Precision stem cell regeneration enabled by a collagen-targeting, fate-directing bi-functional peptide

Precise control over stem cell selection, delivery, and fate remains a key challenge in regenerative medicine. Here, we report a collagen-targeting, fate-directing bi-functional peptide (BiFP) that enhances stem cell therapy for osteoarthritis and severe corneal epithelial defects. BiFP is a synthetic collagen-like peptide composed of a GFOGER motif that binds integrin α2β1 to promote cell adhesion, survival, and chondrogenic differentiation, and a collagen XII-binding sequence that enables tissue-specific targeting. Structural analysis confirmed sequence accuracy by nano-LC-MS/MS, >90 % purity by HPLC, and a single 4434.5 Da band by SDS-PAGE and MALDI-TOF. Biophysical studies showed BiFP forms a stable triple-helix with a melting temperature of 53.2 °C, without higher-order folding, as evidenced by circular dichroism, DSC, and DSF. DLS and TEM revealed nanoparticles (∼1.4 nm) with some aggregation. In rat models, intra-articular or ocular delivery of BiFP-bound multipotent stromal cells (MSCs) significantly improved cell homing, engraftment, and regeneration of multi-layered neocartilage or corneal epithelium compared to MSCs alone. In vivo cell tracking confirmed MSC differentiation into chondrocytes and corneal epithelial cells, partially mediated by FAK-FoxO signaling. Pharmacokinetic analysis showed that free BiFP, when administered subcutaneously, was rapidly cleared, becoming undetectable in plasma and joints within 72h, with the kidneys identified as the primary route of excretion, supporting a favorable safety profile. These findings demonstrate that BiFP provides a modular, ECM-guided delivery platform with defined structural and functional properties, enabling precise localization and fate control of transplanted multipotent stem cells. This approach offers a promising strategy for targeted regenerative therapies with minimal systemic exposure.