A chemical probe to modulate human GID4 Pro/N-degron interactions.

The C-terminal to LisH (CTLH) complex is a ubiquitin ligase complex that recognizes substrates with Pro/N-degrons via its substrate receptor Glucose-Induced Degradation 4 (GID4), but its function and substrates in humans remain unclear. Here, we report PFI-7, a potent, selective and cell-active chemical probe that antagonizes Pro/N-degron binding to human GID4. Use of PFI-7 in proximity-dependent biotinylation and quantitative proteomics enabled the identification of GID4 interactors and GID4-regulated proteins. GID4 interactors are enriched for nucleolar proteins, including the Pro/N-degron-containing RNA helicases DDX21 and DDX50. We also identified a distinct subset of proteins whose cellular levels are regulated by GID4 including HMGCS1, a Pro/N-degron-containing metabolic enzyme. These data reveal human GID4 Pro/N-degron targets regulated through a combination of degradative and nondegradative functions. Going forward, PFI-7 will be a valuable research tool for investigating CTLH complex biology and facilitating development of targeted protein degradation strategies that highjack CTLH E3 ligase activity.

Interaction of the C2C12 myotube contractions and glucose availability on transcriptome and extracellular vesicle microRNAs.

Exercise-like electrical pulse stimulation (EL-EPS) of myotubes mimics many key physiological changes induced by in vivo exercise. Besides enabling intracellular research, EL-EPS allows to study secreted factors, including muscle-specific microRNAs (myomiRs) carried in extracellular vesicles (EVs). These factors can participate in contraction-induced intercellular cross talk and may mediate the health benefits of exercise. However, the current knowledge of these responses, especially under variable nutritional conditions, is limited. We investigated the effects of EL-EPS on C2C12 myotube transcriptome in high- and low-glucose conditions by messenger RNA sequencing, while the expression of EV-carried miRNAs was analyzed by small RNA sequencing and RT-qPCR. We show that higher glucose availability augmented contraction-induced transcriptional changes and that the majority of the differentially expressed genes were upregulated. Furthermore, based on the pathway analyses, processes related to contractility and cytokine/inflammatory responses were upregulated. In addition, we report that EL-EPS increased packing of miR-1-3p into EVs independent of glucose availability. Together our findings suggest that in vitro EL-EPS is a usable tool not only to study contraction-induced intracellular mechanisms but also extracellular responses. The distinct transcriptional changes observed under variable nutritional conditions emphasize the importance of careful consideration of media composition in future exercise-mimicking studies.NEW & NOTEWORTHY The present study examined for the first time the effects of exercise-like electrical pulse stimulation administered under distinct nutritional conditions on 1) the transcriptome of the C2C12 myotubes and 2) their media containing extracellular vesicle-carried microRNAs. We report that higher glucose availability augmented transcriptional responses related especially to contractility and cytokine/inflammatory pathways. Agreeing with in vivo studies, we show that the packing of exercise-responsive miR-1-3p was increased in the extracellular vesicles in response to myotube contractions.

Metabolomics analysis reveals dysregulation in one carbon metabolism in Friedreich Ataxia.

Friedreich Ataxia (FA) is a rare and often fatal autosomal recessive disease in which a mitochondrial protein, frataxin (FXN), is severely reduced in all tissues. With loss of FXN, mitochondrial metabolism is severely disrupted. Multiple therapeutic approaches are in development, but a key limitation is the lack of biomarkers reflecting the activity of FXN in a timely fashion. We predicted this dysregulated metabolism would present a unique metabolite profile in blood of FA patients versus Controls (Con). Plasma from 10 FA and 11 age and sex matched Con subjects was analyzed by targeted mass spectrometry and untargeted NMR. This combined approach yielded quantitative measurements for 540 metabolites and found 59 unique metabolites (55 from MS and 4 from NMR) that were significantly different between cohorts. Correlation-based network analysis revealed several clusters of pathway related metabolites including a cluster associated with one­carbon (1C) metabolism composed of formate, sarcosine, hypoxanthine, and homocysteine. Receiver operator characteristics analyses demonstrated an excellent ability to discriminate between Con and FA with AUC values >0.95. These results are the first reported metabolomic analyses of human patients with FA. The metabolic perturbations, especially those related to 1C metabolism, may serve as a valuable biomarker panel of disease progression and response to therapy. The identification of dysregulated 1C metabolism may also inform the search for new therapeutic targets related to this pathway.

