药学学报

Substrate	Function of USP28 on the substrates	Reference
c-MYC	Deubiquitinating and stabilizing c-MYC by directly binding it or mediated by FBW7	[19, 51, 52]
FBW7	Antagonizing its autocatalytic ubiquitination and the ubiquitination of its substrates	[53]
HIF-1α	Revising the destabilization caused by FBW7	[54]
TCF7L2	Directly interacting with and deubiquitinating it by antagonizing FBW7, further regulating the Wnt/β-catenin signalling	[55]
P53	Deubiquitinating p53 through antagonizing MDM2	[52, 56, 57]
ΔNp63	Removing the K-48 linked ubiquitin chains induced by MDM2	[52, 56, 57]
FOXM1	Directly interacting with it and promoting its stabilization	[21]
LSD1	Deubiquitinating p53 through antagonizing MDM2	[26]
H2A	Decreasing the K119 ubiquitination level	[52, 58]
UCK1	Antagonizing its ubiquitination caused by KLHL2	[59]
NBS1	Promoting its stabilization by deubiquitination	[60]

Substrate	Function of USP28 on the substrates	Reference
c-MYC	Deubiquitinating and stabilizing c-MYC by directly binding it or mediated by FBW7	[19, 51, 52]
FBW7	Antagonizing its autocatalytic ubiquitination and the ubiquitination of its substrates	[53]
HIF-1α	Revising the destabilization caused by FBW7	[54]
TCF7L2	Directly interacting with and deubiquitinating it by antagonizing FBW7, further regulating the Wnt/β-catenin signalling	[55]
P53	Deubiquitinating p53 through antagonizing MDM2	[52, 56, 57]
ΔNp63	Removing the K-48 linked ubiquitin chains induced by MDM2	[52, 56, 57]
FOXM1	Directly interacting with it and promoting its stabilization	[21]
LSD1	Deubiquitinating p53 through antagonizing MDM2	[26]
H2A	Decreasing the K119 ubiquitination level	[52, 58]
UCK1	Antagonizing its ubiquitination caused by KLHL2	[59]
NBS1	Promoting its stabilization by deubiquitination	[60]

Cancer type	Expression level of Usp28	Mechanism/clinicopathological features	Protumoral/antitumoral
Colorectal cancer	Overexpressed	USP28 antagonizes the ubiquitin-dependent degradation of intestinal carcinogens c-MYC, c-JUN and Notch1, and promotes the proliferation of tumor cells.	Protumoral
Melanoma	Overexpressed	USP28 stabilizes the c-MYC level and hence increases ASS1 transcription upon arginine deprivation, and consequently leads to cell survival.	Protumoral
Deleted in a proportion of patients	Loss of Usp28 stabilizes BRAF enhancing downstream MAPK activation, promotes drug resistance of tumor cells and reduces the overall survival rate of patients.	Antitumoral
Breast cancer	Overexpressed	USP28 deubiquitinates and up-regulates LSD1, which can accelerate the formation of tumor stem cell-like features and tumor formation; the indirect suppression function of USP28 in CLDN7, which is mediated by LSD1, can promote the invasiveness of cancers; high serum epinephrine activates LDHA/USP28/MYC/SLUG signaling, associated with poor prognosis of breast cancer, and promotes the survival, proliferation and metastasis of breast cancer cells.	Protumoral
Overexpressed	Overexpression of Usp28 correlates with a better survival in patients with invasive ductal breast carcinoma; lack of Usp28 promotes a more malignant state of breast cancer cells.	Antitumoral
Liver cancer	Overexpressed	The expression levels of USP28 inversely correlate with patient survival; USP28 promotes the activity of the Wnt/β-catenin signaling pathway through maintaining the stability of TCF7L2; USP28 depletion or inhibition by a small molecule inhibitor leads to a halt of growth in liver cancer cells.	Protumoral
/	Lack of Usp28 results in advanced tumor development in a mouse xenograft model as well as in a chemically induced liver cancer mouse model.	Antitumoral
NSCLC	Overexpressed	Overexpression of Usp28 promotes NSCLC cells growth by increasing the stability of STAT3 and is correlated with the poor prognosis and low patient survival rate.	Protumoral
Other cancers	Overexpressed	The overexpression of Usp28 is related to the degree of differentiation and metastasis of gastric cancer cells. Usp28 is highly expressed in PC and is significantly associated with the malignant phenotype and shorter survival. USP28 mediates the activation of Wnt/β-catenin pathway through FOXM1 and promotes the growth of PC. Overexpression of Usp28 can promote the malignant phenotype of OV.	Protumoral

Cancer type	Expression level of Usp28	Mechanism/clinicopathological features	Protumoral/antitumoral
Colorectal cancer	Overexpressed	USP28 antagonizes the ubiquitin-dependent degradation of intestinal carcinogens c-MYC, c-JUN and Notch1, and promotes the proliferation of tumor cells.	Protumoral
Melanoma	Overexpressed	USP28 stabilizes the c-MYC level and hence increases ASS1 transcription upon arginine deprivation, and consequently leads to cell survival.	Protumoral
Deleted in a proportion of patients	Loss of Usp28 stabilizes BRAF enhancing downstream MAPK activation, promotes drug resistance of tumor cells and reduces the overall survival rate of patients.	Antitumoral
Breast cancer	Overexpressed	USP28 deubiquitinates and up-regulates LSD1, which can accelerate the formation of tumor stem cell-like features and tumor formation; the indirect suppression function of USP28 in CLDN7, which is mediated by LSD1, can promote the invasiveness of cancers; high serum epinephrine activates LDHA/USP28/MYC/SLUG signaling, associated with poor prognosis of breast cancer, and promotes the survival, proliferation and metastasis of breast cancer cells.	Protumoral
Overexpressed	Overexpression of Usp28 correlates with a better survival in patients with invasive ductal breast carcinoma; lack of Usp28 promotes a more malignant state of breast cancer cells.	Antitumoral
Liver cancer	Overexpressed	The expression levels of USP28 inversely correlate with patient survival; USP28 promotes the activity of the Wnt/β-catenin signaling pathway through maintaining the stability of TCF7L2; USP28 depletion or inhibition by a small molecule inhibitor leads to a halt of growth in liver cancer cells.	Protumoral
/	Lack of Usp28 results in advanced tumor development in a mouse xenograft model as well as in a chemically induced liver cancer mouse model.	Antitumoral
NSCLC	Overexpressed	Overexpression of Usp28 promotes NSCLC cells growth by increasing the stability of STAT3 and is correlated with the poor prognosis and low patient survival rate.	Protumoral
Other cancers	Overexpressed	The overexpression of Usp28 is related to the degree of differentiation and metastasis of gastric cancer cells. Usp28 is highly expressed in PC and is significantly associated with the malignant phenotype and shorter survival. USP28 mediates the activation of Wnt/β-catenin pathway through FOXM1 and promotes the growth of PC. Overexpression of Usp28 can promote the malignant phenotype of OV.	Protumoral

去泛素化酶USP28在肿瘤中的作用及其抑制剂研究进展

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李幸 ¹ , 张博雅 ¹ , 陆礼婷 ¹ , 王燕 ¹ , 文帅 ¹ , 臧永军 ²^,^* , 徐向明 ¹^,^* , 牟伊 ¹^,^*

药学学报 | 专题报道: 蛋白成熟与翻译后修饰的化学干预 2024,59(11): 2941-2952

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药学学报 | 专题报道: 蛋白成熟与翻译后修饰的化学干预 2024, 59(11): 2941-2952

去泛素化酶USP28在肿瘤中的作用及其抑制剂研究进展

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李幸¹, 张博雅¹, 陆礼婷¹, 王燕¹, 文帅¹, 臧永军²^,^*, 徐向明¹^,^*, 牟伊¹^,^*

作者信息

1.泰州学院医药与化学化工学院, 江苏泰州 225300

2.皖西学院生物与制药工程学院, 安徽六安 237012

通讯作者:

^*臧永军, E-mail: zangyongjun@yeah.net;

徐向明, E-mail: 379648107@qq.com;

牟伊, E-mail: mouyicpu@163.com

Research progress on the roles and inhibitors of deubiquitinase USP28 in tumors

Xing LI¹, Bo-ya ZHANG¹, Li-ting LU¹, Yan WANG¹, Shuai WEN¹, Yong-jun ZANG²^,^*, Xiang-ming XU¹^,^*, Yi MOU¹^,^*

