Real-time detection of the rates of metabolic flux, or exchange rates of endogenous enzymatic reactions, is now feasible in biological systems using Dynamic Nuclear Polarization Magnetic Resonance. Derivation of reaction rate kinetics from this technique typically requires multi-compartmental modeling of dynamic data, and results are therefore model-dependent and prone to misinterpretation. We present a model-free formulism based on the ratio of total areas under the curve (AUC) of the injected and product metabolite, for example pyruvate and lactate. A theoretical framework to support this novel analysis approach is described, and demonstrates that the AUC ratio is proportional to the forward rate constant k. We show that the model-free approach strongly correlates with k for whole cell in vitro experiments across a range of cancer cell lines, and detects response in cells treated with the pan-class I PI3K inhibitor GDC-0941 with comparable or greater sensitivity. The same result is seen in vivo with tumor xenograft-bearing mice, in control tumors and following drug treatment with dichloroacetate. An important finding is that the area under the curve is independent of both the input function and of any other metabolic pathways arising from the injected metabolite. This model-free approach provides a robust and clinically relevant alternative to kinetic model-based rate measurements in the clinical translation of hyperpolarized 13C metabolic imaging in humans, where measurement of the input function can be problematic.

Using low temperature dynamic nuclear polarisation (DNP) in conjunction with dissolution makes it possible to generate highly polarised nuclear spin systems for liquid state applications of nuclear magnetic resonance spectroscopy. However, in its current implementation, which requires the transfer of the solute between two different magnets, the hyperpolarisation strategy is limited to spin systems with relatively long longitudinal relaxation time constants. Here we describe the design and construction of a dedicated spectrometer for DNP applications that is based on a magnet with two isocentres. DNP enhancement is carried out in the upper compartment of this magnet in a low temperature environment at 3.35 T, while a 9.4 T isocentre in the lower compartment is used for high resolution NMR spectroscopy. The close proximity (85 cm) of the two isocentres makes it possible to transfer the sample in the solid state with very little loss of spin polarisation. In first performance tests this novel experimental set-up proved to be superior to the strategy involving two separate magnets.

Background:Dichloroacetate (DCA) has been found to have antitumour properties.Methods:We investigated the cellular and metabolic responses to DCA treatment and recovery in human colorectal (HT29, HCT116 WT and HCT116 Bax-ko), prostate carcinoma cells (PC3) and HT29 xenografts by flow cytometry, western blotting, electron microscopy, 1H and hyperpolarised 13C-magnetic resonance spectroscopy.Results:Increased expression of the autophagy markers LC3B II was observed following DCA treatment both in vitro and in vivo. We observed increased production of reactive oxygen species (ROS) and mTOR inhibition (decreased pS6 ribosomal protein and p4E-BP1 expression) as well as increased expression of MCT1 following DCA treatment. Steady-state lactate excretion and the apparent hyperpolarised [1-13C] pyruvate-to-lactate exchange rate (kPL) were decreased in DCA-treated cells, along with increased NAD+/NADH ratios and NAD+. Steady-state lactate excretion and kPL returned to, or exceeded, control levels in cells recovered from DCA treatment, accompanied by increased NAD+ and NADH. Reduced kPL with DCA treatment was found in HT29 tumour xenografts in vivo.Conclusions:DCA induces autophagy in cancer cells accompanied by ROS production and mTOR inhibition, reduced lactate excretion, reduced kPL and increased NAD+/NADH ratio. The observed cellular and metabolic changes recover on cessation of treatment.

