[CANCER RESEARCH 64, 2572–2579, April 1, 2004] Distribution of Liposomes into Brain and Rat Brain Tumor Models by Convection- Enhanced Delivery Monitored with Magnetic Resonance Imaging Ryuta Saito, 1 John R. Bringas, 1 Tracy R. McKnight, 3 Michael F. Wendland, 3 Christoph Mamot, 2 Daryl C. Drummond, 4,5 Dmitri B. Kirpotin, 4,5 John W. Park, 2,5 Mitchel S. Berger, 1 and Krys S. Bankiewicz 1 1 Department of Neurological Surgery, Brain Tumor Research Center, 2 Division of Hematology-Oncology, and 3 Department of Radiology, University of California, San Francisco, San Francisco, California; 4 California Pacific Medical Center Research Institute, Liposome Research Laboratory, San Francisco, California; and 5 Hermes Biosciences, Inc., South San Francisco, California ABSTRACT Although liposomes have been used as a vehicle for delivery of thera- peutic agents in oncology, their efficacy in targeting brain tumors has been limited due to poor penetration through the blood-brain barrier. Because convection-enhanced delivery (CED) of liposomes may improve the ther- apeutic index for targeting brain tumors, we conducted a three-stage study: stage 1 established the feasibility of using in vivo magnetic reso- nance imaging (MRI) to confirm adequate liposomal distribution within targeted regions in normal rat brain. Liposomes colabeled with gadolin- ium (Gd) and a fluorescent indicator, 1,1-dioctadecyl-3,3,3,3-tetrameth- ylindocarbocyanine-5,5-disulfonic acid [DiI-DS; formally DiIC 18 (3)-DS], were administered by CED into striatal regions. The minimum concen- tration of Gd needed for monitoring, correlation of infused volume with distribution volume, clearance of infused liposome containing Gd and DiI-DS (Lip/Gd/DiI-DS), and potential local toxicity were evaluated. After determination of adequate conditions for MRI detection in normal brain, stage 2 evaluated the feasibility of in vivo MRI monitoring of liposomal distribution in C6 and 9L-2 rat glioma models. In both models, the distribution of Lip/Gd/DiI-DS covering the tumor mass was well defined and monitored with MRI. Stage 3 was designed to develop a clinically relevant treatment strategy in the 9L-2 model by infusing liposome con- taining Gd (Lip/Gd), prepared in the same size as Lip/Gd/DiI-DS, with Doxil, a liposomal drug of similar size used to treat several cancers. MRI detection of Lip/Gd coadministered with Doxil provided optimum CED parameters for complete coverage of 9L-2 tumors. By permitting in vivo monitoring of therapeutic distribution in brain tumors, this technique optimizes local drug delivery and may provide a basis for clinical appli- cations in the treatment of malignant glioma. INTRODUCTION Liposomes (phospholipid bilayers formed into spheres in the pres- ence of water that can be made to incorporate a variety of agents) are a vehicle for administering therapeutic agents, including drugs and genes, to areas of the body afflicted with cancer (1, 2). Recently, efforts have been made to increase the vehicular efficiency of lipo- somes and to direct therapeutic agents to specific target sites (3, 4). Preclinical studies using drug-encapsulated liposomes have shown improvement in the sustained release of the drug, prolongation of the drug’s half-life, and an increase in the therapeutic index of corre- sponding drugs (5). Although immunoliposomes using antibody frag- ments for molecular targeting have shown promising results (6, 7), the potential effectiveness of such immunoliposomes in targeting tumors of the central nervous system (CNS) has not been established. Sys- temic administration may not achieve satisfactory penetration of the blood-brain barrier, and local injection cannot achieve optimum dis- tribution. Convection-enhanced delivery (CED) is a direct intracranial drug delivery technique that utilizes a bulk-flow mechanism to deliver and distribute macromolecules to clinically significant volumes of solid tissues (8, 9). This approach offers a greater volume of distribution than simple diffusion and is designed to direct a drug to a specific target site. As compared with systemic delivery, the CED of lipo- somes carrying chemotherapeutic drugs bypasses the blood-brain bar- rier (8), provides a larger distribution of liposomes within the target site, allows for a locally sustained release of drugs, and minimizes systemic exposure, thereby producing fewer side effects. Despite advances in neurosurgical techniques and in radiation and drug therapies, the mean survival for patients who have a malignant glioma is less than 12 months, and only 20% of patients survive for more than 2 years. More than 80% of patients with this disease experience local recurrence of tumor, which leads to their death (10, 11). Therefore, development of new local management strategies such as the CED of therapeutic liposomes may provide a therapeutic advantage in the adjuvant medical management of gliomas. To develop a CED method for the administration of liposomal therapeutics in the treatment of malignant glioma, our study was designed in three stages. Stage 1 studies were designed to establish the feasibility of using direct in vivo magnetic resonance imaging (MRI) of liposomal delivery to confirm adequate drug distribution within targeted CNS regions. MRI was used to visualize liposomes incorpo- rating the contrast agent gadodiamide [a stable gadolinium (Gd) chelate], which were dispensed by CED to obtain robust distribution in the CNS. The following issues related to the successful application of this technology were assessed in normal brain parenchyma of intact rats: (a) monitoring of the CED infusion of liposomes containing Gd and a fluorescent indicator, 1,1'-dioctadecyl-3,3,3',3'-tetramethylin- docarbocyanine-5,5'-disulfonic acid [DiI-DS; formally DiIC 18 (3)- DS], into rat brain hemispheres to evaluate its feasibility and deter- mination of the lowest concentration of contrast agent required to show the targeted distribution area; (b) evaluation of the liposome containing Gd and DiI-DS (Lip/Gd/DiI-DS) distribution when infus- ing a defined volume at intervals from 5 to 40 l using MRI moni- toring and histological detection of the fluorescence; (c) a time-course evaluation to determine retention of Gd-containing liposomes in the brain after infusion by CED as represented by the liposomal clearance of the magnetic resonance (MR) signal generated by Lip/Gd/DiI-DS; and (d) possible adverse effects of Lip/Gd/DiI-DS infused into the CNS by CED. After several conditions for MRI detection of Lip/Gd/DiI-DS were established in normal brain parenchyma of intact rats, stage 2 studies evaluated the feasibility of in vivo MRI of liposomal distribution in two morphologically dissimilar rat brain tumor models, C6 and 9L-2. These studies addressed the following issues: (a) comparison of Lip/Gd/DiI-DS distribution in normal brain and brain tumor tissues to assess the feasibility of using CED administration of liposomes for the treatment of malignant glioma; and (b) determination of the distribu- Received 11/19/03; revised 1/7/04; accepted 1/28/04. Grant support: National Cancer Institute Specialized Programs of Research Excel- lence grant (M. Berger, K. Bankiewicz, and J. Park) and Accelerate Brain Cancer Cure (K. Bankiewicz). 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. Requests for reprints: Krys S. Bankiewicz, Department of Neurological Surgery, University of California at San Francisco, 1855 Folsom Street, Mission Center Building Room 230, San Francisco, California 94103. Phone: (415) 502-3132; Fax: (415) 514- 2177; E-mail: kbank@itsa.ucsf.edu. 2572 Research. on July 7, 2015. © 2004 American Association for Cancer cancerres.aacrjournals.org Downloaded from