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Due to increased fossil fuel use and fossil fuel limitations, the Indian energy industry is migrating to non-conventional energy resources such as solar power, wind production, and fuel cells. The unpredictability of non-conventional energy sources makes it difficult to balance an electrical system when they are incorporated, necessitating the incorporation of a storage device into the grid. In a microgrid system with wind turbine generation (WTG) and a battery energy storage system (BESS), the BESS may reserve energy during periods of surplus generation and release it to the grid during times of peak demand. The suggested technique establishes the state of charge (SOC) schedule for the BESS by employing an artificial rabbit optimisation (ARO) algorithm that minimises energy costs for customers. The state of health (SOH) of the energy storage is incorporated as an ageing coefficient, which causes the BESS to behave conservatively in order to retain its lifespan. Using a time of use (TOU) tariff, simulation results suggest a substantial possibility to boost the savings of consumers in a grid-connected micro-grid. The simulation findings indicate that by efficiently scheduling the BESS power management technique, the proposed method improves a number of distribution system efficacy.

Autophagy is a conserved metabolic pathway that is central to many diseases. Recently, there has been a lot of interest in targeting autophagy with small molecule inhibitors as a possible therapeutic strategy. However, many of the compounds used for autophagy are non-selective. Here, we explored the inhibition of autophagy in pancreatic cancer cells using established selective small molecule inhibitors and discovered an unexpected link between the autophagy pathway and progression through the cell cycle. Our findings revealed that treatments with inhibitors that have different autophagy pathway targets block cell replication and activate other metabolic pathways to compensate for the blockade in autophagy. An unbiased screen looking for known drugs that might synergize with autophagy inhibition revealed new combination treatments that might provide a blueprint for therapeutic approaches to pancreatic cancer. The drugs quizartinib and THZ1 showed a strong synergistic effect in pancreatic cells with autophagy inhibition.

Wild-type P53-induced phosphatase 1 (WIP1), also known as

Alphascreen monitoring of the Six1/Eya2 interaction demonstrates that compound 9987 has no effect on this interaction.

Compound inhibition of different Eya proteins and Eya mutants.

Eya2 and Six1 expression in different lung cancer cell lines, along with the effects of Eya2 and Eya2 D274N on protein localization and EMT markers.

Eya2 binding to Six1 is critical for its ability to mediate numerous phenotypes in Calu-6 lung cancer cells.

Validation of newly generated Six1 antibody (Ford 1229).

Cellular activity of 9987 in HCC4006 cells. HCC4006 cells were treated with 9987 at 5 or 10 mM and the efficacy of 9987 was tested on overall growth, colony formation, motility, and invasion.

Crystal structure of Eya2 ED-9987 complex shows three Eya2 ED-9987 complexes in one asymmetric unit, with the 9987-binding pocket in the Eya2-9987 crystal structure overlaps with the binding pocket predicted from mutagenesis studies.

The perinucleolar compartment (PNC) is a dynamic subnuclear body found at the periphery of the nucleolus. The PNC is enriched with RNA transcripts and RNA-binding proteins, reflecting different states of genome organization. PNC prevalence positively correlates with cancer progression and metastatic capacity, making it a useful marker for metastatic cancer progression. A high-throughput, high-content assay was developed to identify novel small molecules that selectively reduce PNC prevalence in cancer cells. We identified and further optimized a pyrrolopyrimidine series able to reduce PNC prevalence in PC3M cancer cells at submicromolar concentrations without affecting cell viability. SAR exploration of the structural elements necessary for activity resulted in the discovery of several potent compounds. Analysis of in vitro drug-like properties led to the discovery of the bioavailable analogue, metarrestin, which has shown potent antimetastatic activity with improved survival in rodent models and is currently being evaluated in a first-in-human Phase 1 clinical trial.

Figure legends for all supplemental figures (1-6) describe key findings and experimental details.

