AMPK 激活剂: 作用机制和生理活性

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AMPK activators: mechanisms of action and physiological activities

Abstract

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摘要

AMP-activated protein kinase (AMPK) is a central regulator of energy homeostasis, which coordinates metabolic pathways and thus balances nutrient supply with energy demand. Because of the favorable physiological outcomes of AMPK activation on metabolism, AMPK has been considered to be an important therapeutic target for controlling human diseases including metabolic syndrome and cancer. Thus, activators of AMPK may have potential as novel therapeutics for these diseases. In this review, we provide a comprehensive summary of both indirect and direct AMPK activators and their modes of action in relation to the structure of AMPK. We discuss the functional differences among isoform-specific AMPK complexes and their significance regarding the development of novel AMPK activators and the potential for combining different AMPK activators in the treatment of human disease.

AMP活化蛋白激酶是能量平衡的中枢调节因子,协调代谢途径,从而平衡营养供给和能量需求。由于腺苷酸活化蛋白激酶对代谢的良好生理效应,腺苷酸活化蛋白激酶已被认为是控制人类疾病包括代谢症候群和癌症的重要治疗靶点。因此,腺苷酸活化蛋白激酶可能有潜力作为新的治疗这些疾病。在这篇综述中,我们提供了一个关于 AMPK 结构的间接和直接 AMPK 激活剂及其作用方式的综述。我们讨论了 AMPK 复合物之间的功能差异,以及它们对于开发新的 AMPK 激活剂和组合不同的 AMPK 激活剂治疗人类疾病的潜力的意义。Go to: 去:

Introduction

引言

As a cellular energy sensor, AMP-activated protein kinase (AMPK) is activated in response to a variety of conditions that deplete cellular energy levels, such as nutrient starvation (especially glucose), hypoxia and exposure to toxins that inhibit the mitochondrial respiratory chain complex.12 AMPK is a serine/threonine protein kinase complex consisting of a catalytic α-subunit (α1 and α2), a scaffolding β-subunit (β1 and β2) and a regulatory γ-subunit (γ1, γ2 and γ3; Figure 1). Ubiquitous expression of AMPKα1-, β1- and γ1-subunits in many tissues makes the α1β1γ1 complex a reference for AMPK assays to identify AMPK activators. However, given the unique functions and/or subcellular (or tissue)-specific distribution of the distinct AMPK complex,345 referencing screening to the α1β1γ1 complex may present a limited range of the physiology of AMPK. In line with this notion, increasing evidence shows that inactivating mutations and genetic deletion of specific isoforms produce tissue-specific physiological results.678 Mutations in the AMPKγ2 subunit have frequently been observed in human cardiomyopathies, and deletion of the AMPKα2 subunit, but not α1, has been shown to decrease infarct volume in mouse models of stroke.

作为一种细胞能量传感器,AMP活化蛋白激酶蛋白激酶(AMPK)在各种消耗细胞能量的条件下被激活,例如营养缺乏(特别是葡萄糖)、缺氧和暴露于抑制线粒体呼吸链复合物的毒素。1,2 AMPK 是一种丝氨酸/苏氨酸蛋白激酶复合物,由一个催化亚基(1和2)、一个脚手架亚基(1和2)和一个调节亚基(1,2和3; 图1)组成。AMPK 1-、1-和1-亚基在许多组织中无处不在的表达,使得111复合物成为 AMPK 活化因子鉴定的参考。然而,鉴于 AMPK 复合体的独特功能和/或亚细胞(或组织)特异性分布,3,4,5参考111复合体筛选可能提供 AMPK 生理学的有限范围。与这个概念一致,越来越多的证据表明,失活突变和特定亚型的删除产生组织特异性的生理结果。6,7,8 ampk 2亚基的突变在人类心肌病变中经常被观察到,ampk 2亚基的缺失,但不是1,已被证明可以减少中风小鼠模型的梗死体积。

Functional domains of AMP-activated protein kinase (AMPK) subunits. The mammalian α1/α2 and β1/β2 isoforms are very similar, and their characteristic features are shown. AMPKα subunits: KD, kinase domain containing Thr-172 for the activation by upstream kinases; AID, autoinhibitory domain; two α-RIM, regulatory subunit interacting motifs triggering the conformational changes in response to AMP binding to the AMPKγ subunit; α-CTD, C-terminal domain binding to the β-subunit. AMPKβ subunit: CBM, carbohydrate-binding module, in which Ser108 is important for the action of some direct AMPK activators, such as thienopyridone (A-769662) and salicylate; β-CTD, C-terminal domain containing α-subunit-binding site and immediately followed by the domain for γ-subunit interaction. AMPKγ subunit: three γ-subunit isoforms have variable N-terminal domains (NTDs); four CBS, cystathione-β-synthases domain, which forms two Bateman domains that create four adenosine nucleotide-binding sites (Sites 1–4). Site 2 appears to be always empty and Site 4 to have a tightly bound AMP, whereas Sites 1 and 3 represent the regulatory sites that bind AMP, ADP or ATP in competition.

AMP活化蛋白激酶亚单位的功能域。哺乳动物的1/2和1/2亚型非常相似,并且显示了它们的特征。Ampk 亚基: KD,含有 Thr-172的激酶结构域,用于上游激酶的激活; AID,自抑制结构域; 双 rim,调节亚单位相互作用的模体,触发 AMP 结合 ampk 亚单位的构象变化;-ctd,c- 末端结构域结合 ampk 亚单位。AMPK 亚单位: CBM,碳水化合物结合模块,其中 Ser108对一些直接 AMPK 活化因子如噻吩吡酮(A-769662)和水杨酸盐的作用起重要作用;-ctd,c 末端含有亚单位结合位点的结构域,紧接着是结构域的相互作用。Ampk 亚基: 三个亚基亚基的亚基亚基具有可变的 n 端结构域(NTDs) ,4个 CBS,胱硫蛋白-合成酶结构域,形成两个 Bateman 结构域,产生4个腺苷酸结合位点(位点1-4)。站点2似乎总是空的,站点4有一个紧密结合的 AMP,而站点1和3代表调节站点约束 AMP,ADP 或 ATP 的竞争。

Allosteric activation of AMPK by AMP

AMP 对 AMPK 的变构激活作用

The first class of direct AMPK activators is small molecules that mimic cellular AMP. These molecules trigger a conformational change in the AMPK complex that allows further activation by phosphorylation of Thr-172 in the AMPKα subunit.910 The molecular mechanism underlying allosteric activation of AMPK by AMP binding has been demonstrated by several recent studies of the three-dimensional structure of AMPK.111213 This crystal structure has shown the importance of cystathionine-β-synthase domain repeats within the AMPKγ subunit in the molecular mechanism by which AMPK is activated in response to cellular adenosine nucleotides (AMP, ADP or ATP). Four consecutive cystathionine-β-synthase domains in the AMPKγ subunit provide four potential adenine nucleotide-binding sites. These sites are numbered Sites 1–4, according to the number of the cystathionine-β-synthase domain repeat carrying a conserved aspartate residue involved in ligand binding.111415 In the mammalian AMPKγ1 subunit, Site 2 appears to be always empty and Site 4 to have a tightly bound AMP molecule, whereas Sites 1 and 3 represent the regulatory sites that bind AMP, ADP or ATP, which compete for binding.16 AMP binding to Site 1 appears to cause allosteric activation, whereas binding of AMP or ADP to Site 3 appears to modulate the phosphorylation state of Thr172.13 Although cellular ADP levels are higher than those of AMP, a recent study has shown that AMP is a bona fide activator that enhances LKB1-dependent Thr 172 phosphorylation in vivo.17 AMP binding to the AMPKγ subunit serves as an important regulatory feature of the conformational switch that activates the AMPK complex. The catalytic AMPKα subunit contains an N-terminal kinase domain (KD) immediately followed by an autoinhibitory domain (AID). The three-dimensional structure shows that the AID interacts with the small and large lobes of the KD and causes AMPK to be maintained in an inactive conformation. Once AMP binds to the AMPKγ subunit, the α-RIM (regulatory subunit-interacting motif) between the KD/AID and a globular C-terminal domain of the AMPKα subunit interact with one of the regulatory adenosine nucleotides on the AMPKγ subunit in a manner akin to two arms wrapping around the adenosine. These conformation changes release and expose the KD of AMPKα from its AID to activate the AMPK complex.

