Figure 5

Epidermal growth factor receptor (EGFR) signaling in mice co-injected with SUM149 cells and mesenchymal stem/stromal cells (MSCs) is decreased by erlotinib treatment. SUM149 cells (5 × 10⁵) were injected bilaterally into the cleared mammary fat pad of SCID/Beige mice with or without MSCs (10% of total number of cells injected per mammary gland, see Scheme 1). Mice were treated with erlotinib diet (dose of 40 mg/kg per day). Treatment started on day 1 following injection of cells and ended 17 weeks later. Tumors were resected in a survival surgery when tumors reached a volume of 300 mm³, fixed in formalin, embedded in paraffin, cut, stained and scored by a pathologist specialized in inflammatory breast cancer (IBC). Metastases were collected and processed as the tumors 8 weeks post-resection of primary tumors. (A) Quantification and comparison of E-cadherin staining between control (untreated) and erlotinib-treated groups of mice that were injected only with SUM149 cells. E-cadherin staining of tumor sections from 0% MSC group treated with erlotinib was lower than in untreated 0% MSC group (control diet) (72.0% vs. 10.1%). ^*** P = 0.001, Student’s t test. (B) Quantification and comparison of E-cadherin staining between control (untreated) and erlotinib-treated groups of mice that were co-injected with SUM149 cells and MSCs. E-cadherin staining of tumor sections from 10% MSC group treated with erlotinib was lower than in untreated 10% MSC group (control diet) (70.0% vs. 14.6%). ^*** P = 0.001, Student’s t test. (C) Quantification and comparison of Ki-67 staining between control (untreated) and erlotinib-treated groups of mice that were injected only with SUM149 cells. Ki-67 staining of tumor sections from 0% MSC group treated with erlotinib was lower than in untreated 0% MSC group (control diet) (39.0% vs. 8.9%). ^*** P = 0.001, Student’s t test. (D) Quantification and comparison of Ki-67 staining between control (untreated) and erlotinib-treated groups of mice that were co-injected with SUM149 cells and MSCs. Ki-67 staining of tumor sections from 10% MSC group treated with erlotinib was lower than in untreated 10% MSC group (control diet) (61.8% vs. 15.2%). ^*** P = 0.001, Student’s t test. (E) Quantification and comparison of phospho-EGFR (p-EGFR) staining between control (untreated) and erlotinib-treated groups of mice that were injected only with SUM149 cells. p-EGFR staining of tumor sections from 0% MSC groups was similar regardless of treatment with erlotinib (49.4% vs. 53.6%). P = NS, Student’s t test. (F) Quantification and comparison of p-EGFR staining between control (untreated) and erlotinib-treated groups of mice that were co-injected with SUM149 cells and MSCs. p-EGFR staining of tumor sections from 10% MSC group treated with erlotinib was lower than in untreated 10% MSC group (control diet) (69.3% vs. 44.0%). ^*** P = 0.03, Student’s t test. (G) Ki-67 staining of metastasis section from 10% MSC group in control diet. (H) Ki-67 staining of metastasis section from 10% MSC group in erlotinib diet. (I) p-EGFR staining of metastasis section from 10% MSC group in control diet. (J) p-EGFR staining of metastasis section from 10% MSC group in erlotinib diet. Scale bar is 100 μm in all images.