ABSTRACT
Understanding how and why menopause has evolved is a long-standing challenge across disciplines. Females can typically maximize their reproductive success by reproducing for the whole of their adult life. In humans, however, women cease reproduction several decades before the end of their natural lifespan1,2. Although progress has been made in understanding the adaptive value of menopause in humans3,4, the generality of these findings remains unclear. Toothed whales are the only mammal taxon in which menopause has evolved several times5, providing a unique opportunity to test the theories of how and why menopause evolves in a comparative context. Here, we assemble and analyse a comparative database to test competing evolutionary hypotheses. We find that menopause evolved in toothed whales by females extending their lifespan without increasing their reproductive lifespan, as predicted by the 'live-long' hypotheses. We further show that menopause results in females increasing their opportunity for intergenerational help by increasing their lifespan overlap with their grandoffspring and offspring without increasing their reproductive overlap with their daughters. Our results provide an informative comparison for the evolution of human life history and demonstrate that the same pathway that led to menopause in humans can also explain the evolution of menopause in toothed whales.
Subject(s)
Biological Evolution , Menopause , Models, Biological , Whales , Animals , Female , Databases, Factual , Longevity/physiology , Menopause/physiology , Reproduction/physiology , Whales/classification , Whales/physiology , HumansABSTRACT
BACKGROUND: The monoclonal antibody ublituximab enhances antibody-dependent cellular cytolysis and produces B-cell depletion. Ublituximab is being evaluated for the treatment of relapsing multiple sclerosis. METHODS: In two identical, phase 3, double-blind, double-dummy trials (ULTIMATE I and II), participants with relapsing multiple sclerosis were randomly assigned in a 1:1 ratio to receive intravenous ublituximab (150 mg on day 1, followed by 450 mg on day 15 and at weeks 24, 48, and 72) and oral placebo or oral teriflunomide (14 mg once daily) and intravenous placebo. The primary end point was the annualized relapse rate. Secondary end points included the number of gadolinium-enhancing lesions on magnetic resonance imaging (MRI) by 96 weeks and worsening of disability. RESULTS: A total of 549 participants were enrolled in the ULTIMATE I trial, and 545 were enrolled in the ULTIMATE II trial; the median follow-up was 95 weeks. In the ULTIMATE I trial, the annualized relapse rate was 0.08 with ublituximab and 0.19 with teriflunomide (rate ratio, 0.41; 95% confidence interval [CI], 0.27 to 0.62; P<0.001); in the ULTIMATE II trial, the annualized relapse rate was 0.09 and 0.18, respectively (rate ratio, 0.51; 95% CI, 0.33 to 0.78; P = 0.002). The mean number of gadolinium-enhancing lesions was 0.02 in the ublituximab group and 0.49 in the teriflunomide group (rate ratio, 0.03; 95% CI, 0.02 to 0.06; P<0.001) in the ULTIMATE I trial and 0.01 and 0.25, respectively (rate ratio, 0.04; 95% CI, 0.02 to 0.06; P<0.001), in the ULTIMATE II trial. In the pooled analysis of the two trials, 5.2% of the participants in the ublituximab group and 5.9% in the teriflunomide group had worsening of disability at 12 weeks (hazard ratio, 0.84; 95% CI, 0.50 to 1.41; P = 0.51). Infusion-related reactions occurred in 47.7% of the participants in the ublituximab group. Serious infections occurred in 5.0% in the ublituximab group and in 2.9% in the teriflunomide group. CONCLUSIONS: Among participants with relapsing multiple sclerosis, ublituximab resulted in lower annualized relapse rates and fewer brain lesions on MRI than teriflunomide over a period of 96 weeks but did not result in a significantly lower risk of worsening of disability. Ublituximab was associated with infusion-related reactions. (Funded by TG Therapeutics; ULTIMATE I and II ClinicalTrials.gov numbers, NCT03277261 and NCT03277248.).