Affiliations

1. College of Pharmacy and Chemistry & Chemical Engineering, Taizhou University, Taizhou 225300, China

2. Department of Biological and Pharmaceutical Engineering, West Anhui University, Lu'an 237012, China

出版时间: 2024-11-12 doi: 10.16438/j.0513-4870.2024-0098

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摘要

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去泛素化酶(DUBs) 是蛋白质泛素系统中一类重要的肽水解酶, 对泛素化具有逆转和严格调控作用, 在调节蛋白质稳定性、细胞信号转导等多种生物学过程中发挥重要功能。泛素特异性蛋白酶28 (USP28) 通过增强多种癌症相关蛋白的稳定性, 参与癌症相关的多种信号途径, 与结直肠癌、乳腺癌、肺癌、胰腺癌等多种肿瘤的进展密切相关。USP28已成为一个有吸引力的抗癌药物靶点, 其抑制剂的开发取得了一定进展。本文对USP28的结构及其与底物的相互作用、USP28在肿瘤中的研究进展、USP28抑制剂的发现等方面进行了综述。

关键词

去泛素化酶 / 泛素特异性蛋白酶28 / 肿瘤 / 抑制剂 / 泛素化

Abstract

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The deubiquitinases (DUBs), as the crucial peptidohydrolases in the ubiquitin system, can reverse and strictly regulate ubiquitination and play key roles in various biological processes, including the regulation of protein stability, cell signal transduction. Ubiquitin-specific protease 28 (USP28) involves multiple cancer-related signaling pathways by enhancing the stability of various cancer-related proteins, and is closely associated with the progression of colorectal, breast cancer, lung carcinomas, and pancreatic cancer. USP28 has been considered as a promising drug target in anticancer therapy, and the development of USP28 inhibitors has made some progress. In this article, we review the structure of USP28 and its interaction with substrates, discuss the research progress of USP28 in cancers and summarize the development of USP28 inhibitors.

Key words

deubiquitylating enzyme / ubiquitin-specific protease 28 / cancer / inhibitor / ubiquitination

引用本文

李幸, 张博雅, 陆礼婷, 王燕, 文帅, 臧永军, 徐向明, 牟伊. 去泛素化酶USP28在肿瘤中的作用及其抑制剂研究进展. 药学学报, 2024 , 59 (11) : 2941 -2952 . DOI: 10.16438/j.0513-4870.2024-0098

Xing LI, Bo-ya ZHANG, Li-ting LU, Yan WANG, Shuai WEN, Yong-jun ZANG, Xiang-ming XU, Yi MOU. Research progress on the roles and inhibitors of deubiquitinase USP28 in tumors[J]. Acta Pharmaceutica Sinica, 2024 , 59 (11) : 2941 -2952 . DOI: 10.16438/j.0513-4870.2024-0098

正文

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泛素化是通过一系列的酶促级联反应将泛素以单体或链的形式附着在底物蛋白上的过程^[1-3], 这是一种高度特异性和选择性的翻译后修饰^[4-7], 对底物蛋白的稳定、亚细胞定位和功能等方面起着重要调控作用, 进而广泛参与各种生理过程^{[1, 8]}, 如蛋白质降解^[9]、线粒体自噬、DNA损伤修复^[10]、表观遗传调控^[11]、免疫反应调控、细胞周期和细胞凋亡调控等^{[12, 13]}。相反, 去泛素化是指由去泛素酶(DUBs) 通过特异性地水解肽键或异肽键将泛素从底物蛋白中去除的过程, 继而保护靶蛋白不被降解或失活^{[14, 15]}。目前, 已鉴定的人类DUBs大约有100个, 根据催化域的序列相似性将其分为7个亚家族: 泛素特异性蛋白酶(ubiquitin-specific proteases, USPs)、泛素羧基端水解酶(UCHs)、马查多-约瑟芬结构域蛋白酶(MJDs)、JAB1/MPN/Mov34金属酶(JAMMs)、卵巢肿瘤蛋白酶(OUTs)、与含泛素的新DUB家族相互作用的基序(MINDYs)、含锌指的泛素肽酶1 (ZUP1s)。由DUBs调控的泛素和去泛素化平衡一旦失调, 将会引起包括癌症在内的多种疾病^{[7, 16]}, 因此DUBs有望成为疾病治疗的新靶点^[17]。

USPs是数量最多和研究最为广泛的DUBs亚家族, 泛素特异性蛋白酶28 (USP28) 是其中重要的成员之一^[18]。USP28拥有相对保守的结构, 可以通过逆转多种底物的泛素化而拮抗E3泛素连接酶的功能, 继而参与细胞增殖^{[19, 20]}、细胞凋亡^[21]、应激反应^{[22, 23]}和DNA损伤反应^[24-28]等多种细胞过程。Usp28高表达影响多种肿瘤、神经退行性疾病和免疫系统疾病的进展, 提示USP28可能成为疾病治疗的新靶点^[28]。由于USP28在维持多种癌症相关底物蛋白, 包括c-MYC (c-Myc原癌基因编码的一种转录因子)、赖氨酸特异性组蛋白去甲基化酶1 (LSD1)、缺氧诱导因子-1α (HIF-1α)、c-JUN (一种转录调节因子, 属亮氨酸拉链家族成员) 和缺口受体1 (Notch 1) 的细胞稳定态中起着至关重要的作用^{[19, 20, 23, 25-29]}, 其失调与多种癌症进展密切相关^{[21, 26, 30-32]}。研究表明, Usp28在结直肠癌、乳腺癌等恶性肿瘤中高表达^{[29, 31]}, 且与胶质瘤和非小细胞肺癌等癌症的预后负相关^{[33, 34]}。USP28已成为抗癌药物开发的潜在靶点^{[24, 34]}, 正在引起广泛关注^[32]。近年来, 几种USP28小分子抑制剂已被发现和报道^[35-40]。小分子USP28抑制剂被证明可以下调USP28下游的癌症相关底物蛋白, 如c-MYC、LSD1的表达水平, 并诱导一系列癌细胞系的细胞活力丧失和凋亡。然而, 一些研究也阐明了USP28在某些特定肿瘤中具有抑癌作用。本文简述了USP28的结构和底物相互作用, 系统地介绍了USP28在癌症中的作用, 并总结和分析了USP28抑制剂的研究进展。

1 USP28的结构及其与底物相互作用

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1.1 USP28的结构

2001年, 通过DNA和蛋白质水平的数据库同源性搜索以及cDNA文库筛选, 鉴定出了与Usp25高度同源的Usp28, 该基因位于人类染色体11q23.2上, 主要在肌肉和心脏中表达^[41]。USP28全长蛋白由1 077个氨基酸残基组成(图 1), 相对分子质量约122 kDa^{[42, 43]}, 与其他USP家族成员具有相似的拓扑结构, 可以分为三个结构域, 包括N端的泛素结合区(UBR)、保守的催化域(CD) 以及卷曲的C端结构域(CTD)^{[43, 44]}。其中, USP28的N端结构域由一个泛素相关结构域(UBA)、一个泛素相互作用基序(UIM) 和一个类泛素蛋白修饰分子相互作用基序(SIM) 组成, 这些基序负责泛素底物识别和结合的特异性^{[44, 45]}。USP28的催化域序列与USP25同一性高达57%, 并表现出了与大多数USPs相似的经典结构(图 2), 即类似于一只伸出的右手, 分为手指(finger)、手掌(palm) 和拇指(thumb) 三个子结构域^[45-47], 且都包含一个长度为170个氨基酸(aa) 的插入位点(INS), 这对USP28催化域二聚体的形成具有重要作用^{[44, 48, 49]}。C端部分是一个可变大小的未知折叠区域, 与USP25具有40%的同源性。由于可变剪接, USP28产生1 045~1 077 aa的翻译产物^[44]。

1.2 USP28与底物的相互作用

USP28保守而又特异性的结构使其能够拮抗多种E3泛素连接酶, 其中, 含F-box和WD重复结构域7 (FBW7, 又称FBXW7) 是最重要的一种。FBW7被认为是一个经典的抑癌蛋白, 因其可以泛素化降解多种已知的癌蛋白底物, 如c-MYC、Notch 1、细胞周期蛋白E (cyclin E) 等。一方面, FBW7依赖的底物泛素化可以被USP28去泛素化。c-MYC作为FBW7的泛素化分子之一, 同时也是USP28的底物^[50]。USP28先前被证明通过与FBW7的“背驮式”相互作用与底物结合^[51], 即USP28通过与FBW7a形成二元复合物间接地与c-MYC结合, 在体内去泛素化c-MYC (图 3A), 这个过程是由FBW7的N端区域介导。相应地, 从同时表达USP28和FBW7的细胞中纯化的USP28具有增强c-MYC去泛素化的能力; 删除N端的FBW7α^ΔΝ则不能介导HA-USP28与c-MYC的结合, FBW7α^ΔN在USP28存在时能够促进c-MYC的降解^[19]。另有报道表明, USP28的功能不依赖于FBW7的存在, 而是独立地识别和去泛素化与FBW7相同的底物^[51](图 3A)。在FBW7缺失的情况下, 也观察到USP28与c-MYC的直接相互作用, 此过程中USP28更可能与未磷酸化的肽结合^{[51, 52]}。另一方面, FBW7本身也可以被USP28直接去泛素化, 不过, 去泛素化FBW7所需的USP28蛋白含量明显低于FBW7底物(如c-MYC) 去泛素化所需的含量。半数正常水平的USP28可以维持FBW7的稳定, 但不足以稳定FBW7底物。Usp28的过表达可同时稳定FBW7及其底物; 相反, 完全敲除Usp28会触发FBW7的自催化泛素化和降解, 从而稳定FBW7底物(图 3B)。Usp28的完全缺失和过表达均会促进致癌转化^[53]。

