Angiogenesis, the formation of new blood vessels, is involved in many pathological processes like cancer, ischemic disease, atherosclerosis, and chronic inflammation . Inhibition of angiogenesis in cancer is a novel strategy to stop tumor growth/progression. in the angiogenic cascade different cell surface receptors are expressed at the endothelium of the new vasculature. The avb3-integrin is such a receptor which is associated with angiogenesis. The non-invasive in vivo detection of this integrin would allow one to be able to monitor angiogenesis and to be able to follow the effect of anti-angiogenic therapies. Several studies have used the tripeptide sequence arginineglycine-
aspartic acid (RGD) conjugated with a radiolabel to image the expression of this integrin with nuclear imaging methods [2,3]. In this study, we aim to non-invasively image the expression of avb3 in tumor bearing mice with Magnetic Resonance Imaging (MRI). Therefore, we developed and used MR-detectable and fluorescent liposomes, which carry over 300 RGD-moieties per liposome. The in vivo MRI findings were validated with ex
vivo fluorescence microscopy. Material and Methods Paramagnetic and fluorescent liposomes were prepared as described previously . The cyclic RGD and control RAD-peptide were conjugated to maleimide-PEG-DSPE incorporated in the liposomes. The specificity of the liposomes was determined on HUVEC with MRI and fluorescence microscopy. Tumor bearing mice were anesthetized with an isoflurane/airmixture and an infusion line was put into the tail vein. Next, the mice were placed in a 6.3 T MRI scanner. A T1-map and a T1-weighted image were generated before the contrast agent was injected and at 5 time points after the contrast agent was injected. Next, the mice were euthanized and the tumor was dissected and frozen in isopentane. Sections of tissue (10-lm thickness) were prepared and fluorescence microscopy was performed. Results Both MRI and fluorescence microscopy revealed a strong association of the RGD-liposomes to avb3 expressing HUVEC, while RADliposomes did not. MRI on tumor bearing mice howed contrast enhancement of the tumor after injecting the contrast agent, both for RGD-liposomes and RAD-liposomes. Fluorescence microscopy revealed a distinct difference between tumors of mice that were injected with RGD-liposomes or RAD-liposomes. In case of RADliposomes a diffuse pattern of fluorescence was found around tumor blood vessels in the tumor tissue. This can be ascribed to extravasation of the liposomes in the tumor tissue. In contrast, RGD-liposomes were exclusively found was associated with blood vessels. Conclusions: This study demonstrates that molecular MR imaging of angiogenesis is possible by using a targeted contrast agent specific for the avb3-intergin.
|Title of host publication||Abstracts of the 15th Endothelial Cell Research Symposium Maastricht Auditorium, Maastricht, The Netherlands 23 November 2004|
|Place of Publication||Dordrecht|
|Publication status||Published - 2004|