Inactivation of Protein Tyrosine Phosphatase 1B （PTP1B） Activity by the Aqueous Partition of Guava Leaf Extract

Guava leaf tea has been used as a folk medicine for treating hyperglycemic conditions in Asia and Africa. The hypoglycemic efficacy of guava leaf has been documented by many scientists in these regions, but the hypoglycemic mechanism is poorly understood. Guava leaves were extracted with methanol and the crude extract was partitioned against hexane, ethyl acetate, and butanol in sequence. The leftover in water is defined as the aqueous partition. A second smaller batch was extracted with hot water directly. Oral glucose tolerance test was carried out on healthy mice instead of diabetic mice that lack endogenous insulin. Glucose uptake was examined with 3T3-L1 adipocytes. Oxidative effect on PTP1B （protein tyrosine phosphatase 1b） was carried out with real-time PTP1B enzymatic assay. The aqueous partition of guava leaf extract possesses a potent inhibitory effect on PTP1B enzymatic activity and this PTP1B inhibition is through a slow oxidative but reversible inactivation on the enzyme. The reversible inactivation would suggest guava leaf extract may augment PTP1B inhibition alongside the endogenous H2O2 which itself is induced by insulin. In addition, our study confirmed the hypoglycemic efficacy being associated with guava leaf and found the most effective molecules reside in the aqueous partition which is also less cytotoxic to Chinese hamster ovary cells when compared to other less polar partitions. The guava leaf extract can modulate insulin activity through a redox regulation on PP1B enzymatic activity. It is speculated that a compound similar to gallocatechin in the aqueous partition can reduce an oxygen molecule to hydrogen peroxide which in turn oxidizes the catalytic residue Cys in PTP1B. Therefore, the guava leaf tea can serve as a functional hypoglycemic drink that is suitable for either healthy or diabetic subjects.

Protein tyrosine phosphatase 1B regulates migration of ARPE-19 cells through EGFR/ERK signaling pathway

AIMTo evaluate whether protein tyrosine phosphatase 1B (PTP1B) contributed to initiate human retinal pigment epithelium cells (A)-19 migration and investigate the signaling pathways involved in this process.METHODSARPE-19 cells were cultured and treated with the siRNA-PTP1B. Expression of PTP1B was confirmed by quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR). AG1478 [a selective inhibitor of epidermal growth factor receptor (EGFR)] and PD98059 (a specific inhibitor of the activation of mitogen-activated protein kinase) were used to help to determine the PTP1B signaling mechanism. Western blot analysis verified expression of EGFR and extracellular signal-regulated kinase (ERK) in ARPE-19 cells. The effect of siRNA-PTP1B on cell differentiation was confirmed by immunostaining for α-smooth muscle actin (α-SMA) and qRT-PCR. Cell migration ability was analyzed by transwell chamber assay.RESULTSThe mRNA levels of PTP1B were reduced by siRNA-PTP1B as determined by qRT-PCR assay. SiRNA-PTP1B activated EGFR and ERK phosphorylation. α-SMA staining and qRT-PCR assay demonstrated that siRNA-PTP1B induced retinal pigment epithelium (RPE) cells to differentiate toward better contractility and motility. Transwell chamber assay proved that PTP1B inhibition improved migration activity of RPE cells. Treatment with AG1478 and PD98059 abolished siRNA-PTP1B-induced activation of EGFR and ERK, α-SMA expression and cell migration.CONCLUSIONPTP1B inhibition promoted myofibroblast differentiation and migration of ARPE-19 cells, and EGFR/ERK signaling pathway played important role in migration process.

Structural Insight into the Design on Oleanolic Acid Derivatives as Potent Protein Tyrosine Phosphatase 1B Inhibitors

