Inhibiting WNT and NOTCH in renal cancer stem cells and the implications for human patients

Abstract

Current treatments for clear cell renal cell cancer (ccRCC) are insufficient because two-thirds of patients with metastases progress within two years. Here we report the identification and characterization of a cancer stem cell (CSC) population in ccRCC. CSCs are quantitatively correlated with tumor aggressiveness and metastasis. Transcriptional profiling and single cell sequencing reveal that these CSCs exhibit an activation of WNT and NOTCH signaling. A significant obstacle to the development of rational treatments has been the discrepancy between model systems and the in vivo situation of patients. To address this, we use CSCs to establish non-adherent sphere cultures, 3D tumor organoids, and xenografts. Treatment with WNT and NOTCH inhibitors blocks the proliferation and self-renewal of CSCs in sphere cultures and organoids, and impairs tumor growth in patient-derived xenografts in mice. These findings suggest that our approach is a promising route towards the development of personalized treatments for individual patients.

Establishment of Kidney Cancer Organoid Cultures

“…Since organoids closely recapitulate several properties of the original tumor and since they are scalable for high-throughput drug screens, they present excellent in vitro systems that connect clinical trials with cancer genetics, complement current preclinical model-based drug studies and advance the development of personalized therapy design. This has been successfully demonstrated in colorectal cancer organoids as well as in our study on kidney cancer organoids 5,11 .…”

“…We have shown that organoids exhibit kidney-specific epithelial differentiation, while retaining cancer stem cell populations. Organoids resemble proximal tubule structures -the potential cell of origin of ccRCC -and recapitulate typical characteristics of ccRCC, including glycogen deposits and lipid droplets 5 .…”

“…In this protocol, kidney cancer organoid cultures are grown in defined medium -its composition is summarized in Table 2. Unlike other organoid cultures, kidney cancer organoids exhibited no requirement for R-Spondin 1, since endogenous autocrine mechanisms to upregulate Wnt signaling were found 5 . Kidney cancer organoids showed strong dependence on EGFR and FGF.…”

“…In our work, we focused on the determination of a cancer stem cell population and its implications for patient-specific therapy response in different patient-derived cancer systems 5 . Therefore our focus was to optimize growth conditions for tumor organoids.…”

“…Organoid cultures have increasingly been shown to recapitulate a tumor in vitro 4 . We have developed and characterized a novel ccRCC organoid culture system, which could be used to test the patient-specific response to treatment strategies in vitro 5 . In this protocol, we provide further detailed instructions on how to establish and maintain these culture systems and evaluate them for your individual research needs.…”