Abstract/Summary

Brain metastases (BrMs) are the most common type of brain malignancy and are associated with poor patient outcomes. The emerging field of cancer neuroscience has begun to explore the unexpected ways BrMs colonise the neural microenvironment. However, early colonisation of BrMs has not been effectively characterised; early interactions with the brain microenvironment could shape the trajectory of BrM progression based on the initial plastic responses they exhibit, which could inform BrM prevention. We have developed an ex vivo organotypic brain slice (OBS) model of BrMs to examine early BrM microenvironment interactions and plastic responses, with a particular focus on intracellular Ca2+ oscillations. This involved the co-culture of five mCherry+ cancer cell lines in neocortical organotypic brain slices (NC-OBSs) from 12-day old C57BL/6J mice. Four cell lines representing common primary sites of brain metastases (MDA-MB-231, SK-MEL-28, A549, and DU-145), as well as glioblastoma (U-87 MG). In-model measurements included viability, proliferation, chemokine expression, and cell invasion. Additionally, intracellular Ca2+ imaging was performed on GCaMP6s+ cells before and during co-culture and the variability in oscillation frequencies was assessed via Fast Fourier Transform (FFT) analysis and Principle Component Analysis (PCA). Further phenotypic assessments were conducted on the cancer cells post-model. Results indicated that interactions within the BrM model included cancer cell manipulation based upon altered chemokine secretions and reduced cell death compared to the negative control. FFT analysis and PCA revealed inter- and intracellular variation of spontaneous intracellular Ca2+ oscillation frequencies which changed over the course of the model culture. Further phenotypic changes in the cancer cells indicate a role for polyploidy and anoikis resistance in the early colonisation stage. Future work will utilise the full measurability and manipulability of the model via a variety of specific measurements including functional assessments of the Ca2+ toolkit and cell-cell interactions.