Multi-centre evaluation of variation in cumulative dose assessment in reirradiation scenarios.

BACKGROUND AND PURPOSE: Safe reirradiation relies on assessment of cumulative doses to organs at risk (OARs) across multiple treatments. Different clinical pathways can result in inconsistent estimates. Here, we quantified the consistency of cumulative dose to OARs across multi-centre clinical pathways. MATERIAL AND METHODS: We provided DICOM planning CT, structures and doses for two reirradiation cases: head & neck (HN) and lung. Participants followed their standard pathway to assess the cumulative physical and EQD2 doses (with provided α/ß values), and submitted DVH metrics and a description of their pathways. Participants could also submit physical dose distributions from Course 1 mapped onto the CT of Course 2 using their best available tools. To assess isolated impact of image registrations, a single observer accumulated each submitted spatially mapped physical dose for every participating centre. RESULTS: Cumulative dose assessment was performed by 24 participants. Pathways included rigid (n = 15), or deformable (n = 5) image registration-based 3D dose summation, visual inspection of isodose line contours (n = 1), or summation of dose metrics extracted from each course (n = 3). Largest variations were observed in near-maximum cumulative doses (25.4 - 41.8 Gy for HN, 2.4 - 33.8 Gy for lung OARs), with lower variations in volume/dose metrics to large organs. A standardised process involving spatial mapping of the first course dose to the second course CT followed by summation improved consistency for most near-maximum dose metrics in both cases. CONCLUSION: Large variations highlight the uncertainty in reporting cumulative doses in reirradiation scenarios, with implications for outcome analysis and understanding of published doses. Using a standardised workflow potentially including spatially mapped doses improves consistency in determination of accumulated dose in reirradiation scenarios.

STING-Dependent Signaling in Microglia or Peripheral Immune Cells Orchestrates the Early Inflammatory Response and Influences Brain Injury Outcome.

While originally identified as an antiviral pathway, recent work has implicated that cyclic GMP-AMP-synthase-Stimulator of Interferon Genes (cGAS-STING) signaling is playing a critical role in the neuroinflammatory response to traumatic brain injury (TBI). STING activation results in a robust inflammatory response characterized by the production of inflammatory cytokines called interferons, as well as hundreds of interferon stimulated genes (ISGs). Global knock-out (KO) mice inhibiting this pathway display neuroprotection with evidence that this pathway is active days after injury; yet, the early neuroinflammatory events stimulated by STING signaling remain understudied. Furthermore, the source of STING signaling during brain injury is unknown. Using a murine controlled cortical impact (CCI) model of TBI, we investigated the peripheral immune and microglial response to injury utilizing male chimeric and conditional STING KO animals, respectively. We demonstrate that peripheral and microglial STING signaling contribute to negative outcomes in cortical lesion volume, cell death, and functional outcomes postinjury. A reduction in overall peripheral immune cell and neutrophil infiltration at the injury site is STING dependent in these models at 24 h. Transcriptomic analysis at 2 h, when STING is active, reveals that microglia drive an early, distinct transcriptional program to elicit proinflammatory genes including interleukin 1-ß (IL-1ß), which is lost in conditional knock-out mice. The upregulation of alternative innate immune pathways also occurs after injury in these animals, which supports a complex relationship between brain-resident and peripheral immune cells to coordinate the proinflammatory response and immune cell influx to damaged tissue after injury.

Atypical Neurogenesis, Astrogliosis, and Excessive Hilar Interneuron Loss Are Associated with the Development of Post-Traumatic Epilepsy.

BACKGROUND: Traumatic brain injury (TBI) remains a significant risk factor for post-traumatic epilepsy (PTE). The pathophysiological mechanisms underlying the injury-induced epileptogenesis are under investigation. The dentate gyrus-a structure that is highly susceptible to injury-has been implicated in the evolution of seizure development. METHODS: Utilizing the murine unilateral focal control cortical impact (CCI) injury, we evaluated seizure onset using 24/7 EEG video analysis at 2-4 months post-injury. Cellular changes in the dentate gyrus and hilus of the hippocampus were quantified by unbiased stereology and Imaris image analysis to evaluate Prox1-positive cell migration, astrocyte branching, and morphology, as well as neuronal loss at four months post-injury. Isolation of region-specific astrocytes and RNA-Seq were performed to determine differential gene expression in animals that developed post-traumatic epilepsy (PTE+) vs. those animals that did not (PTE-), which may be associated with epileptogenesis. RESULTS: CCI injury resulted in 37% PTE incidence, which increased with injury severity and hippocampal damage. Histological assessments uncovered a significant loss of hilar interneurons that coincided with aberrant migration of Prox1-positive granule cells and reduced astroglial branching in PTE+ compared to PTE- mice. We uniquely identified Cst3 as a PTE+-specific gene signature in astrocytes across all brain regions, which showed increased astroglial expression in the PTE+ hilus. CONCLUSIONS: These findings suggest that epileptogenesis may emerge following TBI due to distinct aberrant cellular remodeling events and key molecular changes in the dentate gyrus of the hippocampus.

