Induction of IFIT1/IFIT3 and inhibition of Bcl-2 orchestrate the treatment of myeloma and leukemia via pyroptosis.

Induction of pyroptosis is proposed as a promising strategy for the treatment of hematological malignancies, but little is known. In the present study, we find clioquinol (CLQ), an anti-parasitic drug, induces striking myeloma and leukemia cell pyroptosis on a drug screen. RNA sequencing reveals that the interferon-inducible genes IFIT1 and IFIT3 are markedly upregulated and are essential for CLQ-induced GSDME activation and cell pyroptosis. Specifically, IFIT1 and IFIT3 form a complex with BAX and N-GSDME therefore directing N-GSDME translocalization to mitochondria and increasing mitochondrial membrane permeabilization and triggering pyroptosis. Furthermore, venetoclax, an activator of BAX and an inhibitor of Bcl-2, displays strikingly synergistic effects with CLQ against leukemia and myeloma via pyroptosis. This study thus reveals a novel mechanism for mitochondrial GSDME in pyroptosis and it also illustrates that induction of IFIT1/T3 and inhibition of Bcl-2 orchestrate the treatment of leukemia and myeloma via pyroptosis.

Preparation, in vitro anti-tumour activity and in vivo pharmacokinetics of RGD-decorated liposomes loaded with shikonin.

Shikonin (SHK) has been evidenced to possess effects against various cancer cells. However, poor aqueous solubility and high toxicity restrict its application. In the study, RGD-decorated liposomes loaded with SHK (RGD-Lipo-SHK) were prepared via thin-film hydration method. Characterization and cellular uptake of liposomes was evaluated. Cytotoxicity of blank liposomes and different SHK formulations was measured against breast cancer cells (MDA-MB-231, MCF-7, and MCF-10A). Anti-tumour effects and pharmacokinetic parameters of different SHK formulations were appraised in tumour spheroids and in rat model, respectively. Liposomes displayed a particle size of less than 127 nm with a polydispersity index about 0.21. The encapsulation efficiency was about 91% for SHK, and drug leakage rate of liposomes was less than 6%. RGD-Lipo-SHK showed superior cellular internalization in the αvß3-positive MDA-MB-231 cells. Blank liposomes had no cytotoxicity to MDA-MB-231 and MCF-7 cells. Howbeit, different SHK formulations obviously inhibited proliferation of MCF-10A cells, especially free SHK. Meanwhile, RGD-Lipo-SHK significantly inhibited growth inhibition of tumour spheroids. The pharmacokinetics study indicated that the peak concentration, area under plasma concentration-time curves, half-life, and mean residence time of RGD-Lipo-SHK distinctly increased compared with those of free SHK. Altogether, these results demonstrated RGD-Lipo-SHK could reduce cytotoxicity, strengthen the antitumor-targeted effect, and prolong circulation time, which provides a foundation for further in vivo experimentations.

Bruceine B Displays Potent Antimyeloma Activity by Inducing the Degradation of the Transcription Factor c-Maf.

The oncogenic transcription factor c-Maf has been proposed as an ideal therapeutic target for multiple myeloma (MM), a not-yet-curable malignancy of plasma cells. In the present study, we establish a c-Maf-based luciferase screen system and apply it to screen a homemade library composed of natural products from which bruceine B (BB) is identified to display potent antimyeloma activity. BB is a key ingredient isolated from the Chinese traditional medicinal plant Brucea javanica (L.) Merr. (Simaroubaceae). BB inhibits MM cell proliferation and induces MM cell apoptosis in a caspase-3-dependent manner. The mechanism studies showed that BB inhibits c-Maf transcriptional activity and downregulates the expression of CCND2 and ITGB7, the downstream genes typically modulated by c-Maf. Moreover, BB induces c-Maf degradation via proteasomes by inducing c-Maf for K48-linked polyubiquitination in association with downregulated Otub1 and USP5, two proven deubiquitinases of c-Maf. We also found that c-Maf activates STAT3 and BB suppresses the STAT3 signaling. In the in vivo study, BB displays potent antimyeloma activity and almost suppresses the growth of myeloma xenografts in 7 days but shows no overt toxicity to mice. In conclusion, this study identifies BB as a novel inhibitor of c-Maf by promoting its degradation via the ubiquitin-proteasomal pathway. Given the safety and the successful clinical application of bruceine products in traditional medicine, BB is ensured for further investigation for the treatment of patients with MM.

