Abstract 2856: Measuring tumor metabolism in cancer spheroids and biopsies

Abstract

Abstract Introductory Sentence Real-time metabolic flux can be measured non-invasively using hyperpolarized magnetic resonance spectroscopy in cancer spheroids as well as tumor biopsies. Experimental Procedures LnCAP cells were resuspended in a 1:1 mixture of Matrigel:alginate (1x105 cells/ml) and grown in a porous hollow fiber (A/G Technology Corporation, NY). [1-13C] pyruvate was polarized in a SpinLab (General Electric, NY) before dissolving with appropriate buffers. NMR studies were performed on a 1 Tesla Magritek Spectrometer (Magritek, CA). HP spectra were acquired (10o degree flip angle with repetition time of 4s for 25 scans in total). Cell tracking used the manual tracking function in ImageJ. Xenograft biopsies were obtained from LnCAP tumors using a 2mm punch. Data Summary LnCAP cells seeded sparsely enable cancer spheroid formation remain viable as assessed by Live/DEAD fluorescent staining. Histological sectioning revealed spheroids of approximately 200 µm in diameter. Direct deposition of these threads into a 5mm NMR tube allows measurement of metabolic flux using hyperpolarized [1-13C] pyruvate resulting in the formation of [1-13C] lactate. To modulate metabolism, we used an allosteric AKT inhibitor, MK2206. This drug resulted in relocalization of AKT from the plasma membrane as well as a reduction in cell motility from 0.72 ± 0.04 to 0.46 ± 0.03 μm/s (n=5, p<0.05). Administration of hyperpolarized pyruvate resulted in approximately 30% of lactate production 24 hr after treatment. Histological staining of spheroids after treatment revealed sustained decrease of AKT phosphorylation after treatment. These measurements are in good accordance with measurements of lactate secretion in the same cells grown under standard conditions, with MK2206 resulting in loss of AKT Ser473 phosphorylation as early as 2 hr post-treatment, followed by a decrease in lactate secretion in the media from 5.16 ± 0.2 to 1.35 ± 0.05 mM/106 cells (n=3, p>0.05) 24 hr after treatment. To ensure that these methods are applicable clinical settings, LnCAP xenograft biopsies (~60mg) were deposited directly into an NMR tube to measure hyperpolarized [1-13C] pyruvate metabolism. Line-width of the pyruvate peak was measured to be 1.72 ± 0.19 Hz (n=3) in the first spectra, demonstrating good homogeneity of the magnetic field and conversion to lactate was observed. Future experiments will focus on measuring the metabolism of patient-derived xenografts and effects of treatment response. Conclusions Metabolic measurements using hyperpolarized pyruvate can be performed on cancer spheroids as well as biopsies, allowing quantification of metabolism to complement on-going clinical trials utilizing hyperpolarized MR imaging. Citation Format: Sui Seng Tee. Measuring tumor metabolism in cancer spheroids and biopsies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2856. doi:10.1158/1538-7445.AM2017-2856