Self-Assembled Nanoparticles from Xie-Bai-San Decoction: Isolation, Characterization and Enhancing Oral Bioavailability.

Background: Natural nanoparticles have been found to exist in traditional Chinese medicine (TCM) decoctions. However, whether natural nanoparticles can influence the oral bioavailability of active compounds has not been elucidated. Using Xie-Bai-San decoction (XBSD) as an example, the purpose of this study was to isolate, characterize and elucidate the mechanism of the nanoparticles (N-XBSD) in XBSD, and further to explore whether the bioavailability of the main active compounds could be enhanced by N-XBSD. Methods: N-XBSD were isolated from XBSD, and investigated its characterization and study of its formation mechanism, and evaluation of its ability to enhance bioavailability of active compounds. Results: The N-XBSD was successfully isolated with the average particle size of 104.53 nm, PDI of 0.27 and zeta potential of -5.14 mV. Meanwhile, all the eight active compounds were most presented in N-XBSD. Kukoamine B could self-assemble with mulberroside A or liquiritin to form nanoparticles, respectively. And the FT-IR and HRMS results indicated the possible binding of the ammonium group of kukoamine B with the phenolic hydroxyl group of mulberroside A or liquiritin, respectively. The established UPLC-MS/MS method was accurate and reliable and met the quantitative requirements. The pharmacokinetic behaviors of the N-XBSD and decoction were similar in rats. Most notably, compared to that of free drugs, the Cmax, AUC0-∞, AUC0-t, T1/2 and MRT0-∞ values of index compounds were the higher in N-XBSD, with a slower plasma clearance rate in rats. Conclusion: The major active compounds of XBSD were mainly distributed in N-XBSD, and N-XBSD was formed through self-assembly among active compounds. N-XBSD could obviously promote the bioavailability of active compounds, indicating natural nanoparticles of decoctions play an important role in therapeutic effects.

Identification of Kukoamine A, Zeaxanthin, and Clexane as New Furin Inhibitors.

The endogenous protease furin is a key protein in many different diseases, such as cancer and infections. For this reason, a wide range of studies has focused on targeting furin from a therapeutic point of view. Our main objective consisted of identifying new compounds that could enlarge the furin inhibitor arsenal; secondarily, we assayed their adjuvant effect in combination with a known furin inhibitor, CMK, which avoids the SARS-CoV-2 S protein cleavage by means of that inhibition. Virtual screening was carried out to identify potential furin inhibitors. The inhibition of physiological and purified recombinant furin by screening selected compounds, Clexane, and these drugs in combination with CMK was assayed in fluorogenic tests by using a specific furin substrate. The effects of the selected inhibitors from virtual screening on cell viability (293T HEK cell line) were assayed by means of flow cytometry. Through virtual screening, Zeaxanthin and Kukoamine A were selected as the main potential furin inhibitors. In fluorogenic assays, these two compounds and Clexane inhibited both physiological and recombinant furin in a dose-dependent way. In addition, these compounds increased physiological furin inhibition by CMK, showing an adjuvant effect. In conclusion, we identified Kukoamine A, Zeaxanthin, and Clexane as new furin inhibitors. In addition, these drugs were able to increase furin inhibition by CMK, so they could also increase its efficiency when avoiding S protein proteolysis, which is essential for SARS-CoV-2 cell infection.

Kukoamine A Protects against NMDA-Induced Neurotoxicity Accompanied with Down-Regulation of GluN2B-Containing NMDA Receptors and Phosphorylation of PI3K/Akt/GSK-3ß Signaling Pathway in Cultured Primary Cortical Neurons.

Kukoamine (KuA) is a spermine alkaloid present in traditional Chinese medicine Cortex Lycii radices, which possesses various pharmacological properties. Our previous studies have demonstrated that KuA exerts neuroprotective effects against H2O2-induced oxidative stress, radiation-induced neuroinflammation, oxidative stress and neuronal apoptosis, as well as neurotoxin-induced Parkinson's disease through apoptosis inhibition and autophagy enhancement. The present study aimed to investigate the neuroprotective effects of KuA against NMDA-induced neuronal injury in cultured primary cortical neurons and explore the underlying mechanism. Incubation with 200 µM NMDA for 30 min induced excitotoxicity in primary cultured cortical neurons. The results demonstrated that pretreatment with KuA attenuated NMDA induced cell injury, LDH leakage and neuronal apoptosis. KuA also regulated apoptosis-related proteins. Thus, incubation with the alkaloid decreased the ratio of Bax/Bcl-2, and inhibited the release of cytochrome C, the expression of p53 and the cleavage of caspase-3. Moreover, KuA prevented the upregulation of GluN2B-containing NMDA receptors (NMDAR). Additionally, pretreatment with KuA reversed NMDA-induced dephosphorylation of Akt and GSK-3ß and the protective effect of KuA on NMDA-induced cytotoxicity was abolished by wortmannin, a PI3K inhibitor. Taken together, these results indicated that KuA exerted neuroprotective effects against NMDA-induced neurotoxicity in cultural primary cortical neurons and caused the down-regulation of GluN2B-containing NMDARs as well as the phosphorylation of proteins belonging to the PI3K/Akt/GSK-3ß signaling pathway.

