A slow cycling Lgr5 tumour population mediates basal cell carcinoma relapse after therapy
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ABSTRACT:
PROVIDER: PRJNA482149 | ENA |
REPOSITORIES: ENA
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Project description:We used microarray analysis to molecularly profile distinct tumour cell populations present in untreated and vismodegib-treated basal cell carcinomas.
2018-10-11 | GSE117458 | GEO
Project description:Basal cell carcinoma (BCC) is the most frequent cancer in humans and results from constitutive activation of the Hedgehog pathway1. Several Smoothened inhibitors are used to treat Hedgehog-mediated malignancies, including BCC and medulloblastoma2. Vismodegib, a Smoothened inhibitor, leads to BCC shrinkage in the majority of patients with BCC3, but the mechanism by which it mediates BCC regression is unknown. Here we used two genetically engineered mouse models of BCC4 to investigate the mechanisms by which inhibition of Smoothened mediates tumour regression. We found that vismodegib mediates BCC regression by inhibiting a hair follicle-like fate and promoting the differentiation of tumour cells. However, a small population of tumour cells persists and is responsible for tumour relapse following treatment discontinuation, mimicking the situation found in humans5. In both mouse and human BCC, this persisting, slow-cycling tumour population expresses LGR5 and is characterized by active Wnt signalling. Combining Lgr5 lineage ablation or inhibition of Wnt signalling with vismodegib treatment leads to eradication of BCC. Our results show that vismodegib induces tumour regression by promoting tumour differentiation, and demonstrates that the synergy between Wnt and Smoothened inhibitors is a clinically relevant strategy for overcoming tumour relapse in BCC.
| S-EPMC6295195 | biostudies-literature
Project description:SUMMARY: Dormant or slow cycling tumour cells can form a residual chemoresistant reservoir responsible for relapse in patients, years after curative surgery and adjuvant therapy. Since their isolation from human tumors has been a long-stalled technical challenge preventing a direct study of their functional properties, we have adapted the pulse-chase expression and labeling approach using H2BeGFP (Tumbar et al., 2004, DOI:10.1126/science.1092436) to a doxycycline-inducible all-in-one lentiviral vector. In this manner we were able to label and isolate by FACS live slow cycling cancer cells (SCCC) from human colorectal, glioma and melanoma patient-derived tumors as well as from minitumors grown in matrigel (SW1222-cells) or in sphere culture (e225, e216 glioma models). We determined the differentially expressed transcriptional profiles in paired comparisons of slow-cycling cancer cell (SCCC) versus rapid cycling cancer cell (RCCC) sub-populations. H2BeGFP-retention revealed slow cycling as a transient cell state rather than a cellular entity. SCCC showed cancer-initiation potential and enhanced chemoresistance. Cells in this slow cycling status presented a distinctive non-genetic and cell-autonomous gene expression profile shared across different tumour types. We identified TET2 epigenetic enzyme as key factor controlling SCCC numbers and survival. 5-Hydroxymethylcytosine (5hmC), generated by TET2 enzymatic activity, labelled the SCCC genome in carcinomas and was a predictive biomarker of relapse and survival in cancer patients. We demonstrate the enhanced chemoresistance of SCCC, revealed 5hmC as a biomarker for their clinical identification, and identified TET2 as a potential drug-target for SCCC elimination that could extend patients' survival.Experimental Design: Cells from human patient-derived xenografts (PDX) and human cancer cell lines were infected with a doxycycline (DOX)-inducible all-in-one lentivirus coding for the histone2B-eGFP fusion protein. The models were as following: T70 (colorectal cancer); MMPG3 and MMLN9 (melanoma); e225 and e216 (glioma) and human SW1222 colorectal cancer cell line as an infected pool as well as a clone (Clone2) and the following SW1222 derivate cell lines: SW1222-shCTRL (stable expression of nonsilencing shRNA), SW1222-shTET2 (stable expression of TET2-specific shRNA).H2BeGFP-infected cells were isolated and grown/propagated and injected subcutaneously in the flanks of immunocompromised NOD-Scid mice as described (Puig et al., Clin. Cancer Res. 2013; doi: 10.1158/1078-0432.CCR-12-1740.). H2BeGFP-infected SW1222 cells and derivate cell lines were grown in matrigel drops as minitumors. H2BeGFP-infected patient-derived melanoma MMPG3 and MMLN9 minitumor and glioma models e216 and e225 were grown as sphere cultures in their respective conditions as indicated.H2BeGFP expression was induced by DOX treatment in all experimental systems in a pulse-chase fashion and H2BeGFP fusion protein integrated into the host genome labeling all cells. Consecutive cell divisions of proliferating cells diluted and finally eliminated the fluorescent mark in an exponential decay fashion (RCCC or sRCCC), whereas slow-cycling cells (SCCC) retained the fluorescent stain, allowing FACS separation and isolation. Total RNA was separately extracted from both sub-populations of cells of each system and submitted to microarray analyses for characterization.
