Investigating Prostate Cancer Aggressiveness with Hyperpolarized 13C-Urea + 13C-Pyruvate Perfusion & Metabolic Imaging of Transgenic Primary and Metastatic Tumors
Hsin-Yu Chen1, Peder E.Z. Larson1,2, Robert A. Bok2, Cornelius von Morze2, Renuka Sriram2, Romelyn Delos Santos2, Justin Delos Santos2, Jeremy W. Gordon2, John Kurhanewicz1,2, and Daniel B. Vigneron1,2

1Graduate Program in Bioengineering, UCSF and UC Berkeley, University of California, San Francisco, San Francisco, CA, United States, 2Department of Radiology and Biomedical Imaging, University of California, San Francisco, San Francisco, CA, United States


An unmet clinical need facing the management of prostate cancer is an accurate method for distinguishing aggressive prostate cancer from indolent disease. The project investigated the use of hyperpolarized (HP) 13C 3D CS-EPSI imaging of co-polarized 13C-urea + 13C-pyruvate to provide this distinction by simultaneously measuring metabolism and perfusion. Significantly higher pyruvate-to-lactate conversion rates, kPL, (P<0.00001) and significantly (P<0.004) lower urea perfusion were detected in high-grade tumors compared to low-grade tumor. Lymph-node metastases demonstrated high metabolic conversion (P>0.8) and urea perfusion not significantly (P>0.4) different than high-grade primary tumor.


A pressing need facing the clinical management of prostate cancer patients is an accurate method for distinguishing aggressive, potentially lethal, prostate cancer from indolent disease. Prostate cancer (PCa) is the second most prevalent cancer in American men, with 1 in 6 American men being diagnosed, but is fatal for only 12% of these cases [1]. This hyperpolarized 13C MR study of 19 prostate-cancer mice was designed to measure metabolic and perfusion parameters simultaneously by HP 13C-urea & 13C-pyruvate dynamic MRI in primary and metastatic tumors. Correlations with ex vivo measures of aggressiveness were performed with subsequent histochemical, gene expression and enzymatic activity assays.


Sequences: 3D dynamic compressed-sensing 13C-EPSI provided high spatiotemporal resolution for simultaneous imaging of metabolism and perfusion. The sequence employed variable flip angle and multiband excitation, followed by double spin echo refocusing and a compressed-sensing 3D EPSI readout. (Spatial Resolution: 3.3mm x 3.3mm x 5.4 mm, Temporal Resolution = 2s, FOV = 4cm x 4cm x 8.6cm) Anatomical reference images were acquired using T2-FSE.

MRI experiments: 19 transgenic prostate cancer mice (TRAMP) were studied using a clinical 3T scanner with a dual-tuned mouse coil for 13C and proton imaging. The imaging studies were conducted for each animal prior to or on the day of sacrifice. [1-13C]pyruvate and 13C urea were co-polarized using a prototype clinical polarizer for 2 hours, reaching approximately 20-30% percentage polarization. ~350ul of the 13C-labelled compounds (concentration = 80mM for both pyruvate and urea) were rapidly injected into the mice via tail vein catheter over 15 seconds, and the 13C scan began at the end of injection.

Tissue analysis: Following the HP MR studies the animals were sacrificed and the excised tissue was sectioned and histochemical, gene expression and activity analyses were performed. Histochemical assays included H&E, Ki-67 and PIM immunostaining. Gene expression analysis consists of HIF-1α, LDHA, LDHB, VEGF, MCT1 and MCT4. LDH isoenzyme activity was also measured.


