Relevance of the endoplasmic reticulum-mitochondria axis in cancer diagnosis and therapy
内质网-线粒体轴在癌症诊断和治疗中的相关性

Introduction  介绍

An understanding of the cooperation between organelles is crucial for revealing the mechanisms that modulate cellular functions and homeostasis. Among interorganellar networks, the connection between the endoplasmic reticulum (ER) and mitochondria has been extensively studied owing to its diverse functions and impact on the pathogenesis of multiple diseases. The concept of a functional unit comprising the ER and mitochondria was first proposed in 1950¹. The adjacent membrane sites that physically tether the ER and mitochondria are called mitochondria-associated ER membranes (MAMs); technological advances in microscopy have enabled the elucidation of the physiological features of the tethering structures of the MAMs. The ER and mitochondria are separated by a 6–15 nm gap, and the average surface area percentage of mitochondria covered by MAMs was calculated to be 3–5% in mammalian cells².
了解细胞器之间的合作对于揭示调节细胞功能和稳态的机制至关重要。在细胞器间网络中，内质网（ER）和线粒体之间的联系因其多样化的功能和对多种疾病发病机制的影响而被广泛研究。由 ER 和线粒体组成的功能单位的概念于 1950 年首次提出¹ 。物理连接 ER 和线粒体的相邻膜位点称为线粒体相关 ER 膜 (MAM)；显微镜技术的进步使得能够阐明 MAM 系链结构的生理特征。 ER 和线粒体之间有 6–15 nm 的间隙，在哺乳动物细胞中，MAM 覆盖的线粒体平均表面积百分比经计算为 3–5% ² 。

MAMs represent an etiological and therapeutic target in cardiovascular diseases³, neurodegenerative diseases⁴, metabolic disorders^5,6, and cancers. In this review, we discuss the associations between alterations in MAM proteins and cancers and present recent advances in research on these associations. Additionally, we discuss the contribution of MAM proteins to tumorigenesis and cancer progression as well as their possible applications as diagnostic and therapeutic targets.
MAM 代表心血管疾病³ 、神经退行性疾病⁴ 、代谢紊乱^{5 、 6}和癌症的病因学和治疗靶点。在这篇综述中，我们讨论了 MAM 蛋白的改变与癌症之间的关联，并介绍了这些关联研究的最新进展。此外，我们还讨论了 MAM 蛋白对肿瘤发生和癌症进展的贡献以及它们作为诊断和治疗靶点的可能应用。

Structure and functional role of ER–mitochondria contact sites
ER-线粒体接触位点的结构和功能作用

Interaction between the ER–mitochondrial axis and calcium homeostasis
ER-线粒体轴与钙稳态之间的相互作用

Most ER proteins are involved in regulating Ca²⁺ homeostasis⁴³. For instance, the sigma-1 receptor (Sig-1R), a MAM protein, is enriched in the ER vesicles involved in this process. In the resting state, Sig-1R binds to another chaperone in the ER, GRP78. However, this complex dissociates under ER stress conditions, including Ca²⁺ depletion. The dissociated Sig-1R then binds to IP3R, mediating its stabilization and Ca²⁺ influx⁴⁴. PDZ domain-containing protein 8 (PDZD8) in the ER is another example of a Ca²⁺-regulating protein in MAMs. PDZD8 knockdown impairs ER–mitochondria tethering and further inhibits mitochondrial Ca²⁺ uptake in the MAMs of mammalian neurons⁴⁵. Other proteins modulate calcium homeostasis in MAMs, as shown in Table 1.
大多数 ER 蛋白参与调节 Ca ²⁺稳态⁴³ 。例如，sigma-1 受体 (Sig-1R)（一种 MAM 蛋白）在参与该过程的 ER 囊泡中富集。在静息状态下，Sig-1R 与 ER 中的另一个伴侣 GRP78 结合。然而，该复合物在内质网应激条件下（包括 Ca ²⁺耗尽）会解离。然后解离的 Sig-1R 与 IP3R 结合，介导其稳定和 Ca ²⁺流入⁴⁴ 。 ER 中含有 PDZ 结构域的蛋白 8 (PDZD8) 是 MAM 中 Ca ²⁺调节蛋白的另一个例子。 PDZD8 敲低会损害 ER-线粒体束缚，并进一步抑制哺乳动物神经元 MAM 中线粒体 Ca ^{2+ 的}摄取⁴⁵ 。其他蛋白质调节 MAM 中的钙稳态，如表1所示。

