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Background: Amide proton transfer (APT) imaging is a chemical exchange saturation transfer (CEST) technique offering potential clinical applications such as diagnosis, characterization, and treatment planning and monitoring in glioma patients. While APT-CEST has demonstrated high potential, reproducibility remains underexplored. Purpose: To investigate whether cerebral APT-CEST with clinically feasible scan time is reproducible in healthy tissue and glioma for clinical use at 3 T. Study Type: Prospective, longitudinal. Subjects: Twenty-one healthy volunteers (11 females; mean age ± SD: 39 ± 11 years) and 6 glioma patients (3 females; 50 ± 17 years: 4 glioblastomas, 1 oligodendroglioma, 1 radiologically suspected low-grade glioma). Field Strength/Sequence: 3 T, Turbo Spin Echo - ampling perfection with application optimized contrasts using different flip angle evolution - chemical exchange saturation transfer (TSE SPACE-CEST). Assessment: APT-CEST measurement reproducibility was assessed within-session (glioma patients, scan session 1; healthy volunteers scan sessions 1, 2, and 3), between-sessions (healthy volunteers scan sessions 1 and 2), and between-days (healthy volunteers, scan sessions 1 and 3). The mean APT CEST values and standard deviation of the within-subject difference (SD diff) were calculated in whole tumor enclosed by regions of interest (ROIs) in patients, and eight ROIs in healthy volunteers—whole-brain, cortical gray matter, putamen, thalami, orbitofrontal gyri, occipital lobes, central brain—and compared. Statistical Tests: Brown-Forsythe tests and variance component analysis (VCA) were used to assess the reproducibility of ROIs for the three time intervals. Significance was set at P < 0.003 after Bonferroni correction. Results: Intratumoral mean APT CEST was significantly higher than APT CEST in healthy-appearing tissue in patients (0.5 ± 0.46%). The average within-session, between-sessions, and between-days SD diff of healthy control brains was 0.2% and did not differ significantly with each other (0.76 > P > 0.22). The within-session SD diff of whole-brain was 0.2% in both healthy volunteers and patients, and 0.21% in the segmented tumor. VCA showed that within-session factors were the most important (60%) for scanning variance. Data Conclusion: Cerebral APT-CEST imaging may show good scan–rescan reproducibility in healthy tissue and tumors with clinically feasible scan times at 3 T. Short-term measurement effects may be the dominant components for reproducibility. Level of Evidence: 2. Technical Efficacy: Stage 2.

Cancer is one of the most devastating diseases that the world is currently facing, accounting for 10 million deaths in 2020 (WHO). In the last two decades, advanced medical imaging has played an ever more important role in the early detection of the disease, as it increases the chances of survival and the potential for full recovery. To date, dynamic glucose-enhanced (DGE) MRI using glucose-based chemical exchange saturation transfer (glucoCEST) has demonstrated the sensitivity to detect both d-glucose and glucose analogs, such as 3-oxy-methyl-d-glucose (3OMG) uptake in tumors. As one of the recent international efforts aiming at pushing the boundaries of translation of the DGE MRI technique into clinical practice, a multidisciplinary team of eight partners came together to form the “glucoCEST Imaging of Neoplastic Tumors (GLINT)” consortium, funded by the Horizon 2020 European Commission. This paper summarizes the progress made to date both by these groups and others in increasing our knowledge of the underlying mechanisms related to this technique as well as translating it into clinical practice.

Purpose: In this work, we investigated the ability of neural networks to rapidly and robustly predict Lorentzian parameters of multi-pool CEST MRI spectra at 7 T with corresponding uncertainty maps to make them quickly and easily available for routine clinical use.Methods: We developed a deepCEST 7 T approach that generates CEST contrasts from just 1 scan with robustness against B1 inhomogeneities. The input data for a neural feed-forward network consisted of 7 T in vivo uncorrected Z-spectra of a single B1 level, and a B1 map. The 7 T raw data were acquired using a 3D snapshot gradient echo multiple interleaved mode saturation CEST sequence. These inputs were mapped voxel-wise to target data consisting of Lorentzian amplitudes generated conventionally by 5-pool Lorentzian fitting of normalized, denoised, B0 - and B1 -corrected Z-spectra. The deepCEST network was trained with Gaussian negative log-likelihood loss, providing an uncertainty quantification in addition to the Lorentzian amplitudes.Results: The deepCEST 7 T network provides fast and accurate prediction of all Lorentzian parameters also when only a single B1 level is used. The prediction was highly accurate with respect to the Lorentzian fit amplitudes, and both healthy tissues and hyperintensities in tumor areas are predicted with a low uncertainty. In corrupted cases, high uncertainty indicated wrong predictions reliably.Conclusion: The proposed deepCEST 7 T approach reduces scan time by 50% to now 6:42 min, but still delivers both B0 - and B1 -corrected homogeneous CEST contrasts along with an uncertainty map, which can increase diagnostic confidence. Multiple accurate 7 T CEST contrasts are delivered within seconds.

I want to determine how underfitting CEST data in vivo may cause parameter bias due to unmodelled effects

Upper urinary tract obstructions (UTOs) are blockages that inhibit the flow of urine through its normal course, leading to impaired kidney function. Imaging plays a significant role in the initial diagnosis of UTO, with anatomic imaging (primarily ultrasound (US) and non-contrast computed tomography (CT)) serving as screening tools for the detection of the dilation of the urinary collecting systems (i.e., hydronephrosis). Whether hydronephrosis represents UTO or a non-obstructive process is determined by functional imaging (typically nuclear medicine renal scintigraphy). If these exams reveal evidence of UTO but no discernable source, multiphase contrast enhanced CT urography and/or dynamic contrast enhanced MR urography (DCE-MRU) may be performed to delineate a cause. These are often performed in conjunction with direct ureteroscopic evaluation. While contrast-enhanced CT currently predominates, it can induce renal injury due to contrast induced nephropathy (CIN), subject patients to ionizing radiation and is limited in quantifying renal function (traditionally assessed by renal scintigraphy) and establishing the extent to which hydronephrosis is due to functional obstruction. Traditional MRI is similarly limited in its ability to quantify function. DCE-MRU presents concerns regarding nephrogenic systemic fibrosis (NSF), although decreased with newer gadolinium-based contrast agents, and regarding cumulative gadolinium deposition in the basal ganglia. DCE-MR CEST urography is a promising alternative, employing new MRI contrast agents and imaging schemes and allowing for concurrent assessment of renal anatomy and functional parameters. In this review we highlight clinical challenges in the diagnosis and management of UTO, identify key advances in imaging agents and techniques for DCE-MR CEST urography and provide perspective on how this technique may evolve in clinical importance.

A relationship between an acidic pH in the joints, osteoarthritis (OA), and pain has been previously demonstrated. Acidosis Chemical Exchange Saturation Transfer (acidoCEST) indirectly measures the extracellular pH through the assessment of the exchange of protons between amide groups on iodinated contrast agents and bulk water. It is possible to estimate the extracellular pH in the osteoarthritic joint using acidoCEST MRI. However, conventional MR sequences cannot image deep layers of cartilage, meniscus, ligaments, and other musculoskeletal tissues that present with short echo time and fast signal decay. Ultrashort echo time (UTE) MRI, on the other hand, has been used successfully to image those joint tissues. Here, our goal is to compare the pH measured in the knee joints of volunteers without OA and patients with severe OA using acidoCEST-UTE MRI. Patients without knee OA and patients with severe OA were examined using acidoCEST-UTE MRI and the mean pH of cartilage, meniscus, and fluid was calculated. Additionally, the relationship between the pH measurements and the Knee Injury and Osteoarthritis Outcome Score (KOOS) was investigated. AcidoCEST-UTE MRI can detect significant differences in the pH of knee cartilage, meniscus, and fluid between joints without and with OA, with OA showing lower pH values. In addition, symptoms and knee-joint function become worse at lower pH measurements.

Purpose Chemical exchange saturation transfer MRI can provide accurate pH images, but the slow scan time (due to long saturation periods and multiple offsets sampling) reduce both the volume coverage and spatial resolution capability, hence the possibility to interrogate the heterogeneity in tumors and organs. To overcome these limitations, we propose a fast multislice CEST-MRI sequence with high pH accuracy and spatial resolution. Methods The sequence first uses a long saturation pulse to induce the steady-state CEST contrast and a second short saturation pulse repeated after each image acquisition to compensate for signal losses based on an uneven irradiation scheme combined with a single-shot rapid acquisition with refocusing echoes readout. Sequence sensitivity and accuracy in measuring pH was optimized by simulation and assessed by in vitro studies in pH-varying phantoms. In vivo validation was performed in two applications by acquiring multislice pH images covering the whole tumors and kidneys after iopamidol injection. Results Simulated and in vivo data showed comparable contrast efficiency and pH responsiveness by reducing saturation time. The experimental data from a homogeneous, pH-varying, iopamidol-containing phantom show that the sequence produced a uniform CEST contrast across slices and accurate values across slices in less than 10 minutes. In vivo measurements allowed us to quantify the 3D pH gradients of tumors and kidneys, with pH ranges comparable with the literature. Conclusion The proposed fast multislice CEST-MRI sequence allows volumetric acquisitions with good pH sensitivity, accuracy, and spatial resolution for several in vivo pH imaging applications.

