Diselenide Covalent Allosteric Inhibitors of Glutaminase with Strong Anticancer Activity.

Allosteric glutaminase inhibitors demonstrate inhibition of glutamine-dependent cancer cells with low general drug toxicity, but have issues with efficacy . Here, we designed a series of diselenide compounds with 6 atoms in the middle, aiming to target the allosteric site of kidney type glutaminase (KGA) with a covalent linkage to strengthen the interaction. Proteomic analysis demonstrated that the diselenide compounds cross-linked with the Lys320 residue at the KGA allosteric site; this was confirmed by the KGA K320A mutant which showed essentially no binding to the diselenide.

Synthesis of Novel Kidney-Type Glutaminase Allosteric Inhibitors Targeting the Critical Lys-320 Residue.

Reversible allosteric inhibitors of kidney-type glutaminase (GLS1, KGA) showed incomplete inhibition of cancer cell proliferation and poor efficacy. Here, we investigate some irreversible inhibitors targeting the critical K320 residue responsible for GLS1 biological activity. The (trifluoromethoxy)phenylacetic acid motif was replaced by α,β-unsaturated carboxylic acids, and the resulting terminally substituted derivatives (e.

Rapid High-Throughput Assay Identified Gemcitabine and Derivatives As Potent Inhibitors Against Multidrug-Resistant .

Staphylococcus aureus (MRSA) are challenging pathogenic bacteria that can cause severe infection leading to high mortality rates. We found that both the oxacillin- and cefoxitin-resistant strains isolated from clinic showed multidrug-resistant (MDR) characteristics. Through rapid high-throughput screen (HTS) of a compound library, gemcitabine and selen compounds were found to effectively inhibit growth.

Self-Assembled Micellar Glutaminase Allosteric Inhibitor for Effective Therapeutic Intervention.

Introduction: Kidney-type glutaminase (KGA) has been an important anti-tumor drug target, and KGA allosteric inhibitors attracted much interest for their superior enzymatic specificity with good drug safety profiles. For glutaminase allosteric inhibitors such as BPTES, CB-839 and Selen derivatives, the low solubility remains as the main factor that limits in vivo efficacy. The 1,3,4-Selenadiazole compound CPD 23 showed improved in vivo efficacy but worse solubility; however, the graft polymer polyvinyl caprolactam-polyvinyl acetate-polyethylene glycol (PVCap-PVA-PEG), Soluplus (SOL) stood out as an excellent delivery carrier for CPD 23.

Biodegradable self-assembly micelles significantly enhanced the solubility, biological stability and antitumor efficacy of Hexylselen.

Glutaminolysis inhibitors have shown early promise in cancer therapeutics. Specifically, kidney-type glutaminase (KGA) has been a long-standing anti-tumor drug target; KGA allosteric inhibitors have attracted great attention due to their superior enzyme specificity and good drug safety profiles. However, the main issue with allosteric inhibitors-including BPTES, CB-839, and the recently developed KGA allosteric and glutamate dehydrogenase (GDH) dual inhibitor, Hexylselen (CPD-3B)-is their low solubility; it leads to limited efficacy.

Glutamate Dehydrogenase as a Promising Target for Hyperinsulinism Hyperammonemia Syndrome Therapy.

Hyperinsulinism-hyperammonemia syndrome (HHS) is a rare disease characterized by recurrent hypoglycemia and persistent elevation of plasma ammonia, and it can lead to severe epilepsy and permanent brain damage. It has been demonstrated that functional mutations of glutamate dehydrogenase (GDH), an enzyme in the mitochondrial matrix, are responsible for the HHS. Thus, GDH has become a promising target for the small molecule therapeutic intervention of HHS.

A sensitive EZMTT method provides microscale, quantitative and high-throughput evaluation of drug efficacy in the treatment of Mycobacterium tuberculosis infectious diseases.

Drug resistance has become a serious public health problem in mycobacterial infectious diseases. Here, we investigated a water soluble tetrazolium salt (EZMTT)-based detection method to provide an easy, safe and quantitative antimycobacterial susceptibility test (AMST) method, especially for targeting early detection of loss of drug susceptibility in mycobacteria. After a single addition of the EZMTT detection reagent at the inoculation of mycobacteria culture, the AMST was continuously analyzed in a sealed 96-well plate (100 μl), or a sealed tube to ensure biosafety.

