Bioengineered Protein Contrast Agents with Dual-Targeting and Stroma-PrimingCapabilities to Overcome Biophysical Barriers in Pancreatic Cancer

Both pancreatic ductal adenocarcinoma (PDAC) and stroma-rich breast tumors harbor a rigid desmoplastic microenvironment that stifles molecular transport. In these malignancies, the cross-linked dense extracellular matrix (ECM) and elevated interstitial fluid pressure (IFP) function as a biophysical filter, stripping conventional agents of their diagnostic efficacy and rendering these tumors “silent” during routine imaging. Here, a dual- targeted, “transport-enabling” protein scaffold contrast agent, EAH-DOTA-Gd, is engineered to actively sur mount these physiological hurdles. Rationally designed with dual specificity for EGFR and HER2, EAH-DOTA-Gd achieves a remarkable longitudinal relaxivity (r 1 1 ) of 108.92 mM 1 s —approximately 22-fold higher than that of clinical standard Magnevist. Furthermore, the inclusion of an albumin-binding domain extends the systemic circulation half-life to 32.7 h, ensuring a widened diagnostic window. Beyond superior magnetic properties and pharmacokinetics, the contrast agent uniquely functions as a biophysical modulator: by attenuating fibroblast- mediated collagen deposition and reducing tumor IFP, it facilitates deep intratumoral penetration. This active remodeling capability enables robust, high-sensitivity visualization of orthotopic pancreatic and breast tumors that remain undetectable by conventional agents. These findings demonstrate that integrating molecular dual- targeting with active microenvironment modulation establishes a generalizable design paradigm for next- generation MRI contrast agents against transport-limited, desmoplastic malignancies.