Purpose To evaluate the influence of phosphate on amine, amide, and hydroxyl CEST contrast using Bloch-McConnell simulations applied to physical phantom data. Methods Phantom solutions of 4 representative metabolites with exchangeable protons-glycine (α-amine protons), Cr (η-amine protons), egg white protein (amide protons), and glucose (hydroxyl protons)-were prepared at different pH levels (5.6 to 8.9) and phosphate concentrations (5 to 80 mM). CEST images of the phantom were collected with CEST-EPI sequence at 3 tesla. The CEST data were then fitted to full Bloch-McConnell equation simulations to estimate the exchange rate constants. With the fitted parameters, simulations were performed to evaluate the intracellular and extracellular contributions of CEST signals in normal brain tissue and brain tumors, as well as in dynamic glucose-enhanced experiments. Results The exchange rates of α-amine and hydroxyl protons were found to be highly dependent on both pH and phosphate concentrations, whereas the exchange rates of η-amine and amide protons were pH-dependent, albeit not catalyzed by phosphate. With phosphate being predominantly intracellular, CEST contrast of α-amine exhibited a higher sensitivity to changes in the extracellular microenvironment. Simulations of dynamic glucose-enhanced signals demonstrated that the contrast between normal and tumor tissue was mostly due to the extracellular CEST effect. Conclusion The proton exchange rates in some metabolites can be greatly catalyzed by the presence of phosphate at physiological concentrations, which substantially alters the CEST contrast. Catalytic agents should be considered as confounding factors in future CEST-MRI research. This new dimension may also benefit the development of novel phosphate-sensitive imaging methods.

Measuring the magnetization of atomic nuclei as in magnetic resonance imaging (MRI) allows studying molecular systems under chemical exchange. The MRI signal of the water-soluble gas xenon provides a high specificity to its local molecular environment. To increase the sensitivity, hyperpolarization techniques can be applied and also a method called chemical exchange saturation transfer (CEST). Through CEST, molecular hosts that enclose Xe are detected and features of their Xe exchange are revealed. However, the CEST method is slow since multiple acquisitions along a spectral dimension are needed to characterize the exchange. How to accelerate the CEST experiment is a main question of this work which was addressed by introducing a novel method called CAVKA (combined approach of variable flip angle, keyhole, and averaging). It divides the signal domain into two parts, performs time-consuming signal averaging only for the nearly constant part while rapidly updating the second part that encodes image contrast along the spectral dimension. The two are then computationally merged with consideration of the magnetization’s physical properties for MRI scans. CAVKA encodes the magnetization limited information from multiple samples as an image. Main benefit is a fast scan of host systems with relative fast exchange that otherwise causes signal loss. After introducing CAVKA, its application to different host systems is described. The synthetic host cryptophane-A and biogenic gas vesicles were used as a proof of concept to study the method performance and adjust its parameters. Later on, cucurbit[n]urils (CB6 and CB7) were extensively investigated with competitive guests to reveal differences in the host accessibility for Xe. It became clear that the observed CEST signal from commercially available CB7 samples is actually from CB6, an impurity from the synthesis of CB7. This impurity was also quantitatively estimated to be ca. 8.5%. Finally, data denoising by principal component analysis (PCA) and wavelets was explored. Insights from their tailoring to MRI data of hyperpolarized Xe are reported. The two data domains of MRI (k-space and image space) were denoised and the performance was evaluated in terms of the SNR increase., Mit der Magnetresonanztomographie (MRT) lassen sich die Magnetisierung von Atomkernen messen und molekulare Systeme unter chemischem Austausch untersuchen. Das wasserlösliche Gas Xenon hat den Vorteil, dass sein MRT-Signal sehr empfindlich auf die lokale molekulare Umgebung reagiert. Zur Steigerung der Sensitivität können Techniken der Hyperpolarisation angewendet werden und auch das Verfahren CEST (chemical exchange saturation transfer). Mit CEST werden molekulare Wirte, die Xe einschließen, untersucht und die Merkmale ihres Xenon-Austauschs vermessen. Das reguläre CEST-Experiment ist jedoch langsam, da mehrere Aufnahmen entlang der spektralen Dimension notwendig sind, um den Austausch zu charakterisieren. Eine Hauptfrage dieser Dissertation ist, wie man das CEST-Experiment beschleunigen kann. Hierzu wurde die neue Methode CAVKA (combined approach of variable flip angle, keyhole and averaging) entwickelt. Sie teilt die Signaldomäne in zwei Teile, führt eine zeitaufwendige Signalermittelung nur für den fast konstanten Teil durch, während der zweite Teil, der den Bildkontrast entlang der spektralen Dimension kodiert, schnell aktualisiert wird. Beide Teile werden unter Beachtung der physikalischen Eigenschaften der Magnetisierung bei der MRT anschließend wieder rechnergestützt zusammengefügt. CAVKA kodiert die Informationen mehrerer Proben als Bild. Der Hauptvorteil ist der schnelle Scan von Wirtssystemen mit relativ schnellem Austausch, der einen schnellen Signal-Abfall verursacht. CAVKA wurde für verschiedenen Wirtssysteme eingesetzt. Der synthetische Wirt Cryptophan-A und biogene Gasvesikel wurden im Machbarkeitsnachweis verwendet. Später wurden Cucurbit[n]urile (CB6 und CB7) mit kompetitiven Gästen untersucht, um Unterschiede in der Wirtszugänglichkeit für Xe aufzudecken. Es wurde deutlich, dass das beobachtete CEST-Signal von kommerziellen CB7-Proben tatsächlich vom CB6 stammt, und zwar als Verunreinigung, die ein Nebenprodukt bei der CB7-Synthese ist. Diese Verunreinigung wurde auf ca. 8,5% quantifiziert. Schließlich wurden Xe MRT-Daten durch die Techniken Principal Component Analysis (PCA) und Wavelets entrauscht. Die beiden Datendomänen der MRT (k-space und Bildraum) wurden untersucht und die SNR-Steigerung bewertet.

Distinguishing primary central nervous system lymphoma (PCNSL) from glioblastoma, isocitrate dehydrogenase (IDH)-wildtype is sometimes hard. Because the role of operation on them varies, accurate preoperative diagnosis is crucial. In this study, we evaluated whether a specific kind of chemical exchange saturation transfer imaging, i.e., amide proton transfer-weighted (APTw) imaging, was useful to distinguish PCNSL from glioblastoma, IDH-wildtype. A total of 14 PCNSL and 27 glioblastoma, IDH-wildtype cases were evaluated. There was no significant difference in the mean APTw signal values between the two groups. However, the percentile values from the 1st percentile to the 20th percentile APTw signals and the width1–100 APTw signals significantly differed. The highest area under the curve was 0.796, which was obtained from the width1–100 APTw signal values. The sensitivity and specificity values were 64.3% and 88.9%, respectively. APTw imaging was useful to distinguish PCNSL from glioblastoma, IDH-wildtype. To avoid unnecessary aggressive surgical resection, APTw imaging is recommended for cases in which PCNSL is one of the differential diagnoses.

For precise delineation of glioma extent, amino acid PET is superior to conventional MR imaging. Since metabolic MR sequences such as chemical exchange saturation transfer (CEST) imaging and MR spectroscopy (MRS) were developed, we aimed to evaluate the diagnostic accuracy of combined CEST and MRS to predict glioma infiltration. Eighteen glioma patients of different tumor grades were enrolled in this study; 18F-fluoroethyltyrosine (FET)-PET, amide proton transfer CEST at 7 Tesla(T), MRS and conventional MR at 3T were conducted preoperatively. Multi modalities and their association were evaluated using Pearson correlation analysis patient-wise and voxel-wise. Both CEST (R = 0.736, p < 0.001) and MRS (R = 0.495, p = 0.037) correlated with FET-PET, while the correlation between CEST and MRS was weaker. In subgroup analysis, APT values were significantly higher in high grade glioma (3.923 ± 1.239) and IDH wildtype group (3.932 ± 1.264) than low grade glioma (3.317 ± 0.868, p < 0.001) or IDH mutant group (3.358 ± 0.847, p < 0.001). Using high FET uptake as the standard, the CEST/MRS combination (AUC, 95% CI: 0.910, 0.907–0.913) predicted tumor infiltration better than CEST (0.812, 0.808–0.815) or MRS (0.888, 0.885–0.891) alone, consistent with contrast-enhancing and T2-hyperintense areas. Probability maps of tumor presence constructed from the CEST/MRS combination were preliminarily verified by multi-region biopsies. The combination of 7T CEST/MRS might serve as a promising non-radioactive alternative to delineate glioma infiltration, thus reshaping the guidance for tumor resection and irradiation.