Subject(s)
Antibodies, Monoclonal , Multiple Sclerosis, Relapsing-Remitting , Anti-Inflammatory Agents, Non-Steroidal/therapeutic use , Antibodies, Monoclonal/therapeutic use , Crotonates , Double-Blind Method , Gadolinium/therapeutic use , Humans , Hydroxybutyrates , Immunosuppressive Agents/therapeutic use , Magnetic Resonance Imaging , Multiple Sclerosis/complications , Multiple Sclerosis/diagnostic imaging , Multiple Sclerosis/drug therapy , Multiple Sclerosis/pathology , Multiple Sclerosis, Relapsing-Remitting/complications , Multiple Sclerosis, Relapsing-Remitting/diagnostic imaging , Multiple Sclerosis, Relapsing-Remitting/drug therapy , Multiple Sclerosis, Relapsing-Remitting/pathology , Nitriles , ToluidinesABSTRACT
Insulin has long provided a model for studies of protein folding and stability, enabling enhanced treatment of diabetes mellitus via analogue design. We describe the chemical synthesis of a basal insulin analogue stabilized by substitution of an internal cystine (A6-A11) by a diselenide bridge. The studies focused on insulin glargine (formulated as Lantus® and Toujeo®; Sanofi). Prepared at pHâ 4 in the presence of zinc ions, glargine exhibits a shifted isoelectric point due to a basic B chain extension (ArgB31 -ArgB32 ). Subcutaneous injection leads to pH-dependent precipitation of a long-lived depot. Pairwise substitution of CysA6 and CysA11 by selenocysteine was effected by solid-phase peptide synthesis; the modified A chain also contained substitution of AsnA21 by Gly, circumventing acid-catalyzed deamidation. Although chain combination of native glargine yielded negligible product, in accordance with previous synthetic studies, the pairwise selenocysteine substitution partially rescued this reaction: substantial product was obtained through repeated combination, yielding a stabilized insulin analogue. This strategy thus exploited both (a) the unique redox properties of selenocysteine in protein folding and (b) favorable packing of an internal diselenide bridge in the native state, once achieved. Such rational optimization of protein folding and stability may be generalizable to diverse disulfide-stabilized proteins of therapeutic interest.
Subject(s)
Insulin , Selenocysteine , Insulin Glargine , Cystine , DisulfidesABSTRACT
PURPOSE: Evaluation of distribution kinetics is a neglected aspect of pharmacokinetics. This study examines the utility of the model-independent parameter whole body distribution clearance (CLD) in this respect. METHODS: Since mammillary compartmental models are widely used, CLD was calculated in terms of parameters of this model for 15 drugs. The underlying distribution processes were explored by assessment of relationships to pharmacokinetic parameters and covariates. RESULTS: The model-independence of the definition of the parameter CLD allowed a comparison of distributional properties of different drugs and provided physiological insight. Significant changes in CLD were observed as a result of drug-drug interactions, transporter polymorphisms and a diseased state. CONCLUSION: Total distribution clearance CLD is a useful parameter to evaluate distribution kinetics of drugs. Its estimation as an adjunct to the model-independent parameters clearance and steady-state volume of distribution is advocated.
Subject(s)
Metabolic Clearance Rate , Models, Biological , Pharmacokinetics , Humans , Pharmaceutical Preparations/metabolism , Drug Interactions , Tissue DistributionABSTRACT
Charcot-Marie-Tooth disease (CMT) due to GJB1 variants (CMTX1) is the second most common form of CMT. It is an X-linked disorder characterized by progressive sensory and motor neuropathy with males affected more severely than females. Many reported GJB1 variants remain classified as variants of uncertain significance (VUS). In this large, international, multicentre study we prospectively collected demographic, clinical and genetic data on patients with CMT associated with GJB1 variants. Pathogenicity for each variant was defined using adapted American College of Medical Genetics criteria. Baseline and longitudinal analyses were conducted to study genotype-phenotype correlations, to calculate longitudinal change using the CMT Examination Score (CMTES), to compare males versus females, and pathogenic/likely pathogenic (P/LP) variants versus VUS. We present 387 patients from 295 families harbouring 154 variants in GJB1. Of these, 319 patients (82.4%) were deemed to have P/LP variants, 65 had VUS (16.8%) and three benign variants (0.8%; excluded from analysis); an increased proportion of patients with P/LP variants compared with using ClinVar's classification (74.6%). Male patients (166/319, 52.0%, P/LP only) were more severely affected at baseline. Baseline measures in patients with P/LP variants and VUS showed no significant differences, and regression analysis suggested the disease groups were near identical at baseline. Genotype-phenotype analysis suggested c.-17G>A produces the most severe phenotype of the five most common variants, and missense variants in the intracellular domain are less severe than other domains. Progression of disease was seen with increasing CMTES over time up to 8 years follow-up. Standard response mean (SRM), a measure of outcome responsiveness, peaked at 3 years with moderate responsiveness [change in CMTES (ΔCMTES) = 1.3 ± 2.6, P = 0.00016, SRM = 0.50]. Males and females progressed similarly up to 8 years, but baseline regression analysis suggested that over a longer period, females progress more slowly. Progression was most pronounced for mild phenotypes (CMTES = 0-7; 3-year ΔCMTES = 2.3 ± 2.5, P = 0.001, SRM = 0.90). Enhanced variant interpretation has yielded an increased proportion of GJB1 variants classified as P/LP and will aid future variant interpretation in this gene. Baseline and longitudinal analysis of this large cohort of CMTX1 patients describes the natural history of the disease including the rate of progression; CMTES showed moderate responsiveness for the whole group at 3 years and higher responsiveness for the mild group at 3, 4 and 5 years. These results have implications for patient selection for upcoming clinical trials.
