Journal: iScience

Article Title: Pseudo-temporal dynamics of chemoresistant triple negative breast cancer cells reveal EGFR/HER2 inhibition as synthetic lethal during mid-neoadjuvant chemotherapy

doi: 10.1016/j.isci.2023.106064

Figure Lengend Snippet: Pseudo-temporal dynamics of single-cell transcriptome during neoadjuvant chemotherapy (NAC) (A) tSNE plot of cell clusters and associated inferred lineages (I, II and III). The dotted arrows show the inferred tumor trajectories across respective cell clusters labeled by numbers. (B) Heatmap of cluster markers. The log2(TPM +1) of cluster marker expressions across all nuclei are shown. The top panel shows clinical and molecular annotations for individual nuclei. From the top, the first row depicts cell trajectory assignments, the second row shows cell cluster assignments, the third row shows cell groups as defined by patients’ responses (S: chemo-sensitive, R: chemoresistant) and NAC stages (B: pre-NAC, M: mid-MAC, A: post-NAC), the fourth shows the patients from which the nuclei were sourced, and the fifth shows molecular subtypes by PAM50 classifications. On the right, chemo-sensitizing or resistance-promoting gene knockouts from genome-wide CRISPR/Cas9 screening in MCF10A cells are marked (chemo-sensitizing hits: green, resistance-promoting hits: red). The first 5 rows in the bottom panel show key cell type marker genes (epithelial tumor: EPCAM, CDH5, immune cells: CD45, cancer stem-cell marker: THY1, hormone receptors/Her2: ESR1, PGR, and ERBB2 ). The bottom three rows show inferred pseudo-time stamps on individual nuclei per trajectory in A. (C) Pie charts of cell cluster compositions across different NAC stages and NAC sensitivity. Each pie size represents the proportion of cells coming from chemo-responsive (S) or –resistant (R) patients at different stages of NAC (B: pre-NAC, M: mid-MAC, A: post-NAC). The color codes are shown in the bottom legend. (D) Enrichments of chemo-sensitizing (left) or resistance-promoting (right) gene knockouts from genome-wide CRISPR/Cas9 screening for Doxorubicin (top) and Paclitaxel (bottom). Each dot shows enrichment of the screening signatures as evaluated by Gene Set Variation Analysis (GSVA). We applied GSVA Z - score >3 or <−3 thresholds (horizontal red lines) to identify significant enrichment or depletion of the knockout signatures in the cellwise transcriptome. (E) Summary of pseudo-temporal changes in cell populations along trajectory-I. Top panel: Cells by different treatment stages, Bottom panel: Cells by PAM50 molecular subtypes. C9 is marked as the primary cell cluster of interest with therapeutic potentials as indicated by CRISPR-Cas9 screening results in D.

Article Snippet: CRISPR genome-wide loss-of-function (LOF) screens were performed using human CRISPR knockout pooled libraries GeCKOv2 (Addgene Cat #1000000048) and Brunello (Addgene Cat #73179) using a well-established protocol.

Techniques: Labeling, Marker, Genome Wide, CRISPR, Knock-Out

Journal: iScience

Article Title: Pseudo-temporal dynamics of chemoresistant triple negative breast cancer cells reveal EGFR/HER2 inhibition as synthetic lethal during mid-neoadjuvant chemotherapy

doi: 10.1016/j.isci.2023.106064

Figure Lengend Snippet:

Article Snippet: CRISPR genome-wide loss-of-function (LOF) screens were performed using human CRISPR knockout pooled libraries GeCKOv2 (Addgene Cat #1000000048) and Brunello (Addgene Cat #73179) using a well-established protocol.

Techniques: Recombinant, Protease Inhibitor, Cell Viability Assay, Sequencing, Gene Expression, Software