D-Glucose and 3-O-Methyl-D-glucose (3OMG) have been shown to provide contrast in magnetic resonance imaging-chemical exchange saturation transfer (MRI-CEST) images. However, a systematic comparison between these two molecules has yet to be performed. The current study deals with the assessment of the effect of pH, saturation power level (B1 ) and magnetic field strength (B0 ) on the MRI-CEST contrast with the aim of comparing the in vivo CEST contrast detectability of these two agents in the glucoCEST procedure. Phosphate-buffered solutions of D-Glucose or 3OMG (20 mM) were prepared at different pH values and Z-spectra were acquired at several B1 levels at 37°C. In vivo glucoCEST images were obtained at 3 and 7 T over a period of 30 min after injection of D-Glucose or 3OMG (at doses of 1.5 or 3 g/kg) in a murine melanoma tumor model (n = 3-5 mice for each molecule, dose and B0 field). A markedly different pH dependence of CEST response was observed in vitro for D-Glucose and 3OMG. The glucoCEST contrast enhancement in the tumor region following intravenous administration (at the 3 g/kg dose) was comparable for both molecules: 1%-2% at 3 T and 2%-3% at 7 T. The percentage change in saturation transfer that resulted was almost constant for 3OMG over the 30-min period, whereas a significant increase was detected for D-Glucose. Our results show similar CEST contrast efficiency but different temporal kinetics for the metabolizable and the nonmetabolizable glucose derivatives in a tumor murine model when administered at the same doses.

© 2019, Springer Science+Business Media, LLC, part of Springer Nature. Introduction: Non-invasively distinguishing aggressive from non-aggressive brain tumors is an important clinical challenge. Intracellular pH (pHi) regulation is essential for normal cell function and is normally maintained within a narrow range. Cancer cells are characterized by a reversed intracellular to extracellular pH gradient, compared to healthy cells, that is maintained by several distinct mechanisms. Previous studies have demonstrated acute pH modulation in glioblastoma detectable by chemical exchange saturation transfer (CEST) magnetic resonance imaging (MRI) after blocking individual pH regulatory mechanisms. The purpose of the current study was to simultaneously block five pH regulatory mechanisms while also providing glucose as an energy substrate. We hypothesized that this approach would increase the acute pH modulation effect allowing the identification of aggressive cancer. Methods: Using a 9.4 T MRI scanner, CEST spectra were acquired sensitive to pHi using amine/amide concentration independent detection (AACID). Twelve mice were scanned approximately 11 ± 1 days after implanting 105 U87 human glioblastoma multiforme cells in the brain, before and after intraperitoneal injection of a combination of five drugs (quercetin, cariporide, dichloroacetate, acetazolamide, and pantoprazole) with and without glucose. Results: Two hours after combination drug injection there was a significant 0.1 ± 0.03 increase in tumor AACID value corresponding to a 0.4 decrease in pHi. After injecting the drug combination with glucose the AACID value increased by 0.18 ± 0.03 corresponding to a 0.72 decrease in pHi. AACID values were also slightly increased in contralateral tissue. Conclusions: The combined drug treatment with glucose produced a large acute CEST MRI contrast indicating tumor acidification, which could be used to help localize brain cancer and monitor tumor response to chemotherapy.

PurposeThe objective of the current study was to explore the efficacy of using pH-weighted amine CEST-EPI as a potential non-invasive imaging biomarker for treatment response and/or failure in recurrent GBM patients treated with bevacizumab.MethodA total of 11 patients with recurrent GBM treated with bevacizumab were included in this prospective study. CEST-EPI, perfusion MRI, and standardized anatomic MRI were obtained in patients before and after bevacizumab administration. CEST-EPI measures of magnetization transfer ratio asymmetry (MTRasym) at 3ppm were used for pH-weighted imaging contrast. Multiple measures were examined for their association with progression-free survival (PFS).ResultTumor acidity, measured with MTRasym at 3ppm, was significantly reduced in both contrast enhancing and non-enhancing tumor after bevacizumab (p = 0.0002 and p

International journal of molecular sciences 23(13), 6920 (2022). doi:10.3390/ijms23136920 special issue: "Special Issue "Highlights in Pathophysiology of the Musculoskeletal System" / Special Issue Editors: Dr. Marta Anna Szychlinska, Guest Editor; Dr. Rosario Barone, Guest Editor", Published by Molecular Diversity Preservation International, Basel

Dynamic glucose-enhanced (DGE) imaging uses chemical exchange saturation transfer magnetic resonance imaging to retrieve information about the microcirculation using infusion of a natural sugar (D-glucose). However, this new approach is not yet well understood with respect to the dynamic tissue response. DGE time curves for arteries, normal brain tissue, and cerebrospinal fluid (CSF) were analyzed in healthy volunteers and compared with the time dependence of sampled venous plasma blood glucose levels. The arterial response curves (arterial input function [AIF]) compared reasonably well in shape with the time curves of the sampled glucose levels but could also differ substantially. The brain tissue response curves showed mainly negative responses with a peak intensity that was of the order of 10 times smaller than the AIF peak and a shape that was susceptible to both noise and partial volume effects with CSF, attributed to the low contrast-to-noise ratio. The CSF response curves showed a rather large and steady increase of the glucose uptake during the scan, due to the rapid uptake of D-glucose in CSF. Importantly, and contrary to gadolinium studies, the curves differed substantially among volunteers, which was interpreted to be caused by variations in insulin response. In conclusion, while AIFs and tissue response curves can be measured in DGE experiments, partial volume effects, low concentration of D-glucose in tissue, and osmolality effects between tissue and blood may prohibit quantification of normal tissue perfusion parameters. However, separation of tumor responses from normal tissue responses would most likely be feasible.

2D NOESY plays a central role in structural NMR spectroscopy. We have recently discussed methods that rely on solvent‐driven exchanges to enhance NOE correlations between exchangeable and non‐exchangeable protons in nucleic acids. Such methods, however, fail when trying to establish connectivities within pools of labile protons. This study introduces an alternative that also enhances NOEs between such labile sites, based on encoding a priori selected peaks by selective saturations. The resulting selective magnetization transfer (SMT) experiment proves particularly useful for enhancing the imino–imino cross‐peaks in RNAs, which is a first step in the NMR resolution of these structures. The origins of these enhancements are discussed, and their potential is demonstrated on RNA fragments derived from the genome of SARS‐CoV‐2, recorded with better sensitivity and an order of magnitude faster than conventional 2D counterparts., An approach to enhance the sensitivity of 2D NOESY correlations among labile protons is introduced and exemplified with experiments on RNA fragments derived from the SARS‐CoV‐2 genome and with other samples. The approach leads to experiments that are ca. 10‐fold faster and 3‐fold more sensitive than conventional counterparts. The mechanism of this sensitivity boost is explained, and its strengths and limitations are analyzed.

Purpose Chemical exchange saturation transfer MRI provides new approaches for investigating tumor microenvironment, including tumor acidosis that plays a key role in tumor progression and resistance to therapy. Following iopamidol injection, the detection of the contrast agent inside the tumor tissue allows measurements of tumor extracellular pH. However, accurate tumor pH quantifications are hampered by the low contrast efficiency of the CEST technique and by the low SNR of the acquired CEST images, hence in a reduced detectability of the injected agent. This work aims to investigate a novel denoising method for improving both tumor pH quantification and accuracy of CEST-MRI pH imaging. Methods An hybrid denoising approach was investigated for CEST-MRI pH imaging based on the combination of the nonlocal mean filter and the anisotropic diffusion tensor method. The denoising approach was tested in simulated and in vitro data and compared with previously reported methods for CEST imaging and with established denoising approaches. Finally, it was validated with in vivo data to improve the accuracy of tumor pH maps. Results The proposed method outperforms current denoising methods in CEST contrast quantification and detection of the administered contrast agent at several increasing noise levels with simulated data. In addition, it achieved a better pH quantification in in vitro data and demonstrated a marked improvement in contrast detection and a substantial improvement in tumor pH accuracy in in vivo data. Conclusion The proposed approach effectively reduces the noise in CEST images and increases the sensitivity detection in CEST-MRI pH imaging.

Background. With fast-growing evidence in literature for clinical applications of chemical exchange saturation transfer (CEST) magnetic resonance imaging (MRI), this prospective study aimed at applying amide proton transfer-weighted (APTw) CEST imaging in a clinical setting to assess its diagnostic potential in differentiation of intracranial tumors at 3 tesla (T). Methods. Using the asymmetry magnetization transfer ratio (MTRasym) analysis, CEST signals were quantitatively investigated in the tumor areas and in a similar sized region of the normal-appearing white matter (NAWM) on the contralateral hemisphere of 27 patients with intracranial tumors. Area under curve (AUC) analyses were used and results were compared to perfusion-weighted imaging (PWI). Results. Using APTw CEST, contrast-enhancing tumor areas showed significantly higher APTw CEST metrics than contralateral NAWM (AUC = 0.82; p < 0.01). In subgroup analyses of each tumor entity vs. NAWM, statistically significant effects were yielded for glioblastomas (AUC = 0.96; p < 0.01) and for meningiomas (AUC = 1.0; p < 0.01) but not for lymphomas as well as metastases (p > 0.05). PWI showed results comparable to APTw CEST in glioblastoma (p < 0.01). Conclusions. This prospective study confirmed the high diagnostic potential of APTw CEST imaging in a routine clinical setting to differentiate brain tumors.