Discovery of an Inhibitor for Bacterial 3-Mercaptopyruvate Sulfurtransferase that Synergistically Controls Bacterial Survival.

HS-producing enzymes in bacteria have been shown to be closely engaged in the process of microbial survival and antibiotic resistance. However, no inhibitors have been discovered for these enzymes, e.g.

The EZMTT cell proliferation assay provides precise measurement for drug combinations and better correlation between in vitro and in vivo efficacy.

The rate of drug-induced proliferation (DIP) has been proposed as an unbiased alternative drug effect metric. However, current assays are not easy and precise enough to track minor changes in cell growth. Here, we report the optimized EZMTT based detection method which can continuously measure time-dependent growth after drug treatment and reliably detect partial drug resistance for cancer cells.

Design and synthesis of biotinylated Hexylselen as a probe to identify KGA allosteric inhibitors by a convenient biomolecular interaction assay.

Hexylselen is a novel submicromolar dual KGA/GDH inhibitor, which demonstrates potent inhibition of cancer cells with minimal toxicity. To further investigation its mechanism of action, we designed and synthesized its biotinylated derivative 2 as a novel probe. From commercially available starting material, 2 was obtained in 6 steps with 13.

Rapamycin directly activates lysosomal mucolipin TRP channels independent of mTOR.

Rapamycin (Rap) and its derivatives, called rapalogs, are being explored in clinical trials targeting cancer and neurodegeneration. The underlying mechanisms of Rap actions, however, are not well understood. Mechanistic target of rapamycin (mTOR), a lysosome-localized protein kinase that acts as a critical regulator of cellular growth, is believed to mediate most Rap actions.

Design and synthesis of novel tellurodibenzoic acid compounds as kidney-type glutaminase (KGA) inhibitors.

Organotellurium compounds have been reported as an immune-modulator sensitizing chemotherapeutics. Herein, we report the design and synthesis of a series of novel tellurodibenzoic acids as mimics of diphenylarsenic acid (DPAA) and potential selective KGA inhibitors. Representative compound 3B exhibited potent inhibition of KGA and glutamine-dependent HCT-116 cells.

Kidney-Type Glutaminase Inhibitor Hexylselen Selectively Kills Cancer Cells via a Three-Pronged Mechanism.

Tumor metabolism has been deeply investigated for cancer therapeutics. Here, we demonstrate that glutamine deficiency alone could not completely inhibit cancer cell growth and that many potent kidney-type glutaminase (KGA) inhibitors did not show satisfying efficacy. The potent KGA allosteric inhibitor, CB-839, resulted in up to 80% growth inhibition of all tested cell lines, whereas Hexylselen (CPD-3B), a KGA/glutamate dehydrogenase (GDH) inhibitor, showed essentially no toxicity to normal cells up to a 10 μM concentration and could completely inhibit the growth of many aggressive cell lines.

Propylselen inhibits cancer cell growth by targeting glutamate dehydrogenase at the NADP binding site.

High levels of glutamate dehydrogenase (GDH) activity are associated with hypoglycemia, cancer, and Parkinson's disease. Propylselen was synthesized to investigate its mechanism of GDH inhibition in comparison with Ebselen and Epigallocatechin gallate (EGCG). Because Ebselen was found to crosslink with the peptide (AA299-341) at the active site of E.

Novel 1,3,4-Selenadiazole-Containing Kidney-Type Glutaminase Inhibitors Showed Improved Cellular Uptake and Antitumor Activity.

Kidney-type glutaminase [KGA/isoenzyme glutaminase C (GAC)] is becoming an important tumor metabolism target in cancer chemotherapy. Its allosteric inhibitor, CB839, showed early promise in cancer therapeutics but limited efficacy in in vivo cancer models. To improve the in vivo activity, we explored a bioisostere replacement of the sulfur atom in bis-2-(5-phenylacetamido-1,2,4-thiadiazol)ethyl sulfide and CB839 analogues with selenium using a novel synthesis of the selenadiazole moiety from carboxylic acids or nitriles.

FITC-labeled d-glucose analog is suitable as a probe for detecting insulin-dependent glucose uptake.

The detection of the insulin-dependent glucose uptake is a vital part in the research of diabetes. To establish a sensitive assay for measuring glucose uptake in living cells, we synthesized a FITC linked d-glucosamine 2 as a probe. 2 was obtained by the reaction of commercially available d-glucosamine hydrochloride and FITC and was determined as a single anomeric form by H NMR and C NMR.

