ANTI-GLOBULIN RESPONSES TO RAT AND HUMANIZED CAMPATH-1 MONOCLONAL ANTIBODY USED TO TREAT TRANSPLANT REJECTION 1 PERPETUA R.U.B. REBELLO, 2,3 GEOFFREY HALE, 2,4 PETER J. FRIEND, 3 STEPHEN P. COBBOLD, 2 AND HERMAN WALDMANN 2 Sir William Dunn School of Pathology, University of Oxford, UK OX1 3RE, and the Department of Surgery, University of Cambridge, Addenbrookes Hospital, UK CB2 2QQ Background. Antiglobulin responses are a signifi- cant limitation to the repeated use of murine monoclo- nal antibodies for treatment of transplant rejection. It is hoped that these might be largely overcome by using antibodies genetically engineered to resemble human antibodies. Methods. We have compared the responses in pa- tients treated with the CD52 monoclonal antibodies CAMPATH-1G (rat IgG2b) or its humanized derivative, CAMPATH-1H (human immunoglobulin G1). Results. A majority of patients (15 of 17) made re- sponses to the rat antibody, but there were no detect- able responses to the humanized antibody (0 of 12). Conclusions. Although anti-idiotype responses are theoretically possible against humanized therapeutic antibodies and are especially likely to be provoked by cell-binding antibodies, these data show that human- ization offers a significant reduction in immunogenic- ity, potentially allowing repeat courses of treatment. Monoclonal antibodies are becoming accepted as useful agents in the treatment of cancer, autoimmune disease, transplant rejection, and for other indications. From the early days, it was recognized that immune responses against the foreign protein could limit the potential benefit of this therapy. This is most clearly exemplified with the mouse antibody OKT3, licensed since 1986 for the treatment of renal transplant rejection. Despite the high level of immuno- suppressive therapy given to transplant patients, antiglobu- lin responses to OKT3 are seen in about 50% of patients after a single course, and these can block its therapeutic effect if the antibody is used again (1). Various strategies have been proposed to create less immunogenic antibodies. They can be loosely divided into two categories: (1) genetically engineered constructs containing more or less human Ig but with the binding sites from a murine antibody, and (2) completely “human” antibodies (e.g. from phage-display libraries). The practical distinction is not so clear-cut, because every anti- body, whether “human” or “humanized” contains unique id- iotypic determinants. However, there are not many clinical data to indicate the actual likelihood of anti-idiotype re- sponses in a particular setting and few direct comparisons of antiglobulin responses against a chimeric antibody and its murine parent (2). Here we present the results of a study using CAMPATH-1 (CD52) antibodies for treatment of transplant rejection. CAMPATH-1G is a rat immunoglobulin (Ig*)G2b monoclonal antibody directed against the CD52 antigen, which is a GPI- anchored glycoprotein abundantly expressed on lymphocytes and monocytes (3). The antibody has a remarkable ability to lyse cells in vitro and in vivo and has been used to treat kidney transplant rejection and lymphocytic leukemia and to prevent graft-versus-host disease (4, 5). CAMPATH-1H is a human IgG1, the first therapeutic monoclonal antibody to be humanized (6). Like CAMPATH-1G, it depletes lymphocytes in vivo and has been used in a range of clinical trials, includ- ing treatment of transplant rejection (7). This offered a unique opportunity to compare the ability of the rat and the humanized antibodies to provoke anti-isotype or anti-idio- type responses. MONOCLONAL ANTIBODIES CAMPATH-1G was prepared from hybrid myeloma cells cultured in a hollow fibre fermentor (Acusyst-Jr, Cellex, Min- neapolis). It was purified by fractionation with ammonium sulphate. CAMPATH-1H was supplied by the Wellcome Foundation Ltd., Beckenham, UK. It was prepared from Chi- nese hamster ovary transfectants cultured in stirred tanks and purified by affinity chromatography on Protein A, ion exchange chromatography, and size exclusion chromatogra- phy on Superdex-200. PATIENTS There were 17 patients treated with CAMPATH-1G, 5–10 mg/day for 6 –10 days (4). Four were treated for first rejection of kidney allografts and eight for steroid-resistant or recur- rent rejection of kidney allografts. Five patients were treated for rejection of liver allografts, resistant to OKT3, anti-thy- mocyte globulin, and/or high-dose steroids, and three of these were having second transplants. All patients were given hy- drocortisone (100 –200 mg) and chlorpheniramine (10 mg) in advance of the first antibody infusion with the aim of reduc- ing the infusion-related cytokine-release phenomenon. Twelve patients were treated with CAMPATH-1H for first rejection episodes of kidney allografts; 6 with 10 mg/day for 7 days, and 6 with 6 mg/day for 5 days (7). Premedication to prevent infusion-related reactions was not given, but symp- tomatic treatment was at the discretion of the physician. Serum was collected at the beginning of treatment and be- tween 7 and 47 days. Baseline immunosuppression in both groups was the same, a standard triple-drug regimen of cyclosporin, azathioprine and steroids, although in the 6 low-dose CAMPATH-1H pa- tients cyclosporin was stopped during the antibody treat- 1 Supported by the UK Medical Research Council, the Kay Kendall Leukaemia Fund, Wellcome Foundation Ltd., LeukoSite Inc. and the EP Abraham’s Trust. 2 Sir William Dunn School of Pathology, University of Oxford. 3 Department of Surgery, University of Cambridge. 4 Address correspondence to: Geoffrey Hale, PhD, Therapeutic Antibody Centre, Old Road, Headington, Oxford, UK OX3 7JT. * Abbreviation used: Ig, immunoglobulin. BRIEF COMMUNICATIONS November 15, 1999 1417