycles of FOLFOX4 (oxaliplatin 85 mg/m2 intravenously on day 1, leucovorin folinic acid 200 mg/m2 intravenously on days 1 and 2, followed by bolus fluorouracil 400 mg/m2 intravenously and fluorouracil 600 mg/m2 intravenously for 22 h on days 1 and 2, and a chemotherapy-free interval between days 3–14) followed by total mesorectal excision. Choice of CAPOX or FOLFOX4 was per physician discretion or hospital policy. Patients allocated to the standard of care group received 28 daily fractions of 1·8 Gy up to 50·4 Gy or 25 fractions of 2·0 Gy up to 50·0 Gy (per physician discretion or hospital policy), with concomitant twice-daily oral capecitabine 825 mg/m2 followed by total mesorectal excision and, if stipulated by hospital policy, adjuvant chemotherapy with eight cycles of CAPOX or 12 cycles of FOLFOX4. The primary endpoint was 3-year disease-related treatment failure, defined as the first occurrence of locoregional failure, distant metastasis, new primary colorectal tumour, or treatment-related death, assessed in the intention-to-treat population. Safety was assessed by intention to treat. This study is registered with the EudraCT, 2010-023957-12, and ClinicalTrials.gov, NCT01558921, and is now complete.
Between June 21, 2011, and June 2, 2016, 920 patients were enrolled and randomly assigned to a treatment, of whom 912 were eligible (462 in the experimental group; 450 in the standard of care group). Median follow-up was 4·6 years (IQR 3·5–5·5). At 3 years after randomisation, the cumulative probability of disease-related treatment failure was 23·7% (95% CI 19·8–27·6) in the experimental group versus 30·4% (26·1–34·6) in the standard of care group (hazard ratio 0·75, 95% CI 0·60–0·95; p=0·019). The most common grade 3 or higher adverse event during preoperative therapy in both groups was diarrhoea (81 18% of 460 patients in the experimental group and 41 9% of 441 in the standard of care group) and neurological toxicity during adjuvant chemotherapy in the standard of care group (16 9% of 187 patients). Serious adverse events occurred in 177 (38%) of 460 participants in the experimental group and, in the standard of care group, in 87 (34%) of 254 patients without adjuvant chemotherapy and in 64 (34%) of 187 with adjuvant chemotherapy. Treatment-related deaths occurred in four participants in the experimental group (one cardiac arrest, one pulmonary embolism, two infectious complications) and in four participants in the standard of care group (one pulmonary embolism, one neutropenic sepsis, one aspiration, one suicide due to severe depression).
The observed decreased probability of disease-related treatment failure in the experimental group is probably indicative of the increased efficacy of preoperative chemotherapy as opposed to adjuvant chemotherapy in this setting. Therefore, the experimental treatment can be considered as a new standard of care in high-risk locally advanced rectal cancer.
Dutch Cancer Foundation, Swedish Cancer Society, Spanish Ministry of Economy and Competitiveness, and Spanish Clinical Research Network.
Essentials
rVIII‐SingleChain is a unique recombinant factor VIII (FVIII) molecule.
A population pharmacokinetic model was based on FVIII activity of severe hemophilia A patients.
The model was used ...to simulate factor VIII activity–time profiles for various dosing scenarios.
The model supports prolonged dosing of rVIII‐SingleChain with intervals of up to twice per week.
Summary
Background
Single‐chain recombinant coagulation factor VIII (rVIII‐SingleChain) is a unique recombinant coagulation factor VIII molecule.
Objectives
To: (i) characterize the population pharmacokinetics (PK) of rVIII‐SingleChain in patients with severe hemophilia A; (ii) identify correlates of variability in rVIII‐SingleChain PK; and (iii) simulate various dosing scenarios of rVIII‐SingleChain.
Patients/Methods
A population PK model was developed, based on FVIII activity levels of 130 patients with severe hemophilia A (n = 91 for ≥ 12–65 years; n = 39 for < 12 years) who had participated in a single‐dose PK investigation with rVIII‐SingleChain 50 IU kg−1. PK sampling was performed for up to 96 h.
Results
A two‐compartment population PK model with first‐order elimination adequately described FVIII activity. Body weight and predose level of von Willebrand factor were significant covariates on clearance, and body weight was a significant covariate on the central distribution volume. Simulations using the model with various dosing scenarios estimated that > 85% and > 93% of patients were predicted to maintain FVIII activity level above 1 IU dL−1, at all times with three‐times‐weekly dosing (given on days 0, 2, and 4.5) at the lowest (20 IU kg−1) and highest (50 IU kg−1) doses, respectively. For twice weekly dosing (days 0 and 3.5) of 50 IU kg−1 rVIII‐SingleChain, 62–80% of patients across all ages were predicted to maintain a FVIII activity level above 1 IU dL−1 at day 7.