As Ca²⁺ participates in diverse cellular processes, disrupted homeostasis and improper regulation of Ca²⁺ dynamics in MAMs can negatively affect cellular function. The influx of Ca²⁺ into mitochondria is essential for bioenergetics because several intramitochondrial enzymes associated with glycolysis and the tricarboxylic acid cycle are activated in a calcium-dependent manner⁴⁶. The lack of constitutive Ca²⁺ influx through IP3 reduces the enzymatic activity of pyruvate dehydrogenase and thus the production of adenosine triphosphate (ATP), resulting in the activation of autophagy via the AMPK pathway⁴⁷. Translocase of mitochondrial outer membrane 70 (TOM70) also affects constitutive Ca²⁺ shuttling by mediating the linkage between IP3R3 and VDAC, and the depletion of TOM70 results in impaired mitochondrial respiration⁴⁸.
由于 Ca ²⁺参与多种细胞过程，MAM 中体内平衡的破坏和 Ca ²⁺动力学的不当调节会对细胞功能产生负面影响。 Ca ²⁺流入线粒体对于生物能学至关重要，因为几种与糖酵解和三羧酸循环相关的线粒体内酶以钙依赖性方式被激活⁴⁶ 。缺乏通过 IP3 的组成型 Ca ²⁺流入会降低丙酮酸脱氢酶的酶活性，从而降低三磷酸腺苷 (ATP) 的产生，从而导致通过 AMPK 途径激活自噬⁴⁷ 。线粒体外膜转位酶 70 (TOM70) 还通过介导 IP3R3 和 VDAC 之间的连接来影响组成型 Ca ²⁺穿梭，TOM70 的耗竭会导致线粒体呼吸受损⁴⁸ 。

MAM proteins stimulate Ca²⁺-dependent apoptotic pathways. Ca²⁺ overload in the mitochondrial matrix increases mitochondrial permeability by opening mitochondrial permeability transition pores (mPTPs)⁴⁹. One mechanism of permeability transition is Ca²⁺-inducible conformational alteration of F-ATP synthases that bind to and show activity toward mPTPs⁵⁰. The opening of mPTPs disrupts the osmotic balance in mitochondria due to nonselective permeabilization, resulting in an influx of water that induces the release of caspase cofactors⁵¹. Furthermore, alterations in Ca²⁺ levels are closely associated with responses to multiple intracellular stresses, such as ER and oxidative stress.
MAM 蛋白刺激 Ca ²⁺依赖性细胞凋亡途径。线粒体基质中的 Ca ²⁺超载通过打开线粒体通透性转换孔 (mPTP) 来增加线粒体通透性⁴⁹ 。通透性转变的一种机制是 Ca ²⁺诱导的 F-ATP 合酶构象改变，该合酶结合 mPTP 并显示出对 mPTP 的活性⁵⁰ 。由于非选择性透化，mPTP 的开放破坏了线粒体的渗透平衡，导致水流入，诱导 caspase 辅助因子的释放⁵¹ 。此外，Ca ²⁺水平的变化与多种细胞内应激（例如 ER 和氧化应激）的反应密切相关。

ER–mitochondria contacts modulate oxidative stress
ER-线粒体接触调节氧化应激

Oxidative stress results from an imbalance between the production and accumulation of reactive oxygen species (ROS) in cells and is a hallmark of the ability to detoxify or repair reactive products⁵². ROS are produced primarily in mitochondria and play important roles in cell growth, differentiation, and death^53,54. Although low levels of ROS play an essential role in intracellular signaling and pathogen defense, elevated levels can have detrimental effects on cells, such as decreasing the efficiency of mitochondrial respiration and inducing oncogenic stress⁵⁵. Imbalances in ROS accumulation can contribute to the development and progression of several diseases, including cancer, metabolic disorders, diabetes, and cardiovascular diseases^56,57.
氧化应激是由细胞中活性氧 (ROS) 的产生和积累之间的不平衡引起的，是解毒或修复反应产物的能力的标志⁵² 。 ROS 主要在线粒体中产生，在细胞生长、分化和死亡中发挥重要作用^{53 , 54} 。尽管低水平的 ROS 在细胞内信号传导和病原体防御中发挥着重要作用，但水平升高会对细胞产生有害影响，例如降低线粒体呼吸效率并诱导致癌应激⁵⁵ 。 ROS 积累的不平衡可能会导致多种疾病的发生和进展，包括癌症、代谢紊乱、糖尿病和心血管疾病^{56 , 57} 。