The objective of this study was to assess the predictive value of early assessment (after 1 cycle of induction chemotherapy [IC]) with 18 F-FDG PET/CT and diffusion-weighted (DW) MRI for subsequent response to radical chemoradiotherapy in locally advanced head and neck squamous cell carcinoma (HNSCC). Methods: Twenty patients with stage III-IVa HNSCC prospectively underwent 18 F-FDG PET/CT and DW MRI before and 2 wk after each cycle of IC (first cycle, IC1; second cycle, IC2). Response was assessed 3 mo after completion of chemoradiotherapy with clinical examination, MRI, and 18 F-FDG PET/CT. Patients with persistent disease were classed as nonresponders. Changes in functional and molecular imaging parameters after IC1 were compared between responders and nonresponders with the Mann-Whitney U test. The significance threshold was set at a P value of less than 0.05. Results: Responders showed a significantly greater reduction in metabolic tumor volume (P 5 0.03) and total lesion glycolysis (P 5 0.04) after IC1 than nonresponders. Responders also showed a tendency toward a larger but statistically nonsignificant increase in apparent diffusion coefficient after IC1. There was no significant difference in the changes from baseline between the IC1 and IC2 for all functional and molecular imaging parameters, indicating that most biologic response to IC measured by 18 F-FDG PET/CT and DW MRI was observed early after the first cycle of IC. Conclusion: Our preliminary data indicate that the 18 F-FDG PET/CT-derived metabolic tumor volume or total lesion glycolysis, acquired after IC1, are early predictive biomarkers for ultimate response to subsequent chemoradiotherapy. These early biomarkers enable identification of patients at risk of treatment failure at an early time point, permitting treatment individualization and consideration of alternative strategies such as radiotherapy dose escalation or surgery.

Head and neck cancer (HNC) is a challenging tumour site for radiotherapy delivery owing to its complex anatomy and proximity to organs at risk (OARs) such as the spinal cord and optic apparatus. Despite significant advances in radiotherapy planning techniques, radiation-induced morbidities remain substantial. Further improvement would require high-quality imaging and tailored radiotherapy based on intratreatment response. For these reasons, the use of MRI in radiotherapy planning for HNC is rapidly gaining popularity. MRI provides superior soft-tissue contrast in comparison with CT, allowing better definition of the tumour and OARs. The lack of additional radiation exposure is another attractive feature for intratreatment monitoring. In addition, advanced MRI techniques such as diffusion-weighted, dynamic contrast-enhanced and intrinsic susceptibility-weighted MRI techniques are capable of characterizing tumour biology further by providing quantitative functional parameters such as tissue cellularity, vascular permeability/perfusion and hypoxia. These functional parameters are known to have radiobiological relevance, which potentially could guide treatment adaptation based on their changes prior to or during radiotherapy. In this article, we first present an overview of the applications of anatomical MRI sequences in head and neck radiotherapy, followed by the potentials and limitations of functional MRI sequences in personalizing therapy. INTRODUCTIONRadical radiotherapy (RT) is integral to the management of head and neck cancer (HNC) in both the primary and adjuvant settings. Advances in computer-assisted radiological techniques over the past two decades have in turn revolutionized radiotherapy planning. Development of advanced radiotherapy planning techniques such as intensity-modulated radiotherapy (IMRT) and volumetricmodulated arc therapy have allowed for better dose conformation to the tumour target and sparing of surrounding normal tissues. HNC was one of the first tumour sites where IMRT was widely implemented owing to a significant reduction in radiation-induced xerostomia in comparison with three-dimensional conformal planning.

