Figure 3: LYN inhibition prevents ECM stiffness-induced cell invasion and metastatic dissemination in vivo.

A-F) MCF10A cells were grown on 3D-PA gels for 5 days in the presence of 1 μM Bafetinib, Nilotinib or vehicle control.

A) Invasive structures were quantified and represented as percentages of total structures. Data are presented as mean +/− sd, dots represent individual fields.

B) PLA (green dots) was performed to detect TWIST1-G3BP2 endogenous interaction, DAPI is in blue. Scale bar = 25 μm.

C) Quantification of the PLA intensity per cell in B. A.U.: arbitrary units. Data are presented as mean +/− sd, dots represent individual fields.

D) Cells were immunostained for E-cadherin (green), DAPI (blue) and fibronectin (FN, red).

E) Cells were immunostained for TWIST1 (green), G3BP2 (red) and DAPI (blue). Scale bar = 25 μm. Inserts show high magnification images.

F) TWIST1 nuclear signal in (E) is presented as the percentage of total TWIST1 staining on a violin plot showing the distribution of the data with the median (hyphen line) and quartiles (dotted line).

G) Schematic depicting the protocol used for in vivo experiments.

H) Tumor weight (mg) at the end of the experiment are presented as mean +/− sd, dots represent individual animals. N=14 tumors per group.

I) Local (mammary gland, MG) or regional (peritoneum) invasion of the primary tumor. Data are presented as percentage. N=14 mice per group.

J) Representative images of the primary tumor invasion (T: tumor; P: peritoneum). Arrow shows GFP-positive invasive tumor cells. Scale bar = 5 mm.

K) Lung metastatic burden presented as the number of GFP+ nodules in the lungs. Data are presented as mean +/− sd, dots represent individual animals. N=14 mice per group.

L) Representative images. Arrows show metastatic tumor cells. Scale bar = 1 mm.

*** p < 0.001; ns: not significant.

See also Figure S3.