The ER and mitochondrial axes play essential roles in the detection of and response to stress conditions, including oxidative stress, and form interconnected networks⁵⁸. Furthermore, the simultaneous induction of ER stress and overproduction of ROS in several diseases highlights the importance of this axis⁵⁹. The roles of ROS-related MAM proteins, including endoplasmic reticulum oxidoreductase 1 (ERO1), Sig-1R, p66Shc, and MFN2, have been reported (Fig. 2).
ER 和线粒体轴在应激条件（包括氧化应激）的检测和响应中发挥重要作用，并形成互连网络⁵⁸ 。此外，在几种疾病中同时诱导 ER 应激和 ROS 过量产生凸显了该轴的重要性⁵⁹ 。 ROS 相关的 MAM 蛋白，包括内质网氧化还原酶 1 (ERO1)、Sig-1R、p66Shc 和 MFN2 的作用已有报道（图2 ）。

ERO1 is located entirely on the MAMs close to the ER surface and is an essential factor in the ER oxidative folding mechanism through co-localization with protein disulfide isomerase (PDI)⁶⁰. PDI catalyzes the formation of disulfide bonds in unfolded proteins during oxidative protein folding and is then converted to a reduced form⁶¹. Reduced PDI is subsequently oxidized by ERO1 to participate in the catalytic reaction cycle, where reduced ERO1 transfers electrons to an oxygen molecule via flavin adenine dinucleotide, releasing H₂O₂⁶⁰. ERO1α, an ERO1 isoform, is overexpressed in various cancers, and its expression is increased by chronic ER stress, resulting in excessive H₂O₂ production and an increased ROS burden⁶². ERO1 also affects ROS production by regulating other MAM proteins. Under stress conditions, ERO1 oxidizes IP3R1 and induces detachment of the disulfide isomerase–like protein ERp44 from IP3R1⁶³. ERp44 has an inhibitory effect on IP3R1⁶⁴, leading to massive influx of Ca²⁺ through IP3R, which ultimately results in upregulated mitochondrial metabolism and excessive ROS production^65,66.
ERO1 完全位于靠近 ER 表面的 MAM 上，通过与蛋白质二硫键异构酶 (PDI) 共定位，成为 ER 氧化折叠机制的重要因素⁶⁰ 。 PDI 在蛋白质氧化折叠过程中催化未折叠蛋白质中二硫键的形成，然后转化为还原形式⁶¹ 。还原型PDI随后被ERO1氧化，参与催化反应循环，其中还原型ERO1通过黄素腺嘌呤二核苷酸将电子转移到氧分子，释放H ₂ O ₂ ⁶⁰ 。 ERO1α 是一种 ERO1 亚型，在多种癌症中过度表达，并且其表达因慢性 ER 应激而增加，导致 H ₂ O ₂产生过多和 ROS 负担增加⁶² 。 ERO1 还通过调节其他 MAM 蛋白来影响 ROS 的产生。在应激条件下，ERO1 氧化 IP3R1 并诱导二硫键异构酶样蛋白 ERp44 从 IP3R1 上分离⁶³ 。 ERp44 对 IP3R1 具有抑制作用⁶⁴ ，导致 Ca ²⁺通过 IP3R 大量流入，最终导致线粒体代谢上调和 ROS 产生过多^{65 , 66} 。

Sig-1R regulates Ca²⁺ homeostasis and is involved in ROS-related signaling pathways. Although the ROS-regulatory mechanisms of Sig-1R are not fully understood, previous studies have shown that Sig-1R knockdown leads to ROS accumulation^67,68. Furthermore, some Sig-1R agonists exhibit antioxidant activity under pathological conditions⁶⁹.
Sig-1R 调节 Ca ²⁺稳态并参与 ROS 相关信号通路。尽管 Sig-1R 的 ROS 调节机制尚不完全清楚，但先前的研究表明 Sig-1R 敲低会导致 ROS 积累^{67 , 68} 。此外，一些 Sig-1R 激动剂在病理条件下表现出抗氧化活性⁶⁹ 。

p66Shc is located in MAMs, mitochondria, and the cytosol and tetramerizes in response to oxidative stress⁷⁰. Under oxidative stress conditions, its Ser36 residue is phosphorylated by p38MAPK, ERK, and JNK1/2, and phosphorylation of other residues, namely, Ser54 and Thr386, occurs to prevent p66Shc degradation by ubiquitination^71–73. Activated p66Shc translocates through MAMs into mitochondria, where it binds to cytochrome c to generate ROS and ultimately induce cell death⁷⁴. The generation of ROS by activated p66Shc is supported by previous studies showing that both p66Shc knockout mice and cells exhibit reduced oxidative stress levels and a decreased incidence of diseases such as atherosclerosis^75,76.
p66Shc 位于 MAM、线粒体和细胞质中，并响应氧化应激而四聚化⁷⁰ 。在氧化应激条件下，其 Ser36 残基被 p38MAPK、ERK 和 JNK1/2 磷酸化，并且其他残基（即 Ser54 和 Thr386）发生磷酸化，以防止 p66Shc 被泛素化降解^{71 – 73} 。激活的 p66Shc 通过 MAM 易位到线粒体中，在线粒体中与细胞色素 c 结合产生 ROS 并最终诱导细胞死亡⁷⁴ 。先前的研究表明，p66Shc 敲除小鼠和细胞都表现出氧化应激水平降低和动脉粥样硬化等疾病发生率降低^{75 、 76} ，这些研究支持了激活的 p66Shc 产生 ROS 。

