Caspase-3-Responsive Dual-Enhanced 1H/19F MRI Bimodal Probe for In Vivo Tumor Apoptosis Imaging

Caspase-3 (Cas-3) is a crucial cysteine protease involved in executing cell apoptosis, a process that is a hallmark of tumor cell apoptosis response to chemotherapy. Developing molecular imaging probes that specifically detect apoptotic signals during tumor treatment is considered to be one of the most important and effective strategies for reducing cancer-associated death rates and improving treatment outcomes. However, achieving a dual-enhanced bimodal probe in a single molecule remains a significant challenge. In this study, we developed a 1H/19F dual-enhanced magnetic resonance imaging (MRI) probe, CF3DEVDFFFK(Fmoc)-Gd, responsive to caspase-3 for in vivo imaging of apoptotic cells. Upon interaction with caspase-3, CF3DEVDFFFK(Fmoc)-Gd efficiently splits into two components CF3DEVD and FFFK(Fmoc)-C2-Gd, where FFFK(Fmoc)-C2-Gd subsequently self-assembles into nanofibers. This process activates both 19F MRI and 1H MRI, with longitudinal relaxivity (r1) increasing from 9.38 ± 0.22 to 23.24 ± 2.33 mM-1 s-1 at 0.5 T and turning on the 19F MRI signal due to the absence of the paramagnetic relaxation enhancement (PRE) effect. In vivo imaging results demonstrated that, after systemic administration, CF3DEVDFFFK(Fmoc)-Gd effectively accumulated in apoptotic 4T1 tumor tissues, resulting in significantly enhanced 1H MRI signals for visualization of caspase-3 activity in doxorubicin-treated apoptotic 4T1 tumor tissues, with signal intensity three times greater than that of Gd-DTPA. 19F MRI further complemented 1H MRI, with a notable recovery of the 19F MRI signal after intratumoral injection. These results confirm that CF3DEVDFFFK(Fmoc)-Gd effectively reports tumor apoptosis through combined 1H and 19F MRI, offering a promising approach for the preliminary assessment of antitumor efficacy in vivo.