Subject(s)
Charcot-Marie-Tooth Disease , Female , Humans , Male , Charcot-Marie-Tooth Disease/pathology , Connexins/genetics , Mutation/genetics , Mutation, Missense , Phenotype , Gap Junction beta-1 ProteinABSTRACT
INTRODUCTION: High exercise adherence is a key factor for effective exercise programmes. However, little is known about predictors of exercise adherence to a multimodal machine-based training in older retirement home residents. AIMS: To assess exercise adherence and potential predictors of adherence. Furthermore, to evaluate user acceptance of the multimodal training and the change in exercise self-efficacy. METHODS: In this sub-analysis of the bestform-F study, a total of 77 retirement home residents ≥65 years (mean age: 85.6 ± 6.6 years, 77.9% female) participated in a 6-month machine-based resistance, coordination and endurance training. Attendance to the training was documented for each training session. To identify potential predictors a multiple linear regression model was fitted to the data. Analyzed predictors included age, sex, body mass index (BMI), physical function, exercise self-efficacy, and physical activity history. Different domains of user acceptance (e.g. safety aspects, infrastructure) and exercise self-efficacy were assessed by a questionnaire and the exercise self-efficacy scale (ESES), respectively. RESULTS: Mean exercise adherence was 67.2% (median: 74.4%). The regression model (R2 = 0.225, p = 0.033) revealed that the 6-minute walk test (6-MWT) at baseline significantly predicted exercise adherence (ß: 0.074, 95% confidence interval (CI): 0.006-0.142, p = 0.033). Different user domains were rated at least as good by 83.9%-96.9% of participants, reflecting high acceptance. No statistically significant change was found for exercise self-efficacy over 6 months (mean change: 0.47 ± 3.08 points, p = 0.156). CONCLUSION: Retirement home residents attended more than two thirds of offered training sessions and physical function at baseline was the key factor for predicting adherence. User acceptance of the training devices was highly rated. These findings indicate good potential for implementation of the exercise programme.
Subject(s)
Endurance Training , Resistance Training , Humans , Female , Aged , Aged, 80 and over , Male , Retirement , Exercise , Exercise TherapyABSTRACT
Insulin-signaling requires conformational change: whereas the free hormone and its receptor each adopt autoinhibited conformations, their binding leads to structural reorganization. To test the functional coupling between insulin's "hinge opening" and receptor activation, we inserted an artificial ligand-dependent switch into the insulin molecule. Ligand-binding disrupts an internal tether designed to stabilize the hormone's native closed and inactive conformation, thereby enabling productive receptor engagement. This scheme exploited a diol sensor (meta-fluoro-phenylboronic acid at GlyA1) and internal diol (3,4-dihydroxybenzoate at LysB28). The sensor recognizes monosaccharides (fructose > glucose). Studies of insulin-signaling in human hepatoma-derived cells (HepG2) demonstrated fructose-dependent receptor autophosphorylation leading to appropriate downstream signaling events, including a specific kinase cascade and metabolic gene regulation (gluconeogenesis and lipogenesis). Addition of glucose (an isomeric ligand with negligible sensor affinity) did not activate the hormone. Similarly, metabolite-regulated signaling was not observed in control studies of 1) an unmodified insulin analog or 2) an analog containing a diol sensor without internal tethering. Although secondary structure (as probed by circular dichroism) was unaffected by ligand-binding, heteronuclear NMR studies revealed subtle local and nonlocal monosaccharide-dependent changes in structure. Insertion of a synthetic switch into insulin has thus demonstrated coupling between hinge-opening and allosteric holoreceptor signaling. In addition to this foundational finding, our results provide proof of principle for design of a mechanism-based metabolite-responsive insulin. In particular, replacement of the present fructose sensor by an analogous glucose sensor may enable translational development of a "smart" insulin analog to mitigate hypoglycemic risk in diabetes therapy.
