Exploring the interdependence of calcium and chloride activation of O2 evolution in photosystem II.

Calcium and chloride are activators of oxygen evolution in photosystem II (PSII), the light-absorbing water oxidase of higher plants, algae, and cyanobacteria. Calcium is an essential part of the catalytic Mn4CaO5 cluster that carries out water oxidation and chloride has two nearby binding sites, one of which is associated with a major water channel. The co-activation of oxygen evolution by the two ions is examined in higher plant PSII lacking the extrinsic PsbP and PsbQ subunits using a bisubstrate enzyme kinetics approach. Analysis of three different preparations at pH 6.3 indicates that the Michaelis constant, KM, for each ion is less than the dissociation constant, KS, and that the affinity of PSII for Ca2+ is about ten-fold greater than for Cl-, in agreement with previous studies. Results are consistent with a sequential binding model in which either ion can bind first and each promotes the activation by the second ion. At pH 5.5, similar results are found, except with a higher affinity for Cl- and lower affinity for Ca2+. Observation of the slow-decaying Tyr Z radical, YZ•, at 77 K and the coupled S2YZ• radical at 10 K, which are both associated with Ca2+ depletion, shows that Cl- is necessary for their observation. Given the order of electron and proton transfer events, this indicates that chloride is required to reach the S3 state preceding Ca2+ loss and possibly for stabilization of YZ• after it forms. Interdependence through hydrogen bonding is considered in the context of the water environment that intervenes between Cl- at the Cl-1 site and the Ca2+/Tyr Z region.

Dose escalation of tolinapant (ASTX660) in combination with standard radical chemoradiotherapy in cervical cancer : a study protocol for a phase 1b TiTE-CRM clinical trial (CRAIN) in UK secondary care centres.

BACKGROUND: Cervical cancer is the fourth most common cancer in women, with an estimated 342,000 deaths worldwide in 2020. Current standard of care in the UK for locally advanced cervical cancer is concurrent chemoradiotherapy with weekly cisplatin, yet 5-year overall survival rates are only 65% with a distant relapse rate of 50%. Inhibitors of Apoptosis Proteins (IAPs) are often overexpressed in cancer cells and associated with tumour progression and resistance to treatment. Tolinapant, developed by Astex Pharmaceuticals, is an IAP antagonist with an additional mechanism of action via down-regulation of NF-kB, an important regulator in cervical cancer. Preclinical studies performed using tolinapant in combination with cisplatin and radiotherapy showed inhibition of tumour growth and enhanced survival. There is therefore a strong rationale to combine tolinapant with chemoradiotherapy (CRT). METHODS: CRAIN is a phase Ib open-label, dose escalation study to characterise the safety, tolerability and initial evidence for clinical activity of tolinapant when administered in combination with cisplatin based CRT. Up to 42 patients with newly diagnosed cervix cancer will be recruited from six UK secondary care sites. The number of participants and the duration of the trial will depend on toxicities observed and dose escalation decisions, utilising a TiTE-CRM statistical design. Treatment will constist of standard of care CRT with 45 Gy external beam radiotherapy given in 25 daily fractions over 5 weeks with weekly cisplatin 40mg/m2. This is followed by brachytherapy for which common schedules will be 28 Gy in 4 fractions high-dose-rate or 34 Gy in 2 fractions pulsed-dose-rate. Tolinapant will be administered in fixed dose capsules taken orally daily for seven consecutive days as an outpatient on alternate weeks (weeks 1, 3, 5) during chemoradiation. Dose levels for tolinapant which will be assessed are: 60 mg; 90 mg (starting level); 120 mg; 150 mg; 180 mg. Escalation will be guided by emerging safety data and decisions by the Safety Review Committee. DISCUSSION: If this trial determines a recommended phase II dose and shows tolinapant to be safe and effective in combination with CRT, it would warrant future phase trials. Ultimately, we hope to provide a synergistic treatment option for these patients to improve outcome. TRIAL REGISTRATIONS: EudraCT Number: 2021-006555-34 (issued 30th November 2021); ISRCTN18574865 (registered 30th August 2022).

