Simultaneous pH- and oxygen-weighted metabolic MRI of human gliomas at 3T using multi-echo amine proton chemical exchange saturation transfer spin-and-gradient-echo echoplanar imaging (CEST-SAGE-EPI)
Benjamin M Ellingson1, Jingwen Yao1, Ararat Chakhoyan1, Phioanh L Nghiemphu2, Albert Lai2, Whitney B Pope1, Linda Liau3, and Timothy F Cloughesy2

1Radiological Sciences, University of California Los Angeles, Los Angeles, CA, United States, 2Neurology, University of California Los Angeles, Los Angeles, CA, United States, 3Neurosurgery, University of California Los Angeles, Los Angeles, CA, United States


Glycolysis is enhanced in cancers, even in the presence of abundant oxygen, leading to accumulation of lactic acid. We present a technique for fast pH- and oxygen-weighted MR imaging using multi-echo amine proton chemical exchange saturation transfer echo spin-and-gradient echoplanar imaging (CEST-SAGE-EPI) on a clinical 3T MRI system. In phantom and human experiments, we investigate the ability to simultaneously measure MTRasym at 3ppm, a measure dependent on pH, and R2’, which is sensitive to oxygen extraction. Results suggest T2 hyperintense tumor is acidic, but not hypoxic; whereas contrast enhancing tumor is acidic and hypoxic, consistent with known cancer biology.


Abnormal metabolism is a hallmark of cancer and triggers a variety of catastrophic biologic events including increased tumor aggressiveness, resistance to treatments, altered gene expression, formation of new blood vessels, and immunosuppression. Glycolysis is often enhanced in cancers, even in the presence of abundant oxygen (i.e. the Warburg effect1, Fig.1). The chemical exchange between amine and amide protons in bulk water has been shown to be pH dependent using CEST imaging2. The elevated concentration of glutamine inherently within tumors further increases the availability of proton exchange and therefore a higher CEST signal at 3.0 ppm3,4. The level of glycolysis depends on both the accumulation of lactic acid as well as the inefficient use of oxygen. Quantitative blood oxygen level dependent (qBOLD) imaging allow for non-invasive estimation of oxygen consumption. R2’, the reversible transverse relaxation rate, has been shown to correlate with oxygen extraction5-8 and has been used to explore oxygen metabolism in brain tumors7,9. In the current study we present a technique for obtaining fast pH- and oxygen-weighted MR imaging contrast using multi-echo amine proton chemical exchange saturation transfer echo spin-and-gradient echoplanar imaging (CEST-SAGE-EPI) on a clinical 3T MRI system.


Simultaneous acquisition of pH-sensitive information and relaxometry measures were performed using a CEST-SAGE-EPI sequence (Fig.2). The SAGE EPI readout consisted of two gradient echoes (TE1=14.0ms; TE2=34.1ms), an asymmetric spin echo (TE3=58.0ms), and a spin echo (TE4=92.4ms). All phantom and human CEST-SAGE-EPI data were acquired with a CEST saturation pulse train consisting of three 100-ms Gaussian pulses with amplitude B1=6uT, TR>10,000ms, FOV=240x217, matrix size=128x104, partial Fourier encoding=6/8, GRAPPA=3, bandwidth=1630 Hz/pixel, and 25 contiguous slices with a 4mm slice thickness. 29 z-spectral points was acquired with data around +/- 3.0ppm and 0.0ppm. S0 scans were acquired with NEX=4. Total acquisition time was 7 minutes and 30 seconds on a 3T Siemens Prisma. Acquired CEST-SAGE-EPI data were motion and B0 corrected. An integral of width 0.4 ppm was calculated around -3.0 and +3.0 ppm in corrected data and combined with the S0 image to calculate MTRasym at 3.0 ppm. R2’ was calculated from the SAGE-EPI readout10. Relaxometry measures of R2, R2*, and R2’ were performed using CEST-SAGE-EPI, ME-GRE, and CPMG MR acquisition in a series of 36 glutamine phantoms (100mM) with combinations of varying pH (5.0-7.5) and gadopentetic acid (Gd-DTPA; Magnevist®) concentration (0- 2.0mM) in 50mL falcon tubes. All samples were vortexed and pH was re-evaluated prior to MRI acquisition. All experiments were reapeated twice and compared to Bloch-McConnell simultations. 47 histologically proven glioma patients (WHO IV, N=20; WHO III, N=14; WHO II, N=13) were enrolled in the current study prior to initial surgical resection or at first recurrence. All patients provided informed written consent to have advanced imaging and this information included in our IRB-approved research database.