Detailed information (source and working dilution) of antibodies have been used for WB, ICC and IHC.

Detailed description of methods used for compound 8430 synthesis, cell culture and compound 8430 pharmacokinetics.

Supplemental figures (1-6) demonstrate the efficacy of parental SIX1/EYA2 inhibitor, synthesis of compound 8430, endogenous SIX1 and EYA2 expression in tumor lines, as well as 8430's transcriptional impact, pharmacokinetic/toxicity and effect on metastasis.

Metastasis is the major cause of mortality for patients with cancer, and dysregulation of developmental signaling pathways can significantly contribute to the metastatic process. The Sine oculis homeobox homolog 1 (SIX1)/eyes absent (EYA) transcriptional complex plays a critical role in the development of multiple organs and is typically downregulated after development is complete. In breast cancer, aberrant expression of SIX1 has been demonstrated to stimulate metastasis through activation of TGFβ signaling and subsequent induction of epithelial–mesenchymal transition (EMT). In addition, SIX1 can induce metastasis via non-cell autonomous means, including activation of GLI-signaling in neighboring tumor cells and activation of VEGFC–induced lymphangiogenesis. Thus, targeting SIX1 would be expected to inhibit metastasis while conferring limited side effects. However, transcription factors are notoriously difficult to target, and thus novel approaches to inhibit their action must be taken. Here we identified a novel small molecule compound, NCGC00378430 (abbreviated as 8430), that reduces the SIX1/EYA2 interaction. 8430 partially reversed transcriptional and metabolic profiles mediated by SIX1 overexpression and reversed SIX1-induced TGFβ signaling and EMT. 8430 was well tolerated when delivered to mice and significantly suppressed breast cancer–associated metastasis in vivo without significantly altering primary tumor growth. Thus, we have demonstrated for the first time that pharmacologic inhibition of the SIX1/EYA2 complex and associated phenotypes is sufficient to suppress breast cancer metastasis.Significance:These findings identify and characterize a novel inhibitor of the SIX1/EYA2 complex that reverses EMT phenotypes suppressing breast cancer metastasis.

It is extremely difficult to achieve functional recovery after axonal injury in the adult central nervous system. The activation of G-protein coupled receptor 110 (GPR110, ADGRF1) has been shown to stimulate neurite extension in developing neurons and after axonal injury in adult mice. Here, we demonstrate that GPR110 activation partially restores visual function impaired by optic nerve injury in adult mice. Intravitreal injection of GPR110 ligands, synaptamide and its stable analogue dimethylsynaptamide (A8) after optic nerve crush significantly reduced axonal degeneration and improved axonal integrity and visual function in wild-type but not gpr110 knockout mice. The retina obtained from the injured mice treated with GPR110 ligands also showed a significant reduction in the crush-induced loss of retinal ganglion cells. Our data suggest that targeting GPR110 may be a viable strategy for functional recovery after optic nerve injury.

A diverse range of biochemical and cellular assays are used by medicinal chemists to guide compound optimization. The data collected from these assays influence decisions taken on structure-activity relationship (SAR) campaigns. Therefore, it is paramount that medicinal chemists have a solid understanding of the strengths and limitations of each assay being used to characterize synthesized analogs. For the successful execution of a medicinal chemistry campaign, it is our contention that an early partnership among assay biologists, informaticians, and medicinal chemists must exist. Their combined skill sets are necessary to not only design and develop robust assays but also implement an effective screening cascade in which multiple orthogonal and counter assays are selected to validate the activity and target(s) of the synthesized compounds. We review multiple cases of drug and chemical probe discovery from collaborative National Center for Advancing Translational Sciences/National Institutes of Health projects and published scientific literature in which the evaluation of compounds in secondary or orthogonal assays led to the discovery of unexpected activities, forcing a reconsideration of the original assay design that was used to discover the biological activity of the compound. Using these retrospective case studies, the goal of this Perspective is to hedge toward the development of physiologically relevant assays that are able to capture the true bioactivity of compounds being developed in a medicinal chemistry campaign.