Circulating trimethylamine N-oxide levels following fish or seafood consumption.

PURPOSE: Some species of fish and seafood are high in trimethylamine N-oxide (TMAO), which accumulates in muscle where it protects against pressure and cold. Trimethylamine (TMA), the metabolic precursor to TMAO, is formed in fish during bacterial spoilage. Fish intake is promoted for its potential cardioprotective effects. However, numerous studies show TMAO has pro-atherothrombotic properties. Here, we determined the effects of fish or seafood consumption on circulating TMAO levels in participants with normal renal function. METHODS: TMAO and omega-3 fatty acid content were quantified across multiple different fish or seafood species by mass spectrometry. Healthy volunteers (n = 50) were recruited for three studies. Participants in the first study consented to 5 consecutive weekly blood draws and provided dietary recall for the 24 h preceding each draw. In the second study, TMAO levels were determined following defined low and high TMAO diets. Finally, participants consumed test meals containing shrimp, tuna, fish sticks, salmon or cod. TMAO levels were quantified by mass spectrometry in blood collected before and after dietary challenge. RESULTS: TMAO + TMA content varied widely across fish and seafood species. Consumption of fish sticks, cod, and to a lesser extent salmon led to significant increases in circulating TMAO levels. Within 1 day, circulating TMAO concentrations in all participants returned to baseline levels. CONCLUSIONS: We conclude that some fish and seafood contain high levels of TMAO, and may induce a transient elevation in TMAO levels in some individuals. Selection of low TMAO content fish is prudent for subjects with elevated TMAO, cardiovascular disease or impaired renal function.

Indirect treatment comparison of oral versus injectable azacitidine as maintenance therapy for acute myeloid leukemia.

Aim: Evaluate the relative efficacy of oral versus injectable azacitidine (AZA) maintenance therapy in acute myeloid leukemia (AML) after complete remission. Materials & methods: Systematic literature review identified QUAZAR AML-001, HOVON 97 AML, UK NCRI AML16 and QoLESS-AZA-AMLE (sensitivity analysis) trials. Network meta-analysis and matching-adjusted indirect comparisons assessed survival outcomes. Results: In the network meta-analysis, combining the HOVON 97 and UK NCRI trials, oral AZA (QUAZAR) was associated with significantly improved overall survival (OS) versus injectable AZA (hazard ratio: 0.744; 95% credible interval: 0.557-0.998). After matching-adjusted indirect comparisons, to address differences in patient characteristics across trials, OS improvements were maintained with oral versus injectable AZA (hazard ratio: 0.753; credible interval: 0.563-0.998). Conclusion: In AML, maintenance therapy with oral AZA was associated with improved OS versus injectable AZA.

Older people with acute myeloid leukemia (AML) may have remission with or without blood count recovery, after first-line chemotherapy; however, remission is short lived and overall survival is limited (7­12 months). Ongoing treatment (maintenance therapy) after response to initial chemotherapy may prolong remission. Maintenance therapy with azacitidine (AZA) given by injection beneath the skin (subcutaneous) or into a vein (intravenous) can extend disease-free survival compared with no further treatment and best supportive care. However, treatment with intravenous AZA may only extend overall survival in certain patients. ONUREG^® is a novel formulation of AZA that can be taken by mouth (orally), remains in the body for longer periods and has the potential for significant clinical benefits compared with intravenous AZA. Presently, there are no studies directly comparing outcomes of maintenance therapy with oral and injectable AZA in older people with AML. In this analysis, we used an indirect treatment comparison method including four clinical trials to explore the survival benefit associated with ONUREG and injectable AZA when used as maintenance therapies after response to initial chemotherapy in older people with AML. Findings showed ONUREG significantly improved overall survival compared with injectable AZA, with an almost 26% reduction in the risk of death. These results suggest that maintenance therapy with ONUREG significantly improves overall survival compared with injectable AZA in older people with AML who may have remission with or without blood count recovery, after first-line chemotherapy.

