MRI of ASCT2-mediated amino acid uptake in xenograft tumor models

Alanine-serine-cysteine transporter 2 (ASCT2) is a prognostic marker for many cancers. While histological expression is the current clinical metric for ASCT2 evaluation, expression does not scale with activity because post-translational modifications are required for translocation to the plasma membrane. Methods to trace ASCT2 activity ex vivo have not been developed and commercialized. We investigated the utility of imaging alanine as a biomarker for ASCT2 in cancer. Alanine is a major ASCT2 substrate that can be detected in magnetic resonance imaging (MRI) scanners using the chemical exchange saturation transfer (CEST) method. Initially, we demonstrated that its CEST contrast was higher than other major ASCT2 substrates at physiological pH (7.3). We then investigated the sensitivity of alanine-weighted CEST (ALAwCEST) imaging to ASCT2 in vivo using a genetically-engineered patient-derived pancreatic cancer model that we developed. ASCT2 protein expression at the cell surface was 1.6-fold higher in SLC1A5 -overexpressing Pa20c cells than naïve Pa20c cells ( p = 0.0221). Upon bolus injection of alanine (6 mmol/kg i.v.) in tumor-bearing mice, CEST MRI enhancement was higher in SLC1A5 -overexpressing Pa20c tumors than naïve tumors ( p < 0.0001). We next investigated the feasibility of ALAwCEST to distinguish prostate tumors with differing levels of ASCT2 content. Enhancement upon injection of 13 C3-alanine was more pronounced in LNCaP prostate tumors than DU-145 prostate tumors in vivo ( p < 0.0001). Notably, the temporal dynamics of ALAwCEST signal enhancement depended on the volume of the tumor. Mass spectrometric imaging of 13 C-labeled metabolites and CEST imaging of metabolite phantoms revealed these differences in in vivo dynamics could be attributed to differences in alanine uptake and metabolism. Overall, we demonstrated the feasibility of ALAwCEST imaging for reporting ASCT2-mediated amino acid uptake in multiple human xenograft tumor models and its utility for cancer applications.