MTRasym at 3ppm within each phantom, varying by pH, was similar across all four echoes and matched simulation results (Fig.3A; P=0.999). In phantom samples containing varying concentration of Gd-DTPA, CEST-SAGE-EPI estimates of R2,R2*, and R2’ varied linearly with concentration (Fig.3B) and matched ME-GRE and CPMG measurements, with r2=6.24±0.04 mM-1×sec-1 (P<0.0001) and r2*=6.86±0.10 mM-1×sec-1 (P<0.0001). T2 hyperintense lesions in all patients exhibited heterogeneous areas of elevated MTRasym at 3ppm (acidity) and R2’ (hypoxia)(Fig.4). Interestingly, T2 hyperintense lesions exhibited a significantly lower median R2’ (4.8±0.2 sec-1) compared with NAWM (6.2±0.2 sec-1) (Fig.5A; P<0.0001) and did not vary significantly across tumor grade (Fig.5B; P=0.0537), suggesting non-enhancing tumor regions may be adequately oxygenated and undergoing aerobic glycolysis. In glioblastoma, significant differences in median R2’ across tissue types were observed (Fig.5C; P=0.0001), where R2’ was significantly lower in T2 hyperintense lesions (5.3±0.3 sec-1) compared to both NAWM (6.3±0.2 sec-1; P=0.0078) and CE regions (10.0±1.0 sec-1; P=0.0002). MTRasym within T2 hyperintense lesions (1.7±0.1%) were significantly higher than NAWM (0.8±0.03%) (Fig.5D; P<0.0001) and varied across grade (Fig.5E;P=0.0297). Combined information about both median tissue acidity (MTRasym at 3ppm) and oxygen extraction (R2’) helps to further delineate various tissue types and provide additional insights into metabolic behavior.


The current study presents a new CEST-SAGE-EPI sequence for obtaining pH- and oxygen-sensitive image contrasts for brain tumor evaluation. Advantages include speed, in-line B0 correction, whole brain coverage, and simultaneous accurate estimation of CEST effects and relaxation rate Results in tumors showed a high degree of spatial heterogeneity and measurements were consistent with known cancer biology.


We would like to acknowledge Catalina Raymond, M.S., and Saima Charni-Caabane, Ph.D., for their assistance with data collection.


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Fig 1. The Warburg Effect. A) Within normal tissues in the presence of oxygen, glucose
is converted to pyruvate and then used for oxydative phosphorylation within the mitochondria. B) In normal tissues within a hypoxic environment, glucose is converted to pyruvate then to lactate or lactic acid, decreasing extracellular pH. C) In cancer cells, glucose is converted to pyruvate then to lactate and lactic acid (80-85%) and a small portion of pyruvate enters the citric acid cycle (5-15%), regardless of whether oxygen is present.

Fig 2. Pulse sequence diagram for a multi-echo amine chemical exchange saturation transfer spin-and-gradient echo echoplanar imaging (CEST-SAGE-EPI) sequence. After non-selective off-resonance CEST excitation consisting of three 100-ms Gaussian pulses, a spectral-spatial water only excitation pulse is invoked. Following excitation two gradient echo EPI trains are acquired, followed by a 180-degree refocusing pulse. Lastly, an asymmetric spin echo EPI train and a standard spin-echo EPI train are acquired.

Fig 3. Simulation and phantom testing. A) CEST contrast increases with decreasing pH. B) R2 and R2* measured using CEST-SAGE-EPI show a linear increase with increasing Gd-DTPA concentration. C) Comparison between measured R2 using CEST-SAGE-EPI and CPMG show strong correlation (R2=0.9943, P<0.0001), independent of pH (P=0.9915). D) Comparison between measured R2* using CEST-SAGE-EPI and ME-GRE showing a strong correlation (R2=0.9727, P<0.0001) and no dependence on pH (P=0.2184). E) Amine CEST has decreased sensitivity in the presence of contrast agents that shorten T1 and T2. F) COV measured from test-retest is higher in environments with shorter T1 and T2 .

Fig 4. pH- and oxygen-weighted MR images of human gliomas. A) A nonenhancing recurrent WHO II IDH mutant astrocytoma exhibiting a region of focal acidity (MTRasym) and low oxygen consumption (R2'). B) A newly diagnosed nonenhancing IDH mutant WHO III anaplastic astrocytoma with large, heterogeneous regions of abnormally high and low hypoxia and acidity. C) A recurrent IDH wild type WHO IV glioblastoma exhibiting a region of focal acidity and large regions of low oxygen consumption. D) A newly diagnosed IDH wild type WHO IV glioblastoma with ring enhancement, central necrosis, elevated acidity and hypoxia within the enhancing region.

Fig 5. pH and oxygen-weighted MR measurements in human glioma patients. A) R2’ in normal appearing white matter (NAWM) and the T2 hyperintense lesions in all patients; B) in T2 hyperintense regions for different tumor grades; and C) in glioblastoma (GBM). D) MTRasym at 3ppm (%) in NAWM and T2 hyperintense lesions in all patients; E) across tumor grades; and F) in different tissue types in GBM. G) MTRasym versus R2’ for NAWM and T2 hyperintense lesions in all patients pooled across tumor grades; H) for different tumor grades; and in GBM. * = P<0.05, **=P<0.01, ***=P<0.001, ****=P<0.0001.

Proc. Intl. Soc. Mag. Reson. Med. 26 (2018)