Separation and Characterization of Phenolamines and Flavonoids from Rape Bee Pollen, and Comparison of Their Antioxidant Activities and Protective Effects Against Oxidative Stress.

Phenolamines and flavonoids are two important components in bee pollen. There are many reports on the bioactivity of flavonoids in bee pollen, but few on phenolamines. This study aims to separate and characterize the flavonoids and phenolamines from rape bee pollen, and compare their antioxidant activities and protective effects against oxidative stress. The rape bee pollen was separated to obtain 35% and 50% fractions, which were characterized by HPLC-ESI-QTOF-MS/MS. The results showed that the compounds in 35% fraction were quercetin and kaempferol glycosides, while the compounds in 50% fraction were phenolamines, including di-p-coumaroyl spermidine, p-coumaroyl caffeoyl hydroxyferuloyl spermine, di-p-coumaroyl hydroxyferuloyl spermine, and tri-p-coumaroyl spermidine. The antioxidant activities of phenolamines and flavonoids were evaluated by 2,2-diphenyl-1-picrylhydrazyl (DPPH), 2,2'-azino-bis-3-ethylbenzothiazoline-6-sulphonic acid (ABTS), and ferric reducing antioxidant power (FRAP) assays. It was found that the antioxidant activity of phenolamines was significantly higher than that of flavonoids. Moreover, phenolamines showed better protective effects than flavonoids on HepG2 cells injured by AAPH. Furthermore, phenolamines could significantly reduce the reactive oxygen species (ROS), alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels, and increase the superoxide dismutase (SOD) and glutathione (GSH) levels. This study lays a foundation for the further understanding of phenolamines in rape bee pollen.

Inhibitory activities of kukoamines A and B from Lycii Cortex on amyloid aggregation related to Alzheimer's disease and type 2 diabetes.

Alzheimer's diseases (AD) and type 2 diabetes (T2D) are two age-related diseases characterized by amyloid fibrillogenesis. Prevention of amyloid aggregation is a promising therapeutic strategy for AD and T2D. Two spermine alkaloids, kukoamines A and B, isolated from Lycii Cortex (LyC) were investigated for their inhibitory effect on amyloid aggregation. Both kukoamines A and B inhibited aggregation of amyloid ß (Aß) and human islet amyloid polypeptide (hIAPP) in a dose-dependent manner. Kukoamine B showed stronger inhibitory activity than kukoamine A. These results on the inhibitory activity of kukoamines A and B on Aß and hIAPP indicate that the catechol moiety is essential for inhibition of amyloid aggregation.

Intranasal Targeting of Hypothalamic PTP1B and TCPTP Reinstates Leptin and Insulin Sensitivity and Promotes Weight Loss in Obesity.

The importance of hypothalamic leptin and insulin resistance in the development and maintenance of obesity remains unclear. The tyrosine phosphatases protein tyrosine phosphatase 1B (PTP1B) and T cell protein tyrosine phosphatase (TCPTP) attenuate leptin and insulin signaling and are elevated in the hypothalami of obese mice. We report that elevated PTP1B and TCPTP antagonize hypothalamic leptin and insulin signaling and contribute to the maintenance of obesity. Deletion of PTP1B and TCPTP in the hypothalami of obese mice enhances CNS leptin and insulin sensitivity, represses feeding, and increases browning, to decrease adiposity and improve glucose metabolism. The daily intranasal administration of a PTP1B inhibitor, plus the glucocorticoid antagonist RU486 that decreases TCPTP expression, represses feeding, increases browning, promotes weight loss, and improves glucose metabolism in obese mice. Our findings causally link heightened hypothalamic PTP1B and TCPTP with leptin and insulin resistance and the maintenance of obesity and define a viable pharmacological approach by which to promote weight loss in obesity.

