The combination of Bone Marrow Mesenchymal Stromal Cells (BM-MSC) with active injectable carriers brings about innovative solutions to current issues in the field of tissue engineering. In particular, repair of adult articular cartilage lesions remains a clinical challenge because of the limited self-healing capacity of cartilage. We demonstrated previously that the open porosity of homemade collagen microspheres allows for the entrapment and progressive release of TGF-β3, which efficiently triggered the chondrogenic differentiation of BM-MSC in vitro and in vivo, and the production of neo-cartilage tissue. However, one major hurdle in MSC-based therapies for cartilage repair is their late hypertrophic differentiation and subsequent tissue calcification, characterized by the secretion of specific markers such as type X collagen, alkaline phosphatase, osteocalcin and metalloprotease 13 (MMP13). To tackle this challenge, we identified Runx2, which plays a central role in chondrocyte hypertrophy, as the main molecular target to be repressed. Indeed, Runx2 has been widely described to up-regulate the expression of hypertrophic markers. We previously demonstrated that the transient down-regulation of this factor can be achieved with a specific siRNA targeting Runx2. Hence, the strategy of the Spacecart project is to use the transient down-regulation of Runx2 with siRNA (siRunx2) to prevent calcification and ultimately bone formation. We intend to deliver siRunx2 from collagen microspheres also used as an injectable support for BM-MSC and as a TGF-β3 reservoir. However, efficient down-regulation of Runx2 with siRNA requires transfection vectors to bring the nucleic acid to its nuclear target within the cells. Also, to maintain efficient chondrogenic differentiation of BM-MSC, it is important that Runx2 be repressed only after induction of the chondrocyte phenotype, i.e. approximatively day 14, thus calling for delayed delivery of the siRNA. For this project we have designed modified DOTAP-DOPE lipoplexes as the nucleic acid vector. These vectors will be loaded into collagen microspheres and anchored to the matrix via MMP13-sensitive peptides. The action of MMP13 secreted by MSCs will therefore trigger the local delivery of siRNA to the cells at the early stage of hypertrophic differentiation commitment. This cell-elicited spatio-temporal control of siRNA release is expected to help maintain the phenotype of mature chondrocytes in the long term and achieve fully functional hyaline cartilage regeneration. Our work plan includes three experimental tasks dedicated to 1) the design and synthesis of an optimal MMP13 peptide substrate and its use as cleavable linker between the collagen microspheres and siRNA vectors, 2) the investigation of the down-regulation of Runx2 in BM-MSC and its outcome on in vitro chondrogenesis and hypertrophy inhibition and 3) the study of neocartilage production in vivo in the absence of ossification in the long-term. Our consortium gathers complementary expertise in the fields of biomaterial elaboration and functionalization, peptide design and synthesis, and MSC-based therapy of cartilage pathologies. The main originality of our research project is to use the secretion of MMP13 by BM-MSC undergoing hypertrophic differentiation, to locally trigger the delivery of an anti-hypertrophic siRNA. We believe that our integrative approach is original and has a strong innovative potential. In addition, such highly specific self-induced retro-control of the cell behavior can potentially be transposed to other therapeutic indications by adapting the peptides to the enzymatic secretion profile of the specific cells. Therefore, we expect that Spacecart will have an impact on the broad community in the field of biomaterials and tissue engineering.