Publications by authors named "Hongbiao Huang"

59 Publications

USP1-dependent RPS16 protein stability drives growth and metastasis of human hepatocellular carcinoma cells.

J Exp Clin Cancer Res 2021 Jun 21;40(1):201. Epub 2021 Jun 21.

Institute of Digestive Disease of Guangzhou Medical University, The Sixth Affiliated Hospital of Guangzhou Medical University, Qingyuan People's Hospital, 511500, Qingyuan, Guangdong, China.

Background: Hepatocellular carcinoma (HCC) remains a medical challenge due to its high proliferation and metastasis. Although deubiquitinating enzymes (DUBs) play a key role in regulating protein degradation, their pathological roles in HCC have not been fully elucidated.

Methods: By using biomass spectrometry, co-immunoprecipitation, western blotting and immunofluorescence assays, we identify ribosomal protein S16 (RPS16) as a key substrate of ubiquitin-specific peptidase 1 (USP1). The role of USP1-RPS16 axis in the progression of HCC was evaluated in cell cultures, in xenograft mouse models, and in clinical observations.

Results: We show that USP1 interacts with RPS16. The depletion of USP1 increases the level of K48-linked ubiquitinated-RPS16, leading to proteasome-dependent RPS16 degradation. In contrast, overexpression of USP1-WT instead of USP1-C90A (DUB inactivation mutant) reduces the level of K48-linked ubiquitinated RPS16, thereby stabilizing RPS16. Consequently, USP1 depletion mimics RPS16 deficiency with respect to the inhibition of growth and metastasis, whereas transfection-enforced re-expression of RPS16 restores oncogenic-like activity in USP1-deficient HCC cells. Importantly, the high expression of USP1 and RPS16 in liver tissue is a prognostic factor for poor survival of HCC patients.

Conclusions: These findings reveal a previously unrecognized role for the activation of USP1-RPS16 pathway in driving HCC, which may be further developed as a novel strategy for cancer treatment.
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http://dx.doi.org/10.1186/s13046-021-02008-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8215741PMC
June 2021

A new role of GRP75-USP1-SIX1 protein complex in driving prostate cancer progression and castration resistance.

Oncogene 2021 Jun 2;40(25):4291-4306. Epub 2021 Jun 2.

Affiliated Cancer Hospital & institute of Guangzhou Medical University, Guangzhou, Guangdong, China.

Prostate cancer (PC) is the second most common cancer with limited treatment option in males. Although the reactivation of embryonic signals in adult cells is one of the characteristics of cancer, the underlying protein degradation mechanism remains elusive. Here, we show that the molecular chaperone GRP75 is a key player in PC cells by maintaining the protein stability of SIX1, a transcription factor for embryonic development. Mechanistically, GRP75 provides a platform to recruit the deubiquitinating enzyme USP1 to inhibit K48-linked polyubiquitination of SIX1. Structurally, the C-terminus of GRP75 (433-679 aa) contains a peptide binding domain, which is required for the formation of GRP75-USP1-SIX1 protein complex. Functionally, pharmacological or genetic inhibition of the GRP75-USP1-SIX1 protein complex suppresses tumor growth and overcomes the castration resistance of PC cells in vitro and in xenograft mouse models. Clinically, the protein expression of SIX1 in PC tumor tissues is positively correlated with the expression of GRP75 and USP1. These new findings not only enhance our understanding of the protein degradation mechanism, but also may provide a potential way to enhance the anti-cancer activity of androgen suppression therapy.
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http://dx.doi.org/10.1038/s41388-021-01851-0DOI Listing
June 2021

Bioinformatics identification of hub genes and signaling pathways regulated by intravenous immunoglobulin treatment in acute Kawasaki disease.

Exp Ther Med 2021 Jul 19;22(1):784. Epub 2021 May 19.

Institute of Pediatric Research, Children's Hospital of Soochow University, Suzhou, Jiangsu 215025, P.R. China.

Kawasaki disease (KD) is an acute, self-limiting form of vasculitis commonly encountered in infants and young children. Intravenous immunoglobulin (IVIG) is the primary drug used for the treatment of KD, which may significantly reduce the occurrence of coronary artery lesions. However, the specific molecular profile changes of KD caused by IVIG treatment have remained elusive and require further research. The present study was designed to identify key genes, pathways and immune cells affected by IVIG treatment using multiple bioinformatics analysis methods. The results suggested that myeloid cells and neutrophils were affected by IVIG treatment. Kyoto Encyclopedia of Genes and Genomes pathway analysis identified that hematopoietic cell lineages and osteoclast differentiation may have an important role in the mechanism of action of IVIG treatment. Immune cell analysis indicated that the levels of monocytes, M1 macrophages, neutrophils and platelets were markedly changed in patients with KD after vs. prior to IVIG treatment. The key upregulated genes, including ZW10 interacting kinetochore protein, GINS complex subunit 1 and microRNA-30b-3p in whole blood cells of patients with KD following treatment with IVIG indicated that these IVIG-targeted molecules may have important roles in KD. In addition, these genes were further examined by literature review and indicated to be involved in cell proliferation, apoptosis and virus-related immune response in patients with KD. Therefore, the present results may provide novel insight into the mechanisms of action of IVIG treatment for KD.
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http://dx.doi.org/10.3892/etm.2021.10216DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8145699PMC
July 2021

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

Cell Death Dis 2021 May 7;12(5):456. Epub 2021 May 7.

Department of Hematology, the First Affiliated Hospital of Guangzhou Medical University, Guangzhou, 510120, China.

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.
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http://dx.doi.org/10.1038/s41419-021-03732-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8105359PMC
May 2021

Novel variant in BRAT1 with the lethal neonatal rigidity and multifocal seizure syndrome.

Pediatr Res 2021 Mar 31. Epub 2021 Mar 31.

Institute of Pediatric Research, Children's Hospital of Soochow University, Suzhou, Jiangsu Province, People's Republic of China.

Background: Lethal neonatal rigidity and multifocal seizure syndrome (RMFSL) is caused by variants in BRAT1 (BRCA1-associated protein required for ATM activation-1). However, the molecular mechanism of RMFSL is still unclear.

Methods: An RMFSL infant was recruited and the peripheral blood samples from his trio-family were collected. The genomic DNA was extracted, and then the whole-exome sequencing was performed. The expression of BRAT1 was analyzed by Western blotting. The subcellular localization of BRAT1 and MitoSOX (mitochondrial superoxide level) was investigated by confocal microscopy. The RNA samples were obtained from transfected cells, and then the RNA sequencing was performed.

Results: In this study, a novel homozygous BRAT1 variant c.233G > C with amino acid change of R with P at residue 78 (R78P) was identified. This variant altered the peptide structure and subcellular localization, as well as the expression in vitro. However, R78P did not alter the ability of BRAT1 to downregulate MitoSOX in mitochondria. Meanwhile, R78P BRAT1 was positively correlated with temporal lobe epilepsy, autosomal recessive primary microcephaly, defective/absent horizontal voluntary eye movements, and neuron apoptotic process as indicated by gene set enrichment analysis (GSEA).