Higher glucose availability augments the metabolic responses of the C2C12 myotubes to exercise-like electrical pulse stimulation.

The application of exercise-like electrical pulse simulation (EL-EPS) has become a widely used exercise mimetic in vitro. EL-EPS produces similar physiological responses as in vivo exercise, while less is known about the detailed metabolic effects. Routinely, the C2C12 myotubes are cultured in high-glucose medium (4.5 g/L), which may alter EL-EPS responses. In this study, we evaluate the metabolic effects of EL-EPS under the high- and low-glucose (1.0 g/L) conditions to understand how substrate availability affects the myotube response to EL-EPS. The C2C12 myotube, media, and cell-free media metabolites were analyzed using untargeted nuclear magnetic resonance (NMR)-based metabolomics. Furthermore, translational and metabolic changes and possible exerkine effects were analyzed. EL-EPS enhanced substrate utilization as well as production and secretion of lactate, acetate, 3-hydroxybutyrate, and branched-chain fatty acids (BCFAs). The increase in BCFAs correlated with branched-chain amino acids (BCAAs) and BCFAs were strongly decreased when myotubes were cultured without BCAAs suggesting the action of acyl-CoA thioesterases on BCAA catabolites. Notably, not all EL-EPS responses were augmented by high glucose because EL-EPS increased phosphorylated c-Jun N-terminal kinase and interleukin-6 secretion independent of glucose availability. Administration of acetate and EL-EPS conditioned media on HepG2 hepatocytes had no adverse effects on lipolysis or triacylglycerol content. Our results demonstrate that unlike in cell-free media, the C2C12 myotube and media metabolites were affected by EL-EPS, particularly under high-glucose condition suggesting that media composition should be considered in future EL-EPS studies. Furthermore, acetate and BCFAs were identified as putative exerkines warranting more research.NEW & NOTEWORTHY The present study examined for the first time the metabolome of 1) C2C12 myotubes, 2) their growth media, and 3) cell-free media after exercise-like electrical pulse stimulation under distinct nutritional loads. We report that myotubes grown under high-glucose conditions had greater responsiveness to EL-EPS when compared with lower glucose availability conditions and increased media content of acetate and branched-chain fatty acids suggests they might act as putative exerkines warranting further research.

Metabolic profiles identify circulating biomarkers associated with heart failure in young single ventricle patients.

BACKGROUND: Children and young adults with single ventricle (SV) heart disease frequently develop heart failure (HF) that is intractable and difficult to treat. Our understanding of the molecular and biochemical reasons underlying this is imperfect. Thus, there is an urgent need for biomarkers that predict outcome and provide a rational basis for treatment, and advance our understanding of the basis of HF. OBJECTIVE: We sought to determine if a metabolomic approach would provide biochemical signatures of HF in SV children and young adults. If significant, these analytes might serve as biomarkers to predict outcome and inform on the biological mechanism(s) of HF. METHODS: We applied a multi-platform metabolomics approach composed of mass spectrometry (MS) and nuclear magnetic resonance (NMR) which yielded 495 and 26 metabolite measurements respectively. The plasma samples came from a cross-sectional set of young SV subjects, ages 2-19 years with ten control (Con) subjects and 16 SV subjects. Of the SV subjects, nine were diagnosed as congestive HF (SVHF), and 7 were not in HF. Metabolomic data were correlated with clinical status to determine if there was a signature associated with HF. RESULTS: There were no differences in age, height, weight or sex between the 3 cohorts. However, statistical analysis of the metabolomic profiles using ANOVA revealed 44 metabolites with significant differences between cohorts including 41 profiled by MS and 3 by NMR. These metabolites included acylcarnitines, amino acids, and bile acids, which distinguished Con from all SV subjects. Furthermore, metabolite profiles could distinguish between SV and SVHF subjects. CONCLUSION: These are the first data to demonstrate a clear metabolomic signature associated with HF in children and young adults with SV. Larger studies are warranted to determine if these findings are predictive of progression to HF in time to provide intervention.