Subject(s)
Insulin/chemistry , Blotting, Western , Fructose/chemistry , Fructose/metabolism , Hep G2 Cells , Humans , Insulin/metabolism , Ligands , Models, Molecular , Protein Conformation , Signal TransductionABSTRACT
BACKGROUND: High-dose erythropoietin has been shown to have a neuroprotective effect in preclinical models of neonatal brain injury, and phase 2 trials have suggested possible efficacy; however, the benefits and safety of this therapy in extremely preterm infants have not been established. METHODS: In this multicenter, randomized, double-blind trial of high-dose erythropoietin, we assigned 941 infants who were born at 24 weeks 0 days to 27 weeks 6 days of gestation to receive erythropoietin or placebo within 24 hours after birth. Erythropoietin was administered intravenously at a dose of 1000 U per kilogram of body weight every 48 hours for a total of six doses, followed by a maintenance dose of 400 U per kilogram three times per week by subcutaneous injection through 32 completed weeks of postmenstrual age. Placebo was administered as intravenous saline followed by sham injections. The primary outcome was death or severe neurodevelopmental impairment at 22 to 26 months of postmenstrual age. Severe neurodevelopmental impairment was defined as severe cerebral palsy or a composite motor or composite cognitive score of less than 70 (which corresponds to 2 SD below the mean, with higher scores indicating better performance) on the Bayley Scales of Infant and Toddler Development, third edition. RESULTS: A total of 741 infants were included in the per-protocol efficacy analysis: 376 received erythropoietin and 365 received placebo. There was no significant difference between the erythropoietin group and the placebo group in the incidence of death or severe neurodevelopmental impairment at 2 years of age (97 children [26%] vs. 94 children [26%]; relative risk, 1.03; 95% confidence interval, 0.81 to 1.32; P = 0.80). There were no significant differences between the groups in the rates of retinopathy of prematurity, intracranial hemorrhage, sepsis, necrotizing enterocolitis, bronchopulmonary dysplasia, or death or in the frequency of serious adverse events. CONCLUSIONS: High-dose erythropoietin treatment administered to extremely preterm infants from 24 hours after birth through 32 weeks of postmenstrual age did not result in a lower risk of severe neurodevelopmental impairment or death at 2 years of age. (Funded by the National Institute of Neurological Disorders and Stroke; PENUT ClinicalTrials.gov number, NCT01378273.).
Subject(s)
Erythropoietin/administration & dosage , Infant, Extremely Premature , Infant, Premature, Diseases/prevention & control , Neurodevelopmental Disorders/prevention & control , Brain/diagnostic imaging , Child, Preschool , Double-Blind Method , Erythropoietin/adverse effects , Female , Follow-Up Studies , Humans , Infant , Infant, Newborn , Infant, Premature, Diseases/mortality , Male , Neurodevelopmental Disorders/epidemiology , UltrasonographyABSTRACT
Age-related changes in the patterns of local relatedness (kinship dynamics) can be a significant selective force shaping the evolution of life history and social behaviour. In humans and some species of toothed whales, average female relatedness increases with age, which can select for a prolonged post-reproductive lifespan in older females due to both costs of reproductive conflict and benefits of late-life helping of kin. Killer whales (Orcinus orca) provide a valuable system for exploring social dynamics related to such costs and benefits in a mammal with an extended post-reproductive female lifespan. We use more than 40 years of demographic and association data on the mammal-eating Bigg's killer whale to quantify how mother-offspring social relationships change with offspring age and identify opportunities for late-life helping and the potential for an intergenerational reproductive conflict. Our results suggest a high degree of male philopatry and female-biased budding dispersal in Bigg's killer whales, with some variability in the dispersal rate for both sexes. These patterns of dispersal provide opportunities for late-life helping particularly between mothers and their adult sons, while partly mitigating the costs of mother-daughter reproductive conflict. Our results provide an important step towards understanding why and how menopause has evolved in Bigg's killer whales.