Oleanolic acid derivatives act as newer protein tyrosine phosphatase 1B （PTP-1B） inhibitors for type 2 diabetes mellitus （T2DM）. In order to understand the structural requirement of PTP-1B inhibitors, 52 oleanolic acid derivatives were divided into a training set （34 compounds） and a test set （18 compounds）. The highly reliable and predictive 3D-QSAR models were constructed by CoMFA, CoMSIA and topomer CoMFA methods, respectively. The results showed that the cross validated coefficient （q2） and non-cross-validated coefficient （R2） were 0.554 and 0.999 in the CoMFA model, 0.675 and 0.971 in the CoMSIA model, and 0.628 and 0.939 in the topomer CoMFA model, which suggests that three models are robust and have good exterior predictive capabilities. Furthermore, ten novel inhibitors with much higher inhibitory potency were designed. Our design strategy was that （i） the electronegative substituents （Cl, -CH2OH, OH and -CH2Cl） were introduced into the double bond of ring C, （ii） the hydrogen bond acceptor groups （C≡N and N atom）, electronegative groups （C≡N, N atom, -COOH and -COOCH3） and bulky substituents （C6H5N） were connected to the C-3 position, which would result in generating potent and selective PTP-1B inhibitors. We expect that the results in this paper have the potential to facilitate the process of design and to develop new potent PTP-1B inhibitors.

Synthesis and protein tyrosine phosphatase 1B inhibition activities of two new synthetic bromophenols and their methoxy derivatives

3-bromo-4,5-bis（2,3-dibromo-4,5-dihydroxybenzyl）-l,2-benzenediol （1） is a natural bromophenol isolated from the red algae Rhodomela confervoides that exhibits significant inhibition against protein tyrosine phosphatase 1B （PTP1B）. Based on its activity, we synthesized two new synthetic bromophenols and their methoxy derivatives from vanillin using the structure of natural bromophenol 1 as a scaffold. The structures of these bromophenols were elucidated from H NMR, 13C NMR, and high resolution electron ionization mass spectrometry as 2,3-dibromo-1-（2＇-bromo-6＇-（3＂,4＂-dimethoxybenzyl）- 3 ＇,4 ＇-dimethoxybenzyl）-4,5 -dimethoxybenzene （2）, 2,3-dibromo- 1 -（2 ＇-bromo-6＇-（2 ＂-bromo-4＂,5 ＂-dimethoxy- benzyl）-3＇,4＇-dimethoxybenzyl）-4,5-dimethoxybenzene （3）, 3,4-dibromo-5-（2＇-bromo-6＇-（2＂-bromo-4＂,5＂- dihydroxybenzyl）-3＇,4＇-dihydroxybenzyl）pyrocatechol （4） and 3,4-dibromo-5-（2＇-bromo-6＇-（3＂,4＂- dihydroxybenzyl）-3＇,4＇-dihydroxybenzyl）pyrocatechol （5）. PTP1B inhibition activities of these compounds were evaluated using a colorimetric assay, and compounds 3 and 4 demonstrated interesting activity against PTP1B.

A novel protein tyrosine phosphatase 1B inhibitor with therapeutic potential for insulin resistance

Insulin sensitizing medicines are currently limited, and identification of new drug candidate is a chal- lenge. Protein tyrosine phosphatase 1B （PTP1 B） negatively regulates insulin signaling pathway, and its inhibition is anticipated to improve insulin resistance. This study investigated the pharmacological profiles of compound CX08005, a new PTP1B inhibitor, with therapeutic potential for insulin resistance in vivo and in vitro, respective- ly. Recombinant human PTP1B protein was used to measure the enzyme activity. The docking simulation was per- formed to explore the interactions between the compound and the protein. The insulin sensitivity was evaluated in Diet-induced obesity mice and/or T2DM KKAy mice by glucose tolerance test （GTT）, the blood glucose level, glucose stimulated insulin secretion （GSIS）, homeostasis model assessment of insulin resistance index （HOMA-IR） and the whole-body insulin sensitivity （ISwb） index, respectively. The hyperinsulinemic-euglycemic clamp was performed to evaluate the insulin stimulated glucose disposal both in whole body and in insulin-sensitive tissues （muscle and fat）. Furthermore, its direct effect in muscle, fat and liver cells was observed. We found that CX08005 was a competitive inhibitor of PTP1B with dose-dependent activity （IC50=5.95 × 10^-7 M）. Docking simulation demonstrated that CX08005 binds to PTP1B at the catalytic P-loop through hydrogen bonds. In DIO mice, treatment with CX08005 effectively ameliorated glucose intolerance in a dose-dependent manner （50- 200 mg. kg^-1 · d^-l）, and decreased HOMA-IR values. We also demonstrated that oral administration of 50 mg ~ kg^-1· d^-1 CX08005 improved hyperglycemia, hyperinsulinemia, HOMA-IR and ISwb in KKAy mice. In hyperin- sulinemic-euglycemic clamp test, CX08005 increased glucose infusion rate and glucose uptake in muscle and fat of DIO mice. In 3T3-L1 adipocytes and C2C12 myotubes, CX08005 enhanced insulin-induced glucose uptake. In HepG2 hepatocyte, CX08005 enhanced insulin-stimulated tyrosine phosphorylation of IRβ/IRS1 in a dose-depend- ent manner, respectively; furthermore, the phosphorylation of several downstream molecules, including Akt, Foxol and GSK3β was also increased, indicating this compound could augment insulin＇s ability to suppress hepatic glu- cose output （HGO）. Our results strongly suggest that compound CX08005 directly enhances insulin action in vitro and in vivo with therapeutic potential for insulin resistance.

