NSCLC metastases failed to show vascular disruption in response to DMXAA.

<p>(<b>A</b>) BLI of metastatic 344SQ-ELuc tumors prior to DMXAA or DMSO administration and again at 6 and 24 hours (N = 6 and 8 respectively). Whole body regions of interest (ROIs) demonstrate no loss of BLI (<b>B</b>). (<b>C</b>) Representative kidney metastases (tumor = T, kidney = K) did not show evidence of hemorrhagic necrosis after DMXAA treatment. (<b>D–E</b>) BLI of 344SQ-ELuc subcutaneous tumors at day 7 (N = 6) demonstrated a considerable drop in photon emission rates after DMXAA in mice with smaller tumors (*p<0.05), and the latter were accompanied by evidence of hemorrhagic necrosis (<b>F</b>).</p

Evidence of DMXAA-mediated macrophage repolarization <i>in vivo</i>.

<p>(<b>A</b>) Spleen lysates from mice treated with 25 mg/kg DMXAA (N = 3), versus DMSO vehicle (N = 4) demonstrated decreased Arg-1, and elevated iNOS, transcripts. (<b>B</b>) Representative histology of spleen showing Arg-1 down-regulation <i>in vivo</i> in response to DMXAA. (<b>C</b>) 344SQ-ELuc whole tumor lysates from mice treated with 25 mg/kg DMXAA (N = 3), or DMSO vehicle (N = 4), also demonstrate a DMXAA-induced drop in Arg-1 and increase in iNOS transcripts. (<b>D</b>) Representative tumor sections stained with anti-Arg-1 showing a drop in Arg-1 staining as early as 6 hours post DMXAA. Scale bars = 100 µm. Data are the mean ± SEM.</p

DMXAA showed differential tumor site-specific vascular disruption in a human breast cancer xenograft model.

<p>(<b>A</b>) BLI of MDA-MB-231-Luc2 subcutaneous tumors in NIH-III (<i>nu/nu; bg/bg</i>) mice 30 days post-cell inoculation, or metastases at day 21 post-cell inoculation. Mice were randomized into two groups (N = 10 each) and administered DMXAA or vehicle control, and then re-imaged at 6 and 24 hours. ROI encompassing the tumors or whole body were used to quantify photon emission rates. A significant drop in signal intensity in subcutaneous tumors treated with DMXAA, however, there was no change in light emission from DMXAA treated mice with metastatic tumors (<b>B</b>) (***p<0.001). (<b>C</b>) Representative histology of subcutaneous tumors demonstrating the presence of massive hemorrhagic necrosis in DMXAA treated mice (scale bar = 100 µm), with bone metastases (T = tumor, CB = cortical bone, TB = trabecular bone) showing only very limited regions of hemorrhage in response to DMXAA (scale bar = 50 µm). (<b>D</b>) Anti-Iba-1 staining was used to show the presence of macrophages in both subcutaneous and metastatic tumors. Data represent the mean ± SEM.</p

Differences in vascular structure were present between subcutaneous 344SQ-ELuc tumors and metastatic <i>p53^R172HΔg/+ K-ras^LA1/+</i> NSCLC.

<p>(<b>A</b>) Micro-CT 3D rendering of tumors from Microfil-perfused mice harboring 344SQ-ELuc subcutaneous tumors or <i>p53^R172HΔg/+ K-ras^LA1/+</i> lung adenocarcinoma-derived metastases (N = 6). (<b>B</b>) Vessel thickness (V.Th) represented by heat-map, red vessels ≥0.2 mm diameter (<b>C</b>) Vessel separation (V.Sp) represented by the maximal sphere-filling model (red spheres indicating a diameter of ≥2 mm between vessels). Full view and cut-plane through tumor center demonstrates markedly increased avascularity of subcutaneous tumors as compared to metastases. (<b>D</b>) Quantification of vessel density (VV/TV) confirms significantly less vessels present in subcutaneous tumors (**p<0.01). (<b>E</b>) Distribution of V.Th is represented as a percentage of total vessels, indicates similar pattern in both tumor locations (Log10 scale). (<b>F</b>) Distribution of average number of spheres as an indicator of V.Sp demonstrated significantly fewer small-diameter spheres in subcutaneous tumors (***p<0.001), indicative of ischemic and/or necrotic regions (Log¹⁰ scale). Data are represented as the mean ± SEM.</p

No differences in Evans blue permeability was present between primary lung neoplasms and subcutaneous tumors.

<p>(<b>A</b>) Whole-mount images of adenoma- and adenocarcinoma-bearing <i>K-ras^LA1/+</i> lungs and 344SQ-ELuc subcutaneous tumors following Evans Blue dye injection, or PBS control. (<b>B</b>) Fluorescence images of lung sections (phase contrast shown in grey, and reflected light in red) and subcutaneous tumor sections in (<b>C</b>) (Scale bar = 100 µm). The dye was able to extravasate in both tumor locations, suggesting that lack of permeability to small molecules did not account for the failure of DMXAA to disrupt the vasculature of primary lung neoplasms.</p

DMXAA-induced factors in M1- and M2-polarized BMDMs.

<p>Note: Brackets indicate a negative fold-change (**p≤0.01).</p