Subject(s)
Whale, Killer , Humans , Animals , Adult , Male , Female , Aged , Mothers , Reproduction , Longevity , Social BehaviorABSTRACT
Intolerance is the most common reason for kinase inhibitor (KI) discontinuation in chronic lymphocytic leukemia (CLL). Umbralisib, a novel highly selective phosphatidylinositol 3-kinase δ (PI3Kδ)/CK1ε inhibitor, is active and well tolerated in CLL patients. In this phase 2 trial (NCT02742090), umbralisib was initiated at 800 mg/d in CLL patients requiring therapy, who were intolerant to prior BTK inhibitor (BTKi) or PI3K inhibitor (PI3Ki) therapy, until progression or toxicity. Primary end point was progression-free survival (PFS). Secondary end points included time to treatment failure and safety. DNA was genotyped for CYP3A4, CYP3A5, and CYP2D6 polymorphisms. Fifty-one patients were enrolled (44 BTKi intolerant and 7 PI3Kδi intolerant); median age was 70 years (range, 48-96), with a median of 2 prior lines of therapy (range, 1-7), 24% had del17p and/or TP53 mutation, and 65% had unmutated IGHV. Most common adverse events (AEs) leading to prior KI discontinuation were rash (27%), arthralgia (18%), and atrial fibrillation (16%). Median PFS was 23.5 months (95% CI, 13.1-not estimable), with 58% of patients on umbralisib for a longer duration than prior KI. Most common (≥5%) grade ≥3 AEs on umbralisib (all causality) were neutropenia (18%), leukocytosis (14%), thrombocytopenia (12%), pneumonia (12%), and diarrhea (8%). Six patients (12%) discontinued umbralisib because of an AE. Eight patients (16%) had dose reductions and were successfully rechallenged. These are the first prospective data to confirm that switching from a BTKi or alternate PI3Ki to umbralisib in this BTKi- and PI3Ki-intolerant CLL population can result in durable well-tolerated responses.
Subject(s)
Antineoplastic Agents/therapeutic use , Class I Phosphatidylinositol 3-Kinases/antagonists & inhibitors , Heterocyclic Compounds, 4 or More Rings/therapeutic use , Leukemia, Lymphocytic, Chronic, B-Cell/drug therapy , Neoplasm Proteins/antagonists & inhibitors , Protein Kinase Inhibitors/therapeutic use , Adenine/adverse effects , Adenine/analogs & derivatives , Adenine/therapeutic use , Agammaglobulinaemia Tyrosine Kinase/antagonists & inhibitors , Aged , Aged, 80 and over , Antineoplastic Agents/adverse effects , Cardiovascular Diseases/chemically induced , Drug Eruptions/etiology , Drug Resistance, Neoplasm , Female , Gastrointestinal Diseases/chemically induced , Heterocyclic Compounds, 4 or More Rings/adverse effects , Humans , Kaplan-Meier Estimate , Leukemia, Lymphocytic, Chronic, B-Cell/enzymology , Leukemia, Lymphocytic, Chronic, B-Cell/mortality , Male , Middle Aged , Piperidines/adverse effects , Piperidines/therapeutic use , Progression-Free Survival , Protein Kinase Inhibitors/adverse effectsABSTRACT
Clearance concepts are extensively applied in drug development and drug therapy. The well-stirred model (WSM) of hepatic elimination is the most widely adopted physiologic model in pharmacokinetics owing to its simplicity. A common feature of this organ model is its use to relate hepatic clearance of a compound to the physiologic variables: organ blood flow rate, binding within blood, and hepatocellular metabolic and excretory activities. Recently, Kirchhoff's laws of electrical network have been applied to organ clearance (Pachter et al., 2022; Benet and Sodhi, 2023) with the claim that they yield the same equation for hepatic clearance as the WSM, and that the equation is independent of a mechanistic model. This commentary analyzes this claim and shows that implicit in the application of Kirchhoff's approaches are the same assumptions as those of the WSM. Concern is also expressed in the interpretation of permeability or transport parameters and related equations, as well as the inappropriateness of the corresponding equation defining hepatic clearance. There is no value, and some dangers, in applying Kirchhoff's electrical laws to organ clearance. SIGNIFICANCE STATEMENT: This commentary refutes this claim by Pachter et al. (2022), and Benet and Sodhi, (2023), who suggest that the well-stirred model (WSM) of hepatic elimination, the most widely applied physiologic model of hepatic clearance, provides the same equation as Kirchhoff's laws of electrical network that is independent of a physiologic model. A careful review shows that the claim is groundless and fraught with errors. We conclude that there is no place for the application of Kirchhoff's laws to organ clearance models.