POM analysis and computational interactions of 8-hydroxydiospyrin inside active site of protein tyrosine phosphatase 1B

Proteintyrosine phosphatase 1B(PTP1B)inhibitionis consideredas a potentialtherapeuticfor the treatmentof cancer,type2 diabetes,andobesity.Inour presentwork,weinvestigatedtheanti-diabeticpotentialof8-hydroxydiospyrin(8-HDN)from D.lotus against the PTP1B enzyme.It showed significant inhibitory activity of PTP1B with an IC 50 value of 18.37±0.02μM.A detailed molecular docking study was carried out to analyze the binding orientation,binding energy,and mechanism of inhibition.A comparative investigation of 8-HDN in the catalytic,as well as the allosteric site of PTP1B,was performed.Binding energy data showed that compound 8-HDN is more selective for the allosteric site and hence avoids the problems associated with catalytic site inhibition.The inhibition mechanism of 8-HDN can be further investigated as an active lead compound against PTP1B by using in vitro and in vivo models.

Extraction and PTP1B inhibitory activity of bromophenols from the marine red alga Symphyocladia latiuscula

Previously, we had characterized several structurally interesting brominated phenols from the marine red alga Symphyocladia latiuscula collected from various sites. However, Phytochemical investigations on this species collected from the Weihai coastline of Shandong Province remains blank. Therefore, we characterized the chemical constituents of individuals of this species collected from the region. Eight bromophenols were isolated and identified. Using detailed spectroscopic techniques and comparisons with published data, these compounds were identified as 2,3-dibromo-4,5-dihydroxybenzyl methyl ether (1), 3,5-dibromo-4-hydroxybenzoic acid (2), 2,3,6-tribromo-4,5-dihydroxymethylbenzene (3), 2,3,6-tribromo-4,5-dihydroxybenzaldehyde (4), 2,3,6-tribromo-4,5-dihydroxybenzyl methyl ether (5), bis(2,3,6-tribromo-4,5-dihydroxyphenyl)methane (6), 1,2-bis(2,3,6-tribromo-4,5-dihydroxyphenyl)-ethane (7), and 1-(2,3,6-tribromo-4,5-dihydroxybenzyl)-pyrrolidin-2-one (8). Among these compounds, 1 and 2 were isolated for the first time from S. latiuscula. Each compound was evaluated on the ability to inhibit protein tyrosine phosphatase 1B (PTP1B), which is a potential therapeutic target in the treatment of type 2 diabetes. Bromophenols 5, 6, and 7 showed strong activities with IC50 values of 3.9, 4.3, and 3.5 μmol/L, respectively. This study provides further evidence that bromophenols are predominant among the chemical constituents of Symphyocladia, and that some of these compounds may be candidates for the development of anti-diabetes drugs.

Synthesis of mangiferin derivates and study their potent PTP1B inhibitory activity

Protein tyrosine phosphatase 1 B （PTP1 B） has received considerable attention from the drug industry as a potential treatment for diabetes mellitus. Mangiferin has been reported to possess significant antidiabetic activity. Based on the previous study, eight new mangiferin derivates were synthesized and evaluated for their PTP1B inhibitory activity. Some of them displayed good inhibitory activity on PTP1B.

