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Single-cell RNA-seq of OVCAR3 and CCL218 cancer cells with evolved hydroxychloroquine resistance

ABSTRACT: OVCAR3 and CCL218 cells were passaged in the presence or absence of hydroxychloroquine. Drug pulse chases were used, with 48h of drug exposure followed by replacement of fresh media. Cells were allowed to grow to near confluence and then another drug pulse-chase was performed. After 15 hydroxychloroquine pulse-chases, cells were allowed to proliferate for 7 days in drug-free media so that transient drug effects were no longer present.

ORGANISM(S): Homo sapiens

PROVIDER: GSE245329 | GEO | 2024/03/18

REPOSITORIES: GEO

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RNA-seq of OVCAR3 and CCL218 cancer cells with evolved hydroxychloroquine resistance

Project description:OVCAR3 and CCL218 cells were passaged in the presence or absence of hydroxychloroquine. Drug pulse chases were used, with 48h of drug exposure followed by replacement of fresh media. Cells were allowed to grow to near confluence and then another drug pulse-chase was performed. After 15 hydroxychloroquine pulse-chases, cells were allowed to proliferate for 7 days in drug-free media so that transient drug effects were no longer present. Cells were then pelleted and cell pellets were processed for concomitant bulk RNA-seq, whole-exome-sequencing, and single-cell RNA-seq. This project represents the bulk transcriptomic sequencing and analysis.

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Project description:A Brunello CRISPR-Cas9 library screen was performed with OVCAR3 and CCL218 cells in the presence or absence of hydroxychloroquine. Drug pulse chases were used, with 48h of drug exposure followed by replacement of fresh media. Cells were allowed to grow to near confluence and then another drug pulse-chase was performed. After each pulse-chase, a pool of cells were collected for sgRNA quantitation. Three pulse chases were performed. Cell pellets were processed for genomic DNA and sgRNA sequences amplified by PCR for NGS-based quantitation. Once quantified by NGS analysis, MAGeCK MLE analysis was performed to determine which sgRNAs were most associated with resistance or sensitization to hydroxychloroquine.

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Thiouracil Pulse-chase, Tachyzoites and Bradyzoites mRNA stability data

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Hydroxychloroquine inhibits trained immunity

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Hydroxychloroquine inhibits trained immunity

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Project description:To investigate the molecular pathways underlying RNASET2-mediated tumor suppression in vivo, we investigated the gene expression profile of tumor samples obtained from control and RNASET2-silenced OVCAR3 xenografts, respectively. In order to evaluate the contribution of human cancer cells and the murine host stroma to RNASET2-mediated tumorigenesis responses, total RNA extracted from tumor xenografts derived from six independent OVCAR3 cell clones (three parental and three RNASET2-silenced clones) was fluorescently labeled and hybridized to human Agilent whole-genome microarrays. Control and RNASET2-silenced OVCAR3 were injected subcutaneously into nude mice. After 39 days mice were sacrified, tumors were extracted and total RNA purified.

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