USP7 promotes non-small-cell lung cancer cell glycolysis and survival by stabilizing and activating c-Abl.

BACKGROUND: Abelson tyrosine kinase (c-Abl) is frequently mutated and highly expressed, and promotes non-small-cell lung cancer (NSCLC) survival, metastasis and tumorigenesis. c-Abl could also be modified through ubiquitination, but the underlying mechanism is not well understood. METHODS: Mass spectrometry assays were performed to search c-Abl deubiquitination enzymes. The molecular mechanism was determined using Co-IP assays, pull-down assays, Western blotting upon gene knockdown or overexpression. Cell lines and animal models were used to investigate the role of c-Abl and USP7 in NSCLC. EdU staining assay and Transwell assay were performed to evaluate the proliferation and migration ability of NSCLC cells, respectively. RESULTS: Ubiquitin-specific protease 7 (USP7) is found to upregulate c-Abl via the deubiquitinase screen. USP7 interacts with c-Abl and decreases its K48-linked polyubiquitination, thereby increasing the stability of c-Abl. In addition to the wild-type one, c-Abl mutants can also be deubiquitinated and stabilized by USP7. Moreover, USP7 promotes c-Abl accumulation in cytoplasm by increasing its binding to 14-3-3α/ß and activates the oncogenic c-Abl signalling pathway. Furthermore, the USP7/c-Abl axis promotes NSCLC cell glycolysis by direct phosphorylating and stabilizing hexokinase-2 (HK2). Knockdown of USP7 or c-Abl suppresses NSCLC cell glycolysis and reduces lactate production. Further studies revealed that overexpression of USP7 facilitates NSCLC cell growth and metastasis as well as xenograft growth in nude mice, while these activities are suppressed with USP7 or c-Abl being knocked down. CONCLUSIONS: USP7 is a deubiquitinase of c-Abl and upregulates its oncogenic activity. USP7 promotes NSCLC cell metabolism by activating c-Abl and HK2. Targeting the USP7/c-Abl/HK2 axis might be a potential strategy to the precision therapy of NSCLC.

Differences between soluble and insoluble undenatured type II collagen in improving osteoarthritis in rats and their potential mechanisms.

Our previous research showed that soluble (SC II) and insoluble (IC II) undenatured type II collagen had significant differences during gastrointestinal digestion in vitro, and SC II exposed more type II collagen with triple helix structure. However, the differences in their in vivo digestive characteristics, improvement on osteoarthritis (OA), and possible mechanisms have not been elucidated. The aim of this study was to explore these issues. After oral administration of SC II and IC II, the joint swelling of OA rats significantly reduced, and the weight bearing ratio of right hind limb significantly increased, especially in SC II group (raised to 48%). The Mankin and OARSI scores decreased by 35% and 48% in SC II group, respectively. SC II and IC II increased the mRNA expression of anti-inflammatory factors and the proportion of regulatory T cells (Treg). Importantly, type II collagen released by IC II during in vivo gastrointestinal digestion was far less than SC II, which explained the higher ability of SC II to induce immune tolerance in small intestine than IC II. Bioinformatics analysis showed that the differential genes between model and control were significantly enriched in PI3K/AKT, PPAR and AMPK signalling pathways, and 24 hub genes were analyzed. SC II significantly down-regulated the mRNA expression of Il6, Ccl7, NF-κB, AKT and up-regulated the mRNA expression of Scd1. These results showed that SC II was superior to IC II in improving OA by inducing immune tolerance and could regulate key biomarkers and signalling pathways in OA rats.

TRIM26 promotes non-small cell lung cancer survival by inducing PBX1 degradation.

