The deubiquitinase USP28 maintains the expression of the transcription factor MYCN and is essential in neuroblastoma cells.

Neuroblastoma (NB) is one of the most common extracranial solid tumors in children. MYCN gene amplification is highly associated with poor prognosis in high-risk NB patients. In non-MYCN-amplified high-risk NB patients, the expression of c-MYC (MYCC) and its target genes is highly elevated. USP28 as a deubiquitinase is known to regulate the stability of MYCC. We show here USP28 also regulates the stability of MYCN. Genetic depletion or pharmacologic inhibition of the deubiquitinase strongly destabilizes MYCN and stops the growth of NB cells that overexpress MYCN. In addition, MYCC could be similarly destabilized in non-MYCN NB cells by compromising USP28 function. Our results strongly suggest USP28 as a therapeutic target for NB with or without MYCN amplification/overexpression.

The deubiquitinase USP28 stabilizes the expression of RecQ family helicases and maintains the viability of triple negative breast cancer cells.

Triple-negative breast cancer (TNBC) lacks significant expression of the estrogen receptor, the progesterone receptor, and of human epidermal growth factor receptor. It is the most aggressive and malignant of all breast cancers, and for which, there are currently no effective targeted therapies. We have shown previously that the RecQ helicase family member RECQL5 is essential for the proliferation and survival of TNBC cells; however, the mechanism of its involvement in cell viability has not been shown. Here, we report that the expression of RecQ family helicases, including RECQL5, is regulated by the deubiquitinase USP28. We found using genetic depletion or a small molecule inhibitor that like RECQL5, USP28 is also essential for TNBC cells to proliferate in vitro and in vivo. Compromising the function of USP28 by shRNA knockdown or the inhibitor caused TNBC cells to arrest in S/G2 phases, concurrent with DNA-damage checkpoint activation. We further showed that the small molecule inhibitor of USP28 displayed anti-tumor activity against xenografts derived from TNBC cells. Our results suggest that USP28 could be a potential therapeutic target for triple negative breast cancer.

Ubiquitin-specific protease 28 deubiquitinates TCF7L2 to govern the action of the Wnt signaling pathway in hepatic carcinoma.

Overexpression of ubiquitin-specific protease 28 (USP28) is found in hepatic carcinoma. It is unclear whether the deubiquitinase plays a role in hepatocarcinogenesis. Deregulation of the Wnt signaling pathway is frequently associated with liver cancer. Transcription factor 7-like 2 (TCF7L2) is an important downstream transcription factor of the Wnt/ß-catenin signaling pathway, but the mechanisms by which TCF7L2 itself is regulated have not yet been revealed. Here, we report that USP28 promotes the activity of the Wnt signaling pathway through maintaining the stability of TCF7L2. We further show that FBXW7 is the E3 ubiquitin ligase for TCF7L2. By regulating the levels of TCF7L2, USP28 modulates the Wnt/ß-catenin signaling in liver cancer and USP28 depletion or inhibition by a small molecule inhibitor leads to a halt of growth in liver cancer cells. These results suggest that USP28 could be a potential therapeutic target for liver cancer.

USP25 regulates KEAP1-NRF2 anti-oxidation axis and its inactivation protects acetaminophen-induced liver injury in male mice.

Nuclear factor erythroid 2-related factor 2 (NRF2) is a transcription factor responsible for mounting an anti-oxidation gene expression program to counter oxidative stress. Under unstressed conditions, Kelch-like ECH-associated protein 1 (KEAP1), an adaptor protein for CUL3 E3 ubiquitin ligase, mediates NRF2 ubiquitination and degradation. We show here that the deubiquitinase USP25 directly binds to KEAP1 and prevents KEAP1's own ubiquitination and degradation. In the absence of Usp25 or if the DUB is inhibited, KEAP1 is downregulated and NRF2 is stabilized, allowing the cells to respond to oxidative stress more readily. In acetaminophen (APAP) overdose-induced oxidative liver damage in male mice, the inactivation of Usp25, either genetically or pharmacologically, greatly attenuates liver injury and reduces the mortality rates resulted from lethal doses of APAP.