As previously described, both MFN1 and MFN2 are involved in the promotion of mitochondrial fusion. However, the fusion process, which relies primarily on MFN1 and MFN2, is speculated to have additional distinct functions⁷⁷. The possible effects of MFN2 on ROS generation have been suggested to be due to other functions of MFN2. Munoz et al.⁷⁸ reported the possible inhibitory effects of MFN2 on ROS production. MFN2 directly interacts with an ER stress branch, the pancreatic endoplasmic reticulum kinase (PERK) pathway, and inhibits ER stress pathways and ROS production. Other studies have shown that MFN2 overexpression activates the PERK/activating transcription factor 4 (ATF4) pathway and reduces ROS levels in cardiac fibroblasts⁷⁹. However, a recent study showed that MFN2 facilitates the adaptation of macrophages to mitochondrial respiration and ROS generation in response to inflammatory stimuli⁸⁰. Thus, further research is required to fully understand the different functions of MFN2 in different cell types and under specific stress conditions.
如前所述，MFN1 和 MFN2 都参与促进线粒体融合。然而，主要依赖于 MFN1 和 MFN2 的融合过程被推测具有额外的不同功能⁷⁷ 。 MFN2 对 ROS 生成的可能影响被认为是由于 MFN2 的其他功能。穆尼奥斯等人。 ⁷⁸报告了 MFN2 对 ROS 产生可能的抑制作用。 MFN2 直接与 ER 应激分支、胰腺内质网激酶 (PERK) 通路相互作用，并抑制 ER 应激通路和 ROS 产生。其他研究表明，MFN2 过表达会激活 PERK/激活转录因子 4 (ATF4) 通路并降低心脏成纤维细胞中的 ROS 水平⁷⁹ 。然而，最近的一项研究表明，MFN2 促进巨噬细胞适应线粒体呼吸和 ROS 生成，以响应炎症刺激⁸⁰ 。因此，需要进一步的研究来充分了解 MFN2 在不同细胞类型和特定应激条件下的不同功能。

Interaction between ER stress and the ER–mitochondria axis
内质网应激与内质网-线粒体轴之间的相互作用

Protein folding is the main function of the ER. Various conditions, such as disruption of Ca²⁺ homeostasis, inhibition of degradation of unfolded proteins due to proteasome blockade, and genetic mutations, can cause the accumulation of unfolded proteins⁸¹. Under these stress conditions, the unfolded protein response (UPR) is activated by three ER transmembrane proteins: activating transcription factor 6 (ATF6), inositol-requiring enzyme 1α (IRE1α), and PERK⁸². Under normal conditions, the ER chaperone GRP78/BiP binds to the ER lumen region of these transmembrane proteins and inhibits their activity. However, under stress conditions, GRP78 binds to misfolded proteins and induces the activation of these three transmembrane proteins⁸³.
蛋白质折叠是内质网的主要功能。各种条件，例如Ca ²⁺稳态的破坏、由于蛋白酶体阻断而抑制未折叠蛋白的降解以及基因突变，都可能导致未折叠蛋白的积累⁸¹ 。在这些应激条件下，未折叠蛋白反应 (UPR) 由三种 ER 跨膜蛋白激活：激活转录因子 6 (ATF6)、肌醇需求酶 1α (IRE1α) 和 PERK ⁸² 。正常情况下，内质网伴侣 GRP78/BiP 与这些跨膜蛋白的内质网腔区域结合并抑制其活性。然而，在应激条件下，GRP78 与错误折叠的蛋白质结合并诱导这三种跨膜蛋白的激活⁸³ 。