Neomorphic mutations in isocitrate dehydrogenase 1 (IDH1) are oncogenic for a number of malignancies, primarily low-grade gliomas and acute myeloid leukemia (AML). We report a medicinal chemistry campaign around a 7,7-dimethyl-7,8-dihydro-2H-1λ(2)-quinoline-2,5(6H)-dione screening hit against the R132H and R132C mutant forms of isocitrate dehydrogenase (IDH1). Systematic SAR efforts produced a series of potent pyrid-2-one mIDH1 inhibitors, including the atropisomer (+)-119 (NCATS-SM5637, NSC 791985). In an engineered mIDH1-U87-xenograft mouse model; after a single oral dose of 30 mg/kg, 16 h post dose, between 16-48 h, (+)-119 showed higher tumoral concentrations that corresponded to lower 2-HG concentrations, when compared with the approved drug AG-120 (ivosidenib).

Fumarate hydratase (FH) is a metabolic enzyme that is part of the Krebs-cycle, and reversibly catalyzes the hydration of fumarate to malate. Mutations of the FH gene have been associated with fumarate hydratase deficiency (FHD), hereditary leiomyomatosis, renal cell cancer (HLRCC), and other diseases. Currently there are no high-quality small molecule probes for studying human fumarate hydratase. To address this, we developed a quantitative high throughput screening (qHTS) FH assay and screened a total of 57,037 compounds from in-house libraries in dose-response. While no inhibitors of FH were confirmed, a series of phenyl-pyrrolo-pyrimidine-diones were identified as activators of human fumarate hydratase. These compounds were not substrates of fumarate hydratase, were inactive in a malate dehydrogenase counter screen, and showed no detectable reduction–oxidation activity. The binding of two compounds from the series to human fumarate hydratase was confirmed by microscale thermophoresis. The low hit rate in this screening campaign confirmed that FH is a ‘tough target’ to modulate, and the small molecule activators of human fumarate hydratase reported here may serve as a starting point for further optimization and development into cellular probes of human FH and potential drug candidates.

Nontypeable Haemophilus influenzae (NTHi) are clinically important Gram-negative bacteria that are responsible for various human mucosal diseases, including otitis media (OM). Recurrent OM caused by NTHi is common and infections which recur less than two weeks following antimicrobial therapy are largely attributable to recurrence of the same strain of bacteria. Toxin-antitoxin (TA) modules encoded by bacteria enable rapid responses to environmental stresses and are thought to facilitate growth arrest, persistence, and tolerance to antibiotics. The vapBC-1 locus of NTHi encodes a type II TA system, comprising the ribonuclease toxin VapC1 and its cognate antitoxin VapB1. The activity of VapC1 has been linked to the survival of NTHi during antibiotic treatment both in vivo and ex vivo. Therefore, inhibitors of VapC1 might serve as adjuvants to antibiotics, preventing NTHi from entering growth arrest and surviving; however, none have been reported to date. A truncated VapB1 peptide from a crystal structure of the VapBC-1 complex was used to generate pharmacophore queries to facilitate a scaffold hopping approach for the identification of small molecule VapC1 inhibitors. The National Center for Advancing Translational Sciences small molecule library was virtually screened using the shape-based method Rapid Overlay of Chemical Structures (ROCS) and the top-ranking hits were docked into the VapB1 binding pocket of VapC1. Two hundred virtual screening hits with the best docking scores were selected and tested in a biochemical VapC1 activity assay, which confirmed eight compounds as VapC1 inhibitors. An additional sixty compounds were selected with structural similarities to the confirmed VapC1 inhibitors, of which twenty inhibited VapC1 activity. Intracellular target engagement of five inhibitors was indicated by the destabilization of VapC1 within bacterial cells from a cellular thermal shift assay; however, no impact on bacterial growth was observed. Thus, this virtual screening and scaffold hopping approach enabled the discovery of VapC1 ribonuclease inhibitors that might serve as starting points for preclinical development.