与c-MYC类似, HIF-1α也是USP28和FBW7的底物。糖原合成酶激酶-3β (GSK-3β) 可以磷酸化HIF-1α并招募FBW7泛素化和下调HIF-1α, USP28可以去泛素化HIF-1α, 但是两者没有直接的相互作用, 需要由FBW7介导实现USP28对HIF-1α的稳定, 继而影响缺氧和HIF-1α依赖性血管生成^[54]。此外, USP28通过与转录因子7样2 (TCF7L2) 相互作用和去泛素化来提高其稳定性, 并调节Wnt/β-连环蛋白(Wnt/β-catenin) 信号^[55]。USP28还可以逆转鼠双微粒体-2 (MDM2) 的泛素化功能, 通过去除由MDM2诱导的K48连接的泛素链, 继而去泛素化p53和ΔNp63^{[52, 56, 57]}。同样的, USP28可以降低组蛋白H2A的K119泛素化水平(ub-K119-H2A)^{[52, 58]}, 逆转泛素E3连接酶凯尔奇样家族成员2 (KLHL2) 介导的UCK1的K8多泛素化^[59]。此外, USP28对LSD1的蛋白水平和活性有调节作用, 两者的直接相互作用由LSD1的单胺氧化酶结构域和USP28的N端结构域介导完成^[26]。USP28可逆转奈梅亨断裂综合征1 (NBS1) 蛋白的泛素化, 以稳定DNA双链断裂(DSB) 上的MRN复合物, 并在RecQl4缺陷细胞中恢复DSB应答^[60]。USP28去泛素化并稳定Wnt/β-catenin信号传导的关键介质叉头盒蛋白质M1 (FOXM1), 敲降和沉默USP28可以促进FOXM1的多泛素化和蛋白酶体降解^[21]。总之, USP28能够去泛素化多种底物, 并稳定其蛋白水平或功能(表 1)。

2 USP28在不同癌症中的研究进展

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USP28可以直接或间接地作用于多种底物, 包括MYC、c-JUN、HIF-1α和LSD1, 通过调节这些底物的泛素化和稳定性, 参与DNA损伤修复、凋亡、表观遗传调控和细胞周期控制, 潜在地促进各种肿瘤的发展^[55]。USP28的促肿瘤作用及相关研究目前占主导地位, 然而, 近来一些研究证明了它具有抑癌作用, 这可能是基于特定的肿瘤、特定的细胞系或特定的分子而发生的。因此, 明晰USP28在癌症进展中的双重作用, 对于靶向USP28进行治疗具有重要意义, 本文将分别介绍USP28在不同肿瘤中的最新研究进展(表 2)。

2.1 结肠癌

Usp28在结肠癌中高表达^{[19, 29, 51]}, 与肠道的稳态和结肠癌的进展密切相关。Diefenbacher等^[29]使用小鼠遗传模型, 确定了USP28拮抗肠道中致癌因子c-MYC、c-JUN和Notch1的泛素依赖性降解。缺乏Usp28的小鼠没有明显的不良表型, 但表现出肠道增殖减少和分泌谱系细胞分化受损。Usp28在结直肠癌模型中的作用和肠道稳态相似, Usp28缺乏可以促进肿瘤细胞分化, 抑制肿瘤细胞的增殖, 在小鼠结肠直肠癌模型中, 敲除Usp28可以减少肠道肿瘤的数量、缩小肿瘤体积并显著延长荷瘤小鼠的寿命。此外, Usp28是在小鼠和人类肠癌中高度表达的c-MYC靶基因, 这表明Usp28和c-Myc形成了一个正反馈回路, 在肿瘤中维持高c-MYC蛋白水平^[29]。相比Usp28失活的Ls174T结肠癌细胞, 野生型细胞中表达出更高水平的c-MYC蛋白, 促进了肿瘤细胞的增殖^[19]。在APC^Min/+小鼠模型中, Usp28和Fbw7同时失活时可以抑制侵袭性肠道肿瘤的形成^[51]。因此, 抑制USP28的酶活性可能是结肠癌治疗的有效策略。

2.2 黑色素瘤

在对B-Raf原癌基因(BRAF) 抑制剂耐药(BR) 的黑色素瘤细胞中, c-MYC介导的精氨酸琥珀酸合成酶1 (ASS1) 重新表达和自噬相关蛋白水平减弱。此外, 研究发现, 去泛素酶USP28的下调增强c-MYC的蛋白酶体降解, 这是BRAF抑制剂耐药的黑色素瘤细胞在精氨酸剥夺时无法重新表达ASS1的主要机制。BR细胞中Usp28的过表达提高了c-MYC的水平, 继而增加了精氨酸剥夺后ASS1的转录, 从而导致细胞存活^[61]。与之不同的是, Saei等^{[62, 63]}报道, Usp28在9%的黑色素瘤患者中缺失, 且Usp28基因缺失导致黑色素瘤患者的总生存率降低。进一步研究表明, USP28/FBW7复合物直接泛素化和降解BRAF, 而Usp28的缺失通过稳定BRAF增强了丝裂原活化蛋白激酶(MAPK) 的活性, 驱动BRAF (V600E) 肿瘤对BRAF抑制剂治疗的耐药性。通过分析USP28在泛癌中的综合预后发现, USP28可以作为黑色素瘤患者抗细胞毒性T细胞相关蛋白-4 (CTLA4) 治疗反应的重要预测因子^[64]。USP28在黑色素瘤中表现出了促癌和抑癌的双重作用, 值得进一步深入研究靶向USP28进行黑色素瘤治疗的方向。

2.3 乳腺癌

多数研究表明USP28具有促进乳腺癌进展的作用^[52]。Usp28在人类乳腺癌中明显高表达^[28]。在多种癌细胞系和乳腺肿瘤样本中, Usp28过表达与LSD1上调相关, USP28通过去泛素化作用稳定LSD1, Usp28的敲低可下调LSD1水平, 抑制体外肿瘤干细胞样特征和体内致瘤性, 这可以通过LSD1的过表达来逆转^[26]。此外, 在乳腺癌中, USP28通过LSD1介导的对紧密连接蛋白7 (CLDN7) 的下调作用, 有可能促进癌细胞的侵袭性^[52]。

慢性应激诱导的肾上腺素激活乳酸脱氢酶(LDHA) 生成乳酸, 通过局部酸性促进USP28对c-MYC的去泛素化和稳定。SLUG启动子随后被c-MYC激活, 从而促进乳腺癌癌样性状的发展。在乳腺癌患者中, 高血清肾上腺素激活了LDHA/USP28/MYC/SLUG信号轴, 与不良预后相关^[22]。

USP28可以促进三阴性乳腺癌(TNBC) 细胞生存能力, 抑制USP28的功能, 导致TNBC细胞停滞在S/G2期。USP28的小分子抑制剂AZ-1在TNBC异种移植瘤模型中展现了抗肿瘤活性^[65]。USP28可以结合来自癌症相关成纤维细胞的外泌体miR-500a-5p, 促进乳腺癌细胞增殖和转移^[49]。与上述USP28促进乳腺癌研究结果不一致的是, Richter等^[31]发现, 在浸润性导管性乳腺癌患者中, Usp28的过表达与生存率正相关。在小鼠异种移植瘤模型中, 缺乏Usp28促进肿瘤生长和血管生成。此外, 缺乏Usp28会促进乳腺癌细胞的恶性状态, 表现为上皮-间充质(EMT) 转变、增殖、迁移和血管生成增加以及黏附减少。机制上, 由Usp28缺失驱动的血管生成增加和致癌过程可能独立于HIF-1α、p53和P53结合蛋白1 (53BP1)^[31]。研究表明, 较高水平的USP28在预测乳腺癌更好的生存率和TNM分型方面仍具有不可忽视的价值, 这可能对靶USP28治疗癌症具有指导意义^[52]。

2.4 肝癌

生存分析显示, Usp28高表达的肝癌患者总生存期较短, 进一步的研究表明, 过表达miR-216b可以下调肝癌细胞(HCC) 中的USP28/c-MYC信号, 抑制HCC的生长^[66]。USP28通过去除FBW7对TCF7L2的泛素化, 影响TCF7L2的水平和Wnt/β-catenin信号; 抑制或者耗竭USP28可导致肝癌细胞的生长停止^[55]。分子生物学实验证实, Usp28的敲低显著降低了HCC细胞系的增殖和侵袭能力^[64]。另有不同的研究结果表明, 在化学诱导的肝癌模型中, 小鼠缺乏USP28可促进早期发病和更严重的肿瘤形成^[31]。