In the ATF6 pathway of the ER stress response, sensors mediate the UPR, and ATF6 translocates to the Golgi complex after GRP78 is released. ATF6 is first cleaved by site-1 protease, and one half remains at the NH₂-terminus before being cleaved by site-2 protease⁸⁴. Regarding the IRE pathway, GRP78 is normally bound to IRE1α or its homolog, IRE1p, and maintains its inactivation. When GRP78 dissociates from IRE1 in ER-stressed cells, IRE1 is phosphorylated and dimerizes⁸⁵. Finally, activated PERK phosphorylates eIF2α and further increases the translation of selected mRNAs, including ATF4, which then promotes the expression of transcription factors, such as C/EBP homologous protein (CHOP), leading to growth arrest and DNA damage⁸⁶. CHOP overexpression causes apoptosis by translocating B-cell lymphoma 2 (BCL2)-associated X (a proapoptotic protein) to mitochondria and decreasing the expression of BCL2 (an antiapoptotic protein)⁸⁷.
在内质网应激反应的 ATF6 通路中，传感器介导 UPR，GRP78 释放后 ATF6 易位至高尔基复合体。 ATF6 首先被位点 1 蛋白酶切割，一半保留在 NH ₂ -末端，然后被位点 2 蛋白酶切割⁸⁴ 。关于 IRE 通路，GRP78 通常与 IRE1α 或其同源物 IRE1p 结合，并维持其失活状态。当 GRP78 在内质网应激细胞中与 IRE1 解离时，IRE1 被磷酸化并二聚化⁸⁵ 。最后，激活的 PERK 磷酸化 eIF2α 并进一步增加选定 mRNA（包括 ATF4）的翻译，然后促进转录因子（如 C/EBP 同源蛋白 (CHOP)）的表达，导致生长停滞和 DNA 损伤⁸⁶ 。 CHOP 过表达通过将 B 细胞淋巴瘤 2 (BCL2) 相关 X（一种促凋亡蛋白）易位至线粒体并降低 BCL2（一种抗凋亡蛋白）的表达来引起细胞凋亡⁸⁷ 。

The associations between MAM components and ER stress have been widely reported (Table 2), and some UPR-related proteins also function as MAM components. The interaction between PERK and MFN2 is a representative example of the UPR-related MAM pathway. Additionally, some MAM proteins are regulated by ER stress; for instance, Sig-1R is transcriptionally upregulated via the PERK/eIF2α/ATF4 pathway⁸⁸, while another MAM protein, Rab32, is upregulated via the UPR pathway. Rab32 belongs to the Ras-like small GTPase family and is involved in mitochondrial fission via interaction with DRP1⁸⁹. In SH-SY5Y cells, Rab32 expression is elevated upon induction of ER stress (thapsigargin treatment), leading to mitochondrial dysfunction and neuronal death⁹⁰. Furthermore, the ER chaperone GRP78 binds to IP3R1 during the ER stress response, releasing Ca²⁺ for influx into mitochondria and inducing cell death due to mitochondrial dysfunction⁹¹.
MAM 成分与 ER 应激之间的关联已被广泛报道（表2 ），一些 UPR 相关蛋白也起到 MAM 成分的作用。 PERK 和 MFN2 之间的相互作用是 UPR 相关 MAM 途径的代表性例子。此外，一些 MAM 蛋白受 ER 应激调节；例如，Sig-1R 通过 PERK/eIF2α/ATF4 途径转录上调⁸⁸ ，而另一种 MAM 蛋白 Rab32 通过 UPR 途径上调。 Rab32 属于 Ras 样小 GTP 酶家族，通过与 DRP1 相互作用参与线粒体裂变⁸⁹ 。在 SH-SY5Y 细胞中，Rab32 表达在诱导 ER 应激（毒胡萝卜素处理）后升高，导致线粒体功能障碍和神经元死亡⁹⁰ 。此外，ER 伴侣 GRP78 在 ER 应激反应期间与 IP3R1 结合，释放 Ca ²⁺流入线粒体并诱导由于线粒体功能障碍而导致的细胞死亡⁹¹ 。

Further evidence has also revealed that several MAM proteins affect UPR pathways. The ER protein VAPB is an important protein involved in UPR activity, and VABP loss inhibits IRE1/XBP1 activity in response to ER stress⁹². Furthermore, VAPB interacts with ATF6 in response to ER stress, and the terminal domain of ATF6 senses protein accumulation in the ER lumen. VAPB, with no luminal structure, is not directly regulated by ATF6 activation but is indirectly inhibited⁹³. VAPB-induced ER stress has been implicated in inducing mitochondrial dysfunction by releasing Ca²⁺ through interactions with PTPIP51 in the mitochondrial membrane²³.
进一步的证据还表明，几种 MAM 蛋白影响 UPR 途径。 ER 蛋白 VAPB 是参与 UPR 活性的重要蛋白，VABP 损失会抑制 IRE1/XBP1 响应 ER 应激的活性⁹² 。此外，VAPB 与 ATF6 相互作用以响应 ER 应激，并且 ATF6 的末端结构域感知 ER 腔中的蛋白质积累。 VAPB 没有管腔结构，不会直接受 ATF6 激活的调节，而是会被间接抑制⁹³ 。 VAPB 诱导的 ER 应激通过与线粒体膜中的 PTPIP51 相互作用释放 Ca ²⁺来诱导线粒体功能障碍²³ 。