Conclusions: The BRAT1 variant spectrum has been expanded, which will be helpful for genetic counseling. We also explored the molecular mechanism altered by R78P, which will provide a better understanding of the pathogenesis of RMFSL.

Impact: The detailed course of an infant with lethal neonatal RMFSL was depicted. A novel disease-causing variant R78P in BRAT1 for lethal neonatal RMFSL was identified. R78P led to reduced BRAT1 expression and nuclear localization in vitro. R78P did not alter the ability of BRAT1 to downregulate MitoSOX in the mitochondria. The variant R78P in BRAT1 was positively correlated with temporal lobe epilepsy, autosomal recessive primary microcephaly, defective/absent horizontal voluntary eye movements, and neuron apoptotic process as indicated by GSEA.
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http://dx.doi.org/10.1038/s41390-021-01468-9DOI Listing
March 2021

The deubiquitinating enzyme USP15 stabilizes ERα and promotes breast cancer progression.

Cell Death Dis 2021 03 26;12(4):329. Epub 2021 Mar 26.

Affiliated Cancer Hospital & institute of Guangzhou Medical University, Guangzhou, 510095, China.

Breast cancer has the highest incidence and mortality in women worldwide. There are 70% of breast cancers considered as estrogen receptor α (ERα) positive. Therefore, the ERα-targeted therapy has become one of the most effective solution for patients with breast cancer. Whereas a better understanding of ERα regulation is critical to shape evolutional treatments for breast cancer. By exploring the regulatory mechanisms of ERα at levels of post-translational modifications, we identified the deubiquitinase USP15 as a novel protector for preventing ERα degradation and a critical driver for breast cancer progression. Specifically, we demonstrated that USP15 promoted the proliferation of ERα, but not ERα breast cancer, in vivo and in vitro. Meanwhile, USP15 knockdown notably enhanced the antitumor activities of tamoxifen on breast cancer cells. Importantly, USP15 knockdown induced the downregulation of ERα protein via promoting its K48-linked ubiquitination, which is required for proliferative inhibition of breast cancer cells. These findings not only provide a novel treatment for overcoming resistance to endocrine therapy, but also represent a therapeutic strategy on ERα degradation by targeting USP15-ERα axis.
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http://dx.doi.org/10.1038/s41419-021-03607-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7997968PMC
March 2021

Leukocyte proteomics coupled with serum metabolomics identifies novel biomarkers and abnormal amino acid metabolism in Kawasaki disease.

J Proteomics 2021 05 15;239:104183. Epub 2021 Mar 15.

Department of Cardiology, Children's Hospital of Soochow University, Suzhou 215025, China. Electronic address:

Kawasaki disease (KD) is a systemic vasculitis that can lead to severe cardiovascular complications, whereas the development and clinical usage of specific biomarkers might help diagnose KD and avoid certain complications. To this end, the molecular profiles of acute KD patients with coronary artery lesions (CAL) were first investigated through leukocyte proteomics and serum metabolomics assays. A total of 269 differentially abundant proteins and 35 differentially abundant metabolites with the top fold-changed levels were identified in acute KD patients compared to those in the healthy controls. Among them, several highly promising candidate marker proteins and metabolites indicative of KD progression were further analysed, such as the increased proteins ALPL, NAMPT, and S100P, as well as the decreased proteins C1QB and apolipoprotein family members. Moreover, metabolites, including succinic acid, dGMP, hyaluronic acid, L-tryptophan, propionylcarnitine, inosine, and phosphorylcholine, were found to be highly accurate at distinguishing between KD patients and healthy controls. Interestingly, the abnormal expression levels of a distinct set of proteins and metabolites in acute KD patients can be restored to normal levels upon intravenous immunoglobulin (IVIG) treatment. Overall, this work has revealed novel biomarkers and abnormal amino-acid metabolism as a prominent feature involved in KD patients with CAL. SIGNIFICANCE: KD is frequently concomitant with the development of life-threatening coronary vasculitis. Here, the profiles of leukocyte proteomics and serum metabolomics in acute KD patients with CALs were first investigated, and several hub molecules identified here could be used as supplemental biomarkers for KD diagnosis. Moreover, the metabolomic abnormalities especially the amino acids are particularly prominent in KD patients. Interestingly, the abnormal expression levels of a distinct set of proteins and metabolites in acute KD patients can be restored to normal levels upon IVIG treatment. Therefore, these findings might help understand the IVIG activities and also the underlying mechanisms of IVIG-resistant patients, thereby providing a new perspective for the exploration of mechanisms related to KD.
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http://dx.doi.org/10.1016/j.jprot.2021.104183DOI Listing
May 2021

ERα is a target for butein-induced growth suppression in breast cancer.

Am J Cancer Res 2020 1;10(11):3721-3736. Epub 2020 Nov 1.

Affiliated Cancer Hospital & Institute of Guangzhou Medical University Guangzhou 510095, Guangdong, China.

Breast cancer (BCa) has the highest incidence and mortality among malignant diseases in female worldwide. BCa is frequently caused by estrogen receptor α (ERα), a ligand-dependent receptor that highly expressed in about 70% of breast tumors. Therefore, ERα has become a well-characterized and the most effective target for treating ERα-expressing BCa (ERα BCa). However, the acquire resistance was somehow developed in patients who received current ERα signaling-targeted endocrine therapies. Hence, discovery of novel anti-estrogen/ERα strategies is urgent. In the present study, we identified butein as a potential agent for breast cancer treatment by the use of a natural product library. We showed that butein inhibits the growth of ERα BCa both and which is associated with cell cycle arrest that partially triggered by butein-induced ERα downregulation. Mechanically, butein binds to a specific pocket of ERα and promotes proteasome-mediated degradation of the receptor. Collectively, this work reveals that butein is a candidate to diminish ERα signaling which represents a potentially novel strategy for treating BCa.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7716169PMC
November 2020

USP10 deletion inhibits macrophage-derived foam cell formation and cellular-oxidized low density lipoprotein uptake by promoting the degradation of CD36.

Aging (Albany NY) 2020 11 10;12(22):22892-22905. Epub 2020 Nov 10.

Guangzhou Institute of Cardiovascular Disease, Guangdong Key Laboratory of Vascular Diseases, State Key Laboratory of Respiratory Disease, The Second Affiliated Hospital, Guangzhou Medical University, Guangzhou, Guangdong, China.