第一类直接 AMPK 激活剂是模拟细胞 AMP 的小分子。这些分子触发 AMPK 复合体中的构象改变,使得 AMPK 亚基中 Thr-172的磷酸化进一步激活。9,10 AMPK 结合的变构激活的分子机制已经被最近对 AMPK 三维结构的研究所证实。11,12,13这种晶体结构表明,在 AMPK 的分子机制中,胞硫苷-合成酶结构域重复的重要性。四个连续胱硫醚-合酶结构域在 ampk 亚基提供了四个潜在的腺苷酸结合位点。这些位点编号位点1-4,根据携带保守的天冬氨酸残基参与配体结合的胱硫醚-合酶结构域重复的数量。11,14,15在哺乳动物 ampk 1亚基中,位点2似乎总是空的,位点4有一个紧密结合的 AMP 分子,而位点1和3代表结合 AMP,ADP 或 ATP 的调节位点,竞争结合。虽然细胞 ADP 水平高于 AMP 水平,但最近的研究表明 AMP 是一种善意的激活剂,能够在体内增强 lkb1依赖的 Thr172磷酸化。17 AMP 与 AMPK 亚基的结合是构象开关的重要调控特征,激活 AMPK 复合体。催化 ampk 亚基含有一个 n 端激酶结构域(KD) ,紧接着是一个自体抑制结构域(AID)。三维结构表明,AID 与 KD 的大小叶片相互作用,使 AMPK 维持在非活性构象。一旦 AMP 与 ampk 亚单位结合,KD/AID 与 ampk 亚单位的球状 c 末端结构域之间的-rim (调节亚单位-相互作用模序)就会与 ampk 亚单位上的一个调节腺苷核苷酸相互作用,这种作用类似于腺苷环绕在腺苷周围的两条臂。这些构象的变化释放和暴露其援助的 AMPK 的 KD 激活 AMPK 复合物。

Regulation of AMPK activity by upstream kinases

上游激酶对 AMPK 活性的调控

Physiological AMPK activation involves phosphorylation of Thr-172 within the activation loop of the KD in the AMPKα catalytic subunit. Two upstream kinases, LKB118 and CaMKKβ (Ca2+/calmodulin-dependent protein kinase β),19 have been extensively documented to phosphorylate Thr-172 of the AMPKα subunit. Notably, there are lines of evidence showing that the LKB1-dependent AMPKα phosphorylation at Thr172 is greatly enhanced by the binding of AMP to the AMPK γ-subunit, and, at the same time, the AMP-binding inhibits dephosphorylation of this activating phosphorylation by protein phosphatases, such as PP2A and PP2C in vitro.2021 Interestingly, the effect of AMP on Thr172 phosphorylation of the AMPK α-subunit appears to be dependent on N-terminal myristoylation of the β-subunit, although the underlying mechanism remains to be demonstrated.22 In contrast to the LKB1 complex, another upstream AMPK kinase, CaMKKβ, can activate AMPK in response to increases in cellular Ca2+ without any significant change in ATP/ADP/AMP levels. Treatments that deplete cellular ATP do not effectively activate AMPK in LKB1-negative tumors because the basal activity of CaMKKβ is too low to affect the phosphorylation status of AMPKα Thr172, although the increase in AMP due to ATP depletion makes the AMPK α-subunit a better substrate for CaMKKβ. However, these treatments can cause AMPK activation under conditions that elevate intracellular Ca2+. These data indicate that the phosphorylation/dephosphorylation equilibrium at Thr-172 on the AMPK α-subunit involves AMP binding to the AMPKγ subunit and N-terminal modification of the AMPK β-subunit, adding another a level of complexity to the AMPK activation mechanism.

AMPK 的生理活化涉及到 AMPK 催化亚基 KD 活化环中 Thr-172的磷酸化。两个上游激酶 LKB118和 camkk (Ca2 +/钙调蛋白依赖性蛋白激酶) ,19被广泛地记录为磷酸化 ampk 亚基 Thr-172。值得注意的是,一些证据表明,在 Thr172,依赖 lkb1的 AMPK 磷酸化大大增强了 AMP 结合 AMPK 亚基,同时,AMPK 结合抑制这种蛋白磷酸化的去磷酸化,如 PP2A 和 PP2C 体外。20,21有趣的是,AMP 对 AMPK 亚基 Thr172磷酸化的影响似乎依赖于亚基的 n- 端 myristoylation,尽管潜在的机制仍有待证实。22与 LKB1复合体不同,另一种上游 AMPK,camkk,可以激活 AMPK,对细胞内 Ca2 + 的增加作出反应,而 ATP/ADP/AMP 水平没有明显变化。在 lkb1阴性肿瘤中,耗尽细胞 ATP 的处理不能有效地激活 AMPK,因为 camkk 的基本活性太低,不能影响 AMPK Thr172的磷酸化状态。然而,这些处理可以导致 AMPK 激活条件下提高细胞内钙离子。这些数据表明,AMPK 亚基 Thr-172磷酸化/去磷酸化平衡包括 AMP 与 AMPK 亚基的结合和 AMPK 亚基的 n 端修饰,使 AMPK 激活机制更加复杂。

Physiological functions of AMPK

AMPK 的生理功能

As its name suggests, AMPK has a key role in maintaining the balance between anabolic and catabolic programs for cellular homeostasis in response to metabolic stress.232425262728 Given the functional attributes of AMPK in glucose/lipid homeostasis, body weight, food intake, insulin signaling and mitochondrial biogenesis, AMPK is considered to be a major therapeutic target for the treatment of metabolic diseases including type 2 diabetes and obesity.2930

23,24,25,26,27,28鉴于 AMPK 在葡萄糖/脂质平衡、体重、食物摄入、胰岛素信号和线粒体生物合成等方面的功能特性,AMPK 被认为是治疗包括2型糖尿病和肥胖在内的代谢性疾病的主要治疗靶点

