Erythrocytes as Drug Carriers in Medicine PDF
Preview Erythrocytes as Drug Carriers in Medicine
Erythrocytes as Drug Carriers in Medicine Erythrocytes as Drug Carriers in Medicine Edited by Ulrich Sprandel Kreiskrankenhaus Marktoberdorf, Germany and James L. Way Texas A&M University College Station, Texas Springer Science+Business Media, LLC Library of Congress Cataloging-in-Publication Data On file Proceedings of the Sixth Meeting of the International Society for the Use of Resealed Erythrocytes, held July 25-28, 1996, in Irsee, Germany ISBN 978-1-4899-0046-3 ISBN 978-1-4899-0044-9 (eBook) DOI 10.1007/978-1-4899-0044-9 © 1997 Springer Science+Business Media New York Originally published by Plenum Press, New York in 1997 Softcover reprint of the hardcover 1st edition 1997 1098765432 1 All rights reserved No part of this book may be reproduced, stored in a retrieval system, or transmitted in any form or by any means, electronic, mechanical, photocopying, microfilming, recording, or otherwise, without written permission from the Publisher PREFACE The sixth meeting on the use of resealed annealed red blood cells was held in Irsee, Germany by the International Society for the Use of Resealed Erythrocytes (ISURE) on July 25-28, 1996. Although earlier meetings focused on the technology toward develop ment of methods and standardization for efficient, consistent encapsulation, most of the present studies now are directed toward the application use of these carrier blood cells. Basic studies now have been directed toward exploration of commercial applications. In deed, clinical trials were initiated to evaluate the dose-response curves employing L asparagenase in human patients. Also, studies have shown the use of thrombolytic agent in erythrocyte carriers with the use of human red blood cells to provide a new conceptual ap proach in thrombolytic therapy to prevent thrombosis in individuals with higher risk fac tors. For example, with the use of carrier red blood cells, the thrombolytic agents will have a greater potential of acting on clot formation without systemic activation and thus lower the risk of hemorrhage, which is always prevalent in the thrombolytic therapy. Erythrocyte carrier systems are still quite unique and useful as specific targeting agents with a prolonged, sustained action with minimal immunologic or other toxic effects. The stability of this carrier system provides greater applications, especially of enzymes and proteins, by minimizing immunologic reactions and enhancing stability. Each of these studies are directed to minimize the toxicity so that higher doses such as IL2 can be used and it permits the use of more toxic prodrugs such as 3'-azidothymidine homodinucleotide as an anti-HIV drug. The focus of erythrocyte carriers now appears to be in the area of ap plications----especially, commercial area, which is the logical sequence as this area of en deavors matures. v CONTENTS 1. A Model for the Assessment of Human Recombinant Interleukin 2 (RIL2) Coated Erythrocytes as a Delivery System for Immunotherapy ........ . R. B. Moyes and J. R. DeLoach 2. Human Recombinant Interleukin 2 Binds to Mouse Red Blood Cells via the Erythropoietin Receptor ........................................ 13 R. B. Moyes and J. R. DeLoach 3. In Vivo Survival of Human Energy-Replete Carrier Erythrocytes. . . . . . . . . . . . . 25 M. D. Bain, B. E. Bax, P. J. Talbot, E. J. Parker-Williams, and R. A. Chalmers 4. The Entrapment of Polyethylene Glycol-Conjugated Adenosine Deaminase (Pegademase) and Native Adenosine Deaminase in Human Carrier Erythrocytes ................................................. 31 B. E. Bax, L. D. Fairbanks, M. D. Bain, H. A. Simmonds, and R. A. Chalmers 5. Use of Erythrocytes as a New Route of Administration of Fibrinolytic Agents. Preliminary Results. