Development of tools for monitoring the tumor microenvironment by MR techniques
Since several years, we are developing innovative MR technologies to characterize several hallmarks of cancer, including the tumor hemodynamics and their different components: tissue oxygenation, perfusion, oxygen delivery and consumption,听redox status and superoxide production,听as well as the tumor proliferation and metabolic features.
We pioneered developments in EPR oximetry with the characterization of paramagnetic materials possessing favorable features for oximetry. Thanks to these developments, EPR oximetry is routinely used in the laboratory for studying the temporal evolution of tumor pO2. The technique is unique in a sense that it monitors oxygenation inside a tissue non-invasively and repeatidly from the same site over time. In a translational approach, we also developed biocompatible forms of these systems. One clinical EPR system听allows carrying out clinical EPR studies in oncology and diabetology. In the purpose, a clinical study has been conducted听to assess melanin in melanoma with the ultimate goal of stratifying malignant versus bening naevi. We have demonstrated for the first time the ability of EPR to detect noninvasively an endogenous free radical in human skin melanoma. The spectrometer has been upgraded with a unique capability to detect the EPR signal in multiharmonic mode in order to increase the sensitivity of the method. We have also been interested in developing ways to measure oxygen using MRI, namely by using 19F relaxometry in order to map tumor oxygenation, or using endogenous contrast based on R1 and R2*.
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Typical maps of global R鈧� and R鈧� of lipids obtained on the same mice at baseline, and after hyperoxic and hypoxic challenges performed with carbogen and CA4, respectively. For this tumor, actual values of pO鈧� was 6.1 mm Hg at baseline increased to 9.0 mm Hg during the carbogen breathing, and decreased to 5.1 mm Hg 3 hours after CA4 administration.
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Regarding hemodynamics, we are characterizing the tumor perfusion and permeability with Dynamic Contrast-Enhanced (DCE) 鈥� or Dynamic Susceptibility Contrast (DSC) MRI. We are also continuously developing new methodologies to measure tumor oxygen consumption in vivo, using 17O-NMR and EPR oximetry.
We focused more recently on the tumor metabolism, which is a target of new therapeutic strategies. More specifically, studies are assessing in vivo: the extracellular pH, the glycolytic/oxidative tumor phenotypes and their potential role in tumor resistance to treatment, and the link between tumor cell metabolism and cell proliferation, using 1H-MRS, steady-state 13C-MRS, and dynamic hyperpolarized 13C-MRS via 13C-enriched substrates.
We validated mitochondrial redox nitroxide EPR probes to assess tumor redox status in vitro and in vivo, in response to the modulation of gluthatione and thioredoxin status. We are currently assessing innovative probes and combining detection with EPR and MRI for a better characterization of the redox status.
We also recently developed a mitochondrial 鈥榯oolbox鈥� (mito-ToolBox) for measuring mitochondrial superoxide simultaneously to oxygen consumption rate (OCR) measurement. This unique versatile toolbox is presently used to assess the effect of treatments tackling the mitochondrial function of cancer cells as well as the effect of intoxicants on normal cells. In vivo translation of this tool is currently assessed using both EPR and MRI.