Inhibition of protein tyrosine phosphatase 1B activity by triterpenes isolated from Aceriphyllum rossii

An organic layer prepared from the seed of Aceriphyllum rossii was studied to identify the active compounds for protein tyrosine phosphatase 1B（PTP1B） inhibition.Bioassay guided fractionation resulted in the isolation of PTP1B inhibitory activity of triterpenes（1-4）.These four compounds were identified as aceriphyllic acid C（1）,aceriphyllic acid D（2）,aceriphyllic acid E（3） and aceriphyllic acid F（4）.The isolated 1-4 compounds inhibited PTP1B with IC50 values ranged from（2.1±1.5） μmol/L to（11.2±2.5） μmol/L.Kinetic analysis of PTP1B inhibition by aceriphyllic acid C（1） and aceriphyllic acid D（2） suggested that oleanane-type triterpenes inhibited PTP1B activity in a mixed-type manner.

Pachymic acid exerts antitumor activities by modulating the Wnt/β-catenin signaling pathway via targeting PTP1B

Objective:To determine the inhibitory effects of pachymic acid on lung adenocarcinoma(LUAD)cells and elucidate its underlying mechanism.Methods:CCK-8,wound healing,Transwell,Western blot,tube formation,and immunofluorescence assays were carried out to measure the effects of various concentrations of pachymic acid on LUAD cell proliferation,metastasis,angiogenesis as well as autophagy.Subsequently,molecular docking technology was used to detect the potential targeted binding association between pachymic acid and protein tyrosine phosphatase 1B(PTP1B).Moreover,PTP1B was overexpressed in A549 cells to detect the specific mechanisms of pachymic acid.Results:Pachymic acid suppressed LUAD cell viability,metastasis as well as angiogenesis while inducing cell autophagy.It also targeted PTP1B and lowered PTP1B expression.However,PTP1B overexpression reversed the effects of pachymic acid on metastasis,angiogenesis,and autophagy as well as the expression of Wnt3a andβ-catenin in LUAD cells.Conclusions:Pachymic acid inhibits metastasis and angiogenesis,and promotes autophagy in LUAD cells by modulating the Wnt/β-catenin signaling pathway via targeting PTP1B.

Protein tyrosine phosphatase 1B inhibitory activities of ursane-type triterpenes from Chinese raspberry, fruits of Rubus chingii

Protein tyrosine phosphatase 1 B(PTP1 B) has led to an intense interest in developing its inhibitors as anti-diabetes, anti-obesity and anti-cancer agents. The fruits of Rubus chingii(Chinese raspberry) were used as a kind of dietary traditional Chinese medicine. The methanolic extract of R. chingii fruits exhibited significant PTP1 B inhibitory activity. Further bioactivity-guided fractionation resulted in the isolation of three PTP1 B inhibitory ursane-type triterpenes: ursolic acid(1), 2-oxopomolic acid(2), and 2α, 19α-dihydroxy-3-oxo-urs-12-en-28-oic acid(3). Kinetics analyses revealed that 1 was a non-competitive PTP1 B inhibitor, and 2 and 3 were mixed type PTP1 B inhibitors. Compounds 1-3 and structurally related triterpenes(4-8) were further analyzed the structure-activity relationship, and were evaluated the inhibitory selectivity against four homologous protein tyrosine phosphatases(TCPTP, VHR, SHP-1 and SHP-2). Molecular docking simulations were also carried out, and the result indicated that 1, 3-acetoxyurs-12-ene-28-oic acid(5), and pomolic acid-3β-acetate(6) bound at the allosteric site including α3, α6, and α7 helix of PTP1 B.

海藻中溴酚化合物的蛋白酪氨酸磷脂酶1B抑制活性研究

目的:通过对海藻天然产物进行活性筛选,寻找蛋白酪氨酸磷脂酶1B(PTP1B)抑制剂,为2型糖尿病和肥胖症的治疗寻找新的治疗药物。方法:以PTP1B为靶点,采用分子克隆谷胱苷肽巯基转移酶(glutathion Stransferase,GST)融合蛋白的方法,重组表达获得PTP1B,通过高通量筛选模型对海藻中得到的单体化合物进行活性筛选,采用LOGIT法计算LD50。结果:来源于松节藻和小黏膜藻的溴酚类化合物4-二溴-5-(甲氧基甲基)-1,2-二苯酚(1),2-甲基-3-(2,3-二溴-4,5-二羟基)-苯丙醛(2),3-(2,3-二溴-4,5-二羟基苯)-4-溴-5,6-二羟基-1,3-二氢异苯并呋喃(3)表现出显著的PTP1B抑制活性,IC50分别为3.4,4.5,2.8μmol.L-1。结论:首次对溴酚化合物作为PTP1B抑制剂进行研究,海藻中的3个溴酚类化合物表现出显著的PTP1B抑制活性,有开发成新型抗糖尿病海洋药物的潜力。