Ebselen Reversibly Inhibits Human Glutamate Dehydrogenase at the Catalytic Site.

Human glutamate dehydrogenase (GDH) plays an important role in neurological diseases, tumor metabolism, and hyperinsulinism-hyperammonemia syndrome (HHS). However, there are very few inhibitors known for human GDH. Recently, Ebselen was reported to crosslink with Escherichia coli GDH at the active site cysteine residue (Cys321), but the sequence alignment showed that the corresponding residue is Ala329 in human GDH.

Identification of a diverse synthetic abietane diterpenoid library and insight into the structure-activity relationships for antibacterial activity.

A diverse natural product-like (NPL) synthetic abietane diterpenoid library containing 86 compounds were obtained and the SARs were studied based on their antibacterial potential. Further in vitro cytotoxic and in silico drug-like properties evaluation showed that the potent antibacterial compound 84 had good drug-like properties and displayed low cytotoxicity toward noncancerous mammalian cells, indicating the study of AA and DHAA might be a good starting point for the search of novel antimicrobial molecules. Future work should be focused on the optimization of their potency and selectivity.

Ebselen: Mechanisms of Glutamate Dehydrogenase and Glutaminase Enzyme Inhibition.

Ebselen modulates target proteins through redox reactions with selenocysteine/cysteine residues, or through binding to the zinc finger domains. However, a recent contradiction in ebselen inhibition of kidney type glutaminase (KGA) stimulated our interest in investigating its inhibition mechanism with glutamate dehydrogenase (GDH), KGA, thioredoxin reductase (TrxR), and glutathione S-transferase. Fluorescein- or biotin-labeled ebselen derivatives were synthesized for mechanistic analyses.

Click chemistry-based synthesis and anticancer activity evaluation of novel C-14 1,2,3-triazole dehydroabietic acid hybrids.

A concise and efficient synthetic approach has been established to readily access a series of novel C-14 1,2,3-triazole-tethered dehydroabietic acid derivatives in moderate to high yields. In vitro antiproliferative activity evaluation indicated that most of the hybrids exhibited potent inhibitory activities in a variety of cancer cell lines with low micromolar to submicromolar IC values. Further studies demonstrated that some of these analogues such as 20, 21, and 24 were also effective against adriamycin-resistant MCF-7 clone at low concentrations in a dose-dependent manner.

Cationic Polymethacrylate-Modified Liposomes Significantly Enhanced Doxorubicin Delivery and Antitumor Activity.

Liposome (LP) encapsulation of doxorubicin (DOX) is a clinically validated method for cancer drug delivery, but its cellular uptake is actually lower than the free DOX. Therefore, we modified DOX-LP with a cationic polymer (Eudragit RL100; ER) to improve its cellular uptake and antitumor activity. The resulting DOX-ERLP was a 190 nm nanoparticle that was absorbed efficiently and caused cancer cell death in 5 hrs.

Biomolecular Interaction Assays Identified Dual Inhibitors of Glutaminase and Glutamate Dehydrogenase That Disrupt Mitochondrial Function and Prevent Growth of Cancer Cells.

Glutaminase (KGA/isoenzyme GAC) is an emerging and important drug target for cancer. Traditional methods for assaying glutaminase activity are coupled with several other enzymes. Such coupled assays do not permit the direct and stringent characterization of specific glutaminase inhibitors.

Mono-sulfonated tetrazolium salt based NAD(P)H detection reagents suitable for dehydrogenase and real-time cell viability assays.

Glutamate dehydrogenase (GDH) catalyzes the oxidative deamination of L-glutamate and is important for several biological processes. For GDH inhibitor screening, we developed a novel mono-sulfonated tetrazolium salt (EZMTT), which can be synthesized using H2O2 oxidation and purified easily on silica gel in large quantities. The EZMTT detection method showed linear dose responses to NAD(P)H, dehydrogenase concentration and cell numbers.

Biomolecular interaction analysis for carbon nanotubes and for biocompatibility prediction.

The interactions between carbon nanotubes (CNTs) and biologics have been commonly studied by various microscopy and spectroscopy methods. We tried biomolecular interaction analysis to measure the kinetic interactions between proteins and CNTs. The analysis demonstrated that wheat germ agglutinin (WGA) and other proteins have high affinity toward carboxylated CNT (f-MWCNT) but essentially no binding to normal CNT (p-MWCNT).