A number of studies have shed light on the role of AMPK in tumorigenesis.31 An initial report connecting AMPK to cancer biology described the discovery of the tumor suppressor LKB1 as a major AMPK upstream kinase.32 Genetic mutations of the LKB1 gene are responsible for inherited Peutz-Jeghers syndrome, which is characterized by the development of hamartomatous polyps in the intestine.33 Since then, a number of in vitro and in vivo studies have suggested that AMPK indeed mediates the tumor-suppressor effects of LKB1. This is supported by findings that drugs that are capable of activating AMPK (metformin, phenformin, A-769662) delay the onset of tumorigenesis in in vivo models.3435 Much effort has been made to understand the molecular mechanisms underlying the antitumorigenic functions of AMPK. These studies have shown that mTORC13637 and RNA polymerase I transcription factor TIF-1A,38 both of which are required for rapidly proliferating cells, are under the control of AMPK. In addition, AMPK activation has been shown to cause G1 cell cycle arrest, which is associated with activation of p53, followed by induction of the cell cycle inhibitor protein, p21.3940 Similarly, AMPK has been shown to cause cell cycle arrest by inducing the phosphorylation and concomitant stabilization of the cyclin-dependent kinase inhibitor p27kip1 in response to metabolic stress.41 A recent study has described an additional layer of p53–AMPK–mTORC1 regulation via the p53-repsonsive gene products Sestrin1/2.42However, it should be noted that AMPK might protect tumor cells against the action of cytotoxic agents, nutrient limitation and hypoxia, once the tumors are established. Therefore, AMPK activators might be deleterious in the treatment of cancer.

一份将 AMPK 与癌症生物学联系起来的初步报告描述了发现肿瘤抑制基因 LKB1作为 AMPK 上游激酶的主要作用. LKB1基因的32个突变是遗传性珀茨-杰格斯综合征的原因,这是肠道错构性息肉发展的拥有属性. 从那时起,一些体外和体内的研究表明,AMPK 确实介导了 LKB1的抑癌作用。这是支持的发现,药物能够激活 AMPK (二甲双胍,苯乙双胍,A-769662)延缓肿瘤发病的体内模型。这些研究表明,mTORC136,37和 RNA聚合酶I 转录因子 TIF-1A,38都是快速增殖细胞所必需的,在 AMPK 的控制下。另外,AMPK 激活已被证明能够引起 g 1细胞周期阻滞,这与 p53激活有关,接着诱导细胞周期抑制蛋白,p21.39,40。同样,AMPK 也被证明能够引起细胞周期阻滞,因为它能够诱导周期蛋白依赖性激酶抑制剂 p27kip1在代谢应激下的磷酸化并随之保持稳定。41最近的一项研究表明,通过 p53排斥基因产物 Sestrin1/2.42,可以增加一层 p53-AMPK-mTORC1的调节。但是,应该注意的是,一旦肿瘤形成,AMPK 可以保护肿瘤细胞免受细胞毒剂、营养限制和缺氧的作用。因此,AMPK 激活剂在肿瘤治疗中可能是有害的。

Another important aspect of AMPK biology is the role of AMPK in autophagy, a lysosome-dependent catabolic program that maintains cellular homeostasis.43444546 A number of studies have demonstrated that AMPK has important roles in autophagy regulation by directly phosphorylating two autophagy-initiating regulators: a protein kinase complex ULK1 (Unc-51-like autophagy-activating kinase)4748 and a lipid kinase complex PI3KC3/VPS34 (phosphatidylinositol 3-kinase, catalytic subunit type 3; also known as VPS34).49 A number of reports have demonstrated the metabolic significance of autophagy in glycogenolysis (glycophagy)50 and lipolysis (lipophagy)51 and even in regulating adipose mass as well as differentiation in vivo.52 In this regard, elucidating the molecular connection between AMPK and autophagy will provide a novel avenue to expand the functional network of AMPK in cellular homeostasis, including metabolism.

AMPK 生物学的另一个重要方面是 AMPK 在自噬中的作用,自噬是一种维持细胞内稳态的溶酶体依赖的分解代谢程序。43,44,45,46大量研究表明,AMPK 通过直接磷酸化两种自噬启动调节因子,在自噬调节中发挥重要作用: 一种蛋白激酶复合物 ULK1(Unc-51-like autohagy-activating kinase)47,48和一种脂质激酶复合物 PI3KC3/VPS34(磷脂酰肌醇激酶3-kinase,催化亚单位3,也称 VPS34)。49大量报道证实了自噬在糖原分解(glycophagy)50和脂肪分解(lipophagy)51中的代谢意义,甚至在调节体内脂肪量和分化中的作用。52. 在这方面,阐明 AMPK 和自噬之间的分子联系将为扩展 AMPK 在细胞内稳态(包括代谢)中的功能网络提供一条新的途径。

Given these functional attributes, as summarized in Figure 2, much effort has been made to develop robust AMPK assays and to identify AMPK modulators to provide therapies for a variety of human diseases.53,545556 In this review, we present a comprehensive summary of both indirect and direct AMPK activators and their modes of action in relation to the structure of AMPK, and discuss the implications of AMPK as a therapeutic target.

鉴于这些功能属性,如图2所概括的,已经作出了很大努力,以开发健壮的 AMPK 检测,并确定 AMPK 调节剂,以提供治疗各种人类疾病。53,54,55,56在这篇综述中,我们提出了间接和直接 AMPK 激活剂及其行动模式与 AMPK 的结构,并讨论了 AMPK 作为治疗目标的影响。

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Figure 2 图2

A summary of the physiological roles of AMP-activated protein kinase (AMPK).

AMP活化蛋白激酶的生理作用概述。Go to: 去:

Indirect AMPK activators

间接 AMPK 激活剂

Practically, AMPK can be activated by any modulator that causes AMP or calcium accumulation. These are classified as indirect activators because a direct interaction between AMPK and modulators is not necessary. Indirect AMPK activators are listed on Table 1.

实际上,任何引起 AMP 或钙积累的调节剂都可以激活 AMPK。这些被归类为间接激活因为 AMPK 和调节剂之间的直接相互作用是不必要的。间接 AMPK 激活剂列于表1。

Table 1

表一

Indirect AMPK activators 间接 AMPK 激活剂

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Biguanides

双胍类

Metformin is a type of biguanide, a synthetic derivative of guanide that is a natural product from the plant Galega officinalis, and has been used as a first-line antidiabetic drug because of its ability to reduce hepatic glucose production and enhance peripheral insulin sensitivity.57 A number of studies have demonstrated that the actions of metformin are attributable to AMPK. Zhou et al. have revealed the molecular mechanisms by which AMPK mediates the antidiabetic actions of metformin: stimulation of fatty-acid oxidation and glucose uptake, and downregulation of lipogenic genes and hepatic glucose production.58AMPK activation by metformin is not a result of direct activation; instead, metformin inhibits complex I of the mitochondrial respiratory chain, leading to an increased AMP:ATP ratio.59 This indirect mechanism has further been supported by the observation that metformin fails to activate AMPK in cells expressing the AMP-insensitive (R531G) AMPKγ2 subunit.60 Recent findings by Fullerton et al. have also shown that phosphorylation of acetyl-CoA carboxylase by AMPK is required for the lipid-lowering effect and the insulin-sensitizing effects of metformin, thereby supporting the role of AMPK in metformin action. However, the role of AMPK has been called into question by recent work showing that metformin lowers blood glucose levels in animal models of liver-specific AMPKα knockout or LKB1 knockout.61 Thus, further studies are required to distinguish the AMPK-dependent and -independent effects of metformin.

