Establishment of patient-derived 3D tumouroids: Personalised medicine tools for renal cancer.

Abstract

Despite therapeutic advancements in the management of Renal Cell Carcinoma (RCC), there is an unmet clinical need for patient-specific in vitro models that can predict responses to therapy and enhance our understanding of this heterogeneous disease. Here, we established biomimetic 3D in vitro models, termed tumouroids, that incorporate patient-derived tumour cells and a complex stroma to mimic the physiological tumour microenvironment. We isolated tumour cells from RCC surgical specimens (n = 20) using a mechanical-enzymatic method. Two tumouroid types were manufactured: simple tumouroids consisting of patient-derived tumour cells, collagen and matrix proteins, and complex tumouroids that incorporated an additional stromal compartment populated with fibroblasts and endothelial cells. An important feature of tumouroids is that owing to fabrication via plastic compression their density mimics in vivo physiological tissue density. Patient-derived cells and tumouroids were characterised through immunofluorescence and histology to investigate resemblance to original tissues. Finally, tumouroids were subjected to treatment with Pazopanib (Votrient™), a tyrosine kinase inhibitor (TKI) used in the treatment of advanced RCC. Patient-derived tumouroids maintained the expression of characteristic RCC protein markers: carbonic anydrase IX (CA9), cytokeratins (CK7, CK8&18) and EMT markers (αSMA). The subtype-specific histology of the original tumour, e.g., clear cell, was also preserved. We observed Pazopanib-induced cytotoxicity, as measured by ATP production, ranging from none to strong (40 %) for individual patient-derived simple tumouroids (n = 12). In complex tumouroids, endothelial networks were also disrupted. Overall, patient-derived tumouroids mimicked the original tissue and successfully reproduced the response signatures to TKIs. STATEMENT OF SIGNIFICANCE: A significant number of renal cell carcinoma patients do not respond to targeted therapies or develop resistance and succumb to the disease. Here, we developed sophisticated 3D in vitro tumour models, termed tumouroids, from patient samples. Tumouroids mimic the original tumour and its microenvironment and elicit response to drugs that target both the tumour cells and the tumour microenvironment. This is a patient-specific in vitro tool that addresses the unmet clinical need for predicting an individual's response to therapy.