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Julita Chlebowicz,1 Radha Akella,1 John M Humphreys,1 Haixia He,1 Ashari R Kannangara,1 Shuguang Wei,2 Bruce Posner,2 Elizabeth J Goldsmith1 1Department of Biophysics, The University of Texas Southwestern Medical Center, Dallas, TX, USA; 2Department of Biochemistry, The University of Texas Southwestern Medical Center, Dallas, TX, USACorrespondence: Elizabeth J Goldsmith, Department of Biophysics, The University of Texas Southwestern Medical Center, 5323 Harry Hines Boulevard, Dallas, TX, 75390-8816, USA, Tel +1 214 645 6376, Email Elizabeth.goldsmith@utsouthwestern.eduIntroduction: WNK [with no lysine (K)] kinases are serine/threonine kinases associated with familial hyperkalemic hypertension (FHHt). WNKs are therapeutic targets for blood pressure regulation, stroke and several cancers including triple negative breast cancer and glioblastoma. Here, we searched for and characterized novel WNK kinase inhibitors.Methods: We used a ~210,000-compound library in a high-throughput screen, re-acquisition and assay, commercial specificity screens and crystallography to identify WNK-isoform-selective inhibitors.Results: We identified five classes of compounds that inhibit the kinase activity of WNK1: quinoline compounds, halo-sulfones, cyclopropane-containing thiazoles, piperazine-containing compounds, and nitrophenol-derived compounds. The compounds are strongly pan-WNK selective, inhibiting all four WNK isoforms. A class of quinoline compounds was identified that further shows selectivity among the WNK isoforms, being more potent toward WNK3 than WNK1. The crystal structure of the quinoline-derived SW120619 bound to the kinase domain of WNK3 reveals active site binding, and comparison to the WNK1 structure reveals the potential origin of isoform specificity.Discussion: The newly discovered classes of compounds may be starting points for generating pharmacological tools and potential drugs treating hypertension and cancer.Graphical Abstract: Keywords: WNK, inhibition screen, kinase inhibitor, WNK3-specific, structure–activity relationship, crystallography

In this paper, we develop a linear programming framework for computing a quadratic approximation to the value function, which constitutes the off-line computation of a hierarchical FMS scheduling approach previously developed by us. In contrast to previous work, where relatively crude value functions were used, we develop a quadratic approximation that is a prior fit. We consider the multiple part multiple machine discounted cost case and illustrate the approach via a simulation example in the context of an industrial setting. [ABSTRACT FROM AUTHOR]

Abstract Fabry disease (FD) is a treatable X linked lysosomal storage disorder with a wide phenotypic spectrum. There is a scarcity of published data on the burden of FD in India. This study evaluates the clinical and molecular spectrum of Indian patients with FD. In this multicentric study involving 10 tertiary referral centers in India, we analyzed the clinical course and genotype of 54 patients from 37 families. Family screening identified 19 new patients (35%) from 12 index cases. Then, 33 GLA gene variants were identified in 49/54 (90.7%) which included 11 novel and 22 known pathogenic variants. Of the 54 patients in our cohort, 40 patients had “classical” and 10 patients had a “nonclassical” presentation. The symptoms and signs included kidney dysfunction in 38/54 (70.3%), neuropathic pain in 34/54 (62.9%), left ventricular hypertrophy in 22/49 (44.8%) and stroke in 5/54 (9.2%). Female heterozygotes were 10/54 (18.5%) of whom 2 were index cases. There was a significant delay in reaching the diagnosis of 11.7 years. Enzyme replacement therapy was initiated in 28/54 (51.8%) patients with significant improvement of neuropathic pain and gastrointestinal symptoms. This study highlights the clinical presentation and mutational spectrum of FD in India and suggests that family screening and screening of high‐risk groups (hypertrophic cardiomyopathy, idiopathic chronic renal failure and cryptogenic stroke) could be the most cost‐effective strategies for early identification of FD.