Numerous studies have established the involvement of lysosomal and mitochondrial dysfunction in the pathogenesis of neurodegenerative disorders such as Alzheimer's and Parkinson diseases. Building on our previous studies of the neurodegenerative lysosomal lipidosis Niemann-Pick C1 (NPC1), we have unexpectedly discovered that activation of the mitochondrial chaperone tumor necrosis factor receptor-associated protein 1 (TRAP1) leads to the correction of the lysosomal storage phenotype in patient cells from multiple lysosomal storage disorders including NPC1. Using small compound activators specific for TRAP1, we find that activation of this chaperone leads to a generalized restoration of lysosomal and mitochondrial health. Mechanistically, we show that this process includes inhibition of oxidative phosphorylation and reduction of oxidative stress, which results in activation of AMPK and ultimately stimulates lysosome recycling. Thus, TRAP1 participates in lysosomal-mitochondrial crosstalk to maintain cellular homeostasis and could represent a potential therapeutic target for multiple disorders.

Classic galactosemia is a rare disease caused by inherited deficiency of galactose-1 phosphate uridylyltransferase (GALT). Accumulation of galactose-1 phosphate (gal-1P) is thought to be the major cause of the chronic complications associated with this disease, which currently has no treatment. Inhibiting galactokinase (GALK1), the enzyme that generates galactose-1 phosphate, has been proposed as a novel strategy for treating classic galactosemia. Our previous work identified a highly selective unique dihydropyrimidine inhibitor against GALK1. With the determination of a co-crystal structure of this inhibitor with human GALK1, we initiated a structure-based structure–activity relationship (SAR) optimization campaign that yielded novel analogs with potent biochemical inhibition (IC(50) < 100 nM). Lead compounds were also able to prevent gal-1P accumulation in patient-derived cells at low micromolar concentrations and have pharmacokinetic properties suitable for evaluation in rodent models of galactosemia.

Dystonias are a group of chronic movement-disabling disorders for which highly effective oral medications or disease-modifying therapies are lacking. The most effective treatments require invasive procedures such as deep brain stimulation (DBS). In this study, we used a high-throughput assay based on a monogenic form of dystonia, DYT1 (DYT-TOR1A), to screen a library of compounds approved for use in humans, the NCATS Pharmaceutical Collection (NPC; 2816 compounds) and identify drugs able to correct mislocalization of the disease-causing protein variant, ΔE302/3 hTorsinA. The HIV protease inhibitor, ritonavir, was among 18 compounds found to normalize hTorsinA mislocalization. Using a DYT1 knock-in mouse model to test efficacy on brain pathologies, we found that ritonavir restored multiple brain abnormalities across development. Ritonavir acutely corrected striatal cholinergic interneuron physiology in the mature brain and yielded sustained correction of diffusion tensor magnetic resonance imaging (MRI) signals when delivered during a discrete early developmental window. Mechanistically, we found that across the family of HIV protease inhibitors, efficacy correlated with integrated stress response (ISR) activation. These preclinical results identify ritonavir as a drug candidate for dystonia with disease-modifying potential.

Nonstructural protein 1 (NS1) plays a crucial function in the replication, spread, and pathogenesis of influenza virus by inhibiting the host innate immune response. Here we report the discovery and optimization of novel pyrazolopyridine NS1 antagonists that can potently inhibit influenza A/PR/8/34 replication in MDCK cells, rescue MDCK cells from cytopathic effects of seasonal influenza A strains, reverse NS1-dependent inhibition of IFN-β gene expression, and suppress the slow growth phenotype in NS1-expressing yeast. These pyrazolopyridines will enable researchers to investigate NS1 function during infection and how antagonists can be utilized in the next generation of treatments for influenza infection.