Computational identification of surface markers for isolating distinct subpopulations from heterogeneous cancer cell populations [scRNA-seq]
Ontology highlight
ABSTRACT: Intratumoral heterogeneity reduces treatment efficacy and complicates our understanding of tumor progression. There is a pressing need to understand how heterogeneous tumor cell subpopulations coexist within a tumor, yet biological systems to study these processes in vitro are limited. With the advent of single-cell RNA sequencing (scRNA-seq), it has been found that some cancer cell lines contain distinct subpopulations. These heterogeneous cell lines provide a unique opportunity to study coexistence and evolution between genetically similar cancer cell subpopulations in controlled experimental settings. Here, we present scEMD, a computational package that returns candidate surface makers maximally unique to transcriptomic subpopulations in scRNA-seq which may be used for FACS isolation. scEMD was experimentally validated using the MDA-MB-231 and MDA-MB-436 cell lines. ESAM and BST2/Tetherin were experimentally confirmed to identify and separate scRNA-seq cluster-matched subpopulations of MDA-MB-231 and MDA-MB-436 cells, respectively. Identification and enrichment of distinct subpopulations within cell line models using this computationally efficient and experimentally validated workflow paves the way for studies on the generation and maintenance of cellular heterogeneity in low-complexity in vitro systems.
Project description:Intratumoral heterogeneity reduces treatment efficacy and complicates our understanding of tumor progression. There is a pressing need to understand how heterogeneous tumor cell subpopulations coexist within a tumor, yet biological systems to study these processes in vitro are limited. With the advent of single-cell RNA sequencing (scRNA-seq), it has been found that some cancer cell lines contain distinct subpopulations. These heterogeneous cell lines provide a unique opportunity to study coexistence and evolution between genetically similar cancer cell subpopulations in controlled experimental settings. Here, we present scEMD, a computational package that returns candidate surface makers maximally unique to transcriptomic subpopulations in scRNA-seq which may be used for FACS isolation. scEMD was experimentally validated using the MDA-MB-231 and MDA-MB-436 cell lines. ESAM and BST2/Tetherin were experimentally confirmed to identify and separate scRNA-seq cluster-matched subpopulations of MDA-MB-231 and MDA-MB-436 cells, respectively. Identification and enrichment of distinct subpopulations within cell line models using this computationally efficient and experimentally validated workflow paves the way for studies on the generation and maintenance of cellular heterogeneity in low-complexity in vitro systems.
Project description:We performed gene expression profiling on the MDA-MB-231 and MDA-MB-436 human breast cancer cell lines following siRNA-mediated inhibition of Fn14 expression as an approach to identify the mechanistic basis for Fn14 regulation of invasive capacity. Keywords: siRNA-mediated inhibition
Project description:In order to identify the microRNAs differentially expressed according to the estrogen receptor status, total RNAs in triplicate from six breast cancer cell lines with different ERalpha status (ER+: T47D, BT-474, MDA-MB-483; ER-: MDA-MD-436, MDA-MB-231, MDA-MB-468) was extracted by using Trizol reagent and used for microarray experiments.
Project description:Five Triple Negative Breast Cancer cell lines were exposed to increasing concentration of Paclitaxel untill they acquired resistance. In order to identify changes in gene expression associated with resistance to PTX, we performed gene expression profiling on parental and resistant cell lines. Of ~22000 genes surveyed by microarray analysis, 5.0%, 3.7%, 9.0%, 7.3%, and 5.4% of the genes showed changes in expression of 2-fold or greater (p value < 0.05) in BT20, SUM149, MDA-MB-231, MDA-MB-436 and MDA-MB-468 cell lines, respectively.
Project description:Using CRISPR-Cas9 technology, we stably disrupted an insulator element in two different TNBC cell lines. The aim of this experiment was to characterize the chromatin accessibility profile using ATAC-seq. For this experiment, we selected three biological replicates of MDA-MB-231 (WT and IE8dis) and MDA-MB-436 (WT and IE8dis).
Project description:We performed gene expression profiling on the MDA-MB-231 and MDA-MB-436 human breast cancer cell lines following siRNA-mediated inhibition of Fn14 expression as an approach to identify the mechanistic basis for Fn14 regulation of invasive capacity. Experiment Overall Design: Each cell lines was transfected with etiher an Fn14 or firefly luciferase-specific (GL2) siRNA, with untreated cells used as a reference. 2 biological replicates were performed for each array, independently grown, treated and harvested.
Project description:The project profiled the expression patterns in hypoxia induced secretomes between MDA-MB-231 parental and MDA-MB-231 Bone Tropic (BT) breast cancer cell lines which have been previously generated by Massague and colleagues (Kang et al. Cancer Cell 2003).
Project description:Indole-3-carbinol (I3C) is a natural anti-carcinogenic compound found at high concentrations in Brassica vegetables. ER-positive cell lines demonstrated the greatest sensitivity to the anti-tumor effects of I3C compared to ER-negative breast cancer cell lines. Gene expression analysis was performed to identify genes and pathways that accounted for sensitivity to I3C. Microarray analysis performed using Illumina HT-12 v4 expression arrays A total of 36 samples were analyzed with six breast cancer cell lines treated with either the vehicle control or the drug Indole-3-carbinol in triplicate. The cell lines were: MCF-7, T47D, ZR751(sensitive to the drug, apoptosis/growth arrest) and MDA-MB-231, MDA-MB-157, and MDA-MB-436 (insensitive to the drug). Sensitive cell lines are of the luminal subtype and insensitive cell lines are of the basal subtype.
Project description:The project profiled the expression patterns in hypoxia induced secretomes between MDA-MB-231 parental and MDA-MB-231 Bone Tropic (BT) breast cancer cell lines which have been previously generated by Massague and colleagues (Kang et al. Cancer Cell 2003).