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Computational protein design of an ensemble of conformations for one protein-i.e., multi-state design-determines the side chain identity by optimizing the energetic contributions of that side chain in each of the backbone conformations. Sampling the resulting large sequence-structure search space limits the number of conformations and the size of proteins in multi-state design algorithms. Here, we demonstrated that the REstrained CONvergence (RECON) algorithm can simultaneously evaluate the sequence of large proteins that undergo substantial conformational changes. Simultaneous optimization of side chain conformations across all conformations increased sequence conservation when compared to single-state designs in all cases. More importantly, the sequence space sampled by RECON MSD resembled the evolutionary sequence space of flexible proteins, particularly when confined to predicting the mutational preferences of limited common ancestral descent, such as in the case of influenza type A hemagglutinin. Additionally, we found that sequence positions which require substantial changes in their local environment across an ensemble of conformations are more likely to be conserved. These increased conservation rates are better captured by RECON MSD over multiple conformations and thus multiple local residue environments during design. To quantify this rewiring of contacts at a certain position in sequence and structure, we introduced a new metric designated 'contact proximity deviation' that enumerates contact map changes. This measure allows mapping of global conformational changes into local side chain proximity adjustments, a property not captured by traditional global similarity metrics such as RMSD or local similarity metrics such as changes in φ and ψ angles.

Proton minibeam therapy (PMBT) is a form of spatially fractionated radiotherapy wherein broad beam radiation is replaced with segmented minibeams-either parallel, planar minibeam arrays generated by a multislit collimator or scanned pencil beams that converge laterally at depth to create a uniform dose layer at the tumor. By doing so, the spatial pattern of entrance dose is considerably modified while still maintaining tumor dose and efficacy. Recent studies using computational modeling, phantom experiments, and preclinical models, and early clinical feasibility assessments suggest that unique physical and biological attributes of PMBT can be exploited for future clinical benefit. We outline some of the guiding principle of PMBT in this concise overview of this emerging area of preclinical and clinical research inquiry.

The single photon response (SPR) in vertebrate photoreceptors is inherently variable due to several stochastic events in the phototransduction cascade, the main one being the shutoff of photoactivated rhodopsin. Deactivation is driven by a random number of steps, each of random duration with final quenching occurring after a random delay. Nevertheless, variability of the SPR is relatively low, making the signal highly reliable. Several biophysical and mathematical mechanisms contributing to variability suppression have been examined by the authors. Here we investigate the contribution of local depletion of cGMP by PDE*, the non linear dependence of the photocurrent on cGMP, Ca2+ feedback by making use of a fully space resolved (FSR) mathematical model, applied to two species (mouse and salamander), by varying the cGMP diffusion rate severalfold and rod outer segment diameter by an order of magnitude, and by introducing new, more refined, and time dependent variability functionals. Globally well stirred (GWS) models, and to a lesser extent transversally well stirred models (TWS), underestimate the role of nonlinearities and local cGMP depletion in quenching the variability of the circulating current with respect to fully space resolved models (FSR). These distortions minimize the true extent to which SPR is stabilized by locality in cGMP depletion, nonlinear effects linking cGMP to current, and Ca2+ feedback arising from the physical separation of E* from the ion channels located on the outer shell, and the diffusion of these second messengers in the cytoplasm.

The presynaptic dopamine transporter mediates rapid reuptake of synaptic dopamine. Although cell surface DAT trafficking recently emerged as an important component of DAT regulation, it has not been systematically investigated. Here, we apply our single quantum dot (Qdot) tracking approach to monitor DAT plasma membrane dynamics in several heterologous expression cell hosts with nanometer localization accuracy. We demonstrate that Qdot-tagged DAT proteins exhibited highly heterogeneous membrane diffusivity dependent on the local membrane topography. We also show that Qdot-tagged DATs were localized away from the flat membrane regions and were dynamically retained in the membrane protrusions and cell edges for the duration of imaging. Single quantum dot tracking of wildtype DAT and its conformation-defective coding variants (R60A and W63A) revealed a significantly accelerated rate of dysfunctional DAT membrane diffusion. We believe our results warrant an in-depth investigation as to whether compromised membrane dynamics is a common feature of brain disorder-derived DAT mutants.