2018-06-28 | E-MTAB-4004 | biostudies-arrayexpress
Project description:Gastric cancer is amongst the most prevalent and deadliest cancers globally. To derive mechanistic insight into the pathways governing this disease, we generated a stomach-specific Claudin18-IRES-CreERT2 mouse line to model advanced human gastric cancer and investigated the role of tumour resident Lgr5+ stem-like cells in gastric cancer. Mouse lines incorporating Cldn18-driven conditional dysregulation of the Wnt, RTK and Trp53 pathways modelled metastatic Chromosomal Instable-type gastric cancer. In parallel, we supplemented these cancer models with orthotopic transplantation of cancer organoids to derive detailed mechanistic insight into the roles of Lgr5+ cancer stem cell in gastric cancer. Here we show, we recapitulated advanced human gastric cancer in terms of disease phenotype, aberrant gene expression profile, molecular markers, and sites of distant metastases. Importantly, we established that tumour-resident Lgr5+ stem-like cells are critical to the initiation and maintenance of tumour burden, and are obligatory for the establishment of metastases. These models will be invaluable for deriving clinically-relevant mechanistic insights into cancer progression and as pre-clinical models for evaluating novel therapeutics.
2021-11-26 | GSE184613 | GEO
Project description:Epimorphic regeneration commonly relies on the activation of quiescent stem cells to drive new cell production. The planarian Schmidtea mediterranea is among the best regenerators in nature, thanks to its large population of adult stem cells, called neoblasts. While neoblasts have long been known to drive regeneration, whether a subset of neoblasts is reserved for this purpose is unknown. Here, we revisited the idea of reserved neoblasts by approaching neoblast heterogeneity from a regulatory perspective. By implementing a new fluorescence-activated cell sorting strategy in planarians, we identified a population of neoblasts defined by low transcriptional activity. These RNAlow neoblasts are relatively slow-cycling at homeostasis and undergo a morphological regeneration response characterized by cell growth at 48 hours post-amputation. At this time, RNAlow neoblasts proliferated in a TOR-dependent manner. Additionally, knockdown of the tumour suppressor Lrig-1, which is enriched in RNAlow neoblasts, resulted in RNAlow neoblast growth and hyperproliferation at homeostasis, and ultimately delayed regeneration. We propose that slow-cycling RNAlow neoblasts represent a regeneration-reserved neoblast population.
2020-12-31 | GSE157860 | GEO
Project description:A slow cycling, dormant cell state is thought to contribute to the therapeutic resistance of GBM tumour cells, yet not much is known about the biology of this slow-cycling cell population in GBM. By using somatic mouse model of GBM, combined with the inducible H2B-GFP label retention system, we isolated dormant tumour cells from their in vivo niches, and characterised them by single cell RNA sequencing.
2023-04-03 | GSE228785 | GEO
Project description:We identified slow-cycling cells (SCCs) in Ewing sarcoma using a label retention assay with CFSE. We labeled cells of SK-ES-1, an Ewing sarcoma cell line, with CFSE. After 5 days culture, we isolated cells retaining strong fluorescence (upper, ~10%) as SCCs and other cells (lower, ~90%) as non-slow-cycling cells (non-SCCs) using FACS AriaTM Ⅲ cell sorter.