2.5 肺癌

USP28在非小细胞肺癌(NSCLC) 原代组织和细胞系中均上调, USP28 mRNA和蛋白水平与患者生存率均呈负相关, Usp28过表达促进NSCLC细胞生长, 并与NSCLC患者的不良预后相关; 反之, 下调USP28通过信号转导和转录激活因子3 (STAT3) 信号通路在体外抑制NSCLC细胞的生长, 在体内延缓NSCLC肿瘤的生长^{[34, 67]}。Ren等^[68]指出, miR-3940-5p通过靶向cyclin D1和USP28可抑制NSCLC细胞增殖并诱导细胞凋亡。类似研究表明, 内源性USP28对于小鼠肺鳞状细胞癌(LSCC) 的诱发和维持都是必需的^[57], Usp28的基因失活可诱导小鼠LSCC肿瘤的消退, USP28抑制剂FT206治疗显著下调了c-MYC、c-JUN和ΔNp63蛋白水平, 从而诱导自发性小鼠LSCC肿瘤和人类LSCC异种移植瘤的实质性消退, 进一步证实了基因缺失所观察到的表型^[69]。

2.6 其他癌症

Usp28在胰腺癌(PC) 肿瘤中的高表达与PC患者的恶性表型和较短的生存期显著相关^[21]。在体外和体内实验中, Usp28过表达通过促进细胞周期进程和抑制细胞凋亡来加速PC细胞的生长, 而Usp28敲低则抑制了PC细胞的生长。从机制上讲, USP28介导的FOXM1稳定显著促进了细胞核β-catenin反式激活, 进而导致Wnt/β-catenin通路的激活。与此一致的是, FOXM1表达的恢复消除了Usp28沉默的抗肿瘤作用^[21]。与正常卵巢组织相比, Usp28在卵巢癌(OV) 样本中高表达, 且与OV患者的不良预后相关。β-catenin/YAP1/TBX5介导的Usp28异常表达可促进OV的恶性表型, 而对Usp28进行基因消融或药物抑制均可损害OV细胞的体外和体内增殖能力^[70]。Usp28在胃癌中过表达, 且与胃癌分化和转移程度有关。体外抑制Usp28表达可通过下调LSD1抑制胃癌细胞的增殖和侵袭^[71]。上市药物西地兰可以减弱USP28对c-MYC的去泛素化, 下调c-MYC并抑制Wnt/β-catenin信号, 抑制人胃癌细胞MKN-45增殖^[72]。

3 USP28抑制剂的研究进展

收起

DUBs具有广泛的去泛素化底物蛋白, 与多种癌症的发生、转移、预后等方面密切相关。因此, 靶向DUBs的小分子抑制剂发现及研究已经成为抗肿瘤候选药物的热点领域之一^[17]。目前研究较多的是USPs, 其中USP7抑制剂研究较为深入, 最近报到了一些有潜力的候选化合物^{[73, 74]}。近年来, USP28的晶体结构及其在肿瘤中的重要作用被揭示, USP28的失调已被证明可以加速多种癌症的进展, 将其作为癌症治疗的靶点可能是一个潜在的方向。虽然, USP28抑制剂的研究引起了关注, 但由于USP28和USP25高度同源且USP28与小分子抑制剂的结构生物学报道有限, 使针对该酶的有效抑制剂的开发仍处于早期阶段, 尚无分子进入临床试验。目前, 仅有少数USP28抑制剂被报道, 根据其结构母核和来源可以分为以下六类: 二氨基吡啶类、苄氨基乙醇类、三氮唑并嘧啶类、甲酰胺噻吩并吡啶类、vismodegib及其衍生物类、天然产物和其他类。

3.1 二氨基吡啶类

2011年, Altun等^[39]利用基于活性的高通量筛选进行USP7活性调节剂开发时, 发现了苗头化合物PR-619 (图 4)。体外实验比较了PR-619对DUBs、半胱氨酸蛋白酶和其他蛋白水解酶家族的抑制活性, 数据表明, PR-619对多种DUBs表现出广泛的抑制作用, 但对其他蛋白酶家族的活性有限, 其中对USP28的IC₅₀是6.24 µmol·L^-1。进一步研究了PR-619在细胞裂解液中对DUBs的作用, 结果显示, PR-619在20 µmol·L^-1时可抑制大部分DUBs, 在浓度为5 µmol·L^-1时抑制程度较低。在低摩尔范围内诱导了HCT-116结直肠癌和HEK293T细胞的死亡。细胞通透性和活细胞中DUBs抑制效力实验表明, PR-619在浓度为5 mmol·L^-1或更高时干扰探针标记。

3.2 苄氨基乙醇类

2017年, Wrigley等^[35]利用高通量筛选方法发现了以AZ1~4为代表的一系列苄氨基乙醇类USP28抑制剂(图 5), 采用以泛素-罗丹明110 (Ub-Rh110) 为底物的测试方法, 4个化合物的IC₅₀值分别为0.7、1.1、59.6和2.0 μmol·L^-1。等温滴定量热法(ITC) 和微量热泳动(MST) 两种实验均测定AZ1、AZ2和AZ4可以直接有效地结合USP28, 且结合方式是可逆的, 与活性结果一致; 低活性的AZ3没有测出有效结合力。除了可以同等程度地抑制USP28同源度最高的USP25, 这些抑制剂对大多数DUBs具有高度的体外选择性。

随后, 它们被用作化学工具分子进行USP28细胞水平的生物学概念验证。用AZ1和AZ2处理HCT116结肠癌细胞后, c-MYC总蛋白水平出现了快速的浓度依赖性下降, 这种效果由蛋白酶体降解介导。在AZ1和AZ2的最高测试浓度下观察到c-MYC水平均显著降低, 而AZ3仅在最高浓度为100 μmol·L^-1时对c-MYC水平有影响(60 μmol·L^-1及以下没有检测到明显的影响), 这三种化合物的在靶生化指标与它们的体外相对活性一致。AZ1和AZ2能够以时间和浓度依赖的方式诱导包括HCT116在内的一系列癌细胞系的细胞凋亡和细胞活力丧失, EC₅₀值集中在20 μmol·L^-1左右。然而, 进一步的分析表明, 在肿瘤细胞和正常细胞之间没有观察到明显的活性差异。这种细胞杀伤缺乏选择性可能是由于所涉及的途径复杂、USP28和USP25之间潜在功能冗余造成的, 进一步提高抑制剂的效力和选择性是提高治疗指数的有效解决途径之一。

Sun等^[55]发现AZ1可以抑制肝癌细胞HepG2和HuH6 (EC₅₀分别为12.65和5.85 μmol·L^-1), 并显著降低了USP28自身和TCF7L2的水平, 以及TCF7L2、AXIN2和cyclin D1的靶基因水平。AZ1还抑制了HepG2和HuH6细胞的集落形成; 不过, 其抗增殖作用不完全依赖于TCF7L2。这些工具分子和数据促进了对DUBs作为潜在治疗靶点的理解。

3.3 三氮唑并嘧啶类衍生物

2020年, Liu等^[38]报道了一系列新型的[1,2,3]三氮唑[4,5-d]嘧啶衍生物作为USP28抑制剂。以AZ1为阳性对照, 采用Ub-AMC为底物的活性测试方法对目标化合物开展了体外活性研究, 化合物19 (图 6) 对USP28具有较强的抑制活性和亲和力(IC₅₀ = 1.10 μmol·L^-1, K_d = 40 nmol·L^-1), 且可逆结合胃癌细胞中内源性USP28, 对USP7和LSD1几乎没有抑制活性(IC₅₀ > 100 μmol·L^-1)。进一步研究表明, USP28在胃癌MGC-803、MKN45和HGC27细胞系中过表达, 体外细胞活力实验结果显示, 化合物19对HGC-27和MKN45细胞具有良好的抑制活性(IC₅₀值分别为0.61和1.49 μmol·L^-1)。对于MGC-803细胞和胃上皮永生化GES-1细胞抑制活性较低, 相应的IC₅₀值分别为4.95和9.19 μmol·L^-1。化合物19在HGC-27细胞中浓度依赖性地诱导USP28的底物LSD1和c-MYC的蛋白酶体降解, 从而抑制胃癌细胞系的增殖、S期细胞周期和EMT进程^[75]。

构效关系和分子对接研究表明, 母核中的三氮唑和嘧啶环均与USP28中的Phe370发生π-π相互作用, 同时, 氯苯环与Asn597形成π-H相互作用(图 7)。结合活性数据推测, 与三唑环相连的苄基是活性必需基团, 将化合物19的4-Cl-苄基替换为羟乙基所得化合物12 (图 6) 导致失活^[38]。同时, 嘧啶环上的氨基对USP28的抑制活性也至关重要。这些数据为发现新的USP28抑制剂提供了一定的指导意义。

