泛素-蛋白酶体抑制剂用于肿瘤治疗

目的:综述近几年国内外有关蛋白酶体抑制剂的研究进展。方法:查阅相关文献,并进行分析、归纳、综述。结果:蛋白酶体抑制剂已成为肿瘤治疗的一个新靶点,在临床前期及临床试验中已显示出了抗肿瘤疗效。结论:泛素-蛋白酶体抑制剂有望成为抗肿瘤、抗炎、抗病毒以及神经系统疾病治疗的有效药物。     

泛素-蛋白酶体抑制剂对大鼠肺纤维化的影响

目的:探讨泛素-蛋白酶体抑制剂MG-132对大鼠肺纤维化的影响。方法:45只SD大鼠随机分成对照组、模型组、干预组和治疗组。模型组和干预、治疗组大鼠气管内滴入博来霉素(5mg.kg-1)建立肺纤维化动物模型,对照组以等量生理盐水气管内滴入。干预组于第造模后第1天起给予MG-132腹腔内注射(0.1mg.kg-1.d-1),模型组和对照组注射等量生理盐水;治疗组于造模后第28天起给予MG-132(0.1mg.kg-1.d-1)腹腔内注射。于不同时间点测肺系数、肺组织行病理学检查及肺组织羟脯氨酸(HYP)含量测定。结果:干预组及治疗组肺系数及肺组织HYP含量各时间点均低于模型组(P<0.01,P<0.05),肺泡炎的程度也轻于模型组;虽然干预组和治疗组肺纤维化程度均明显重于对照组,但干预组28d和治疗组肺纤维化较模型组轻。结论:泛素-蛋白酶体抑制剂MG-132对大鼠肺纤维化有一定的干预、治疗作用。     

蛋白酶体与蛋白酶体抑制剂

蛋白酶体是一种存在于细胞质和细胞核内的蛋白水解酶复合物,是降解细胞内蛋白质的主要酶系统。蛋白酶体涉及机体的多种生理功能和许多疾病的发病机制,近年来已成为基础和临床研究的热点之一。在此对蛋白酶体的结构、功能及其抑制剂的研究进展进行综述。     

泛素-蛋白酶体抑制剂对血管平滑肌细胞凋亡的影响

血管平滑肌细胞(Vascular smooth muscle,VSMC)是构成血管壁的重要成分。在动脉粥样硬化(Atherosclerosis,AS)病变中,VSMC在病理状态下凋亡失平衡引起血管重塑,导致动脉粥样硬化的发生。目前研究发现,细胞凋亡在动脉粥样硬化形成的发病和治疗中具有重要意义,而泛素-蛋白酶体抑制剂(The ubiquitinproteasome inhibitor,UPI)能够促进VSM C凋亡进而影响动脉粥样硬化。本文对泛素-蛋白酶体抑制剂影响VSM C凋亡的机制进行了综述。     

泛素-蛋白酶体通路:从理论到临床

真核细胞内大多数蛋白质降解过程的调控是通过泛素-蛋白酶体通路(UPP)进行的。该过程分为两个步骤:①蛋白质的泛素化:一个或多个泛素分子通过共价键连接在目标蛋白质上;②多泛素化的蛋白质被26S蛋白酶体降解成寡肽。26S蛋白酶体是一个多亚单位的蛋白酶复合体,它可分为一个19S具     

去泛素化酶小分子抑制剂抗肿瘤作用研究:新进展和新思路

泛素-蛋白酶体通路异常是导致蛋白质内稳态失控的重要因素。而在该过程中,负责移除蛋白底物泛素链的去泛素化酶至关重要,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。基于此,靶向去泛素化酶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。本综述将重点介绍泛素-蛋白酶体通路、尤其是去泛素化酶对肿瘤的调控作用和机制,介绍去泛素化酶小分子抑制剂在肿瘤治疗研究中的应用,并针对小分子抑制剂的研究现状和最新进展展开讨论,为基于去泛素化酶的抗肿瘤新策略研究提供思路。     

泛素-蛋白酶体通路在心血管疾病中的作用

近年来,泛素-蛋白酶体通路(ubiquitin-proteasome pathway,UPP)作为细胞内蛋白质非溶解体性降解的主要途径已受到人们的日益重视.UPP广泛参与机体多种代谢活动,在调节细胞的各种生物途径中发挥重要作用.新近的研究资料表明UPP与动脉粥样硬化及冠心病介入治疗(PCI)后再狭窄有关.但UPP在心血管系统的研究目前仍处于起步阶段.本文就UPP在心血管系统疾病中作用的研究进展作一综述.     

