Fig. 1: In vivo epigenetic screening implicates PBAF and BRD7 in breast cancer lung metastatic dormancy.

A Design of in vivo loss of function screen. After transduction with the mouse epigenetic shRNA libraries, the 4T07-TGL cells were injected i.v. into syngeneic Balb/cJ mice. Candidate suppressors of metastasis were ascertained by sequencing analysis of lung metastatic lesions and were further validated by additional shRNA studies. B, C 4T07-TGL cells with scrambled sgRNA vector (sgCtr) or Brd7 knockout were injected i.v. into syngeneic mice. (4T07-TGL-sgctr, n = 15 mice; 4T07-TGL-Brd7 KO1, n = 15 mice; 4T07-TGL-Brd7 KO2, n = 15 mice). The extent of lung colonization was measured by bioluminescent imaging. Panels show representative images (B), and quantified normalized photon flux at the indicated time points, error bars represent SD (C). D Kaplan-Meier survival analysis for mice injected with the indicated cell lines. E Representative H&E-stained images of lungs extracted from mice injected with the indicated cell lines at the experimental endpoint. F Quantified lung metastatic lesions in control and Brd7 KO mice (mean ± SEM.; 4T07-TGL-sgctr, n = 15 mice; 4T07-TGL-Brd7 KO1, n = 15 mice; 4T07-TGL-Brd7 KO2, n = 15 mice). Error bars represent SEM. G Representative images of lung sections subjected to IF for the indicated markers. Control (sgCtr) and Brd7 knockout (KO1 and KO2) 4T07-TGL cells were inoculated i.v., and lung sections stained with anti-Ki-67, anti-GFP and DAPI (G). H Bar graph showing the percentage of tumor cells with positive Ki-67 IF at the indicated times. (4T07-TGL-sgctr, n = 5 mice; 4T07-TGL-Brd7 KO1, n = 5 mice; 4T07-TGL-Brd7 KO2, n = 5 mice) *p < 0.05, **p < 0.01. Error bars represent SEM. I, J Kaplan-Meier analysis of distant metastasis-free survival (I) and relapse-free survival (J) in publicly available breast cancer datasets of the indicated subtype (source: KM Plotter for breast cancer). Patients were divided according to BRD7 expression level with the cutoff at 50%. HR, hazard ratio. K PBAF subunit (BRD7, PBRM1, and ARID2) expression in breast cancer primary and metastatic tumors; dataset indicated. *p < 0.05, **p < 0.01. [BRD7: Primary (n = 15 patients)- Minima = 6.966, Maxima = 10.996, Median expression = 9.627, Metastasis (n = 60 patients) – Minima = 8.380, Maxima = 10.759, Median expression = 9.354; PBRM1: Primary (n = 15 patients)- Minima = 10.087, Maxima = 12.162, Median expression = 11.349, Metastasis (n = 60 patients)- Minima = 9.471, Maxima = 12.045, Median expression = 10.115; ARID2: Primary (n = 15 patients)- Minima = 8.755, Maxima = 12.656, Median expression = 11.490, Metastasis (n = 60 patients)- Minima = 8.272, Maxima = 11.718, Median expression = 9.574 (L) PBAF specific subunit expression in normal breast tissue, breast primary tumor and metastases from TCGA invasive breast cancer dataset [Normal (n = 113 patients)- Minima = 775,Maxima = 5805, Median expression = 2018; Primary (n = 1210 patients)- Minima = 110,Maxima = 10285, Median expression = 1563; and Metastasis (n = 7 patients)- Minima = 611,Maxima = 1960, Median expression = 836). M Representative immunohistochemistry (IHC) staining for paired primary tumor and lung metastasis sections in breast cancer tissue microarrays. BRD7 expression was quantified by stain intensity across n = 3 independent tumor regions obtained from n = 1 female breast cancer patient. ****p < 0.0001. Statistical analysis by unpaired two-tailed t test, in all panels, error bars represent SD. 95% CI was used to analyze the results. Source data are provided as a Source Data file.