Foam cell formation process is involved in the pathogenesis of atherosclerosis (AS). Activation of this biological process depends on lipid uptake by scavenger receptors, such as CD36, SR-A and SR-B1. Among these receptors, CD36 is the principal one because it dominates roughly 50% lipid uptake in monocytes. In this study, our western blotting and RT-qPCR assays revealed that USP10 inhibition promotes the degradation of CD36 protein but does not change its mRNA level. In addition, Co-IP results showed that USP10 interacts with CD36 and stabilizes CD36 protein by cleaving poly-ubiquitin on CD36. Significantly, USP10 promotes foam cell formation. Immunofluorescence and Oil red O staining assays show that inhibition or knockdown of USP10 suppresses lipid uptake and foam cell formation by macrophages. In conclusion, USP10 promotes the development and progression of atherosclerosis through stabilizing CD36 protein expression. The regulation of USP10-CD36 may provide a significant therapeutic scheme in atherosclerosis.
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http://dx.doi.org/10.18632/aging.104003DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7746336PMC
November 2020

Deubiquitination of CD36 by UCHL1 promotes foam cell formation.

Cell Death Dis 2020 08 15;11(8):636. Epub 2020 Aug 15.

Guangzhou Institute of Cardiovascular Disease, Guangdong Key Laboratory of Vascular Diseases, State Key Laboratory of Respiratory Disease, The Second Affiliated Hospital of Guangzhou Medical University, 510260, Guangzhou, China.

Atherosclerosis-associated cardiovascular diseases are main causes leading to high mortality worldwide. Macrophage-derived foam cell formation via uptaking modified lipoproteins is the initial and core step in the process of atherosclerosis. Meanwhile, scavenger receptor is indispensable for the formation of foam cells. UCHL1, a deubiquitinase, has been widely studied in multiple cancers. UCHL1 could be an oncogene or a tumor suppressor in dependent of tumor types. It remains unknown whether UCHL1 influences cellular oxLDL uptake. Herein we show that UCHL1 deletion significantly inhibits lipid accumulation and foam cell formation. Subsequently, we found that UCHL1 inhibitor or siRNA downregulates the expression of CD36 protein whereas SR-A, ABCA1, ABCG1, Lox-1, and SR-B1 have no significant change. Furthermore, the treatment of UCHL1 inhibition increases the abundance of K48-polyubiquitin on CD36 and the suppression of lipid uptake induced by UCHL1 deficiency is attenuated by blocking CD36 activation. Our study concluded that UCHL1 deletion decreases foam cell formation by promoting the degradation of CD36 protein, indicating UCHL1 may be a potential target for atherosclerosis treatment.
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http://dx.doi.org/10.1038/s41419-020-02888-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7429868PMC
August 2020

NCAPG confers trastuzumab resistance via activating SRC/STAT3 signaling pathway in HER2-positive breast cancer.

Cell Death Dis 2020 07 18;11(7):547. Epub 2020 Jul 18.

State Key Laboratory of Oncology in South China, Collaborative Innovation Center for Cancer Medicine, Sun Yat-sen University Cancer Center, 510060, Guangzhou, China.

HER2+ breast cancer (BC) is characterized by rapid growth, early recurrence, early metastasis, and chemoresistance. Trastuzumab is the most effective treatment for HER2+ BC and effectively reduces the risk of recurrence and death of patients. Resistance to trastuzumab results in cancer recurrence and metastasis, leading to poor prognosis of HER2+ BC. In the present study, we found that non-structural maintenance of chromosome condensin 1 complex subunit G (NCAPG) expression was highly upregulated in trastuzumab-resistant HER2+ BC. Ectopic NCAPG was positively correlated with tumor relapse and shorter survival in HER2+ BC patients. Moreover, overexpression of NCAPG promoted, while silencing of NCAPG reduced, the proliferative and anti-apoptotic capacity of HER2+ BC cells both in vitro and in vivo, indicating NCAPG reduces the sensitivity of HER2+ BC cells to trastuzumab and may confer trastuzumab resistance. Furthermore, our results suggest that NCAPG triggers a series of biological cascades by phosphorylating SRC and enhancing nuclear localization and activation of STAT3. To summarize, our study explores a crucial role for NCAPG in trastuzumab resistance and its underlying mechanisms in HER2+ BC, and suggests that NCAPG could be both a potential prognostic marker as well as a therapeutic target to effectively overcome trastuzumab resistance.
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http://dx.doi.org/10.1038/s41419-020-02753-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7368860PMC
July 2020

Targeting SKP2/Bcr-Abl pathway with Diosmetin suppresses chronic myeloid leukemia proliferation.

Eur J Pharmacol 2020 Sep 15;883:173366. Epub 2020 Jul 15.

Affiliated Cancer Hospital & Institute of Guangzhou Medical University, Guangzhou, 510095, China; Guangzhou Municipal and Guangdong Provincial Key Laboratory of Protein Modification and Degradation, School of Basic Medical Sciences, Guangzhou Medical University, Guangzhou, 511436, China. Electronic address:

Bcr-Abl is the primary cause as well as currently key therapeutic target of chronic myeloid leukemia (CML). SKP2, an E3 ligase, is a downstream factor of Bcr-Abl to motivate the cell cycle transition of CML and also found to bind and activate Bcr-Abl in reverse. Therefore, SKP2/Bcr-Abl pathway is an attractive target for CML treatment. This study aims to identify an inhibitor of the SKP2/Bcr-Abl pathway based on a large screening of the natural products. We demonstrate that Diosmetin, a kind of phytoestrogens, notably downregulates the expression of SKP2, Bcr-Abl phosphorylation, and moderately downregulates the Bcr-Abl level. Furthermore, Diosmetin displays a favorable anti-tumor activity in CML cells and xenograft models. Collectively, our study reveals a natural compound in the treatment of CML on the basis of SKP2/Bcr-Abl signaling pathway.
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http://dx.doi.org/10.1016/j.ejphar.2020.173366DOI Listing
September 2020

Targeting ERα degradation by L-Tetrahydropalmatine provides a novel strategy for breast cancer treatment.

Int J Biol Sci 2020 18;16(12):2192-2204. Epub 2020 May 18.

Affiliated Cancer Hospital & institute of Guangzhou Medical University, Guangzhou Municipal and Guangdong Provincial Key Laboratory of Protein Modification and Degradation, School of Basic Medical Sciences, Guangzhou, Guangdong 511436, China.

The incidence and mortality of breast cancer (BCa) are the highest among female cancers. There are approximate 70% BCa that are classified as estrogen receptor alpha (ERα) positive. Therefore, targeting ERα is the most significantly therapeutic schedule. However, patients with breast cancer develop resistance to ERα or estrogen (E2) antagonists such as fulvestrant and tamoxifen. In the present study, we found that L-Tetrahydropalmatine (L-THP) significantly suppressed cell proliferation in ERα BCa cells via inducing cell cycle arrest rather than apoptosis. Additionally, L-THP enhanced the sensitivity of ERα BCa cells to tamoxifen and fulvestrant. Mechanically, the application of L-THP promotes ERα degradation through accumulating ubiquitin chains on ERα. Overexpressing ERα abrogates L-THP induced-antiproliferation in ERα BCa cells. Collectively, our work indicates that L-THP may represent a potentially novel therapeutic medicine for ERα breast cancer patient.
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http://dx.doi.org/10.7150/ijbs.44005DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7294940PMC
May 2020

Targeting GRP78-dependent AR-V7 protein degradation overcomes castration-resistance in prostate cancer therapy.