二甲双胍是一种双胍,是鸟苷的一种人工合成衍生物,是植物山羊豆的天然产物,由于能够降低肝糖产生和提高外周胰岛素敏感性而被用作一线抗糖尿病药物。已经揭示了 AMPK 介导二甲双胍抗糖尿病作用的分子机制: 刺激脂肪酸氧化和葡萄糖摄取,降低脂肪基因和肝脏葡萄糖产量。58二甲双胍激活 AMPK 不是直接激活的结果,相反,二甲双胍抑制线粒体呼吸链的复合物 i,导致 AMP: ATP 比例增加。59观察到二甲双胍不能激活表达 AMPK 不敏感亚单位(R531G) AMPK 2的细胞中的 AMPK,进一步证实了这一间接机制。60 Fullerton 等人最近的发现也表明,AMPK 的乙酰辅酶A羧化酶磷酸化作用对于二甲双胍的降脂作用和胰岛素增敏作用是必需的,从而支持 AMPK 在二甲双胍作用中的作用。然而,AMPK 的作用已经受到质疑,因为最近的研究表明,在肝脏特异性 AMPK 基因敲除或 LKB1基因敲除的动物模型中,二甲双胍降低血糖水平。因此,需要进一步的研究来区分二甲双胍的 AMPK 依赖性和独立性效应。

Thiazolidinedione

噻唑烷二酮类

Thiazolidinediones (TZDs), also known as glitazones, are a class of insulin-sensitizing drugs including troglitazone, pioglitazone and rosiglitazone. TZDs act primarily by activating the nuclear hormone receptor peroxisome proliferator-activated receptors (PPARs), notably PPARγ, for which their affinity is highest. They are also known to exert their antidiabetic effect in part through AMPK activation. TZDs rapidly activate AMPK in a variety of tissues including skeletal muscle,6263 liver and adipose tissue,64 and the activation mechanisms are associated with accumulation of AMP as a result of inhibiting complex I of the mitochondrial respiratory chain.65 In addition, TZD treatment induces the expression and release of adiponectin from adipocytes,63 which in turn activates AMPK in skeletal muscle and the liver, resulting in increased glucose uptake and fatty-acid oxidation, and decreased hepatic glucose production. Thus, AMPK can be activated by TZDs through at least two different mechanisms.

噻唑烷二酮类药物又称格列酮,是一类胰岛素增敏药物,包括曲格列酮、吡格列酮和罗格列酮。TZDs 的主要作用是激活核受体过氧化物酶体增殖物激活受体(PPARs) ,尤其是 ppar,它们对 ppar 的亲和力最高。它们也被认为可以通过活化 AMPK 发挥部分的抗糖尿病作用。TZD 能迅速激活包括骨骼肌、62、63肝脏和脂肪组织在内的多种组织中的 AMPK,其激活机制与线粒体呼吸链复合物 i 的抑制作用有关。因此,AMPK 可以通过至少两种不同的机制被 tzd 激活。

Polyphenols

多酚

In addition to pharmaceutical agents, numerous naturally occurring compounds and phytochemicals have been shown to activate AMPK. Among them are polyphenols, a structural class of natural or synthetic products characterized by the presence of multiples of phenol structure units. Despite the structural variance, numerous polyphenols are capable of activating AMPK, and they exert beneficial effects on type 2 diabetes and metabolic syndrome. These include resveratrol from red grapes,6667 quercetin from many plant units including fruits, vegetables and grains,68 genistein found in a number of plants such as soybeans,69 epigallocatechin gallate from green tea,69 berberine from Coptis chinensis70 and curcumim from Curcuma longa.71 Mechanisms of activation of AMPK by these compounds appear to require the elevation of AMP levels because many of these compounds are known to inhibit mitochondrial ATP production. Resveratrol, quercetin, epigallocatechin-3-gallate and curcumin target and inhibit the mitochondrial F1F0–ATPase/ATP synthase,7273 whereas berberine is associated with the inhibition of respiratory chain complex I.74 The molecular mechanism of AMPK activation by resveratrol, berberine and quercetin has further been supported by the observation that these compounds fail to activate AMPK in cells expressing AMP-insensitive (R531G) AMPKγ2 subunit.60

除了药剂,许多自然存在的化合物和植物化学物质已被证明可以激活 AMPK。其中包括多酚类物质,这是一类天然或人工合成的产物,拥有属性中含有多种酚类结构单元。尽管结构不同,但是大量的多酚类物质能够激活 AMPK,并且对2型糖尿病和代谢症候群有益。这些化合物包括从红葡萄中提取的白藜芦醇、从包括水果、蔬菜和谷物在内的许多植物单元中提取的66,67种槲皮素、在大豆等一些植物中发现的68种染料木素、绿茶中的69种表没食子儿茶素没食子酸酯、从黄连中提取的69种黄连素和从 Curcuma 长叶姜黄素。白藜芦醇、槲皮素、表没食子儿茶素没食子酸酯和姜黄素作用靶点,抑制线粒体 F1F0-atp 酶/atp 合酶,72,73而小檗碱与呼吸链复合物 i. 74白藜芦醇、小檗碱和槲皮素激活 AMPK 的分子机制进一步得到证实,这些化合物不能激活表达 AMPK 不敏感(R531G) AMPK 2.60的细胞中的 AMPK

Ginsenoside

人参皂苷

Panax ginseng has been long known to have favorable effects in type 2 diabetes and metabolic syndrome. Ginsenosides, a class of tetracyclic triterpene glycosides, are the major pharmacological ingredients in ginseng. To date, more than 80 structurally different ginsenosides have been isolated from the plant genus Panax, and a number of ginsenosides, including Rb1, Rb2, Rc, Re, Rg1, Rg2 and Rg3, have been reported to activate AMPK, resulting in an increased glucose uptake, decreased hepatic triglyceride and cholesterol levels, and the inhibition of lipogenesis and hepatic glucose production.75 The mechanisms for AMPK activation by ginsenosides are largely unknown; however, presumably these compounds are likely to activate AMPK via AMP-dependent mechanisms because the ginsenoside, Rb1, has been reported to increase the intracellular AMP:ATP ratio.76

人参长期以来被认为对2型糖尿病和代谢症候群有良好的作用。人参皂苷是一类四环三萜皂苷,是人参的主要药理成分。到目前为止,已从人参属植物中分离出80多种结构不同的人参皂苷,其中包括 Rb1、 Rb2、 Rc、 Re、 Rg1、 Rg2和 Rg3,据报道它们能够激活 AMPK,导致葡萄糖摄取增加,肝脏甘油三酯和胆固醇水平降低,并抑制脂肪生成和肝脏葡萄糖产生

α-Lipoic acid

– 硫辛酸

α-Lipoic acid (ALA), a naturally occurring dithiol compound derived from octanoic acid, has a critical role in mitochondrial bioenergetics reactions by acting as a cofactor for pyruvate dehydrogenase and α-ketoglutarate dehydrogenase. Owing to its powerful antioxidant property, ALA has gained substantial attention for use in managing diabetic complications.77 Recent studies have also demonstrated that ALA exerts beneficial effects on metabolic syndrome, lipotoxic cardiomyopathy and endothelial dysfunction through the activation of AMPK in various tissues.787980 Although the underlying mechanisms for AMPK regulation by ALA are poorly understood, Shen et al. have reported that ALA increases the intracellular calcium level in C2C12 myotubes, suggesting that CaMKK, but not LKB1, is responsible for AMPK activation.81 In the hypothalamus, where AMPK is implicated in the regulation of appetite, ALA suppresses AMPK activity, leading to reduced food intake.82 Further examination is required to understand the molecular mechanism of the regulation of AMPK by ALA.

