Imaging in vivo extracellular pH with a single paramagnetic chemical exchange saturation transfer magnetic resonance imaging contrast agent

Abstract

The measurement of extracellular pH (pHe) has potential utility for cancer diagnoses and for assessing the therapeutic effects of pH-dependent therapies. A single magnetic resonance imaging (MRI) contrast agent that is detected through paramagnetic chemical exchange saturation transfer (PARACEST) was designed to measure tumor pH(e) throughout the range of physiologic pH and with magnetic resonance saturation powers that are not harmful to a mouse model of cancer. The chemical characterization and modeling of the contrast agent Yb(3+)-1,4,7,10-tetraazacyclododecane-1,4,7-triacetic acid,10-o-aminoanilide (Yb-DO3A-oAA) suggested that the aryl amine of the agent forms an intramolecular hydrogen bond with a proximal carboxylate ligand, which was essential for generating a practical chemical exchange saturation transfer (CEST) effect from an amine. A ratio of CEST effects from the aryl amine and amide was linearly correlated with pH throughout the physiologic pH range. The pH calibration was used to produce a parametric pH map of a subcutaneous flank tumor on a mouse model of MCF-7 mammary carcinoma. Although refinements in the in vivo CEST MRI methodology may improve the accuracy of pHe measurements, this study demonstrated that the PARACEST contrast agent can be used to generate parametric pH maps of in vivo tumors with saturation power levels that are not harmful to a mouse model of cancer.

Figure 1

A model of Yb-DO3A-oAA. The proximity of Yb(III) to the amide and the amine causes a shift in the magnetic resonance frequencies of these chemical groups. The proximities of the amine and carboxylate ligands indicate hydrogen bonds between these groups.

Figure 2

The extraction of PARACEST effects of the amine and amide from experimental data by three lorentzian line shapes centered at 0, −11, and +8 ppm. The CEST spectrum (solid line, no symbol) of Yb-DO3A-oAA at pH 6.5 and 37°C was acquired with selective saturation applied at 12.2 μT for 6 s, from +20 ppm to −20 ppm in 1 ppm increments. The line-fitting output showed a lorentzian line shape of water that was symmetric about 0 ppm (solid line with triangles) and lorentzian line shapes of the CEST effects of the amine (solid line with open circles) and the amide (solid line with filled circles).

Figure 3

The exchange rate dependencies on pH of the amine and amide. The measured exchange rates of the amide (circles) and the amine (squares) were fit with equation 7 to extract the acid-base-catalyzed exchange constants. The thin line was the modeled behavior for the amide exchange process. The thick line was the modeled behavior for base-catalyzed exchange of amine protons in the pH range of 6.1 to 7.0 and for acid-catalyzed exchange in the pH range of 7.2 to 8.0.

Figure 4

A, The pH dependencies of amide CEST (diamonds) and amine CEST (squares). B, The ratio of the CEST effects relative to pH. C, The log₁₀ of the CEST ratio relative to pH. The phantoms consisted of 30 mM Yb-DO3A-oAA in phosphate-buffered saline at 37°C, with pH values ranging from 6.1 to 8.0 in increments of 0.25 pH units. The NMR method was the same as described in Figure 2.

Figure 5

The precision of pH measurements. A, Solutions of Yb-DO3A-oAA with T_1Wsat greater than 1.5 s were used to measure pH at 600 MHz magnetic field strength. The standard deviation was determined from three trials per concentration. B, Solutions of Yb-DO3A-oAA at 40 mM concentration were used to measure pH at 300 MHz magnetic field strength. The standard deviation was determined from three trials per T_1Wsat value. The average standard deviation of pH measurements was 0.11 pH units for samples with at least 10 mM concentration and 0.5 s T_1Wsat.

Figure 6

The parametric map of the CEST effect of the amine divided by the CEST effect of the amide (A) was used to create an initial parametric pH map (B). Pixels that showed a statistically significant CEST effect from the amine and amide were retained to create (C) a filtered parametric pH map of a mouse tumor model. The filtered parametric pH map is overlaid on an anatomic MRI. A solution of 60 mM of Yb-DO3A-oAA in 50 μL volume was directly injected into the center of the tumor. The result showed an acidic environment in the tumor region.