Kayvan R. Keshari: HP Metabolic Dynamics

Contrary to popular belief, before gene transcription and proteins, metabolism is acutely sensitive to change. In some cases, rapid changes in substrate availability can then become a signal to initiate signaling cascades or gene transcription. Since metabolism is dynamic and the probes we develop require a relevant system, we utilize custom-designed MR-compatible bioreactors to investigate changes in the metabolism of our HP probes and compare them to other multimodality approaches.

Recently we’ve shown that the cell’s ability to transport lactate, a by-product of increased glycolytic metabolism, could inform on renal cell carcinoma aggressiveness(1). This we could visualize in an MR-compatible bioreactor system with HP pyruvate (Figure 2). We can then develop schemes to translate this phenomenon in vivo, for instance by using diffusion(2).

While changes in the metabolism of immortal cells and animal models have been used as the basis for probe and drug discovery, it has been difficult to translate these mechanisms to the clinic. This is possibly due to the deviation between the metabolism of these cells and the actual human condition. For this reason, we’ve begun to explore the use of human tissue slice cultures (TSCs) with our hyperpolarized probes for translation. In some recent work (3), we show that the metabolism of primary human prostate tissue slices are very different from that of immortal cells and show hyperpolarized lactate as a biomarker for prostate cancer (Figure 3). We are in the process of developing new probes for study in TSCs and their translation to the clinic.

Figure 2 Enlarge Image Figure 2 (A) Scheme of 13C-labeled carbon atom transitions used to detect C1-labeled pyruvate metabolism during the hyperpolarized experiment. (B) Fitted pyruvate-to-lactate flux and representative spectra (inset) of 13C pyruvate and lactate in the UMRC6 cells. (C) Schematic illustrating the relationship between flow rates and observed HP pyruvate-to-lactate flux in the bioreactor. At high flow rates, the extracellular lactate is more likely to flow out of the NMR coil’s sensitive volume and not contribute to the MR signal, thereby decreasing the observed pyruvate-to-lactate flux. The dotted square represents NMR sensitive region. О denotes encapsulated microspheres containing cells. █ denotes extracellular lactate. (D) HP pyruvate-to-lactate flux of UOK262 and UMRC6 cells at three different flow rates (N=5 for each). There is a decreasing trend in observed pyruvate-to-lactate flux with increasing flow rate for UOK262 cells, which transport lactate out via MCT4 at significantly higher rates (approximately 30% decrease in flux). All values are reported as mean ± std. err. * denotes significance (p<0.05). Figure 3 Enlarge Image Figure 3 Representative 31P MR spectra of living benign and malignant TSCs (A) with accompanying histology after perfusion in the bioreactor (B). These resonances represent quantitative differences in bioenergetics (E). Hyperpolarized dynamics (C, D) demonstrate the increased production of labeled HP lactate as well as the total area under the curve (AUC, F). These are correlated to changes in LDH activity (G) as well as to expression of LDHA and relevant monocarboxylate transporters (MCT1 and 4, H). All values are reported as mean ± standard error of the mean.