3.4 甲酰胺噻吩并吡啶类

2021年, Ruiz等^[69]报道了一种有效的USP28抑制剂FT206 (图 8), 这是一种甲酰胺噻吩并吡啶类衍生物, 源于FORMA Therapeutics公司在之前申请的专利(EP3414252B1) 和最近更新的(US11524966B1、EP3833661A1) 的专利。其中, FT206结构在2020年的专利中结构明确公开, 并给出了其对USP28的活性抑制范围, 虽然对USP25表现出交叉性抑制, 但对其他DUBs表现出了高度的选择性^[69]。使用FT206 (浓度0.2和0.4 μmol·L^-1) 抑制USP28可降低人LUDLU-1 LSCC细胞中c-MYC、c-JUN和ΔNp63蛋白水平, 并抑制人LSCC细胞系的细胞增殖。在异种移植模型中, FT206展现了抗LSCC肿瘤进展效应, 并通过阻断USP28介导的底物去泛素化, 降低了c-MYC蛋白水平。

2022年, Peng等^[37]通过高通量筛选获得了先导化合物CT1002 (图 8), 初步构效关系探索发现噻吩环上的氨基单甲基化有利于活性提高, 得到了化合物CT1018 (图 8)。然而, CT1018在细胞的实验中无活性, 继续优化获得了USP25/28双重抑制剂CT1073和CT1113 (图 8), 化学结构与FT206具有明显的相似性, 都含有噻吩并吡啶母核和酰胺结构, CT1113的对映异构体CT1113S (图 8) 对USP28和USP25的活性都有所降低。表面等离子体共振(SPR) 实验测定这两个化合物对USP25和USP28具有相似的结合动力学参数, 但是对其他DUBs表现出了很高的选择性。CT1073和CT1113都能够在多种癌细胞系中显著降低USP28和USP25本身及其底物c-MYC、LSD1和Tac-MYC水平, CT1113的抗增殖作用在很大程度上是一种靶效应, 尽管不能排除潜在的脱靶效应。CT1113显著抑制了人胰腺癌异种移植肿瘤的生长, 并下调了c-MYC水平, 在结肠癌CDX模型上观察到类似的效应。

3.5 Vismodegib及其衍生物

Wang等^[36]采用Ub-AMC水解活性测定实验联合高通量筛选的方法从市售药物库中发现vismodegib (图 9) 可以抑制USP28, 这是一种FDA批准的治疗基底细胞癌的hedgehog通路抑制剂药物。体外Ub-AMC水解实验测得vismodegib对USP28和USP25的IC₅₀值分别是4.41和1.42 μmol·L^-1。随后, 采用多种技术和途径表征了vismodegib和USP28之间的相互作用, 并采用氢-氘交换质谱(HDX-MS) 实验绘制了vismodegib在USP28中的结合位点。数据分析显示, vismodegib在USP28中占据的结合腔与报道的USP7抑制剂中一个结合口袋(PDB: 5UQV) 能很好地对齐。此外, 细胞实验证实vismodegib可以与USP28和USP25相互作用, 并下调这两种酶的底物蛋白c-MYC、Notch 1和Tankyrase-1/2的水平。

随后, Zhou等^[40]首次解析了USP28与小分子抑制剂vismodegib结合的共晶结构(PDB: 8HJE, 图 10)。共晶结构显示vismodegib位于USP28的一个变构结合位点, 并通过酰胺基团和氯原子分别于与附近Cys644的Ser257残基之间形成了氢键相互作用, 苯甲磺酰基部分延伸到溶剂区未观察到与USP28蛋白的相互作用, 吡啶部分与Leu264形成了H-π相互作用, 吡啶环周围有足够的空间进行进一步的结构修饰。

随后基于结构的优化获得了一系列vismodegib衍生物作为有效的USP28抑制剂。其中, 代表性的化合物9l、9o和9p (图 9) 对USP28具有较高的活性, 对USP25也展现了相似程度的抑制作用, 对其他6个DUBs具有较高的选择性。细胞实验表明, 化合物9l、9o和9p对人结直肠癌和肺鳞癌细胞均具有细胞毒性, 并显著增强结直肠癌细胞对瑞戈非尼的敏感性。进一步研究表明, 化合物9l、9o和9p可通过泛素蛋白酶体系统剂量依赖性地下调c-MYC的细胞水平, 其抗癌作用可能主要与抑制USP28有关, 而不涉及Hedgehog-Smoothened通路。

3.6 天然产物和其他

SGH (图 11) 是一种新的戊二酰亚胺类放线菌次级代谢物。研究表明, SGH可下调c-MYC、USP28和细胞周期和核苷酸合成相关蛋白, 阻断G0/G1期细胞周期, 降低糖酵解代谢酶的表达水平, 显著抑制不同类型肺癌细胞的增殖^[76]。

咖啡酸衍生物(CADPE, 图 11) 是从中药提取物中分离出来的天然多酚酯^[77], 在Caki-1小鼠肿瘤模型中, CADPE能抑制肿瘤中STAT3的磷酸化、HIF-1α和VEGF的基因表达, 继而延缓肿瘤生长^[78]。此外, CADPE通过激活GSK-3β和下调USP28来触发FBW7对c-MYC的泛素化, 促进c-MYC的降解, 进而下调细胞周期调节因子和抗凋亡蛋白, 最终导致细胞周期阻滞和细胞凋亡, 对不同白血病细胞系均有较强杀伤性, 对正常细胞的毒性较低^[77]。

毛花苷C (lanatoside C, 图 11) 是由毛花洋地黄中提取的一种速效强心苷, 临床上被批准广泛用于治疗抗心律失常和心力衰竭^[79]。毛花苷C可以通过减弱USP28与c-MYC之间的相互作用, 促进c-MYC去泛素化和降解。此外, 毛花苷C可以诱导G2/M细胞周期阻滞, 阻碍MKN-45细胞迁移, 促进其凋亡, 对MKN-45细胞具有抗增殖活性(IC₅₀ = 32.94 nmol·L^-1) ^[72]。

Xu等^[80]通过高通量筛选和验证发现, 经批准治疗肠易激综合征的药物奥替溴铵(otilonium brimide, 图 11) 对USP28有抑制作用(IC₅₀ = 6.90 μmol·L^-1), 其对USP28的抑制选择性是USP25的3~4倍。酶动力学和HDX-MS数据分析结果表明, 奥替溴铵可能与vismodegib结合在USP28中相同的变构口袋上, 并以可逆和非竞争的方式抑制其活性。细胞实验显示, 奥替溴铵可能通过下调USP28的致癌底物c-MYC和/或ΔNp63, 对人类结直肠癌细胞和肺鳞癌细胞具有细胞毒性, 并显著增强结直肠癌细胞对瑞戈非尼的敏感性。

4 总结与展望

收起

泛素化和去泛素化是机体中调控蛋白质降解的重要分子机制, 在该过程中, 负责移除蛋白底物泛素链的DUBs至关重要^{[81, 82]}。DUBs通过多种分子在多种细胞生命活动中均扮演着重要角色, 其活性或表达异常可造成关键致癌/抑癌蛋白的功能变化, 直接或间接导致肿瘤发生发展和恶性演进^{[83, 84]}。

USP28是一种与USP25高度同源的去泛素酶, 主要调控多种蛋白的泛素化降解, 继而产生促进细胞增殖、启动侵袭转移、刺激细胞存活、抑制细胞分化、诱导血管生成等多种作用, 这些都有助于肿瘤的进展。USP28的失调已被证明可以加速许多癌症的进展, 将其作为癌症治疗的靶点可能是一个潜在的方向。然而, 除了促进癌症的进展, 最近的一些研究发现USP28可以参与抑制一些涉及黑色素瘤的癌症。因此, 仍需进一步研究USP28及其对癌症的影响。USP28在影响预后和癌症对各种治疗的敏感性方面的关键作用也已得到证实。然而, 一些相关的机制尚不清楚, 其临床应用的效果和安全性仍需要更多的实验。

尽管USP28抑制剂的研究取得了一定的进展, 一些化合物通过抑制USP28展现了抗癌作用, 但需要指出的是, USP28抑制剂存在结构种类较少、活性不足、选择性较低、缺乏成药性研究等问题。USP家族成员间的高度同源性, 为开发高选择性抑制剂带来了极大的挑战。近年来, 文献相继报道了USP25和USP28的蛋白结构和分子特征差异与活性调节的关系^{[43, 44]}、USP28与小分子抑制剂的共晶^[40]。根据以上研究, 发掘对USP28活性调节具有重要作用的泛素特异性结合新口袋, 通过竞争性削弱泛素与USP28的结合, 可能为实现USP28的选择性抑制提供新策略, 并有望获得活性高、成药性良好的USP28抑制剂。针对USP28进行癌症治疗的临床试验和研究仍然不足, 这突出了探索更深层机制以阐明USP28抑制剂对癌症患者是否安全有效的重要性。

作者贡献: 李幸是本文的第一作者, 负责文献资料的收集及文章的撰写; 张博雅、陆礼婷负责图表的绘制和完善; 王燕、文帅提出文章修改建议; 牟伊为本文的通讯作者, 负责提供撰写思路和内容框架组织等工作; 徐向明、臧永军为本文的共同通讯作者, 负责稿件修改完善等工作。

利益冲突: 所有作者均声明无利益冲突。

参考文献

收起

文献

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参考文献引证文献

排序方式：

[1]

Swatek KN, Komander D. Ubiquitin modifications [J]. Cell Res, 2016, 26: 399-422.