Enhancing Nutrition Support for Esophageal Cancer Patients: Understanding Factors Influencing Feeding Tube Utilization.

Objective: We sought to identify factors that can predict esophageal cancer (EC) patients at high risk of requiring feeding tube insertion. Methods: A retrospective cohort review was conducted, including all patients diagnosed with EC at our cancer center from 2013 to 2018. Multivariate logistic regression was performed comparing the group that required a reactive feeding tube insertion to those who did not require any feeding tube insertion to identify risk factors. Results: A total of 350 patients were included in the study, and 132/350 (38%) patients received a feeding tube. 50 out of 132 (38%) patients had feeding tube inserted reactively. Severe dysphagia (OR 19.9, p < 0.001) at diagnosis and decision to undergo chemotherapy (OR 2.8, p = 0.008) appeared to be predictors for reactive feeding tube insertion. The reactive insertion group had a 7% higher rate of complications relating to feeding tube. Conclusion: Severe dysphagia at diagnosis and undergoing chemotherapy were identified as risk factors for requiring a feeding tube. Ultimately, the aim is to create a predictive tool that utilizes these risks factors to accurate identify high-risk patients who may benefit from prophylactic feeding tube insertion.

BRCA1/Trp53 heterozygosity and replication stress drive esophageal cancer development in a mouse model.

BRCA1 germline mutations are associated with an increased risk of breast and ovarian cancer. Recent findings of others suggest that BRCA1 mutation carriers also bear an increased risk of esophageal and gastric cancer. Here, we employ a Brca1/Trp53 mouse model to show that unresolved replication stress (RS) in BRCA1 heterozygous cells drives esophageal tumorigenesis in a model of the human equivalent. This model employs 4-nitroquinoline-1-oxide (4NQO) as an RS-inducing agent. Upon drinking 4NQO-containing water, Brca1 heterozygous mice formed squamous cell carcinomas of the distal esophagus and forestomach at a much higher frequency and speed (∼90 to 120 d) than did wild-type (WT) mice, which remained largely tumor free. Their esophageal tissue, but not that of WT control mice, revealed evidence of overt RS as reflected by intracellular CHK1 phosphorylation and 53BP1 staining. These Brca1 mutant tumors also revealed higher genome mutation rates than those of control animals; the mutational signature SBS4, which is associated with tobacco-induced tumorigenesis; and a loss of Brca1 heterozygosity (LOH). This uniquely accelerated Brca1 tumor model is also relevant to human esophageal squamous cell carcinoma, an often lethal tumor.

Evaluation of a rapid sequence induction technique in dogs with or without rocuronium.

OBJECTIVE: To determine, using a rapid sequence induction (RSI) technique, whether rocuronium improves the quality and speed of endotracheal intubation in healthy dogs. STUDY DESIGN: Randomized, crossover, experimental study. ANIMALS: Six adult intact male Beagles (12.3 ± 0.4 kg). METHODS: Dogs were premedicated with intravenous acepromazine (0.03 mg kg-1) and hydromorphone (0.1 mg kg-1). Ten minutes later, anesthesia was induced with intravenous propofol (2 mg kg-1 over 5 seconds), followed by saline (0.06 mL kg-1, CT group) or rocuronium (0.6 mg kg-1, RT group), with orotracheal intubation attempted after 45 seconds. Intubation time (IT) and conditions (IC) were assessed. PaO2, PaCO2, arterial blood pH and serum cortisol were obtained before and after RSI. After endotracheal intubation, saline (0.04 mL kg-1) or sugammadex (4 mg kg-1) were administered intravenously in CT or RT groups, respectively. Spontaneous ventilation restoration was noted. RESULTS: The IT was 54.3 ± 6.9 (mean ± SD) and 57.8 ± 5.2 seconds for CT and RT, respectively (p = 0.385). All laryngoscopies indicated good IC in both treatment groups. Heart rate was lower in CT group than in RT group (66 ± 16 versus 103 ± 39 beats minute-1, p = 0.016). PaCO2, pH, PaO2 and cortisol did not differ between treatments. Compared with baseline, PaCO2 increased from 47.7 ± 6.2 to 58.8 ± 5.8 (p < 0.001) and pH decreased from 7.35 ± 0.04 to 7.28 ± 0.04 (p = 0.003), independent of treatment. Dogs in both treatment groups returned to spontaneous ventilation within 30 seconds of RSI. CONCLUSIONS AND CLINICAL RELEVANCE: RSI resulted in respiratory acidosis without hypoxemia or increased cortisol. Rocuronium did not improve IT or IC. Spontaneous ventilation was observed immediately after administering saline or sugammadex. The co-administration of rocuronium showed no clinical benefits over propofol alone in RSI in healthy dogs.