– 硫辛酸(lipoic acid,ALA)是由辛酸衍生而来的二硫醇化合物,在线粒体生物能量学反应中起着重要作用,它可以作为丙酮酸脱氢酶和-酮戊二酸脱氢酶的辅助因子。由于 ALA 具有强大的抗氧化功能,因此在治疗糖尿病并发症方面受到广泛关注。77. 最近的研究也表明 ALA 通过激活不同组织中的 AMPK 对代谢症候群、脂毒性心肌病和内皮功能障碍有益。78,79,80尽管人们对 ALA 调节 AMPK 的潜在机制知之甚少,沈等人已经报道 ALA 增加了 C2C12肌管的细胞内钙水平,这表明 CaMKK 而不是 LKB1负责 AMPK 的激活。81在 AMPK 参与调节食欲的下丘脑,ALA 抑制 AMPK 活性,导致食物摄入量减少。82需要进一步研究,以了解 ala. 调节 AMPK 的分子机制。

Other AMPK modulators

其他 AMPK 调制器

Although intracellular energy levels are a major determinant of AMPK activity, AMPK is highly sensitive to the cellular level of reactive oxygen species (ROS).83 In many cases, oxidative stress results in intracellular ATP depletion. However, recent studies have revealed that ROS can stimulate AMPK activity even without a decrease in cellular ATP.8485 Oxidative modification of the AMPKα subunit appears to be a major mechanism by which AMPK is activated under conditions of oxidative stress.86 Therefore, any modulators capable of inducing intracellular ROS generation can activate AMPK without an associated decrease in ATP levels. Such a modulator is cryptotanshinone from Salvia miltiorrhiza Bunge, which exerts antidiabetic87 and anticancer effects88 through ROS-dependent AMPK activation. DNA-damaging agents, such as cisplatin89 or metals, including arsenite, vanadate and cobalt,90 activate AMPK through ROS generation.

虽然细胞内能量水平是 AMPK 活性的主要决定因素,但 AMPK 对细胞内活性活性氧类高度敏感。83在许多情况下,氧化应激会导致细胞内 ATP 损耗。84,85 AMPK 亚基的氧化修饰似乎是 AMPK 在氧化应激条件下被激活的主要机制。这样的一个调节剂是来自丹参的隐丹参酮,它通过依赖于 ros 的 AMPK 激活来发挥抗糖尿病和抗癌作用。Dna 破坏剂,如顺铂89或金属,包括亚砷酸盐,钒酸盐和钴,90通过活性氧产生活化 AMPK。Go to: 去:

Direct AMPK activators

直接 AMPK 激活剂

Several AMPK activators directly bind to and activate AMPK without any significant change in cellular ATP, ADP or AMP levels. Instead, these activators induce conformation changes in the AMPK complex, leading to activation, possibly through a direct interaction with a specific subunit of AMPK (Table 2). The identification of A-769662 by Abbott Laboratories in 2006 provided a novel insight into the development of direct AMPK activators by demonstrating that AMPK activation with non-nucleotide ligands is possible. In addition, it opened up the possibility of developing an activator with AMPK heterotrimer specificity. Since then, numerous studies reporting direct AMPK activators have provided meaningful advances regarding isoform-specific modulators.

几个 AMPK 激活剂直接结合和激活 AMPK 没有任何明显的变化,在细胞 ATP,ADP 或 AMP 水平。相反,这些激活剂诱导 AMPK 复合物的构象变化,导致激活,可能通过直接相互作用与一个特定的 AMPK 亚单位(表2)。美国雅培在2006年鉴定了 A-769662,通过证明非核苷酸配体激活 AMPK 是可能的,为直接 AMPK 激活剂的发展提供了一个新的见解。此外,它开辟了开发 AMPK 异三聚体特异性激活剂的可能性。从那时起,大量的研究报告直接 AMPK 激活剂提供了有意义的进展等形式特异性调节剂。

Direct AMPK activators 直接 AMPK 激活剂

5-Aminoimidazole-4-carboxamide riboside

5- 氨基咪唑 -4- 羧酰胺核苷

The first direct AMPK activator, 5-aminoimidazole-4-carboxamide riboside (AICAR), is an adenosine analog taken up into cells by adenosine transporters and phosphorylated by adenosine kinase, thus generating the AMP-mimetic, AICAR monophosphate (ZMP).9192 Similarly to cellular AMP, ZMP binds to site 3 on the AMPKγ subunit. ZMP does not change the ADP:ATP ratio or alter oxygen uptake, which occurs with many AMPK activators through the inhibition of mitochondrial function.11 Although ZMP is a much less potent AMPK activator than AMP in cell-free systems, AICAR directly activates AMPK in most cells because ZMP can accumulate to millimolar concentrations in cells. ZMP is a natural intermediate in the purine nucleotide synthetic pathway and is metabolized by AICAR transformylase, which catalyzes synthesis of the purine nucleotide inosinate.93 Therefore, the effect of AICAR seems to be more apparent in quiescent, primary cells than in rapidly proliferating cells. Consistently with this notion, anticancer agents that inhibit AICAR transformylase, such as methotrexate and Pemetrexed, sensitize tumor cells to the AMPK-activating and growth-inhibitory effects of AICAR.9495 These results indicate that AMPK participates in the chemotherapeutic effects of antifolate drugs to treat cancers. However, it should be noted that, as an AMP analog, AICAR is able to activate many other AMP-dependent enzymes, such as fructose-1,6-bisphosphatase.9697

第一个直接 AMPK 激活剂5- 氨基咪唑 -4- 羧酰胺核苷(AICAR)是一种腺苷类似物,通过腺苷转运体被腺苷激酶磷酸化,从而产生 AMPK 模拟物 AICAR 单磷酸(ZMP) . 91,92与细胞 AMP 相似,ZMP 与 AMPK 亚单位3位点结合。ZMP 通过抑制线粒体功能,不改变 ADP: ATP 比值,也不改变氧摄取,而许多 AMPK 活化因子通过抑制线粒体功能发生这种作用。ZMP 是嘌呤核苷酸合成途径中的天然中间体,通过 AICAR 转化酶代谢,催化嘌呤核苷酸的合成。与这一概念相一致的是,抑制 AICAR 转化酶的抗癌药物,如甲氨蝶呤和培美曲塞,使肿瘤细胞对 AICAR 的 AMPK 激活和生长抑制作用敏感。然而,值得注意的是,作为 AMP 类似物,AICAR 能够激活许多其他依赖 AMP 的酶,如果糖 -1,6- 二磷酸酶