Anti-Osteoporotic Effects of Kukoamine B Isolated from Lycii Radicis Cortex Extract on Osteoblast and Osteoclast Cells and Ovariectomized Osteoporosis Model Mice.

Osteoporosis is an abnormal bone remodeling condition characterized by decreased bone density, which leads to high risks of fracture. Previous study has demonstrated that Lycii Radicis Cortex (LRC) extract inhibits bone loss in ovariectomized (OVX) mice by enhancing osteoblast differentiation. A bioactive compound, kukoamine B (KB), was identified from fractionation of an LRC extract as a candidate component responsible for an anti-osteoporotic effect. This study investigated the anti-osteoporotic effects of KB using in vitro and in vivo osteoporosis models. KB treatment significantly increased the osteoblastic differentiation and mineralized nodule formation of osteoblastic MC3T3-E1 cells, while it significantly decreased the osteoclast differentiation of primary-cultured monocytes derived from mouse bone marrow. The effects of KB on osteoblastic and osteoclastic differentiations under more physiological conditions were also examined. In the co-culture of MC3T3-E1 cells and monocytes, KB promoted osteoblast differentiation but did not affect osteoclast differentiation. In vivo experiments revealed that KB significantly inhibited OVX-induced bone mineral density loss and restored the impaired bone structural properties in osteoporosis model mice. These results suggest that KB may be a potential therapeutic candidate for the treatment of osteoporosis.

Comparative Study of the Chemical Constituents and Bioactivities of the Extracts from Fruits, Leaves and Root Barks of Lycium barbarum.

The fruits, leaves and root barks of L. barbarum plant are widely used as functional foods and as ingredients in traditional Chinese prescriptions and patent medicines. They are considered to have different pharmacological activities and health benefits because of their diverse constituents. Here, the chemical constituents of the extracts from fruits, leaves and root barks of L. barbarum were compared by ultra-high performance liquid chromatography coupled with high resolution mass spectrometry (UPLC-HR-MS). A total of 131 compounds were identified and seven of them were quantified. Among them, 98, 28 and 35 constituents were detected in fruits, leaves and root barks respectively. Dicaffeoylspermidine/spermine derivatives were the most detected compounds (74/131); among them, dicaffeoylspermine isomers and propionyl-dicaffeoylspermidine were found in root barks in very large amounts (e.g., kukoamine B = 10.90 mg/g dry powder); dicaffeoyl-spermidine isomers were detected in fruits/leaves in a high amount, and many of their glycosylated derivatives were mainly detected in fruits. In addition, six saponins from L. barbarum fruits were reported for the first time, and 5,6-dihydrosolasonine was reported for the first time in plants. The activity assays showed that the root bark extract possessed the strongest antioxidative activity and cytotoxicity, which was presumed due to the large amount of dicaffeoylspermine/spermidines in root barks. Fourteen potential bioactive components from fruits were identified by a target cell-based screening method. These results will help to understand the different biological activities of these three parts of L. barbarum plant and will benefit the discovery of new functional components.

Trodusquemine enhances Aß42 aggregation but suppresses its toxicity by displacing oligomers from cell membranes.

Transient oligomeric species formed during the aggregation process of the 42-residue form of the amyloid-ß peptide (Aß42) are key pathogenic agents in Alzheimer's disease (AD). To investigate the relationship between Aß42 aggregation and its cytotoxicity and the influence of a potential drug on both phenomena, we have studied the effects of trodusquemine. This aminosterol enhances the rate of aggregation by promoting monomer-dependent secondary nucleation, but significantly reduces the toxicity of the resulting oligomers to neuroblastoma cells by inhibiting their binding to the cellular membranes. When administered to a C. elegans model of AD, we again observe an increase in aggregate formation alongside the suppression of Aß42-induced toxicity. In addition to oligomer displacement, the reduced toxicity could also point towards an increased rate of conversion of oligomers to less toxic fibrils. The ability of a small molecule to reduce the toxicity of oligomeric species represents a potential therapeutic strategy against AD.

Curcumin mediates polyamine metabolism and sensitizes gastrointestinal cancer cells to antitumor polyamine-targeted therapies.