Quantitative SWATH-based proteomic profiling of urine for the identification of endometrial cancer biomarkers in symptomatic women.

BACKGROUND: A non-invasive endometrial cancer detection tool that can accurately triage symptomatic women for definitive testing would improve patient care. Urine is an attractive biofluid for cancer detection due to its simplicity and ease of collection. The aim of this study was to identify urine-based proteomic signatures that can discriminate endometrial cancer patients from symptomatic controls. METHODS: This was a prospective case-control study of symptomatic post-menopausal women (50 cancers, 54 controls). Voided self-collected urine samples were processed for mass spectrometry and run using sequential window acquisition of all theoretical mass spectra (SWATH-MS). Machine learning techniques were used to identify important discriminatory proteins, which were subsequently combined in multi-marker panels using logistic regression. RESULTS: The top discriminatory proteins individually showed moderate accuracy (AUC > 0.70) for endometrial cancer detection. However, algorithms combining the most discriminatory proteins performed well with AUCs > 0.90. The best performing diagnostic model was a 10-marker panel combining SPRR1B, CRNN, CALML3, TXN, FABP5, C1RL, MMP9, ECM1, S100A7 and CFI and predicted endometrial cancer with an AUC of 0.92 (0.96-0.97). Urine-based protein signatures showed good accuracy for the detection of early-stage cancers (AUC 0.92 (0.86-0.9)). CONCLUSION: A patient-friendly, urine-based test could offer a non-invasive endometrial cancer detection tool in symptomatic women. Validation in a larger independent cohort is warranted.

Proteomic analysis identifies subgroups of patients with active systemic lupus erythematosus.

OBJECTIVE: Systemic lupus erythematosus (SLE) is a clinically and biologically heterogenous autoimmune disease. We aimed to investigate the plasma proteome of patients with active SLE to identify novel subgroups, or endotypes, of patients. METHOD: Plasma was collected from patients with active SLE who were enrolled in the British Isles Lupus Assessment Group Biologics Registry (BILAG-BR). The plasma proteome was analysed using a data-independent acquisition method, Sequential Window Acquisition of All theoretical mass spectra mass spectrometry (SWATH-MS). Unsupervised, data-driven clustering algorithms were used to delineate groups of patients with a shared proteomic profile. RESULTS: In 223 patients, six clusters were identified based on quantification of 581 proteins. Between the clusters, there were significant differences in age (p = 0.012) and ethnicity (p = 0.003). There was increased musculoskeletal disease activity in cluster 1 (C1), 19/27 (70.4%) (p = 0.002) and renal activity in cluster 6 (C6) 15/24 (62.5%) (p = 0.051). Anti-SSa/Ro was the only autoantibody that significantly differed between clusters (p = 0.017). C1 was associated with p21-activated kinases (PAK) and Phospholipase C (PLC) signalling. Within C1 there were two sub-clusters (C1A and C1B) defined by 49 proteins related to cytoskeletal protein binding. C2 and C6 demonstrated opposite Rho family GTPase and Rho GDI signalling. Three proteins (MZB1, SND1 and AGL) identified in C6 increased the classification of active renal disease although this did not reach statistical significance (p = 0.0617). CONCLUSIONS: Unsupervised proteomic analysis identifies clusters of patients with active SLE, that are associated with clinical and serological features, which may facilitate biomarker discovery. The observed proteomic heterogeneity further supports the need for a personalised approach to treatment in SLE.

The impact of phosphorylated PTEN at threonine 366 on cortical connectivity and behaviour.