[2]

Pickart CM, Fushman D. Polyubiquitin chains: polymeric protein signals [J]. Curr Opin Chem Biol, 2004, 8: 610-616.

[3]

Nandi D, Tahiliani P, Kumar A, et al. The ubiquitin-proteasome system [J]. J Biosci, 2006, 31: 137-155.

[4]

Zheng N, Shabek N. Ubiquitin ligases: structure, function, and regulation [J]. Annu Rev Biochem, 2017, 86: 129-157.

[5]

Senft D, Qi J, Ronai ZA. Ubiquitin ligases in oncogenic transformation and cancer therapy [J]. Nat Rev Cancer, 2018, 18: 69-88.

[6]

Qi SM, Cheng G, Cheng XD, et al. Targeting USP7-mediated deubiquitination of MDM2/MDMX-p53 pathway for cancer therapy: are we there yet? [J]. Front Cell Dev Biol, 2020, 8: 233.

[7]

Stone SL. Role of the ubiquitin proteasome system in plant response to abiotic stress [J]. Int Rev Cell Mol Biol, 2019, 343: 65-110.

[8]

Dittmar G, Winklhofer KF. Linear ubiquitin chains: cellular functions and strategies for detection and quantification [J]. Front Chem, 2019, 7: 915.

[9]

Hicke L. Protein regulation by monoubiquitin [J]. Nat Rev Mol Cell Biol, 2001, 2: 195-201.

[10]

Jackson SP, Durocher D. Regulation of DNA damage responses by ubiquitin and SUMO [J]. Mol Cell, 2013, 49: 795-807.

[11]

Muratani M, Tansey WP. How the ubiquitin-proteasome system controls transcription [J]. Nat Rev Mol Cell Biol, 2003, 4: 192-201.

[12]

Jesenberger V, Jentsch S. Deadly encounter: ubiquitin meets apoptosis [J]. Nat Rev Mol Cell Biol, 2002, 3: 112-121.

[13]

McClellan AJ, Laugesen SH, Ellgaard L. Cellular functions and molecular mechanisms of non-lysine ubiquitination [J]. Open Biol, 2019, 9: 190147.

[14]

Fennell LM, Rahighi S, Ikeda F. Linear ubiquitin chain-binding domains [J]. FEBS J, 2018, 285: 2746-2761.

[15]

Komander D, Clague MJ, Urbé S. Breaking the chains: structure and function of the deubiquitinases [J]. Nat Rev Mol Cell Biol, 2009, 10: 550-563.

[16]

Sowa ME, Bennett EJ, Gygi SP, et al. Defining the human deubiquitinating enzyme interaction landscape [J]. Cell, 2009, 138: 389-403.

[17]

Harrigan JA, Jacq X, Martin NM, et al. Deubiquitylating enzymes and drug discovery: emerging opportunities [J]. Nat Rev Drug Discov, 2018, 17: 57-78.

[18]

Prieto-Garcia C, Tomašković I, Shah VJ, et al. USP28: oncogene or tumor suppressor? A unifying paradigm for squamous cell carcinoma [J]. Cells, 2021, 10: 2652.

[19]

Popov N, Wanzel M, Madiredjo M, et al. The ubiquitin-specific protease USP28 is required for MYC stability [J]. Nat Cell Biol, 2007, 9: 765-774.

[20]

Meitinger F, Anzola JV, Kaulich M, et al. 53BP1 and USP28 mediate p53 activation and G1 arrest after centrosome loss or extended mitotic duration [J]. J Cell Biol, 2016, 214: 155-166.

[21]

Chen L, Xu Z, Li Q, et al. USP28 facilitates pancreatic cancer progression through activation of Wnt/β-catenin pathway via stabilising FOXM1 [J]. Cell Death Dis, 2021, 12: 887.

[22]

Cui B, Luo Y, Tian P, et al. Stress-induced epinephrine enhances lactate dehydrogenase A and promotes breast cancer stem-like cells [J]. J Clin Invest, 2019, 129: 1030-1046.

[23]

Du SC, Zhu L, Wang YX, et al. SENP1-mediated deSUMOylation of USP28 regulated HIF-1α accumulation and activation during hypoxia response [J]. Cancer Cell Int, 2019, 19: 4.

[24]

Zhang D, Zaugg K, Mak TW, et al. A role for the deubiquitinating enzyme USP28 in control of the DNA-damage response [J]. Cell, 2006, 126: 529-542.

[25]

Lambrus BG, Daggubati V, Uetake Y, et al. A USP28-53BP1-p53-p21 signaling axis arrests growth after centrosome loss or prolonged mitosis [J]. J Cell Biol, 2016, 214: 143-153.

[26]

Wu Y, Wang Y, Yang XH, et al. The deubiquitinase USP28 stabilizes LSD1 and confers stem-cell-like traits to breast cancer cells [J]. Cell Rep, 2013, 5: 224-236.

[27]

Close V, Close W, Kugler S, et al. Notch1 signaling is activated in CLL by mutations of FBXW7 and low expression of USP28 at 11q23 [J]. Blood, 2018, 132: 946.

[28]

Popov N, Herold S, Llamazares M, et al. Fbw7 and usp28 regulate MYC protein stability in response to DNA damage [J]. Cell Cycle, 2014, 6: 2327-2331.

[29]

Diefenbacher ME, Popov N, Blake SM, et al. The deubiquitinase USP28 controls intestinal homeostasis and promotes colorectal cancer [J]. J Clin Invest, 2014, 12: 3407-3418.

[30]

Han H, Sun D, Li W, et al. A c-MYC-microRNA functional feedback loop affects hepatocarcinogenesis [J]. Hepatology, 2013, 57 : 2378-2389.

[31]

Richter K, Paakkola T, Mennerich D, et al. USP28 deficiency promotes breast and liver carcinogenesis as well as tumor angiogenesis in a HIF-independent manner [J]. Mol Cancer Res, 2018, 16: 1000-1012.

[32]

Wang X, Liu Z, Zhang L, et al. Targeting deubiquitinase USP28 for cancer therapy [J]. Cell Death Dis, 2018, 9: 186.

[33]

Wang Z, Song Q, Xue J, et al. Ubiquitin-specific protease 28 is overexpressed in human glioblastomas and contributes to glioma tumorigenicity by regulating MYC expression [J]. Exp Biol Med (Maywood), 2016, 241: 255-264.

[34]

Zhang L, Xu B, Qiang Y, et al. Overexpression of deubiquitinating enzyme USP28 promoted non-small cell lung cancer growth [J]. J Cell Mol Med, 2015, 19: 799-805.

[35]

Wrigley JD, Gavory G, Simpson I, et al. Identification and characterization of dual inhibitors of the USP25/28 deubiquitinating enzyme subfamily [J]. ACS Chem Biol, 2017, 12: 3113-3125.

[36]

Wang H, Meng Q, Ding Y, et al. USP28 and USP25 are downregulated by vismodegib in vitro and in colorectal cancer cell lines [J]. FEBS J, 2020, 288: 1325-1342.

[37]

Peng J, Jiang K, Sun X, et al. Identification of a class of potent USP25/28 inhibitors with broad-spectrum anti-cancer activity [J]. Signal Transduct Target Ther, 2022, 7: 393.

[38]

Liu Z, Zhao T, Li Z, et al. Discovery of [1,2,3]triazolo[4,5-d]pyrimidine derivatives as highly potent, selective, and cellularly active USP28 inhibitors [J]. Acta Pharm Sin B, 2020, 10: 1476-1491.

[39]

Altun M, Kramer HB, Willems LI, et al. Activity-based chemical proteomics accelerates inhibitor development for deubiquitylating enzymes [J]. Chem Biol, 2011, 18: 1401-1412.

[40]

Zhou D, Xu Z, Huang Y, et al. Structure-based discovery of potent USP28 inhibitors derived from vismodegib [J]. Eur J Med Chem, 2023, 254: 115369.

[41]

Valero R, Bayés M, Francisca Sánchez-Font M, et al. Characterization of alternatively spliced products and tissue-specific isoforms of USP28 and USP25 [J]. Genome Biol, 2001, 2: Research0043.

[42]

Zhen Y, Knobel PA, Stracker TH, et al. Regulation of USP28 deubiquitinating activity by SUMO conjugation [J]. J Biol Chem, 2014, 289: 34838-34850.