Characteristics and diagnostic role of ER–mitochondria contact sites in cancers
癌症中内质网-线粒体接触位点的特征和诊断作用

Cancer cells require a substantial amount of energy for their rapid proliferation and acquisition of malignant phenotypes and use various methods, such as increases in glucose uptake and glycolytic activity (a phenomenon known as the Warburg effect), lipid synthesis and lipolysis, and modulation of Ca²⁺ signaling, to meet these requirements^94–96. Therefore, MAMs play important roles in cancer cell function and metabolism, as they regulate the aforementioned pathways (Fig. 3).
癌细胞需要大量的能量来快速增殖和获得恶性表型，并使用各种方法，例如增加葡萄糖摄取和糖酵解活性（一种称为瓦尔堡效应的现象）、脂质合成和脂肪分解以及 Ca 的调节²⁺信令，以满足这些要求^{94 – 96} 。因此，MAM在癌细胞功能和代谢中发挥着重要作用，因为它们调节上述途径（图3 ）。

The regulation of Ca²⁺ signaling is crucial in cancers, as it is involved in cancer progression, epithelial-to-mesenchymal transition, invasion, and resistance to apoptosis⁹⁷. Therefore, Ca²⁺-regulating proteins in MAMs play various roles in cancer development (Table 3). The IP3R-GRP75-VDAC-MCU complex, which plays an important role in Ca²⁺ transport, is regulated by oncoproteins such as PTEN, BRCA1, and BCL2⁹⁸. In MAMs, PTEN binds to IP3R and prevents its degradation, thereby promoting Ca²⁺ transport to mitochondria, which is important for apoptosis^98,99. However, in various cancers, PTEN loses functionality and triggers inappropriate Ca²⁺ transport, leading to apoptosis resistance^100,101. BCL2, another oncoprotein in MAMs, interacts with IP3R and VDAC and prevents the translocation of Ca²⁺ from the ER to mitochondria. Furthermore, the interaction between BCL2 and VDAC1 interferes with the export of cytochrome c from mitochondria and thus hinders apoptosis^98,102. Therefore, BCL2 overexpression in cancers results in resistance to apoptosis.
Ca ²⁺信号传导的调节在癌症中至关重要，因为它参与癌症进展、上皮间质转化、侵袭和细胞凋亡抵抗⁹⁷ 。因此，MAM中的Ca ²⁺调节蛋白在癌症发展中发挥着多种作用（表3 ）。 IP3R-GRP75-VDAC-MCU 复合物在 Ca ²⁺转运中发挥重要作用，受 PTEN、BRCA1 和 BCL2 等癌蛋白的调节⁹⁸ 。在 MAM 中，PTEN 与 IP3R 结合并防止其降解，从而促进 Ca ²⁺转运至线粒体，这对于细胞凋亡很重要^{98 , 99} 。然而，在各种癌症中，PTEN 失去功能并触发不适当的 Ca ²⁺转运，导致细胞凋亡抵抗^{100 , 101} 。 BCL2 是 MAM 中的另一种癌蛋白，与 IP3R 和 VDAC 相互作用，并阻止 Ca ²⁺从 ER 易位至线粒体。此外，BCL2 和 VDAC1 之间的相互作用会干扰细胞色素 c 从线粒体的输出，从而阻碍细胞凋亡^{98 , 102} 。因此，BCL2在癌症中的过度表达会导致细胞凋亡抵抗。