The lipid phosphatase PTEN (phosphatase and tensin homologue on chromosome 10) is a key tumour suppressor gene and an important regulator of neuronal signalling. PTEN mutations have been identified in patients with autism spectrum disorders, characterized by macrocephaly, impaired social interactions and communication, repetitive behaviour, intellectual disability, and epilepsy. PTEN enzymatic activity is regulated by a cluster of phosphorylation sites at the C-terminus of the protein. Here, we focused on the role of PTEN T366 phosphorylation and generated a knock-in mouse line in which Pten T366 was substituted with alanine (PtenT366A/T366A). We identify that phosphorylation of PTEN at T366 controls neuron size and connectivity of brain circuits involved in sensory processing. We show in behavioural tests that PtenT366/T366A mice exhibit cognitive deficits and selective sensory impairments, with significant differences in male individuals. We identify restricted cellular overgrowth of cortical neurons in PtenT366A/T366A brains, linked to increases in both dendritic arborization and soma size. In a combinatorial approach of anterograde and retrograde monosynaptic tracing using rabies virus, we characterize differences in connectivity to the primary somatosensory cortex of PtenT366A/T366A brains, with imbalances in long-range cortico-cortical input to neurons. We conclude that phosphorylation of PTEN at T366 controls neuron size and connectivity of brain circuits involved in sensory processing and propose that PTEN T366 signalling may account for a subset of autism-related functions of PTEN.

Accuracy, precision, and interobserver and intraobserver agreements related to pressure-measurement devices.

OBJECTIVE: To evaluate the accuracy, precision, and observer agreement of three pressure measurement devices. STUDY DESIGN: In vitro model study. SAMPLE POPULATION: Water manometer with built-in gauge (WMg), arterial pressure transducer (APT), and Compass CT (CCT). METHODS: The model was set to five predetermined pressures (4, 8, 13, 17, and 24 cm H2 O) using a water manometer with a ruler (WMr) as the gold standard. Each device was tested at each pressure in a randomized order by three investigators. Bland-Altman plots were used to assess agreement between devices. Intraclass correlation coefficients (ICC) were calculated for interobserver and intraobserver agreements. RESULTS: The mean differences (cm H2 O) ± SEM in comparison with the set pressure were -0.020 ± 0.010 (WMg), -0.390 ± 0.077 (APT), and -1.267 ± 0.213 (CCT). Pressures measured by WMg did not differ from those measured by WMr. Pressures measured by all devices did not differ from each other (p > .062 for all comparisons). Interobserver agreement was excellent (1.000), and intraobserver agreement was excellent (0.985, 0.990, 0.998 for each observer). CONCLUSION: Compared to the WMr, the WMg was the most accurate and precise, followed by the APT; the CCT was the least accurate and precise. Interobserver and intraobserver agreements for all three devices were excellent. CLINICAL SIGNIFICANCE: The largest mean difference of all devices was within 1.3 cm H2 O of the set pressure, indicating possible clinical utility of any of the devices. However, WMr or WMg should be considered first due to their high precision and accuracy.

Longitudinal evaluation of fentanyl concentrations in equine plasma and synovial fluid following application of transdermal fentanyl patches over one carpal joint.

OBJECTIVE: To determine if transdermally delivered fentanyl can achieve greater concentrations of fentanyl in synovial fluid when applied over a synovial structure. STUDY DESIGN: Randomized, experimental study. ANIMALS: Six healthy adult horses. METHODS: Each horse had two 100 µg/h fentanyl matrix patches applied on the dorsal aspect of one, randomly assigned, carpometacarpal joint (CMCJ) for 48 h. Whole blood and bilateral synovial samples from the intercarpal joint were obtained at 0, 2, 6, 12, 24, 36 and 48 h. Fentanyl concentrations were measured with liquid chromatography-mass spectrometry. RESULTS: All subjects achieved detectable concentrations of fentanyl in both plasma and synovial fluid. Time to peak synovial and plasma concentration was 12 h. At 6 h, the synovial concentration in the untreated carpus (0.104 ng/mL ± 0.106) was lower than plasma fentanyl concentrations 0.31 ± 0.27 (p = .036). At 12 h, both treated (0.55 ng/mL ± 0.3) and untreated (0.53 ng/mL ± 0.28) synovial fluid fentanyl concentrations were lower than plasma (0.87 ng/mL ± 0.48) concentrations (p < .001 and p = .001, respectively). Synovial concentrations of fentanyl did not differ between treated and untreated joints (p > 0.608 for all time points). CONCLUSION: Application of fentanyl matrix patches directly over the CMCJ did not result in increased fentanyl concentrations in the synovial fluid of the treated intercarpal joint in normal horses. CLINICAL SIGNIFICANCE: There is likely no analgesic advantage to placing fentanyl patches directly over the affected joint, as it did not result in increased synovial concentrations at the tested site.

