Elena Delgrosso1, Franca Scocozza2, Laura Cansolino1,3, Federica Riva4, Michele Conti2, Ian Postuma3, Silva Bortolussi3,5, Cinzia Ferrari1,3,6
1Department of Clinical Surgical Sciences, Integrated Unit of Experimental Surgery, Advanced Microsurgery and Regenerative Medicine, University of Pavia, Pavia, Italy;
2Department of Civil Engineering and Architecture, University of Pavia, Pavia, Italy;
3National Institute of Nuclear Physics (INFN), Unit of Pavia, Pavia, Italy;
4Department of Public Health, Experimental and Forensic Medicine, Histology and Embryology, University of Pavia, Pavia, Italy;
5Department of Physics, University of Pavia, Pavia, Italy;
6Animal Care and Radiobiology Centre, University of Pavia, Pavia, Italy
Correspondence to: Elena Delgrosso, PhD. Department of Clinical Surgical Sciences, Integrated Unit of Experimental Surgery, Advanced Microsurgery and Regenerative Medicine, University of Pavia, via Ferrata 9, 27100 Pavia, Italy. Email: elena.delgrosso01@universitadipavia.it.
Background: Osteosarcoma is the most common primary malignant bone tumor, primarily affecting children and young adults. Although combined chemotherapy and surgical resection have improved outcomes in localized cases, prognosis remains poor in patients with metastases due to the tumor’s infiltrative nature, which leads to high rates of recurrence. Boron neutron capture therapy (BNCT) is an experimental radiotherapy that offers a selective approach: boron-10 accumulates in tumor cells, and upon neutron irradiation, it triggers a reaction that destroys cancer cells while sparing healthy tissue. However, current preclinical models—two-dimensional (2D) cultures and animal studies—do not accurately replicate tumor complexity or are limited by ethical and logistical constraints. The objective of this study is to develop and optimize a three-dimensional (3D) bioprinted osteosarcoma model for experimental BNCT applications, providing a physiologically relevant, reproducible, and ethical in vitro platform that better mimics the tumor microenvironment.
Methods: A 3D osteosarcoma model was created using a pneumatic extrusion bioprinter with a bioink of 8% sodium alginate and 4% gelatin, encapsulating UMR-106 cells. Constructs were cultured for 28 days; cell distribution and proliferation were assessed by Hoechst staining. Boron uptake and distribution for BNCT were analyzed via neutron autoradiography.
Results: We successfully generated a fully colonized 3D living cell construct using the rat osteosarcoma cell line UMR-106 cells within 28 days of culture. The evaluation of intracellular boron levels is crucial for BNCT studies. However, the method for quantifying boron in cells within the constructs needs to be further refined. A critical point is the interference of the gel matrix with the boron measurements.
Conclusions: The proposed 3D model could serve as an alternative or parallel approach to 2D culture and in vivo animal models for BNCT experimental studies. By providing a more complex and representative tumor microenvironment, this model has the potential to enhance our understanding of BNCT treatment efficacy and could ultimately aid in the development of more effective therapies for osteosarcoma.
Keywords: Three-dimensional bioprinting (3D bioprinting); tumor model; osteosarcoma in vitro model; 3D cell culture; boron neutron capture therapy (BNCT)