蛋白酶体抑制剂的研究进展

作为体内蛋白质降解的途径之一,蛋白酶体具有非常重要的生理作用,并与多种疾病密切相关。抑制蛋白酶体的功能已经成为肿瘤治疗的叉一有前景的新途径,并受到越来越多的关注。本文对蛋白酶体的组成结构、病理生理作用和现有抑制剂进行归纳总结。     

蛋白酶体抑制剂MG132诱导HepG2细胞凋亡及其机制研究

目的观察蛋白酶体抑制剂MG132对人肝癌细胞HepG2的致凋亡作用,并从泛素蛋白酶体途径(UPP)相关基因E1、E2和E3及天冬氨酸特异的半胱氨酸蛋白酶3(Caspase3)表达初步探讨MG132的致细胞凋亡机制。方法采用多个浓度(2、5、10μmol·L-1)的蛋白酶体抑制剂MG132处理HepG2细胞;流式细胞术检测细胞周期和细胞凋亡率,DNA琼脂糖凝胶电泳检测细胞凋亡;逆转录聚合酶链反应(RTPCR)检测UPP相关基因E1、E2、E3和凋亡相关基因Caspase3的转录水平;免疫细胞化学检测Caspase3蛋白表达。结果对照组HepG2细胞凋亡率低于5%,在2、5和10μmol·L-1MG132作用下,细胞凋亡率分别为42.9%、66.1%、72.8%,MG132诱导HepG2细胞凋亡具有量效关系;RTPCR检测发现细胞内UPP相关基因E1、E2、E3mRNA表达下降,而凋亡相关基因Caspase3mRNA表达上调;免疫组化检测Caspase3蛋白表达水平升高。结论蛋白酶体抑制剂MG132能够诱导HepG2细胞凋亡,其机制可能与MG132抑制UPP活性,使细胞内Caspase3蛋白降解减少,同时上调Caspase3基因转录,促进细胞凋亡。     

抗肿瘤治疗新药蛋白酶体抑制剂Bortezomib的临床应用

蛋白酶体抑制剂(proteasom e inh ib itor)通过抑制26S蛋白酶体活性,激活胱冬肽酶(caspase)-3诱导凋亡,抑制核因子(NF)-κB依赖性基因转录,影响细胞内多种信号级联,破坏维持机体正常内稳态的机制导致肿瘤细胞生长延迟或死亡。V e lcade(化学名Bortezom ib,前用名PS-341)是美国FDA批准的第一个已供临床应用的蛋白酶体抑制剂。临床用于治疗初治或复发性、难治性多发性骨髓瘤,其次为造血组织恶性肿瘤及进展性实体瘤等显示具有良好的抗瘤活性,安全性好,不良反应少。其临床应用研究正在迅速扩展中。     

Proteasome inhibitor inhibits proliferation and induces apoptosis in renal interstitial fibroblasts

BackgroundThe ubiquitin proteasome pathway plays a pivotal role in controlling cell proliferation, apoptosis and differentiation in a variety of normal and tumor cells. This study aimed to investigate the role of a proteasome inhibitor on proliferation, apoptosis and related proteins in renal interstitial fibroblasts (NRK-49F).MethodsNRK-49F cells were induced using transforming growth factor-β1 (TGF-β1) and pretreated with the proteasome inhibitor MG-132. Cell proliferation was measured using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT). The cell cycle and apoptosis were analyzed using flow cytometry. Apoptosis was also analyzed using a DNAladder. The protein expression of p53, p27, p21, caspase-3, Bcl-2 and Bax was examined using western blots.ResultsThe results showed that TGF-β1 (5 ng/ml) can stimulate the proliferation of NRK-49F cells.MG-132 (0.25–5 μM) inhibited TGF-β1-induced proliferation in a dose-dependent manner through G1-arrest; TGF-β1 alone did not induce apoptosis (3.8 ± 0.4% vs. 4.7 ± 1.6%). However, pretreatment with MG-132 significantly induced apoptosis in TGF-β1-stimulated NRK-49F cells in a dosedependentmanner. AtypicalDNAladderwas also confirmed in these two groups.Western blot analysis showed thatMG-132 activated p53, p21, caspase-3 and Bax, and inhibited Bcl-2 in a dose-dependent manner, while p27 expression remained unchanged.ConclusionsA proteasome inhibitor inhibited proliferation and induced apoptosis in renal interstitial fibroblasts stimulated by TGF-β1. The mechanism may relate to the p53, p21, caspase-3, Bcl-2 and Bax pathways. Our results suggest that a proteasome inhibitor could be a new strategy to treat renal interstitial fibrosis.     