Antinociceptive effects of bupivacaine injected within the internal abdominis rectus sheath in standing healthy horses.

OBJECTIVE: To evaluate a regional anesthetic technique for blocking the abdominal midline in horses. STUDY DESIGN: Anatomical description and prospective, crossover, placebo-controlled, blinded study. ANIMALS: Adult horses; two cadavers, six healthy animals. METHODS: In stage 1, 0.5% methylene blue with 0.25% bupivacaine (0.5 mL kg-1) was injected using ultrasonography into the internal rectus abdominis sheath (RAS) of two cadavers with a one-point or two-point technique. The dye spread was described after the dissection of the abdomens. In stage 2, each horse was injected with 1 mL kg-1 of 0.9% NaCl (treatment PT) or 0.2% bupivacaine (treatment BT) using a two-point technique. The abdominal midline mechanical nociceptive threshold (MNT) was measured with a 1 mm blunted probe tip and results analyzed with mixed-effect anova. Signs of pelvic limb weakness were recorded. RESULTS: The cadaver dissections showed staining of the ventral branches from the eleventh thoracic (T11) to the second lumbar (L2) nerve with the one-point technique and T9-L2 with the two-point technique. Baseline MNTs were, mean ± standard deviation, 12.6 ± 1.6 N and 12.4 ± 2.4 N in treatments PT and BT, respectively. MNT increased to 18.9 ± 5.8 N (p = 0.010) at 30 minutes, and MNT was between 9.4 ± 2.0 and 15.3 ± 3.4 N from 1 to 8 hours (p > 0.521) in treatment PT. MNTs in treatment BT were 21.1 ± 5.9 to 25.0 ± 0.1 N from 30 minutes to 8 hours (p < 0.001). MNTs after the RAS injections were higher in treatment BT than PT (p = 0.007). No pelvic limb weakness was observed. CONCLUSIONS AND CLINICAL RELEVANCE: Antinociception of at least 8 hours without pelvic limb weakness was observed in the abdominal midline in standing horses after the RAS block. Further investigations are necessary to evaluate suitability for ventral celiotomies.

Comparison of Traditional Lecture-Based Learning versus Interactive Electronic Book Learning in Veterinary Student Comprehension of Inhalant Anesthetic Administration, Uptake, and Elimination.

The administration, uptake, and elimination of inhalant anesthetics is a challenging topic in the veterinary curriculum, and lecture-based learning is often insufficient to ensure that students understand these concepts. We hypothesized that the use of an interactive electronic book (e-book) would enhance student comprehension of the material. Two sequential Doctorate of Veterinary Medicine student cohorts participated in a prospective controlled study. The first cohort received traditional lecture-based learning while the second cohort was taught the topic using an interactive e-book. Student comprehension of the material was assessed twice during the course via multiple-choice questions: five questions in a midcourse quiz and seven within the final exam. At the end of the course, students also completed a Likert survey assessing their confidence regarding the topic. Averaged across assessment types, students taught using the interactive e-book scored higher than those taught via the traditional method (p < .001). Final exam scores were significantly higher in the e-book cohort compared with the lecture-based cohort (p < .001). However, there was no difference in quiz scores between groups (p = .109). No significant difference was found between groups in responses to the Likert survey. In conclusion, students using the interactive e-book had better comprehension of the material than students in the traditional lecture group as measured by their scores on multiple-choice question assessments. Future studies are needed to determine whether this advantage persists later in the curriculum when students apply these concepts in the clinical year.