// Jonathan Klein 1, 2, 3, 4 , Wilfred W. Lam 1 , Gregory J. Czarnota 1, 2, 3, 4 and Greg J. Stanisz 1, 2, 5 1 Physical Sciences, Sunnybrook Research Institute, Toronto, Ontario, Canada 2 Department of Medical Biophysics, University of Toronto, Toronto, Ontario, Canada 3 Department of Radiation Oncology, University of Toronto, Toronto, Ontario, Canada 4 Department of Radiation Oncology, Odette Cancer Centre, Sunnybrook Health Sciences Centre, Toronto, Ontario, Canada 5 Department of Neurosurgery and Pediatric Neurosurgery, Medical University of Lublin, Lublin, Poland Correspondence to: Greg J. Stanisz, email: stanisz@sri.utoronto.ca Keywords: CEST; MRI; breast cancer; cell death; response monitoring Received: March 14, 2018 Accepted: July 12, 2018 Published: July 31, 2018 ABSTRACT Purpose: Detecting cell death and predicting tumor response early in a course of chemotherapy could help optimize treatment regimens and improve clinical outcomes. Chemical exchange saturation transfer (CEST) MRI was investigated in vivo to study properties that may be able to detect cancer death. Results: Using a magnetization transfer ratio (MTR) cutoff of 0.12 at 1.8 ppm was able to differentiate between viable tumor and cell death regions. Comparison of MTR values at this frequency showed significant differences ( p < 0.0001) between viable tumor and cell death regions, matching patterns seen on histology. Using this cutoff, the mean increase in cell death index (± standard error of the mean) after chemotherapy was 4 ± 4%, 10% ± 7%, 10 ± 8%, and 4 ± 9% at 4, 8, 12, and 24 h, respectively. Conclusions: CEST MRI can detect cell death in MDA-231 xenografts but further work is needed to characterize the clinical applications of this finding. Maximum response to chemotherapy occurred at 8–12 h after chemotherapy injection in this in vivo tumor model. Materials and Methods: Breast cancer xenografts (MDA-MB-231) were scanned using 7 T MRI before and after chemotherapy. As a measure of CEST effect at 0.5 μT saturation amplitude, MTR values at frequency offsets of 1.8 and −3.3 ppm were evaluated. CEST signals after chemotherapy treatment were compared to cell-death histopathology of tumors.

Glioblastoma is an aggressive brain cancer that is very difficult to treat. Clinically, it is important to be able to distinguish aggressive from non-aggressive brain tumors. Previous studies have shown that some drugs can induce a rapid change in intracellular pH that could help to identify aggressive cancer. The sodium proton exchanger (NHE1) plays a significant role in maintaining pH balance in the tumor microenvironment. Cariporide is a sodium proton exchange inhibitor that is well tolerated by humans in cardiac applications. We hypothesized that cariporide could selectively acidify brain tumors. The purpose of this study was to determine whether amine/amide concentration-independent detection (AACID) chemical exchange saturation transfer (CEST) MRI measurement of tumor pHi could detect acidification after cariporide injection. Using a 9.4T MRI scanner, CEST spectra were acquired in six mice approximately 14 days after implanting 105 U87 human glioblastoma multiforme cells in the brain, before and after administration of cariporide (dose: 6 mg/kg) by intraperitoneal injection. Three additional mice were studied as controls and received only vehicle injection (DMSO + PBS). Repeated measures t test was used to examine changes in tumor and contralateral tissue regions of interest. Two hours after cariporide injection, there was a significant 0.12 ± 0.03 increase in tumor AACID value corresponding to a 0.48 decrease in pHi and no change in AACID value in contralateral tissue. A small but significant increase of 0.04 ± 0.017 in tumor AACID value was also observed following vehicle injection. This study demonstrates that acute CEST MRI contrast changes, indicative of intracellular acidification, after administration of cariporide could help localize glioblastoma.

Alzheimer’s disease (AD) is one of the most common causes of dementia and difficult to study as the pool of subjects is highly heterogeneous. Saturation transfer (ST) magnetic resonance imaging (MRI) methods are quantitative modalities with potential for non-invasive identification and tracking of various aspects of AD pathology. In this review we cover ST-MRI studies in both humans and animal models of AD over the past 20 years. A number of magnetization transfer (MT) studies have shown promising results in human brain. Increased computing power enables more quantitative MT studies, while access to higher magnetic fields improves the specificity of chemical exchange saturation transfer (CEST) techniques. While much work remains to be done, results so far are very encouraging. MT is sensitive to patterns of AD-related pathological changes, improving differential diagnosis, and CEST is sensitive to particular pathological processes which could greatly assist in the development and monitoring of therapeutic treatments of this currently incurable disease.

Chemical Exchange Saturation Transfer (CEST) approach is a novel tool within magnetic resonance imaging (MRI) that allows visualization of molecules possessing exchangeable protons with water. Many molecules, employed as excipients for the formulation of finished drug products, are endowed with hydroxyl, amine or amide protons, thus can be exploitable as MRI-CEST contrast agents. Their high safety profiles allow them to be injected at very high doses. Here we investigated the MRI-CEST properties of several excipients (ascorbic acid, sucrose, N-acetyl-D-glucosamine, meglumine and 2-pyrrolidone) and tested them as tumor-detecting agents in two different murine tumor models (breast and melanoma cancers). All the investigated molecules showed remarkable CEST contrast upon i.v. administration in the range 1-3 ppm according to the type of mobile proton groups. A marked increase of CEST contrast was observed in tumor regions up to 30 min post injection. The combination of marked tumor contrast enhancement and lack of toxicity make these molecules potential candidates for the diagnosis of tumors within the MRI-CEST approach. (C) 2017 Elsevier B.V. All rights reserved.

© 2018, Springer Science+Business Media, LLC, part of Springer Nature. The response of tumor intracellular pH to a pharmacological challenge could help identify aggressive cancer. Chemical exchange saturation transfer (CEST) is an MRI contrast mechanism that is dependent on intracellular pH (pHi). pHi is important in the maintenance of normal cell function and is normally maintained within a narrow range by the activity of transporters located at the plasma membrane. In cancer, changes in pHi have been correlated with both cell proliferation and cell death. Quercetin is a bioflavonoid and monocarboxylate transporter (MCT) inhibitor. Since MCTs plays a significant role in maintaining pH balance in the tumor microenvironment, we hypothesized that systemically administered quercetin could selectively acidify brain tumors. The goals of the current study were to determine whether CEST MRI measurements sensitive to tumor pH could detect acidification after quercetin injection and to measure the magnitude of the pH change (ΔpH). Using a 9.4 T MRI, amine and amide concentration independent detection (AACID) CEST spectra were acquired in six mice approximately 15 ± 1 days after implanting 105 U87 human glioblastoma multiforme cells in the brain, before and after administration of quercetin (dose: 200 mg/kg) by intraperitoneal injection. Three additional mice were studied as controls and received only vehicle dimethyl sulfoxide (DMSO) injection. Repeated measures t-test was used to compare AACID changes in tumor and contralateral tissue regions of interest. Two hours after quercetin injection there was a significant increase in tumor AACID by 0.07 ± 0.03 corresponding to a 0.27 decrease in pHi, and no change in AACID in contralateral tissue. There was also a small average increase in AACID in tumors within the three mice injected with DMSO only. The use of the natural compound quercetin in combination with pH weighted MRI represents a unique approach to cancer detection that does not require injection of an imaging contrast agent.

Fluid biphasic systems are one of the most interesting dynamic systems in chemistry and biochemistry. In nuclear magnetic resonance (NMR) spectroscopy, the study of the solute dynamics across fluid biphasic systems requires the introduction of dedicated NMR methods, due to their intrinsic heterogeneity. Diffusion and spatially resolved NMR techniques represent a useful approach for dealing with the study of solutes in biphasic systems and have been applied lately with success. Nevertheless, other potential applications of NMR spectroscopy for biphasic systems remain to be explored. In this proof of-concept communication, we specifically aimed to investigate whether solute exchange between two immiscible phases can be followed by NMR experiments involving transfer of magnetization. To that aim, we have used spatially resolved saturation transfer difference NMR (SR-STD NMR) experiments to analyze solute exchange by transfer of saturation from one phase to the other in a biphasic system and have explored which are the underlying mechanisms leading to the transfer of magnetization between phases and the limits of the approach. We hereby demonstrate that SR-STD NMR is feasible and that it might be implemented in pharmacological screening for binders of biological receptors or in the study of chemical and biochemical reactions occurring at interfaces.

