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. 2019 Oct 28;217(1):e20181184. doi: 10.1084/jem.20181184

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© 2019 Haeger et al.

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Figure 4. — Compromised tumor integrity and persistence of the invasion niche by RNAi- and antibody-based targeting of β1/β3 integrins in HT-1080 tumors. (A) Experimental procedure for administration of anti-β1 integrin mAb 4B4 or IgG1 and sequential intravital microscopy of the tumor response to integrin interference. Fluo, epifluorescence overview microscopy. MPM, subcellular-resolved multiphoton microscopy. (B) Time course of tumor growth or regression in control tumors transduced with empty vectors (p-puro/p-neo), β1RNAi or β1/β3RNAi in the absence or presence of IgG1 or anti-β1 integrin mAb 4B4. White arrowheads, onset of collective invasion. Numbers (right column), percentage mean regression of the tumor core (day 13 compared with day 6) from three to four independent tumors. Scale bars, 1 mm. (C) Fractions of mitotic and dead cells (day 6) quantified based on nuclear morphology for different interference schemes displayed as medians, 25th/75th percentiles (box), and 5th/95th percentiles (whiskers) from 20 independent fields from three to four independent tumors. Per condition, 19–20 nuclei were analyzed for the core and ∼10 nuclei for the invasion zone. *, P = 0.01; ***, P < 0.0001; ns, not significant. Statistics, Mann–Whitney U test (Bonferroni-corrected threshold: P = 0.0125). (D) Tumor development in response to the indicated interference procedures. Data show the means ± SD from three to four independent tumors. *, P = 0.0286. Statistics, Mann–Whitney U test. (E) Preferential survival of invading collective strands after combined β1/β3 integrin targeting. Z-projections of the same tumor region. Black box in upper left corner (day 6) results from stitching of adjacent images without complete overlap. Insets, mitotic figures in collective strands. Arrowheads, detachment of cell groups and individualized cells. Scale bar, 250 µm. (F) Median residual volume of tumor core and collective invasion (CI) zones after β1/β3RNAi combined with mAb 4B4 (day 13 compared with day 6) from three independent tumors. *, P < 0.05. Statistics, paired t test.