The transcription factor PBX1 is regarded as an oncogene in various cancers, but its role in non-small cell lung cancer (NSCLC) and the detailed mechanism is not known. In the present study, we found that PBX1 is downregulated in NSCLC tissues and inhibits NSCLC cell proliferation and migration. Subsequently, we performed an affinity purification-coupled tandem mass spectrometry (MS/MS) and found the ubiquitin ligase TRIM26 in the PBX1 immunoprecipitates. Moreover, TRIM26 binds to and mediates PBX1 for K48-linked polyubiquitination and proteasomal degradation. Noticeably, TRIM26 activity depends on its C-terminal RING domain when it is deleted TRIM26 loses its function towards PBX1. TRIM26 further inhibits PBX1 transcriptional activity and downregulates the PBX1 downstream genes, such as RNF6. Moreover, we found that overexpression of TRIM26 significantly promotes NSCLC proliferation, colony formation, and migration in contradiction to PBX1. TRIM26 is highly expressed in NSCLC tissues and predicts poor prognosis. Lastly, the growth NSCLC xenografts is promoted by overexpression of TRIM26 but is suppressed by TRIM26 knockout. In conclusion, TRIM26 is a ubiquitin ligase of PBX1 and it promotes while PBX1 inhibits NSCLC tumor growth. TRIM26 might be a novel therapeutic target for the treatment of NSCLC.

The ubiquitin ligase HERC4 suppresses MafA transcriptional activity triggered by GSK3ß in myeloma by atypical K63-linked polyubiquitination.

MafA and c-Maf are close members of the Maf transcription factor family and indicators of poor prognosis of multiple myeloma (MM). Our previous study finds that the ubiquitin ligase HERC4 induces c-Maf degradation but stabilizes MafA, and the mechanism is elusive. In the present study, we find that HERC4 interacts with MafA and mediates its K63-linked polyubiquitination at K33. Moreover, HERC4 inhibits MafA phosphorylation and its transcriptional activity triggered by glycogen synthase kinase 3ß (GSK3ß). The K33R MafA variant prevents HERC4 from inhibiting MafA phosphorylation and increases MafA transcriptional activity. Further analyses reveal that MafA can also activate the STAT3 signaling, but it is suppressed by HERC4. Lastly, we demonstrate that lithium chloride, a GSK3ß inhibitor, can upregulate HERC4 and synergizes dexamethasone, a typical anti-MM drug, in inhibiting MM cell proliferation and xenograft growth in nude mice. These findings thus highlight a novel regulation of MafA oncogenic activity in MM and provide the rationale by targeting HERC4/GSK3ß/MafA for the treatment of MM.

Inhibition of USP10 induces myeloma cell apoptosis by promoting cyclin D3 degradation.

The cell cycle regulator cyclin D3 (CCND3) is highly expressed in multiple myeloma (MM) and it promotes MM cell proliferation. After a certain phase of cell cycle, CCND3 is rapidly degraded, which is essential for the strict control of MM cell cycle progress and proliferation. In the present study, we investigated the molecular mechanisms regulating CCND3 degradation in MM cells. By utilizing affinity purification-coupled tandem mass spectrometry, we identified the deubiquitinase USP10 interacting with CCND3 in human MM OPM2 and KMS11 cell lines. Furthermore, USP10 specifically prevented CCND3 from K48-linked polyubiquitination and proteasomal degradation, therefore enhancing its activity. We demonstrated that the N-terminal domain (aa. 1-205) of USP10 was dispensable for binding to and deubiquitinating CCND3. Although Thr283 was important for CCND3 activity, it was dispensable for CCND3 ubiquitination and stability modulated by USP10. By stabilizing CCND3, USP10 activated the CCND3/CDK4/6 signaling pathway, phosphorylated Rb, and upregulated CDK4, CDK6 and E2F-1 in OPM2 and KMS11 cells. Consistent with these findings, inhibition of USP10 by Spautin-1 resulted in accumulation of CCND3 with K48-linked polyubiquitination and degradation that synergized with Palbociclib, a CDK4/6 inhibitor, to induce MM cell apoptosis. In nude mice bearing myeloma xenografts with OPM2 and KMS11 cells, combined administration of Spautin-l and Palbociclib almost suppressed tumor growth within 30 days. This study thus identifies USP10 as the first deubiquitinase of CCND3 and also finds that targeting the USP10/CCND3/CDK4/6 axis may be a novel modality for the treatment of myeloma.

Proteasomal inhibitors induce myeloma cell pyroptosis via the BAX/GSDME pathway.