[43]

Gersch M, Wagstaff JL, Toms AV, et al. Distinct USP25 and USP28 oligomerization states regulate deubiquitinating activity [J]. Mol Cell, 2019, 74: 436-451.e7.

[44]

Sauer F, Klemm T, Kollampally RB, et al. Differential oligomerization of the deubiquitinases USP25 and USP28 regulates their activities [J]. Mol Cell, 2019, 74: 421-435.e10.

[45]

Zhou L, Qin B, Yassine DM, et al. Structure and function of the highly homologous deubiquitinases ubiquitin specific peptidase 25 and 28: insights into their pathophysiological and therapeutic roles [J]. Biochem Pharmacol, 2023, 213: 115624.

[46]

Hu M, Li P, Song L, et al. Structure and mechanisms of the proteasome-associated deubiquitinating enzyme USP14 [J]. EMBO J, 2005, 24: 3747-3756.

[47]

Hu M, Li P, Li M, et al. Crystal structure of a UBP-family deubiquitinating enzyme in isolation and in complex with ubiquitin aldehyde [J]. Cell, 2002, 111: 1041-1054.

[48]

Ye Y, Scheel H, Hofmann K, et al. Dissection of USP catalytic domains reveals five common insertion points [J]. Mol Biosyst, 2009, 5: 1797-1808.

[49]

Chen B, Sang Y, Song X, et al. Exosomal miR-500a-5p derived from cancer-associated fibroblasts promotes breast cancer cell proliferation and metastasis through targeting USP28 [J]. Theranostics, 2021, 11: 3932-3947.

[50]

Chakravorty D, Ghosh A, Saha S. Computational approach to target USP28 for regulating MYC [J]. Comput Biol Chem, 2020, 85: 107208.

[51]

Diefenbacher ME, Chakraborty A, Blake SM, et al. Usp28 counteracts Fbw7 in intestinal homeostasis and cancer [J]. Cancer Res, 2015, 75: 1181-1186.

[52]

Ren X, Jiang M, Ding P, et al. Ubiquitin-specific protease 28: the decipherment of its dual roles in cancer development [J]. Exp Hematol Oncol, 2023, 12: 27.

[53]

Schülein-Völk C, Wolf E, Zhu J, et al. Dual regulation of Fbw7 function and oncogenic transformation by Usp28 [J]. Cell Rep, 2014, 9: 1099-1109.

[54]

Flügel D, Görlach A, Kietzmann T. GSK-3β regulates cell growth, migration, and angiogenesis via Fbw7 and USP28-dependent degradation of HIF-1α [J]. Blood, 2012, 119: 1292-1301.

[55]

Sun X, Cai M, Wu L, et al. Ubiquitin‐specific protease 28 deubiquitinates TCF7L2 to govern the action of the Wnt signaling pathway in hepatic carcinoma [J]. Cancer Sci, 2022, 113: 3463-3475.

[56]

Müller I, Strozyk E, Schindler S, et al. Cancer cells employ nuclear caspase-8 to overcome the p53-dependent G2/M checkpoint through cleavage of USP28 [J]. Mol Cell, 2020, 77: 970-984.e7.

[57]

Prieto‐Garcia C, Hartmann O, Reissland M, et al. Maintaining protein stability of ∆Np63 via USP28 is required by squamous cancer cells [J]. EMBO Mol Med, 2020, 12: e11101.

[58]

Li F, Han H, Sun Q, et al. USP28 regulates deubiquitination of histone H2A and cell proliferation [J]. Exp Cell Res, 2019, 379: 11-18.

[59]

Zhang H, Huang H, Feng X, et al. Deubiquitinase USP28 inhibits ubiquitin ligase KLHL2-mediated uridine-cytidine kinase 1 degradation and confers sensitivity to 5'-azacytidine-resistant human leukemia cells [J]. Theranostics, 2020, 10: 1046-1059.

[60]

Kim H, Choi H, Im JS, et al. Stable maintenance of the Mre11-Rad50-Nbs1 complex is sufficient to restore the DNA double-strand break response in cells lacking RecQL4 helicase activity [J]. Biol Chem, 2021, 297: 101148.

[61]

Li YY, Wu C, Chen SM, et al. BRAF inhibitor resistance enhances vulnerability to arginine deprivation in melanoma [J]. Oncotarget, 2016, 7: 17665-17680.

[62]

Saei A, Palafox M, Benoukraf T, et al. Loss of USP28 drives resistance to BRAF targeted therapy in melanoma [J]. Ann Oncol, 2017, 28: 442.

[63]

Saei A, Palafox M, Benoukraf T, et al. Loss of USP28-mediated BRAF degradation drives resistance to RAF cancer therapies [J]. J Exp Med, 2018, 215: 1913-1928.

[64]

Zhou W, Chen J, Wang J. Comprehensive prognostic and immunological analysis of ubiquitin specific peptidase 28 in pan-cancers and identification of its role in hepatocellular carcinoma cell lines [J]. Aging (Albany NY), 2023, 15: 6545-6576.

[65]

Wang J, Dong Y, Ma H, et al. The deubiquitinase USP28 stabilizes the expression of RecQ family helicases and maintains the viability of triple negative breast cancer cells [J]. J Biol Chem, 2022, 298: 101443.

[66]

Zhang JF. MicroRNA-216b suppresses the cell growth of hepatocellular carcinoma by inhibiting ubiquitin-specific peptidase 28 expression [J]. Kaohsiung J Med Sci, 2020, 36: 423-428.

[67]

Li P, Huang Z, Wang J, et al. Ubiquitin-specific peptidase 28 enhances STAT3 signaling and promotes cell growth in non-small-cell lung cancer [J]. Onco Targets Ther, 2019, 12: 1603-1611.

[68]

Ren K, Li Y, Lu H, et al. miR-3940-5p functions as a tumor suppressor in non-small cell lung cancer cells by targeting cyclin D1 and ubiquitin specific peptidase-28 [J]. Transl Oncol, 2017, 10: 80-89.

[69]

Ruiz EJ, Pinto-Fernandez A, Turnbull AP, et al. USP28 deletion and small-molecule inhibition destabilizes c-MYC and elicits regression of squamous cell lung carcinoma [J]. Elife, 2021, 10: e71596.

[70]

Shen J, Xie M, Xu Y, et al. Identification of the deubiquitinase USP28 as a novel molecular therapeutic target of ovarian cancer [J]. Biochem Biophys Res Commun, 2023, 638: 184-191.

[71]

Zhao LJ, Zhang T, Feng XJ, et al. USP28 contributes to the proliferation and metastasis of gastric cancer [J]. J Cell Biochem, 2019, 120: 7657-7666.

[72]

Hu Y, Yu K, Wang G, et al. Lanatoside C inhibits cell proliferation and induces apoptosis through attenuating Wnt/β-catenin/c-MYC signaling pathway in human gastric cancer cell [J]. Biochem Pharmacol, 2018, 150: 280-292.

[73]

Wu J, Kumar S, Wang F, et al. Chemical approaches to intervening in ubiquitin specific protease 7 (USP7) function for oncology and immune oncology therapies [J]. J Med Chem, 2017, 61: 422-443.

[74]

Li X, Yang S, Zhang H, et al. Discovery of orally bioavailable N-benzylpiperidinol derivatives as potent and selective USP7 inhibitors with in vivo antitumor immunity activity against colon cancer [J]. J Med Chem, 2022, 65: 16622-16639.

[75]

Liu ZZ. Screening and Antitumor Activity of USP28 Inhibitors (USP28抑制剂的筛选及抗肿瘤活性研究) [D]. Zhengzhou: Zhengzhou University, 2019.

[76]

Ge H, Zhang D, Shi M, et al. Antiproliferative activity and potential mechanism of marine-sourced streptoglutarimide H against lung cancer cells [J]. Mar Drugs, 2021, 19: 79.

[77]

Tang M, Xie X, Shi M, et al. Antileukemic effect of caffeic acid 3, 4-dihydroxyphenetyl ester. Evidences for its mechanisms of action [J]. Phytomedicine, 2021, 80: 153383.

[78]

Jung JE, Kim HS, Lee CS, et al. Caffeic acid and its synthetic derivative CADPE suppress tumor angiogenesis by blocking STAT3-mediated VEGF expression in human renal carcinoma cells [J]. Carcinogenesis, 2007, 28: 1780-1787.

[79]

Prassas I, Diamandis EP. Novel therapeutic applications of cardiac glycosides [J]. Nat Rev Drug Discov, 2008, 7: 926-935.

[80]

Xu Z, Wang H, Meng Q, et al. Otilonium bromide acts as a selective USP28 inhibitor and exhibits cytotoxic activity against multiple human cancer cell lines [J]. Biochem Pharmacol, 2023, 215: 115746.

[81]

Liu X, Du J, Qian M, et al. Research on antitumor effects of small molecule inhibitors of deubiquitinases: new progress and new ideas [J]. Acta Pharm Sin (药学学报), 2022, 57: 547-556.

