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In the process of tumorigenesis, metabolic pathways are altered in support of proliferation. A frequent adaptation in the development of melanoma is the downregulation of arginionsuccinate synthetase 1 (ASS1), the rate limiting enzyme in the conversion of citrulline and aspartate to the amino acid arginine (Arg). Our data supports previous knowledge that the growth of ASS1 deficient cells is inhibited in the absence of Arg. Consequently, depletion of Arg has been considered as a potential therapy for the treatment of ASS1 deficient cancers.

Looking to further exploit potential vulnerabilities that arise with ASS1 downregulation, we performed a functional genomics screen using CRISPR to identify synthetic lethalities with loss of ASS1 and Arg starvation. A top gene hit on the screen encodes for the mostly endosome localized polyamine (PA) transporter ATP13A3. As a downstream product of Arg catabolism, PAs have essential roles in cellular functions including RNA splicing and nucleic acid stabilization. Therefore, we hypothesized that ASS1 deficient cells grow poorly in Arg depleted conditions due to a lack of PAs and poorer growth will result with loss of ATP13A3.

To test our hypothesis, we grew cells with deficient ASS1 at increasing Arg concentrations with putrescine (Put), a PA, added to RPMI media. Subsequently, we found improved growth with Put at about 20-110 µM Arg. Additionally, no growth improvement was seen in cells with ATP13A3 knocked out, further supporting its function as a PA transporter. Interestingly, the improved growth seen with ornithine, a precursor to PAs and other metabolites (e.g., proline and glutamate), was greater than that seen with Put. This suggests that PA deficiency is not the primary factor that inhibits the growth of ASS1 deficient cells in low Arg. Instead, the metabolic versatility of ornithine presents as a point worthy of future inquiry. Understanding the metabolic pathways essential to the growth of cancers that are deficient in ASS1 could reveal new vulnerabilities used in their treatment. (https://tinyurl.com/2ku63f6x)