Proteasomes are overexpressed in multiple myeloma (MM) and proteasomal inhibitors (PIs) have been widely used for the treatment of MM. PIs are reported to induce MM cell apoptosis but impair necroptosis. In the present study, we found that PIs MG132 and bortezomib induce MM cell pyroptosis, a novel type of cell death, in a GSDME-dependent manner. Lack of GSDME totally blocks PI-induced pyroptosis. Interestingly, we found that Caspase-3/6/7/9 are all involved in pyroptosis triggered by PIs because the specific inhibitor of each caspase ablates GSDME activation. PIs markedly reduce mitochondrial membrane potential. Moreover, PIs disrupt the interaction of Bcl-2 and BAX, induce cytochrome c release from mitochondria to cytosol and activate GSDME. Furthermore, we found that overexpression of an N-terminal portion of GSDME suffices to release cytochrome c from mitochondria and to activate Caspase-3/9, suggesting N-GSDME might penetrate the mitochondrial membrane. Consistent with Bcl-2 inhibition, BAX can induce MM cell pyroptosis in a GSDME-dependent manner. In accordance with these findings, inhibition of Bcl-2 synergizes with PIs to induce MM cell pyroptosis. Therefore, the present study indicates that PIs trigger MM cell pyroptosis via the mitochondrial BAX/GSDME pathway and provides a rationale for combined treatment of MM with Bcl-2 and proteasome inhibitors to increase therapeutic efficiency via induction of pyroptosis.

Associations between Dietary Antioxidant Vitamin Intake and the Changes in Bone Mass in Chinese Adolescents: A 2.5-Year Longitudinal Study.

(1) Background: Optimal bone mass accumulation during adolescence is crucial for maximising peak bone mass during adulthood. Dietary antioxidant vitamins may contribute to bone mass accumulation. This 2.5-year-long longitudinal study aimed to evaluate the relationships between dietary vitamin A, C, and E intakes and the annual changes in bone parameters among Chinese adolescents. (2) Method: Subjects aged 10-18 years (n = 1418) were recruited from a secondary school in Jiangmen, China. Dietary vitamin A, C, and E intakes were assessed using 24 h dietary records over 3 consecutive days. The Sahara Clinical Bone Sonometer was used to measure the broadband ultrasound attenuation (BUA) and the speed of sound (SOS). Their annual changes were then calculated (i.e., BUA%/year, SOS%/year). The associations were detected after adjusting for the baseline bone phenotype; age; sex; weight; height; pubertal stage; physical activity; and dietary intakes of vitamin D, calcium and energy. (3) Results: A curvilinear relationship was found between the dietary intake of vitamin C and BUA%/year (p = 0.026); further analyses in the subgroups revealed that this relationship was observed in male adolescents (p = 0.012). A positive association was observed only in boys with a dietary vitamin C intake of ≥159.01 mg/day (ß = 0.395, p = 0.036). Moreover, a linear positive association was shown between the dietary intake of vitamin E and BUA%/year in female adolescents (ß = 0.082, p = 0.033). (4) Conclusion: Our findings indicated that dietary vitamin C intake has a threshold effect on bone mass gain in male adolescents and that dietary vitamin E intake could be a positive predictor of bone mass gain in female adolescents.

Induction of zinc finger protein RNF6 auto-ubiquitination for the treatment of myeloma and chronic myeloid leukemia.

The zinc finger ubiquitin ligase RNF6 has been proposed as a potential therapeutic target in several cancers, but understanding its molecular mechanism of degradation has been elusive. In the present study, we find that RNF6 is degraded via auto-ubiquitination in a manner dependent on its Really Interesting New Gene (RING) domain. We determine that when the RING domain is deleted (ΔRING) or the core cysteine residues in the zinc finger are mutated (C632S/C635S), the WT protein, but not the ΔRING or mutant RNF6 protein, undergoes polyubiquitination. We also identify USP7 as a deubiquitinase of RNF6 by tandem mass spectrometry. We show that USP7 interacts with RNF6 and abolishes its K48-linked polyubiquitination, thereby preventing its degradation. In contrast, we found a USP7-specific inhibitor promotes RNF6 polyubiquitination, degradation, and cell death. Furthermore, we demonstrate the anti-leukemic drug Nilotinib and anti-myeloma drug Panobinostat (LBH589) induce RNF6 K48-linked polyubiquitination and degradation in both multiple myeloma (MM) and leukemia cells. In agreement with our hypothesis on the mode of RNF6 degradation, we show these drugs promote RNF6 auto-ubiquitination in an in vitro ubiquitination system without other E3 ligases. Consistently, reexpression of RNF6 ablates drug-induced MM and leukemia cell apoptosis. Therefore, our results reveal that RNF6 is a RING E3 ligase that undergoes auto-ubiquitination, which could be abolished by USP7 and induced by anti-cancer drugs. We propose that chemical induction of RNF6 auto-ubiquitination and degradation could be a novel strategy for the treatment of hematological malignancies including MM and leukemia.