Searching through functional space reveals distributed visual, auditory, and semantic coding in the human brain.

The extent to which brain functions are localized or distributed is a foundational question in neuroscience. In the human brain, common fMRI methods such as cluster correction, atlas parcellation, and anatomical searchlight are biased by design toward finding localized representations. Here we introduce the functional searchlight approach as an alternative to anatomical searchlight analysis, the most commonly used exploratory multivariate fMRI technique. Functional searchlight removes any anatomical bias by grouping voxels based only on functional similarity and ignoring anatomical proximity. We report evidence that visual and auditory features from deep neural networks and semantic features from a natural language processing model, as well as object representations, are more widely distributed across the brain than previously acknowledged and that functional searchlight can improve model-based similarity and decoding accuracy. This approach provides a new way to evaluate and constrain computational models with brain activity and pushes our understanding of human brain function further along the spectrum from strict modularity toward distributed representation.

Applications of Lipidomics to Age-Related Musculoskeletal Disorders.

PURPOSE OF REVIEW: The goal of this review is to highlight the need for new biomarkers for the diagnosis and treatment of musculoskeletal disorders, especially osteoporosis and sarcopenia. These conditions are characterized by loss of bone and muscle mass, respectively, leading to functional deterioration and the development of disabilities. Advances in high-resolution lipidomics platforms are being used to help identify new lipid biomarkers for these diseases. RECENT FINDINGS: It is now well established that bone and muscle have important endocrine functions, including the release of bioactive factors in response to mechanical and biochemical stimuli. Bioactive lipids are a prominent set of these factors and some of these lipids are directly related to the mass and function of bone and muscle. Recent lipidomics studies have shown significant dysregulation of lipids in aged muscle and bone, including alterations in diacylglycerols and ceramides. Studies have shown that alterations in some types of plasma lipids are associated with aging including reduced bone mineral density and the occurrence of osteoporosis. Musculoskeletal disorders are a major burden in our society, especially for older adults. The development and application of new lipidomics methods is making significant advances in identifying new biomarkers for these diseases. These studies will not only lead to improved detection, but new mechanistic insights that could lead to new therapeutic targets and interventions.

Nicardipine: When high dose nitrates fail in treating heart failure.

Sympathetic Crashing Acute Pulmonary Edema (SCAPE) describes patients who present with acute hypertensive cardiogenic pulmonary edema. These patients present in respiratory distress, and requiring immediate medical and airway management. The treatment of SCAPE includes non-invasive positive pressure ventilation (NIPPV) to maintain oxygenation, and high dose nitrates to lower blood pressure and reduce afterload. We present a case report of a patient with refractory hypertension to high dose nitrates likely due to nitroglycerin resistance or an attenuated response. The addition of nicardipine led to marked clinical improvement, normalized blood pressure and spared the patient from endotracheal intubation and admission to the intensive care unit.

PDK4 drives metabolic alterations and muscle atrophy in cancer cachexia.

