Taha-Mehlitz Stephanie, Bianco Gaia, Coto-Llerena Mairene, Kancherla Venkatesh, Bantug Glenn R, Gallon John, Ercan Caner, Panebianco Federica, Eppenberger-Castori Serenella, von Strauss Marco, Staubli Sebastian, Bolli Martin, Peterli Ralph, Matter Matthias S, Terracciano Luigi M, von Flüe Markus, Ng Charlotte K Y, Soysal Savas D, Kollmar Otto, Piscuoglio Salvatore
Visceral Surgery and Precision Medicine Research Laboratory, Department of Biomedicine, University of Basel, Basel, Switzerland.
Clarunis, Department of Visceral Surgery, University Center for Gastrointestinal and Liver Diseases, St. Clara Hospital and University Hospital Basel, Switzerland.
Theranostics. 2021 Feb 15;11(9):4011-4029. doi: 10.7150/thno.50051. eCollection 2021.
Adenylosuccinate lyase (ADSL) is an essential enzyme for purine biosynthesis. Here we sought to investigate the putative role of ADSL in colorectal carcinoma (CRC) carcinogenesis and response to antimetabolites. ADSL expression levels were assessed by immunohistochemistry or retrieved from The Cancer Genome Atlas (TCGA) dataset. The effects of ADSL silencing or overexpression were evaluated on CRC cell proliferation, cell migration and cell-cycle. tumor growth was assessed by the chicken chorioallantoic membrane (CAM). Transfected cell lines or patient-derived organoids (PDO) were treated with 5-fluorouracil (5-FU) and 6-mercaptopurine (6-MP) and drug response was correlated with ADSL expression levels. Metabolomic and transcriptomic profiling were performed to identify dysregulated pathways and ADSL downstream effectors. Mitochondrial respiration and glycolytic capacity were measured using Seahorse; mitochondrial membrane potential and the accumulation of ROS were measured by FACS using MitoTracker Red and MitoSOX staining, respectively. Activation of canonical pathways was assessed by immunohistochemistry and immunoblotting. ADSL expression is significantly increased in CRC tumors compared to non-tumor tissue. ADSL-high CRCs show upregulation of genes involved in DNA synthesis, DNA repair and cell cycle. Accordingly, ADSL overexpression accelerated progression through the cell cycle and significantly increased proliferation and migration in CRC cell lines. Additionally, ADSL expression increased tumor growth and sensitized CRCs to 6-MP , (PDOs) and (CAM model). ADSL exerts its oncogenic function by affecting mitochondrial function via alteration of the TCA cycle and impairment of mitochondrial respiration. The KEAP1-NRF2 and mTORC1-cMyc axis are independently activated upon ADSL overexpression and may favor the survival and proliferation of ROS-accumulating cells, favoring DNA damage and tumorigenesis. Our results suggest that ADSL is a novel oncogene in CRC, modulating mitochondrial function, metabolism and oxidative stress, thus promoting cell cycle progression, proliferation and migration. Our results also suggest that ADSL is a predictive biomarker of response to 6-mercaptopurine in the pre-clinical setting.
腺苷酸琥珀酸裂解酶（ADSL）是嘌呤生物合成所必需的一种酶。在此，我们试图研究ADSL在结直肠癌（CRC）致癌作用及对抗代谢物反应中的假定作用。通过免疫组织化学评估ADSL表达水平，或从癌症基因组图谱（TCGA）数据集中获取相关信息。评估ADSL沉默或过表达对CRC细胞增殖、细胞迁移和细胞周期的影响。通过鸡胚绒毛尿囊膜（CAM）评估肿瘤生长。用5-氟尿嘧啶（5-FU）和6-巯基嘌呤（6-MP）处理转染的细胞系或患者来源的类器官（PDO），并将药物反应与ADSL表达水平相关联。进行代谢组学和转录组学分析以鉴定失调的途径和ADSL下游效应物。使用海马分析仪测量线粒体呼吸和糖酵解能力；分别使用MitoTracker Red和MitoSOX染色通过流式细胞术测量线粒体膜电位和活性氧（ROS）的积累。通过免疫组织化学和免疫印迹评估经典途径的激活。与非肿瘤组织相比，ADSL在CRC肿瘤中的表达显著增加。ADSL高表达的CRC显示出参与DNA合成､DNA修复和细胞周期的基因上调。因此，ADSL过表达加速了细胞周期进程，并显著增加了CRC细胞系中的增殖和迁移。此外，ADSL表达增加了肿瘤生长，并使CRC对6-MP敏感（在PDO和CAM模型中）。ADSL通过改变三羧酸循环和损害线粒体呼吸来影响线粒体功能，从而发挥其致癌功能。在ADSL过表达时，KEAP1-NRF2和mTORC1-cMyc轴被独立激活，这可能有利于ROS积累细胞的存活和增殖，促进DNA损伤和肿瘤发生。我们的结果表明，ADSL是CRC中的一种新型癌基因，调节线粒体功能、代谢和氧化应激，从而促进细胞周期进程、增殖和迁移。我们的结果还表明，在临床前环境中，ADSL是对6-巯基嘌呤反应的预测生物标志物。