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 35 B. Delahousse, R. Kravtzoff, and C. Ropars 6. Synthesis, Characterization and Erythrocyte Encapsulation of an Azidothymidine Homodinuc1eotide ............................... 43 M. Giovine, S. Scarfi, A. Gasparini, E. Millo, G. Damonte, A. De Flora, M. Magnani, A. Fratemale, L. Rossi, R. Williams, and U. Benatti 7. Erythrocyte-Based Targeted Release to Macrophages of an Azidothymidine Homodinuc1eotide Prevents Retroviral Infection . . . . . . . . . . . . . . . . . . . . . 51 U. Benatti, M. Giovine, G. Damonte, A. De Flora, R. Williams, S. Gessani, G. Brandi, A. Casabianca, A. Fratemale, and y M. Magnani 8. The Entrapment of Mannose-Terminated Glucocerebrosidase (Alglucerase) in Human Carrier Erythrocytes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 59 B. E. Bax, M. D. Bain, C. P. Ward, A. H. Fensom, and R. A. Chalmers 9. Macrophage Protection by Nucleoside and Nucleotide Analogue Administration 63 L. Rossi, A. Casabianca, A. Fratemale, G. F. Schiavano, G. Brandi, A. Antonelli, and M. Magnani vii viii Contents 10. Inhibition of Murine AIDS by Combination of AZT and DDCTP-Loaded Erythrocytes ................................................. 73 A. Fraternale, A. Casabianca, L. Rossi, L. Chiarantini, G. Brandi, G. Aluigi, G. F. Schiavano, and M. Magnani 11. Red Blood Cells as a Glucocorticoids Delivery System .................... 81 M. D'Ascenzo, A. Antonelli, L. Chiarantini, U. Mancini, and M. Magnani 12. Organophosphorus Antagonism by Resealed Erythrocytes Containing Recombinant Paraoxonase ...................................... 89 J. L. Way, L. Pei, I. Petrikovics, D. McGuinn, C. Tamalinas, Q. Z. Hu, E. P. Cannon, and A. Zitzer 13. Biotinylation of Erythrocytes Prepares to Allow to Circulation-Stable Immunoerythrocytes Capable of Recognizing the Antigen ............. 93 1. C. Murciano, V. R. Muzykantov, and A. Herniez 14. Biochemical Properties of Alcohol Dehydrogenase and Glutamate Dehydrogenase Encapsulated into Human Erythrocytes by a Hypotonic-Dialysis Procedure ................................... 10 1 S. Sanz, C. Lizano, M. I. Garin, 1. Luque, and M. Pinilla 15. Influence of Chemical Modification on "in Vivo" and "in Vitro" Mouse Carrier Erythrocyte Survival and Recognition ............................. 109 J. A. Jordan, F. 1. Alvarez, J. C. Murciano, A. Lotero, A. Herraez, M. C. Tejedor, J. Luque, J. R. DeLoach, and 1. C. Diez 16. Rat Carrier Erythrocytes Circulate and Arrive to Organs ................... 119 F. 1. Alvarez, 1. A. Jordan, 1. C. Murciano, J. Luque, A. Herraez, J. C. Diez, and M. C. Tejedor 17. Encapsulation of Alcohol Dehydrogenase and Acetaldehyde Dehydrogenase into Human Erythrocytes by an Electroporation Procedure ................ 129 C. Lizano, S. Sanz, P. Sancho, 1. Luque, and M. Pinilla 18. Pharmacokinetics of Doxorubicin in Patients with Lymphoproliferative Disorders after Infusion of Doxorubicin-Loaded Erythrocytes. . . . . . . . .. 137 F. I. Ataullakhanov, V. G. Isaev, A. V. Kohno, E. V. Kulikova, E. N. Parovichnikova, V. G. Savchenko, and V. M. Vitvitsky 19. Binding of Daunorubicin and Doxorubicin to Erythrocytes Treated with Glutaraldehyde ............................................... 