Cachexia is frequently accompanied by severe metabolic derangements, although the mechanisms responsible for this debilitating condition remain unclear. Pyruvate dehydrogenase kinase (PDK)4, a critical regulator of cellular energetic metabolism, was found elevated in experimental models of cancer, starvation, diabetes, and sepsis. Here we aimed to investigate the link between PDK4 and the changes in muscle size in cancer cachexia. High PDK4 and abnormal energetic metabolism were found in the skeletal muscle of colon-26 tumor hosts, as well as in mice fed a diet enriched in Pirinixic acid, previously shown to increase PDK4 levels. Viral-mediated PDK4 overexpression in myotube cultures was sufficient to promote myofiber shrinkage, consistent with enhanced protein catabolism and mitochondrial abnormalities. On the contrary, blockade of PDK4 was sufficient to restore myotube size in C2C12 cultures exposed to tumor media. Our data support, for the first time, a direct role for PDK4 in promoting cancer-associated muscle metabolic alterations and skeletal muscle atrophy.-Pin, F., Novinger, L. J., Huot, J. R., Harris, R. A., Couch, M. E., O'Connell, T. M., Bonetto, A. PDK4 drives metabolic alterations and muscle atrophy in cancer cachexia.

Transthoracic echocardiographic findings in patients admitted with SARS-CoV-2 infection.

INTRODUCTION: Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2)-infected patients commonly have elevated troponin and D-dimer levels, but limited imaging exists to support most likely etiologies in efforts to avoid staff exposure. The purpose of this study was to report transthoracic echocardiographic (TTE) findings in SARS-CoV-2 patients with correlating troponin and D-dimer levels. METHODS: We identified 66 SARS-CoV-2 patients (mean age 60 ± 15.7 years) admitted within a large, eight-hospital healthcare system over a 6-week period with a TTE performed. TTE readers were blinded to laboratory data with intra-observer and inter-observer analysis assessed. RESULTS: Sixty-six of 1780 SARS-CoV-2 patients were included and represented a high-risk population as 38 (57.6%) were ICU-admitted, 47 (71.2%) had elevated D-dimer, 41 (62.1%) had elevated troponin, and 25 (37.9%) died. Right ventricular (RV) dilation was present in 49 (74.2%) patients. The incidence and average D-dimer elevation was similar between moderate/severe vs. mild/no RV dilation (69.6% vs 67.6%, P = 1.0; 3736 ± 2986 vs 4141 ± 3351 ng/mL, P = .679). Increased left ventricular (LV) wall thickness was present in 46 (69.7%) with similar incidence of elevated troponin and average troponin levels compared to normal wall thickness (66.7% vs 52.4%, P = .231; 0.88 ± 1.9 vs 1.36 ± 2.4 ng/mL, P = .772). LV dilation was rare (n = 6, 9.1%), as was newly reduced LV ejection fraction (n = 2, 3.0%). CONCLUSION: TTE in SARS-CoV-2 patients is scarce, technically difficult, and reserved for high-risk patients. RV dilation is common in SARS-CoV-2 but does not correlate with elevated D-dimer levels. Increased LV wall thickness is common, while newly reduced LV ejection fraction is rare, and neither correlates with troponin levels.

Sperm function, protein phosphorylation, and metabolism differ in mice lacking successive sperm-specific glycolytic enzymes.

Glyceraldehyde 3-phosphate dehydrogenase-S (GAPDHS) and phosphoglycerate kinase 2 (PGK2), two isozymes restricted to the male germline, catalyze successive steps in the glycolytic pathway in mammalian sperm. Although gene targeting of each isozyme demonstrated that glycolysis is required for normal sperm motility and male fertility, the phenotype of mice lacking GAPDHS is more severe than that of mice lacking PGK2. This study examined sperm function, signaling pathways, and metabolism to investigate factors that contribute to the phenotypic differences between these knockout models. Sperm from the two knockouts exhibited comparable deficits in zona binding, in vitro fertilization with or without zona drilling, and capacitation-dependent tyrosine phosphorylation. In contrast, signaling and metabolic differences were apparent prior to capacitation. Phosphorylation of sperm protein phosphatase 1, which has been associated with the acquisition of motile capacity during epididymal maturation, was deficient only in GAPDHS-null sperm. Carnitine, choline, phosphocholine, and taurine were elevated in sperm from both knockouts immediately after collection from the epididymis. However, only carnitine levels in PGK2-null sperm were significantly different from wild-type sperm, while all four metabolites were significantly higher in GAPDHS-null sperm. We confirmed that glycolysis is required for robust hyperactivation, but found that the motility of PGK2-null sperm improved to levels comparable to wild-type sperm with pyruvate as the sole metabolic substrate. This nonglycolysable substrate did not improve progressive motility in GAPDHS-null sperm. These results identify multiple signaling and metabolic defects that are likely contributors to male infertility and the absence of progressive sperm motility seen in mice lacking GAPDHS.

