[CANCER RESEARCH 64, 3155–3161, May 1, 2004] High-Resolution Magnetic Resonance Imaging of Disparities in the Transcapillary Transfer Rates in Orthotopically Inoculated Invasive Breast Tumors Maya Dadiani, Raanan Margalit, Noa Sela, and Hadassa Degani Department of Biological Regulation, Weizmann Institute of Science, Rehovot, Israel ABSTRACT In vivo mapping of the transcapillary fluxes in tumors can help predict the efficacy of delivery of blood-borne anticancer drugs. These fluxes are primarily affected by the vascular permeability and the pressure gradi- ents across the blood vessels’ walls. We describe herein high-resolution dynamic contrast-enhanced magnetic resonance imaging of the influx and outflux transcapillary transfer rates in vivo in invasive MDA-MB-231 tumors orthotopically inoculated in severe combined immunodeficient mice. The tumors were noted for rapid growth, impaired drainage of fluid, and subsequent formation of cysts. Consequently, the time evolution of the contrast enhancement, induced by i.v. injection of Gadolinium diethylene- triamine-penta-acetate, exhibited two distinct patterns: transcapillary transfer in the cellular regions and simple diffusion in the cyst fluid. Both processes were analyzed at pixel resolution applying to each a physiolog- ical model and a corresponding algorithm. In the cellular region, the influx and outflux transcapillary transfer rates decreased during tumor growth; however, an increased disparity between the transfer constants was observed, with the outflux rate exceeding the influx rate. This quan- titative spatial and temporal mapping of this disparity can provide a means to assess the physiological barriers to tracer delivery. It is hypoth- esized that both the increased disparity in transcapillary transfer rates and impaired fluid drainage in these tumors could arise from the devel- opment of interstitial hypertension. INTRODUCTION Tumor growth and development relies on the formation of a cap- illary network that serves to supply nutrients and oxygen to the cancerous tissue, as well as to provide escape routes by which inva- sive cells can metastasize to distant sites (1). Tumor vasculature, however, is not an ideal network; its rapid and irregular formation results in a disorganized and tortuous architecture, as well as fragile and leaky vessels that lead to both impaired oxygen and nutrient perfusion and impaired drainage of fluids (2). As a result, tumor tissue often develops several abnormalities such as hypoxic and acidic loci and large necrotic areas, as well as the formation of cysts or edemas and regions of elevated interstitial fluid pressure (IFP) (3). The latter feature, in particular, is a major obstacle to the successful delivery of chemotherapeutic agents to the tumors. Comprehensive studies by Jain et al. (3, 4) have demonstrated that experimental tumors in animal models and several human tumors develop elevated levels of IFP during growth. Overall, the aforementioned vascular and tissue abnormalities lead to the inadequate delivery of diagnostic and therapeutic agents to solid tumors (3). Therefore, quantifying these abnormalities is of major importance for predicting the efficacy of chemotherapeutic drug de- livery, in particular, in breast cancer because neoadjuvant, preopera- tive treatment is increasingly being used (5, 6). Quantification of transport-related parameters in vivo during tumor growth and progres- sion, as well as in response to therapy requires the use of high- resolution noninvasive imaging methods. Although in vivo micros- copy provides the desired spatial resolution required to monitor events at the cellular level, this method is depth limited. Dynamic contrast- enhanced magnetic resonance imaging (MRI) effectively provides means to measure the physiological parameters of perfusion in a noninvasive manner (Refs. 7–11 and references cited therein). This method has been extensively used to diagnose breast and other tumors in preclinical (12, 13) and clinical (9, 14) studies. Dynamic acquisition of images following injection of a contrast agent enables the tracking of tracer uptake and clearance over time, as well as providing infor- mation about tissue perfusion. In a recent study, the uptake kinetics of Gadolinium-diethylene-triamino-pentaacetic acid (Gd-DTPA) as measured by dynamic contrast-enhanced MRI were shown to corre- late with the delivery of the anticancer agent phenyl acetate (15). The uptake of diffusible tracers such as Gd-DTPA has been exten- sively studied, using model-based equations to describe physiological parameters (16 –18). Here, we used the generalized scheme proposed by Tofts (18) that uses a compartmental model to describe the transfer of the contrast agent from the intravascular to the interstitial space and back, thus yielding the transcapillary influx and outflux transfer constants, respectively. In most previous studies, it was assumed that the transfer constant is the same in both directions; however, disparate influx and outflux transfer constants more accurately describes the dispersal of the contrast agent into and out of the interstitium, partic- ularly when there are differences in diffusion or pressure on either side of the capillary walls (18). This disparity is more likely to be present in tumors characterized by high interstitial pressure. In this article, we present data from MRI studies of Gd-DTPA perfusion and diffusion in MDA-MB-231 breast tumors orthotopically inoculated in severe combined immunodeficient (SCID) mice. These tumors exhibited rapid growth accompanied with impaired drainage and formation of cysts and revealed invasive features which includes the formation of metastases. Analyses of dynamic contrast-enhanced MR images of the tumors revealed two distinct patterns of the contrast agent uptake: transcapillary exchange in the cellular regions and diffusion in the cyst. Each process was, therefore, analyzed according to a related model-based equation to yield the appropriate physiolog- ical parameters. Monitoring changes in these parameters enabled us to follow alterations in tumor perfusion during growth. MATERIALS AND METHODS Mice and Tumors. Human MDA-MB-231 breast cancer cells were ob- tained from the American Type Culture Collection (Rockville, MD) and were cultured as recommended by the supplier. Cells (2  10 6 to 1  10 7 ) suspended in 0.2– 0.5 ml PBS, were inoculated into the mammary fat pad of female CB-17 SCID mice, 7–10 weeks old. During the experiments, mice were anesthetized by inhalation of 1% Isoflu- rane (Medeva Pharmaceuticals PA, Inc., Bethlehem, PA) in an O 2 :N 2 O (3:7) mixture, applied through a nose cone. After euthanasia, tumors were surgically removed, fixed with 4% formaldehyde, and sectioned in a plane parallel to that of the MRI. Tumors were then embedded in paraffin, sectioned to obtain 4-m Received 8/26/03; revised 2/5/04; accepted 2/19/04. Grant support: Israel Science Foundation Grant 648/01 and by Sir David Alliance CBE, United Kingdom. H. Degani is the incumbent of the Fred and Andrea Fallek Professorial Chair for Breast Cancer Research and heads the Willner Family Center for Vascular Biology. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. Note: Supplementary data for this article can be found at Cancer Research Online (http://cancerres.aacrjournals.org). Requests for reprints: Hadassa Degani, Department of Biological Regulation, Weiz- mann Institute of Science, Rehovot 76100, Israel. Phone: 972-8-934-3920; Fax: 972-8- 934-4186; E-mail: hadassa.degani@weizmann.ac.il. 3155 on July 4, 2015. © 2004 American Association for Cancer Research. cancerres.aacrjournals.org Downloaded from