A review of the proteasome inhibitor bortezomib in multiple myeloma

Multiple myeloma (MM) represented 14% of new haematological malignancies in the US in 2003 and almost 19% of anticipated deaths. Treatment with standard chemotherapy has resulted in a median survival of about 3 years and despite the improvements in survival seen with the use of intensive therapy supported by autologous stem cell transplantation, MM remains incurable; hence, new therapeutic strategies are urgently needed. One novel approach to the treatment of MM is the use of proteasome inhibitors. Proteasomes are ubiquitous protease complexes involved in diverse aspects of cell biology, such as protein homeostasis, cell cycle progression, apoptosis and inflammation, as well as resistance to antineoplastic therapy. The first-in-class proteasome inhibitor, bortezomib was recently approved in the US for the treatment of patients with MM who have received at least two prior therapies and are progressing on their last therapy. Its use in earlier-stage MM, other haematological malignancies and in solid tumours as monotherapy and in combination therapy is currently under investigation.     

蛋白酶体抑制剂MG132诱导人血管平滑肌细胞凋亡及其机制研究

目的：观察蛋白酶体抑制剂MG132对人脐静脉血管平滑肌细胞（vascular smooth musele cells,VSMC）的致凋亡作用及其对凋亡相关的天冬氨酸特异的半胱氨酸蛋白酶3（caspase 3）表达的影响。方法：采用多个浓度（2．5、5、10μmol／L）的蛋白酶体抑制剂MG132处理人脐静脉VSMC 24h和48h;流式细胞术分析细胞凋亡率;RT-PCR检测泛素-蛋白酶体途径（ubiquitin-proteasome pathway,UPP）相关基因泛素、泛素活化酶（E1）、泛素缀合酶（E2）、泛素连接酶（E3）、26S蛋白酶体和caspase 3基因转录水平;免疫细胞化学检测细胞caspase 3蛋白表达。结果：蛋白酶体抑制剂MG132处理48h后细胞凋亡率增加：细胞内UPP相关的基因rnRNA表达降低,而caspase 3 mRNA表达增高;免疫细胞化学检测细胞caspase 3蛋白表达水平升高。结论：蛋白酶体抑制剂MG132能够诱导人脐静脉VSMC凋亡,其作用呈量-效关系;MG132诱导vSMC凋亡与下调UPP相关基因和上调caspase 3基因及蛋白的表达有关。     

Proteasome inhibitor MG-132 induces C6 glioma cell apoptosis via oxidative stress

Aim:

Proteasome inhibitors have been found to suppress glioma cell proliferation and induce apoptosis, but the mechanisms are not fully elucidated. In this study we investigated the mechanisms underlying the apoptosis induced by the proteasome inhibitor MG-132 in glioma cells.

Methods:

C6 glioma cells were used. MTT assay was used to analyze cell proliferation. Proteasome activity was assayed using Succinyl-LLVY-AMC, and intracellular ROS level was evaluated with the redox-sensitive dye DCFH-DA. Apoptosis was detected using fluorescence and transmission electron microscopy as well as flow cytometry. The expression of apoptosis-related proteins was investigated using Western blot analysis.

