PEO-PPO-PEO micelles as effective rAAV-mediated gene delivery systems to target human mesenchymal stem cells without altering their differentiation potency.

Unlabelled: Recombinant adeno-associated viral (rAAV) vectors are clinically adapted gene transfer vectors for direct human cartilage regenerative medicine. Their appropriate use in patients is still limited by a relatively low efficacy of vector penetration inside the cells, by the pre-existing humoral immune responses against the viral capsid proteins in a large part of the human population, and by possible inhibition of viral uptake by clinical compounds such as heparin. The delivery of rAAV vectors to their targets using optimized vehicles is therefore under active investigation. Here, we evaluated the possibility of providing rAAV to human bone marrow-derived mesenchymal stem cells (hMSCs), a potent source of cartilage regenerative cells, via self-assembled poly(ethylene oxide) (PEO) and poly(propylene oxide) (PPO) triblock copolymers as linear poloxamers or X-shaped poloxamines. Encapsulation in poloxamer PF68 and poloxamine T908 polymeric micelles allowed for an effective, durable, and safe modification of hMSCs via rAAV to levels similar to or even higher than those noted upon direct vector application. The copolymers were capable of restoring the transduction of hMSCs with rAAV in conditions of gene transfer inhibition, i.e. in the presence of heparin or of a specific antibody directed against the rAAV capsid, enabling effective therapeutic delivery of a chondrogenic sox9 sequence leading to an enhanced chondrocyte differentiation of the cells. The present findings highlight the value of PEO-PPO copolymers as powerful tools for rAAV-based cartilage regenerative medicine.

Statement Of Significance: While recombinant adeno-associated viral (rAAV) vectors are adapted vectors to treat a variety of human disorders, their clinical use is still restricted by pre-existing antiviral immune responses, by a low efficacy of natural vector entry in the target cells, and by inhibition of viral uptake by clinically used compounds like heparin. The search for alternative routes of rAAV delivery is thus becoming a new field of investigation. In the present study, we describe the strong benefits of providing rAAV to human mesenchymal stem cells, a potent source of cells for regenerative medicine, encapsulated in polymeric micelles based on poly(ethylene oxide) (PEO) and poly(propylene oxide) (PPO) triblock copolymers as novel, effective and safe delivery systems for human gene therapy.