Dataset Information

Characterization of Renal Cell Carcinoma Heterotypic 3D Co-Cultures with Immune Cell Subsets.

ABSTRACT: Two-dimensional cell culture-based platforms are easy and reproducible, however, they do not resemble the heterotypic cell-cell interactions or the complex tumor microenvironment. These parameters influence the treatment response and the cancer cell fate. Platforms to study the efficacy of anti-cancer treatments and their impact on the tumor microenvironment are currently being developed. In this study, we established robust, reproducible, and easy-to-use short-term spheroid cultures to mimic clear cell renal cell carcinoma (ccRCC). These 3D co-cultures included human endothelial cells, fibroblasts, immune cell subsets, and ccRCC cell lines, both parental and sunitinib-resistant. During spheroid formation, cells induce the production and secretion of the extracellular matrix. We monitored immune cell infiltration, surface protein expression, and the response to a treatment showing that the immune cells infiltrated the spheroid co-cultures within 6 h. Treatment with an optimized drug combination or the small molecule-based targeted drug sunitinib increased immune cell infiltration significantly. Assessing the therapeutic potential of this drug combination in this platform, we revealed that the expression of PD-L1 increased in 3D co-cultures. The cost- and time-effective establishment of our 3D co-culture model and its application as a pre-clinical drug screening platform can facilitate the treatment validation and clinical translation.

SUBMITTER: Rausch M

PROVIDER: S-EPMC8197009 | biostudies-literature | 2021 May

REPOSITORIES: biostudies-literature

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Characterization of Renal Cell Carcinoma Heterotypic 3D Co-Cultures with Immune Cell Subsets.

Rausch Magdalena M Blanc Léa L De Souza Silva Olga O Dormond Olivier O Griffioen Arjan W AW Nowak-Sliwinska Patrycja P

Cancers 20210522 11

Two-dimensional cell culture-based platforms are easy and reproducible, however, they do not resemble the heterotypic cell-cell interactions or the complex tumor microenvironment. These parameters influence the treatment response and the cancer cell fate. Platforms to study the efficacy of anti-cancer treatments and their impact on the tumor microenvironment are currently being developed. In this study, we established robust, reproducible, and easy-to-use short-term spheroid cultures to mimic cl ...[more]

PMID: 34067456

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Project description:Background: Head and neck squamous cell carcinomas (HNSCC) are phenotypically and molecularly heterogeneous and frequently develop therapy resistance. Reliable patient-derived 3D tumor models are urgently needed to further study the complex pathogenesis of these tumors and to overcome treatment failure. Methods: We developed a three-dimensional organotypic co-culture (3D-OTC) model for HNSCC that maintains the architecture and cell composition of the individual tumor. A dermal equivalent (DE), composed of healthy human-derived fibroblasts and viscose fibers, served as a scaffold for the patient sample. DEs were co-cultivated with 13 vital HNSCC explants (non-human papillomavirus (HPV) driven, n = 7; HPV-driven, n = 6). Fractionated irradiation was applied to 5 samples (non-HPV-driven, n = 2; HPV-driven n = 3). To evaluate expression of ki-67, cleaved caspase-3, pan-cytokeratin, p16INK4a, CD45, ∝smooth muscle actin and vimentin over time, immunohistochemistry and immunofluorescence staining were performed Patient checkup data were collected for up to 32 months after first diagnosis. Results: All non-HPV-driven 3D-OTCs encompassed proliferative cancer cells during cultivation for up to 21 days. Proliferation indices of primaries and 3D-OTCs were comparable and consistent over time. Overall, tumor explants displayed heterogeneous growth patterns (i.e., invasive, expansive, silent). Cancer-associated fibroblasts and leukocytes could be detected for up to 21 days. HPV DNA was detectable in both primary and 3D-OTCs (day 14) of HPV-driven tumors. However, p16INK4a expression levels were varying. Morphological alterations and radioresistant tumor cells were detected in 3D-OTC after fractionated irradiation in HPV-driven and non-driven samples. Conclusions: Our 3D-OTC model for HNSCC supports cancer cell survival and proliferation in their original microenvironment. The model enables investigation of invasive cancer growth and might, in the future, serve as a platform to perform sensitivity testing upon treatment to predict therapy response.