Targeting the oncogenic transcription factor c-Maf for the treatment of multiple myeloma.

Multiple myeloma (MM) is a hematologic malignancy derived from clonal expansion of plasma cells within the bone marrow and it may progress to the extramedullary region in late stage of the disease course. c-Maf, an oncogenic zipper leucine transcription factor, is overexpressed in more than 50% MM cell lines and primary species in association with chromosomal translocation, aberrant signaling transduction and modulation of stability. By triggering the transcription of critical genes including CCND2, ITGB7, CCR1, ARK5, c-Maf promotes MM progress, proliferation, survival and chemoresistance. Notably, c-Maf is usually expressed at the embryonic stage to promote cell differentiation but less expressed in healthy adult cells. c-Maf has long been proposed as a promising therapeutic target of MM and a panel of small molecule compounds have been identified to downregulate c-Maf and display potent anti-myeloma activities. In the current article, we take a concise summary on the advances in c-Maf biology, pathophysiology, and targeted drug discovery in the potential treatment of MM.

TRIM25 activates AKT/mTOR by inhibiting PTEN via K63-linked polyubiquitination in non-small cell lung cancer.

The PTEN/AKT/mTOR signaling pathway is frequently dysregulated in non-small cell lung cancer (NSCLC), but the mechanisms are not well-understood. The present study found that the ubiquitin ligase TRIM25 is highly expressed in NSCLC tissues and promotes NSCLC cell survival and tumor growth. Mechanistic studies revealed that TRIM25 binds to PTEN and mediates its K63-linked ubiquitination at K266. This modification prevents the plasma membrane translocation of PTEN and reduces its phosphatase activity therefore accumulating PI(3,4,5)P3. TRIM25 thus activates the AKT/mTOR signaling. Moreover, we found that the antibacterial nitroxoline can activate PTEN by reducing its K63-linked polyubiquitination and sensitizes NSCLC to cisplatin-induced apoptosis. This study thus identified a novel modulation on the PTEN signaling pathway by TRIM25 and provides a potential target for NSCLC treatment.

Deubiquitinase USP5 promotes non-small cell lung cancer cell proliferation by stabilizing cyclin D1.

BACKGROUND: Cyclin D1 (CCND1) is overexpressed in non-small cell lung cancer (NSCLC) and contributes to its tumorigenesis and progression. Accumulating evidence shows that ubiquitin-specific protease 5 (USP5), an important member of the USP family, acts as a tumor promoter by deubiquitinating and stabilizing oncoproteins. However, neither the mechanism for dysregulated turnover of CCND1 protein nor the association of CCND1 with USP5 in NSCLC is well understood. METHODS: The association of USP5 with CCND1 in human NSCLC cells and clinical tissues was determined by immunoprecipitation/immunoblotting, immunohistochemistry (IHC), and The Cancer Genome Atlas database analyses. The effect of USP5 knockdown or overexpression on NSCLC cell proliferation in vitro was assessed by Cell Counting Kit-8, flow cytometry-based cell cycle, and colony formation assays. The effect of the USP5 inhibitor EOAI3402143 (G9) on NSCLC proliferation in vitro was analyzed by CCK-8 assay. The effect of G9 on NSCLC xenograft tumor growth was also examined in vivo, using athymic BALB/c nude mice. RESULTS: USP5 physically bound to CCND1 and decreased its polyubiquitination level, thereby stabilizing CCND1 protein. This USP5-CCND1 axis promoted NSCLC cell proliferation and colony formation. Further, knockdown of USP5 led to CCND1 degradation and cell cycle arrest in NSCLC cells. Importantly, this tumor-suppressive effect elicited by USP5 knockdown in NSCLC cells was validated in vitro and in vivo through chemical inhibition of USP5 activity using G9. Consistently, G9 downregulated the protein levels of CCND1 in NSCLC cells and xenograft tumor tissues. Also, the expression level of USP5 was positively associated with the protein level of CCND1 in human clinical NSCLC tissues. CONCLUSIONS: This study has provided the first evidence that CCND1 is a novel substrate of USP5. The USP5-CCND1 axis could be a potential target for the treatment of NSCLC.

