Objectives The aim of this study was to demonstrate the feasibility of in vivo three-dimensional (3D) relaxation time normalT2∗ mapping of a dicarboxy-PROXYL radical using continuous-wave electron paramagnetic resonance (CW-EPR) imaging. Materials and methods Isotopically substituted dicarboxy-PROXYL radicals, 3,4-dicarboxy-2,2,5,5-tetra(2H3)methylpyrrolidin-(3,4-2H2)-(1-15N)-1-oxyl (2H,15N-DCP) and 3,4-dicarboxy-2,2,5,5-tetra(2H3)methylpyrrolidin-(3,4-2H2)-1-oxyl (2H-DCP), were used in the study. A clonogenic cell survival assay was performed with the 2H-DCP radical using squamous cell carcinoma (SCC VII) cells. The time course of EPR signal intensities of intravenously injected 2H,15N-DCP and 2H-DCP radicals were determined in tumor-bearing hind legs of mice (C3H/HeJ, male, n = 5). CW-EPR-based single-point imaging (SPI) was performed for 3D normalT2∗ mapping. Results 2H-DCP radical did not exhibit cytotoxicity at concentrations below 10 mM. The in vivo half-life of 2H,15N-DCP in tumor tissues was 24.7 ± 2.9 min (mean ± standard deviation [SD], n = 5). The in vivo time course of the EPR signal intensity of the 2H,15N-DCP radical showed a plateau of 10.2 ± 1.2 min (mean ± SD) where the EPR signal intensity remained at more than 90% of the maximum intensity. During the plateau, in vivo 3D maps with 2H,15N-DCP were obtained from tumor-bearing hind legs, with a total acquisition time of 7.5 min. Conclusion EPR signals of 2H,15N-DCP persisted long enough after bolus intravenous injection to conduct in vivo 3D normalT2∗ mapping with CW-EPR-based SPI.

This article reports a method of simultaneous T 2 * mapping of 14 N-and 15 N-labeled dicarboxy-PROXYLs using 750-MHz continuous-wave electron paramagnetic resonance (CW-EPR) imaging. To separate the spectra of 14 N-and 15 N-labeled dicarboxy-PROXYLs under magnetic field gradients, an optimization problem for spectral projections was formulated with the spatial total variation as a regularization term and solved using a local search based on the gradient descent algorithm. Using the single-point imaging (SPI) method with spectral projections of each radical, simultaneous T 2 * mapping was performed for solution samples. Simultaneous T 2 * mapping enabled visualization of the response of T 2 * values to the level of dissolved oxygen in the solution. Simultaneous T 2 * mapping applied to a mouse tumor model demonstrated the feasibility of the reported method for potential application to in vivo oxygenation imaging.