Curcumin, a natural polyphenol that contributes to the flavor and yellow pigment of the spice turmeric, is known for its antioxidant, anti-inflammatory, and anticarcinogenic properties. Capable of affecting the initiation, promotion, and progression of carcinogenesis through multiple mechanisms, curcumin has potential utility for both chemoprevention and chemotherapy. Previous studies demonstrated that curcumin can inhibit ornithine decarboxylase (ODC) activity in human leukemia and breast cancer cells, and pretreatment with dietary curcumin blocks carcinogen-induced ODC activity in rodent models of skin, colon, and renal cancer. The current study investigated the regulation of polyamine metabolism in human gastric and colon carcinoma cell lines in response to curcumin. Curcumin treatment significantly induced spermine oxidase (SMOX) mRNA and activity, which results in the generation of hydrogen peroxide, a source of ROS. Simultaneously, curcumin down regulated spermidine/spermine N1-acetyltransferase (SSAT) activity and the biosynthetic enzymes ODC and S-adenosylmethionine decarboxylase (SAMDC), thereby diminishing intracellular polyamine pools. Combination treatments using curcumin with the ODC inhibitor 2-difluoromethylornithine (DFMO), an agent currently in clinical chemoprevention trials, significantly enhanced inhibition of ODC activity and decreased growth of GI cancer cell lines beyond that observed with either agent alone. Similarly, combining curcumin with the polyamine analogue bis(ethyl)norspermine enhanced growth inhibition that was accompanied by enhanced accumulation of the analogue and decreased intracellular polyamine levels beyond those observed with either agent alone. Importantly, cotreatment with curcumin permitted the lowering of the effective dose of ODC inhibitor or polyamine analogue. These studies provide insight into the polyamine-related mechanisms involved in the cancer cell response to curcumin and its potential as a chemopreventive or chemotherapeutic agent in the GI tract.

Detection of Thermospermine and Spermine by HPLC in Plants.

Thermospermine, a structural isomer of spermine, is widely spread in the plant kingdom and has recently been shown to play a key role in the repression of xylem differentiation in vascular plants. However, a standard high-performance liquid chromatography (HPLC) protocol for detecting polyamines as their dansyl derivative cannot distinguish themospermine from spermine. These isomers become separated from each other after benzoylation. In this chapter, we describe a simple protocol for extraction, benzoylation, and HPLC detection of thermospermine and spermine with other polyamines from plant material.

Determination of S-Adenosylmethionine Decarboxylase Activity in Plants.

The synthesis of spermidine, spermine and thermospermine requires the addition of aminopropyl groups from decarboxylated S-adenosyl-methionine (dSAM). The synthesis of dSAM is catalyzed by S-adenosylmethionine decarboxylase. dSAM levels are usually low, which constitutes a rate-limiting factor in the synthesis of polyamines. In this chapter, we provide a protocol for the determination of SAMDC activity in plants through the detection of radiolabelled CO2 released during the SAMDC reaction.

The LIM domain only 4 protein is a metabolic responsive inhibitor of protein tyrosine phosphatase 1B that controls hypothalamic leptin signaling.

Protein tyrosine phosphatase 1B (PTP1B) counteracts leptin signaling and is a therapeutic target for obesity and diabetes. Here we found that LIM domain only 4 (LMO4) inhibits PTP1B activity by increasing the oxidized inactive form of PTP1B. Mice with neuronal ablation of LMO4 have elevated PTP1B activity and impaired hypothalamic leptin signaling, and a PTP1B inhibitor normalized PTP1B activity and restored leptin control of circulating insulin levels. LMO4 is palmitoylated at its C-terminal cysteine, and deletion of this residue prevented palmitoylation and retention of LMO4 at the endoplasmic reticulum and abolished its inhibitory effect on PTP1B. Importantly, LMO4 palmitoylation is sensitive to metabolic stress; mice challenged with a brief high-fat diet or acute intracerebroventricular infusion of saturated fatty acid had less palmitoylated LMO4, less oxidized PTP1B, and increased PTP1B activity in the hypothalamus. Thus, unleashed PTP1B activity attributable to loss of LMO4 palmitoylation may account for rapid loss of central leptin signaling after acute exposure to saturated fat.

Nitric oxide rescues thalidomide mediated teratogenicity.

Thalidomide, a sedative drug given to pregnant women, unfortunately caused limb deformities in thousands of babies. Recently the drug was revived because of its therapeutic potential; however the search is still ongoing for an antidote against thalidomide induced limb deformities. In the current study we found that nitric oxide (NO) rescues thalidomide affected chick (Gallus gallus) and zebrafish (Danio rerio) embryos. This study confirms that NO reduced the number of thalidomide mediated limb deformities by 94% and 80% in chick and zebrafish embryos respectively. NO prevents limb deformities by promoting angiogenesis, reducing oxidative stress and inactivating caspase-3 dependent apoptosis. We conclude that NO secures angiogenesis in the thalidomide treated embryos to protect them from deformities.