ABSTRACT
BACKGROUND: Neuroprognostication in neonates with neonatal encephalopathy (NE) may be enhanced by early serial measurement of a panel of four brain-specific biomarkers. METHODS: To evaluate serum biomarkers, 40 NE samples and 37 healthy neonates from a biorepository were analyzed. Blood samples were collected at 0-6, 12, 24, 48, and 96 h of life. MRI provided a short-term measure of injury. Long-term outcomes included death or a Bayley III score at 17-24 months of age. RESULTS: Glial fibrillary acidic protein (GFAP), ubiquitin c-terminal hydrolase-L1 (UCH-L1), and Tau peaked at 0-6 h of life, while neurofilament light chain (NFL) peaked at 96 h of life. These four marker concentrations at 96 h of life differentiated moderate/severe from none/mild brain injury by MRI, while GFAP and Tau showed early discrimination. For long-term outcomes, GFAP, NFL, Tau, and UCH-L1 could differentiate a poor outcome vs good outcome as early as 0-6 h of life, depending on the Bayley domain, and a combination of the four markers enhanced the sensitivity and specificity. Machine learning trajectory analyses identified upward trajectory patients with a high concordance to poor outcomes. CONCLUSION: GFAP, NFL, Tau, and UCH-L1 may be of neuroprognostic significance after NE. IMPACT: Serial measurements of GFAP, NFL, Tau, and UCH-L1 show promise in aiding the bedside clinician in making treatment decisions in neonatal encephalopathy. The panel of four neuroproteins increased the ability to predict neurodevelopmental outcomes. The study utilized a trajectory analysis that enabled predictive modeling. A panel approach provides the bedside clinician with objective data to individualize care. This study provides the foundation to develop a point of care device in the future.
Subject(s)
Brain Injuries , Intermediate Filaments , Infant, Newborn , Humans , Glial Fibrillary Acidic Protein , Ubiquitin Thiolesterase , BiomarkersABSTRACT
BRAF/MEK inhibition remains standard of care for treatment of BRAF-mutated non-small cell lung cancer (NSCLC). Although common adverse events (AEs) have been reported through clinical trials and ongoing clinical practice, only a handful of reports have detailed unusual adverse events associated with these medications. This report presents a patient with BRAF-mutated NSCLC treated with dabrafenib and trametinib who experienced 2 unusual AEs-Sweet syndrome and MEK-associated retinopathy-that responded to steroid treatment. The patient was able to continue BRAF/MEK inhibition through a coordinated multidisciplinary approach. This case highlights the importance for all clinicians to recognize unusual AEs associated with BRAF/MEK inhibition, particularly in the setting of expanded use for all BRAF V600E-mutated solid tumors.
Subject(s)
Carcinoma, Non-Small-Cell Lung , Lung Neoplasms , Humans , Carcinoma, Non-Small-Cell Lung/drug therapy , Proto-Oncogene Proteins B-raf/genetics , Lung Neoplasms/drug therapy , Mitogen-Activated Protein Kinase Kinases/therapeutic use , Oximes/therapeutic use , MutationABSTRACT
PURPOSE: The one-compartment model with first order absorption (ka1C) has been extensively used to fit oral data. But when the disposition parameters of the drug are not available, the bias in the parameter estimates remains unclear. In this paper, the effect of potential misspecification of the area under the curve (AUC) and the mean absorption time (MAT) was evaluated for three relatively slowly absorbed drugs/formulations. METHODS: Assuming a three-compartment disposition model with an input (absorption) rate described as a sum of two inverse Gaussian functions (2IG3C) as the true model, the deviations of AUC and MAT estimated with simpler models were analyzed. Simpler models, as the ka1C model (Bateman function), the one-compartment model with IG input function (IG1C) and the gamma density function were fitted to the oral data alone, and compared to the fits obtained with the 2IG3C model which also uses the 3C disposition parameters of the drug. Data from pharmacokinetic studies of trospium, propiverine and ketamine in healthy volunteers were analyzed using a population approach. RESULTS: The Bateman function (ka1C) allowed a robust estimation of the population mean AUC, but the individual estimates were highly biased. It failed in evaluating MAT. The simple alternative models did not improve the situation. CONCLUSIONS: The Bateman function appears to be useful for estimating the population mean value of AUC after oral administration. The results reemphasize the fact that insight into the absorption process can be only gained when also intravenous reference data are available.
ABSTRACT
INTRODUCTION: Currently, the rate of bile duct injury and leak following laparoscopic cholecystectomy (LC) is still higher than for open surgery. Diverse investigative algorithms were suggested for bile leak, shifting from hepatobiliary scintigraphy (HBS) toward invasive and more sophisticated means. We aimed to analyze the use of biliary scan as the initial modality to investigate significant bile leak in the drain following LC, attempting to avoid potential unnecessary invasive means when the scan demonstrate fair passage of nuclear substance to the intestine, without leak. METHODS: We have conducted a prospective non-randomized study, mandating hepatobiliary scintigraphy first, for asymptomatic patients harboring drain in the gallbladder fossa, leaking more than 50 mL/day following LC. Analysis was done based on medical data from the surgical, gastroenterology, and the nuclear medicine departments. RESULTS: Among 3,124 patients undergoing LC, significant bile leak in the drain was seen in 67 subjects, of whom we started with HBS in 50 patients, presenting our study group. In 27 of whom, biliary scan was the only investigative modality, showing fair passage of the nuclear isotope to the duodenum and absence of leak in the majority. The leak stopped spontaneously within a mean of 3.6 days, and convalescence as well as outpatient clinic follow-up was uneventful. In 23 patients, biliary scan that was interpreted as abnormal was followed by endoscopic retrograde cholangio-pancreatography (ERCP). However, ERCP did not demonstrate any bile leak in 13 subjects. In 17 patients, ERCP was used initially, without biliary scan, suggesting the possibility of avoiding invasive modalities in 7 patients. CONCLUSIONS: Based on a negative predictive value of 91%, we suggest that in cases of asymptomatic significant bile leak through a drain following LC, a normal HBS as the initial modality can safely decrease the rate of using invasive modalities.