Caenorhabditis elegans as a Model System for Discovering Bioactive Compounds Against Polyglutamine-Mediated Neurotoxicity.

Age-related misfolding and aggregation of pathogenic proteins are responsible for several neurodegenerative diseases. For example, Huntington's disease (HD) is principally driven by a CAG nucleotide repeat that encodes an expanded glutamine tract in huntingtin protein. Thus, the inhibition of polyglutamine (polyQ) aggregation and, in particular, aggregation-associated neurotoxicity is a useful strategy for the prevention of HD and other polyQ-associated conditions. This paper introduces generalized experimental protocols to assess the neuroprotective capacity of test compounds against HD using established polyQ transgenic Caenorhabditis elegans models. The AM141 strain is chosen for the polyQ aggregation assay as an age-associated phenotype of discrete fluorescent aggregates can be easily observed in its body wall at the adult stage due to muscle-specific expression of polyQ::YFP fusion proteins. In contrast, the HA759 model with strong expression of polyQ-expanded tracts in ASH neurons is used to examine neuronal death and chemoavoidance behavior. To comprehensively evaluate the neuroprotective capacity of target compounds, the above test results are ultimately presented as a radar chart with profiling of multiple phenotypes in a manner of direct comparison and direct viewing.

Current clinical developments in curcumin-based therapeutics for cancer and chronic diseases.

The last decade has seen an unprecedented rise in the prevalence of chronic diseases worldwide. Different mono-targeted approaches have been devised to treat these multigenic diseases, still most of them suffer from limited success due to the off-target debilitating side effects and their inability to target multiple pathways. Hence a safe, efficacious, and multi-targeted approach is the need for the hour to circumvent these challenging chronic diseases. Curcumin, a natural compound extracted from the rhizomes of Curcuma longa, has been under intense scrutiny for its wide medicinal and biological properties. Curcumin is known to manifest antibacterial, antiinflammatory, antioxidant, antifungal, antineoplastic, antifungal, and proapoptotic effects. A plethora of literature has already established the immense promise of curcuminoids in the treatment and clinical management of various chronic diseases like cancer, cardiovascular, metabolic, neurological, inflammatory, and infectious diseases. To date, more than 230 clinical trials have opened investigations to understand the pharmacological aspects of curcumin in human systems. Still, further randomized clinical studies in different ethnic populations warrant its transition to a marketed drug. This review summarizes the results from different clinical trials of curcumin-based therapeutics in the prevention and treatment of various chronic diseases.

The deubiquitinase USP10 restores PTEN activity and inhibits non-small cell lung cancer cell proliferation.

The phosphatase and tensin homolog deleted on chromosome 10 (PTEN) protein is a key player in tumorigenesis of non-small cell lung cancer (NSCLC) and was recently found to be inactivated by tripartite motif containing 25 (TRIM25)-mediated K63-linked polyubiquitination. However, the deubiquitinase (Dub) coordinate TRIM25 in PTEN ubiquitination is still elusive. In the present study, we found that this K63-linked polyubiquitination could be ablated by the ubiquitin-specific protease 10 (USP10) in a screen against a panel of Dubs. We found using coimmununoprecipitation/immunoblotting that USP10 interacted with PTEN and reduced the K63-linked polyubiquitination of PTEN mediated by TRIM25 in NSCLC cells. Moreover, USP10, but not its inactive C424A deubiquitinating mutant or other Dubs, abolished PTEN from K63-linked polyubiquitination mediated by TRIM25. In contrast to TRIM25, USP10 restored PTEN phosphatase activity and reduced the production of the secondary messenger phosphatidylinositol-3,4,5-trisphosphate, thereby inhibiting AKT/mammalian target of rapamycin progrowth signaling transduction in NSCLC cells. Moreover, USP10 was downregulated in NSCLC cell lines and primary tissues, whereas TRIM25 was upregulated. Consistent with its molecular activity, re-expression of USP10 suppressed NSCLC cell proliferation and migration, whereas knockout of USP10 promoted NSCLC cell proliferation and migration. In conclusion, the present study demonstrates that USP10 coordinates TRIM25 to modulate PTEN activity. Specifically, USP10 activates PTEN by preventing its K63-linked polyubiquitination mediated by TRIM25 and suppresses the AKT/mammalian target of rapamycin signaling pathway, thereby inhibiting NSCLC proliferation, indicating that it may be a potential drug target for cancer treatment.