Theranostics 2020 10;10(8):3366-3381. Epub 2020 Feb 10.

Affiliated Cancer Hospital & institute of Guangzhou Medical University, Protein Modification and Degradation Key Lab of Guangzhou and Guangdong, State Key Laboratory of Respiratory Disease, School of Basic Medical Sciences, Guangzhou Medical University, Guangzhou, Guangdong 510095, China.

: Androgen receptor splice variant 7 (AR-V7) is a leading cause of the development of castration-resistant prostate cancer (CRPC). However, the regulation and function of AR-V7 at levels of post-translational modifications in prostate cancer therapy remain poorly understood. Here, we conducted a library screen of natural products to identify potential small molecules responsible for AR-V7 protein degradation in human prostate cancer cell lines. : A natural product library was used to screen the inhibitor of AR-V7. Co-IP and biomass spectrum assays were used to identify the AR-V7-interacting proteins, whereas western blot, confocal microscopy, RNA interfering, and gene transfection were used to validate these interactions. Cell viability, EDU staining, and colony formation assays were employed to detect cell growth and proliferation. Flowcytometry assays were used to detect the distribution of cell cycle. Mouse xenograft models were used to study the anti-CRPC effects . : This screen identified rutaecarpine, one of the major components of the Chinese medicine , as a novel chemical that selectively induces AR-V7 protein degradation via K48-linked ubiquitination. Mechanically, this effect relies on rutaecarpine inducing the formation of a GRP78-AR-V7 protein complex, which further recruits the E3 ligase SIAH2 to directly promote the ubiquitination of AR-V7. Consequently, the genetic and pharmacological activation of the GRP78-dependent AR-V7 protein degradation restores the sensitivity of castration-resistant prostate cancer to anti-androgen therapy in cell culture and animal models. : These findings not only provide a new approach for overcoming castration-resistance in prostate cancer therapy, but also increase our understanding about the interplay between molecular chaperones and ubiquitin ligase in shaping protein stability.
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http://dx.doi.org/10.7150/thno.41849DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7069092PMC
April 2021

Short Term Exposure to Bilirubin Induces Encephalopathy Similar to Alzheimer's Disease in Late Life.

J Alzheimers Dis 2020 ;73(1):277-295

Guangzhou Municipal and Guangdong Provincial Key Laboratory of Protein Modification and Degradation, State Key Lab of Respiratory Disease, School of Basic Medical Sciences, Affiliated Cancer Hospital of Guangzhou Medical University, Guangdong, People's Republic of China.

Hyperbilirubinemia may increase the risk of Alzheimer's disease (AD) but its mechanistic role in AD pathogenesis remains obscure. Here, we used animal models to investigate the short- and long-term effects of neonatal systemic exposure to bilirubin on brain histology and function as well as the acute effect of lateral ventricle injection of bilirubin in adult rats. We found that three days exposure to bilirubin in newborn rats could induce AD-like pathological changes in late life, including tau protein hyperphosphorylation at multiple sites, increased Aβ production in brain tissues, and spatial learning and memory injury. Bilirubin activated the activities of several protein kinases (GSK-3β, CDK5, and JNK), which were positively correlated with hyperphosphorylated tau; simultaneously increased the expression of AβPP γ-secretase PS2 and decreased the expression of α-secretase ADAM17, which were positively correlated with Aβ production. The above results were well replicated in primary hippocampal cell cultures. These data demonstrate that bilirubin encephalopathy is an AD-like disease, suggesting a potent role of bilirubin in AD.
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http://dx.doi.org/10.3233/JAD-190945DOI Listing
April 2021

Parkin facilitates proteasome inhibitor-induced apoptosis via suppression of NF-κB activity in hepatocellular carcinoma.

Cell Death Dis 2019 09 26;10(10):719. Epub 2019 Sep 26.

Affiliated Cancer Hospital & Institute of Guangzhou Medical University, Key Laboratory of Protein Modification and Degradation, State Key Laboratory of Respiratory Disease, School of Basic Medical Sciences, Guangzhou Medical University, 511436, Guangzhou, China.

The ubiquitin-proteasome system (UPS) is a tight homeostatic control mechanism of intracellular protein degradation and turnover involved in many human diseases. Proteasome inhibitors were initially developed as anticancer agents with potential benefits in the suppression of tumor growth. However, clinical trials of patients with solid tumors fail to demonstrate the same efficacy of these proteasome inhibitors. Here, we show that Parkin, an E3 ubiquitin ligase, is implicated in tumorigenesis and therapy resistance of hepatocellular carcinoma (HCC), the most common type of primary liver cancer in adults. Lower Parkin expression correlates with poor survival in patients with HCC. Ectopic Parkin expression enhances proteasome inhibitor-induced apoptosis and tumor suppression in HCC cells in vitro and in vivo. In contrast, knockdown of Parkin expression promotes apoptosis resistance and tumor growth. Mechanistically, Parkin promotes TNF receptor-associated factor (TRAF) 2 and TRAF6 degradation and thus facilitates nuclear factor-kappa-B (NF-κB) inhibition, which finally results in apoptosis. These findings reveal a direct molecular link between Parkin and protein degradation in the control of the NF-κB pathway and may provide a novel UPS-dependent strategy for the treatment of HCC by induction of apoptosis.
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http://dx.doi.org/10.1038/s41419-019-1881-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6763437PMC
September 2019

Deubiquitination and stabilization of estrogen receptor α by ubiquitin-specific protease 7 promotes breast tumorigenesis.

Cancer Lett 2019 11 10;465:118-128. Epub 2019 Sep 10.

Affiliated Cancer Hospital & Institute of Guangzhou Medical University, School of Basic Medical Sciences, Guangzhou Medical University, Guangzhou, Guangdong, 510095, China; Key Laboratory of Protein Modification and Degradation, School of Basic Medical Sciences, Guangzhou Medical University, Guangzhou, Guangdong, 511436, China; State Key Laboratory of Respiratory Disease, Guangzhou Medical University, Guangzhou, Guangdong, China. Electronic address:

Breast cancer is the most common malignancy in women around the world. Estrogen receptor α (ERα) is expressed in approximately 70% of breast tumors, and considered as one of most effective targets in breast cancer therapy. It has been reported that the degradation of ERα protein is mediated by ubiquitin-proteasome system. However, little is known about the regulation of ERα deubiquitination, a critical constituent of its degradation control. The current study first reports that there is a positive correlation between ERα and ubiquitin specific protease 7 (USP7) protein levels in human breast tumor tissues. Subsequent studies showed that USP7 physically interacted with the ERα, thereby mediating the deubiquitination and stabilization of ERα. In addition, USP7 inhibition or silencing led to growth inhibition and apoptosis of ERα-positive breast cancer cells both in vitro and in vivo. Furthermore, overexpression of ERα rescued the USP7 silencing-induced cell cycle arrest and apoptosis, supporting that ERα status is essential to the function of USP7 in breast carcinogenesis. Overall, this study suggests that targeting USP7-ERα complex could be a potential strategy to treat ERα-positive breast cancer.
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http://dx.doi.org/10.1016/j.canlet.2019.09.003DOI Listing
November 2019

Synergistic effects of gefitinib and thalidomide treatment on EGFR-TKI-sensitive and -resistant NSCLC.