基于天然产物的蛋白酪氨酸磷酸酶1B抑制剂的虚拟筛选(英文)

蛋白酪氨酸磷酸酶1B(protein tyrosine phosphatase 1B,PTP1B)是治疗Ⅱ型糖尿病的靶点之一,筛选PTP1B抑制剂具有十分重要的意义.本文采用分子对接虚拟筛选方法,构建共含有42 296个小分子的天然产物库,分别与PTP1B靶点蛋白进行分子对接,以原配体的结合能量为阈值,经过三轮筛选选取打分值高于阈值的小分子进行药代动力学参数和毒性参数预测,最终筛选出3个PTP1B抑制剂,对苯醌类化合物7、异香豆素类衍生物10和Clavepictine类似物11.结合方式研究表明,3个候选抑制剂类药性良好,均具有较好的PTP1B抑制活性,其中化合物10和11的PTP1B抑制活性未见报道.对化合物10进行体外抑制活性检测,其IC50为(74.58±1.23)μmol/L,可作为潜在Ⅱ型糖尿病治疗药物.

桑叶多糖SJB的结构分析和蛋白酪氨酸磷酸酯酶PTP1B抑制活性

通过DEAE-纤维素和凝胶过滤柱色谱对桑叶碱提粗多糖进行分级分离,获得均一多糖SJB,进行结构鉴定.采用蛋白酪氨酸磷酸酯酶PTP1B体外模型对SJB进行降血糖活性测定.结果表明:SJB的相对分子质量为5.4×104,由鼠李糖、阿拉伯糖、葡萄糖、半乳糖、半乳糖醛酸组成的酸性杂多糖;主链由1,2-、1,2,4-连接的鼠李糖和1,4-、1,3,4-连接的半乳糖醛酸组成;侧链包括末端、1,5-、1,3,5-连接的阿拉伯糖;末端、1,4-连接的葡萄糖以及末端、1,3-、1,4-、1,6-连接的半乳糖,主要通过鼠李糖的O4位和半乳糖醛酸的O3位与主链相连.该多糖为首次从桑叶中获得的酸性杂多糖.20μg/mL SJB对PTP1B的抑制率为31.7%.

阿勒泰黄芪提取物对蛋白酪氨酸磷酸酯酶1B的抑制作用

本文探讨了阿勒泰黄芪不同提取物对蛋白酪氨酸磷酸酯酶1B(PTP1B)的抑制作用。采用分光光度法测定了提取物中的黄酮和皂苷含量;通过体外酶促动力学方法检测了不同提取物对PTP1B的影响,并确定了抑制类型;并采用氧化酶法检测了阿勒泰黄芪提取物对细胞利用葡萄糖能力的作用。结果表明,阿勒泰黄芪8种提取物(E1～8)中黄酮含量分别为5.09、10.46、3.58、3.23、53.91、21.77、5.76和7.49 mg/mL,其中E1、E2、E6、E7、E8皂苷含量分别为16.53、27.45、21.90、10.21和8.96 mg/mL;各提取物对PTP1B活性均表现出抑制作用,其中E1、E2、E7、E8的IC50分别为34.8、4.7、7.35和7.15μg/mL,E1、E7和E8是竞争性抑制,E2是混合型竞争性抑制。E1、E2、E5、E7和E8较明显的提高了CHO-K1细胞对葡萄糖的利用。提示皂苷可能是阿勒泰黄芪抑制PTP1B活性的主要物质,通过PTP1B途径有效了提高细胞利用葡萄糖的能力。本研究为阿勒泰黄芪开发为防治糖尿病及改善胰岛素抵抗的药物或保健品提供实验依据。

蛋白酪氨酸磷酸酶1B及其小分子抑制剂的研究进展

蛋白酪氨酸磷酸酶1B(PTP-1B)是胰岛素和瘦素信号传导通路的负调节因子。目前,作为糖尿病和肥胖症治疗的新靶点,PTP-1B抑制剂的研究引起了广泛的关注。现从PTP-1B简介、生理功能以及抑制剂的结构性质等方面进行综述。