ABSTRACT
Proteins have evolved to be foldable, and yet determinants of foldability may be inapparent once the native state is reached. Insight has emerged from studies of diseases of protein misfolding, exemplified by monogenic diabetes mellitus due to mutations in proinsulin leading to endoplasmic reticulum stress and ß-cell death. Cellular foldability of human proinsulin requires an invariant Phe within a conserved crevice at the receptor-binding surface (position B24). Any substitution, even related aromatic residue TyrB24, impairs insulin biosynthesis and secretion. As a seeming paradox, a monomeric TyrB24 insulin analog exhibits a native-like structure in solution with only a modest decrement in stability. Packing of TyrB24 is similar to that of PheB24, adjoining core cystine B19-A20 to seal the core; the analog also exhibits native self-assembly. Although affinity for the insulin receptor is decreased â¼20-fold, biological activities in cells and rats were within the range of natural variation. Together, our findings suggest that the invariance of PheB24 among vertebrate insulins and insulin-like growth factors reflects an essential role in enabling efficient protein folding, trafficking, and secretion, a function that is inapparent in native structures. In particular, we envision that the para-hydroxyl group of TyrB24 hinders pairing of cystine B19-A20 in an obligatory on-pathway folding intermediate. The absence of genetic variation at B24 and other conserved sites near this disulfide bridge-excluded due to ß-cell dysfunction-suggests that insulin has evolved to the edge of foldability. Nonrobustness of a protein's fitness landscape underlies both a rare monogenic syndrome and "diabesity" as a pandemic disease of civilization.
Subject(s)
Insulin/metabolism , Amino Acid Substitution/physiology , Animals , Cell Line , Cell Line, Tumor , Diabetes Mellitus/metabolism , Disulfides/metabolism , Gene Regulatory Networks/physiology , HEK293 Cells , Humans , Insulin-Secreting Cells/metabolism , MCF-7 Cells , Proinsulin/metabolism , Protein Binding/physiology , Protein Folding , Rats , Receptor, Insulin/metabolism , Structure-Activity RelationshipABSTRACT
PURPOSE: To describe the process of developing, and evaluating the feasibility and acceptability of, an EMR-based transition readiness assessment. DESIGN AND METHODS: A Cerner-based version of the UNC TRxANSITION Index was implemented across four pediatric subspecialty clinics: epilepsy, inflammatory bowel disease; type 1 diabetes, oncology survivorship. The feasibility was assessed by each's clinic's ability to meet form completion goals and their assessment rate. Acceptability was assessed via family refusal rate, a staff-completed feedback questionnaire, and whether the form was adopted into routine clinical care after completion of the pilot study. RESULTS: All clinics met form completion goals (N = 10/clinic). The assessment rate ranged from 66 to 100% across clinics. No families refused completion of the form. Most staff (70%) reported completing the form in <10 min. Staff reported on challenges experienced and provided recommendations to streamline administration and enhance clinical care. All staff reported the form helped them identify knowledge gaps in their patients. Two clinics continued using the form following completion of the pilot study. CONCLUSIONS: Implementation was most feasible in clinics that were well-staffed and had lengthier patient visits, however, time and staff resources were the biggest challenges to implementation across clinics. Based on staff feedback to improve efficiency and developmentally-tailor assessment, the form will be divided into Beginner Skills and Advanced Skills. PRACTICAL IMPLICATIONS: Integrating transition readiness assessment into the EMR has the potential to improve clinical care by facilitating staff's ability to efficiently identify knowledge gaps in their transition-aged patients and intervene.