[82]

Fan G, Zhu H, Fang Y, et al. The roles of deubiquitinating enzymes in cancer therapy [J]. Acta Pharm Sin (药学学报), 2018, 53: 169-176.

[83]

Xu J, Li H, Ba R, et al. Advances in developing small molecule inhibitors of ubiquitin-specific protease 1 [J]. Acta Pharm Sin (药学学报), 2024, 59: 866-885.

[84]

Fang Y, Jiang L, He Q, et al. Deubiquitination complex platform: a plausible mechanism for regulating the substrate specificity of deubiquitinating enzymes [J]. Acta Pharm Sin B, 2023, 13: 2955-2962.

2024年第59卷第11期

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doi: 10.16438/j.0513-4870.2024-0098

接收时间：2024-01-30
首发时间：2025-11-24
出版时间：2024-11-12

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收稿日期：2024-01-30
修回日期：2024-05-13

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1.泰州学院医药与化学化工学院, 江苏泰州 225300

2.皖西学院生物与制药工程学院, 安徽六安 237012

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^*臧永军, E-mail: zangyongjun@yeah.net;

徐向明, E-mail: 379648107@qq.com;

牟伊, E-mail: mouyicpu@163.com

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2种不同金属材料的力学参数

科 Family	属数 Number of genus	种数 Number of species	占总种数比例 Percentage of total species (%)	属 Genus	种数 Number of species	占总种数比例 Percentage of total species (%)
鹅膏菌科Amanitaceae	2	11	5.26	鹅膏菌属 Amanita	10	4.78
小菇科 Mycenaceae	2	12	5.74	丝盖伞属 Inocybe	5	2.39
多孔菌科 Polyporaceae	8	14	6.70	蜡蘑属 Laccaria	5	2.39
红菇科 Russulaceae	3	23	11.00	小皮伞属 Marasmius	6	2.87
				小菇属 Mycena	11	5.26
				光柄菇属 Pluteus	5	2.39
				红菇属 Russula	17	8.13
				栓菌属 Trametes	5	2.39

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Substrate	Function of USP28 on the substrates	Reference
c-MYC	Deubiquitinating and stabilizing c-MYC by directly binding it or mediated by FBW7	[19, 51, 52]
FBW7	Antagonizing its autocatalytic ubiquitination and the ubiquitination of its substrates	[53]
HIF-1α	Revising the destabilization caused by FBW7	[54]
TCF7L2	Directly interacting with and deubiquitinating it by antagonizing FBW7, further regulating the Wnt/β-catenin signalling	[55]
P53	Deubiquitinating p53 through antagonizing MDM2	[52, 56, 57]
ΔNp63	Removing the K-48 linked ubiquitin chains induced by MDM2	[52, 56, 57]
FOXM1	Directly interacting with it and promoting its stabilization	[21]
LSD1	Deubiquitinating p53 through antagonizing MDM2	[26]
H2A	Decreasing the K119 ubiquitination level	[52, 58]
UCK1	Antagonizing its ubiquitination caused by KLHL2	[59]
NBS1	Promoting its stabilization by deubiquitination	[60]

Substrate

Function of USP28 on the substrates

Reference

c-MYC

Deubiquitinating and stabilizing c-MYC by directly binding it or mediated by FBW7

[19, 51, 52]

FBW7

Antagonizing its autocatalytic ubiquitination and the ubiquitination of its substrates

[53]

HIF-1α

Revising the destabilization caused by FBW7

[54]

TCF7L2

Directly interacting with and deubiquitinating it by antagonizing FBW7, further regulating the Wnt/β-catenin signalling

[55]

P53

Deubiquitinating p53 through antagonizing MDM2

[52, 56, 57]

ΔNp63

Removing the K-48 linked ubiquitin chains induced by MDM2

[52, 56, 57]

FOXM1

Directly interacting with it and promoting its stabilization

[21]

LSD1

Deubiquitinating p53 through antagonizing MDM2

[26]

H2A

Decreasing the K119 ubiquitination level

[52, 58]

UCK1

Antagonizing its ubiquitination caused by KLHL2

[59]

NBS1

Promoting its stabilization by deubiquitination

[60]

Cancer type	Expression level of Usp28	Mechanism/clinicopathological features	Protumoral/antitumoral
Colorectal cancer	Overexpressed	USP28 antagonizes the ubiquitin-dependent degradation of intestinal carcinogens c-MYC, c-JUN and Notch1, and promotes the proliferation of tumor cells.	Protumoral
Melanoma	Overexpressed	USP28 stabilizes the c-MYC level and hence increases ASS1 transcription upon arginine deprivation, and consequently leads to cell survival.	Protumoral
Deleted in a proportion of patients	Loss of Usp28 stabilizes BRAF enhancing downstream MAPK activation, promotes drug resistance of tumor cells and reduces the overall survival rate of patients.	Antitumoral
Breast cancer	Overexpressed	USP28 deubiquitinates and up-regulates LSD1, which can accelerate the formation of tumor stem cell-like features and tumor formation; the indirect suppression function of USP28 in CLDN7, which is mediated by LSD1, can promote the invasiveness of cancers; high serum epinephrine activates LDHA/USP28/MYC/SLUG signaling, associated with poor prognosis of breast cancer, and promotes the survival, proliferation and metastasis of breast cancer cells.	Protumoral
Overexpressed	Overexpression of Usp28 correlates with a better survival in patients with invasive ductal breast carcinoma; lack of Usp28 promotes a more malignant state of breast cancer cells.	Antitumoral
Liver cancer	Overexpressed	The expression levels of USP28 inversely correlate with patient survival; USP28 promotes the activity of the Wnt/β-catenin signaling pathway through maintaining the stability of TCF7L2; USP28 depletion or inhibition by a small molecule inhibitor leads to a halt of growth in liver cancer cells.	Protumoral
/	Lack of Usp28 results in advanced tumor development in a mouse xenograft model as well as in a chemically induced liver cancer mouse model.	Antitumoral
NSCLC	Overexpressed	Overexpression of Usp28 promotes NSCLC cells growth by increasing the stability of STAT3 and is correlated with the poor prognosis and low patient survival rate.	Protumoral
Other cancers	Overexpressed	The overexpression of Usp28 is related to the degree of differentiation and metastasis of gastric cancer cells. Usp28 is highly expressed in PC and is significantly associated with the malignant phenotype and shorter survival. USP28 mediates the activation of Wnt/β-catenin pathway through FOXM1 and promotes the growth of PC. Overexpression of Usp28 can promote the malignant phenotype of OV.	Protumoral

Cancer type

Expression level of Usp28

Mechanism/clinicopathological features

Protumoral/antitumoral

Colorectal cancer

Overexpressed

USP28 antagonizes the ubiquitin-dependent degradation of intestinal carcinogens c-MYC, c-JUN and Notch1, and promotes the proliferation of tumor cells.

Protumoral

Melanoma

Overexpressed

USP28 stabilizes the c-MYC level and hence increases ASS1 transcription upon arginine deprivation, and consequently leads to cell survival.

Protumoral

Deleted in a proportion of patients

Loss of Usp28 stabilizes BRAF enhancing downstream MAPK activation, promotes drug resistance of tumor cells and reduces the overall survival rate of patients.

Antitumoral

Breast cancer

Overexpressed

USP28 deubiquitinates and up-regulates LSD1, which can accelerate the formation of tumor stem cell-like features and tumor formation; the indirect suppression function of USP28 in CLDN7, which is mediated by LSD1, can promote the invasiveness of cancers; high serum epinephrine activates LDHA/USP28/MYC/SLUG signaling, associated with poor prognosis of breast cancer, and promotes the survival, proliferation and metastasis of breast cancer cells.

Protumoral

Overexpressed

Overexpression of Usp28 correlates with a better survival in patients with invasive ductal breast carcinoma; lack of Usp28 promotes a more malignant state of breast cancer cells.

Antitumoral

Liver cancer

Overexpressed

The expression levels of USP28 inversely correlate with patient survival; USP28 promotes the activity of the Wnt/β-catenin signaling pathway through maintaining the stability of TCF7L2; USP28 depletion or inhibition by a small molecule inhibitor leads to a halt of growth in liver cancer cells.

Protumoral

Lack of Usp28 results in advanced tumor development in a mouse xenograft model as well as in a chemically induced liver cancer mouse model.

Antitumoral

NSCLC

Overexpressed

Overexpression of Usp28 promotes NSCLC cells growth by increasing the stability of STAT3 and is correlated with the poor prognosis and low patient survival rate.

Protumoral

Other cancers

Overexpressed

The overexpression of Usp28 is related to the degree of differentiation and metastasis of gastric cancer cells. Usp28 is highly expressed in PC and is significantly associated with the malignant phenotype and shorter survival. USP28 mediates the activation of Wnt/β-catenin pathway through FOXM1 and promotes the growth of PC. Overexpression of Usp28 can promote the malignant phenotype of OV.

Protumoral