Amide-to-ester substitution as a stable alternative to N-methylation for increasing membrane permeability in cyclic peptides.

Naturally occurring peptides with high membrane permeability often have ester bonds on their backbones. However, the impact of amide-to-ester substitutions on the membrane permeability of peptides has not been directly evaluated. Here we report the effect of amide-to-ester substitutions on the membrane permeability and conformational ensemble of cyclic peptides related to membrane permeation. Amide-to-ester substitutions are shown to improve the membrane permeability of dipeptides and a model cyclic hexapeptide. NMR-based conformational analysis and enhanced sampling molecular dynamics simulations suggest that the conformational transition of the cyclic hexapeptide upon membrane permeation is differently influenced by an amide-to-ester substitution and an amide N-methylation. The effect of amide-to-ester substitution on membrane permeability of other cyclic hexapeptides, cyclic octapeptides, and a cyclic nonapeptide is also investigated to examine the scope of the substitution. Appropriate utilization of amide-to-ester substitution based on our results will facilitate the development of membrane-permeable peptides.

The role of STING signaling in central nervous system infection and neuroinflammatory disease.

The cyclic guanosine monophosphate-adenosine monophosphate (GMP-AMP) synthase-Stimulator of Interferon Genes (cGAS-STING) pathway is a critical innate immune mechanism for detecting the presence of double-stranded DNA (dsDNA) and prompting a robust immune response. Canonical cGAS-STING activation occurs when cGAS, a predominantly cytosolic pattern recognition receptor, binds microbial DNA to promote STING activation. Upon STING activation, transcription factors enter the nucleus to cause the production of Type I interferons, inflammatory cytokines whose primary function is to prime the host for viral infection by producing a number of antiviral interferon-stimulated genes. While the pathway was originally described in viral infection, more recent studies have implicated cGAS-STING signaling in a number of different contexts, including autoimmune disease, cancer, injury, and neuroinflammatory disease. This review focuses on how our understanding of the cGAS-STING pathway has evolved over time with an emphasis on the role of STING-mediated neuroinflammation and infection in the nervous system. We discuss recent findings on how STING signaling contributes to the pathology of pain, traumatic brain injury, and stroke, as well as how mitochondrial DNA may promote STING activation in common neurodegenerative diseases. We conclude by commenting on the current knowledge gaps that should be filled before STING can be an effective therapeutic target in neuroinflammatory disease. This article is categorized under: Neurological Diseases > Molecular and Cellular Physiology Infectious Diseases > Molecular and Cellular Physiology Immune System Diseases > Molecular and Cellular Physiology.

Monocyte proinflammatory phenotypic control by ephrin type A receptor 4 mediates neural tissue damage.

Circulating monocytes have emerged as key regulators of the neuroinflammatory milieu in a number of neuropathological disorders. Ephrin type A receptor 4 (Epha4) receptor tyrosine kinase, a prominent axon guidance molecule, has recently been implicated in the regulation of neuroinflammation. Using a mouse model of brain injury and a GFP BM chimeric approach, we found neuroprotection and a lack of significant motor deficits marked by reduced monocyte/macrophage cortical infiltration and an increased number of arginase-1+ cells in the absence of BM-derived Epha4. This was accompanied by a shift in monocyte gene profile from pro- to antiinflammatory that included increased Tek (Tie2 receptor) expression. Inhibition of Tie2 attenuated enhanced expression of M2-like genes in cultured Epha4-null monocytes/macrophages. In Epha4-BM-deficient mice, cortical-isolated GFP+ monocytes/macrophages displayed a phenotypic shift from a classical to an intermediate subtype, which displayed reduced Ly6chi concomitant with increased Ly6clo- and Tie2-expressing populations. Furthermore, clodronate liposome-mediated monocyte depletion mimicked these effects in WT mice but resulted in attenuation of phenotype in Epha4-BM-deficient mice. This demonstrates that monocyte polarization not overall recruitment dictates neural tissue damage. Thus, coordination of monocyte proinflammatory phenotypic state by Epha4 is a key regulatory step mediating brain injury.