Thienopyridone (A-769662) and benzimidazole (Compound 911) derivatives

噻吩吡啶酮(A-769662)和苯并咪唑(化合物911)衍生物

Abbott Laboratories has developed a thienopyridone compound, A-769662, which causes allosteric activation of purified AMPK in cell-free assays.98 This compound shows many of the metabolic effects that would be expected with AMPK activation in vivo (increase in fat oxidation in normal rats; decreases in body weight, plasma glucose/triglycerides and liver triglycerides in obese mice). Unlike AICAR, A-769662 shows high specificity toward AMPK. A-769662, similar to AMP, allosterically activates the AMPK complex and inhibits dephosphorylation of Thr-172 in the AMPKα subunit.99100 However, A-769662 appears to use a different molecular mechanism to activate AMPK.101 Notably, it allosterically activates AMPK without Thr172 phosphorylation on the AMPKα subunit, which is absolutely required for AMP-dependent AMPK activation. Importantly, it requires phosphorylation of Ser108 on the AMPKβ1 subunit. Moreover, the strong synergic AMPK activation by AMP and A-769662 has been observed both in vitro and in vivo, clearly demonstrating that A-769662 and AMP have different binding sites on the AMPK complex and different mechanisms of activation.102 Another direct AMPK activator, compound 911, has recently been identified. 911 has been reported to be 5–10-fold more potent than A-769662 in allosterically activating AMPK and preventing dephosphorylation.12 Similarly to A-769662, 991 does not activate AMPK complexes containing the Ser108 mutation of the AMPKβ subunit, suggesting that these two AMPK modulators share a similar molecular mechanism of AMPK activation. Xiao B et al.12 have solved the crystal structure of the full-length human AMPK complex in the presence of A-769662 or 991. In this structure, both A-769662 and 911 are located at a site between the KD of the AMPKα subunit and the carbohydrate-binding module (CBM) of the β-subunit, a site distinct from the adenine nucleotide-binding sites on the AMPKγ subunit. Interestingly, both chemicals exhibit specificity toward AMPK complexes containing the β1 rather than the β2 isoform.

美国雅培已经开发出一种噻吩吡啶酮化合物 A-769662,它能在无细胞实验中引起纯化 AMPK 的变构激活。98这种化合物显示了在体内 AMPK 激活所预期的许多代谢效应(正常大鼠的脂肪氧化增加; 肥胖小鼠的体重、血糖/甘油三酯和肝脏甘油三酯下降)。与 AICAR 不同,A-769662对 AMPK 具有较高的特异性。A-769662与 AMP 相似,在 AMPK 亚基中异构激活 AMPK 复合体并抑制 Thr-172的去磷酸化。然而,A-769662似乎使用了一种不同的分子机制来激活 AMPK。101值得注意的是,它异构激活 AMPK 而没有激活 AMPK 亚基上的 Thr172磷酸化,这是绝对需要的 AMPK 依赖的活化。重要的是,它需要 ampk 1亚基的 Ser108磷酸化。此外,在体内和体外都观察到 AMP 和 A-769662对 AMPK 的强协同活化作用,清楚地表明 A-769662和 AMP 在 AMPK 复合物上有不同的结合位点和不同的激活机制。12与 A-769662相似,991不激活 AMPK Ser108突变的 AMPK 复合物,这表明这两种 AMPK 调节剂具有相似的 AMPK 激活分子机制。肖博等人12已经解决了全长人 AMPK 复合物的晶体结构存在的 A-769662或991。在这个结构中,A-769662和911都位于 ampk 亚基 KD 和-亚基碳水化合物结合模块(CBM)之间的一个位点,这个位点与 ampk 亚基上的腺嘌呤核苷酸结合位点不同。有趣的是,这两种化学物质对包含1而不是2亚型的 AMPK 复合物表现出特异性。

Salicylate (pro-drug of Asprin)

水杨酸盐(阿司匹林前药)

Salicylate is a natural compound traditionally extracted from willow bark. Acetyl salicylate (aspirin) is a derivative that is easier than salicylate to take orally and is rapidly broken down to salicylate upon entering the circulation. Although cyclo-oxygenases (COX1 and COX2) are the established targets for aspirin, it has been reported recently that salicylate (although not aspirin) is a direct activator of AMPK.103 In line with its structural similarity to A-769662, salicylate appears to bind at a site that overlaps with the site targeted by A-769662. Both compounds cause allosteric activation, with salicylate antagonizing the effect of A-769662. In addition, the effects of both compounds are highly dependent on the AMPKβ1 subunit but not on AMPKβ2. Neither compound activates AMPK complexes with the Ser108 mutation of the AMPKβ1 subunit. Considering that thienopyridone (A-769662), benzimidazole (Compound 911) and salicylate derivatives activate AMPK by mechanisms different from most AMP-mimetics or ATP-depleting AMPK activators, the combination of these molecules with the indirect AMPK activators is expected to augment the effect of AMPK on pathophysiological conditions, such as metabolic disorders and cancers.104105106,107

水杨酸盐是一种天然化合物,传统上提取自柳树皮。乙酰水杨酸盐(阿司匹林)是一种比水杨酸盐更容易口服的衍生物,进入循环后迅速分解为水杨酸盐。虽然环氧化酶(COX1和 COX2)是阿司匹林的既定目标,但最近有报道称水杨酸盐(虽然不是阿司匹林)是 AMPK. 103的直接激活剂。与其对 A-769662的结构相似性一致,水杨酸盐似乎在与 A-769662的目标位点重叠的位点结合。两种化合物均能引起变构激活,水杨酸盐能拮抗 A-769662的作用。此外,这两个化合物的影响是高度依赖于 ampk 1亚基,而不是 ampk 2。这两种化合物都不能激活 AMPK 复合物与 AMPK 1的 Ser108突变。考虑到噻吩吡啶酮(A-769662)、苯并咪唑(化合物911)和水杨酸衍生物通过不同于大多数 AMP-mimetics 或 atp-klocollolyampk 激活剂的机制激活 AMPK,这些分子与 AMPK 间接激活剂的结合可望增强 AMPK 对代谢紊乱和癌症等病理生理状态的作用

Compound-13

化合物 -13

Recent screening of a chemical library containing 1,200 AMP mimetics has identified 5-(5-hydroxyl-isoxazol-3-yl)-furan-2-phosphonic acid, termed Compound-2 (C-2), and its pro-drug C-13, as potent allosteric activators of AMPK.108 A subsequent study has demonstrated the molecular mechanism by which C-2 mimics the effects of AMP to stimulate AMPK.109 One concern, as observed with AICAR, is the possibility that C-2 may affect AMP-regulated enzymes other than AMPK (PFK1, FBP1 and glycogen phosphorylase). However, C-2 does not affect any of these enzymes or several enzymes that use AMP as a substrate. In vitro cell-free assays using several AMPK complexes have revealed that C-2 is a potent allosteric activator of AMPK (EC50 of 10–30 nm). In fact, C-2 has been reported to be >20-fold more potent than A769662 and more than two orders of magnitude more potent than AMP.98110 In addition, C-2 and C-13 do not induce any significant change in adenine nucleotide levels. Although the precise C-2-binding sites have not been identified, evidence presented by Hunter et al.109 has suggested that C-2 competes with AMP for binding on the AMPKγ subunit. Surprisingly, the AMPK activators C-2 and C-13 exhibit isoform specificity toward the AMPKα1 subunit. Structural analyses of AMPK complexes1213indicate that different sequences of AMPKα1 and α2 subunits in the α-regulatory subunit-interacting motif-2 (α-RIM2) region, which is used to generate AMPKα isoform-specific antibodies, result in unique interactions of C-2 with one face of AMP bound at Site 3 of the γ-subunit, accounting for the selectivity of C-2 toward AMPKα isoforms. Identification of C-2/C-13 represents an example of the development of a direct and isoform-specific AMPK modulator that is distinct from A-769662 that shows a CBM-dependent AMPKβ subunit specificity.109

