Tumour spheroids are common in vitro experimental models of avascular tumour growth.,Compared with traditional two-dimensional culture, tumour spheroids more closely mimic the avascular tumour microenvironment where spatial differences in nutrient availability strongly influence growth.,We show that spheroids initiated using significantly different numbers of cells grow to similar limiting sizes, suggesting that avascular tumours have a limiting structure; in agreement with untested predictions of classical mathematical models of tumour spheroids.,We develop a novel mathematical and statistical framework to study the structure of tumour spheroids seeded from cells transduced with fluorescent cell cycle indicators, enabling us to discriminate between arrested and cycling cells and identify an arrested region.,Our analysis shows that transient spheroid structure is independent of initial spheroid size, and the limiting structure can be independent of seeding density.,Standard experimental protocols compare spheroid size as a function of time; however, our analysis suggests that comparing spheroid structure as a function of overall size produces results that are relatively insensitive to variability in spheroid size.,Our experimental observations are made using two melanoma cell lines, but our modelling framework applies across a wide range of spheroid culture conditions and cell lines.
Cutaneous melanoma is a highly aggressive cancer with a propensity for distant metastasis to various organs.,In contrast, melanoma arising in pigmented uveal layers of the eye metastasizes mostly in the liver.,The mechanisms of these metastases, which are ultimately resistant to therapy, are still unclear.,Metastasis via intravascular dissemination of tumour cells is widely accepted as a central paradigm.,However, we have previously described an alternative mode of tumour dissemination, extravascular migratory metastasis, based on clinical and experimental data.,This mechanism is characterised by the interaction of cancer cells with the abluminal vascular surface, which defines angiotropism.,Here, we employed our 3D co-culture approach to monitor cutaneous and uveal human melanoma cells dynamics in presence of vascular tubules.,Using time-lapse microscopy, we evaluated angiotropism, the migration of tumour cells along vascular tubules and the morphological changes occurring during these processes.,Cutaneous and uveal melanoma cells were injected in zebrafish embryos in order to develop xenografts.,Employing in vivo imaging coupled with 3D reconstruction, we monitored the interactions between cancer cells and the external surface of zebrafish vessels.,Overall, our results indicate that cutaneous and uveal melanoma cells spread similarly along the abluminal vascular surfaces, in vitro and in vivo.