BRCA1-associated protein 1 (BAP1), a tumor suppressor protein in MAMs, facilitates Ca²⁺ influx into mitochondria by interacting with IP3R¹⁰³. Abnormalities in the function of BAP1 can induce inappropriate Ca²⁺ influx into mitochondria, which may affect the regulation of apoptosis and lead to carcinogenesis¹⁰⁴. Mutations in BAP1 have been observed in various cancers, including renal cell carcinoma, cutaneous melanoma, and uveal melanoma¹⁰⁴. GRP75 also plays an important role in the regulation of Ca²⁺ homeostasis². Transglutaminase type 2 modulates GRP75 function by binding to GRP75 and increasing Ca²⁺ flux between the ER and mitochondria, which affects cancer growth and metastasis. Upregulation of transglutaminase type 2 is a hallmark of breast cancer^105,106. Additionally, TOM70, a protein that links IP3R3 to VDAC, exhibits notably high expression levels in breast cancer cells, and its potential as a therapeutic target has been duly recognized in previous studies^48,107.
BRCA1 相关蛋白 1 (BAP1) 是 MAM 中的一种肿瘤抑制蛋白，通过与 IP3R 相互作用促进 Ca ²⁺流入线粒体¹⁰³ 。 BAP1功能异常可诱导Ca ²⁺不适当地流入线粒体，从而影响细胞凋亡的调节并导致癌变¹⁰⁴ 。 BAP1 突变已在多种癌症中观察到，包括肾细胞癌、皮肤黑色素瘤和葡萄膜黑色素瘤¹⁰⁴ 。 GRP75 在 Ca ²⁺稳态的调节中也发挥着重要作用² 。 2 型转谷氨酰胺酶通过与 GRP75 结合并增加 ER 和线粒体之间的 Ca ²⁺通量来调节 GRP75 功能，从而影响癌症生长和转移。 2 型转谷氨酰胺酶的上调是乳腺癌的标志^{105 , 106} 。此外，TOM70（一种将 IP3R3 与 VDAC 连接的蛋白质）在乳腺癌细胞中表现出显着高的表达水平，其作为治疗靶点的潜力已在之前的研究中得到充分认识^{48 , 107} 。

In addition to its proapoptotic role in mitochondria, Ca²⁺ is important for energy production, progression, and metastasis in cancer^108,109. Ca²⁺ influx into mitochondria mediated by MCU promotes mitochondrial biogenesis and colon cancer proliferation¹⁰⁸, and impairment of Ca²⁺ uptake by MCU knockdown inhibits the proliferation of embryonal rhabdomyosarcoma¹¹⁰. Other types of cancers with high MCU expression include prostate, ovarian, and breast cancers, indicating the diagnostic utility of MCU expression in cancer¹¹¹. Moreover, PDZD8, another Ca²⁺-regulating protein in MAMs, was found to exhibit increased expression levels in stomach cancer tissue compared with normal tissue and is involved in the proliferation and metastasis of stomach cancer¹¹².
除了其在线粒体中的促凋亡作用之外，Ca ²⁺对于癌症的能量产生、进展和转移也很重要^{108 , 109} 。 MCU 介导的 Ca ²⁺流入线粒体可促进线粒体生物合成和结肠癌增殖¹⁰⁸ ，而 MCU 敲低导致的 Ca ²⁺摄取受损可抑制胚胎横纹肌肉瘤的增殖¹¹⁰ 。其他具有高 MCU 表达的癌症类型包括前列腺癌、卵巢癌和乳腺癌，这表明 MCU 表达在癌症中的诊断效用¹¹¹ 。此外，MAM中的另一种Ca ²⁺调节蛋白PDZD8被发现在胃癌组织中与正常组织相比表现出表达水平升高，并且参与胃癌的增殖和转移¹¹² 。

Although research on the exact role of ROS in cancers is still underway, ROS are known to be involved in cancer progression and metastasis⁹⁸. Several MAM proteins, including p66hsc, are regulated by ROS, and p66hsc and the oncoprotein p53 regulate each other¹¹³. Furthermore, p66hsc can be activated by steroid hormones, and activated p66hsc interacts with cytochrome c to increase ROS production. These alterations, including oxidative stress, have been reported to result in poor prognosis in patients with prostate cancer^72,114–116. These characteristics of p66hsc have also been observed in other cancers, including breast and lung cancers, indicating its potential as a diagnostic and therapeutic target^117–119. Furthermore, ERO1, which controls ROS production through the regulation of MAM proteins, is overexpressed in cholangiocarcinoma and is involved in proliferation and metastasis, leading to poor prognosis in patients¹²⁰. Notably, ERO1 is also overexpressed in various other cancers, including breast cancer, lung cancer, and hepatocellular carcinoma, in which it ultimately results in poor prognosis^121–123.
尽管关于 ROS 在癌症中的确切作用的研究仍在进行中，但已知 ROS 参与癌症进展和转移⁹⁸ 。包括 p66hsc 在内的多种 MAM 蛋白受 ROS 调节，并且 p66hsc 和癌蛋白 p53 相互调节¹¹³ 。此外，p66hsc可以被类固醇激素激活，激活的p66hsc与细胞色素c相互作用以增加ROS的产生。据报道，这些改变（包括氧化应激）会导致前列腺癌患者预后不良^{72 , 114 – 116} 。 p66hsc 的这些特征也在其他癌症（包括乳腺癌和肺癌）中观察到，表明其作为诊断和治疗靶点的潜力^{117 – 119} 。此外，ERO1通过调节MAM蛋白来控制ROS的产生，在胆管癌中过度表达，并参与增殖和转移，导致患者预后不良¹²⁰ 。值得注意的是，ERO1 在多种其他癌症中也过度表达，包括乳腺癌、肺癌和肝细胞癌，最终导致预后不良^{121 – 123} 。

