Journal: iScience
Article Title: Epigenomic analysis identifies DTP subpopulation using HOPX to develop targeted therapy resistance in lung adenocarcinoma
doi: 10.1016/j.isci.2025.112387
Figure Lengend Snippet: Deletion of HOPX impairs the progress of DTP regrowth (A) Western blot showing reduction of HOPX expression in PC9 cells by CRISPR-Cas9 mediated deletion of HOPX gene. (B) Cell growth curve of PC9 cells transfected with sgRNAs targeting HOPX or non-target sgRNAs. (C) Cell viability of PC9 cells transfected with sgRNAs targeting HOPX or non-target sgRNAs under the treatment of 150nM osimertinib at indicated time points (normalized to day0). Right: Bar graph showing cell viability at day 28 ( n = 4, including duplicates for sgRNA1 and duplicates for sgRNA2). (D) Colony formation assay on PC9 cells, at seeding density of 150,000 cells/well under 150nM osimertinib starting at 24 h after seeding. Left: Two representative images taken at 20 days after treatment per condition. Right: Quantification of colony numbers based on the representative fields shown on the left ( n = 4), including duplicates for sgRNA1 and duplicates for sgRNA2. (E) Western blot showing reduction of HOPX expression in NCI-H358 cells. (F) Cell growth of NCI-H358 cells transfected with sgRNAs targeting HOPX or non-target sgRNAs. (G) Cell viability of NCI-H358 cells transfected with sgRNAs targeting HOPX or non-target sgRNAs under the treatment of 500nM sotorasib at indicated time points (normalized to day 0). Right: Bar graph showing cell viability at day 25 ( n = 4, including duplicates for sgRNA1 and duplicates for sgRNA2). (H) Colony formation assay on NCI-H358 cells, at seeding density of 150,000 cells/well under 500nM sotorasib starting at 24 h after seeding. Left: Two representative images taken at 20 days after treatment per condition. Right: Quantification of colony numbers based on the representative fields shown on the left ( n = 4), including duplicates for sgRNA1 and duplicates for sgRNA2. Data are presented as means ± S.D. ( n = 4). Statistical significance was determined using Mann-Whitney test. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001.
Article Snippet: Cells stably expressing human codon-optimized S. pyogenes Cas9 expression were generated by infection with the lentiCas9-Blast plasmid (a gift from Feng Zhang, Addgene #52962). sgRNAs targeting HOPX (selected from Brunello library) and non-target sgRNAs (selected from the Gecko library v2) were synthesized by Integrated DNA Technologies and listed in sgRNAs were cloned into lentiGuide-Puro (Addgene #52963) at Esp3I site.
Techniques: Western Blot, Expressing, CRISPR, Transfection, Colony Assay, MANN-WHITNEY
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