143 F. I. Ataullakhanov, E. V. Kulikova, and V. M. Vitvitsky Index 149 1 A MODEL FOR THE ASSESSMENT OF HUMAN RECOMBINANT INTERLEUKIN 2 (RIL2) COATED ERYTHROCYTES AS A DELIVERY SYSTEM FOR IMMUNOTHERAPY R. B. Moyes and J. R. DeLoach USDA-ARS-FAPRL College Station, Texas, 77845 1. ABSTRACT A carrier system for IL2 is needed in order to circumvent the toxicity associated with high dose interleukin 2 (lL2) administration and its rapid clearance from circulation. Erythrocytes (RBC) coated with recombinant interleukin 2 (rIL2) provide a means of de livering IL2 into the system in a continuous, low dose manner which in turn maintains a low, potentially non-toxic, IL2 concentration. Murine RBC coated with rIL2 (RBC-rIL2) induce cytotoxicity (21-31%) upon cytotoxic testing of spleens cells stimulated in vivo. Using the murine Meth A sarcoma model, the effectiveness of this RBC-rIL2 vehicle is demonstrated in vivo by a 84% reduction in tumor size as compared to the soluble rIL2 treated mice. Moreover, the RBC-rIL2 vehicle is able to induce tumoricidal cytotoxicity with very low rIL2 concentrations (about 10,000 LU. ofrIL2 per mouse). These results in dicate that rIL2 retains its biological activity when bound to the RBC and therefore could prove useful as a therapeutic delivery system for cancer treatment. 2. INTRODUCTION Interleukin 2 (lL2), a lymphocytotrophic hormone secreted exclusively by stimulated T cells, has been characterized to be of major importance in both cell mediated and humoral im munity. IL2 acts on several different cell types including natural killer (NK) cells, B cells, and T cells resulting in cellular proliferation and augmentation of their immunological functions. In the case ofNK cells, both secretory and cytolytic capabilities are markedly enhanced. NK cells release a repertoire of cytokines as a result of IL2 introduction including interferon gamma (IFN-y), granulocyte-macrophage colony stimulating factor (GM-CSF), and tumor necrosis factor (TNF-a.,P)(Ritz 1988, Kaplan 1992), all which in turn stimulate monocyte and 2 R. B. Moyes and J. R. DeLoach macrophage responses. Thus IL2, in concert with an array of stimulated cells and lymphoki nes, is an important immunological defense against infection and tumorigenic growth. Lymphokine therapy, especially IL2 therapy, is being used experimentally in high dose regimes to treat cancer (Lotze 1986, Rosenberg 1987). These clinical tests have shown that IL2 administration can mediate the regression of established tumors especially renal cell carcinoma and malginant melanoma (Sosman 1993) in both experimental animal models and humans. However, high dose IL2 immunotherapy is toxic to the patient result ing in capillary leak syndrome due to the release of other cytokines, most notably TNF-a, eventually leading to hypotension (Mier 1990). It is difficult to maintain plasma IL2 levels due to the rapid clearance of IL2 from circulation. It has been previously shown that, regardless of route of injection, by two hours post injection, no IL2 activity is detectable in the serum (Cheever 1986). Anti- tu mor effects of IL2 correlate more strongly with low sustained IL2 plasma levels than with intermittent peak IL2 plasma levels (Cheever 1985). Therefore, any method that sustains delivery of IL2 at a low constant level would circumvent high dose toxicity and at the same time probably increase the therapeutic index ofIL2. The purpose of this investigation was to observe the biological effects produced by IL2-coated erythrocytes both in vitro and in vivo in the murine system. 3. MATERIALS AND METHODS 3.1. Media RPMI 1640 (JRH Biosciences, Lenexa,KS) was supplemented with 2mM L-glutamine (Sigma, St. Louis, MO), and 5% Antimycotic/Antibiotic solution (Sigma) in all cases, and with 10% fetal bovine serum (FBS)(JRH Biosciences) (RPMI-FBS) for cell culture. 3.2. Cell Lines The WEHI 164 cell line, a fibrosarcoma originally induced by subcutaneous injec tion of3-methylcholanthrene in Balb/c mice (ATCC CRL 1751, Rockville, MD) served as a syngeneic tumor line for in vivo tumor studies. The malignant murine cell line, Y AC-l (ATCC TIB 160) was used in the in vitro cytotoxicity assays. Both cell lines were main tained in culture in RPMI-FBS. An IL2-dependent cytotoxic T cell line, CTLL (ATCC TIB 214) was maintained on RPMI-FBS with the addition of T-stim (Collaborative Biomedical Products, Bedford, MA), an IL2 supplement. 