Eur J Pharmacol 2019 Aug 25;856:172409. Epub 2019 May 25.

Affiliated Cancer Hospital and Institute of Guangzhou Medical University, Key Laboratory of Protein Modification and Degradation, State Key Laboratory of Respiratory Disease, School of Basic Medical Sciences, Guangzhou Medical University, Guangzhou, Guangdong, 510095, China. Electronic address:

EGF receptor tyrosine kinase inhibitors (EGFR-TKIs) have been widely used as a standard therapy in non-small cell lung cancer (NSCLC) patients with EGFR mutations. However, most if not all of the patients who initially have responded to EGFR-TKIs later experience progression or deterioration of the disease while still on the treatment. Drug resistance becomes inevitable due to the emergence of the second-site EGFR T790M mutation within exon 20, MET and HER2 amplification, small cell histologic transformation and rare secondary BRAF mutations. The acquired drug resistance limits the efficacy of EGFR-TKIs in patients. Thalidomide is a widely used anti-angiogenic and immunomodulatory drug with anticancer effects. The current study was aimed to explore the combined effects of gefitinib and thalidomide on NSCLC. The combination of thalidomide and gefitinib induced antiproliferative and proapoptotic effects in HCC827, PC9, and PC9GR cells. The inhibition of EGFR phosphorylation and downstream signaling was more pronounced in the thalidomide and gefitinib co-treatment group as compared with the single agent treatment groups. Further study revealed that the inhibitors of AKT, MEK/ERK, and p38 increased the antiproliferative and proapoptotic effects of the combined treatment of thalidomide and gefitinib. However, JNK inhibition moderately abrogated cell apoptosis induced by the co-treatment. In conclusion, thalidomide and gefitinib exhibit synergistic effects on both TKI-sensitive and -resistant NSCLC cells by targeting the EGFR signaling pathways, suggesting that the combination strategy is promising for the treatment of NSCLC.
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http://dx.doi.org/10.1016/j.ejphar.2019.172409DOI Listing
August 2019

Inhibition of USP14 enhances the sensitivity of breast cancer to enzalutamide.

J Exp Clin Cancer Res 2019 May 24;38(1):220. Epub 2019 May 24.

Affiliated Cancer Hospital and institute of Guangzhou Medical University; Key Laboratory of Protein Modification and Degradation; State Key Laboratory of Respiratory Disease, School of Basic Medical Sciences, Guangzhou Medical University, Guangzhou, 510095, Guangdong, China.

Background: Androgen receptor (AR) is expressed in approximately 70% of breast tumors. Recent studies increasingly support AR as a potential therapeutic target of AR-positive breast cancer. We have previously reported that deubiquitinase USP14 stabilizes AR proteins by deubiquitination and USP14 inhibition results in inhibition of cell growth and tumor progression in AR-positive prostate cancer and breast cancer. The current study aims to explore the anticancer effect of a treatment combining AR antagonist enzalutamide with USP14 inhibition on breast cancer cells.

Methods: The combining effects of enzalutamide and USP14 inhibition on breast cancer cell proliferation and apoptosis and associated cell signaling were evaluated in vitro and in vivo.

Results: USP14 inhibition via administration of IU1 or USP14-specific siRNA/shRNA enhanced cell growth inhibition and apoptosis induction by enzalutamide in breast cancer cell lines in vitro and in vivo. Additionally, the combination of enzalutamide with USP14 inhibition/knockdown induced significant downregulation of AR proteins and suppression of AR-related signaling pathways, including Wnt/β-catenin and PI3K/AKT pathways. Moreover, AKT inhibition via MK2206 increased the antiproliferative and proapoptotic effects of enzalutamide+IU1 combined treatment.

Conclusion: Collectively, our data suggest that USP14 inhibition in combination with enzalutamide represents a potentially new therapeutic strategy for breast cancer.
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http://dx.doi.org/10.1186/s13046-019-1227-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6534920PMC
May 2019

USP10 modulates the SKP2/Bcr-Abl axis via stabilizing SKP2 in chronic myeloid leukemia.

Cell Discov 2019 30;5:24. Epub 2019 Apr 30.

1Affiliated Cancer Hospital and institute of Guangzhou Medical University; Key Laboratory of Protein Modification and Degradation, State Key Laboratory of Respiratory Disease, School of Basic Medical Sciences, Guangzhou Medical University, Guangzhou, Guangdong 511436 China.

Constitutive activation of tyrosine kinase Bcr-Abl is the leading cause of the development and progression of chronic myeloid leukemia (CML). Currently, the application of tyrosine kinase inhibitors (TKIs) targeting the Bcr-Abl is the primary therapy for CML patients. However, acquired resistance to TKIs that develops overtime in the long-term administration renders TKIs ineffective to patients with advanced CML. Therefore, increasing studies focus on the amplified expression or activation of Bcr-Abl which is proposed to contribute to the advanced phase. Here, we show that S-phase kinase-associated protein 2 (SKP2) acts as a co-regulator of Bcr-Abl by mediating its K63-linked ubiquitination and activation. Further investigations show that USP10 as a novel deubiquitinase of SKP2 amplifies the activation of Bcr-Abl via mediating deubiquitination and stabilization of SKP2 in CML cells. Moreover, inhibition of USP10 significantly suppresses the proliferation of both imatinib-sensitive and imatinib-resistant CML cells, which likely depends on SKP2 status. This findings are confirmed in primary CML cells because these cells are over-expressed with USP10 and SKP2 and are sensitive to a USP10 inhibitor. Taken together, the present study not only provides a novel insight into the amplified activation of Bcr-Abl in CML, but also demonstrates that targeting the USP10/SKP2/Bcr-Abl axis is a potential strategy to overcome imatinib resistance in CML patients.
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http://dx.doi.org/10.1038/s41421-019-0092-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6488640PMC
April 2019

Overexpression of PIMREG promotes breast cancer aggressiveness via constitutive activation of NF-κB signaling.

EBioMedicine 2019 May 9;43:188-200. Epub 2019 Apr 9.