Image-guided drug delivery is of great clinical interest. Here, we explored a direct way, namely CEST theranostics, to detect diamagnetic anticancer drugs simply through their inherent Chemical Exchange Saturation Transfer (CEST) MRI signal, and demonstrated its application in image-guided drug delivery of nanoparticulate chemotherapeutics. We first screened 22 chemotherapeutic agents and characterized the CEST properties of representative agents and natural analogs in three major categories, i.e., pyrimidine analogs, purine analogs, and antifolates, with respect to chemical structures. Utilizing the inherent CEST MRI signal of gemcitabine, a widely used anticancer drug, the tumor uptake of the i.v.-injected, drug-loaded liposomes was successfully detected in CT26 mouse tumors. Such label-free CEST MRI theranostics provides a new imaging means, potentially with an immediate clinical impact, to monitor the drug delivery in cancer.

Semisolid magnetization transfer contrast (MTC) and chemical exchange saturation transfer (CEST) MRI based on MT phenomenon have shown potential to evaluate brain development, neurological, psychiatric, and neurodegenerative diseases. However, a qualitative MT ratio (MTR) metric commonly used in conventional MTC imaging is limited in the assessment of quantitative semisolid macromolecular proton exchange rates and concentrations. In addition, CEST signals measured by MTR asymmetry analysis are unavoidably contaminated by upfield nuclear Overhauser enhancement (NOE) signals of mobile and semisolid macromolecules. To address these issues, we developed an MTC-MR fingerprinting (MTC-MRF) technique to quantify tissue parameters, which further allows an estimation of accurate MTC signals at a certain CEST frequency offset. A pseudorandomized RF saturation scheme was used to generate unique MTC signal evolutions for different tissues and a supervised deep neural network was designed to extract tissue properties from measured MTC-MRF signals. Through detailed Bloch equation-based digital phantom and in vivo studies, we demonstrated that the MTC-MRF can quantify MTC characteristics with high accuracy and computational efficiency, compared to a conventional Bloch equation fitting approach, and provide baseline reference signals for CEST and NOE imaging. For validation, MTC-MRF images were synthesized using the tissue parameters estimated from the deep-learning method and compared with experimentally acquired MTC-MRF images as the reference standard. The proposed MTC-MRF framework can provide quantitative 3D MTC, CEST, and NOE imaging of the human brain within a clinically acceptable scan time.

Detecting tissue pH in vivo is extremely vital for medical diagnosis and formulation of treatment decisions. To this end, many investigations have been carried out to develop an accurate and efficient method of in vivo pH measurement. Most of the techniques developed so far suffer from inadequate accuracy, due to poor sensitivity at low concentration of the target or nonspecific interactions within the tissue matrix. To overcome these issues, we describe herein the development of a simple, yet reliable, way to estimate pH with high precision using a Gd(III)-DOTA-silyl-based acid-labile group as a pH-sensitive contrast agent with Magnetic Resonance Imaging (MRI). With this method, a change in T 1 weighted image intensity of the newly developed pH-sensitive contrast is directly linked to the proton concentration in the media. As a result, we were able estimate the pH of the target with 95% reliability.

PURPOSE:To introduce a new pH-sensitive and oxygen-sensitive MRI technique using amine proton CEST echo spin-and-gradient echo (SAGE) EPI (CEST-SAGE-EPI). METHODS:pH-weighting was obtained using CEST estimations of magnetization transfer ratio asymmetry (MTRasym ) at 3 ppm, and oxygen-weighting was obtained using R2' measurements. Glutamine concentration, pH, and relaxation rates were varied in phantoms to validate simulations and estimate relaxation rates. The values of MTRasym and R2' in normal-appearing white matter, T2 hyperintensity, contrast enhancement, and macroscopic necrosis were measured in 47 gliomas. RESULTS:Simulation and phantom results confirmed an increase in MTRasym with decreasing pH. The CEST-SAGE-EPI estimates of R2 , R2*, and R2' varied linearly with gadolinium diethylenetriamine penta-acetic acid concentration (R2 = 6.2 mM-1 ·sec-1 and R2* = 6.9 mM-1 ·sec-1 ). The CEST-SAGE-EPI and Carr-Purcell-Meiboom-Gill estimates of R2 (R2 = 0.9943) and multi-echo gradient-echo estimates of R2* (R2 = 0.9727) were highly correlated. T2 lesions had lower R2' and higher MTRasym compared with normal-appearing white matter, suggesting lower hypoxia and high acidity, whereas contrast-enhancement tumor regions had elevated R2' and MTRasym , indicating high hypoxia and acidity. CONCLUSION:The CEST-SAGE-EPI technique provides simultaneous pH-sensitive and oxygen-sensitive image contrasts for evaluation of the brain tumor microenvironment. Advantages include fast whole-brain acquisition, in-line B0 correction, and simultaneous estimation of CEST effects, R2 , R2*, and R2' at 3 T.

To explore the capability of the amide-proton-transfer weighted (APTW) magnetic resonance imaging (MRI) in the evaluation of clinical neurological deficit at the time of hospitalization and assessment of long-term daily functional outcome for patients with acute ischemic stroke (AIS). We recruited 55 AIS patients with brain MRI acquired within 24–48 h of symptom onset and followed up with their 90-day modified Rankin Scale (mRS) score. APT weighted MRI was performed for all the study subjects to measure APTW signal quantitatively in the acute ischemic area (APTWipsi) and the contralateral side (APTWcont). Change of the APT signal between the acute ischemic region and the contralateral side (ΔAPTW) was calculated. Maximum APTW signal (APTWmax) and minimal APTW signal (APTWmin) were also acquired to demonstrate APTW signals heterogeneity (APTWmax−min). In addition, all the patients were divided into 2 groups according to their 90-day mRS score (good prognosis group with mRS score

Cerebral acidosis is a consequence of secondary injury mechanisms following traumatic brain injury (TBI), including excitotoxicity and ischemia, with potentially significant clinical implications. However, there remains an unmet clinical need for technology for non-invasive, high resolution pH imaging of human TBI for studying metabolic changes following injury. The current study examined 17 patients with TBI and 20 healthy controls using amine chemical exchange saturation transfer echoplanar imaging (CEST EPI), a novel pH-weighted molecular MR imaging technique, on a clinical 3T MR scanner. Results showed significantly elevated pH-weighted image contrast (MTRasym at 3 ppm) in areas of T2 hyperintensity or edema (P, Highlights • Cerebral acidosis is a consequence of traumatic brain injury (TBI) with significant clinical implications. • Amine CEST EPI is a novel pH-weighted molecular MR imaging technique for clinical 3T MR scanners. • Results showed significantly elevated pH-weighted image contrast (MTRasym at 3 ppm) in T2 hyperintense lesions. • This pH-weighted MR contrast correlated with Glasgow Coma Scale (GCS), 6 month clinical outcome, and time from initial injury to MRI exam.

PurposeThere is an increased interest to determine the exchange rate using CEST to provide pH information. However, current CEST quantification methods require lengthy scan times and do not address magnetization transfer effects. The purpose of this work was to apply the magnetic resonance fingerprinting (MRF) concept to CEST to achieve more efficient and accurate exchange rate quantification.MethodsThe proposed CEST fingerprinting method used varying saturation powers and saturation times to create unique signal evolutions for different exchange rates. The acquired signal was matched to a predefined dictionary to determine the exchange rate. The magnetization transfer effects were also addressed in the framework of CEST fingerprinting: The simulated dictionary could predict the signal curves without magnetization transfer effects, and comparing the dictionary to the acquired signals allowed the correction of the magnetization transfer effects. The CEST fingerprinting method was compared with the conventional pulsed quantitative CEST method using omega plots in the creatine phantom study.ResultsThe CEST fingerprinting method has a significantly reduced scan time (10 minutes versus 50 minutes) while providing more accurate exchange rate quantification using literature values as the reference.ConclusionIn this study, we demonstrate that CEST fingerprinting is more efficient (5 times faster) compared with pulsed quantitative CEST. It is also shown that the results of the proposed CEST fingerprinting technique are much closer to the literature values than pulsed quantitative CEST at 3 T.

A novel MRI contrast is proposed which enables the selective detection of endogenous bulk mobile proteins in vivo. Such a non-invasive imaging technique may be of particular interest for many diseases associated with pathological alterations of protein expression, such as cancer and neurodegenerative disorders. Specificity to mobile proteins was achieved by the selective measurement of intramolecular spin diffusion and the removal of semi-solid macromolecular signal components by a correction procedure. For this purpose, the approach of chemical exchange saturation transfer (CEST) was extended to a radiofrequency (RF) irradiation scheme at two different frequency offsets (dualCEST). Using protein model solutions, it was demonstrated that the dualCEST technique allows the calculation of an image contrast which is exclusively sensitive to changes in concentration, molecular size and the folding state of mobile proteins. With respect to application in humans, dualCEST overcomes the selectivity limitations at relatively low magnetic field strengths, and thus enables examinations on clinical MR scanners. The feasibility of dualCEST examinations in humans was verified by a proof-of-principle examination of a brain tumor patient at 3 T. With its specificity for the mobile fraction of the proteome, its comparable sensitivity to conventional water proton MRI and its applicability to clinical MR scanners, this technique represents a further step towards the non-invasive imaging of proteomic changes in humans.