Conclusions
The population PK model adequately characterized rVIII‐SingleChain PK, and the model can be utilized to simulate FVIII activity–time profiles for various dosing scenarios.
The right optic nerve of adult, 6 month to 1 year old, female and male Danio rerio were crushed and collected three days after. Matching controls of uninjured left optic nerves were also collected. ...The tissue was dissected from euthanized fish and frozen on dry ice. Samples were pooled for each category (female crush, female control, male crush, male control) n = 24 to obtain sufficient tissue for analysis. The brain from one male fish was also collected for control/calibration. Lipid extraction was done with the Bligh and Dyer 1 method, followed by untargeted liquid chromatography-mass spectrometry (LC MS-MS) lipid profiling using a Q-Exactive Orbitrap instrument coupled with Vanquish Horizon Binary UHPLC LC-MS system. The lipids were identified and quantified with LipidSearch 4.2.21 and the statistical analysis was conducted through Metaboanalyst 5.0. This data is available at Metabolomics Workbench, Study ID ST001725.
Zebrafish (Danio rerio) have the capacity for successful adult optic nerve regeneration. In contrast, mammals lack this intrinsic ability and undergo irreversible neurodegeneration seen in glaucoma ...and other optic neuropathies. Optic nerve regeneration is often studied using optic nerve crush, a mechanical neurodegenerative model. Untargeted metabolomic studies within successful regenerative models are deficient. Evaluation of tissue metabolomic changes in active zebrafish optic nerve regeneration can elucidate prioritized metabolite pathways that can be targeted in mammalian systems for therapeutic development. Female and male (6 month to 1 year old wild type) right zebrafish optic nerves were crushed and collected three days after. Contralateral, uninjured optic nerves were collected as controls. The tissue was dissected from euthanized fish and frozen on dry ice. Samples were pooled for each category (female crush, female control, male crush, male control) and pooled at n = 31 to obtain sufficient metabolite concentrations for analysis. Optic nerve regeneration at 3 days post crush was demonstrated by microscope visualization of GFP fluorescence in Tg(gap43:GFP) transgenic fish. Metabolites were extracted using a Precellys Homogenizer and a serial extraction method: (1) 1:1 Methanol/Water and (2) 8:1:1 Acetonitrile/Methanol/Acetone. Metabolites were analyzed by untargeted liquid chromatography-mass spectrometry (LC MS-MS) profiling using a Q-Exactive Orbitrap instrument coupled with Vanquish Horizon Binary UHPLC LC-MS system. Metabolites were identified and quantified using Compound Discoverer 3.3 and isotopic internal metabolites standards.
A human therapeutic that specifically modulates skeletal muscle growth would potentially provide a benefit for a variety of conditions including sarcopenia, cachexia, and muscular dystrophy. ...Myostatin, a member of the TGF-β family of growth factors, is a known negative regulator of muscle mass, as mice lacking the myostatin gene have increased muscle mass. Thus, an inhibitor of myostatin may be useful therapeutically as an anabolic agent for muscle. However, since myostatin is expressed in both developing and adult muscles, it is not clear whether it regulates muscle mass during development or in adults. In order to test the hypothesis that myostatin regulates muscle mass in adults, we generated an inhibitory antibody to myostatin and administered it to adult mice. Here we show that mice treated pharmacologically with an antibody to myostatin have increased skeletal muscle mass and increased grip strength. These data show for the first time that myostatin acts postnatally as a negative regulator of skeletal muscle growth and suggest that myostatin inhibitors could provide a therapeutic benefit in diseases for which muscle mass is limiting.
There is great potential for host-based gene expression analysis to impact the early diagnosis of infectious diseases. In particular, the influenza pandemic of 2009 highlighted the challenges and ...limitations of traditional pathogen-based testing for suspected upper respiratory viral infection. We inoculated human volunteers with either influenza A (A/Brisbane/59/2007 (H1N1) or A/Wisconsin/67/2005 (H3N2)), and assayed the peripheral blood transcriptome every 8 hours for 7 days. Of 41 inoculated volunteers, 18 (44%) developed symptomatic infection. Using unbiased sparse latent factor regression analysis, we generated a gene signature (or factor) for symptomatic influenza capable of detecting 94% of infected cases. This gene signature is detectable as early as 29 hours post-exposure and achieves maximal accuracy on average 43 hours (p = 0.003, H1N1) and 38 hours (p-value = 0.005, H3N2) before peak clinical symptoms. In order to test the relevance of these findings in naturally acquired disease, a composite influenza A signature built from these challenge studies was applied to Emergency Department patients where it discriminates between swine-origin influenza A/H1N1 (2009) infected and non-infected individuals with 92% accuracy. The host genomic response to Influenza infection is robust and may provide the means for detection before typical clinical symptoms are apparent.