Conditional Deletion of EphA4 on Cx3cr1-Expressing Microglia Fails to Influence Histopathological Outcome and Blood Brain Barrier Disruption Following Brain Injury.

Erythropoietin-producing human hepatocellular receptors play a major role in central nervous system injury. Preclinical and clinical studies revealed the upregulation of erythropoietin-producing human hepatocellular A4 (EphA4) receptors in the brain after acute traumatic brain injury. We have previously reported that Cx3cr1-expressing cells in the peri-lesion show high levels of EphA4 after the induction of controlled cortical impact (CCI) injury in mice. Cx3cr1 is a fractalkine receptor expressed on both resident microglia and peripheral-derived macrophages. The current study aimed to determine the role of microglial-specific EphA4 in CCI-induced damage. We used Cx3cr1 CreER/+ knock-in/knock-out mice, which express EYFP in Cx3cr1-positive cells to establish microglia, EphA4-deficient mice following 1-month tamoxifen injection. Consistent with our previous findings, induction of CCI in wild-type (WT) Cx3cr1 CreER/+ EphA4 +/+ mice increased EphA4 expression on EYFP-positive cells in the peri-lesion. To distinguish between peripheral-derived macrophages and resident microglia, we exploited GFP bone marrow-chimeric mice and found that CCI injury increased EphA4 expression in microglia (TMEM119+GFP-) using immunohistochemistry. Using Cx3cr1 CreER/+ EphA4 f/f (KO) mice, we observed that the EphA4 mRNA transcript was undetected in microglia but remained present in whole blood when compared to WT. Finally, we found no difference in lesion volume or blood-brain barrier (BBB) disruption between WT and KO mice at 3 dpi. Our data demonstrate a nonessential role of microglial EphA4 in the acute histopathological outcome in response to CCI.

Geometrically Diverse Lariat Peptide Scaffolds Reveal an Untapped Chemical Space of High Membrane Permeability.

Constrained, membrane-permeable peptides offer the possibility of engaging challenging intracellular targets. Structure-permeability relationships have been extensively studied in cyclic peptides whose backbones are cyclized from head to tail, like the membrane permeable and orally bioavailable natural product cyclosporine A. In contrast, the physicochemical properties of lariat peptides, which are cyclized from one of the termini onto a side chain, have received little attention. Many lariat peptide natural products exhibit interesting biological activities, and some, such as griselimycin and didemnin B, are membrane permeable and have intracellular targets. To investigate the structure-permeability relationships in the chemical space exemplified by these natural products, we generated a library of scaffolds using stable isotopes to encode stereochemistry and determined the passive membrane permeability of over 1000 novel lariat peptide scaffolds with molecular weights around 1000. Many lariats were surprisingly permeable, comparable to many known orally bioavailable drugs. Passive permeability was strongly dependent on N-methylation, stereochemistry, and ring topology. A variety of structure-permeability trends were observed including a relationship between alternating stereochemistry and high permeability, as well as a set of highly permeable consensus sequences. For the first time, robust structure-permeability relationships are established in synthetic lariat peptides exceeding 1000 compounds.

Abrogation of atypical neurogenesis and vascular-derived EphA4 prevents repeated mild TBI-induced learning and memory impairments.

Brain injury resulting from repeated mild traumatic insult is associated with cognitive dysfunction and other chronic co-morbidities. The current study tested the effects of aberrant neurogenesis in a mouse model of repeated mild traumatic brain injury (rmTBI). Using Barnes Maze analysis, we found a significant reduction in spatial learning and memory at 24 days post-rmTBI compared to repeated sham (rSham) injury. Cell fate analysis showed a greater number of BrdU-labeled cells which co-expressed Prox-1 in the DG of rmTBI-injured mice which coincided with enhanced cFos expression for neuronal activity. We then selectively ablated dividing neural progenitor cells using a 7-day continuous infusion of Ara-C prior to rSham or rmTBI. This resulted in attenuation of cFos and BrdU-labeled cell changes and prevented associated learning and memory deficits. We further showed this phenotype was ameliorated in EphA4f./f/Tie2-Cre knockout compared to EphA4f./f wild type mice, which coincided with altered mRNA transcript levels of MCP-1, Cx43 and TGFß. These findings demonstrate that cognitive decline is associated with an increased presence of immature neurons and gene expression changes in the DG following rmTBI. Our data also suggests that vascular EphA4-mediated neurogenic remodeling adversely affects learning and memory behavior in response to repeated insult.

Using a neural network to predict deviations in mean heart dose during the treatment of left-sided deep inspiration breath hold patients.