Activated lipid metabolism and the accumulation of lipid droplets are hallmarks of various cancer cells⁹⁵. Elevated lipid levels in cancer cells promote proliferation and serve as energy reserves and messengers in oncogenic pathways^95,124. Furthermore, various enzymes involved in lipid synthesis are upregulated in various cancers, including lung, ovarian, and prostate cancers^95,125,126. Various enzymes involved in lipid synthesis, such as fatty acid CoA ligase, which catalyzes the ligation of triacylglycerols and ceramide, and acyl-coenzyme A:cholesterol acyltransferase-1 (ACAT-1), which catalyzes the synthesis of cholesterol, are mainly located in MAMs^98,127,128. Therefore, alterations in the expression of these enzymes in MAMs are strongly associated with cancer. For example, after passing through mitochondria, ceramide plays an important role as an apoptosis inducer and can inhibit cancer growth and cell death^129,130. Cholesterol metabolism is strongly associated with cancer. ACAT-1 in MAMs converts cholesterol to cholesteryl esters, which accumulate in the lipid droplets of cancer cells^98,131. These accumulated cholesteryl esters have a considerable impact on the proliferation and metastasis of cancer cells¹³². Caveolin-1, located in MAMs, is involved in cholesterol efflux, and its overexpression has been identified in a variety of cancers, such as lung, liver, kidney, and colon cancers¹³³. These expression patterns of caveolin-1 are closely related to cancer progression, metastasis, and drug resistance^134–136. Therefore, various MAM proteins play major roles in cancer and can potentially be used in diagnosis and treatment. The association between ER stress and cancer has been established¹³⁷. Sig-1R, a MAM protein regulated by ER stress, has been reported to be overexpressed in myelogenous leukemia and colon cancer¹³⁸. This increased expression promotes angiogenesis and facilitates cancer cell migration, resulting in poor prognosis in patients. Consequently, Sig-1R is considered a promising therapeutic target¹³⁸. Another MAM protein associated with ER stress, VAP-B, has been reported to play a key role in breast cancer progression, highlighting its potential as a diagnostic marker for this malignancy¹³⁹.
激活的脂质代谢和脂滴的积累是各种癌细胞的标志⁹⁵ 。癌细胞中脂质水平升高会促进增殖并充当致癌途径中的能量储备和信使^{95 , 124} 。此外，参与脂质合成的各种酶在各种癌症中上调，包括肺癌、卵巢癌和前列腺癌^{95 , 125 , 126} 。参与脂质合成的各种酶，如催化三酰甘油和神经酰胺连接的脂肪酸CoA连接酶，以及催化胆固醇合成的酰基辅酶A：胆固醇酰基转移酶-1（ACAT-1），主要位于^{MAM 98、127、128 。}因此，MAM 中这些酶表达的改变与癌症密切相关。例如，通过线粒体后，神经酰胺作为细胞凋亡诱导剂发挥着重要作用，可以抑制癌症生长和细胞死亡^{129 、 130} 。胆固醇代谢与癌症密切相关。 MAM 中的 ACAT-1 将胆固醇转化为胆固醇酯，胆固醇酯积聚在癌细胞的脂滴中^{98 、 131} 。这些积累的胆固醇酯对癌细胞的增殖和转移有相当大的影响¹³² 。 Caveolin-1 位于 MAM 中，参与胆固醇流出，其过度表达已在多种癌症中发现，如肺癌、肝癌、肾癌和结肠癌¹³³ 。 Caveolin-1 的这些表达模式与癌症进展、转移和耐药性密切相关^{134 – 136} 。因此，各种MAM蛋白在癌症中发挥着重要作用，并有可能用于诊断和治疗。 ER 应激与癌症之间的关联已被确定¹³⁷ 。 Sig-1R 是一种受 ER 应激调节的 MAM 蛋白，据报道在骨髓性白血病和结肠癌中过度表达¹³⁸ 。这种表达增加会促进血管生成并促进癌细胞迁移，导致患者预后不良。因此，Sig-1R 被认为是一个有前途的治疗靶点¹³⁸ 。据报道，另一种与 ER 应激相关的 MAM 蛋白 VAP-B 在乳腺癌进展中发挥着关键作用，凸显了其作为这种恶性肿瘤的诊断标志物的潜力¹³⁹ 。

The ER–mitochondrial axis as a therapeutic target
ER-线粒体轴作为治疗靶点