青钱柳叶化学成分及其PTP1B抑制活性的研究

目的研究青钱柳叶的化学成分及其PTP1B抑制活性。方法利用反向色谱法分离水溶性成分,利用正向色谱法分离脂溶性成分,根据化合物的化学性质与波谱数据(如IR、ESI-MS、~1H-NMR、^(13)C-NMR、HMQC、HMBC等)鉴定化合物结构,PTP1B抑制活性模型测定化学成分的抑制活性。结果得到16个化合物(1~16),分别为山柰酚3-O-β-D-葡萄糖醛酸苷(1),山柰酚-3-O-α-L-鼠李糖苷(2),山柰酚-3-O-β-D-葡萄糖苷(3),槲皮素3-O-α-L-鼠李糖(4),山柰酚(5),苯甲酸(6),没食子酸(7),(+)-2α-落叶松脂素硫酸钠(8),山柰酚-3-O-α-D-半乳糖苷(9),邻苯二甲酸二(2-乙基己基)酯(10),六氢姜黄素(11),5S-5-羟基-1-(4-羟基-3-甲氧基苯基)-7-(4-羟苯基)-3-庚酮(12),(+)-丁香脂素(13),浙贝素(14),鹅掌揪树脂酚A(15),烟酰胺(16)。结论化合物8~16为首次从青钱柳属植物中分得。化合物10具有PTP1B抑制活性,IC50为(4.89±0.37)μmol·L^(-1)。

Inhibition of protein tyrosine phosphatase 1B by triterpenes isolated from Potentilla discolor Bge

Seven oleanene triterpenes were isolated from the roots of Potentilla discolor Bge and their structures were identified as 3-oxoolean-12-en-27-oic acid（1）, gypsogenic acid（2）, 3α-hydroxyolean-12-en-27-oic acid（3）, 3β-hydroxyolean-12-en-27-oic acid（4）, aceriphyllic acid A（5）, aceriphyllic acid A methyl ester（6）, and oleanolic acid（7）. Compounds 1–7 inhibited protein tyrosine phosphatase 1B（PTP1B） activity, with IC50 values ranging from（7.5±0.5） to（22.7±0.5） μmol/L. Among the isolates, compounds 1, 2, 3 and 7 from the Potentilla discolor Bge were found to exhibit selective PTP1 B inhibitory activity.

PTP-1B基因突变的PCR-RFLP与肥胖的相关性

采用多聚酶联反应-限制性片段长度多态性方法,对110例单纯肥胖者、110例肥胖合并2型糖尿病者、120例正常对照者PTP1B基因ISV5+3666、303位编码子进行酶切研究。结果显示PTP1B基因的P303P突变可能与肥胖发生相关,ISV5+3666delT突变与肥胖发生不相关。

黄连中木脂素类化合物的分离鉴定及其对蛋白酪氨酸磷酸酶1B的抑制活性

目的：对黄连中木脂素类化合物进行分离鉴定,并考察其对蛋白酪氨酸磷酸酶1B（PTP1B）的抑制活性。方法：利用高效液相色谱及凝胶柱色谱等技术对黄连95%乙醇提取物的乙酸乙酯部位进行分离,将得到的化合物采用紫外（UV）、红外（IR）、质谱（MS）、核磁共振（NMR）等方法鉴定其化学结构;以RK-682为阳性药物,考察分离的各化合物对PTP1B的抑制活性（阳性药物和分离化合物的浓度均为0、15、30、50、65、80、100、120、140、170、200μmol/L）,并计算其抑制率和半数抑制浓度（IC50）。结果：共分离得到5个木脂素类化合物,分别鉴定为9-acetyl lanicepside B、Lanicepside A、Woorenogenin、（＋）-isolariciresinol和（＋）-lariciresinol gluciside（即化合物1-5）,其中化合物1、2首次从黄连中分离得到。化合物1-5对PTP1B均有一定抑制作用,且与浓度呈正相关,其IC50分别为57、49、71、58、51μmol/L,但其作用均不及RK-682（IC50为32μmol/L）。结论：从黄连中共分离鉴定出了5个木质素类化合物,且均具有一定的PTP1B抑制活性。