For an antibody to be a successful therapeutic many competing factors require optimization, including binding affinity, biophysical characteristics, and immunogenicity risk. Additional constraints ...may arise from the need to formulate antibodies at high concentrations (>150 mg/ml) to enable subcutaneous dosing with reasonable volume (ideally <1.0 mL). Unfortunately, antibodies at high concentrations may exhibit high viscosities that place impractical constraints (such as multiple injections or large needle diameters) on delivery and impede efficient manufacturing. Here we describe the optimization of an anti-PDGF-BB antibody to reduce viscosity, enabling an increase in the formulated concentration from 80 mg/ml to greater than 160 mg/ml, while maintaining the binding affinity. We performed two rounds of structure guided rational design to optimize the surface electrostatic properties. Analysis of this set demonstrated that a net-positive charge change, and disruption of negative charge patches were associated with decreased viscosity, but the effect was greatly dependent on the local surface environment. Our work here provides a comprehensive study exploring a wide sampling of charge-changes in the Fv and CDR regions along with targeting multiple negative charge patches. In total, we generated viscosity measurements for 40 unique antibody variants with full sequence information which provides a significantly larger and more complete dataset than has previously been reported.
We investigate class differences in youth activity participation with interview, survey, and archival data from a diverse sample of parents (n = 51) in two schools. Findings point toward structural ...rather than cultural explanations. Working-and middle-class parents overlap in parenting logics about participation, though differ in one respect: middle-class parents are concerned with customizing children's involvement in activities, while working-class parents are concerned with ensuring safety and social mobility for children through participation. Like financial constraints among the working class, social institutions are implicated in class differences. Schools act as an equalizing institution by offering low-cost activities; yet, contribute to class differences by offering different activities to working-and middle-class youth. Opportunities for working-class youth to participate in non-school activities are few due to the scarcity of community organizations in their neighborhood, while middle-class families are heavily involved in out-of-school activities. We explore the implications of our findings for educational stratification.
Indoleamine-2,3-dioxygenase 1 (IDO1) inhibition and its combination with immune checkpoint inhibitors like pembrolizumab have drawn considerable attention from both academia and the pharmaceutical ...industry. Here, we describe the discovery of a novel class of highly potent IDO1 heme-displacing inhibitors featuring a unique bicyclo1.1.1pentane motif. Compound 1, evolving from an ALIS (automated ligand identification system) hit, exhibited excellent potency but lacked the desired pharmacokinetic profile due to extensive amide hydrolysis of the benzamide moiety. Replacing the central phenyl ring in 1 with a bicyclo1.1.1pentane bioisostere effectively circumvented the amide hydrolysis issue, resulting in the discovery of compound 2 with a favorable overall profile such as excellent potency, selectivity, pharmacokinetics, and a low predicted human dose.
Alzheimer's disease (AD) is the major cause of adult-onset dementia and is characterized in its pre-diagnostic stage by reduced cerebral cortical glucose metabolism and in later stages by reduced ...cortical oxygen uptake, implying reduced mitochondrial respiration. Using quantitative PCR we determined the mitochondrial DNA (mtDNA) gene copy numbers from multiple groups of 15 or 20 pyramidal neurons, GFAP(+) astrocytes and dentate granule neurons isolated using laser capture microdissection, and the relative expression of mitochondrial biogenesis (mitobiogenesis) genes in hippocampi from 10 AD and 9 control (CTL) cases. AD pyramidal but not dentate granule neurons had significantly reduced mtDNA copy numbers compared to CTL neurons. Pyramidal neuron mtDNA copy numbers in CTL, but not AD, positively correlated with cDNA levels of multiple mitobiogenesis genes. In CTL, but not in AD, hippocampal cDNA levels of PGC1α were positively correlated with multiple downstream mitobiogenesis factors. Mitochondrial DNA copy numbers in pyramidal neurons did not correlate with hippocampal Aβ1-42 levels. After 48 h exposure of H9 human neural stem cells to the neurotoxic fragment Aβ25-35, mtDNA copy numbers were not significantly altered. In summary, AD postmortem hippocampal pyramidal neurons have reduced mtDNA copy numbers. Mitochondrial biogenesis pathway signaling relationships are disrupted in AD, but are mostly preserved in CTL. Our findings implicate complex alterations of mitochondria-host cell relationships in AD.
Many morphological, physiological and ecological traits of trees scale with diameter, shaping the structure and function of forest ecosystems. Understanding the mechanistic basis for such scaling ...relationships is key to understanding forests globally and their role in Earth's changing climate system. Here, we evaluate theoretical predictions for the scaling of nine variables in a mixed‐age temperate deciduous forest (CTFS‐ForestGEO forest dynamics plot at the Smithsonian Conservation Biology Institute, Virginia, USA) and compare observed scaling parameters to those from other forests world‐wide. We examine fifteen species and various environmental conditions. Structural, physiological and ecological traits of trees scaled with stem diameter in a manner that was sometimes consistent with existing theoretical predictions – more commonly with those predicting a range of scaling values than a single universal scaling value. Scaling relationships were variable among species, reflecting substantive ecological differences. Scaling relationships varied considerably with environmental conditions. For instance, the scaling of sap flux density varied with atmospheric moisture demand, and herbivore browsing dramatically influenced stem abundance scaling. Thus, stand‐level, time‐averaged scaling relationships (e.g., the scaling of diameter growth) are underlain by a diversity of species‐level scaling relationships that can vary substantially with fluctuating environmental conditions. In order to use scaling theory to accurately characterize forest ecosystems and predict their responses to global change, it will be critical to develop a more nuanced understanding of both the forces that constrain stand‐level scaling and the complexity of scaling variation across species and environmental conditions.
