是时候重新书写线粒体和细胞内多药耐药癌症之间的通信故事了。在此，我们描述了线粒体网络融合在多药耐药（MDR）乳腺癌中的程度和细胞优势，并设计了一种新型纳米医学，破坏线粒体网络融合并系统操纵细胞器融合和功能。组合细胞器线粒体内质网疗法（COMET）是一种创新的纳米医学，用于治疗MDR三阴性乳腺癌（TNBC），其安全性优于目前的标准治疗（紫杉醇），疗效相当。我们的研究表明，在MDR TNBC中，增加的线粒体网络有助于凋亡抵抗，并且网络融合由线粒体外膜的线粒体融合蛋白2（MFN2）介导。COMET由三个组成部分组成：装载有抗-MFN2肽、链霉菌素和Bam7的破坏线粒体网络（MiND）纳米颗粒（NPs）。COMET的治疗原理是通过使用MiND NPs降低MDR细胞的凋亡阈值，然后通过链霉菌素使MDR细胞受到内质网介导的未折叠蛋白应答（UPR）的刺激，最后使用Bam7直接诱导线粒体凋亡，Bam7是一种特异性的bcl-2 Bax激活剂。MiND NPs是含有21个氨基酸（2577.98 MW）抗-MFN2肽隔室化在水核心中的PEG化脂质体。我们使用缺氧（0.5%氧气）来制备MDR的MDA-MB-231细胞和BT-549细胞。使用Keyence BZ-X710显微镜获取的60×活细胞图像进行的三维分析来量化线粒体网络，MiND NPs有效地破坏了药物敏感和MDR TNBC细胞中的线粒体网络。剂量反应研究得到的IC50值、组合指数和剂量减少指数表明，MiND NPs降低了药物敏感和MDR TNBC细胞的凋亡阈值，并且COMET是一种协同作用的药物组合。我们使用从牛心脏线粒体中提取的复合物V（ATP合酶）评估MiND NPs对氧化磷酸化作用（OXPHOS）的影响；MiND NPs和抗-MFN2肽溶液都显著降低了线粒体复合物V的活性，并降低了OXPHOS的能力。我们使用基于BacMam病毒载体的荧光生物传感器来量化内质网上的未折叠蛋白应答（UPR），链霉菌素特异性地诱导了药物敏感和MDR TNBC细胞中的UPR。我们使用caspase 3 ELISA证明了COMET的协同三药组合增加了Bam7特异性诱导凋亡的能力。剂量限制性毒性和靶点效应是包括紫杉醇在内的当前化疗方案面临的重大挑战。COMET在人类胚胎肾上皮细胞中的细胞毒性明显低于紫杉醇，并有潜力满足更安全的癌症治疗的临床需求。COMET是一种有前景的早期转化性纳米医学，用于治疗MDR TNBC。操纵细胞内通信和细胞器融合是治疗MDR癌症的一种新方法。本研究的数据重写了线粒体、细胞器融合和细胞内通信的故事，而COMET则是癌症治疗的一个令人兴奋的新篇章，可能改变MDR TNBC的临床结果。版权所有 © 2023 Elsevier B.V. 保留所有权利。

It is time for the story of mitochondria and intracellular communication in multidrug resistant cancer to be rewritten. Herein we characterize the extent and cellular advantages of mitochondrial network fusion in multidrug resistant (MDR) breast cancer and have designed a novel nanomedicine that disrupts mitochondrial network fusion and systematically manipulates organelle fusion and function. Combination Organelle Mitochondrial Endoplasmic reticulum Therapy (COMET) is an innovative translational nanomedicine for treating MDR triple negative breast cancer (TNBC) that has superior safety and equivalent efficacy to the current standard of care (paclitaxel). Our study has demonstrated that the increased mitochondrial networks in MDR TNBC contribute to apoptotic resistance and network fusion is mediated by mitofusin2 (MFN2) on the outer mitochondrial membrane. COMET consists of three components; Mitochondrial Network Disrupting (MiND) nanoparticles (NPs) that are loaded with an anti-MFN2 peptide, tunicamycin, and Bam7. The therapeutic rationale of COMET is to reduce the apoptotic threshold in MDR cells with MiND NPs, followed by inducing the endoplasmic reticulum mediated unfolded protein response (UPR) by stressing MDR cells with tunicamycin, and finally, directly inducing mitochondrial apoptosis with Bam 7 which is a specific bcl-2 Bax activator. MiND NPs are PEGylated liposomes with the 21 amino acid (2577.98 MW) anti-MFN2 peptide compartmentalized in the aqueous core. Hypoxia (0.5% oxygen) was used to create MDR derivatives of MDA-MB-231 cells and BT-549 cells. Mitochondrial networks were quantified using 3D analysis of 60× live cell images acquired with a Keyence BZ-X710 microscope and MiND NPs effectively fragmented mitochondrial networks in drug sensitive and MDR TNBC cells. The IC50 values, combination index, and dose reduction index derived from dose response studies demonstrate that MiND NPs decrease the apoptotic threshold of both drug sensitive and MDR TNBC cells and COMET is a synergistic drug combination. Complex V (ATP synthase) extracted from bovine cardiac mitochondria was used to assess the effect of MiND NPs on OXPHOS; both MiND NPs and anti-MFN2 peptide solution significantly decrease the activity of mitochondrial complex V and decrease the capacity of OXPHOS. A BacMam viral vector based fluorescent biosensor was used to quantify the unfolded protein response (UPR) at the level of endoplasmic reticulum and tunicamycin specifically induces the UPR in drug sensitive and MDR TNBC cells. A caspase 3 ELISA demonstrated that the synergistic triple drug combination of COMET increases the ability of Bam7 to specifically induce apoptosis. Dose limiting toxicity and off target effects are a significant challenge for current chemotherapy regimens including paclitaxel. COMET has significantly lower cytotoxicity than paclitaxel in human embryonic kidney epithelial cells and has the potential to fulfill the clinical need for safer cancer therapeutics. COMET is a promising early stage translational nanomedicine for treating MDR TNBC. Manipulating intracellular communication and organelle fusion is a novel approach to treating MDR cancer. The data from this study has rewritten the story of mitochondria, organelle fusion, and intracellular communication and COMET is an exciting new chapter in cancer therapeutics that could transform the clinical outcome of MDR TNBC.Copyright © 2023 Elsevier B.V. All rights reserved.