L-5-[¹¹C]-glutamine PET of breast cancer: Preclinical studies in mouse models

Glutamine is an important metabolic substrate in many aggressive tumors, with comparable importance to glucose metabolism. Utilizing human breast cancer mouse xenograft models, we studied the kinetics of the PET imaging agent, L-5-[¹¹C]-glutamine ([¹¹C]glutamine or [¹¹C]GLN) a biochemical authentic substrate for glutamine metabolism, to further characterize the metabolism of glutamine and downstream labeled metabolites. Studies were performed with and without inhibition of the enzyme, glutaminase (GLS), the first step in glutamine catabolism that generates glutamate, and key target for therapy directed to glutamine-metabolizing cancers.

The study used xenograft mouse models for two breast cancer cell lines, HCC1806, a highly glutaminolytic triple-negative cell line, and MCF-7, a hormone receptor positive line with only low levels of glutaminolysis. Mice were injected with [¹¹C]glutamine and either underwent metabolite analysis or dynamic PET imaging. The contributions of individual metabolites to the total ¹¹C-activity signal in blood and tumor tissue were measured at 10, 20, and 30 min via HPLC. We measured fractional activity in the form of [¹¹C]glutamine versus labeled metabolites, focusing on L-5-[¹¹C]-glutamate ([¹¹C]glutamate or [¹¹C]GLU), and any activity in the other metabolite small molecules labeled with ¹¹C (¹¹C-other or ¹¹C-OTH). Additionally, the contribution of [¹¹C]CO₂ to total ¹¹C-activity was measured. Together with image-based uptake curves, this generated estimated time activity curves for [¹¹C]glutamine and downstream metabolites in both xenograft models treated with vehicle or GLS inhibitor (CB-839).

We found that, out to 30 min post-injection, the majority of radioactivity in highly glutaminolytic tumors (HCC1806) was in the form of [¹¹C]glutamine and [¹¹C]glutamate, with relatively low amounts of radioactivity in metabolites downstream of glutamate including [¹¹C]CO₂. In HCC1806 tumors, [¹¹C]glutamate was retained in the large cellular glutamate pool leading to a majority fraction of total radioactivity in tumor tissue that is greater than the fraction within the blood, with this tumoral fractional pattern reversing with CB-839. This phenomenon leads to a total tumor time-activity curve that is only marginally different before and after CB-839. The radioactivity patterns of MCF-7 tumors after vehicle treatment were similar HCC1806 tumors after CB-839 treatment.

Our studies on [¹¹C]glutamine in breast cancer models show significant retention of ¹¹C-activity in the form of [¹¹C]glutamate in tumors with high GLS activity that confounds non-invasive inference of GLS activity. This suggests limited utility for [¹¹C]glutamine PET for inferring tumor GLS activity and its specific antagonism by drug inhibitors. Our analysis of labeled metabolites in mouse models does, however, yield insights that include the retention of glutamate generated by GLS-mediated catabolism in a large cellular pool and also provide data that is the basis for a compartmental model of glutamine metabolism that is the subject of a companion paper.