Checkpoint inhibitors have demonstrated unprecedented efficacy and are evolving to become standard of care for certain types of cancers. However, low overall response rates often hamper the broad ...utility and potential of these breakthrough therapies. Combination therapy strategies are currently under intensive investigation in the clinic, including the combination of PD-1/PD-L1 agents with IDO1 inhibitors. Here, we report the discovery of a class of IDO1 heme-binding inhibitors featuring a unique amino-cyclobutarene motif, which was discovered through SBDD from a known and weakly active inhibitor. Subsequent optimization efforts focused on improving metabolic stability and were greatly accelerated by utilizing a robust SNAr reaction of a facile nitro-furazan intermediate to quickly explore different polar side chains. As a culmination of these efforts, compound 16 was identified and demonstrated a favorable overall profile with superior potency and selectivity. Extensive studies confirmed the chemical stability and drug-like properties of compound 16, rendering it a potential drug candidate.
Origins of onset and progression of motor neurodegeneration in amyotrophic lateral sclerosis (ALS) are not clearly known, but may include impairment of mitochondrial bioenergetics. We used ...quantitative PCR approaches to analyze the mitochondrial oxidative phosphorylation (OXPHOS) transcriptomes of spinal cord tissue and peripheral blood mononuclear cells (PBMC) from persons with sporadic ALS compared with those without neurological disease. Expression measurements of 88 different nuclear (n) and mitochondrial (mt) DNA-encoded OXPHOS genes showed mtDNA-encoded respiratory gene expression was significantly decreased in ALS spinal cord by 78–84 % (ANOVA
p
< 0.002). We observed the same phenomenon in freshly isolated PBMC from ALS patients (reduced 24–35 %, ANOVA
p
< 0.001) and reproduced it in a human neural stem cell model treated with 2′,3′-dideoxycytidine (ddC) (reduced 52–78 %, ANOVA
p
< 0.001). nDNA-encoded OXPHOS genes showed heterogeneously and mostly decreased expression in ALS spinal cord tissue. In contrast, ALS PBMC and ddC-treated stem cells showed no significant change in expression of nDNA OXPHOS genes compared with controls. Genes related to mitochondrial biogenesis (PGC-1α, TFAM, ERRα, NRF1, NRF2 and POLG) were queried with inconclusive results. Here, we demonstrate there is a systemic decrease in mtDNA gene expression in ALS central and peripheral tissues that support pursuit of bioenergetic-enhancing therapies. We also identified a combined nDNA and mtDNA gene set (
n
= 26), downregulated in spinal cord tissue that may be useful as a biomarker in the development of cell-based ALS models.
Sleep is essential to life. Accurate measurement and classification of sleep/wake and sleep stages is important in clinical studies for sleep disorder diagnoses and in the interpretation of data from ...consumer devices for monitoring physical and mental well-being. Existing non-polysomnography sleep classification techniques mainly rely on heuristic methods developed in relatively small cohorts. Thus, we aimed to establish the accuracy of wrist-worn accelerometers for sleep stage classification and subsequently describe the association between sleep duration and efficiency (proportion of total time asleep when in bed) with mortality outcomes. We developed a self-supervised deep neural network for sleep stage classification using concurrent laboratory-based polysomnography and accelerometry. After exclusion, 1448 participant nights of data were used for training. The difference between polysomnography and the model classifications on the external validation was 34.7 min (95% limits of agreement (LoA): -37.8-107.2 min) for total sleep duration, 2.6 min for REM duration (95% LoA: -68.4-73.4 min) and 32.1 min (95% LoA: -54.4-118.5 min) for NREM duration. The sleep classifier was deployed in the UK Biobank with 100,000 participants to study the association of sleep duration and sleep efficiency with all-cause mortality. Among 66,214 UK Biobank participants, 1642 mortality events were observed. Short sleepers (<6 h) had a higher risk of mortality compared to participants with normal sleep duration of 6-7.9 h, regardless of whether they had low sleep efficiency (Hazard ratios (HRs): 1.58; 95% confidence intervals (CIs): 1.19-2.11) or high sleep efficiency (HRs: 1.45; 95% CIs: 1.16-1.81). Deep-learning-based sleep classification using accelerometers has a fair to moderate agreement with polysomnography. Our findings suggest that having short overnight sleep confers mortality risk irrespective of sleep continuity.
PDF