Hippocampal polyamine levels and transglutaminase activity are paralleling spatial memory retrieval in the C57BL/6J mouse.

Although a potential role for polyamines and transglutaminases (TGs) in memory mechanisms have been proposed, hippocampal spermine (SPM) and spermidine (SPD) levels as well as transamidating activity of TG in spatial memory have not been addressed yet. It was therefore the aim of the study to assess hippocampal polyamines and TG activity at the probe trial in a spatial memory paradigm. C57BL/6J mice (20 animals per group) were used for the experiments and divided into a trained and a yoked (untrained) group. The Morris water maze (MWM) was selected as the memory test, animals were sacrificed within 5 min following the probe trial and hippocampi were taken for biochemical analysis. SPD and SPM levels were assessed by an analytical procedure according to Gismondi et al. Transamidating activity of TG was determined following the method described by Chung and Folk using [14C] methylamine as substrate. γ-(Glutamyl)-polyamine levels were evaluated by ion exchange chromatography according to Folk et al. Animals learned the task in the MWM as latencies and pathlengths were significantly reduced. At the probe trial mice showed significantly higher preference for the target quadrant. Free SPD and SPM levels were manifold decreased in the trained as compared to the yoked group. Transamidating activity of TG was fourfold increased in trained as compared to yoked controls. γ-(Glutamyl)-SPD was comparable while γ-(glutamyl)-SPM was significantly higher in the trained group. The findings show a potential role for polyamines, their derivative γ-(glutamyl)-SPM and transamidating activity of TG at memory retrieval or formation. Results from this study are extending and knowledge on polyamines and report for the first time involvement of γ-(glutamyl)-SPM and transamidating activity of TG that may form the basis for future neurochemical and pharmacological studies and indeed, modulation of polyamine and TG activity has been already proposed as a tentative therapeutical concept.

Application of a molecularly imprinted polymer for the extraction of kukoamine a from potato peels.

A molecularly imprinted polymer (MIP) for the purification of N(1),N(12)-bis(dihydrocaffeoyl)spermine (kukoamine A) was computationally designed and tested. The properties of the polymer were characterized. The protocol of the solid phase extraction (SPE) of kukoamine A from potato peels was optimized. A HPLC-MS method for the quantification of kukoamine A was developed and used for all optimization studies. The capacity of the MIP in relation to kukoamine A from the potato peels extract was estimated at 54 mg/g of the polymer. The kukoamine A purified from potato extract using MIP was exceptionally pure (≈ 90%). Although the corresponding blank polymer was less selective than the MIP for the extraction of kukoamine A from the potato extract, it was shown that the blank polymer could be effectively used for the purification of the crude synthetic kukoamine (polymer capacity = 80 mg of kukoamine A/g of the adsorbent, kukoamine A purity ≈ 86%). Therefore, selective adsorbents could be computationally designed for other plant products, allowing their purification in quantities that would be sufficient for more detailed studies and potential practical applications.

Initial testing (stage 1) of the polyamine analog PG11047 by the pediatric preclinical testing program.

BACKGROUND: PG11047 is a novel conformationally restricted analog of the natural polyamine, spermine that lowers cellular endogenous polyamine levels and competitively inhibits natural polyamine functions leading to cancer cell growth inhibition. The activity of PG11047 was evaluated against the PPTP's in vitro and in vivo panels. PROCEDURES: PG11047 was evaluated against the PPTP in vitro panel using 96 hr exposure at concentrations ranging from 10 nM to 100 µM. It was tested against the PPTP in vivo panels at a dose of 100 mg/kg administered by the intraperitoneal route weekly for 6 weeks. RESULTS: In vitro PG11047 demonstrated a concentration-response pattern consistent with cytostatic activity. The median EC(50) for PG11047 was 71 nM. Cell lines of the Ewing sarcoma panel had a lower median EC(50) value compared to the remaining cell lines in the panel, while cell lines of the neuroblastoma panel had a higher median EC(50) value. In vivo PG11047 induced significant differences in EFS distribution compared to control in 5 of 32 (15.6%) of the evaluable solid tumor xenografts and in 0 of 7 (0%) of the evaluable ALL xenografts. The single case of tumor regression occurred in an ependymoma xenograft. CONCLUSIONS: Further pediatric development of PG11047 will require better defining a target population and identifying combinations for which there is a tumor-selective cytotoxic effect. The regression observed for an ependymoma xenograft and the exquisite sensitivity of some Ewing sarcoma cell lines to the antiproliferative effects of PG11047 provide leads for further preclinical investigations.