In glioma, the acidification of the extracellular tumor microenvironment drives proliferation, angiogenesis, immunosuppression, invasion and chemoresistance. Therefore, quantification of glioma extracellular pH (pHe) is of crucial importance. This study is focused on the application of the YbHPDO3A (ytterbium 1,4,7-triscarboxymethyl-1,4,7,10-tetraazacyclododecane) probe for in vivo glioma pHe quantification using chemical exchange saturation transfer (CEST)-MRI and its correlation with tumor metabolism assessed by immunohistochemistry. The U87 glioma mouse model was used (n = 18) and MRI performed at 4.7 T. CEST-MRI of YbHPDO3A solutions at different pH values showed two resolved CEST spectra at 71 ppm and 99 ppm, both sensitive to pH variations, allowing therefore calculation of the ratiometric curve for in vivo pH quantification. In vivo MRI sequences consisted of T2w for tumor localization, T2w * to assess YbHPDO3A biodistribution by exploiting its magnetic susceptibility effect and CEST for glioma pHe mapping. T2w * images show that YbHPDO3A extravasates in tumor in regions with damaged blood-brain barrier. The pHe is calculated only in these regions. Hematoxylin/eosin histology and Ki-67, CA-IX (carbonic anhydrase 9) and NHE-1 immunohistochemical staining were performed; their expression rates were compared with the in vivo pHe values. On the basis of the cell proliferation marker Ki-67, two groups were defined: one group with a lower mitotic index (MI% 20% and an acidic pHe of 6.6 (high-proliferation/low-pH group). CA-IX and NHE-1 were over-expressed in the high-proliferation/low-pH group (CA-IX, 92 ± 7% versus 30 ± 13%; NHE-1, 84 ± 8% versus 35 ± 11%), indicating an acidic/hypoxic microenvironment. These immunohistochemical results are consistent with our pHe mapping (Pearson correlation coefficient > 0.70) and provide evidence for the feasibility of the CEST-MRI method with the YbHPDO3A probe for glioma pHe quantification at 4.7 T. Importantly, the YbHPDO3A probe has similar chemical and biological properties to the clinically approved MRI contrast agent GdHPDO3A. This makes the method promising for a clinical translation.

Intracellular pH (pHi) plays an important role in the maintenance of normal cell function, and is maintained within a narrow range by the activity of transporters located at the plasma membrane. Modulation of tumor pHi may influence proliferation, apoptosis, chemotherapy resistance, and thermosensitivity. Chemical exchange saturation transfer (CEST) is a novel MRI contrast mechanism that is dependent on cellular pH. Amine and amide concentration-independent detection (AACID) is a recently developed CEST contrast method that is intracellular pH (pHi) weighted. Dichloroacetate (DCA) can alter tumor pHi by inhibiting the enzyme pyruvate dehydrogenase kinase causing reduced lactate (increasing pHi), or by decreasing the expression of monocarboxylate transporters and vacuolar ATPase leading to reduced pHi. Since the net in vivo effect of DCA on pHi is difficult to predict, the purpose of this study was to quantify the magnitude of acute pHi change in glioblastoma after a single DCA injection using AACID CEST MRI. Using a 9.4T MRI scanner, CEST spectra were acquired in six mice approximately 14 days after implanting 105 U87 human glioblastoma multiforme (GBM) cells in the brain, before and after intravenous injection of DCA (dose: 200 mg/kg). Three additional mice received only phosphate buffered saline (PBS) injection and were studied as controls. Repeated measures t test was used to compare AACID changes in tumor and contralateral tissue regions of interest. One hour after DCA injection there was a significant increase in tumor AACID level by 0.04 ± 0.01 corresponding to a 0.16 decrease in pHi, and no change in AACID in contralateral tissue. Inspection of AACID maps following PBS injection showed no differences. The use of DCA to induce a tumor specific pH change detectable by AACID CEST MRI is consistent with previous studies that have shown similar effects for lonidamine and topiramate. This study demonstrates that a single dose of DCA can be used as a pharmacological challenge to induced rapid tumor intracellular acidification.

Several molecules have been exploited for developing MRI pH sensors based on the chemical exchange saturation transfer (CEST) technique. A ratiometric approach, based on the saturation of two exchanging pools at the same saturation power, or by varying the saturation power levels on the same pool, is usually needed to rule out the concentration term from the pH measurement. However, all these methods have been demonstrated by using a continuous wave saturation scheme that limits its translation to clinical scanners. This study shows a new ratiometric CEST-MRI pH-mapping approach based on a pulsed CEST saturation scheme for a radiographic contrast agent (iodixanol) possessing a single chemical exchange site. This approach is based on the ratio of the CEST contrast effects at two different flip angles combinations (180 degrees/360 degrees and 180 degrees/720 degrees), keeping constant the mean irradiation RF power (B-avg power). The proposed ratiometric approach index is concentration independent and it showed good pH sensitivity and accuracy in the physiological range between 6.0 and 7.4. (C) 2017 Elsevier Inc. All rights reserved.

Purpose Magnetic resonance imaging has been used extensively to track in vivo implanted cells that have been previously labeled with relaxation enhancers. However, this approach is not suitable to track multiple cell populations, as it may lead to confounding results in case the contrast agent is released from the labeled cells. This paper demonstrates how the use of CEST agents can overcome these issues. After encapsulating paramagnetic lanthanide shift reagents, we may shift the absorption frequency of the intracellular water resonance (δIn ), thus generating frequency-encoding CEST responsive cells that can be visualized in the MR image by applying the proper RF irradiation. Methods Eu-HPDO3A, Dy-HPDO3A, and Tm-HPDO3A were used as shift reagents for labeling murine breast cancer cells and murine macrophages by hypotonic swelling and pinocytosis. The CEST-MR images were acquired at 7 T, and the saturation transfer effect was measured. Samples at different dilution of cells were analyzed to quantify the detection threshold. In vitro experiments of cell proliferation were carried out. Finally, murine breast cancer cells were injected subcutaneously in mice, and MR images were acquired to assess the proliferation index in vivo. Results It was found that entrapment of the paramagnetic complexes into endosomes (i.e., using the pinocytosis route) leads to an enhanced shift of the intracellular water resonance. δIn appears to be proportional to the effective magnetic moment (μeff ) and to the concentration of the loaded lanthanide complex. Moreover, a higher shift is present when the complexes are entrapped in the endosomes. The cell proliferation index was assessed both in vitro and in vivo by evaluating the reduction of δIn value in the days after the cell labeling. Conclusion Cells can be visualized by CEST MRI after loading with paramagnetic shift reagent, by exploiting the large ensemble of the properly shifted intracellular water molecules. A better performance is obtained when the complexes are entrapped inside the endosomes. The observed (δIn ) value is strongly correlated to the chemical nature of the probe, and to its concentration and cellular localization. Two applications of this method are reported in this paper: (1) for in vivo cell visualization and (2) for the monitoring of the cellular proliferation process, as this method is accompanied by a change in δIn that may be exploited as a longitudinal reporter of the proliferation rate.

Purpose To detect neuronal activity–evoked pH changes by amide proton transfer–chemical exchange saturation transfer (APT‐CEST) MRI at 7 T. Methods Three healthy subjects participated in the study. A low‐power 3‐dimensional APT‐CEST sequence was optimized through the Bloch‐McConnell equations. pH sensitivity of the sequence was estimated both in phantoms and in vivo. The feasibility of pH–functional MRI was tested in Bloch‐McConnell‐simulated data using the optimized sequence. In healthy subjects, the visual stimuli were used to evoke transient pH changes in the visual cortex, and a 3‐dimensional APT‐CEST volume was acquired at the pH‐sensitive frequency offset of 3.5 ppm every 12.6 s. Results In theory, a three‐component general linear model was capable of separating the effects of blood oxygenation level–dependent contrast and pH. The Bloch‐McConnell equations indicated that a change in pH of 0.03 should be measurable at the experimentally determined temporal signal‐to‐noise ratio of 108. However, only a blood oxygenation level–dependent effect in the visual cortex could be discerned during the visual stimuli experiments performed in the healthy subjects. Conclusions The results of this study suggest that if indeed there are any transient brain pH changes in response to visual stimuli, those are under 0.03 units pH change, which is extremely difficult to detect using the existent techniques. Magn Reson Med 80:126–136, 2018. © 2017 The Authors Magnetic Resonance in Medicine published by Wiley Periodicals, Inc. on behalf of International Society for Magnetic Resonance in Medicine. This is an open access article under the terms of the Creative Commons Attribution NonCommercial License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited and is not used for commercial purposes.

