ASS1 and ASL suppress growth in clear cell renal cell carcinoma via altered nitrogen metabolism

Sanika Khare; Laura C Kim; Graham Lobel; Paschalis-Thomas Doulias; Harry Ischiropoulos; Itzhak Nissim; Brian Keith; M Celeste Simon

doi:10.1186/s40170-021-00271-8

ASS1 and ASL suppress growth in clear cell renal cell carcinoma via altered nitrogen metabolism

Cancer Metab. 2021 Dec 3;9(1):40. doi: 10.1186/s40170-021-00271-8.

Authors

Sanika Khare¹, Laura C Kim¹, Graham Lobel¹, Paschalis-Thomas Doulias², Harry Ischiropoulos², Itzhak Nissim^{3

4}, Brian Keith⁵, M Celeste Simon⁶

Affiliations

¹ Abramson Family Cancer Research Institute, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, 19104, USA.
² Children's Hospital of Philadelphia Research Institute and Departments of Pediatrics and Pharmacology, Perelman School of Medicine at the University of Pennsylvania, Philadelphia, PA, 19104, USA.
³ Division of Genetics and Metabolism, Children's Hospital of Philadelphia, Philadelphia, PA, 19104, USA.
⁴ Department of Pediatrics, Biochemistry, and Biophysics, University of Pennsylvania, Philadelphia, PA, 19104, USA.
⁵ The Wistar Institute, Philadelphia, PA, 19104, USA.
⁶ Abramson Family Cancer Research Institute, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, 19104, USA. celeste2@pennmedicine.upenn.edu.

Abstract

Background: Kidney cancer is a common adult malignancy in the USA. Clear cell renal cell carcinoma (ccRCC), the predominant subtype of kidney cancer, is characterized by widespread metabolic changes. Urea metabolism is one such altered pathway in ccRCC. The aim of this study was to elucidate the contributions of urea cycle enzymes, argininosuccinate synthase 1 (ASS1), and argininosuccinate lyase (ASL) towards ccRCC progression.

Methods: We employed a combination of computational, genetic, and metabolomic tools along with in vivo animal models to establish a tumor-suppressive role for ASS1 and ASL in ccRCC.

Results: We show that the mRNA and protein expression of urea cycle enzymes ASS1 and ASL are reduced in ccRCC tumors when compared to the normal kidney. Furthermore, the loss of ASL in HK-2 cells (immortalized renal epithelial cells) promotes growth in 2D and 3D growth assays, while combined re-expression of ASS1 and ASL in ccRCC cell lines suppresses growth in 2D, 3D, and in vivo xenograft models. We establish that this suppression is dependent on their enzymatic activity. Finally, we demonstrate that conservation of cellular aspartate, regulation of nitric oxide synthesis, and pyrimidine production play pivotal roles in ASS1+ASL-mediated growth suppression in ccRCC.

Conclusions: ccRCC tumors downregulate the components of the urea cycle including the enzymes argininosuccinate synthase 1 (ASS1) and argininosuccinate lyase (ASL). These cytosolic enzymes lie at a critical metabolic hub in the cell and are involved in aspartate catabolism and arginine and nitric oxide biosynthesis. Loss of ASS1 and ASL helps cells redirect aspartate towards pyrimidine synthesis and support enhanced proliferation. Additionally, reduced levels of ASS1 and ASL might help regulate nitric oxide (NO) generation and mitigate its cytotoxic effects. Overall, our work adds to the understanding of urea cycle enzymes in a context-independent of ureagenesis, their role in ccRCC progression, and uncovers novel potential metabolic vulnerabilities in ccRCC.

Keywords: Argininosuccinate lyase; Argininosuccinate synthase 1; Aspartate; DNA synthesis; Nitric oxide metabolism; Urea cycle; ccRCC.

Abstract

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