A DNA-Based MRI Contrast Agent for Quantitative pH Measurement

Abstract

Extracellular pH is important in clinical measurements due to its correlation to cell metabolism and disease progression. In MRI, T₁/T₂ ratiometric analysis and other methods have been previously applied to quantify pH using conventional pulse sequences. However, for nanoparticle-based approaches, heterogeneity in size and surface functionalization tends toward qualitative rather than quantitative results. To address this limitation, we developed a novel DNA-based MRI contrast agent, pH-DMRCA, which utilizes a highly programmable and reproducible nanostructure. The pH-DMRCA is a dendritic DNA scaffold that is functionalized with a pH-responsive MRI-sensitive construct, Gd(NP-DO3A), at the end of each DNA arm. We first evaluated the r₁ and r₂ response of our pH-DMRCA over a range of pH values (pH = 5-9) to establish a relaxometric model of pH. These MRI-based assessments of pH were validated in a separate set of samples using a pH electrode (n = 18) and resulted in a good linear correlation (R² = 0.99, slope = 0.98, intercept = 0). A Bland-Altman analysis of the results also showed reasonable agreement between the calculated pH and measured pH. Moreover, these pH comparisons were consistent across three different pH-DMRCA concentrations, demonstrating concentration-independence of the method. This MRI-based pH quantification methodology was further verified in human blood plasma. Given the versatility of the DNA-based nanostructures, the contrast agent has a potential to be applied to a wide variety of imaging applications where extracellular pH is important including cancer, stroke, cardiovascular disease, and other important diseases.

Keywords: DNA dendrimer; DNA nanostructure; MRI contrast agent; bioimaging; pH quantification.

Figure 1.

Scheme of pH-DMRCA fabrication. (a) Annealing of amino-functionalized oligonucleotide strands S1–S5. (b) Conjugation of NP-DO3A to the DNA scaffold 1 via amide coupling. (c) Complexation of Gd³⁺ to 2 in water.

Figure 2.

(a) Relaxivity profiles of pH-DMRCA in HEPES as a function of pH. Relaxivities (r₁ and r₂) were calculated from concentration of pH-DMRCA and water relaxation times, n = 3. The inset figure is the r₁/r₂ ratiometric response at pH range 7.0–8.0. (b) T₁ and T₂ color maps of pH-DMRCA as a response to pH. Both (a) and (b) were measured with 2 μM pH-DMRCA in HEPES at 3.0 T, 25 °C.

Figure 3.

Unity plot comparison of pH measured using ratiometric MR analysis and conventional pH electrode. Measurements were performed at three different pH-DMRCA concentrations (n = 6 for 1 μM, n = 4 for 2 μM, n = 8 for 3 μM). Analysis of all measurements together (n = 18) shows linear correlation between the two methods (intercept set to 0). All data sets lie within the 95% prediction interval (mean ± 2σ).