Subject(s)
Electronic Health Records , Inflammatory Bowel Diseases , Child , Humans , Aged , Pilot ProjectsABSTRACT
Insulin is a mainstay of therapy for diabetes mellitus, yet its thermal stability complicates global transportation and storage. Cold-chain transport, coupled with optimized formulation and materials, prevents to some degree nucleation of amyloid and hence inactivation of hormonal activity. These issues hence motivate the design of analogs with increased stability, with a promising approach being single-chain insulins (SCIs), whose C domains (foreshortened relative to proinsulin) resemble those of the single-chain growth factors (IGFs). We have previously demonstrated that optimized SCIs can exhibit native-like hormonal activity with enhanced thermal stability and marked resistance to fibrillation. Here, we describe the crystal structure of an ultrastable SCI (C-domain length 6; sequence EEGPRR) bound to modules of the insulin receptor (IR) ectodomain (N-terminal α-subunit domains L1-CR and C-terminal αCT peptide; "microreceptor" [µIR]). The structure of the SCI-µIR complex, stabilized by an Fv module, was determined using diffraction data to a resolution of 2.6 Å. Remarkably, the αCT peptide (IR-A isoform) "threads" through a gap between the flexible C domain and the insulin core. To explore such threading, we undertook molecular dynamics simulations to 1) compare threaded with unthreaded binding modes and 2) evaluate effects of C-domain length on these alternate modes. The simulations (employing both conventional and enhanced sampling simulations) provide evidence that very short linkers (C-domain length of -1) would limit gap opening in the SCI and so impair threading. We envisage that analogous threading occurs in the intact SCI-IR complex-rationalizing why minimal C-domain lengths block complete activity-and might be exploited to design novel receptor-isoform-specific analogs.
Subject(s)
Insulin , Receptor, Insulin , Receptor, Insulin/metabolism , Insulin/metabolism , Models, Molecular , Protein Binding , Peptides/chemistryABSTRACT
SRY is the Y-chromosomal gene that determines male sex development in humans and most other mammals. After three decades of study, we still lack a detailed understanding of which domains of the SRY protein are required to engage the pathway of gene activity leading to testis development. Some insight has been gained from the study of genetic variations underlying differences/disorders of sex determination (DSD), but the lack of a system of experimentally generating SRY mutations and studying their consequences in vivo has limited progress in the field. To address this issue, we generated a mouse model carrying a human SRY transgene able to drive testis determination in XX mice. Using CRISPR-Cas9 gene editing, we generated novel genetic modifications in each of SRY's three domains (N-terminal, HMG box, and C-terminal) and performed a detailed analysis of their molecular and cellular effects on embryonic testis development. Our results provide new functional insights unique to human SRY and present a versatile and powerful system in which to functionally analyze variations of SRY including known and novel pathogenic variants found in DSD.
Subject(s)
Gene Editing , Mice, Transgenic , Sex-Determining Region Y Protein , Testis , Animals , Humans , Male , Mice , Protein Domains , Sex-Determining Region Y Protein/genetics , Testis/metabolismABSTRACT
PURPOSE OF REVIEW: Diabetes mellitus (DM) due to toxic misfolding of proinsulin variants provides a monogenic model of endoplasmic reticulum (ER) stress. The mutant proinsulin syndrome (also designated MIDY; Mutant INS-gene-induced Diabetes of Youth or Maturity-onset diabetes of the young 10 (MODY10)) ordinarily presents as permanent neonatal-onset DM, but specific amino-acid substitutions may also present later in childhood or adolescence. This review highlights structural mechanisms of proinsulin folding as inferred from phenotype-genotype relationships. RECENT FINDINGS: MIDY mutations most commonly add or remove a cysteine, leading to a variant polypeptide containing an odd number of thiol groups. Such variants are associated with aberrant intermolecular disulfide pairing, ER stress, and neonatal ß-cell dysfunction. Non-cysteine-related (NCR) mutations (occurring in both the B and A domains of proinsulin) define distinct determinants of foldability and vary in severity. The range of ages of onset, therefore, reflects a "molecular rheostat" connecting protein biophysics to quality-control ER checkpoints. Because in most mammalian cell lines even wild-type proinsulin exhibits limited folding efficiency, molecular barriers to folding uncovered by NCR MIDY mutations may pertain to ß-cell dysfunction in non-syndromic type 2 DM due to INS-gene overexpression in the face of peripheral insulin resistance. Recent studies of MIDY mutations and related NCR variants, combining molecular and cell-based approaches, suggest that proinsulin has evolved at the edge of non-foldability. Chemical protein synthesis promises to enable comparative studies of "non-foldable" proinsulin variants to define key steps in wild-type biosynthesis. Such studies may create opportunities for novel therapeutic approaches to non-syndromic type 2 DM.