Project description:Invasive lobular carcinoma (ILC) is the second most common subtype of breast cancer following invasive ductal carcinoma (IDC) and characterized by the loss of E-cadherin-mediated adherens junctions. Despite displaying unique histologic and clinical features, ILC still remains a chronically understudied disease, with limited knowledge gleaned from available laboratory research models. Here we report a comprehensive 2D and 3D phenotypic characterization of four estrogen receptor-positive human ILC cell lines: MDA-MB-134, SUM44, MDA-MB-330, and BCK4. Compared with the IDC cell lines MCF7, T47D, and MDA-MB-231, ultra-low attachment culture conditions revealed remarkable anchorage independence unique to ILC cells, a feature not evident in soft-agar gels. Three-dimensional Collagen I and Matrigel culture indicated a generally loose morphology for ILC cell lines, which exhibited differing preferences for adhesion to extracellular matrix proteins in 2D. Furthermore, ILC cells were limited in their ability to migrate and invade in wound-scratch and transwell assays, with the exception of haptotaxis to Collagen I. Transcriptional comparison of these cell lines confirmed the decreased cell proliferation and E-cadherin-mediated intercellular junctions in ILC while uncovering the induction of novel pathways related to cyclic nucleotide phosphodiesterase activity, ion channels, drug metabolism, and alternative cell adhesion molecules such as N-cadherin, some of which were differentially regulated in ILC versus IDC tumors. Altogether, these studies provide an invaluable resource for the breast cancer research community and facilitate further functional discoveries toward understanding ILC, identifying novel drug targets, and ultimately improving the outcome of patients with ILC.Significance: These findings provide the breast cancer research community with a comprehensive assessment of human invasive lobular carcinoma (ILC) cell line signaling and behavior in various culture conditions, aiding future endeavors to develop therapies and to ultimately improve survival in patients with ILC. Cancer Res; 78(21); 6209-22. ©2018 AACR.

Project description:BackgroundSLFN11 has been found to regulate the development and progression of a variety of cancers and is associated with drug therapy, while its role in clear cell renal cell carcinoma (ccRCC) remains unclear; therefore, the aim of this study was to investigate SLFN11 expression in ccRCC patients and its correlations with clinicopathological and immunological features.MethodsGene profiles of ccRCC and the clinicopathological information of patients were downloaded from the TCGA database. Microarrays from the GEO database were used as a validation set for SLFN11 expression, which was experimentally verified in renal cancer cell lines by quantitative polymerase chain reaction (qPCR); protein expression and methylation levels were obtained from the HPA database and the UALCAN database. ROC curves, Kaplan-Meier survival analysis and Cox analysis were used to assess the diagnostic and predictive value of SLFN11 in ccRCC. Protein-protein interaction (PPI) networks for SLFN11 were obtained from the STRING website, and the TISIDB and TIMER 2.0 databases were used to study the relationship between SLFN11 and immune infiltration in the tumour microenvironment (TME).ResultsSLFN11 was significantly overexpressed in ccRCC tissues and renal cancer cell lines, which may be closely related to its hypermethylation status (P < 0.001). SLFN11 was positively correlated with a highly aggressive disease state, with the ROC curve showing an AUC value of 0.910 for SLFN11 in diagnosing ccRCC, and Kaplan-Meier and Cox analyses also revealed that upregulation of SLFN11 predicted a poor prognosis for ccRCC patients (P < 0.05). In addition, enrichment analysis showed that SLFN11 was closely associated with immune-related signalling pathways, and further exploration comprehensively demonstrated strong positive correlations with tumour immune lymphocytes, immune checkpoint genes, chemokines and chemokine receptors.ConclusionOverall, our data analysis shows that SLFN11 is a strong diagnostic and prognostic biomarker for ccRCC and is also associated with immune infiltration in the TME.

Dataset Information

Characterization of Renal Cell Carcinoma Heterotypic 3D Co-Cultures with Immune Cell Subsets.

Publications

Characterization of Renal Cell Carcinoma Heterotypic 3D Co-Cultures with Immune Cell Subsets.

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