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Theranostics 2021; 11(9):4011-4029. doi:10.7150/thno.50051 This issue Cite

Research Paper

Adenylosuccinate lyase is oncogenic in colorectal cancer by causing mitochondrial dysfunction and independent activation of NRF2 and mTOR-MYC-axis

Stephanie Taha-Mehlitz^1,2*, Gaia Bianco^1*, Mairene Coto-Llerena^1,3*, Venkatesh Kancherla³, Glenn R. Bantug⁴, John Gallon¹, Caner Ercan³, Federica Panebianco¹, Serenella Eppenberger-Castori³, Marco von Strauss², Sebastian Staubli², Martin Bolli², Ralph Peterli², Matthias S. Matter³, Luigi M. Terracciano^5,6, Markus von Flüe², Charlotte K.Y. Ng⁷, Savas D Soysal², Otto Kollmar², Salvatore Piscuoglio^1,3✉

1. Visceral Surgery and Precision Medicine Research Laboratory, Department of Biomedicine, University of Basel, Basel, Switzerland.
2. Clarunis, Department of Visceral Surgery, University Center for Gastrointestinal and Liver Diseases, St. Clara Hospital and University Hospital Basel, Switzerland.
3. Institute of Medical Genetics and Pathology, University Hospital Basel, Basel, Switzerland.
4. Department of Biomedicine, Immunobiology, University of Basel, Basel, Switzerland.
5. Department of Pathology, Humanitas Clinical and Research Center, IRCCS, Milan, Italy.
6. Humanitas University, Department of Biomedical Sciences, Milan, Italy.
7. Department for BioMedical Research, University of Bern, Bern, Switzerland.
*These authors equally contributed to this work.

✉ Corresponding author: Dr. Salvatore Piscuoglio, Visceral Surgery and Precision Medicine Research Laboratory, Department of Biomedicine, Hebelstrasse 20, 4031 Basel, Switzerland. Tel: +41613286874; Fax: +41612653194. E-mail: s.piscuoglioch.

Abstract

Rationale: Adenylosuccinate lyase (ADSL) is an essential enzyme for de novo purine biosynthesis. Here we sought to investigate the putative role of ADSL in colorectal carcinoma (CRC) carcinogenesis and response to antimetabolites.

Methods: 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. In vivo 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.

Results: 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 in vivo and sensitized CRCs to 6-MP in vitro, ex vivo (PDOs) and in vivo (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.

Conclusions: 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.

Keywords: colorectal cancer, ADSL, mitochondria, fumarate, mTOR-MYC-axis

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