Affiliated Cancer Hospital & Institute of Guangzhou Medical University, Key Laboratory of Protein Modification and Degradation, State Key Laboratory of Respiratory Disease, School of Basic Medical Sciences, Guangzhou Medical University, Guangzhou 511436, China; Department of Biochemistry, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou 510080, China. Electronic address:

Background: It is well-established that activation of nuclear factor-kappa B (NF-κB) signaling plays important roles in cancer development and progression. However, the underlying mechanism by which the NF-κB pathway is constitutively activated in cancer remains largely unclear. The present study aimed to investigate the effect of PICALM interacting mitotic regulator (PIMREG) on sustaining NF-κB activation in breast cancer.

Methods: The underlying mechanisms in which PIMREG-mediated NF-κB constitutive activation were determined via immunoprecipitation, EMSA and luciferase reporter assays. The expression of PIMREG was examined by quantitative PCR and western blotting analyses and immunohistochemical assay. The effect of PIMREG on aggressiveness of breast cancer cell was measured using MTT, soft agar clonogenic assay, wound healing and transwell matrix penetration assays in vitro and a Xenografted tumor model in vivo.

Findings: PIMREG competitively interacted with the REL homology domain (RHD) of NF-κB with IκBα, and sustained NF-κB activation by promotion of nuclear accumulation and transcriptional activity of NF-κB via disrupting the NF-κB/IκBα negative feedback loop. PIMREG overexpression significantly enhanced NF-κB transactivity and promoted the breast cancer aggressiveness. The expression of PIMREG was markedly upregulated in breast cancer and positively correlated with clinical characteristics of patients with breast cancer, including the clinical stage, tumor-node-metastasis classification and poorer survival.

Interpretation: PIMREG promotes breast cancer aggressiveness via disrupting the NF-κB/IκBα negative feedback loop, which suggests that PIMREG might be a valuable prognostic factor and potential target for diagnosis and therapy of metastatic breast cancer. FUND: The science foundation of China, Guangdong Province, Guangzhou Education System, and the Science and Technology Program of Guangzhou.
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http://dx.doi.org/10.1016/j.ebiom.2019.04.001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6557765PMC
May 2019

Inhibition of EGFR signaling with Spautin-1 represents a novel therapeutics for prostate cancer.

J Exp Clin Cancer Res 2019 Apr 11;38(1):157. Epub 2019 Apr 11.

Affiliated Cancer Hospital and institute of Guangzhou Medical University; Key Laboratory of Protein Modification and Degradation; State Key Laboratory of Respiratory Disease, School of Basic Medical Sciences, Guangzhou Medical University, Guangzhou, 511436, Guangdong, China.

Background: Prostate cancer (PCa) remains a challenge worldwide. Due to the development of castration-resistance, traditional first-line androgen deprivation therapy (ADT) became powerlessness. Epidermal growth factor receptor (EGFR) is a well characterized therapeutic target to treat colorectal carcinoma and non-small cell lung cancer. Increasing studies have unraveled the significance of EGFR and its downstream signaling in the progression of castration-resistant PCa.

Method: MTS, colony formation and Edu staining assays were used to analyze the cell proliferation of PCa cells. Flow cytometry was used to analyze PCa cell cycle distribution and cell apoptosis. Western blot was used to measure the expression of key proteins associated with cell cycle progression, apoptosis and EGFR signaling pathways. Transfection of exogenous small interfering RNA (siRNA) or plasmid was used to intervene specific gene expression. Nude mouse model was employed to test the in vivo effect of Spautin-1.

Results: The current study reveals that Spautin-1, a known inhibitor of ubiquitin-specific peptidase 10 (USP10) and USP13, inhibits EGFR phosphorylation and the activation of its downstream signaling. Inhibition of EGFR signaling induced by Spautin-1 leads to cell cycle arrest and apoptosis of PCa in a USP10/USP13 independent manner. The application of Spautin-1 reduces the expression of glucose transporter 1 (Glut1) and dramatically induces cell death under glucose deprivation condition. In vivo experiments show a potent anti-tumor effect of Spautin-1 alone and in combination with Enzalutamide.

Conclusion: This study demonstrates the therapeutic potential of EGFR signaling inhibition by the use of Spautin-1 for PCa treatment.
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http://dx.doi.org/10.1186/s13046-019-1165-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6460657PMC
April 2019

MiR-454-3p-Mediated Wnt/β-catenin Signaling Antagonists Suppression Promotes Breast Cancer Metastasis.

Theranostics 2019 1;9(2):449-465. Epub 2019 Jan 1.

Affiliated Cancer Hospital & Institute of Guangzhou Medical University; Key Laboratory of Protein Modification and Degradation; State Key Laboratory of Respiratory Disease, School of Basic Medical Sciences, Guangzhou Medical University, Guangzhou 511436, China.

The Wnt/β-catenin pathway is constitutively active and promotes multiple tumor processes, including breast cancer metastasis. However, the underlying mechanism by which the Wnt/β-catenin pathway is constitutively activated in breast cancer metastasis remains unclear. Inhibition of Wnt antagonists is important for Wnt/β-catenin signaling activation, and post-transcriptional regulation of these antagonists by microRNAs (miRNAs) might be a possible mechanism underlying signaling activation. Regulation of nuclear pre-mRNA domain-containing 1A (RPRD1A) is a known inhibitor of cell growth and Wnt/β-catenin signaling activity, but the function and regulatory mechanism of RPRD1A in breast cancer have not been clarified. The aim of this study was to understand how regulators of the Wnt/β-catenin pathway may play a role in the metastasis of this cancer. RPRD1A expression and its association with multiple clinicopathological characteristics was analyzed immunohistochemically in human breast cancer specimens. miR-454-3p expression was analyzed using real-time PCR. RPRD1A or miR-454-3p knockdown and overexpression were used to determine the underlying mechanism of their functions in breast cancer cells. Xenografted tumor model, 3D invasive culture, cell migration and invasion assays and sphere formation assay were used to determine the biofunction of RPRD1A and miR-454-3p in breast cancer. Electrophoretic mobility shift assay (EMSA), luciferase reporter assay, and RNA immunoprecipitation (RIP) were performed to study the regulation and underlying mechanisms of RPRD1A and miR-454-3p expression and their correlation with the Wnt/β-catenin pathway in breast cancer. The Wnt/β-catenin signaling antagonist RPRD1A was downregulated and its upstream regulator miR-454-3p was amplified and overexpressed in metastatic breast cancer, and both were correlated with overall and relapse-free survival in breast cancer patients. The suppression by miR-454-3p on RPRD1A was found to activate Wnt/β-catenin signaling, thereby promoting metastasis. Simultaneously, three other negative regulators of the Wnt/β-catenin pathway, namely, AXIN2, dickkopf WNT signaling pathway inhibitor (DKK) 3 and secreted frizzled related protein (SFRP) 1, were also found to be targets of miR-454-3p and were involved in the signaling activation. miR-454-3p was found to be involved in early metastatic processes and to promote the stemness of breast cancer cells and early relapse under both and conditions. The findings indicate that miR-454-3p-mediated suppression of Wnt/β-catenin antagonist RPRD1A, as well as AXIN2, DKK3 and SFRP1, sustains the constitutive activation of Wnt/β-catenin signaling; thus, miR-454-3p and RPRD1A might be potential diagnostic and therapeutic targets for breast cancer metastasis.
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http://dx.doi.org/10.7150/thno.29055DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6376193PMC
December 2019

Auranofin lethality to prostate cancer includes inhibition of proteasomal deubiquitinases and disrupted androgen receptor signaling.