Suppression of USP7 induces BCR-ABL degradation and chronic myelogenous leukemia cell apoptosis.

Chronic myelogenous leukemia (CML) is a clonal malignancy of hematopoietic stem cells featured with the fusion protein kinase BCR-ABL. To elicit the mechanism underlying BCR-ABL stability, we perform a screen against a panel of deubiquitinating enzymes (DUBs) and find that the ubiquitin-specific protease 7 (USP7) drastically stabilizes the BCR-ABL fusion protein. Further studies show that USP7 interacts with BCR-ABL and blocks its polyubiquitination and degradation. Moreover, USP7 knockdown triggers BCR-ABL degradation and suppresses its downstream signaling transduction. In line with this finding, genetic or chemical inhibition of USP7 leads to BCR-ABL protein degradation, suppresses BCR/ABL signaling, and induces CML cell apoptosis. Furthermore, we find the antimalarial artesunate (ART) significantly inhibits USP7/BCR-ABL interaction, thereby promoting BCR-ABL degradation and inducing CML cell death. This study thus identifies USP7 as a putative Dub of BCR-ABL and provides a rationale in targeting USP7/BCR-ABL for the treatment of CML.

Inhibition of the Otub1/c-Maf axis by the herbal acevaltrate induces myeloma cell apoptosis.

BACKGROUND: The oncogenic transcript factor c-Maf is stabilized by the deubiquitinase Otub1 and promotes myeloma cell proliferation and confers to chemoresistance. Inhibition of the Otub1/c-Maf axis is a promising therapeutic target, but there are no inhibitors reported on this specific axis. METHODS: A luciferase assay was applied to screen potential inhibitors of Otub1/c-Maf. Annexin V staining/flow cytometry was applied to evaluate cell apoptosis. Immunoprecipitation was applied to examine protein ubiquitination and interaction. Xenograft models in nude mice were used to evaluate anti-myeloma activity of AVT. RESULTS: Acevaltrate (AVT), isolated from Valeriana glechomifolia, was identified based on a bioactive screen against the Otub1/c-Maf/luciferase system. AVT disrupts the interaction of Otub1/c-Maf thus inhibiting Otub1 activity and leading to c-Maf polyubiquitination and subsequent degradation in proteasomes. Consistently, AVT inhibits c-Maf transcriptional activity and downregulates the expression of its target genes key for myeloma growth and survival. Moreover, AVT displays potent anti-myeloma activity by triggering myeloma cell apoptosis in vitro and impairing myeloma xenograft growth in vivo but presents no marked toxicity. CONCLUSIONS: The natural product AVT inhibits the Otub1/c-Maf axis and displays potent anti-myeloma activity. Given its great safety and efficacy, AVT could be further developed for MM treatment. Video Abstract.

Targeting the Otub1/c-Maf axis for the treatment of multiple myeloma.

The oncogenic transcription factor c-Maf has been proposed as an ideal therapeutic target for multiple myeloma (MM), but how to achieve it is still elusive. In the present study, we found the Otub1/c-Maf axis could be a potential target. Otub1, an OTU family deubiquitinase, was found to interact with c-Maf by mass spectrometry. Otub1 abrogates c-Maf K48-linked polyubiquitination, thus preventing its degradation and enhancing its transcriptional activity. Specifically, this deubiquitinating activity depends on its Lys71 and the N terminus but is independent of UBE2O, a known E2 of c-Maf. Otub1 promotes MM cell survival and MM tumor growth. In contrast, silence of Otub1 leads to c-Maf degradation and c-Maf-expressing MM cell apoptosis. Therefore, the Otub1/c-Maf axis could be a therapeutic target of MM. In order to explore this concept, we performed a c-Maf recognition element-driven luciferase-based screen against US Food and Drug Administration-approved drugs and natural products, from which the generic cardiac glycoside lanatoside C (LanC) is found to prevent c-Maf deubiquitination and induces its degradation by disrupting the interaction of Otub1 and c-Maf. Consequently, LanC inhibits c-Maf transcriptional activity, induces c-Maf-expressing MM cell apoptosis, and suppresses MM growth and prolongs overall survival of model mice, but without apparent toxicity. Therefore, the present study identifies Otub1 as a novel deubiquitinase of c-Maf and establishes that the Otub1/c-Maf axis is a potential therapeutic target for MM.