Adopting a systems approach, we devise a general workflow to define actionable subtypes in human cancers. Applied to small cell lung cancer (SCLC), the workflow identifies four subtypes based on global gene expression patterns and ontologies. Three correspond to known subtypes (SCLC-A, SCLC-N, and SCLC-Y), while the fourth is a previously undescribed ASCL1+ neuroendocrine variant (NEv2, or SCLC-A2). Tumor deconvolution with subtype gene signatures shows that all of the subtypes are detectable in varying proportions in human and mouse tumors. To understand how multiple stable subtypes can arise within a tumor, we infer a network of transcription factors and develop BooleaBayes, a minimally-constrained Boolean rule-fitting approach. In silico perturbations of the network identify master regulators and destabilizers of its attractors. Specific to NEv2, BooleaBayes predicts ELF3 and NR0B1 as master regulators of the subtype, and TCF3 as a master destabilizer. Since the four subtypes exhibit differential drug sensitivity, with NEv2 consistently least sensitive, these findings may lead to actionable therapeutic strategies that consider SCLC intratumoral heterogeneity. Our systems-level approach should generalize to other cancer types.

BACKGROUND - Management algorithms for adult severe traumatic brain injury (sTBI) were omitted in later editions of the Brain Trauma Foundation's sTBI Management Guidelines, as they were not evidence-based.
METHODS - We used a Delphi-method-based consensus approach to address management of sTBI patients undergoing intracranial pressure (ICP) monitoring. Forty-two experienced, clinically active sTBI specialists from six continents comprised the panel. Eight surveys iterated queries and comments. An in-person meeting included whole- and small-group discussions and blinded voting. Consensus required 80% agreement. We developed heatmaps based on a traffic-light model where panelists' decision tendencies were the focus of recommendations.
RESULTS - We provide comprehensive algorithms for ICP-monitor-based adult sTBI management. Consensus established 18 interventions as fundamental and ten treatments not to be used. We provide a three-tier algorithm for treating elevated ICP. Treatments within a tier are considered empirically equivalent. Higher tiers involve higher risk therapies. Tiers 1, 2, and 3 include 10, 4, and 3 interventions, respectively. We include inter-tier considerations, and recommendations for critical neuroworsening to assist the recognition and treatment of declining patients. Novel elements include guidance for autoregulation-based ICP treatment based on MAP Challenge results, and two heatmaps to guide (1) ICP-monitor removal and (2) consideration of sedation holidays for neurological examination.
CONCLUSIONS - Our modern and comprehensive sTBI-management protocol is designed to assist clinicians managing sTBI patients monitored with ICP-monitors alone. Consensus-based (class III evidence), it provides management recommendations based on combined expert opinion. It reflects neither a standard-of-care nor a substitute for thoughtful individualized management.