Eur J Pharmacol 2019 Mar 9;846:1-11. Epub 2019 Jan 9.

Key Laboratory of Protein Modification and Degradation, State Key Laboratory of Respiratory Disease, School of Basic Medical Sciences; Affiliated Cancer Hospital and institute of Guangzhou Medical University, Guangzhou, Guangdong 511436, China. Electronic address:

Auranofin (Aur) inhibits thioredoxin reductases and is also an inhibitor of 19S proteasome associated deubiquitinases, targeting USP14 and UCHL5. Androgen receptor is often over-expressed in prostate cancer (PCa) and is strongly linked to PCa growth and progression. Consequently, androgen deprivation therapy (ADT) that reduces androgen has been applied to treat androgen receptor-mediated PCa for decades. Nevertheless, most ADT treated patients experience relapse due to the development of the castration-resistant PCa. Numerous studies have shown that down-regulation of cellular androgen receptor level, including inhibiting its transcription and promoting its protein degradation, is lethal to PCa cells. Here we report that Aur arrested cell cycle progression and induced apoptosis of PCa cells. Co-inhibition of USP14 and UCHL5 with Aur facilitated the ubiquitination and degradation of androgen receptors in LNcap and 22RV1 PCa cells. Our results also show that Aur decreases the mRNA level of androgen receptors. In conclusion, our findings suggest that Aur is a promising agent for clinical translation to treat PCa.
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http://dx.doi.org/10.1016/j.ejphar.2019.01.004DOI Listing
March 2019

Targeting proteasome-associated deubiquitinases as a novel strategy for the treatment of estrogen receptor-positive breast cancer.

Oncogenesis 2018 Sep 24;7(9):75. Epub 2018 Sep 24.

Key Laboratory of Protein Modification and Degradation of Guangdong Higher Education Institutes, School of Basic Medical Sciences; Guangzhou Institute of Oncology, Tumor Hospital, Guangzhou Medical University, Guangzhou, Guangdong, 511436, China.

Estrogen receptor α (ERα) is expressed in ~67% of breast cancers and is critical to their proliferation and progression. The expression of ERα is regarded as a major prognostic marker, making it a meaningful target to treat breast cancer (BCa). However, hormone receptor-positive BCa was sometimes irresponsive or even resistant to classic anti-hormonal therapies (e.g., fulvestrant and tamoxifen). Hence, novel anti-endocrine therapies are urgent for ERα BCa. A phase II study suggested that bortezomib, an inhibitor blocking the activity of 20 S proteasomes, intervenes in cancer progression for anti-endocrine therapy in BCa. Here we report that proteasome-associated deubiquitinases (USP14 and UCHL5) inhibitors b-AP15 and platinum pyrithione (PtPT) induce growth inhibition in ERα BCa cells. Further studies show that these inhibitors induce cell cycle arrest and apoptosis associated with caspase activation, endoplasmic reticulum (ER) stress and the downregulation of ERα. Moreover, we suggest that b-AP15 and PtPT block ERα signaling via enhancing the ubiquitin-mediated degradation of ERα and inhibiting the transcription of ERα. Collectively, these findings demonstrate that proteasome-associated deubiquitinases inhibitors b-AP15 and PtPT may have the potential to treat BCa resistant to anti-hormonal therapy.
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http://dx.doi.org/10.1038/s41389-018-0086-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6155249PMC
September 2018

Growth arrest and apoptosis induction in androgen receptor-positive human breast cancer cells by inhibition of USP14-mediated androgen receptor deubiquitination.

Oncogene 2018 04 22;37(14):1896-1910. Epub 2018 Jan 22.

Affiliated Cancer Hospital of Guangzhou Medical University; Protein Modification and Degradation Lab, SKLRD, School of Basic Medical Sciences, Guangzhou Medical University, Guangzhou, Guangdong, China.

It has been well known that androgen receptor (AR) is critical to prostate cancer development and progression. It has also been documented that AR is expressed in more than 60% of breast tumors, which promotes the growth of estrogen receptor-negative (ER)/AR-positive (AR) breast cancer cells. Thus, AR might be a potential therapeutic target for AR-positive/ER-negative breast cancer patients. Previously we reported that in prostate cancer cells proteasome-associated deubiquitinase ubiquitin-specific protease 14 (USP14) stabilized AR protein level by removing its ubiquitin chain. In the current study, we studied the USP14-AR protein interaction and cell proliferation status after USP14 reduction or inhibition in breast cancer cells, and our results support the conclusion that targeting USP14 is a novel strategy for treating AR-responsive breast cancer. We found that inhibition of USP14 accelerated the K48-ubiquitination and proteasome-mediated degradation of AR protein. Additionally, both genetic and pharmacological inhibition of USP14 significantly suppressed cell proliferation in AR-responsive breast cancer cells by blocking G/G to S phase transition and inducing apoptosis. Moreover, AR overexpression inhibited USP14 inhibition-induced events, suggesting that AR deubiquitination by USP14 is critical for breast cancer growth and USP14 inhibition is a possible strategy to treat AR-positive breast cancer.
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http://dx.doi.org/10.1038/s41388-017-0069-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5886989PMC
April 2018

A novel deubiquitinase inhibitor b-AP15 triggers apoptosis in both androgen receptor-dependent and -independent prostate cancers.

Oncotarget 2017 Sep 28;8(38):63232-63246. Epub 2017 Jun 28.

Protein Modification and Degradation Lab, SKLRD, School of Basic Medical Sciences, The Affiliated Cancer Hospital of Guangzhou Medical University, Guangdong 511436, China.