ObjectiveTo assess the optimal timing and predictive value of early intra-treatment changes in multimodality functional and molecular imaging (FMI) parameters as biomarkers for clinical remission in patients receiving chemoradiation for head and neck squamous cell carcinoma (HNSCC).MethodsThirty-five patients with stage III-IVb (AJCC 7th edition) HNSCC prospectively underwent 18F–FDG-PET/CT, and diffusion-weighted (DW), dynamic contrast-enhanced (DCE) and susceptibility-weighted MRI at baseline, week 1 and week 2 of chemoradiation. Patients with evidence of persistent or recurrent disease during follow-up were classed as non-responders. Changes in FMI parameters at week 1 and week 2 were compared between responders and non-responders with the Mann–Whitney U test. The significance threshold was set at a p value of <0.05.ResultsThere were 27 responders and 8 non-responders. Responders showed a greater reduction in PET-derived tumor total lesion glycolysis (TLG40%; p = 0.007) and maximum standardized uptake value (SUVmax; p = 0.034) after week 1 than non-responders but these differences were absent by week 2. In contrast, it was not until week 2 that MRI-derived parameters were able to discriminate between the two groups: larger fractional increases in primary tumor apparent diffusion coefficient (ADC; p < 0.001), volume transfer constant (Ktrans; p = 0.012) and interstitial space volume fraction (Ve; p = 0.047) were observed in responders versus non-responders. ADC was the most powerful predictor (∆ >17%, AUC 0.937).ConclusionEarly intra-treatment changes in FDG-PET, DW and DCE MRI-derived parameters are predictive of ultimate response to chemoradiation in HNSCC. However, the optimal timing for assessment with FDG-PET parameters (week 1) differed from MRI parameters (week 2). This highlighted the importance of scanning time points for the design of FMI risk-stratified interventional studies.Electronic supplementary materialThe online version of this article (10.1007/s00259-017-3890-2) contains supplementary material, which is available to authorized users.

Purpose: To evaluate intrinsic susceptibility (IS) MRI for the identification of cycling hypoxia, and the assessment of its extent and spatial distribution, in head and neck squamous cell carcinoma (HNSCC) xenografts and patients.Experimental Design: Quantitation of the transverse relaxation rate, R 2 Ã , which is sensitive to paramagnetic deoxyhemoglobin, using serial IS-MRI acquisitions, was used to monitor temporal oscillations in levels of paramagnetic deoxyhemoglobin in human CAL R xenografts and patients with HNSCC at 3T. Autocovariance and power spectrum analysis of variations in R 2 Ã was performed for each imaged voxel, to assess statistical significance and frequencies of cycling changes in tumor blood oxygenation. Pathologic correlates with tumor perfusion (Hoechst 33342), hypoxia (pimonidazole), and vascular density (CD31) were sought in the xenografts, and dynamic contrast-enhanced (DCE) MRI was used to assess patient tumor vascularization. The prevalence of fluctuations within patient tumors, DCE parameters, and treatment outcome were reported.Results: Spontaneous R 2 Ã fluctuations with a median periodicity of 15 minutes were detected in both xenografts and patient tumors. Spatially, these fluctuations were predominantly associated with regions of heterogeneous perfusion and hypoxia in the

PurposeTo determine whether quantitation of T2* is sufficiently repeatable and sensitive to detect clinically relevant oxygenation levels in head and neck squamous cell carcinoma (HNSCC) at 3T.Materials and MethodsTen patients with newly diagnosed locally advanced HNSCC underwent two magnetic resonance imaging (MRI) scans between 24 and 168 hours apart prior to chemoradiotherapy treatment. A multiple gradient echo sequence was used to calculate T2* maps. A quadratic function was used to model the blood transverse relaxation rate as a function of blood oxygenation. A set of published coefficients measured at 3T were incorporated to account for tissue hematocrit levels and used to plot the dependence of fractional blood oxygenation (Y) on T2* values, together with the corresponding repeatability range. Repeatability of T2* using Bland–Altman analysis, and calculation of limits of agreement (LoA), was used to assess the sensitivity, defined as the minimum difference in fractional blood oxygenation that can be confidently detected.Results T2* LoA for 22 outlined tumor volumes were 13%. The T2* dependence of fractional blood oxygenation increases monotonically, resulting in increasing sensitivity of the method with increasing blood oxygenation. For fractional blood oxygenation values above 0.11, changes in T2* were sufficient to detect differences in blood oxygenation greater than 10% (Δ T2* > LoA for ΔY > 0.1).ConclusionQuantitation of T2* at 3T can detect clinically relevant changes in tumor oxygenation within a wide range of blood volumes and oxygen tensions, including levels reported in HNSCC. J. Magn. Reson. Imaging 2016;44:72–80.