High‐Resolution Imaging of Extracellular pH in Mouse Liver Tumor

ABSTRACT

Extracellular acidosis is a biologically important feature of the tumor microenvironment in the liver, promoting immune evasion, angiogenesis, and resistance to therapy, and representing a mechanistically important and potentially targetable axis in liver cancer. Imaging extracellular pH (pH _e ) at high resolution is needed to better understand the immuno‐metabolic interplay, especially at the transition regions between the tumor core, tumor margin, and background liver, which is critical for any pharmacological or image‐guided intervention. Yet, there is a paucity of imaging techniques capable of providing pH _e mapping at high resolution. Here, we demonstrate high‐resolution pH _e imaging in a mouse Hepa1–6 liver tumor model using ¹ H Biosensor Imaging of Redundant Deviation in Shifts (BIRDS) with REduced Spherical Encoding with GAussian Weighting (RESEGAW). Eight tumor‐bearing C57BL/6J mice were used to demonstrate pH _e imaging with RESEGAW using the macrocyclic agent TmDOTP ⁵⁻ at 0.6 mm isotropic resolution on a 9.4 T scanner, which was validated using ³¹ P‐MRSI with 3‐aminopropylphosphonate (3‐APP). pH _e imaging with ¹ H‐BIRDS‐RESEGAW consistently showed acidic tumor regions (pH _e  = 6.77 ± 0.14) relative to adjacent normal liver (pH _e  = 7.14 ± 0.07). Mean pH _e values measured by ³¹ P‐MRSI with 3‐APP and ¹ H‐BIRDS‐RESEGAW with TmDOTP ⁵⁻ show no significant differences in tumors (pH _e  = 6.81 ± 0.13) and normal liver (pH _e  = 7.14 ± 0.06). Voxelwise comparison after co‐registration of ³¹ P‐MRSI with 3‐APP to ¹ H‐BIRDS‐RESEGAW using Bland–Altman analysis demonstrated excellent agreement between the two methods, with minimal mean bias (−0.005 pH units) and variance of less than 0.1 pH units. These results demonstrate the feasibility and quantitative reliability of ¹ H‐BIRDS‐RESEGAW for imaging extracellular acidosis in liver tumors at submillimeter resolution, establishing a technical foundation for studying the immuno‐metabolic interplay in liver cancer and its response to therapy.