Prostate cancer (PCa) remains a leading cause of cancer-related death in men. Especially, a subset of patients will eventually progress to the metastatic castrate-resistant prostate cancer (CRPC) which is currently incurable. Deubiquitinases (DUBs) associated with the 19S proteasome regulatory particle are increasingly emerging as significant therapeutic targets in numerous cancers. Recently, a novel small molecule b-AP15 is identified as an inhibitor of the USP14/UCHL5 (DUBs) of the 19S proteasome, resulting in cell growth inhibition and apoptosis in several human cancer cell lines. Here, we studied the therapeutic effect of b-AP15 in PCa, and our results indicate that (i) b-AP15 decreases viability, proliferation and triggers cytotoxicity to both androgen receptor-dependent and -independent PCa cells and , associated with caspase activation, inhibition of mitochondria function, increased reactive oxygen species (ROS) generation and endoplasmic reticulum (ER) stress; (ii) pan-caspase inhibitor z-VAD-FMK and ROS scavenger N-acetyl-L-cysteine (NAC) efficiently block apoptosis but not proteasome inhibition induced by exposure of b-AP15; (iii) treatment with b-AP15 in androgen-dependent prostate cancer (ADPC) cells down-regulates the expression of androgen receptor (AR), which is degraded the ubiquitin proteasome system. Hence, the potent anti-tumor effect of b-AP15 on both androgen receptor-dependent and -independent PCa cells identifies a new promising therapeutic strategy for prostate cancer.
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http://dx.doi.org/10.18632/oncotarget.18774DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5609916PMC
September 2017

Platinum pyrithione induces apoptosis in chronic myeloid leukemia cells resistant to imatinib via DUB inhibition-dependent caspase activation and Bcr-Abl downregulation.

Cell Death Dis 2017 07 6;8(7):e2913. Epub 2017 Jul 6.

Protein Modification and Degradation Lab, SKLRD, School of Basic Medical Sciences, The affiliated Tumor Hospital of Guangzhou Medical University, Guangdong 511436, China.

Chronic myelogenous leukemia (CML) is characterized by the chimeric tyrosine kinase Bcr-Abl. T315I Bcr-Abl is the most notorious point mutation to elicit acquired resistance to imatinib (IM), leading to poor prognosis. Therefore, it is urgent to search for additional approaches and targeting strategies to overcome IM resistance. We recently reported that platinum pyrithione (PtPT) potently inhibits the ubiquitin-proteasome system (UPS) via targeting the 26 S proteasome-associated deubiquitinases (DUBs), without effecting on the 20 S proteasome. Here we further report that (i) PtPT induces apoptosis in Bcr-Abl wild-type and Bcr-Abl-T315I mutation cells including the primary mononuclear cells from CML patients clinically resistant to IM, as well as inhibits the growth of IM-resistant Bcr-Abl-T315I xenografts in vivo; (ii) PtPT downregulates Bcr-Abl level through restraining Bcr-Abl transcription, and decreasing Bcr-Abl protein mediated by DUBs inhibition-induced caspase activation; (iii) UPS inhibition is required for PtPT-induced caspase activation and cell apoptosis. These findings support that PtPT overcomes IM resistance through both Bcr-Abl-dependent and -independent mechanisms. We conclude that PtPT can be a lead compound for further drug development to overcome imatinib resistance in CML patients.
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http://dx.doi.org/10.1038/cddis.2017.284DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5550844PMC
July 2017

Platinum-containing compound platinum pyrithione suppresses ovarian tumor proliferation through proteasome inhibition.

J Exp Clin Cancer Res 2017 06 15;36(1):79. Epub 2017 Jun 15.

Key Laboratory of Protein Modification and Degradation, School of Basic Medical Sciences and Affiliated Cancer Hospital of Guangzhou Medical University, Guangzhou, Guangdong, 511436, China.

Background: Ovarian carcinoma is one of the most aggressive gynecological malignant neoplasms and makes up 25-30% of all cancer cases of the female genital tract. Currently, resistance to traditional chemotherapy is a great challenge for patients with Epithelial ovarian cancer (EOC). Therefore, identifying novel agents for EOC treatment is essential and urgent.

Method: MTS assay was used to analyze the cell viability and proliferation of cancer cells. Flow cytometry was employed to analyze cell cycle distribution and cell apoptosis. Protein signaling pathways were detected by western blot and immunohistochemical staining. Nude mouse experiment was performed to test the in vivo effect of platinum pyrithione (PtPT).

Results: PtPT is a chemically well-characterized synthetic complex of platinum that potently inhibits proteasome-associated deubiquitinases USP14 and UCHL5 activity and shows selective cytotoxicity to multiple cancer cells without damaging DNA. We found that PtPT significantly accumulated ubquitinated-proteins and suppressed the proliferation of multiple EOC cells. Additionally, PtPT induced G2 phase arrest and apoptosis in both A2780 and SKOV3 cells. More importantly, animal experiments showed that PtPT dramatically suppressed the growth of EOC xenografts without obvious side effects.

Conclusion: These results suggest that through proteasome inhibition, PtPT significantly suppressed the proliferation of EOC in vitro and in vivo and could be developed as a novel agent for EOC treatment in the future.
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http://dx.doi.org/10.1186/s13046-017-0547-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5471884PMC
June 2017

Bilirubin neurotoxicity is associated with proteasome inhibition.

Cell Death Dis 2017 06 15;8(6):e2877. Epub 2017 Jun 15.

Protein Modification and Degradation Lab, SKLRD, School of Basic Medical Sciences, Affiliated Cancer Hospital of Guangzhou Medical University, Guangdong, People's Republic of China.

The molecular mechanism underlying bilirubin neurotoxicity remains obscure. Ubiquitin-proteasome system-mediated proteolysis is pivotal to virtually all cellular processes and cell survival. Here we report for the first time that bilirubin at a clinically relevant elevated level impairs proteasomal function via inhibiting both the 19S proteasome-associated deubiquitinases (USP14 and UCHL5) and the chymotrypsin-like (CT-like) peptidase activity of 20S proteasomes, thereby contributing to bilirubin neurotoxicity. This is supported by multiple lines of evidence. First, sera from patients with hyperbilirubinemia were able to inhibit the peptidase activity of purified 20S proteasome in vitro in a bilirubin concentration-dependent manner; meanwhile, the blood cells of these patients showed significantly increased levels of ubiquitinated proteins (Ub-prs), consistent with proteasome inhibition. Second, intracerebroventricular injection to adult rats or intraperitoneal injections to neonatal rats of bilirubin-induced neural accumulation of Ub-prs, concurrent with other neural pathology; and brain malfunction and pathology induced by neonatal exposure to hyperbilirubinemia were detectable in the rats during their adulthood. Third, in primary cultures of hippocampal neurons, bilirubin strikingly induced Ub-pr accumulation before the activation of cell death pathway becomes discernible. Finally, bilirubin in vitro directly inhibited both the deubiquitination activity of proteasome-associated USP14 and UCHL5 and the CT-like peptidase activity of purified 20S proteasomes, in a dose-dependent manner. Hence, this study has discovered that increased bilirubin at a clinically achievable level can act as a proteasome inhibitor via targeting the 19S proteasome-associated deubiquitinases (DUBs) and, perhaps to a less extent, the 20S proteasome, identifying a novel mechanism for bilirubin neurotoxicity.
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http://dx.doi.org/10.1038/cddis.2017.274DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5520929PMC
June 2017
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