The Head Control Scale: Inter-Rater Reliability among Therapy Students.

Research has demonstrated the Head Control Scale to have almost perfect inter-rater reliability when utilized by experienced clinicians. This study examines if further clinical experience or additional training is required to use the scale for assessment of head control in a reliable and effective manner. First and second year physical and occupational therapy students were shown five videotaped subjects of varying ages and abilities in four positions (supine, prone, supported sitting, and pull to sit). Students then utilized each subscale of the HCS to rate every subject on the rating scale in each of the positions. When utilizing the kappa coefficient statistic, the inter-rater reliability among student participants was "almost perfect" on each of the subscales. Thus, the Head Control Scale was found to be reliable with no significant difference in interrater reliability when used by clinicians or students suggesting that additional training or expertise is not required to reliably use this scale in clinical practice.

Novel manifestations of immune dysregulation and granule defects in gray platelet syndrome.

Gray platelet syndrome (GPS) is a rare recessive disorder caused by biallelic variants in NBEAL2 and characterized by bleeding symptoms, the absence of platelet α-granules, splenomegaly, and bone marrow (BM) fibrosis. Due to the rarity of GPS, it has been difficult to fully understand the pathogenic processes that lead to these clinical sequelae. To discern the spectrum of pathologic features, we performed a detailed clinical genotypic and phenotypic study of 47 patients with GPS and identified 32 new etiologic variants in NBEAL2. The GPS patient cohort exhibited known phenotypes, including macrothrombocytopenia, BM fibrosis, megakaryocyte emperipolesis of neutrophils, splenomegaly, and elevated serum vitamin B12 levels. Novel clinical phenotypes were also observed, including reduced leukocyte counts and increased presence of autoimmune disease and positive autoantibodies. There were widespread differences in the transcriptome and proteome of GPS platelets, neutrophils, monocytes, and CD4 lymphocytes. Proteins less abundant in these cells were enriched for constituents of granules, supporting a role for Nbeal2 in the function of these organelles across a wide range of blood cells. Proteomic analysis of GPS plasma showed increased levels of proteins associated with inflammation and immune response. One-quarter of plasma proteins increased in GPS are known to be synthesized outside of hematopoietic cells, predominantly in the liver. In summary, our data show that, in addition to the well-described platelet defects in GPS, there are immune defects. The abnormal immune cells may be the drivers of systemic abnormalities such as autoimmune disease.

Development of the Auburn Induction Scale for evaluating induction quality in dogs.

OBJECTIVE: To develop and begin establishing evidence for validity of an instrument to assess the quality of induction in dogs. STUDY DESIGN: Cross-sectional survey and video scoring. ANIMALS AND POPULATION: A total of 51 veterinary anesthesia personnel, four board-certified anesthesiologists and videos of induction of anesthesia in 18 dogs. METHODS: In Part 1, an online survey was sent to veterinary anesthesia personnel to solicit expressions and words that they associate with induction of anesthesia. These expressions were evaluated by four anesthesiologists to create a composite scale (Auburn Induction Scale). In Part 2, 18 videos were reviewed by the same four anesthesiologists on two separate occasions. The videos were scored using the Auburn Induction Scale, a simple descriptive scale (SDS) and a visual analog scale (VAS). Intra-rater and inter-rater reliability was measured using an intraclass correlation coefficient (ICC). Significance was set at p < 0.05. RESULTS: The survey yielded 51 responses that were condensed into 133 expressions. The four anesthesiologists created 18 items incorporating the 133 expressions. The mean ± standard deviation intra-rater reliability ICC was 0.81 ± 0.08 for the Auburn Induction Scale, 0.71 ± 0.02 for the SDS and 0.71 ± 0.08 for the VAS for all raters. The mean ± standard deviation inter-rater reliability ICC was 0.69 ± 0.04 for the Auburn Induction Scale, 0.61 ± 0.05 for the SDS and 0.60 ± 0.06 for the VAS. CONCLUSIONS AND CLINICAL RELEVANCE: In a research setting, widespread use of this scale may be helpful in increasing the accuracy of data and improving agreement between studies assessing induction of anesthesia in dogs. The results of this study have yielded a composite scale that is more reliable between and among raters than a unidimensional scale.