An analysis of three levels of scaled-up coverage for 28 interventions to avert stillbirths and maternal, newborn and child mortality in 27 countries in Latin America and the Caribbean with the Lives Saved Tool (LiST).

BACKGROUND: Action to avert maternal and child mortality was propelled by the Millennium Development Goals (MDGs) in 2000. The Latin American and Caribbean (LAC) region has shown promise in achieving the MDGs in many countries, but preventable maternal, neonatal and child mortality persist. Furthermore, preventable stillbirths are occurring in large numbers in the region. While an effective set of maternal, newborn and child health (MNCH) interventions have been identified, they have not been brought to scale across LAC. METHODS: Baseline data for select MNCH interventions for 27 LAC countries that are included in the Lives Saved Tool (LiST) were verified and updated with survey data. Three LiST projections were built for each country: baseline, MDG-focused, and All Included, each scaling up a progressively larger set of interventions for 2015 - 2030. Impact was assessed for 2015 - 2035, comparing annual and total lives saved, as projected by LiST. RESULTS: Across the 27 countries 235,532 stillbirths, and 752,588 neonatal, 959,393 under-five, and 60,858 maternal deaths would be averted between 2015 and 2035 by implementing the All-Included intervention package, representing 67 %, 616 %, 807 % and 101 % more lives saved, respectively, than with the MDG-focused interventions. 25 % neonatal deaths averted with the All-Included intervention package would be due to asphyxia, 42 % from prematurity and 24 % from sepsis. CONCLUSIONS: Our modelling suggests a 337 % increase in the number of lives saved, which would have enormous impacts on population health. Further research could help clarify the impacts of a comprehensive scale-up of the full range of essential MNCH interventions we have modelled.

Synthesizing qualitative and quantitative evidence on non-financial access barriers: implications for assessment at the district level.

INTRODUCTION: A key element of the global drive to universal health coverage is ensuring access to needed health services for everyone, and to pursue this goal in an equitable way. This requires concerted efforts to reduce disparities in access through understanding and acting on barriers facing communities with the lowest utilisation levels. Financial barriers dominate the empirical literature on health service access. Unless the full range of access barriers are investigated, efforts to promote equitable access to health care are unlikely to succeed. This paper therefore focuses on exploring the nature and extent of non-financial access barriers. METHODS: We draw upon two structured literature reviews on barriers to access and utilization of maternal, newborn and child health services in Ghana, Bangladesh, Vietnam and Rwanda. One review analyses access barriers identified in published literature using qualitative research methods; the other in published literature using quantitative analysis of household survey data. We then synthesised the key qualitative and quantitative findings through a conjoint iterative analysis. RESULTS: Five dominant themes on non-financial access barriers were identified: ethnicity; religion; physical accessibility; decision-making, gender and autonomy; and knowledge, information and education. The analysis highlighted that non-financial factors pose considerable barriers to access, many of which relate to the acceptability dimension of access and are challenging to address. Another key finding is that quantitative research methods, while yielding important findings, are inadequate for understanding non-financial access barriers in sufficient detail to develop effective responses. Qualitative research is critical in filling this gap. The analysis also indicates that the nature of non-financial access barriers vary considerably, not only between countries but also between different communities within individual countries. CONCLUSIONS: To adequately understand access barriers as a basis for developing effective strategies to address them, mixed-methods approaches are required. From an equity perspective, communities with the lowest utilisation levels should be prioritised and the access barriers specific to that community identified. It is, therefore, critical to develop approaches that can be used at the district level to diagnose and act upon access barriers if we are to pursue an equitable path to universal health coverage.

