Angiogenesis Inhibitors in a Murine Neuroblastoma Model: Quantitative Assessment of Intratumoral Blood Flow with Contrast-enhanced Gray-Scale US

Abstract

To quantify intratumoral ultrasonographic (US) contrast agent flow at gray-scale imaging as a measure of functional tumor vascularity in an orthotopic murine neuroblastoma model treated with angiogenesis inhibitors. After Institutional Animal Care and Use Committee approval, retroperitoneal neuroblastomas were established in mice with unmodified NXS2 cells (n = 13) or with cells engineered to overexpress an angiogenesis inhibitor--either tissue inhibitor of matrix metalloproteinase-3 (n = 22) or a truncated soluble form of the vascular endothelial growth factor receptor-2 (truncated soluble fetal liver kinase-1; n = 13). When tumors were approximately 600 mm3, contrast material-enhanced gray-scale US was performed, and the imaging was recorded on cine clips. Regions of interest within tumors were analyzed off-line to determine postcontrast change in signal intensity (SI) from baseline to initial peak (deltaSI), rate of SI increase from baseline to initial peak (RSI), and contrast material washout. The Mann-Whitney test was used to evaluate potential differences in these US parameters between treatment groups. The mean intratumoral endothelial cell (CD34) and pericyte (smooth muscle actin [SMA]) counts at immunohistochemical analysis were also evaluated. Spearman correlation test was used to investigate the relation between US parameters and these histologic markers. The deltaSI and RSI were lower in tumors overexpressing an angiogenesis inhibitor than in control tumors (all P < .03). Contrast material washout did not differ between groups. For the entire cohort, the RSI correlated with the immunohistochemical assessment of tumor vascularity (SMA and CD34 counts) (P < .003). Quantification of intratumoral flow of a US contrast agent at gray-scale imaging shows promise for monitoring tumor vascular response to antiangiogenic therapy.