PURPOSE: We investigated if a neural network could be used to predict the change in mean heart dose when a patient's heart deviates from its planned position during radiotherapy treatment. METHODS: Predictions were made based on parameters available at the time of treatment planning. The dose prescription, deep inspiration breath-hold (DIBH) amplitude, heart volume, lung volume, V90% and mean heart dose were used to predict the increase in dose to the heart when a shift towards the treatment field was undertaken. The network was trained using 3 mm, 5 mm and 7 mm shifts in heart positions for 50 patients' giving 150 data points in total. The neural network architecture was also varied to find the most optimal network design. The final neural network was then tested using cross-validation to evaluate the model's ability to generalise to new data. RESULTS: The optimal neural network found was comprised of a single hidden layer of 30 neurons. Based on twenty train/test splits, 94% of all prediction errors were below 0.2 Gy, 97.3% were below 0.3 Gy and 100% were below 0.5 Gy. The average RMSE and maximum prediction error over all train/test splits were 0.13 Gy and 0.5 Gy respectively. CONCLUSIONS: Our approach using a neural network provides a clinically acceptable estimate of the increase in Mean Heart Dose (MHD), without the need for further imaging, contouring or evaluation. The trained neural network gives clinicians the information and tools required to evaluate what shift in heart position would be acceptable and which scenarios require immediate action before treatment continues.

Lactobacillus rescues postnatal neurobehavioral and microglial dysfunction in a model of maternal microbiome dysbiosis.

Increasing reports of pregnancy events leading to maternal microbiome dysbiosis (MMD) show strong correlates with atypical neurodevelopmental outcomes. However, the mechanism(s) driving microbiome-mediated behavioral dysfunction in offspring remain understudied. Here, we demonstrate the presence of a novel gut commensal bacterium strain, Lactobacillus murinus HU-1, was sufficient to rescue behavioral deficits and brain region-specific microglial activationobserved in MMD-reared murine offspring. We furtheridentified a postnatal window of susceptibility that could prevent social impairments with timed maternal administration of the symbiotic bacterium. Moreover, MMD increased expression of microglial senescence genes, Trp53 and Il1ß, and Cx3cr1 protein in the prefrontal cortex, which correlated with dysfunctional modeling of synapses and accompanied dysbiosis-induced microglial activation. MMD male offspring harboring Lactobacillus murinus HU-1 or lacking Cx3cr1 showed amelioration of these effects. The current study describes a new avenue of influence by which maternally transferred Lactobacillus drives proper development of social behavior in the offspring through microglia-specific regulation of Cx3cr1 signaling.

Lipophilic Permeability Efficiency Reconciles the Opposing Roles of Lipophilicity in Membrane Permeability and Aqueous Solubility.

As drug discovery moves increasingly toward previously "undruggable" targets such as protein-protein interactions, lead compounds are becoming larger and more lipophilic. Although increasing lipophilicity can improve membrane permeability, it can also incur serious liabilities, including poor water solubility, increased toxicity, and faster metabolic clearance. Here we introduce a new efficiency metric, especially relevant to "beyond rule of 5" molecules, that captures, in a simple, unitless value, these opposing effects of lipophilicity on molecular properties. Lipophilic permeability efficiency (LPE) is defined as log D7.4dec/w - mlipocLogP + bscaffold, where log D7.4dec/w is the experimental decadiene-water distribution coefficient (pH 7.4), cLogP is the calculated octanol-water partition coefficient, and mlipo and bscaffold are scaling factors to standardize LPE values across different cLogP metrics and scaffolds. Using a variety of peptidic and nonpeptidic macrocycle drugs, we show that LPE provides a functional assessment of the efficiency with which a compound achieves passive membrane permeability at a given lipophilicity.

A Manganese Pre-Catalyst: Mild Reduction of Amides, Ketones, Aldehydes, and Esters.

A new (N-phosphinoamidinate)manganese complex is shown to be a useful pre-catalyst for the hydrosilative reduction of carbonyl compounds, and in most cases at room temperature. The Mn-catalyzed reduction of tertiary amides to tertiary amines, with a useful scope, is demonstrated for the first time by use of this catalyst, and is competitive with the most effective transition-metal catalysts known for such transformations. Ketones, aldehydes, and esters were also successfully reduced under mild conditions by using this new Mn catalyst.

Dehydrogenative B-H/C(sp3 )-H Benzylic Borylation within the Coordination Sphere of Platinum(II).

