Size-Dependent Cascade Enhancement of T₁-T₂ Dual-Modal MRI in Tumors

Currently, there is no conclusive evidence indicating that in situ self-assembled Gd nanostructures of varying sizes demonstrate distinct T₁ and T₂ signal enhancement capabilities. Furthermore, it remains uncertain whether size adjustment can effectively achieve enhanced T₁-T₂ dual-modal MRI. To address these uncertainties, a two-step in situ self-assembly strategy is developed. This approach began with a small-sized nanoprobe, Gd-TCO-P, with a hydrodynamic diameter (dH) of 16 ± 3 nm. This nanoprobe underwent alkaline phosphatase (ALP) cleavage and self-assembled intracellularly into short nanofibers termed Gd-NFs (dH: 200 ± 51 nm). The subsequent introduction of tetrazine-tetrazine crosslinked these Gd-NFs, leading to the formation of larger two-stage dendritic nanofibers known as Gd-TS-NFs (dH: 4371 ± 236 nm). This process achieves size-dependent enhancement of both T₁ and T₂ signals, which is validated through both in vitro and in vivo experiments, enabling precise long-term imaging of ALP-overexpressing tumors. This study not only provides valuable insights into the relationship between the size of in situ formed Gd nanostructures and T₁/T₂ MRI contrast enhancement, but also suggests a promising strategy for clinical applications of T₁-T₂ dual-modal MRI.