The development of anti-angiogenesis agents and the clinical evaluation of anti-angiogenic therapy have produced a need for non-invasive imaging methods to monitor response. The fundamental hypothesis of this project is that tumor neovascularity can be quantified and monitored non-invasively using contrast-enhanced ultrasound imaging modes. Studies will be performed in rats implanted with human breast and glioma tumors (cell lines NMU and C6, respectively). These tumors are rapidly growing with marked neovascularity and typically reach an average diameter of 10 mm by 3 to 4 weeks. Half the rats will receive orthotopic tumor implantations and in the remaining animals, heterotopic implantations will be performed. A microbubble based ultrasound contrast agent will be used in conjunction with power Doppler, flash echo imaging (FEI), excitation enhanced imaging (EEl) and pulse inversion harmonic imaging (HI) to detect and quantify tumor perfusion and vascular volume. Contrast enhanced ultrasound evaluation of tumors will be performed on day 7, 14 or 21 after tumor inoculation. Initial experiments will be performed on 432 mice (4 ultrasound modes x 2 tumors x 2 implantation techniques x 3 time points x 9 rats/group). Immediately after the ultrasound study, the animals will be sacrificed. Tumor vascularity will be assessed by staining for endothelial ceils (CD31), vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF) as well as cyclooxygenase-2 (COX-2), and correlated with the ultrasound results. Based on this analysis the best implantation and the best ultrasound technique will be selected. Next, 54 rats will be studied after treatment with anti-angiogenesis therapy (1 ultrasound mode x 2 tumors x 1 implantation technique x 3 time points x 9 rats/group). The same comparison between contrast enhanced ultrasound and pathology will be performed. If successful, this project will permit non-invasive ultrasound evaluations of anti-angiogenic tumor therapies in animals as well as humans.