Results:

MG-132 inhibited C6 glioma cell proliferation in a time- and dose-dependent manner (the IC₅₀ value at 24 h was 18.5 μmol/L). MG-132 (18.5 μmol/L) suppressed the proteasome activity by about 70% at 3 h. It induced apoptosis via down-regulation of antiapoptotic proteins Bcl-2 and XIAP, up-regulation of pro-apoptotic protein Bax and caspase-3, and production of cleaved C-terminal 85 kDa PARP). It also caused a more than 5-fold increase of reactive oxygen species. Tiron (1 mmol/L) effectively blocked oxidative stress induced by MG-132 (18.5 μmol/L), attenuated proliferation inhibition and apoptosis in C6 glioma cells, and reversed the expression pattern of apoptosis-related proteins.

Conclusion:

MG-132 induced apoptosis of C6 glioma cells via the oxidative stress.     

PD-1抑制剂信迪利单抗的临床研究进展

信迪利单抗（商品名：达伯舒^®）是一种全人源化IgG4单克隆抗体,可与程序性细胞死亡受体-1（PD-1）结合,从而阻断PD-1与其配体（PD-L1和PL-L2）相互作用,从而有助于恢复内源性抗肿瘤T细胞反应。信迪利单抗由信达生物制药和礼来制药共同合作研发,已在我国批准用于至少经过二线系统化疗的复发或难治性经典型霍奇金淋巴瘤的治疗。目前正在我国进行多项Ⅰ期、Ⅱ期和Ⅲ期临床试验,开发用于多种实体肿瘤的治疗,包括非小细胞肺癌和食道癌。该药用于治疗实体瘤的Ⅰ/Ⅱ期临床试验也正在美国进行,美国FDA已于2018年1月接受信迪利单抗的研究性新药申请。本文对该药的作用机制、药动学和临床研究等进行总结,为更好地指导临床应用提供建议。     

半乳糖凝集素抑制剂及其抗肿瘤作用研究进展

目的：总结半乳糖凝集素（galectin ,Gal）相关抑制剂及其抗肿瘤作用的研究进展。方法查阅、分析相关文献并综述。结果 Gal是凝集素家族中的一员,在多种肿瘤细胞中都有表达,并参与肿瘤发生、发展和转移的多个过程。因此,可作为疾病,尤其是肿瘤的诊断、治疗的新靶点,其抑制剂也成为肿瘤治疗的新选择。结论 Gal与肿瘤的形成高度相关,可成为肿瘤治疗的新靶点。     

Proteasome inhibitor lactacystin ablates liver injury induced by intestinal ischaemia-reperfusion

1. The aim of the present study was to investigate the role of proteasome in the pathogenesis of liver injury induced by intestinal ischaemia-reperfusion (I/R) and the effect of the proteasome inhibitor lactacystin on neutrophil infiltration, intracellular adhesion molecule (ICAM)-1 and nuclear factor (NF)-kappaB expression in the liver tissues of rats. 2. Thirty-two Wistar rats were randomly divided into four groups (n = 8 in each group) as follows: (i) a control, sham-operated group; (ii) an I/R group subjected to 1 h intestinal ischaemia and 4 h reperfusion; (iii) a group pretreated with 0.2 mg/kg lactacystin 1 h before intestinal I/R; and (iv) a group pretreated with 0.6 mg/kg lactacystin 1 h before intestinal I/R. Liver and intestine histology were observed. Serum levels of aspartate aminotransferase (AST), alanine aminotransferase (ALT) and lactate dehydrogenase (LDH), as well as 20S proteasome activity in circulating white blood cells, were measured. Myeloperoxidase (MPO) activity in liver tissues and the immunohistochemical expression of liver NF-kappaB and ICAM-1 were assayed. In addition, a western blot of liver NF-kappaB was performed. 3. Compared with the sham-operated control group, liver and intestine injury was induced by intestinal I/R, characterized as histological damage including oedema, haemorrhage and infiltration by inflammatory cells, as well as a significant increase in serum AST (365 +/- 121 vs 546 +/- 297 IU/L, respectively; P < 0.05), ALT (65 +/- 23 vs 175 +/- 54 IU/L, respectively; P < 0.01) and LDH levels (733 +/- 383 vs 1434 +/- 890 IU/L, respectively; P < 0.05). Compared with the control group, MPO activity in the liver tissues increased significantly in the I/R group (2.05 +/- 0.69 vs 3.42 +/- 1.11 U/g, respectively; P < 0.05). Strong positive expression of liver ICAM-1 and NF-kappaB p65 was observed. 4. Compared with the intestinal I/R group, administration of 0.6 mg/kg lactacystin markedly reduced 20S proteasome activity in circulating white blood cells (15.47 +/- 4.00 vs 2.07 +/- 2.00 pmol 7-amino-4-methylcoumarin (AMC)/s per mg, respectively; P < 0.01) and ameliorated liver injury, which was demonstrated by decreased levels of serum AST (546 +/- 297 vs 367 +/- 86 IU/L, respectively; P < 0.05), ALT (175 +/- 54 vs 135 +/- 26 IU/L, respectively; P < 0.05) and LDH (1434 +/- 890 vs 742 +/- 218 IU/L, respectively; P < 0.05) and a reduced liver pathological score (2.13 +/- 0.64 vs 1.25 +/- 0.46, respectively; P < 0.01). Compared with the intestinal I/R group, MPO activity in liver tissues decreased significantly following lactacystin pretreatment (3.42 +/- 1.11 vs 2.58 +/- 0.61 U/g, respectively; P < 0.05) and the expression of liver NF-kappaB and ICAM-1 was markedly ameliorated. 5. The present study reveals that the proteasome inhibitor lactacystin ablates liver injury induced by intestinal I/R. One possible mechanism responsible for this effect is the inhibition of enhanced ICAM-1 and neutrophil infiltration by inhibition of NF-kappaB activity. The results suggest the feasibility of using proteasome inhibitor clinically in the treatment of intestinal I/R.     

