Abstract It was recently demonstrated that newly invented positronium imaging may be used for improving cancer diagnostics by providing additional information about tissue pathology with respect to the standardized uptake value currently available in positron emission tomography (PET). Positronium imaging utilizes properties of a positronium atoms, which are built from the electron and positron produced in the body during PET examinations. We hypothesized whether positronium imaging would be sensitive to in vitro discrimination of tumour-like three-dimensional structures (spheroids) build of melanoma cell lines with different cancer activity and biological properties. The lifetime of ortho-Positronium (o-Ps) was evaluated in melanoma spheroids from two cell lines (WM266-4 and WM115) differing in the stage of malignancy. Additionally, we considered such parameters: as cell size, proliferation rate and malignancy to evaluate their relationship with o-Ps lifetime. We demonstrate the pilot results for the o-Ps lifetime measurement in extracellular matrix free spheroids. With the statistical significance of two standard deviations, we demonstrated that the higher the degree of malignancy and the rate of proliferation of neoplastic cells the shorter the lifetime of ortho-positronium. In particular we observed following indications encouraging further research: (i) WM266-4 spheroids characterized with higher proliferation rate and malignancy showed shorter o-Ps lifetime compared to WM115 spheroids characterized by lower growth rate, (ii) Both cell lines showed a decrease in the lifetime of o-Ps after spheroid generation in 8 th day comparing to 4 th day in culture and the mean o-Ps lifetime is longer for spheroids formed from WM115 cells than these from WM266-4 cells, regardless spheroid age. The results of these study revealed that positronium is a promising biomarker that may be applied in PET diagnostics for the assessment of the degree of cancer malignancy.

3D melanoma spheroid model for the development of positronium biomarker