GPR177 in A-fiber sensory neurons drives diabetic neuropathic pain via WNT-mediated TRPV1 activation.

Diabetic neuropathic pain (DNP) is a common and devastating complication in patients with diabetes. The mechanisms mediating DNP are not completely elucidated, and effective treatments are lacking. A-fiber sensory neurons have been shown to mediate the development of mechanical allodynia in neuropathic pain, yet the molecular basis underlying the contribution of A-fiber neurons is still unclear. Here, we report that the orphan G protein-coupled receptor 177 (GPR177) in A-fiber neurons drives DNP via WNT5a-mediated activation of transient receptor potential vanilloid receptor-1 (TRPV1) ion channel. GPR177 is mainly expressed in large-diameter A-fiber dorsal root ganglion (DRG) neurons and required for the development of DNP in mice. Mechanistically, we found that GPR177 mediated the secretion of WNT5a from A-fiber DRG neurons into cerebrospinal fluid (CSF), which was necessary for the maintenance of DNP. Extracellular perfusion of WNT5a induced rapid currents in both TRPV1-expressing heterologous cells and nociceptive DRG neurons. Computer simulations revealed that WNT5a has the potential to bind the residues at the extracellular S5-S6 loop of TRPV1. Using a peptide able to disrupt the predicted WNT5a/TRPV1 interaction suppressed DNP- and WNT5a-induced neuropathic pain symptoms in rodents. We confirmed GPR177/WNT5A coexpression in human DRG neurons and WNT5A secretion in CSF from patients with DNP. Thus, our results reveal a role for WNT5a as an endogenous and potent TRPV1 agonist, and the GPR177-WNT5a-TRPV1 axis as a driver of DNP pathogenesis in rodents. Our findings identified a potential analgesic target that might relieve neuropathic pain in patients with diabetes.

Design, Synthesis, and Potential Antidepressant-like Activity of 7-prenyloxy-2,3-dihydroflavanone Derivatives.

A series of 7-prenyloxy-2, 3-dihydroflavanone derivatives were synthesized and screened for their antidepressant-like activity. Among them, it was observed that compounds 5j and 5k were found to be the most antidepressant-like activity. In addition, it was found that compounds 5j and 5k significantly increased the concentrations of the main neurotransmitters 5-HT and NE in the hippocampus, hypothalamus, and cortex. Compounds 5j and 5k also significantly increased the contents of 5-HIAA in the hippocampus and cortex, shut down 5-HT metabolism compared with mice treated with stress vehicle. These results suggested that compounds 5j and 5k displayed potent antidepressant-like properties that were mediated via neurochemical systems.

Practical Synthesis, Antidepressant, and Anticonvulsant Activity of 3-Phenyliminoindolin-2-one Derivatives.

Herein, a series of 3-phenyliminoindolin-2-one derivatives were designed, synthesized, and screened for their antidepressant and anticonvulsant activities. The IR spectra of the compounds afforded NH stretching (3340-3346 cm(-1)) bands and C=O stretching (1731-1746 cm(-1)). In the (1)H-NMR spectra of the compounds, N-H protons of indoline ring were observed at 10.65-10.89 ppm generally as broad bands, and (13)C-NMR spectra of the compounds C=O were seen at 161.72-169.27 ppm. Interestingly, compounds 3o, 3p and 3r significantly shortened immobility time in the The forced swimming test (FST) and The tail suspension test (TST) at 50 mg/kg dose levels. In addition, compound 3r exhibited higher levels of efficacy than the reference standard fluoxetine but had no effect on locomotor activity in the open-field test. Compound 3r significantly increased serotonin and norepinephrine and the metabolite 5-hydroxyindoleacetic acid in mouse brain, suggesting that the effects of compound 3r may be mediated through these neurotransmitters. In the seizure screen, 15 compounds showed some degree against PTZ-induced seizure at a dose of 100 mg/kg, and the tested compounds did not show any neurotoxicity at a dose of 300 mg/kg in the rotarod test.