Dual targets guided screening and isolation of Kukoamine B as a novel natural anti-sepsis agent from traditional Chinese herb Cortex lycii.

Treating sepsis remains challenging at present. Bacterial lipopolysaccharide (LPS) and bacterial DNA/CpG DNA are important pathogenic molecules and drug targets for sepsis. It is thus a promising strategy to treat sepsis by discovering agents that neutralize LPS and CpG DNA simultaneously. In this study, we present evidences of the biosensor based screening and isolation of active anti-sepsis fractions and monomers from traditional Chinese herbs using dual targets (LPS and CpG DNA) guided drug discovery strategy. Firstly, LPS or CpG DNA was immobilized on surfaces of cuvettes in the biosensor to establish a screening platform. Then, Cortex lycii with both highest affinities was selected out from one hundred and fourteen traditional Chinese herbs. In subsequent experiments, chromatography was utilized and coupled with the biosensor to purify fractions with a higher affinity for LPS and CpG DNA. In line with affinity assay, these fractions were shown to neutralize LPS and CpG DNA and inhibit their activity in vitro and in vivo. Lastly, the contributing monomer Kukoamine B (KB) was purified. KB neutralized LPS and CpG DNA in vitro. It inhibited TLR4, TLR9 and MyD88 mRNA expressions up-regulated by LPS and CpG DNA, and also attenuated the LPS and CpG DNA elicited nuclear translocation of NF-κB p65 protein in RAW264.7 cells. It also protected mice from lethal challenge of heat-killed E. coli, a mixture of LPS and CpG DNA. In conclusion, we presented a dual target guided discovery of a novel anti-sepsis agent KB from traditional Chinese herbs via combination of biosensor technology and chromatography methods.

Quantitative analysis of plant polyamines including thermospermine during growth and salinity stress.

Arabidopsis thaliana was thought to contain two spermine synthase genes, ACAULIS 5 (ACL5) and SPMS. Recent investigations, however, revealed that the ACL5 gene encodes thermospermine synthase. In this study, we have established a simple method to separate two isomers of tetraamine, spermine and thermospermine, in extracts from plant tissues of less than 500 mg. Polyamines (PAs) extracted from plant tissues were benzoylated, and the derivatives were completely resolved by high-performance liquid chromatography on a C18 reverse-phase column, by eluting with 42% (v/v) acetonitrile in water in an isocratic manner at 30 degrees C and monitoring at 254 nm. The relevance of the method was confirmed by co-chromatography with respective PAs and by the PA analysis of the single- and double-mutants of acl5 and spms, which could not synthesize thermospermine and/or spermine, respectively. Furthermore, with this method, we monitored the thermospermine contents in various tissues of A. thaliana and found that stems and flowers contain two- to three-fold more thermospermine compared to whole seedlings and mature leaves. The presence of thermospermine was confirmed in Oryza sativa and Lycopersicon pesculentum. Finally we addressed whether salinity stress changes the contents of PAs including thermospermine in Arabidopsis.

Inhibition of PTP1B by trodusquemine (MSI-1436) causes fat-specific weight loss in diet-induced obese mice.

Trodusquemine (MSI-1436) causes rapid and reversible weight loss in genetic models of obesity. To better predict the potential effects of trodusquemine in the clinic, we investigated the effects of trodusquemine treatment in a murine model of diet-induced obesity (DIO). Trodusquemine suppressed appetite, reduced body weight (BW) in a fat-specific manner, and improved plasma insulin and leptin levels in mice. Screening assays revealed that trodusquemine selectively inhibited protein-tyrosine phosphatase 1B (PTP1B), a key enzyme regulating insulin and leptin signaling. Trodusquemine significantly enhanced insulin-stimulated tyrosine phosphorylation of insulin receptor (IR) beta and STAT3, direct targets of PTP1B, in HepG2 cells in vitro and/or hypothalamic tissue in vivo. These data establish trodusquemine as an effective central and peripheral PTP1B inhibitor with the potential to elicit noncachectic fat-specific weight loss and improve insulin and leptin levels.