Synthesis and Evaluation of Derivatives of the Proteasome Deubiquitinase Inhibitor b‐AP15

The ubiquitin–proteasome system (UPS) is increasingly recognized as a therapeutic target for the development of anticancer therapies. The success of the 20S proteasome core particle (20S CP) inhibitor bortezomib in the clinical management of multiple myeloma has raised the possibility of identifying other UPS components for therapeutic intervention. We previously identified the small molecule b‐AP15 as an inhibitor of 19S proteasome deubiquitinase (DUB) activity. Building upon our previous data, we performed a structure–activity relationship (SAR) study on b‐AP15 and identified VLX1570 as an analog with promising properties, including enhanced potency and improved solubility in aqueous solution. In silico modeling was consistent with interaction of VLX1570 with key cysteine residues located at the active sites of the proteasome DUBs USP14 and UCHL5. VLX1570 was found to inhibit proteasome deubiquitinase activity in vitro in a manner consistent with competitive inhibition. Furthermore, using active‐site‐directed probes, VLX1570 also inhibited proteasome DUB activity in exposed cells. Importantly, VLX1570 did not show inhibitory activity on a panel of recombinant non‐proteasome DUBs, on recombinant kinases, or on caspase‐3 activity, suggesting that VLX1570 is not an overtly reactive general enzyme inhibitor. Taken together, our data shows the chemical and biological properties of VLX1570 as an optimized proteasome DUB inhibitor.     

姜黄素前体药物的合成及其体外抗肿瘤活性研究

目的制备姜黄素前体药物,使其在肿瘤细胞内高选择性活化,为进一步开展肿瘤靶向性化疗奠定基础。方法以姜黄素分子结构为基础,化学合成N-马来酰-L-缬氨酸酯姜黄素、N-马来酰-甘氨酸酯姜黄素,红外光谱法进行鉴定;MTT比色分析法比较两种姜黄素前体药物作用6~24 h后,人膀胱癌EJ细胞及肾小管上皮HKC细胞生长抑制的差异。结果20~40μmol.L-1的N-马来酰-L-缬氨酸酯姜黄素、N-马来酰-甘氨酸酯姜黄素作用6~24 h后,EJ细胞生长抑制率分别为6.71%~65.13%(P<0.05)、10.96%~73.01%(P<0.05),呈浓度、时间依赖性。与同浓度姜黄素比较,两种前体药物对HKC细胞生长的抑制作用均降低(P<0.01)。结论本研究成功的合成了两种姜黄素的前体药物,N-马来酰-L-缬氨酸酯姜黄素、N-马来酰-甘氨酸酯姜黄素;二者均能体外抑制人膀胱癌EJ细胞增殖,其对人肾小管上皮HKC细胞的抑制毒性作用低于姜黄素。