3.3. Cytokines Recombinant human interleukin 2 (rIL2) was purchased from Cellular Products (Buffalo, NY; supplied by Cetus Corporation). Each vial contained 100ug of rIL2 (15.4 kD) and was stabilized in a 2% FBS solution. 3.4. Mice Mouse spleens were obtained from female Balb/c mice which had been purchased from., Harlan Laboratories (Houston, Texas). The mice were housed and fed ad libitum ac cording to the USDA guidelines established by the Animal Care and Use Committee. A Model for the Assessment of Human Recombinant Interleukin 2 (rIL2) 3 3.5. Mitogens Phytohemagglutin P (PHA) and lipopolysaccharide (LPS) were both obtained from Sigma and used at a concentration of 1u glwell in Falcon 96 well plates (Baxter Scientific Products) or 1u g/ml when added to tissue culture flasks. 3.6. Isolation of Erythrocytes Mice were anesthetized with CO and blood was collected via cardiac puncture. The 2 RBC were washed 3 times with a PBS buffer containing 10 mM glucose, 4 mM MgCI2, 5 mM adenosine, and 5 mM inosine (Sigma) (complete buffer referred to as MBA) at 400 x g at 4°C for 10 minutes. The buffy coat was removed and discarded after the first wash. Erythrocytes were counted using a computerized Coulter counter (Hileah, FL) and were resuspended in MBA to a concentration of 1010 cells per ml. 3.7. Adsorption ofIL2 to Erythrocytes RBC were resuspended in sterile eppendorf tubes (USA Scientific, Ocala, FL) to 1010 cells per ml and incubated with rIL2 at a ratio of 72,000 LU.l109 erythrocytes (unless otherwise stated) with gentle rotation for 30 min at 4°C. The cells were then washed 3 times with 4 volumes of MBA. This technique yielded approximately 8000-12,000 LU. of rIL2 coated on the surface per 109 erythrocytes as determined by the CTLL bioassay. 3.8. In Vitro Cytotoxicity Assay For radiolabeling of the target cells, YAC-l mouse myeloma cells were suspended to 107 cells/ml of RPMI-FBS and incubated with 100 ul/ml of 51 Chromium (200-900 Ci/g, ICN Radiochemicals, Irvine, CA) at 37°C for 120 minutes. The target cells were washed 3 times, diluted to a concentration of 106 cells/ml, and dispensed at 100 ul per well in Falcon 96 well U bottom microtiter plates (Baxter Scientific Products). One hundred ul of the spleen cells at fi nal effector: target ratio of 25: 1 were then dispensed in the microtiter plate. Background or spontaneous release was determined by incubating labeled target cells in RPMI-FBS alone, and total release was determined by incubating labeled target cells in 100 ul of 1% sodium do decyl sulfate (Biorad, Richmond, CA) solution. Plates were centrifuged at 900 rpm for 5 min utes and then incubated for 24 hr in a 5% CO incubator. After incubation, the plates were 2 again centrifuged and 150 ul from each well was collected and counted in a LKB 1282 Com pugamma gamma counter (Gaithersburg, MD). Cytotoxicity was quantitated as follows: )/ f ·fi C I experimental release cpm - spontaneous release cpm x 100 ~oo spec I IC r re ease = ---''---------=-----''-----------''---- total release cpm - spontaneous release cpm 3.9. In Vivo Activation Balb/c mice were injected i.p. with 100 ul of one of the following preparations: nor mal RBC (109RBC/mouse), RBC-rIL2 (l09 RBC + 8000-12,000 I. U. IL2/mouse), soluble rlL2 (72,000 IV/mouse), or MBA (lOOullmouse), unless otherwise indicated. Depending on the experiment, mice received either 1 to 4 injections at one injection per week, or 4 in-
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