Journal: bioRxiv
Article Title: USP7 inhibition perturbs proteostasis and tumorigenesis in triple negative breast cancer
doi: 10.1101/2025.01.28.635372
Figure Lengend Snippet: (A) Western Blot analysis showing USP7 protein expression levels in malignant breast cancer cells, including TNBC, compared to the non-malignant MCF10A mammary epithelial cell line used as control. Quantification relative to MCF10A expression shown at the bottom after β-actin normalization. β-actin was used as loading control. (B) Representative Western blot analysis of USP7 in CRISPR/Cas9-mediated USP7 knockout (USP7 KO) clones of TNBC cell lines in comparison to non-targeting vector control (Ctrl). β-actin was used as loading control. (C) Bar graph of colony counts from clonogenic assays comparing USP7 KO to Ctrl. (D) Chemical structures of USP7 inhibitors that were used for in vitro assays. (E) Representative images of colonies stained with crystal violet comparing SUM159 treated with XL177A or FT671. DMSO used as vehicle control. (F) Bar graph of colony counts from clonogenic assays comparing TNBC cells treated with XL177A or FT671. DMSO as control. (G) Representative image from scratch-wound assay comparing wound closure of SUM159 non-targeting vector control (Ctrl) and USP7 KO clone over 8 hours. White lines represent the wound edge. (H) Quantification of the distance cells migrated in scratch-wound assays for each TNBC cell line. (I) Representative images from Transwell invasion assays with cells stained with crystal violet comparing SUM159 Ctrl and USP7 KO clone. (J) Quantification of the number of cells invaded across a Transwell membrane for each of the TNBC cell lines. (K) Quantification of the distance cells migrated for TNBC cells treated with XL177A or FT671. DMSO used as vehicle control. (L) Quantification of cell invasion for TNBC cells treated with XL177A or FT671. DMSO used as vehicle control. (M) Quantification of the distance cells migrated normalized to DMSO for MDA-MB-231 Ctrl and USP7 KO. (N) Quantification of cell invasion normalized to DMSO for MDA-MB-231 Ctrl and USP7 KO. For all bar graphs, values were normalized to control. Each bar is shown as mean ± SD, with each dot representing an independent replicate, n= 3. For all graphs: ns=not significant, *p<0.05, **p<0.01, ***p<0.001, ****p<0.0001 by unpaired two-tailed t test.
Article Snippet: Tissue microarray (TMA) slides containing 110 TNBC cases were obtained from US Biomax (BRE1201; TissueArray.com ).
Techniques: Western Blot, Expressing, Control, CRISPR, Knock-Out, Clone Assay, Comparison, Plasmid Preparation, In Vitro, Staining, Scratch Wound Assay Assay, Membrane, Two Tailed Test