PO-435 Photoactivation of nanoparticles delivered by mesenchymal stem cells induces osteosarcoma cell death in in vitro 3D co-culture models

Abstract

IntroductionOsteosarcoma (OS) is a rare and aggressive tumour that mainly affects long bones of adolescents. Currently, OS patients are treated with a combination of multi-agent chemotherapy and surgery. However, 30% of patients do not respond to standard treatment. Therefore, innovative therapeutic agents are needed. Mesenchymal stem cells (MSCs) display a specific tumour-tropism and have been previously used in successful preclinical studies to deliver several therapeutic agents. Furthermore, the safety of genetically engineered MSCs was demonstrasted in ongoing clinical trial. The goal of the present study was to test in vitro whether MSCs could uptake photoactivable nanoparticles (NPs) and induce cell death of OS cells upon photoactivation.Material and methodsPtl@PMMA NPs were produced by adding tetrasulfonate aluminium phthalocyanine (Ptl) to an aqueous solution of positively charged poly-methylmethacrylate (PMMA) nanoparticles. The photosensitizer Ptl is activated in near-infrared light allowing a deep tissue penetration. Human MSC lines, isolated from the bone marrow of multiple donors, were loaded with Ptl@PMMA NPs. The MSCs’ ability to internalise and retain NPs, along with their migratory properties, were tested. Cell death upon photoactivation (PDT) was evaluated in vitro, on a monolayer co-culture of MSCs and OS cells and in 3D multicellular spheroids, generated via cell suspension in ultralow attachment platesResults and discussionsMSCs showed an internalisation rate of Plt@PMMA>95%, which did not alter cell viability and migratory capacity. When Ptl@PMMA-MSCs were co-cultured with a human OS cell line (SaOS-2) in monolayers, they efficiently triggered cell death upon PDT. In particular, AnnexinV/PI and CalceinAM/EthD staining showed 70% of cell death in the co-culture system. These results were also validated by a metabolic assay. Interestingly, in a 3D co-culture of the OS cell line MG63 and Ptl@PMMA-MSCs, we observed a marked reduction of the viability (<5%) measured by the ATP content 24 hours after PDT. A massive cell necrosis induced by the photoactivation of the Ptl in the whole spheroid mass was confirmed by CalceinAM/EthD staining and TEM imaging.ConclusionFor the first time, we demonstrated that photoactivation of MSCs loaded with Ptl@PMMA NPs can successfully induce OS cell death in a three-dimensional OS model. These results encourage further in vivo evaluation to demonstrate the specific targeting of Plt@PMMA loaded MSCs to the tumour stroma and the efficacy of PDT treatment