Comparison of hydromorphone and butorphanol for management of pain in equine patients undergoing elective arthroscopy: a randomized clinical trial.

OBJECTIVE: To compare the effects of hydromorphone and butorphanol in horses undergoing arthroscopy and describe the pharmacokinetics of hydromorphone in anesthetized horses. STUDY DESIGN: Randomized controlled clinical trial. ANIMALS: A total of 40 adult horses admitted for elective arthroscopy. METHODS: Horses were randomly assigned to be administered intravenous hydromorphone (0.04 mg kg-1; group TxH; n = 19) or butorphanol (0.02 mg kg-1; group TxB; n = 21) prior to surgery as part of a standardized anesthetic protocol. Pain was scored by two observers unaware of group assignment using the Equine Utrecht University Scale for Facial Assessment of Pain (EQUUS-FAP) and a composite pain scale (CPS) prior to surgery (baseline), 2 hours (P2) and 4 hours (P4) following recovery from anesthesia. Blood samples were collected at various time points for determination of plasma hydromorphone concentration using liquid chromatography-tandem mass spectrometry. Data were analyzed with a mixed-effect model. RESULTS: Median (range) baseline EQUUS-FAP was 1.2 (0.0-4.0) with no effect of group, time points or interaction. Baseline CPS was similar between groups. Group TxH baseline CPS was 2.5 (0.0-10.0), increased at P2 [4.5 (0-10.0); p = 0.046] and returned to baseline values at P4 [3.0 (0.0-11.0)]. Group TxB baseline CPS was 2.0 (0.0-8.0), increased at P2 [3.5 (0.0-11.0); p = 0.009] and P4 [5.0 (0.0-11.0); p < 0.001]. Pharmacokinetic terminal half-life was 774 ± 82.3 minutes, area under the curve was 1362 ± 314 ng minutes mL-1, clearance was 30.7 ± 7.23 mL minute-1 kg-1 and volume of distribution at steady state was 884 ± 740 mL kg-1. CONCLUSIONS: Hydromorphone, but not butorphanol, decreased CPS back to baseline at P4 after recovery. CLINICAL RELEVANCE: Hydromorphone may provide superior postoperative analgesia compared with butorphanol in horses undergoing arthroscopy.

Results of a UK National Cancer Research Institute Phase II study of brentuximab vedotin using a response-adapted design in the first-line treatment of patients with classical Hodgkin lymphoma unsuitable for chemotherapy due to age, frailty or comorbidity (BREVITY).

Standard treatment for classical Hodgkin lymphoma (cHL) is poorly tolerated in older patients and results disappointing. We assessed safety and efficacy of brentuximab vedotin (BV), in previously untreated patients with cHL unfit for standard treatment due to age, frailty or comorbidity. The primary outcome was complete metabolic response (CMR) by positron emission tomography/computed tomography after four BV cycles (PET4). The secondary outcomes included progression-free survival (PFS), overall survival (OS), and toxicity. In all, 35 patients with a median age of 77 years and median total Cumulative Illness Rating Scale for Geriatrics (CIRS-G) score of 6 were evaluable for toxicity and 31 for response. A median of four cycles were given (range one-16). In all, 14 patients required dose reduction due to toxicity and 11 patients stopped treatment due to adverse events (AEs). A total of 716 AEs were reported, of which 626 (88%) were Grade 1/2 and 27 (77%) patients had at least one AE Grade ≥3. At PET4, CMR was 25·8% [95% confidence interval (CI) 13·7-42.2%] and objective response rate 83·9% (95% CI 63·7-90·8%). Median PFS was 7·3 months (95% CI 5·2-9·0), and OS 19·5 months. Our results suggest that BV monotherapy is tolerable but suboptimal in the front-line therapy of elderly or comorbid patients with cHL. Combining BV with other agents may be more effective. Trial Registration: Clinicaltrials.gov identifier: NCT02567851.

The use of missing values in proteomic data-independent acquisition mass spectrometry to enable disease activity discrimination.