最近对一个含有1,200 AMP 的化学文库的筛选鉴定出5-(5- 羟基异噁唑 -3- 基)-呋喃 -2- 磷酸化合物2(C-2)及其前药 C-13为 AMPK 的强变构活化剂。108随后的一项研究证实了 C-2模拟 AMP 刺激 AMPK 的分子机制。然而,C-2不会影响任何这些酶或几种以 AMP 为底物的酶。用几种 AMPK 配合物进行的体外无细胞实验表明,C-2是 AMPK 的强变构激活剂(EC50为10-30nm)。实际上,据报道 C-2的效力是 A769662的20倍以上,比 AMP. 98、110的效力高出2倍以上。此外,C-2和 C-13不会引起腺嘌呤核苷酸水平的显著变化。虽然精确的 C-2结合位点尚未确定,证据提交亨特等人109表明,C-2竞争与 AMP 的结合 ampk 亚单位。令人惊讶的是,AMPK 活化因子 C-2和 C-13对 AMPK 1亚单位表现出异型特异性。AMPK 复合物12,13的结构分析表明,用于产生 AMPK 亚型特异性抗体的调节亚单位相互作用 motif-2(- rim2)区域的 AMPK 1和2个亚基的不同序列,导致了 C-2与 AMPK 亚单位3结合面的特异性相互作用,从而解释了 C-2对 AMPK 亚型的选择性。C-2/C-13的鉴定代表了直接和异形特异性 AMPK 调节剂的发展实例,该调节剂不同于 A-769662,显示了 cbm 依赖的 AMPK 亚单位的特异性

PT-1

Another small molecule activator of AMPK, PT-1, was initially isolated via a screen of compounds that activated the truncated AMPKα1 construct containing only the KD and the AID.111 PT-1 activates the complete AMPK α1β1γ1 as well as the AMPKα1 KD-AID construct but not the AMPKα1 KD construct, suggesting that PT-1 directly binds to the cleft between the KD and the AID, thereby relieving autoinhibition. Consistently with results from a cell-free kinase assay, PT-1 has been shown to increase the phosphorylation of ACC at Ser79, a well-characterized substrate of AMPK, in L6 myotubes without any significant change in cellular AMP:ATP ratio. However, this result has been questioned by a recent report by Jensen et al.112 showing that PT-1 indirectly activates AMPK via inhibition of the mitochondrial respiratory chain complex, thereby increasing cellular AMP:ATP and/or ADP:ATP ratios, instead of binding directly to the AMPKα1 subunit, as previously suggested.111 In line with the notion that PT-1 increases intracellular AMP levels, PT-1 does not activate AMPK in HEK293 cells expressing an AMP-insensitive AMPKγ1 R299G mutant, suggesting that PT-1 functions as an indirect activator. Furthermore, this study has shown that PT-1 selectively activates the AMPK complex containing the γ1-subunit but not γ3 in incubated mouse muscle. The authors have proposed that the failure of PT-1 to activate γ3-containing complexes in muscle is not an intrinsic feature of such complexes but occurs because PT-1 does not increase cellular AMP:ATP ratios in the distinct subcellular compartments containing γ3-complexes. Therefore, the molecular details of PT-1 action should be further studied to address the questions raised by these contradictory results.

另一个 AMPK 的小分子激活剂,PT-1,最初是通过筛选的化合物激活了仅含 KD 和 AID 的截短 AMPK 1结构而分离出来的。111 PT-1激活了完整的 AMPK 111结构和 AMPK 1 KD-AID 结构,但没有激活 AMPK 1 KD 结构,这表明 PT-1直接结合在 KD 和 AID 之间的裂隙上,从而缓解了自我抑制。与无细胞激酶试验结果一致的是,PT-1已被证明能够增加 L6肌管中 AMPK 的底物 Ser79处 ACC 的磷酸化,而细胞内 AMP: ATP 比例没有任何显著变化。然而,Jensen 等人最近的一份报告对这一结果提出了质疑,该报告表明 PT-1通过抑制线粒体呼吸链复合物间接激活 AMPK,从而提高细胞 AMP: ATP 和/或 ADP: ATP 比率,而不是像先前提出的直接结合 AMPK 1亚单位。此外,这项研究表明,PT-1在孵化的小鼠肌肉中选择性地激活包含1- 亚基但不是3的 AMPK 复合体。作者提出,PT-1不能激活肌肉中的3-含有复合物不是这种复合物的固有特征,而是因为 PT-1不能提高含有3-复合物的不同亚细胞区室中的细胞 AMP: ATP 比率。因此,PT-1作用的分子细节应该进一步研究,以解决这些矛盾的结果所提出的问题。

MT 63–78 (Debio0930)

Mt63-78(Debio0930)

Another AMPK direct modulator, MT 63–78 (Debio0930), has recently been identified to allosterically activate AMPK.113 Biochemical analysis has shown that the effect of MT 68–78 is highly selective for the AMPK complex containing the AMPKβ1 subunit, as was seen for A-769662 and salicylate. Notably, MT 63–78 strongly suppresses the growth of prostate cancer cell lines with a concomitant activation of AMPK but without any significant change in cellular ATP, ADP and AMP levels. Importantly, the growth-inhibitory effects of MT 63–78 on prostate cancers are at least 10–40 times higher than those of A-769662. In many prostate cancer models, androgen is believed to drive tumorigenesis and progression of the cancers.114 Therefore, androgen deprivation therapy is a first option to treat this cancer. However, in many cases, the androgen-signaling cascade is re-activated after chemotherapeutic treatments that target the androgen receptor, for example, the androgen receptor antagonist MDV3100.115 Upregulation of de novolipogenesis by androgen in prostate cancer is also closely related to cancer development.116117Considering that AMPK negatively regulates de novo lipogenesis,92108118119 the combination treatment of AMPK activators and androgen receptor inhibitors may function cooperatively as antiprostate cancer drugs. The clinical potential of this concept has been shown in a therapeutic trial. This trial showed that the suppression of de novo lipogenesis is the key mechanism of AMPK inhibition of growth and that MT 63–78 enhances the inhibitory effect of androgen receptor antagonist (MDV3100) on the growth of prostate cancer cells. In addition, the inhibitory effect of MT 63–78 on growth is not limited to prostate cancer cells and has also been observed in LKB1-null A549 cells and in B-RAF-mutated (V600E) KTC-1 cells. These results suggest that MT 63–78 slows the growth of a wide spectrum of cancers, thus increasing the chemotherapeutic effects of current anticancer drugs.