Background Image contrast enhanced by exogenous contrast agents plays a crucial role in the early detection, characterization, and determination of the precise location of cancers. Here, we investigate the feasibility of using a non-nutritive sweetener, sucralose (commercial name, Splenda), as magnetic resonance imaging (MRI) contrast agent for cancer studies. Methods High-resolution nuclear-magnetic-resonance spectroscopy and MR studies on sucralose solution phantom were performed to detect the chemical exchange saturation transfer (CEST) property of sucralose hydroxyl protons with bulk water (sucCEST). For the animal experiments, female Fisher rats (F344/NCR) were used to generate 9L-gliosarcoma model. MRI with CEST experiments were performed on anesthetized rats at 9.4 T MR scanner. Following the baseline CEST scans, sucralose solution was intravenously administered in control and tumor bearing rats. CEST acquisitions were continued during and following the administration of sucralose. Following the sucCEST, Gadolinium-diethylenetriamine pentaacetic acid was injected to perform Gd-enhanced imaging for visualizing the tumor. Results The sucCEST contrast in vitro was found to correlate positively with the sucralose concentration and negatively with the pH, indicating the potential of this technique in cancer imaging. In a control animal, the CEST contrast from the brain was found to be unaffected following the administration of sucralose, demonstrating its blood–brain barrier impermeability. In a 9L glioma model, enhanced localized sucCEST contrast in the tumor region was detected while the unaffected brain region showed unaltered CEST effect implying the specificity of sucralose toward the tumorous tissue. The CEST asymmetry plots acquired from the tumor region before and after the sucralose infusion showed elevation of asymmetry at 1 ppm, pointing towards the role of sucralose in increased contrast. Conclusions We show the feasibility of using sucralose and sucCEST in study of preclinical models of cancer. This study paves the way for the potential development of sucralose and other sucrose derivatives as contrast agents for clinical MRI applications. Electronic supplementary material The online version of this article (doi:10.1186/s12967-017-1221-9) contains supplementary material, which is available to authorized users.

We present MRiLab, a new comprehensive simulator for large-scale realistic MRI simulations on a regular PC equipped with a modern graphical processing unit (GPU). MRiLab combines realistic tissue modeling with numerical virtualization of an MRI system and scanning experiment to enable assessment of a broad range of MRI approaches including advanced quantitative MRI methods inferring microstructure on a sub-voxel level. A flexible representation of tissue microstructure is achieved in MRiLab by employing the generalized tissue model with multiple exchanging water and macromolecular proton pools rather than a system of independent proton isochromats typically used in previous simulators. The computational power needed for simulation of the biologically relevant tissue models in large 3D objects is gained using parallelized execution on GPU. Three simulated and one actual MRI experiments were performed to demonstrate the ability of the new simulator to accommodate a wide variety of voxel composition scenarios and demonstrate detrimental effects of simplified treatment of tissue micro-organization adapted in previous simulators. GPU execution allowed ∼ 200× improvement in computational speed over standard CPU. As a cross-platform, open-source, extensible environment for customizing virtual MRI experiments, MRiLab streamlines the development of new MRI methods, especially those aiming to infer quantitatively tissue composition and microstructure.

Uptake of administered D-glucose (Glc) or 2-deoxy-D-glucose (2DG) has been indirectly mapped through the chemical exchange (CE) between glucose hydroxyl and water protons using CE-dependent saturation transfer (glucoCEST) magnetic resonance imaging (MRI). We propose an alternative technique—on-resonance CE-sensitive spin-lock (CESL) MRI—to enhance responses to glucose changes. Phantom data and simulations suggest higher sensitivity for this ‘glucoCESL’ technique (versus glucoCEST) in the intermediate CE regime relevant to glucose. Simulations of CESL signals also show insensitivity to B0-fluctuations. Several findings are apparent from in vivo glucoCESL studies of rat brain at 9.4 Tesla with intravenous injections. First, dose-dependent responses are nearly linearly for 0.25-, 0.5-, and 1-g/kg Glc administration (obtained with 12-second temporal resolution), with changes robustly detected for all doses. Second, responses at a matched dose of 1 g/kg are much larger and persist for a longer duration for 2DG versus Glc administration, and are minimal for mannitol as an osmolality control. And third, with similar increases in steady-state blood glucose levels, glucoCESL responses are ~2.2 times higher for 2DG versus Glc, consistent with their different metabolic properties. Overall, we show that glucoCESL MRI could be a highly sensitive and quantifiable tool for glucose transport and metabolism studies.

International audience; Sparsely populated and transiently formed protein conformers can play key roles in many biochemical processes. Understanding the structure function paradigm requires, therefore, an atomic-resolution description of these rare states. However, they are difficult to study because they cannot be observed using standard biophysical techniques. In the past decade, NMR methods have been developed for structural studies of these elusive conformers, focusing primarily on backbone H-1, N-15 and C-13 nuclei. Here we extend the methodology to include side chains by developing a C-13-based chemical exchange saturation transfer experiment for the assignment of side-chain aliphatic C-13 chemical shifts in uniformly C-13 labeled proteins. A pair of applications is provided, involving the folding of beta-sheet Fyn SH3 and alpha-helical FF domains. Over 96% and 89% of the side-chain C-13 chemical shifts for excited states corresponding to the unfolded conformation of the Fyn SH3 domain and a folding intermediate of the FF domain, respectively, have been obtained, providing insight into side-chain packing and dynamics.

Reversal of the intracellular/extracellular pH gradient is a hallmark of malignant tumors and is an important consideration in evaluating tumor growth potential and the effectiveness of anticancer therapies. Glioblastoma multiforme (GBM) brain tumors have increased expression of the carbonic anhydrase (CA) isozymes CAII, CAIX and CAXII that contribute to the altered regulation of intracellular pH (pHi). The anti-epileptic drug topiramate (TPM) inhibits CA action and may acidify the tumor intracellular compartment. In-vivo detection of acute tumor acidification could aid in cancer diagnosis and monitoring treatment response. Chemical exchange saturation transfer (CEST) magnetic resonance imaging (MRI) has been used to measure tissue pH. Using a recently developed CEST-MRI method called amine/amide concentration independent detection (AACID), we have previously shown intracellular acidification caused by single dose of lonidamine. The current study aims to evaluate the intracellular acidification induced by a single dose of the clinically approved drug TPM. Brain tumors were induced in NU/NU mice by injecting 105 U87 human glioblastoma multiforme cells into the right frontal lobe. Using a 9.4T MRI scanner AACID measurements were acquired, before and after administration of TPM (dose: 120 mg/kg, intraperitoneal), 15 ± 2 days after tumor cell implantation. TPM administration induced acute intracellular acidification (average ± SD: baseline AACID = 1.14 ± 0.05; post AACID = 1.19 ± 0.05, paired ttest p = 0.02) in implanted brain tumors. In contrast, contralateral tissue showed no change in AACID value. These results suggest that topiramate can rapidly induce a tumor specific physiological change detectable by AACID CEST. This pH challenge paradigm could be exploited to aid in tumor detection and monitoring treatment response.

Three different 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetamide (DOTAM) derivatives bearing as amide N-substituents phenyl, p-methoxyphenyl and p-ethylbenzoate groups were synthesized and the 1H and 17O NMR relaxometric behaviour of the Gd(III)-chelates and chemical exchange saturation transfer (CEST) effect of the Eu(III) complexes were evaluated. The electronic properties of the substituents were shown to strongly influence the coordinated water exchange rate (kex), resulting in five times faster kex for the electron donating phenylmethoxy group compared to the electron withdrawing ethylbenzoate group.

Acidity within the extracellular milieu is a hallmark of cancer. There is a current need for fast, high spatial resolution pH imaging techniques for clinical evaluation of cancers, including gliomas. Chemical exchange saturation transfer (CEST) MRI targeting fast-exchanging amine protons can be used to obtain high-resolution pH-weighted images, but conventional CEST acquisition strategies are slow. There is also a need for more accurate MR simulations to better understand the effects of amine CEST pulse sequence parameters on pH-weighted image contrast. In the current study we present a simulation of amine CEST contrast specific for a newly developed CEST echoplanar imaging (EPI) pulse sequence. The accuracy of the simulations was validated by comparing the exchange rates and Z-spectrum under a variety of conditions using physical phantoms of glutamine with different pH values. The effects of saturation pulse shapes, pulse durations, pulse train lengths, repetition times, and relaxation rates of bulk water and exchangeable amine protons on the CEST signal were explored for normal-appearing white matter (NAWM), glioma, and cerebrospinal fluid. Last, 18 patients with WHO II-IV gliomas were evaluated. Results showed that the Z-spectrum was highly dependent on saturation pulse shape, repetition time, saturation amplitude, magnetic field strength, and T2 within bulk water; however, the Z-spectrum was only minimally influenced by saturation pulse duration and the specific relaxation rates of amine protons. Results suggest that a Gaussian saturation pulse train consisting of 3×100ms pulses using the minimum allowable repetition time is optimal for achieving over 90% available contrast across all tissues. Results also demonstrate that high saturation pulse amplitude and scanner field strength (>3 T) are necessary for adequate endogenous pH-weighted amine CEST contrast. pH-weighted amine CEST contrast increased with increasing tumor grade, with glioblastoma showing significantly higher contrast compared with WHO II or III gliomas.