In this paper, we solve a production planning problem that has a direct bearing on the cost to a facility of vendor quality and reliability. This solution is also an important first step in solving the multi-plant coordination problem. We consider a manufacturing facility with one critical raw material, and one or more products whose demand needs to be met. It is assumed that demand for finished goods and the arrival process for raw material are stochastic. We calculate simultaneously the optimal production level, and the raw material order quantity which minimize the total expected cost subject to capacity and release level constraints. The results obtained by our model (the integrated model) show that the production level should be no larger than the production level obtained by solving the traditional (nonintegrated) models, and that major savings are obtained by solving the integrated model. We are also able to characterize and evaluate the cost of reliable supplies of raw material. In addition we solve the above problem in the presence of capacity constraints. A procedure which allocates capacity to different products is presented. [ABSTRACT FROM AUTHOR]

Osmotic stress and chloride regulate the autophosphorylation and activity of the WNK1 and WNK3 kinase domains. The kinase domain of unphosphorylated WNK1 (uWNK1) is an asymmetric dimer possessing water molecules conserved in multiple uWNK1 crystal structures. Conserved waters are present in two networks, referred to here as conserved water networks 1 and 2 (CWN1 and CWN2). Here, we show that PEG400 applied to crystals of dimeric uWNK1 induces de-dimerization. Both the WNK1 the water networks and the chloride-binding site are disrupted by PEG400. CWN1 is surrounded by a cluster of pan-WNK-conserved charged residues. Here, we mutagenized these charges in WNK3, a highly active WNK isoform kinase domain, and WNK1, the isoform best studied crystallographically. Mutation of E314 in the Activation Loop of WNK3 (WNK3/E314Q and WNK3/E314A, and the homologous WNK1/E388A) enhanced the rate of autophosphorylation, and reduced chloride sensitivity. Other WNK3 mutants reduced the rate of autophosphorylation activity coupled with greater chloride sensitivity than wild-type. The water and chloride regulation thus appear linked. The lower activity of some mutants may reflect effects on catalysis. Crystallography showed that activating mutants introduced conformational changes in similar parts of the structure to those induced by PEG400. WNK activating mutations and crystallography support a role for CWN1 in WNK inhibition consistent with water functioning as an allosteric ligand., Competing Interests: LT, RA, JH, HH, EG No competing interests declared, (© 2023, Teixeira et al.)

Previous study has demonstrated that the WNK kinases 1 and 3 are direct osmosensors consistent with their established role in cell-volume control. WNK kinases may also be regulated by hydrostatic pressure. Hydrostatic pressure applied to cells in culture with N 2 gas or to Drosophila Malpighian tubules by centrifugation induces phosphorylation of downstream effectors of endogenous WNKs. In vitro, the autophosphorylation and activity of the unphosphorylated kinase domain of WNK3 (uWNK3) is enhanced to a lesser extent than in cells by 190 kPa applied with N 2 gas. Hydrostatic pressure measurably alters the structure of uWNK3. Data from size exclusion chromatography in line with multi-angle light scattering (SEC-MALS), SEC alone at different back pressures, analytical ultracentrifugation (AUC), NMR, and chemical crosslinking indicate a change in oligomeric structure in the presence of hydrostatic pressure from a WNK3 dimer to a monomer. The effects on the structure are related to those seen with osmolytes. Potential mechanisms of hydrostatic pressure activation of uWNK3 and the relationships of pressure activation to WNK osmosensing are discussed.

Background-Previous studies on coronavirus disease 2019 (COVID-19) were limited to specific geographical locations and small sample sizes. Therefore, we used the National Inpatient Sample (NIS) 2020 database to determine the risk factors for severe outcomes and mortality in COVID-19. Methods-We included adult patients with COVID-19. Univariate and multivariate logistic regression was performed to determine the predictors of severe outcomes and mortality in COVID-19. Results-1,608,980 (95% CI 1,570,803-1,647,156) hospitalizations with COVID-19 were included. Severe complications occurred in 78.3% of COVID-19 acute respiratory distress syndrome (ARDS) and 25% of COVID-19 pneumonia patients. The mortality rate for COVID-19 ARDS was 54% and for COVID-19 pneumonia was 16.6%. On multivariate analysis, age > 65 years, male sex, government insurance or no insurance, residence in low-income areas, non-white races, stroke, chronic kidney disease, heart failure, malnutrition, primary immunodeficiency, long-term steroid/immunomodulatory use, complicated diabetes mellitus, and liver disease were associated with COVID-19 related complications and mortality. Cardiac arrest, septic shock, and intubation had the highest odds of mortality. Conclusions-Socioeconomic disparities and medical comorbidities were significant determinants of mortality in the US in the pre-vaccine era. Therefore, aggressive vaccination of high-risk patients and healthcare policies to address socioeconomic disparities are necessary to reduce death rates in future pandemics.