Synthesis and Evaluation of Phenyliminoindolin-Containing Phenylacetamide Derivatives with the Antidepressant and Anticonvulsant Effects.

BACKGROUND: To discover a novel antidepressant-like effect and anticonvulsant compound, seventeen new 2-oxo-3-phenyliminoindolin-1-Nphenylacetamide compounds were synthesized and screened for the antidepressant activity and anticonvulsant effects. METHOD: 2-oxo-3-phenyliminoindolin-1-N-phenylacetamide derivatives were synthesized with indoline-2, 3-dione as the starting material, through a nucleophilic substitution reaction and a nucleophilic addition-elimination reaction. The target derivatives 2a-2q were evaluated the antidepressant-like activity using the FST, TST, and evaluated anticonvulsant effect by MES test. The main monoamine neurotransmitters and their metabolites in mouse brain regions were also simultaneously determined by HPLC-ECD. RESULTS: It was observed that 13 compounds showed significant reductions in the immobility time in the FST at a concentration of 50 mg/kg. Compound 2b was found to have the most potent antidepressant activity in the FST and the TST for 30 min after treatment. Compound 2b significantly increased the concentrations of the main neurotransmitters 5-HT, NE and the metabolite (5-HIAA, suggesting that the effects of compound 2b may be mediated through these neurotransmitters. As assessed using maximal electroshock, 13 compounds showed the anticonvulsant effects administered at the concentration levels of 100 or 300 mg/kg. Compound 2b showed anticonvulsant effect. Compound 2b could potentially be used as adjuncts of the antidepressants to treat depression in patients with epilepsy. CONCLUSION: In conclusion, compound 2b produced significant antidepressant-like activity and the mechanism of action may be due to increased 5-HT and NE in the mouse. Compound 2b showed more anticonvulsant effect. Compound 2b could potentially be used as adjuncts of the antidepressants to treat depression in patients with epilepsy.

Synthesis, potential anticonvulsant and antidepressant effects of 2-(5-methyl-2,3-dioxoindolin-1-yl)acetamide derivatives.

A new series of 2-(5-methyl-2,3-dioxoindolin-1-yl)acetamide derivatives were synthesized and evaluated for their anticonvulsive activity in a pentylenetetrazole (PTZ)-evoked convulsion model and antidepressant activity in the forced swimming test (FST) model. Eleven synthesized compounds were found to be protective against PTZ-induced seizure and showed the anticonvulsant activity. In addition, four of the synthesized compounds (4l, 4m, 4p and 4q) showed potent antidepressant-like activity. Among these compounds, compound 4l was found to have the most potent antidepressant-like activity, and significantly reduced the duration of immobility time at 100 mg/kg dose level when compared to the vehicle control, which is similar to the reference drug fluoxetine.

Fucosterol, a sterol extracted from Sargassum fusiforme, shows antidepressant and anticonvulsant effects.

We previously showed that extracts of Sargassum fusiforme significantly reduce immobility time in the forced swim test and tail suspension test, suggesting that these extracts possess antidepressant-like effects. Here, fucosterol extracted from S. fusiforme was evaluated for antidepressant and anticonvulsant activities in mice. Fucosterol (10, 20, 30 and 40mg/kg) significantly shortened immobility time in the forced swim test and tail suspension test for30min after treatment but had no effect on locomotor activity in the open field test. Fucosterol significantly increased serotonin, norepinephrine and the metabolite 5-hydroxyindoleacetic acid in mouse brain, suggesting that the effects of fucosterol may be mediated through these neurotransmitters. As assessed using maximal electroshock, fucosterol (20, 40, 100mg/kg) possessed anticonvulsant activity, whereas rotarod toxicity test results indicated that fucosterol did not induce neurotoxicity at the same dose levels in mice. Thus, fucosterol may be a useful antidepressant adjunct candidate for treating depression in patients with epilepsy. A significant increase in hippocampal brain-derived neurotrophic factor (BDNF) levels was found in the fucosterol 20mg/kg group (P<0.05). Our findings suggested that fucosterol may possess an antidepressant-like effect, which may be mediated by increasing central BDNF levels.