Dissociation of salience-driven and content-driven spatial attention to scene category with predictive decoding of gaze patterns.

Scene content is thought to be processed quickly and efficiently to bias subsequent visual exploration.Does scene content bias spatial attention during task-free visual exploration of natural scenes?If so, is this bias driven by patterns of physical salience or content-driven biases formed through previous encounters with similar scenes? We conducted two eye-tracking experiments to address these questions. Using a novel gaze decoding method, we show that fixation patterns predict scene category during free exploration. Additionally, we isolate salience-driven contributions using computational salience maps and content-driven contributions using gaze-restricted fixation data. We find distinct time courses for salience-driven and content-driven effects. The influence of physical salience peaked initially but quickly fell off at 600 ms past stimulus onset. The influence of content effects started at chance and steadily increased over the 2000 ms after stimulus onset. The combination of these two components significantly explains the time course of gaze allocation during free exploration.

Solution-based indirect affinity selection mass spectrometry--a general tool for high-throughput screening of pharmaceutical compound libraries.

We show here that an automated solution-based affinity selection mass spectrometry (ASMS) system can be built exclusively from commercially available parts. The value of this technology lies in the throughput (~1 × 10(5) compounds/day) coupled with a low hit rate. The system, being a binding assay, requires little development time yielding a fast timeline between target availability and hit identification. In addition, the use of exact mass simplifies the hit identification. We demonstrate this system using carbonic anhydrase as the target and a library of 144,000 proprietary compounds.

Plasma metabolites and physical function in patients undergoing hemodialysis.

Impaired physical function contributes to falls, fractures, and mortality among patients undergoing dialysis. Using a metabolomic approach, we identified metabolite alterations and effect size-based composite scores for constructs of impaired gait speed and grip strength. 108 participants incident to dialysis had targeted plasma metabolomics via liquid chromatography-mass spectrometry and physical function assessed (i.e., 4 m walk, handgrip strength). Physical function measures were categorized as above/ below median, with grip utilizing sex-based medians. To develop composite scores, metabolites were identified via Wilcoxon uncorrected p < 0.05 and effect size > 0.40. Receiver operating characteristic analyses tested whether scores differentiated between above/below function groups. Participants were 54% male, 77% Black and 53 ± 14 y with dialysis vintage of 101 ± 50 days. Median (IQR) grip strength was 35.5 (11.1) kg (males) and 20 (8.4) kg (females); median gait speed was 0.82 (0.34) m/s. Of 246 measured metabolites, composite scores were composed of 22 and 12 metabolites for grip strength and gait speed, respectively. Area under the curve for metabolite composite was 0.88 (gait) and 0.911 (grip). Composite scores of physical function performed better than clinical parameters alone in patients on dialysis. These results provide potential pathways for interventions and needed validation in an independent cohort.

Chemical tools for the Gid4 subunit of the human E3 ligase C-terminal to LisH (CTLH) degradation complex.

We have developed a novel chemical handle (PFI-E3H1) and a chemical probe (PFI-7) as ligands for the Gid4 subunit of the human E3 ligase CTLH degradation complex. Through an efficient initial hit-ID campaign, structure-based drug design (SBDD) and leveraging the sizeable Pfizer compound library, we identified a 500 nM ligand for this E3 ligase through file screening alone. Further exploration identified a vector that is tolerant to addition of a linker for future chimeric molecule design. The chemotype was subsequently optimized to sub-100 nM Gid4 binding affinity for a chemical probe. These novel tools, alongside the suitable negative control also identified, should enable the interrogation of this complex human E3 ligase macromolecular assembly.