BACKGROUND - Checkpoint inhibitor therapy is a standard of care for patients with metastatic renal cell carcinoma. Treatment options after checkpoint inhibitor therapy include vascular endothelial growth factor receptor (VEGF-R) tyrosine kinase inhibitors, although no prospective data regarding their use in this setting exist. Axitinib is a VEGF-R inhibitor with clinical data supporting increased activity with dose titration. We aimed to investigate the activity of dose titrated axitinib in patients with metastatic renal cell carcinoma who were previously treated with checkpoint inhibitor.
METHODS - We did a multicentre, phase 2 trial of axitinib given on an individualised dosing algorithm. Patients at least 18 years of age with histologically or cytologically confirmed locally recurrent or metastatic renal cell carcinoma with clear cell histology, a Karnofsky Performance Status of 70% or more, and measurable disease who received checkpoint inhibitor therapy as the most recent treatment were eligible. There was no limit on number of previous therapies received. Patients received oral axitinib at a starting dose of 5 mg twice daily with dose titration every 14 days in 1 mg increments (ie, 5 mg twice daily to 6 mg twice daily, up to 10 mg twice daily maximum dose) if there was no axitinib-related grade 2 or higher mucositis, diarrhoea, hand-foot syndrome, or fatigue. If one or more of these grade 2 adverse events occurred, axitinib was withheld for 3 days before the same dose was resumed. Dose reductions were made if recurrent grade 2 adverse events despite treatment breaks or grade 3-4 adverse events occurred. The primary outcome was progression-free survival. Analyses were done per protocol in all patients who received at least one dose of axitinib. Recruitment has been completed and the trial is ongoing. This trial is registered with ClincalTrials.gov, number NCT02579811.
FINDINGS - Between Jan 5, 2016 and Feb 21, 2018, 40 patients were enrolled and received at least one dose of study treatment. With a median follow-up of 8·7 months (IQR 3·7-14·2), the median progression-free survival was 8·8 months (95% CI 5·7-16·6). Fatigue (83%) and hypertension (75%) were the most common all-grade adverse events. The most common grade 3 adverse event was hypertension (24 patients [60%]). There was one (3%) grade 4 adverse event (elevated lipase) and no treatment-related deaths occurred. Serious adverse events that were likely related to therapy occurred in eight (20%) patients; the most common were dehydration (n=4) and diarrhoea (n=2).
INTERPRETATION - Individualised axitinib dosing in patients with metastatic renal cell inoma previously treated with checkpoint inhibitors did not meet the prespecified threshold for progression free survival, but these data show that this individualised titration scheme is feasible and has robust clinical activity. These prospective results warrant consideration of axitinib in this setting.
FUNDING - Pfizer.
Copyright © 2019 Elsevier Ltd. All rights reserved.

A novel, common, and potent cardiovascular risk factor has recently emerged: clonal hematopoiesis of indeterminate potential (CHIP). CHIP arises from somatic mutations in hematopoietic stem cells that yield clonal progeny of mutant leukocytes in blood. Individuals with CHIP have a doubled risk of coronary heart disease and ischemic stroke, and worsened heart failure outcomes independent of traditional cardiovascular risk factors. The recognition of CHIP as a nontraditional risk factor challenges specialists in hematology/oncology and cardiovascular medicine alike. Should we screen for CHIP? If so, in whom? How should we assess cardiovascular risk in people with CHIP? How should we manage the excess cardiovascular risk in the absence of an evidence base? This review explains CHIP, explores the clinical quandaries, strives to provide reasonable recommendations for the multidisciplinary management of cardiovascular risk in individuals with CHIP, and highlights current knowledge gaps.
Copyright © 2019 American College of Cardiology Foundation. Published by Elsevier Inc. All rights reserved.

BACKGROUND AND PURPOSE - The purpose of the study is to characterize diffusion tensor imaging indices in the developing spinal cord, evaluating differences based on age and cord region. Describing the progression of DTI indices in the pediatric cord increases our understanding of spinal cord development.
MATERIALS AND METHODS - A retrospective analysis was performed on DTI acquired in 121 pediatric patients (mean, 8.6 years; range, 0.3-18.0 years) at Monroe Carell Jr. Children's Hospital at Vanderbilt from 2017 to 2018. Diffusion-weighted images (15 directions; = 750 s/mm; slice thickness, 5 mm; in-plane resolution, 1.0 × 1.0 mm) were acquired on a 3T scanner in the cervicothoracic and/or thoracolumbar cord. Manual whole-cord segmentation was performed. Images were masked and further segmented into cervical, upper thoracic, thoracolumbar, and conus regions. Analyses of covariance were performed for each DTI-derived index to investigate how age affects diffusion across cord regions, and 95% confidence intervals were calculated across age for each derived index and region. Post hoc testing was performed to analyze regional differences.
RESULTS - Analyses of covariance revealed significant correlations of age with axial diffusivity, mean diffusivity, and fractional anisotropy (all, < .001). There were also significant differences among cord regions for axial diffusivity, radial diffusivity, mean diffusivity, and fractional anisotropy (all, < .001).
CONCLUSIONS - This research demonstrates that diffusion evolves in the pediatric spinal cord during development, dependent on both cord region and the diffusion index of interest. Future research could investigate how diffusion may be affected by common pediatric spinal pathologies.
© 2019 by American Journal of Neuroradiology.