Tissue-engineered heart valves with self-repair and regeneration properties may overcome the problem of long-term degeneration of currently used artificial prostheses. The aim of this study was the development and in vivo proof-of-concept of next-generation off-the-shelf tissue-engineered sinus valve (TESV) for transcatheter pulmonary valve replacement (TPVR). Transcatheter implantation of off-the-shelf TESVs was performed in a translational sheep model for up to 16 weeks. Transapical delivery of TESVs was successful and showed good acute and short-term performance (up to 8 weeks), which then worsened over time most likely due to a non-optimized in vitro valve design. Post-mortem analyses confirmed the remodelling potential of the TESVs, with host cell infiltration, polymer degradation, and collagen and elastin deposition. TESVs proved to be suitable as TPVR in a preclinical model, with encouraging short-term performance and remodelling potential. Future studies will enhance the clinical translation of such approach by improving the valve design to ensure long-term functionality.