Single cell RNA-seq from 10X for a living biobank of ovarian cancer ex vivo models reveals profound mitotic heterogeneity
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PROVIDER: PRJEB36333 | ENA |
REPOSITORIES: ENA
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Project description:We validated fifteen models from “living biobank” to provide models that support interrogation of Chromosome Instability and to evaluate novel therapeutics. Single cell RNA-seq was performed on tumour and stromal cells from 4 patients with ovarian cancer to establish gene expression profile differences between the two cell types and also heterogeneity within the tumour population. The samples used were AS38b, AS59, AS74-1, AS79, they are grown in OCMI media supplemented with 5% hyclone serum (AS38, AS59) or 5% FBS (AS74, AS79) in 5% CO2 and 5% O2. At 37C
2020-01-23 | E-MTAB-8559 | biostudies-arrayexpress
Project description:High-grade serous ovarian carcinoma is characterised by TP53 mutation and extensive chromosome instability (CIN). Because our understanding of CIN mechanisms is based largely on analysing established cell lines, we developed a workflow for generating ex vivo cultures from patient biopsies to provide models that support interrogation of CIN mechanisms in cells not extensively cultured in vitro. Here, we describe a "living biobank" of ovarian cancer models with extensive replicative capacity, derived from both ascites and solid biopsies. Fifteen models are characterised by p53 profiling, exome sequencing and transcriptomics, and karyotyped using single-cell whole-genome sequencing. Time-lapse microscopy reveals catastrophic and highly heterogeneous mitoses, suggesting that analysis of established cell lines probably underestimates mitotic dysfunction in advanced human cancers. Drug profiling reveals cisplatin sensitivities consistent with patient responses, demonstrating that this workflow has potential to generate personalized avatars with advantages over current pre-clinical models and the potential to guide clinical decision making.
| S-EPMC7018727 | biostudies-literature
Project description:In Rspondin-based 3D cultures, Lgr5 stem cells from multiple organs form ever-expanding epithelial organoids that retain their tissue identity. We report the establishment of tumor organoid cultures from 20 consecutive colorectal (CRC) patients. For most, organoids were also generated from adjacent normal tissue. The organoids closely resemble the original tumor. The spectrum of genetic changes observed within the 'living biobank' agrees well with previous large-scale mutational analyses of CRC. Gene expression analysis indicates that the major CRC molecular subtypes are represented. Tumor organoids are amenable to robotized, high-throughput drug screens allowing detection of gene-drug associations. As an example, a single organoid culture was exquisitely sensitive to Wnt secretion (porcupine) inhibitors and carried a mutation in the negative Wnt feedback regulator RNF43 (rather than in APC). Organoid technology may fill the gap between cancer genetics and patient trials, complement cell line- and xenograft-based drug studies and allow personalized therapy design. We generated organoids from healthy tissue and coloncarcinoma tissue. The organoids were trypsinized, plated in matrigel and overlaid with medium. After three days, RNA was isolated using Qiagen RNAeasy. Medium conditions are the same for all organoids, irrespective of their origin.
2015-05-07 | E-GEOD-64392 | biostudies-arrayexpress
Project description:A living biobank of ovarian cancer ex vivo models reveals profound mitotic heterogeneity
| PRJEB28664 | ENA
Project description:In Rspondin-based three-dimensional cultures, Lgr5 stem cells from multiple organs form ever-expanding epithelial organoids that retain their tissue identity. Here we report the establishment of tumor organoid cultures from 20 consecutive colorectal carcinoma (CRC) patients. For most, organoids were also generated from adjacent normal tissue. Organoids closely resemble the original tumor. The spectrum of genetic changes within the 'living biobank' agrees well with previous large-scale mutational analyses of CRC. Gene expression analysis indicates that the major CRC molecular subtypes are represented. Tumor organoids are amenable to high-throughput drug screens allowing detection of gene-drug associations. As an example, a single organoid culture was exquisitely sensitive to Wnt secretion (porcupine) inhibitors and carried a mutation in the negative Wnt feedback regulator RNF43, rather than in APC. Organoid technology may fill the gap between cancer genetics and patient trials, complement cell line- and xenograft-based drug studies and allow personalized therapy design.
2015-05-07 | GSE65253 | GEO
Project description:In Rspondin-based 3D cultures, Lgr5 stem cells from multiple organs form ever-expanding epithelial organoids that retain their tissue identity. We report the establishment of tumor organoid cultures from 20 consecutive colorectal (CRC) patients. For most, organoids were also generated from adjacent normal tissue. The organoids closely resemble the original tumor. The spectrum of genetic changes observed within the 'living biobank' agrees well with previous large-scale mutational analyses of CRC. Gene expression analysis indicates that the major CRC molecular subtypes are represented. Tumor organoids are amenable to robotized, high-throughput drug screens allowing detection of gene-drug associations. As an example, a single organoid culture was exquisitely sensitive to Wnt secretion (porcupine) inhibitors and carried a mutation in the negative Wnt feedback regulator RNF43 (rather than in APC). Organoid technology may fill the gap between cancer genetics and patient trials, complement cell line- and xenograft-based drug studies and allow personalized therapy design.
2015-05-07 | GSE64392 | GEO
Project description:In Rspondin-based three-dimensional cultures, Lgr5 stem cells from multiple organs form ever-expanding epithelial organoids that retain their tissue identity. Here we report the establishment of tumor organoid cultures from 20 consecutive colorectal carcinoma (CRC) patients. For most, organoids were also generated from adjacent normal tissue. Organoids closely resemble the original tumor. The spectrum of genetic changes within the 'living biobank' agrees well with previous large-scale mutational analyses of CRC. Gene expression analysis indicates that the major CRC molecular subtypes are represented. Tumor organoids are amenable to high-throughput drug screens allowing detection of gene-drug associations. As an example, a single organoid culture was exquisitely sensitive to Wnt secretion (porcupine) inhibitors and carried a mutation in the negative Wnt feedback regulator RNF43, rather than in APC. Organoid technology may fill the gap between cancer genetics and patient trials, complement cell line- and xenograft-based drug studies and allow personalized therapy design. Self-renewal of the intestinal epithelium is driven by Lgr5 stem cells located in crypts. We have recently developed a long-term culture system that maintains basic crypt physiology. Wnt signals are required for the maintenance of active crypt stem cells. Indeed, the Wnt agonist R-spondin1 induces dramatic crypt hyperplasia in vivo. R-spondin-1 is the ligand for Lgr5. Epidermal growth factor (EGF) signaling is associated with intestinal proliferation, while transgenic expression of Noggin induces a dramatic increase in crypt numbers. The combination of R-spondin-1, EGF, and Noggin in Matrigel sustains ever-expanding small intestinal organoids, which display all hallmarks of the original tissue in terms of architecture, cell type composition, and self-renewal dynamics. We adapted the culture condition for long-term expansion of human colonic epithelium and primary colonic adenocarcinoma, by adding nicotinamide, A83-01 (Alk inhibitor), Prostaglandin E2 and the p38 inhibitor SB202190. Of note, a two-dimensional culture method for cells from normal and malignant primary tissue has been described by Schlegel and colleagues. Here, we explore organoid technology to routinely establish and phenotypically annotate ‘paired organoids’ derived from adjacent tumor and healthy epithelium from CRC patients.
2015-05-07 | E-GEOD-65253 | biostudies-arrayexpress
Project description:A biobank collection of carotid plaque samples taken from patients undergoing endarterectomy operations.
2012-03-28 | GSE21545 | GEO