Applications of MR (EPR and NMR) to characterize the tumor micro-environment
Our goal is to characterize how the tumor microenvironment influences the response to therapy. We are testing novel approaches using the modulation of the vascular network and/or the inhibition of the oxygen consumption by tumor cells to increase the response to radiation therapy and/or chemotherapy. In this way, we are trying to define optimal schedule for an optimal therapy. We are also characterizing the evolution of the tumor microenvironment after therapies that are targeting the tumor metabolism. Thanks to the unique tools that have been developed in our laboratory, we propose new strategies to optimize radiation therapy, chemotherapy, and targeted therapies. As an illustrative example, we are studying the effect of statins on the tumor hemodynamics and response to therapies. Another field of interest is the application of pH assessment using both CEST-MRI and EPR to assist in therapeutic guidance of treatments targeting proton extruders overexpressed by glycolytic cancer cells. Recent studies were focused on inhibitors of monocarboxylate transporters (MCTs) and mitochondrial pyruvate carrier (MPC).听A more recent research activity of the laboratory is focused on the anti-cancer strategies targeting the tumor metabolism. Using 13C-NMR spectroscopy, we are assessing the effect of PDK, BRAF, EGFR, and CDK4/6 inhibitors on glycolytic flux and tumor metabolism. The identification of alternative metabolic pathways used by tumor cells to sustain their proliferation can be considered as a major mechanism of resistance to this type of treatment. This research will provide a rationale for innovative combination of therapies targeting tumor metabolism. In collaboration with the Metabolism and nutrition research group of the LDRI, we recently assessed the metabolic status of breast tumors growing on obese mice in comparison with lean mice and identified a unique metabolic feature linked to obesity, using hyperpolarized 13C-pyruvate combined with 13C-glucose tracing experiments involving both 13C-MRS and mass spectrometry (collaborative work involving the NEST and MASMET platform of the LDRI).
Development of biomarkers predictive of the response to a treatment
In the field of radiation therapy, ongoing studies assess the use of imaging biomarkers (18F-FAZA, EPR oximetry, 19F-MRI) to evaluate the efficacy of anti-cancer strategies such as dose painting and dose escalation. In the field of chemotherapy, we are currently implementing methods that might be predictive of tumor response early in the treatment regimen and comparing their respective value: diffusion MRI (cellularity), 1H-spectroscopy of choline (membrane turnover), 13C-MRS (metabolism), 18F-FDG (glucose uptake), 18F-FLT PET (cell proliferation).
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Typical ADCw (Apparent Diffusion Coefficient of water) maps obtained on mice xenografts in response to the multi-kinase inhibitor sorafenib. Note the increase in global ADCw in the tumor region at day 5 post therapy.
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A Dynamic Nuclear Polarization (DNP, 鈥淗ypersense鈥�) system allows the study of metabolic fluxes using 13C-MRS. We are looking to the value of 13C enriched substrates (i.e. pyruvate-lactate exchange) as biomarkers of response to anti-cancer treatment, including EGFR (epidermal growth factor) inhibitors, MAPKinase inhibitors, as well as CDK4/6 inhibitors.
The steady-state assessment of 13C-enriched substrates is also used in combination with hyperpolarized studies for identification of resistance mechanisms to currently available therapies, as well as for the stratification of tumors that may benefit from innovative therapies that modulate the metabolism of cancer cells. This multi-modal strategy significantly contributes to the identification of early non-invasive imaging markers of tumor response to combined targeted therapies in the transition towards individualized cancer therapy, with a special focus on the resistance to first line therapy in advanced breast cancer, in advanced melanoma, and in Head & Neck tumors, in collaborations with medical oncologists of the Experimental and Clinical Research Institute (UCL, Profs. J-P. Machiels, S. Schmidt, J.F. Baurain, F. Duhoux and C. van Marcke). The ultimate goal of this type of studies is to spare patient鈥檚 cycles of futile therapy, and possibly allow them to move to other, possibly experimental therapies. These metabolic studies, by identifying resistance mechanisms to targeted therapies (such as glutaminolysis, fatty acid oxidation, or glycolysis inhibition), thanks to the developed imaging metabolic tracers, will provide rationale for new therapeutic combinations involving metabolic targeted therapies.
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Schematic representation of an imaging session including 13C-MRS of hyperpolarized 13C-pyruvate. The 13C-pyruvate substrate is first hyperpolarized and then directly injected intravenously to the tumor-bearing mouse that is concomitantly imaged in a small-animal MRI scanner for detection of exchange with lactate and alanine using 13C-MRS, to assess metabolic fluxes in vivo in real time.
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In the last five years, the influence of obesity on breast cancer progression and tumor response to treatment is also being studied in collaboration with Prof. P.D. Cani of the Metabolism and Nutrition group of the LDRI institute. This project involves the study of the role of adipokines and gut microbiota in breast cancer and melanoma progression and metastatization.
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