2022-10-17 | GSE204671 | GEO
Project description:We analyzed gene expression of 3 lines LGR5-GFP, 2 lines KRT20-GFP knock-in colorectal tumor organdies. The cancer stem cell (CSC) theory highlights a self-renewing subpopulation of cancer cells that fuels tumour growth. The existence of human CSCs is mainly supported by xenotransplantation of prospectively isolated cells, but their clonal dynamics and plasticity remain unclear. Here, we show that human LGR5+ colorectal cancer cells serve as CSCs in growing cancer tissues. Lineage-tracing experiments with a tamoxifen inducible Cre knock-in allele of LGR5 reveal the self-renewal and differentiation capacity of LGR5+ tumour cells. Selective ablation of LGR5+ CSCs in LGR5-iCaspase9 knock-in organoids leads to tumour regression, followed by tumour regrowth driven by re-emerging LGR5+ CSCs. KRT20 knock-in reporter marks differentiated cancer cells that constantly diminish in tumour tissues, while reverting to LGR5+ CSCs and contributing to tumour regrowth after LGR5+ CSC ablation. We also show that combined chemotherapy potentiates LGR5+ CSCs targeting. These data provide insights into the plasticity of CSCs and their potential as a therapeutic target in human colorectal cancer.
2017-03-30 | GSE83513 | GEO
Project description:Molecular stratification using gene-level transcriptional data has identified subtypes with distinctive genotypic and phenotypic traits, as exemplified by the consensus molecular subtypes (CMS) in colorectal cancer (CRC). Here, rather than gene-level data, we make use of gene ontology and biological activation state information for initial molecular class discovery. In doing so, we defined three pathway-derived subtypes (PDS) in CRC; PDS1 tumours are canonical/LGR5+ stem-rich, highly proliferative and display good prognosis. PDS2 which is regenerative/ANXA1+ stem-rich, with elevated stromal/immune tumour microenvironmental lineages. PDS3 tumours represent a previously overlooked slow-cycling subset of tumours within CMS2, with reduced stem populations and increased differentiated lineages, particularly enterocytes and enteroendocrine cells, yet display the worst prognosis in locally advanced disease. These PDS3 phenotypic traits are evident across numerous bulk and single-cell datasets, and demark a series of subtle biological states that are under-represented in pre-clinical models currently and are not identified using existing subtyping classifiers Mutations: https://www.s-cort.org/sites/default/files/exports/scort_spinal_focus_paper/ws4_spinal_mutations.txt, CNA per gene : https://www.s-cort.org/sites/default/files/exports/scort_spinal_focus_paper/ws4_spinal_cna_gene_subselection_for_paper.txt, CNA segments SPINAL https://www.s-cort.org/sites/default/files/exports/scort_spinal_focus_paper/ws4_spinal_cna.zip, CNA segments FOCUS https://www.s-cort.org/sites/default/files/exports/scort_spinal_focus_paper/ws2.focus_cna.zip
2023-12-12 | GSE248381 | GEO
Project description:The identification of Lgr5 as an intestinal stem cell marker has made it possible to isolate and study primary stem cells from small intestine. Using the cell cycle specific expression og the mKi67 gene, we generated a novel Ki67-RFP knock-in allele which identifies dividing cells. Using Lgr5-GFP;Ki67-RFP mice, we isolated CBCs with distinct Wnt signaling levels and cell cycle features, and analyzed their global gene expression pattern using microarrays. We concluded that the cycling Lgr5hi stem cells exit the cell cycle in transition into the secretory lineage. Lgr5med Ki67low intermediate precursors reside in the zone of differentiation, resemble quiescent stem cells and generate the Dll1+ secretory precursors and the label retaining cells. Our findings support the cycling stem cell hypothesis and highlight the heterogeneity of early progenitors during lineage commitment. We used cell fractions of intestines from Lgr5-EGFP-ires-CreERT2 mice, expressing GFP under the control of the Lgr5 promoter, and Ki67-TagRFP mice where the RFP is fused to the C-terminus of the endogenous Ki67 gene. RNA was isolated from several FACS sorted cell populations of combinations expressing different levels of GFP and RFP: GFP high RFP high (Lgr5hi Ki67hi), GFP high RFP low (Lgr5hi Ki67low), GFP medium RFP high (Lgr5med Ki67high) and GFP medium RFP low (Lgr5med Ki67low). Purified RNA was processed, hybridized, and scanned according to the manufacturerM-bM-^@M-^Ys protocol and were hybridized on Affymetrix Mouse Gene ST 1.1 arrays).
2014-09-08 | E-GEOD-52813 | biostudies-arrayexpress