Two relatively new types of exogenous magnetic resonance imaging contrast agents may provide greater impact for molecular imaging by providing greater specificity for detecting molecular imaging biomarkers. Exogenous chemical exchange saturation transfer (CEST) agents rely on the selective saturation of the magnetization of a proton on an agent, followed by chemical exchange of a proton from the agent to water. The selective detection of a biomarker-responsive CEST signal and an unresponsive CEST signal, followed by the ratiometric comparison of these signals, can improve biomarker specificity. We refer to this improvement as a “double-agent” approach to molecular imaging. Exogenous T2-exchange agents also rely on chemical exchange of protons between the agent and water, especially with an intermediate rate that lies between the slow exchange rates of CEST agents and the fast exchange rates of traditional T1 and T2 agents. Because of this intermediate exchange rate, these agents have been relatively unknown and have acted as “secret agents” in the contrast agent research field. This review exposes these secret agents and describes the merits of double agents through examples of exogenous agents that detect enzyme activity, nucleic acids and gene expression, metabolites, ions, redox state, temperature, and pH. Future directions are also provided for improving both types of contrast agents for improved molecular imaging and clinical translation. Therefore, this review provides an overview of two new types of exogenous contrast agents that are becoming useful tools within the armamentarium of molecular imaging.

The vast majority of cancers exhibit increased glucose uptake and glycolysis regardless of oxygen availability. This metabolic shift leads to an enhanced production of lactic acid that decreases extracellular pH (pHe), a hallmark of the tumor microenvironment. In this way, dysregulated tumor pHe and upregulated glucose metabolism are linked tightly and their relative assessment may be useful to gain understanding of the underlying biology. Here we investigated noninvasively the in vivo correlation between tumor 18F-FDG uptake and extracellular pH values in a murine model of HER2+ breast cancer. Tumor extracellular pH and perfusion were assessed by acquiring MRI-CEST (chemical exchange saturation transfer) images on a 3T scanner after intravenous administration of a pH-responsive contrast agent (iopamidol). Static PET images were recorded immediately after MRI acquisitions to quantify the extent of 18F-FDG uptake. We demonstrated the occurrence of tumor pHe changes that report on acidification of the interstitial fluid caused by an accelerated glycolysis. Combined PET and MRI-CEST images reported complementary spatial information of the altered glucose metabolism. Notably, a significant inverse correlation was found between extracellular tumor pH and 18F-FDG uptake, as a high 18F-FDG uptake corresponds to lower extracellular pH values. These results show how merging the information from 18F-FDG-uptake and extracellular pH measurements can improve characterization of the tumor microenvironment. Cancer Res; 76(22); 6463–70. ©2016 AACR.

X-ray iodinated contrast media have been shown to generate contrast in MR images when used with the chemical exchange saturation transfer (CEST) approach. The aim of this study is to compare contrast enhancement (CE) capabilities and perfusion estimates between radiographic molecules and a Gd-based contrast agent in two tumour murine models with different vascularization patterns. MRI-CEST and MRI-CE T1w images were acquired in murine TS/A and 4 T1 breast tumours upon sequential i.v. injection of iodinated contrast media (iodixanol, iohexol, and iopamidol) and of gadoteridol. The signal enhancements observed in the two acquisition modalities were evaluated using Pearson’s correlation, and the correspondence in the spatial distribution was assessed by a voxelwise comparison. A significant, positive correlation was observed between iodinated contrast media and gadoteridol for tumour contrast enhancement and perfusion values for both tumour models (r = 0.51-0.62). High spatial correlations were observed in perfusion maps between iodinated molecules and gadoteridol (r = 0.68-0.86). Tumour parametric maps derived by iodinated contrast media and gadoteridol showed high spatial similarities. A good to strong spatial correlation between tumour perfusion parameters derived from MRI-CEST and MRI-CE modalities indicates that the two procedures provide similar information. • Gd-based agents are the standard of reference for contrast-enhanced MRI. • Iodinated contrast media provides MRI-CEST contrast enhancement in animal tumour models. • Contrast enhancements were positively correlated between iodinated agents and gadoteridol. • Tumour perfusion map showed similar spatial distribution between iodinated agents and gadoteridol. • MRI-CEST with iodinated agents provide similar information to gadoteridol.

The purpose of this study was to develop realistic phantom models of the intracellular environment of metastatic breast tumour and naïve brain, and using these models determine an analysis metric for quantification of CEST MRI data that is sensitive to only labile proton exchange rate and concentration. The ability of the optimal metric to quantify pH differences in the phantoms was also evaluated. Novel phantom models were produced, by adding perchloric acid extracts of either metastatic mouse breast carcinoma cells or healthy mouse brain to bovine serum albumin. The phantom model was validated using 1H NMR spectroscopy, then utilized to determine the sensitivity of CEST MRI to changes in pH, labile proton concentration, T 1 time and T 2 time; six different CEST MRI analysis metrics (MTRasym, APT*, MTRRex, AREX and CESTR* with and without T 1/T 2 compensation) were compared. The new phantom models were highly representative of the in vivo intracellular environment of both tumour and brain tissue. Of the analysis methods compared, CESTR* with T 1 and T 2 time compensation was optimally specific to changes in the CEST effect (i.e. minimal contamination from T 1 or T 2 variation). In phantoms with identical protein concentrations, pH differences between phantoms could be quantified with a mean accuracy of 0.6 pH units. We propose that CESTR* with T 1 and T 2 time compensation is the optimal analysis method for these phantoms. Analysis of CEST MRI data with T 1/T 2 time compensated CESTR* is reproducible between phantoms, and its application in vivo may resolve the intracellular alkalosis associated with breast cancer brain metastases without the need for exogenous contrast agents.

Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disease involving cytotoxic conformations of Cu, Zn superoxide dismutase (SOD1). A major challenge in understanding ALS disease pathology has been the identification and atomic-level characterization of these conformers. Here, we use a combination of NMR methods to detect four distinct sparsely populated and transiently formed thermally accessible conformers in equilibrium with the native state of immature SOD1 (apoSOD12SH). Structural models of two of these establish that they possess features present in the mature dimeric protein. In contrast, the other two are non-native oligomers in which the native dimer interface and the electrostatic loop mediate the formation of aberrant intermolecular interactions. Our results show that apoSOD12SH has a rugged free energy landscape that codes for distinct kinetic pathways leading to either maturation or non-native association and provide a starting point for a detailed atomic-level understanding of the mechanisms of SOD1 oligomerization. DOI: http://dx.doi.org/10.7554/eLife.07296.001, eLife digest Amyotrophic lateral sclerosis (or ALS) is a disease that affects the nerve cells in the brain and spinal cord, such that more and more of these cells die as the disease progresses. Since nerve cells direct muscle movement, people with ALS increasingly lose control of their muscles, and may therefore lose the ability to speak, eat, move, and breathe. Around one in five people who have an inherited form of ALS also have mutations in a gene that encodes an enzyme called SOD1. This antioxidant enzyme detoxifies harmful chemicals that are the byproducts of normal cellular activity. The mature active form of this enzyme contains two SOD1 proteins, each of which binds one copper ion and one zinc ion. Each protein in a pair also contains a strong bond that helps to stabilize its three-dimensional structure. However, immature copies of this protein, which lack the strong bond and metal ions, often fold into the wrong shape. These misfolded proteins can clump together into clusters, and potentially lead to the development of ALS. Efforts to study misfolded proteins are limited by the fact that misfolding is a rare event. As a result, protein samples generally contain very small fractions of misfolded molecules and most techniques for investigating protein structure are ill-suited to probe the process of misfolding. Nuclear magnetic resonance spectroscopy (or NMR for short) is one technique that has been used to visualize flexible proteins, and protein folding and misfolding, down to the level of individual atoms. Recent improvements in NMR spectroscopy have opened up the possibility of studying protein structures that exist at levels as low as 1% of the sample. Now, Sekhar et al. have used NMR to document the dynamics of immature SOD1 proteins. The experiments show that the protein is flexible and readily switches back and forth between its ‘default’ shape and one of at least four different shapes; and at any one time, the fraction of molecules in solution that were in each of these shapes was around 1%. Two of the shapes have many of the same features as those in a mature SOD1 protein pair. But the other two shapes involve SOD1 proteins interacting in unusual ways, and may resemble the cluster-forming misfolded SOD1 proteins. By using the NMR data to create three-dimensional models of the SOD1 proteins, Sekhar et al. were able to identify two sites on immature SOD1 proteins involved in these unusual interactions that are inaccessible in mature enzymatically active SOD1. These findings suggest that targeting these two sites with drugs could possibly block the formation of toxic versions of SOD1 early in the course of ALS, and thus prevent the progression of the disease. DOI: http://dx.doi.org/10.7554/eLife.07296.002