MOTIVATION: Data-independent acquisition mass spectrometry allows for comprehensive peptide detection and relative quantification than standard data-dependent approaches. While less prone to missing values, these still exist. Current approaches for handling the so-called missingness have challenges. We hypothesized that non-random missingness is a useful biological measure and demonstrate the importance of analysing missingness for proteomic discovery within a longitudinal study of disease activity. RESULTS: The magnitude of missingness did not correlate with mean peptide concentration. The magnitude of missingness for each protein strongly correlated between collection time points (baseline, 3 months, 6 months; R = 0.95-0.97, confidence interval = 0.94-0.97) indicating little time-dependent effect. This allowed for the identification of proteins with outlier levels of missingness that differentiate between the patient groups characterized by different patterns of disease activity. The association of these proteins with disease activity was confirmed by machine learning techniques. Our novel approach complements analyses on complete observations and other missing value strategies in biomarker prediction of disease activity. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Cooperation between somatic mutation and germline-encoded residues enables antibody recognition of HIV-1 envelope glycans.

Viral glycoproteins are a primary target for host antibody responses. However, glycans on viral glycoproteins can hinder antibody recognition since they are self glycans derived from the host biosynthesis pathway. During natural HIV-1 infection, neutralizing antibodies are made against glycans on HIV-1 envelope glycoprotein (Env). However, such antibodies are rarely elicited with vaccination. Previously, the vaccine-induced, macaque antibody DH501 was isolated and shown to bind to high mannose glycans on HIV-1 Env. Understanding how DH501 underwent affinity maturation to recognize glycans could inform vaccine induction of HIV-1 glycan antibodies. Here, we show that DH501 Env glycan reactivity is mediated by both germline-encoded residues that contact glycans, and somatic mutations that increase antibody paratope flexibility. Only somatic mutations in the heavy chain were required for glycan reactivity. The paratope conformation was fragile as single mutations within the immunoglobulin fold or complementarity determining regions were sufficient for eliminating antibody function. Taken together, the initial germline VHDJH rearrangement generated contact residues capable of binding glycans, and somatic mutations were required to form a flexible paratope with a cavity conducive to HIV-1 envelope glycan binding. The requirement for the presence of most somatic mutations across the heavy chain variable region provides one explanation for the difficulty in inducing anti-Env glycan antibodies with HIV-1 Env vaccination.

Influence of doxapram and intermittent 10% carbon dioxide inspiration on cardiovascular and laryngeal functions in anesthetized dogs.

OBJECTIVE: To compare the effects of two doses of doxapram intravenous injection and carbon dioxide inhalation on the cardiovascular and laryngeal functions of anesthetized hounds. STUDY DESIGN: Experimental study. ANIMALS: Six healthy adult dogs. METHODS: In a Latin-square design, the mean arterial blood pressure (MABP) and heart rate (HR) were recorded continuously. The inspiratory normalized glottic gap areas (iNGGA) were measured before and after each stimulation with 0.55 mg/kg of doxapram (L-DOX), 2.2 mg/kg of doxapram (H-DOX), or 90 s of inhalation of 10% carbon dioxide in oxygen (I-CO2 ). The stimulations were tested in duplicate or triplicate. Video clips of the laryngeal movement were scored by board-certified surgeons masked to the treatment. RESULTS: The MABP increased with L-DOX and H-DOX up to 81% (both p < .001 compared to I-CO2 ), and persisted during the other stimulations (both p < .001). An intermittent tachycardic effect of up to 79% increase in HR was observed with doxapram. The HR following H-DOX was higher than L-DOX and I-CO2 (both p < .016). Neither hypertension nor tachycardia was observed with I-CO2 . The iNGGA increased with all treatments (p < .001). The iNGGA was greater with H-DOX than L-DOX and I-CO2 (both p < .007). All treatments received higher scores (all p < .001) with acceptable inter- and intra-observers Krippendorff's alphas. CONCLUSION: All treatments were effective respiratory stimulants in anesthetized dogs; however, doxapram caused hypertension and tachycardia. CLINICAL SIGNIFICANCE: Carbon dioxide inhalation might improve arytenoid motion without cardiovascular effects in dogs during clinical airway examinations.