In vitro cell cultures are crucial research tools for modeling human development and diseases. Although the conventional monolayer cell cultures have been widely used in the past, the lack of tissue architecture and complexity of such model fails to inform the true biological processes in vivo. Recent advances in the organoid technology have revolutionized the in vitro culture tools for biomedical research by creating powerful three-dimensional (3D) models to recapitulate the cellular heterogeneity, structure, and functions of the primary tissues. Such organoid technology enables researchers to recreate human organs and diseases in a dish and thus holds great promises for many translational applications such as regenerative medicine, drug discovery, and precision medicine. In this review, we provide an overview of the organoid history and development. We discuss the strengths and limitations of organoids as well as their potential applications in the laboratory and the clinic.

Characterised by high intra‐ and inter‐tumor heterogeneity, metastatic renal cell carcinoma (RCC) is resistant to chemo‐ and radiotherapy. Therefore, the development of new prognostic and diagnostic markers for RCC patients is needed. Cancer stem cells (CSCs) are a small population of neoplastic cells within a tumor which present characteristics reminiscent of normal stem cells. CSCs are characterised by unlimited cell division, maintenance of the stem cell pool (self‐renewal), and capability to give rise to all cell types within a tumor; and contribute to metastasis in vivo (tumourigenicity), treatment resistance and recurrence. So far, many studies have tried to establish unique biomarkers to identify CSC populations in RCC. At the same time, different approaches have been developed with the aim to isolate CSCs. Consequently, several markers were found to be specifically expressed in CSCs and cancer stem‐like cells derived from RCC such as CD105, ALDH1, OCT4, CD133, and CXCR4. However, the contribution of genetic and epigenetic mechanisms, and tumor microenvironment, to cellular plasticity have made the discovery of unique biomarkers a very difficult task. In fact, contrasting results regarding the applicability of such markers to the isolation of renal CSCs have been reported in the literature. Therefore, a better understanding of the mechanism underlying CSC may help dissecting tumor heterogeneity and drug treatment efficiency.

Bi-allelic inactivation of the VHL gene on chromosome 3p is the characteristic feature in most clear cell renal cell carcinomas (ccRCC). Frequent gene alterations were also identified in SETD2, BAP1 and PBRM1, all of which are situated on chromosome 3p and encode histone/chromatin regulators. The relationship between gene mutation, loss of protein expression and the correlations with clinicopathological parameters is important for the understanding of renal cancer progression. We analyzed PBRM1 and BAP1 protein expression as well as the tri-methylation state of H3K36 as a surrogate marker for SETD2 activity in more than 700 RCC samples. In ccRCC loss of nuclear PBRM1 (68%), BAP1 (40%) and H3K36me3 (47%) expression was significantly correlated with each other, advanced tumor stage, poor tumor differentiation (P < .0001 each), and necrosis (P < .005) Targeted next generation sequencing of 83 ccRCC samples demonstrated a significant association of genetic mutations in PBRM1, BAP1, and SETD2 with absence of PBRM1, BAP1, and HEK36me3 protein expression (P < .05, each). By assigning the protein expression patterns to evolutionary subtypes, we revealed similar clinical phenotypes as suggested by TRACERx Renal. Given their important contribution to tumor suppression, we conclude that combined functional inactivation of PBRM1, BAP1, SETD2 and pVHL is critical for ccRCC progression.

Background: Extensive DNA sequencing has led to an unprecedented view of the diversity of individual genomes and their evolution among patients with clear cell renal cell carcinoma (ccRCC). Objective: To understand subclonal architecture and dynamics of patient-derived two-dimensional (2D) and three-dimensional (3D) ccRCC models in vitro, in order to determine whether they mirror ccRCC inter-and intratumor heterogeneity. Design, setting, and participants: We have established a comprehensive platform of living renal cancer cell models from ccRCC surgical specimens. Outcome measurements and statistical analysis: We confirmed the concordance of 2D and 3D patient-derived cell (PDC) models with the original tumor tissue in terms of histology, biomarker expression, cancer driver mutations, and copy number alterations. We addressed inter-and intrapatient heterogeneity by analyzing clonal dynamics during serial passaging. Results and limitations: In-depth genetic characterization verified the presence of heterogeneous cell populations, and revealed a high degree of similarity between subclonal compositions of monolayer and organoid cell cultures and the corresponding parental ccRCCs. Clonal dynamics were evident during serial passaging of cells in vitro, suggesting that PDC cultures can offer insights into evolutionary potential and treatment susceptibility of ccRCC subclones in vivo. Proof-of-concept drug profiling using selected ccRCC-targeted therapy agents highlighted patient-specific vulnerabilities in PDC models that could not be anticipated by interrogating commercially available cell lines. Conclusions: We demonstrate that PDC models mirror inter-and intratumor heterogeneity of ccRCC in vitro. Based on our findings, we envision that the use of these models will advance our understanding of the trajectories that cause genetic diversity and their consequences for treatment on an individual level. Patient summary: In this study, we developed two-and three-dimensional patient-derived models from clear cell renal cell carcinoma (ccRCC) as "mini-tumors in a dish." We show that these cell models retain important features of the human ccRCCs such as the profound tumor heterogeneity, thus highlighting their importance for cancer research and precision medicine.