A total of 19 TRAMP mice were studied. 9 were histo-pathologically categorized as low-grade, while 10 were high-grade. Lymph node metastases were found in 5 of the high-grade mice. The mean tumor size in low-grade animal was 0.184±0.082(cm3), whereas that of high-grade was 4.03±3.60(cm3), and of metastasis was 0.11±0.09(cm3). Rate of pyruvate-to-lactate conversion (kPL) was significantly (P<0.00001) higher in high-grade (0.0560±0.0047s-1) and metastatic cancers (0.0593±0.0261s-1) versus low-grade tumor (0.0197±0.0012s-1) as shown in Figures 1-3. No overlap in kPL was observed between low- and high-grade tumors. Perfusion area-under-curve (AUC) measurements significantly reduced in high-grade disease (high-: 640.5±94.1, low-: 1407.4±221.9 A.U., High grade/Low grade=45.5%, P<0.004), while ktrans significantly increased (high-: 358.4±38.5, low-: 180.1±24.1 s-1, P<0.002) as shown in Fig1.D). Fig1.B) shows significantly higher LDH activity in high grade tumors (High grade/Low grade=196.9%). As shown in Fig.2&5, both Ki-67 staining for proliferation and PIM staining for hypoxia increased in high-grade tumors. No significant difference in kPL(P>0.8) or perfusion AUC(P>0.4) was found between high-grade and metastatic tissue (Figures 3-4). ktrans, as a combined measure of perfusion and permeability, indicated a loss of morphology/function and increased leakiness in the vasculature of high-grade cancers.


Histologically high-grade tumor exhibited significantly higher kPL, with no overlap from the low-grade cases, whereas perfusion AUC was significantly lower. Lymph-node metastases demonstrated similar kPL and perfusion indices as high-grade tumor, indicating high aggressiveness in these metastatic sites as well. These data are highly consistent with previous findings but the 3D dynamic HP 13C imaging is much less susceptible than single-timepoint studies to timing variability of bolus infusion and arrival, thus offering a more robust way to quantitatively analyze metabolism and perfusion in vivo. In conclusion, this 3D dynamic CS-EPSI acquisition and modeling approach not only allows identification and assessment of aggressiveness in preclinical primary and metastatic tumors, but also shows great potential for clinical translation.


This work was supported by grants from the DOD Prostate Cancer Research Program and the NIH (P41EB013598 & R01EB017449).


[1] (Cancer Facts and Figures 2015. Atlanta, GA: American Cancer Society; (2015)

[2] Albers M et al., Hyperpolarized 13C Lactate, Pyruvate, and Alanine: Noninvasive Biomarkers for Prostate Cancer Detection and Grading, Cancer Res. 2008. October 15, 2008 68; 8607

[3] Kurhanewicz J et al., Analysis of Cancer Metabolism by Imaging Hyperpolarized Nuclei: Prospects for Translation to Clinical Research, Neoplasia. 2011. 13(2):81-97

[4] Xing Y et al., Optimal variable flip angle schemes for dynamic acquisition of exchanging hyperpolarized substrates, JMR. 2013. 234:75–81


A) kPL images overlaid on corresponding T2 images acquired from low and high grade TRAMP tumors. A graphical comparison of kPL (B right) LDH (B left), Ki-67 staining (C) urea perfusion AUC (D right), ktrans (D left) and PIM staining (E) in high and low grade TRAMP tumors.

For N=19 TRAMPs, (9 low-grade, 10 high-grade) A) mRNA expression analysis of Hif-1α, LDHA, LDHB, VEGF, MCT1 and MCT4 in low- and high-grade tumors B) Comparison of LDHA/LDHB ratio in low- and high-grade tumors

Overlaid HP-13C MR spectra (A) and kPL image (B) showing high lactate levels and pyruvate to lactate flux in both aggressive primary tumor (N=5) and metastasis (N=5). C) No significant difference in kPL was observed between high-grade primary and lymph-node metastasis. (Primary tumor kPL =0.0572±0.0114, metastasis 0.0593±0.0261 sec-1, P>0.8)

A) Map shows estimates of urea perfusion area under curve (AUC) overlaid on T2-FSE anatomical references. B) Urea perfusion area under curve (AUC) for high-grade tumor versus metastases. (Primary tumor AUC = 679.4 ± 254.2, metastasis 909.1 ± 520.2 , P>0.4)

Representative H&E (A) Ki-67 (B) and PIM staining (C) for low and high grade tumors, and lymph node metastases. (100x magnification)

Proc. Intl. Soc. Mag. Reson. Med. 24 (2016)