另一种 AMPK 直接调节剂 mt63-78(Debio0930)最近被证实可以异构地激活 AMPK。113生化分析表明,mt68-78对含有 AMPK 1亚单位的 AMPK 复合物具有高度选择性,如同对 A-769662和水杨酸盐的作用一样。值得注意的是,MT 63-78强烈抑制前列腺癌细胞系的生长,同时活化 AMPK,但是在细胞 ATP,ADP 和 AMP 水平没有任何显著变化。重要的是,MT 63-78对前列腺癌的生长抑制作用至少比 A-769662高10-40倍。在许多前列腺癌模型中,雄激素被认为是促使肿瘤发生和进展的因素。因此,雄激素剥夺疗法是治疗这种癌症的第一选择。然而,在许多情况下,雄激素信号级联反应在针对雄激素受体的化疗治疗后被重新激活,例如,雄激素受体拮抗剂 MDV3100.115上调雄激素在前列腺癌中的新生脂肪形成也与癌症的发展密切相关。这一概念的临床潜力已经在治疗试验中显示出来。本实验表明,抑制新生脂质合成是 AMPK 抑制前列腺癌细胞生长的关键机制,MT 63-78增强雄激素受体拮抗剂(MDV3100)对前列腺癌细胞生长的抑制作用。此外,MT 63-78对前列腺癌细胞生长的抑制作用并不仅限于 LKB1-null A549细胞和 b-raf 突变(V600E) KTC-1细胞。这些结果表明,MT 63-78减缓了广泛的癌症的生长,从而增加了当前抗癌药物的化疗效果。Go to: 去:

Perspective

透视法

Most of the current agents that have been shown to activate AMPK in physiological trials, such as metformin, TZDs and 2-deoxyglucose, are indirect activators that inhibit oxidative phosphorylation and glycolysis, thereby increasing the ADP (AMP):ATP ratio. However, it is not always clear whether the effects of these agents are mediated by AMPK. In this sense, much effort has been focused on demonstrating the molecular mechanisms of AMPK activators and on validating the resulting physiologies on many human diseases.2120 Another concern when developing AMPK activators is that there are functional differences between isoform-specific AMPK complexes. For instance, the AMPK α2β2γ3 complex is predominantly activated by exercise in skeletal muscle,5 and therefore specific targeting of the AMPK α2β2γ3. Recent studies reporting direct AMPK activators have provided meaningful advances in developing isoform-specific modulators. For the AMPKα subunit, C-2 (or a pro-drug C-13) has a preference for AMPK complexes containing the AMPKα1 subunit.108 Similarly to A-769662,98 several compounds including 911,12 salicylate (a pro-drug of aspirin)103 and MT 68–78113 specifically activate AMPKβ1-containing complexes but not those containing AMPKβ2. In the case of the AMPKγ subunit, although further studies at the cellular level are required, in vitro biochemical data have shown that PT-1 has a specificity toward AMPK complexes harboring the AMPKγ1 subunit.111 In addition to these activators, a number of pharmaceutical companies have filed patent applications for novel AMPK activators, which are structurally unrelated to AMP. Some representative compounds from each pharmaceutical company are listed in Table 3. Comprehensive lists of AMPK activators in the patent literature are available elsewhere.121122 It is highly intriguing that, although they have been claimed to be novel, the majority of the direct AMPK activators listed in Table 3 show a close resemblance to the original thienopyridone core structure of A-769662, except for the alkene oxindole derivative reported from F. Hoffmann-La Roche AG. Given the recent reports suggesting the AMPK-independent effects of A-769662,100123 further studies are needed to clarify the molecular basis of the accumulating number of direct AMPK activators, by comparing their activation mechanisms and by analyzing their profiles of selectivity across AMPK complex combinations.

目前在生理实验中已被证明能够激活 AMPK 的大多数药物,如二甲双胍、 TZDs 和2- 脱氧葡萄糖,都是抑制氧化磷酸化和糖酵解的间接激活剂,从而提高 ADP (AMP) : ATP 的比例。然而,它并不总是清楚这些代理人的影响是否调解 AMPK。在这个意义上,许多努力已经集中在证明 AMPK 活化因子的分子机制和在许多人类疾病上验证产生的生理学。2,120当开发 AMPK 活化因子时,另一个关注的问题是,在同形特异性 AMPK 复合物之间有功能上的差异。例如,AMPK 223复合物主要是由骨骼肌中的运动激活的,5因此是 AMPK 223的特定靶点。最近的研究报告直接 AMPK 激活剂提供了有意义的进展,开发异形特异性调节剂。108与 a-769662相似,98个化合物包括911,12水杨酸盐(阿司匹林的前药)103和 MT 68-78113特异性激活 AMPK 1的配合物,但不激活 AMPK 2。在 AMPK 亚单位的情况下,虽然需要在细胞水平进一步的研究,体外生化数据表明,PT-1对包含 AMPK 1亚单位的 AMPK 复合物具有特异性。表3列出了每个制药公司的一些代表性化合物。121,122这是非常有趣的,尽管它们被声称是新颖的,但是表3中列出的大多数直接 AMPK 激活剂与原始的 A-769662噻吩利酮核心结构非常相似,除了从罗氏报道的烯氧吲哚衍生物。鉴于最近的报告表明 A-769662,100,123进一步的研究需要澄清的直接 AMPK 激活剂的积累数量的分子基础,通过比较它们的激活机制和分析他们的选择性跨 AMPK 复合物组合。

One interesting aspect of AMPK activators revealed by preclinical studies is the enhanced therapeutic effects of the combination of different AMPK activators. As a master regulator of lipogenic pathway,25AMPK may be an additional chemotherapeutic target because the upregulation of fatty-acid synthesis is a hallmark of many cancers.124 Evidence has shown that the combination of aspirin (salicylate) and Metformin effectively decreases clonogenic survival of prostate and lung cancer cells.104 Consistently with this finding, the addition of fatty acids and/or cholesterol into the culture medium reverses the suppressive effects of salicylate and metformin on cell survival, indicating that the inhibition of de novo lipogenesis is important.105106 Similarly, direct AMPK activators may open new therapeutic avenues for antichemotherapeutic reagents. In the case of the conventional indirect AMPK activators, the mechanism of action requires the upstream kinase LKB1 for physiological AMPK activation. Therefore, the potential of indirect AMPK activators as anticancer drugs is limited to LKB1-deficient tumors, especially for non-small cell lung cancers, of which more than 30% have LKB1-inactivating mutations. In this aspect, direct AMPK activators may overcome this limitation. The evidence shows that the growth-inhibitory response to the AMPK activator, MT 63–78, is not affected by the status of the upstream AMPK-activating kinase LKB1.

临床前研究揭示的 AMPK 激活剂的一个有趣的方面是不同 AMPK 激活剂联合使用的增强治疗效果。124有证据表明,阿司匹林(水杨酸盐)和二甲双胍联合有效地降低前列腺癌和肺癌细胞的克隆生存率。与这一发现一致的是,在培养基中添加脂肪酸和/或胆固醇可逆转水杨酸盐和二甲双胍对细胞存活的抑制作用,这表明抑制新生细胞的生成是重要的。在传统的间接 AMPK 激活剂的情况下,作用机制需要上游激酶 LKB1的生理 AMPK 激活。因此,AMPK 间接活化剂作为抗肿瘤药物的潜力仅限于 lkb1缺陷型肿瘤,尤其是非小细胞肺癌,其中30% 以上具有 lkb1失活突变。在这方面,直接的 AMPK 活化剂可以克服这个限制。有证据表明,AMPK 激活剂 mt63-78的生长抑制反应不受 AMPK 激活激酶 LKB1上游位置的影响。

In conclusion, the recent advances identifying direct AMPK activators make AMPK a ‘druggable’ target for many human diseases, although further studies are required to gain insight into the molecular mechanisms by which AMPK regulates its distinct and diverse downstream targets to produce physiological outcomes.

总之,最近确定 AMPK 直接激活因子的研究进展使 AMPK 成为许多人类疾病的可药物靶点,尽管还需要进一步的研究来深入了解 AMPK 调节其独特和多样的下游靶点以产生生理结果的分子机制。Go to: 去:

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