To report endothelial cell loss (ECL) caused by a novel S-stamp preparation technique for Descemet membrane endothelial keratoplasty (DMEK).
Six cadaveric human corneas were prepared for DMEK ...transplantation using a single standardized technique, including the application of a dry ink gentian violet S-stamp to the stromal side of Descemet membrane. Endothelial cell death was evaluated and quantified using computerized analysis of vital dye staining.
ECL caused by the S-stamp was 0.6% (range 0.1%-1.0%), which comprised less than one-tenth of the total ECL caused by our preparation of the DMEK graft from the start to finish, including recovery, prestripping, S-stamping, and trephination (13.7% total ECL, range 9.9%-17.6%).
Our novel S-stamp donor tissue preparation technique is intuitive to learn and holds the promise of preventing iatrogenic primary graft failure due to upside-down grafts without causing unacceptable increases in ECL.
Essentials
rVIII‐SingleChain is a novel recombinant factor VIII with covalently bonded heavy and light chains.
Efficacy, safety and pharmacokinetics were studied in pediatric patients with severe ...hemophilia A.
Across all prophylaxis regimens, the median annualized spontaneous bleeding rate was 0.00.
rVIII‐SingleChain showed excellent hemostatic efficacy and a favorable safety profile.
Summary
Background
rVIII‐SingleChain is a novel B‐domain truncated recombinant factor VIII (rFVIII) comprised of covalently bonded FVIII heavy and light chains, demonstrating a high binding affinity to von Willebrand factor.
Objectives
This phase III study investigated the safety, efficacy and pharmacokinetics of rVIII‐SingleChain in previously treated pediatric patients < 12 years of age with severe hemophilia A.
Patients/Methods
Patients could be assigned to prophylaxis or on‐demand therapy by the investigator. For patients assigned to prophylaxis, the treatment regimen and dose were based on the bleeding phenotype. For patients receiving on‐demand therapy, dosing was guided by World Federation of Hemophilia recommendations. The primary endpoint was treatment success, defined as a rating of ‘excellent’ or ‘good’ on the investigator's clinical assessment of hemostatic efficacy for all treated bleeding events.
Results
The study enrolled 84 patients (0 to < 6 years, n = 35; ≥ 6 to < 12 years, n = 49); 81 were assigned to prophylaxis and three to an on‐demand regimen. Patients accumulated a total of 5239 exposure days (EDs), with 65 participants reaching > 50 EDs. In the 347 bleeds treated and evaluated by the investigator, hemostatic efficacy was rated as excellent or good in 96.3%. The median annualized spontaneous bleeding rate was 0.00 (Q1, Q3: 0.00, 2.20), and the median annualized bleeding rate was 3.69 (Q1, Q3: 0.00, 7.20) across all prophylaxis regimens. No participant developed an inhibitor.
Conclusions
rVIII‐SingleChain is a novel rFVIII molecule showing excellent hemostatic efficacy and a favorable safety profile in a clinical study in children < 12 years of age with severe hemophilia A.
Objectives: The purpose of this work was to study the genetic determinants responsible for extended-spectrum β-lactamase (ESBL) resistance of Salmonella isolated from Dutch poultry, poultry meat and ...hospitalized humans. Methods: Thirty-four ESBL-resistant Salmonella isolates from The Netherlands were tested towards 21 antimicrobial agents. PCR and sequencing were used to determine the underlying genetic determinants responsible for the ESBL phenotypes. The transferability of the ESBL phenotypes was tested by conjugation to a susceptible Salmonella enterica serovar Dublin and plasmid purification, restriction fragment length polymorphism (RFLP) and pulsed-field gel electrophoresis (PFGE) were employed to further characterize a subset of the isolates. Results: A great genetic diversity was seen among the isolates. The blaTEM-52 gene was most predominant and was found among Salmonella enterica serovars Blockley, Thomson, London, Enteritidis phage type 14b, Paratyphi B, Virchow and Typhimurium phage types 11 and 507. We also found the blaTEM-20 gene in S. Paratyphi B var. Java and the blaTEM-63 gene in S. Isangi. Furthermore, we detected the blaCTX-M-28 gene in S. Isangi and the blaCTX-M-3 gene in S. Typhimurium phage type 507. The blaCTX-M-2 gene was identified in S. Virchow, which also contained a copy of the blaSHV-2 gene and a copy of the blaTEM-1 gene. The blaSHV-12 gene was found alone in S. Concord and together with the blaTEM-52 gene in S. Typhimurium. Finally, the blaACC-1 gene was cloned from a S. Bareilly isolate and was found to be present on indistinguishable plasmids in all S. Bareilly isolates examined as well as in a S. Braenderup isolate and a S. Infantis isolate. Conclusions: Our data underscore the diversity of ESBL genes in Salmonella enterica isolated from animals, food products and human patients.
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