The first examples of stoichiometric dehydrogenative B-H/C(sp3 )-H benzylic borylation reactions, which are of relevance to catalytic methylarene (di)borylation, are reported. These unusual transformations involving a (κ2 -P,N)Pt(η3 -benzyl) complex, and either pinacolborane or catecholborane, proceed cleanly at room temperature. Density functional calculations suggest that borylation occurs via successive σ-bond metathesis steps, whereby a PtII -H intermediate engages in C(sp3 )-H bond activation-induced dehydrogenation.

Synthesis and Reactivity of a Neutral, Three-Coordinate Platinum(II) Complex Featuring Terminal Amido Ligation.

A crystallographically characterized three-coordinate, formally 14 electron Pt(II) complex 1 featuring terminal amido ligation is reported. Computational analysis revealed relatively weak π donation from the amide lone pair to platinum and supports a 14-electron assignment for 1. Stoichiometric reactivity studies confirmed the viability of net O-H and C-H addition across, as well as isonitrile insertion into, the terminal platinum-amido linkage of 1.

Automatic extraction of property norm-like data from large text corpora.

Traditional methods for deriving property-based representations of concepts from text have focused on either extracting only a subset of possible relation types, such as hyponymy/hypernymy (e.g., car is-a vehicle) or meronymy/metonymy (e.g., car has wheels), or unspecified relations (e.g., car--petrol). We propose a system for the challenging task of automatic, large-scale acquisition of unconstrained, human-like property norms from large text corpora, and discuss the theoretical implications of such a system. We employ syntactic, semantic, and encyclopedic information to guide our extraction, yielding concept-relation-feature triples (e.g., car be fast, car require petrol, car cause pollution), which approximate property-based conceptual representations. Our novel method extracts candidate triples from parsed corpora (Wikipedia and the British National Corpus) using syntactically and grammatically motivated rules, then reweights triples with a linear combination of their frequency and four statistical metrics. We assess our system output in three ways: lexical comparison with norms derived from human-generated property norm data, direct evaluation by four human judges, and a semantic distance comparison with both WordNet similarity data and human-judged concept similarity ratings. Our system offers a viable and performant method of plausible triple extraction: Our lexical comparison shows comparable performance to the current state-of-the-art, while subsequent evaluations exhibit the human-like character of our generated properties.

(1S*,2R*,3S*,4R*,5R*)-5-Tetra-decyloxy-methyl-7-oxabicyclo-[2.2.1]heptane-2,3-dicarb-oxy-lic anhydride.

In the title compound, C23H38O5, the oxabicyclo-[2.2.1]heptane-2,3-dicarb-oxy-lic anhydride unit has a normal geometry and the tetra-decoxymethyl side chain is fully extended. In the crystal, mol-ecules are linked head-to-head by C-H⋯O hydrogen bonds, forming two-dimensional networks propagating along the a and c-axis directions.

A method for the prediction of late organ-at-risk toxicity after radiotherapy of the prostate using equivalent uniform dose.

PURPOSE: To evaluate the predictive value of equivalent uniform doses (EUD) for late bladder and rectal toxicity after high-dose three-dimensional conformal radiation therapy (3D-CRT) to the prostate. MATERIALS AND METHODS: Using the method developed by Kutcher et al., EUDs for whole bladder and rectum were calculated from the dose-volume histograms of 180 patients with localized prostate cancer treated to 70-74 Gy with 3D-CRT. Late complications were recorded using the Radiation Therapy Oncology Group scale, correlated against EUD and known physical predictive indicators. RESULTS: EUD is an independent prognostic factor for Grade 2+ long-term rectal and bladder toxicity after radiation treatment to the prostate. Patients receiving an EUD >63.1 Gy to the rectum have a statistically significant (10% vs. 30%; p = 0.002) higher risk of developing Grade 2+ late complications. Patients receiving an EUD >53.4 Gy to the bladder have a statistically significant (10% vs. 33%; p = 0.001) higher risk of developing Grade 2+ late complications. CONCLUSIONS: It has been demonstrated that EUD is a strong independent predictive factor for Grade 2+ late complications after 3D-CRT to the prostate. Threshold values have been demonstrated for both bladder and rectum, above which there is a clinically significant increased risk of complications.

A randomised comparison of two brachytherapy devices for the treatment of uterine cervical carcinoma.

Two brachytherapy applicators commonly used in the treatment of uterine cervical carcinoma, the Henschke shielded and Fletcher-Suit-Declos, were randomly and prospectively compared in vivo in 20 patients. Based on two-dimensional planning and the ICRU-38 bladder and rectal reference points, an advantage for the shielded Henschke applicator was demonstrated.